Genomic characterization of malonate positive Cronobacter sakazakii serotype O:2, sequence type 64 strains, isolated from clinical, food, and environment samples
Gopal R. Gopinath,
Hannah R. Chase,
Jayanthi Gangiredla,
Athmanya Eshwar,
Hyein Jang,
Isha Patel,
Flavia Negrete,
Samantha Finkelstein,
Eunbi Park,
TaeJung Chung,
YeonJoo Yoo,
JungHa Woo,
YouYoung Lee,
Jihyeon Park,
Hyerim Choi,
Seungeun Jeong,
Soyoung Jun,
Mijeong Kim,
Chaeyoon Lee,
HyeJin Jeong,
Séamus Fanning,
Roger Stephan,
Carol Iversen,
Felix Reich,
Günter Klein,
Angelika Lehner,
Ben D. Tall
Affiliations
Gopal R. Gopinath
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Hannah R. Chase
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Jayanthi Gangiredla
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Athmanya Eshwar
Institute for Food Safety and Hygiene, University of Zurich
Hyein Jang
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Isha Patel
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Flavia Negrete
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Samantha Finkelstein
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Eunbi Park
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
TaeJung Chung
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
YeonJoo Yoo
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
JungHa Woo
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
YouYoung Lee
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Jihyeon Park
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Hyerim Choi
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Seungeun Jeong
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Soyoung Jun
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Mijeong Kim
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Chaeyoon Lee
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
HyeJin Jeong
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Séamus Fanning
UCD Centre for Food Safety, School of Public Health, Physiotherapy & Population Science, University College, Dublin & WHO Collaborating Centre for Cronobacter
Roger Stephan
Institute for Food Safety and Hygiene, University of Zurich
Carol Iversen
Institute for Food Safety and Hygiene, University of Zurich
Felix Reich
Institute for Food Quality and Safety, University of Veterinary Medicine Hannover
Günter Klein
Institute for Food Quality and Safety, University of Veterinary Medicine Hannover
Angelika Lehner
Institute for Food Safety and Hygiene, University of Zurich
Ben D. Tall
Center of Food Safety and Applied Nutrition, U. S. Food and Drug Administration
Abstract Background Malonate utilization, an important differential trait, well recognized as being possessed by six of the seven Cronobacter species is thought to be largely absent in Cronobacter sakazakii (Csak). The current study provides experimental evidence that confirms the presence of a malonate utilization operon in 24 strains of sequence type (ST) 64, obtained from Europe, Middle East, China, and USA; it offers explanations regarding the genomic diversity and phylogenetic relatedness among these strains, and that of other C. sakazakii strains. Results In this study, the presence of a malonate utilization operon in these strains was initially identified by DNA microarray analysis (MA) out of a pool of 347 strains obtained from various surveillance studies involving clinical, spices, milk powder sources and powdered infant formula production facilities in Ireland and Germany, and dried dairy powder manufacturing facilities in the USA. All ST64 C. sakazakii strains tested could utilize malonate. Zebrafish embryo infection studies showed that C. sakazakii ST64 strains are as virulent as other Cronobacter species. Parallel whole genome sequencing (WGS) and MA showed that the strains phylogenetically grouped as a separate clade among the Csak species cluster. Additionally, these strains possessed the Csak O:2 serotype. The nine-gene, ~ 7.7 kbp malonate utilization operon was located in these strains between two conserved flanking genes, gyrB and katG. Plasmidotyping results showed that these strains possessed the virulence plasmid pESA3, but in contrast to the USA ST64 Csak strains, ST64 Csak strains isolated from sources in Europe and the Middle East, did not possess the type six secretion system effector vgrG gene. Conclusions Until this investigation, the presence of malonate-positive Csak strains, which are associated with foods and clinical cases, was under appreciated. If this trait was used solely to identify Cronobacter strains, many strains would likely be misidentified. Parallel WGS and MA were useful in characterizing the total genome content of these Csak O:2, ST64, malonate-positive strains and further provides an understanding of their phylogenetic relatedness among other virulent C. sakazakii strains.
