Phenotypic and molecular characterization of ß-lactamase-producing Klebsiella species among children discharged from hospital in Western Kenya.

BACKGROUND: The emergence and spread of ß-lactamase-producing Klebsiella spp. has been associated with a substantial healthcare burden resulting in therapeutic failures. We sought to describe the proportion of phenotypic resistance to commonly used antibiotics, characterize ß-lactamase genes among isolates with antimicrobial resistance (AMR), and assess the correlates of phenotypic AMR in Klebsiella spp. isolated from stool or rectal swab samples collected from children being discharged from hospital. METHODS: We conducted a cross-sectional study involving 245 children aged 1-59 months who were being discharged from hospitals in western Kenya between June 2016 and November 2019. Whole stool or rectal swab samples were collected and Klebsiella spp. isolated by standard microbiological culture. ß-lactamase genes were detected by PCR whilst phenotypic antimicrobial susceptibility was determined using the disc diffusion technique following standard microbiology protocols. Descriptive analyses were used to characterize phenotypic AMR and carriage of ß-lactamase-producing genes. The modified Poisson regression models were used to assess correlates of phenotypic beta-lactam resistance. RESULTS: The prevalence of ß-lactamase carriage among Klebsiella spp. isolates at hospital discharge was 62.9% (154/245). Antibiotic use during hospitalization (adjusted prevalence ratio [aPR] = 4.51; 95%CI: 1.79-11.4, p < 0.001), longer duration of hospitalization (aPR = 1.42; 95%CI: 1.14-1.77, p < 0.002), and access to treated water (aPR = 1.38; 95%CI: 1.12-1.71, p < 0.003), were significant predictors of phenotypically determined ß-lactamase. All the 154 ß-lactamase-producing Klebsiella spp. isolates had at least one genetic marker of ß-lactam/third-generation cephalosporin resistance. The most prevalent genes were blaCTX-M 142/154 (92.2%,) and blaSHV 142/154 (92.2%,) followed by blaTEM 88/154 (57.1%,) and blaOXA 48/154 (31.2%,) respectively. CONCLUSION: Carriage of ß-lactamase producing Klebsiella spp. in stool is common among children discharged from hospital in western Kenya and is associated with longer duration of hospitalization, antibiotic use, and access to treated water. The findings emphasize the need for continued monitoring of antimicrobial susceptibility patterns to inform the development and implementation of appropriate treatment guidelines. In addition, we recommend measures beyond antimicrobial stewardship and infection control within hospitals, improved sanitation, and access to safe drinking water to mitigate the spread of ß-lactamase-producing Klebsiella pathogens in these and similar settings.

Typhoidal salmonella disease in Mukuru informal settlement, Nairobi Kenya; carriage, diversity, and antimicrobial resistant genes.

INTRODUCTION: Multiple studies have shown that typhoid fever is endemic in developing countries characterized by poor hygiene. A unique way of Salmonella Typhi (S.Typhi) pathogenicity is establishing a persistent, usually asymptomatic carrier state in some infected individuals who excrete large numbers of bacteria in faeces. This study aimed to determine the isolation rate of S.Typhi from blood and stool samples among cases and asymptomatic individuals in the Mukuru informal settlement and identify antibiotic resistance patterns within the same population. MATERIALS AND METHODS: We recruited 1014 outpatient participants presenting with typhoid-like symptoms in selected health centres in Nairobi, Kenya. Bacterial isolation was done on Xylose Lysine Deoxycholate agar (XLD) and Mac Conkey agar (Oxoid), followed by standard biochemical tests. Identification was done using API20E, and S.Typhi was confirmed by serotyping using polyvalent antisera 0-9 and monovalent antisera d. The Kirby-Bauer disc diffusion method was used to test the antimicrobial susceptibility of S.Typhi isolates, while Multi-Drug Resistant (MDR) strains were characterized using conventional PCR. RESULTS: Of 1014 participants, 54 (5%) tested positive for S.Typhi. Thirty-eight (70%) of the S.Typhi isolated were from stool samples, while sixteen (30%) were from blood. Three (0.2%) of the isolates were from asymptomatic carriers. Of the 54 S.Typhi isolates, 20 (37%) were MDR. Resistance to ciprofloxacin and nalidixic acid was 43% and 52%, respectively. Resistance to amoxicillin-clavulanic acid (a beta-lactam inhibitor) was 2%. The BlaTEM-1 gene was present in 19/20 (95%) MDR isolates. CONCLUSION: MDR S.Typhi is prevalent in Mukuru Informal settlement. The sharp increase in nalidixic acid resistance is an indication of reduced susceptibility to fluoroquinolones, which are currently the recommended drugs for the treatment of typhoid fever. This study highlights the need for effective antimicrobial stewardship and routine surveillance of antimicrobial resistance (AMR) to inform policy on the prevention and control of MDR Typhoid disease.

