Serogroup O1 Research Articles

Гены и белки холодового шока у Vibrio cholerae различных серогрупп

553 whole genome sequences of Vibrio cholerae strains O1, O139 and nonO1/nonO139 serogroups were studied for the presence of cold-shock genes cspA, cspV and csh1. The cspA and cspV genes were present in almost all strains. The csh1 gene was present in 99/449 strains of the serogroup O1 and in 21/86 cultures of nonO1/nonO139 taken in the study. The absence of the csh1 gene was revealed in strains of the O139 serogroup and strains of the O1 serogroup with ctxAB and tcpA genes. The study of the nucleotide composition identified 13 different variants of the csh1 gene which cause structural differences in the cold-shock protein Csh1. The two main types of Csh1 protein were represented by the reference type (68 genomes) and the major type (35 genomes), 11 minor variants included single genomes. According to the NCBI database, mainly representatives of minor types of Csh1 protein circulate abroad. The reference type Csh1 was mainly detected in O1 serogroup strains, while 10 strains with the reference type Csh1 protein were identified before 2000, then in 2001-2022 their number reached 55 strains. It is assumed that the new cold shock gene csh1 gives vibrions a selective advantage by ensuring survival at low temperatures in reservoirs.

Genomic and Evolutionary Insights into Australian Toxigenic Vibrio cholerae O1 Strains.

Vibrio cholerae O1 is the causative agent of cholera, a severe diarrheal disease which can cause death if left untreated. In this study, a collection of clinical and environmental V. cholerae serogroup O1 isolates from Australia (1977 to 1987) (from local cases and cases acquired through international travel) and publicly available international isolates were characterized for genotypic features (virulence genes, mobile genetic elements [MGEs], and antimicrobial resistance gene profiles). Whole-genome sequencing (WGS) was used to investigate and compare the genetic relatedness between the 44 Australian and nine travel-associated isolates and the 60 publicly available international V. cholerae sequences representing pre-seventh-pandemic (pre-7PET) isolates and different waves of 7PET isolates. In this study, 36 (81%) Australian clinical and aquatic isolates harbored the cholera toxin-producing genes located in the CTX bacteriophage region. All the Australian environmental and clinical isolates lacked the seventh-pandemic virulence-associated genomic islands (VSP-I and -II). In silico multilocus sequence typing (MLST) classified all nine internationally acquired isolates as sequence type 69 (ST69), 36 clinical and aquatic isolates as ST70, and eight isolates from Australia as ST71. Most of the nontoxigenic clinical and aquatic isolates of ST71 had diverse genetic variations compared to ST70 Australian strains. The antimicrobial resistance-associated genes gyrA, parC, and parE had no mutations in all the environmental and clinical isolates from Australia. The SXT genetic element and class 1 integron gene sequences were not detected in Australian strains. Moreover, in this study, a Bayesian evolutionary study suggests that two distinct lineages of ST71 (new set of strains) and ST70 strains were prevalent around similar times in Australia, in ~1973 and 1969. IMPORTANCE Australia has its own indigenous V. cholerae strains, both toxigenic and nontoxigenic, that are associated with disease. Exotic strains are also detected in Australian patients returning from overseas travel. The clinical and aquatic V. cholerae O1 toxin gene-positive isolates from Australia responsible for cases in 1977 to 1987 were linked to acquisition from Queensland waterways but until now had not been characterized genetically. It is important to determine the genetic relatedness of Australian strains to international strains to assist in understanding their origin. This is the first extensive study to provide sequences and genomic analysis focused on toxigenic O1 V. cholerae clinical and environmental strains from Australia and its possible evolutionary relationship with other publicly available pre-7PET and 7PET V. cholerae strains. It is important to understand the population genetics of Australian V. cholerae from a public health perspective to assist in devising control measures and management plans for reducing V. cholerae exposure in Australia, given previous Australian disease clusters.

