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Aim: This study aimed to characterize ciprofloxacin-resistance genes in clinical Escherichia coli isolates obtained from a six-month antimicrobial resistance (AMR) surveillance from Ghana. Methods: Eighty-three of 440 archived E. coli isolates were confirmed by biochemical reactions and resistance profiles by the disc diffusion method. These isolates were cultured from urine (42), stool (23), vaginal swabs (12), wounds (5) and heart valve (1) during AMR surveillance. Minimum Inhibition Concentration (MIC) by E-test method was performed on all E. coli isolates that were resistant to ciprofloxacin by the disc diffusion method. Additionally, all isolates with reduced MIC to ciprofloxacin (>32 µg/ml) were selected for molecular assays. Three chromosomal and nine plasmid-mediated resistance genes were screened in all Ciprofloxacin resistant E. coli (CRE) by polymerase chain reaction (PCR). Randomly selected amplified genes were commercially sequenced and analyzed. Results: In total, 47/83 (56.6%) E. coli isolates were resistant to ciprofloxacin and 29 (61.7%) had MIC values greater than 32 µg/ml. Chromosomal mediated genes (gyrA, gyrB and parC) were present in all 29 CRE isolates (100%). Distribution of the plasmid-mediated genes were as follows; qnrA 16/29 (55.1%), qnrB 16/29(55.1%), qnrC 22/29(75.8%), qnrS 26/29(89.6%), qepA 5/29(17.2%) and oqxB 19/29(65.5%). Genes encoding for altered aminoglycoside acetyltransferase [aac(6’)1bcr] were also present in all 29 CRE isolates. The majority (72.4%) of the CRE isolates had gyrA mutations at codons 83 and 87. In parC, the mutations were at codons 71 and 80. Five isolates had mutations at codon 56 and four each had mutations at positions 79 and 80. Conclusion: In this study, fluoroquinolone resistance genes were identified in all CRE isolates, mostly with putative mutations in the Quinolone Resistance Determining Region (QRDR). These chromosomal and plasmid-mediated genes may be widespread in Ghana and associated with CRE from the AMR surveillance. Although new mutations points were identified in parC, they may not be linked to the CRE.

Background Carriage of multidrug resistant (MDR) Gram-negative bacteria (GN) in hospitalized neonates may increase the risk of difficult-to-treat invasive infections at neonatal intensive care units (NICUs). Data on MDRGN carriage among hospitalized newborns in Africa are limited. Methods We conducted a cross-sectional study at the NICUs of 2 tertiary hospitals in Ghana. Swabs from the axilla, groin, perianal region, and the environment were cultured, GN were identified, and antibiotic susceptibility was tested. We obtained blood culture isolates from neonates with sepsis. Whole-genome sequencing was used to characterize carbapenemase-producing Klebsiella pneumoniae. Typing was done by multilocus sequence typing (MLST) and single nucleotide polymorphism (SNP) analysis. Results A total of 276 GN were isolated from 228 screened neonates. Pathogenic GN were cultured in 76.8% (175 of 228) of neonates. Klebsiella spp (41.7%; 115 of 276) and Escherichia coli (26.4%; 73 of 276) were the commonest organisms. Carriage rates of MDRGN and third-generation cephalosporin resistant organisms were 49.6% (113 of 228) and 46.1% (105 of 228), respectively. Among Klebsiella spp, 75.6% (87 of 115) phenotypically expressed extended-spectrum β-lactamase activity, whereas 15.6% expressed carbapenemase and harbored bla-OXA-181 and bla-CTX-M-15. Overall, 7.0% (16 of 228) of neonates developed GN bloodstream infection. In 2 of 11 neonates, sequencing showed the same identity between carriage and the bloodstream isolate. Length of stay before specimen collection and antibiotic use were independently associated with carriage rates, which increased from 13% at admission to 42% by day 2 and reached a plateau at 91% by day 15. Conclusions High carriage rates of MDRGN, including carbapenemase-producing enterobacterales may be an emerging problem in NICUs in Africa., We show carriage rate of MDR enterobacterales among neonates in 2 NICUs in Ghana. MDR carriage is associated with duration of stay and antibiotic use. We also report OXA-181 carbapenemase-producing organism from Ghana. WGS showed identity between carriage and BSI.

We sequenced 29 carbapenemase-producing Klebsiella pneumoniae isolates from a neonatal intensive care unit in Ghana. Twenty-eight isolates were sequence type 17 with blaOXA-181 and differed by 0-32 single-nucleotide polymorphisms. Improved surveillance and infection control are needed to characterize and curb the spread of multidrug-resistant organisms in sub-Saharan Africa.

SUMMARYPoultry are possible sources of non-typhoidalSalmonellaserovars which may cause foodborne human disease. We conducted a cross-sectional study to determine the prevalence ofSalmonellaserovars in egg-laying hens and broilers at the farm level and their susceptibility to antimicrobials commonly used in the poultry industry in Ghana. Sampling of faeces by a sock method (n= 75), dust (n= 75), feed (n= 10) and drinking water (n= 10) was performed at 75 commercial egg-laying and broiler farms in two regions of Ghana and skin neck (n= 30) at a local slaughterhouse from broilers representing different flocks.Salmonellawas detected in 94/200 (47%) samples with an overall flock prevalence of 44·0%. Sixteen different serovars were identified withS. Kentucky (18·1%),S. Nima (12·8%),S. Muenster (10·6%),S. Enteritidis (10·6%) andS. Virchow (9·6 %) the most prevalent types. The predominant phage type ofS. Enteritidis was PT1. All strains were susceptible to cefotaxime, ceftazidime and cefoxitin. Fifty-seven (60·6%) strains were resistant to one or more of the remaining nine antimicrobials tested by disk diffusion, of which 23 (40·4%) showed multi-resistance (resistance to ⩾3 classes of antimicrobials). Of the resistant strains (n= 57), the most significant were to nalidixic acid (89·5%), tetracycline (80·7%), ciprofloxacin (64·9%), sulfamethazole (42·1%), trimethoprim (29·8%) and ampicillin (26·3%). AllS.Kentucky strains were resistant to more than two antimicrobials and shared common resistance to nalidixic acid or ciprofloxacin and tetracycline, often in combinations with other antimicrobials. PFGE analysis usingXbaI ofS. Kentucky demonstrated one dominant clone in the country. In conclusion, poultry produced in Ghana has a high prevalence of multi-resistantSalmonellaand the common finding of clonalS.Kentucky in the Kumasi area warrants further investigations into the epidemiology of this serovar. There is an urgent need for surveillance and control programmes onSalmonellaand use of antimicrobials in the Ghanaian poultry industry to protect the health of consumers.