Age-dependent acquisition of IgG antibodies to Shigella serotypes-a retrospective analysis of seroprevalence in Kenyan children with implications for infant vaccination.

Background: Shigellosis mainly affects children under 5 years of age living in low- and middle-income countries, who are the target population for vaccination. There are, however, limited data available to define the appropriate timing for vaccine administration in this age group. Information on antibody responses following natural infection, proxy for exposure, could help guide vaccination strategies. Methods: We undertook a retrospective analysis of antibodies to five of the most prevalent Shigella serotypes among children aged <5 years in Kenya. Serum samples from a cross-sectional serosurvey in three Kenyan sites (Nairobi, Siaya, and Kilifi) were analyzed by standardized ELISA to measure IgG against Shigella sonnei and Shigella flexneri 1b, 2a, 3a, and 6. We identified factors associated with seropositivity to each Shigella serotype, including seropositivity to other Shigella serotypes. Results: A total of 474 samples, one for each participant, were analyzed: Nairobi (n = 169), Siaya (n = 185), and Kilifi (n = 120). The median age of the participants was 13.4 months (IQR 7.0-35.6), and the male:female ratio was 1:1. Geometric mean concentrations (GMCs) for each serotype increased with age, mostly in the second year of life. The overall seroprevalence of IgG antibodies increased with age except for S. flexneri 6 which was high across all age subgroups. In the second year of life, there was a statistically significant increase of antibody GMCs against all five serotypes (p = 0.01-0.0001) and a significant increase of seroprevalence for S. flexneri 2a (p = 0.006), S. flexneri 3a (p = 0.006), and S. sonnei (p = 0.05) compared with the second part of the first year of life. Among all possible pairwise comparisons of antibody seropositivity, there was a significant association between S. flexneri 1b and 2a (OR = 6.75, 95% CI 3-14, p < 0.001) and between S. flexneri 1b and 3a (OR = 23.85, 95% CI 11-54, p < 0.001). Conclusion: Children living in low- and middle-income settings such as Kenya are exposed to Shigella infection starting from the first year of life and acquire serotype-specific antibodies against multiple serotypes. The data from this study suggest that Shigella vaccination should be targeted to infants, ideally at 6 or at least 9 months of age, to ensure children are protected in the second year of life when exposure significantly increases.

Enteric bacterial agents associated with diarrhea and their antimicrobial sensitivity profiles in children under 5 years from mukuru informal settlement, Nairobi, Kenya.

