Diarrhoeagenic Escherichia Coli Research Articles

Multi- and extensively drug resistant Escherichia coli isolated from South African children under 5 years old with diarrhoea

BackgroundGlobally, millions of children die as a result of diarrhoea and/or antimicrobial resistant infections. Diarrhoeagenic Escherichia coli (DEC) are responsible for a substantial proportion of cases of diarrhoea in South Africa and sub-Saharan Africa. Effective treatments (including the use of antimicrobials) are therefore essential. MethodologyE. coli isolated from children under the age of five were subjected to antimicrobial susceptibility testing using the Vitek 2® compact automated system (bioMérieux Inc., France) and categorized as multidrug or extensively drug resistant (MDR or XDR). ResultsAlmost all isolates (164/166, 98.8 %) were categorized as MDR with 4.9 % (9/166) categorized as XDR. The majority of isolates (153/166, 92.2 %) were also phenotypically classified as extended-spectrum β-lactamase (ESBL) producers. More than half of these isolates (78/153, 51.0 %) were subjected to PCR for genes associated with ESBL production. More than half (45/78, 57.7 %) of the isolates tested were PCR positive for at least one ESBL gene or gene group and 11.5 % (9/78) were positive for two ESBL genes or gene groups. DiscussionThere is a need to strengthen antimicrobial resistance surveillance in South Africa and improve infection prevention and control measures. There is also a need to review the current South African Treatment Guidelines as outlined by the South African Essential Drugs Programme.

Application of a Novel Proteomic Microarray Reveals High Exposure to Diarrhoeagenic Escherichia coli among Children in Zambia Participating in a Phase I Clinical Trial.

Diarrhoeagenic E. coli (DEC) significantly contributes to the burden of diarrhoea among children. Currently, there is no approved vaccine against DEC, but several vaccines against the enterotoxigenic E. coli (ETEC) pathotype are in advanced clinical trial stages, including the ETVAX® vaccine, undergoing evaluation in Zambia. This study reports on the reactivity of antibodies from ETVAX® vaccine and placebo recipients in a phase I clinical trial to proteins derived from (DEC) other than ETEC. Plasma samples collected at two time points (prior to any vaccination and post-third dose vaccination) from 16 vaccinated and 4 placebo participants in a phase 1 clinical trial examining the safety, tolerability, and immunogenicity of ETVAX® with dmLT adjuvant were evaluated for IgG response to E. coli antigens other than ETEC using the Pan-DEC protein microarray. This was the first field application of the novel pan-DEC array as a new tool in assessing the antigenic breadth of antibody responses induced by the ETVAX vaccine, as well as to assess early life exposure to DEC pathotypes and other bacterial enteric pathogens. We observed that plasma obtained from ETVAX® and placebo recipients had high antibody reactivity to Ipa, SseC and EspB proteins. These findings suggest that there is high exposure early in life to DEC pathogens, like EPEC, EHEC, EAEC and EIEC in addition to ETEC, in the Zambian population. These immunological observations are consistent with the results of recent epidemiological studies assessing the etiology of diarrheal disease among infants and young children in Zambia.

Efficacy evaluation of anti-DEC-IgY against antibiotic-resistant diarrhoeagenic Escherichia coli.

Introduction. The rise of multi-drug-resistant bacteria poses a global threat. In 2017, the World Health Organization identified 12 antibiotic-resistant 'priority pathogens', including Enterobacteriaceae, highlighting the menace of Gram-negative bacteria. Diarrhoeagenic Escherichia coli (DEC)-induced diarrhoea is particularly problematic for travellers and infants. In contrast to other antibiotic alternatives, passive immunotherapy is showing promise by providing immediate and precise protection. However, mammalian-sourced antibodies are costly, hindering large-scale production. Egg-laying chicken-derived IgY antibodies present a cost-effective, high-yield solution, revolutionizing antibody-based therapeutics compared to mammalian IgG.Hypothesis/Gap Statement. This study hypothesized that developing anti-DEC-IgY could combat DEC infections effectively.Aim. The primary aim was to develop anti-DEC-IgY and assess its potential in DEC-induced diarrhoeal management.Method. Chickens were immunized with DEC antigens to induce an immune response. IgY antibodies were extracted from immune eggs and purified using ion-exchange column chromatography. Anti-DEC-IgY's ability to inhibit DEC growth was evaluated through growth inhibition assays. Anti-DEC-IgY's capacity to prevent E. coli adhesion was assessed using mice intestinal mucosa. In vivo experiments measured pathogen colonization reduction and infection severity reduction. P values were calculated to confirm statistical significance.Result. The antibacterial efficacy of anti-DEC-IgY by growth inhibition assay demonstrated that 25 mg ml-1 of IgY could inhibit the DEC growth. The anti-adherence-property was tested using mice intestinal mucosa and found that anti-DEC-IgY could prevent the E. coli adhesion. In vivo results suggest that 12 mg ml-1 of IgY will reduce the pathogen colonization in intestine and reduce the severity of the infection. The P values between the experimental groups confirm the statistical significance of the findings.Conclusion. The study findings suggest that IgY-based passive immunotherapy could be a potential strategy for managing the risks associated with antibiotic-resistant bacterial infections. Additionally, this study paves the way for the development of IgY-related research and applications in India.

