International Journal of Agriculture: Research and Review. Vol., 1 (3), 107-115, 2011Available online at http://www.ecisi.comISSN 2228-7973 2011 ECISI Journals

EFFECTS OF ARBUSCULAR MYCORRHIZA FUNGI (AMF), PLANT GROWTH PROMOTING

BACTERIA (PGPR) AND INTERACTION ON STRIGA HERMONTHICA MANAGEMENT IN

SORGHUM

MOHAMMED MAHGOUB HASSAN1, TILAL SAYED ABDELHALIM; SAMIA OSMAN YAGOUB3; AWAD GALAL OSMAN1;

MIGDAM EL SHEIK ABDEL GAIN1; ABDEL GABAR EL TAYEB BABIKER3

National Centre for Research Khartoum, Sudan 1Environment and Natural Resources Research Institutes Research

Institute, National Centre for Research, Khartoum, Sudan.2 Agricultural Research Corporation, Wad madni, Sudan.

3College of Agricultural Studies, Department of Crop Sciences, Sudan University of Science and Technology, Khartoum,

Sudan.

Corresponding author: Email: mohkadis@yahoo.com

Abstract: Our study was performed to test the effects of mycorrhiza fungi and plant growth promoting

bacteria on Striga control in sorghum. AM fungi negatively impacted on Striga germination. It reduced the

number of Striga seedlings attaching and emerging, and delayed the emergence time of Striga in pot

experiment. The performance of the AMF alone or in combinations with bacterial strains was significantly

better than that of the bacteria alone in terms of reduction of Striga infestation, plant height and dry matter.

AM fungi enhanced the performance of the sorghum host, allowing it to withstand Striga damage better. The

percent reduction (82%) of Striga emergence after AM inoculation resulted in about a 28% increase in total

dry matter of sorghum over the control. The plant height and dry matter of sorghum were significantly higher

in the combinations between AM + bacteria strains. Moreover, results of this experiment suggest that

mycorrhizal management might be an important factor in an integrated management with bacteria for Striga

control.

Key words: Striga hermonthica, AM, bacterial strains, suppression, PGPR

INTRODUCTION

A large number of different interactions

between fungi and bacteria occur in association with

plants, and depending on the nature of the species

involved, the plant can be positively or negatively

affected. There is also evidence that combined

interactions between AM fungi and PGPB strains can

enhance plant growth and that some of these

interactions may be very specific (Artursson et al.,

2006).Management of Striga spp. is often difficult due

to several reasons. These include the high amount of

Intl. J. Agric: Res & Rev. Vol., 1 (3), 107-115, 2011

108

seed production, viability of seeds in the soil over

several years, lack of seed germination in the absence of

a chemical trigger from a suitable host, vigorous growth

habit after emergence, and close association with the

host crop (Joel at al., 2007 and Press MC, 2008).

Several means for managing Striga spp. have been tried

over the years, albeit with somewhat limited

effectiveness. The development of new control

strategies should preferably focus on the initial steps

(germination and haustorium initiation) in the host

parasite interaction. Although Striga and Orobanche

spp. parasitize different hosts in different parts of the

world, their life cycles are very similar and involve

seeds germination in response to root host stimuli called

strigolactones. Strigolactones are signaling molecules

that play a double role in the rhizosphere as host

detection signals for arbuscular mycorrhizal (AM) fungi

and root parasitic plants (Akiyama et al.,2005). It has

been suggested that soil fertility and mineral nutrients

not only stimulate growth of the host, but also adversely

affect germination, attachment and subsequent

development of the parasitic weed (Abunyewa & Padi,

2003). Yoneyama and co workers reported that nutrient

deficiency, which in some cases is mitigated by AMF,

can increase strigolactone production by potential host

plants (Yoneyama et al., 2007). Recognition that

strigolactones that induce parasitic plant seeds to

germinate also recruit nutrient-supplying AMF suggests

that manipulating mycorrhizal colonization could be

used to manage parasitic plants (Akiyama et al., 2005).

Altogether, the positive effect of AM colonization on

plant fitness (facilitating the uptake of mineral nutrients

by plants) and their effect in reducing strigolactone

production and on the induction of plant defence genes

make AM fungi a suitable and promising tool for

controlling parasitic plants. Furthermore, a new group

of microorganisms have gained much attention because

of their effect on enhancement and health of crops

(Bloemberg and Lugtenberg 2001). These

microorganisms can influence plant growth by

producing one or more substances that act as growth

stimulators and are referred as plant growth promoting

bacteria/rhizobacteria (Xaio et al. 2002). One of the

alternative to control soilborne pathogens is the use of

antagonistic microorganisms (fungi or bacteria)

naturally found in soils, that could restrict pathogen

development and/or spread, thus reducing plant diseases.

These microorganisms use different biocontrol

strategies including production of antibiotics and

enzymes that could degrade the pathogen cell wall or

effective competition for nutrients (Verma et al. 2001).

In spite of intensive research, adequate strategies for

controlling parasitic plants remain elusive, and these

weeds continue to threaten agricultural crops worldwide.

