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.

blaVIM- and blaOXA-mediated carbapenem resistance among Acinetobacter baumannii and Pseudomonas aeruginosa isolates from the Mulago hospital intensive care unit in Kampala, Uganda.

BACKGROUND: Between January 2015 and July 2017, we investigated the frequency of carbapenem resistant Acinetobacter baumannii (CRAB) and carbapenem resistant Pseudomonas aeruginosa (CRPA) at the Mulago Hospital intensive care unit (ICU) in Kampala, Uganda. Carbapenemase production and carbapenemase gene carriage among CRAB and CRPA were determined; mobility potential of carbapenemase genes via horizontal gene transfer processes was also studied. METHODS: Clinical specimens from 9269 patients were processed for isolation of CRAB and CRPA. Drug susceptibility testing was performed with the disk diffusion method. Carriage of carbapenemase genes and class 1 integrons was determined by PCR. Conjugation experiments that involved blaVIM positive CRAB/CRPA (donors) and sodium azide resistant Escherichia coli J53 (recipient) were performed. RESULTS: The 9269 specimens processed yielded 1077 and 488 isolates of Acinetobacter baumannii and Pseudomonas aeruginosa, respectively. Of these, 2.7% (29/1077) and 7.4% (36/488) were confirmed to be CRAB and CRPA respectively, but 46 were available for analysis (21 CRAB and 25 CRPA). Majority of specimens yielding CRAB and CRPA were from the ICU (78%) while 20 and 2% were from the ENT (Ear Nose & Throat) Department and the Burns Unit, respectively. Carbapenemase assays performed with the MHT assay showed that 40 and 33% of CRPA and CRAB isolates respectively, were carbapenemase producers. Also, 72 and 48% of CRPA and CRAB isolates respectively, were metallo-beta-lactamase producers. All the carbapenemase producing isolates were multidrug resistant but susceptible to colistin. blaVIM was the most prevalent carbapenemase gene, and it was detected in all CRAB and CRPA isolates while blaOXA-23 and blaOXA-24 were detected in 29 and 24% of CRAB isolates, respectively. Co-carriage of blaOXA-23 and blaOXA-24 occurred in 14% of CRAB isolates. Moreover, 63% of the study isolates carried class 1 integrons; of these 31% successfully transferred blaVIM to E. coli J53. CONCLUSIONS: CRAB and CRPA prevalence at the Mulago Hospital ICU is relatively low but carbapenemase genes especially blaVIM and blaOXA-23 are prevalent among them. This requires strengthening of infection control practices to curb selection and transmission of these strains in the hospital.

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.

Species, antibiotic susceptibility profiles and van gene frequencies among enterococci isolated from patients at Mulago National Referral Hospital in Kampala, Uganda.

BACKGROUND: The increase in drug resistance to affordable antibiotics used to treat Gram positive bacterial infections has complicated the management of enterococcal infections. Resistance to vancomycin, one of the most powerful antibiotics, is of utmost concern as both intrinsic and acquired forms of resistance do occur in enterococci. This cross-sectional study aimed to determine the species, antibiotic susceptibility profiles and vanA/vanB gene frequencies among enterococci isolated from patients at Mulago Hospital in Kampala, Uganda. METHODS: Between November 2011 and October 2012, stool, urine, sputum and blood samples, as well as vaginal, endocervical, pus, ear and urethra swabs from 3229 patients were processed for isolation of bacteria, yielding 162 enterococci of which 115 were available for analysis (one isolate per specimen/patient). Species-level confirmation and susceptibility testing were determined with the Phoenix™ AST/ID Automated System, while vanA/vanB gene carriage was determined by PCR. RESULTS: Species-level identification revealed 72 isolates of E. faecalis, 20 E. gallinarum/casseliflavus, 5 E. faecium, 4 E. raffinosus and 2 isolates each for E. hirae and E. durans. Ten isolates could not be identified to species level. Antibiotic resistance was generally low especially to ampicillin, quinolones, nitrofurantoin, glycopeptides and linezolid, but high for erythromycin and tetracycline. Equally, vanA and vanB gene frequencies were low (i.e. 15.8 and 7.9%, respectively) and detected only in E. casseliflavus/gallinarum species that are intrinsically resistant to vancomycin. Vancomycin resistant isolates of E. faecalis and E. faecium were not detected. CONCLUSIONS: Enterococcus species are frequent in clinical specimens at Mulago Hospital but they are highly susceptible to common antibiotics especially ampicillin. While vancomycin resistant enterococcal (VRE) isolates of E. faecium and E. faecalis are rare in the hospital and frequency of multidrug resistance is low, non-faecium and non-faecalis VRE isolates (i.e. E. gallinarum/casseliflavus) are frequent, some with VanA/VanB (high-level) vancomycin resistance. Therefore, species-level identification of enterococci is necessary in resource limited settings to guide infection control and treatment of enterococcal infections.

