Epidemiology and multiple colonization of gastrointestinal pathogens in rural Tanzanian children with and without diarrhea: A case-control study.

Diarrheal diseases are important causes of morbidity and mortality, worldwide. The occurrence of multiple pathogens in stool samples of symptomatic and asymptomatic individuals in resource-limited countries have been repeatedly described. In this study, we assessed the differentiated effects of combined pathogen detections on recorded symptoms. A case-control study was conducted among 620 under-five-year-old children in rural northeastern Tanzania with emphasis of multiple detection. The median age of children was 11 months (IQR = 7, 20), and 52.1% were male. Cases (50.2%, n = 157) were less likely than controls (64.5%, n = 198) to have multiple colonization with gastrointestinal tract (GIT) pathogens. The children's age was positively associated with the likelihood of harboring multiple GIT pathogens [OR, 1.02, 95% CI = 1.01, 1.04]. Shigella spp./enteroinvasive Escherichia coli (EIEC) [OR = 2.80, 95% CI 1.62, 4.83] and norovirus [OR = 2.04, 95% CI 1.23, 3.39] were more common in cases and were strongly associated with diarrhea, while enteroaggregative E. coli (EAEC) [OR = 0.23, 95%CI 0.17-0.33] were more common in controls. Diarrheal diseases in under-five children from rural Tanzania are likely to be due to infections with Shigella spp./EIEC, and norovirus with strongly age-dependent associations.

Understanding pre-hospital disease management of fever and diarrhoea in children-Care pathways in rural Tanzania.

OBJECTIVE: Many children in sub-Saharan Africa die from infectious diseases like malaria, pneumonia, and diarrhoea that can be prevented by early diagnosis, effective and targeted treatment. This study aimed to gain insights into case management practices by parents before they present their children to hospital. METHODS: We conducted a cross-sectional study among 332 parents attending a district hospital with their under-fives symptomatic with fever and/or diarrhoea between November 2019 and July 2020 in rural Tanzania. Timely and targeted treatment was defined as seeking health care within 24 h of fever onset, and continued fluid intake in case of diarrhoea. RESULTS: The main admission diagnoses were acute respiratory infections (61.8%), malaria (25.3%), diarrhoea (18.4%) and suspected sepsis (8.1%). The majority of children (91%) received treatment prior to admission, mostly antipyretics (75.6%), local herbal medicines (26.8%), and antibiotics (17.8%)-half of them without prescription from a clinician. For diarrhoea, the use of oral rehydration solution was rare (9.0%), although perceived as easily accessible and affordable. 49.4% of the parents presented their children directly to the hospital, 23.2% went to a pharmacy/drug shop and 19.3% to a primary health facility first. Malaria symptoms began mostly 3 days before the hospital visit; only 25.4% of febrile children visited any health facility within 24 h of disease onset. Prior use of local herbal medicine (AOR = 3.2; 95% CI 1.4-7.3), visiting the pharmacy (adjusted Odds Ratio [AOR] = 3.1; 95% confidence interval [CI]: 1.0-9.8), the dispensary being the nearest health facility (AOR = 3.0; 95% CI: 1.5-6.2), and financial difficulties (AOR = 2.2; 95% CI 1.1-4.5) were associated with delayed treatment. CONCLUSION: This study suggests that antipyretics and antibiotics dispensed at pharmacies/drug shops, as well as use of local herbal medicines, delay early diagnosis and treatment, which can be life-threatening. Pharmacies/drug shops could be integrated as key focal points for sensitising community members on how to respond to paediatric illnesses and encourage the use of oral rehydration solutions.

Antibiotic-Resistant Arcobacter spp. in commercial and smallholder farm animals in Asante Akim North Municipality, Ghana and Korogwe Town Council, Tanzania: a cross-sectional study.

