AGROECOLOGICAL ANALYSIS OF ARTHROPODS INVOLVED IN …

AGROECOLOGICAL ANALYSIS OF ARTHROPODS INVOLVED IN MANGO POLLINATION IN SOUTH FLORIDA

By

MATTHEW QUENAUDON

A THESIS PRESENTED TO THE GRADUATE SCHOOL

OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF

MASTER OF SCIENCE

UNIVERSITY OF FLORIDA

2019

To my parents

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ACKNOWLEDGMENTS

I am grateful to my major professor Dr. Daniel Carrillo, for his guidance, support,

and prowess during my time as a graduate student at the University of Florida. Dr.

Carrillo was always patient, thoughtful, and provided his insights while allowing me the

intellectual freedom to shape my own research. I also want to thank the other members

of my committee, Dr. Zachary Brym, Dr. Jonathan Crane, Dr. Rachel Mallinger, and Dr.

Catharine Mannion whose expertise and contributions greatly improved this study. I

thank Alejandra Canon and Mariane Ruviéri for their contributions to data collecting and

analyzing. Thank you to Dr. Gary Steck for his aid in the identification of insects and Dr.

Alexandra Revynthi for her statistical help. I am grateful to everyone in the Tropical Fruit

Entomology lab, including Jose Alegria, Luisa Cruz, Rita Duncan, and Octavio Menocal

who helped and created a positive work environment. Lastly, I am thankful to my family

for their support and loving encouragement, providing me the motivation and mental

fortitude to complete my study.

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TABLE OF CONTENTS page

ACKNOWLEDGMENTS .................................................................................................. 4

LIST OF TABLES ............................................................................................................ 7

LIST OF FIGURES .......................................................................................................... 8

ABSTRACT ................................................................................................................... 10

CHAPTER

1 LITERATURE REVIEW .......................................................................................... 12

Origin, Distribution, and Importance of Mangifera indica ........................................ 12

Reproductive Physiology and Floral Biology ........................................................... 14

Insect Pollinators .................................................................................................... 16 Objectives of Master of Science Thesis Research .................................................. 22

2 MOST FREQUENT ARTHROPOD VISITORS ON ‘KEITT’ MANGO (MANGIFERA INDICA) FLOWERS IN SOUTH FLORIDA ...................................... 23

Introduction ............................................................................................................. 23

Material and Methods ............................................................................................. 25

Results .................................................................................................................... 28

Order Diptera .................................................................................................... 29 Chloropidae ................................................................................................ 30

Drosophilidae ............................................................................................. 30 Sciaridae .................................................................................................... 31 Muscidae.................................................................................................... 31

Syrphidae ................................................................................................... 31 Calliphoridae .............................................................................................. 31 Ceratopogonidae ....................................................................................... 32

Order Coleoptera .............................................................................................. 32 Cryptophagidae .......................................................................................... 32 Coccinellidae .............................................................................................. 32

Curculionidae ............................................................................................. 33 Order Hemiptera ............................................................................................... 33

Miridae ....................................................................................................... 34 Cicadellidae ............................................................................................... 34

Aphididae ................................................................................................... 34 Anthocoridae .............................................................................................. 35 Other Hemiptera ........................................................................................ 35

Order Hymenoptera .......................................................................................... 35 Apidae ........................................................................................................ 36 Formicidae ................................................................................................. 36

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Eulophidae ................................................................................................. 36

Other Hymenoptera ................................................................................... 37

Order Lepidoptera ............................................................................................ 37 Order Thysanoptera ......................................................................................... 37 Order Araneae .................................................................................................. 38 Insect Dependency on Bloom Period ............................................................... 38

Discussion .............................................................................................................. 39

Pollinator Candidates Based on Population Density ........................................ 39 Differences in Orchards .................................................................................... 42

3 INSECT BEHAVIOR AND POLLEN COLLECTION DURING FLOWER VISITATIONS ......................................................................................................... 60

Introduction ............................................................................................................. 60 Materials and Methods............................................................................................ 62 Results .................................................................................................................... 64

Discussion .............................................................................................................. 68

4 IMPORTANCE OF ARTHROPODS IN POLLINATION AND FRUIT SET AND PRODUCTION OF MANGIFERA INDICA .............................................................. 76

Introduction ............................................................................................................. 76 Material and Methods ............................................................................................. 78

Results .................................................................................................................... 80 Discussion .............................................................................................................. 80

5 CONCLUDING SUMMARY ON PRIMARY INSECTS INVOLVED IN MANGO POLLINATION IN THE SOUTH-FLORIDA REGION .............................................. 94

LIST OF REFERENCES ............................................................................................... 97

BIOGRAPHICAL SKETCH .......................................................................................... 102

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LIST OF TABLES

Table page 2-1 Insect sampling dates and times from Mangifera indica over the entire 8-

week blooming period at three orchard sites in Miami-Dade County, Florida. .... 45

2-2 Total number of insects collected throughout the 8-week blooming period ........ 46

2-3 Insects most prevalent throughout the 8-week mango blooming period (Jan. 23 to March 16, 2018) at 3 mango orchards in south Florida. ............................ 47

2-4 The percentage of Diptera collected throughout the 8-week blooming period .... 48

2-5 The percentage of Coleoptera collected throughout the 8-week blooming period ................................................................................................................. 49

2-6 The percentage of Hemiptera collected throughout the 8-week blooming period ................................................................................................................. 50

2-7 The percentage of Hymenoptera collected throughout the 8-week blooming period ................................................................................................................. 51

2-8 The percentage of Thysanoptera collected throughout the 8-week blooming period ................................................................................................................. 52

3-1 Observed insects on ‘Keitt’ mango flowers (Mangifera indica) ........................... 72

3-2 Quantification of mango (Mangifera indica) pollen on insects collected from ‘Keitt’ mango trees .............................................................................................. 73

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LIST OF FIGURES

Figure page 2-1 Orchard 1 (25°30’22.04” N 80°29’56.4 W) on January 1, 2018, during the

beginning of panicle emergence. ........................................................................ 53

2-2 Orchard 2 (25°29’50.15” N 80°29’25.64 W) a commercial orchard on February 13, 2018, during the completion of panicle emergence and flower opening ............................................................................................................... 54

2-3 Orchard 3 (25°35’58.96” N 80°26’43.96 W), commercial orchard on January 25, 2018, during early panicle emergence and flowering which was sparse. ..... 55

2-4 Most abundant insect orders collected from three mango orchards in south Florida during the 2018 8-week blooming period in south Florida ...................... 56

2-5 The five most prevalent Dipteran families collected on Mangifera indica throughout the 8-week blooming period across the three orchards in south Florida. ............................................................................................................... 57

2-6 Comparison of the four most prevalent Chloropidae genera collected from three mango (Mangifera indica) orchards in south Florida during the 2018 8-week blooming period ......................................................................................... 58

2-7 Comparison of the two most prevalent Chloropidae genera collected from three mango (Mangifera indica) orchards in south Florida ................................. 59

3-1 The mean number of flowers visited on ‘Keitt’ mango (Mangifera indica) at the Tropical Research and Education Center, Homestead, Florida .................... 74

3-2 The mean visual observation time insects visited ‘Keitt’ mango (Mangifera indica) inflorescences ......................................................................................... 75

4-1 Pollinator exclusion bags (middle-right side) in the canopy of ‘Keitt’ mango trees ................................................................................................................... 83

4-2 Pollination exclusion bag placed around a mango inflorescence prior to anthesis on January 30, 2018. ............................................................................ 84

4-3 A developing mango inflorescence on March 1, 2018, inside an exclusion bag. .................................................................................................................... 85

4-4 A bagged panicle with no fruit-set or vegetative growth (March 15, 2018). ........ 86

4-5 Initial fruit set on ‘Keitt’ mango. Fruit set varies throughout the panicle (March 1, 2018). ................................................................................................. 87

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4-6 Fruit enlarging after initial fruit set (March 15, 2018). ......................................... 88

4-7 Fully developed fruit on July 28, 2018 at the Tropical Research Center, Homestead, Florida. ........................................................................................... 89

4-8 The mean number (± SE) of fruit per panicle on non-bagged and bagged (insects excluded) mango inflorescences on March 2, 2018 .............................. 91

4-9 The mean number (± SE) of fruit on non-bagged and bagged inflorescences on May 10, 2018, 140 days after bagging. ......................................................... 93

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Abstract of Thesis Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Master of Science

AGROECOLOGICAL ANALYSIS OF ARTHROPODS INVOLVED IN MANGO

POLLINATION IN SOUTH FLORIDA

By

Matthew Quenaudon

May 2019

Chair: Daniel Carrillo Major: Entomology and Nematology

The role of insects on pollination of Mangifera indica is poorly understood. We

identified the most abundant arthropods visiting mango flowers, their interaction with

mango flowers, and how much mango pollen they are carrying. A total of 4,564 insects

were collected from mango flowers during the entire mango bloom period (8 weeks) in

three mango orchards located in Homestead, Florida. Hippelates sp., Liohippelates sp.,

and Oscinella sp. were the most abundant insects during the peak flowering period

when mango flowers are more receptive to pollination. Drosophilids, Sciarids,

Cryptophagus sp., and Cicadellids were present across the entire mango blooming

period. Cardiastethus sp., Dagbertus sp., Microtechnites sp., Zaprionus sp., and

Frankliniella sp. were abundant during the last two weeks of the mango bloom. Apis

mellifera carried large amounts of pollen but was rare in mango orchards. Muscids,

Allograpta obliqua, and Camponotus planatus were also observed to average high

pollen counts. Camponotus floridanus visited more mango flowers per unit of time,

followed by Calliphorids, Syrphids, and Apis mellifera. An arthropod exclusion test

revealed that insects may increase fruit set up to 17%. Our results indicate that a wide

diversity of insects pollinate mango in Florida and there are temporal shifts in insects

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throughout the mango bloom. Musca domestica, Allograpta oblique, Forcipomyia

genualis, Liohippelates sp., Hippelates sp., Camponotus floridanus, Camponotus

plantatus, and Apis mellifera are the most important insects providing pollination

services in mango in Florida as indicated by their visitation frequency and/or their pollen

loads. Differences in insect populations in separate orchards suggest that cultural

practices may influence insect populations and could be used to augment pollinator

populations.

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CHAPTER 1 LITERATURE REVIEW

Origin, Distribution, and Importance of Mangifera indica

Mango (Mangifera indica) is a major fruit crop of the tropics and subtropics

worldwide. The center of origin of mango is eastern India and southern Asia. There are

two basic types of mango, which are distinguished by mode of reproduction and

generalized fruit characteristics. The Indian mango type is indigenous to the subtropics

and northeastern India and typically possesses a monoembryonic seed while the

southeastern Asian mango is indigenous to tropical Asia and possesses a

polyembryonic seed (Iyer and Schnell, 2009). A monoembryonic seed contains only a

single zygotic embryo, with characteristics of its male and female parents.

Polyembryonic seeds possess multiple embryos, one of which may be zygotic but all

the rest are clones of the female parent. ‘Keitt’ is a monoembryonic-type mango that

requires grafting in order to be perpetuated. Grafting is used to reduce the time it takes

a tree to produce fruit and is a more economical approach (Campbell, et al. 2002).

Through the European voyages of the 15th and 16th centuries, mango spread

globally. Mango transportation had to occur as ripe fruit, seedlings, or grafted plants

because mango seeds could not survive freezing or drying (Mukherjee, 2009). The

Portuguese were responsible for introducing the mango from their Indian colonies to

Africa and later to the Americas (Mukherjee, 2009). Over time the polyembryonic mango

varieties were brought through the Pacific trading ports of Mexico and Panama into the

New World colonies, and from there to the West Indies. The first introduction into

Florida was a polyembryonic seedling brought from Cuba in 1861 (Mukherjee, 2009).

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The diversity of the U.S. mango gene pool continued to increase and by the 20th

century there was mango germplasm from India, Cambodia, the Philippines, and other

areas in southeast Asia (Mukherjee, 2009). Despite the vast amount of mango

germplasm introductions into Florida, it is estimated that most of the Florida cultivars are

descended from four monoembryonic Indian mango cultivars (‘Amini’, ‘Sandersha’,

‘Mulgoba’, and ‘Bombay’) and one polyembryonic cultivar (‘Turpentine’) from the West

Indies (Schnell et al., 1995).

Mango production in the U.S. is restricted to California, Florida, Hawaii, and

Puerto Rico (Marzolo and Lee, 2016). Despite having many influential cultivars, the

U.S. is not a major producer or exporter of mangoes and produces a mere 3,000 metric

tons annually (Evans, 2008). India however, accounted for 38.6% (10.79 million metric

tons) of world production between 2003 and 2005 and is the largest producer. The U.S.

is, however, the top importer of fresh mangoes with 459,936 metric tons in 2017

according to USDA market news (Mango Volume & Price History, 2018).

Florida is the largest producer of mangoes in the U.S. (Draper, 2014). Florida

grows about 150,000 mango trees on approximately 1,350 acres, producing an

estimated 370,000 bushels (~20.4 million pounds) with an estimated value of $5.6

million (Crane, 2018). Production of mango in Florida is limited largely due to climate

requirements. The mango season in Florida extends up to 6 months from early May to

October (Marzolo and Lee, 2016). Flowering is affected by inherent genetics, previous

and current weather conditions, soil moisture, and cultural practices. Panicle emergence

and flowering may begin anytime from late December through April. However, one way

of increasing mango production in existing plantings is to increase fruit set through

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improved pollination. The relationship between pollination and fruit set is important to

understand, as pollination and fertilization are essential yield-limiting constraints, as

evidenced by the high density of flowers compared to fruit set quantities per tree

(Davenport, 2009). Insects play a major role as pollinators of many agricultural crops

including mango (Ramirez and Davenport, 2016). In Florida mangoes, however, the role

of insects in pollination is poorly understood.

Reproductive Physiology and Floral Biology

In order to best understand the role and impact of insects in pollination, it is

essential to understand the floral biology of mango. The sex ratio amongst perfect

(pistil and staminate structures) to staminate flowers (only male structures) varies with

cultivar and climate (weather conditions) and within each panicle (Davenport, 2009).

Physical and environmental conditions can also play a role in this variability, although

the terminals of inflorescences contain more perfect flowers compared to the panicle

axis, where staminate flowers are more clustered (Davenport, 2009). Perfect flowers

seem to make up the final vertical spike of a panicle, however following anthesis, the

flowers closer to the panicle axis fall off and the sex ratio fluctuates (Davenport, 2009).