Antimicrobial Drug Resistance in Blood Culture Isolates at a Tertiary Hospital, Uganda.

We summarize antimicrobial drug resistance (AMR) patterns from blood cultures at a tertiary hospital in Uganda. High rates of resistance to first-line antibiotic drugs were observed among Staphylococcus aureus and gram-negative organisms. Microbiology services with susceptibility testing should be strengthened to support standardized reporting of AMR data in sub-Saharan Africa.

Nasopharyngeal carriage and antibiotic susceptibility patterns of streptococcus pneumoniae, haemophilus influenzae, moraxella catarrhalis and staphylococcus aureus among urban Ugandan children post-PCV10 introduction: a cross-sectional study.

In 2013, Uganda introduced the PCV10 pneumococcal vaccine and it is given to children at 6, 10 and 14 weeks after birth. Carriage prevalence studies post PCV10-introduction are necessary for monitoring the impact of vaccination and trends in antibiotic resistance. Here, we studied carriage/antibiotic resistance of Streptococcus pneumoniae (pneumococcus), Haemophilus influenzae, Moraxella catarrhalis, and Staphylococcus aureus isolated from 194 children at the Mulago Assessment Centre clinic in Kampala-Uganda, 5 years post-PCV10 introduction. Almost all the children were vaccinated with PCV10 (98.5%, 191/194). The overall carriage prevalence (any species) was 62% (120/194), and it was associated with a history of antibiotics use (p=0.0159) and having respiratory symptoms (p=0.0003). The pneumococcus, H. influenzae, M. catarrhalis, and S. aureus carriage prevalence was 46% (90/194), 21% (40/194), 7% (14/194), and 6% (12/194), respectively. Species co-carriage occurred in 32 children (17%, 32/194), predominantly multidrug resistant pneumococcus + H. influenzae (23 children). Furthermore, pneumococci were highly resistant to cotrimoxazole (100%), erythromycin (76%), and tetracycline (52%), 42% being multidrug-resistant. Overall, we note an increase in antibiotic resistance post-PCV10 introduction, and microbial shifts i.e., a decrease in pneumococcus, M. catarrhalis and S. aureus carriage and an increase in H. influenzae carriage suggesting vaccine-associated perturbation of the respiratory ecology.

Source-tracking ESBL-producing bacteria at the maternity ward of Mulago hospital, Uganda.

INTRODUCTION: Escherichia coli, Klebsiella pneumoniae and Enterobacter (EKE) are the leading cause of mortality and morbidity in neonates in Africa. The management of EKE infections remains challenging given the global emergence of carbapenem resistance in Gram-negative bacteria. This study aimed to investigate the source of EKE organisms for neonates in the maternity environment of a national referral hospital in Uganda, by examining the phenotypic and molecular characteristics of isolates from mothers, neonates, and maternity ward. METHODS: From August 2015 to August 2016, we conducted a cross-sectional study of pregnant women admitted for elective surgical delivery at Mulago hospital in Kampala, Uganda; we sampled (nose, armpit, groin) 137 pregnant women and their newborns (n = 137), as well as health workers (n = 67) and inanimate objects (n = 70 -beds, ventilator tubes, sinks, toilets, door-handles) in the maternity ward. Samples (swabs) were cultured for growth of EKE bacteria and isolates phenotypically/molecularly investigated for antibiotic sensitivity, as well as ß-lactamase and carbapenemase activity. To infer relationships among the EKE isolates, spatial cluster analysis of phenotypic and genotypic susceptibility characteristics was done using the Ridom server. RESULTS: Gram-negative bacteria were isolated from 21 mothers (15%), 15 neonates (11%), 2 health workers (3%), and 13 inanimate objects (19%); a total of 131 Gram-negative isolates were identified of which 104 were EKE bacteria i.e., 23 (22%) E. coli, 50 (48%) K. pneumoniae, and 31 (30%) Enterobacter. Carbapenems were the most effective antibiotics as 89% (93/104) of the isolates were susceptible to meropenem; however, multidrug resistance was prevalent i.e., 61% (63/104). Furthermore, carbapenemase production and carbapenemase gene prevalence were low; 10% (10/104) and 6% (6/104), respectively. Extended spectrum ß-lactamase (ESBL) production occurred in 37 (36%) isolates though 61 (59%) carried ESBL-encoding genes, mainly blaCTX-M (93%, 57/61) implying that blaCTX-M is the ideal gene for tracking ESBL-mediated resistance at Mulago. Additionally, spatial cluster analysis revealed isolates from mothers, new-borns, health workers, and environment with similar phenotypic/genotypic characteristics, suggesting transmission of multidrug-resistant EKE to new-borns. CONCLUSION: Our study shows evidence of transmission of drug resistant EKE bacteria in the maternity ward of Mulago hospital, and the dynamics in the ward are more likely to be responsible for transmission but not individual mother characteristics. The high prevalence of drug resistance genes highlights the need for more effective infection prevention/control measures and antimicrobial stewardship programs to reduce spread of drug-resistant bacteria in the hospital, and improve patient outcomes.

