Antibiotic-Resistant Vibrio cholerae O1 and Its SXT Elements Associated with Two Cholera Epidemics in Kenya in 2007 to 2010 and 2015 to 2016.

Multidrug-resistant Vibrio cholerae O1 strains have long been observed in Africa, and strains exhibiting new resistance phenotypes have emerged during recent epidemics in Kenya. This study aimed to determine the epidemiological aspects, drug resistance patterns, and genetic elements of V. cholerae O1 strains isolated from two cholera epidemics in Kenya between 2007 and 2010 and between 2015 and 2016. A total of 228 V. cholerae O1 strains, including 226 clinical strains isolated from 13 counties in Kenya during the 2007-2010 and 2015-2016 cholera epidemics and two environmental isolates (from shallow well water and spring water isolates) isolated from Pokot and Kwale Counties, respectively, in 2010 were subjected to biotyping, serotyping, and antimicrobial susceptibility testing, including the detection of antibiotic resistance genes and mobile genetic elements. All V. cholerae isolates were identified as El Tor biotypes and susceptible to ceftriaxone, gentamicin, and ciprofloxacin. The majority of isolates were resistant to trimethoprim-sulfamethoxazole (94.6%), streptomycin (92.8%), and nalidixic acid (64.5%), while lower resistance was observed against ampicillin (3.6%), amoxicillin (4.2%), chloramphenicol (3.0%), and doxycycline (1.8%). Concurrently, the integrating conjugative (SXT) element was found in 95.5% of the V. cholerae isolates; conversely, class 1, 2, and 3 integrons were absent. Additionally, 64.5% of the isolates exhibited multidrug resistance patterns. Antibiotic-resistant gene clusters suggest that environmental bacteria may act as cassette reservoirs that favor resistant pathogens. On the other hand, the 2015-2016 epidemic strains were found susceptible to most antibiotics except nalidixic acid. This revealed the replacement of multidrug-resistant strains exhibiting new resistance phenotypes that emerged after Kenya's 2007-2010 epidemic. IMPORTANCE Kenya is a country where cholera is endemic; it has experienced three substantial epidemics over the past few decades, but there are limited data on the drug resistance patterns of V. cholerae at the national level. To the best of our knowledge, this is the first study to investigate the antimicrobial susceptibility profiles of V. cholerae O1 strains isolated from two consecutive epidemics and to examine their associated antimicrobial genetic determinants. Our study results revealed two distinct antibiotic resistance trends in two separate epidemics, particularly trends for multidrug-associated mobile genetic elements and chromosomal mutation-oriented resistant strains from the 2007-2010 epidemic. In contrast, only nalidixic acid-associated chromosomal mutated strains were isolated from the 2015-2016 epidemic. This study also found similar patterns of antibiotic resistance in environmental and clinical strains. Continuous monitoring is needed to control emerging multidrug-resistant isolates in the future.

Impact of integrated water, sanitation, hygiene, health and nutritional interventions on diarrhoea disease epidemiology and microbial quality of water in a resource-constrained setting in Kenya: A controlled intervention study.

OBJECTIVES: We assessed the impact of water, hygiene and sanitation (WASH), maternal, new-born and child health (MNCH), nutrition and early childhood development (ECD) on diarrhoea and microbial quality of water in a resource-constrained rural setting in Kenya. METHODS: Through a controlled intervention study, we tested faecal and water samples collected from both the intervention and control sites before and after the interventions using microbiological, immunological and molecular assays to determine the prevalence of diarrhoeagenic agents and microbial quality of water. Data from the hospital registers were used to estimate all-cause diarrhoea prevalence. RESULTS: After the interventions, we observed a 58.2% (95% CI: 39.4-75.3) decline in all-cause diarrhoea in the intervention site versus a 22.2% (95% CI: 5.9-49.4) reduction of the same in the control site. Besides rotavirus and pathogenic Escherichia coli, the rate of isolation of other diarrhoea-causing bacteria declined substantially in the intervention site. The microbial quality of community and household water improved considerably in both the intervention (81.9%; 95% CI: 74.5%-87.8%) and control (72.5%; 95% CI: 64.2%-80.5%) sites with the relative improvements in the intervention site being slightly larger. CONCLUSIONS: The integrated WASH, MNCH, nutrition and ECD interventions resulted in notable decline in all-cause diarrhoea and improvements in water quality in the rural resource-limited population in Kenya. This indicates a direct public health impact of the interventions and provides early evidence for public health policy makers to support the sustained implementation of these interventions.

