Unravelling patient pathways in the context of antibacterial resistance in East Africa.

BACKGROUND: A key factor driving the development and maintenance of antibacterial resistance (ABR) is individuals' use of antibiotics (ABs) to treat illness. To better understand motivations and context for antibiotic use we use the concept of a patient treatment-seeking pathway: a treatment journey encompassing where patients go when they are unwell, what motivates their choices, and how they obtain antibiotics. This paper investigates patterns and determinants of patient treatment-seeking pathways, and how they intersect with AB use in East Africa, a region where ABR-attributable deaths are exceptionally high. METHODS: The Holistic Approach to Unravelling Antibacterial Resistance (HATUA) Consortium collected quantitative data from 6,827 adult outpatients presenting with urinary tract infection (UTI) symptoms in Kenya, Tanzania, and Uganda between February 2019- September 2020, and conducted qualitative in-depth patient interviews with a subset (n = 116). We described patterns of treatment-seeking visually using Sankey plots and explored explanations and motivations using mixed-methods. Using Bayesian hierarchical regression modelling, we investigated the associations between socio-demographic, economic, healthcare, and attitudinal factors and three factors related to ABR: self-treatment as a first step, having a multi-step treatment pathway, and consuming ABs. RESULTS: Although most patients (86%) sought help from medical facilities in the first instance, many (56%) described multi-step, repetitive treatment-seeking pathways, which further increased the likelihood of consuming ABs. Higher socio-economic status patients were more likely to consume ABs and have multi-step pathways. Reasons for choosing providers (e.g., cost, location, time) were conditioned by wider structural factors such as hybrid healthcare systems and AB availability. CONCLUSION: There is likely to be a reinforcing cycle between complex, repetitive treatment pathways, AB consumption and ABR. A focus on individual antibiotic use as the key intervention point in this cycle ignores the contextual challenges patients face when treatment seeking, which include inadequate access to diagnostics, perceived inefficient public healthcare and ease of purchasing antibiotics without prescription. Pluralistic healthcare landscapes may promote more complex treatment seeking and therefore inappropriate AB use. We recommend further attention to healthcare system factors, focussing on medical facilities (e.g., accessible diagnostics, patient-doctor interactions, information flows), and community AB access points (e.g., drug sellers).

Biofilm formation, antimicrobial susceptibility and virulence genes of Uropathogenic Escherichia coli isolated from clinical isolates in Uganda.

INTRODUCTION: Uropathogenic E. coli is the leading cause of Urinary tract infections (UTIs), contributing to 80-90% of all community-acquired and 30-50% of all hospital-acquired UTIs. Biofilm forming Uropathogenic E. coli are associated with persistent and chronic inflammation leading to complicated and or recurrent UTIs. Biofilms provide an environment for poor antibiotic penetration and horizontal transfer of virulence genes which favors the development of Multidrug-resistant organisms (MDRO). Understanding biofilm formation and antimicrobial resistance determinants of Uropathogenic E. coli strains will provide insight into the development of treatment options for biofilm-associated UTIs. The aim of this study was to determine the biofilm forming capability, presence of virulence genes and antimicrobial susceptibility pattern of Uropathogenic E. coli isolates in Uganda. METHODS: This was a cross-sectional study carried in the Clinical Microbiology and Molecular biology laboratories at the Department of Medical Microbiology, Makerere University College of Health Sciences. We randomly selected 200 Uropathogenic E. coli clinical isolates among the stored isolates collected between January 2018 and December 2018 that had significant bacteriuria (> 105 CFU). All isolates were subjected to biofilm detection using the Congo Red Agar method and Antimicrobial susceptibility testing was performed using the Kirby disk diffusion method. The isolates were later subjected PCR for the detection of Urovirulence genes namely; Pap, Fim, Sfa, Afa, Hly and Cnf, using commercially designed primers. RESULTS: In this study, 62.5% (125/200) were positive biofilm formers and 78% (156/200) of these were multi-drug resistant (MDR). The isolates were most resistant to Trimethoprim sulphamethoxazole and Amoxicillin (93%) followed by gentamycin (87%) and the least was imipenem (0.5%). Fim was the most prevalent Urovirulence gene (53.5%) followed by Pap (21%), Sfa (13%), Afa (8%), Cnf (5.5%) and Hyl (0%). CONCLUSIONS: We demonstrate a high prevalence of biofilm-forming Uropathogenic E. coli strains that are highly associated with the MDR phenotype. We recommend routine surveillance of antimicrobial resistance and biofilm formation to understand the antibiotics suitable in the management of biofilm-associated UTIs.

Nasopharyngeal carriage, spa types and antibiotic susceptibility profiles of Staphylococcus aureus from healthy children less than 5 years in Eastern Uganda.