BACKGROUND: In Kenya, diarrhoeal disease is the third leading cause of child mortality after malaria and pneumonia, accounting for nearly 100 deaths daily. We conducted a cross-sectional study in Mukuru informal settlements to determine the bacteria associated with diarrhea and their ASTs to provide data essential for implementing appropriate intervention measures. METHODS: Diarrheagenic children (≤ 5 years) were purposively recruited from outpatient clinics of Municipal City Council, Mukuru kwa Reuben, Medical Missionaries of Mary, and Mama Lucy Kibaki Hospital, Nairobi. A total of 219 stool samples were collected between May 2021 and August 2021. Stool culture was done on MacConkey and Salmonella Shigella agar, while the recovered bacteria were identified using VITEK®2GNID and polymerase chain reaction (PCR) used for E. coli pathotyping. Antibiotic Susceptibility Testing was done using VITEK®2AST-GN83. RESULTS: At least one bacterial organism was recovered from each of the 213 (97%) participants, with 115 (56%) participants having only one bacterial type isolated, 90 (43%) with two types of bacteria, and 2 (1%) with three types of bacteria recovered. The most predominant bacteria recovered was 85% (93/109) non-pathogenic E.coli and 15% (16/109)of pathogenic E.coli, with 2 (1%) were Enterohemorrhagic E.coli (EHEC), 6 (3%) were Enteroaggregative E.coli (EAEC), and 8 (4%) were Enteropathogenic E.coli (EPEC). Other potentially pathogenic bacteria included Enterobacter sp (27.8%), Klebsiella sp 33(11%), and Citrobacter sp 15(4.7%). Pathogenic isolates such as Salmonella 7 (2%), Proteus mirabilis 16 (6%), Providencia alcalifaciens 1 (0.3%), and Shigella 16 (4.7%) were detected. Isolates such as Pantoea spp 2(0.67%), Raoultella planticola 1(0.33%), and Kluyvera 6(2%) rarely reported but implicated with opportunistic diarrhoeal disease were also recovered. Ampicillin, cefazolin, and sulfamethoxazole-trimethoprim were the least effective antimicrobials at 64%, 57%, and 55% resistance, respectively, while meropenem (99%), amikacin (99%), tazobactam piperacillin (96%), and cefepime (95%) were the most effective. Overall, 33(21%) of all enterics recovered were multidrug-resistant. CONCLUSION: The study documented different bacteria potentially implicated with childhood diarrhea that were not limited to E. coli, Shigella, and Salmonella, as previously observed in Kenya. The strains were resistant to the commonly used antibiotics, thus narrowing the treatment options for diarrheal disease.

Antimicrobial resistance and the great divide: inequity in priorities and agendas between the Global North and the Global South threatens global mitigation of antimicrobial resistance.

To limit the catastrophic effects of the increasing bacterial resistance to antimicrobials on health, food, environmental, and geopolitical security, and ensure that no country or region is left behind, a coordinated global approach is required. In this Viewpoint, we argue that the diverging resource availabilities, needs, and priorities of the Global North and the Global South in terms of the actions required to mitigate the antimicrobial resistance pandemic are a direct threat to success. We argue that evidence suggests a need to prioritise and support infection prevention interventions (ie, clean water and safe sanitation, increased vaccine coverage, and enhanced infection prevention measures for food production in the Global South contrary to the focus on research and development of new antibiotics in the Global North) and to recalibrate global funding resources to address this need. We call on global leaders to redress the current response, which threatens mitigation of the antimicrobial resistance pandemic.

Typhoid fever.

Typhoid fever is an invasive bacterial disease associated with bloodstream infection that causes a high burden of disease in Africa and Asia. Typhoid primarily affects individuals ranging from infants through to young adults. The causative organism, Salmonella enterica subsp. enterica serovar Typhi is transmitted via the faecal-oral route, crossing the intestinal epithelium and disseminating to systemic and intracellular sites, causing an undifferentiated febrile illness. Blood culture remains the practical reference standard for diagnosis of typhoid fever, where culture testing is available, but novel diagnostic modalities are an important priority under investigation. Since 2017, remarkable progress has been made in defining the global burden of both typhoid fever and antimicrobial resistance; in understanding disease pathogenesis and immunological protection through the use of controlled human infection; and in advancing effective vaccination programmes through strategic multipartner collaboration and targeted clinical trials in multiple high-incidence priority settings. This Primer thus offers a timely update of progress and perspective on future priorities for the global scientific community.

The epidemiology of fecal carriage of nontyphoidal Salmonella among healthy children and adults in three sites in Kenya.

