Identification of Shiga-Toxin-Producing Shigella Infections in Travel and Non-Travel Related Cases in Alberta, Canada

It has long been accepted that Shiga toxin (Stx) only exists in Shigella dysenteriae serotype 1. However, in recent decades, the presence of Shiga toxin genes (stx) in other Shigella spp. have been reported. We screened 366 Shigella flexneri strains from Alberta, Canada (2003 to 2016) for stx and 26 positive strains were identified. These isolates are highly related with the majority originating from the Dominican Republic and three isolates with Haiti origin. Both phylogenetic and spanning tree analysis of the 26 Alberta and 29 stx positive S. flexneri originating from the U.S., France, Canada (Quebec) and Haiti suggests that there are geographic specific distribution patterns (Haiti and Dominican Republic clades). This study provides the first comprehensive whole genome based phylogenetic analysis of stx positive S. flexneri strains as well as their global transmission, which signify the public health risks of global spreading of these strains.

PREVALENCE OF ENTEROHAEMORRHAGIC ESCHERICHIA COLI O157:H7, SHIGA TOXIN GENES AND ANTIBIOTIC RESISTANCE IN CHILDREN AGED 0-5 YEARS WITH DIARRHOEA IN IBADAN, OYO STATE

Diarrhoea is a predominant cause of childhood illness and death, Enterohaemorrhagic Escherichia coli (EHEC) have increasingly been recognised as an important cause of diarrhea all over the world. This study investigates the prevalence, presence of shiga toxin (Stx) and antibiotic resistance of Enterohaemorrhagic Escherichia coli (EHEC) O157:H7 in children with diarrhoea. Stool samples collected from children less than five year with diarrhoea were cultured, bacteria isolated were identified and antibiotics susceptibility testing was performed using standard methods. Entrohaemorrhagic E. coli O157:H7 and Stx were screened using serology and Polymerase Chain Reaction respectively. Structured questionnaires were administered to determine factors that predispose the children to diarrhoea. A total of 301/370 (81.4%) stool samples yielded bacterial growth, 261/301 (86.7%) were gram negative bacilli, 188/261 (72.0%) Escherichia coli, 163/188 (86.7%) were EHEC O157: H7. Out of the 5 EHEC O157:H7 only 2 possessed Stx genes, 1 have Stx 2 while the other have both Stx 1 and Stx 2 gene. Escherichia coli were resistant to Tetracycline 98.4%, Ampicillin 83.0%, Cefuroxime 76.5%, Augmentin 62.9% and Gentamycin 51.4%, all the EHEC O157:H7 were resistant to Tetracycline and Ampicillin. Diarrhoea in the children were significantly associated with hand wash after toilet, eating pastries, sources of drinking water and the educational level of parent/caretaker (p-values = 0.04, 0.00, 0.00 and 0.03 respectively). The presence of EHEC O157:H7 carrying Stx 1 and Stx 2 gene as well as resistant to all antibiotics tested is a pointer for more stringent and better screening for diarrhoea infections in children.

Genetic Characterization of Escherichia coli O157 Isolated From Human Stool Specimens in Wassit Province of Iraq

The goal of this study was to assess the severity of diarrhea caused by Shiga toxins produced by Escherichia coli ( STEC) isolated from human infections in Wassit province of Iraq. Stool specimens from (161) sporadic cases of diarrheal Patients with the mean age of (20 years), and range from 1 month to 75 years, were collected in AL-Karamma Teaching Hospital between March to July 2019. Then they were processed by culture, microscopic tests and VITEK which were used for the identification as E. coli. PCR was performed for detecting shiga toxin genes in E. coli isolates, and rfb gene (encoding O-antigene ) of STEC- O157. DNA sequencing was done on some positive isolates. The results of PCR detected stx genes in 19 (12 %) culture isolates of E. coli isolated from human diarrheal specimens. While 9 cases were positive for stx genes and have rfb gene. DNA sequences that depend on the sequence of the vtx2 gene have shown a highly homologous sequencing identity with NCBI-Blast Escherichia coli strain( O157:H7) isolates from humans and animals. The phylogenetic study revealed a clear genetic relationship between human and animal E. coli isolates and then gene sequencing was deposited with accession number into NCBI-Genbank (0MN944014.1). In conclusion, prevalence of E. coli O157 in humans remains undiagnosed because there are no traditional O157 detection methods in all our hospitals. Our study showed that the use of molecular methods in the detection can be used to detect STECs which cannot be detected using routine methods.

