Serogroup O1 Research Articles

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.

Age related differences in phylogenetic diversity, prevalence of Shiga toxins, Intimin, Hemolysin genes and select serogroups of Escherichia. coli from pastured meat goats detected in a longitudinal cohort study

BackgroundLittle is known on significance, diversity and characteristics of gut E. coli in goats despite their importance as food animals globally. We characterized the temporal dynamics in diversity of E. coli in fecal samples from a cohort of goat kids and adult meat goats on pasture over a one-year period. Isolates were characterized based on phylogenetic grouping, virulence genes; shiga toxins 1 and 2 (Stx1&Stx2) (STEC), intimin (eaeA), hemolysin (hly) and select important sero-groups (026, 045, 0103, 0126 and 0146) using molecular methods.ResultsA total of 516 E. coli isolates were screened. Prevalence of virulence genes and STEC was 65 and 56% respectively. Prevalence of virulence genes and STEC was significantly higher in goat kids less than six months (76% /66%) than adults (48% /28%). Isolates with virulence profiles of two or more genes were also higher in young goat kids (50%) than adults (20%). Entero-pathogenic E. coli (EPEC-eaeA gene only) were mostly from pre-weaned goat kids while hly gene only isolates were significantly higher in adults. The stx1, stx2 and hly genes peaked around weaning (60, 63 and 52%) respectively. Goats kids were mostly hosts to group D (59%) while adults older than one year had B1 (75%) isolates. Group D isolates were most abundant at weaning (64%) and diarrhea samples (74%). Group B2 isolates overall (6%) were mostly detected around weaning (63%) while A isolates were 4% overall. Twenty-four isolates belonged to sero-groups 026, 0103 and 0146 with 70% of the isolates detected around weaning. Nineteen of these isolates were STEC with most harboring the stx1/stx2/hly/eae (25%) profile. Most belonged to O26 sero-group (75%) and phylogroup D (75%).ConclusionTo our knowledge this is the first study to highlight longitudinal age related differences in E. coli phylogenetic diversity, abundance of virulence genes and select important sero-groups in goats. Differences detected suggest a possible role of age and weaning stress in influencing E. coli diversity in the gut of goats. The findings are relevant to both animal and public health to advise on further studies on caprine E. coli isolates as animal and human pathogens.

Cholera: Monitoring of Epidemiological Situation around the World and in Russia (2010–2019). Forecast for 2020

Objectiveof the study was to monitor the spread of cholera in the world, in the CIS countries and in Russia with an assessment of risks and emergencies that contribute to the activation of the epidemic process. Despite the downward trend in the global incidence rate of cholera during the period between 2010 and 2019, epidemics and major outbreaks occurred in 96 countries. WHO has reported 2013 imported cases of cholera to countries in Asia, the Americas, including the Caribbean, Europe, and Australia with Oceania; cholera-endemic administrative territories are identified in 24 countries. According to WHO, cholera burden reduction in Asia and Africa is associated with large-scale vaccination. During epidemiological surveillance of cholera, 705 strains ofV. choleraeO1 and O139 serogroups were isolated from surface reservoirs in 26 constituent entities of the Russian Federation, including 10 strains of V. cholerae O1 ctxA+tcpA+, 35 strains ofV. choleraeO1ctxA–tcpA+, 655 strains ofV. cholerae O1 ctxA–tcpA–, and five strains ofV. choleraeO139ctxA–tcpA–. Identification of strains with unique, previously unknown INDEL genotypes testifies to their imported nature. The forecast for cholera in the world for 2020, given the proven high degree of epidemic process activation at the expense of social and environmental risks caused by emergencies of different origin, the presence of endemic foci, infection import and other risk factors is unfavorable. For Russia, the forecast for cholera will be determined by the presence of external risks created by the ongoing 7-th pandemic, possible importation of infection to constituent entities of the Russian Federation that differ by the types of epidemic manifestations.

A molecular and epidemiological study of Vibrio cholerae isolates from cholera outbreaks in southern Ghana.