Diarrhoeagenic Escherichia coli isolated from children with acute diarrhoea at Rakai hospital, Southern Uganda.

Background: Diarrhoeagenic Escherichia coli (DEC) is a leading cause of childhood diarrhoea. This study estimated the prevalence of DEC and DEC pathotypes among children with acute diarrhoea in Southern Uganda. Methods: A cross-sectional study was conducted on 267 children less than 5 years with acute diarrhoea, admitted to Rakai General Hospital in Southern Uganda. Faecal samples were collected from the children and processed for isolation of E. coli. The presence of DEC and the distribution of DEC pathotypes were determined by polymerase chain reaction. Results: A total of 102 (38.2%, 102/267) children had DEC of various pathotypes - enteroaggregative E. coli (EAEC) (14.2%); enteropathogenic E. coli (EPEC) (6.7%); enterotoxigenic E. coli (ETEC) (6%); enteroinvasive E. coli (EIEC) (7.5%); enterohemorrhagic E. coli (EHEC) (3%); and cell-detaching E. coli (CDEC) (0.75%). The difference in the overall prevalence of DEC was not significant regarding HIV but individually, EAEC and CDEC were associated with HIV-positive status while ETEC was associated with HIV-negative status. Conclusions: DEC is prevalent in children with acute diarrhoea in Southern Uganda and its identification in children should be considered among strategies for combatting childhood diarrhoea in Africa.

Species and drug susceptibility profiles of staphylococci isolated from healthy children in Eastern Uganda.

Staphylococci are a key component of the human microbiota, and they mainly colonize the skin and anterior nares. However, they can cause infection in hospitalized patients and healthy individuals in the community. Although majority of the Staphylococcus aureus strains are coagulase-positive, some do not produce coagulase, and the isolation of coagulase-positive non-S. aureus isolates in humans is increasingly being reported. Therefore, sound knowledge of the species and characteristics of staphylococci in a given setting is important, especially isolates from children and immunocompromised individuals. The spectrum of Staphylococcus species colonizing children in Uganda is poorly understood; here, we aimed to determine the species and characteristics of staphylococci isolated from children in Eastern Uganda. Seven hundred and sixty four healthy children less than 5 years residing in Iganga and Mayuge districts in Eastern Uganda were enrolled. A total of 513 staphylococci belonging to 13 species were isolated from 485 children (63.5%, 485/764), with S. aureus being the dominant species (37.6%, 193/513) followed by S. epidermidis (25.5%, 131/513), S. haemolyticus (2.3%, 12/513), S. hominis (0.8%, 4/513) and S. haemolyticus/lugdunensis (0.58%, 3/513). Twenty four (4.95%, 24/485) children were co-colonized by two or more Staphylococcus species. With the exception of penicillin, antimicrobial resistance (AMR) rates were low; all isolates were susceptible to vancomycin, teicoplanin, linezolid and daptomycin. The prevalence of methicillin resistance was 23.8% (122/513) and it was highest in S. haemolyticus (66.7%, 8/12) followed by S. aureus (28.5%, 55/193) and S. epidermidis (23.7%, 31/131). The prevalence of multidrug resistance was 20.3% (104/513), and 59% (72/122) of methicillin resistant staphylococci were multidrug resistant. Four methicillin susceptible S. aureus isolates and a methicillin resistant S. scuiri isolate were mupirocin resistant (high-level). The most frequent AMR genes were mecA, vanA, ant(4')-Ia, and aac(6')-Ie- aph(2'')-Ia, pointing to presence of AMR drivers in the community.

Prevalence of plasmid-mediated AmpC beta-lactamases in Enterobacteria isolated from urban and rural folks in Uganda.

