This study aimed to determine the distribution of plasmid-mediated quinolone resistance (PMQR) and class 1, and 2 integrons in ciprofloxacin-resistant Enterobacterales isolated at the Aristide le Dantec university teaching hospital in Dakar, Senegal. In the E. coli strains, the most prevalent PMQR gene was aac(6’)-Ib-cr [59/73; 80.8

Senegal has witnessed the emergence and spread of carbapenem-resistant Enterobacterales (CRE), which often cause deadly infections. Accordingly, this study aimed to determine the antimicrobial susceptibility and prevalence of carbapenemases, as well as to perform a whole-genome sequence analysis of clinical CRE isolates from a university hospital in Dakar, Senegal. MALDI-TOF MS and VITEK2 systems were used for bacterial identification and antimicrobial susceptibility testing (AST). Carbapenemase- and cephalosporinase-encoding genes were screened using simplex end-point polymerase chain reaction. Whole-genome sequencing (WGS) was performed using the Illumina MiSeq platform. The CRE isolates were resistant to almost all the 34 antimicrobials tested. Nevertheless, colistin and amikacin remained active, with susceptibility rates of 96% and 71%, respectively. Only the carbapenemase genes blaOXA-48 (53.8%; 15/28) and blaNDM (35.7%; 10/28) and the cephalosporinase gene blaCMY-1 (25%; 7/28) were identified. In this context, two extensively drug-resistant Enterobacter hormaechei isolates were subjected to WGS analysis. These isolates were assigned as sequence type (ST) 182 and carried several genes related to antimicrobial resistance (AMR), metal tolerance, and virulence. An IncL/M plasmid with 61,054 bp in length was identified as carrying the blaOXA-48 gene, whereas an IncFIB(pECLA)/IncFII(pECLA)/IncX3 mutireplicon plasmid with 217,745 bp in length was detected as harboring the blaNDM-5 gene and other genes related to AMR and metal tolerance. Our study presents the first landscape of clinical CRE circulating in Senegal, along with additional genomic analysis of E. hormaechei ST182 strains, which could be useful for mitigating the burden associated with CRE in this country.IMPORTANCEThe investigation of global critical priority CRE isolates has become crucial to reduce morbidity and mortality associated with AMR. This study revealed that colistin and amikacin can be considered good alternatives for treating CRE-associated infections in Dakar. In addition, the genomic approach revealed that the CRE isolates carried both a wide resistome and virulome. Moreover, the abundance of horizontal gene transfer regions in the genomes suggests the great implications of mobile genetic elements in the spread of AMR in Dakar. Furthermore, this study reported the complete sequences of chromosomes and blaOXA-48 and blaNDM-5-carrying plasmids. Our findings are of great importance because complete genome sequences are still rarely characterized in the West African region. Finally, this study highlights the importance of strengthening genomic surveillance of CRE in sub-Saharan African countries to mitigate the burden associated with these pathogens.

Senegal has witnessed the emergence and spread of carbapenem-resistant Enterobacterales (CRE), which often cause deadly infections. Accordingly, this study aimed to determine the antimicrobial susceptibility and prevalence of carbapenemases, as well as to perform a whole-genome sequence analysis of clinical CRE isolates from a university hospital in Dakar, Senegal. MALDI-TOF MS and VITEK2 systems were used for bacterial identification and antimicrobial susceptibility testing (AST). Carbapenemase- and cephalosporinase-encoding genes were screened using simplex end-point polymerase chain reaction. Whole-genome sequencing (WGS) was performed using the Illumina MiSeq platform. The CRE isolates were resistant to almost all the 34 antimicrobials tested. Nevertheless, colistin and amikacin remained active, with susceptibility rates of 96% and 71%, respectively. Only the carbapenemase genes bla OXA-48 (53.8%; 15/28) and bla NDM (35.7%; 10/28) and the cephalosporinase gene bla CMY-1 (25%; 7/28) were identified. In this context, two extensively drug-resistant Enterobacter hormaechei isolates were subjected to WGS analysis. These isolates were assigned as sequence type (ST) 182 and carried several genes related to antimicrobial resistance (AMR), metal tolerance, and virulence. An IncL/M plasmid with 61,054 bp in length was identified as carrying the bla OXA-48 gene, whereas an IncFIB(pECLA)/IncFII(pECLA)/IncX3 mutireplicon plasmid with 217,745 bp in length was detected as harboring the bla NDM-5 gene and other genes related to AMR and metal tolerance. Our study presents the first landscape of clinical CRE circulating in Senegal, along with additional genomic analysis of E. hormaechei ST182 strains, which could be useful for mitigating the burden associated with CRE in this country. The investigation of global critical priority CRE isolates has become crucial to reduce morbidity and mortality associated with AMR. This study revealed that colistin and amikacin can be considered good alternatives for treating CRE-associated infections in Dakar. In addition, the genomic approach revealed that the CRE isolates carried both a wide resistome and virulome. Moreover, the abundance of horizontal gene transfer regions in the genomes suggests the great implications of mobile genetic elements in the spread of AMR in Dakar. Furthermore, this study reported the complete sequences of chromosomes and bla OXA-48 and bla NDM-5 -carrying plasmids. Our findings are of great importance because complete genome sequences are still rarely characterized in the West African region. Finally, this study highlights the importance of strengthening genomic surveillance of CRE in sub-Saharan African countries to mitigate the burden associated with these pathogens.