BACKGROUND: Staphylococcus aureus carriage is a known risk factor for staphylococcal disease. However, the carriage rates vary by country, demographic group and profession. This study aimed to determine the S. aureus carriage rate in children in Eastern Uganda, and identify S. aureus lineages that cause infection in Uganda. METHODS: Nasopharyngeal samples from 742 healthy children less than 5 years residing in the Iganga/Mayuge Health and Demographic Surveillance Site in Eastern Uganda were processed for isolation of S. aureus. Antibiotic susceptibility testing based on minimum inhibitory concentrations (MICs) was determined by the BD Phoenix™ system. Genotyping was performed by spa and SCCmec typing. RESULTS: The processed samples yielded 144 S. aureus isolates (one per child) therefore, the S. aureus carriage rate in children was 19.4% (144/742). Thirty one percent (45/144) of the isolates were methicillin resistant (MRSA) yielding a carriage rate of 6.1% (45/742). All isolates were susceptible to rifampicin, vancomycin and linezolid. Moreover, all MRSA were susceptible to vancomycin, linezolid and clindamycin. Compared to methicillin susceptible S. aureus (MSSA) isolates (68.8%, 99/144), MRSA isolates were more resistant to non-beta-lactam antimicrobials -trimethoprim/sulfamethoxazole 73.3% (33/45) vs. 27.3% (27/99) [p < 0.0001]; erythromycin 75.6% (34/45) vs. 24.2% (24/99) [p < 0.0001]; chloramphenicol 60% (27/45) vs. 19.2% (19/99) [p < 0.0001]; gentamicin 55.6% (25/45) vs. 25.3% (25/99) [p = 0.0004]; and ciprofloxacin 35.6% (16/45) vs. 2% (2/99) [p < 0.0001]. Furthermore, 42 MRSA (93.3%) were multidrug resistant (MDR) and one exhibited high-level resistance to mupirocin. Overall, 61 MSSA (61.6%) were MDR, including three mupirocin and clindamycin resistant isolates. Seven spa types were detected among MRSA, of which t037 and t064 were predominant and associated with SCCmec types I and IV, respectively. Fourteen spa types were detected in MSSA which consisted mainly of t645 and t4353. CONCLUSIONS: S. aureus carriage rate in healthy children in Eastern Uganda is high and comparable to rates for hospitalized patients in Kampala. The detection of mupirocin resistance is worrying as it could rapidly increase if mupirocin is administered in a low-income setting. S. aureus strains of spa types t064, t037 (MRSA) and t645, t4353 (MSSA) are prevalent and could be responsible for majority of staphylococcal infections in Uganda.

Epidemiology and antibiotic susceptibility of Vibrio cholerae associated with the 2017 outbreak in Kasese district, Uganda.

BACKGROUND: Uganda is among the 51 countries where cholera outbreaks are common with epidemics occurring predominantly along the western border with Democratic Republic of Congo (DRC), Kampala city slums, Busia district which is a border town with Western Kenya, Mbale district and the Karamoja Sub-region. This report summarizes findings from the epidemiologic investigation, which aimed at identifying the mode of transmission and antibiotic susceptibility patterns of the Vibrio cholerae isolated in Kasese district, Uganda. METHODS: A descriptive cross-sectional study was carried out between 2017 and 2018 to describe the epidemiology of the cholera epidemic in Kasese district, Uganda. Rectal swabs were collected from 69 suspected case-persons and cultured on Thiosulphate-Citrate-Bile-Salts Sucrose (TCBS™; SEIKEN Japan) agar and incubated at 37 °C for 18-24 h. The isolates were serotyped with polyvalent 01 antiserum and monovalent serotype Inaba and Ogawa antisera (Denka Seiken, Tokyo, Japan) to determine which serotype was responsible for the outbreak. Antimicrobial susceptibility testing was performed using the Kirby-Bauer disk diffusion method on Mueller-Hinton agar. A list of discharged patients was obtained from the isolation units of Bwera hospital and Kagando hospital and the individuals were followed to the community where they live. Questionnaires were administered to a total of 75 participants who were either the cases or relatives to the case. GPS points of the homes of the cases and pictures of potential source infection were also taken and cases were mapped. RESULTS: A total of 222 cases were recorded in the Kasese District outbreak between the month of September 2017 and January 2018 with the case fatality rate (CFR) of 1.4%. Children below the age of 14 years contributed the biggest proportion of the cases (70%) and out of these, 33% were aged below 5 years. Culture isolated 69 V. cholerae 01 serotype Inaba from the total of 71 samples. Salmonella typhi was Isolated from the other two samples which were negative for V. cholerae. Antibiotic susceptibility using Kirby-Bauer disc diffusion method was done on isolates from 69 participants and showed 100% resistance to Ampicillin and over 50% were resistant to trimethoprim/Sulfamethoxazole whereas gentamicin showed 100% susceptibility. Environmental assessment revealed rampant cases of open defecation. CONCLUSION: Though we did not culture water to confirm contamination with Vibrio cholerae, we hypothesize that the cholera epidemic in Kasese 2017 was sparked off by consumption of contaminated water following the heavy floods that washed away latrines into water sources in Bwera, Isango and Nakiyumbu sub-counties. V. cholerae was also highly resistant to the commonly used antibiotics.

Mycobacterium tuberculosis thymidylate kinase antigen assays for designating incipient, high-risk latent M.tb infection.

