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

BACKGROUND: Globally, 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. METHODOLOGY: E. 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). RESULTS: Almost 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 ß-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. DISCUSSION: There 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.

Epidemiology of Cefotaxime-Hydrolysing ß-Lactamase-Producing Escherichia coli in Children with Diarrhoea Reported Globally between 2012 and 2022.

The global spread of cefotaxime-hydrolysing ß-lactamase (CTX-M)-producing Escherichia coli (E. coli) and its associated impact on paediatric diarrhoeal treatment and management has become a public health concern. This review assessed surveillance studies on CTX-M-producing E. coli associated with diarrhoea in children published between 2012 and 2022 globally. A total of thirty-eight studies were included for data analysis, categorised into continental regions, and tabulated. The majority (68%) of studies were conducted in Asian countries while few studies were conducted in Europe (11%) and Africa (18%), respectively. On the African continent, the majority (11%) of studies were conducted in Northern Africa while no studies were reported in East Africa. On the American continent, 3% of the studies were reported from South America. The studies included were classified into diarrheagenic E. coli (74%; 28/38) and faecal carriage (26%; 10/38). Of all the E. coli pathotypes associated with CTX-M production, EPEC was frequently reported. The prevalence of CTX-M-producing E. coli including the CTX-M-15-producing variants ranged between 1% and 94%. About 37% of the studies generalised the report as blaCTX-M-positive E. coli. The use of sequencing in characterising the CTX-M-producing E. coli was reported in only 32% of all the studies. This review provides information on the epidemiology of CTX-M-15-producing E. coli in paediatric diarrhoea and the extent to which surveillance is being performed. This is relevant in informing clinical practice for the management of diarrhoea as well as the design of future surveillance studies.

Norovirus-Associated Gastroenteritis Vesikari Score and Pre-Existing Salivary IgA in Young Children from Rural South Africa.

Norovirus (NoV) is the leading cause of viral gastroenteritis, mostly affecting young children worldwide. However, limited data are available to determine the severity of norovirus-associated AGE (acute gastroenteritis) and to correlate it with the NoV-specific IgA antibodies' level. Between October 2019 and September 2021, two hundred stool samples were randomly collected from symptomatic cases for the vesikari score and NoV-specific IgA assessment in young children from rural South Africa. Additionally, one hundred saliva specimens were concomitantly sampled within the same cohort to evaluate the NoV-specific salivary IgA levels. In addition, 50 paired saliva and stool samples were simultaneously collected from asymptomatic children to serve as controls. NoV strains in stool samples were detected using real-time RT-PCR, amplified, and genotyped with RT-PCR and Sanger sequencing. ELISA using NoV VLP (virus-like particles) GII.4 as antigens was performed on the saliva specimens. Dehydrated children were predominantly those with NoV infections (65/74, 88%; p < 0.0001). NoV-positive infections were significantly associated with the severe diarrhea cases having a high vesikari score (55%, 33/60) when compared to the non-severe diarrheal score (29.3%, 41/140; p < 0.0308). NoV of the GII genogroup was mainly detected in severe diarrhea cases (50.9%, 30/59; p = 0.0036). The geometric means of the NoV-specific IgA level were higher in the asymptomatic NoV-infected group (0.286) as compared to the symptomatic group (0.174). This finding suggests that mucosal immunity may not protect the children from the NoV infection. However, the findings indicated the contribution of the pre-existing NoV-specific IgA immune response in reducing the severity of diarrheal disease. A high vesikari score of AGE associated with the NoV GII genogroup circulating in the study area underscores the need for an appropriate treatment of AGE based on the severity level of NoV-associated clinical symptoms in young children.

