Simultaneous detection of Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae by quantitative PCR from CSF samples with negative culture in Morocco.

Bacteriological cultures from cerebrospinal fluids (CSF) have less sensitivity and specificity compared to quantitative PCR (RT-PCR), and multiple facts still conduct to the increase of negative culture. The aims of this study are to determine the molecular epidemiology and the simultaneous detection of bacterial meningitis in Morocco by using RT-PCR and compared this molecular approach with culture method to improve the etiological diagnosis of meningitis. The CSFs were collected over one-year period in 2018 in different hospitals covering all regions of the Kingdom of Morocco, from patients with suspected meningitis. The results showed the confirmation rate per culture recorded a rate of 33% and the RT-PCR of 70%. Molecular epidemiology is predominant of Neisseria meningitidis followed by Streptococcus pneumoniae and a dramatic reduction in meningitis due to Haemophilus influenzae following the introduction of conjugate vaccine in 2007. Also, the epidemiological profile shows a sex ratio M/F of 1.4 and a median age of 2 years. The national distribution showed a predominant of meningococcal disease followed by pneumococcal disease, especially a dominance of N. meningitidis over S. pneumoniae in two regions and a slight predominance of S. pneumoniae in the other two regions over N. meningitidis. Our research shows that culture in our country has less sensitivity and specificity than RT-PCR in diagnosis of bacterial meningitis and that molecular biology technique at bacteriology laboratories is desirable for diagnosis, early management of meningitis cases and in the context of the surveillance of meningitis in Morocco in parallel with culture.

The novel biphasic medium for transport, culture and conservation at an ambient temperature of Neisseria meningitidis, Streptococcus pneumoniae and Haemophilus influenzae.

Bacterial meningitis remains a very important disease worldwide, and the major causative pathogens were Neisseria meningitidis (N. meningitidis), Streptococcus pneumoniae (S. pneumoniae) and Haemophilus influenzae (H. influenzae). In our context, the technical difficulties encountered in the routine practice were associated with the fragility of these bacteria, the high rates of negative culture and the demanding transport conditions. That's why the need to look for a solution to its technical problems and to propose a new proper solution with the local situation. The aim of this study was to develop, perform and evaluate a novel biphasic medium used for the transport, culture and conservation at an ambient temperature of N. meningitidis, S. pneumoniae and H. influenzae. The results showed that this biphasic medium provided more, novels and easy nutriments through the addition of liquid phase and solid phase medium and it was found to be conducive to the growth and conservation of N. meningitidis, S. pneumoniae and H. influenzae at an ambient temperature of a minimum of 40 days. And the ingredients used in the medium are readily available at a low cost as well as the components prepared in large quantities, they could be stored at + 4 ± 1 °C for 2 years without significantly altering their growth and conservation supporting their potential. The survival and recovery for the fastidious bacteria on the biphasic medium and the other media used for comparison in this study were significantly different (P < 0.05). In addition, the Sensitivity, Specificity, Positive and Negative Predictive Value of biphasic medium showed highest among the three bacteria at least 40 days of storage at room temperature in this study. In conclusion, we found the biphasic medium to be low cost and suitable for previously mentioned bacteria from suspected meningitis patients, offering an optimal condition and an increase in the viability of the isolates at ambient temperature. And it was concluded that this biphasic medium could be used as a technical solution in laboratories for the management of meningitis.

Epidemiology and antibiotic resistance profile of bacterial meningitis in Morocco from 2015 to 2018.

Over a 4-year study period from 2015 to 2018, altogether 183 isolates of bacterial meningitis were collected from 12 hospitals covering the entire Moroccan territory. Neisseria meningitidis represented 58.5%, Streptococcus pneumoniae 35.5%, and Haemophilus influenzae type b 6%. H. influenzae type b mainly affected 5-year-olds and unvaccinated adults. N. meningitidis serogroup B represented 90.7% followed by serogroup W135 with 6.5%. Decreased susceptibility to penicillin G (DSPG) for all isolates accounted for 15.7%, with 11.6% being resistant to penicillin G (PG) and 4.1% decreased susceptibility. Cumulative results of all strains showed 2.7% decreased susceptibility to amoxicillin and 3.3% resistant, 2.2% of isolates were resistant to third-generation cephalosporin and 2.2% were decreased susceptible, 5.5% were resistant to chloramphenicol and 2.7% were resistant to rifampin. The frequency of DSPG observed in our study is more common in S. pneumoniae than in N. meningitidis (P < 0.05). These isolates have been found to be highly susceptible to antibiotics used for treatment and prophylaxis chemotherapy and the observed resistance remains rare. The impact of introduction of conjugate vaccines against H. influenzae type b and S. pneumoniae (PCVs) is an advantage in reducing meningitis cases due to these two species.