BACKGROUND: Precise designation of high risk forms of latent Mycobacterium tuberculosis-M.tb infections (LTBI) is impossible. Delineation of high-risk LTBI can, however, allow for chemoprophylaxis and curtail majority cases of active tuberculosis (ATB). There is epidemiological evidence to support the view that LTBI in context of HIV-1 co-infection is high-risk for progression to ATB relative to LTBI among HIV-ve persons. We recently showed that assays of M.tb thymidylate kinase (TMKmt) antigen and host specific IgG can differentiate ATB from LTBI and or no TB (NTB, or healthy controls). In this study, we aimed to expose the differential levels of TMKmt Ag among HIV+ve co-infected LTBI relative to HIV-ve LTBI as a strategy to advance these assays for designating incipient LTBI. METHODS: TMKmt host specific IgM and IgG detection Enzyme Immuno-Assays (EIA) were conducted on 40 TB exposed house-hold contacts (22 LTBI vs. 18 no TB (NTB) by QunatiFERON-TB GOLD®); and TMKmt Ag detection EIA done on 82 LTBI (46 HIV+ve vs 36 HIV-ve) and 9 NTB (American donors). Purified recombinant TMKmt protein was used as positive control for the Ag assays. RESULTS: IgM levels were found to be equally low across QuantiFERON-TB GOLD® prequalified NTB and TB exposed house-hold contacts. Higher TMKmt host specific IgG trends were found among TB house-hold contacts relative to NTB controls. TMKmt Ag levels among HIV+ve LTBI were 0.2676 ± 0.0197 (95% CI: 0.2279 to 0.3073) relative to 0.1069 ± 0.01628 (95% CI: 0.07385 to 0.14) for HIV-ve LTBI (supporting incipient nature of LTBI in context of HIV-1 co-infection). NTB had TMKmt Ag levels of 0.1013 ± 0.02505 (5% CI: 0.0421 to 0.1606) (intimating that some were indeed LTBI). CONCLUSIONS: TMKmt Ag levels represent a novel surrogate biomarker for high-risk LTBI, while host-specific IgG can be used to designate NTB from LTBI.

Prevalence and molecular characteristics of Staphylococcus aureus, including methicillin resistant strains, isolated from bulk can milk and raw milk products in pastoral communities of South-West Uganda.

BACKGROUND: Staphylococcus aureus strains are now regarded as zoonotic agents. In pastoral settings where human-animal interaction is intimate, multi-drug resistant microorganisms have become an emerging zoonotic issue of public health concern. The study of S. aureus prevalence, antimicrobial resistance and clonal lineages in humans, animals and food in African settings has great relevance, taking into consideration the high diversity of ethnicities, cultures and food habits that determine the lifestyle of the people. Little is known about milk carriage of methicillin resistant S. aureus strains (MRSA) and their virulence factors in Uganda. Here, we present the prevalence of MRSA in bulk can milk and raw milk products in pastoral communities of south-west Uganda. We also present PFGE profiles, spa-types, as well as frequency of enterotoxins genes. METHODS: S. aureus was identified by the coagulase test, susceptibility testing by the Kirby-Bauer disc diffusion and E-test methods and MRSA by detection of the mecA gene and SCCmec types. The presence of Panton - Valentine Leucocidin (PVL) genes and staphylococcal enterotoxins was determined by PCR, while genotyping was by PFGE and spa typing. RESULTS: S. aureus were isolated from 30/148 (20.3%) milk and 11/91(12%) sour milk samples. mecA gene carriage, hence MRSA, was detected in 23/41 (56.1%) of the isolates, with 21 of the 23 (91.3%) being SCCmec type V; while up to 30/41 (73.2%) of the isolates were resistant to tetracycline. Only five isolates carried the PVL virulence gene, while PFGE typing revealed ten clusters (ranging from two seven isolates each) that comprised 83% of the sample, and only eight isolates with unique pulsotypes. The largest PFGE profile (E) consisted of seven isolates while t7753, t1398, and t2112 were the most common spa-types. Thirty seven of the 41 strains (90.2%) showed at least one of the eight enterotoxin genes tested, with sem 29 (70.7%), sei 25 (61%) and seg 21 (51.2%) being the most frequently observed genes. CONCLUSION: This is the first study to demonstrate MRSA and enterotoxin genes in raw milk and its products in Uganda. The fact that over 90% of the isolates carried at least one gene encoding enterotoxins shows a high risk of spread of foodborne diseases through milk in this setting.

Molecular epidemiology of Panton-Valentine Leukocidin-positive community-acquired methicillin resistant Staphylococcus aureus isolates in pastoral communities of rural south western Uganda.

