A proposed harmonized LPS molecular-subtyping scheme for Cronobacter species

Yan, Qiongqiong; Jarvis, Karen G.; Chase, Hannah R.; Hébert, Karine; Trach, Larisa H.; Lee, Chloe; Sadowski, J. A.; Lee, Boram; Hwang, Seongeun; Sathyamoorthy, Venugopal; Mullane, Niall; Pava‐Ripoll, Monica; Iversen, Carol; Pagotto, Franco; Fanning, Séamus; Tall, Ben D.

doi:10.1016/j.fm.2015.03.003

Cited by 22 publications

(27 citation statements)

References 36 publications

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“…sakazakii is an opportunistic pathogen causing life-threatening infections in neonates. The pathogen can be isolated from clinical, environmental, and a wide range of food sources, predominantly from PIF and its associated production environments (21,29,45,46).…”

Section: Discussionmentioning

confidence: 99%

“…To identify Cronobacter strains at the genus, species, and serotype levels, the proposed schema for Cronobacter spp. (29) was applied using molecular detection techniques: an alpha-glucosidase-based PCR assay was performed to confirm the bacterial genus (37), and the species of the Cronobacter strains were identified using rpoB and cgcA as the gene targets for the PCR amplification (7,38). Only strains that were confirmed to be C. sakazakii were then subjected to multiplex PCR to identify the serotype.…”

Section: Methodsmentioning

confidence: 99%

“…Due to the recently described limitation of PCR-based methods for the determination of the different variations of genomic O-antigen gene clusters of C. sakazakii strains (22,30), it has been proposed that C. sakazakii serotypes can reliably be determined only after the strains have been identified as members of the genus Cronobacter and species sakazakii (29). Therefore, the genus and species of a collection of 76 strains that were previously identified as C. sakazakii at our institute by classical culture methods, according to the ISO standard in reference 25, were verified using a PCR method (7,37,38).…”

Section: Identification Of C Sakazakii Serotypesmentioning

confidence: 99%

“…In light of the need for a reliable and inexpensive rapid detection method, several PCRbased protocols for the identification of C. sakazakii at the genus, species, and serotype levels have been established (28). Despite their rapidity, some molecular methods lack specificity, depending on the chosen primers (22,29). The recent reassignment of C. sakazakii serotypes O5 and O6 to C. malonaticus highlights the unreliability of PCR-based serotyping for the detection of all sequence-based variations of the O antigen of C. sakazakii (21,22,30).…”

mentioning

confidence: 99%

“…or C. sakazakii, regardless of the serotype (31)(32)(33). Due to the lack of availability of high-affinity MAbs, until now, there has been no rapid detection method available for the individual serotypes of C. sakazakii (29). The objective of this study was to develop highly sensitive MAbs that are reactive with C. sakazakii serotypes O1, O2, and O3 in order to establish sandwich EIAs for the specific detection and identification of these serotypes.…”

mentioning

confidence: 99%

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Simultaneous Rapid Detection and Serotyping of Cronobacter sakazakii Serotypes O1, O2, and O3 by Using Specific Monoclonal Antibodies

Scharinger

Dietrich

Kleinsteuber

et al. 2016

Appl Environ Microbiol

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Cronobacter sakazakii is a foodborne pathogen associated with rare but often lethal infections in neonates. Powdered infant formula (PIF) represents the most frequent source of infection. Out of the identified serotypes (O1 to O7), O1, O2, and O3 are often isolated from clinical and PIF samples. Serotype-specific monoclonal antibodies (MAbs) suitable for application in enzyme immunoassays (EIAs) for the rapid detection of C. sakazakii have not yet been developed. In this study, we created specific MAbs with the ability to bind to C. sakazakii of serotypes O1, O2, and O3. Characterization by indirect EIAs, immunofluorescence, motility assays, and immunoblotting identified lipopolysaccharide (LPS) and exopolysaccharide (EPS) as the antigenic determinants of the MAbs. The established sandwich EIAs were highly sensitive and were able to detect between 2 ؋ 10 3 and 9 ؋ 10 6 CFU/ml. Inclusivity tests confirmed that 93% of serotype O1 strains, 100% of O2 strains, and 87% of O3 strains were detected at low cell counts. No cross-reactivity with >100 strains of Cronobacter spp. and other Enterobacteriaceae was observed, except for that with C. sakazakii serotype O3 and Cronobacter muytjensii serotype O1. Moreover, the sandwich EIAs detected C. sakazakii in PIF samples artificially contaminated with 1 to 10 bacterial cells per 10 g of sample after 15 h of preenrichment. The use of these serotype-specific MAbs not only allows the reliable detection of C. sakazakii strains but also enables simultaneous serotyping in a simple sandwich EIA method. Cronobacter spp. are Gram-negative opportunistic foodborne pathogens of the family Enterobacteriaceae that cause rare but severe infections in patients of all age groups. In adults, Cronobacter spp. are often associated with nosocomial infections, including pneumonia, septicemia, wound infections, and osteomyelitis, while causing invasive disease in young infants and neonates (1-4). Among the seven identified Cronobacter species, C. sakazakii, C. malonaticus, C. muytjensii, C. turicensis, C. dublinensis, C. condimenti, and C. universalis (5-8), C. sakazakii plays a prominent role due to it causing life-threatening infections in neonates (9-11). Clinically manifested infections present as necrotizing enterocolitis, sepsis, and meningitis, with a mortality rate as high as 80% (1, 12, 13). Although C. sakazakii has been isolated from a variety of different plant-and animal-based food products (14, 15), the presence in powdered infant formula (PIF) seems crucial in the infection of neonates (9, 12, 16). According to an established O-antigen serotyping scheme based on rabbit antisera and a PCRbased serotyping method (17-21), seven serotypes (O1 to O7) have been identified for C. sakazakii. Serotypes O1 and O2 seem to be most prevalent in PIF samples and in clinical cases, whereas serotype O3 has been isolated quite frequently from PIF but not as often from clinical cases (19,(22)(23)(24).Today, the contamination of PIF by C. sakazakii is being detected using conventional microbiological m...

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Section: Discussionmentioning

confidence: 99%