Isoniazid and Rifampicin Resistance-Conferring Mutations in Mycobacterium tuberculosis Isolates from South Africa.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (M. tb), remains a significant global health issue, with high morbidity and mortality rates. The emergence of drug-resistant strains, particularly multidrug-resistant TB (MDR-TB), poses difficult challenges to TB control efforts. This comprehensive review and meta-analysis investigated the prevalence of and molecular insights into isoniazid (INH) and rifampicin (RIF) resistance-conferring mutations in M. tb isolates from South Africa. Through systematic search and analysis of 11 relevant studies, we determined the prevalence of gene mutations associated with RIF and INH resistance, such as rpoB, katG, and inhA. The findings demonstrated a high prevalence of specific mutations, including S450L in rpoB, and S315T, which are linked to resistance against RIF and INH, respectively. These results contribute to the understanding of drug resistance mechanisms and provide valuable insights for the development of targeted interventions against drug-resistant TB.

Prevalence of Human Norovirus GII.4 Sydney 2012 [P31] between 2019 and 2021 among Young Children from Rural Communities in South Africa.

Acute gastroenteritis (AGE) accounts for considerable morbidity and mortality in the paediatric population worldwide, especially in low-income countries. Human norovirus (HNoV), particularly GII.4 strains, are important agents of AGE. This study aimed to detect and characterise HNoV in children with and without AGE. Between 2019 and 2021, 300 stool samples (200 AGE and 100 without AGE) were collected from children below 5 years of age referred to the healthcare facilities of the rural communities of Vhembe District, South Africa. After detection using real-time RT-PCR, HNoV positive samples were subjected to RT-PCR and Sanger sequencing. Partial nucleotide sequences (capsid/RdRp) were aligned using the Muscle tool, and phylogenetic analysis was performed using MEGA 11. The nucleotides' percent identity among HNoV strains was compared using ClustalW software. A significant difference in HNoV prevalence between AGE children (37%; 74/200) and non-AGE (14%; 14/100) was confirmed (p < 0.0001). Genogroup II (GII) HNoV was predominant in AGE children (80%; 59/74), whereas most non-AGE children were infected by the GI norovirus genogroup (64%; 9/14). GII.4 Sydney 2012 [P31] strains were dominant (59%; 19/32) during the study period. A phylogenetic analysis revealed a close relationship between the HNoV strains identified in this study and those circulating worldwide; however, ClustalW showed less than 50% nucleotide similarity between strains from this study and those from previously reported norovirus studies in the same region. Our findings indicate significant changes over time in the circulation of HNoV strains, as well as the association between high HNoV prevalence and AGE symptoms within the study area. The monitoring of HuNoV epidemiology, along with stringent preventive measures to mitigate the viral spread and the burden of AGE, are warranted.

Prevalence and Antimicrobial Resistance Profile of Diarrheagenic Escherichia coli from Fomites in Rural Households in South Africa.

Diarrheagenic Escherichia coli (DEC) pathotypes are the leading cause of mortality and morbidity in South Asia and sub-Saharan Africa. Daily interaction between people contributes to the spreading of Escherichia coli (E. coli), and fomites are a common source of community-acquired bacterial infections. The spread of bacterial infectious diseases from inanimate objects to the surrounding environment and humans is a serious problem for public health, safety, and development. This study aimed to determine the prevalence and antibiotic resistance of diarrheagenic E. coli found in toilets and kitchen cloths in the Vhembe district, South Africa. One hundred and five samples were cultured to isolate E. coli: thirty-five samples were kitchen cloths and seventy-five samples were toilet swabs. Biochemical tests, API20E, and the VITEK®-2 automated system were used to identify E. coli. Pathotypes of E. coli were characterised using Multiplex Polymerase Chain Reaction (mPCR). Nine amplified gene fragments were sequenced using partial sequencing. A total of eight antibiotics were used for the antibiotic susceptibility testing of E. coli isolates using the Kirby-Bauer disc diffusion method. Among the collected samples, 47% were positive for E. coli. DEC prevalence was high (81%), with ETEC (51%) harboring lt and st genes being the most dominant pathotype found on both kitchen cloths and toilet surfaces. Diarrheagenic E. coli pathotypes were more prevalent in the kitchen cloths (79.6%) compared with the toilet surfaces. Notably, hybrid pathotypes were detected in 44.2% of the isolates, showcasing the co-existence of multiple pathotypes within a single E. coli strain. The antibiotic resistance testing of E. coli isolates from kitchen cloths and toilets showed high resistance to ampicillin (100%) and amoxicillin (100%). Only E. coli isolates with hybrid pathotypes were found to be resistant to more than three antibiotics. This study emphasizes the significance of fomites as potential sources of bacterial contamination in rural settings. The results highlight the importance of implementing proactive measures to improve hygiene practices and antibiotic stewardship in these communities. These measures are essential for reducing the impact of DEC infections and antibiotic resistance, ultimately safeguarding public health.