Genomic characterization of an African G4P[6] human rotavirus strain identified in a diarrheic child in Kenya: Evidence for porcine-to-human interspecies transmission and reassortment.

Human rotavirus strains having the unconventional G4P[6] genotype have been sporadically identified in diarrheic patients in different parts of the world. However, the whole genome of only one human G4P[6] strain from Africa (central Africa) has been sequenced and analyzed, and thus the exact origin and evolutionary pattern of African G4P[6] strains remain to be elucidated. In this study, we characterized the full genome of an African G4P[6] strain (RVA/Human-wt/KEN/KCH148/2019/G4P[6]) identified in a stool specimen from a diarrheic child in Kenya. Full genome analysis of strain KCH148 revealed a unique Wa-like genogroup constellation: G4-P[6]-I1-R1-C1-M1-A1-N1-T7-E1-H1. NSP3 genotype T7 is commonly found in porcine rotavirus strains. Furthermore, phylogenetic analysis showed that 10 of the 11 genes of strain KCH148 (VP7, VP4, VP6, VP1-VP3, NSP1, and NSP3-NSP5) appeared to be of porcine origin, the remaining NSP2 gene appearing to be of human origin. Therefore, strain KCH148 was found to have a porcine rotavirus backbone and thus is likely to be of porcine origin. Furthermore, strain KCH148 is assumed to have been derived through interspecies transmission and reassortment events involving porcine and human rotavirus strains. To our knowledge, this is the first report on full genome-based characterization of a human G4P[6] strain from east Africa. Our observations demonstrated the diversity of human G4P[6] strains in Africa, and provide important insights into the origin and evolutionary pattern of zoonotic G4P[6] strains on the African continent.

Characterization of Vibrio cholerae O1 isolates responsible for cholera outbreaks in Kenya between 1975 and 2017.

Kenya is endemic for cholera with different waves of outbreaks having been documented since 1971. In recent years, new variants of Vibrio cholerae O1 have emerged and have replaced most of the traditional El Tor biotype globally. These strains also appear to have increased virulence, and it is important to describe and document their phenotypic and genotypic traits. This study characterized 146 V. cholerae O1 isolates from cholera outbreaks that occurred in Kenya between 1975 and 2017. Our study reports that the 1975-1984 strains had typical classical or El Tor biotype characters. New variants of V. cholerae O1 having traits of both classical and El Tor biotypes were observed from 2007 with all strains isolated between 2015 and 2017 being sensitive to polymyxin B and carrying both classical and El Tor type ctxB. All strains were resistant to Phage IV and harbored rstR, rtxC, hlyA, rtxA and tcpA genes specific for El Tor biotype indicating that the strains had an El Tor backbone. Pulsed field gel electrophoresis (PFGE) genotyping differentiated the isolates into 14 pulsotypes. The clustering also corresponded with the year of isolation signifying that the cholera outbreaks occurred as separate waves of different genetic fingerprints exhibiting different genotypic and phenotypic characteristics. The emergence and prevalence of V. cholerae O1 strains carrying El Tor type and classical type ctxB in Kenya are reported. These strains have replaced the typical El Tor biotype in Kenya and are potentially more virulent and easily transmitted within the population.

Genomic characterization of uncommon human G3P[6] rotavirus strains that have emerged in Kenya after rotavirus vaccine introduction, and pre-vaccine human G8P[4] rotavirus strains.

A monovalent rotavirus vaccine (RV1) was introduced to the national immunization program in Kenya in July 2014. There was increased detection of uncommon G3P[6] strains that coincided temporally with the timing of this vaccine introduction. Here, we sequenced and characterized the full genomes of two post-vaccine G3P[6] strains, RVA/Human-wt/KEN/KDH1951/2014/G3P[6] and RVA/Human-wt/KEN/KDH1968/2014/G3P[6], as representatives of these uncommon strains. On full-genomic analysis, both strains exhibited a DS-1-like genotype constellation: G3-P[6]-I2-R2-C2-M2-A2-N2-T2-E2-H2. Phylogenetic analysis revealed that all 11 genes of strains KDH1951 and KDH1968 were very closely related to those of human G3P[6] strains isolated in Uganda in 2012-2013, indicating the derivation of these G3P[6] strains from a common ancestor. Because the uncommon G3P[6] strains that emerged in Kenya are fully heterotypic as to the introduced vaccine strain regarding the genotype constellation, vaccine effectiveness against these G3P[6] strains needs to be closely monitored.