Dynamic Interactions between Diarrhoeagenic Enteroaggregative Escherichia coli and Presumptive Probiotic Bacteria: Implications for Gastrointestinal Health.

This study delves into the temporal dynamics of bacterial interactions in the gastrointestinal tract, focusing on how probiotic strains and pathogenic bacteria influence each other and human health. This research explores adhesion, competitive exclusion, displacement, and inhibition of selected diarrhoeagenic Escherichia coli (D-EAEC) and potential probiotic strains under various conditions. Key findings reveal that adhesion is time-dependent, with both D-EAEC K2 and probiotic L. plantarum FS2 showing increased adhesion over time. Surprisingly, L. plantarum FS2 outperformed D-EAEC K2 in adhesion and exhibited competitive exclusion and displacement, with inhibition of adhesion surpassing competitive exclusion. This highlights probiotics' potential to slow pathogen attachment when not in competition. Pre-infecting with L. plantarum FS2 before pathogenic infection effectively inhibited adhesion, indicating probiotics' ability to prevent pathogen attachment. Additionally, adhesion correlated strongly with interleukin-8 (IL-8) secretion, linking it to the host's inflammatory response. Conversely, IL-8 secretion negatively correlated with trans-epithelial electrical resistance (TEER), suggesting a connection between tight junction disruption and increased inflammation. These insights offer valuable knowledge about the temporal dynamics of gut bacteria interactions and highlight probiotics' potential in competitive exclusion and inhibiting pathogenic bacteria, contributing to strategies for maintaining gastrointestinal health and preventing infections.

Pentavalent outer membrane vesicles immunized mice sera confers passive protection against five prevalent pathotypes of diarrhoeagenic Escherichia coli in neonatal mice

Diarrhoeagenic Escherichia coli (DEC) pathotypes are one of the major causative agents of diarrhoea induced childhood morbidity and mortality in developing countries. Licensed vaccines providing broad spectrum protection against DEC mediated infections are not available. Outer membrane vesicles (OMVs) are microvesicles released by gram-negative bacteria during the growth phase and contain multiple immunogenic proteins. Based on prevalence of infections, we have formulated a pentavalent outer-membrane vesicles (POMVs) based immunogen targeting five main pathotypes of DEC responsible for diarrhoeal diseases. Following isolation, OMVs from five DEC pathotypes were mixed in equal proportions to formulate POMVs and 10 µg of the immunogen was intraperitoneally administered to adult BALB/c mice. Three doses of POMVs induced significant humoral immune response against whole cell lysates (WCLs), outer membrane proteins (OMPs) and lipopolysaccharides (LPS) isolated from DEC pathotypes along with significant induction of cellular immune response in adult mice. Passive transfer of POMVs immunized adult mice sera protected neonatal mice significantly against DEC infections. Overall, this study finds POMVs to be immunogenic in conferring broad-spectrum passive protection to neonatal mice against five main DEC pathotypes. Altogether, these findings suggest that POMVs can be used as a potent vaccine candidate to ameliorate the DEC-mediated health burden.

Rotavirus and bacterial diarrhoea among children in Ile-Ife, Nigeria: Burden, risk factors and seasonality.