Neisseria gonorrhoeae antimicrobial resistance (AMR) surveillance is essential for tracking the emergence and spread of AMR strains in local, national and international populations. This is crucial for developing or refining treatment guidelines. N. gonorrhoeae multiantigen sequence typing (NG-MAST) is beneficial for describing the molecular epidemiology of gonococci at national and international levels. Elucidation of AMR determinants to β-lactam drugs, is a means of monitoring the development of resistance. In Ghana, little is known about the current gonococcal AMR prevalence and no characterization of gonococcal isolates has been previously performed. In this study, gonococcal isolates (n = 44) collected from five health facilities in Ghana from 2012 to 2015, were examined using AMR testing, NG-MAST and sequencing of penA. High rates of resistance were identified to tetracycline (100%), benzylpenicillin (90.9%), and ciprofloxacin (81.8%). One isolate had a high cefixime MIC (0.75 μg/ml). Twenty-eight NG-MAST sequence types (STs) were identified, seventeen of which were novel. The isolate with the high cefixime MIC contained a mosaic penA-34 allele and belonged to NG-MAST ST1407, an internationally spreading multidrug-resistant clone that has accounted for most cefixime resistance in many countries. In conclusion, AMR testing, NG-MAST, and sequencing of the AMR determinant penA, revealed high rates of resistance to tetracycline, benzylpenicillin, and ciprofloxacin; as well as a highly diverse population of N. gonorrhoeae in Ghana. It is imperative to continue with enhanced AMR surveillance and to understand the molecular epidemiology of gonococcal strains circulating in Ghana and other African countries.

BackgroundBuruli ulcer (BU) disease, a skin condition caused by Mycobacterium ulcerans (M. ulcerans) is endemic in remote rural areas. Disease diagnosis on clinical basis alone can be misleading, requiring definitive diagnosis based on laboratory tests. Resource constraints in BU endemic areas make microscopy for the detection of acid fast bacilli (AFB) an important and useful method. It is rapid, user-friendly, convenient and cheap. Despite its usefulness, its performance is relatively low. This study investigated modifications of the current method aimed at improving its performance. Forty (IS) 2404 polymerase chain reactions (PCR) positive BU samples were processed by eight physical (centrifugation and overnight sedimentation) and chemical (phenol ammonium sulphate and sodium hypochlorite) modifications of the current direct method. Assessments were based on standard AFB evaluation coupled with in house criteria; positivity (P), clarity and contrast (C) release of bacilli from specimen (R). Overall AFB positivity rate was 64% (409/640). Each protocol had 80 smears. The percentage positivity (P) for the conventional method was 58% (46/80) smears. The highest positivity rate of 57/80 (%) was by protocol 7 (5% phenol in 4% ammonium sulphate (PhAS) and concentrated by overnight gravitational sedimentation). The least positivity rate at 35% (28/80) was by protocol 1 (smears from direct application of swab tips). The differences in performance between the two chemical tested; 5% phenol in 4% ammonium sulphate (PhAS) and 3.5% NaHOCl was significant (p0.05). This study concluded that BU samples treated with a solution of 5% phenol in 4% ammonium sulphate and concentrated by either centrifugation or overnight sedimentation is useful for maximizing AFB detection by bright field microscopy. This can be useful in rural health facilities with resource constraints.

Streptococcus pneumoniae is a major cause of morbidity and mortality worldwide. Pneumococcal strains are classified according to their capsular polysaccharide and more than 90 different serotypes are currently known. In this project, three distinct groups of pneumococcal carriage isolates from Ghana were investigated; isolates from healthy children in Tamale and isolates from both healthy and children attending the outpatient department at a hospital in Accra. The isolates were previously identified and characterized by Gram staining, serotyping and susceptibility to penicillin. In this study, isolates of the common serotype 19F were further investigated by Multi-Locus Sequence Typing (MLST). Overall, 14 different Sequence Types (STs) were identified by MLST, of which nine were novel based on the international MLST database. Two clones within serotype 19F seem to circulate in Ghana, a known ST (ST 4194) and a novel ST (ST 9090). ST 9090 was only found in healthy children in Accra, whereas ST 4194 was found equally in all children studied. In the MLST database, other isolates of ST 4194 were also associated with serotype 19F, and these isolates came from other West African countries. The majority of isolates were penicillin intermediate resistant. In conclusion, two clones within serotype 19F were found to be dominating in pneumococcal carriage in Accra and Tamale in Ghana. Furthermore, it seems as though the clonal distribution of serotype 19F may be different from what is currently known in Ghana in that many new clones were identified. This supports the importance of continued monitoring of pneumococcal carriage in Ghana and elsewhere when vaccines, e.g., PCV-13, have been introduced to monitor the possible future spread of antimicrobial resistant clones.

