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Front. Microbiol. | doi: 10.3389/fmicb.2018.02201

High-Throughput flaA Short Variable Region Sequencing to Assess Campylobacter Diversity from Fecal Samples from Birds

 Qian Zhang1, 2,  Gabriel A. Al-Ghalith2, Mayumi Kobayashi3,  Takahiro Segawa4, 5, Mitsuto Maeda3,  Satoshi Okabe3,  Dan Knights1, 2 and  Satoshi Ishii1, 2, 3, 6*
  • 1BioTechnology Institute, University of Minnesota Twin Cities, United States
  • 2Bioinformatics and Computational Biology, University of Minnesota Twin Cities, United States
  • 3Division of Environmental Engineering, Hokkaido University, Japan
  • 4Center for Life Science Research, University of Yamanashi, Japan
  • 5National Institute of Polar Research, Japan
  • 6Soil, Water, and Climate, University of Minnesota Twin Cities, United States

Current approach to identify sources of human pathogens is largely dependent on the cultivation and isolation of target bacteria. For rapid pathogen source identification, culture-independent strain typing method is necessary. In this study, we designed new primer set that broadly covers flaA short variable region (SVR) of various Campylobacter species, and applied the flaA SVR sequencing method to examine the diversity of Campylobacter spp. in geese fecal samples with and without bacteria cultivation. We also developed and optimized the sequence data analysis pipeline to identify as many genotypes as possible, while minimizing the detection of genotypes generated by sequencing errors. Our results suggest that the flaA SVR sequencing can cluster Campylobacter strains with higher discriminant power than conventional flaA restriction fragment length polymorphism (RFLP) method. Culture-independent flaA SVR MiSeq sequencing identified almost all flaA genotypes obtained by culture-based method. In addition, more flaA genotypes were identified probably due to high throughput nature of the MiSeq sequencing. These results suggest that the flaA SVR sequencing could be used to analyze the diversity of Campylobacter spp. without bacteria isolation. This method is promising to rapidly identify potential sources of Campylobacter pathogens.

Keywords: Campylobacter diversity, pathogens in environment, source tracking, genotyping, flaA, next generation sequencing

Received: 14 Feb 2018; Accepted: 28 Aug 2018.

Edited by:

Abd El-Latif Hesham, Assiut University, Egypt

Reviewed by:

Beatrix Stessl, Veterinärmedizinische Universität Wien, Austria
Shannon D. Manning, Michigan State University, United States  

Copyright: © 2018 Zhang, Al-Ghalith, Kobayashi, Segawa, Maeda, Okabe, Knights and Ishii. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Prof. Satoshi Ishii, University of Minnesota Twin Cities, BioTechnology Institute, 140 Gortner Lab, 1479 Gortner Avenue, St. Paul, Minneapolis, 55108, MN, United States,