Wastewater from healthcare centers in Burkina Faso is a source of ESBL, AmpC-ß-lactamase and carbapenemase-producing Escherichia coli and Klebsiella pneumoniae.

BACKGROUND: Extended-spectrum ß-lactamase (ESBL), plasmid-mediated AmpC-ß-lactamase and carbapenemase-producing Escherichia coli and Klebsiella pneumoniae have spread into the environment worldwide posing a potential public health threat. However, the prevalence data for low- and middle-income countries are still scarce. The aim of this study was to evaluate the presence of ESBL, AmpC-ß-lactamase and carbapenemase-producing and multidrug-resistant E. coli and K. pneumoniae in wastewaters from healthcare centers in Burkina Faso. RESULTS: Eighty-four (84) wastewater samples were collected from five healthcare centers and plated on selective ESBL ChromAgar. E. coli and Klebsiella pneumoniae isolates were identified using API20E. ESBL-producing bacteria were detected in 97.6% of the samples and their average concentration per hospital ranged from 1.10 × 105 to 5.23 × 106 CFU/mL. Out of 170 putative ESBL-producing isolates (64% of them were E. coli) and 51 putative AmpC-ß-lactamase-producing isolates, 95% and 45% were confirmed, respectively. Carbapenemase production was detected in 10 isolates, of which 6 were NDM producers, 3 were OXA-48 producers and 1 was NDM and OXA-48 producer. All isolates were multidrug resistant and, moreover, all of them were resistant to all tested ß-lactams. Resistance to ESBL inhibitors was also common, up to 66% in E. coli and 62% in K. pneumoniae. Amikacin, fosfomycin and nitrofurantoin were the antibiotics to which the least resistance was detected. CONCLUSIONS: This study showed that wastewater from healthcare centers constitutes a reservoir of multidrug-resistant bacteria in Burkina Faso, including carbapenemase producers. Untreated healthcare wastewater entering the environment exposes people and animals to infections caused by these multi-resistant bacteria, which are difficult to treat, especially in the resource-poor settings.

Isolation and characterization of three novel Acinetobacter baumannii phages from Beninese hospital wastewater.

Acinetobacter baumannii is an opportunistic pathogen that is mostly associated with hospital-acquired infections. The rapid emergence of multi- and pan-drug-resistant Acinetobacter strains poses an increasing challenge in hospitals. Phage therapy offers one treatment option for infections caused by A. baumannii. We isolated three phages from Beninese hospital wastewater - fBenAci001, fBenAci002, and fBenAci003 - that infected clinical A. baumannii strains from Finnish patients. Phylogenetic analysis showed that these phages resemble phages of the genus Friunavirus, family Autographiviridae. The isolated phages meet the requirements set for phages used for phage therapy. However, they were found to have a narrow host range, which may limit their therapeutic use.

Characterization of Salmonella Typhimurium isolates from domestically acquired infections in Finland by phage typing, antimicrobial susceptibility testing, PFGE and MLVA.

BACKGROUND: Salmonella enterica spp. enterica serotype Typhimurium (STM) is the most common agent of domestically acquired salmonellosis in Finland. Subtyping methods which allow the characterization of STM are essential for effective laboratory-based STM surveillance and for recognition of outbreaks. This study describes the diversity of Finnish STM isolates using phage typing, antimicrobial susceptible testing, pulsed-field gel electrophoresis (PFGE) and multilocus variable-number tandem repeat analysis (MLVA), and compares the discriminatory power and the concordance of these methods. RESULTS: A total of 375 sporadic STM isolates were analysed. The isolates were divided into 31 definite phage (DT) types, dominated by DT1 (47 % of the isolates), U277 (9 % of the isolates) and DT104 (8 % of the isolates). Of all the isolates, 62 % were susceptible to all the 12 antimicrobials tested and 11 % were multidrug resistant. Subtyping resulted in 83 different XbaI-PFGE profiles and 111 MLVA types. The three most common XbaI-PFGE profiles (STYM1, STYM7 and STYM8) and one MLVA profile with three single locus variants accounted for 56 % and 49 % of the STM isolates, respectively. The studied isolates showed a genetic similarity of more than 70 % by XbaI-PFGE. In MLVA, 71 % of the isolates lacked STTR6 and 77 % missed STTR10p loci. Nevertheless, the calculated Simpson's diversity index for XbaI-PFGE was 0.829 (95 % CI 0.792-0.865) and for MLVA 0.867 (95 % CI 0.835-0.898). However, the discriminatory power of the 5-loci MLVA varied among the phage types. The highest concordance of the results was found between XbaI-PFGE and phage typing (adjusted Wallace coefficient was 0.833 and adjusted Rand coefficient was 0.627). CONCLUSIONS: In general, the calculated discriminatory power was higher for genotyping methods (MLVA and XbaI-PFGE) than for phenotyping methods (phage typing). Overall, comparable diversity indices were calculated for PFGE and MLVA (both DI > 0.8). However, MLVA was phage type dependent providing better discrimination of the most common phage types. Furthermore, 5-loci MLVA was a less laborious method and easier to interpret than XbaI-PFGE. Thus, the laboratory-based surveillance of the Finnish human STM infections has been conducted with a combination of phage typing, antimicrobial susceptibility testing and 5-loci MLVA since January 2014.