Diarrhoea is a leading cause of death among under-five children globally, with sub-Saharan Africa alone accounting for 1/3 episodes yearly. Viruses, bacteria and parasites may cause diarrhoea. Rotavirus is the most common viral aetiology of diarrhoea in children less than five years globally. In Nigeria, there is scarce data on the prevalence/importance, burden, clinical/risk factors and seasonality of rotavirus and bacteria and this study aims to determine the role of rotavirus and bacteria on diarrhoea cases in children less than five years in Ile-Ife, Nigeria. Socio-demographic data, environmental/risk factors and diarrhoiec stool samples were collected from children less than five years presenting with acute diarrhoea. Rotavirus was identified using ELISA. Bacteria pathogens were detected using cultural technique and typed using PCR. Diarrhoeagenic E. coli (DEC) isolates were subjected to antimicrobial susceptibility testing. Pathogen positive and negative samples were compared in terms of gender, age-group, seasonal distribution, and clinical/risk factors using chi-square with two-tailed significance. SPSS version 20.0.1 for Windows was used for statistical analysis. At least one pathogen was detected from 63 (60.6%) children having gastroenteritis while 28 (44.4%) had multiple infections. Rotavirus was the most detected pathogen. Prevalence of rotavirus mono-infection was 22%, multiple infection with bacteria was 45%. Mono-infection prevalence of DEC, Shigella spp., and Salmonella spp. were 5.8% (6/104), 5.8% (6/104), and 2.9% (3/104) and co-infection with RVA were 23.1% (24/104), 21.2% (22/104) and 10.6% (11/104) respectively. All rotaviral infections were observed in the dry season. The pathotypes of DEC detected were STEC and EAEC. Parent earnings and mid-upper arm circumference measurement have statistical correlation with diarrhoea (p = 0.034; 0.035 respectively). In this study, rotavirus was more prevalent than bacteria and occurred only in the dry season. Among bacteria aetiologies, DEC was the most common detected. Differences in seasonal peaks of rotavirus and DEC could be employed in diarrhoea management in Nigeria and other tropical countries to ensure optimal limited resources usage in preventing diarrhoea transmission and reducing indiscriminate use of antibiotics.

Are non-lactose-fermenting Escherichia coli important diarrhoeal pathogens in children and adults?

Diarrhoeagenic Escherichia coli (DEC) remains one of the major causes of acute diarrhoea episodes in developing countries. The percentage of acute diarrhoea cases caused by DEC is 30-40 % in these countries. Approximately 10% of E. coli isolates obtained from stool specimens have been reported to be non-lactose-fermenting (NLF). The available literature is sparse regarding the pathogenicity of NLF E. coli causing infectious diarrhoea. We aimed to elucidate the importance of NLF E. coli in causing diarrhoea in both adults and children by detecting various DEC pathotypes among NLF E. coli in stool samples taken from gastroenteritis cases. A total of 376 NLF E. coli isolates from 3110 stool samples from diarrhoea/gastroenteritis patients were included in the study. Up to three NLF colonies that were not confirmed as Vibrio cholerae , Aeromonas spp., Salmonella spp. or Shigella spp., but were identified as E. coli using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF), were carefully picked up from each MacConkey agar plate and then meticulously streaked onto freshly prepared, sterilized nutrient agar plates, and biochemical reactions were conducted. Multiplex PCR was conducted for the EAEC, EPEC, ETEC and EHEC pathotypes and PCR for the ipaH gene was conducted for EIEC. The disc diffusion method was used for antibiotic sensitivity testing. Using multiplex PCR and ipaH PCR, a total of 63 pathotypes of DEC were obtained, with EAEC being the most predominant (n=31) followed by EIEC (n=22), EPEC (n=8) and ETEC (n=2). To further differentiate EIEC from Shigella , additional biochemical tests were performed, including acetate utilization, mucate and salicin fermentation, and aesculin hydrolysis. Antimicrobial susceptibility testing (AST) showed that maximum resistance was seen against ciprofloxacin (82.5 %) followed by ampicillin (77.8 %) and cotrimoxazole (68.2 %), and minimum resistance was seen against ertapenem (4.8 %). In our study two pathotypes (EAEC, EIEC) were predominant among NLF E. coli and these were not only important aetiological agents in children, but also in adults. Our study also sheds light on the epidemiology of EIEC, which is one of the most neglected DEC pathotypes, as hardly any microbiological laboratories process NLF E. coli for EIEC.

Characterization of diarrhoeagenic Escherichia coli with special reference to antimicrobial resistance isolated from hospitalized diarrhoeal patients in Kolkata (2012-2019), India.