Abstract Background Bacterial virulence is a key factor determining the outcome of each bacterial infection and virulent bacteria are often associated to high-risk infections. Extraintestinal pathogenic Escherichia coli (ExPEC) is the most implicated bacterium in human bacterial infections and its virulence factors are classified into five categories: adhesins, toxins, iron capture systems, protectins and invasins. Furthermore, bacterial biofilms are the main cause of hospital-acquired infections like urinary catheter-associated infections, valve endocarditis, Otitis and cystic fibrosis.Results For the sixteen virulence genes sought by standard polymerase chain reaction (PCR), all the 78 ExPECs isolates carried at least four virulence genes. Following prevalences of virulence genes were reported: adhesins genes fimH (98.7%), mrkD (98.7%), papC (46.2%), afaC (9%), sfa / focDE (1.3%); iron acquisition systems genes entB (98.7%), fepA (98.7%), ybtS (93.6%), fyuA (91%), iucA (91%), iucB (91%), iutA (34.6), iroB (6.4%), iroN (6.4%) and toxins genes hlyA (10.3%), cnf (1 & 2) (10.3%). Seventy-five out of 78 isolates (96.2%) carried at least the combination of two adhesins genes and two iron capture systems genes whereas 8 out of 78 (10.3%) harbored the combination of (adhesins genes + iron acquisition systems genes + toxins genes). Among the 78 strains studied, one hospital-acquired strain isolated from urine harbored 15 virulence genes out of 16 sought. The evaluation of biofilm-formation capacity revealed that all (29/29) hospital-acquired isolates were biofilm producers with (6/29; 20.7%) strong biofilm producers, (15/29; 51.7%) moderate biofilm producers and (8/29; 27.6%) weak biofilm producers. Isolates carrying papC had greater biofilm formation capacity than those not carrying papC (p < 0.001).Conclusions Most of our strains had moderate biofilm-formation capacity and carried an average of 9 virulence genes out of 16 sought. These eight strains carrying a combination of genes (adhesins + iron acquisition systems genes + toxins genes) may be hypervirulent isolates. Additional studies may confirm this. The deepening of this kind of study on bacterial virulence and hospital bacterial biofilms could lead to the improvement of infections investigation, prevention and therapeutic protocols.