Prevalence and diversity of Salmonella enterica in water, fish and lettuce in Ouagadougou, Burkina Faso.

BACKGROUND: This study investigated the prevalence, serotypes and antimicrobial sensitivity patterns of Salmonella enterica in environment in Ouagadougou, Burkina Faso. A total of 476 samples, consisting of 36 samples of tap water, 51 samples of well water, 87 samples of channel water, 44 samples of reservoir water, 238 samples of fish, and 20 samples of lettuce were examined using standard bacteriological procedures for Salmonella. RESULTS: Salmonella were isolated from 98 samples. Salmonella were rare in drinking water, since they were not found at all from the tap water, and only in 2 % of well water. Salmonella were more common in the water of reservoir of Tanghin (15 %), reservoir of Yamtenga (20 %), and in the water channels in the city (from 20 to 31 %). Salmonella were commonly isolated from the fish (24 %) caught from the reservoir of Tanghin and from the lettuce (50 %) irrigated with water from Tanghin. The Salmonella isolates were found to represent 50 different serotypes. The 11 most common serotypes were Salmonella Bredeney and S. Colindale (both 8.2 %), S. Muenster (6.1 %), S. Korlebu (5.1 %), S. Eastbourne and S. Poona (both 4.1 %), and S. Agona, S. Derby, S. Drac, S. Senftenberg, S. Waycross (each 3.1 %), accounting for 51.3 % of all the isolates. In general, the Salmonella strains were sensitive to the antimicrobials tested, but two strains were resistant to streptomycin and many more intermediate to streptomycin or sulphonamide. CONCLUSION: This study highlights the common prevalence of Salmonella and the high diversity of Salmonella serotypes in aquatic environment in Ouagadougou, Burkina Faso. Therefore, various human activities linked to water and consumption of water-related products, such as fish and lettuce, can lead to human Salmonella infections.

Development of microbiological field methodology for water and food-chain hygiene analysis of Campylobacter spp. and Yersinia spp. in Burkina Faso, West Africa.

Field-adaptable research methods for identifying Campylobacter sp., Yersinia sp. and other pathogenic and indicator bacteria were designed in Finland and tested in Burkina Faso. Several bacterial groups were also validated from artificially contaminated samples. Campylobacter strains were cultivated using an innovative gas generation system: The 'Portable Microbe Enrichment Unit' (PMEU) which provides microaerobic gas flow into the enrichment broth. This enhanced cultivation system produced rapid growth of several isolates of campylobacteria from water and chicken samples. The latter were obtained from local marketplace samples. No yersinias were found in the field studies, whereas they were readily recovered from the spiked samples, as well as Salmonella sp. and Escherichia coli strains. The PMEU method turned out to be reliable for monitoring of water and food hygiene in remote locations.

Strengthening clinical bacteriology laboratory diagnostics to combat sepsis and antimicrobial resistance in Benin: a train-the-trainer approach.

Introduction: Improved laboratory diagnostics is needed to support sepsis diagnosis and combat increasing antibiotic resistance in Benin. We trained clinical laboratory experts and technicians to improve their skills in accurate and up-to-date diagnostics. Methods: A Train-the-Trainer (TtT) approach was used to design the course that combines theoretical and practical laboratory skills, specifically addressing the knowledge gaps we had previously identified in our national survey. Pedagogical methods were student-centered, including peer learning, use of online materials, practical laboratory work and pre-and post-course tests. Results: We first trained 10 trainers who in turn trained 40 laboratory technicians from across the country, from both public and private clinical and veterinary laboratories. The trainers also prepared standard operation procedures for blood culture and antibiotic susceptibility testing based on international standards. Three months after the training, follow-up visits were made to the laboratories where the implementation of the new skills was evaluated. The progress of the participants observed during the course and the implementation of the new skills afterwards proved the training to be effective. Discussion: The professional networks created during the training, the empowerment that utilizes local knowledge resources, and the government support for our initiative can be expected to bring sustainability to the initiative and support the participation of Beninese laboratories in international surveillance programs in the future.