Background: AmpC beta-lactamase-producing bacteria are associated with increased resistance to third-generation cephalosporins. Here, we describe plasmid-mediated AmpC beta-lactamase-producing enterobacteria isolated from urban and rural dwellers in Uganda. Methods: Stool and urine from 1,448 individuals attending outpatient clinics in Kampala and two rural districts in central Uganda were processed for isolation of Escherichia coli and Klebsiella. Following antibiotic susceptibility testing, cefoxitin resistant isolates, and amoxicillin/clavulanate resistant but cefoxitin susceptible isolates, were tested for AmpC beta-lactamase production using the cefoxitin-cloxacillin double-disc synergy test. Carriage of plasmid-mediated AmpC beta-lactamase-encoding genes (pAmpC) and extended spectrum beta-lactamase (ESBL) encoding genes was determined by PCR. Results: Nine hundred and thirty E. coli and 55 Klebsiella were recovered from the cultured samples, yielding 985 isolates investigated (one per participant). One hundred and twenty-nine isolates (13.1%, 129/985) were AmpC beta-lactamase producers, of which 111 were molecularly characterized for pAmpC and ESBL gene carriage. pAmpC genes were detected in 60% (67/111) of the AmpC beta-lactamase producers; pAmpC genes were also detected in 18 AmpC beta-lactamase non-producers and in 13 isolates with reduced susceptibility to third-generation cephalosporins, yielding a total of 98 isolates that carried pAmpC genes. Overall, the prevalence of pAmpC genes in cefoxitin resistant and/or amoxicillin/clavulanate resistant E. coli and Klebsiella was 59% (93/157) and 26.1% (5/23), respectively. The overall prevalence of pAmpC-positive enterobacteria was 10% (98/985); 16.4% (45/274) in Kampala, 6.2% (25/406) Kayunga, and 9.2% (28/305) Mpigi. Ciprofloxacin use was associated with carriage of pAmpC-positive bacteria while residing in a rural district was associated with protection from carriage of pAmpC-positive bacteria. Conclusion: pAmpC beta-lactamase producing enterobacteria are prevalent in urban and rural dwellers in Uganda; therefore, cefoxitn should be considered during routine susceptibility testing in this setting.

Bacteremia in febrile cancer patients in Uganda.

OBJECTIVE: The aim of this study was to determine the predominant bacterial species causing bacteremia among febrile cancer patients, and their antibacterial resistance profiles at the Uganda Cancer Institute. RESULTS: We enrolled in-patients with a documented fever (≥ 37.5 °C). Bacteria from positive blood cultures were identified using standard methods biochemically. Antibacterial susceptibility testing was performed with the Kirby-Bauer disc diffusion method. From a total of 170 febrile episodes, positive blood cultures were obtained from 24 (14.1%). A positive culture was more likely to be obtained from a patient with neutropenia (P = 0.017). Of 22 (66.7%) Gram-negative bacteria isolated, half were E. coli (n = 11). Gram-negative compared to Gram-positive bacteria were most likely to be isolated from patients with a hematologic malignancy (P = 0.02) or patients with neutropenia (P = 0.006). Of the isolated Enterobacteriaceae 85% (n = 20) were resistant to three or more classes of antibiotic and 41% (n = 7) had extended spectrum beta-lactamases. Of the 11 Gram-positive bacteria isolated, the S. aureus isolate was methicillin resistant but susceptible to vancomycin. Multidrug resistant Gram-negative bacteria are the main cause of bacteremia in febrile cancer patients at the Uganda Cancer Institute. There is need for ongoing microbial surveillance, infection prevention and control, and antibiotic stewardship programs.

CA-MRSA and HA-MRSA coexist in community and hospital settings in Uganda.

Background: Methicillin resistant Staphylococcus aureus (MRSA) strains were once confined to hospitals however, in the last 20 years MRSA infections have emerged in the community in people with no prior exposure to hospitals. Strains causing such infections were novel and referred to as community-associated MRSA (CA-MRSA). The aim of this study was to determine the MRSA carriage rate in children in eastern Uganda, and to investigate coexistence between CA-MRSA and hospital-associated (HA-MRSA). Methods: Between February and October 2011, nasopharyngeal samples (one per child) from 742 healthy children under 5 years in rural eastern Uganda were processed for isolation of MRSA, which was identified based on inhibition zone diameter of ≤19 mm on 30 µg cefoxitin disk. SCCmec and spa typing were performed for MRSA isolates. Results: A total of 140 S. aureus isolates (18.9%, 140/742) were recovered from the children of which 5.7% (42/742) were MRSA. Almost all (95.2%, 40/42) MRSA isolates were multidrug resistant (MDR). The most prevalent SCCmec elements were types IV (40.5%, 17/42) and I (38.1%, 16/42). The overall frequency of SCCmec types IV and V combined, hence CA-MRSA, was 50% (21/42). Likewise, the overall frequency of SCCmec types I, II and III combined, hence HA-MRSA, was 50% (21/42). Spa types t002, t037, t064, t4353 and t12939 were detected and the most frequent were t064 (19%, 8/42) and t037 (12%, 5/42). Conclusion: The MRSA carriage rate in children in eastern Uganda is high (5.7%) and comparable to estimates for Mulago Hospital in Kampala city. Importantly, HA-MRSA (mainly of spa type t037) and CA-MRSA (mainly of spa type t064) coexist in children in the community in eastern Uganda, and due to high proportion of MDR detected, outpatient treatment of MRSA infection in eastern Uganda might be difficult.