Mango shoots undergo different phenological stages, beginning with cell division

in the apical and lateral meristems (Ramirez and Davenport, 2010). This cell division

results in stem flushes that are either synchronous or asynchronous throughout the

canopy (Ramirez and Davenport, 2010). Davenport (2007, 2009) stated there are

three main shoot types as a result of cell division. This includes vegetative shoots that

form leaves and stems, generative shoots that form the inflorescence, and mixed shoots

that may possess both leaves and inflorescences within the same node (Davenport,

2007, 2009; Ramirez and Davenport, 2010). The vegetative appearance of leaves

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changes as the leaves mature; initially light green and then turning reddish generally

two weeks after initial bud break (Ramirez and Davenport, 2010). Mango tree

development is tied to genetic predisposition, climate, and other biotic factors.

Vegetative growth flushes generally take place during warmer temperatures, 25°C or

higher, and take roughly three to six weeks for full maturation (Ramirez and Davenport,

2010). This can be exhibited when comparing subtropical to tropical mango

development, where a distinct time gap between vegetative and reproductive stages is

present under subtropical conditions but not tropical conditions under which fruit,

flowers, and vegetation can be intermingled on the same canopy at the same time

(Ramirez and Davenport, 2010).

The sex ratio, i.e. ratio of perfect to male flowers, is determined prior to and

during flowering by both environmental and physiological factors. Cool weather, which

is more common during the early flowering period, may limit perfect flower development

(Davenport, 2009). In contrast, warmer temperatures promote the occurrence of perfect

flowers sometimes reaching a two to seven-fold increase (Majumder and Mukherjee,

1961; Davenport, 2009). Endogenous factors such as hormones and exogenously

applied plant growth regulators may modify the sex ratio of inflorescences (Davenport,

2009). The combination of gibberellic acid (GA3) and urea for instance, when applied

right before inflorescence shoot initiation, will result in a decline of the number of perfect

flowers (Rajput and Singh, 1989; Davenport, 2009). In comparison, the application of

paclobutrazol (an inhibitor of GA3) to the soil and naphthalene acetic acid (NAA) has

been shown to increase the perfect to staminate flower ratios, and foliar applications of

BA (benzylaminopurine) with 2% calcium ion increased the percentage of perfect

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flowers (Singh and Rajput, 1990; Kurian and Iyer, 1993; Mallik et al., 1959; Singh et al.,

1965; Davenport, 2009). Despite the ability to manipulate sex ratios and increase

perfect to staminate flower ratios, there has been no evidence to suggest increased fruit

yield from chemically increased sex ratios, implying perfect flower ratios are not the

limiting factor in crop performance (Schaffer et al., 1994; Davenport, 2009). Davenport

(2009) theorized that pollen viability, inflorescence growth, and ovule fertilization are the

main factors resulting in low fruit set. Thus, an increased understanding of the role of

insects involved in mango pollination and fruit set may greatly improve mango fruit

production.

Insect Pollinators

A deeper understanding of the insects involved in the pollination of mango

cultivars may lead to cultural practices that improve pollination and result in greater fruit

set. Popenoe (1917) and Davenport (2009) first suggested that pollen transfer amongst

mango flowers was accomplished primarily by insects as opposed to earlier ideas that

mangoes were primarily wind pollinated. Early observations suggested the most

efficient pollinators listed in order of importance included wasps, bees, large ants, and

large flies (Anderson et al., 1982; Davenport, 2009). Depending on various abiotic

factors such as wind, rain, and temperature, differences in insect pollination rates may

occur. Young (1942) observed that insects visited only 10-12% of mango flowers

predominantly in the morning and evening, with some visitation at night. In general,

knowledge about the role of insects in cross-pollination is limited (Anderson et al.,

1982), as key species were not identified and no observations were made of pollen

transfer and pollen deposition by insects in previous studies (Anderson et al., 1982;

Davenport, 2009).

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Ne’eman et al. (2010) postulated that one of the main proxies for evaluating a

pollinator’s impact on fruit set is to look at pollination efficiency, which depends on the

‘frequency’ of flower visitation and ‘effectiveness’ of pollen deposition. Pollination

deposition effectiveness can be estimated by the sheer number of pollen grains

deposited on the stigmas. However, pollen deposition effectiveness also depends on

how likely the pollen deposition will result in seed set per flower, which is influenced by

stigma receptivity and pollen quality. If a stigma is not receptive, it will be unable to

recognize the pollen, and/or the pollen will not adhere to the stigmatic surface. If pollen

has been tainted or degraded, the viable pollen to ovule ratio may be decreased,

resulting in a reduction in seed set (Ne’eman et al., 2010).

Pollen deposition effectiveness is also tied to the insect’s morphology and pollen

carrying capacity. With this understanding, most bees, given their distinct

morphological trait of having a corbicula or scopa to allow for carrying relatively large

amounts of pollen, would be high on the pollen deposition effectiveness scale.

However, just because an insect does not have the capability to transfer large amounts

of pollen does not mean it is not a good pollinator, as some insects may have a much

higher population density or higher preference for a plant, leading to an increase in

flower visitation frequency. Ne’eman et al. (2010) foresaw this issue, taking into

consideration that continuous visitation from a pollinator to a plant species may

contribute to effective pollination even when per-visit pollen deposition is low. However,

King et al. (2013) postulated that frequency of visitation is a poor representation for

overall pollination effectiveness (PE) and proposed the single visit deposition (SVD)

method to distinguish ‘true’ pollinators from insects just visiting flowers.

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The SVD method focuses on quantifying pollen deposition on virgin stigmas

during a single visit by a given insect, achieved through bagging flowers and excluding

other visitors. Nevertheless, King et al. (2013) recognized the potential limitation of the

SVD method to evaluate the importance of different pollinators, namely that the method

may result in unnatural flower visitor identity or behavior due to delayed removal of bags

on the flowers. Moreover, King et al. (2013) suggested that the SVD method provide

better PE assessments for those insects that are similar in size to the flower, feed

rapidly, and gather pollen on their body quickly. Using the SVD method to identify PE

may be even more skewed in a single inflorescence versus panicle inflorescences,

given that there may be a minimum threshold of pollen grains required in order for fruit

set to occur. The determinant inflorescence on mangoes, or cymes, contain an apex

bud which is the primary bloom followed by lateral buds which may lead to another set

of cymes and delayed bloom. This process is heavily dependent on both biotic and

abiotic factors, and thousands of blooms generally span over several months, allowing

for a much longer timeframe in which insects and other pollinators may contribute to

pollination. An overall high abundance of insects depositing less pollen per visit could

surpass the total amount of pollen deposited by larger but less frequently visiting insects

such as bees. For this reason, it is crucial to observe and understand the unmanaged

insect fauna that may be contributing to pollination.

The collective role of unmanaged insects can be just as important and useful for

the pollination of crops as compared to managed pollinators. Rader et al. (2012) looked

at the importance of unmanaged insects including bees on crop pollination and found

varying degrees of evidence for the importance of unmanaged insects. Insects visiting

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Brassica rapa flowers in four different fields were observed over the course of 4 years.

After 42,032 visits, the most prevalent visitors included Apis melifera (honey bee) and

seven unmanaged insect species. While honeybees were responsible for 40-60% of all

visits compared to 39.2% of the unmanaged insects, in two out of the four years

unmanaged insects were able to deliver more efficient and consistent pollination

services compared to the honeybee.

Decreases in bee populations due to limited gene pools, insecticides, and the

presence of disease and parasites (Bartomeus et al., 2013), reinforce the importance of

a better understanding of the other insects involved in mango pollination. As

landscapes and crops change, some populations of insects will falter while others will

rise, making it important to understand whether insects that can thrive in human-altered

ecosystems will deliver the pollination services previously provided by other insects.

These developments have motivated studies examining looking at the local fauna of a

given location to see what is really pollinating crops, including both managed and

unmanaged pollinators.

Sung et al. (2006) looked at pollinators of mango flowers in southern Taiwan.

The most common insects collected during 60 min intervals between 9 am and 1 pm

included bees, Apis cerana, A. mellifera, Braunsapis hewitti, Halictus sp., and flies,

Chrysomya megacephala, Musca domestica, Menochilus sexmaculatus, and

Indioscopus sp. (Sung et al., 2006). Of all the insects collected, 69.1% were female and

42.0% were flies (Diptera). The most dominant insect pollinators included Apis sp.,

Halictus sp., and C. megacephala. Diptera were considered unmanaged insects that

often congregated in larger densities than Hymenoptera while the bee species A. cerna

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and A. mellifera were considered managed. Small arthropods such as mites, thrips,

small flies, and parasitoids were all disregarded and not accounted for, and thus their

sample size contained only 126 insects observed at nine different locations over a 32-

day period (Sung et al., 2006). However, there was insufficient data to conclude which

insects were most responsible for pollination and fruit set.

Huda et al. (2015) determined that large flies, such as Eristalinus sp. and

Chrysomya sp. in the Syrphidae and Calliphoridae families respectively, were pollen

carriers and efficient pollinators of mangoes in Malaysia. Insect morphological traits

and size effects in pollination and pollen carrying capacity were investigated. The most

important Dipteran was an Eristalinus sp., possessing the greatest number of pollen

grains on its body with a Stomorhina sp. and Chrysomya sp. following. However, the

Eristalinus sp. was rarely found in mango orchards. Both Sarcophaga sp. and

Camponotus sp., a large fly and ant, contained very few pollen grains on their bodies

(Huda et al., 2015). A smaller ant, Iridomyrmex sp., did not carry any pollen, possibly

because of its often-observed grooming habits. When comparing males to females,

anthophilous females often had less pollen on their bodies than males (Huda et al.,

2015). This could be attributed to differences in head size between sexes, considering

no differences in body length between sexes were found (Huda et al., 2015).

Interestingly, insects with a large head width, as opposed to head length, had greater

pollen reserves on them, and overall larger pollinators contained more pollen (Huda et

al., 2015). This relationship holds true for several insect genera, but varies amongst the

Diptera, where size of the insect seemed to vary with the pollen capacity of an individual

insect (Huda et al., 2015). Augmentation of Eristalinus spp., Chrysomya spp.,

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Stomohina spp., Sarcophaga spp., and Camponotus spp. is believed to improve

pollination, as these insects have relatively large body parts and hairy bodies (Huda et

al., 2015).

Pollinator effectiveness may be higher for those insects that occur in high

densities in an area and actively forage amongst flowers with high visitations rates,

allowing them to encounter the stigma and pollen grains (Rader et al., 2009). In

addition to insect morphology, the behavior of an insect can influence its ability to be an

efficient pollinator (Huda et al., 2015). Insects that have high visitation rates but spend

less time per flower may allow for a greater spread of pollen throughout the field,

however, the amount of pollen delivered per visit may vary. This can be compared to

insects that have a lower visitation frequency, but a longer duration of interaction on the

flower and stigma, possibly resulting in more pollen deposition per visit. Additionally,

insects show different methods of foraging for nectar and may side-work (behavior in

which an insect approaches) a flower resulting in less stigmatic contact leading to

reduced seed set per visit (Park et al., 2016). This suggests that understanding the

individual insect’s behavior is just as important as evaluating the density and frequency

of an insect in an orchard.

Research has helped further shape an understanding of the importance of

insects and the impact that they have in the pollination of mangoes. Previous reports

indicated the presence and importance of Hymenoptera, Diptera, and other insects in

mango production. Although certain insects such as Syrphidae, Apis sp., and

Calliphoridae, make an appearance across different mango production regions, some of

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the more recent studies reveal the significance of understanding the insect fauna at an

orchard level as insect communities can vary across regions.

Objectives of Master of Science Thesis Research

1. Determine and identify the most frequent arthropod visitors on ‘Keitt’ mango

flowers at three separate orchards over the entire mango blooming period.

2. Determine the behavior of the most common arthropods during flower

visitation; duration and interaction with flowers and flower structures and the amount of

pollen they are transporting.

3. Determine the importance of arthropods in pollination and crop production

through comparison of fruit set within bagged and non-bagged inflorescences.

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CHAPTER 2 MOST FREQUENT ARTHROPOD VISITORS ON ‘KEITT’ MANGO (MANGIFERA

INDICA) FLOWERS IN SOUTH FLORIDA

Introduction

Mango (Mangifera indica) is one of the world’s major fruit crops in the tropics and

subtropics. The monoembryonic and polyembryonic seed are the two main types of

mangoes, with the latter pertaining to tropical Asia and the former being significant to

India (Litz, 2009). ‘Keitt’ mango is a monoembryonic-type mango cultivar and is

especially important to Florida, being a dual-purpose fruit and a major commercial

variety (Crane, 2018). Dual-purpose signifies the ability of the fruit to be eaten either as

a green fruit that is popular amongst Asian-Americans or as a ripe fruit. Production in

Florida is less about volume but rather more focused on producing a diverse array of

mango cultivars and specialty types (e.g., green market, fresh market, specialty food

service market). Puerto Rico and Florida are the largest producers of mangoes in the

U.S. (Draper, 2014).

Despite producing an estimated value of $5.6 million annually in Florida (Crane,

2018), two of the main limiting factors to production are weather (i.e., potential freeze

events) and land availability. One way to increase mango production is through

improved pollination to increase fruit set. Florida growers have attempted using honey

bees to increase pollination, however, mango flowers do not appear to be overly

attractive to honey bees (Popenoe, 1917). Around the world different insects including

Musca domestica, Chrysomya megacephala, Cantharis sp., Apis cera, and Apis

mellifera have been found to be important pollinators of mangoes (Sung et al., 2006). In

Florida mangoes, the insects responsible for pollination are poorly understood and this

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lack of understanding limits the development of cultivation strategies to improve

pollination.

Further insights into the insects pollinating mango cultivars in the south Florida

region could result in changes to cultural practices that improve pollination and increase

fruit set. The population of the well-known and studied honey bee, Apis melifera, has

been in decline worldwide due to Colony Collapse Disorder, the Varroa destructor

parasitic mite, loss of habitat, in-breeding, and insecticides (Rader et al., 2009). It is

therefore imperative to look to other, unknown and unmanaged insects for “pollination

insurance” (Winfree et al., 2007). These are insects that can pollinate the intended

target in the absence of the managed pollinators such as honey bees. While studies

have focused on the dieback of honeybees and the subsequent native bees that may

become more significant in the pollination of crops, much of this focus has been on

different unmanaged hymenopteran pollinators.

While hymenoptera, especially Apidae, are known for their strong pollination

capabilities, non-hymenopteran insects such as Stratiomyidae and Syrphidae (Diptera)

have been shown to carry pollen up to 400 m, 100 m more than bees including

Halictidae and Apidae (Rader et al., 2011). The species richness of crop pollinators

may often be overlooked, though numerous studies have found that a diversity of

unmanaged pollinators results in greater pollination and crop yields (Klein et al., 2007).