The African Wastewater Resistome: Identifying Knowledge Gaps to Inform Future Research Directions.

Antimicrobial resistance (AMR) is a growing global public health threat. Furthermore, wastewater is increasingly recognized as a significant environmental reservoir for AMR. Wastewater is a complex mixture of organic and inorganic compounds, including antibiotics and other antimicrobial agents, discharged from hospitals, pharmaceutical industries, and households. Therefore, wastewater treatment plants (WWTPs) are critical components of urban infrastructure that play a vital role in protecting public health and the environment. However, they can also be a source of AMR. WWTPs serve as a point of convergence for antibiotics and resistant bacteria from various sources, creating an environment that favours the selection and spread of AMR. The effluent from WWTPs can also contaminate surface freshwater and groundwater resources, which can subsequently spread resistant bacteria to the wider environment. In Africa, the prevalence of AMR in wastewater is of particular concern due to the inadequate sanitation and wastewater treatment facilities, coupled with the overuse and misuse of antibiotics in healthcare and agriculture. Therefore, the present review evaluated studies that reported on wastewater in Africa between 2012 and 2022 to identify knowledge gaps and propose future perspectives, informing the use of wastewater-based epidemiology as a proxy for determining the resistome circulating within the continent. The study found that although wastewater resistome studies have increased over time in Africa, this is not the case in every country, with most studies conducted in South Africa. Furthermore, the study identified, among others, methodology and reporting gaps, driven by a lack of skills. Finally, the review suggests solutions including standardisation of protocols in wastewater resistome works and an urgent need to build genomic skills within the continent to handle the big data generated from these studies.

High Burden of Co-Infection with Multiple Enteric Pathogens in Children Suffering with Diarrhoea from Rural and Peri-Urban Communities in South Africa.

Infectious diarrhoea contributes to high morbidity and mortality in young children from sub-Saharan Africa. The aim of this study was to assess the prevalence of single and multiple diarrhoeal-causing pathogen combinations in children suffering from diarrhoea from rural and peri-urban communities in South Africa. A total of 275 diarrhoea stool specimens were collected between 2014 and 2016 from Hospitals and Primary Health Care clinics. The BioFire® FilmArray® Gastrointestinal panel was used to simultaneously detect 22 diarrhoea pathogens (viruses, bacteria, parasites) known to cause diarrhoea. A total of 82% (226/275) enteric pathogens were detected in the stool specimens. The two most detected bacterial, viral and parasitic pathogens each included: EAEC (42%), EPEC (32%), Adenovirus F40/41 (19%), Norovirus (15%), Giardia (8%) and Cryptosporidium (6%), respectively. Single enteric pathogen infections were recorded in 24% (65/275) specimens with EAEC, and Norovirus was found in 26% (17/65) and 14% (9/65) of the specimens, respectively. Multiple enteric pathogen combinations were recorded in 59% (161/275) of the stool specimens with 53% (85/161) containing two pathogens, 22% (35/161) containing three pathogens and 25% (41/161) containing four or more pathogens. The results from this study demonstrated the complex nature of pathogen co-infections in diarrhoeal episodes which could have an impact on treatment effectiveness.

Enteropathogenic Escherichia coli (EPEC) expressing a non-functional bundle-forming pili (BFP) also leads to increased growth failure and intestinal inflammation in C57BL/6 mice.