The objective of this study was to provide baseline information on circulating methicillin-resistant Staphylococcus aureus (MRSA) clones in Ghana. Thirty MRSA isolates collected between 2010 and 2013 from patients and healthy carriers were characterised by DNA microarray analysis, staphylococcal protein A (spa) typing, multilocus sequence typing (MLST) and minimum inhibitory concentration (MIC) determination to 21 antimicrobial agents. Phenotypic resistance was detected to tetracycline (67%), norfloxacin (40%), moxifloxacin (37%), erythromycin (37%), clindamycin (33%), gentamicin (30%), kanamycin (30%) and ceftaroline (20%), whereas no resistance was observed for glycopeptides, linezolid, daptomycin and tigecycline. DNA microarray analysis showed that tet(M) (43%), tet(K) (33%), aphA3 (23%), aacA-aphD (17%) and erm(C) (13%) were the most prevalent resistance genes. ST88-IV (WA MRSA-2) (n=8), ST8-IV (USA300) (n=5) containing arginine catabolic mobile element (ACME) and Panton-Valentine leukocidin (PVL), and ST247-I (North German/Iberian EMRSA) (n=4) were the most frequent clones detected. All MRSA contained sak and scn genes, one isolate (ST36-II) harboured the gene encoding the toxic shock syndrome toxin (TSST) and none contained exfoliative toxin genes. In conclusion, the relatively high levels of resistance to easily accessible non-β-lactam agents further complicate the treatment of MRSA infections in Ghana. The occurrence of USA300 and other epidemic multidrug-resistant MRSA clones in this African country is a matter of public health concern due to the lack of adequate infrastructures for MRSA surveillance and control in this geographical setting.

Background Blood stream infections (BSI) are critical medical conditions with high morbidity and mortality. There is paucity of information on BSI from surveillance studies in Ghana. Aim This study sought to demonstrate how useful BSI data can be gleaned from population-based surveillance, especially from resource-limited settings. Methods Data from a nationwide surveillance of antimicrobial drug resistance (AMR) in Ghana were extracted and analyzed. Secondly, we revived archived Staphylococcus aureus isolates from blood cultures that were cefoxitin resistant (CRSA), and screened these for protein A (spa) and mec A genes. Results Overall blood culture positivity was 11.2% (714/6351). All together, participating laboratories submitted 100 multidrug resistant blood culture isolates (Gram-negative = 49 and Gram-positive = 51). Prevalence of some Gram-negative isolates was as follows; Escherichia coli (20.4%), Pseudomonas aeruginosa (16.3%), Enterobacter spp. (14.3%), Salmonella serotype Typhi (8.2%) and Non-typhoidal Salmonella [NTS] (8.2%). Gram-positive pathogens included Staphylococcus aureus (66.7%), coagulase negative S. aureus [CoNS] (17.6%) and Streptococcus pneumoniae (11.8%). No methicillin resistance was confirmed in our CRSA isolates. Most blood stream associated infections were from inpatients (75%) and cultured bacteria were resistant to common and cheaper antimicrobials. Conclusion E. coli and S. aureus are common pathogens associated with BSI in Ghana and they are resistant to several antimicrobials. Active and continuous AMR surveillance can serve multiple purposes, including data generation for BSI.

Background.The current definitive treatment of Buruli ulcer with antibiotics makes the issue of antimicrobial drug resistance an unavoidable one. This is as a result of drug misuse by health personnel and patients’ noncompliance to treatment regimen. Monitoring of these factors and screening for new effective antimicrobials are crucial to effective management of Buruli ulcer disease. This study therefore investigated the inhibitory activity of some antibiotics against isolates ofMycobacterium ulcerans.Methods.Activity of eight antibiotics was tested against twelveM. ulceransisolates (2 reference strains and 10 clinical isolates). The anti-M. ulceransactivities were determined by the agar dilution method and the minimum inhibitory concentrations (MICs) were determined by the agar proportion method.Results.All antimicrobials investigated had activity againstM. ulceransisolates tested. The MICs ranged from 0.16 μg/mL to 2.5 μg/mL. Azithromycin recorded the highest inhibitory activity at a mean MIC of 0.39 μg/mL, whilst clofazimine a second-line antileprosy drug, recorded the lowest at a mean MIC of 2.19 μg/mL. Among the four antituberculosis drugs, rifampicin had the highest activity with a mean MIC of 0.81 μg/mL.Conclusion.Azithromycin could be considered as a lucrative alternative to existing treatment methods for inhibitingM. ulceransin Ghana.

BackgroundNon-typhoidal Salmonella (NTS) is a public health problem worldwide and particularly in Africa with high disease burden. This study characterized Salmonella isolates from humans in Ghana to determine serovar distribution, phage types, and antimicrobial resistance. Further, the clonal relatedness among isolates was determined.MethodsOne hundred and thirty-seven Salmonella isolates (111 clinical and 26 public toilet) were characterized using standard serotyping, phage typing, and antimicrobial susceptibility testing methods. The molecular epidemiology of common serovars (Salmonella Typhimurium and Salmonella Enteritidis) was established by pulsed field gel electrophoresis (PFGE).ResultsTwenty-two serovars were identified with S. Enteritidis, S. Typhimurium, and Salmonella Derby as the most dominant. One hundred and twelve isolates showed resistance to more than one antimicrobial. Fifty-eight (n = 58/112; 54.5%) strains were multi-resistant with low resistance to cephalosporins ceftazidime (8.0%), cefotaxime (4.5%), and cefoxitin (2.7%) with synergy to clavulanic acid indicating possible ESBLs. Isolates showed high resistance to trimethoprim (66.1%), tetracycline (61.6%), ampicillin (57.1%), sulfamethoxazole (46.4%), chloramphenicol (33.9%), and ciprofloxacin (25.0%). The most common resistance pattern of multi-resistant serovars was to ampicillin, chloramphenicol, sulphonamide, and trimethoprim. S. Enteritidis (18/43) strains reacted with typing phages but did not conform to any phage type with PT14B and PT4 as predominant definitive phage types. Six S. Typhimurium strains reacted but did not conform to any recognized phage type while seven were non-typable. The predominant definitive phage types were DT1 and DT22. PFGE patterns of human S. Enteritidis were closely related to patterns of poultry isolates obtained in a previous study in Ghana.ConclusionsCephalosporin resistance is uncommon among Salmonella from humans in Ghana. Poultry may be an important source of human salmonellosis. There is an urgent need for the implementation of routine surveillance of antimicrobial use and bacterial resistance among humans in Ghana.