This study analyses the prevalence and antimicrobial resistance (AMR) of major diarrhoeagenic Escherichia coli (DEC) pathotypes detected in hospitalized diarrhoeal patients in Kolkata, India, during 2012-2019. A total of 8891 stool samples were collected from the Infectious Diseases Hospital, Kolkata and screened for the presence of enteric pathogens. Multiplex PCR identified the presence of DEC in 7.8% of the samples, in which ETEC was most common (47.7%) followed by EAEC (38.4%) and EPEC (13.9%). About 54% cases were due to sole DEC infections. Majority of the mixed DEC infections were caused by the Vibrio spp. (19.1%) followed by Rotavirus (14.1%) and Campylobacter spp. (8.4%). ETEC and EAEC were associated significantly with diarrhoea in children <5 years of age, whereas EPEC and also ETEC were prevalent in patients aged between 5 and 14 years. AMR profile showed high prevalence of multidrug resistance (MDR) among DEC (56.9%) in which 9% were resistant to antibiotics of six different antimicrobial classes. Screening of the AMR conferring genes of DEC showed the presence of blaCTX-M3 (30.2%) in highest number followed by blaTEM (27.5%), tetB (18%), sul2 (12.6%), strA (11.8%), aadA1 (9.8%), blaOXA-1 (9%), dfrA1 (1.6%) and blaSHV (1.2%). These findings highlighted the high prevalence of MDR in major DEC pathotypes that could be considered as the leading aetiological bacterial agent responsible for diarrhoea and suggests a significant public health threat. The results of this study can help to improve the understanding of the epidemiology of DEC infections in patients with diarrhoea. Monitoring of AMR surveillance needs special attention because the DEC isolates were highly resistant to commonly used antimicrobials in the treatment of diarrhoea.

Diarrhoeagenic E. coli occurrence and antimicrobial resistance of Extended Spectrum Beta-Lactamases isolated from diarrhoea patients attending health facilities in Accra, Ghana.

IntroductionDiarrhoea accounts for high morbidity and mortality in children and adults worldwide. Extended Spectrum Beta-Lactamase-Producing Enterobacteriaceae (ESBL-PE) and Diarrhoeagenic Escherichia coli (DEC) contribute to prolonged hospitalization because of their resistance and virulence properties aiding in the spread of diarrhoeal disease and delayed treatment.AimTo determine DEC and the antimicrobial resistance of ESBL-PE isolated among diarrhoea patients attending two health facilities in Ghana.MethodsStool samples were collected from 122 diarrhoeal patients who attended Maamobi General Hospital and Kaneshie Polyclinic between January 2019 and March 2020. Identification of bacteria was performed by using the Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Using disk diffusion, antimicrobial susceptibility testing (AST) was conducted and interpreted according to the 2018 CLSI guidelines. Detection of ESBL and DEC genes was performed using Polymerase chain reaction (PCR).ResultsA total of 80.3% (98/122) Enterobacteriaceae was recovered from the patients in the study with an overall ESBL occurrence of 20.4% (20/98), predominantly among E. coli showed 13.2% (10/76), Klebsiella pneumoniae,35.7%(5/14) and Proteus mirabilis, 57.1%(4/7). Among the ESBL genes detected, blaTEM (n = 14) was common, followed by blaCTX-M (n = 13) and blaSHV (n = 4). Thirty-four E. coli isolates possessed the heat labile (Lt) gene of Enterotoxigenic E. coli (ETEC).ConclusionOur findings confirm the existence of DEC and the antimicrobial resistance patterns of ESBL-PE among stool isolates, limiting the options of commonly used drugs for diarrhoeal treatment in Ghana. Routine laboratory testing in health care facilities and strengthened surveillance systems among hospital networks are encouraged for a better understanding of their epidemiology and clinical implications.

Use of long read sequencing to characterise the genomic architecture of mobile genetic elements encoding blaCTX-M-15 in Escherichia coli causing travellers’ diarrhoea

Increasing levels of antimicrobial resistance (AMR) have been documented in Escherichia colicausing travellers’ diarrhoea, particularly to the third-generation cephalosporins. Diarrhoeagenic E. coli (DEC) can act as a reservoir for the exchange of AMR genes between bacteria residing in the human gut, enabling them to survive and flourish through the selective pressures of antibiotic treatments. Using Oxford Nanopore Technology (ONT), we sequenced eight different sequence types (ST) belonging to five different pathotypes of extended-beta-lactamase-producing DEC harbouring blaCTX-M-15from four patients recently returned to the UK from Pakistan. The aim of the study was to determine whether blaCTX-M-15 was chromosome or plasmid-encoded to better understand the mechanisms of onward transmission of AMR determinants. In Patient A, blaCTX-M-15was plasmid-encoded in both DEC isolates (ST504/ST3032), whereas in Patient B blaCTX-M-15was located on the chromosome in both DEC isolates (ST227/1283). Patients C and D both had one isolate where blaCTX-M-15 was located on the plasmid and one chromosomally encoded (ST443/182 and ST38/99, respectively). The two plasmids associated with Patient A were different although one exhibited high similarity to the plasmid from Patient C. In the four isolates where blaCTX-M-15 was chromosomally encoded, the site of insertion and the characteristics of the inserted plasmid segment differed. Analysis of long-read sequencing data enables us to characterise the genomic architecture of mobile genetic elements encoding AMR determinants. These data may contribute to a better understanding of persistence and onward transmission of AMR determinants in MDR E. coli causing gastrointestinal and extra-intestinal infections.