Hand Colonization with Gram-Negative Organisms of Healthcare Workers Accessing the Cardiac Intensive Care Unit: A Cross-Sectional Study at the Uganda Heart Institute.

BACKGROUND: Hands of healthcare workers (HCWs) are vehicles for pathogens responsible for healthcare-associated infections (HAIs). Following the identification of Gram-negative organisms (GNOs) in all cases of HAIs in the cardiac intensive care unit (ICU), we sought to determine the burden of hand colonization with GNOs among healthcare workers who access the cardiac ICU. METHODS: We retrospectively reviewed results from surveillance cultures of fingertip imprints of HCWs who access the cardiac ICU at the Uganda Heart Institute. We collected data on staff category, isolates, and susceptibility to antibiotics. We analyzed the data using Microsoft Excel, and the results are summarized in proportions and percentages and presented in charts and tables. RESULTS: Fifty-six healthcare workers participated in the surveillance. 21 were ICU clinicians, 21 non-ICU clinicians, and 14 nonclinicians. GNOs were cultured in 19 (33.9%) HCWs, in which 8/19 (42.1%) were non-ICU clinicians, 6/19 (31.2%) ICU clinicians, and 5/19 (26.3%) nonclinicians. 32 isolates were identified, of which 47%, 28%, and 25% were cultured from non-ICU clinicians, nonclinicians, and ICU clinicians, respectively. Predominant isolates were Acinetobacter (34%), Citrobacter (21.9%), and Pseudomonas (21.9%). Antimicrobial resistance ranged from 4% to 90%. 9/28 (32.1%) isolates, predominantly Acinetobacter species (spp), were carbapenem resistant. 8/28 (28.6%) isolates, predominantly Citrobacter spp, were multidrug resistant. Resistance to ciprofloxacin and cefepime was low at 3.6% and 4.4%, respectively. CONCLUSION: Gram-negative organisms, predominantly Acinetobacter, Citrobacter, and Pseudomonas spp, were prevalent on the hands of HCWs who access the cardiac ICU irrespective of the staff category. Antimicrobial resistance was high, with multidrug resistance and carbapenem resistance common among Citrobacter spp and Acinetobacter spp, respectively. Resistance to cefepime and ciprofloxacin was low.

Genotypic diversity among multidrug resistant Pseudomonas aeruginosa and Acinetobacter species at Mulago Hospital in Kampala, Uganda.

BACKGROUND: Multidrug resistant Pseudomonas aeruginosa and Acinetobacter species are common causes of nosocomial infections worldwide. Recently we reported the occurrence of carbapenem resistant Enterobacteriaceae, P. aeruginosa and Acinetobacter species at Mulago National Referral Hospital in Kampala, Uganda, but the isolates were not analyzed for genetic relatedness. Herein we report the intra-species genotypic diversity among P. aeruginosa and Acinetobacter baumannii isolated from hospitalized patients and the environment at Mulago Hospital, using repetitive elements-based PCR (Rep-PCR) genotyping. RESULTS: A total of 736 specimens from hospitalized patients were processed for culture and sensitivity testing yielding 9 (1.2%) P. aeruginosa and 7 (0.95%) A. baumannii. Similarly, 100 samples from the hospital environment were processed yielding 33 (33%) P. aeruginosa and 13 (13%) A. baumannii. Altogether, 62 non-repetitive isolates were studied (42 P. aeruginosa and 20 A. baumannii), of which 38% (16/42) P. aeruginosa and 40% (8/20) A. baumannii were multidrug resistant (isolates resistant to three or more classes of antimicrobials). Carbapenem resistance prevalence was 33 and 21% for P. aeruginosa from patients and the environment, respectively, while it was 14 and 86% for A. baumannii from patients and environment, respectively. Cluster analysis of the Rep-PCR fingerprints revealed a high level of genetic diversity among the isolates within each species as few isolates were clustered (at 100% level of similarity). More to this, the clustered isolates revealed a complex nature of multidrug resistant P. aeruginosa and A. baumannii clones circulating at Mulago Hospital. Importantly, certain isolates from the environment and patients were clustered, implying that hospitalized patients at Mulago were probably infected with strains from the environment. CONCLUSIONS: The prevalence of multidrug resistant P. aeruginosa and A. baumannii is high at Mulago Hospital but carbapenem resistance prevalence remains relatively low in isolates from hospitalized patients. Importantly, the prevalence of carbapenem resistance in isolates from the environment is high implying the infection control practices at the hospital might be inadequate.