Dual-Use Vaccine for Diarrhoeal Diseases: Cross-Protective Immunogenicity of a Cold-Chain-Free, Live-Attenuated, Oral Cholera Vaccine against Enterotoxigenic Escherichia coli (ETEC) Challenge in BALB/c Mice.

In low- and middle-income countries, diarrhoeal diseases are the second most common cause of mortality in children, mainly caused by enterotoxin-producing bacteria, such as Shigella, Vibrio, Salmonella, and Escherichia coli. Cholera and traveller's diarrhoea are caused by Vibrio cholerae (O1 and O139 serogroups) and enterotoxigenic Escherichia coli (ETEC), respectively. The cholera toxin (CT) produced by V. cholerae and the heat-labile enterotoxin (LT) of ETEC are closely related by structure, function, and the immunological response to them. There is no exclusive vaccine for ETEC; however, cholera vaccines based on the CT-B component elicit a short-term cross-protection against ETEC infection. In this context, the cross-protective efficacy of MyCholTM, a prototype cold-chain-free, live-attenuated, oral cholera vaccine against V. cholerae O139 was evaluated in BALB/c mice. The 100% lethal dose (LD100) of 109 CFU/mL of the ETEC H10407 strain was used for the challenge studies. The mice immunised with MyChol™ survived the challenge by producing anti-CT antibodies, which cross-neutralised the LT toxin with no body weight loss and no sign of diarrhoea. Compared to unimmunised mice, the immunised mice elicited the neutralising antitoxin that markedly decreased ETEC colonisation and fluid accumulation caused by ETEC H10407 in the intestines. The immunised mice recorded higher antibody titres, including anti-CT IgG, anti-LT IgG, anti-CT-B IgG, and anti-LTB IgG. Only a two-fold rise in anti-CT/CT-B/LT/LT-B IgA was recorded in serum samples from immunised mice. No bactericidal antibodies against ETEC H10407 were detected. This investigation demonstrates the safety, immunogenicity, and cross-protective efficacy of MyCholTM against the ETEC H10407 challenge in BALB/c mice.

Cholera Outbreak 2022 in Karachi: A Report on Serotype and Antibiotic Susceptibility Pattern.

To identify the bacterial agent responsible for the surge of cases of acute diarrhoeal disease in Karachi, Pakistan, and assess the antimicrobial susceptibility pattern of the isolates. Observational study. Department of Microbiology, Indus Hospital & Health Network, Karachi, Pakistan, from 1st March to 31st May 2022. Vibrio cholerae (V. Cholerae) isolates from stool samples received in the laboratory in the months of March, April, and May were identified and antibiotic susceptibility testing was performed by the Kirby-Bauer disk diffusion method. Serology was performed to identify the serotype causing a surge in cases of acute watery diarrhoea. A total of three-hundred and seventy-eight stool samples were received during the study period, out of which seventy-eight were positive for V. cholerae serogroup O1 serotype Ogawa. The isolate tested 100% sensitive for Ciprofloxacin, Tetracycline, Doxycycline, and Azithromycin, while 74% resistance was observed in Trimethoprim/Sulfamethoxazole and 25% resistance in Ampicillin. V. cholerae serogroup O1 serotype Ogawa was the causative agent responsible for cholera outbreak in Karachi during the early summer of 2022. Ciprofloxacin, Tetracycline, Doxycycline, and Azithromycin are appropriate antimicrobials for treatment; whereas resistance to Trimethoprim/Sulfamethoxazole, and Ampicillin was higher. Vibrio cholerae, Outbreak, Acute diarrhoeal disease.

Cholera Vaccine: Recommendations of the Advisory Committee on Immunization Practices, 2022.