Bundle-forming pili (BFP) are implicated in the virulence of typical enteropathogenic E. coli (EPEC), resulting in enhanced colonization and mild to severe disease outcomes; hence, non-functional BFP may have a major influence on disease outcomes in vivo. Weaned antibiotic pre-treated C57BL/6 mice were orally infected with EPEC strain UMD901 (E2348/69 bfpA C129S); mice were monitored daily for body weight; stool specimens were collected daily; and intestinal tissues were collected at the termination of the experiment on day 3 post-infection. Real-time PCR was used to quantify fecal shedding and tissue burden. Intestinal inflammatory biomarkers lipocalin-2 (LCN-2) and myeloperoxidase (MPO) were also assessed. Infection caused substantial body weight loss, bloody diarrhea, and intestinal colonization with fecal and intestinal tissue inflammatory biomarkers that were comparable to those previously published with the wild-type typical EPEC strain. Here we further report on the evaluation of an EPEC infection model, showing how disruption of bfp function does not impair, and may even worsen diarrhea, colonization, and intestinal disruption and inflammation. More research is needed to understand the role of bfp in pathogenicity of EPEC infections in vivo.

Partial Analysis of the Capsid Protein (VP1) of Human Sapovirus Isolated from Children with Diarrhoea in Rural Communities of South Africa.

Background: Viral diarrhoea is a concern in acute gastroenteritis cases among children younger than 5 years of age. Sapovirus has been noted as an emerging causative agent of acute gastroenteritis worldwide. Objective/Study Design. The aim of this study was to characterize human sapoviruses targeting the VP1 (NVR and N-terminal) region. Twenty-five samples were randomly selected from 40 sapovirus-positive samples previously detected and analyzed for the VP1 region using the One-Step RT-PCR assay. The PCR products were subjected to Sanger sequencing analysis. Results: The polyprotein segment (NVR and N-terminal) was successfully amplified from 10/25 samples. Sapovirus GI.1 was the most predominant strain (6/10; 60%), followed by SV-GII.1 (2/10; 20%) and 10% of each GI.3 and GII.3. Conclusion: Through the partial analysis of the VP1 region, this study provides more data to add on the human sapovirus genetic characterization of circulating strains in South Africa, with the proposition of further analysis of sapovirus VP1 fragments for the viral structure and function.

Effectiveness of the Novel Anti-TB Bedaquiline against Drug-Resistant TB in Africa: A Systematic Review of the Literature.

BACKGROUND: In 2018, an estimated 10.0 million people contracted tuberculosis (TB), and 1.5 million died from it, including 1.25 million HIV-negative persons and 251,000 HIV-associated TB fatalities. Drug-resistant tuberculosis (DR-TB) is an important contributor to global TB mortality. Multi-drug-resistant TB (MDR-TB) is defined as TB resistant to at least isoniazid (INH) and rifampin (RMP), which are recommended by the WHO as essential drugs for treatment. OBJECTIVE: To investigate the effectiveness of bedaquiline addition to the treatment of drug-resistant TB infections on the African continent. METHODOLOGY: The search engine databases Medline, PubMed, Google Scholar, and Embase were used to obtain published data pertaining to DR-TB between 2012 and 2021 in Africa. Included studies had to document clinical characteristics at treatment initiation and outcomes at the end of treatment (i.e., success, failure, recurrence, loss to follow-up, and death). The included studies were used to conduct a meta-analysis. All data analysis and visualization were performed using the R programming environment. The log risk ratios and sample variances were calculated for DR-TB patients treated with BBQ monotherapy vs. BDQ and other drug therapy. To quantify heterogeneity among the included studies, random effect sizes were calculated. RESULTS: A total of 16 studies in Africa from Mozambique (N = 1 study), Eswatini (N = 1 study), Democratic Republic of the Congo (N = 1 study), South Africa (N = 12 studies), and a multicenter study undertaken across Africa (N = 1 study) were included. In total, 22,368 individuals participated in the research studies. Among the patients, (55.2%; 12,350/22,368) were male while 9723/22,368 (44%) were female. Overall, (9%; 2033/22,368) of patients received BDQ monotherapy, while (88%; 19,630/22,368) patients received bedaquiline combined with other antibiotics. In total, (42%; 9465/22,368) of the patients were successfully treated. About (39%; 8653/22,368) of participants finished their therapy, meanwhile (5%; 1166/22,368) did not finish their therapy, while people (0.4%; 99/22,368) were lost to follow up. A total of (42%; 9265/22,368) patients died. CONCLUSION: Very few studies on bedaquiline usage in DR-TB in Africa have been published to date. Bedaquiline has been shown to enhance DR-TB results in clinical studies and programmatic settings. Hence, the World Health Organization (WHO) has recommended that it be included in DR-TB regimens. However, in the current study limited improvement to DR-TB treatment results were observed using BDQ on the continent. Better in-country monitoring and reporting, as well as multi-country collaborative cohort studies of DR-TB, can expand the knowledge of bedaquiline usage and clinical impact, as well as the risks and benefits throughout the continent.