Cholera remains a major global public health threat and continuous emergence of new Vibrio cholerae strains is of major concern. We conducted a molecular epidemiological study to detect virulence markers and antimicrobial resistance patterns of V. cholerae isolates obtained from the 2012–2015 cholera outbreaks in Ghana. Archived clinical isolates obtained from the 2012, 2014 and 2015 cholera outbreaks in Ghana were revived by culture and subjected to microscopy, biochemical identification, serotyping, antibiotic susceptibility testing, molecular detection of distinct virulence factors and Multi-Locus Variable-Number of Tandem-Repeat Analysis (MLVA). Of 277 isolates analysed, 168 (60.6%) were confirmed to be V. cholerae and 109 (39.4%) isolates constituted other bacteria (Escherichia coli, Aeromonas sobria, Pseudomonas aeruginosa, Enterobacter cloacae and Enterococci faecalis). Serotyping the V. cholerae isolates identified 151 (89.9%) as Ogawa, 3 (1.8%) as Inaba and 14 (8.3%) as non-O1/O139 serogroup. The O1 serogroup isolates (154/168, 91.7%) carried the cholera toxin ctxB gene as detected by PCR. Additional virulence genes detected include zot, tcpA, ace, rtxC, toxR, rtxA, tcpP, hlyA and tagA. The most common and rare virulence factors detected among the isolates were rtxC (165 isolates) and tcpP (50 isolates) respectively. All isolates from 2014 and 2015 were multidrug resistant against the selected antibiotics. MLVA differentiated the isolates into 2 large unique clones A and B, with each predominating in a particular year. Spatial analysis showed clustering of most isolates at Ablekuma sub-district. Identification of several virulence genes among the two different genotypes of V. cholerae isolates and resistance to first- and second-line antibiotics, calls for scaleup of preventive strategies to reduce transmission, and strengthening of public health laboratories for rapid antimicrobial susceptibility testing to guide accurate treatment. Our findings support the current WHO licensed cholera vaccines which include both O1 Inaba and Ogawa serotypes.

Immunogenicity and protective efficacy of a live, oral cholera vaccine formulation stored outside-the-cold-chain for 140\u2009days

BackgroundCholera, an acute watery diarrhoeal disease caused by Vibrio cholerae serogroup O1 and O139 across the continents. Replacing the existing WHO licensed killed multiple-dose oral cholera vaccines that demand ‘cold chain supply’ at 2–8 °C with a live, single-dose and cold chain-free vaccine would relieve the significant bottlenecks and cost determinants in cholera vaccination campaigns. In this direction, a prototype cold chain-free live attenuated cholera vaccine formulation (LACV) was developed against the toxigenic wild-type (WT) V. cholerae O139 serogroup. LACV was found stable and retained its viability (5 × 106 CFU/mL), purity and potency at room temperature (25 °C ± 2 °C, and 60% ± 5% relative humidity) for 140 days in contrast to all the existing WHO licensed cold-chain supply (2–8 °C) dependent killed oral cholera vaccines.ResultsThe LACV was evaluated for its colonization potential, reactogenicity, immunogenicity and protective efficacy in animal models after its storage at room temperature for 140 days. In suckling mice colonization assay, the LACV recorded the highest recovery of (7.2 × 107 CFU/mL) compared to those of unformulated VCUSM14P (5.6 × 107 CFU/mL) and the WT O139 strain (3.5 × 107 CFU/mL). The LACV showed no reactogenicity even at an inoculation dose of 104–106 CFU/mL in a rabbit ileal loop model. The rabbits vaccinated with the LACV or unformulated VCUSM14P survived a challenge with WT O139 and showed no signs of diarrhoea or death in the reversible intestinal tie adult rabbit diarrhoea (RITARD) model. Vaccinated rabbits recorded a 275-fold increase in anti-CT IgG and a 15-fold increase in anti-CT IgA antibodies compared to those of rabbits vaccinated with unformulated VCUSM14P. Vibriocidal antibodies were increased by 31-fold with the LACV and 14-fold with unformulated VCUSM14P.ConclusionThe vaccine formulation mimics a natural infection, is non-reactogenic and highly immunogenic in vivo and protects animals from lethal wild-type V. cholerae O139 challenge. The single dose LACV formulation was found to be stable at room temperature (25 ± 2 °C) for 140 days and it would result in significant cost savings during mass cholera vaccination campaigns.

Cholera Toxin Production in Vibrio cholerae O1 El Tor Biotype Strains in Single-Phase Culture.