The eradication of neonatal Group B Streptococcus (GBS) infections, considered as a major public health priority, necessarily requires a mastery of the data on vaginal carriage in pregnant women. The aims of this study were to determine the prevalence of vaginal carriage of GBS in pregnant women, antibiotic susceptibility, and associated risk factors. This was a cross-sectional, descriptive study conducted over a period of 9 months (July 2020 to March 2021) in pregnant women between 34 and 38 weeks of gestation (WG) followed at the Nabil Choucair health center in Dakar. Identification and antibiotic susceptibility of GBS isolates were performed on the Vitek 2 from vaginal swabs cultured on Granada medium. Demographic and obstetric interview data were collected and analyzed on SPSS (version 25). The level of significance for all statistical tests was set at P < .05. The search of GBS vaginal carriage had involved 279 women aged 16 to 46 years, with a median pregnancy age of 34 (34-37) weeks' gestation. GBS was found in 43 women, for a vaginal carriage rate of 15.4%. In 27.9% (12/43) of volunteers screened, this carriage was monomicrobial, while in 72.1% (31/43) of women, GBS was associated with other pathogens such as Candida spp. (60.5%), Trichomonas vaginalis (2.3%), Gardnerella vaginalis (34.9%) and/or Mobiluncus spp. (11.6%). The level of resistance was 27.9% (12/43) for penicillin G, 53.5% (23/43) for erythromycin, 25.6% (11/43) for clindamycin and 100% for tetracycline. However, the strains had retained fully susceptible to vancomycin and teicoplanin. The main risk factor associated with maternal GBS carriage were ectocervical inflammation associated with contact bleeding (OR = 3.55; P = .005). The high rate of maternal vaginal GBS carriage and the levels of resistance to the various antibiotics tested confirm the importance of continuous GBS surveillance in our resource-limited countries.

Background: The emergence and spread of carbapenem-resistant Acinetobacter baumannii are critical in hospitals, particularly in intensive care units (ICUs), which represents a public health concern worldwide. In this study, we investigated the molecular epidemiology of multi-drug resistant A. baumannii (MDR-AB) in Dakar, Senegal. Methods. The A. baumannii was isolated from Eosin Methylene Blue Agar culture and identified using API 20NE strip test and MALDI-TOF. The antimicrobial susceptibility testing was performed using the disk diffusion method. Simplex and multiplex-polymerase chain reactions with appropriate primers were used to detect and sequence the following β-lactamase genes: Two class D carbapenem hydrolyzing oxacillinases (blaOXA-51 and blaOXA-23), three class B metallo-β-lactamase genes (blaIMP, blaVIM and blaNDM), and five class A β-lactamase genes (blaPER, blaSHV, blaVEB, blaTEM, and blaGES). Results: A total of 29 strains of MDR-AB were isolated from patients hospitalized at Aristide Le Dantec University teaching hospital in Dakar, Senegal. Among the 29 MDR-AB strains isolated, 11 (37.9%) were isolated from ICUs, 5 (17.2%) from pediatric surgery, and 13 (44.8%) from other departments. The MDR strains were isolated from urine and pus samples with 12 (41.4%) and 9 (31.0%), respectively. All isolates were positive for the A. baumannii specific gene blaOXA-51. The blaOXA-51 and blaOXA-23 genes coexisted in 26 (89.65%) of the strains. The blaIMP and blaVIM genes were not detected among the selected strains. Moreover1 (3.4%) strain elicited the gene coding for metallo-β-lactamase NDM-1. 2 (6.9%) isolates turned out to produce the penicillinase TEM-2. Conclusions: Carbapenem resistance in Senegalese strains of A. baumannii is predominantly due to the worldwide disseminated gene blaOXA-23, with a subset of strains due to NDM-1 and TEM-2. Systemic molecular surveillance network should be established for further efficient monitoring of MDR strains in Senegal.

Background: Bacterial vaginosis (BV) is associated with a high risk of complications, particularly during pregnancy. Objectives: The aims of this study were to determine the prevalence of BV in pregnant women in Dakar, Senegal, and to identify its associated risk factors. Patients and Methods: This prospective cross-sectional study was conducted from July 2020 to March 2021 in pregnant women with 34 to 38 weeks of gestation and seen for their routine prenatal consultation at the Nabil Choucair health center in Dakar, Senegal. Vaginal swabs were taken and examined using the Nugent scoring system for the diagnosis of the BV. Data analysis on SPSS (version 25) was done using the chi-square test to measure the strength of association. A value of p ≤ 0.05 was considered statistically significant. Results: BV was found in 28.0% (112/400) of the screened women with a median age 24 (21-29) years. A pH >4.5 predictive of BV was found in 80.4% (90/112) of samples. BV was associated with vaginal candidiasis in 49.1% (55/112) while Trichomonas vaginitis was found in 1.8% (02/112). In 80.4% (90/112) of affected women, BV was caused by Gardnerella vaginalis. Mobiluncus spp. was found in 19.6% (22/112) pregnant women in association with G. vaginalis. Symptoms characterized by episodes of pruritus, pelvic pain, burning and/or dyspareunia were seen among 59.8% (67/112) of these women with BV while 40.1% (45/112) of them reported no symptoms. Conclusion: In view of these results and in order to reduce gestational complications and adverse outcomes in the newborn, screening for BV in pregnant women should be favored in developing countries.