Background Mycobacterium ulcerans (M. ulcerans) is the causative agent of Buruli Ulcer (BU) disease. In order to inhibit the growth of the microbial contaminants during culture of M. ulcerans, it is necessary to decontaminate BU samples with effective chemical agents. This study aimed at investigating some selected chemicals as potential decontamination agents for the isolation of M. ulcerans from swabs. Results Povidone iodine at 0.5 and 1% exhibited the lowest contamination and recovery rate for microbial contaminants and M. ulcerans. The most effective decontamination method was the protocol using 2% cetylpyridinium chloride/4% sodium chloride (recovery rate = 53%, contamination rate = 14%). The observed difference between the recovery rate of 2% CPC/4% NaC and the other protocols was however not statistically significant (p = 0.76). Conclusions Two percent (2%) cetylpyridinium chloride/4% sodium chloride can be conveniently used as an alternative decontamination method for the isolation of M. ulcerans from swabs.

Infections with bacteria producing extended-spectrum beta-lactamases (ESBLs) are increasing across Africa. This study reports on ESBL-producing Enterobacteriaceae as significant causes of infections and antibiotic resistance at Korle-Bu Teaching Hospital in Accra, Ghana. Of 300 isolates examined, 49.3% produced ESBLs. The prevalence of ESBLs was significantly high among isolates from neonates (28 of 43, 65.1%; relative risk = 1.62, 95% confidence interval = 1.33-2.13, P = 0.002) and adult patients > 65 years of age (36 of 51, 70.5%; relative risk = 1.89, 95% confidence interval = 1.41-2.40, P = 0.001). A marked increase in minimum inhibitory concentrations of ESBL-positive species was noticed compared with those for the other strains. Using these concentrations, we found that 26 (17%) ESBL producers were resistant to two or more antibiotics (aminoglycosides, fluoroquinolones, sulfonamide, and carbapenems) whereas 5 (3.2%) non-ESBL producers were multidrug resistant. Regular ESBL detection and evaluation of antibiotic resistance may help reduce the spread of ESBLs and antibiotic resistance in Ghana.

This study was carried out primarily to evaluate the public health burden related to Streptococcus pneumoniae in Ghana and to provide related preliminary molecular epidemiological data on the organism. Invasive and nasopharyngeal specimens were screened for S. pneumoniae, and isolates were subjected to serotyping, multilocus sequence typing (MLST) and antibiotic susceptibility testing. Overall, the prevalence of S. pneumoniae in cerebrospinal fluid (CSF) was 1.7%, in blood was 0.2%, and in nasopharyngeal specimens was 15.3%. The prevalence of multiple drug resistance among the isolates was 48.6%, while the percentage resistance to various drugs was in the range of 11.1-84.0%. Serotyping of the S. pneumoniae isolates showed 7 different serotypes (3, 6B, 9, 10, 14, 16 and 23F). The extent of coverage of serotypes by the 7-valent pneumococcal conjugate vaccine was 57.1%, for the 10-valent vaccine was 57.1%, and for the 13-valent vaccine was 71.4%. MLST of 7 housekeeping genes of the organism showed a high level of genetic diversity among the isolates. S. pneumoniae appears to be an important organism in invasive infections in Ghana, being the most prevalent organism in CSF in this study. The high multiple drug resistance of the organism observed heightens the public health burden, which may be controlled by pneumococcal conjugate vaccines to a large extent.

Objectives: Vibrio cholerae O1 from a 2006 outbreak in Accra were commonly resistant to multiple antimicrobials and, in particular, to trimethoprim/sulfamethoxazole, drugs commonly used in the treatment of cholera. We sought to determine the genetic basis for trimethoprim/sulfamethoxazole resistance in outbreak isolates. Methods: Twenty-seven isolates from the outbreak were screened by PCR and sequencing for class 1 and 2 integrons and for the SXT element. Results: Twenty-one of the 27 isolates examined, all from the Accra metropolitan area, carried both SXT, an integrated chromosomal element, and a class 2 integron bearing dfrA1, sat and aadA1 cassettes. All these isolates had identical random amplification of polymorphic DNA profiles and two of them also carried a class 1 integron. Conclusions: Most strains characterized carried multiple elements conferring resistance to trimethoprim. This suggests that trimethoprim/sulfamethoxazole should not be used empirically in cholera treatment.