The facts that, the dual inoculation of AM and specific

PGPR can enhance the activity of AM during the

symbiosis with the host plant (Artursson et al. 2006;

Richardson et al., 2009; Linderman 1997) and

synergetic effects in plant born diseases make the use of

both PGPR and mycorrhiza as promising biological

agent for Striga management. To our knowledge, there

is little information regarding the simultaneous and

enhancing effects of AM fungi and PGPR in terms of

parasitic weed management. The aim of this study;

therefore, is to investigate the effects of both a single

and co-inoculation of selected PGPR and AMF as

efficient biocontrol agent for Striga hermonthica in

Sorghum plants used through pot experiments

MATERIALS AND METHODS

The experiment used a randomized complete-

block design with two factors: one factor, AM fungal

inoculums, consisted of a mixed population of AM

mycorrhizal treatments: Glomus intraradices , G.

geosporunz, G. clarodium and Paraglomus spp.. The

second factor, microbial inoculants compoensations,

consisted of the application of bacterial strains

(Flavobacteria spp. (F), F+ Azotobacter vienlandi. (A),

Intl. J. Agric: Res & Rev. Vol., 1 (3), 107-115, 2011

109

F+ Bacillus megatherium var phosphaticum (BMP),

Mycorrihzae (AM), AM+F+A, AM+F+BMP, AM+F).

The experiments were conducted at Sudan

University during November 2010 January 2011 in a

greenhouse. Sorghum cv Abu Sabeen (Striga -

sensitive) was grown in a mixture of 2:1 of river silt and

sand. Plants were grown in black plastic bags (19 cm

diameter) filled with 4kg of the non-sterilized soil/sand

mixture, with drainage holes in the bottom. Sorghum

was inoculated with AM fungal inoculums, bacterial

and the combinations between AM fungi and bacteria

strains. Each treatment was replicates five times.

The mycorrhizal inocula obtained from pot

cultures of Sudan grass. as the host plant and

maintained by storage for 36 months in polyethylene

bags at 4_C. Two free-living nitrogen fixing

microorganisms, belonging to the genera Azotobacter,

Flavobacterium in addition to phosphorus solubilizing

microorganism belong to genera Bacillus, were assayed.

The bacterial strains were grown in Meat Peptone Broth

Media (MPBM), incubated overnight at 280C with

shaking. The bacterial strains were obtained from the

Environment and Natural Resources Research Institute

(ENRRI), the National Centre for Research and

University of Khartoum, Khartoum, Sudan, respectively.

Striga infestation was accomplished by

mixing 10 mg of sterilized Striga seeds (Ca 1500 seeds)

in the top 6 cm soil in each bag. Surface sterilized

sorghum seeds (7/bag) were sown and immediately

irrigated. Aliquots of the respective bacterial

suspensions (15 ml each) were injected into the soil

surface in each bag. However, mycorrhizal inoculum

was treated at sowing by spot application of 5 g of the

fungi inoculum to each pot and mixed with the soil

sand mixture. Subsequent irrigations were made every 2

days. Striga infested sorghum control was included for

comparison. Emergent Striga plants (Striga incidence)

were counted weekly starting from three weeks after

crop emergence. Sorghum height was measured at 6, 8,

10 and 12 weeks after sowing (WAS).

Measurements of the sorghum plants biomass

Upon harvest, root and shoot dry weights

were determined and the plant material dried at 80C

for 24 h in an oven to determine dry matter content. The

numbers of colony forming units (CFU) of bacterial

were determined in MPA medium using serial dilutions.

Statistical analysis

Data from the greenhouse experiments were

transformed to log (x + 0.5) in which x is the number of

Striga plants/bag and then subjected to analysis of

variance (ANOVA). Means were tested for significance

by LSD at 5%. The data were tabulated. Data on

sorghum biomass were subjected to ANOVA using the

statistical package SAS System for Windows (8th

edition).

RESULTS

Striga incidence

Striga infestation was influenced by the AM

fungi and bacteria (Table 1 and Fig. 1). At 4 WAS,

Striga was observed in all treatments except AMF

treatment. Striga emergence was very low as only 4.75

Striga plants emerged on the un-inoculated control (Fig.

1). All treatments reduced emergence of the parasite,

except the combinations between F+A which increased

the number of Striga emergence as compared to control.

Sorghum inoculated with the combinations between

AM+F+A sustained less Striga emergence than the

respective un-inoculated control.

At five WAS, un-inoculated sorghum

sustained the highest infestation (10 Striga plants/ bag).

Intl. J. Agric: Res & Rev. Vol., 1 (3), 107-115, 2011

110

Sorghum treated with the bacterial combinations F+A

and F+BMP had no adverse effects on Striga

emergence. In among all treatments, AM fungi was the

most suppressive. Inoculation of sorghum with AM

fungi reduced Striga infestation by 90%. While the AM

fungi incorporated with F+A reduced the parasite

emergence significantly but not more affective as

compared to AM fungi alone. It reduced Striga

infestation by 67.5%.

At six WAS, Striga emergence increased,

substantially, and was heighest on the uninoculated

control (12 Striga plants/ bag). All treatments reduced

Striga infestation, except the combinations of F+A.

Sorghum inoculated with AM fungi was most inhibitory.

It reduced Striga infestation by 79%. The combination

of AM fungi plus F displayed more reductions of the

parasite emergence (Fig. 1).

