Cronobacter spp. (formerly known as Enterobacter sakazakii) are foodborne pathogens that cause rare but life-threatening diseases in neonates and infants through consumption of contaminated powdered infant formula. This study was conducted to investigate the occurrence of Cronobacter spp. in powdered formula in China and to further characterize Cronobacter isolates. Isolates were identified to the species level based on the fusA gene sequence, and strains of C. sakazakii were further subtyped by applying the polymerase chain reaction (PCR)–based serotyping method. A total of 23 strains of Cronobacter spp. isolated from 530 powdered formula samples were identified using conventional biochemical methods and duplex PCR. Cronobacter spp. were detected in 6.25%, 1.82%, 3.64%, 5.45%, and 2.50% of the general formula, infant formula (age <6 months), follow-up formula (6–12 months of age), growing-up formula (1–3 years of age), and children's formula (3–6 years of age), respectively. The individual species were identified as C. sakazakii (22 isolates) and C. malonaticus (1 isolate). Among 22 C. sakazakii isolates, representatives of all but two O-antigen serotypes (serotypes O5 and O6) were recognized.
Get full access to this article
View all access options for this article.
References
1.
BaldwinA, LoughlinM, Caubilla-BarronJ, et al.Multilocus sequence typing of Cronobacter sakazakii and Cronobacter malonaticus reveals stable clonal structures with clinical significance which do not correlate with biotypes. BMC Microbiol, 2009; 9:223.
2.
BaumgartnerA, GrandM, LinigerM, et al.Detection and frequency of Cronobacter spp. (Enterobacter sakazakii) in different categories of ready-to-eat foods other than infant formula. Int J Food Microbiol, 2009; 136:189–192.
Caubilla-BarronJ, HurrellE, TownsendS, et al.Genotypic and phenotypic analysis of Enterobacter sakazakii strains from an outbreak resulting in fatalities in a neonatal intensive care unit in France. J Clin Microbiol, 2007; 45:3979–3985.
5.
[CDC] Centers for Disease Control and Prevention. Cronobacter species isolation in two infants—New Mexico, 2008. Morb Mortal Weekly Rep, 2009; 58:1179–1183.
6.
CDC. CDC update: Investigation of Cronobacter infections among infants in the United States. January13, 2012. Available at: http://www.cdc.gov/foodsafety/deseases/cronobacter/investigation.html, accessed August22, 2012.
7.
ChapJ, JacksonP, SiqueiraR, et al.International survey of Cronobacter sakazakii and other Cronobacter spp. in follow up formulas and infant foods. Int J Food Microbiol, 2009; 136:185–188.
8.
ChenY, NoeKE, ThompsonS, et al.Evaluation of a revised U.S. Food and Drug Administration method for the detection of Cronobacter in powdered infant formula: A collaborative study. J Food Prot, 2012; 75:1144–1147.
9.
DebRoyC, RobertsE, KundratJ, et al.Detection of Escherichia coli serogroups O26 and O113 by PCR amplification of the wzx and wzy genes. Appl Environ Microbiol, 2004; 70:1830–1832.
10.
DennisonSK, MorrisJ. Multiresistant Enterobacter sakazakii wound infection in an adult. Infect Med, 2002; 19:533–535.
11.
[FAO/WHO] Food and Agriculture Organization of the United Nations/World Health Organization. Enterobacter sakazakii (Cronobacter spp.) in powdered follow-up formulae. Microbiological Risk Assessment Series No. 15. Rome: Food and Agriculture Organization of the United Nations/World Health Organization, 2008.
GosneyM. Enterobacter sakazakii bacteraemia with multiple splenic abscesses in a 75-year-old woman: A case report. Age Ageing, 2008; 37:236–237.
14.
HaririS, JosephS, ForsytheSJ. Cronobacter sakazakii ST4 strains and neonatal meningitis, United States. Emerg Infect Dis, 2013; 19:175.
15.
HealyB, CooneyS, O'BrienS, et al.Cronobacter(Enterobacter sakazakii): An opportunistic foodborne pathogen. Foodborne Pathog Dis, 2010; 7:339–350.
16.
HealyB, HuynhS, MullaneN, et al.Microarray-based comparative genomic indexing of the Cronobacter genus (Enterobacter sakazakii). Int J Food Microbiol, 2009; 136:159–164.
17.
HochelI, RuzickovaH, KrasnyL, et al.Occurrence of Cronobacter spp. in retail foods. J Appl Microbiol, 2012; 112:1257–1265.
18.
[ISO] International Organization for Standardization. Milk and milk products—Detection of Enterobacter sakazakii. 2006. ISO/TS22964: 2006 and IDF/RM 210:2006. Geneva, Switzerland: International Organization for Standardization, 2006.
19.
IversenC, ForsytheS. Isolation of Enterobacter sakazakii and other Enterobacteriaceae from powdered infant formula milk and related products. Food Microbiol, 2004; 21:771–777.
20.