Biosafety and biosecurity capacity building: insights from implementation of the NUITM-KEMRI biosafety training model.

The NUITM-KEMRI biosafety training program was developed for capacity building of new biosafety level three (BSL-3) laboratory users. The training program comprehensively covers biosafety and biosecurity theory and practice. Its training curriculum is based on the WHO biosafety guidelines, local biosafety standards, and ongoing biosafety level three research activities in the facility, also taking into consideration the emerging public health issues. The program's training approach enhances the participant's biosafety and biosecurity knowledge and builds their skills through the hands-on practice sessions and mentorship training. Subsequently, the trainees are able to integrate acquired knowledge and good practices into their routine laboratory procedures. This article describes implementation of the NUITM-KEMRI biosafety training program.

Impact of rotavirus vaccination on rotavirus hospitalisation rates among a resource-limited rural population in Mbita, Western Kenya.

OBJECTIVES: A two-dose oral monovalent rotavirus vaccine (RV1) was introduced into the Kenyan National Immunization Program in July 2014. We assessed trends in hospitalisation for rotavirus-specific acute gastroenteritis (AGE) and strain distribution among children <5 years in a rural, resource-limited setting in Kenya before and after the nationwide implementation of the vaccine. METHODS: Data on rotavirus AGE and strain distribution were derived from a 5-year hospital-based surveillance. We compared rotavirus-related hospitalisations and strain distribution in the 2-year post-vaccine period with the 3-year pre-vaccine baseline. Vaccine administrative data from the Unit of Vaccines and Immunization Services (UVIS) for Mbita sub-county were used to estimate rotavirus immunisation coverage in the study area. RESULTS: We observed a 48% (95% CI: 27-64%) overall decline in rotavirus-related hospitalisations among children aged <5 years in the post-vaccine period. Coverage with the last dose of rotavirus vaccine increased from 51% in year 1% to 72% in year 2 of the vaccine implementation. Concurrently, reductions in rotavirus hospitalisations increased from 40% in the first year to 53% in the second year of vaccine use. The reductions were most pronounced among the vaccine-eligible group, with the proportion of cases in this age group dropping to 14% in post-vaccine years from a high of 51% in the pre-vaccine period. A diversity of rotavirus strains circulated before the introduction of the vaccine with G1P[8] being the most dominant strain. G2P[4] replaced G1P[8] as the dominant strain after the vaccine was introduced. CONCLUSIONS: Rotavirus vaccination has resulted in a notable decline in hospital admissions for rotavirus infections in a rural resource-limited population in Kenya. This provides early evidence for continued use of rotavirus vaccines in routine childhood immunisations in Kenya. Our data also underscore the need for expanding coverage on second dose so as to maximise the impact of the vaccine.

Impact of rotavirus vaccination on rotavirus and all-cause gastroenteritis in peri-urban Kenyan children.

A monovalent rotavirus vaccine (RV1) was introduced into the National Immunization Program in Kenya in July 2014. We examined the impact of the vaccine on hospitalization for all-cause acute gastroenteritis (AGE) and rotavirus-specific AGE and strain distribution at a large referral hospital which serves a predominantly peri-urban population in Central Kenya. Data on rotavirus AGE and strain distribution were derived from ongoing hospital-based AGE surveillance. Hospital administrative data were used to compare trends in all-cause AGE. Pre-vaccine (July 2009-June 2014) and post-vaccine (July 2014-June 2016) periods were compared for changes in hospitalization for all-cause AGE and rotavirus AGE and strain distribution. Following the vaccine introduction, the proportion of children aged <5years hospitalized for rotavirus declined by 30% (95% CI: 19-45%) in the first year and 64% (95% CI: 49-77%) in the second year. Reductions in rotavirus positivity were most pronounced among the vaccine-eligible group (<12months) in the first year post-vaccination at 42% (95% CI: 28-56%). Greater reductions of 67% (95% CI: 51-79%) were seen in the second year in the 12-23months age group. Similarly, hospitalizations for all-cause AGE among children <5years of age decreased by 31% (95% CI: 24-40%) in the first year and 58% (95% CI: 49-67%) in the second year of vaccine introduction. Seasonal peaks of rotavirus and all-cause AGE were reduced substantially. There was an increased detection of G2P[4], G3P[6] and G3P[8], which coincided temporally with the timing of the vaccine introduction. Thus, introducing the rotavirus vaccine into the routine immunization program in Kenya has resulted in a notable decline in rotavirus and all-cause AGE hospitalizations in Central Kenya. This provides early evidence for public health policy makers in Kenya to support the sustained use of the rotavirus vaccine in routine immunizations.