SummaryThis report summarizes all recommendations from CDC’s Advisory Committee on Immunization Practices (ACIP) for the use of lyophilized CVD 103-HgR vaccine (CVD 103-HgR) (Vaxchora, Emergent BioSolutions, Gaithersburg, MD) in the United States. The live attenuated oral cholera vaccine is derived fromVibrio cholerae O1 and is administered in a single dose. Cholera is a toxin-mediated bacterial gastrointestinal illness caused by toxigenic V. cholerae serogroup O1 or, uncommonly, O139. Up to 10% of infections manifest as severe cholera (i.e., cholera gravis), profuse watery diarrhea that can cause severe dehydration and death within hours. Fluid replacement therapy can reduce the fatality rate to <1%. Risk factors for cholera gravis include high dose exposure, blood group O, increased gastric pH (e.g., from antacid therapy), and partial gastrectomy. Cholera is rare in the United States, but cases occur among travelers to countries where cholera is endemic or epidemic and associated with unsafe water and inadequate sanitation. Travelers might be at increased risk for poor outcomes from cholera if they cannot readily access medical services or if they have a medical condition that would be worsened by dehydration, such as cardiovascular or kidney disease. This report describes previously published ACIP recommendations about use of CVD 103-HgR for adults aged 18–64 years and introduces a new recommendation for use in children and adolescents aged 2–17 years. ACIP recommends CVD 103-HgR, the only cholera vaccine licensed for use in the United States, for prevention of cholera among travelers aged 2–64 years to an area with active cholera transmission. Health care providers can use these guidelines to develop the pretravel consultation for persons traveling to areas with active cholera transmission.

Shiga Toxin Subtypes, Serogroups, Phylogroups, RAPD Genotypic Diversity, and Select Virulence Markers of Shiga-Toxigenic Escherichia coli Strains from Goats in Mid-Atlantic US.

Understanding Shiga toxin subtypes in E. coli from reservoir hosts may give insight into their significance as human pathogens. The data also serve as an epidemiological tool for source tracking. We characterized Shiga toxin subtypes in 491 goat E. coli isolates (STEC) from the mid-Atlantic US region (stx1 = 278, stx2 = 213, and stx1/stx2 = 95). Their serogroups, phylogroups, M13RAPD genotypes, eae (intimin), and hly (hemolysin) genes were also evaluated. STEC-positive for stx1 harbored Stx1c (79%), stx1a (21%), and stx a/c (4%). Those positive for Stx2 harbored stx2a (55%) and Stx2b (32%), while stx2a/stx2d and stx2a/stx2b were each 2%. Among the 343 STEC that were serogrouped, 46% (n = 158) belonged to O8, 20% (n = 67) to 076, 12% (n = 42) to O91, 5% (n = 17) to O5, and 5% (n = 18) to O26. Less than 5% belonged to O78, O87, O146, and O103. The hly and eae genes were detected in 48% and 14% of STEC, respectively. Most belonged to phylogroup B1 (73%), followed by D (10%), E (8%), A (4%), B2 (4%), and F (1%). M13RAPD genotyping revealed clonality of 091, O5, O87, O103, and O78 but higher diversity in the O8, O76, and O26 serogroups. These results indicate goat STEC belonged to important non-O157 STEC serogroups, were genomically diverse, and harbored Shiga toxin subtypes associated with severe human disease.

Acute exacerbation of chronic suppurative otitis media by Vibrio cholerae O39 – A case report

Vibrio cholerae is gram-negative bacilli belonging to the Vibrionaceae family generally found in brackish and marine water. Based on the O antigen on the cell wall of the bacteria, they are divided into three major subgroups – V. cholerae O1, V. cholerae O139, and V. cholerae non-O1/non-O139. The serogroups O1 and O139 harboring cholera toxin gene cause diarrhea, while the non-O1/non-O139 can cause a milder diarrhea and extraintestinal infections. We report a case of acute exacerbation of chronic suppurative otitis media by V. cholerae O39, which is classified in the non-O1/non-O139 group. The patient presented with slowly progressive painful ear discharge which evolved over a period of 3 months. V. cholerae O39 was isolated from the ear discharge by culture and confirmed by serologic and molecular testing. The patient improved symptomatically with 1 week of treatment with intravenous ciprofloxacin and tobramycin ear drops.

