Serogroup O1 Research Articles

Proteomic analysis of typical and genetically altered strains of <i>Vibrio cholerae</i> serogroup O1, biovar El Tor

Objective — comparative study of protein expression in typical and genetically altered Vibrio cholerae strains of O1 serogroup, biovar El Tor by means of proteomic analysis.Materials and methods. Clinical V. cholerae strains — typical strain, M106 (Astrakhan, 1970) and genetically altered one, M1509 (Moscow, 2012) — were used as model ones. Strains were cultivated in LB broth (pH7.2). Then, cell and exoprotein lysate fractions were obtained and investigated in 2D electrophoresis. Different protein stains were examined using mass spectrometry. Survivability of V. cholerae strains under osmotic and oxidative stresses was studied during incubation of the strains in 3 M NaCl solution or 20 mM H2O2 solution.Results and discussion. When analyzing cell lysates, significant differences in protein expression with known function between studied strains were not detected. The great majority of identified proteins in the lysates is functionally associated with carbohydrate metabolism, amino acid metabolism, and energy processes in a cell. At the same time, exoprotein fraction of M1509 genovariant contained increased amount of proteins (peroxidase, superoxide dismutase, thioredoxin, outer membrane proteins OmpW, OmpT) protecting the cells of cholera vibrio from effect of stress factors of the environment. Further study of the resistance to osmotic and oxidative stresses revealed better survivability in the genovariant when exposed to the stated factors.Conclusion. The data of proteomic analysis of the typical and genetically altered V. cholera strains, biovar El Tor, testify to high levels of expression of the proteins that provide for vibrio resistance to the effect of environmental stress factors in genovariants, which is possibly one of the causes of their wide dissemination. In addition, the results obtained will allow for identification of new biomarkers which can be used for differentiation of typical strains and genovariants of V. cholerae, biovar El Tor.

Evolution, distribution and genetics of atypical Vibrio cholerae - A review.

Vibrio cholerae is the etiological agent of cholera, a severe diarrheal disease, which can occur as either an epidemic or sporadic disease. Cholera pandemic-causing V. cholerae O1 and O139 serogroups originated from the Indian subcontinent and spread globally and millions of lives are lost each year, mainly in developing and underdeveloped countries due to this disease. V. cholerae O1 is further classified as classical and El Tor biotype which can produce biotype specific cholera toxin (CT). Since 1961, the current seventh pandemic El Tor strains replaced the sixth pandemic strains resulting in the classical biotype strain that produces classical CT. The ongoing evolution of Atypical El Tor V. cholerae srains encoding classical CT is of global concern. The severity in the pathophysiology of these Atypical El Tor strains is significantly higher than El Tor or classical strains. Pathogenesis of V. cholerae is a complex process that involves coordinated expression of different sets of virulence-associated genes to cause disease. We are yet to understand the complete virulence profile of V. cholerae, including direct and indirect expression of genes involved in its survival and stress adaptation in the host. In recent years, whole genome sequencing has paved the way for better understanding of the evolution and strain distribution, outbreak identification and pathogen surveillance for the implementation of direct infection control measures in the clinic against many infectious pathogens including V. cholerae. This review provides a synopsis of recent studies that have contributed to the understanding of the evolution, distribution and genetics of the seventh pandemic Atypical El Tor V. cholerae strains.

Retrospective Analysis of the Serovars and Antibiogram of Vibrio cholerae Isolates of the 2017 Ilorin Cholera Outbreak, Nigeria.

In this retrospective study, we determined the incidence, serovars, and antibiogram of Vibrio cholerae isolated from 102 clinical stool samples collected from rice water diarrheic patients during an outbreak (May - July 2017) in Ilorin metropolis, Nigeria. The culture positive rate of the V. cholerae isolates was 41.2%, with 41 and 1 isolates from O1 (Inaba) and non-O1/O139 serogroups, respectively. The isolates were the most susceptible to ciprofloxacin (76.2%) followed by amoxicillin-clavulanate (71.4%). However, all isolates were resistant to ampicillin and tetracycline. In conclusion, V. cholerae O1 was the predominant circulating serogroup exhibiting multi-drug resistance during the outbreak.