ISOLATION, CHARACTERIZATION AND ANTIBIOTIC SUSCEPTIBILITY PATTERN OF DIARRHEAGENIC ESCHERICHIA COLI (DEC) AMONG CHILDREN ATTENDING SOME SELECTED HOSPITALS WITHIN KADUNA METROPOLIS

Diarrhoea has been associated with significant morbidity and mortality, ranked the second cause of death in children aged 0 to 5 years. Bacteria, viruses, protozoa, and helminths have all been implicated in diarrhoea, however rotavirus and diarrhoeagenic E. coli (DEC) are the most common cause. This study aimed to determine the antibiotic susceptibility pattern of diarrheagenic Escherichia coli isolated from children 0-5 years attending selected hospitals in Kaduna metropolis. A total of 264 stool samples were collected from children attending four selected hospitals in Kaduna metropolis. Standard culture procedures and molecular techniques such as PCR and 16s rRNA were employed in isolation and characterization of diarrhoeagenic E. coli from the stool samples. The study established a prevalence of 24.2% for diarrheagenic Escherichia coli, and all the isolates demonstrated multiple antimicrobial resistance index (MARI) of 0.5 and above, and showed significant resistance against augmentin (100%), amoxacillin (100%), ampicillin-cloxacillin (ampiclox) (100%), erythromycin (100%), gentamycin (100%), cefoxitin (100%), cefuroxime (95%), ceftriazone (95%) and ceftaxidime (85%). The least (60%) resistance was observed against imipenem. The study concluded that antibiotics have not been very effective in the treatment of E. coli-related diarrhoea. The study, therefore, recommends the implementation of programmes geared towards good hygiene, good nutrition and good health

Use of Nanopore Sequencing to Characterise the Genomic Architecture of Mobile Genetic Elements Encoding bla CTX-M-15 in Escherichia coli Causing Travellers' Diarrhoea.

Increasing levels of antimicrobial resistance (AMR) have been documented in Escherichia coli causing travellers’ diarrhoea, particularly to the third-generation cephalosporins. Diarrhoeagenic E. coli (DEC) can act as a reservoir for the exchange of AMR genes between bacteria residing in the human gut, enabling them to survive and flourish through the selective pressures of antibiotic treatments. Using Oxford Nanopore Technology (ONT), we sequenced eight isolates of DEC from four patients’ specimens who had all recently returned to the United Kingdome from Pakistan. Sequencing yielded two DEC harbouring blaCTX-M-15 per patient, all with different sequence types (ST) and belonging to five different pathotypes. The study aimed to determine whether blaCTX-M-15 was located on the chromosome or plasmid and to characterise the drug-resistant regions to better understand the mechanisms of onward transmission of AMR determinants. Patients A and C both had one isolate where blaCTX-M-15 was located on the plasmid (899037 & 623213, respectively) and one chromosomally encoded (899091 & 623214, respectively). In patient B, blaCTX-M-15 was plasmid-encoded in both DEC isolates (786605 & 7883090), whereas in patient D, blaCTX-M-15 was located on the chromosome in both DEC isolates (542093 & 542099). The two blaCTX-M-15-encoding plasmids associated with patient B were different although the blaCTX-M-15-encoding plasmid isolated from 788309 (IncFIB) exhibited high nucleotide similarity to the blaCTX-M-15-encoding plasmid isolated from 899037 (patient A). In the four isolates where blaCTX-M-15 was chromosomally encoded, two isolates (899091 & 542099) shared the same insertion site. The blaCTX-M-15 insertion site in isolate 623214 was described previously, whereas that of isolate 542093 was unique to this study. Analysis of Nanopore sequencing data enables us to characterise the genomic architecture of mobile genetic elements encoding AMR determinants. These data may contribute to a better understanding of persistence and onward transmission of AMR determinants in multidrug-resistant (MDR) E. coli causing gastrointestinal and extra-intestinal infections.

Burden of Diarrhoeagenic Escherichia coli in Santa Rosa, Guatemala in active health-services surveillance during 2008-2009 and 2014-2015.