BACKGROUND: The emergence of multidrug resistant Staphylococcus aureus strains, including methicillin resistant (MRSA), is a global concern. Treatment of bacterial infections in Uganda's health care settings is largely empirical, rarely accompanied by laboratory confirmation. Here we show the burden, characteristics of MRSA and epidemiology of Panton-Valentine Leukocidin (PVL) positive strains in asymptomatic carriers in pastoral households of south-west Uganda. METHODS: Nasal swabs from 253 participants were cultured following standard methodology. MRSA strains were identified by detection of the mecA gene and SCCmec typing, and PVL genes detected by PCR. Pulsed Field Gel Electrophoresis (PFGE) was done to evaluate possible transmission patterns. Spa typing of PVL positive isolates was done to study the epidemiology of virulent strains in this setting. RESULTS: S. aureus was isolated in 29% (n = 73) of the participants, of which 48 were MRSA by mecA typing. PVL-encoding genes were found in 49.3% (n = 36) of the 73 isolates, of which 25 were also mecA positive. Among the PVL negative strains (n = 37), 62.2% (n = 23) carried the mecA gene. The most common SCCmec type was V, detected in 39 (18 PVL positive and 21 PVL negative) isolates. PFGE clustered 21/36 (58.3%) PVL positive isolates divided in four pulsotypes and 18/37 (48.6%) PVL negative isolates divided in eight pulsotypes. The most prevalent Spa types were t318 (26.5%, n = 9) and t645 (20.6%, n = 7); while other common Spa types were t11656 (n = 3), t127 (n = 3) and t355 (n = 3). CONCLUSION: The study shows a high prevalence of community acquired (CA)-MRSA, and PVL-positive isolates with two predominant spa types in rural Uganda, further complicating infection control strategies in these underprivileged communities.

Noninvasive test for tuberculosis detection among primates.

Traditional testing methods have limited epidemiologic studies of tuberculosis among free-living primates. PCR amplification of insertion element IS6110 of Mycobacterium tuberculosis from fecal samples was evaluated as a noninvasive screening test for tuberculosis in primates. Active tuberculosis was detected among inoculated macaques and naturally exposed chimpanzees, demonstrating the utility of this test.

The T2 Mycobacterium tuberculosis genotype, predominant in Kampala, Uganda, shows negative correlation with antituberculosis drug resistance.

Surveillance of the circulating Mycobacterium tuberculosis complex (MTC) strains in a given locality is important for understanding tuberculosis (TB) epidemiology. We performed molecular epidemiological studies on sputum smear-positive isolates that were collected for anti-TB drug resistance surveillance to establish the variability of MTC lineages with anti-TB drug resistance and HIV infection. Spoligotyping was performed to determine MTC phylogenetic lineages. We compared patients' MTC lineages with drug susceptibility testing (DST) patterns and HIV serostatus. Out of the 533 isolates, 497 (93.2%) had complete DST, PCR, and spoligotyping results while 484 (90.1%) participants had results for HIV testing. Overall, the frequency of any resistance was 75/497 (15.1%), highest among the LAM (34.4%; 95% confidence interval [CI], 18.5 to 53.2) and lowest among the T2 (11.5%; 95% CI, 7.6 to 16.3) family members. By multivariate analysis, LAM (adjusted odds ratio [OR(adj)], 5.0; 95% CI, 2.0 to 11.9; P < 0.001) and CAS (OR(adj), 2.9; 95% CI, 1.4.0 to 6.3; P = 0.006) families were more likely to show any resistance than was T2. All other MTC lineages combined were more likely to be resistant to any of the anti-TB drugs than were the T2 strains (OR(adj), 1.7; 95% CI, 1.0 to 2.9; P = 0.040). There were no significant associations between multidrug resistance and MTC lineages, but numbers of multidrug-resistant TB strains were small. No association was established between MTC lineages and HIV status. In conclusion, the T2 MTC lineage negatively correlates with anti-TB drug resistance, which might partly explain the reported low levels of anti-TB drug resistance in Kampala, Uganda. Patients' HIV status plays no role with respect to the MTC lineage distribution.

Clinico-pathological features of tuberculosis due to Mycobacterium tuberculosis Uganda genotype in patients with tuberculous lymphadenitis: a cross sectional study.

BACKGROUND: Tuberculous lymphadenitis is next to pulmonary tuberculosis as the most common cause of tuberculosis. Uganda genotype, one of the sub-lineages of Mycobacterium tuberculosis, is the most prevalent cause of pulmonary tuberculosis in Uganda. We here investigate the clinicopathological characteristics of patients with tuberculous lymphadenitis infected with M. tuberculosis Uganda genotype compared with those infected with M. tuberculosis non-Uganda genotype strains. METHODS: Between 2010 and 2012, we enrolled 121 patients (mean age 28.5 yrs, male 48%; female 52%) with tuberculous lymphadenitis, and categorized them by their M. tuberculosis genotypes. The clinical features and lymph node cytopathological parameters were compared between patients in the Uganda and non-Uganda categories using a crude and multivariable logistic regression model with adjustment for confounding factors. RESULTS: Of the 121participants, 56 (46%) were infected with strains of Uganda genotype. Patients infected with this genotype had significantly lower frequency of abdominal lymphadenopathy (odds ratio 0.4, p = 0.046) after adjusting for sex, age and HIV. Abdominal lymphadenopathy was also significantly associated with abnormal chest X-ray (p = 0.027). CONCLUSION: Tuberculous lymphadenitis patients infected with M. tuberculosis Uganda genotype were significantly less prone to have abdominal lymphadenopathy indicating potential reduced ability to disseminate and supporting the concept that differences in M. tuberculosis genotype may have clinical implications.