Virulence and phylogenetic analysis of enteric pathogenic Escherichia coli isolated from children with diarrhoea in South Africa.

BACKGROUND: Diarrhoeagenic Escherichia coli pose a significant risk to human health. As such, determining the source(s) of these bacteria when isolated from patients with diarrhoea is an important step in disease prevention. OBJECTIVES: To identify the presence of genes coding for virulence and phylogroups among E. coli isolated from children hospitalized due to diarrhoea in Limpopo Province, South Africa. METHODS: E. coli isolates were identified using the VITEK-2 automated system. An 11-gene multiplex polymerase chain reaction (PCR) was used to differentiate five pathogenic types of E. coli: enteroaggregative (EAEC), enteroinvasive (EIEC), enterohaemorrhagic (EHEC), enteropathogenic (EPEC) and enterotoxigenic (ETEC). The Clermont quadruplex PCR method was used to identify phylogroups of isolates. RESULTS: From the 133 isolates tested, 79 were confirmed as E. coli. Of these, 19.0% (15/79) were commensals and 81.0% (64/79) were positive for at least one pathotype, of which ETEC was predominant (16.5%, 13/79), followed by EAEC (10.1%, 8/79), EPEC (7.6%, 6/79) and EHEC (2.5%, 2/79). Hybrid pathotypes were also detected and EAEC/ETEC was predominant (25.3%, 20/79). Phylogroup B2 was predominant (30.4%, 24/79), followed by phylogroup B1 (22.8%, 18/79), and phylogroups C and E (both 12.7%, 10/79). Just over 6% (5/79) of isolates were non-typable. CONCLUSION: There was a high distribution of diarrhoeagenic E. coli associated with different phylogroups among children living in Limpopo Province, South Africa. This emphasizes the importance of future monitoring of virulence and phylogroup distribution of E. coli isolates in this province in particular and in South Africa as a whole.

Co-Existence of Free-Living Amoebae and Potential Human Pathogenic Bacteria Isolated from Rural Household Water Storage Containers.

This study investigated the co-existence of potential human pathogenic bacteria and free-living amoebae in samples collected from stored water in rural households in South Africa using borehole water as a primary water source. Over a period of 5 months, a total of 398 stored water and 392 biofilm samples were collected and assessed. Free-living amoebae were identified microscopically in 92.0% of the water samples and 89.8% of the biofilm samples. A further molecular identification using 18S rRNA sequencing identified Vermamoeba vermiformis, Entamoeba spp., Stenamoeba spp., Flamella spp., and Acanthamoeba spp. including Acanthamoeba genotype T4, which is known to be potentially harmful to humans. Targeted potential pathogenic bacteria were isolated from the water samples using standard culture methods and identified using 16S rRNA sequencing. Mycobacterium spp., Pseudomonas spp., Enterobacter spp., and other emerging opportunistic pathogens such as Stenotrophomonas maltophilia were identified. The results showed the importance of further studies to assess the health risk of free-living amoebae and potential human pathogenic bacteria to people living in rural communities who have no other option than to store water in their homes due to water shortages.

Human Astrovirus in Symptomatic and Asymptomatic Children: A Cross-Sectional Study on Hospitalized and Outpatients from Rural Communities of South Africa between 2017-2021.