Streptococcus pneumoniae is the cause of high mortality among children worldwide. Antimicrobial treatment and vaccination are used to control pneumococcal infections. In Ghana, data on antimicrobial resistance and the prevalence of multidrug-resistant pneumococcal clones are scarce; hence, the aim of this study was to determine the antibiogram of S. pneumoniae recovered from Ghanaian children younger than six years of age and to what extent resistances were due to the spread of certain sero- and multilocus sequence typing (MLST) types. The susceptibility of 115 pneumococcal isolates, recovered in a previous study, to six antimicrobials was determined by disk diffusion test. Overall, 90.4% of isolates were intermediate penicillin resistant, 99.1% were trimethoprim resistant, 73.0% were tetracycline resistant, and 33.9% were sulfamethoxazole resistant. Low resistance was recorded for erythromycin (2.6%) and cefotaxime (5.2%). Overall, 72.2% of isolates were resistant to penicillin (I or R) and at least two other antimicrobials. MLST of 20 isolates showing resistance to at least four antimicrobials revealed a high diversity documented by 16 different clones, none of which had previously been associated with multidrug resistance. The resistances found may have emerged due to nonprudent antimicrobial use practices and there is a need to monitor and promote prudent antimicrobial usage in Ghana.

Background Buruli ulcer (BU) is one of the neglected tropical diseases. Mycobacterium ulcerans is the aetiologic agent of Buruli ulcer. Many extensive studies have failed to isolate M. ulcerans in pure culture from the environment, even in highly endemic areas of BU. We investigated the role of macro-invertebrates as possible hosts or vectors for M. ulcerans by attempting to cultivate M. ulcerans from these organisms. Methods The study was conducted in 5 villages in the Asante Akim District of Ghana for 10 months. Primary detection of M. ulcerans was done by real-time PCR targeting insertion sequence IS2404 coupled with the detection of IS2606 and Ketone reductase genes for increased sensitivity and specificity. Primary cultures were done using routine bacteriological media for culturing mycobacteria, L-J and special enrichment liquid broth, BACTEC®. Results The overall rate of detection of IS2404 in the general macro-invertebrate population was 12.8%. Cluster of CT-values was observed around a mean value of 35.88 and range values of 28.35–38.61. Statistically, there were no significant differences between the various CT-values obtained, p>0.05. The difference in ∆CT values (IS2606-IS2404) for homogenate sample obtained from Naucoridae which was positive for the three targets on M. ulcerans genome was estimated to be 1.77. The present study reports the cultivation of two IS2404 positive Mycobacteria spp. from two aquatic macro-invertebrates of the families Belostomidae and Notonectidae both of the order Hemiptera. The isolate from Belostomidae was identified as either M. ulcerans or M. marinum with 98% identities that from Notonectidae was 98% identical to M. neoaurum. The organisms are yet to be passaged through mice footpad and fully characterised. Conclusions For the first time M. neoaurum species was reported to have harboured IS2404 element. Aquatic Hemiptera are highly suspected to be vectors or hosts for M. ulcerans and they may transmit the pathogen to humans through biting.

Background The objective of this study was to determine the prevalence of nasopharyngeal carriage, serotype distribution, and penicillin resistance of Streptococcus pneumoniae in children ≤6 years of age in Ghana. Methods A cross-sectional study was carried out on a cluster-randomized sample of children ≤6 years of age attending nurseries and kindergartens in Accra and Tamale, Ghana. Basic data on age, sex and exposure to antimicrobials in the previous month were collected on all study subjects. Nasopharyngeal swabs were obtained from participants and all pneumococcal isolates were characterized by serotyping and their penicillin resistance determined. Results The overall prevalence of pneumococcal carriage among the children was 34% in Accra and 31% in Tamale. The predominant serotypes were 19F, 6B, 23F, and 6A with 23% of the isolates being non-typable in Accra and 12% in Tamale. Only two isolates (serotypes 19F and 6B) from Tamale had a MIC >2 μg/ml and were classified as fully penicillin resistant with 45% of the isolates having intermediate resistance. Conclusions These findings indicate that the 13-valent pneumococcal conjugate vaccine (PCV-13) recently introduced in Ghana will cover 48% and 51% of the serotypes identified in Accra and Tamale, respectively. The 23-valent pneumococcal polysaccharide vaccine (PPV-23) will cover 54% of all serotypes detected. The two penicillin resistant isolates (MIC 32 μg/ml) were serotypes included in both PCV-13 and PPV-23. A nationwide monitoring system of penicillin susceptibility patterns and pneumococcal serotypes is recommended.

There is a paucity of data on Staphylococcus aureus epidemiology in Africa. Prevalence of nasal carriage and genetic diversity of S. aureus were determined among hospital staff (HS) and inpatients (IP) at the largest hospital in Ghana. In total, 632 nasal swabs were obtained from 452 IP and 180 HS in the Child Health Department (CHD) and Surgical Department (SD). S. aureus carriage prevalences were 13.9% in IP and 23.3% in HS. The chance of being a carrier was higher in HS ( P = 0.005) and IP staying ≤7 days in hospital ( P = 0.007). Resistance to penicillin (93%), tetracycline (28%) and fusidic acid (12%) was more common than for other agents ( S. aureus carriage was observed among IP compared with HS ( P = 0.01). High genetic diversity was shown by spa typing, with 55 spa types found among 105 isolates; the predominant spa types were t355 (10%) and t084 (10%). MRSA was detected in six IP with an overall carriage prevalence of ca. 1.3%, but not in HS. All three MRSA isolates from SD belonged to ST88–SCC mec IV, and two of them displayed the same spa type and antibiograms; three MRSA isolates from CHD belonged to distinct lineages (ST88–SCC mec IV, ST8–SCC mec V and ST72–SCC mec V). Altogether, these data indicate a high diversity of S. aureus , low levels of MRSA carriage, and a higher chance of nasal carriage of multidrug-resistant S. aureus among IP compared with HS in this hospital.