Vibrio cholerae O139 genomes provide a clue to why it may have failed to usher in the eighth cholera pandemic

Cholera is a life-threatening infectious disease that remains an important public health issue in several low and middle-income countries. In 1992, a newly identified O139 Vibrio cholerae temporarily displaced the O1 serogroup. No study has been able to answer why the potential eighth cholera pandemic (8CP) causing V. cholerae O139 emerged so successfully and then died out. We conducted a genomic study, including 330 O139 isolates, covering emergence of the serogroup in 1992 through to 2015. We noted two key genomic evolutionary changes that may have been responsible for the disappearance of genetically distinct but temporally overlapping waves (A-C) of O139. Firstly, as the waves progressed, a switch from a homogenous toxin genotype in wave-A to heterogeneous genotypes. Secondly, a gradual loss of antimicrobial resistance (AMR) with the progression of waves. We hypothesize that these two changes contributed to the eventual epidemiological decline of O139.

Genomic Characterization of Escherichia coli O8 Strains Producing Shiga Toxin 2l Subtype.

Shiga toxin-producing Escherichia coli (STEC) can cause diseases ranging from mild diarrhea to fatal extra-intestinal hemolytic uremic syndrome (HUS). Shiga toxin (Stx) is the key virulence factor in STEC, two Stx types (Stx1 and Stx2) and several subtypes varying in sequences, toxicity, and host specificity have been identified. Stx2l is a newly-designated subtype related to human disease but lacks thorough characterization. Here, we identified Stx2l from five STEC strains (Stx2l-STECs) recovered from raw mutton and beef in China. Whole-genome sequencing (WGS) was used to characterize the Stx2l-STECs in this study together with Stx2l-STECs retrieved from public databases. Our study revealed that all the analyzed Stx2l-STEC strains belonged to the same serogroup O8. Multilocus sequencing typing (MLST) showed two sequence types (ST88 and ST23) among these strains. Stx2l-converting prophages from different sources shared a highly similar structure and sequence. Single-nucleotide polymorphism (SNP)-based analysis revealed genetic relatedness between the human-derived and food-derived strains belonging to ST23. To conclude, our study supported the designation of Stx2l and demonstrated diverse host range and geographical distribution of Stx2l-STECs.Stx2l-STEC strains from different sources showed a high genetic similarity with an identical O8 serogroup. Further studies are needed to investigate the epidemiological trait and pathogenic potential of Stx2l-STEC strains.

Diversity of Vibrio cholerae and prevalence of biomarker genes in the ballast water, Port Blair, South Andaman, India

Ship's ballast water is the primary carriers for global transport of non-indigenous marine species, harmful aquatic organisms and pathogens. The rates of non-indigenous species introduced to marine environments via ballast water were estimated to be 30% during the last 20–30 years. In this study, with an aim to detect the transportable bacteriological risk of ships' ballast water for the Andaman sea, the composition and frequency of Vibrio cholerae O1 and O139 isolates, abundance of virulence factors and antibiotic resistance pattern were investigated in the ballast water of ships coming from various marine environments to Port Blair, Andaman Islands. The in-situ use of chromogenic agar were investigated to isolate the V. cholerae strains in the ships' ballast water from different regions to the Port Blair. The samples were collected from 10 ships coming from various marine environments were tested. Seventy eight V. cholerae strains were isolated from the ballast water of five ships, out of 10 ballast water samples V. cholerae was not detected in the five samples. Out of 78 isolates, 27 were O1 and 17 were O139 serogroups.