At seven WAS, the un-inoculated control

supported the highest Striga emergence (14 plants/ bag)

(Fig. 1). All treatments reduced Striga infestation as

compared to the control. In among all treatments AM

fungi alone or in combination with F+A were the most

inhibitory. They reduced the Striga infestation by 82-59,

respectively%. At eight WAS, the uninoculated control

supported the highest Striga emergence (17.25 plants/

bag) (Fig.1). Inoculation of sorghum with AM Fungi

reduced Striga infestation by 82 %. While sorghum

treated with the combinations of M+F+AZ or

M+F+BMP reduced Striga emergence by 52%. At 9

WAS, Striga incidence followed the same trend as at

eight WAS. The counts begin to decline in Striga

incidence with obvious trends (Fig. 1).

0

2

4

6

8

10

12

14

16

18

20

CONT

ROL

F+AZ

OTOB

ACTE

R

F+BM

P

FAVO

BACT

ERIA

M+F

M+F+

AZOT

M+F+

BMP

MYC

ORRH

IZA

Treatments

No.

of S

trig

a

4 WAS

5 WASk

6 WAS

8 WAS

9 WAS

sixth week

Fig. 1. Effects of AM fungi and bacterial strains on Striga incidence on sorghum Abu Sabeen. Vertical bar indicates LSD.

Plant height

Sorghum growth, as indicated by height, was

differentially affected by AM, bacteria and their

combinations. Results indicated that all treatments

increased sorghum height in comparison with the Striga

infested control albeit not significantly (Table 1 and Fig

2).

At 6 WAS, Striga infested uninoculated

sorghum displayed 15.97 cm in height. Sorghum

inoculation with Flavobacterium spp. alone or in

Intl. J. Agric: Res & Rev. Vol., 1 (3), 107-115, 2011

111

combinations with Mycorrhizae plus BMP increased

plant height significantly as compared to the un-

inoculated control. They increased sorghum height by

50%. Inoculation with the combinations between

Mycohrrizae plus Flavobacterium plus Azotobacter did

not have an effect on plant growth (Fig. 2). While

sorghum inoculated with AM fungi displayed slight

increment as compared to the control.

At 8 WAS, all treatments increase sorghum

growth as compare to the control. In among all

treatments the combinations of AM+F and

AM+F+BMP resulted in heighest increment (18 and

25 %, respectively) in sorghum height. At 10 WAS,

Striga infested uninoculated sorghum displayed 23 cm

in height. The combinations of AM plus

Flavobacterium ; AM plus Flavobacterium plus BMP

and AM alone increased sorghum height in comparison

to un-inoculated control. They increased sorghum

height by 40-50%, respectively. At 12 WAS, sorghum

height followed the same trend as at 10 WAS (Fig. 2).

05

101520253035

CONT

ROL

F+AZ

OTOB

ACTE

R

F+BM

P

FAVO

BACT

ERIA M+

F

M+F+

AZOT

M+F+

BMP

MYCO

RRHI

ZA

Treatments

Avera

ge p

lan

t h

eig

ht

(cm

)

6 WAS

8 WAS

10 WAS

12 WAS

02468

101214161820

CONT

ROL

F+AZ

OTOB

ACTE

R

F+BM

P

FAVO

BACT

ERIA M+

F

M+F+

AZOT

M+F+

BMP

MYCO

RRHI

ZA

Treatments

No

. o

f S

trig

a p

lan

ts/p

ot

4 WAS

5 WAS

6 WAS

7 WAS

8 WAS

9 WAS

Fig. 2. Effect of AM and bacterial strains on plant height of sorghum Abu sabeen. Vertical bar indicates S.E.

Intl. J. Agric: Res & Rev. Vol., 1 (3), 107-115, 2011

112

Table 1. Table 1 Effects of AMF, PGPR and their interaction on S. hermonthica management and sorghum height

F: Flavobavterium, A: Azotobacter vienlandi. BMP: Bacillus megatherium var phosphaticum

M-: without Mycorrihzae, M+: with Mycorrihzae, n.s.: no significant differences

( ) indicates back transformed data, data without () indicates square root transformed data (x+0.5 x: variable).

Total dry weight matter

Significant differences were observed among

the treatments in shoot, root and total dry matter of

sorghum (Table 2). At harvest, all treatments were

increased sorghum root dry weight except, F+BMP as

compared to the control. In among all treatments

sorghum inoculated with the combination of AM+F

recorded significantly higher dry weight accumulation

in root (5.82 g plant-1) followed by F+ A (5.22g plant-1)

as compared to the control. Significantly lowest dry

weight accumulation in root was recorded in a

combination of AM+F+ BMP (2.42 g plant-1).

Furthermore, results indicated that all treatments

increased sorghum shoot dry weight, except the

combinations between FandBMP as compared to the

control. Flavobacterium plus A (10.27 g plant -1) or in

combination with AM+BMP (9.4 g plant -1) displayed

the highest dry weight as compared to other treatments

(Table 2). Also results displayed that, all treatments

increased total dry weight, significantly, in comparison

with the infested control. The combinations between

F+A, AM+F and AM fungi alone displayed the highest

dry weight as compared to control. Moreover, sorghum

inoculated with the combination of F+ BMP or AM+F

increased the root: shoot ratio as compared to the

control (Table 2).