Vibrio cholerae O1 serogroup strains have been classified into classical and El Tor biotypes. Cholera, a life-threatening diarrheal disease, can be caused by either biotype through the cholera toxin (CT) that they produce. To increase our knowledge of the pathogenicity of bacteria, we must understand the toxigenicity of bacteria. CT production by classical biotype strains in simple single-phase cell cultures has been established; however, special culture media and growth conditions that are not appropriate for mass production of CT are required to facilitate CT production in El Tor biotype strains. In this report, we produced CT in El Tor biotype strains using simple media and single-phase culture conditions. A single point mutation in ToxT, a transcriptional activator of toxin co-regulated pilus (TCP) and CT, enabled the El Tor biotype strains to produce CT in similar quantities as classical biotype strains in single-phase laboratory culture conditions. CT production capacity varied between El Tor biotype strains. Wave 2 and 3 atypical El Tor strains tended to produce more CT than prototype Wave 1 strains. Wave 2 and 3 strains lack neutral fermentation; however, the capacity for neutral fermentation was not associated with significant differences in CT production by El Tor biotype strains. The Wave 3 strain that caused the 2010 cholera outbreak in Haiti produced CT only when neutral fermentation was abolished. The disparity in CT production between the seventh cholera pandemic strains highlight the differences in virulence between strains and the cause of population changes in V. cholerae.

Nonmetabolizable Arabinose Inhibits Vibrio cholerae Growth in M9 Medium with Gluconate as the Sole Carbon Source.

The serogroups O1 and O139 of the marine bacterium Vibrio cholerae are responsible for causing cholera in humans. The pentose sugar arabinose is nonmetabolizable by the pathogen and is present in environmental niches as well as in the human intestine. In this study, arabinose-mediated V. cholerae growth interference was assessed in M9 minimal medium containing gluconate as the sole carbon source in the light of Entner-Doudoroff (ED) pathway, an obligatory metabolic route for gluconate utilization. V. cholerae O1 and O139 strains failed to grow in the presence of ≥ 0.3% arabinose in M9 with 0.2% gluconate, but there was no growth inhibition in the presence of arabinose in M9 with 0.2% glucose. Transcriptional analysis of edd and eda, the genes constituting the ED pathway, showed ~100- and ~17-fold increases, respectively, in M9-gluconate. Minor increases of ~4- and ~2-fold for edd and eda, respectively, were noted in AKI medium supplemented with 0.5% arabinose. The observed arabinose-mediated growth inhibition can contribute toward deepening the understanding of altered phenotypes, if any, via complementation/expression studies in V. cholerae with pBAD vectors and arabinose as an inducer.

Genomic characterization of antibiotic resistance-encoding genes in clinical isolates of Vibrio cholerae non-O1/non-O139 strains from Kolkata, India: generation of novel types of genomic islands containing plural antibiotic resistance genes.

Non-O1/non-O139 nontoxigenic Vibrio cholerae associated with cholera-like diarrhea has been reported in Kolkata, India. However, the property involved in the pathogenicity of these strains has remained unclear. The character of 25 non-O1/non-O139 nontoxigenic V. cholerae isolated during 8 years from 2007 to 2014 in Kolkata was examined. Determination of the serogroup showed that the serogroups O6, O10, O35, O36, O39, and O70 were represented by two strains in each serogroup, and the remaining isolates belonged to different serogroups. To clarify the character of antibiotic resistance of these isolates, an antibiotic resistance test and the gene analysis were performed. According to antimicrobial drug susceptibility testing, 13 strains were classified as drug resistant. Among them, 10 strains were quinolone resistant and 6 of the 13 strains were resistant to more than three antibiotics. To define the genetic background of the antibiotic character of these strains, whole-genome sequences of these strains were determined. From the analysis of these sequences, it becomes clear that all quinolone resistance isolates have mutations in quinolone resistance-determining regions. Further research on the genome sequence showed that four strains possess Class 1 integrons in their genomes, and that three of the four integrons are found to be located in their genomic islands. These genomic islands are novel types. This indicates that various integrons containing drug resistance genes are spreading among V. cholerae non-O1/non-O139 strains through the action of newly generated genomic islands.

Deciphering the possible role of ctxB7 allele on higher production of cholera toxin by Haitian variant Vibrio cholerae O1.