BACKGROUND: Arcobacter species are considered emerging foodborne pathogens that can potentially cause serious infections in animals and humans. This cross-sectional study determined the frequency of potentially pathogenic Arcobacter spp. in both commercial and smallholder farm animals in Ghana and Tanzania. A total of 1585 and 1047 (poultry and livestock) samples were collected in Ghana and Tanzania, respectively. Selective enrichment media, along with oxidase and Gram testing, were employed for isolation of suspected Arcobacter spp. and confirmation was done using MALDI-TOF MS. Antibiotic susceptibility was assessed through disk diffusion method and ECOFFs were generated, for interpretation, based on resulting inhibition zone diameters. RESULTS: The overall Arcobacter frequency was higher in Ghana (7.0%, n = 111) than in Tanzania (2.0%, n = 21). The frequency of Arcobacter in commercial farms in Ghana was 10.3% (n/N = 83/805), while in Tanzania, it was 2.8% (n/N = 12/430). Arcobacter was detected in only 3.6% (n/N = 28/780) of the samples from smallholder farms in Ghana and 1.5% (n/N = 9/617) of the samples from Tanzania. For commercial farms, in Ghana, the presence of Arcobacter was more abundant in pigs (45.1%, n/N = 37/82), followed by ducks (38.5%, n/N = 10/26) and quails (35.7%, n/N = 10/28). According to MALDI-TOF-based species identification, Arcobacter butzleri (91.6%, n/N = 121/132), Arcobacter lanthieri (6.1%, n/N = 8/132), and Arcobacter cryaerophilus (2.3%, n/N = 3/132) were the only three Arcobacter species detected at both study sites. Almost all of the Arcobacter from Ghana (98.2%, n/N = 109/111) were isolated during the rainy season. The inhibition zone diameters recorded for penicillin, ampicillin, and chloramphenicol allowed no determination of an epidemiological cut-off value. However, the results indicated a general resistance to these three antimicrobials. Multidrug resistance was noted in 57.1% (n/N = 12/21) of the Arcobacter isolates from Tanzania and 45.0% (n/N = 50/111) of those from Ghana. The type of farm (commercial or smallholder) and source of the sample (poultry or livestock) were found to be associated with multi-drug resistance. CONCLUSIONS: The high levels of MDR Arcobacter detected from farms in both countries call for urgent attention and comprehensive strategies to mitigate the spread of antimicrobial resistance in these pathogens.

A genomic appraisal of invasive Salmonella Typhimurium and associated antibiotic resistance in sub-Saharan Africa.

Invasive non-typhoidal Salmonella (iNTS) disease manifesting as bloodstream infection with high mortality is responsible for a huge public health burden in sub-Saharan Africa. Salmonella enterica serovar Typhimurium (S. Typhimurium) is the main cause of iNTS disease in Africa. By analysing whole genome sequence data from 1303 S. Typhimurium isolates originating from 19 African countries and isolated between 1979 and 2017, here we show a thorough scaled appraisal of the population structure of iNTS disease caused by S. Typhimurium across many of Africa's most impacted countries. At least six invasive S. Typhimurium clades have already emerged, with ST313 lineage 2 or ST313-L2 driving the current pandemic. ST313-L2 likely emerged in the Democratic Republic of Congo around 1980 and further spread in the mid 1990s. We observed plasmid-borne as well as chromosomally encoded fluoroquinolone resistance underlying emergences of extensive-drug and pan-drug resistance. Our work provides an overview of the evolution of invasive S. Typhimurium disease, and can be exploited to target control measures.

Carriage of ESBL-producing Klebsiella pneumoniae and Escherichia coli among children in rural Ghana: a cross-sectional study.