Prevalence and characterization of Salmonella enterica from the feces of cattle, poultry, swine and hedgehogs in Burkina Faso and their comparison to human Salmonella isolates.

BACKGROUND: Production and wild animals are major sources of human salmonellosis and animals raised for food also play an important role in transmission of antimicrobial resistant Salmonella strains to humans. Furthermore, in sub-Saharan Africa non-typhoidal Salmonella serotypes are common bloodstream isolates in febrile patients. Yet, little is known about the environmental reservoirs and predominant modes of transmission of these pathogens. The purpose of this study was to discover potential sources and distribution vehicles of Salmonella by isolating strains from apparently healthy slaughtered food animals and wild hedgehogs and by determining the genetic relatedness between the strains and human isolates. For this purpose, 729 feces samples from apparently healthy slaughtered cattle (n = 304), poultry (n = 350), swine (n = 50) and hedgehogs (n = 25) were examined for the presence of Salmonella enterica in Burkina Faso. The isolates were characterized by serotyping, antimicrobial-susceptibility testing, phage typing, and pulsed-field gel electrophoresis (PFGE) with XbaI and BlnI restriction enzymes. RESULTS: Of the 729 feces samples, 383 (53%) contained Salmonella, representing a total of 81 different serotypes. Salmonella was present in 52% of the cattle, 55% of the poultry, 16% of the swine and 96% of the hedgehog feces samples. Antimicrobial resistance was detected in 14% of the isolates. S. Typhimurium isolates from poultry and humans (obtained from a previous study) were multiresistant to the same antimicrobials (ampicillin, chloramphenicol, streptomycin, sulfonamides and trimethoprim), had the same phage type DT 56 and were closely related in PFGE. S. Muenster isolates from hedgehogs had similar PFGE patterns as the domestic animals. CONCLUSIONS: Based on our results it seems that production and wild animals can share the same Salmonella serotypes and potentially transmit some of them to humans. As the humans and animals often live in close vicinity in Africa and the hygiene control of the meat retail chain is defective, high Salmonella carriage rates of the animals can pose a major public health risk in Burkina Faso. This underlines the necessity for a joint and coordinated surveillance and monitoring programs for salmonellosis in Africa.

Bacterial and viral etiology of childhood diarrhea in Ouagadougou, Burkina Faso.

BACKGROUND: Diarrhea is the most frequent health problem among children in developing countries. This study investigated the bacterial and viral etiology and related clinical and epidemiological factors in children with acute diarrhea in Ouagadougou, Burkina Faso. METHODS: Stool specimens were collected from 283 children under 5 years of age visiting hospital due to acute diarrhea and from 60 healthy controls of similar age. Pathogens were investigated by using conventional culture techniques, PCR and immunochromatographic testing. Salmonella and Shigella strains were serotyped and their susceptibility to 23 antimicrobial agents was determined by the agar dilution method. RESULTS: At least one pathogen was detected in 64% of the 283 patients and in 8% of the 60 controls (p < 0.001). Rotavirus was found in 30% of the patients, followed by diarrheagenic Escherichia coli (24%), Salmonella enterica ssp. enterica (9%), Shigella spp. (6%), adenovirus (5%) and Campylobacter spp. (2%). Multiple pathogens were found in 11% of the patients and in 2% of the controls (p = 0.028). Viruses were found mainly in children of ≤ 2 years of age, whereas bacteria were equally prevalent among all the age groups. Viral infections occurred mostly during the cool dry season and the bacterial infections during the rainy season. Fever (64%) and vomiting (61%) were the most common symptoms associated with diarrhea. Only one Salmonella strain was resistant to nalidixic acid and ciprofloxacin. Of the Shigella strains, one was resistant to nalidixic acid but 81% to trimethoprim- sulfamethoxazole, 63% to streptomycin and 50% to ampicillin. Most of all the other Salmonella and Shigella strains were sensitive to all antimicrobials tested. CONCLUSION: Rotaviruses and diarrheal E. coli were the most predominant pathogens associated with acute diarrhea in Burkinabe children. Constant antimicrobial surveillance is warranted to observe for the emergence of enteric bacteria resistant to antimicrobials that are important in treatment also of severe infections.