Total Synthesis of a Structurally Complex Tetrasaccharide Repeating Unit of <i>Vibrio cholerae</i> O43

Vibrio cholerae is a causative agent of the deadly pandemic - cholera. Among ~200 serogroups of V. cholerae, the O1 and O139 serogroups are mainly responsible for virulent cholera. Some other serogroups are also found to be associated with diarrhoea and minor outbreaks of cholera type diseases. The deadly pathogen is a threat for many developing countries, predominately in Asia and Africa, and still causes millions of deaths every year. A multicomponent glycoconjugate vaccine covering all the strains of V. cholerae is highly desired to prevent the spread of this disease. Here, we report the first total synthesis of a densely functionalized tetrasaccharide repeating unit of V. cholerae O43, which contains rare deoxy amino sugars D-quinosamine and D-viosamine attached with a rare amino acid N-acetyl-L-allothreonine. Synthesis of orthogonally protected rare sugars and unnatural amino acid building blocks, stereoselective construction of three consecutive 1,2-cis glycosidic linkages and the presence of five nitrogen atoms dispersed over four sugar units as well as carboxylic acid functionality makes the total synthesis a formidable task. The tetrasaccharide was assembled via a highly regio- and stereoselctive [3+1] glycosylation of fully functionalized, amino acid-bearing D-viosamine donor with the trisaccharide diol. The assembly of all-1,2-cis linked trisaccharide diol was expeditiously achieved in one-pot manner. Judicious selection of protecting groups and reaction conditions enabled access to the structurally complex, amino acid-attached tetrasaccharide repeating unit, which can be used for future biological studies directed towards development of vaccine. The synthetic strategy will be useful for accessing other structurally related complex bacterial glycans.

Association between antimicrobial treatment and resistance of pathogenic Escherichia coli isolated from diseased swine in Kagoshima Prefecture, Japan.

Pathogenic Escherichia coli is an important cause of diarrhea, edema disease, and septicemia in swine. In Japan, the volume of antimicrobial drugs used for animals is highest in swine, but information about the prevalence of antimicrobial-resistant bacteria is confined to apparently healthy animals. In the present study, we determined the O serogroups, virulence factors, and antimicrobial resistance of 360 E. coli isolates from swine that died of disease in Kagoshima Prefecture, Japan, between 1999 and 2017. The isolates of the predominant serogroups O139, OSB9, O149, O8, and O116 possessed virulence factor genes typically found in diarrheagenic E. coli. We further found five strains of third-generation cephalosporin-resistant E. coli that each produced an extended-spectrum β-lactamase encoded by blaCTX-M-14, blaCTX-M-15, blaCTX-M-24, blaCTX-M-61, or blaSHV-12. In 218 swine with a clear history of antimicrobial drug use, we further analyzed associations between the use of antimicrobials for the treatment of diseased swine and the isolation of resistant E. coli. We found significant associations between antimicrobial use and selection of resistance to the same class of antimicrobials, such as the use of ceftiofur and resistance to cefotaxime, cefazolin, or ampicillin, the use of aminoglycosides and resistance to streptomycin, and the use of phenicols and resistance to chloramphenicol. A significant association between antimicrobial use and the resistance of E. coli isolates to structurally unrelated antimicrobials, such as the use of ceftiofur and resistance to chloramphenicol, was also observed.

An outbreak investigation of acute Diarrheal Disease, Nagpur District, Maharashtra, India.