Burden of Rotavirus and Enteric Bacterial Pathogens among Children under 5 Years of Age Hospitalized with Diarrhea in Suburban and Rural Areas in Kenya.

This cross-sectional descriptive study aimed to investigate the incidence of rotavirus and enteric bacterial infections among children up to 5 years old with diarrhea living in suburban and rural areas of Kenya. Between August 2011 and December 2013, a total of 1,060 diarrheal fecal specimens were obtained from 722 children at Kiambu County Hospital (KCH), located in a suburban area, and from 338 children from Mbita District Hospital (MDH), located in a rural part of western Kenya. Of the 1,060 isolates, group A rotavirus was detected in 29.6% (214/722) and 11.2% (38/338) fecal specimens from KCH and MDH, respectively. Diarrheagenic Escherichia coli (DEC) was found to be the most frequently isolated bacterial pathogens in both study areas (32.8% at KCH and 44.1% at MDH). Two different mixed infection patterns (virus/bacteria and bacteria/bacteria) were observed among patients. A significantly higher infection rate of rotavirus (17.6%, p = 0.001) and DEC (10.5%, p = 0.007) were observed during the dry season. Our study found that in both suburban and rural settings in Kenya, rotavirus and DEC are the principal cause of pediatric diarrhea and exhibit higher incidence during the dry season.

Molecular epidemiology of rotavirus gastroenteritis in Central Kenya before vaccine introduction, 2009-2014.

Between July 2009 and June 2014, a total of 1,546 fecal specimens were collected from children <5 years of age with acute gastroenteritis admitted to Kiambu County Hospital, Central Kenya. The specimens were screened for group A rotavirus (RVA) using ELISA, and RVA-positive specimens were subjected to semi-nested RT-PCR to determine the G and P genotypes. RVA was detected in 429/1,546 (27.5%) fecal specimens. RVA infections occurred in all age groups <59 months, with an early peak at 6-17 months. The infections persisted year-round with distinct seasonal peaks depending on the year. G1P[8] (28%) was the most predominant genotype, followed by G9P[8] (12%), G8P[4] (7%), G1P[4] (5%), G9P[4] (4%), and G12P[6] (3%). In the yearly change of G and P genotypes, a major shift from G9P[8] to G1P[8] was found in 2012. Phylogenetic analysis of the nucleotide sequences of the VP7 and VP4 genes of seven strains with unusual G8 or P[6] showed that the VP7 nucleotide sequences of G8 were clustered in lineage 6 in which African strains are included, and that there are at least two distinct VP4 nucleotide sequences of P[6] strains. These results represent basic data on RVA strains circulating in this region before vaccine introduction. J. Med. Virol. 89:809-817, 2017. © 2016 Wiley Periodicals, Inc.

An Outbreak of Diarrhea in Mandera, Kenya, Due to Escherichia coli Serogroup O-Nontypable Strain That Had a Coding Gene for Enteroaggregative E. coli Heat-Stable Enterotoxin 1.

In an outbreak of gastroenteritis in December 2009, in Mandera, Kenya, Escherichia coli O-nontypable (ONT) strain was isolated from stool specimens of patients (18/24, 75%). The E. coli ONT organisms could not be assigned to any of the recognized diarrheagenic groups of E. coli However, they possessed the enteroaggregative E. coli heat-stable enterotoxin-1 gene. The cell-free culture filtrates of the E. coli ONT strain isolated from the outbreak cases induced considerable amount of fluid accumulation in suckling mouse intestine, indicating production of an enterotoxic factor(s). These results identify E. coli that did not have any diarrheagenic characteristics except astA as the etiological agent of the diarrheal outbreak in Mandera. It is however considered necessary to characterize the fluid accumulation factor(s) to determine whether any novel toxins were responsible for the fluid accumulation. Moreover, it is important to study dissemination of strains producing the enterotoxic factor(s) to assess their public health significance distribution in the environment.