Knowledge and perceptions of brucellosis in the pastoral communities adjacent to Lake Mburo National Park, Uganda.

BACKGROUND: Brucellosis is one of the most common zoonotic infections globally. Lack of knowledge about brucellosis may affect the health-seeking behavior of patients, thus leading to sustained transmission in these communities. Our study assessed knowledge and perceptions of brucellosis among pastoral communities adjacent to Lake Mburo National Park (LMNP), Kiruhura District, Uganda. METHODS: A community cross-sectional questionnaire survey involving 371 randomly selected household heads from three sub-counties neighboring LMNP were interviewed between June and August 2012. Data collected included communities' knowledge on causes, symptoms, transmission, treatment, prevention and risk factors of brucellosis. Multivariable logistic regression analysis was performed to explore strength of association between overall knowledge of brucellosis and various individual factors using odds ratios and 95% confidence intervals. RESULTS: Only 70 (19%) knew the symptoms of brucellosis in animals, and three quarters (279, 75.5%) mentioned joint and muscle pain as a common symptom in humans. Almost all participants (370, 99.3%) had ever heard about brucellosis, majority (311, 84.7%) believed it affects all sexes and two thirds (67.7%) of the respondents believed close proximity to wildlife contributes to the presence of the disease. Almost all (352, 95.4%) knew that brucellosis in humans could be treatable using modern drugs. The main routes of infection in humans such as consumption of unpasteurized dairy products were known by 97% (360/371); eating of half-cooked meat by 91.4% and eating contaminated pasture in animals by 97.4%. There was moderate overall knowledge of brucellosis 197 (53.1%). Factors associated with higher overall knowledge were being agro-pastoralists (aOR: 2.08, CI: 1.17-3.71) compared to pure pastoralists while those who reported that the disease was a health problem (aOR: 0.18, CI: 0.06-0.56) compared to those who said it was not were less likely to be knowledgeable. CONCLUSIONS: There was moderate overall knowledge of human and animal brucellosis among the participants. Majority of the participants believed that close proximity to wildlife contributes to the presence of the disease in the area. There is a need for collaboration between the public health, veterinary and wildlife sectors to provide health education on brucellosis for better management of the disease in the communities.

Early changes in the gut microbiome among HIV-infected Individuals in Uganda initiating daily TMP/SMX.

Daily cotrimoxazole (TMP/SXT) prophylaxis is part of the HIV treatment package for all new HIV-infected individuals in Uganda. Although this treatment has shown reduced morbidity and mortality in HIV, it remains controversial due to its contribution to developing antibiotic-resistant bacteria. Moreover, the effects of daily use of a broad-spectrum antibiotic on the gut microbiome remain unknown. To study the early effects, we analysed shotgun metagenome sequence data from stool samples of five newly HIV-infected individuals initiating TMP/SXT prophylaxis longitudinally for the first 30 days of treatment. Using shotgun metagenomics sequencing, we generated both taxonomic and functional profiles from each patient and compared gut microbial changes Pre- TMP/SXT and post-TMP/SXT on Day 5, Day 14, and Day 30. Daily TMP/SXT prophylaxis resulted in a shift characterised by an enrichment of Prevetollea and Ruminococcus genera members and the depletion of Lactococcus and Bacteroides genera members. Furthermore, these microbial shifts were associated with changes in the functional profile revealed by a differential abundance of pathways of amino acid metabolism, carbohydrate metabolism, and nucleotide biosynthesis linked to members of the Bacteroidaceae and Enterobacteriaceae families. TMP/SXT daily prophylaxis in HIV-infected individuals is associated with dramatic changes in microbial composition and functional profiles; however, other factors such as Age, Gender, HIV clinical stage, and ART regiment are at play. Further investigation is needed to examine the implication of these shifts on clinical management and outcomes among HIV patients.

Evaluation of three protocols for direct susceptibility testing for gram negative-Enterobacteriaceae from patient samples in Uganda with SMS reporting.