Although some crops may see better fruit set with particular bees, and some bees may

prefer select crops, there is a tremendous number of insects being overlooked for their

pollination contribution.

25

The use of unmanaged insects supports the idea that numerous insect species

with high visitation frequency are capable of providing the “pollination insurance”

previously discussed. Winfree (2007) determined through both empirical and simulation

results that native bees were the most important and alone were capable of adequately

pollinating watermelon crops. Manipulating an insect’s population by maintaining

natural sites and implementing sustainable cultural practices (e.g., not mowing row-

middles all year and not applying pesticides within a window before and during the

flowering period) may increase pollinator frequency and improve pollination rates. Prior

to manipulating an environment to better favor specific insects, it is vital to understand

which insects are present in the agroecosystem and visiting the crop flowers.

In this study we hypothesize that numerous Hymenoptera and Diptera insects

are the main pollinators for mangoes in the south Florida region. The objectives of this

investigation were to: (1) determine the most frequent arthropod visitors on ‘Keitt’

mango flowers, (2) examine variation in insect visitation across three separate orchards

over the entire mango blooming period, and (3) determine how arthropod visitation rates

differed depending on the time of day.

Material and Methods

To gain a better understanding of the diversity of species contributing to the

pollination of mango flowers, three ‘Keitt’ mango orchards were identified and sampled

over the entire 2018 mango blooming period (eight weeks). Orchard 1 (25°30’22.04” N

80°29’56.4 W) was located at the Tropical Research and Education Center (TREC)

(Figure 2-1). TREC maintains a regular weed control (i.e., mowing and herbicide

applications) and fertilizer program (granular NPK-Mg, foliar minor element applications,

and soil-drench chelated-Fe) throughout the year. The disease control program includes

26

fungicide-spraying from flowering to harvest as needed. The application of the fungicide

Bravo Weather Stik (chlorothalonil) occured January 2, 10, and 22, 2018 and the

herbicide Roundup Max (glyphosate) was made on January 11, 2018. Between

February 5 and June 4, 2018, the fungicide Penncozeb 75 DF (mancozeb) with the

adjuvant Nu Film 17 (pinene sticker) was sprayed intermittently eight different times.

Another fungicide, Satori (azoxystrobin) was sprayed on April 30 and May 9, 2018. The

off-season cultural program is comprised mostly of weed control and one application of

the insecticide, malathion on December 19, 2017. All products were applied at

recommended label rates. Orchard 2 (25°29’50.15” N 80°29’25.64 W) is a commercial

orchard located within 2 miles of TREC, where similar maintenance is undertaken but it

contains more weeds and wild flowers in the understory (Figure 2-2). Boron and Bravo

(chlorothalonil) were sprayed multiple times through the 8-week blooming period.

Penncozeb 75 DF (mancozeb) with Mn, ZnNO3, MgSO4 were applied on February 26,

2018. Penncozeb was sprayed again on March 6, 13, and 20, 2018. On March 20, a

fungicide and bactericide Kphite 7LP (mono and dipotassium salts of phosophorous

acid) was sprayed. The fungicide Switch (cyprodinil and fludioxonil) was applied on

March 30, 2018. Orchard 3 (25°35’58.96” N 80°26’43.96 W) was located roughly 10

miles from TREC, where a wide variety of fungicides and herbicides were applied

(Figure 2-3). Manzate Pro-Stick (mancozeb), Nu Film 17 (pinene polymer sticker),

Urea, Micorthiol Disperss (sulfur), Gramoxone SL 2.0 (paraquat), Liberty 280SL

(glufosinate ammonium), Freeway (organosilicone surfactant), Level 7 (nonionic

surfactant, spreader), and Cuprofix Ultra 40 (copper) were all applied throughout the

27

blooming period. This orchard had interconnecting canopies so the area beneath the

canopy received very little sunlight.

Each orchard was sampled once per week but differed in the sampling frequency

per day. Orchard 1 was sampled 3 times in a day (8-10 am, 1-3 pm, and 8-10 pm) and

Orchards 2 and 3 were sampled two times in a day (8-10 am and 1-3 pm) (Table 2-1).

Ten ‘Keitt’ mango trees were selected randomly throughout each orchard and five

randomly selected inflorescences within seven feet of the soil surface per tree were

used for sweep net collection. As there were insufficient trees with inflorescences

during the first week, blooming trees were initially selected. Inflorescences were swept

in a single motion using an insect sweep net, where the entire bag quickly engulfed the

branch of the inflorescence. Once completely covered, the sweep net was pulled to the

side and twisted, as to not allow any arthropods to escape. Prior to sweeping the next

inflorescence, the sweep net was briefly shaken, untwisted, and then swept over the

next inflorescence. Once five randomly selected inflorescences per tree had been

swept, the contents in the sweep net were immediately placed into a plastic bag that

was stored in a freezer (0C) for further identification. On the same trees, an additional

sample was collected using a beat cloth to capture smaller arthropods not collected

from the sweep net or missed due to small size and/or clinging to the flowers. A white

collection tray measuring 32 x 45 cm was lined with paper towel and misted with water

to help keep the insects docile. Each inflorescence was lightly shaken over the

collection tray to catch falling insects. Two of the five inflorescences used for the sweep

netting were chosen at random and this comprised a single beat cloth sample.

28

The time of collection at each site can be viewed in Table 2-1. Orchard 1 was

the only site sampled at night due to logistical reasons and availability. Due to weather

and lack of insects collected, nighttime collections (7-9 pm) took place for 7 of the 8

weeks. Over the course of the 8-week blooming period, inflorescence accessibility

changed from the first to the last samples collected. During weeks 1 through 4 there

was an increase in new blooms while a healthy bloom population was maintained during

weeks 5 and 6. During weeks 7 and 8 healthy blooms decreased and limited flowers

were available for pollination. Once a collection was completed at each location,

specimens were brought back to the lab for identification and study. Some samples

were sent to a state taxonomist, Dr. Gary Steck, Florida Department of Agriculture and

Consumer Services, for identification. The data collected includes:

Number of organisms collected

Number of different species (species richness)

Species evenness (how close in numbers are the species)

Location of collection (orchard)

Results

The total number of insects collected from inflorescences in three ‘Keitt’ mango

orchards using both sweep net and beat cloth methods over the 8-week blooming

period was 4,564. A total of 14 orders and 78 families were identified (Figure 2-4).

Thysanoptera was the most abundant order with a total of 1,663 insects which

comprised 36% of all insects collected. However, almost all thrips (1,160 of the 1,663)

were collected in week 8 of the collection. Diptera was the second most abundant order

with 1,293 insects (28% of total insects collected) and was distributed across all three

orchards with 885 from Orchard 1, 201 from Orchard 2, and 207 from Orchard 3. The

next most abundant order was Hemiptera with 766 insects at 17% of the total collected.

29

There were 406 (9%) coleopterans, 184 (4%) hymenopterans, and 137 (3%) arachnids.

The remaining orders were Neuroptera at 54 insects, Lepidoptera at 48, Pscoptera at 6,

Trichoptera at 3, Orthoptera at 2, Odonata at 1, and Collembola at 1.

The total number of insects collected differed among the three orchards (Table 2-

2). Orchard 2 was the most abundant with 2,377 total insects followed by Orchard 1

with 1,811 insects and Orchard 3 with 376 insects. Distribution of the most abundant

orders was even across the orchards with the exception of Thysanoptera, which was

more abundant in Orchard 2 with 1,471 thrips compared to Orchard 3 with only 32

thrips. Insects from nine orders were collected in all orchards but only Orchard 1

included insects from the Collembola, Odonata, Orthoptera, and Trichoptera orders

Order Diptera

The top 5 most prevalent families of dipterans collected throughout the 8-week

mango blooming period included Chloropidae, Drosophilidae, Sciaridae,

Ceratopogonidae, and Muscidae (Figure 2-5). The chloropids comprised 54% of the

total, with 698 insects followed by Drosophilidae, Sciaridae, and Ceratopogonidae with

approximately133-169 insects each. Muscidae had the fewest insects but were present

in all three mango orchards (Table 2-3). Muscids were collected all weeks except for

week 7 and were mainly collected in Orchards 1 and 2 (n = 10 and 9, respectively)

(Table 2-3). Chloropidae were the most abundant during the first 3 weeks, declining

from week 3 through week 8. Drosophilidae were consistently present across all 8

weeks, with 96 out of the 169 collected in Orchard 3. Sciaridae peaked in week 4 with

82 insects before decreasing in weeks 5 through 8 to population numbers similar to

weeks 1 through 3. Ceratopogonidae were predominantly found in Orchard 1, with 98

out of 137, and well represented across all 8 weeks.

30

Chloropidae

The Chloropids collected consisted of 4 genera: 246 Hippelates sp., 301

Liohippelates sp., 121 Oscinella sp., 15 Ceratobarys sp., and 15 unidentified species;

representing 54% of all Diptera collected. The two most prevalent Chloropids,

Hippelates sp. and Liohippelates sp., were very similar in size, shape, and color and

could be mistaken for each other at first observation. Three out of the 4 genera of

Chloropidae were collected predominantly in the morning sampling, with fewer in the

afternoon and nightly collections even lower (Figure 2-6). Unlike the other genera, the

number of Oscinella sp. collected was similar throughout the three orchards and the

time of day.

The chloropids were highly abundant and well represented in all three orchards.

Both Hippelates sp. and Liohippelates sp. were captured at each location in relatively

equal numbers (Figure 2-7). These two genera represent a large number of individual

species that are both abundant and appear to be well distributed in mango orchards of

south Florida.

Drosophilidae

The second most abundant dipteran family, Drosophilidae, consisted of four

species, Drosophila sp., Zaprionus indianus, Scaptomyza sp., and an unidentified

species (Table 2-4). These four species were captured primarily during the morning.

Drosophila sp. was found in all three orchards 7 out of the 8 weeks, Z. indianus in two

orchards 3 out of the 8 weeks, and the unidentified species in all three orchards 7 out of

the 8 sampled weeks. However, 27 out of the 28 Z. indianus were collected in Orchard

3 on a single inflorescence.

31

Sciaridae

Two species of Sciaridae, Odontosciara sp. and an unidentified fungus gnat,

were found visiting mango flowers mostly at night and primarily from one grove with 121

out of a total of 133 collected at Orchard 1. The remaining 12 sciarids were collected at

Orchard 2; none were collected at Orchard 3. Week 4 had the highest populations with

81 specimens collected at Orchard 1, a time when the majority of Diptera and overall

insect populations were lower (Table 2-4).

Muscidae

Atherigona reversura, Musca domestica, and an unidentified species were the

three Muscidae collected visiting mango flowers. Atherigona reversura captures were

split between morning and night in Orchard 1 and 2 whereas other muscids were

collected throughout the day in all three orchards (Table 2-4). Orchard 1 contained 10

muscids, with 9 in Orchard 2 and 2 in Orchard 3.

Syrphidae

An assortment of syrphid flies including Ornidia obesa, Allograpta obliqua,

Toxomerus watsoni, Toxomerus marginatus, Copestylum violaceum, and Palpada

alhambra were collected primarily in the morning and afternoon (Table 2-4). Orchard 1

contained the most syrphids with 8 total, followed by Orchard 2 with 3 and Orchard 3

with 2.

Calliphoridae

The blowfly Lucilia coeruleiviridis was collected in two orchards (Table 2-4). The

4 calliphorids collected were split between Orchard 2 and 3 and were collected in weeks

1, 4, and 6.

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Ceratopogonidae

Four species of Ceratopogonidae were found during the day and night in all three

orchards and throughout the entire 8-week blooming period; Forcipomyia genualis,

Forcipomyia biannulate, Forcipomyia spp and Artichopogon warmkei (Table 2-4). Most

ceratopogonids (106 of 154) were collected from Orchard 1.

Order Coleoptera

A total of 406 beetles were collected, comprising 9% of the total insects collected

(Figure 2-4). The majority of these (346) were collected in Orchard 1, generally

scattered throughout the sample weeks. Cryptophagus sp. were the most abundant

with a total of 330 insects (Table 2-3). Species collected include Cryptophagus sp.,

Diabrotica balteata, Diaprepres abbreviatus, Delphastus sp., Myllocerus

undecimpustulatus, Haromia axyridis, Euphoria sepulcralis, Melanophthalma sp.,

Hypothenemus sp., Brachiacantha barberi, Scymnus cervicalis, Cryptocephalus

irroratus, Cycloneda sanguinea, Diomus sp., and Diachus auratus (Table 2-5).

Cryptophagidae

A total of 330 Cryptophagus sp. were collected, during the morning, afternoon,

and night sampling (29.1, 33.9, and 37.0% of all samples, respectively) (Table 2-3).

Although 302 out of the 330 Cryptophagus sp. were collected in Orchard 1, the other

two orchards had approximately the same number collected (n = 13 and 15 specimens,

respectively). Cryptophagidae numbers gradually increased until weeks 6 and 7, when

populations were 101 and 99 specimens collected, respectively.

Coccinellidae

Seven coccinellid species were collected which included B. barberi, C.

sanguinea, Delphastus sp., Diomus sp., Harmonia axyridis, and Scymnus cervicalis and

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an unidentified species. These insects were found in relatively low abundance, only

comprising a total of 22 specimens (Table 2-5). Thirteen coccinellids were found in

Orchard 1, 7 in Orchard 2 and 2 in Orchard 3. All Coccinellidae were found in the

morning and afternoon sampling and were mostly collected starting in week 4 and

through week 8.

Curculionidae

Five curculionid species were collected including D. abbreviatus, Hypothenemus

sp., Myllocerus undecimpustulatus, Scolytinae sp., and one unidentified species (Table

2-5). The majority of these insects were rarely found and were mostly comprised of M.

undecimpustulatus (n = 18 out of the 28 curculionids collected). All 18 M.

undecimpustulatus were found in orchard 3 and were caught between weeks 2 and 6.

The damage caused by this curculionid was highly noticeable in orchard 3 and it was

evident the pest was well established in this location. Feeding damage on the mango

foliage was obvious with leaf notching and feeding alongside leaf veins.

Order Hemiptera

Thirteen different families with 23 species of Hemiptera were collected over the

8-week blooming period (Table 2-6). Families include Anthocoridae, Aphididae,

Cercopidae, Cicadellidae, Delphacidae, Flatidae, Geocoridae, Lygaidae, Miridae,

Pentatomidae, Psyllidae, Reduviidae, and Rhyparochromidae. Hemiptera represented

16.8% of all insects with 766 collected (Figure 2-4). Most hemipterans were collected

during weeks 7 and 8 with a total of 476 specimens (62% of the total) collected during

those weeks.

34

Miridae

The mirids represented 59.5% of all Hemiptera and 10% of all insects collected.

