Emily E. Jackson

Genotypic and phenotypic characteristics of Cronobacter species, with particular attention to the newly reclassified species Cronobacter helveticus, Cronobacter pulveris, and Cronobacter zurichensis.

In 2013, Enterobacter helveticus, Enterobacter pulveris and Enterobacter turicensis, were reclassified as Cronobacter helveticus, Cronobacter pulveris and Cronobacter zurichensis, respectively. Previously these species had been used as negative controls for some Cronobacter detection assays. This study examined cultural, biochemical and molecular Cronobacter detection and identification assays, with emphasis on the new species. Additionally, 32 Cronobacter genomes were examined for the presence of PCR target genes using the BLAST function of the online Cronobacter PubMLST facility. The results of the cultural methods varied and no single medium was able to correctly detect all Cronobacter spp. Since the supporting databases have not been updated to include the Cronobacter genus, Enterobacter sakazakii was returned for four strains of the newly reclassified species with ID32E and none with API 20E. PCR probes targeting rpoB and ompA could not correctly identify the new Cronobacter spp., due to primer specificity or absent target genes. As neonates have been identified as a high-risk group for infection, international standards require the absence of all Cronobacter species in powdered infant formula. However, many conventional detection methods cannot correctly identify the newly recognized species. Conversely, DNA sequence-based methods can adapt to taxonomic revisions and will likely become more common.

Reevaluation of a Suspected Cronobacter sakazakii Outbreak in Mexico.

In 2010, two infants became ill at a hospital in Mexico. Subsequently, a range of clinical, environmental, and powdered and rehydrated infant formula isolates were identified by using a combination of phenotyping and PCR probes. The strains were clustered according to pulsed-field gel electrophoresis. The causative agent was reported as Cronobacter sakazakii, with powdered infant formula (PIF) identified as the likely source of the infections. This new study further characterized the isolates from this outbreak by using multilocus sequence typing and whole genome sequencing of selected strains. Though four PIF isolates and one hospital environmental isolate were identified as C. sakazakii sequence type 297 by multilocus sequence typing, they were isolated 6 months prior to the outbreak. Genotypic analyses of patient isolates identified them as Enterobacter hormaechei and Enterobacter spp. The pulsed-field gel electrophoresis profile of the Enterobacter spp. isolates matched those of isolates from previously unopened tins of PIF. E. hormaechei was only isolated from the two infants and not PIF. The reevaluation of this outbreak highlights the need for accurate detection and identification assays, particularly during outbreak investigations in which incorrect identifications may mislead the investigation and attribution of the source. Though the species responsible for the symptoms could not be determined, this outbreak demonstrated the possible transmission of Enterobacter spp. from PIF to infants. These are possibly the first reported cases of Enterobacter spp. infection of infants from bacterial-contaminated PIF.

Description of Siccibacter colletis sp. nov., a novel species isolated from plant material, and emended description of Siccibacter turicensis

A re-evaluation of the taxonomic position of two strains, 1383(T) and 2249, isolated from poppy seeds and tea leaves, which had been identified as Siccibacter turicensis (formerly Cronobacter zurichensis ), was carried out. The analysis included phenotypic characterization, 16S rRNA gene sequencing, multilocus sequence analysis (MLSA) of five housekeeping genes (atpD, fusA, glnS, gyrB and infB; 2034 bp) and ribosomal MLSA (53 loci; 22 511 bp). 16S rRNA gene sequence analysis and MLSA showed that the strains formed an independent phylogenetic lineage, with Siccibacter turicensis LMG 23730(T) as the closest neighbour. Average nucleotide identity analysis and phenotypic analysis confirmed that these strains represent a novel species, for which the name Siccibacter colletis sp. nov. is proposed. The type strain is 1383(T) ( = NCTC 14934(T) = CECT 8567(T) = LMG 28204(T)). An emended description of Siccibacter turicensis is also provided.

Draft Genome Sequence of “Candidatus Cronobacter colletis” NCTC 14934T, a New Species in the Genus Cronobacter

ABSTRACT Members of the Cronobacter genus are associated with serious infections in neonates. This is the first report of the draft genome sequence for the newly proposed species Cronobacter colletis.

Riesgo de contaminación por Cronobacter Sakazakii en leches en polvo para la nutrición de lactantes

Cronobacter spp es un genero bacteriano con 7 especies, siendo C. sakazakii la especie clinica mas reportada y asociada a meningitis y septicemia en lactantes. Es transmitida por leche en polvo (LP) por lo que la OMS recomienda su ausencia en este producto. En Chile, el reglamento sanitario de los alimentos (RSA) no lo considera. Objetivo: Evaluar el riesgo por Cronobacter sakazakii en LP destinadas al consumo de lactantes. Metodologia: Se analizaron 72 muestras de LP de 3 marcas y 3 paises. El recuento de bacterias mesofilas (RAM), Enterobacteriaceae (ENT) y numero mas probable (NMP) se realizo con la metodologia de Puch and Ito (2001). Se utilizo agar diferencial Cronobacter para aislamiento (DFI, Oxoid, England) y kit bioquimico ID32E (Biomeriux, Francia) para fenotipo. El patogeno fue identificado y genotipificado por multilocus sequence typing (MLST) utilizando criterios de http://www.pubmlst.org/cronobacter. Resultados: La medianas de RAM para LP etapa 1 y prematuros fueron 300 UFC/g (10-36 000) y 650 UFC/g (70-30 000), siendo mayor en las muestras de Chile (p=0,016). Para ENT de 75 UFC/g (10-1 060) y 200 UFC/g (30-1 000), no existiendo diferencias significativas por tipo, pais o marca de LP (p>0,05). Dos cepas de 2 lotes diferentes caracteristicas en agar DFI se identificaron como C. sakazaki con 0,23 y 2,3 NMP/g. Ademas de Franconibacter helveticus en otras 2 cepas, especie relacionada estrechamente con Cronobacter spp. Conclusiones: La prevalencia de C. sakazakii en todas las muestras fue de 2,7% y aislo solo en LP de elaborados en Chile. La ausencia de Cronobacter spp en 25 g debe ser incorporado en el RSA de Chile

Draft Genome Sequence of an Enterobacter Species Associated with Illnesses and Powdered Infant Formula

ABSTRACT This is the first report of the draft genome sequence of an Enterobacter species that may have been transmitted from powdered infant formula (PIF) to infants, resulting in illness. Enterobacter spp. are currently permitted in PIF, but the transmission of this strain indicates that the microbiological criteria for PIF may need revision.