BACKGROUND: Extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae (ESBL-KP) and Escherichia coli (ESBL-EC) present a high burden in both communities and healthcare sectors, leading to difficult-to-treat infections. Data on intestinal carriage of ESBL-KP and ESBL-EC in children is scarce, especially in sub-Saharan African countries. We provide data on faecal carriage, phenotypic resistance patterns, and gene variation of ESBL-EC and ESBL-KP among children in the Agogo region of Ghana. METHODS: From July to December 2019, fresh stool samples were collected within 24 h from children < 5 years with and without diarrhoea attending the study hospital. The samples were screened for ESBL-EC and ESBL-KP on ESBL agar and confirmed using double-disk synergy testing. Bacterial identification and an antibiotic susceptibility profile were performed using the Vitek 2 compact system (bioMérieux, Inc.). ESBL genes, blaSHV, blaCTX-M, and blaTEM were identified by PCR and further sequencing. RESULTS: Of the 435 children recruited, stool carriage of ESBL-EC and ESBL-KP was 40.9% (n/N = 178/435) with no significant difference in prevalence between children with diarrhoea and non-diarrhoea. No association between ESBL carriage and the age of the children was found. All isolates were resistant to ampicillin and susceptible to meropenem and imipenem. Both ESBL-EC and ESBL-KP isolates showed over 70% resistance to tetracycline and sulfamethoxazole-trimethoprim. Multidrug resistance was observed in over 70% in both ESBL-EC and ESBL-KP isolates. The blaCTX-M-15 was the most prevalent ESBL gene detected. blaCTX-M-27, blaCTX-M-14, and blaCTX-M-14b were found in non-diarrhoea stools of children, whereas blaCTX-M-28 was found in both the diarrhoea and non-diarrhoea patient groups. CONCLUSIONS: The carriage of ESBL-EC and ESBL-KP among children with and without diarrhoea in the Agogo community with a high prevalence of blaCTX-M-15 is noteworthy, highlighting the importance of both the population as a possible reservoir. This study reports for the first time the ESBL gene blaCTX-M-28 among the studied populations in Ghana.

Prevalence, Characterization, and Antimicrobial Resistance of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli from Domestic Free-Range Poultry in Agogo, Ghana.

Poultry has been suggested as an important source for extended-spectrum beta-lactamase (ESBL)-producing bacteria that can lead to difficult-to treat infections in humans. Therefore, this study aims to determine the frequency, the genetics, and antimicrobial resistance profiles of ESBL-producing Escherichia coli in domestic free-range poultry in Agogo, Ghana. The study was set up and piloted from January 2019 until June 2019. Between June and December 2019, fecal samples (N = 144) were collected from free-roaming chickens from domestic farms in the regions of Sukuumu, Bontodiase, and Freetown and cultured on ESBL screening agar. Strain identification and antibiotic susceptibility were performed using the VITEK 2 compact system. ESBL-producing E. coli were confirmed using the double disk synergy test. Molecular characterization of ESBL-associated genes (blaTEM, blaSHV, and blaCTX-M) were performed using conventional polymerase chain reaction (PCR) and further sequencing of obtained PCR amplicons. The result showed that 56.2% (n/N = 81/144) of collected fecal samples were positive for ESBL-producing E. coli. Majority of the isolates showed resistance to tetracycline (93.8%, n/N = 76/81) and trimethoprim-sulfamethoxazole (66.7, n/N = 54/81), whereas resistance to carbapenems was not found. The majority of ESBL-producing E. coli carried the blaCTX-M genes, with blaCTX-M-15 being the dominant (95.1%, n/N = 77/81) genotype. In this study, we report high frequencies of ESBL-producing E. coli in smallholder free-range poultry representing a potential source of infection, highlighting the need for control of antibiotic use and animal hygiene/sanitation measures, both important from a One Health perspective.

High anti-SARS-CoV-2 seroprevalence among unvaccinated mother-child pairs from a rural setting in north-eastern Tanzania during the second wave of COVID-19.

Background: The reported infection rates and burden of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in low- and middle-income countries, including those in sub-Saharan Africa, are relatively low compared to the rates and burden in Europe and America, partly due to limited testing capability. Unlike many countries, Tanzania has implemented neither mass screening nor restrictive measures such as lockdowns to date. The prevalence of SARS-CoV-2 infection in rural mainland Tanzania is largely unknown. Methods: A cross-sectional study was conducted between April and October 2021 to assess the anti-SARS-CoV-2 seroprevalence among mother-child pairs (n = 634 children, n = 518 mothers) in a rural setting in north-eastern Tanzania. Results: A very high prevalence of anti-SARS-CoV-2 antibody titres was found, with seroprevalence rates ranging from 29% among mothers and 40% among children, with a dynamic peak in seropositivity incidence at the end of July/early August being revealed. Significant differences in age, socioeconomic status, and body composition were associated with seropositivity in mothers and children. No significant associations were observed between seropositivity and comorbidities, including anaemia, diabetes, malaria, and HIV. Conclusions: The transmission of SARS-CoV-2 in a rural region of Tanzania during 2021 was high, indicating a much higher infection rate in rural Tanzania compared to that reported in the UK and USA during the same period. Ongoing immune surveillance may be vital to monitoring the burden of viral infection in rural settings without access to molecular genotyping, where the load of communicable diseases may mask COVID-19. Surveillance could be implemented in tandem with the intensification of vaccination strategies.