Six species of Miridae were present throughout the 8 weeks (Table 2-6). Miridae

collected included Camplyomma verbasci, Dagbertus sp., Lygocoris sp., Microtechnites

bractatus, Pcynoderes atratus and an unidentified species. Dagbertus sp., were

especially abundant during the last two weeks, (n = 293 out of 304 total specimens),

and were found primarily in Orchard 2. Interestingly, Dagbertus sp. were the most

prevalent Hemiptera collected and peaked in abundance during late bloom and fruit set

(weeks 7 and 8) (Table 2-6). Their feeding has been documented to cause flower and

fruit abscission. Initial numbers of Miridae across all three orchards were between 7

and 17 insects per week, which quickly increased to 168 and 222 insects per week for

weeks 7 and 8, respectively.

Cicadellidae

The Cicadellidae were the second most abundant hemipteran family, with a total

of 157 insects (Table 2-6). Two different species were present, Protalebrella

brasiliensis and an unidentified species. The majority of these insects came from

Orchard 1 (n = 107), followed by Orchard 2 (n = 47), and Orchard 3 (n = 10). Unlike the

mirids whose population drastically increased in the last 2 weeks, the cicadellids were

evenly present throughout the 8 weeks. The cicadellid collections increased as the day

progressed with 39 collected in the morning, 55 in the afternoon, and 63 at night (Table

2-6).

Aphididae

Aphids were not well represented and only accounted for 57 total insects and

were predominantly collected in the morning; 32 collected in the morning, 14 in the

35

afternoon, and 10 at night (Table 2-6). Uroleucon sp., Tetraneura sp., and an

unidentified species comprised the aphids collected. Twenty-four aphids were collected

in Orchard 1, 10 in Orchard 2, and 23 in Orchard 3. Aphid populations were most

abundant during the first 3 weeks and increased again in weeks 6 and 7, with lower

abundances in mid-bloom.

Anthocoridae

Four species of Anthocoridae were collected; Amphiareus sp., Cardiastethus sp.,

Orius sp., and an unidentified species (Table 2-6). Cardiastethus sp. were the most

abundant, representing 32 out of the 50 anthocorids, with 25 of these 32 specimens

collected during week 7. Forty-eight of the 50 anthocorids were collected during the

morning and afternoon sampling. Forty-two of the 50 anthocorids collected in Orchard 2

were associated with a high abundance of Frankliniella sp. (Table 2-6).

Other Hemiptera

Individuals from other hemipteran families, including Cercopidae, Delphacidae,

Flatidae, Geocoridae, Lygaeidae, Pentatomidae, Psyllidae, Reduviidae, and

Rhyparochromidae, were less frequent and collectively only represent 5% of Hemiptera

collected.

Order Hymenoptera

The Hymenoptera, a typically important order for pollinators, only accounted for

184 insects; or 4% of all insects collected (Figure 2-4). Families collected include

Apidae, Braconidae, Chalcidoidea, Enyrtidae, Eulophidae, Figitidae, Formicidae,

Halictidae, Ichneumonidae, and Pteromalidae. The most prevalent of these species

were Pheidole sp., Brachymyrmex sp., Quadrastichus sp., Apis mellifera, an

unidentified braconid species, and an unidentified eulophid species. Hymenoptera were

36

evenly represented throughout the 8 weeks, with the highest total in week 7 (n = 43

specimens) (Table 2-7). Most of hymenopterans (166 of 184) were collected in the

morning or afternoon. In addition, 107 of the 184 were collected in Orchard 2, with 47

collected in Orchard 1 and 30 in Orchard 3.

Apidae

Twelve Apis mellifera were collected between weeks 1 and 5, with 8 collected in

Orchard 3, 3 in Orchard 2, and 1 in Orchard 1 (Table 2-7). Most of these insects were

collected in the morning. Honeybees were seldom seen across all three orchards

(personal observations).

Formicidae

The formicids were the most numerous family, encompassing 35% of the total

Hymenoptera with 65 insects collected (Table 2-7). Species include Brachymyrmex sp.,

Camponotus floridanus, Camponotus planatus, Technomyrmex difficilis, Pheidole sp.,

and Tapinoma melanocephalum (Table 2-7). Twenty-five of these insects were

Pheidole sp., all collected from Orchard 2. More formicids were collected from Orchard

2 (n = 46) than Orchards 1 (10 insects) and 3 (9 insects). Most of the formicids were

collected in the morning and afternoon.

Eulophidae

Two species of eulophids were collected; Quadrastichus sp., and an unidentified

species (Table 2-7). There was a total of 45 eulophids, with 37 collected in Orchard 2, 7

in Orchard 1, and 1 in Orchard 3. Most of the eulophids (91%) were collected in the

morning and afternoon. Although no eulophids were collected in week 1, the remaining

weeks had a similar number of specimens collected.

37

Other Hymenoptera

The remaining families of Braconidae, Chalcidoidea, Encyrtidae, Figitidae,

Halictidae, Ichneumonidae, and Pteromalidae and an unidentified species combined

represented 38.5% of all Hymenoptera (Table 2-7). Halictidae, another family known to

be an excellent pollinator, was only found once in Orchard 3. Of all Hymenoptera

collected, all insect families were more numerous in the morning and afternoon, except

for Braconidae, which was collected throughout the morning, afternoon, and night.

Order Lepidoptera

Adults of nine Lepidoptera families collected from mango inflorescences include

Crambidae, Gelechiidae, Geometridae, Gracillariidae, Hesperiidae, Noctuidae,

Tineidae, Tortricidae, and an unidentified species. Forty-eight lepidoptera were

collected, evenly spread out throughout the day with 18 in the morning, 16 in the

afternoon, and 14 at night. Orchard 1 had 18, Orchard 2 had 21, and Orchard 3 had 9.

Populations of Lepidoptera remained low from weeks 1 through 5, then gradually

increased during the last 3 weeks. An unidentified species of Geometridae was the

most numerous with a total of 16 insects. Six were collected in the morning, 9 in the

afternoon, and only 1 at night. Twelve of the 16 geometrids were collected in Orchard

2, with 2 from both Orchard 1 and Orchard 3.

Order Thysanoptera

Four families comprising 7 species were collected, Franklinothrips sp.,

Franklinothrips vespiformis, Frankliniella sp., Frankliniella occidentalis, Frankliniella

fusca, a species of Phlaeothripidae, and an unidentified species were collected

throughout the 8 weeks (Table 2-8). Thysanoptera were the most abundant near the

end of the blooming season with a total of 1,663 specimens; 70.4% of collected thrips

38

were found in week 8 and 1,462 were collected from Orchard 2 alone during weeks 7

and 8. It is important to note that Hemiptera and Thysanoptera populations were

comprised mostly of pests, which increased in the last 3 weeks of collecting when

flowers were no longer receptive and began showing signs of deformity and injury.

Order Araneae

Five different families of spiders were collected during the 8 weeks, with the

majority being several unidentified minute spiders accounting for 128 out of the 137

collected specimens. Araneidae, Linyphiidae, Salticidae, Thomisidae, and the

unidentified small spiders were found predominantly in the afternoon with 62

specimens, followed by morning with 47, and night with 28 specimens. Most spiders

were collected at orchard 1 (90 specimens) followed by orchard 2 (33 specimens) and

orchard 3 (14 specimens).

Insect Dependency on Bloom Period

The most abundant insects collected during the first 3 weeks of bloom, when

pollination is critical, include Hippelates sp., Liohippelates sp., Oscinella sp.,

Drosophilidae, and Apis mellifera (Table 2-3). Those insects most abundant throughout

the entire bloom period include Ceratopogonidae, Drosophila, Sciaridae, Cicadellidae,

Braconidae, and Cryptophagus sp. Insects collected predominantly at the end of the

bloom season during weeks 6 through 8 include Zaprionus sp., Atherigona sp.,

Frankliniella sp., Cardiastethus sp., Campylomma sp., Dagbertus sp., Microtechnites

sp., and Eulophidae sp. Apidae and Syrphidae, two families often associated with

pollination, were seldom caught across the entire bloom period (Table 2-3).

39

Discussion

Pollinator Candidates Based on Population Density

Our findings are similar to those reported previously from India, Malaysia, and

Taiwan (Singh, 1989; Huda et al., 2015; Sung et al., 2006 and Kumar et al., 2012).

Diptera accounted for 28% of the insects collected from the three orchards, including

Syrphidae, Sarcophagidae, Calliphoridae, and Muscidae. Kumar et al. (2016)

concluded that Diptera are neglected pollinators, but studies are continually showing

their importance in pollination. Flies are responsible for pollinating more than 550

different plant species (Kumar et al., 2016), and both Kumar et al. (2016) and Singh

(1989) found that Syrphidae is the most prominent family found visiting mangoes in

India. In addition, Huda (2015) reported Eristalinus spp. (Sirphidae) and Chrysomya

spp. (Calliphoridae) as the most prevalent and important pollinators of mango in

Taiwan. In our study, neither Syrphidae nor Calliphoridae were frequently caught. Out

of 1,293 Diptera caught, only 13 were Syrphidae and 4 were Calliphoridae. However,

these insects move quickly and are difficult to catch with an insect sweep net, which

captures more crawling insects, and may be an underrepresentation of the actual

abundance of these flies in the field.

In contrast, relatively large numbers of Chloropidae, Drosophilidae,

Ceratopogonidae, and Sciaridae were collected. Ceratopogonidae have never been

reported in previous mango studies as most of those studies simply focused on the

large, hairy-bodied flies known to be good pollinators. A species collected during our

sampling, Atrichopogon warmkei, has been stated as a pollinator of Hevea brasiliensis

(Wilkening et al., 1985). Another biting midge in our sampling, Forcipomyia genualis, is

listed as a potential pollinator of mangoes that feeds on flower nectar (Borkent and

40

Spinelli, 2000). Although these insects are small, their sheer population densities and

widespread abundance begs the question as to how important these insects are for

pollination, especially on mango. Chloropidae, Ceratopogonidae, and Drosophilidae

were all found in each of the 3 mango orchards sampled, providing a pattern of

importance. The Chloropidae, Ceratopogonidae, and Sciaridae could often be

observed sitting on an individual flower for several minutes, moving around feeding on

the nectar and pollen. Despite taking longer to visit several flowers, unlike the

Syrphidae and Muscidae that move faster, these small flies displayed a long duration of

interaction with the stigma and stamens that could result in effective pollination.

Three of the most prevalent Chloropidae genera collected were Hippelates sp.,

Liohippelates sp., and Oscinella sp. These insect species have not been reported in

any previous mango pollination study, and thus our results may offer new insights into

the differences in mango pollinators across environments and localities. Liohippelates

sp. are often regarded as an animal pest, given their ability to mechanically transmit

disease in both livestock and humans (Machtinger and Kaufman, 2011). Although

regarded as a nuisance and pest, perhaps new understandings of these irritating flies

could show them to be an important pollinator of mangoes. Large numbers of

Hippelates sp. Liohippelates, and Oscinella sp., were collected in the first 3 weeks of

the mango bloom, a time regarded as a crucial pollination period given the freshness

and receptivity of mango flowers during this time. In total these 3 genera represent

51.7% of all Diptera caught. These insects were caught almost exclusively during the

day, with higher representations during the morning collection (8-10 am) than afternoon

(2-4 pm) collection period. This could be due to greater flower aroma (freshness) during

41

that time of day, i.e. prior to drying of the stigmatic surfaces. Furthermore, morning is

regarded as a crucial pollination period given the higher pollen viability and receptivity of

mango inflorescences (Davenport, 2009). Our findings contribute to a greater

understanding of when insects may be more active and foraging for nectar and pollen,

given the receptivity and biology of the flower in the earlier hours of the day.

Singh (1989) found that after Diptera, Coleopterans (beetles) were the most

numerous insects collected in his study, with 7 out of the 25 total insects belonging to

this order. Despite this small sample size, it offers a portrayal of perhaps other insects’

involvement in mango pollination aside from flies and bees. Similarly, our study

mirrored this and shed light on Cryptophagus sp. beetles and their interaction with

mango flowers. A total of 330 Cryptophagus sp., were collected, with similar numbers

collected in the morning, afternoon, and night, but mostly during weeks 6 and 7 of the

bloom period when fewer flowers were receptive.

Miridae and Thripidae captures increased significantly during the last two to three

weeks of bloom but are generally disregarded as important pollinators due to their pest

attributes and lack of abundance early in the blooming period. Although highly

abundant, these two insect families were almost exclusively found in one orchard, and

therefore act more as an outlier and not a good indicator of common species across

mango orchards. Orchard 2 had the most mirids and thrips, which may be due to

differences in cultural and pest control methods as further described below.

These results differ slightly from other studies, which could be a result of

collecting techniques influencing captures. All insects were collected with either a

sweep net on the inflorescence or a beat sheet, which favors crawling insects and may

42

lead to a collecting bias based on the insects’ behavior and speed at which it is able to

avoid collection. Another potential influence was the observation that some Syrphidae,

such as Copestylum violaceum, Palpada alhambra, and Ornidia obesa, were often seen

flying higher than other syrphids, roughly 6 feet up or higher and outside of our

collecting region. These 3 species were observed to be territorial and were far more

numerous than the data may suggest, especially in Orchard 2, but were outside of our

sampling area and were clumped in distribution. Futhermore, Muscidae, Calliphoridae,

and Syrphidae are quick to move and are difficult to catch with an insect sweep net.

Therefore, the fly numbers may be underrepresented in terms of how numerous they

are.

Differences in Orchards

All insects were collected in the same manner and with minimal bias, however all

three orchards varied in upkeep and maintenance. Differences between orchard

environments may not only impact insect diversity but the presence of key pollinators

during the bloom period. This is similar to findings from Carvalheiro et al. (2012) that

indicate the application of pesticides and seclusion from natural habitats lead to a

decrease in flying insects in mango orchards.

Orchard 1 is a non-commercial site with minimal insecticide use and limited

weeds. Mango trees were spaced 25’ x 25’ and the sides pruned to maintain an 8’

middle and tree height to maintain 15’, with open skies for plenty of sunlight to shine

through the canopy to reach branches and panicles lower down in the understory. The

greatest diversity of insects was found in Orchard 1, with low pest numbers at the end of

the blooming season.

43

Orchard 2 is a commercial orchard, with tall grasses, wild flowers, and weeds,

and has the greatest density of inflorescences amongst the three orchards when in

bloom. Although the data collected may not support it due to collecting difficulties,

Orchard 2 had the most observed Syrphidae present, perhaps due to having many

weeds infested with aphids that the syrphid larva could feed on. The trees were spaced

24’ x 24’ and pruned to maintain an open center (~8’ wide) and tree height of

~15’allowing sunlight to shine down through the canopy to the orchard floor. The

majority of pests were collected from Orchard 2, which resulted in most of the flowers

distorted and disfigured by weeks 7 and 8 from feeding by high populations of

Dagbertus sp. and Frankliniella sp.