Acute diarrheal disease (ADD) accounts for 12 million cases and 1216 deaths annually in India. On July 13, 2016, an ADD outbreak was reported from Sawargaon village from Nagpur district, Maharashtra. The outbreak was investigated to describe the epidemiology and suggest control and preventive measures. A case was defined as a person experiencing at least one loose stool in Sawargaon village between July 9, 2016, and July 31, 2016. We searched for cases by enhanced passive surveillance. We collected stool samples for bacterial culture and tested water from multiple water sources for fecal coliforms. We also reviewed sanitary practices and rainfall data. A total of 889 cases were identified, with 51% female, 280 hospitalizations (31%), and two deaths. The median age was 27 years (range 6 months to 90 years). Cases started on July 9, a week after heavy rains. District authorities started chlorination of water sources on July 13 and cases declined soon after. Two of nine stool samples tested positive for Vibrio cholera O1 serogroup. Of the 18 water samples collected, 16 (88%) samples from multiple sources, including wells, hand pumps, and taps, were positive for fecal coliforms. Of 1,885 households in the village, 450 (24%) households had no toilets and open defecation was commonly observed in the nearby river bed. This ADD outbreak was likely associated with drinking contaminated groundwater, which probably occurred after heavy rainfall in an area of open defecation. We recommended providing chlorinated drinking water, promoting safe sanitation practices, including building more public and private toilets, and enhancing diagnostic laboratory capacity.

Identification of chironomid species as natural reservoirs of toxigenic Vibrio cholerae strains with pandemic potential.

Vibrio cholerae causes the fatal cholera diarrhea. Chironomids (Diptera; Chironomidae) are abundant in freshwater aquatic habitats and estuaries and are natural reservoirs of V. cholerae. Until now, only the non-O1/O139 serogroups of V. cholerae were identified in chironomids. Here, we explored whether chironomids are natural reservoirs of V. cholerae O1/O139 serogroups, which are associated with cholera endemics and pandemics. All four life stages of chironomids were sampled from two rivers, and a laboratory culture in Pune, India, and from a pond in Israel. In total, we analyzed 223 chironomid samples. The presence of V. cholerae O1/O139 serogroups was verified using molecular tools. Nine chironomid species were identified; of them, Chironomus circumdatus was the most abundant. The presence of V. cholerae serogroup O1 and the cholera toxin genes were detected in samples from all chironomid species. However, serogroup O139 was detected in only two chironomid species. Besides PCR to detect specific genes, a metagenomic analysis that was performed in three selected C. ramosus larvae, identified a list of virulence genes associated with V. cholerae. The findings provide evidence that chironomids are natural reservoirs of toxigenic V. cholerae O1/O139. Chironomid populations and V. cholerae show biannual peak patterns. A similar pattern is found for cholera epidemics in the Bengal Delta region. Thus, we hypothesize that monitoring chironomids in endemic areas of the disease may provide a novel tool for predicting and preventing cholera epidemics. Moreover, serogroup O139 was detected only in two chironomid species that have a restricted distribution in the Indian subcontinent, possibly explaining why the distribution of the O139 serogroup is limited.

DksA coordinates bile-mediated regulation of virulence-associated phenotypes in type three secretion system-positive Vibrio cholerae.

In order to cause disease, pathogenic strains of Vibrio cholerae rely on intricate regulatory networks to orchestrate the transition between their native aquatic environment and the human host. For example, bacteria in a nutrient-starved environment undergo a metabolic shift called the stringent response, which is mediated by the alarmone ppGpp and an RNA-polymerase binding transcriptional factor, DksA. In O1 serogroup strains of V. cholerae, which use the toxin co-regulated pilus (TCP) and cholera toxin (CT) as primary virulence factors, DksA was reported to have additional functions as a mediator of virulence gene expression. However, little is known about the regulatory networks coordinating virulence phenotypes in pathogenic strains that use TCP/CT-independent virulence mechanisms. We therefore investigated whether functions of DksA outside of the stringent response are conserved in type three secretion system (T3SS)-positive V. cholerae. In using the T3SS-positive clinically isolated O39 serogroup strain AM-19226, we observed an increase in dksA expression in the presence of bile at 37 °C. However, DksA was not required for wild-type levels of T3SS structural gene expression, or for colonization in vivo. Rather, data indicate that DksA positively regulates the expression of master regulators in the motility hierarchy. Interestingly, the ΔdksA strain forms a less robust biofilm than the WT parent strain at both 30 and 37 °C. We also found that DksA regulates the expression of hapR, encoding a major regulator of biofilm formation and protease expression. Athough DksA does not appear to modulate T3SS virulence factor expression, its activity is integrated into existing regulatory networks governing virulence-related phenotypes. Strain variations therefore may take advantage of conserved ancestral proteins to expand regulons responding to in vivo signals and thus coordinate multiple phenotypes important for infection.