Prevalence and characterization of ESBL-producing Escherichia coli in healthy pregnant women and hospital environments in Benin: an approach based on Tricycle.

Introduction: Extended-Spectrum Beta-Lactamase (ESBL)-producing Enterobacterales are recognized as significant pathogens due to their resistance to multiple antibiotics. This study aimed to determine the prevalence of ESBL-producing Escherichia coli (E. coli) in different settings, including healthy pregnant women, the food chain, and the environment of tertiary hospitals in Benin. Methods: Samples were collected from various sources, including fecal samples from healthy pregnant women, food samples from hospital canteens, and hospital effluents from four tertiary hospitals in southern Benin. Fecal samples were plated on MacConkey agar supplemented with cefotaxime (4 µg/mL), while food and water samples were plated on Tryptone Bile X agar supplemented with cefotaxime (4 µg/mL). Urea indole tests were used for preliminary identification of E. coli colonies, followed by confirmation of ESBL production using the double disk synergy technique. Antibiotic susceptibility testing of ESBL-producing E. coli strains was conducted using the disk diffusion method on MH agar. Polymerase Chain Reaction (PCR) was used to investigate the presence of ESBL-encoding genes. Results: Among the 296 fecal samples collected from four tertiary hospitals, ESBL-producing E. coli was isolated from 22.30% (66) of the samples. All E. coli isolates from hospital effluents exhibited ESBL production, while ESBL-producing E. coli was not detected in food and drinking water samples. The analysis of variable associations showed no significant associations (p > 0.05) for the studied factors. Antibiotic susceptibility testing revealed high resistance rates among the ESBL-Ec isolates against several tested antibiotics, including amoxicillin, aztreonam, ceftriaxone, ciprofloxacin, and trimethoprim-sulfamethoxazole. However, most isolates remained susceptible to ertapenem, amoxicillin-clavulanate, and imipenem. The most prevalent ESBL-encoding genes were blaTEM (37.50%), blaOXA-1 (19.44%), and blaSHV (11.11%), while a smaller proportion of isolates carried blaCTXM-1/blaCTXM-15 (5.55%) and blaCTXM-9. Discussion: This study provides insights into the prevalence of ESBL-producing E. coli carriage in the feces of healthy pregnant women in southern Benin. Additionally, it highlights hospital wastewater as a potential reservoir of ESBL-producing bacteria in the environment. The detection of ESBL-producing E. coli in hospital effluents raises concerns about the dissemination of antibiotic resistance genes into the environment. The high resistance rates observed among ESBL-Ec isolates against commonly used antibiotics emphasize the urgent need for antimicrobial stewardship and infection control measures. The identification of prevalent ESBL-encoding genes contributes to understanding the genetic basis of ESBL resistance in the studied population. Further research is warranted to explore the mechanisms of transmission and potential interventions to mitigate the spread of ESBL-producing Enterobacterales.

Metagenomic Analysis of the Abundance and Composition of Antibiotic Resistance Genes in Hospital Wastewater in Benin, Burkina Faso, and Finland.

Antibiotic resistance is a global threat to human health, with the most severe effect in low- and middle-income countries. We explored the presence of antibiotic resistance genes (ARGs) in the hospital wastewater (HWW) of nine hospitals in Benin and Burkina Faso, two low-income countries in West Africa, with shotgun metagenomic sequencing. For comparison, we also studied six hospitals in Finland. The highest sum of the relative abundance of ARGs in the 68 HWW samples was detected in Benin and the lowest in Finland. HWW resistomes and mobilomes in Benin and Burkina Faso resembled each other more than those in Finland. Many carbapenemase genes were detected at various abundances, especially in HWW from Burkina Faso and Finland. The blaGES genes, the most widespread carbapenemase gene in the Beninese HWW, were also found in water intended for hand washing and in a puddle at a hospital yard in Benin. mcr genes were detected in the HWW of all three countries, with mcr-5 being the most common mcr gene. These and other mcr genes were observed in very high relative abundances, even in treated wastewater in Burkina Faso and a street gutter in Benin. The results highlight the importance of wastewater treatment, with particular attention to HWW. IMPORTANCE The global emergence and increased spread of antibiotic resistance threaten the effectiveness of antibiotics and, thus, the health of the entire population. Therefore, understanding the resistomes in different geographical locations is crucial in the global fight against the antibiotic resistance crisis. However, this information is scarce in many low- and middle-income countries (LMICs), such as those in West Africa. In this study, we describe the resistomes of hospital wastewater in Benin and Burkina Faso and, as a comparison, Finland. Our results help to understand the hitherto unrevealed resistance in Beninese and Burkinabe hospitals. Furthermore, the results emphasize the importance of wastewater management infrastructure design to minimize exposure events between humans, HWW, and the environment, preventing the circulation of resistant bacteria and ARGs between humans (hospitals and community) and the environment.