Orchard 3 is a commercial orchard and was the most sprayed orchard, with

fungicides and insecticides applied weekly. The fewest insects were collected here,

possibly from the large amounts of insecticidal spraying and the limited number of

inflorescences. Tree density was greater than in the other sampled orchards, which

resulted in loss of much of the lower tree canopies and dense shade along the orchard

floor. All of the mango trees were connected and touching with over locking canopies,

leaving limited sunlight to shine through and limited branches and flowers in the

understory. Despite having 376 insects collected out of the total 4,564, Orchard 3 had

the highest population of Apis melifera and Drosophilidae. The prevalence of

honeybees was most likely due to the installation of managed honeybee hives in this

orchard. Especially prominent was Zaprionus sp., with the vast majority of specimens

collected at this site. In addition, all 18 Myllocerus undecimpustulatus were collected at

this site. There were limited weeds, grass and wild flowers. Interesting enough, despite

44

having far lower numbers of insects collected, this orchard had several insects not

found at other orchards. Several times no insects were found on multiple

inflorescences, followed by an inflorescence containing 20 or more flies clustered

together. This indicates perhaps a response to the lethal effects on insects in some

parts of the orchards and trees more heavily sprayed.

In conclusion, a wide diversity of insects visit mango flowers in south Florida and

there is a succession of insects throughout the mango bloom. Hippelates sp.,

Liohippelates sp., and Oscinella sp. were the most abundant insects during the first 3

weeks when mango flowering (i.e., the number of individual flowers opening).

Drosophilids, Sciarids, Cryptophagus sp., and Cicadellids were present across the

entire mango blooming period. Cardiastethus sp., Dagbertus sp., Microtechnites sp.,

Zaprionus sp., Frankliniella sp. were abundant during the last two weeks of the mango

bloom. Apis mellifera was rare in the three orchards. Differences in insect populations in

separate orchards suggest that cultural practices may affect populations of insects

visiting mango flowers.

45

Table 2-1. Insect sampling dates and times from Mangifera indica over the entire 8-week blooming period at three orchard sites in Miami-Dade County, Florida. Orchard 1 – (25°30’22.04” N 80°29’56.4 W); Orchard 2 – (25°29’50.15” N 80°29’25.64 W); Orchard 3 – (25°35’58.96” N 80°26’43.96 W).

Week Date Time

8-10 am 1-3 pm 8-10 pm 1 (1/23/18) Orchard 1 Orchard 1 Orchard 1

(1/24/18) Orchard 2 Orchard 2 (1/25/18) Orchard 3 Orchard 3 2 (1/30/18) Orchard 1 Orchard 1 Orchard 1

(1/31/18) Orchard 2 Orchard 2 (2/2/18) Orchard 3 Orchard 3 3 (2/6/18) Orchard 3 Orchard 3 (2/7/18) Orchard 1 Orchard 1 Orchard 1

(2/8/18) Orchard 2 Orchard 2 4 (2/13/18) Orchard 2 Orchard 2 (2/14/18) Orchard 3 Orchard 3 (2/15/18) Orchard 1 Orchard 1 Orchard 1

5 (2/21/18) Orchard 1 Orchard 1 Orchard 1

(2/22/18) Orchard 2 Orchard 2 (2/23/18) Orchard 3 Orchard 3 6 (2/27/18) Orchard 2 Orchard 2 (2/28/18) Orchard 3 Orchard 3 (3/1/18) Orchard 1 Orchard 1 Orchard 1

7 (3/6/18) Orchard 3 (3/7/18) Orchard 2 Orchard 2 (3/8/18) Orchard 1 Orchard 1 Orchard 1

8 (3/14/18) Orchard 2 Orchard 2 (3/16/18) Orchard 1 Orchard 1

46

Table 2-2. Total number of insects collected throughout the 8-week blooming period (23 January to 16 March 2018) from 3 mango (Mangifera indica) orchards in south Florida.

Orders Orchard 1 Orchard 2 Orchard 3

Araneae 90 33 14

Coleoptera 346 22 38

Collembola 1

Diptera 885 201 207

Hemiptera 204 518 44

Hymenoptera 47 107 30

Lepidoptera 18 21 9

Neuroptera 50 3 1

Odonata 1

Orthoptera 2

Psocoptera 4 1 1

Thysanoptera 160 1471 32

Trichoptera 3

Grand Total 1811 2377 376

47

Table 2-3. Insects most prevalent throughout the 8-week mango blooming period (Jan. 23 to March 16, 2018) at 3 mango orchards in south Florida. Orchard numbers represent total caught, while numbers under time of day and weeks are percent.

Percent Per Week Orchards

Family Species Total Insects % Morning % Afternoon% Night 1 2 3 4 5 6 7 8 Orchard 1Orchard 2Orchard 3

Curculionidae Myllocerus

undecimpustulatus 18 55.6 44.4 0.0 0.0 27.8 22.2 11.1 5.6 33.3 0.0 0.0 0 0 18

Cryptophagidae Cryptophagus sp. 330 29.1 33.9 37.0 2.4 0.0 10.9 17.3 5.2 30.6 29.7 3.9 302 13 15

Calliphoridae complex 4 25 75 0 25 0 0 50 0 25 0 0 2 2 0

Ceratopogonidae Forcipomyia sp. 1 100 0 0 100 0 0 0 0 0 0 0 1 0 0

complex 136 31 31 38 6 28 15 7 6 21 10 7 97 28 11

Chloropidae Ceratobarys sp. 15 73 27 0 47 0 27 0 27 0 0 0 14 1 0

Hippelates sp. 246 72 25 3 61 11 17 1 5 2 2 1 187 20 39

Liohippelates sp. 301 60 39 1 14 19 46 9 10 2 1 0 230 32 39

Oscinella sp. 121 31 31 38 10 50 21 6 10 3 0 0 106 12 3

Drosophilidae Drosophila sp. 74 65 30 5 1 0 9 19 14 14 41 3 9 41 24

Zaprionus indianus 28 93 4 4 0 0 0 4 0 75 21 0 1 0 27

complex 66 85 5 11 15 33 45 2 2 3 0 0 16 6 44

Muscidae Atherigona reversura 7 14 43 43 0 0 14 14 0 14 0 57 3 4 0

complex 14 50 29 21 7 29 14 7 7 7 0 29 7 5 2

Sciaridae complex 132 14 7 80 5 2 7 62 2 12 8 2 121 11 0

Syrphidae Allograpta obliqua 4 50 25 25 0 0 0 50 50 0 0 0 3 1 0

Copestylum violaceum 1 0 100 0 0 0 0 100 0 0 0 0 0 1 0

Ornidia obesa 1 100 0 0 100 0 0 0 0 0 0 0 0 0 1

Palpada mexicana 2 100 0 0 0 0 0 50 50 0 0 0 1 1 0

Toxomerus marginatus 3 100 0 0 0 0 33 0 67 0 0 0 3 0 0

complex 2 0 100 0 0 50 0 0 0 50 0 0 1 0 1

Anthocoridae Cardiastethus sp. 31 38.7 61.3 0.0 0.0 0.0 0.0 0.0 0.0 6.5 77.4 16.1 0 31 0

Aphididae complex 56 57.1 25.0 17.9 17.9 1.8 23.2 7.1 7.1 25.0 14.3 3.6 24 9 23

Cicadellidae complex 152 25.0 34.2 40.8 9.2 3.3 34.2 22.4 7.9 10.5 8.6 3.9 101 42 9

Miridae Camplyomma verbasci 27 63.0 37.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 22.2 77.8 0 27 0

Dagbertus sp. 304 57.2 41.4 1.3 0.0 0.0 0.0 0.0 0.3 0.0 36.8 62.8 10 294 0

Microtechnites bractatus 15 46.7 53.3 0.0 0.0 0.0 6.7 6.7 6.7 0.0 60.0 20.0 10 4 1

complex 98 56.1 38.8 5.1 10.2 17.3 6.1 6.1 13.3 8.2 38.8 0.0 33 60 5

Apidae Apis mellifera 12 66.7 33.3 0.0 8.3 33.3 25.0 8.3 25.0 0.0 0.0 0.0 1 4 8

Braconidae complex 18 27.8 33.3 38.9 0.0 0.0 16.7 44.4 0.0 0.1 16.7 11.1 10 4 4

Eulophidae Quadrastichus sp. 12 41.7 58.3 0.0 0.0 0.0 41.7 16.7 0.0 0.4 0.0 0.0 0 11 1

complex 33 39.4 48.5 12.1 0.0 12.1 3.0 9.1 15.2 0.1 24.2 27.3 7 26 0

Thripidae Frankliniella occidentalis 1643 65.7 31.8 2.6 0.1 0.0 0.1 0.0 0.0 1.6 27.8 70.4 152 1460 31

48

Table 2-4. The percentage of Diptera collected throughout the 8-week blooming period (23 January to 16 March 2018) in 3 mango orchard locations in south Florida by time of day and week.

Percent Per Week

Family Species % Morning % Afternoon% Night 1 2 3 4 5 6 7 8

Agromyzidae Ophiomyia sp. 1 100 0 0 0 0 0 0 0 0 100 0

unidentfied 4 25 50 25 0 0 0 50 0 25 0 25

Calliphoridae complex 4 25 75 0 25 0 0 50 0 25 0 0

CeratopogonidaeForcipomyia sp. 1 100 0 0 100 0 0 0 0 0 0 0

complex 136 31 31 38 6 28 15 7 6 21 10 7

Chironomidae Tanytarsus sp. 9 56 11 33 0 0 0 0 22 67 11 0

complex 17 6 12 82 0 12 65 0 12 12 0 0

Chloropidae Ceratobarys sp. 15 73 27 0 47 0 27 0 27 0 0 0

Hippelates sp. 246 72 25 3 61 11 17 1 5 2 2 1

Liohippelates sp. 301 60 39 1 14 19 46 9 10 2 1 0

Oscinella sp. 121 31 31 38 10 50 21 6 10 3 0 0

complex 15 40 60 0 0 13 0 60 13 7 0 7

Chyromyidae complex 4 25 0 75 0 75 0 25 0 0 0 0

Culicidae complex 2 0 50 50 0 0 0 50 0 50 0 0

Drosophilidae Drosophila sp. 74 65 30 5 1 0 9 19 14 14 41 3

Scaptomyza sp. 1 100 0 0 0 0 0 0 0 100 0 0

Zaprionus indianus 28 93 4 4 0 0 0 4 0 75 21 0

complex 66 85 5 11 15 33 45 2 2 3 0 0

Ephydridae Leptopsilopa sp. 1 0 0 100 0 0 100 0 0 0 0 0

Notiphila sp. 1 100 0 0 0 0 0 100 0 0 0 0

complex 15 47 40 13 0 13 27 7 13 13 7 20

Fanniidae complex 1 0 100 0 100 0 0 0 0 0 0 0

Lauxaniidae Camptoprosopella sp. 1 100 0 0 0 0 0 0 100 0 0 0

complex 1 0 100 0 0 0 0 0 0 0 0 100

Limoniidae complex 1 0 0 100 0 0 0 0 0 100 0 0

Micropezidae complex 1 100 0 0 0 0 100 0 0 0 0 0

Muscidae Atherigona reversura 7 14 43 43 0 0 14 14 0 14 0 57

complex 14 50 29 21 7 29 14 7 7 7 0 29

Phoridae complex 4 0 75 25 0 0 25 0 25 25 25 0

Sarcophagidae complex 4 25 50 25 0 0 0 25 25 50 0 0

Sciaridae Odontosciara sp. 1 100 0 0 0 0 0 0 0 0 0 100

complex 132 14 7 80 5 2 7 62 2 12 8 2

Syrphidae Allograpta obliqua 4 50 25 25 0 0 0 50 50 0 0 0

Copestylum violaceum 1 0 100 0 0 0 0 100 0 0 0 0

Ornidia obesa 1 100 0 0 100 0 0 0 0 0 0 0

Palpada mexicana 2 100 0 0 0 0 0 50 50 0 0 0

Toxomerus marginatus 3 100 0 0 0 0 33 0 67 0 0 0

complex 2 0 100 0 0 50 0 0 0 50 0 0

Tephritidae Dioxyna picciola 1 0 100 0 0 0 0 100 0 0 0 0

Evaresta 1 0 100 0 0 0 0 100 0 0 0 0

Xanthaciura sp. 7 43 57 0 0 0 14 0 0 71 14 0

complex 1 100 0 0 100 0 0 0 0 0 0 0

Tipulidae complex 10 10 10 80 0 40 20 0 10 30 0 0

Ulidiidae Seioptera sp. 1 0 100 0 0 100 0 0 0 0 0 0

complex complex 29 48 10 41 14 48 7 3 21 3 3 0

Total

Insects

49

Table 2-5. The percentage of Coleoptera collected throughout the 8-week blooming period (23 January to 16 March 2018) in 3 mango orchard locations in south Florida by time of day and week.

Percent Per Week


Chrysomelidae Cryptocephalus irroratus 2 50 50 0 0 0 0 0 0 50 50 0

Diabrotica balteata 1 100 0 0 100 0 0 0 0 0 0 0

Diachus auratus 2 100 0 0 0 0 0 0 0 0 0 100

Coccinellidae Brachiacantha sp. 1 0 100 0 0 0 0 0 100 0 0 0

Cycloneda sanguinea 3 67 33 0 0 0 0 0 0 33 67 0

Delphastus sp. 2 0 100 0 0 0 0 100 0 0 0 0

Diomus sp. 3 100 0 0 0 0 0 0 0 33 67 0

Harmonia axyridis 2 100 0 0 0 0 0 50 0 0 50 0

Scymnus sp. 2 0 100 0 0 0 0 0 50 0 50 0

complex 9 22 44 33 11 0 11 0 22 44 11 0

Cryptophagidae Cryptophagus sp. 330 29 34 37 2 0 11 17 5 31 30 4

complex 5 40 40 20 20 20 20 20 0 0 20 0

Curculionidae Diaprepes abbreviatus 1 0 100 0 100 0 0 0 0 0 0 0

Hypothenemus sp. 2 50 0 50 0 0 0 0 50 50 0 0

Myllocerus

undecimpustulatus

18 56 44 0 0 28 22 11 6 33 0 0

Scolytinae sp. 1 100 0 0 0 0 0 0 0 100 0 0

complex 6 33 33 33 0 17 50 17 0 17 0 0

Kateretidae complex 1 100 0 0 0 0 0 100 0 0 0 0

Latridiidae Melanophthalma sp. 6 0 67 33 0 0 0 17 0 17 67 0

complex 2 0 100 0 0 0 0 0 50 50 0 0

Scarabeidae Euphoria sepulcralis 3 0 33 67 0 0 0 33 33 33 0 0

complex 1 100 0 0 0 0 100 0 0 0 0 0

Staphylinidae complex 3 33 33 33 0 0 67 0 0 0 33 0

Total

Insects

50

Table 2-6. The percentage of Hemiptera collected throughout the 8-week blooming period (23 January to 16 March 2018) in 3 mango orchard locations in south Florida by time of day and week.

