{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,27]],"date-time":"2026-05-27T13:49:07Z","timestamp":1779889747687,"version":"3.53.1"},"reference-count":33,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2024,9,29]],"date-time":"2024-09-29T00:00:00Z","timestamp":1727568000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Genes"],"abstract":"<jats:p>Cronobacter sakazakii is associated with the ingestion of contaminated reconstituted powdered infant formula (PIF), resulting in necrotizing enterocolitis, sepsis and meningitis in neonatal infants. Potential virulence determinants include the variable capsular polysaccharides; K-antigen and colanic acid (CA). Strains encoding for the capsule variant K2:CA2 have been strongly associated with neonatal meningitis cases. This study aimed to develop and apply a multiplex PCR assay to determine C. sakazakii K-antigen and colanic acid types. Twenty-six strains of C. sakazakii which had previously been isolated from food and environmental sources were used. These cover 18 multilocus sequence types and four serotypes. Based on our research findings, we have identified two K-antigen types present. Specifically, the K1-antigen was observed in sequence types ST1, ST8, ST20, ST23, ST64, ST198, ST263, ST264 and ST406, while the K2-antigen was present in ST4, ST9, ST12, ST13, ST136, ST233, ST245 and ST405. Additionally, we detected colanic acid (CA) type 1 in sequence types ST1, ST8, ST9, ST20, ST245 and ST405, and colanic acid (CA) type 2 in ST4, ST12, ST13, ST23, and ST64. We compared the predicted K-antigen and colanic acid types with the entire genome sequences of the strains. The comparison showed complete agreement between the PCR amplification results and the genomic analysis of the K-antigen and colanic acid-encoding regions. This assay is a useful tool for rapid identification of C. sakazakii, K-antigen and colanic acid types, in routine diagnoses and foodborne investigations. In addition, it will contribute to our knowledge of virulence factors associated with life-threatening neonatal meningitis.<\/jats:p>","DOI":"10.3390\/genes15101282","type":"journal-article","created":{"date-parts":[[2024,9,30]],"date-time":"2024-09-30T03:12:28Z","timestamp":1727665948000},"page":"1282","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Multiplex Determination of K-Antigen and Colanic Acid Capsule Variants of Cronobacter sakazakii"],"prefix":"10.3390","volume":"15","author":[{"given":"Khaled M.","family":"Ibrahim","sequence":"first","affiliation":[{"name":"Microbiology Department, Libyan Biotechnology Research Center, Tripoli P.O. Box 30313, Libya"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7668-9137","authenticated-orcid":false,"given":"Abdlrhman M.","family":"Alsonosi","sequence":"additional","affiliation":[{"name":"Microbiology Department, Faculty of Medicine, Sebah University, Sebha P.O. Box 1000, Libya"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mahmoud B.","family":"Agena","sequence":"additional","affiliation":[{"name":"Libyan Medical Research Center, Zawia P.O. Box 20311, Libya"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6372-5197","authenticated-orcid":false,"given":"Bassam A.","family":"Elgamoudi","sequence":"additional","affiliation":[{"name":"Institute for Biomedicine and Glycomics, Griffith University, Southport, QLD 4215, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6709-0712","authenticated-orcid":false,"given":"Stephen J.","family":"Forsythe","sequence":"additional","affiliation":[{"name":"Foodmicrobe.com Ltd., Nottingham NG12 5GY, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2024,9,29]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1979","DOI":"10.1007\/s10096-015-2440-8","article-title":"The speciation and genotyping of Cronobacter isolates from hospitalised patients","volume":"34","author":"Alsonosi","year":"2015","journal-title":"Eur. J. Clin. Microbiol. Infect. Dis."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"435","DOI":"10.1007\/s10482-018-1178-6","article-title":"Characterization of the pathogenicity of Clinical Cronobacter malonaticus strains based on the tissue culture investigations","volume":"112","author":"Alsonosi","year":"2019","journal-title":"Antonie Leeuwenhoek"},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Bla\u017ekov\u00e1, M., Javurkov\u00e1, B., Vlach, J., G\u00f6selov\u00e1, S., Karamonov\u00e1, L., Ogrodzki, P., Forsythe, S., and Fukal, L. (2015). Diversity of O antigens within the genus Cronobacter: From disorder to order. Appl. Environ. Microbiol., 81.","DOI":"10.1128\/AEM.00277-15"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1185","DOI":"10.3201\/eid1208.051509","article-title":"Invasive Enterobacter sakazakii disease in infants","volume":"12","author":"Bowen","year":"2006","journal-title":"Emerg. Infect. Dis."