The prevalence and genomic context of Shiga toxin 2a genes in E. coli found in cattle

Shiga toxin-producing Escherichia coli (STEC) that cause severe disease predominantly carry the toxin gene variant stx2a. However, the role of Shiga toxin in the ruminant reservoirs of this zoonotic pathogen is poorly understood and strains that cause severe disease in humans (HUSEC) likely constitute a small and atypical subset of the overall STEC flora. The aim of this study was to investigate the presence of stx2a in samples from cattle and to isolate and characterize stx2a-positive E. coli. In nationwide surveys in Sweden and Norway samples were collected from individual cattle or from cattle herds, respectively. Samples were tested for Shiga toxin genes by real-time PCR and amplicon sequencing and stx2a-positive isolates were whole genome sequenced. Among faecal samples from Sweden, stx1 was detected in 37%, stx2 in 53% and stx2a in 5% and in skin samples in 64%, 79% and 2% respectively. In Norway, 79% of the herds were positive for stx1, 93% for stx2 and 17% for stx2a. Based on amplicon sequencing the most common stx2 types in samples from Swedish cattle were stx2a and stx2d. Multilocus sequence typing (MLST) of 39 stx2a-positive isolates collected from both countries revealed substantial diversity with 19 different sequence types. Only a few classical LEE-positive HUSEC were found among the stx2a-positive isolates, notably a single O121:H19 and an O26:H11. Known LEE-negative HUSEC lineages were also recovered including O113:H21 (ST-223), O130:H11 (ST-297), and O101:H33 (ST-330). We conclude that E. coli encoding stx2a in cattle are ranging from well-known HUSEC to unknown STEC variants. Comparison of isolates from human HUS cases to related STEC from the ruminant reservoirs can help identify combinations of virulence attributes necessary to cause HUS, as well as provide a better understanding of the routes of infection for rare and emerging pathogenic STEC.

Isolation and Culture Dependent Characterization of Escherichia coli from the Sewage Waste Water of Lahore, Pakistan

Aims: Infectious disease haemorrhagic uremic syndrome (HUS), present in sewage water, its cross contamination with drinking water may affect the community. This study was conducted to isolate the pathogenic strain of E. coli O157 from the sewage waste water of Lahore, Pakistan. The aim of this study was to evaluate biofilm formation and antibiotic susceptibility pattern for the isolated strains. Selective Hichrome EC O157 agar media was used for the isolation of E. coli. Methodology: Taxonomic status of strain was confirmed by 16S rRNA gene sequencing. Serotyping of E. coli O157 strains was accomplished by ProlexTM E. coli O157 Latex Test Reagent Kit results for O157. Bacterial strains were also evaluated for biofilm formation, toxin related genes (stx1, stx2, stx2c, stx2d) and antibiotic sensitivity. Results: Bacterial strains showed resistance against amoxicillin, tobramycin, tetracycline and nitrofurantoin antibiotics. Maximum biofilm formation was shown by strain E124 when used as monoculture. In cocultures, strains E35 and E101 were the most efficient biofilm formers. PCR amplification recorded negative results for shiga toxin genes. Conclusion: Shiga toxin genes were not present in any of the E. coli strain, from which we can suggest that our environment is free of shiga toxin genes. E. coli was present in sewage water, its cross contamination with drinking water may affect the community. Therefore, waste water should be treated properly before discarding it into the common water bodies.

Contribution and Interaction of Shiga Toxin Genes to Escherichia coli O157:H7 Virulence

Escherichia coli O157:H7 is the predominant cause of diarrhea-associated hemolytic uremic syndrome (HUS) worldwide. Its cardinal virulence traits are Shiga toxins, which are encoded by stx genes, the most common of which are stx1a, stx2a, and stx2c. The toxins these genes encode differ in their in vitro and experimental phenotypes, but the human population-level impact of these differences is poorly understood. Using Shiga toxin-encoding bacteriophage insertion typing and real-time polymerase chain reaction, we genotyped isolates from 936 E. coli O157:H7 cases and verified HUS status via chart review. We compared the HUS risk between isolates with stx2a and those with stx2a and another gene and estimated additive interaction of the stx genes. Adjusted for age and symptoms, the HUS incidence of E. coli O157:H7 containing stx2a alone was 4.4% greater (95% confidence interval (CI) −0.3%, 9.1%) than when it occurred with stx1a. When stx1a and stx2a occur together, the risk of HUS was 27.1% lower (95% CI −87.8%, −2.3%) than would be expected if interaction were not present. At the population level, temporal or geographic shifts toward these genotypes should be monitored, and stx genotype may be an important consideration in clinically predicting HUS among E. coli O157:H7 cases.