BACKGROUND: Despite the importance of non-Typhoidal Salmonella (NTS) disease in Africa, epidemiologic data on carriage and transmission are few. These data are important to understand the transmission of NTS in Africa and to design control strategies. METHOD: To estimate the prevalence of stool carriage of NTS in Kenya, we conducted a cross-sectional study in Kilifi, Nairobi, and Siaya, sites with a low, moderate and high incidence of invasive NTS disease, respectively. At each site, we randomly selected 100 participants in each age-group of 0-11 months, 12-59 months, 5-14 years, 15-54 years and ≥55 years. We collected stool, venous blood (for hemoglobin and malaria rapid tests), anthropometric measurements, and administered a questionnaire on Water Access Sanitation and Hygiene (WASH) practices. Stool samples were cultured on selective agar for Salmonella; suspect isolates underwent serotyping and antimicrobial susceptibility testing. RESULT: Overall, 53 (3.5%) isolates of NTS were cultured from 1497 samples. Age-adjusted prevalence was 13.1% (95%CI 8.8-17.4) in Kilifi, 0.4% (95%CI 0-1.3) in Nairobi, and 0.9% (95%CI 0-2.0) in Siaya. Prevalence was highest among those aged 15-54 years (6.2%). Of 53 isolates; 5 were S. Enteritidis, 1 was S. Typhimurium. No S. Typhi was isolated. None of the risk factors were associated with carriage of NTS. All isolates were susceptible to all antibiotics tested, including ampicillin, chloramphenicol, ciprofloxacin and co-trimoxazole. CONCLUSION: Prevalence of fecal carriage was high in Kilifi, an area of low incidence of invasive NTS disease and was low in areas of higher incidence in Nairobi and Siaya. The age-prevalence, risk factors, geographical and serotype distribution of NTS in carriage differs from invasive disease.

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.

Evaluating the relationship between community water and sanitation access and the global burden of antibiotic resistance: an ecological study.

BACKGROUND: Antibiotic resistance is a leading cause of death, with the highest burden occurring in low-resource settings. There is little evidence on the potential for water, sanitation, and hygiene (WASH) access to reduce antibiotic resistance in humans. We aimed to determine the relationship between the burden of antibiotic resistance in humans and community access to drinking water and sanitation. METHODS: In this ecological study, we linked publicly available, geospatially tagged human faecal metagenomes (from the US National Center for Biotechnology Information Sequence Read Archive) with georeferenced household survey datasets that reported access to drinking water sources and sanitation facility types. We used generalised linear models with robust SEs to estimate the relationship between the abundance of antibiotic resistance genes (ARGs) in human faecal metagenomes and community-level coverage of improved drinking water and sanitation within a defined radii of faecal metagenome coordinates. FINDINGS: We identified 1589 metagenomes from 26 countries. The mean abundance of ARGs, in units of log10 ARG fragments per kilobase per million mapped reads classified as bacteria, was highest in Africa compared with Europe (p=0·014), North America (p=0·0032), and the Western Pacific (p=0·011), and second highest in South-East Asia compared with Europe (p=0·047) and North America (p=0·014). Increased access to improved water and sanitation was associated with lower ARG abundance (effect estimate -0·22, [95% CI -0·39 to -0·05]) and the association was stronger in urban (-0·32 [-0·63 to 0·00]) than in rural (-0·16 [-0·38 to 0·07]) areas. INTERPRETATION: Although additional studies to investigate causal effects are needed, increasing access to water and sanitation could be an effective strategy to curb the proliferation of antibiotic resistance in low-income and middle-income countries. FUNDING: Bill & Melinda Gates Foundation.

Risk Factors for Colonization With Extended-Spectrum Cephalosporin-Resistant and Carbapenem-Resistant Enterobacterales Among Hospitalized Patients in Kenya: An Antibiotic Resistance in Communities and Hospitals (ARCH) Study.