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"2111","DOI":"10.4315\/0362-028X-70.9.2111","article-title":"Dry stress and survival time of Enterobacter sakazakii and other Enterobacteriaceae","volume":"70","author":"Forsythe","year":"2007","journal-title":"J. Food Prot."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1044","DOI":"10.1111\/j.1365-2672.2010.04733.x","article-title":"Distribution, prevalence and persistence of Cronobacter (Enterobacter sakazakii) in the nonprocessing and processing environments of five milk powder factories","volume":"109","author":"Craven","year":"2010","journal-title":"J. Appl. Microbiol."},{"key":"ref_7","first-page":"1","article-title":"Identification of Klebsiella pneumoniae K1 and K2 capsular types by PCR and quellung test","volume":"9","author":"Feizabadi","year":"2013","journal-title":"Jundishapur J. Microbiol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1146\/annurev-food-030117-012246","article-title":"Updates on the Cronobacter genus","volume":"9","author":"Forsythe","year":"2018","journal-title":"Annu. Rev. Food Sci. Technol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"7661","DOI":"10.1128\/AEM.71.12.7661-7669.2005","article-title":"Performance of media for recovering stressed cells of Enterobacter sakazakii as determined using spiral plating and ecometric techniques","volume":"71","author":"Gurtler","year":"2005","journal-title":"Appl. Environ. Microbiol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"225","DOI":"10.1089\/fpd.2008.0175","article-title":"Temporal and spatial distribution of Cronobacter isolates in a milk powder processing plant determined by pulsed-field gel electrophoresis","volume":"6","author":"Hein","year":"2009","journal-title":"Foodborne Pathog."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"1713","DOI":"10.1038\/s41396-020-0628-0","article-title":"Diversity and evolution of surface polysaccharide synthesis loci in Enterobacteriales","volume":"14","author":"Holt","year":"2020","journal-title":"ISME J."},{"key":"ref_12","first-page":"69","article-title":"Epidemiology of Cronobacter spp. isolates from patients admitted to the Olomouc University Hospital (Czech Republic)","volume":"63","author":"Holy","year":"2014","journal-title":"Epidemiol. Mikrobiol. Imunol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"169","DOI":"10.1016\/j.jhin.2013.09.011","article-title":"Cronobacter spp. as emerging causes of healthcare-associated infection","volume":"86","author":"Holy","year":"2014","journal-title":"J. Hosp. Infect."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"378","DOI":"10.1111\/j.1472-765X.2004.01507.x","article-title":"The growth profile, thermotolerance and biofilm formation of Enterobacter sakazakii grown in infant formula milk","volume":"38","author":"Iversen","year":"2004","journal-title":"Lett. Appl. Microbiol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"771","DOI":"10.1016\/j.fm.2004.01.009","article-title":"Isolation of Enterobacter sakazakii and other Enterobacteriaceae from powdered infant formula milk and related products","volume":"21","author":"Iversen","year":"2004","journal-title":"Food Microbiol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"3801","DOI":"10.3168\/jds.2011-4318","article-title":"Reservoir and routes of transmission of Enterobacter sakazakii (Cronobacter spp.) in a milk powder-producing plant","volume":"94","author":"Jacobs","year":"2011","journal-title":"J. Dairy Sci."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Joseph, S., Desai, P., Ji, Y., Cummings, C.A., Shih, R., Degoricija, L., Rico, A., Brzoska, P., Hamby, S., and Masood, N. (2012). Comparative analysis of genome sequences covering the seven Cronobacter species. PLoS ONE, 16.","DOI":"10.1371\/journal.pone.0049455"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"419","DOI":"10.1007\/s12223-014-0315-5","article-title":"Detection of K1 antigen of Escherichia coli rods isolated from pregnant women and neonates","volume":"5","author":"Kaczmarek","year":"2014","journal-title":"Folia Microbiol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"5913","DOI":"10.1128\/AEM.00745-08","article-title":"Dissemination of Cronobacter spp. (Enterobacter sakazakii) in a powdered milk protein manufacturing facility","volume":"74","author":"Mullane","year":"2008","journal-title":"Appl. Environ."