Vibrio cholerae: a pathogen shared by human and aquatic animals

Vibrio cholerae: a pathogen shared by human and aquatic animals

Identifying long-term colonization factors of Vibrio cholerae O1 El Tor in the zebrafish natural host model

The human disease cholera, marked by acute, voluminous watery diarrhea, is caused by the gram-negative, aquatic bacterium Vibrio cholerae. All seven cholera pandemics since 1817 were identified as being caused by just 2 of over 155 known V. choleraeserogroups: O1 and O139. The O1 serogroup is divided into two biotypes: classical and El Tor. Classical biotype is associated with pandemics 1 through 6, but the El Tor biotype has since displaced classical as the causative agent of the ongoing 7th cholera pandemic over the past 60 years. The El Tor genome resembles that of classical but has acquired two unique pathogenicity islands known as Vibrio Seventh Pandemic (VSP) -1 and -2. El Tor biotype has been associated with prolonged colonization, infection, and disease both in humans and in the zebrafish natural host model. The zebrafish model allows for complete observation of Vibrio cholerae infection in a system undisrupted by antibiotic use or immune suppression. El Tor strains colonize the zebrafish intestine for up to 10 days longer than classical strains. Preliminary studies demonstrate VSP-2 is required to observe this phenotype, but VSP-1 is not. By creating targeted regional knockouts of the VSP-2 island, the specific gene(s) essential for enabling prolonged colonization will be identified and applied to the understanding of how El Tor interacts with a natural host. By identifying the genes used by El Tor to colonize a natural host for prolonged periods, we gain insight into how this pathogen may persist in the environment and perpetuate disease.

Fragmentation analysis of O-specific polysaccharide from bacteria Vibrio cholerae O139 by MALDI-TOF and LC/ESI-MS/MS.

Cholera is a life-threatening diarrhoeal disease caused by ingestion of Vibrio cholerae. There are at least 200 serogroups of V. cholerae but only two of them are causing epidemics - O1 and O139 serogroups. Fragmentation analysis of O-antigen, also known as O-specific polysaccharide (OSP), from lipopolysaccharide (LPS) is important to obtain new information about its structure, such as fragmentation patterns and fragment structures. In the present study, OSP and core (OSPc) structure from V. cholerae O139 was studied using matrix-assisted laser desorption ionization (MALDI)-time of flight (TOF) and direct injection electrospray ionization (ESI)-MS methods. MALDI-TOF analysis was performed in positive-ion reflectron mode, while ESI-MS was performed in negative ionization mode. ESI-MS analysis was followed by ESI-MS/MS analysis. Using this analytical approach, we managed to obtain two possible fragmentation pathways of OSP from V. cholerae O139. Mutual sign of these two pathways is shortening the length of the oligosaccharide by neutral loss of monosaccharide residues. Additionally, liquid chromatography-MS analysis was performed to separate depicted molecular forms of OSPc.

Investigation of Serotype Prevalence of Escherichia coli Strains Isolated from Layer Poultry in Greece and Interactions with Other Infectious Agents.

Colibacillosis is the most common bacterial disease in poultry and it is caused by avian pathogenic Escherichia coli (APEC), which is assigned to various O-serogroups. Previous studies have shown that APEC strains are more often related to certain O-serogroups such asO78, O2 and O1. E. coli has been reported to act either as a primary or secondary agent in complicating other infections. The aim of this study was to investigate the occurrence of and characterize the O-serogroups of E. coli strains isolated from commercial layer and layer breeder flocks showing macroscopic lesions of colibacillosis and increased or normal mortality in Greece. Furthermore, we attempted to assess the interaction between infectious agents such as Mycoplasma gallisepticum (MG), Mycoplasma synoviae (MS), infectious bronchitis (IBV) and infectious laryngotracheitis (ILT) with E. coli infections in layer flocks with increased mortality. Our study revealed that in addition to the common serogroups (O78, O2), many other, and less common serogroups were identified, including O111. The O78, O111 and O2 serogroups were frequently detected in flocks with lesions of colibacillosis and increased mortality whereas O2, O88 and O8 were reported more commonly in birds with colibacillosis lesions but normal mortality rates. These data provide important information for colibacillosis monitoring and define preventative measures, especially by using effective vaccination programs because E. coli vaccines are reported to mainly offer homologous protection. Finally, concerning the association of the four tested infectious agents with E. coli mortality, our study did not reveal a statistically significant effect of the above infectious agents tested with E. coli infection mortality.