To describe the epidemiology of laboratory-confirmed Diarrhoeagenic Escherichia coli (DEC) cases from active facility-based surveillance in Guatemala. We collected clinical and risk factor data on enrolled patients (aged 0-52years) with acute diarrhoea at government healthcare facilities (1hospital and 6clinics) in Santa Rosa, Guatemala, during 2008-2009 and 2014-2015. Stool samples were analysed, E. coli identified through culture and biochemical tests, PCR amplification of genes encoding pathotype-specific virulence factors identified specific DEC pathotypes. Healthcare-seeking adjusted incidence rates were calculated. A total of 3041 diarrhoea cases were captured by surveillance (647hospitalisations (H), 2394clinic visits (CV)); general E. coli prevalence was 17.9%. DEC pathotypes were identified in 19% (n=95/497) and 21% (n=450/2113) in diarrhoea H and CV, respectively. Enteropathogenic E. coli (EPEC) was most frequently isolated (8.2% (n=41) in diarrhoea H, 12.0% (n=255) in diarrhoea CV), followed by ETEC (6.8% (n=34) in H, 6% (n=128) in CV) and STEC (0.6% (n=3) in H, 0.6% (n=13) in CV). We did not find evidence of a difference in severity between DEC and non-DEC diarrhoea. Incidence of DEC clinic visits and hospitalisations was 648.0 and 29.3, respectively, per 10,000 persons aged ≤5years and 36.8 and 0.4, respectively, per 10,000 persons aged >5years. DEC pathotypes, especially EPEC and ETEC, were detected frequently from patients presenting with diarrhoeal illness in Santa Rosa, Guatemala. Our findings suggest that preventive interventions should be prioritised for young children.

Diarrhoeagenic Escherichia coli isolated from children with acute diarrhoea at Rakai hospital, Southern Uganda.

Diarrhoeagenic Escherichia coli (DEC) is a leading cause of childhood diarrhoea. This study estimated the prevalence of DEC and DEC pathotypes among children with acute diarrhoea in Southern Uganda. A cross-sectional study was conducted on 267 children less than 5 years with acute diarrhoea, admitted to Rakai General Hospital in Southern Uganda. Faecal samples were collected from the children and processed for isolation of E. coli. The presence of DEC and the distribution of DEC pathotypes were determined by polymerase chain reaction. A total of 102 (38.2%, 102/267) children had DEC of various pathotypes - enteroaggregative E. coli (EAEC) (14.2%); enteropathogenic E. coli (EPEC) (6.7%); enterotoxigenic E. coli (ETEC) (6%); enteroinvasive E. coli (EIEC) (7.5%); enterohemorrhagic E. coli (EHEC) (3%); and cell-detaching E. coli (CDEC) (0.75%). The difference in the overall prevalence of DEC was not significant regarding HIV but individually, EAEC and CDEC were associated with HIV-positive status while ETEC was associated with HIV-negative status. DEC is prevalent in children with acute diarrhoea in Southern Uganda and its identification in children should be considered among strategies for combatting childhood diarrhoea in Africa.

Real-time multiplex PCR assay reveals the increased prevalence of Campylobacter spp and diarrhoeagenic Escherichia coli in humans from Vellore, South India.

Introduction. Bacterial dysentery is one of the greatest causes of morbidity and mortality worldwide. Campylobacter spp. and diarrhoeagenic Escherichia coli (DEC) are recognised as the most common causes of bacterial enteritis in developing countries including India.Hypothesis/Gap statement. Rapid and accurate identification of dysentery causing organisms using molecular methods is essential for better disease management, epidemiology and outbreak investigations.Aim. In view of the limited information available on the dysentery causing agents like Campylobacter spp., enterohemorrhagic E. coli (EHEC)/enteropathogenic E. coli (EPEC) and enteroinvasive E. coli (EIEC)/Shigella in India, this study was undertaken to investigate the presence of these pathogens in human and poultry stool samples by molecular methods.Methodology. In total, 400 human stool samples and 128 poultry samples were studied. Microaerophilic culture along with real-time multiplex PCR with the targets specific to the genus Campylobacter, Campylobacter jejuni, Campylobacter coli, EHEC, EPEC and EIEC/Shigella was performed. Further species confirmation was done using MALDI-TOF MS.Results. On microaerophilic culture, C. coli was isolated in one human sample and two C. jejuni and one C. fetus in poultry samples. On PCR analysis, among human stool samples, typical EPEC (42%) was predominantly seen followed by Campylobacter spp. (19%) and EIEC/Shigella (10%). In contrast, Campylobacter spp. (41%) was predominant in poultry samples, followed by typical EPEC (26%) and EIEC/Shigella (9%). Poly-infections with Campylobacter spp. and DEC were also observed among both sources.Conclusion. The present study documented the increased prevalence of Campylobacter spp. in humans compared with the results of previous studies from India. Typical EPEC was found to be predominant in children less than 5 years of age in this study. The high prevalence of coinfections in the current study indicates that a multiple aetiology of diarrhoea is common in our settings.

