Assessment of pathogenicity and tissue distribution of infectious bronchitis virus strains (Italy 02 genotype) isolated from moroccan broiler chickens.

BACKGROUND: Avian infectious bronchitis (IB) is one of the most important viral diseases of poultry, affecting chickens of all ages and causing major economic losses in poultry flocks. Mass vaccination is conducted in Morocco using a vaccine against Massachusetts, which is the most dominant serotype; however no information about the pathogenesis and tissue distribution of the Moroccan Italy 02 genotype was reported. 40 one-day-old specific pathogen free chickens were divided randomly into four groups. Group1, 2 and 3 were inoculated intra oculo-nasally with 103.5 EID50 of Italy02 viruses, and group 4 was kept as control. Chickens in each group were monitored for 14 days post-infection (pi). RESULTS: Chickens in all infected groups showed severe respiratory signs, which most of them have been reproduced on 2dpi, with varying times of appearance and disappearance. The infected birds appeared lethargic, reluctant to move, with specific respiratory clinical signs and macroscopic lesions. However no nephritis lesions or mortality were recorded in all groups. The specific histological lesions finding in all infected birds, exhibited tracheal lesions with mucosal thickening, hyperplasia of the surface epithelium, mononuclear inflammatory cell infiltrate of lamina propria. Primary and secondary bronchi, epithelial hyperplasia and mononuclear inflammatory cell infiltrate of the lamina propria were also observed. Tracheal lesions developed in all infected birds, confirm the ability of the three tested strains to induce respiratory disease. The results at 14 dpi also revealed that all strains were able to induce serological response. Virus re-isolation from infected organs and amplification of the viral RNA by real-time PCR proved the presence of the virus in lung and trachea of infected chicks. Neither re-isolation nor significant viral RNA detection were detected in the kidney. CONCLUSION: The results demonstrated that the three strains Italy02 genotype emerging in Moroccan poultry farms have a wide distribution for respiratory system, without kidney damage and without causing mortality.

Phylogenetic analysis of avian infectious bronchitis virus S1 glycoprotein regions reveals emergence of a new genotype in Moroccan broiler chicken flocks.

BACKGROUND: Infectious bronchitis virus (IBV), a major pathogen of commercial poultry flocks, circulates in the form of several serotypes/genotypes. Only a few amino-acid changes in the S1 subunit of wild-type IBVS proteins may result in mutants unaffected by current vaccines. METHODS: Partial S1 gene sequences of 3 IBV isolates of the Moroccan Italy 02 genotype from vaccinated and unvaccinated broiler chicken flocks, located in southern and central regions of Morocco, were amplified by RT-PCR, sequenced, and aligned for phylogenetic and amino-acid similarity analyses. RESULTS: The three isolates were found genetically highly distant from known avian IBV based on partial sequences of their S1 genes: gammaCoV/chicken/Morocco/I01/2011(IBV/Morocco/01), gammaCoV/chicken/Morocco/I30/2010 (IBV/Morocco/30), and gammaCoV/chicken/Morocco/I38/2013 (IBV/Morocco/38), nucleotide sequence identities reached 89.5 % to 90.9 % among the three isolates. The deduced protein sequence identities ranged from 29.7 % (between IBV/Morocco/38 and Egypt SCU-14/2013-1) to 78.2 % (between IBV/Morocco/01 and Spain/05/866). Amino acid sequence comparison and phylogenetic analysis indicated the emergence of a new Moroccan genotype, clustering with regionally related isolates from Spain (Spain/05/866) and belonging to a new sub-genotype. CONCLUSION: Our sequencing results demonstrate a co-circulation of wild-type infectious bronchitis viruses in broiler chickens. These results justify permanent monitoring of circulating strains in order to rationally modify vaccination strategies to make them appropriate to the evolving field situation.

Prevalence and molecular characterization of avian infectious bronchitis virus in poultry flocks in Morocco from 2010 to 2014 and first detection of Italy 02 in Africa.

The aim of this study was to investigate the prevalence and diversity of infectious bronchitis virus (IBV) genotypes in poultry flocks in 16 areas of Morocco between 2010 and 2014. A total of 360 chicken flocks suspected of being infected by IBV were screened for the IBV N gene using real-time reverse transcriptase-polymerase chain reaction (RT-PCR). Flocks were classified into four groups according to their IBV vaccination programme. Group 1 contained unvaccinated birds. Group 2 received a single application of live H120 vaccine. Groups 3 and 4 birds received one or two booster vaccination(s), respectively, mostly using the H120 vaccine. The real-time RT-PCR results showed that 51.7% of the flocks were positive for the IBV genome with geographical disparities. Molecular characterization of IBV was performed on 50 RT-PCR positive samples by partially sequencing the S1 gene, including the hypervariable regions (nucleotides 705-1097). Two predominant genotypes were detected, with the Massachusetts type dominating (66%), among which 25% of the samples were identical to the H120 vaccine. The second most common genotype (present in 32% of the flocks) was surprisingly Italy 02, revealing the first detection of this genotype in Morocco and also in Africa. 793B, the predominant genotype in the late 1990s in Morocco, was only detected on one occasion and was identical to the 4/91 vaccine strain. This study highlights the high prevalence of IBV in poultry farms in Morocco and confirms its continuous dynamic changes and evolution.