Inhibitory activities of Typhonium trilobatum (L.) Schott on virulence potential of multi-drug resistant toxigenic Vibrio cholerae

Cholera is a serious epidemic disease caused by the toxigenic strains of Vibrio cholerae belonged to O1 or O139 serogroups. The emergence of antibacterial resistance in V. cholerae is an increasing concern. Natural product drug invention and Ethnopharmacology may demonstrate a considerable expectation under this circumstance. Traditionally, leaves of Typhonium trilobatum (L.) Schott (locally known as Ghatkanchu or Bengal Arum) are employed for treatment of gastrointestinal disorder in different region of India.The objective of the present study was to evaluate the antibacterial, and antibiofilm activities of methanol extract of T. trilobatum leaves (METTL) against the strains of multi-drug resistant (MDR) Vibrio cholerae (serotypes O1, O139, non-O1, and non-O139) which are responsible for watery diarrhea such as cholera.MIC, MBC and time-kill kinetic studies were used for evaluation of In vitro antibacterial activity of METTL. Microdilution method and Confocal laser scanning microscopy were used to evaluate biofilm-inhibitory activities. The gene expression was analyzed by performing Quantitative real-time PCR (qRT-PCR).METTL showed antibacterial activity with MIC and MBC at 1–32 mg/mL and 8–32 mg/mL, respectively against the clinical strains of Vibrio cholerae belonged to different serogroups. METTL showed significant (P < 0.05) inhibitory activity on the formation of biofilm by V. cholerae SG24, with 81.3, 75.8, and 69.6% of inhibition at MIC, ½ MIC and ¼ MIC, respectively. METTL showed also significant (P < 0.05) inhibitory activity on the formation of extracellular polymeric substances (EPS) formation by V. cholerae SG24, with 89.41, and 99.26% of inhibition of EPS protein and EPS carbohydrate at MIC, respectively.METTL significantly (p < 0.01) inhibited the Cholera toxin (CT) production by the V. cholerae strain SG24 evaluated by the CT - ELISA assay. The cholera toxin production was reduced by 76.26%, 48.76% and 29.93 at MIC (8 mg/mL), ½ MIC (4 mg/mL) and ¼ MIC (2 mg/mL), respectively. METTL was shown to repress ctxAB gene transcription 1.76 fold (p < 0.05) at sub-bactericidal concentration (¼ MIC). We also found that the expression of cholera toxin activator genes, toxT and tcpP was reduced by 11.56- fold (p < 0.001) and 23.52- fold (p < 0.001), respectively, at sub-bactericidal concentration (¼ MIC). Transcription of the following genes was repressed: vpsR (1.8-fold; p < 0.05), Bap1 (1.53-fold; p ≤ 0.05), and rmbA (2.89-fold) by METTL at sub-bactericidal concentration. The expression of vpsT was also repressed by 1.5-fold (p < 0.01) at sub-bactericidal concentration.The active Typhonium trilobatum (L.) leaves extract may be suggested as an substitute for the treatment of MDR V. cholerae infection and could be used as prospective source for the development of novel antimicrobial compound/s and biofilm-inhibitory drug/s useful for the treatment of cholera and diarrheal patients. The results obtained here also validate scientifically the traditional uses of Typhonium trilobatum (L.) in India employed for the treatment of gastrointestinal disorder. Further studies should be directed at purifying and characterizing these antibacterial principles against Vibrio cholerae.