Carbapenem resistant Pseudomonas aeruginosa and Acinetobacter baumannii at Mulago Hospital in Kampala, Uganda (2007-2009).

BACKGROUND: Multidrug resistant Pseudomonas aeruginosa and Acinetobacter baumannii are common causes of health care associated infections worldwide. Carbapenems are effective against infections caused by multidrug resistant Gram-negative bacteria including Pseudomonas and Acinetobacter species. However, their use is threatened by the emergence of carbapenemase-producing strains. The aim of this study was to determine the prevalence of carbapenem-resistant P. aeruginosa and A. baumannii at Mulago Hospital in Kampala Uganda, and to establish whether the hospital environment harbors carbapenem-resistant Gram-negative rods. RESULTS: Between February 2007 and September 2009, a total of 869 clinical specimens were processed for culture and sensitivity testing yielding 42 (5 %) P. aeruginosa and 29 (3 %) A. baumannii isolates, of which 24 % (10/42) P. aeruginosa and 31 % (9/29) A. baumannii were carbapenem-resistant. Additionally, 80 samples from the hospital environment were randomly collected and similarly processed yielding 58 % (46/80) P. aeruginosa and 14 % (11/80) A. baumannii, of which 33 % (15/46) P. aeruginosa and 55 % (6/11) A. baumannii were carbapenem-resistant. The total number of isolates studied was 128. Carbapenemase genes detected were bla IMP-like (36 %, 9/25), bla VIM-like (32 %, 8/25), bla SPM-like (16 %, 4/25); bla NDM-1-like (4 %, 1/25) in carbapenem-resistant P. aeruginosa, and bla OXA-23-like (60 %, 9/15), bla OXA-24-like (7 %, 1/15), bla OXA-58-like (13 %, 2/15), and bla VIM-like (13 %, 2/15) in carbapenem-resistant A. baumannii. Furthermore, class 1 integrons were detected in 38 % (48/128) of P. aeruginosa and Acinetobacter, 37 % (26/71) of which were in clinical isolates and 39 % (22/57) in environment isolates. Gene cassettes were found in 25 % (12/48) of integron-positive isolates. These were aminoglycoside adenylyltransferase ant(4')-IIb (3 isolates); trimethoprim-resistant dihydrofolate reductase dfrA (2 isolates); adenyltransferase aadAB (3 isolates); QacE delta1 multidrug exporter (2 isolates); quinolone resistance pentapeptide repeat protein qnr (1 isolate); and metallo-ß-lactamase genes bla VIM-4-like, bla IMP-19-like, and bla IMP-26-like (1 isolate each). Gene cassettes were missing in 75 % (36/48) of the integron-positive isolates. CONCLUSIONS: The prevalence of carbapenem-resistant P. aeruginosa and Acinetobacter among hospitalized patients at Mulago Hospital is low compared to rates from South-East Asia. However, it is high among isolates and in the environment, which is of concern given that the hospital environment is a potential source of infection for hospitalized patients and health care workers.

Antimicrobial susceptibility profiles of Escherichia coli and Klebsiella pneumoniae isolated from outpatients in urban and rural districts of Uganda.

BACKGROUND: Antimicrobial resistance is a global public health concern contributing to increased morbidity and mortality particularly in low-income countries. Studies on commensal bacteria are important as they reflect the state of antimicrobial susceptibility patterns in populations. However, susceptibility data on potentially pathogenic commensal bacteria from individuals in communities are still limited. The aim of this cross-sectional study was to determine the susceptibility profiles of Escherichia coli and Klebsiella species isolated from clients attending outpatient clinics in Kampala (urban district) and two rural districts of Uganda, Kayunga and Mpigi. Factors associated with such carriage are also reported. RESULTS: A total of 1448 participants were recruited into the study with 985 yielding organisms of interest from stool or urine samples (one per client). Most growth occurred from stool samples (636/985, 87%), of which 620/636 (97%) grew E. coli while 16 (3%) were Klebsiella pneumoniae. Growth from urine was 349/985 (35%) of which 310/349 (89%) were E. coli while 39 (11%) K. pneumoniae. High rates of antimicrobial resistance were detected among E. coli and Klebsiella isolates combined: sulphamethoxazole/trimethoprim 70%, amoxicillin/clavulanate 36%, chloramphenicol 20%, ciprofloxacin 11%, gentamicin 11%, nitrofurantoin 4%, ceftriaxone 3%, piperacillin/tazobactam 27%, cefoxitin 22%, and cefepime 15%. Multidrug resistance was noted in 33% of the isolates. None of the isolates were resistant to imipenem. Overall, isolates from Kampala were more resistant to antimicrobials. Across the three districts combined, isolates producing beta-lactamase enzymes extended spectrum ß-lactamase-(ESBL) and AmpC comprised 5.3 and 13.2%, respectively. Further, medical procedures involving inoculation were independent risk factors [aOR 50.76 (1.80, 1432.90)] while residing in a rural district and use of sulphamethoxazole/trimethoprim 3 months prior to visiting the outpatient clinics were protective against carriage of multidrug resistant isolates. Furthermore, use of gentamicin was protective against AmpC producing isolates while clients attending HIV/AIDs clinics were less likely to carry such isolates. No factor was independently associated with carriage of ESBL-producing isolates. CONCLUSION: Antimicrobial resistance is prevalent among E. coli and K. pneumoniae carried in the gut of clients attending outpatient clinics in Kampala and two rural districts in Uganda. This could complicate treatment options for community-acquired infections caused by the Enterobacteriaceae.