Simultaneous Quantification of Vibrio metoecus and Vibrio cholerae with Its O1 Serogroup and Toxigenic Subpopulations in Environmental Reservoirs.

Vibrio metoecus is a recently described aquatic bacterium and opportunistic pathogen, closely related to and often coexisting with Vibrio cholerae. To study the relative abundance and population dynamics of both species in aquatic environments of cholera-endemic and cholera-free regions, we developed a multiplex qPCR assay allowing simultaneous quantification of total V. metoecus and V. cholerae (including toxigenic and O1 serogroup) cells. The presence of V. metoecus was restricted to samples from regions that are not endemic for cholera, where it was found at 20% of the abundance of V. cholerae. In this environment, non-toxigenic O1 serogroup V. cholerae represents almost one-fifth of the total V. cholerae population. In contrast, toxigenic O1 serogroup V. cholerae was also present in low abundance on the coast of cholera-endemic regions, but sustained in relatively high proportions throughout the year in inland waters. The majority of cells from both Vibrio species were recovered from particles rather than free-living, indicating a potential preference for attached versus planktonic lifestyles. This research further elucidates the population dynamics underpinning V. cholerae and its closest relative in cholera-endemic and non-endemic regions through culture-independent quantification from environmental samples.

Heterogeneity by antibiotic sensitivity and genotype of Vibrio Cholerae El Tor strains isolated from environmental objects in Russia

BACKGROUND: Waterborne diseases kill 3.4 million people worldwide each year. Cholera is one such disease. Up to 4 million cases of this infection occur in the world every year, leading to more than 100,000 deaths. The plasticity of the genome of the causative agent of cholera, the mobility of genetic elements carrying factors of pathogenicity and antibiotic resistance, contribute to the variability and unpredictability of the spectrum of resistance, the formation of new pheno- and genotypes. A single Vibrio cholerae isolate can contain up to 40 different genes that can confer resistance to 22 antibiotics, representing nine different classes of antimicrobial drugs. The ability of Vibrio cholerae to long-term survival in aquatic ecosystems in which there is an active exchange of genetic information and new ecological lines may arise that have potential advantages in the adaptation of microorganisms to adverse conditions, emphasizes the complexity of the ways of transmission of this infection and the need for studies at the environmental level. Periodic deliveries of cholera to the territory of the Russian Federation with (without) the spread of the infectious agent, V. cholerae contamination of surface water bodies used as sources of water supply and for recreational water use, the possibility of implementing the main route for cholera (water) the pathway for the spread of the pathogen of the infection indicate the need for conducting annual monitoring of antimicrobial resistance as part of the epidemiological surveillance of cholera in order to obtain information on the distribution, nature and dynamics of resistance in a specific period of time in a given territory.
 AIM: Analysis of the spectrum of antibiotic resistance of Vibrio cholerae O1 serogroup strains isolated from environmental objects in various territories of the Russian Federation in 2020.
 MATERIALS AND METHODS: 25 strains V. cholerae О1 El Tor, isolated from environmental objects in the Russian Federation in 2020. The sensitivity / resistance of the studied strains to 13 antibacterial drugs was determined by the method of serial dilutions on a solid nutrient medium in accordance with guidelines. PCR-genotyping of strains V. cholerae El Tor was performed for 14 target genes, followed by cluster analysis.
 RESULTS: The strains showed resistance markers to furazolidone, trimethoprim/sulfamethoxazole, ampicillin, nalidixic acid, ceftriaxone, which formed 5 phenotypes. PCR-genotyping genotyping of 14 target genes divided the strains into five genotypes (A1A5), corresponding to certain territories. The antibiotic resistance profiles within the same genotype in V. cholerae О1 El Tor belonging to different territories were both the same and different.
 CONCLUSION: The genotypic diversity of isolated strains was revealed, the variability of resistance markers even in one region, which indicates both changes in the V. cholerae population and the possibility of circulation of various geno- and phenotypes, which emphasizes the importance of constant monitoring of these pathogens.