Percent Per Week


Anthocoridae Amphiareus sp. 2 0 0 100 0 0 0 0 0 100 0 0

Cardiastethus sp. 31 39 61 0 0 0 0 0 0 6 77 16

Orius sp. 10 40 60 0 20 0 0 0 20 10 20 30

complex 6 83 17 0 0 17 17 0 0 0 17 50

Aphididae Tetraneura sp. 1 0 100 0 0 0 100 0 0 0 0 0

complex 56 57 25 18 18 2 23 7 7 25 14 4

Cercopidae complex 8 63 38 0 0 25 38 13 13 13 0 0

Cicadellidae Protalebrella brasiliensis 5 20 60 20 0 20 0 0 0 0 80 0

complex 152 25 34 41 9 3 34 22 8 11 9 4

Delphacidae complex 21 38 52 10 0 0 29 5 10 19 19 19

Flatidae Ormenoides sp. 1 100 0 0 0 0 0 0 0 0 100 0

Geocoridae complex 1 100 0 0 100 0 0 0 0 0 0 0

Lygaeidae complex 1 100 0 0 100 0 0 0 0 0 0 0

Miridae Campylomma verbasci 27 63 37 0 0 0 0 0 0 0 22 78

Dagbertus sp. 304 57 41 1 0 0 0 0 0 0 37 63

Lygocoris sp. 3 67 33 0 67 0 0 0 0 0 0 33

Microtechnites bractatus 15 47 53 0 0 0 7 7 7 0 60 20

Pcynoderes sp. 9 0 100 0 0 0 0 0 0 0 33 67

complex 98 56 39 5 10 17 6 6 13 8 39 0

Pentatomidae Proxys punctualis 1 0 100 0 0 0 0 0 0 0 100 0

Psyllidae complex 2 100 0 0 0 100 0 0 0 0 0 0

Reduviidae complex 3 33 67 0 0 0 0 33 0 33 0 33

Rhyparochromidae complex 2 0 0 100 0 0 0 50 50 0 0 0

Blank complex 6 50 50 0 33 0 0 17 0 0 17 33

Total

Insects

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Table 2-7. The percentage of Hymenoptera collected throughout the 8-week blooming period (23 January to 16 March 2018) in 3 mango orchard locations in south Florida by time of day and week.

Percent Per Week

Family Species

Total

Insects % Morning % Afternoon% Night 1 2 3 4 5 6 7 8

Apidae Apis mellifera 12 67 33 0 8 33 25 8 25 0 0 0

Unidentified 1 100 0 0 0 100 0 0 0 0 0 0

Apocrita complex 3 0 100 0 0 0 0 0 33 0 0 33

Braconidae complex 18 28 33 39 0 0 17 44 0 0 17 11

Chalcidoidea complex 1 0 100 0 0 0 0 100 0 0 0 0

Encyrtidae Enyrtus sp. 1 100 0 0 0 0 0 100 0 0 0 0

Eulophidae Quadrastichus sp. 12 42 58 0 0 0 42 17 0 0 0 0

complex 33 39 48 12 0 12 3 9 15 0 24 27

Figidae Aganaspis sp. 1 100 0 0 100 0 0 0 0 0 0 0

Ealata sp. 1 0 100 0 100 0 0 0 0 0 0 0

complex 1 0 100 0 100 0 0 0 0 0 0 0

Formicidae Brachymyrmex sp. 11 9 91 0 0 0 36 0 0 0 0 45

Camponotus floridanus 1 0 0 100 0 0 0 0 0 1 0 0

Pheidole sp. 25 72 28 0 0 0 0 20 4 0 72 0

Pseudomyrmex gracilis 2 50 50 0 0 0 0 50 0 0 0 50

Tapinoma melanocephalum 7 29 71 0 0 0 0 0 0 0 100 0

Technomyrmex difficilis 2 0 0 100 0 0 0 0 0 1 0 0

complex 17 47 47 6 35 24 18 6 6 0 0 6

Halictidae complex 1 100 0 0 0 0 0 0 0 1 0 0

Ichneumonidae complex 4 25 25 50 0 0 0 0 50 0 50 0

Pteromalidae complex 4 25 75 0 0 0 0 50 50 0 0 0

Unidentified complex 26 35 62 4 19 12 4 15 8 0 19 8

52

Table 2-8. The percentage of Thysanoptera collected throughout the 8-week blooming period (23 January to 16 March 2018) in 3 mango orchard locations in south Florida by time of day and week.

Percent Per Week


Aeolothripidae Franklinothrips vespiformis 2 100 0 0 0 0 0 0 0 100

complex 1 100 0 0 0 0 0 100 0 0

Phlaeothripidae complex 8 38 50 13 0 0 0 13 0 13 63 13

Thripidae Frankliniella occidentalis 1643 66 32 3 0 0 0 0 0 2 28 70

complex 4 75 25 0 0 0 0 0 0 50 50 0

complex complex 5 40 40 20 40 20 20 0 0 0 0 20

Totat

Insects

53

Figure 2-1. Orchard 1 (25°30’22.04” N 80°29’56.4 W) on January 1, 2018, during the beginning of panicle emergence.

Photo courtesy of Matthew Quenaudon.

54

Figure 2-2. Orchard 2 (25°29’50.15” N 80°29’25.64 W) a commercial orchard on February 13, 2018, during the

completion of panicle emergence and flower opening. Photo courtesy of Matthew Quenaudon.

55

Figure 2-3. Orchard 3 (25°35’58.96” N 80°26’43.96 W), commercial orchard on January 25, 2018, during early panicle

emergence and flowering which was sparse. Orchard 3 had a heavy canopy with limited inflorescence in the understory. Photo courtesy of Matthew Quenaudon.

56

Figure 2-4. Most abundant insect orders collected from three mango orchards in south Florida during the 2018 8-week blooming period in south Florida from 23 January to 16 March 2018.

57

Figure 2-5. The five most prevalent Dipteran families collected on Mangifera indica throughout the 8-week blooming period across the three orchards in south Florida.

58

Figure 2-6. Comparison of the four most prevalent Chloropidae genera collected from three mango (Mangifera indica) orchards in south Florida during the 2018 8-week blooming period (23 January to 16 March) based on the time of day. Nighttime collections were only conducted at one location (TREC) for the first 7 weeks.

59

Figure 2-7. Comparison of the two most prevalent Chloropidae genera collected from three mango (Mangifera indica) orchards in south Florida from 23 January to 16 March 2018.

60

CHAPTER 3 INSECT BEHAVIOR AND POLLEN COLLECTION DURING FLOWER VISITATIONS

Introduction

The morphology of an insect and its behavioral interactions with flower sexual

organs help form an understanding of how important an insect may be in contributing to

successful pollination. Many scientists believe efficient pollinators have a high

population density and are in constant movement between flowers resulting in a high

frequency of interaction (Rader et al., 2009). The problem with determining which

insects fit this narrative in mangoes stems from the ambiguity in determining which

flowers are visited because of the small size of mango flowers (Huda et al., 2015). Prior

to looking at the duration and interaction of an insect on a flower, it is important to

comprehend the insect’s behavior.

The behavior of an insect is considered an important factor in pollen deposition

success, and influences whether pollen comes into contact with the stigma. Insect

behavior differs among orders, families, and species so it is important to determine

which insects are visiting the flowers. For example, ants (Formicidae) are present in

both frequency and density on mango but are often viewed as poor pollinators because

ants have little to no interaction with the stigma (Huda et al., 2015). In addition, ants

have grooming habits, which possibly remove any pollen adhering to their body. In

addition, the presence of ants often discourages other insects such as small flies from

visiting the flower (Huda et al., 2015). Understanding these small subtleties will help

define key pollinators and further pollination research.

Another aspect of behavior includes the duration and interaction of the most

prevalent insects with floral sexual organs on mango flowers. Although visitation rate is

61

divided into two parts: 1. how often an individual is observed on a flower per unit time;

and 2. how many individuals per flower are present per unit of time (Ne’eman et al.,

2010), this does not include the insects’ interaction with the flower’s sexual organs,

which is vital to ensuring pollination. As previously discussed, visitation frequency and

population density can be important indicators of valuable pollinators but understanding

behavior on the flower can provide knowledge as to whether they are actually

contributing to pollination. Monzon et al. (2004) determined that the behavior of an

important mason bee pollinator of pear, Osmia cornuta, directly affects pollination.

Osmia cornuta repeatedly landed on the reproductive structures of pear flowers

gathering pollen from anthers while moving their proboscis around in search of nectar.

In another study looking at the importance of bees, Peponapis pruinosa, on summer

squash (Cucurbita pepo), it was determined that males visited flowers twice as

frequently as the female bees, thereby increasing their role in pollination (Cane et al.,

2011. These studies both demonstrate how differences in behavior may play a role in

pollination and the value of understanding the contribution of behavior of the species

being observed.

Another important aspect for successful pollination, is the amount of pollen an

insect can carry and deposit. If consistent contact with the stigma is occurring and

pollen is present, there is a good chance for pollination and subsequent fertilization.

Estimating how much pollen an insect may be depositing on a mango inflorescence can

be difficult to determine due to the small flower size. Howlett et al. (2011) concluded

that collecting an insect directly from a flower and measuring the quantity of pollen

grains on its body is an easy, fast, and accurate technique to gage pollen carrying

62

capacity and deposition. The amount of pollen an insect can carry can depend on

multiple factors, including larger abdominal surface size, tongue length, presence of a

scopa, hairiness, and grooming behavior (Willmer and Finlayson, 2014).

In this study we hypothesize that insects of larger size will carry higher amounts

of pollen as shown in previous studies (Huda et al., 2015) and that Diptera will be

important pollinators based on past research (Sung et al., 2006; Huda et al., 2015;

Kumar et al., 2016). Although Apis mellifera is considered a key pollinator of mangoes

in Sao Francisco, Brazil, Diptera are the primary pollinators of mango in other tropical

areas (Ramirez and Davenport, 2016). The objective of this study was to determine

three different aspects of pollinator importance: (1) the behavior of the most common

arthropods during mango flower visitation; (2) duration and interaction with flowers and

flower structures; and (3) the amount of mango pollen transported.

Materials and Methods

A mango orchard, Mangifera indica ‘Keitt’, located at the Tropical Research and

Education Center (TREC) was selected to determine the number of arthropod visits to

mango flowers, observation of visiting behavior, and identification of those arthropods.

Observational data of insects visiting flowers were recorded in a different year than

when insects were collected for pollen quantification. The observational data were

collected between March 4 and April 20, 2018, during a second flush of newly formed

inflorescences, whereas insects collected for pollen determination were collected

between February 5 to 7, 2019. Trees were selected based on their availability of

inflorescences with pollen receptive flowers. At this time of year, there were only a few

trees with new inflorescences blooming within the same area. Over the course of 6

weeks, inflorescences with flower blooms on these selected trees were used for

63

observational field studies. Observations were conducted every other day in the

morning between 8 and 11 am or in the afternoon from 1 to 3 pm when insect activity

was at its peak and flowers were still receptive.

Once an insect was present on an inflorescence, it was observed for up to 2

minutes or until the insect flew away and could no longer be observed. Flowers

observed were those recently opened or considered receptive for pollination. During

that time the number of flowers visited and the insect's behavior and interaction with the

flower was recorded. Behavior data collected included movement speed, flight pattern,

if the insect groomed itself, and interaction with other insects. Insect visit duration per

flower and whether nectar or pollen was fed on or collected were recorded. Insects

found feeding on the outside portion of the flowers were marked as foraging on pollen

while those positioned in the middle were marked as foraging on nectar. Assumption of

foraging behavior based on the location of the insects on the flowers was due to

observations of where pollen and nectar are located within mango flowers. In our

observations, nectar was observed closer to the middle of the flower near the floral

disks, whereas pollen was found gathered near the edges of the flower. Insects

observed feeding between the two locations were marked as both. After the 2-minute

observation, the insect was collected using a kill jar and immediately placed in a vial

following death with minimal contact. At times, the insect was not able to be collected.

Collected insects were frozen at -20°C and held for further identification and pollen

acquisition at TREC.

For pollen quantification, frozen insects were thawed for 15 minutes after which

they were submerged in 50 microliters of 70% ethanol and vortexed for 30 seconds in

64

micro-centrifuge tubes to wash off any pollen. The same procedures were used for all

insects with the exception of A. mellifera, where the corbicula was removed prior to

washing the specimen. Removal of the corbicula is a standard practice for studies

involving pollination success, due to the pollen found in the corbicula being unavailable

for pollination services (Delaplane et al., 2013). Ten microliters were immediately

removed from the solution and pipetted into a haemocytometer for counting. The insect

was vortexed for another 30 seconds before a second extraction of 10 microliters was

placed into the second loading section of the haemocytometer. The insect and the

remaining 30 microliters were then labeled and kept for further analysis.

Transmitted light microscopy was used for pollen identification. The number of

mango pollen grains per insect sample was counted. Mango pollen was distinguished

from other pollen grains by comparing with samples directly derived from mango

flowers. Due to the relatively low number of pollen grains, nine alternating squares of

the haemocytometer were counted. If no pollen grains were in these 9 squares, the

number of pollen grains in the entire 10 microliter sample was counted. The insect was

identified prior to pollen collection, and the number of pollen grains for each insect was

recorded with the date collected.

Results

Eight different genera of insects were observed visiting mangoes between March

4 and April 20, 2018. With a few exceptions, insect identification during observations

were to the family level which included the Chloropidae, Syrphidae, Muscidae,

Sciaridae, and Calliphoridae. Individuals identified to genus or species included Apis

mellifera, Brachymyrmex sp., and Camponotus floridanus.

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The insects observed and the average number of flowers visited, average time

spent on each flower, total number of individuals collected, total number of flowers

visited, and number of visits for pollen and nectar were tabulated (Table 3-1). Most

insects observed foraged on pollen and nectar and include the Chloropidae, Syrphidae,

and Musicdae whereas insects from the remaining families mostly foraged on pollen

alone.