Extra-Intestinal Pathogenic Escherichia coli (ExPEC) is a predominant Gram-negative bacterial pathogen and is responsible of several diseases including Urinary Tract Infections (UTI), nosocomial pneumonia, and neonatal meningitis. ExPEC isolates are often multidrug resistant and clones producing Extended-Spectrum Beta-Lactamases (ESBL) are increasingly reported all over the world. Seventy-eight clinical ExPEC strains (49 Community-Acquired (CA) and 29 Hospital-Acquired (HA)) were selected for this study. The majority was from UTIs (n=51), while the rest (n=27) was from pus, sputum, bronchial fluid and vaginal samples (non-uropathogenic ExPEC). Antibiotic susceptibility testing was performed using the Kirby-Bauer disc diffusion method. Standard polymerase chain reaction was used to screen major ESBL genes (blaCTX-M, blaOXA-1, blaTEM, blaSHV) and blaCTX-M variants (blaCTX-M-1, blaCTX-M-9, blaCTX-M-15, blaCTX-M-25). All the tested isolates were resistant to ampicillin, ticarcillin, amoxicillin/ clavulanic acid combination, cefalotin, cefotaxime, ceftazidime, cefepime and aztreonam, but showed a high susceptibityto fosfomycin (98.7%, n = 77), ertapenem (96.2%, n = 75), and imipenem (100%). Moreover, isolates harbored at least one ESBL gene, including blaCTX-M (98.7%), blaOXA-1 (78.2%), blaTEM (44.9%) and blaSHV (3.8%). The CTX-M variants were also found with the predominance of blaCTX-M-1 (89.7%) and blaCTX-M-15 (89.7%) followed by blaCTX-M-9 (11.5%). Despite the resistance to most of the tested antibiotics, ExPEC isolates showed fortunately good susceptibility to fosfomycin and carbapenems. blaCTX-M1, blaCTX-M15 and blaOXA-1 seem to be E.coli major ESBL genes circulating in Senegal. The high level of resistance to antimicrobials observed stresses the need of establishing an epidemiological surveillance of antimicrobial resistance in both community and hospital settings.

Extended spectrum β-lactamase (ESBL) and carbapenemase-producing Enterobacteriaceae (CPE) have been increasingly reported worldwide. The objective of this study is to determine the prevalence of these multidrug-resistant strains in a major university teaching hospital in Dakar, Senegal. A total of 1205 Enterobacteriaceae stains were tested for ESBL and carbapenemase production. Antibiotics susceptibility test was performed with disk diffusion method. ESBL was detected using a double-disk synergy method. Carbapenemase production was detected with ertapenem 10 µg disk charge. The overall prevalence of ESBL- and carbapenemase-producing Enterobacteriaceae was 26.2 (316/1205) and 5.1% (62/1205), respectively. Interestingly, 3.8% of these pathogens were both ESBL-carbapenemase producers. Among the Enterobacteriaceae ESBL positive, Escherichia coli (45.2%, 143/316), Klebsiella pneumoniae (26.3%, 83/316), Enterobacter cloacae (12.7%, 40/316), and Proteus vulgaris (9.2%, 29/316) were the most prevalent. These strains were mainly isolated from urine (56.6%) and pus (22.7%) specimen. The most prevalent CPEs were E. coli (45.2%, 28/62), K. pneumoniae (27.4%, 17/62), and E. cloacae (16.1%, 10/62), particularly isolated from urine (58%) and pus (19.3%). The majority of these MDR strains were isolated from patients hospitalized in urology (32.4%), surgery (27.7%), internal medicine (18.5%), and intensive care units (10%). ESBL-producing Enterobacteriaceae remain highly susceptible to fosfomycin (94.1%), amikacin (92.5%) and ertapenem (88.6%), while carbapenemase producers were fully susceptible to amikacin (100%), and to a lesser extent, fosfomycin (66.7%) and colistin (60%). This study revealed increasing prevalence of ESBL- and carbapenemase-producing Enterobacteriaceae with limited therapeutic options, suggesting a need for continuous multi-drug resistant (MDR) surveillance patterns particularly in hospital settings. Key words: Extended spectrum β-lactamase, carbapenemase, Enterobacteriaceae.