Atypicity of <i>Vibrio cholerae </i> O1 strains based on agglutability

The review analyzes literature data on the phenotypic variability of Vibrio cholerae of the O1 serogroup. Three types of polysaccharide structures are distinguished in the cholera pathogen: lipopolysaccharide, or O-antigen, capsular polysaccharide and exopolysaccharide. The rugous form of V. cholerae strains is able to synthesize exopolysaccharide which is highly resistant to antimicrobials, and to form wrinkled colonies that can be agglutinated with cholera sera in various combinations. Serological classification of V. cholerae is based on the specificity of the lipopolysaccharide. V. cholerae of serogroups O1 and O139 causes cholera epidemics, although strains from non-O1/non-O139 serogroups with the main virulence factors are known. Upon transition from the Sto the R-form, lipopolysaccharide loses the O-polysaccharide, and the central region begins to fulfill the function of somatic antigen, exhibiting R-specificity. The serological differences in the strains of V. cholerae are based on a change in the regulation or structural organization of genes encoding the biosynthesis of O-antigen (rfb). From 1988 to 2019, 168 strains of V. cholerae that are atypical for agglutinability and were isolated from the R variant were isolated from environmental objects in the territory of the former USSR and subjects of the Russian Federation; the genetic organization of these strains has not been fully studied. The evolutionary relationships of atypical strains of cholera vibrios isolated from environmental objects in Siberia and the Far East are determined on the basis of a comparative analysis of housekeeping genes. The analysis of the results of basic and applied research indicates that the problem of variability in agglutinability of V. cholerae strains isolated from humans and from environmental objects remains relevant at the current stage of development of the seventh cholera pandemic. The atypicality of strains of V. cholerae O1 on this basis is considered in aspects of the environmental conditions of their existence and the conditionality of phenotypic manifestations by molecular biological determination.

Total Synthesis and Immunological Evaluation of the Tri-d-glycero-d-manno-heptose Antigen of the Lipopolysaccharide as a Vaccine Candidate against Helicobacter pylori.

Helicobacter pylori, the most common cause of chronic gastritis, peptic ulcers, and gastric cancers, infects around half of the world's population. Although the drawbacks of antibiotic-based combination therapy are emerging, no effective vaccine is available to prevent H. pylori infections. Here, we describe the total synthesis of the unique α-(1→3)-linked tri-d-glycero-d-manno-heptose antigen from the lipopolysaccharide of H. pylori serogroups O3 and O6 and strains MO19, D2, D4, and D5 based on de novo synthesis of the differentially protected d-glycero-d-manno-heptosyl building blocks. Immunization of mice with the semisynthetic glycoconjugate elicited a very robust T-cell-dependent antigen-specific immune response, resulting in very high titers of IgG1 and IgG2b protective antibody isotypes. The postimmune sera recognized H. pylori NCTC 11637 and bound strongly to the surface of the intact bacteria.

Characterization of Plasmid-Mediated Quinolone Resistance and Serogroup Distributions of Uropathogenic Escherichia coli among Iranian Kidney Transplant Patients.