Prevalence, seasonal variation, and antibiotic resistance pattern of enteric bacterial pathogens among hospitalized diarrheic children in suburban regions of central Kenya.

BACKGROUND: The epidemiology of enteric pathogens has not been well studied in Kenya because of wide disparities in health status across the country. Therefore, the present study describes the prevalence of enteropathogenic bacteria, their seasonal variation, and antibiotic resistance profiles among hospitalized diarrheic children in a suburban region of central Kenya. METHODS: Fecal samples were collected between July 2009 and December 2013 from a total of 1410 children younger than 5 years, hospitalized with acute diarrhea in Kiambu County Hospital, Kenya. Conventional culture, biochemical, and molecular methods were conducted to identify causative bacterial pathogens and their virulence factors. Antimicrobial susceptibility tests were performed using E-test strips and VITEK-2 advanced expert system (AES) to evaluate the drug-resistance pattern of the isolates. RESULTS: Of the 1410 isolates, bacterial infections were identified in 474 cases. Diarrheagenic Escherichia coli (DEC) was the most frequently isolated pathogen (86.5%). Other pathogens such as Aeromonas (5.5%), Shigella (4%), Salmonella (3.4%), Providencia (3.2%), Vibrio spp. (1.1%), Yersinia enterocolitica (1.1%), and Plesiomonas shigelloides (0.2%) were also identified. Mixed bacterial infection was observed among 11.1% of the cases. The highest infection rate was found during the dry season (59.3%, p = 0.04). Most of the DEC was found to be multidrug resistant to trimethoprim/sulfamethoxazole 97.6%, amoxicillin 97.6%, erythromycin 96.9%, ampicillin 96.6%, and streptomycin 89%. CONCLUSIONS: This study suggests that DEC is the leading diarrhea-causing bacterial pathogen circulating in central Kenya, and seasonality has a significant effect on its transmission. Proper antibiotic prescription and susceptibility testing is important to guide appropriate antimicrobial therapy.

First Report of a Foodborne Providencia alcalifaciens Outbreak in Kenya.

Providencia alcalifaciens is an emerging bacterial pathogen known to cause acute gastroenteritis in children and travelers. In July 2013, P. alcalifaciens was isolated from four children appearing for diarrhea at Kiambu District Hospital (KDH) in Kenya. This study describes the outbreak investigation, which aimed to identify the source and mechanisms of infection. We identified seven primary and four secondary cases. Among primary cases were four mothers who had children and experienced mild diarrhea after eating mashed potatoes. The mothers reported feeding children after visiting the toilet and washing their hands without soap. P. alcalifaciens was detected from all secondary cases, and the isolates were found to be clonal by random amplified polymorphic DNA (RAPD) fingerprinting. Our study suggests that the outbreak was caused by P. alcalifaciens, although no fluid accumulation was observed in rabbit ileal loops. The vehicle of the outbreak was believed to be the mashed potato dish, but the source of P. alcalifaciens could not be confirmed. We found that lack of hygiene, inadequate food storage, and improper hand washing before food preparation was the likely cause of the current outbreak. This is the first report of a foodborne infection caused by P. alcalifaciens in Kenya.

BSL-3 Laboratory User Training Program at NUITM-KEMRI.

Pathogens handled in a Biosafety Level 3 (BSL-3) containment laboratory pose significant risks to laboratory staff and the environment. It is therefore necessary to develop competency and proficiency among laboratory workers and to promote appropriate behavior and practices that enhance safety through biosafety training. Following the installation of our BSL-3 laboratory at the Center for Microbiology Research-Kenya Medical Research Institute in 2006, a biosafety training program was developed to provide training on BSL-3 safety practices and procedures. The training program was developed based on World Health Organization specifications, with adjustments to fit our research activities and biosafety needs. The program is composed of three phases, namely initial assessment, a training phase including theory and a practicum, and a final assessment. This article reports the content of our training program.