The smallest of the insects observed (Sciaridae, Chloropidae, and

Brachymyrmex sp.) were also the 3 insects with the lowest average number of flowers

visited per 120 seconds (Figure 3-1). Sciaridae visited the least number of flowers with

an average of 1.09 flowers per 120 seconds. These insects were often found stationary

or moving slowly around and inside mango flowers feeding on both pollen and nectar. In

comparison, larger insects moved quickly amongst the flowers with Camponotus

floridanus averaging 7.16 flowers per 120 seconds. This, however, may be skewed

slightly as these ants were almost always observed for the full 120 seconds, due to their

inability to fly away upon approach. Although the muscids and A. mellifera averaged

3.56 and 4.60 flowers per visitation, respectively, they moved more quickly and thus

were almost never observed for a full 120 seconds as the observers lost sight of them

before the period was done.

Apis mellifera was recorded to have the lowest interaction time with mango

flowers, averaging 42 seconds per visit, which in contrast to Brachymyrmex sp., that

were observed on a single flower for the full 120 seconds (Figure 3-2). However, a low

number (5) of honey bees were observed. Other insects observed for longer periods of

time included Camponotus floridanus, Chloropidae, and Sciaridae, which were also

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those groups that had the fewest numbers of flowers visited on average per 120 second

observation period. Insect activity was correlated with the size of the insect. Smaller

insects may be more likely to remain stationary while feeding while larger insects moved

quickly and frequently amongst flowers.

From Feb 5 to 7, 2019, 1 to 9 insects of 15 species of insects were collected from

mango flowers for pollen quantification. These species include Brachiacantha barberi,

Cryptophagus sp., Euphoria sepulcralis, Hippelates sp., Liohippelates sp., Oscinella sp.,

Chrysotus sp., Ephydridae, Muscidae, Platypeza sp., Sarcophagidae, Sciaridae,

Allograpta obliqua, Apis mellifera, and Camponotus planatus.

Apis mellifera had pollen counts greater than any other insect collected,

averaging 788 pollen grains per insect (Table 3-2). Muscids averaged 47 pollen grains

per insect, followed by the syrphid, Allograpta oblique, with an average of 16 pollen

grains per insect. Camponotus planatus averaged 15 pollen grains followed by

Sarcophagidae averaging 14.6. Insects with an average of less than 10 pollen grains

per insect included two different beetles, Euphoria sepulcralis and Brachiacantha

barberi and 3 flies, Ephydridae, Hippelates sp., and Oscinella sp.

Chloropidae and Sciaridae displayed similar behavior characteristics of time

spent on the flower. These insects were found to move between the petals and the

ovary, with their abdomen rubbing up against any anthers present while searching for

nectar amongst the floral disks. Chloropidae moved at a brisk pace, whereas Sciaridae

moved leisurely amongst the flowers. Grooming habits were not observed, and insects

were viewed several times flying away upon intrusion by larger insects such as muscids

or Camponotus floridanus.

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Muscidae and Calliphoridae were often seen zipping between flower to leaf and

resting on nearby leaves. Muscidae were observed grooming their legs on occasion

and both insects were sporadic with their movement. Interaction with other insects was

rarely noted. Muscids would often be found landing on the edge of the flower, with its

legs on the petals and outer boundary of the flower. The dorsal side of the abdomen

could be viewed contacting the reproductive structures of the flower. Syrphids were

typically smaller and would touch down briefly on a flower often landing on the edge or

middle of the flower. With smaller legs, syrphids appeared down in the flower, rubbing

up against the stamen and stigma. Syrphid flight behavior can be compared to a

helicopter, often hovering in place a few inches to a few feet from a potential flower,

before carefully landing and either moving from flower to flower or flying back into the

air. Not as quick as the calliphorids or muscids, syrphids were the most hesitant in

approaching a flower.

Apis mellifera was seldom seen on mangoes during observation and were quick

to fly away. A. mellifera lacked interest in flowers despite the hundreds of available

flowers for nectar or pollen collecting and would roam around flowers before dispersing

to some other location. Brachymyrmex sp. are much smaller than C. floridanus and

would often be found moving slowly within the mango flowers whereas C. floridanus

were actively moving from flower to flower. Brachymyrmex sp. favored feeding near the

floral disks, presumably feeding on nectar while C. floridanus was observed feeding on

the edges of the flower where potential pollen may have fallen. C. floridanus were often

territorial while feeding, causing other insects to leave or fly away when approached.

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Discussion

Further insight into the primary insects visiting mango flowers, Mangifera indica,

in the south Florida region and the amount of pollen each may be carrying was explored

in this chapter. An observational approach provides an opportunity to identify those

insects associated with mangoes that may have been missed with a sweep net. Similar

to Huda et al. (2015) observations of 5 insect genera pollinating mangoes in Malaysia,

our findings show similar insects, including Camponotus sp., Sarcophaga sp.,

Calliphoridae, and Brachymyrmex sp. This coincides with another report from southern

Taiwan, where 15 species were found visiting mango flowers of which 42% were

Diptera (Sung et al., 2006).

Although Apis mellifera may carry the most pollen grains, it is often the flies that

are most prevalent and found on mango inflorescences. The lack of honeybees, sweat

bees, and other solitary bees visiting mango flowers in this study demonstrates that

either bees are not attracted to mango flowers or that other factors play a role in the

shortage of Hymenoptera within mango orchards. These data agree with other reports

that Diptera are the most prevalent and important insects in mango orchards, whether it

be Syrphidae and Bombyliidae (Kumar et al., 2016), Chrysomya megacephala and

Musca domestica (Sung et al., 2006), or Eristalinus sp. and Stomorhina sp. (Huda et al.,

2015).

Insect morphology and size appear to influence the average time an insect visits

an individual flower. Average flower visit time seemed to increase as body size of the

insect decreased. Smaller insects such as Sciaridae, Chloropidae, and Brachymyrmex

sp. averaged the fewest flowers visited per observation period (120 seconds) indicating

their slow movement while navigating the mango flowers (Figure 3-1). This is indicative

69

of the potential attractiveness and/or size compatibility of mango flowers as well as

general insect mobility and behavior. Larger insects such as Muscidae, Apis mellifera,

Syrphidae, Calliphoridae, and Camponotus floridanus averaged more flowers per visit in

the 120 seconds demonstrating their active behavior while foraging. Depending on the

morphology of the insect and interaction of its appendages with the anthers or stamen,

this can be a benefit or detriment to pollination efficiency.

Long visitation time may result in greater pollen deposition rates, however, less

visits per time could result in less cross pollination. Body size and how the pollen is

stored on an insect are also factors that play a role in pollination. A large insect for

instance, will not fit inside a small mango flower allowing it to touch the stigma or anther.

In this case, it would be important that pollen adheres to an insect’s legs or ventral side

of the abdomen if that is the only part of the body contacting plant reproductive parts.

Similarly, a very small insect may spend minutes in one flower but never contact any

pollen or the stigma while feeding on nectar. Insect setae help pollen adhere to the

body or appendages of an insect and are key factors in how successful an insect may

be in the transfer of pollen. Future observational studies based on insect size could

further elaborate which insects may be efficient pollinators of mangoes due to the

morphological traits that are associated with different pollinators.

Although it has been reported that smaller ants such as Iridomyrmex sp. carry

very few pollen grains on their body; larger ants such as Camponotus sp. have been

reported to be key participants in mango pollination (Huda et al., 2015). Camponotus

planatus averaged 15 pollen grains per sample, which was roughly equivalent to both

the Sarcophagidae and Allograpta obliqua pollinators that carried 14.6 and 16 pollen

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grains on average, respectively. The variance in insects collected suggests the best

pollinators for each orchard may differ due to location. This is to suggest each orchard

may have different insect pollinators better suited for it’s specific geography and climate.

Based on the collective data from observations, behavioral traits, and pollen

carrying capacity, the insects of most importance include Syrphidae, specifically A.

oblique, Muscidae, Sarcophagidae, Drosophila sp., Cryptophagus sp., Camponotus

planatus, Hippelates sp., Liohippelates sp., and Ceratopogonidae. Most of these

insects have been shown to carry mango pollen and those that haven’t are extremely

prevalent and need to be further looked at to demonstrate their importance in mango

pollination. Although most insects collected in our insect diversity study (Chapter 2)

were present in our observational and pollen quantification studies, there were a few

insects that were not caught that could be of importance in pollination. The ants, in

particular Brachymyrmex, Camponotus floridanus, and Camponotus planatus were

extremely prevalent during our observational and pollen quantification studies but were

seldom collected with sweep net sampling. However, an ant that was present during

our sweet net sampling, Pheidole sp., was not present during our observational study.

Most of the larger flies were often seen during our sampling but seldom collected in

sweep nets, whereas these insects were better represented during our observational

and pollen study. On the flipside, smaller flies such as Sciaridae, Ceratopogonidae, and

Drosophila were commonly collected during our sweep net sampling but not in the

observational study likely due to difficulty in seeing these small insects.

Method of insect sampling influenced the number and diversity of insects

collected on mangoes. In order to better understand the insects of most importance in

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mango pollination, it is important to consider multiple methods of insect collection.

Number of samples and sample size also influences the number and diversity of insects

collected. Increasing either of these will provide increased accuracy and help pinpoint

the most effective pollinators of mangoes. The number of insects collected in this study

was low and observations were only conducted during one bloom season for a few days

during each sample week. This brief sampling period could lead to a bias in the insects

collected based on what insects are present during that timeframe. Increased sampling

across multiple orchards and over multiple blooming seasons could strengthen the data

and provide improved insight on the key pollinators of mangoes.

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Table 3-1. Observed insects on ‘Keitt’ mango flowers (Mangifera indica) from March 4 to April 20, 2018 at the Tropical Research and Education Center, Homestead, Florida.

Insect

Avg Number Of

Flowers

Visited per 2 min

Average Time

Per

Visit (sec)

Total # of

Insects

Observed

Total # of

Flowers

Visited

Pollen

Collecting / Feeding

(# of insects)

Nectar Collecting

/ Feeding (# of

insects)

Pollen & Nectar

Collecting / Feeding

(# of insects)

Chloropidae 1.25 ± 0.07 112.91 ± 3 55 69 54 12 11

Syrphidae 5.03± 0.45 103.81 ± 6 31 156 30 23 22

Muscidae 3.56 ± 0.40 96.92 ± 5 39 139 38 15 14

Apis mellifera 4.6 ± 0.54 42 ± 20 5 23 5 1 1

Sciaridae 1.09 ± 0.09 110 ± 21 10 12 10 1 1

Calliphoridae 5.38 ± 0.55 109.52 ± 8 21 113 21 2 2

Brachymyrmex sp. 1.65 ± 0.16 120 ± 0 23 38 23 1 1

Camponotus floridanus 7.15 ± 0.66 118.95 ± 6 19 136 19 3 3

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Table 3-2. Quantification of mango (Mangifera indica) pollen on insects collected from ‘Keitt’ mango trees from Feb 5 and Feb 7, 2019 at the Tropical Research and Education Center, Homestead, Florida.

InsectTotal Insects

Collected

Mean Number (±SE) of

Pollen Grains Per Insect

Total Pollen Grains

Counted

Brachiacantha barberi 5 3.7 ± 3.38 37

Cryptophagus sp. 1 0 0

Euphoria sepulcralis 2 7.5 ± 7.03 30

Hippelates sp. 1 1 ± 1.41 2

Liohippelates sp. 2 0 0

Oscinella sp. 6 0.33 ± 0.36 4

Chrysotus sp. 1 0 0

Ephydridae 1 3 ± 2.83 6

Muscidae 9 46.83 ± 20.96 843

Platypeza sp. 1 0 0

Sarcophagidae 4 14.625 ± 9.09 117

Sciaridae 1 0 0

Allograpta obliqua 6 16.17 ± 5.31 194

Apis mellifera 4 788.25 ± 209.49 6306

Camponotus planatus 3 15.33 ± 8.43 92

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Figure 3-1. The mean number of flowers visited on ‘Keitt’ mango (Mangifera indica) at the Tropical Research and Education Center, Homestead, Florida, during a 120 second visual observation from March 4 and April 20, 2018.

75

Figure 3-2. The mean visual observation time insects visited ‘Keitt’ mango (Mangifera

indica) inflorescences up to 120 second time period from March 4 to April 20, 2018 at the Tropical Research and Education Center, Homestead, Florida.

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CHAPTER 4 IMPORTANCE OF ARTHROPODS IN POLLINATION AND FRUIT SET AND

PRODUCTION OF MANGIFERA INDICA

Introduction

The significance of insects and the impact they have on pollination in agricultural

landscapes has been researched and further supports the ways in which non-managed

insects increase fruit set. A total of 41 crop systems worldwide revealed a positive

correlation of fruit set with wild insects (Garibaldi et al., 2013). They concluded that wild

insects more effectively pollinated crops than did honey bees, boosting fruit set by up to

twice as much. In addition, results indicate that lack of wild pollinator species richness

leads to a decrease in visitation from these wild insects and lower fruit set quantities

(Garibaldi et al., 2013). Therefore, to attribute a single species as a main pollinator or

justify augmenting a population of honeybees for a select crop is continuously revealed

as an incomplete practice or, in the case of mango, ineffective. The overall role of

pollinator diversity in influencing crop yields is continually being tested, demonstrating

the shift in strategic approach to pollination effectiveness and efficiency. The reliance

on insect pollination for agricultural crops is vital, with 39 out of the leading 57 single

crops seeing an increase in production with aid from these pollinators (Klein et al.,

2007). These crops make up 35% of global food production, demonstrating once again

the role of pollinator diversity in influencing crop yields.

Previous studies have suggested mangoes are anemophilous (wind pollinated)

plants (Kumar et al., 2016), however, Popenoe (1917) and Davenport (2009) brought

forth the idea that insects were responsible for the pollination of mangoes.

Hymenoptera and Diptera have been shown to be key orders of insect pollinators of

mangoes in various parts of the world. Apidae, Syrphidae, Tachinidae, and

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Bombyliidae are just a few of the families containing species involved in mango

pollination (Larson et al., 2001).

Davenport (2009) indicated that self-pollination is a likely occurrence in flowers

while the pollen is still damp, and this self-pollination may be aided by insects moving

pollen between floral sexual organs. Others have also claimed that depending on the

cultivar self-pollination can occur (Dijkman & Soule, 1951). Despite these assertions, it

was established by Singh et al. (1962) that self-incompatibility occurs in monoembryonic

‘Dashehair’, ‘Langra’, ‘Chausa’, and ‘Bombay Green’ (Mukherjee et al., 1968; Sharma &

Singh, 1970) to name a few.

