Bess FleischmanAmulya Makkapati

Muhlenberg College Biology Department

Prevalence of Human pathogens in Ixodes scapularis ticks

Collected in the Lehigh Valley

Blacklegged Ticks

http://bugguide.net/node/view/108373

http://www.tickencounter.org/tick_identification/deer_tick

Winter

Fall Spring

Summer http://www.tickencounter.org/

Lyme Disease

Total U.S Lyme Disease Cases

1982 2004 2013

(Prior to 2014 press-release reporting likely CDC underestimate)

Duik-Wasser et al.Am. J. Trop Med. Hyg 2012

Predictive ModelElevationClimate Variables (Temperature, Humidity)GIS-based estimates of forest fragmentation.

Field Collected DataDensity of infected nymphs per 1000 m2

Lehigh Valley is in a transition zoneNo samples were taken from this region in this study, or any other published study

Duik-Wasser et al.Am. J. Trop Med. Hyg 2012

LowerHudson Valley

http://www.nsf.gov/news/special_reports/ecoinf/lyme.jsp

Conservation Biology 2003

Borrelia burgdorferiSpirocheteLyme Disease

Borrelia miyamotoiSpirocheteTick Borne Relapsing Fever

Anaplasma phagocytophilumBacteria that live inside white blood cellsAnaplasmosis

Babesia microtiProtozoa that lives in red blood cellsBabesiosis

More than one pathogenare transmitted by blacklegged ticks!

Babesiosis Anaplasmosis

1999 2010 1994 2010

Our Research Questions

Are the tick-borne pathogens that are abundant in the Lower Hudson Valley already established in the Lehigh Valley?

Do the lower rates of tick-borne disease in the Lehigh Valleycorrespond with lower rates of infection in ticks?

Can we establish a baseline of the current rates of infection to inform future studies in the Lehigh Valley?

LowerHudson Valley

Our Study

Nine sites428 Adult and Nymph Ticks

Average of 54 ticks per site (range 11-133)

DNAExtraction

Yittrium-ZirconiumCeramic Beads 2 mm and 0.1 mm

Pathogen DNA detected using Real-Time PCR

Advantages Real-Time PCRSensitive: Few false negatives for lack of detectionSpecific: Few false positives as a result of DNA contamination

Pathogen N positive/N tested

% Positive (Our

Study)

% Positive (Hudson

Valley)

B. burgdorferi Total

Nymphs

Adults

98/428 22.9

20/111 18.0 67%

78/317 24.6 60%

Lyme Disease

Results (Continued) Pathogen N positive/N

tested % Positive

(Our Study) % Positive

(Hudson Valley)

B. miyamotoi 2/428 0.5

Unidentified Borrelia spp. 14/428 3.3

A. phagocytophilum Total 8/428 1.9 8.5%

AP-ha 2/428 0.5

Ap-Variant 1 6/428 1.4

B. microti 2/428 0.5 1.2%

B. odocoilei 1/428 0.2

Coinfections

Pathogens N positive/N

tested

% Positive

B. burgdorferi, A. phagocytophilum (AP-ha) 1/428 0.2

B. burgdorferi, A. phagocytophilulm (Ap- Variant 1) 1/428 0.2B. burgdorferi, B. microti 2/428 0.5

Conclusions

Acknowledgements• Dr. Marten Edwards• Current Members of the Edwards Lab: Laura Barbalato, Kristina Fioretti, Rachel Heist

• Dr. Luther V. Rhodes Endowment for Infectious Disease Research through the LVHN

• Lehigh Valley Hospital Scholars Program• Muhlenberg College Department of Biology • James Vaughan Fund for Summer Research• Louise Bugbee, Meaghan Bird, Katerina Pham, Sarah Richardson

2 ul 0.12 ul

1 ul

0.25 ul

Control experiments with serial dilutions of pathogen control DNA showed that we could detect a very small number of pathogens in our tick samples.

PCR Assays

Ticks: ITS2 Ribosomal Gene

Borrelia burgdorferi

23S Ribosomal RNA: Common to several Borrelia spp.

Outer Surface Protein A (OspA): Only found in B. burgdorferi

PCR AssaysAnaplasma phagocytophilum

Msp2: bacterial surface proteinPositive samples were used in a sequencing reaction with a

different gene

B. microti

18S ribosomal gene sequence only found in B. microtiPositive samples were used in a sequencing reaction with a

different gene

B. miyamotoi

16S ribosomal gene sequence only found in B. miyamotoi Positive samples were used in a sequencing reaction with a

different gene

Lyme Disease

Babesiosis

Human Granulocytic Anaplasmosis

Tick-Borne Relapsing Fever

State/Area of residenceCalifornia 1 (0.1)Connecticut 74 (6.6)Delaware 1 (0.1)Indiana 0 (0.0)Maine 9 (0.8)Maryland 4 (0.4)Massachusetts 208 (18.5)Minnesota 73 (6.5)Nebraska 0 (0.0)New Hampshire 13 (1.2)New Jersey 166 (14.8)New York 361 (32.1)Oregon 1 (0.1)Rhode Island 73 (6.5)Tennessee 1 (0.1)Vermont 2 (0.2)Washington 0 (0.0)Wisconsin 80 (7.1)

Three cases in Eastern PA

http://www.cdc.gov/anaplasmosis/stats/