BACKGROUND: The spread of extended-spectrum cephalosporin-resistant Enterobacterales (ESCrE) and carbapenem-resistant Enterobacterales (CRE) represents a significant global public health threat. We identified putative risk factors for ESCrE and CRE colonization among patients in 1 urban and 3 rural hospitals in Kenya. METHODS: During a January 2019 and March 2020 cross-sectional study, stool samples were collected from randomized inpatients and tested for ESCrE and CRE. The Vitek2 instrument was used for isolate confirmation and antibiotic susceptibility testing, and least absolute shrinkage and selection operator (LASSO) regression models were used to identify colonization risk factors while varying antibiotic use measures. RESULTS: Most (76%) of the 840 enrolled participants received ≥1 antibiotic in the 14 days preceding their enrollment, primarily ceftriaxone (46%), metronidazole (28%), or benzylpenicillin-gentamycin (23%). For LASSO models that included ceftriaxone administration, ESCrE colonization odds were higher among patients hospitalized for ≥3 days (odds ratio, 2.32 [95% confidence interval, 1.6-3.37]; P < .001), intubated patients (1.73 [1.03-2.91]; P = .009), and persons living with human immunodeficiency virus (1.70 [1.03-2.8]; P = .029). CRE colonization odds were higher among patients receiving ceftriaxone (odds ratio, 2.23 [95% confidence interval, 1.14-4.38]; P = .025) and for every additional day of antibiotic use (1.08 [1.03-1.13]; P = .002). CONCLUSIONS: While CRE colonization was strongly associated with ceftriaxone use and duration of antibiotic use, the odds of ESCrE colonization increased with exposure to the hospital setting and invasive medical devices, which may reflect nosocomial transmission. These data suggest several areas where hospitals can intervene to prevent colonization among hospitalized patients, both through robust infection prevention and control practices and antibiotic stewardship programs.

Risk Factors for Colonization With Multidrug-Resistant Bacteria in Urban and Rural Communities in Kenya: An Antimicrobial Resistance in Communities and Hospitals (ARCH) Study.

BACKGROUND: Colonization with antimicrobial-resistant bacteria increases the risk of drug-resistant infections. We identified risk factors potentially associated with human colonization with extended-spectrum cephalosporin-resistant Enterobacterales (ESCrE) in low-income urban and rural communities in Kenya. METHODS: Fecal specimens, demographic and socioeconomic data were collected cross-sectionally from clustered random samples of respondents in urban (Kibera, Nairobi County) and rural (Asembo, Siaya County) communities between January 2019 and March 2020. Presumptive ESCrE isolates were confirmed and tested for antibiotic susceptibility using the VITEK2 instrument. We used a path analytic model to identify potential risk factors for colonization with ESCrE. Only 1 participant was included per household to minimize household cluster effects. RESULTS: Stool samples from 1148 adults (aged ≥18 years) and 268 children (aged <5 years) were analyzed. The likelihood of colonization increased by 12% with increasing visits to hospitals and clinics. Furthermore, individuals who kept poultry were 57% more likely to be colonized with ESCrE than those who did not. Respondents' sex, age, use of improved toilet facilities, and residence in a rural or urban community were associated with healthcare contact patterns and/or poultry keeping and may indirectly affect ESCrE colonization. Prior antibiotic use was not significantly associated with ESCrE colonization in our analysis. CONCLUSIONS: The risk factors associated with ESCrE colonization in communities include healthcare- and community-related factors, indicating that efforts to control antimicrobial resistance in community settings must include community- and hospital-level interventions.

Seroprevalence of non-typhoidal Salmonella disease and associated factors in children in Mukuru settlement in Nairobi County, Kenya.

Non-typhoidal Salmonella (NTS) infections remain a significant public health challenge especially in sub-Saharan Africa. NTS disease is endemic in Kenya and is associated with sporadic fatal outbreaks in several regions of the country with poor resource setting. Data is limited on background exposure of NTS in the population in endemic areas and the general immune status of the community most affected by NTS. The aim of the study was to determine the proportion of children exposed to Salmonella Enteritidis or Salmonella Typhimurium O antigen among the apparently healthy children and patients and the associated host and environmental factors among children attending selected healthcare facilities in Mukuru, Nairobi County, Kenya. A cross-sectional case-control study was conducted among patients and apparently healthy participants aged 0-5 years. Blood was collected and centrifuged to obtain serum. The serum was used to test for the presence of antibodies (IgA, IgG, IgM) against NTS using ELISA. A questionnaire was administered to obtain relevant demographic, socio-economic and healthcare utilization information. A total of 382 children were recruited into the study. The NTS seroprevalence was 12.6%. Among the apparently healthy participants, mean age of those exposed to NTS was 36 months and those not exposed was 27 months. Among patients, the mean age was 39 months and those not exposed was 30 months. The seroprevalence of NTS infection among the apparently healthy was significantly associated with cooking water, washing water and age of the child. Treating water using chlorine or boiling method was identified as being protective against contracting Salmonella Typhimurium/Enteritidis. Among the patients, the proportion of exposure was significantly associated with keeping animals and the chicken count. There is a high exposure to NTS among young children below five years of age and the population has developed immunity to the disease.