Population structure and antimicrobial resistance among Klebsiella isolates sampled from human, animal, and environmental sources in Ghana: a cross-sectional genomic One Health study.

BACKGROUND: One Health approaches to address the increasing threat of antimicrobial resistance (AMR) are gaining attention. However, data on the distribution and movement of bacteria and their AMR-associated genes between clinical and non-clinical sources are scarce, especially from low-income and middle-income countries. We aimed to analyse Klebsiella isolates from various sources in Ghana and compare the prevalence of AMR with datasets from two other countries. METHODS: We conducted a cross-sectional genomic One Health study. Multiple clinical, environmental, and animal sources were sampled from 78 locations (eg, hospitals, residential areas, and farms) in and around Tamale, Ghana. Clinical samples were collected through routine screening and in cases of suspected infection between March 15 and Sept 15, 2019, and samples from the wider environment were collected during a dedicated sampling effort between the dates of Aug 19, 2018, and Sept 26, 2019. Sampling locations were approximately evenly distributed from the centre of the city and steadily outwards to capture both rural and urban locations. Samples with positive growth for Klebsiella were included. Isolates of Klebsiella were obtained from the samples using Simmons citrate agar medium and characterised by antimicrobial susceptibility testing and whole-genome sequencing. A comparative analysis with Klebsiella population surveys from Pavia, Italy, and Tromsø, Norway, was performed. AMR-associated and virulence genes were detected, and the population distribution of these genes was studied. FINDINGS: Of 957 samples collected around Tamale, Ghana, 620 were positive for Klebsiella spp. 573 Klebsiella isolates were successfully sequenced, of which 370 were Klebsiella pneumoniae. Only two hospital isolates were carbapenem-resistant. Extended-spectrum ß-lactamase (ESBL) genes were relatively common among the Ghanaian clinical isolates but rare in the environmental samples. Prevalence of ESBL genes in human-hospital disease samples was 64% (14 of 22 isolates) in Ghana and 44% (four of nine isolates) in Italy, and prevalence in human-hospital carriage samples was 7% (eight of 107) in Ghana and 13% (54 of 428) in Italy; the prevalence was higher in human-hospital disease samples than in human-hospital carriage samples in both countries, and prevalence across both samples in both countries was higher than in Norway. Ghanaian isolates showed evidence of high recombination rates (recombination events compared with point mutations [r/m] 9·455) and a considerable accessory gene overlap with isolates from Italy and Norway. INTERPRETATION: Although several AMR-associated gene classes were observed relatively frequently in non-clinical sources, ESBL, carbapenemase, and virulence genes were predominantly present only in hospital samples. These results suggest that interventions should be focused on clinical settings to have the greatest effect on the prevalence and dissemination of AMR-associated genes. FUNDING: European Research Council (742158), Academy of Finland EuroHPC grant, Trond Mohn Foundation (BATTALION grant), and Wellcome Trust.

Phenotypic Detection of Carbapenemase and AmpC-ß-Lactamase Production among Extended Spectrum ß-Lactamase (ESBL)-Producing Escherichia coli and Klebsiella spp. Isolated from Clinical Specimens.