To provide evidence to support the importance of insects in mango pollination,

Popenoe (1917) conducted a bagging experiment to exclude insects and found that

when insects were excluded, it resulted in less fruit set. However, Wester (1920)

considered wind pollination a viable means of pollination in mangoes. Free and

Williams (1976) showed that bagged panicles still produced fruit set, suggesting the

transfer of mango pollen by wind occurred despite the fact that mangoes do not show

any physiological or morphological adaptations for wind pollination (Kumar, 2016).

Furthermore, mangoes have two types of flowers, hermaphroditic and staminate, both

with sticky-pollen, which may allow for pollen grains to adhere to insect structures

during nectar feeding, suggesting some type of insect interaction is necessary for

pollination (Usman et al., 2001, Huda et al., 2015; Willmer and Finlayson, 2014).

An increase in mango fruit set by cross pollination shown in previous studies is

evidence for arthropod pollination and indicates that despite potential self-pollination,

mango fruit set is often limited by pollen incompatibility and stigma deficiencies (Singh

78

et al., 1962, Huda et al., 2015). Although less than 50% of flowers receive pollen in

nature (Iyer & Schnell, 2009), augmenting insect pollinators could potentially alter this.

The stigma is most receptive within the first 6 hours of anthesis, but will remain viable

for 72 hours (Singh, 1960). Once pollen is received by the stigma, germination

transpires within 90 min (Singh, 1954). The physiology of pollination has been

documented (Sandip et al., 2015; Usman et al., 2001) in mangoes, however, how pollen

moves around and the role of insects in cross-pollination is still ongoing research.

To investigate the impact or benefit of insects in pollination of mangoes,

comparison studies similar to Popenoe (1917) and Free and Williams (1976) were

conducted in which inflorescences were bagged prior to anthesis restricting the

presence of insects. This is a common method often used to compare between a

positive and negative control in which the insect visitors are excluded for the entire

duration until fruit is present (Delaplane et al., 2013).

Material and Methods

The ‘Keitt’ mango orchard at the Tropical Research and Education Center

(TREC) was used to determine the importance of arthropods in fruit set. Twenty-two

trees were selected based on their availability of inflorescences that shared similarity in

panicle length and flower development (Figure 4-1). Eight inflorescences per tree were

selected for a total of 4 control and 4 bagged inflorescences. A total of 176

inflorescences were selected, 88 bagged and 88 controls that were spread throughout

the mango orchard at TREC.

Four arthropod excluding bags were placed on each tree over an inflorescence

that had not yet gone through anthesis. The mesh nylon bags were constructed of “No

Thrips” Insect Screen 75 Mesh, 134" Wide” from Greenhouse Megastore and were

79

sewn individually to measure 97 cm long and 116 cm wide (Figure 4-2). The screen

used for these bags allows the entry of air and light but excludes insects. Each bag

contains a drawstring located at the top to secure the bag around an inflorescence

rachis and a wire frame to limit contact between the screen and the flowers. The frame

was constructed of chicken wire that provided support of the mesh bags but was not too

heavy for the branches. A chicken wire structure (61 cm long and 64 cm wide) was

placed inside each bag. Galvanized utility wire was added around the base of the

branch to further secure the mesh bags around the inflorescence. The bottom of the

bag was left open for placement over the inflorescence while minimizing disturbance of

the flowers. The bottom of the bag was then tied to the limb around the base of the

inflorescence using zip ties. In some cases when the bag was too heavy, a string was

used to secure the branch to surrounding branches or the trunk, which provided

additional support to the branch (Figures 4-2, 3, 4). To exclude ants and other small

arthropods, tangle foot was applied at the base of each bagged inflorescence. Four

other inflorescences were tagged but not bagged for comparison. These inflorescences

were similar in size, maturity, and located near the bagged counterpart. Mesh bags

were placed on the trees over the course of a month during bloom and prior to fruit

development. Mesh bags remained in place during the entire bloom period until fruit

maturation when they were removed mid-July.

The number of fruit per bagged and control inflorescence was recorded and

compared from initial fruit set until mature fruit were present (Figure 4-5, 6, 7). Data

were collected twice between March 2nd and July 13th representing initial fruit set and

mature fruit set. A Welch Two Sample t-test was conducted to compare the relationship

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between the fruit on bagged and control branches at both sampling points using

RStudio.

Results

Fruit set on the first sampling date was 6.7 times higher in inflorescences

exposed to insects compared to those where insects were excluded. Inflorescences

exposed to insects (un-bagged) averaged 2.69 ± 0.51 fruit per inflorescence which was

significantly greater than inflorescences where insects were excluded (bagged) which

averaged 0.4 ± 0.11 fruit per inflorescence (Figure 4-8) (t-value = -4.32, df = 181.78, P =

2.54e-05).

There was significant fruit drop between the first and the second sampling dates.

The number of fruit per inflorescence decreased 92% and 97% in un-bagged and

bagged inflorescences, respectively. However, the number of mature fruit on the second

sample date was 17 times higher in inflorescences exposed to insects as compared to

those where insects were excluded. There was an average of 0.17 ± 0.04 fruit per

inflorescence when exposed to insects (un-bagged) and 0.01 ± 0.01 fruit per

inflorescence when bagged as measured on the second sampling date when fruit were

mature (Figure 4-9) (t-value = -3.62, df = 97.62, P = 0.00047). Overall, the arthropod

exclusion tests revealed that insects can increase fruit set up to

Discussion

Arthropod exclusion tests suggest insects play an important role in the pollination

of mangoes and that wind pollination and/or self-pollination may be less of a factor than

previously noted. Free and Williams (1976) conducted a similar study in which mango

inflorescences that were bagged prior to anthesis to exclude insects did set fruit,

concluding pollen may also be transferred by wind. Although our findings do not entirely

81

refute this, as fruit set did occur in bagged inflorescences, our findings do reveal the

importance of arthropod interactions with mango flowers to improve fruit set.

There was a dramatic decrease in the amount of fruit set between the first and

second evaluation of March 2 and April 3, 2018. Fruitlet abscission in mangoes is

known to occur several weeks after anthesis resulting in a high percentage of panicles

to lose their fruit (Nunez-Elisea, 1986; Davenport, 2009). Different abiotic and biotic

factors may influence fruit set, including cold temperatures, high winds, low relative

humidity, limited nutrients, or lack of pollination. Research indicates that less than 1%

of fruit reaches maturity, which arises from the 8-13% of perfect flowers that set fruit

(Bijhouwer, 1937). Fruitlet abscission is random and may affect any fruit independent of

size or location on the panicle (Davenport, 2009). This suggests that in our study,

fruitlet abscission should be random, or that no pattern in mature fruit set between bag

vs. non-bagged treatments should exist if insects do not play a role in fruit set and

pollination. However, our results indicate arthropod involvement is important if not

essential to increased fruit set. Our data helps strengthen the importance of insect

involvement in mango pollination and provides more insights for future studies.

The sheer number of inflorescences on a mango tree coupled with naturally low

fruit set pose a challenge for investigating mango pollinators and their effect on fruit set.

Although 176 inflorescences were chosen, with 88 bagged and 88 non-bagged spread

across 22 trees, the low ratio of flowers to fruit set may limit a complete understanding

of how important insect pollination is for mango. It is also possible that bagged

inflorescences for prolonged periods may have limited some of the fruit present due to

factors other than insect exclusion. While the mesh bags were used to exclude

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arthropods from mango pollination, these bags may also limit the amount of pollen

being transferred by wind. The idea of wind pollination has been previously postulated,

and thus the statistically significant difference between bagged and non-bagged may be

due to both reduced insect and wind pollination. Additional studies with more replication

may better indicate how important insect pollination is to mango fruit set and crop yields.

The absence of insects during anthesis lead to reduced fruit set. Given the low

percentage of fruit set as previously discussed, maximizing every tree in an orchard to

produce maximum fruit is important for efficiency, especially given the lack of land

available for planting more mango trees. Although our sample size of 176

inflorescences is relatively small, the difference in fruit production between bagged (1%)

and non-bagged (17%) was significant. In 2016, the U.S. Department of Agriculture’s

Economic Research Service (ERS), listed the average cost of a fresh mango at $1.32

and a dried mango at $10.16 per pound. Our results suggest insects are significantly

contributing to the profitability of mango production systems in Florida. Moreover,

management tactics targeted to improve pollination services could lead to a greater

income for farmers. For instance, implementation of native flowers in small clusters can

help increase pollination when complimented with other factors such as maintenance of

natural habitats that serve as pollinator reservoirs (Carvalheiro et al., 2012). Moreover,

environmentally friendly management approaches to reduce pesticide pressure and

increase insect populations may also improve pollination. Further studies aimed at

identifying effective tactics to preserve and augment mango pollinators in Florida could

increase the sustainability and profitability of mango production systems in Florida.

83

Figure 4-1. Pollinator exclusion bags (middle-right side) in the canopy of ‘Keitt’ mango

trees at the Tropical Research and Education Center, Homestead, Florida, on March 1, 2018. Photo courtesy of Matthew Quenaudon.

84

Figure 4-2. Pollination exclusion bag placed around a mango inflorescence prior to

anthesis on January 30, 2018. The panicle can be seen upright without any contact to the mesh nylon bag. Photo courtesy of Matthew Quenaudon.

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Figure 4-3. A developing mango inflorescence on March 1, 2018, inside an exclusion

bag. Photo courtesy of Matthew Quenaudon.

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Figure 4-4. A bagged panicle with no fruit-set or vegetative growth (March 15, 2018).

Photo courtesy of Matthew Quenaudon.

87

Figure 4-5. Initial fruit set on ‘Keitt’ mango. Fruit set varies throughout the panicle

(March 1, 2018). Photo courtesy of Matthew Quenaudon.

88

Figure 4-6. Fruit enlarging after initial fruit set (March 15, 2018). Photo courtesy of

Matthew Quenaudonon.

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Figure 4-7. Fully developed fruit on July 28, 2018 at the Tropical Research Center,

Homestead, Florida. Photo courtesy of Matthew Quenaudon.

90

p = 2.54e^-05

91

Figure 4-8. The mean number (± SE) of fruit per panicle on non-bagged and bagged (insects excluded) mango

inflorescences on March 2, 2018, 50 days after bagging

92

p = 0.00047

93

Figure 4-9. The mean number (± SE) of fruit on non-bagged and bagged inflorescences on May 10, 2018, 140 days after bagging.

94

CHAPTER 5 CONCLUDING SUMMARY ON PRIMARY INSECTS INVOLVED IN MANGO

POLLINATION IN THE SOUTH-FLORIDA REGION

The notion that a good pollinator is subject to several criteria helps us process

which insects may be good candidates for mango pollination. Population density and

high frequency of insect to flower interactions are characteristics of the most effective

insect pollinators. Although pollination efficiency is a complex interaction influenced by

insect morphology, behavior, and flower characteristics, there is now a greater

understanding of the key insects visiting “Keitt” mangoes in south Florida. In addition to

insect population dynamics, understanding the ecological diversity and species

evenness as affected by time of day, location, and sampling period during bloom will

improve the identification and knowledge of the insects involved in pollination of

mangoes. Sampling throughout the entire blooming period provides a more reliable

representation of insects and life stages visiting mangoes in south Florida.

Primary insects included Musca domestica, Allograpta obliqua, Sarcophagidae,

and Camponotus plantatus that were all found to carry large amounts of pollen and

were prevalent throughout the mango orchards independent of location. In contrast, the

distribution and density of A. mellifera was significantly less than the Dipteran species.

This highlights the importance of Diptera in mango pollination and subsequently the

production of fruit. Our results indicated a large presence of flies within the first three

weeks of anthesis, critical time for pollination and early fruit set. Most of the insects

observed were Diptera and this may be the ideal time for augmentation of these

pollinators. Other potential pollinators included Brachiacantha baberi, Euphoria

sepulcralis, and Hippelates sp. Despite low numbers of B. baberi collected, this insect

was highly abundant in visual observations, suggesting further study.

95

Although A. mellifera contained the highest amount of pollen per insect, it’s lack

of visitation and seemingly disinterest in mango flowers limits it as a key pollinator. In

contrast, the density and prevalence of flies found to carry mango pollen is highly

significant. This suggests Hippelates sp., Liohippelates sp., Oscinella sp., Forcipomyia

genualis, Sciaridae, and Cryptophagus sp. may be important pollinators, however,

future research is needed to confirm this. Other insects that are potential pollinators

included Copestylum violaceum, Palpada mexicana, Ornidia obesa, Toxomerus

marginatus, and Lucilia coeruleiviridis. These five species of insects were not well

represented in our samples due to their speed and difficulty in collecting them with a

standard sweep net. Copestylum violaceum, Palpada mexicana, Ornidia obesa were all

observed to interact with mango flowers in the upper mango canopy (trees were up to

15ft tall). In addition to these insects there may be other pollinators that were outside

our sampling area.

In conclusion, this investigation identified the density, frequency, and behavior of

insects interacting with mango flowers during the 8-week bloom period of ‘Keitt’ mango

trees in three orchards in Homestead, Florida. Our results indicate that a wide diversity

of insects pollinate mango and there is a temporal shift of insects throughout the mango

bloom. When insects were excluded, there was an increase in fruit set up to 17%.

Moreover, differences in insect populations in separate orchards suggest that cultural

practices may have affected the populations of these insects and could be used to

increase these populations. Future research focusing on less observed and collected

insects such as C. violaceum, P. mexicana, O. obesa, and L. coeruleiviridis would help

to further fill the gaps in information of key pollinators for mango. Additional

96

investigations to increase pollination by augmenting mango orchards with the insects

shown in this research could be of practical and scientific value. Improvements of

cultural, biological, and physical methods to increase the natural populations of Musca

domestica, Allograpta obliqua, Sarcophagidae, Forcipomyia genualis, Hippelates sp.,

Liohippelates sp., Oscinella sp., Apis mellifera, and Camponotus plantatus may produce

an increase of pollination services in mango orchards.

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BIOGRAPHICAL SKETCH

Mr. Matthew R. Quenaudon finished his undergraduate degree of entomology at

Michigan State University in May of 2012. Matt went on to work as curator at The

Original Butterfly House & Insect World on Mackinac Island from May to Oct 2012. In

June of 2013, Matt worked as an integrated pest management intern at Phipps

Conservatory and Botanical Gardens, before working as integrated pest management

specialist in October 2013. Matt worked as an integrated pest management specialist

for 3 years until July 2016 before starting a master’s program at the University of Florida

in August of 2016. Matt is an entomologist with wide-ranging academic and

professional experience, who hopes to continue to translate his passion for nature with

a strong work ethic into a career grounded in positive ecological change. Matt strives to

use his expertise to tackle entomologic challenges both economic and environmental,

while challenging commonly-held stigmas about insects and emphasizing the

importance of insects to our world and way of living.

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