Human astroviruses are considered acute gastroenteritis agents (AGE) and are largely reported in children worldwide. There are limited data on astrovirus prevalence in rural communities, especially in hospitalized and asymptomatic cases. This study was a cross-sectional survey aiming to investigate the prevalence of classic human astroviruses in symptomatic and asymptomatic cases and hospitalized and outpatient children in rural communities of the Vhembe District, South Africa. A total of 236 stool samples (166 symptomatic and 70 asymptomatic) were collected from young children under 5 years of age. Real-time RT-PCR for astrovirus detection, RT-PCR amplification of capsid and polymerase partial genes as well as Sanger sequencing were performed. The classic astrovirus prevalence in symptomatic patients (7.23%, 12/166) as compared to healthy controls (4.29%, 3/70) was not statistically different (t-value: 1.782, p = 0.141: 95% CI). We did not observe a significant difference of classic astrovirus prevalence rate between the hospitalized group (6.52%, 3/46) and outpatient group (7.5%, 9/120). Symptomatic children below 6 months old were the most affected group (18.18%, 6/33). This study characterized human astrovirus genotype 2 and a putative recombinant strain (polymerase genotype 1/capsid genotype 2). Phylogenetic analysis revealed these genotypes are closely related to the strains circulating elsewhere within the African continent. The findings suggest that astrovirus is a common enteric pathogen in the study area. The results highlight the exposure of children and the need to monitor astroviruses for their potential impact in diarrhoeal diseases.

Prevalence and Genetic Characterisation of Human Sapovirus from Children with Diarrhoea in the Rural Areas of Vhembe District, South Africa, 2017-2020.

Diarrhoeal disease is considered an important cause of morbidity and mortality in developing areas, and a large contributor to the burden of disease in children younger than five years of age. This study investigated the prevalence and genogroups of human sapovirus (SV) in children ≤5 years of age in rural communities of Vhembe district, South Africa. Between 2017 and 2020, a total of 284 stool samples were collected from children suffering with diarrhoea (n = 228) and from children without diarrhoea (n = 56). RNA extraction using Boom extraction method, and screening for SV using real-time PCR were done in the lab. Positive samples were subjected to conventional RT-PCR targeting the capsid fragment. Positive sample isolates were genotyped using Sanger sequencing. Overall SV were detected in 14.1% (40/284) of the stool samples (16.7% (38/228) of diarrhoeal and 3.6% (2/56) of non-diarrhoeal samples). Significant correlation between SV positive cases and water sources was noted. Genogroup-I was identified as the most prevalent strain comprising 81.3% (13/16), followed by SV-GII 12.5% (2/16) and SV-GIV 6.2% (1/16). This study provides valuable data on prevalence of SV amongst outpatients in rural and underdeveloped communities, and highlights the necessity for further monitoring of SV circulating strains as potential emerging strains.

Detection of Free-Living Amoebae and Their Intracellular Bacteria in Borehole Water before and after a Ceramic Pot Filter Point-of-Use Intervention in Rural Communities in South Africa.

Free-living amoebae (FLA) are ubiquitous in nature, whereas amoeba-resistant bacteria (ARB) have evolved virulent mechanisms that allow them to resist FLA digestion mechanisms and survive inside the amoeba during hostile environmental conditions. This study assessed the prevalence of FLA and ARB species in borehole water before and after a ceramic point-of-use intervention in rural households. A total of 529 water samples were collected over a five-month period from 82 households. All water samples were subjected to amoebal enrichment, bacterial isolation on selective media, and molecular identification using 16S PCR/sequencing to determine ARB species and 18S rRNA PCR/sequencing to determine FLA species present in the water samples before and after the ceramic pot intervention. Several FLA species including Acanthamoeba spp. and Mycobacterium spp. were isolated. The ceramic pot filter removed many of these microorganisms from the borehole water. However, design flaws could have been responsible for some FLA and ARB detected in the filtered water. FLA and their associated ARB are ubiquitous in borehole water, and some of these species might be potentially harmful and a health risk to vulnerable individuals. There is a need to do more investigations into the health risk of these organisms after point-of-use treatment.

Human Enteric Pathogens in Eight Rivers Used as Rural Household Drinking Water Sources in the Northern Region of South Africa.