A high-risk carbapenem-resistant Pseudomonas aeruginosa clone detected in red deer (Cervus elaphus) from Portugal

Pseudomonas aeruginosa is a ubiquitous bacterium, successfully exploiting a variety of environmental niches due to its remarkable metabolic versatility. The World Health Organization classifies P. aeruginosa as a “priority pathogen” due to its a great ability to overcome the action of antimicrobials, including carbapenems. Hitherto, most studies have focused on clinical settings from humans, but much less on animal and environmental settings, particularly on wildlife. In this work, we report the isolation of a carbapenem-resistant Pseudomonas aeruginosa strain recovered from the faeces of a red deer adult female sampled in a humanized area. This isolate was obtained during a nationwide survey on antimicrobial resistance in wildlife aimed to determine the occurrence of carbapenem-resistant bacteria among 181 widely distributed wild ungulates. This P. aeruginosa isolate was found to be a high-risk clone, belonging to the sequence type (ST) 274. The genomic analysis of P. aeruginosa isolate UP4, classified this isolate as belonging to serogroup O3, which was also found to harbour the genes blaPAO, blaPDC-24, blaOXA-486 (encoding resistance to beta-lactams), aph(3′)-IIb (aminoglycosides resistance), fosA (fosfomycin resistance) and catB7 (chloramphenicol resistance). Antimicrobial susceptibility screening, according to EUCAST, showed resistance to imipenem and intermediate resistance to meropenem and doripenem. To our knowledge, this is the first description of carbapenem-resistant P. aeruginosa in deer in Europe. Our results highlight the importance of wild ungulates either as victims of human activity or amplifiers of AMR, either way with potential impacts on animal, human and ecosystem health, since excretion of AMR bacteria might directly or indirectly contaminate other animals and the surrounding environment, perpetuating the spill-over and chain dissemination of AMR determinants.

Characterization of Uropathogenic Escherichia coli Reveals Hybrid Isolates of Uropathogenic and Diarrheagenic (UPEC/DEC) E. coli.

(1) Background: Pathogenic Escherichia coli are divided into two groups: diarrheagenic (DEC) and extraintestinal pathogenic (ExPEC) E. coli. ExPEC causing urinary tract infections (UTIs) are termed uropathogenic E. coli (UPEC) and are the most common cause of UTIs worldwide. (2) Methods: Here, we characterized 112 UPEC in terms of phylogroup, serotype, the presence of virulence factor-encoding genes, and antimicrobial resistance. (3) Results: The majority of the isolates were assigned into the phylogroup B2 (41.07%), and the serogroups O6 (12.5%) and O25 (8.9%) were the most frequent. Five hybrid UPEC (4.5%), with markers from two DEC pathotypes, i.e., atypical enteropathogenic (aEPEC) and enteroaggregative (EAEC) E. coli, were identified, and designated UPEC/aEPEC (one isolate) and UPEC/EAEC (four isolates), respectively. Three UPEC/EAEC harbored genes from the pap operon, and the UPEC/aEPEC carried ibeA. The highest resistance rates were observed for ampicillin (46.4%) and trimethoprim/sulfamethoxazole (34.8%), while 99.1% of the isolates were susceptible to nitrofurantoin and/or fosfomycin. Moreover, 9.8% of the isolates were identified as Extended Spectrum β-Lactamase producers, including one hybrid UPEC/EAEC. (4) Conclusion: Our data reinforce that hybrid UPEC/DEC are circulating in the city of Botucatu, Brazil, as uropathogens. However, how and whether these combinations of genes influence their pathogenicity is a question that remains to be elucidated.

Assay for Evaluating the Abundance of Vibrio cholerae and Its O1 Serogroup Subpopulation from Water without DNA Extraction.

Cholera is a severe diarrheal disease caused by Vibrio cholerae, a natural inhabitant of brackish water. Effective control of cholera outbreaks depends on prompt detection of the pathogen from clinical specimens and tracking its source in the environment. Although the epidemiology of cholera is well studied, rapid detection of V. cholerae remains a challenge, and data on its abundance in environmental sources are limited. Here, we describe a sensitive molecular quantification assay by qPCR, which can be used on-site in low-resource settings on water without the need for DNA extraction. This newly optimized method exhibited 100% specificity for total V. cholerae as well as V. cholerae O1 and allowed detection of as few as three target CFU per reaction. The limit of detection is as low as 5 × 103 CFU/L of water after concentrating biomass from the sample. The ability to perform qPCR on water samples without DNA extraction, portable features of the equipment, stability of the reagents at 4 °C and user-friendly online software facilitate fast quantitative analysis of V. cholerae. These characteristics make this assay extremely useful for field research in resource-poor settings and could support continuous monitoring in cholera-endemic areas.