Awareness and knowledge regarding of cervical cancer, Pap smear screening and human papillomavirus infection in Gabonese women.

BACKGROUND: Cervical cancer is the commonest cancer and the leading cause of cancer mortality in women in Gabon. The age-standardized incidence of cervical cancer is 19.9 per 100 000 women and the mortality rate is 8.4 per 100 000. Various international studies have identified the lack of awareness and knowledge about cervical cancer as barriers to use preventive methods. This article assesses the awareness and knowledge about cervical cancer, Pap smear testing and its use and HPV among women living in Libreville, Gabon. METHODS: This study was conducted in October 2014 in Libreville. A total of 452 women aged 16 years and above were recruited from different town locations. Logistic regression analysis was used to identify the effect of demographic characteristics on the level of knowledge about cervical cancer, Pap smear testing and HPV. Odds ratio and 95% confidence intervals were used to identify the strength of association. Associations were considered statistically significant at p < 0.05. RESULTS: Of all the women interviewed, 91.6% (414/452) had heard about cervical cancer and only 27.9% (126/452) had heard of Pap smear test. Of these 126 women, only 65.1% (82/126) had done cervical cancer screening and 68.3% (56/82) on the suggestion of a doctor. The most common reason for not undergoing Pap smear testing was neglect (50%, 22/44) followed by lack of financial resources (13.6%, 6/44), fear of discovering a serious disease (13.6%, 6/44) and deeming it unimportant (13.6%, 6/44). Only 8% (40/452) of the participants had heard about HPV and their knowledge of HPV was fair. There is a very poor level of knowledge about cervical cancer among Gabonese women. CONCLUSION: This study demonstrates a very low level of knowledge about cervical cancer, Pap smear testing and HPV in a sample of Gabonese women. There is a critical need for Gabonese women to be informed about cervical cancer and the Pap smear test to improve the use of this preventive method. The implication of health staff and Gabonese media should be included as a centerpiece in the effort to inform the population in order to reduce the burden of cervical cancer in Gabon and save women lives.

Expression profile of undifferentiated cell transcription factor 1 in normal and cancerous human epithelia.

Undifferentiated cell Transcription Factor 1 (UTF1) is a chromatin-bound protein involved in stem cell differentiation. It was initially reported to be restricted to stem cells or germinal tissues. However, recent work suggests that UTF1 is also expressed in somatic cells and that its expression may increase during carcinogenesis. To further clarify the expression profile of UTF1, we evaluated UTF1 expression levels immunohistochemically in eight normal human epithelia (from breast, prostate, endometrium, bladder, colon, oesophagus, lung and kidney) and their corresponding tumours as well as in several epithelial cell lines. We showed UTF1 staining in normal and tumour epithelial tissues, but with varying intensities according to the tissue location. In vitro analyses also revealed that UTF1 is expressed in somatic epithelial cell lines even in the absence of Oct4A and Sox2, its two main known regulators. The comparison of UTF1 levels in normal and tumoral tissues revealed significant overexpression in endometrial and prostatic adenocarcinomas, whereas lower intensity of the staining was observed in renal and colic tumours, suggesting a potential tissue-specific function of UTF1. Altogether, these results highlight a potential dual role for UTF1, acting either as an oncogene or as a tumour suppressor depending on the tissue. These findings also question its role as a specific marker for stem cells.

Analysis of mutations in the E6 oncogene of human papillomavirus 16 in cervical cancer isolates from Moroccan women.

BACKGROUND: Worldwide, cervical cancer is the second most common cancer in women. High-risk human papillomavirus (HPV) play a crucial role in the etiology of cervical cancer and the most prevalent genotype is HPV16. HPV 16 intratypic variants have been reported to differ in their prevalence, biological and biochemical properties. The present study was designed to analyze and identify HPV type 16 E6 variants among patients with cervical cancer in Morocco. METHODS: A total of 103 HPV16 positive samples were isolated from 129 cervical cancer cases, and variant status was subsequently determined by DNA sequencing of the E6 gene. RESULTS: Isolates from patients were grouped into the European (E), African (Af) and North-American (NA1) phylogenetic clusters with a high prevalence of E lineage (58.3%). The Af and NA1 variants were detected in 31.1% and 11.6% of the HPV16 positive specimens, respectively, whereas, only 3% of cases were prototype E350T. No European-Asian (EA), Asian (As) or Asian-American (AA) variants were observed in our HPV16-positive specimens. At the amino acid level, the most prevalent non-synonymous variants were L83V (T350G), H78Y (C335T), E113D (A442C), Q14D (C143G/G145T) and R10I (G132T), and were observed respectively in 65%, 41.8%, 38.8%, 30.1% and 23.3% of total samples.Moreover, HPV16 European variants were mostly identified in younger women at early clinical diagnosis stages. Whereas, HPV16 Af variants were most likely associated with cervical cancer development in older women with pronounced aggressiveness. CONCLUSION: This study suggests a predominance of E lineage strains among Moroccan HPV 16 isolates and raises the possibility that HPV16 variants have a preferential role in progression to malignancy and could be associated with the more aggressive nature of cervical cancer.