Cholera continues to be an important public health concern in developing countries where proper hygiene and sanitation are compromised. This severe diarrheal disease is caused by the Gram-negative pathogen Vibrio cholerae belonging to serogroups O1 and O139. Cholera toxin (CT) is the prime virulence factor and is directly responsible for the disease manifestation. The ctxB gene encodes cholera toxin B subunit (CTB) whereas the A subunit (CTA) is the product of ctxA gene. Enzymatic action of CT depends on binding of B pentamers to the lipid-based receptor ganglioside GM1. In recent years, emergence of V. cholerae Haitian variant strains with ctxB7 allele and their rapid spread throughout the globe has been linked to various cholera outbreaks in Africa and Asia. These strains produce classical type (WT) CTB except for an additional mutation in the signal sequence region where an asparagine (N) residue replaces a histidine (H) at the 20th amino acid position (H20N) of CTB precursor (pre-CTB). Here we report that Haitian variant V. cholerae O1 strains isolated in Kolkata produced higher amount of CT compared to contemporary O1 El Tor variant strains under in vitro virulence inducing conditions. We observed that the ctxB7 allele, itself plays a pivotal role in higher CT production. Based on our in silico analysis, we hypothesized that higher accumulation of toxin subunits from ctxB7 allele might be attributed to the structural alteration at the CTB signal peptide region of pre-H20N CTB. Overall, this study provides plausible explanation regarding the hypertoxigenic phenotype of the Haitian variant strains which have spread globally, possibly through positive selection for increased pathogenic traits.

An outbreak of diarrhoeal disease of El Tor Vibrio cholerae O1 Ogawa in and around Yavatmal district, Maharashtra, India in 2018

Background: Epidemics of cholera have been reported from various parts of India. We investigated the epidemic of cholera that occurred in and around Yavatmal district in Maharashtra, India 2018, reported during March to July.Methods: 711 stool samples collected from diarrhea patients were bacteriologically analyzed for their identification and antibiogram of Vibrio cholera.Results: The cholera outbreak was caused by V. cholerae O1 Ogawa biotype El Tor. All the V. cholerae isolates from the stool samples were sensitive to tetracycline, doxycycline, ofloxacin, ciprofloxacin gentamycin, cotrimoxazole and resistant to ampicillin and ceftazidime.Conclusions: The present outbreak was due to V. cholerae O1 Ogawa El Tor which seems to have completely replaced O139 serogroup of the previous outbreaks during the last decade.

Genomic recombination of the vibrio cholerae serogroup O1 El Tor pandemic strains

Objective: To analyze the genomic recombination of the vibrio cholerae serogroup O1 El Tor pandemic strains. Methods: A total of 292 complete or draft genome sequences of Vibrio cholerae O1 serogroup El Tor strains isolated from 1937 to 2015 were selected from National Biotechnology Information Center database. The genome alignment of strains was computed by snippy software by using N16961 as reference sequence. Then ClonalFrameML software was used to do the recombinant analysis. The wilcox.test function in agricolae package was used to compare the number recombinant segments and the total length of recombinant regions between small and large chromosomes. The kruskal function was used to compare the number recombinant segments and the total length of recombinant regions among different isolation continents. The KOBAS tool was used to do the gene ontology enrichment analysis of recombinant hotspot genes. Results: Of all 292 strains of Vibrio cholerae, 163 strains (55.8%) were recombined. The median of normalized recombinant segment number of small chromosome was 4.7×10(-6) (9.3×10(-7), 2.0×10(-5)), which was significantly larger than that of large chromosome [2.4×10(-6) (3.4×10(-7), 5.7×10(-6))] (P<0.001). The median (P(25),P(75)) of recombinant segment number of strains isolated from Africa, Asia, Europe, North America and South America were 23(1.0,33.0), 1.0(0.0,34.0), 6.0(2.0,13.0), 0.0(0.0,1.0) and 29.5(6.8,56.8), respectively, and the difference was statistically significant (P<0.001). The median (P(25),P(75)) of total length of recombinant regions of strains isolated from Africa, Asia, Europe, North America and South America were 233.0(4.0, 461.0), 11.0(0.0, 695.5), 56.0(4.0,111.0), 0.0(0.0,9.0) and 347.5(132.8,1 323.5) bp, respectively, and the difference was statistically significant (P<0.001). Gene ontology Enrichment analysis showed that the functions of 62 recombinant hotspot genes were mainly enrichment in chemotaxis, taxis, response to external stimulus, receptor activity and molecular transducer activity. Conclustion: In this study, we found that there were significant differences in the number of recombinant fragments and the length of recombinant regions between large and small chromosomes of Vibrio cholerae El Tor. We also found significant differences in the number of recombinant fragments and the total length of recombinant regions among different continents.