Introduction: the challenge facing developing countries is the availability of methods for rapid and accurate diagnosis of tuberculosis. Some molecular techniques offer this advantage, so we used GeneXpert MTB / RIF test in the diagnosis of extra-pulmonary tuberculosis to evaluate its performance compared with conventional methods. Methods: between 2010 and 2015, 544 extrapulmonary clinical specimens were collected and analyzed by microscopy, culture and GeneXpert. The evaluation of antitubercular susceptibility testing was performed using the MGIT 960 system. Genotype MTBDRplus was used to confirm the cases of rifampicin resistance detected by the GX system. Results: the study population included 544 patients, 55.15% men and 44.85% women. Patients age ranged from 1-92 years with the majority in the 18-45 age group. The sensitivity and the overall specificity of microscopy was 43.86% and 98.36%, 94.74% and 97.95% for GeneXpert r respectively ( 95% CI). There were two discrepant rifampicin-resistant results between GeneXpert test and phenotypic method. Among these cases MTBDRplus test results showed 100% agreement with those of the MGIT 960. Conclusion: this study shows that the GeneXpert test exhibits high sensitivity for routine diagnosis of extra-pulmonary tuberculosis and should be used instead of microscopy. The cases of rifampicin resistance detected by GeneXpert should be confirmed by other molecular testing methods before initiating treatment.

INTRODUCTION: le défi des pays en voie de développement est la disponibilité de méthodes de diagnostic rapide et précis pour le management de la tuberculose. Des techniques moléculaires offrent cet avantage et nous avons utilisé le test GeneXpert MTB/RIF dans le diagnostic de la tuberculose extra-pulmonaire pour évaluer sa performance par rapport aux méthodes conventionnelles. METHODES: entre 2010 et 2015, 544 échantillons cliniques extra-pulmonaires ont été recueillis et traitées par la microscopie, la culture et le GeneXpert. L'étude de la sensibilité aux antituberculeux a été effectué avec le MGIT 960. Le Génotype MTBDRplus a été utilisé pour confirmer les cas de résistance à la rifampicine détectés par le système GX. RESULTATS: la population d'étude de 544 patients incluait 55,15% d'hommes et 44,85% de femmes. L'âge des patients variait entre 1 à 92 avec la majorité dans le groupe d'âge 18-45 ans. La sensibilité et la spécificité globale de la microscopie étaient de 43,86% et 98,36%, et pour le GeneXpert, 94,74% et 97,95% respectivement avec 95% IC. Deux résultats de résistance à la rifampicine discordants ont été trouvées entre le test GeneXpert et la méthode phénotypique. Les résultats du test MTBDRplus ont montré une concordance de 100% avec ceux du MGIT 960 pour les cas discordants de résistance à la rifampicine. CONCLUSION: cette étude a montré que le test GeneXpert a une plus grande sensibilité pour le diagnostic de routine de la tuberculose extra-pulmonaire et devrait être utilisé à la place de la microscopie. Les cas de résistance à la rifampicine détectés par le GeneXpert doivent être confirmés par d'autres tests moléculaires avant d'initier un traitement.