IversenC, MullaneN, McCardellB, et al.Cronobacter gen. nov., a new genus to accommodate the biogroups of Enterobacter sakazakii, and proposal of Cronobacter sakazakii gen. nov., comb. nov., Cronobacter malonaticus sp. nov., Cronobacter turicensis sp. nov., Cronobacter muytjensii sp. nov., Cronobacter dublinensis sp. nov., Cronobacter genomospecies 1, and of three subspecies, Cronobacter dublinensis subsp. dublinensis subsp. nov., Cronobacter dublinensis subsp. lausannensis subsp. nov. and Cronobacter dublinensis subsp. lactaridi subsp. nov. Int J Syst Evol Microbiol, 2008; 58:1442–1447.
21.
JarvisKG, GrimCJ, FrancoAA, et al.Molecular characterization of Cronobacter lipopolysaccharide O-antigen gene clusters and development of serotype-specific PCR assays. Appl Environ Microbiol, 2011; 77:4017–4026.
22.
JosephS, CetinkayaE, DrahovskaH, et al.Cronobacter condimenti sp. nov., isolated from spiced meat, and Cronobacter universalis sp. nov., a species designation for Cronobacter sp. genomospecies 1, recovered from a leg infection, water and food ingredients. Int J Syst Evol Microbiol, 2012a;62:1277–1283.
23.
JosephS, ForsytheSJ. Predominance of Cronobacter sakazakii sequence type 4 in neonatal infections. Emerg Infect Dis, 2011; 17:1713–1715.
24.
JosephS, SonbolH, HaririS, et al.Diversity of the Cronobacter genus as revealed by multilocus sequence typing. J Clin Microbiol, 2012b;50:3031–3039.
25.
KandhaiM, HeuvelinkA, ReijM, et al.A study into the occurrence of Cronobacter spp. in the Netherlands between 2001 and 2005. Food Control, 2010; 21:1127–1136.
26.
KucerovaE, CliftonSW, XiaXQ, et al.Genome sequence of Cronobacter sakazakii BAA-894 and comparative genomic hybridization analysis with other Cronobacter species. PLoS ONE, 2010; 5:e9556.
27.
LehnerA, Fricker-FeerC, GschwendK, et al. Identification of Enterobacteriaceae and Cronobacter spp. in raw milk, milk concentrate and milk powder: Prevalence and genotyping. Archiv Lebensmittelhygiene, 2010; 5:22–26.
28.
MacLeanLL, PagottoF, FarberJM, et al. The structure of the O-antigen in the endotoxin of the emerging food pathogen Cronobacter (Enterobacter) muytjensii strain 3270. Carbohydr Res, 2009; 344:667–671.
29.
MullaneN, O'GaoraP, NallyJE, et al.Molecular analysis of the Enterobacter sakazakii O-antigen gene locus. Appl Environ Microbiol, 2008; 74:3783–3794.
30.
MullerA, StephanR, Fricker-FeerC, et al.Genetic diversity of Cronobacter sakazakii isolates collected from a Swiss infant formula production facility. J Food Prot, 2013; 76:883–887.
31.
MuytjensHL, ZanenHC, SonderkampHJ, et al. Analysis of eight cases of neonatal meningitis and sepsis due to Enterobacter sakazakii. J Clin Microbiol, 1983; 18:115–120.
32.
Nazarowec-WhiteM, FarberJM. Incidence, survival, and growth of Enterobacter sakazakii in infant formula. J Food Prot, 1997; 60:226–230.
33.
RayP, DasA, GautamV, et al. Enterobacter sakazakii in infants: Novel phenomenon in India. Indian J Med Microbiol, 2007; 25:408–410.
34.
StrydomA, CameronM, WitthuhnRC. Phylogenetic analysis of Cronobacter isolates based on the rpoA and 16S rRNA genes. Curr Microbiol, 2012; 64:251–258.
35.
SunY, WangM, LiuH, et al. Development of an O-antigen serotyping scheme for Cronobacter sakazakii. Appl Environ Microbiol, 2011; 77:2209–2214.
36.
SunY, WangM, WangQ, et al.Genetic analysis of the Cronobacter sakazakii O4 to O7 O-antigen gene clusters and development of a PCR assay for identification of all C. sakazakii O serotypes. Appl Environ Microbiol, 2012; 78:3966–3974.
37.
TamuraK, PetersonD, PetersonN, et al.MEGA5: Molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol, 2011; 28:2731–2739.
38.
ThompsonJD, GibsonTJ, PlewniakF, et al.The CLUSTAL_X windows interface: Flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res, 1997; 25:4876–4882.
39.
TsaiHY, LiaoCH, HuangYT, et al.Cronobacter infections not from infant formula, Taiwan. Emerg Infect Dis, 2013; 19:167–169.
40.
ZhouY, WuQ, XuX, et al.Development of an immobilization and detection method of Enterobacter sakazakii from powdered infant formula. Food Microbiol, 2008; 25:648–652.
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