Genomic characterization of the non-O1/non-O139 Vibrio cholerae strain that caused a gastroenteritis outbreak in Santiago, Chile, 2018.

Vibrio cholerae is a human pathogen, which is transmitted by the consumption of contaminated food or water. V. cholerae strains belonging to the serogroups O1 and O139 can cause cholera outbreaks and epidemics, a severe life-threatening diarrheal disease. In contrast, serogroups other than O1 and O139, denominated as non-O1/non-O139, have been mainly associated with sporadic cases of moderate or mild diarrhea, bacteremia and wound infections. Here we investigated the virulence determinants and phylogenetic origin of a non-O1/non-O139 V. cholerae strain that caused a gastroenteritis outbreak in Santiago, Chile, 2018. We found that this outbreak strain lacks the classical virulence genes harboured by O1 and O139 strains, including the cholera toxin (CT) and the toxin-coregulated pilus (TCP). However, this strain carries genomic islands (GIs) encoding Type III and Type VI secretion systems (T3SS/T6SS) and antibiotic resistance genes. Moreover, we found these GIs are wide distributed among several lineages of non-O1/non-O139 strains. Our results suggest that the acquisition of these GIs may enhance the virulence of non-O1/non-O139 strains that lack the CT and TCP-encoding genes. Our results highlight the pathogenic potential of these V. cholerae strains.

Genomic epidemiology of Vibrio cholerae reveals the regional and global spread of two epidemic non-toxigenic lineages.

Non-toxigenic Vibrio cholerae isolates have been found associated with diarrheal disease globally, however, the global picture of non-toxigenic infections is largely unknown. Among non-toxigenic V. cholerae, ctxAB negative, tcpA positive (CNTP) isolates have the highest risk of disease. From 2001 to 2012, 71 infectious diarrhea cases were reported in Hangzhou, China, caused by CNTP serogroup O1 isolates. We sequenced 119 V. cholerae genomes isolated from patients, carriers and the environment in Hangzhou between 2001 and 2012, and compared them with 850 publicly available global isolates. We found that CNTP isolates from Hangzhou belonged to two distinctive lineages, named L3b and L9. Both lineages caused disease over a long time period with usually mild or moderate clinical symptoms. Within Hangzhou, the spread route of the L3b lineage was apparently from rural to urban areas, with aquatic food products being the most likely medium. Both lineages had been previously reported as causing local endemic disease in Latin America, but here we show that global spread of them has occurred, with the most likely origin of L3b lineage being in Central Asia. The L3b lineage has spread to China on at least three occasions. Other spread events, including from China to Thailand and to Latin America were also observed. We fill the missing links in the global spread of the two non-toxigenic serogroup O1 V. cholerae lineages that can cause human infection. The results are important for the design of future disease control strategies: surveillance of V. cholerae should not be limited to ctxAB positive strains.

A novel pre-CTX prophage in the Vibrio cholerae serogroup O139 strain

In Vibrio cholerae, the lysogenic bacteriophage CTXΦ carries the cholera toxin genes ctxAB, which can be transferred from toxigenic strains to nontoxigenic strains through infection and lysogenic conversion of CTXΦ. This phage also has the precursor genome which does not harbor ctxAB, named pre-CTXΦ. Based on the sequences of the transcriptional regulator-encoding gene rstR alleles in CTXΦ/pre-CTXΦ, multiple types of these prophages have been classified and identified in toxigenic and nontoxigenic V. cholerae strains. In this study, by combining the short-read and long-read sequencing approaches of next generation sequencing, we obtained the complete genome sequence of the studied V. cholerae toxigenic serogroup O139 strain and identified the CTXΦ and a pre-CTXΦ genome type encoding a novel rstR allele, pre-CTXZHJΦ. This pre-CTX prophage integrates into the small chromosome of the V. cholerae host strain and coexists with a typical CTXETΦ prophage present in the large chromosome, which is commonly present in the seventh pandemic serogroup O1 and toxigenic serogroup O139 strains. RstRZHJ could bind to the ig-2 region in the RstAB promotor in the pre-CTXZHJΦ genome, and could repress the expression of its own rstAB genes but could not repress rstAB expression in CTXETΦ and CTXclassΦ, suggesting that the V. cholerae strains carrying the pre-CTXZHJΦ prophage cannot prevent the infection of these epidemic CTXΦs, hence have the potentiality to become toxigenic by acquiring and lysogenic conversion of CTXΦs. Our study identified a novel pre-CTXΦ type, and presents the new evidence for the complexity and diversity of the CTXΦ/pre-CTXΦ family in V. cholerae.