Table 2. Effect of AMF and bacterial strains on plant dry mattersTreatments (bacteria)

Plant parametersRDW (g) SHDW TDM (g) R:SH ratio

M- M+ M- M+ M- M+ M- M+Control 3.33 3.6 6.38 3.33 3.6 6.38 0.52 0.41F 3.93 5.8 7.6 3.93 5.8 7.6 0.52 0.77F+A 5.23 3.08 10.28 5.23 3.08 10.28 0.51 0.35F+BMP 4.05 2.43 5.48 4.05 2.43 5.475 0.74 0.26LSD for Interaction 1.16

0.410.20

3.211.140.57

3.511.240.62

n.s.n.s.n.s.

LSD for MycorrhizaeLSD for bacteria

F: Flavobavterium, A: Azotobacter vienlandi. BMP: Bacillus megatherium var phosphaticum

M-: without Mycorrihzae, M+: with Mycorrihzae, n.s.: no significant differences

DISCUSSION

Generally, the results of this study displayed

that AM fungi alone or in combinations with bacterial

strains significantly reduced and delayed Striga

infestation in sorghum, which resulted in taller plants,

higher shoot weight, total biomass, and higher

root:shoot ratio. This shows the potential of AM fungi

Plant parameters

Treatments (bacteria)Striga No. plant heightM- M+ M- M+

Control (24.25) 12.37 (3.33) 1.92 (40.15) 20.33 (43.69) 22.09F (14.42) 7.46 (16.62) 8.56 (43.39) 21.95 (45.15) 22.83F+AZOTO (21.33) 10.92 (10.98) 5.74 (43.65) 22.07 (41.39) 20.94F+BMP (17.75) 9.13 (13.75) 7.13 (42.32) 21.41 (47.38) 23.94LSD for Interaction 1.36LSD for Mycorrhizae 0.48LSD for Bacteria 0.24

n.sn.sn.s

Intl. J. Agric: Res & Rev. Vol., 1 (3), 107-115, 2011

113

for the control of Striga on sorghum. Gworgwor and

Weber (2003) have reported the successful use of G.

fasciculatum in controlling S. hermonthica in developed

resistant varieties of sorghum. F plus A increased Striga

emergence at 2 and 5 WAS, where the growth and

reproduction of the hemiparasite attacking the hosts

were significantly stimulated. On the other hand these

bacterial strains enhanced germination. Hence, they

enhance depletion of the parasite seed bank (Babiker et

al., 2000). These results are consistent with earlier

reports. Several mechanisms have been suggested by

which AM fungi could decrease damage by Striga

(Borowicz, 2001;Taylor and Harrier, 2003; Lendzemo

et al., 2007). These mechanisms include increased

nutritional status of the plant, changes in root exudates,

competition for colonization sites, and mobilization of

plant defence mechanisms after initial colonization by

the AM fungus. Like wise, strigolactones are required

by AM fungi for rapid colonization of their hosts,

selection of crop cultivars for lower induction of Striga

germination may be traded off against selection for

reduced mycorrhizal colonization. Reports showed that

lower biomass accumulation in Striga infested plant has

been attributed to competition between the host and

parasites for solutes, including carbon, lower rates of

photosynthesis in the leaves of infected plants (Press et

al., 1999), reduction in the level of growth promoting

hormones and increase in growth inhibiting hormones.

The reduction in root growth could have been due to the

reduction in available nutrient for root growtha

consequence of the nutrient flow into the parasite shoots

growing on their roots (Alonge et al., 2002). In the

absence of AM fungi, bacteria and their combinations,

Striga reduced sorghum biomass. Root dry matter of

sorghum increased in presence of F + A and AM+F.

Frankenberger and Arshad, (1995) observed that

infection by the mycorrhizal fungus, Glomus

fasciculatus resulted in significantly increased

Gibberellins (GA) activity in the leaves of Boutelona

gracilis, with a tendency for decreased GA activity in

the roots. Whereas shoot dry matter was significantly

increased in all treatments, except in the presence of

F+A. Furthermore, there was a high root: shoot ratio

with all treatments except in the combination between

AM+F+A; AM+F+BMP, which had a reduced root:

shoot ratio in comparison with to the control.

Frankenberger and Arshad, (1995) reported that two

bacterial strains belong to Enterobacteriaceae were

capable of producing copious amounts of IAA, leading

to reduced plant root elongation and an increased

shoot/root ratio in sugar beet when applied as seeds

inoculum. Moreover, this mycorrhizosphere effect

could be attributed to AM fungi causing nutrient

leakage from roots (quantitative changes in root

exudates) or specific changes in the quality of root

exudates. The results showed that mycorrhizae are the

only AM fungal treatment in this study proved to be in

improving plant growth and in controlling Striga. The

high shoot weight produced by sorghum inoculated with

AM alone or in combinations with bacterial strains

indicates an efficient compensatory effect of the AM, as

well as the significant role in control of Striga.

Frankenberger and Arshad (1995) reported that plant

hormones interact (synergistic and antagonistic) with

each other. Under different environmental and soil

conditions, the concentration of bacterial-produced

plant hormones may vary, which could impinge on the

effectiveness of inocula. The potential use of AM fungi

to control Striga or compensate for the negative effects

of the parasite will be important for soil management,

especially in the tropics, where herbicides and fertilizers

are an expensive input for farmers. These results are

agreement with the outcomes of previous studies under

controlled conditions (Gworgwor and Weber, 2003;

Lendzemo, 2004). Edriss et al., (1984) reported that

dry weight and cytokinine levels of the mycorrhizal

plants however, were still twice those of the non-

mycorrhizal plants. The plant height and dry matter of

sorghum were significantly higher in the combinations

between AM + bacteria strains. While the results of this

Intl. J. Agric: Res & Rev. Vol., 1 (3), 107-115, 2011

114

experiment suggest that mycorrhizal management might

be an important factor in an integrated management

(bacteria) of Striga, several issues need further studies.