Aetiology of hospital-acquired diarrhoea in under-five children from an urban hospital in East Delhi, India.

Majority of the studies of hospital-acquired diarrhoea conducted in Western countries have focused on the detection of Clostridium difficile in stool samples. Limited Asian and Indian literature is available on hospital-acquired diarrhoea. This study was aimed to describe the aetiological profile for hospital-acquired diarrhoea in children aged below five years. One hundred children aged one month to five years who developed diarrhoea (≥3 loose stools for >12 h) after hospitalization for at least 72 h were enrolled. Children who were prescribed purgatives or undergoing procedures such as enema and endoscopy or those with underlying chronic gastrointestinal disorders such as celiac disease and inflammatory bowel disease were excluded from the study. Stool samples from the enrolled children were subjected to routine microscopic examination, modified Ziel-Nielson (ZN) staining for Cryptosporidium and culture for various enteropathogens. Multiplex PCR was used to identify the strains of diarrhoeagenic Escherichia coli. Rotavirus detection was done using rapid antigen kit. Toxins (A and B) of C. difficile were detected using enzyme immunoassay. Of the 100 samples of hospital-acquired diarrhoea analysed, diarrhoeagenic E. coli (DEC) was found to be the most common organism, detected in 37 per cent of cases (enteropathogenic E. coli-18%, enterotoxigenic E. coli-8%, enteroaggregative E. coli-4% and mixed infections-7%). Cryptosporidium was detected in 10 per cent of cases. Rotavirus was detected in six per cent and C. difficile in four per cent of cases. The findings of this study suggest that the aetiological profile of hospital-acquired diarrhoea appears to be similar to that of community-acquired diarrhoea, with DEC and Cryptosporidium being the most common causes. The efforts for the prevention and management of hospital-acquired diarrhoea should, thus, be directed towards these organisms.

Pathotyping and antimicrobial susceptibility testing of Escherichia coli isolates from neonatal calves.

Neonatal calf mortality is a major concern to livestock sector worldwide. Neonatal calf diarrhoea (NCD), an acute severe condition causes morbidity and mortality in calves. Amongst various pathogens involved in NCD, E. coli is considered as one of the major causes. The study was targeted to characterize E. coli isolates from neonatal calves for diarrhoeagenic Escherichia coli (DEC) types (pathotyping), antimicrobial resistance (AMR) profiling and to correlate with epidemiological parameters. From neonates, a total of 113 faecal samples were collected, out of that 308, lactose fermenting colonies were confirmed as E. coli. Pathotypable isolates (12.3%) were represented by STEC (6.1%), EPEC (2.9%), ETEC (1.9%), EAEC (0.9%) and EHEC (0.3%). Occurrence of STEC was more in non-diarrhoeic calves, whereas ETEC was observed more in diarrhoeic calves. EPEC occurrence was observed in both diarrhoeic and non-diarrhoeic calves. Fishers extract test showed no significant association for occurrence of DEC types to type of dairies, health status, species, breed, age and sex of neonatal calves. Two hundred and eighty isolates were tested for antimicrobial susceptibility. The isolates showed maximum resistance towards ampicillin (55.4%) followed by tetracycline (54.3%), while minimum resistance was observed towards meropenem (2.5%). Multidrug resistant E. coli isolates were found to be 139 (49.6%), and Extended-spectrum beta-lactamase (ESBL) producers were 120 (42.9%). DEC pathotypes like STEC, ETEC, EHEC and EAEC that are also multidrug resistant present in neonatal calves have zoonotic potential and hence are of public health significance.Supplementary InformationThe online version contains supplementary material available at 10.1007/s11259-021-09857-5.

Occurrence of Hybrid Diarrhoeagenic Escherichia coli Associated with Multidrug Resistance in Environmental Water, Johannesburg, South Africa.