Nontyphoidal Salmonella Invasive Disease: Challenges and Solutions.

Nontyphoidal Salmonella are a leading cause of community-onset bacteremia and other serious infections in sub-Saharan African countries where large studies indicate that they are an uncommon cause of moderate-to-severe diarrhea. Approximately 535 000 nontyphoidal Salmonella invasive disease illnesses and 77 500 deaths were estimated to occur in 2017; 422 000 (78.9%) illnesses and 66 500 (85.9%) deaths in countries in sub-Saharan Africa. Lineages of Salmonella enterica serovar Typhimurium sequence type (ST) 313 and lineages of Salmonella enterica serovar Enteritidis ST11 dominate as causes of invasive disease. A major reservoir for these specific strains outside of humans has not been identified to date. Human fecal shedding of such strains is common in areas where nontyphoidal Salmonella invasive disease incidence is high. The case-fatality ratio of nontyphoidal Salmonella invasive disease is approximately 15%. Early diagnosis and treatment are needed to avert fatal outcomes. Antimicrobial resistance, including multiple drug resistance, decreased fluoroquinolone susceptibility, and resistance to third-generation cephalosporins, is increasing in prevalence and is likely to further compromise patient outcomes. Naturally acquired immunity against invasive disease develops in children aged >3 years in endemic areas, likely mediated in part by the sequential acquisition of T-cell immunity, followed by antigen-specific immunoglobulin G antibodies. Vaccines in preclinical or clinical development include live-attenuated S. enterica serovar Typhimurium, nontyphoidal S. enterica core and O-polysaccharide glycoconjugates, multiple antigen-presenting system complexes, and generalized modules for membrane antigens vaccines. The latter are in phase I trials in Europe and Africa. Both vaccine use, and other effective, evidence-based nonvaccine interventions, are needed to prevent and control nontyphoidal Salmonella invasive disease.

Plasmid-mediated quinolone resistance genes detected in Ciprofloxacin non-susceptible Escherichia coli and Klebsiella isolated from children under five years at hospital discharge, Kenya.

BACKGROUND: The increasing spread of fluoroquinolone resistant enteric bacteria is a global public health concern. Children recently discharged from the hospital are at high risk of carriage of antimicrobial resistance (AMR) due to frequent exposure to antimicrobials during inpatient stays. This study aimed to determine the prevalence, correlates of ciprofloxacin (CIP) non-susceptibility, and distribution of plasmid-mediated quinolone resistance (PMQR) genes in Escherichia coli (E. coli) and Klebsiella spp isolated from children under five years being discharged from two Kenyan Hospitals. METHODS: E. coli and Klebsiella spp were isolated from fecal samples from children discharged from hospital and subjected to antimicrobial susceptibility testing (AST) by disc diffusion and E-test. CIP non-susceptible isolates were screened for seven PMQR genes using multiplex polymerase chain reaction (PCR). Poisson regression was used to determine the association between the carriage of CIP non-susceptible isolates and patient characteristics. RESULTS: Of the 280 CIP non-susceptible isolates: 188 E. coli and 92 Klebsiella spp isolates identified among 266 discharged children, 195 (68%) were CIP-non-susceptible with minimum inhibitory concentrations (MICs) of ≥ 1 µg/mL. Among these 195 isolates, 130 (67%) had high-level CIP MIC = ≥ 32 µg/mL). Over 80% of the isolates had at least one PMQR gene identified: aac(6')lb-cr (60%), qnrB (24%), oqxAB (22%), qnrS (16%), and qepA (6%), however, qnrA was not identified in any isolates tested. Co-carriage of qnrB with acc(6')-lb-cr was the most predominant accounting for 20% of all the isolates. Ceftriaxone use during hospital admission and the presence of extended spectrum beta-lactamase (ESBL) production were significantly associated with the carriage of CIP non-susceptible E. coli and Klebsiella spp. CONCLUSION: CIP non-susceptibility is common among E. coli and Klebsiella spp isolated from hospital discharged children in Kenya. Carriage and co-carriage of PMQR, including the newly identified qepA gene, were frequently observed. These findings suggest that children leaving the hospital may serve as an important reservoir for transmission of resistant E. coli and Klebsiella spp to the community. Enhanced surveillance for AMR determinants is critical to inform interventions to control antimicrobial-resistant bacteria.

