Project description:The genus Cronobacter (formerly called Enterobacter sakazakii) is composed of five species; C. sakazakii, C. malonaticus, C. turicensis, C. muytjensii, and C. dublinensis. The genus includes opportunistic human pathogens, and the first three species have been associated with neonatal infections. The most severe diseases are caused in neonates and include fatal necrotizing enterocolitis and meningitis. The genetic basis of the diversity within the genus is unknown, and few virulence traits have been identified. We report here the first sequence of a member of this genus, C. sakazakii strain BAA-894. The genome of Cronobacter sakazakii strain BAA-894 comprises a 4.4 Mb chromosome (57% GC content) and two plasmids; 31 Kb (51% GC) and 131 Kb (56% GC). The genome was used to construct a 385,000 probe oligonucleotide tiling DNA microarray covering the whole genome. Comparative genomic hybridization (CGH) was undertaken on five other C. sakazakii strains, and representatives of the four other Cronobacter species. Among 4,382 annotated genes inspected in this study, about 55% of genes were common to all C. sakazakii strains and 43% were common to all Cronobacter strains, with 10 - 17% absence of genes. CGH highlighted 15 clusters of genes in C. sakazakii BAA-894 that were divergent or absent in more than half of the tested strains; six of these are of probable prophage origin. Putative virulence factors were identified in these prophage and in other variable regions. A number of genes unique to Cronobacter species associated with neonatal infections (C. sakazakii, C. malonaticus and C. turicensis) were identified. These included a copper and silver resistance system known to be linked to invasion of the blood-brain barrier by neonatal meningitic strains of Escherichia coli. In addition, genes encoding for multidrug efflux pumps and adhesins were identified that were unique to C. sakazakii strains from outbreaks in neonatal intensive care units. Comparative genomic hybridization highlighted 15 clusters of genes in C. sakazakii BAA-894 that were divergent or absent in more than half of the tested strains; six of these are of probable prophage origin. Putative virulence factors were identified in these prophage and in other variable regions. A number of genes unique to Cronobacter species associated with neonatal infections (C. sakazakii, C. malonaticus and C. turicensis) were identified. These included a copper and silver resistance system known to be linked to invasion of the blood-brain barrier by neonatal meningitic strains of Escherichia coli. In addition, genes encoding for multidrug efflux pumps and adhesins were identified that were unique to C. sakazakii strains from outbreaks in neonatal intensive care units. Ten Cronobacter samples were analyzed, including total genomic DNA of six C. sakazakii strains, one C. malonaticus strain, one C. muytjensii strain, one C. dublinensis strain and one C. turicensis strain.

Project description:The genus Cronobacter (formerly called Enterobacter sakazakii) is composed of five species; C. sakazakii, C. malonaticus, C. turicensis, C. muytjensii, and C. dublinensis. The genus includes opportunistic human pathogens, and the first three species have been associated with neonatal infections. The most severe diseases are caused in neonates and include fatal necrotizing enterocolitis and meningitis. The genetic basis of the diversity within the genus is unknown, and few virulence traits have been identified. We report here the first sequence of a member of this genus, C. sakazakii strain BAA-894. The genome of Cronobacter sakazakii strain BAA-894 comprises a 4.4 Mb chromosome (57% GC content) and two plasmids; 31 Kb (51% GC) and 131 Kb (56% GC). The genome was used to construct a 385,000 probe oligonucleotide tiling DNA microarray covering the whole genome. Comparative genomic hybridization (CGH) was undertaken on five other C. sakazakii strains, and representatives of the four other Cronobacter species. Among 4,382 annotated genes inspected in this study, about 55% of genes were common to all C. sakazakii strains and 43% were common to all Cronobacter strains, with 10 - 17% absence of genes. CGH highlighted 15 clusters of genes in C. sakazakii BAA-894 that were divergent or absent in more than half of the tested strains; six of these are of probable prophage origin. Putative virulence factors were identified in these prophage and in other variable regions. A number of genes unique to Cronobacter species associated with neonatal infections (C. sakazakii, C. malonaticus and C. turicensis) were identified. These included a copper and silver resistance system known to be linked to invasion of the blood-brain barrier by neonatal meningitic strains of Escherichia coli. In addition, genes encoding for multidrug efflux pumps and adhesins were identified that were unique to C. sakazakii strains from outbreaks in neonatal intensive care units. Comparative genomic hybridization highlighted 15 clusters of genes in C. sakazakii BAA-894 that were divergent or absent in more than half of the tested strains; six of these are of probable prophage origin. Putative virulence factors were identified in these prophage and in other variable regions. A number of genes unique to Cronobacter species associated with neonatal infections (C. sakazakii, C. malonaticus and C. turicensis) were identified. These included a copper and silver resistance system known to be linked to invasion of the blood-brain barrier by neonatal meningitic strains of Escherichia coli. In addition, genes encoding for multidrug efflux pumps and adhesins were identified that were unique to C. sakazakii strains from outbreaks in neonatal intensive care units.

Project description:BackgroundThe Cronobacter genus is composed of seven species, and can cause infections in all age groups. Of particular concern is C. sakazakii, as this species is strongly associated with severe and often fatal cases of necrotizing enterocolitis and meningitis in neonates and infants. Whole genome sequencing has revealed that the nanAKT gene cluster required for the utilisation of exogenous sialic acid is unique to the C. sakazakii species (ESA_03609-13).Sialic acid is found in breast milk, infant formula, intestinal mucin, and gangliosides in the brain, hence its metabolism by C. sakazakii is of particular interest. Therefore its metabolism could be an important virulence factor. To date, no laboratory studies demonstrating the growth of C. sakazakii on sialic acid have been published nor have there been reports of sialidase activity. The phylogenetic analysis of the nan genes is of interest to determine whether the genes have been acquired by horizontal gene transfer.ResultsPhylogenetic analysis of 19 Cronobacter strains from 7 recognised species revealed the nanAKTR genes formed a unique cluster, separate from other Enterobacteriaceae such as E. coli K1 and Citrobacter koseri, which are also associated with neonatal meningitis. The gene organisation was similar to Edwardsiella tarda in that nanE gene (N-acetylmannosamine-6-phosphate-2epimerase) was not located within the nanATK cluster. Laboratory studies confirmed that only C. sakazakii, and not the other six Cronobacter species, was able to use sialic acid as a carbon source for growth. Although the ganglioside GM1 was also used as carbon source, no candidate sialidase genes were found in the genome, instead the substrate degradation is probably due to β-galactosidase activity.ConclusionsGiven the relatively recent evolution of both C. sakazakii (15-23 million years ago) and sialic acid synthesis in vertebrates, sialic acid utilization may be an example of co-evolution by one species of the Cronobacter genus with the mammalian host. This has possibly resulted in additional virulence factors contributing to severe life-threatening infections in neonates due to the utilization of sialic acid from breast milk, infant formula, milk (oligosaccharides), mucins lining the intestinal wall, and even gangliosides in the brain after passing through the blood-brain barrier.

Dataset Information

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.

Publications

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.

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