CONCLUSIONS

The high specificity of many organisms (AM

fungi and bacteria), feeding exclusively on

selected hosts, in our case parasitic weeds,

can be considered an advantage because these

organisms may work as biocontrol agents

where other weed control options have failed.

Future research should focuson i) re-

screening of the effective microorganisms and

rank them according to their ability to

suppress or promote specific stages in Striga

life cycle, ii) screen various inexpensive

compounds and local materials as

physiological precursors for the respective

phytohormones.

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International Journal of Agriculture: Research and Review. Vol., 1 (3), 107-115, 2011

Available online at http://www.ecisi.com

ISSN 2228-7973 2011 ECISI Journals

Effects of Arbuscular Mycorrhiza Fungi (AMF), Plant Growth Promoting Bacteria (PGPR) and Interaction on Striga hermonthica management in sorghum

Mohammed Mahgoub Hassan1, Tilal Sayed Abdelhalim; Samia Osman Yagoub3; Awad Galal Osman1; Migdam El sheik Abdel gain1; Abdel Gabar El Tayeb Babiker3

National Centre for Research Khartoum, Sudan 1Environment and Natural Resources Research Institutes Research Institute, National Centre for Research, Khartoum, Sudan.

2 Agricultural Research Corporation, Wad madni, Sudan.

3College of Agricultural Studies, Department of Crop Sciences, Sudan University of Science and Technology, Khartoum, Sudan.

Corresponding author: Email: mohkadis@yahoo.com

Abstract: Our study was performed to test the effects of mycorrhiza fungi and plant growth promoting bacteria on Striga control in sorghum. AM fungi negatively impacted on Striga germination. It reduced the number of Striga seedlings attaching and emerging, and delayed the emergence time of Striga in pot experiment. The performance of the AMF alone or in combinations with bacterial strains was significantly better than that of the bacteria alone in terms of reduction of Striga infestation, plant height and dry matter. AM fungi enhanced the performance of the sorghum host, allowing it to withstand Striga damage better. The percent reduction (82%) of Striga emergence after AM inoculation resulted in about a 28% increase in total dry matter of sorghum over the control. The plant height and dry matter of sorghum were significantly higher in the combinations between AM + bacteria strains. Moreover, results of this experiment suggest that mycorrhizal management might be an important factor in an integrated management with bacteria for Striga control.

Key words: Striga hermonthica, AM, bacterial strains, suppression, PGPR

Introduction

A large number of different interactions between fungi and bacteria occur in association with plants, and depending on the nature of the species involved, the plant can be positively or negatively affected. There is also evidence that combined interactions between AM fungi and PGPB strains can enhance plant growth and that some of these interactions may be very specific (Artursson et al., 2006).Management of Striga spp. is often difficult due to several reasons. These include the high amount of seed production, viability of seeds in the soil over several years, lack of seed germination in the absence of a chemical trigger from a suitable host, vigorous growth habit after emergence, and close association with the host crop (Joel at al., 2007 and Press MC, 2008). Several means for managing Striga spp. have been tried over the years, albeit with somewhat limited effectiveness. The development of new control strategies should preferably focus on the initial steps (germination and haustorium initiation) in the hostparasite interaction. Although Striga and Orobanche spp. parasitize different hosts in different parts of the world, their life cycles are very similar and involve seeds germination in response to root host stimuli called strigolactones. Strigolactones are signaling molecules that play a double role in the rhizosphere as host detection signals for arbuscular mycorrhizal (AM) fungi and root parasitic plants (Akiyama et al.,2005). It has been suggested that soil fertility and mineral nutrients not only stimulate growth of the host, but also adversely affect germination, attachment and subsequent development of the parasitic weed (Abunyewa & Padi, 2003). Yoneyama and co workers reported that nutrient deciency, which in some cases is mitigated by AMF, can increase strigolactone production by potential host plants (Yoneyama et al., 2007). Recognition that strigolactones that induce parasitic plant seeds to germinate also recruit nutrient-supplying AMF suggests that manipulating mycorrhizal colonization could be used to manage parasitic plants (Akiyama et al., 2005). Altogether, the positive effect of AM colonization on plant tness (facilitating the uptake of mineral nutrients by plants) and their effect in reducing strigolactone production and on the induction of plant defence genes make AM fungi a suitable and promising tool for controlling parasitic plants. Furthermore, a new group of microorganisms have gained much attention because of their effect on enhancement and health of crops (Bloemberg and Lugtenberg 2001). These microorganisms can influence plant growth by producing one or more substances that act as growth stimulators and are referred as plant growth promoting bacteria/rhizobacteria (Xaio et al. 2002). One of the alternative to control soilborne pathogens is the use of antagonistic microorganisms (fungi or bacteria) naturally found in soils, that could restrict pathogen development and/or spread, thus reducing plant diseases. These microorganisms use different biocontrol strategies including production of antibiotics and enzymes that could degrade the pathogen cell wall or effective competition for nutrients (Verma et al. 2001). In spite of intensive research, adequate strategies for controlling parasitic plants remain elusive, and these weeds continue to threaten agricultural crops worldwide. The facts that, the dual inoculation of AM and specific PGPR can enhance the activity of AM during the symbiosis with the host plant (Artursson et al. 2006; Richardson et al., 2009; Linderman 1997) and synergetic effects in plant born diseases make the use of both PGPR and mycorrhiza as promising biological agent for Striga management. To our knowledge, there is little information regarding the simultaneous and enhancing effects of AM fungi and PGPR in terms of parasitic weed management. The aim of this study; therefore, is to investigate the effects of both a single and co-inoculation of selected PGPR and AMF as efficient biocontrol agent for Striga hermonthica in Sorghum plants used through pot experiments