Seroprevalence of SARS-CoV-2 antibodies among children and adolescents recruited in a malariometric survey in north-eastern Tanzania July 2021.

BACKGROUND: African countries stand out globally as the region seemingly least affected by the COVID-19 pandemic, caused by the virus SARS-CoV-2. Besides a younger population and potential pre-existing immunity to a SARS-CoV-2-like virus, it has been hypothesized that co-infection or recent history of Plasmodium falciparum malaria may be protective of COVID-19 severity and mortality. The number of COVID-19 cases and deaths, however, may be vastly undercounted. Very little is known about the extent to which the Tanzanian population has been exposed to SARS-CoV-2. Here, we investigated the seroprevalence of IgG to SARS-CoV-2 spike protein in two Tanzanian rural communities 1½ years into the pandemic and the association of coinciding malaria infection and exposure. METHODS: During a malariometric survey in July 2021 in two villages in north-eastern Tanzania, blood samples were taken from 501 participants (0-19 years old). Malaria was detected by mRDT and microscopy. Levels of IgG against the spike protein of SARS-CoV-2 were measured by ELISA as well as IgG against five different antigens of P. falciparum; CIDRα1.1, CIDRα1.4 and CIDRα1.5 of PfEMP1 and GLURP and MSP3. RESULTS: The seroprevalence of SARS-CoV-2 IgG was 39.7% (106/267) in Kwamasimba and 32.5% (76/234) in Mkokola. In both villages the odds of being seropositive increased significantly with age (AOR = 1.12, 95% CI 1.07-1.17, p < 0.001). P. falciparum malaria prevalence by blood smear microscopy was 7.9% in Kwamasimba and 2.1% in Mkokola. 81.3% and 70.5% in Kwamasimba and Mkokola, respectively, showed recognition of minimum one malaria antigen. Residing in Kwamasimba was associated with a broader recognition (AOR = 1.91, 95% CI 1.34-2.71, p < 0.001). The recognition of malaria antigens increased significantly with age in both villages (AOR = 1.12; 95% CI 1.08-1.16, p < 0.001). Being SARS-CoV-2 seropositive did not associate with the breadth of malaria antigen recognition when adjusting for age (AOR = 0.99; 95% CI 0.83-1.18; p = 0.91). CONCLUSION: More than a third of the children and adolescents in two rural communities in Tanzania had antibodies to SARS-CoV-2. In particular, the adolescents were seropositive but being seropositive did not associate with the status of coinciding malaria infections or previous exposure. In Tanzania, natural immunity may have developed fast, potentially protecting a substantial part of the population from later variants.

Antibiotic-resistant Campylobacter coli and Campylobacter jejuni in commercial and smallholder farm animals in the Asante Akim North Municipality of Ghana.