In Uganda, the challenge of generating and timely reporting essential antimicrobial resistance (AMR) data has led to overreliance on empirical antibiotic therapy, exacerbating the AMR crisis. To address this issue, this study aimed to adapt a one-step AMR testing protocol alongside an SMS (Short Message Service) result relay system (SRRS), with the potential to reduce the turnaround time for AMR testing and result communication from 4 days or more to 1 day in Ugandan clinical microbiology laboratories. Out of the 377 samples examined, 54 isolates were obtained. Notably, E. coli (61%) and K. pneumoniae (33%) were the most frequently identified, majority testing positive for ESBL. Evaluation of three AMR testing protocols revealed varying sensitivity and specificity, with Protocol A (ChromID ESBL-based) demonstrating high sensitivity (100%) but no calculable specificity, Protocol B (ceftazidime-based) showing high sensitivity (100%) and relatively low specificity (7.1%), and Protocol C (cefotaxime-based) exhibiting high sensitivity (97.8%) but no calculable specificity. ESBL positivity strongly correlated with resistance to specific antibiotics, including cefotaxime, ampicillin, and aztreonam (100%), cefuroxime (96%), ceftriaxone (93%), and trimethoprim sulfamethoxazole (87%). The potential of integrating an SRRS underscored the crucial role this could have in enabling efficient healthcare communication in AMR management. This study underscores the substantial potential of the tested protocols for accurately detecting ESBL production in clinical samples, potentially, providing a critical foundation for predicting and reporting AMR patterns. Although considerations related to specificity warrant careful assessment before widespread clinical adoption.

Treatment seeking and antibiotic use for urinary tract infection symptoms in the time of COVID-19 in Tanzania and Uganda.

Background: There is still little empirical evidence on how the outbreak of coronavirus disease 2019 (COVID-19) and associated regulations may have disrupted care-seeking for non-COVID-19 conditions or affected antibiotic behaviours in low- and middle-income countries (LMICs). We aimed to investigate the differences in treatment-seeking behaviours and antibiotic use for urinary tract infection (UTI)-like symptoms before and during the pandemic at recruitment sites in two East African countries with different COVID-19 control policies: Mbarara, Uganda and Mwanza, Tanzania. Methods: In this repeated cross-sectional study, we used data from outpatients (pregnant adolescents aged >14 and adults aged >18) with UTI-like symptoms who visited health facilities in Mwanza, Tanzania and Mbarara, Uganda. We assessed the prevalence of self-reported behaviours (delays in care-seeking, providers visited, antibiotics taken) at three different time points, labelled as 'pre-COVID-19 phase' (February 2019 to February 2020), 'COVID-19 phase 1' (March 2020 to April 2020), and 'COVID-19 phase 2' (July 2021 to February 2022). Results: In both study sites, delays in care-seeking were less common during the pandemic than they were in the pre-COVID phase. Patients in Mwanza, Tanzania had shorter care-seeking pathways during the pandemic compared to before it, but this difference was not observed in Mbarara, Uganda. Health centres were the dominant sources of antibiotics in both settings. Over time, reported antibiotic use for UTI-like symptoms became more common in both settings. During the COVID-19 phases, there was a significant increase in self-reported use of antibiotics like metronidazole (<30% in the pre-COVID-19 phase to 40% in COVID phase 2) and doxycycline (30% in the pre-COVID-19 phase to 55% in COVID phase 2) that were not recommended for treating UTI-like symptoms in the National Treatment Guidelines in Mbarara, Uganda. Conclusions: There was no clear evidence that patients with UTI-like symptoms attending health care facilities had longer or more complex treatment pathways despite strict government-led interventions related to COVID-19. However, antibiotic use increased over time, including some antibiotics not recommended for treating UTI, which has implications for future antimicrobial resistance.

Predominance of multidrug-resistant bacteria causing urinary tract infections among symptomatic patients in East Africa: a call for action.

Background: In low- and middle-income countries, antibiotics are often prescribed for patients with symptoms of urinary tract infections (UTIs) without microbiological confirmation. Inappropriate antibiotic use can contribute to antimicrobial resistance (AMR) and the selection of MDR bacteria. Data on antibiotic susceptibility of cultured bacteria are important in drafting empirical treatment guidelines and monitoring resistance trends, which can prevent the spread of AMR. In East Africa, antibiotic susceptibility data are sparse. To fill the gap, this study reports common microorganisms and their susceptibility patterns isolated from patients with UTI-like symptoms in Kenya, Tanzania and Uganda. Within each country, patients were recruited from three sites that were sociodemographically distinct and representative of different populations. Methods: UTI was defined by the presence of >104 cfu/mL of one or two uropathogens in mid-stream urine samples. Identification of microorganisms was done using biochemical methods. Antimicrobial susceptibility testing was performed by the Kirby-Bauer disc diffusion assay. MDR bacteria were defined as isolates resistant to at least one agent in three or more classes of antimicrobial agents. Results: Microbiologically confirmed UTI was observed in 2653 (35.0%) of the 7583 patients studied. The predominant bacteria were Escherichia coli (37.0%), Staphylococcus spp. (26.3%), Klebsiella spp. (5.8%) and Enterococcus spp. (5.5%). E. coli contributed 982 of the isolates, with an MDR proportion of 52.2%. Staphylococcus spp. contributed 697 of the isolates, with an MDR rate of 60.3%. The overall proportion of MDR bacteria (n = 1153) was 50.9%. Conclusions: MDR bacteria are common causes of UTI in patients attending healthcare centres in East African countries, which emphasizes the need for investment in laboratory culture capacity and diagnostic algorithms to improve accuracy of diagnosis that will lead to appropriate antibiotic use to prevent and control AMR.

Intersecting social and environmental determinants of multidrug-resistant urinary tract infections in East Africa beyond antibiotic use.