This study was undertaken to determine the virulence and antibiotic resistance profiles of diarrhoeagenic Escherichia coli (DEC) in environmental waters of Johannesburg, South Africa. Samples were collected and cultured on selective media. An 11-plex PCR assay was used to differentiate five DEC, namely: enteroaggregative (EAEC), enterohaemorrhagic (EHEC), enteroinvasive (EIEC), enteropathogenic (EPEC) and enterotoxigenic (ETEC). The antibiotic resistance profile of isolates was determined using the VITEK®-2 automated system. The virulence profiles of 170 E. coli tested showed that 40% (68/170) were commensals and 60% (102/170) were pathogenic. EPEC had a prevalence of 19.2% (32/170), followed by ETEC 11.4% (19/170), EAEC 6% (10/170) and EHEC 3% (5/170). Hybrid DEC carrying a combination of simultaneously two and three pathogenic types was detected in twenty-eight and nine isolates, respectively. The antibiotic susceptibility testing showed isolates with multidrug resistance, including cefuroxime (100%), ceftazidime (86%), cefotaxime (81%) and cefepime (79%). This study highlighted the widespread occurrence of DEC and antibiotic resistance strains in the aquatic ecosystem of Johannesburg. The presence of hybrid pathotypes detected in this study is alarming and might lead to more severe diseases. There is a necessity to enhance surveillance in reducing the propagation of pathogenic and antibiotic-resistant strains in this area.

Extended-spectrum beta-lactamase-producing strains among diarrhoeagenic Escherichia coli-prospective traveller study with literature review.

BackgroundAntibiotics are no longer the primary approach for treating all travellers’ diarrhoea (TD): most cases resolve without antibiotics and using them predisposes to colonization by multidrug-resistant bacteria. Data are accumulating on increasing resistance among TD pathogens, yet research into the most common agents, diarrhoeagenic Escherichia coli (DEC), remains limited.MethodsA total of 413 travellers to the (sub)tropics were analyzed for travel-acquired diarrhoeal pathogens and ESBL-PE. To identify ESBL-producing DEC, ESBL-producing E. coli (ESBL-EC) isolates were subjected to multiplex qPCR for various DEC pathotypes: enteroaggregative (EAEC), enteropathogenic (EPEC), enterotoxigenic (ETEC), enteroinvasive (EIEC) and enterohaemorrhagic (EHEC) E. coli.For a literature review, we screened studies among travellers and locals in low- and middle-income countries (LMICs) on the frequency of ESBL-producing DEC, and among travellers, also DEC with resistance to ciprofloxacin, azithromycin, and rifamycin derivatives.ResultsOur rate of ESBL-EC among all DEC findings was 2.7% (13/475); among EAEC 5.7% (10/175), EPEC 1.1% (2/180), ETEC 1.3% (1/80) and EHEC (0/35) or EIEC 0% (0/5). The literature search yielded three studies reporting ESBL-EC frequency and thirteen exploring resistance to TD antibiotics among travel-acquired DEC. For EAEC and ETEC, the ESBL-EC rates were 10–13% and 14–15%, resistance to fluoroquinolones 0–42% and 0–40%, azithromycin 0–29% and 0–61%, and rifaximin 0% and 0–20%. The highest rates were from the most recent collections. Proportions of ESBL-producing DEC also appear to be increasing among locals in LMICs and even carbapenemase-producing DEC were reported.ConclusionESBL producers are no longer rare among DEC, and the overall resistance to various antibiotics is increasing. The data predict decreasing efficacy of antibiotic treatment, threatening its benefits, for disadvantages still prevail when efficacy is lost.

Molecular typing and prevalence of extended-spectrum β-lactamase genes in diarrhoeagenic Escherichia coli strains isolated from foods and humans in Mashhad, Iran.

Present study was aimed to determine ESBL-encoding genes distribution in Diarrhoeagenic Escherichia coli (DEC) isolated from animal-source food products and human clinical samples in Mashhad, Iran. The strains were also further studied to analyse genotypic diversity and find genetic relationships between them. The number of 85 DEC strains including 52 and 33 strains isolated from 300 food and 520 human stool samples, respectively. Randomly amplified polymorphic DNA (RAPD), and repetitive extragenic palindromic-PCR (rep-PCR) typing methods were used to track their genetic relationships. The ESBL-encoding genes prevalence was approximately 70% in both groups of isolates. The blaTEM , blaCTX-M and blaSHV were prevalent in 67·1, 20 and 10·6% of isolates, respectively. The ESBL-positives showed significantly higher resistance rates to gentamicin, co-trimoxazole, tetracycline, aztreonam and chloramphenicol (P<0·05). Fingerprinting patterns-based dendrograms divided DEC strains into separate clusters irrespective of their sources and pathotypes. In typing field, rep-PCR provided more discriminatory power (Simpson's index of diversity (SID)=0·925) than RAPD (SID=0·812). Molecular similarity between certain animal-sourced food products and clinical sample strains supported food-borne transmission routes for genotypic elements such as ESBL-encoding genes. Findings emphasize the importance of resistance issues, the need to improve treatment guidelines and routine surveillance of hygienic measures during food processing.