Introduction Urinary tract infection (UTI) is one of the most frequent infections in kidney transplant patients (KTPs). This infection is mainly caused by uropathogenic Escherichia coli (UPEC). Plasmid-mediated quinolone resistance (PMQR) was also increasingly identified in UPEC. This study proposed to investigate the frequency of quinolone-resistance plasmid genes and the O-antigen serogroup among UPEC isolated from KTPs and non-KTP with UTI. Methods Totally, 114 UPEC isolates from 49 KTPs and 65 non-KTPs patients diagnosed with an UPEC-associated UTI were obtained from June 2019 to December 2019 at three laboratory centers in Isfahan, Iran. The isolates were confirmed through phenotypic and genotypic methods. Moreover, the antimicrobial susceptibility test to nalidixic acid, ciprofloxacin, norfloxacin, and ofloxacin was performed using a disk diffusion method. The presence of the qnr gene as well as the serogroup distribution was identified using the PCR method. Result According to data, the distribution of O1, O2, O4, O16, and O25 serogroups were 3.5%, 2.6, 3.5, 3.5, and 20.2%, respectively. Antibiotic susceptibility pattern revealed that the highest and lowest resistance rates were to nalidixic acid (69.3%) and norfloxacin (43.9%), respectively. Also, the frequency of qnrS and qnrB genes were 33.3% and 15.8%, respectively, while none of the isolates was found to be positive for the qnrA gene. There was no significant association between the presence of qnr genes and higher antibiotic resistance. Conclusion This study recognized that the qnrS gene, O25 serotype, and resistance to nalidixic acid had the highest frequencies in UPEC strains isolated from UTI patients.

Pheno- and Genotypical Features of Non-Toxigenic Strains of Cholera Vibrios of Different Origins, Isolated in the Territory of Russia

Aim. Analysis of the phenotypic characteristics and identification of peculiarities of the genotypic organization in non-toxigenic strains of cholera vibrios having different origin, isolated in Russia. Materials and methods. A sample of 548 non-toxigenic strains obtained using the author’s updated GIS “Cholera 1989–2014” was used. PCR genotyping was carried out in accordance with the patented “Method for the identification of non-toxigenic strains of cholera vibrio O1 serogroup using PCR to isolate genetic determinants.” Cluster analysis was performed applying the UPGMA method. The dendrogram was constructed using MEGA 5 software package.Results and discussion. Representative cultural-morphological, serological and biochemical properties of V. cholerae strains have been specified. The variability of the studied strains on the basis of phagolizability has been revealed. Unique phage-types not previously encountered in Russia have been identified. The population of non-toxigenic strains of cholera vibrio O139 serogroup is genetically homogeneous in contrast to V. cholerae O1 El Tor isolates and has identical PCR genotypes. The universality of the PCR genotyping by 14 target genes has been shown to differentiate the studied strains of V. cholerae O1 and O139, as well as to identify disparities among O139 strains isolated in different geographical regions of the country.

Genetic relation and virulence factors of carbapenemase-producing Uropathogenic Escherichia coli from urinary tract infections in Iraq

BackgroundUropathogenic Escherichia coli (E. coli) or UPEC is among major nosocomial pathogens causing urinary tract infections (UTIs). The emergence of carbapenem- resistant strains is a major concern regarding the UTI treatment. The subjective of this study included assessment of genetic relation and screening of virulence factors among carbapenemase- producing E. coli from UTI. MethodsThree-hundred E. coli isolates were collected from cystitis and pyelonephritis. Antibiotic susceptibility test was conducted by disk diffusion as provided by clinical and laboratory standards institute (CLSI) version 2017. Carbapenemase production and related genes and virulence factors were screened by polymerase chain reaction (PCR) technique. Genetic relation of isolates was evaluated using phylogrouping and serogrouping. ResultsOf 300 isolates, 11 (3.66%) of them were carbapenemase-producing E. coli (CP-E. coli). Imipenem minimum inhibitory concentration ranged 4-128 μg/ml. The blaOXA-48 and blaIMP genes were co-existed in three (1%) isolates (imipenem MIC: 64-128 μg/ml), but the blaKPC, blaNDM and blaVIM were not amplified. Predominant virulence genes included iutA (n = 293, 97.66%), fyuA (n = 256, 85.33%), inh (n = 249, 83%), traT (n = 247, 82.33%), papП (n = 96, 32%), fimH (n = 93, 31%), csgA (n = 92, 30.66%). All the CP-E. coli contained the iutA, fyuA, traT, papII, fimH and csgA genes. O1 (32%), O16 (15%) and O25 (7%) serogroups were predominant and 6/11, 4/11 and 1/11 of CP-E. coli belonged to O1, O25 and O75 serogroups, respectively. Among eleven CP-E. coli isolates, nine of them were belonged to the B2 phylogroup and two isolates were belonged to B1 phylogroup. ConclusionCP-E. coli (or CP-UPEC) contained the blaIMP and blaOXA-48 genes and belonged to O25/B2. High rate of virulence factors among CP-E. coli from UTI is a concern in Baghdad hospitals. The spread of isolates with resistance to last-line antibiotics must be controlled.