Worldwide, farm animals, in particular poultry, are an important reservoir for Campylobacter spp. However, information on Campylobacter colonization in farm animals in Africa is scarce. Hence, this cross-sectional study determined antibiotic-resistant Campylobacter from both commercial and smallholder farm animals in the Asante Akim North Municipality of Ghana. Fecal samples from poultry and livestock kept by commercial and smallholder farms were collected and analyzed using standard microbiological methods. The overall Campylobacter frequency was 20.3% (n/N = 322/1,585), and frequencies detected were similarly high in isolates from commercial (21.0%, n/N = 169/805) and smallholder (19.6%, n/N = 153/780) farms. Species isolated were C. coli (67.7%, n/N = 218/322) and C. jejuni (32.3%, n/N = 104/322). However, the frequency of C. coli was 2.1 (95% CI: 1.8-2.5) times higher than what was found for C. jejuni. Campylobacter frequencies in the rainy season was 22.2% (n/N = 258/1,160) and 15.1% (n/N = 64/425) in the dry season (prevalence ratio = 1.48, 95% CI: 1.2-1.9). About 1.7% (n/N = 6/322) of the Campylobacter isolates, all from smallholder farms, were susceptible to all antibiotics tested. Multidrug resistance was observed for 4.7% (n/N = 15/322) of the Campylobacter isolates, of which 93.3% (n/N = 14/15) occurred in isolates from commercial farms. This study highlights the need for the implementation of control programs, in commercial farming but also at the smallholder farm level, to formulate clear guidelines aimed at decreasing Campylobacter contamination of meat products and reducing the use of antibiotics in the farming sector.

Comparative genomics revealed adaptive admixture in Cryptosporidium hominis in Africa.

Cryptosporidiosis is a major cause of diarrhoeal illness among African children, and is associated with childhood mortality, malnutrition, cognitive development and growth retardation. Cryptosporidium hominis is the dominant pathogen in Africa, and genotyping at the glycoprotein 60 (gp60) gene has revealed a complex distribution of different subtypes across this continent. However, a comprehensive exploration of the metapopulation structure and evolution based on whole-genome data has yet to be performed. Here, we sequenced and analysed the genomes of 26 C. hominis isolates, representing different gp60 subtypes, collected at rural sites in Gabon, Ghana, Madagascar and Tanzania. Phylogenetic and cluster analyses based on single-nucleotide polymorphisms showed that isolates predominantly clustered by their country of origin, irrespective of their gp60 subtype. We found a significant isolation-by-distance signature that shows the importance of local transmission, but we also detected evidence of hybridization between isolates of different geographical regions. We identified 37 outlier genes with exceptionally high nucleotide diversity, and this group is significantly enriched for genes encoding extracellular proteins and signal peptides. Furthermore, these genes are found more often than expected in recombinant regions, and they show a distinct signature of positive or balancing selection. We conclude that: (1) the metapopulation structure of C. hominis can only be accurately captured by whole-genome analyses; (2) local anthroponotic transmission underpins the spread of this pathogen in Africa; (3) hybridization occurs between distinct geographical lineages; and (4) genetic introgression provides novel substrate for positive or balancing selection in genes involved in host-parasite coevolution.

Transmission of Cryptosporidium Species Among Human and Animal Local Contact Networks in Sub-Saharan Africa: A Multicountry Study.

BACKGROUND: Cryptosporidiosis has been identified as one of the major causes of diarrhea and diarrhea-associated deaths in young children in sub-Saharan Africa. This study traces back Cryptosporidium-positive children to their human and animal contacts to identify transmission networks. METHODS: Stool samples were collected from children < 5 years of age with diarrhea in Gabon, Ghana, Madagascar, and Tanzania. Cryptosporidium-positive and -negative initial cases (ICs) were followed to the community, where stool samples from households, neighbors, and animal contacts were obtained. Samples were screened for Cryptosporidium species by immunochromatographic tests and by sequencing the 18S ribosomal RNA gene and further subtyped at the 60 kDa glycoprotein gene (gp60). Transmission clusters were identified and risk ratios (RRs) calculated. RESULTS: Among 1363 pediatric ICs, 184 (13%) were diagnosed with Cryptosporidium species. One hundred eight contact networks were sampled from Cryptosporidium-positive and 68 from negative ICs. Identical gp60 subtypes were detected among 2 or more contacts in 39 (36%) of the networks from positive ICs and in 1 contact (1%) from negative ICs. In comparison to Cryptosporidium-negative ICs, positive ICs had an increased risk of having Cryptosporidium-positive household members (RR, 3.6 [95% confidence interval {CI}, 1.7-7.5]) or positive neighboring children (RR, 2.9 [95% CI, 1.6-5.1]), but no increased risk of having positive animals (RR, 1.2 [95% CI, .8-1.9]) in their contact network. CONCLUSIONS: Cryptosporidiosis in rural sub-Saharan Africa is characterized by infection clusters among human contacts, to which zoonotic transmission appears to contribute only marginally.