Materials and methods

The experiment used a randomized complete-block design with two factors: one factor, AM fungal inoculums, consisted of a mixed population of AM mycorrhizal treatments: Glomus intraradices , G. geosporunz, G. clarodium and Paraglomus spp.. The second factor, microbial inoculants compoensations, consisted of the application of bacterial strains (Flavobacteria spp. (F), F+ Azotobacter vienlandi. (A), F+ Bacillus megatherium var phosphaticum (BMP), Mycorrihzae (AM), AM+F+A, AM+F+BMP, AM+F).

The experiments were conducted at Sudan University during November 2010 January 2011 in a greenhouse. Sorghum cv Abu Sabeen (Striga - sensitive) was grown in a mixture of 2:1 of river silt and sand. Plants were grown in black plastic bags (19 cm diameter) filled with 4kg of the non-sterilized soil/sand mixture, with drainage holes in the bottom. Sorghum was inoculated with AM fungal inoculums, bacterial and the combinations between AM fungi and bacteria strains. Each treatment was replicates five times.

The mycorrhizal inocula obtained from pot cultures of Sudan grass. as the host plant and maintained by storage for 36 months in polyethylene bags at 4_C. Two free-living nitrogen fixing microorganisms, belonging to the genera Azotobacter, Flavobacterium in addition to phosphorus solubilizing microorganism belong to genera Bacillus, were assayed. The bacterial strains were grown in Meat Peptone Broth Media (MPBM), incubated overnight at 280C with shaking. The bacterial strains were obtained from the Environment and Natural Resources Research Institute (ENRRI), the National Centre for Research and University of Khartoum, Khartoum, Sudan, respectively.

Striga infestation was accomplished by mixing 10 mg of sterilized Striga seeds (Ca 1500 seeds) in the top 6 cm soil in each bag. Surface sterilized sorghum seeds (7/bag) were sown and immediately irrigated. Aliquots of the respective bacterial suspensions (15 ml each) were injected into the soil surface in each bag. However, mycorrhizal inoculum was treated at sowing by spot application of 5 g of the fungi inoculum to each pot and mixed with the soilsand mixture. Subsequent irrigations were made every 2 days. Striga infested sorghum control was included for comparison. Emergent Striga plants (Striga incidence) were counted weekly starting from three weeks after crop emergence. Sorghum height was measured at 6, 8, 10 and 12 weeks after sowing (WAS).

Measurements of the sorghum plants biomass

Upon harvest, root and shoot dry weights were determined and the plant material dried at 80C for 24 h in an oven to determine dry matter content. The numbers of colony forming units (CFU) of bacterial were determined in MPA medium using serial dilutions.

Statistical analysis

Data from the greenhouse experiments were transformed to log (x + 0.5) in which x is the number of Striga plants/bag and then subjected to analysis of variance (ANOVA). Means were tested for significance by LSD at 5%. The data were tabulated. Data on sorghum biomass were subjected to ANOVA using the statistical package SAS System for Windows (8th edition).

Results

Striga incidence

Striga infestation was influenced by the AM fungi and bacteria (Table 1 and Fig. 1). At 4 WAS, Striga was observed in all treatments except AMF treatment. Striga emergence was very low as only 4.75 Striga plants emerged on the un-inoculated control (Fig. 1). All treatments reduced emergence of the parasite, except the combinations between F+A which increased the number of Striga emergence as compared to control. Sorghum inoculated with the combinations between AM+F+A sustained less Striga emergence than the respective un-inoculated control.

At five WAS, un-inoculated sorghum sustained the highest infestation (10 Striga plants/ bag). Sorghum treated with the bacterial combinations F+A and F+BMP had no adverse effects on Striga emergence. In among all treatments, AM fungi was the most suppressive. Inoculation of sorghum with AM fungi reduced Striga infestation by 90%. While the AM fungi incorporated with F+A reduced the parasite emergence significantly but not more affective as compared to AM fungi alone. It reduced Striga infestation by 67.5%.

At six WAS, Striga emergence increased, substantially, and was heighest on the uninoculated control (12 Striga plants/ bag). All treatments reduced Striga infestation, except the combinations of F+A. Sorghum inoculated with AM fungi was most inhibitory. It reduced Striga infestation by 79%. The combination of AM fungi plus F displayed more reductions of the parasite emergence (Fig. 1).