Buruli ulcer (BU), caused by Mycobacterium ulcerans is a chronic necrotizing skin disease. It usually starts with a subcutaneous nodule or plaque containing large clusters of extracellular acid-fast bacilli. Surrounding tissue is destroyed by the cytotoxic macrolide toxin mycolactone produced by microcolonies of M. ulcerans. Skin covering the destroyed subcutaneous fat and soft tissue may eventually break down leading to the formation of large ulcers that progress, if untreated, over months and years. Here we have analyzed the bacterial flora of BU lesions of three different groups of patients before, during and after daily treatment with streptomycin and rifampicin for eight weeks (SR8) and determined drug resistance of the bacteria isolated from the lesions. Before SR8 treatment, more than 60% of the examined BU lesions were infected with other bacteria, with Staphylococcus aureus and Pseudomonas aeruginosa being the most prominent ones. During treatment, 65% of all lesions were still infected, mainly with P. aeruginosa. After completion of SR8 treatment, still more than 75% of lesions clinically suspected to be infected were microbiologically confirmed as infected, mainly with P. aeruginosa or Proteus miriabilis. Drug susceptibility tests revealed especially for S. aureus a high frequency of resistance to the first line drugs used in Ghana. Our results show that secondary infection of BU lesions is common. This could lead to delayed healing and should therefore be further investigated., Author Summary Buruli ulcer (BU) can lead to large ulcerative lesions due to extensive skin loss caused by the necrotizing effect of the main virulence factor mycolactone. For a long time the general perception was that BU lesions are not infected by other bacteria because of a postulated antimicrobial effect of the macrolide toxin, mycolactone. In this study, we analyzed laboratory confirmed BU lesions before, during, and after streptomycin/rifampicin treatment. Contrary to popular belief, our findings show that BU lesions are frequently co-colonized with other potential bacterial pathogens before, during, and after antibiotic treatment. For example, 75% of cases that were clinically indicative of being infected after treatment were microbiologically confirmed as infected. Most microbiologically infected cases were also confirmed by histopathological analysis. The most prominent bacterial species isolated included Pseudomonas aeruginosa, S. aureus, and P. mirabilis. When we tested the isolates against first line drugs used in Ghana, the isolates were found to be resistant to most of these drugs. This study indicates that wound care practices need to be improved and that wound infection may be a common cause of wound healing delay in BU.

AIMS--Suppurative keratitis is a serious problem in all tropical countries, but very little information is available about the causative organisms in Africa. The objectives were to identify the causative organisms and the proportion of cases caused by fungi in southern Ghana, and to determine whether correct decisions about treatment could be made on the basis of Gram stain in the eye clinic. METHODS--Scrapings were taken from corneal ulcers of consecutive new patients presenting at Korle Bu Hospital, Accra, and inoculated on 'chocolate' and Sabouraud's agars. Further scrapings were taken for Gram staining and interpretation in the eye clinic. Duplicate slides were assessed by an experienced microbiologist in the UK. RESULTS--One or more organisms were cultured from 114 of 199 patients (57.3%), the most common being Fusarium species, Pseudomonas aeruginosa, and Staphylococcus epidermidis. Fungi, alone or in combination, were isolated from 56% of the patients who had positive cultures. In total, 122 patients (61.3%) had their treatment either determined or altered based on the results of the microbiological diagnosis; in 87 of these solely on the basis of direct microscopic examination. CONCLUSIONS--Infection by filamentous fungi accounted for more than half of the ulcers from which cultures were obtained. Both training in technique and experience in interpretation are necessary for microscopy based diagnosis by staff in the clinic to be of greatest value. Direct microscopy was particularly useful for detecting fungi.

Introduction: Oropharyngeal candidiasis is a common occurrence in the course of human immunodeficiency virus (HIV) disease progression. Changes in the clinical severity of oropharyngeal candidiasis and type of Candida species profile may be a reflection of immunological changes in patients. The aim of this study was to undertake a baseline Candida species identification for future reference. Methodology: Oral swabs of 267 HIV-infected patients with oropharyngeal candidiasis were cultured and Candida species were identified by API 32 C. Results: A total of 201 (75.3%) Candida species and 10 (3.7%) non candida fungi were identified. Twenty different Candida species were isolated. Candida albicans was the most prevalent species (68.5%) followed by C. tropicalis (7.4%), C. krusei (6.4%), C. parapsilosis (3.0%) and C. sake (2.5%). Other species ranged from 0.5% to 1.5%. Positive culture was independent of whether patients were on anti-retroviral therapy or not. Conclusion: Of all Candida isolates, 68.5% were identified as C. albicans. Since other uncommon species were also isolated, it may be necessary in this group of patients to identify Candida species causing severe infections.

Diarrhea is a major public health problem that affects the development of children. Anthropometric data were collected from 274 children with (N = 170) and without (N = 104) diarrhea. Stool specimens were analyzed by conventional culture, polymerase chain reaction for enteroaggregative Escherichia coli (EAEC), Shigella, Cryptosporidium, Entamoeba, and Giardia species, and by enzyme-linked immunosorbent assay for fecal lactoferrin levels. About 50% of the study population was mildly to severely malnourished. Fecal lactoferrin levels were higher in children with diarrhea (P = 0.019). Children who had EAEC infection, with or without diarrhea, had high mean lactoferrin levels regardless of nutritional status. The EAEC and Cryptosporidium were associated with diarrhea (P = 0.048 and 0.011, respectively), and malnourished children who had diarrhea were often co-infected with both Cryptosporidium and EAEC. In conclusion, the use of DNA-biomarkers revealed that EAEC and Cryptosporidium were common intestinal pathogens in Accra, and that elevated lactoferrin was associated with diarrhea in this group of children.