Prevalence and Characterization of Carbapenem-Resistant Enterobacteriaceae Isolated from Mulago National Referral Hospital, Uganda.

INTRODUCTION: Carbapenemases have increasingly been reported in enterobacteriaceae worldwide. Most carbapenemases are plasmid encoded hence resistance can easily spread. Carbapenem-resistant enterobacteriaceae are reported to cause mortality in up to 50% of patients who acquire bloodstream infections. We set out to determine the burden of carbapenem resistance as well as establish genes encoding for carbapenemases in enterobacteriaceae clinical isolates obtained from Mulago National Referral Hospital, Uganda. METHODS: This was a cross-sectional study with a total of 196 clinical isolates previously collected from pus swabs, urine, blood, sputum, tracheal aspirates, cervical swabs, endomentrial aspirates, rectal swabs, Vaginal swabs, ear swabs, products of conception, wound biopsy and amniotic fluid. All isolates were subjected to phenotypic carbapenemase screening using Boronic acid-based inhibition, Modified Hodge and EDTA double combined disk test. In addition, all the isolates were subjected to PCR assay to confirm presence of carbapenemase encoding genes. RESULTS: The study found carbapenemase prevalence of 22.4% (44/196) in the isolates using phenotypic tests, with the genotypic prevalence slightly higher at 28.6% (56/196). Over all, the most prevalent gene was blaVIM (21,10.7%), followed by blaOXA-48 (19, 9.7%), blaIMP (12, 6.1%), blaKPC (10, 5.1%) and blaNDM-1 (5, 2.6%). Among 56 isolates positive for 67 carbapenemase encoding genes, Klebsiella pneumonia was the species with the highest number (52.2%). Most 32/67(47.7%) of these resistance genes were in bacteria isolated from pus swabs. CONCLUSION: There is a high prevalence of carbapenemases and carbapenem-resistance encoding genes among third generation cephalosporins resistant Enterobacteriaceae in Uganda, indicating a danger of limited treatment options in this setting in the near future.

Prevalence and antimicrobial susceptibility patterns of bacteria from milkmen and cows with clinical mastitis in and around Kampala, Uganda.

BACKGROUND: Identification of pathogens associated with bovine mastitis is helpful in treatment and management decisions. However, such data from sub-Saharan Africa is scarce. Here we describe the distribution and antimicrobial susceptibility patterns of bacteria from cows with clinical mastitis in Kampala, Uganda. Due to high concern of zoonotic infections, isolates from milkmen are also described. METHODOLOGY/PRINCIPAL FINDINGS: Ninety seven milk samples from cows with clinical mastitis and 31 nasal swabs from milkmen were collected (one sample per cow/human). Fifty eight (60%) Gram-positive isolates namely Staphylococci (21), Enterococci (16), Streptococci (13), Lactococci (5), Micrococci (2) and Arcanobacteria (1) were detected in cows; only one grew Staphylococcus aureus. Furthermore, 24 (25%) coliforms namely Escherichia coli (12), Klebsiella oxytoca (5), Proteus vulgaris (2), Serratia (2), Citrobacter (1), Cedecea (1) and Leclercia (1) were identified. From humans, 24 Gram-positive bacteria grew, of which 11 were Staphylococci (35%) including four Staphylococcus aureus. Upon susceptibility testing, methicillin-resistant coagulase-negative staphylococci (CoNS) were prevalent; 57%, 12/21 in cows and 64%, 7/11 in humans. However, methicillin-resistant Staphylococcus aureus was not detected. Furthermore, methicillin and vancomycin resistant CoNS were detected in cows (Staphylococcus hominis, Staphylococcus lugdunensis) and humans (Staphylococcus scuiri). Also, vancomycin and daptomycin resistant Enterococci (Enterococcus faecalis and Enterococcus faecium, respectively) were detected in cows. Coliforms were less resistant with three pan-susceptible isolates. However, multidrug resistant Klebsiella, Proteus, Serratia, Cedecea, and Citrobacter were detected. Lastly, similar species grew from human and bovine samples but on genotyping, the isolates were found to be different. Interestingly, human and bovine Staphylococcus aureus were genetically similar (spa-CC435, spa-type t645 corresponding to ST121) but with different susceptibility patterns. CONCLUSIONS/SIGNIFICANCE: CoNS, Enterococci, Streptococci, and Escherichia coli are the predominant pathogens associated with clinical bovine-mastitis in Kampala, Uganda. Multidrug resistant bacteria are also prevalent. While similar species occurred in humans and cows, transmission was not detected.