Antimicrobial resistance monitoring in Neisseria gonorrhoeae and strategic use of funds from the Global Fund to set up a systematic Moroccan gonococcal antimicrobial surveillance programme.

OBJECTIVES: The aims of this study were to assess antimicrobial resistance in Neisseria gonorrhoeae infections and update the treatment in the national guidelines for the syndromic management of sexually transmitted infections in Morocco. METHODS: 171 men complaining of urethral discharge were recruited from basic health services during 2009. Urethral swab samples were collected and N gonorrhoeae identification was performed by culture. Antimicrobial susceptibility testing was performed using the Etest method and the antimicrobial agents tested were ciprofloxacin, penicillin, spectinomycin, tetracycline, ceftriaxone and cefixime. RESULTS: A total of 72 isolates were examined. Significant resistance to tetracycline (92.8%) and ciprofloxacin (86.8%), which was used as first-line treatment in gonococcal infections, was noted. No resistance to spectinomycin, ceftriaxone or cefixime was detected in all the isolates. CONCLUSIONS: Following these results the Ministry of Health of Morocco replaced ciprofloxacin and introduced ceftriaxone 250 mg as a single dose in the treatment of gonococcal infections. Using funds from the Global Fund to Fight AIDS, Tuberculosis and Malaria (the Global Fund), a surveillance programme was set up for antimicrobial resistance testing in N gonorrhoeae.

Assessment of Chlamydia trachomatis, Neisseria gonorrhoeae and Trichomonas vaginalis prevalence using a molecular Point of Care: Findings from a respondent driven sampling study among MSM.

Objectives: Due to the limited data available within the Moroccan context, the aim of the study was therefore to estimate the prevalence of Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG) and Trichomonas vaginalis (TV) infection and co-infection among men who have sex with men (MSM) as well as to update the behavioral indicators for this population. Methods: During the period of November 2020 to January 2021, 275 and 303 MSM in Agadir and Fes respectively, were recruited by using respondent-driven sampling protocol (RDS). Eligibility criteria for participants included men identified as having anal sex with another man in the last 6 months, aged 18 years or older and residing in either Agadir or Fes, regardless of their nationality, for the past 6 months.Anal swabs were collected from 445 respondents for molecular investigation of CT, NG, and TV. GeneXpert (Cepheid, USA) was used to test all samples. A survey on the socio-demographic, and risk behavior was then administered to participants. Results: Most MSM subjects were identified as being young, and homosexual. CT prevalence was 11.3% (95%CI, 7.2 to 15.4) and 12.5% (95%CI, 7.5 to 17.5) in Agadir and Fes respectively; NG was 13.3% (95%CI, 8.5 to 18.1) in Agadir and 5.5% (95%CI, 1.9 to 9.2) in Fes. Meanwhile, TV prevalence was 0.4% (95%CI, 0 to 1.1) in Agadir and 0.2% (95%CI, -0.2 to 0.6) in Fes. A CT/NG co-infection was found in 4.5% (95%CI, 3.5 to 5.9) of cases in Agadir and 2.7% (95%CI, 1.9 to 3.9), in Fes. Conclusion: It follows that a regular risk assessment and Sexually Transmitted Infectious (STIs) screening should be administered in these two cities as part of a global strategy to enhance the sexual health of the key populations in question.

[Microbiological quality of wheat flour consumed in Morocco]. / Qualité microbiologique des farines de blé consommées au Maroc.

Cereal products (soft and hard wheat) are a basic staple food in the Moroccan diet. A total of 60 samples of two types of wheat flours used for human consumption were collected; 30 samples among this collection were obtained from various households using Moroccan varieties of wheat produced in traditional flour mills. The rest of the samples were purchased from retail wheat flour sources in the Rabat and Sale city markets. Standard plate counts (SPC), total and faecal coliforms, Clostridium, Salmonella spp., Shigella spp., Staphylococcus aureus, Listeria monocytogenes, yeast, lactic acid bacteria, and molds, were carried out to assess the microbiological quality of wheat flour. Microbiological interpretation of the criteria was performed according to standards implemented by the Codex Alimentarius Commission. Most frequent counts, in traditional and industrial wheat flour, were total aerobic mesophilic bacteria with an average 4 × 104 and 2.5 × 104 cfu/g, respectively. The results showed higher coliform and fungi counts in house than in commercial samples. Pathogenic flora as Salmonella spp., Shigella spp., S. aureus, L. monocytogenes, and Clostridium were not detected in all investigated samples. Bacterial strains isolated from both flours belong to the following genera: Enterobacter spp., Serratia spp., Klebsiella spp., Pantoea spp., Leclercia spp., Proteus spp. The most frequent genus of the investigated isolates was Aspergillus (81 %). Microbial counts were lower than the limit laid down in the Codex Alimentarius, attributing to these flours a satisfactory microbiological quality.