Population Analysis of O26 Shiga Toxin-Producing Escherichia coli Causing Hemolytic Uremic Syndrome in Italy, 1989-2020, Through Whole Genome Sequencing.

Shiga toxin-producing Escherichia coli (STEC) belonging to the O26 serogroup represent an important cause of Hemolitic Uremic Syndrome (HUS) in children worldwide. The localization of STEC virulence genes on mobile genetic elements allowed the emergence of clones showing different assets of this accessory genomic fraction. A novel O26 STEC clone belonging to Sequence Type (ST) 29 and harboring stx2a, ehxA and etpD plasmid-borne genes has emerged and spread in Europe since the mid-1990s, while another ST29 clone positive for stx2d and lacking plasmid-borne virulence genes was recently described as emerging in France. In Italy, O26 has been the most frequently detected STEC serogroup from HUS cases since the late 1990s. In this study we describe the genomic characterization and population structure of 144 O26 STEC strains isolated from human sources in Italy in the period 1989-2020. A total of 89 strains belonged to ST21, 52 to ST29, two to ST396 and one to ST4944. ST29 strains started to be isolated from 1999. 24 strains were shown to harbour stx1a, alone (n=20) or in combination with stx2a (n=4). The majority of the strains (n=118) harbored stx2a genes only and the two ST396 strains harbored stx2d. A Hierarchical Clustering on Principal Components (HCPC) analysis, based on the detection of accessory virulence genes, antimicrobial resistance (AMR) genes and plasmid replicons, classified the strains in seven clusters identified with numbers from 1 to 7, containing two, 13, 39, 63, 16, 10 and one strain, respectively. The majority of the genetic features defining the clusters corresponded to plasmid-borne virulence genes, AMR genes and plasmid replicons, highlighting specific assets of plasmid-borne features associated with different clusters. Core genome Multi Locus Sequence Typing grouped ST21 and ST29 strains in three clades each, with each ST29 clade exactly corresponding to one HCPC cluster. Our results showed high conservation of either the core or the accessory genomic fraction in populations of ST29 O26 STEC, differently from what observed in ST21 strains, suggesting that a different selective pressure could drive the evolution of different populations of these pathogens possibly involving different ecological niches.

Phylogenetic Analysis of Serogroup O5 Vibrio cholerae that Caused Successive Cholera Outbreaks - Guangdong Province, China, 2020-2021.

IntroductionGastroenteritis caused by non-O1/non-O139 Vibrio cholerae exhibited an increasing trend in recent years in China. Whole genome sequence (WGS) data could play an important role both in the identification of the outbreaks and in the determination of the serogroup. Here, we present the employment of WGS data in the investigation of two outbreaks caused by non-O1/non-O139 V. cholerae in Guangdong, China, 2020–2021. MethodsWe obtained the whole genome sequence of 66 V. cholerae strains isolated in two outbreaks with next generation sequencing technology. We retrieved the publicly available WGS data of non-O1/non-O139 V. cholerae from public database. We used a pipeline integrated in China Pathogen Identification Net (PIN) to complete the phylogenetic analysis. ResultsTwo outbreaks caused by non-O1/non-O139 V. cholerae were identified using WGS data. These V. cholerae strains were determined as serogroup O5. Type 3 and 6 secretion systems were detected in these serogroup O5 strains. These serogroup O5 strains belonged to sequence type (ST) 88. ConclusionsOur analysis indicated the risk of non-O1/non-O139 V. cholerae leading to outbreaks of diarrheal diseases. The application of genomic data played an important role in the identification of the serogroup of non-O1/non-O139 V. cholerae in the lack of antiserum, which gave an example of the application of genome data in disease surveillance and public health emergency response.