Virulence Characterization of Salmonella enterica by a New Microarray: Detection and Evaluation of the Cytolethal Distending Toxin Gene Activity in the Unusual Host S. Typhimurium

Abstract

Salmonella enterica is a zoonotic foodborne pathogen that causes acute gastroenteritis in humans. We assessed the virulence potential of one-hundred and six Salmonella strains isolated from food animals and products. A high through-put virulence genes microarray demonstrated Salmonella Pathogenicity Islands (SPI) and adherence genes were highly conserved, while prophages and virulence plasmid genes were variably present. Isolates were grouped by serotype, and virulence plasmids separated S. Typhimurium in two clusters. Atypical microarray results lead to whole genome sequencing (WGS) of S. Infantis Sal147, which identified deletion of thirty-eight SPI-1 genes. Sal147 was unable to invade HeLa cells and showed reduced mortality in Galleria mellonella infection model, in comparison to a SPI-1 harbouring S. Infantis. Microarray and WGS of S. Typhimurium Sal199, established for the first time in S. Typhimurium presence of cdtB and other Typhi-related genes. Characterization of Sal199 showed cdtB genes were upstream of transposase IS911, and co-expressed with other Typhi-related genes. Cell cycle arrest, cytoplasmic distension, and nuclear enlargement were detected in HeLa cells infected by Sal199, but not with S. Typhimurium LT2. Increased mortality of Galleria was detected on infection with Sal199 compared to LT2. Thus, Salmonella isolates were rapidly characterized using a high through-put microarray; helping to identify unusual virulence features which were corroborated by further characterisation. This work demonstrates that the use of suitable screening methods for Salmonella virulence can help assess the potential risk associated with certain Salmonella to humans. Incorporation of such methodology into surveillance could help reduce the risk of emergence of epidemic Salmonella strains.

Identification of Genetic Features for Attenuation of Two Salmonella Enteritidis Vaccine Strains and Differentiation of These From Wildtype Isolates Using Whole Genome Sequencing

“…As for AviPro SALMONELLA VAC E, the attenuation may also be the result of multiple factors: e.g., the missing envR gene ( Table 2) and point mutations in 20 virulence-associated genes (Table 4). EnvR as a potential candidate for attenuation (Row 13, Table 2) is a regulator in the Salmonella pathogenicity island 2 (SPI2) that encodes type III secretion system (T3SS) that changes the host cell functions and facilitate intracellular replication (65,66). The RpoB H526D mutant (Rif) displayed reduced survival compared with control strains in Mycobacterium tuberculosis (36).…”

Microarray Evaluation of Antimicrobial Resistance and Virulence of Escherichia coli Isolates from Portuguese Poultry

“…The E. coli isolates were cultured overnight on Luria Bertani agar plates at 37 °C and DNA extracted by lysis as described previously [ 10 , 28 ]. A microarray was then used to screen each DNA sample for the presence of 88 genes encoding resistance determinants [ 10 ] and 60 genes encoding virulence factors [ 29 ].…”

“…Among the virulence array there were genes encoding to several factors, like increased serum survival, siderophere receptors, bacteriocins, toxins, fimbria and other adhesins, among others. The DNA was labeled as described previously [ 10 , 28 , 29 ], and hybridized to the microarray using the HybPlus Kit (Alere Technologies, Jena, Germany), according to the manufacturer's instructions. Microarray analysis was performed according Figueiredo et al [ 28 ].…”

The Cytolethal Distending Toxin Contributes to Microbial Virulence and Disease Pathogenesis by Acting As a Tri-Perditious Toxin

“…In contrast to all other bacteria expressing Cdt, Salmonella enterica serotype Typhi ( S . Typhi) lack genes encoding CdtA and CdtC (Haghjoo and Galán, 2004; Skyberg et al, 2006; Suez et al, 2013; Mezal et al, 2014; Figueiredo et al, 2015). …”