At seven WAS, the un-inoculated control supported the highest Striga emergence (14 plants/ bag) (Fig. 1). All treatments reduced Striga infestation as compared to the control. In among all treatments AM fungi alone or in combination with F+A were the most inhibitory. They reduced the Striga infestation by 82-59, respectively%. At eight WAS, the uninoculated control supported the highest Striga emergence (17.25 plants/ bag) (Fig.1). Inoculation of sorghum with AM Fungi reduced Striga infestation by 82 %. While sorghum treated with the combinations of M+F+AZ or M+F+BMP reduced Striga emergence by 52%. At 9 WAS, Striga incidence followed the same trend as at eight WAS. The counts begin to decline in Striga incidence with obvious trends (Fig. 1).

0

2

4

6

8

10

12

14

16

18

20

CONTROL

F+AZOTOBACTER

F+BMP

FAVOBACTERIA

M+F

M+F+AZOT

M+F+BMP

MYCORRHIZA

Treatments

No. of Striga

4 WAS

5 WASk

6 WAS

8 WAS

9 WAS

sixth week

Fig. 1. Effects of AM fungi and bacterial strains on Striga incidence on sorghum Abu Sabeen. Vertical bar indicates LSD.

Plant height

Sorghum growth, as indicated by height, was differentially affected by AM, bacteria and their combinations. Results indicated that all treatments increased sorghum height in comparison with the Striga infested control albeit not significantly (Table 1 and Fig 2).

At 6 WAS, Striga infested uninoculated sorghum displayed 15.97 cm in height. Sorghum inoculation with Flavobacterium spp. alone or in combinations with Mycorrhizae plus BMP increased plant height significantly as compared to the un-inoculated control. They increased sorghum height by 50%. Inoculation with the combinations between Mycohrrizae plus Flavobacterium plus Azotobacter did not have an effect on plant growth (Fig. 2). While sorghum inoculated with AM fungi displayed slight increment as compared to the control.

At 8 WAS, all treatments increase sorghum growth as compare to the control. In among all treatments the combinations of AM+F and AM+F+BMP resulted in heighest increment (18 and 25 %, respectively) in sorghum height. At 10 WAS, Striga infested uninoculated sorghum displayed 23 cm in height. The combinations of AM plus Flavobacterium ; AM plus Flavobacterium plus BMP and AM alone increased sorghum height in comparison to un-inoculated control. They increased sorghum height by 40-50%, respectively. At 12 WAS, sorghum height followed the same trend as at 10 WAS (Fig. 2).

0

5

10

15

20

25

30

35

CONTROL

F+AZOTOBACTER

F+BMP

FAVOBACTERIA

M+F

M+F+AZOT

M+F+BMP

MYCORRHIZA

Treatments

Average plant height (cm)

6 WAS

8 WAS

10 WAS

12 WAS

0

2

4

6

8

10

12

14

16

18

20

CONTROL

F+AZOTOBACTER

F+BMP

FAVOBACTERIA

M+F

M+F+AZOT

M+F+BMP

MYCORRHIZA

Treatments

No. of Striga plants/pot

4 WAS

5 WAS

6 WAS

7 WAS

8 WAS

9 WAS

Fig. 2. Effect of AM and bacterial strains on plant height of sorghum Abu sabeen. Vertical bar indicates S.E.

Table 1. Table 1 Effects of AMF, PGPR and their interaction on S. hermonthica management and sorghum height

Plant parameters

Treatments (bacteria)

Striga No.

plant height

M-

M+

M-

M+

Control

(24.25)

12.37

(3.33)

1.92

(40.15)

20.33

(43.69)

22.09

F

(14.42)

7.46

(16.62)

8.56

(43.39)

21.95

(45.15)

22.83

F+AZOTO

(21.33)

10.92

(10.98)

5.74

(43.65)

22.07

(41.39)

20.94

F+BMP

(17.75)

9.13

(13.75)

7.13

(42.32)

21.41

(47.38)

23.94

LSD for Interaction 1.36

LSD for Mycorrhizae 0.48

LSD for Bacteria 0.24

n.s

n.s

n.s

F: Flavobavterium, A: Azotobacter vienlandi. BMP: Bacillus megatherium var phosphaticum

M-: without Mycorrihzae, M+: with Mycorrihzae, n.s.: no significant differences

( ) indicates back transformed data, data without () indicates square root transformed data (x+0.5 x: variable).

Total dry weight matter

Significant differences were observed among the treatments in shoot, root and total dry matter of sorghum (Table 2). At harvest, all treatments were increased sorghum root dry weight except, F+BMP as compared to the control. In among all treatments sorghum inoculated with the combination of AM+F recorded significantly higher dry weight accumulation in root (5.82 g plant-1) followed by F+ A (5.22g plant-1) as compared to the control. Significantly lowest dry weight accumulation in root was recorded in a combination of AM+F+ BMP (2.42 g plant-1). Furthermore, results indicated that all treatments increased sorghum shoot dry weight, except the combinations between FandBMP as compared to the control. Flavobacterium plus A (10.27 g plant -1) or in combination with AM+BMP (9.4 g plant -1) displayed the highest dry weight as compared to other treatments (Table 2). Also results displayed that, all treatments increased total dry weight, significantly, in comparison with the infested control. The combinations between F+A, AM+F and AM fungi alone displayed the highest dry weight as compared to control. Moreover, sorghum inoculated with the combination of F+ BMP or AM+F increased the root: shoot ratio as compared to the control (Table 2).