Frequency and diversity of carbapenemase-producing Enterobacterales recovered from untreated wastewater impacted by selective media containing cefotaxime and meropenem in Ohio, USA.

As safe agents of last resort, carbapenems are reserved for the treatment of infections caused by multidrug-resistant organisms. The impact of ß-lactam antibiotics, cefotaxime, and meropenem on the frequency and diversity of carbapenemase-producing organisms recovered from environmental samples has not been fully established. Therefore, this methodological study aimed at determining ß-lactam drugs used in selective enrichment and their impact on the recovery of carbapenemase-producing Enterobacterales (CPE) from untreated wastewater. We used a longitudinal study design where 1L wastewater samples were collected weekly from wastewater treatment plant (WWTP) influent and quarterly from contributing sanitary sewers in Columbus, Ohio USA with 52 total samples collected. Aliquots of 500 mL were passed through membrane filters of decreasing pore sizes to enable all the water to pass through and capture bacteria. For each sample, the resulting filters were placed into two modified MacConkey (MAC) broths, one supplemented with 0.5 µg/mL of meropenem and 70 µg/mL of ZnSO4 and the other supplemented with 2 µg/mL cefotaxime. The inoculated broth was then incubated at 37° C overnight, after which they were streaked onto two types of correspondingly-modified MAC agar plates supplemented with 0.5 µg/mL and 1.0 µg/mL of meropenem and 70 µg/mL of ZnSO4 and incubated at 37°C overnight. The isolates were identified based on morphological and biochemical characteristics. Then, up to four distinct colonies of each isolate's pure culture per sample were tested for carbapenemase production using the Carba-NP test. Matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) MALDI-TOF MS was used to identify carbapenemase-producing organisms. In total 391 Carba-NP positive isolates were recovered from the 52 wastewater samples: 305 (78%) isolates had blaKPC, 73 (19%) carried blaNDM, and 14 (4%) harbored both blaKPC and blaNDM resistance genes. CPE genes of both blaKPC and blaNDM were recovered in both types of modified MAC broths, with 84 (21%) having a blaKPC gene, 22 (6%) carrying blaNDM and 9 (2%) harbored both a blaKPC and blaNDM of isolates recovered from MAC medium incorporated with 0.5ug/mL meropenem and 70ug/mL ZnSO4. The most prevalent isolates were Klebsiella pneumoniae, Escherichia coli, and Citrobacter spp.

Prevalence of colonization with multidrug-resistant bacteria in communities and hospitals in Kenya.

We estimated the prevalence of extended-spectrum cephalosporin-resistant Enterobacterales (ESCrE), carbapenem-resistant Enterobacterales (CRE), and methicillin-resistant Staphylococcus aureus (MRSA) in communities and hospitals in Kenya to identify human colonization with multidrug-resistant bacteria. Nasal and fecal specimen were collected from inpatients and community residents in Nairobi (urban) and Siaya (rural) counties. Swabs were plated on chromogenic agar to presumptively identify ESCrE, CRE and MRSA isolates. Confirmatory identification and antibiotic susceptibility testing were done using the VITEK®2 instrument. A total of 1999 community residents and 1023 inpatients were enrolled between January 2019 and March 2020. ESCrE colonization was higher in urban than rural communities (52 vs. 45%; P = 0.013) and in urban than rural hospitals (70 vs. 63%; P = 0.032). Overall, ESCrE colonization was ~ 18% higher in hospitals than in corresponding communities. CRE colonization was higher in hospital than community settings (rural: 7 vs. 1%; urban: 17 vs. 1%; with non-overlapping 95% confidence intervals), while MRSA was rarely detected (≤ 3% overall). Human colonization with ESCrE and CRE was common, particularly in hospitals and urban settings. MRSA colonization was uncommon. Evaluation of risk factors and genetic mechanisms of resistance can guide prevention and control efforts tailored to different environments.