Prevalence, genetic analysis and CRISPR typing of Cronobacter spp. isolated from meat and meat products in China

Cronobacter spp. are important foodborne pathogens that infections occur in all age groups, especially cause serious life-threatening diseases in infants. This study aimed to acquire data on Cronobacter spp. contamination of meat and meat products (n = 588) in China during 2011 to 2016, and investigated the use of CRISPR typing technology as an approach for characterizing the genetics of Cronobacter spp. The overall contamination rate for Cronobacter spp. was determined to be 9.18% (54/588). Of the positive samples, 90.74% (49/54) had <10 MPN/g, with duck samples had a relatively high contamination rate (15.69%, 8/51) and highest contamination level (28.90 MPN/g). Four species and nine serotypes were identified among 69 isolates, of which C. sakazakii was the major species (n = 50) and C. sakazakii serogroup O1 and O2 (n = 17) were the primary serotypes. The majority of Cronobacter spp. strains were found to be susceptible to most antibiotics except exhibited high resistance to cephalothin (76.81%, 53/69), and total two multi-drug resistant C. sakazakii strains were isolated from duck. The genetic diversity of Cronobacter spp. was remarkably high, as evidenced by the identification of 40 sequence types (STs) and 60 CRISPR types (CTs). C. sakazakii ST64 (n = 7) was the predominant genotype and was further divided into two sub-lineages based on CRISPR diversity, showing different antibiotic resistance profile. These results demonstrate that CRISPR typing results have a good correspondence with bacterial phenotypes, and it will be a tremendously useful approach for elucidating inter-subtyping during molecular epidemiological investigations while interpreting the divergent evolution of Cronobacter. The presence of Cronobacter spp. in meat and meat product is a potential threat to human public health.

Construction of Polymeric Antigenic Diagnosticum Based on &lt;i&gt;Vibrio cholera&lt;/i&gt; О1 Lipopolysaccharide

Representatives of the genus Vibrio cholerae differ in the structure of lipopolysaccharide, in particular, its O-polysaccharide chains (O-antigen), which determines the serological specificity of vibrios. Currently, the water-phenolic method is used to obtain the lipopolysaccharide preparation. However, this technique relates to harsh chemical methods, leads to a change in original molecular organization of biopolymer, violating its structure and biological properties. Modern technologies in the development of diagnostic kits for the immunosuspension reaction of volume agglomeration allow for obtaining synthetic carriers with different reaction groups on the particle surface capable to bind antigens/antibodies. The aim of this study was to construct cholera antigenic polymeric diagnostic kit based on the lipopolysaccharide of Vibrio cholerae O1 serogroup. Materials and methods . The lipopolysaccharide was used as a sensitizer obtained through the author's modification of enzymatic purification from the cell membranes of Vibrio cholerae using ultrasonic disintegration. Results and discussion . The resulting sensitin contains small impurities of protein (1.5 %) and nucleic acids (0.1 %). Diagnosticum is characterized by high analytical sensitivity in agglomeration reaction with commercial and experimental rabbit serum to Vibrio cholerae O1 serogroup (1:640 - 1:5120) and analytical specificity (the diagnosticum does not interact with heterologous sera, with serums to pathogens of acute intestinal infections, as well as with sera from healthy donors). A polymeric antigenic cholera diagnosticum designed to detect antibodies to lipopolysaccharide of Vibrio cholerae in the blood serum of patients who were ill, suspected of the disease or vaccinated people has been constructed.

Alterations in glucose metabolism in Vibrio cholerae serogroup O1 El Tor biotype strains

The 2 biotypes of Vibrio cholerae O1 serogroup strains—classical and El Tor—use glucose in distinct ways. Classical biotype strains perform organic acid-producing fermentation and eventually lose viability due to the self-induced creation of an acidic environment, whereas El Tor biotype strains use an alternative neutral fermentation pathway, which confers them with survival advantages. However, we report that the neutral fermentation pathway has only been recruited in prototype Wave 1 El Tor biotype strains, which have not been isolated since the mid-1990s. Current Wave 2 and Wave 3 atypical El Tor strains contain a single-base deletion in a gene that directs bacteria toward neutral fermentation, resulting in the loss of neutral fermentation and an appearance that is similar to classical biotype strains. Moreover, when sufficient glucose was supplied, Wave 1 El Tor strains maintained their use of acid-producing fermentation, in parallel with neutral fermentation, and thus lost viability in the late stationary phase. The global replacement of Wave 1 El Tor strains by Wave 2 and 3 atypical El Tor strains implies that the acidic fermentation pathway may not be disadvantageous to V. cholerae. The characteristics that we have reported might improve oral rehydration in the treatment of cholera.