Background: Chemoprophylaxis of contacts of infectious tuberculosis (TB) cases is recommended for TB control, particularly in endemic countries, but is hampered by the difficulty to diagnose latent TB infection (LTBI), classically assessed through response to the Tuberculin Skin Test (TST).Interferon-gamma release assays (IGRA) are proposed new tools to diagnose LTBI, but there are limited data on their ability to predict the development of active TB disease.To address this, we investigated the response to TST and IGRA in household contacts of infectious TB cases in a TB high-burden country and the potential correlation with development of TB.Methodology/Principal Findings: Prospective household contacts study conducted in two health centres in Dakar, Senegal.A total of 2679 household contacts of 206 newly detected smear and/or culture positive index TB cases aged 18 years or greater were identified A TST was performed in each contact and an ESAT6/CFP10 ELISPOT assay performed in a random sample of those.Contacts were followed-up for 24 months.TB was diagnosed in 52 contacts, an incidence rate of 9.27/1000 person-years.In univariable analysis, the presence of positive TST ($10 mm) and ELISPOT (.32 SFC/million PBMC) responses at baseline were associated with active TB during follow-up: Rate Ratio [RR] = 2.32 (95%CI:1.12-4.84)and RR = 2.09 (95%CI:0.83-5.31),respectively.After adjustment for age, sex and proximity to index case, adjusted RRs were 1.51 (95%CI:0.71-3.19)and 1.98 (95%CI:0.77-5.09),respectively.Restricting analysis to the 40 microbiologically confirmed cases, the adjusted RR for positive ELISPOT was 3.61 (95%CI:1.03-12.65).The median ELISPOT response in contacts who developed TB was 5-fold greater than in those who did not develop TB (p = 0.02).Conclusions/Significance: TST and IGRAs are markers of a contact of the immune system with tubercle bacilli.In a TB endemic area, a high ELISPOT response may reflect increased bacterial replication that may subsequently be associated with development of TB disease and may have a prognostic value.Further longitudinal data are needed to assess whether IGRAs are reliable markers to be used for targeting chemoprophylaxis.

BACKGROUND:Detection and treatment of latent TB infection (LTBI) in HIV infected individuals is strongly recommended to decrease morbidity and mortality in countries with high levels of HIV.OBJECTIVE:To assess the validity of a newly developed in-house ELISPOT interferon-gamma release assay (IGRA) for the detection of LTBI amongst HIV infected individuals, in comparison with the Tuberculin Skin Test (TST).METHODOLOGY/PRINCIPAL FINDINGS:ESAT6/CFP10 (EC) ELISPOT assays were performed, together with a TST, in 285 HIV infected individuals recruited in HIV clinics in Dakar, Senegal, who had no signs of active TB at time of enrolment. Thirty eight of the subjects (13.3%) failed to respond to PHA stimulation and were excluded from the analysis. In the 247 remaining patients, response to PHA did not vary according to CD4 cell count categories (p = 0.51). EC ELISPOT was positive in 125 (50.6%) subjects, while 53 (21.5%) had a positive TST. Concordance between EC ELISPOT and TST was observed in 151 patients (61.1%) (kappa = 0.23). The proportion of subjects with a positive response to the EC ELISPOT assay decreased with declining CD4 counts (p trend = 0.001), but were consistently higher than the proportion of TST responders. In multivariate analysis, the risk of being EC-ELISPOT positive in HIV infected individuals was associated with age, CD4 count and HIV-1 strain.CONCLUSION:Our study indicates that IGRAs using M. tuberculosis specific antigens are likely to retain their validity for the diagnosis of LTBI among HIV positive individuals, but may be impaired by T-cell anergy in severely immuno-suppressed individuals.

Flow cytometry is an accurate but expensive method to determine absolute CD4 cell counts. We compared different methods to measure absolute CD4 counts in blood samples from HIV-infected and uninfected subjects using a research/clinical flow cytometer (FACScan); a dedicated clinical instrument (FACSCount); and a volumetric, mobile, open-system flow cytometer equipped with 3 fluorescence and 2 light scatter detectors (Cyflow SL blue). The FACScan and Cyflow were used as single-platform instruments, but they differ in running cost, which is a central factor for resource-poor settings. Direct volumetric and bead-based CD4 measurements on the Cyflow were compared with 2 bead-based single-platform CD4 measurements on the FACSCount and on FACScan (TruCount) in "Le Dantec" Hospital, Dakar, Senegal, using whole blood samples from 102 HIV+ and 28 HIV- subjects. The agreement between the various measurement methods was evaluated by Bland-Altman analysis. Volumetric CD4 measurements on the Cyflow using a no-lyse-no-wash (NLNW) procedure and a lyse-no-wash (LNW) procedure correlated well with each other (R2 = 0.98) and with CD4 measurements on the FACSCount (R2 = 0.97) and FACScan (R2 = 0.97), respectively. Red blood cell lysis had no negative effect on the accuracy of absolute CD4 counting on the Cyflow. An excellent correlation was observed between bead-based CD4 measurements on the Cyflow and CD4 measurements on the FACSCount (R2 = 0.99) and FACScan (R2 = 0.99). Rigid internal and external quality control monitoring and adequate training of technicians were considered essential to generate accurate volumetric CD4 measurements on the Cyflow.