Introduction: Data on antimicrobial resistance (AMR) are sparse across numerous African countries, as microbiological analyses are not routinely conducted and surveillance data are not collected. Accordingly, clinical samples are not routinely tested for carbapenem-resistant bacteria and, therefore, the general understanding of their prevalence in the region remains limited. Methods: Between January 2020 and June 2022, we collected extended spectrum ß-lactamase (ESBL)-producing Enterobacterales (ESBL-PE) isolates from five hospitals in Burkina Faso. After an initial culture on ESBL-selective media, the species were identified using API20E and isolates were tested against 13 antimicrobial agents using the disc diffusion method on Mueller-Hinton (MH) agar. ESBL production was confirmed via a double-disc synergy test. Production of carbapenemases and AmpC-ß-lactamases and phenotypic co-resistance were determined. Results: Among the 473 ESBL-PE, 356 were ESBL-E. coli (ESBL-Ec) and 117 were Klebsiella spp. (ESBL-K). Of these isolates, 5.3% were carbapenemase and 5.3% were AmpC-ß-lactamase-positive. Three types of carbapenemases were identified: 19 NDM, 3 OXA-48-like and 1 VIM. Two isolates produced both NDM and OXA-48-like carbapenemases. Carbapenemase producers were detected at all levels of healthcare. Co-resistance rates were up to 85% for aminoglycosides, 90% for sulfonamides, 95% for fluoroquinolones and 25% for chloramphenicol. Fosfomycin resistance was 6% for ESBL-Ec and 49% for ESBL-K (49%). Conclusions: Some of the ESBL-Ec and ESBL-K co-produced carbapenemases and/or AmpC-ß-lactamases at all healthcare levels and in various sample types with high co-resistance rates to non-betalactams. Carbapenem resistance is no longer rare, calling for testing in routine diagnostics, a comprehensive resistance surveillance system and infection control within healthcare.

Clinical isolates of Yersinia enterocolitica biotype 1A represent two phylogenetic lineages with differing pathogenicity-related properties.

BACKGROUND: Y. enterocolitica biotype (BT) 1A strains are often isolated from human clinical samples but their contribution to disease has remained a controversial topic. Variation and the population structure among the clinical Y. enterocolitica BT 1A isolates have been poorly characterized. We used multi-locus sequence typing (MLST), 16S rRNA gene sequencing, PCR for ystA and ystB, lipopolysaccharide analysis, phage typing, human serum complement killing assay and analysis of the symptoms of the patients to characterize 298 clinical Y. enterocolitica BT 1A isolates in order to evaluate their relatedness and pathogenic potential. RESULTS: A subset of 71 BT 1A strains, selected based on their varying LPS patterns, were subjected to detailed genetic analyses. The MLST on seven house-keeping genes (adk, argA, aroA, glnA, gyrB, thrA, trpE) conducted on 43 of the strains discriminated them into 39 MLST-types. By Bayesian analysis of the population structure (BAPS) the strains clustered conclusively into two distinct lineages, i.e. Genetic groups 1 and 2. The strains of Genetic group 1 were more closely related (97% similarity) to the pathogenic bio/serotype 4/O:3 strains than Genetic group 2 strains (95% similarity). Further comparison of the 16S rRNA genes of the BT 1A strains indicated that altogether 17 of the 71 strains belong to Genetic group 2. On the 16S rRNA analysis, these 17 strains were only 98% similar to the previously identified subspecies of Y. enterocolitica. The strains of Genetic group 2 were uniform in their pathogenecity-related properties: they lacked the ystB gene, belonged to the same LPS subtype or were of rough type, were all resistant to the five tested yersiniophages, were largely resistant to serum complement and did not ferment fucose. The 54 strains in Genetic group 1 showed much more variation in these properties. The most commonly detected LPS types were similar to the LPS types of reference strains with serotypes O:6,30 and O:6,31 (37%), O:7,8 (19%) and O:5 (15%). CONCLUSIONS: The results of the present study strengthen the assertion that strains classified as Y. enterocolitica BT 1A represent more than one subspecies. Especially the BT 1A strains in our Genetic group 2 commonly showed resistance to human serum complement killing, which may indicate pathogenic potential for these strains. However, their virulence mechanisms remain unknown.

Characterization of Salmonella enterica and detection of the virulence genes specific to diarrheagenic Escherichia coli from poultry carcasses in Ouagadougou, Burkina Faso.