Evaluation of the Knowledge and Attitudes of Dental Students toward Occupational Blood Exposure Accidents at the End of the Dental Training Program.

AIMS AND OBJECTIVES: This study was conducted to assess knowledge, attitudes, and behavior regarding occupational blood exposure accidents (OBEA) among dental students at the end of the whole dental education program. MATERIALS AND METHODS: The survey was performed using a student self-administered questionnaire during July 2015 in the dental teaching hospital in Rabat. It was conducted on 117 dental students registered in the 5th year. The results were analyzed by SPSS Software, Version 13.0. Univariate analysis was performed by descriptive statistics, and bivariate analysis was used to identify correlations between different variables. RESULTS: Eighty-three students answered the questionnaire (participation rate: 70%). Most participants had incomplete knowledge about the management and the risks of OBEA. Thirty-five participants (42%) experienced at least one occupational exposure to patients' blood. Needle recapping has been the main cause of such injuries (41%). Most accidents occurred, respectively, in the oral surgery (51%) and the restorative dentistry (17%) departments. According to many participants, they had received insufficient education concerning this topic. CONCLUSIONS: More effective education on prevention and management of OBEA is highly recommended to reduce the prevalence of such injuries. Dental schools should set up a local management unit for following and supporting the exposed students.

Analysis of isoniazid and rifampicin resistance in Mycobacterium tuberculosis isolates in Morocco using GenoType® MTBDRplus assay.

OBJECTIVES: The number of multidrug-resistant tuberculosis (MDR-TB) cases is rising worldwide. The present investigation aimed to evaluate, using the GenoType® MTBDRplus assay, the most common mutations associated with rifampicin (RIF) and isoniazid (INH) resistance among resistant strains in Morocco. METHODS: A total of 319 Mycobacterium tuberculosis isolates sent to the National Tuberculosis Reference Laboratory between 2013 and 2015 were subjected to GenoType® MTBDRplus for detecting M. tuberculosis and determination of drug susceptibility. Correlation of the minimum inhibitory concentrations (MICs) of INH with genotypic assay results was carried out for 97 MDR-TB strains. Various concentrations of INH were tested. RESULTS: The most frequent mutations observed were rpoBS531L (67.2%) and katGS315T1/2 (66.5%). Isolates with inhA gene mutation, katG gene mutation, and dual mutations in katG and inhA had MICs ranging from 0.5-1µg/mL, 2-10µg/mL and ≥12µg/mL, respectively. CONCLUSION: In Morocco, 66.5% and 76.7% of M. tuberculosis strains carried mutations causing high-level resistance to INH and RIF, respectively.

Synthesis and antibacterial activity of new 1,2,3-triazolylmethyl-2H-1,4-benzothiazin-3(4H)-one derivatives.

BACKGROUND: A novel series of 1,2,3-triazole derivatives containing 1,4-benzothiazin-3-one ring (7a-9a, 7b-9b), (10a-12a, 10b-12b) and (13-15) were synthesized by 1,3-dipolar cycloaddition reactions of azides α-D-galactopyranoside azide F, 2,3,4,6-tetra-O-acetyl-(D)-glucopyranosyl azide G and methyl-N-benzoyl-α-azidoglycinate H with compounds 4-6. FINDINGS: Initially, the reactions were conducted under thermal conditions in ethanol. The reaction leads, each time, to the formation of two regioisomers: (Schemes 2, 3) with yields of 17 to 21% for 1,5-disubstituted 1,2,3-triazole-regioisomers (7b-12b) and yields ranging from 61 to 65% for the 1,4-disubstituted regioisomers (7a-12a). In order to report an unequivocal synthesis of the 1,4-regioisomers and confirm the structures of the two regioisomers obtained in thermal conditions (Huisgen reactions), the method click chemistry (Copper-Catalyzed Azide-Alkyne Cycloaddition) has been used. CONCLUSIONS: The newly synthesized compounds using cycloaddition reactions were evaluated in vitro for their antibacterial activities against some Gram positive and Gram negative microbial strains. Among the compounds tested, the compound 8a showed excellent antibacterial activities against PA ATCC and Acin ESBL (MIC = 31.2 µg/ml).

Extensively drug-resistant tuberculosis (XDR-TB) in Morocco.