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"276","DOI":"10.1111\/1750-3841.15577","article-title":"Advances in our understanding and distribution of the Cronobacter genus in China","volume":"86","author":"Ling","year":"2021","journal-title":"J. Food Sci."},{"key":"ref_21","first-page":"721","article-title":"Temperature has reciprocal effects on colanic acid and polysialic acid biosynthesis in E. Coli K92","volume":"4","author":"Navasa","year":"2009","journal-title":"Appl. Microbiol."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Navasa, N., Rodr\u00edguez-Aparicio, L., Ferrero, M., Monteagudo-Mera, A., and Mart\u00ednez-Blanco, H. (2013). Polysialic and colanic acids metabolism in Escherichia coli K92 is regulated by RcsA and RcsB. Biosci. Rep., 3.","DOI":"10.1042\/BSR20130018"},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Ogrodzki, P., and Forsythe, S. (2015). Capsular profiling of the Cronobacter genus and the association of specific Cronobacter sakazakii and C. Malonaticus capsule types with neonatal meningitis and necrotizing enterocolitis. BMC Genom., 16.","DOI":"10.1186\/s12864-015-1960-z"},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Ogrodzki, P., and Forsythe, S.J. (2017). DNA-sequence based typing of the Cronobacter genus using MLST, CRISPR-cas array and capsular profiling. Front. Microbiol., 8.","DOI":"10.3389\/fmicb.2017.01875"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1520","DOI":"10.3201\/eid2009.140545","article-title":"Incidence of Cronobacter spp. infections, United States, 2003\u20132009","volume":"20","author":"Patrick","year":"2014","journal-title":"Emerg. Infect. Dis."},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Qin, X., Wang, H., Chao, M., Yang, X., Zhang, Y., Feng, J., Forsythe, S.J., Man, C., and Jiang, Y. (J. Dairy Sci., 2021). Stress resistance of Cronobacter spp. affecting its control during food production, J. Dairy Sci., in print.","DOI":"10.3168\/jds.2021-20591"},{"key":"ref_27","first-page":"323","article-title":"Structure and genetics of the O-antigen of Cronobacter turicensis G3882 from a new serotype, C. turicensis O2, and identification of a serotype-specific gene","volume":"66","author":"Sun","year":"2012","journal-title":"FEMS Microbiol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"247","DOI":"10.1111\/j.1574-6968.2008.01208.x","article-title":"Evaluation of a multiplex PCR for detection of serotypes K1, K2 and K5 in Klebsiella sp. and comparison of isolates within these serotypes","volume":"2","author":"Turton","year":"2008","journal-title":"FEMS Microbiol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"293","DOI":"10.1128\/JCM.39.1.293-297.2001","article-title":"Outbreak of necrotising enterocolitis associated with Enterobacter sakazakii in powdered milk formula","volume":"39","author":"Smet","year":"2001","journal-title":"J. Clin. Microbiol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2491","DOI":"10.1016\/j.carres.2003.08.010","article-title":"Biosynthesis and assembly of group 1 capsular polysaccharides in Escherichia coli and related extracellular polysaccharides in other bacteria","volume":"23","author":"Whitfield","year":"2003","journal-title":"Carbohydr. Res."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"39","DOI":"10.1146\/annurev.biochem.75.103004.142545","article-title":"Biosynthesis and assembly of capsular polysaccharides in Escherichia coli","volume":"75","author":"Whitfield","year":"2006","journal-title":"Annu. Rev. Biochem."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"35","DOI":"10.1016\/j.carres.2013.05.007","article-title":"Structure, biosynthesis, and function of bacterial capsular polysaccharides synthesized by ABC transporter-dependent pathways","volume":"378","author":"Willis","year":"2013","journal-title":"Carbohydr. Res."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Wijetunge, S., Gongati, S., DebRoy, C., Kim, S., Couraud, O., Romero, A., Weksler, B., and Kariyawasam, S. (2015). Characterizing the pathotype of neonatal meningitis causing Escherichia coli (NMEC). BMC Microbiol., 15.","DOI":"10.1186\/s12866-015-0547-9"}],"container-title":["Genes"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4425\/15\/10\/1282\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T16:07:06Z","timestamp":1760112426000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4425\/15\/10\/1282"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,9,29]]},"references-count":33,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2024,10]]}},"alternative-id":["genes15101282"],"URL":"https:\/\/doi.org\/10.3390\/genes15101282","relation":{},"ISSN":["2073-4425"],"issn-type":[{"value":"2073-4425","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,9,29]]}}}