Background: Compliance with hand hygiene recommendations is the most important measure in preventing health care-associated infections. The objective of this study was to assess the nature of patient contact and the hand hygiene practices of nurses and physicians in the neonatal intensive care unit in a tertiary hospital in Ghana. Methodology: Unobtrusive observation of patient contact, hand hygiene practices, and hand washing technique among nurses and physicians attending randomly selected newborns for five hours daily for two weeks. Patient contact categorized as low-risk or high-risk. Hand hygiene practice before and after patient contact categorized as clean uncontaminated, clean recontaminated, new gloves, unchanged gloves. Compliance to alcohol rub use assessed. Results: The patient to nurse/physician ratio varied from 9:1 to 12:1. There were 97 patient contacts of which 49 were high-risk and 48 low-risk. Most (73%) patient contacts were from nurses. Compliance to hand hygiene recommendations before versus after patient contact was 15.4% versus 38.5% for physicians and 14.1% versus 9.9% for nurses. Gloves were used for 60.8% patient contacts (85.7% high-risk, 35.4% low-risk); however, compliance to recommended procedure occurred in only 12.2% of high-risk contacts and none of the low-risk contacts. Gloves were not changed between patients in 43.7% of high-risk contacts and 88.2% of low-risk contacts. Hand washing protocol was generally followed. Alcohol hand rub was always available but was not used for hand hygiene. Conclusions: Hand hygiene compliance of physicians and nurses was low. Gloves and alcohol rub were not used according to recommended guidelines. Incorporating effective education programs that improve adherence to hand hygiene guidelines into the continuing education curriculum of health professionals is recommended.

Japheth A Opintan,1 Mercy J Newman,1 Reuben E Arhin,1 Eric S Donkor,1 Martha Gyansa-Lutterodt,2 William Mills-Pappoe3 1Department of Medical Microbiology, School of Biomedical and Allied Health Sciences, University of Ghana, 2Pharmaceutical Services, Ministry of Health, Ghana Health Services, 3Clinical Laboratory Unit, Institutional Care Division, Ghana Health Service, Accra, Ghana Abstract: Global efforts are underway to combat antimicrobial resistance (AMR). A key target in this intervention is surveillance for local and national action. Data on AMR in Ghana are limited, and monitoring of AMR is nonexistent. We sought to generate baseline data on AMR, and to assess the readiness of Ghana in laboratory-based surveillance. Biomedical scientists in laboratories across Ghana with capacity to perform bacteriological culture were selected and trained. In-house standard operating protocols were used to perform microbiological investigations on clinical specimens. Additional microbiological tests and data analyses were performed at a centralized laboratory. Surveillance data were stored and analyzed using WHONET program files. A total of 24 laboratories participated in the training, and 1,598 data sets were included in the final analysis. A majority of the bacterial species were isolated from outpatients (963 isolates; 60.3%). Urine (617 isolates; 38.6%) was the most common clinical specimen cultured, compared to blood (100 isolates; 6.3%). Ten of 18 laboratories performed blood culture. Bacteria isolated included Escherichia coli (27.5%), Pseudomonas spp. (14.0%), Staphylococcus aureus (11.5%), Streptococcus spp. (2.3%), and Salmonella enterica serovar Typhi (0.6%). Most of the isolates were multidrug-resistant, and over 80% of them were extended-spectrum beta-lactamases-producing. Minimum inhibitory concentration levels at 50% and at 90% for ciprofloxacin, ceftriaxone, and amikacin on selected multidrug-resistant bacteria species ranged between 2 µg/mL and >256 µg/mL. A range of clinical bacterial isolates were resistant to important commonly used antimicrobials in the country, necessitating an effective surveillance to continuously monitor AMR in Ghana. With local and international support, Ghana can participate in global AMR surveillance. Keywords: antimicrobial resistance, ESBL-producing, quinolone, multiple drug resistance

Improvement in the care and treatment of neonates had contributed to their increased survival. Nosocomial infection remains an important problem in intensive care units. Hospital wards had been shown to act as reservoirs of pathogenic microorganisms associated with infection. To assess the prevalence of pathogenic organisms in the environment of the neonatal unit, 92 swabs were randomly collected from cots, incubators and various equipments in the unit and were cultured on Blood agar and MacConkey agar plates. Air contamination was detected by exposing the same types of agar plates for 3 hours in several areas of the unit. After 48 hours incubation, isolates were identified biochemically. There is marked congestion in the unit. Ninety one percent of swabs yielded growth, with coagulase negative Staphylococcus being the predominant organism (44%), followed by Bacillus species (20%), E. coli (12.5%), and Klebsiella (8.5%), Pseudomonas species (3.6%) and moulds (3.6%). Sedimentation plates had colony counts of from 10 - 100 per plate and the majority of the cultures were polymicrobial cultures. The presence of various Gram-negative bacili including known neonatal pathogens (like E. Coli and Pseudomonas) especially on ward equipment and congestion in the ward has the potential to cause nosocomial infection. Keywords: Neonates, Pathogenic microorganisms, Intensive care unit. Résumé Le progrèss dans le soin et le traitement des néo-natals avait contribué à leur survivance angmentée. L'infection nosocomique demeure un problème important dans les centres de soins intensifs. Les salles d'hôpital avaient été témoignées (d'agir) comme des réservoirs de microorganisms pathogéniques associés à l'infection. Pour évaluer la prévalence des organismes pathogéniques dans l'environnement du center néo- natal, 92 tampons furent, par occurrenœ, accumulés de couches, couveuses et diverses installations dans le center, et furent cultivés sur la gélose de sang et (la gélose) de plaque MacConkey. La contamination d'air fut découverte en exposant les mêmes types de plaques de culture pour 3 heures dans les endroits variés du center. Après 48 heures d'incubation, les isolés furent, biochimiquement identifiés. II y a une congestion jalonnée dans le center. Quatre-vingt-onze pour cent de tampons rapportèrent une croissanœ, d'avec staphylocoque négatif coagulum, étant l'organisme (44%) prédominant, suivi par l'espece de bacilli (20%), E.coli (colique) (12.5%), et Klebsiella (8.5%), l' espece de Pseudomonas (3.6%) et les moisissures (3.6%). Les plaques de sédimentation avaient des calculs de colonie de 10-100 par plaque et a majorité des cultures furent des cultures polymicrobiques. La présence de grammes variés-les bacilli négatifs, comprenant les pathogènes néo-natals (comme E. coli et Pseudomonas) particulièrement sur l'installation de salle d'hôpital et la congestion dans la salle d'hôpital a du potentiel de causer l'infection nosocomique. West African Journal of Medicine Vol.21(4) 2002: 310-312