Serogrouping, phylotyping, and virulence genotyping of commensal and avian pathogenic Escherichia coli isolated from broilers in Hamedan, Iran

In the present study, 100 Avian-Pathogenic Escherichia coli (APEC) isolates from colibacillosis-suspected broilers and 100 Avian Faecal Escherichia coli (AFEC) isolates from healthy broilers in Iran were examined by PCR for confirmation of their serogroups and phylogenetic background, and their association with ten virulence-associated genes (VAG) including fimC, iutA, chuA, sitA, iss, cvaA/B, hylA, stx1, stx2, and yjaA. Serogroups O78, O1, O2 and O18 were the prominent strains including 54 % of the APEC and 23 % of the AFEC strains. At phylotyping, the majority of APEC strains belonged to phylogenetic group E (22 %) while for the AFEC strains, half of the isolates were not assigned to any group but the predominant phylogroup was E (27 %). Virulence genotyping, revealed that the predominant VAGs were iutA (97 %), fimC (87 %) and iss (84 %) among APEC strains, and fimC (95 %), iss (93 %) and sitA (87 %) in AFEC strains. This is the first time that phylogroup E is described as predominant phylogroup among APEC strains also, this is the first report on the presence of the stx1 gene in APEC strains isolated from broilers in Iran. The results of the present study indicate that VAGs are more prevalent in APEC strains belonging to O2 and O78 serogroups, also phylogroups E and D have more frequency of VAGs than other phylogroups. Therefore, the APEC strains belonging to O2 and O78 serogroups and phylogroups E and D probably have more pathogenicity to broilers.

Cronobacter spp. isolated from aquatic products in China: Incidence, antibiotic resistance, molecular characteristic and CRISPR diversity

Cronobacter species (Cronobacter spp.) are important foodborne pathogens that can infect and cause serious life-threatening diseases in infants and immunocompromised elderly. This study aimed to acquire data on Cronobacter spp. contamination of aquatic products in China from 2011 to 2016. In total, 800 aquatic products were tested, and the overall contamination rate for Cronobacter spp. was 3.9% (31/800). The average contamination level of the positive samples was 2.05 MPN/g. Four species and nine serotypes were identified among 33 isolates, of which the C. sakazakii serogroup O1 (n = 9) was the primary serotype. The majority of Cronobacter spp. strains harbored highest resistance against cephalothin (84.8%), followed by tetracycline (6.1%), trimethoprim/sulfameth-oxazole (3.0%) and chloramphenicol (3.0%). Two isolates were resistant to three antibiotics. In total, 26 sequence types and 33 CRISPR types (including 6 new STs and 26 new CTs) were identified, which indicates the extremely high diversity of Cronobacter spp. in aquatic products. Pathogenic C. sakazakii ST4, ST1, and C. malonaticus ST7 were also observed. Overall, this large-scale study revealed the relatively low prevalence and high genetic diversity of Cronobacter spp. in aquatic products in China, and the findings provide valuable information that can guide the establishment of effective measures for the control and precaution of Cronobacter spp. in aquatic products during production processes.

Characterization of Vibrio cholerae isolates from freshwater sources in northwest Ohio.