The global health crisis of antibacterial resistance (ABR) poses a particular threat in low-resource settings like East Africa. Interventions for ABR typically target antibiotic use, overlooking the wider set of factors which drive vulnerability and behaviours. In this cross-sectional study, we investigated the joint contribution of behavioural, environmental, socioeconomic, and demographic factors associated with higher risk of multi-drug resistant urinary tract infections (MDR UTIs) in Kenya, Tanzania, and Uganda. We sampled outpatients with UTI symptoms in healthcare facilities and linked their microbiology data with patient, household and community level data. Using bivariate statistics and Bayesian profile regression on a sample of 1610 individuals, we show that individuals with higher risk of MDR UTIs were more likely to have compound and interrelated social and environmental disadvantages: they were on average older, with lower education, had more chronic illness, lived in resource-deprived households, more likely to have contact with animals, and human or animal waste. This suggests that interventions to tackle ABR need to take account of intersectional socio-environmental disadvantage as a priority.

Treatment seeking behaviours, antibiotic use and relationships to multi-drug resistance: A study of urinary tract infection patients in Kenya, Tanzania and Uganda.

Antibacterial resistance (ABR) is a major public health threat. An important accelerating factor is treatment-seeking behaviour, including inappropriate antibiotic (AB) use. In many low- and middle-income countries (LMICs) this includes taking ABs with and without prescription sourced from various providers, including health facilities and community drug sellers. However, investigations of complex treatment-seeking, AB use and drug resistance in LMICs are scarce. The Holistic Approach to Unravel Antibacterial Resistance in East Africa (HATUA) Consortium collected questionnaire and microbiological data from adult outpatients with urinary tract infection (UTI)-like symptoms presenting at healthcare facilities in Kenya, Tanzania and Uganda. Using data from 6,388 patients, we analysed patterns of self-reported treatment seeking behaviours ('patient pathways') using process mining and single-channel sequence analysis. Among those with microbiologically confirmed UTI (n = 1,946), we used logistic regression to assess the relationship between treatment seeking behaviour, AB use, and the likelihood of having a multi-drug resistant (MDR) UTI. The most common treatment pathway for UTI-like symptoms in this sample involved attending health facilities, rather than other providers like drug sellers. Patients from sites in Tanzania and Uganda, where over 50% of patients had an MDR UTI, were more likely to report treatment failures, and have repeat visits to providers than those from Kenyan sites, where MDR UTI proportions were lower (33%). There was no strong or consistent relationship between individual AB use and likelihood of MDR UTI, after accounting for country context. The results highlight the hurdles East African patients face in accessing effective UTI care. These challenges are exacerbated by high rates of MDR UTI, suggesting a vicious cycle of failed treatment attempts and sustained selection for drug resistance. Whilst individual AB use may contribute to the risk of MDR UTI, our data show that factors related to context are stronger drivers of variations in ABR.

Long-term dominance of Mycobacterium tuberculosis Uganda family in peri-urban Kampala-Uganda is not associated with cavitary disease.

BACKGROUND: Previous studies have shown that Mycobacterium tuberculosis (MTB) Uganda family, a sub-lineage of the MTB Lineage 4, is the main cause of tuberculosis (TB) in Uganda. Using a well characterized patient population, this study sought to determine whether there are clinical and patient characteristics associated with the success of the MTB Uganda family in Kampala. METHODS: A total of 1,746 MTB clinical isolates collected from 1992-2009 in a household contact study were genotyped. Genotyping was performed using Single Nucleotide Polymorphic (SNP) markers specific for the MTB Uganda family, other Lineage 4 strains, and Lineage 3, respectively. Out of 1,746 isolates, 1,213 were from patients with detailed clinical data. These data were used to seek associations between MTB lineage/sub-lineage and patient phenotypes. RESULTS: Three MTB lineages were found to dominate the MTB population in Kampala during the last two decades. Overall, MTB Uganda accounted for 63% (1,092/1,746) of all cases, followed by other Lineage 4 strains accounting for 22% (394/1,746), and Lineage 3 for 11% (187/1,746) of cases, respectively. Seventy-three (4 %) strains remained unclassified. Our longitudinal data showed that MTB Uganda family occurred at the highest frequency during the whole study period, followed by other Lineage 4 strains and Lineage 3. To explore whether the long-term success of MTB Uganda family was due to increased virulence, we used cavitary disease as a proxy, as this form of TB is the most transmissible. Multivariate analysis revealed that even though cavitary disease was associated with known risk factors such as smoking (adjusted odds ratio (aOR) 4.8, 95% confidence interval (CI) 3.33-6.84) and low income (aOR 2.1, 95% CI 1.47-3.01), no association was found between MTB lineage and cavitary TB. CONCLUSION: The MTB Uganda family has been dominating in Kampala for the last 18 years, but this long-term success is not due to increased virulence as defined by cavitary disease.

Species and genotypic diversity of non-tuberculous mycobacteria isolated from children investigated for pulmonary tuberculosis in rural Uganda.