OBJECTIVES: Extensively drug-resistant tuberculosis (XDR-TB) has recently been identified as a major global health threat. The aim of this study was to evaluate the presence of XDR-TB among Mycobacterium tuberculosis isolates in Morocco and its association with demographic, clinical and epidemiological features. METHODS: A total of 524 patients from the Moroccan National Tuberculosis Reference Laboratory, representative of all of the geographic regions, were subject to first-line drug susceptibility testing (DST). Subsequently, 155 isolates found to be multidrug-resistant tuberculosis (MDR-TB) underwent second-line DST. Moreover, to enhance our understanding of the genetic basis of these drug-resistant strains, drug resistance-associated mutations were investigated in isolates either identified as pre-XDR- and XDR-TB or suspected resistant using the GenoType® MTBDRsl V1.0 assay. RESULTS: In this study, 4 (2.6%) XDR-TB and 18 (11.6%) pre-XDR-TB isolates were identified. Agreement between the MTBDRsl assay results and phenotypic DST was 95.2% for ofloxacin, 81.0% for kanamycin and 95.2% for amikacin. CONCLUSIONS: To the best of our knowledge, this is the first study to evaluate the frequency of XDR-TB in Morocco. These results highlight the need to reinforce the TB management policy in Morocco with regard to control and detection strategies in order to prevent further spread of XDR-TB isolates.

Molecular Typing of Mycobacterium Tuberculosis Complex by 24-Locus Based MIRU-VNTR Typing in Conjunction with Spoligotyping to Assess Genetic Diversity of Strains Circulating in Morocco.

BACKGROUND: Standard 24-locus Mycobacterial Interspersed Repetitive Unit Variable Number Tandem Repeat (MIRU-VNTR) typing allows to get an improved resolution power for tracing TB transmission and predicting different strain (sub) lineages in a community. METHODOLOGY: During 2010-2012, a total of 168 Mycobacterium tuberculosis Complex (MTBC) isolates were collected by cluster sampling from 10 different Moroccan cities, and centralized by the National Reference Laboratory of Tuberculosis over the study period. All isolates were genotyped using spoligotyping, and a subset of 75 was genotyped using 24-locus based MIRU-VNTR typing, followed by first line drug susceptibility testing. Corresponding strain lineages were predicted using MIRU-VNTRplus database. PRINCIPAL FINDINGS: Spoligotyping resulted in 137 isolates in 18 clusters (2-50 isolates per cluster: clustering rate of 81.54%) corresponding to a SIT number in the SITVIT database, while 31(18.45%) patterns were unique of which 10 were labelled as "unknown" according to the same database. The most prevalent spoligotype family was LAM; (n = 81 or 48.24% of isolates, dominated by SIT42, n = 49), followed by Haarlem (23.80%), T superfamily (15.47%), >Beijing (2.97%), > U clade (2.38%) and S clade (1.19%). Subsequent 24-Locus MIRU-VNTR typing identified 64 unique types and 11 isolates in 5 clusters (2 to 3isolates per cluster), substantially reducing clusters defined by spoligotyping only. The single cluster of three isolates corresponded to two previously treated MDR-TB cases and one new MDR-TB case known to be contact a same index case and belonging to a same family, albeit residing in 3 different administrative regions. MIRU-VNTR loci 4052, 802, 2996, 2163b, 3690, 1955, 424, 2531, 2401 and 960 were highly discriminative in our setting (HGDI >0.6). CONCLUSIONS: 24-locus MIRU-VNTR typing can substantially improve the resolution of large clusters initially defined by spoligotyping alone and predominating in Morocco, and could therefore be used to better study tuberculosis transmission in a population-based, multi-year sample context.

Antimicrobial resistance in Salmonella enterica serovar Enteritidis in Morocco.

INTRODUCTION: Salmonella enterica is recognised worldwide as one of the major agents of human gastrointestinal infections. The aim of the present work is to ascertain the antimicrobial susceptibilities of 150 Salmonella enterica serovar Enteritidis isolates from humans in Morocco during the period from 2000 to 2008. METHODOLOGY: Antimicrobial resistance determination was performed by disk diffusion method using seven antibiotics. The minimal inhibitory concentration (MIC) of ciprofloxacin was determined for nalidixic acid-resistant (NAR) isolates using E-test strips. RESULTS: Sixty-one (42%) isolates were resistant to at least one class of antimicrobial agent. The largest numbers of resistant isolates were observed for nalidixic acid with 53 isolates (36%) followed by ampicillin with 7 isolates (5%), tetracycline with 6 isolates (4%), and trimethoprim/sulfamethoxazole with 2 isolates (1%).The resistant isolates were grouped in seven different resistance patterns of which two isolates were resistant to three antibiotics. Among the 53 (36%) NAR isolates, 37 (76%) had a reduced susceptibility to ciprofloxacin. CONCLUSION: Resistance rates of Salmonella enterica serovar Enteritidis from Morocco are generally low but the resistance to nalidixic acid is worryingly common. Continual surveillance of antibiotic resistance is of primary importance.