Ranga Sampath

Genotypic Variation and Mixtures of Lyme Borrelia in Ixodes Ticks from North America and Europe

Background Lyme disease, caused by various species of Borrelia, is transmitted by Ixodes ticks in North America and Europe. Studies have shown the genotype of Borrelia burgdorferi sensu stricto (s.s.) or the species of B. burgdorferi sensu lato (s.l.) affects the ability of the bacteria to cause local or disseminated infection in humans. Methodology/Principal Findings We used a multilocus PCR electrospray mass spectrometry assay to determine the species and genotype Borrelia from ticks collected in New York, Connecticut, Indiana, Southern Germany, and California and characterized isolates from parts of the United States and Europe. These analyses identified 53 distinct genotypes of B. burgdorferi sensu stricto with higher resolution than ospC typing. Genotypes of other members of the B. burgdorferi sensu lato complex were also identified and genotyped including B. afzelii, B. garinii, B. lusitaniae, B. spielmanii, and B. valaisiana. While each site in North America had genotypes unique to that location, we found genotypes shared between individual regions and two genotypes found across the United States. Significant B. burgdorferi s.s. genotypic diversity was observed between North America and Europe: only 6.6% of US genotypes (3 of 45) were found in Europe and 27% of the European genotypes (3 of 11) were observed in the US. Interestingly, 39% of adult Ixodes scapularis ticks from North America were infected with more than one genotype of B. burgdorferi s.s. and 22.2% of Ixodes ricinus ticks from Germany were infected with more than one genotype of B. burgdorferi s.l. Conclusions/Significance The presence of multiple Borrelia genotypes in ticks increases the probability that a person will be infected with more than one genotype of B. burgdorferi, potentially increasing the risks of disseminated Lyme disease. Our study indicates that the genotypic diversity of Borrelia in ticks in both North America and Europe is higher then previously reported and can have potential clinical consequences.

Usefulness of multilocus polymerase chain reaction followed by electrospray ionization mass spectrometry to identify a diverse panel of bacterial isolates

Polymerase chain reaction electrospray ionization mass spectrometry (PCR/ESI-MS) was tested for its ability to accurately identify a blinded panel of 156 diverse bacterial isolates, mostly human and/or animal pathogens. Here, 142/156 (91%) isolates were correctly identified to the genus level and 115/156 (74%) were correctly identified to the species level. Only 9% were misidentified. This study shows that multilocus PCR/ESI-MS has the potential to be a useful technique for identifying a broad range of bacteria.

Detection of heartworm infection in dogs via PCR amplification and electrospray ionization mass spectrometry of nucleic acid extracts from whole blood samples

OBJECTIVE To develop and evaluate a rapid and accurate assay involving PCR amplification and electrospray ionization mass spectrometry of nucleic acid extracts from whole blood samples for the detection of Dirofilaria immitis infection in dogs. SAMPLE Whole blood nucleic acid extracts from 29 dogs experimentally infected with D immitis (and in which circulating D immitis antigen was detected) and 10 uninfected dogs. PROCEDURES 16 of the 29 whole blood samples from infected dogs were examined at the time of collection for circulating microfilaria. Nucleic acids were extracted from all whole blood specimens and underwent PCR amplification with 12 PCR primer pairs designed to detect a wide range of pathogens (including the Wolbachia endosymbiont of D immitis) and electrospray ionization mass spectrometry. RESULTS On the basis of assay results, heartworm infection was detected in 13 of 13 antigen-positive dogs of unknown microfilaria status, 11 of 11 antigen-positive dogs with circulating microfilaria, 0 of 3 antigen-positive dogs tested at 3 months after larval infection, 0 of 2 antigen-positive dogs with occult infections, and 0 of 10 uninfected dogs. CONCLUSIONS AND CLINICAL RELEVANCE With the assay under investigation, it was possible to identify D immitis infection in dogs with circulating microfilaria via detection of the obligate Wolbachia endosymbiont of D immitis. It was not possible to identify dogs with occult infections, which suggested that circulating microfilaria must be present to detect infection with this assay, although further studies would be required to verify that finding.

608Can Rapid Molecular Diagnostics Assist in the Choice of b-Lactam Antibiotics? An Analysis of Data from PRIMERS-II of the Antibiotic Resistance Leadership Group (ARLG)

608. Can Rapid Molecular Diagnostics Assist in the Choice of b-Lactam Antibiotics? An Analysis of Data from PRIMERS-II of the Antibiotic Resistance Leadership Group (ARLG) Andrea M. Hujer, BS; Scott Evans, PhD; Hongyu Jiang, PhD; Kristine M. Hujer, BS; Thomas Hall, PhD; Christine Marzan, PhD; Michael Jacobs, MD, PhD; Ranga Sampath, PhD; David J. Ecker, PhD; T. Nicholas Domitrovic, BA; Claudia Manca, PhD; Kalyan Chavda, PhD; Pan Zhang, MD, PhD; Liang Chen, PhD; Carol Hill, PhD; Federico Perez, MD; Barry Kreiswirth, PhD; Vance Fowler, MD; Henry F. Chambers, MD; Robert A. Bonomo, MD; Antimicrobial Resistance Leadership Group; Case Western Reserve University, Cleveland, OH; Center for Biostatistics in AIDS Research, Harvard University, Boston, MA; Biostatistics, Harvard School of Public Health, Boston, MA; Louis Stokes Cleveland Veterans Affairs Medical Center, Cleveland, OH; Ibis Biosciences, Carlsbad, CA; Case Western Reserve University/University Hospitals of Cleveland, Cleveland, OH; Ibis Biosciences, Inc., A Division of Abbott, Carlsbad, CA; PHRI UMDNJ, Newark, NJ; Weill Cornell Medical College, New York, NY; PHRI and UMDNJ, Newark, NJ; Duke University Medical Center, Durham, NC; Cleveland VAMC Case Western Reserve University, Cleveland Heights, OH; University of Medicine and Dentistry of NJ, PHRI TB Center, Newark, NJ; University of California, San Francisco General Hospital, San Francisco, CA; Medicine, Pharmacology and Molecular Biology and Microbiology, Case Western Reserve University School of Medicine, Cleveland, OH