One hundred chicken carcasses purchased from three markets selling poultry in Ouagadougou, Burkina Faso, between June 2010 and October 2010 were examined for their microbiological quality. The presence of Salmonella was investigated using standard bacteriological procedures, and the isolates obtained were serotyped and tested for antimicrobial susceptibility. The presence of virulence-associated genes of the five main pathogroups of diarrheagenic Escherichia coli-Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli (EPEC), enteroaggregative E. coli (EAEC), enterotoxigenic E. coli, and enteroinvasive E. coli-was investigated using 16-plex polymerase chain reaction (PCR) on the mixed bacterial cultures from the poultry samples. Of the 100 chicken carcasses studied, 57 were contaminated by Salmonella; 16 different serotypes were identified, the most frequent being Salmonella Derby, found in 28 samples. Four Salmonella strains were resistant to tetracycline, and two were resistant to streptomycin. Based on the PCR detection of the virulence genes, in total, 45 carcasses were contaminated by three pathogroups of E. coli: STEC, EPEC, or EAEC. The STEC and EPEC virulence genes were detected on six and 39 carcasses, respectively. EAEC virulence genes were only detected in combination with those of EPEC (on 11 carcasses) or STEC (on two carcasses). The STEC-positive carcasses contained the genes stx(1), stx(2), eaeA, escV, and ent in different combinations. None of the EPEC-positive carcasses contained the bfp gene, indicating that only atypical EPEC was present. EAEC virulence genes detected were aggR and/or pic. The high proportion of chicken carcasses contaminated by Salmonella and diarrheagenic E. coli indicates a potential food safety risk for consumers and highlights the necessity of public awareness of these pathogens.

Characterization of Shigatoxigenic Escherichia coli strains from Burkina Faso.

Shigatoxigenic Escherichia coli (STEC) cause serious foodborne infections that lead to diarrheal disease and sequelae worldwide. In Burkina Faso, West Africa, STEC strains from environmental and human sources have not been isolated and characterized before. In this study, 21 STEC strains were isolated from food samples of animal origin and human feces using colony hybridization of the Shiga toxin gene stx. The STEC strains belonged to 15 different serotypes, including O43:H2, O8:H(-), and O2:H2. All strains were positive for stx(1) and 10 also for stx(2). The most common stx(1) subtype was stx(1a), and the most common stx(2) subtype was stx(2b). In five strains, stx(2) subtypes stx(2a) and/or stx(2c), which were previously associated with hemolytic uremic syndrome, were present. Some of the strains possessed the gene saa, encoding autoagglutinating adhesin. None of the strains possessed the gene eae, encoding intimin. Two STEC strains carried also an enterotoxigenic E. coli-associated gene estIa, encoding heat-stable enterotoxin. The STEC isolated from food in Burkina Faso are potentially pathogenic for humans based on the virulence gene combinations that they possess and phenotypes that they express.

Assessment of blood cultures and antibiotic susceptibility testing for bacterial sepsis diagnosis and utilization of results by clinicians in Benin: A qualitative study.

Objectives: We assessed the current status of blood culture and antibiotic susceptibility testing (AST) practices in clinical laboratories in Benin, and how the laboratory results are used by physicians to prescribe antibiotics. Methods: The qualitative study covered twenty-five clinical laboratories with a bacteriology unit and associated hospitals and pharmacies. Altogether 159 laboratory staff, physicians and pharmacists were interviewed about their perceptions of the state of laboratory diagnostics related to sepsis and the use of antibiotics. Face-to-face interviews based on structured questionnaires were supported by direct observations when visiting five laboratories in across the country. Results: Only 6 laboratories (24%) conducted blood cultures, half of them with a maximum of 10 samples per month. The most common gram-negative bacteria isolated from blood cultures were: Escherichia coli, Salmonella spp. and Salmonella enterica serovar Typhi while the most common gram-positives were Enterococcus spp. and Staphylococcus aureus. None of the laboratories listed Klebsiella pneumoniae among the three most common bacteria isolated from blood cultures, although other evidence indicates that it is the most common cause of sepsis in Benin. Due to limited testing capacity, physicians most commonly use empirical antibiotic therapy. Conclusions: More resources are needed to develop laboratory testing capacity, technical skills in bacterial identification, AST, quality assurance, and communication of results must be strengthened.

Multilocus variable-number tandem-repeat analysis, pulsed-field gel electrophoresis, and antimicrobial susceptibility patterns in discrimination of sporadic and outbreak-related strains of Yersinia enterocolitica.