Vibrio cholerae is a natural inhabitant of aquatic ecosystems worldwide, typically residing in coastal or brackish water. While more than 200 serogroups have been identified, only serogroups O1 and O139 have been associated with epidemic cholera. However, infections other than cholera can be caused by nonepidemic, non-O1/non-O139 V. cholerae strains, including gastroenteritis and extraintestinal infections. While V. cholerae can also survive in freshwater, that is typically only observed in regions of the world where cholera is endemic. We recently isolated V. cholerae from several locations in lakes and rivers in northwest Ohio. These isolates were all found to be non-O1/non-O139 V. cholerae strains, that would not cause cholera. However, these isolates contained a variety of virulence genes, including ctxA, rtxA, rtxC, hlyA, and ompU. Therefore, it is possible that some of these isolates have the potential to cause gastroenteritis or other infections in humans. We also investigated the relative motility of the isolates and their ability to form biofilms as this is important for V. cholerae survival in the environment. We identified one isolate that forms very robust biofilms, up to 4x that of our laboratory strains. Finally, we investigated the susceptibility of these isolates to a panel of antibiotics. We found that many of the isolates showed decreased susceptibility to some of the antibiotics tested, which could be of concern. While we do not know if these isolates are pathogenic to humans, increased surveillance to better understand the public health risk to the local community should be considered.

Generation and Characterization of Monoclonal Antibodies to the Ogawa Lipopolysaccharide of Vibrio cholerae O1 from Phage-Displayed Human Synthetic Fab Library.

Vibrio cholerae, cause of the life-threatening diarrheal disease cholera, can be divided into different serogroups based on the structure of its lipopolysaccharide (LPS), which consists of lipid-A, corepolysaccharide and O-antigen polysaccharide (O-PS). The O1 serogroup, the predominant cause of cholera, includes two major serotypes, Inaba and Ogawa. These serotypes are differentiated by the presence of a single 2-O-methyl group in the upstream terminal perosamine of the Ogawa O-PS, which is absent in the Inaba O-PS. To ensure the consistent quality and efficacy of the current cholera vaccines, accurate measurement and characterization of each of these two serotypes is highly important. In this study, we efficiently screened a phage-displayed human synthetic Fab library by bio-panning against Ogawa LPS and finally selected three unique mAbs (D9, E11, and F7) that specifically react with Ogawa LPS. The mAbs bound to Vibrio cholerae vaccine in a dose-dependent fashion. Sequence and structure analyses of antibody paratopes suggest that IgG D9 might have the same fine specificity as that of the murine mAbs, which were shown to bind to the upstream terminal perosamine of Ogawa O-PS, whereas IgGs F7 and E11 showed some different characteristics in the paratopes. To our knowledge, this study is the first to demonstrate the generation of Ogawa-specific mAbs using phage display technology. The mAbs will be useful for identification and quantification of Ogawa LPS in multivalent V. cholerae vaccines.

A new plasmid carrying mphA causes prevalence of azithromycin resistance in enterotoxigenic Escherichia coli serogroup O6

BackgroundAt present, azithromycin has become an effective treatment for severe diarrhea caused by Enterotoxigenic Escherichia coli (ETEC) infection. However, enterobacteria have begun to develop resistance to azithromycin and have attracted attention in recent years. This study conducted to described the emergence of a high proportion of azithromycin-resistant ETEC serogroup O6 strains in Shanghai and to analyzed the mechanisms of azithromycin resistance.ResultsStrains from adult diarrhea patients with ETEC serogroup O6 infections were collected by Shanghai Diarrhea Surveillance Network and the Foodborne Surveillance Network from 2016 to 2018. We tested 30 isolates of ETEC O6 serogroup, 26 of which were resistant to azithromycin. Phylogenetic analysis revealed that these ETEC serogroup O6 strains have formed an independent dominant clone. S1-PFGE and southern blotting revealed the presence of the mphA gene on the 103 kb plasmid. Illumina and Nanopore sequencing and plasmid coverage analysis further confirmed that azithromycin-resistant strains carried a novel IncFII plasmid harboring mphA and blaTEM-1 resistance genes.ConclusionsThis is the first study to report a high proportion of azithromycin resistance in a particular ETEC serogroup due to a specific plasmid carrying mphA. Our findings indicate the rapid spread of azithromycin resistance, highlighting the urgency of stringent surveillance and control measure.