Multi-Drug Resistant Staphylococcus aureus Carriage in Abattoir Workers in Busia, Kenya.

Abattoir workers have been identified as high-risk for livestock-associated Staphylococcus aureus carriage. This study investigated S. aureus carriage in abattoir workers in Western Kenya. Nasal swabs were collected once from participants between February-November 2012. S. aureus was isolated using bacterial culture and antibiotic susceptibility testing performed using the VITEK 2 instrument and disc diffusion methods. Isolates underwent whole genome sequencing and Multi Locus Sequence Types were derived from these data. S. aureus (n = 126) was isolated from 118/737 (16.0%) participants. Carriage was higher in HIV-positive (24/89, 27.0%) than HIV−negative participants (94/648, 14.5%; p = 0.003). There were 23 sequence types (STs) identified, and half of the isolates were ST152 (34.1%) or ST8 (15.1%). Many isolates carried the Panton-Valentine leucocidin toxin gene (42.9%). Only three isolates were methicillin resistant S. aureus (MRSA) (3/126, 2.4%) and the prevalence of MRSA carriage was 0.4% (3/737). All MRSA were ST88. Isolates from HIV-positive participants (37.0%) were more frequently resistant to sulfamethoxazole/trimethoprim compared to isolates from HIV-negative participants (6.1%; p < 0.001). Similarly, trimethoprim resistance genes were more frequently detected in isolates from HIV-positive (81.5%) compared to HIV-negative participants (60.6%; p = 0.044). S. aureus in abattoir workers were representative of major sequence types in Africa, with a high proportion being toxigenic isolates. HIV-positive individuals were more frequently colonized by antimicrobial resistant S. aureus which may be explained by prophylactic antimicrobial use.

Genomic epidemiology of Escherichia coli: antimicrobial resistance through a One Health lens in sympatric humans, livestock and peri-domestic wildlife in Nairobi, Kenya.

BACKGROUND: Livestock systems have been proposed as a reservoir for antimicrobial-resistant (AMR) bacteria and AMR genetic determinants that may infect or colonise humans, yet quantitative evidence regarding their epidemiological role remains lacking. Here, we used a combination of genomics, epidemiology and ecology to investigate patterns of AMR gene carriage in Escherichia coli, regarded as a sentinel organism. METHODS: We conducted a structured epidemiological survey of 99 households across Nairobi, Kenya, and whole genome sequenced E. coli isolates from 311 human, 606 livestock and 399 wildlife faecal samples. We used statistical models to investigate the prevalence of AMR carriage and characterise AMR gene diversity and structure of AMR genes in different host populations across the city. We also investigated household-level risk factors for the exchange of AMR genes between sympatric humans and livestock. RESULTS: We detected 56 unique acquired genes along with 13 point mutations present in variable proportions in human and animal isolates, known to confer resistance to nine antibiotic classes. We find that AMR gene community composition is not associated with host species, but AMR genes were frequently co-located, potentially enabling the acquisition and dispersal of multi-drug resistance in a single step. We find that whilst keeping livestock had no influence on human AMR gene carriage, the potential for AMR transmission across human-livestock interfaces is greatest when manure is poorly disposed of and in larger households. CONCLUSIONS: Findings of widespread carriage of AMR bacteria in human and animal populations, including in long-distance wildlife species, in community settings highlight the value of evidence-based surveillance to address antimicrobial resistance on a global scale. Our genomic analysis provided an in-depth understanding of AMR determinants at the interfaces of One Health sectors that will inform AMR prevention and control.