BACKGROUND: We assessed the potential of multilocus variable-number tandem-repeat analysis (MLVA), pulsed-field gel electrophoresis (PFGE), and antimicrobial susceptibility testing for discriminating 104 sporadic and outbreak-related Yersinia enterocolitica (YE) bio/serotype 3-4/O:3 and 2/O:9 isolates. MLVA using six VNTR markers was performed in two separate multiplex PCRs, and the fluorescently labeled PCR products were accurately sized on an automated DNA sequencer. RESULTS: MLVA discriminated 82 sporadic YE 3-4/O:3 and 2/O:9 strains into 77 types, whereas PFGE with the restriction enzyme NotI discriminated the strains into 23 different PFGE pulsotypes. The discriminatory index for a sporadic strain was 0.862 for PFGE and 0.999 for MLVA. MLVA confirmed that a foodborne outbreak in the city of Kotka, Finland in 2003 had been caused by a multiresistant YE 4/O:3 strain that was distinctly different from those of epidemiologically unrelated strains with an identical PFGE pulsotype. The multiresistance of Y. enterocolitica strains (19% of the sporadic strains) correlated significantly (p = 0.002) with travel abroad. All of the multiresistant Y. enterocolitica strains belonged to four PFGE pulsotypes that did not contain any susceptible strains. Resistance to nalidixic acid was related to changes in codons 83 or 87 that stemmed from mutations in the gyrA gene. The conjugation experiments demonstrated that resistance to CHL, STR, and SUL was carried by a conjugative plasmid. CONCLUSIONS: MLVA using six loci had better discriminatory power than PFGE with the NotI enzyme. MLVA was also a less labor-intensive method than PFGE and the results were easier to analyze. The conjugation experiments demonstrated that a resistance plasmid can easily be transferred between Y. enterocolitica strains. Antimicrobial multiresistance of Y. enterocolitica strains was significantly associated with travel abroad.

Genotype diversity of group A rotavirus strains in children with acute diarrhea in urban Burkina Faso, 2008-2010.

In this study, the diversity of G and P genotypes of rotavirus strains in Burkinabe children were examined. Between November 2008 and February 2010, 447 stool samples were collected from children <5 years of age with acute diarrhea visiting hospital in Ouagadougou. Group A rotavirus was previously detected in 151/447 (33.8%) of the samples tested by an immunochromatographic test and these samples were now tested further for rotavirus G and P genotypes by RT-PCR. Of these, the rotavirus type genes were amplified by RT-PCR for 140/151 (92.7%) samples and G and P genotypes were successfully determined for 81 (57.9%) and 130 (92.9%) samples, respectively. The most prevalent G genotypes were G1, 34/140 (24.3%), and G9, 21/140 (15%), while the predominant P genotypes were P[6], 56/140 (40%), and P[8], 54/140 (38.6%). Among the single infections, 63/140 (45%), the predominant G/P combinations were: G1P[8] (33%), G9P[8] (29%), and G2P[6] (14%). The unusual strains G1P[9] (3%), G12P[6] (3%), G10P[6] (2%), and G2P[8] (2%) were also detected. In a high number of strains 61/140 (43.6%), the G genotype could not be determined and mixed infections were determined in 17/140 (12.1%) of strains identified. This study highlights the high diversity and presence of unusual rotavirus strains in children in Burkina Faso.

Shiga toxin-producing Escherichia coli O100:H⁻: stx2e in drinking water contaminated by waste water in Finland.

In November 2007, 450 m(3) of treated wastewater leaked into the drinking water distribution system contaminating the drinking water of over 10,000 inhabitants of Nokia, Southern Finland. Nearly 1,000 people visited the health centre because of gastroenteritis during the following 5 weeks. A wide range of enteric pathogens was found in the patients. The authors used the 16-plex PCR to investigate whether the five major diarrheagenic Escherichia coli pathotypes (EPEC, ETEC, STEC, EIEC or EAEC) were present in the contaminated drinking water and in the patients' stool samples. The contaminated drinking water was positive for genes characteristic of various E. coli pathotypes: pic, invE, hlyA, ent, escV, eae, aggR, stx(2) , estIa and astA. These genes, except stx(2), hlyA and invE, were also detected in the stool samples of the patients linked to this outbreak. A sorbitol positive, streptomycin resistant STEC strain was isolated from the drinking water, and belonged to the serotype O100:H(-), produced Stx2 toxin (titre 1:8 by reversed-passive latex agglutination method), and carried the genes stx(2e), estIa and irp2.

The ail gene is present in some Yersinia enterocolitica biotype 1A strains.

One chromosomal virulence marker of Yersinia is the gene ail, which encodes Ail, an outer membrane protein that promotes attachment and invasion. A high correlation has been found between the ail gene and the virulence of Yersinia. Here, we report two Yersinia enterocolitica biotype 1A strains that are usually nonpathogenic and carry the ail gene. The ail gene sequences of biotype 1A strains displayed similarity to the bioserotype 1B/O:8 strain 8081. The finding suggests that ail-based detection methods for Y. enterocolitica alone are insufficient to detect real pathogenic strains.