Phenotypic Pattern of Vibrio choleraeIsolates from a Tertiary Care Hospital in Vadodara, Gujarat, India

Introduction: Cholera is an acute diarrhoeal disease caused by Vibrio cholerae (V.cholerae). Based on antigenic differences of O antigen, O1 serogroup can be divided into three serotypes. In addition, by performing various biochemical reactions, O1 Serogroup can be differentiated into two biotypes. Outbreaks of Cholera occur seasonally. It is associated with monsoon season, warm temperature, heavy rainfall and increased plankton population. Aim: The aim was to determine the trends in resistance pattern and phenotypic Pattern of Vibrio cholerae. Materials and Methods: A retrospective study was conducted during the period from June 2019-December 2019. Culture of Stool specimens were done on different agar media. Biotyping was done by conventional methods. Serotyping and phage typing was also done along with the Antibiotic susceptibility testing. Descriptive analysis was used and presented in terms of percentage. Results:V.cholerae was isolated in 72 patients and they belonged to serogroup O1 and biotype El Tor. The most common serotype was Ogawa. The predominant phage type were T2 by old scheme and T27 by new scheme of phage typing. The maximum number of V. cholerae isolates was seen in the month of November, 2019 followed by October, 2019. Conclusion: The phenotypic pattern and fluctuating seasonal trend of V. cholerae and antimicrobial resistance encourage the continued epidemiological and microbiological surveillance of the disease.

Phylogeny of Vibrio cholerae Isolates from Patients with Cholera Disease in Babylon Province

In the present study, a total of 35 stool samples were collected in the Central Health Laboratory of Babylon Province from patients presenting invasive cholera disease. The period of collection was from November 2017 to December 2018. Identification of Vibrio cholerae species was carried out by conventional methods, biochemical tests, diagnostic kits and then confirmed by PCR-based assay targeting the ompW gene. The results of this study reported that O1 serogroups were the predominant serogroup among all clinical samples with a high rate of 94.3% (N = 33), while only two isolates of non-O1/non-O139 (NAG) (5.7%) were documented as a causative agent to cholera or cholera-like disease. The phylogenetic relationship among all 35 studied strains elucidated by using polymerase chain reaction (PCR)-based fingerprinting assay (ISSR-PCR). The results of this assay showed grouping of Inaba strains into different clusters indicating that these strains were genetically diverse. Furthermore, V. cholerae El Tor O1 Ogawa strain (OG1) was closely related to strains of Inaba serotype. In contrast, NAG strains (NAG1 and NAG2) were not genetically similar to any of Inaba or Ogawa strains indicating different clone origin.

Molecular Characterization of Uropathogenic Escherichia coli Reveals Emergence of Drug Resistant O15, O22 and O25 Serogroups

Background and Objectives: Uropathogenic Escherichia coli (UPEC) are common pathogens causing urinary tract infections (UTIs). We aimed to investigate the relationship among clinical manifestation, serogroups, phylogenetic groups, and antimicrobial resistance among UPEC. Materials and Methods: One-hundred Escherichia coli isolates recovered from urine and ureteral scrapings were used for the study. The prevalence of antimicrobial resistance was determined by using European Committee on Antimicrobial Susceptibility Testing (EUCAST) recommendations. E. coli serogroups associated with UTI, as well as phylogenetic diversity were analyzed using multiplex PCR reactions. Results: Eighty-seven strains (87%) were isolated from females, while 13 (13%) from males. A high frequency of resistance to cephalosporins (43%) and fluoroquinolones (31%) was observed. Among UTI-associated serogroups O15 (32.8%), O22 (23.4%), and O25 (15.6%) were dominant and demonstrated elevated resistance rates. The E. coli phylogenetic group B2 was most common. These observations extended to pregnant patients with asymptomatic bacteriuria. Conclusions: Due to high rates of resistance, strategies using empirical therapy of second-generation cephalosporins and fluoroquinolones should be reconsidered in this population.