Characterization of the Malonate Utilization Operon in <i>Cronobacter sakazakii</i> Csak O:2, Sequence Type 64 Strains Using a Custom-Designed DNA Microarray and Whole Genome Sequencing

Gopinath, Gopal

Wednesday, August 3, 2016

America's Center - St. Louis

Gopal Gopinath, U.S. Food and Drug Administration, Laurel, MD

Jayanthi Gangiredla, U.S. Food and Drug Administration, Laurel, MD

Isha Patel, U.S. Food and Drug Administration, Laurel, MD

Qiongqiong Yan, University College Dublin, Dublin, Ireland

Venugopal Sathyamoorthy, U.S. Food and Drug Administration, Laurel, MD

Mahendra Kothary, U.S. Food and Drug Administration, Laurel, MD

Hediye Nese Cinar, U.S. Food and Drug Administration, Laurel, MD

Laurenda Carter, U.S. Food and Drug Administration, Laurel, MD

Hannah Chase, U.S. Food and Drug Administration, Laurel, MD

Eunbi Park, U.S. Food and Drug Administration, Laurel, MD

TaeJung Chung, U.S. Food and Drug Administration, Laurel, MD

YeonJoo Yoo, U.S. Food and Drug Administration, Laurel, MD

Jihyeon Park, U.S. Food and Drug Administration, Laurel, MD

Hyerim Choi, U.S. Food and Drug Administration, Laurel, MD

Seungeun Jeong, U.S. Food and Drug Administration, Laurel, MD

Soyoung Jun, U.S. Food and Drug Administration, Laurel, MD

MiJeong Kim, U.S. Food and Drug Administration, Laurel, MD

Seamus Fanning, University College Dublin, Dublin, Ireland

Carol Iversen, University of Dundee, Dundee, Scotland

Roger Stephan, University of Zurich, Zurich, Switzerland

Angelika Lehner, University of Zurich, Zurich, Switzerland

Guenter Klein, Tieraerztliche Hochschule Hannover, Hannover, Germany

Franco Pagotto, Health Canada, Ottawa, Canada

Jeffrey Farber, University of Guelph, Guelph, Canada

Ben D. Tall, U.S. Food and Drug Administration, Laurel, MD

Introduction: Malonate utilization, is an important differential trait, recognized as being possessed by six of the seven species of the foodborne pathogen Cronobacter and thought to be largely absent in Cronobacter sakazakii (Csak).

Purpose: In this study, a diverse group of 357 Cronobacter strains obtained during environmental sampling of five powdered infant formula production facilities in Ireland and Germany, and 93 milk powder and cheese manufacturing facilities in the USA were analyzed. This study describes the genomic diversity and phylogenetic relationships among these strains.

Methods: Genomic DNA was isolated using a QIAcube workstation and hybridizations were performed using a previously described pan-genome DNA microarray. Whole genome sequencing (WGS) was carried out on a MiSeq platform using Nextera XT chemistry.

Results: Microarray analysis (MA) determined that 22 phylogenetically-related Csak strains possessed genes of the C. malonaticus-like malonate utilization operon; and all, but one strain, could utilize malonate. The strains phylogenetically grouped as a separate clade among the Csak cluster. Additionally, these strains possessed the Csak O:2 serotype and 21 of the strains were identified as sequence type (ST) ST64. The malonate-negative strain was shown by MA and WGS to lack the transcriptional regulator, mdcR; and its ST was ST1. WGS confirmed that the nine gene, ~7.7 kbp malonate utilization operon was located in these strains between two flanking genes, gyrB and katG.

Significance: Until this investigation, the presence of malonate-positive Csak strains that are associated with foods was underappreciated, possibly leading to misidentification when relying on phenotypic identification alone. A custom designed pan genome microarray was useful in characterizing the total genome content of these CsakO:2, ST64/ST1, malonate-positive strains; and showed for the first time that these Csak strains were highly related in total gene content, serotype and ST. This study further establishes MA as a powerful platform for genomics research of Cronobacter.

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P3-63 Characterization of the Malonate Utilization Operon in Cronobacter sakazakii Csak O:2, Sequence Type 64 Strains Using a Custom-Designed DNA Microarray and Whole Genome Sequencing