The objective of this study was to determine the antimicrobial susceptibility patterns and clonal diversity of clinical Staphylococcus aureus isolates from Ghana. A total of 308 S. aureus isolates from six healthcare institutions located across Northern, Central and Southern Ghana were characterized by antibiotyping, spa typing and PCR detection of Panton Valentine leukocin (PVL) genes. Methicillin-resistant S. aureus (MRSA) were confirmed by PCR detection of mecA gene and further characterized by SCCmec and multi-locus sequence typing (MLST). The prevalence of antimicrobial resistance was below 5% for all agents tested except for penicillin (97%), tetracycline (42%) and erythromycin (6%). Ninety-one spa types were found, with t355 (ST152, 19%), t084 (ST15, 12%) and t314 (ST121, 6%) being the most frequent types. Based on established associations between spa and MLST types, isolates were assigned to 16 clonal complexes (CCs): CC152 (n = 78), CC15 (n = 57), CC121 (n = 39), CC8 (n = 36), CC5 (n = 33), CC1 (n = 29), CC45 (n = 9), CC88 (n = 8), CC30 (n = 4), CC9 (n = 3), CC25 (n = 2), CC97 (n = 2) CC20 (n = 2), CC707 (n = 2), CC7 (n = 3) and CC522 (n = 1). Most isolates (60%) were PVL-positive, especially those belonging to ST152, ST121, ST5, ST15, ST1, ST8, and ST88. Nine (3%) isolates were MRSA belonging to seven distinct clones: ST88-IV (n = 2), ST250-I (n = 2), ST8-IV (n = 1), ST72-V (n = 1), ST789-IV (n = 1), ST2021-V (n = 1), and ST239-III (n = 1). The study confirmed a high frequency of PVL-positive S. aureus in Africa, low prevalence of antimicrobial resistance and high diversity of MRSA lineages in Ghana compared to developed countries and other African countries. The detection of known pandemic MRSA clones in the absence of routine MRSA identification in most Ghanaian clinical microbiology laboratories calls for capacity building to strengthen surveillance and prevent spread of these clones.

Background Antimicrobial resistance is a growing international problem. We observed a 50% increase in the prevalence of trimethoprim resistance among fecal Escherichia coli from healthy Nigerian students between 1998 and 2005, a trend to increase that continued in 2009. Methods and Findings A PCR-based screen revealed that 131 (43.1%) of isolates obtained in Nigeria in 2005 and 2009 carried integron-borne dfrA cassettes. In the case of 67 (51.1%) of these isolates, the cassette was a class 1-integron-borne dfrA7 gene, which has been reported at high prevalence from E. coli isolates from other parts of Africa. Complete sequencing of a 27 Kb dfrA7-bearing plasmid from one isolate located the dfrA7 gene within a Tn21-type transposon. The transposon also contained an IS26-derived bla/sul/str element, encoding resistance to β-lactams, sulphonamides and streptomycin, and mercury resistance genes. Although the plasmid backbone was only found in 12 (5.8%) of trimethoprim-resistant isolates, dfrA7 and other transposon-borne genes were detected in 14 (16.3%) and 32 (26.3%) of trimethoprim resistant isolates collected in Nigeria in 2005 and 2009, respectively. Additionally, 37 (19.3%) of trimethoprim-resistant E. coli isolates collected between 2006 and 2008 from Ghana were positive for the dfrA7 and a transposon marker, but only 4 (2.1%) harbored the plasmid backbone. Conclusions Our data point to transposition as a principal mechanism for disseminating dfrA7 among E. coli from Nigeria and Ghana. On-going intensive use of the affordable broad-spectrum antibacterials is likely to promote selective success of a highly prevalent transposable element in West Africa.

Objective To examine the frequency of community-acquired salmonella bloodstream infection in Ghanaian children and the occurrence of antibiotic resistance in salmonellae. Methods The study comprised 472 patients with a blood culture obtained within 48 h of admission to the pediatric department of Korle Bu Teaching Hospital in Accra, Ghana, over a 3-month period. All Salmonella isolates from blood cultures were speciated and antibiotic susceptibility tests were performed. Clinical data of children with salmonella bloodstream infection were compared to those of controls. Two control groups were identified: all children enrolled in the study without salmonella bloodstream infection (group 1), and those with bloodstream infection due to other organisms (group 2). Results A pathogen was isolated from 111 children (23.5%), and salmonellae were among the most common isolates ( n =24; 21.6%). Among Salmonella strains, S. enteritidis ( n = 14; 59%) predominated over S. typhi ( n =6; 25%). Resistance to several antibiotics was only found in S. enteritidis isolates ( n =8; 57%). Children with salmonella bloodstream infection presented more often than controls with severe anemia, jaundice, abdominal pain and distension as well as hepatomegaly and splenomegaly. They were also hospitalized for a significantly longer period, but the case-fatality rate was similar. Conclusions Salmonella bloodstream infection, especially due to non-typhoidal strains, is a potential health problem for Ghanaian children and may be complicated by resistance to the commonly available antibiotics.