Performance of a rapid diagnostic test for the detection of Cryptosporidium spp. in African children admitted to hospital with diarrhea.

BACKGROUND: Cryptosporidium is a protozoan parasite that causes mild to severe diarrhoeal disease in humans. To date, several commercial companies have developed rapid immunoassays for the detection of Cryptosporidium infection. However, the challenge is to identify an accurate, simple and rapid diagnostic tool for the estimation of cryptosporidiosis burden. This study aims at evaluating the accuracy of CerTest Crypto, a commercialized rapid diagnostic test (RDT) for the detection of Cryptosporidium antigens in the stool of children presenting with diarrhoea. METHODS: A cross-sectional study was conducted in four study sites in Sub-Saharan Africa (Gabon, Ghana, Madagascar, and Tanzania), from May 2017 to April 2018. Stool samples were collected from children under 5 years with diarrhoea or a history of diarrhoea within the last 24 hours. All specimens were processed and analyzed using CerTest Crypto RDT against a composite diagnostic panel involving two polymerase chain reaction (PCR) tests (qPCR and RFLP-PCR,) as the gold standard. RESULTS: A total of 596 stool samples were collected. Evaluation of the RDT yielded a very low overall sensitivity of 49.6% (confidence interval (CI) 40.1-59.0), a specificity of 92.5% (CI 89.8-94.7), positive predictive value of 61.3% (CI 50.6-71.2), and negative predictive value of 88.5% (85.3-91.1) when compared to the composite reference standard of qPCR and RFLP-PCR for the detection of Cryptosporidium species. Moreover, the performance of this test varied across different sites. CONCLUSION: The weak performance of the studied RDT suggests the need to carefully evaluate available commercial RDTs before their use as standard tools in clinical trials and community survey of Cryptosporidium infections in pediatric cohorts.

Epidemiology and Antimicrobial Susceptibility of Salmonella enterica Bloodstream Isolates Among Febrile Children in a Rural District in Northeastern Tanzania: A Cross-sectional Study.

BACKGROUND: Salmonella enterica including Salmonella Typhi and nontyphoidal Salmonella (NTS) are the predominant cause of community-acquired bloodstream infections in sub-Saharan Africa (sSA). Multiple-drug resistance and emerging fluoroquinolone resistance are of concern. Data on the age distribution of typhoid fever in sSA are scarce but essential for typhoid conjugate vaccine policy. We sought to describe Salmonella bloodstream infections, antimicrobial resistance, and age distribution at a rural district hospital in northeastern Tanzania. METHODS: From 2008 to 2016, febrile children or children with a history of fever aged 1 month to 5 years admitted to Korogwe District Hospital were enrolled. Demographic, clinical data and blood cultures were collected. Organisms were identified by conventional microbiological methods, and antimicrobial susceptibility test was done by disc diffusion. RESULTS: Of 4176 participants receiving blood cultures, 383 (9.2 %) yielded pathogens. Of pathogens, 171 (44.6%) were Salmonella enterica of which 129 (75.4%) were Salmonella Typhi, and 42 (24.6%) were NTS. The median (interquartile range age of participants was 13.1 (6.3-28.0) months for those with Salmonella Typhi and 11.5 (8.5-23.4) months for NTS. Of 129 Salmonella Typhi, 89 (89.9%) were resistant to amoxicillin, 85 (81.0%) to chloramphenicol, and 93 (92.1%) to trimethoprim-sulfamethoxazole compared with 22 (62.9%), 15 (39.4%), and 27 (79.4%), respectively, for NTS. Multidrug resistance was present in 68 (81.0%) of Salmonella Typhi and 12 (41.4%) of NTS. CONCLUSION: Salmonella Typhi was the leading cause of bloodstream infection among infants and young children <2 years of age admitted to Korogwe District Hospital. Multidrug resistance was common, highlighting a role for typhoid conjugate vaccine into routine infant vaccine schedules.