Table 2. Effect of AMF and bacterial strains on plant dry matters

Treatments (bacteria)

Plant parameters

RDW (g)

SHDW

TDM (g)

R:SH ratio

M-

M+

M-

M+

M-

M+

M-

M+

Control

3.33

3.6

6.38

3.33

3.6

6.38

0.52

0.41

F

3.93

5.8

7.6

3.93

5.8

7.6

0.52

0.77

F+A

5.23

3.08

10.28

5.23

3.08

10.28

0.51

0.35

F+BMP

4.05

2.43

5.48

4.05

2.43

5.475

0.74

0.26

LSD for Interaction

1.16

0.41

0.20

3.21

1.14

0.57

3.51

1.24

0.62

n.s.

n.s.

n.s.

LSD for Mycorrhizae

LSD for bacteria

F: Flavobavterium, A: Azotobacter vienlandi. BMP: Bacillus megatherium var phosphaticum

M-: without Mycorrihzae, M+: with Mycorrihzae, n.s.: no significant differences

Discussion

Generally, the results of this study displayed that AM fungi alone or in combinations with bacterial strains significantly reduced and delayed Striga infestation in sorghum, which resulted in taller plants, higher shoot weight, total biomass, and higher root:shoot ratio. This shows the potential of AM fungi for the control of Striga on sorghum. Gworgwor and Weber (2003) have reported the successful use of G. fasciculatum in controlling S. hermonthica in developed resistant varieties of sorghum. F plus A increased Striga emergence at 2 and 5 WAS, where the growth and reproduction of the hemiparasite attacking the hosts were significantly stimulated. On the other hand these bacterial strains enhanced germination. Hence, they enhance depletion of the parasite seed bank (Babiker et al., 2000). These results are consistent with earlier reports. Several mechanisms have been suggested by which AM fungi could decrease damage by Striga (Borowicz, 2001;Taylor and Harrier, 2003; Lendzemo et al., 2007). These mechanisms include increased nutritional status of the plant, changes in root exudates, competition for colonization sites, and mobilization of plant defence mechanisms after initial colonization by the AM fungus. Like wise, strigolactones are required by AM fungi for rapid colonization of their hosts, selection of crop cultivars for lower induction of Striga germination may be traded off against selection for reduced mycorrhizal colonization. Reports showed that lower biomass accumulation in Striga infested plant has been attributed to competition between the host and parasites for solutes, including carbon, lower rates of photosynthesis in the leaves of infected plants (Press et al., 1999), reduction in the level of growth promoting hormones and increase in growth inhibiting hormones. The reduction in root growth could have been due to the reduction in available nutrient for root growtha consequence of the nutrient flow into the parasite shoots growing on their roots (Alonge et al., 2002). In the absence of AM fungi, bacteria and their combinations, Striga reduced sorghum biomass. Root dry matter of sorghum increased in presence of F + A and AM+F. Frankenberger and Arshad, (1995) observed that infection by the mycorrhizal fungus, Glomus fasciculatus resulted in significantly increased Gibberellins (GA) activity in the leaves of Boutelona gracilis, with a tendency for decreased GA activity in the roots. Whereas shoot dry matter was significantly increased in all treatments, except in the presence of F+A. Furthermore, there was a high root: shoot ratio with all treatments except in the combination between AM+F+A; AM+F+BMP, which had a reduced root: shoot ratio in comparison with to the control. Frankenberger and Arshad, (1995) reported that two bacterial strains belong to Enterobacteriaceae were capable of producing copious amounts of IAA, leading to reduced plant root elongation and an increased shoot/root ratio in sugar beet when applied as seeds inoculum. Moreover, this mycorrhizosphere effect could be attributed to AM fungi causing nutrient leakage from roots (quantitative changes in root exudates) or specific changes in the quality of root exudates. The results showed that mycorrhizae are the only AM fungal treatment in this study proved to be in improving plant growth and in controlling Striga. The high shoot weight produced by sorghum inoculated with AM alone or in combinations with bacterial strains indicates an efficient compensatory effect of the AM, as well as the significant role in control of Striga. Frankenberger and Arshad (1995) reported that plant hormones interact (synergistic and antagonistic) with each other. Under different environmental and soil conditions, the concentration of bacterial-produced plant hormones may vary, which could impinge on the effectiveness of inocula. The potential use of AM fungi to control Striga or compensate for the negative effects of the parasite will be important for soil management, especially in the tropics, where herbicides and fertilizers are an expensive input for farmers. These results are agreement with the outcomes of previous studies under controlled conditions (Gworgwor and Weber, 2003; Lendzemo, 2004). Edriss et al., (1984) reported that dry weight and cytokinine levels of the mycorrhizal plants however, were still twice those of the non-mycorrhizal plants. The plant height and dry matter of sorghum were significantly higher in the combinations between AM + bacteria strains. While the results of this experiment suggest that mycorrhizal management might be an important factor in an integrated management (bacteria) of Striga, several issues need further studies.

Conclusions

The high specificity of many organisms (AM fungi and bacteria), feeding exclusively on selected hosts, in our case parasitic weeds, can be considered an advantage because these organisms may work as biocontrol agents where other weed control options have failed.

Future research should focuson i) re-screening of the effective microorganisms and rank them according to their ability to suppress or promote specific stages in Striga life cycle, ii) screen various inexpensive compounds and local materials as physiological precursors for the respective phytohormones.

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