Molecular characterization of Staphylococcus aureus from patients with surgical site infections at Mulago Hospital in Kampala, Uganda.

BACKGROUND: The prevalence of Methicillin resistant Staphylococcus aureus (MRSA) is progressively increasing globally with significant regional variation. Understanding the Staphylococcus aureus lineages is crucial in controlling nosocomial infections. Recent studies on S. aureus in Uganda have revealed an escalating burden of MRSA. However, the S. aureus genotypes circulating among patients are not known. Here, we report S. aureus lineages circulating in patients with surgical site infections (SSI) at Mulago National hospital, Kampala, Uganda. METHODS: A cross-sectional study involving 314 patients with SSI at Mulago National Hospital was conducted from September 2011 to April 2012. Pus swabs from the patients' SSI were processed using standard microbiological procedures. Methicillin sensitive Staphylococcus aureus (MSSA) and MRSA were identified using phenotypic tests and confirmed by PCR-detection of the nuc and mecA genes, respectively. SCCmec genotypes were determined among MRSA isolates using multiplex PCR. Furthermore, to determine lineages, spa sequence based-genotyping was performed on all S. aureus isolates. RESULTS: Of the 314 patients with SSI, S. aureus accounted for 20.4% (64/314), of which 37.5% (24/64) were MRSA. The predominant SCCmec types were type V (33.3%, 8/24) and type I (16.7%, 4/24). The predominant spa lineages were t645 (17.2%, 11/64) and t4353 (15.6%, 10/64), and these were found to be clonally circulating in all the surgical wards. On the other hand, lineages t064, t355, and t4609 were confined to the obstetrics and gynecology wards. A new spa type (t10277) was identified from MSSA isolate. On multivariate logistic regression analysis, cancer and inducible clindamycin resistance remained as independent predictors of MRSA-SSI. CONCLUSION: SCCmec types I and V are the most prevalent MRSA mecA types from the patients' SSI. The predominant spa lineages (t645 and t4353) are clonally circulating in all the surgical wards, calling for strengthening of infection control practices at Mulago National Hospital.

Antimicrobial resistance in hospitalized surgical patients: a silently emerging public health concern in Uganda.

BACKGROUND: Surgical site infections (SSIs) are difficult to treat and are associated with substantially longer hospital stay, higher treatment cost, morbidity and mortality, particularly when the etiological agent is multidrug-resistant (MDR). To address the limited data in Uganda on SSIs, we present the spectrum of bacteria isolated from hospitalized patients, the magnitude and impact of MDR bacterial isolates among patients with SSIs. METHODS: A descriptive cross sectional study was conducted from September 2011 through April 2012 involving 314 patients with SSIs in the obstetrics & gynecology, general surgery and orthopedic wards at Mulago National Hospital in Kampala, Uganda. Wound swabs were taken and processed using standard microbiological methods. Clinico-demographic characteristics of patients were obtained using structured questionnaires and patients' files. RESULTS: Of the 314 enrolled patients with SSIs (mean age 29.7 ±13.14 years), 239 (76.1%) were female. More than half of the patients were from obstetrics and gynecology (62.1%, 195/314). Of 314 wound swabs taken, 68.8% (216/314) were culture positive aerobically, yielding 304 bacterial isolates; of which 23.7% (72/304) were Escherichia coli and 21.1% (64/304) were Staphylococcus aureus. More than three quarters of Enterobacteriaceae were found to be extended spectrum beta lactamase (ESBL) producers and 37.5% of S. aureus were Methicillin resistant S. aureus (MRSA). MDR occurred in 78.3% (238/304) of the isolates; these were more among Gram-negative bacteria (78.6%, 187/238) compared to Gram-positive bacteria (21.4%, 51/238), (p-value < 0.0001, χ2 = 49.219). Amikacin and imepenem for ESBL-producing Enterobacteriacea and vancomycin for MRSA showed excellent performance except that they remain expensive drugs in Uganda. CONCLUSION: Most SSIs at Mulago National Hospital are due to MDR bacteria. Isolation of MRSA and ESBL-producing Enterobacteriaceae in higher proportions than previously reported calls for laboratory guided SSIs- therapy and strengthening of infection control surveillance in this setting.