People living in rural areas still rely on the use of environmental water that is contaminated by human and animal activities. This study assessed the occurrence of human enteric pathogens in rivers that are used by rural communities Vhembe District of South Africa as a source of drinking water covering two seasons (winter and summer) over a one-year period. Water quality was assessed using physico characteristics and indicator organisms (total coliforms, E. coli, Clostridium perfringens). Pathogens tested included bacteria (Pathogenic E. coli, Salmonella-, Shigella- and Vibrio spp.), protozoa (Cryptosporidium- and Giardia spp.), and enteric viruses (Rota-, Noro-, Entero-, and Adenoviruses) while using published molecular protocols. The results showed that the indicator bacteria counts exceeded South African drinking water quality guideline limits and pathogenic E. coli was detected in the samples. No Shigella spp. were isolated, while Vibrio spp. and Salmonella spp. were present; parasites were detected in four rivers and Enteric viruses were predominantly detected in the winter season. The results indicated the poor condition of water and the potential health risks to consumers highlighting the need for implementing river catchment management strategies for continued sustainability in these rivers.

Predominance of Human Bocavirus Genotype 1 and 3 in Outpatient Children with Diarrhea from Rural Communities in South Africa, 2017-2018.

Human bocavirus (HBoV) is an emerging virus globally associated with diarrhea in young children. This study aims to investigate the prevalence of HBoV genotypes in children (≤5 years) from rural communities in South Africa (SA) suffering from acute gastroenteritis (AGE). A total of 141 fecal samples of children ≤5 years with acute gastroenteritis (AGE) were collected from rural primary health care facilities in the Vhembe district of SA between June 2017 and July 2018. Clinical symptoms and demographic data were also recorded. A total of 102 (72%) were outpatients, and 39 (28%) were hospitalized patients. Human bocavirus (HBoV) genotypes were determined using real-time multiplex PCR. DNA extracts of positive samples were confirmed by conventional PCR targeting the NS1 gene. Co-infection with other enteric viruses were determined in HBoV-positive samples using real-time PCR. HBoV was detected in eight (5.7%) children with AGE, of which three (37.5%) were HBoV1, three (37.5%) were HBoV3, and two (25%) were HBoV2. The majority of positive cases were identified in outpatients (62%) between the ages of 1 and 24 months. Co-infection in HBoV-positive samples with other enteric viruses included rotavirus (37.5%), adenovirus (37.5%), norovirus (25%), and astrovirus (12.5%). HBoV infections could be seen as a potential emerging diarrheal pathogen in South Africa. However, more studies are needed to understand the role of HBoV infections in children with AGE.

Enteropathogenic Escherichia coli Infection Induces Diarrhea, Intestinal Damage, Metabolic Alterations, and Increased Intestinal Permeability in a Murine Model.

Enteropathogenic E. coli (EPEC) are recognized as one of the leading bacterial causes of infantile diarrhea worldwide. Weaned C57BL/6 mice pretreated with antibiotics were challenged orally with wild-type EPEC or escN mutant (lacking type 3 secretion system) to determine colonization, inflammatory responses and clinical outcomes during infection. Antibiotic disruption of intestinal microbiota enabled efficient colonization by wild-type EPEC resulting in growth impairment and diarrhea. Increase in inflammatory biomarkers, chemokines, cellular recruitment and pro-inflammatory cytokines were observed in intestinal tissues. Metabolomic changes were also observed in EPEC infected mice with changes in tricarboxylic acid (TCA) cycle intermediates, increased creatine excretion and shifts in gut microbial metabolite levels. In addition, by 7 days after infection, although weights were recovering, EPEC-infected mice had increased intestinal permeability and decreased colonic claudin-1 levels. The escN mutant colonized the mice with no weight loss or increased inflammatory biomarkers, showing the importance of the T3SS in EPEC virulence in this model. In conclusion, a murine infection model treated with antibiotics has been developed to mimic clinical outcomes seen in children with EPEC infection and to examine potential roles of selected virulence traits. This model can help in further understanding mechanisms involved in the pathogenesis of EPEC infections and potential outcomes and thus assist in the development of potential preventive or therapeutic interventions.