BACKGROUND: Smear microscopy, a mainstay of tuberculosis (TB) diagnosis in developing countries, cannot differentiate M. tuberculosis complex from NTM infection, while pulmonary TB shares clinical signs with NTM disease, causing clinical and diagnostic dilemmas. This study used molecular assays to identify species and assess genotypic diversity of non-tuberculous mycobacteria (NTM) isolates from children investigated for pulmonary tuberculosis at a demographic surveillance site in rural eastern Uganda. METHODS: Children were investigated for pulmonary tuberculosis as part of a TB vaccine surveillance program (2009-2011). Two cohorts of 2500 BCG vaccinated infants and 7000 adolescents (12-18 years) were recruited and followed up for one to two years to determine incidence of tuberculosis. Induced sputum and gastric aspirates were processed by the standard N-acetyl L-cysteine (NALC)-NaOH method. Sediments were cultured in the automated MGIT (Becton Dickson) liquid culture system and incubated at 37°C for at least six weeks. Capilia TB assay was used to classify mycobacteria into MTC and NTM. The GenoType CM/AS assays were performed to identify species while Enterobacterial Repetitive Intergenic Consensus (ERIC) PCR genotyping was used to assess genetic diversity of the strains within each species. RESULTS: Among 2859 infants and 2988 adolescents screened, the numbers of TB suspects were 710 and 1490 infants and adolescents respectively. The prevalence of NTM in infant suspects was 3.7% (26/710) (95% CI 2.5-5.2) while that in adolescent suspects was 4.6% (69/1490) (95% CI 3.6-5.8). On culture, 127 isolates were obtained, 103 of which were confirmed as mycobacteria comprising of 95 NTM and eight M. tuberculosis complex. The Genotype CM/AS assay identified 63 of the 95 NTM isolates while 32 remained un-identified. The identified NTM species were M. fortuitum (40 isolates, 63.5%), M. szulgai (9 isolates, 14.3%), M. gordonae (6 isolates, 9.5%), M. intracellulare (3 isolates, 4.7%), M. scrofulaceum (2 isolates, 3.2%), M. lentiflavum (2 isolates, 3.2%), and M. peregrinum (1 isolate, 1.6%). Genotyping did not reveal any clustering in M. intracellulare, M. gordonae and M. szulgai species. M. fortuitum, on the other hand, had two clusters, one with three isolates of M. fortuitum 1 and the other with two isolates of M. fortuitum 2 subspecies. The remaining 35 of the 40 isolates of M. fortuitum had unique fingerprint patterns. CONCLUSION: M. fortuitum is the most common cause of infection by NTM among Infants and adolescents in rural Uganda. There is a varied number of species and genotypes, with minimal clustering within species, suggesting ubiquitous sources of infection to individuals in this community.

Klebsiella pneumoniae carbapenamases in Escherichia coli isolated from humans and livestock in rural south-western Uganda.

BACKGROUND: The accumulation of resistance genes in Escherichia coli (E. coli) strains imposes limitations in the therapeutic options available for the treatment of infections caused by E.coli. Production of Klebsiella pneumoniae carbapenemase (KPC) by E. coli renders it resistant to broad-spectrum ß-lactam antibiotics. Globally there is existing evidence of spread of carbapenem-resistant E. coli in both humans and livestock driven by acquisition of the several other carbapenemase genes. Overall, there is little information regarding the extent of KPC gene distribution in E. coli. We set out to determine the prevalence, and evaluate the phenotypic and genotypic patterns of KPC in E. coli isolated from humans and their livestock in rural south western Uganda. METHODS: A laboratory-based, descriptive cross-sectional study was conducted involving 96 human and 96 livestock isolates collected from agro-pastoralist communities in Mbarara district in south western Uganda. Phenotypic and molecular methods (PCR) were used for presence and identification of KPC genes in the E. coli isolates. A chi-square test of independence was used to evaluate the differences in resistant patterns between carbapenems and isolates. RESULTS: The overall prevalence of carbapenem resistance by disk diffusion susceptibility testing (DST) for both humans and livestock isolates were 41.7% (80/192). DST-based resistance was identical in both human and livestock isolates (41.7%). The prevalence of carbapenem resistance based on Modified Hodge Test (MHT) was 5% (2/40) and 10% (4/40) for humans and livestock isolates respectively. Both human and livestock isolates, 48.7% (95/192) had the KPC gene, higher than phenotypic expression; 41.7% (80/192). blaKPC gene prevalence was overall similar in human isolates (51%; 49/96) vs livestock isolates (47.9%; 46/96). Approximately, 19% (15/80) of the isolates were phenotypically resistant to carbapenems and over 70% (79/112) of the phenotypically sensitive strains harbored the blaKPC gene. CONCLUSION: Our results suggest that both human and livestock isolates of E. coli in our setting carry the blaKPC gene with a high percentage of strains not actively expressing the blaKPC gene. The finding of fewer isolates carrying the KPC gene than those phenotypically resistant to carbapenems suggests that other mechanisms are playing a role in this phenomenon, calling for further researcher into this phenomenon.