Prevalence and risk factors of hepatitis B and C virus infections among the general population and blood donors in Morocco.

BACKGROUND: Viral hepatitis is a serious public health problem affecting billions of people globally. Limited information is available on this issue in Morocco. This cross-sectional study was undertaken with the aim of determining the seroprevalence and risk factors of hepatitis B virus (HBV) and hepatitis C virus (HCV) among the general population and among blood donors. METHODS: Blood samples from volunteers, have been screened with ELISA tests for detecting the hepatitis-B surface antigen (HBsAg) and anti-HCV. Within the seroreactive patients for HCV in the general population, RT-PCR was performed by the Cobas Ampliprep/Cobas Amplicor. RESULTS: HCV and HBV-seropositivity was documented in 1.58% and 1.81% out of 41269 and 23578 participants respectively from the general population. Two patients were found to be co-infected. HCV-RNA was detected by PCR in 70.9% of the 195 anti-HCV positive subjects. The anti-HCV prevalence was not different among males and females (P = 0.3). It increased with age; the highest prevalence was observed among subjects with >50 years old (3.12%). Various risk factors for acquiring HCV infection were identified; age, dental treatment, use of glass syringes and surgical history. In addition to these factors, gender and sexual risk behaviors were found to be associated with higher prevalence of hepatitis B. The HBV positivity was significantly higher among males than females participants in all age groups (P < 0.01). The peak was noticed among males aged 30-49 years (2.4%). None of the 152 persons younger than 20 years had HBsAg or anti-HCV. The prevalence of anti-HCV and HBsAg among 169605 blood donors was 0.62% and 0.96% respectively. CONCLUSIONS: Our study provided much important information concerning hepatitis B and C prevalence and risk factors; it confirmed the intermediate endemicity for HCV infection and pointed to a decreasing trend of HBV incidence, which might reclassify Morocco in low HBV endemicity area. This could be attributed primarily to the universal HBV vaccination among infants and healthcare workers over the past 13 years. HCV and HBV infections in the present survey were mainly associated with nosocomial exposures. Prevention and control of HBV infection are needed to reduce HBV transmission between adults.

Impact of rotavirus and hepatitis A virus by worldwide climatic changes during the period between 2000 and 2013.

Enteric viruses are present in the environment as a result of the discharge of poorly or untreated wastewater. The spread of enteric viruses in the environment depend to human activities like stools of infected individuals ejected in the external environment can be transmitted by water sources and back to susceptible individuals for other cycles of illness. Among the enteric viruses Rotaviruses (RV) and Hepatitis A viruses (HAV) is the most detected in wastewater causing gastroenteritis and acute hepatitis. Therefore, it is of interest to climate change, mainly temperature and carbon Dioxide (CO2) variations, on Rotavirus and Hepatitis A as a model of enteric viruses present in the aquatic environment using computational modelling tools. The results of genetic ratio showed a negative correlation between the epidemiological data and the mutation rate. However, the correlation was positive between the temperature, CO2 increase, and the rate of mutation. The positive correlation is explained by the adaptation of the viruses to the climatic changes, the RNA polymerase of the RV induces errors to adapt to the environmental conditions. The simultaneous increase in number of infections and temperature in 2010 has been demonstrated in previous studies deducing that viral pathogenicity increase with temperature increase.

Naturally occurring capsid protein variants L1 of human papillomavirus genotype 16 in Morocco.

HPV L1 protein is a corner stone in HPV structure, it's involved in the formation of the viral capsid; widely used as a systematic material and considered as the main component in vaccines development and production. The present study aims to characterize genetic variation of L1 gene of HPV 16 specimens and to evaluate in silico the impact of major variants on the epitope change affecting its conformational structure. A fragment of L1 gene from 35 HPV 16 confirmed specimens were amplified by PCR and sequenced. Overall, five amino acids residues changes were reported: T390P in 16 specimens, M425I and M431I in 2 cases, insertion of Serine at 460 and aspartic acid deletion at position 477 in all analyzed cases. The 3D generated model showed that T389P amino acid substitution is located in the H-I loop; the two substitutions M424I and M430I are both located in the H2 helice. The Serine insertion and aspartic acid deletion are located in the H4 helice and B-C loop, respectively. Superimposition of sequences' structures showed that they share a very similar conformation highlighting that the reported amino acids variations don't affect the structure of the L1 protein. However T389P, located in the H-I loop identified as an immunogenetic region of L1 capsid, was reported in 51.4% of cases could interact with vaccines induced monoclonal antibodies suggesting a potential impact on the efficacy of available anti-HPV vaccines.

Current situation, genetic relationship and control measures of infectious bronchitis virus variants circulating in African regions.

Infectious bronchitis virus (IBV) is a major viral pathogen of commercial poultry, affecting chickens of all ages and causing major economic losses in poultry industry worldwide. Frequent points of mutations and recombination events in the S1 gene region, result in the emergence of new IBVs variants circulating in the form of several serotypes/genotypes that can be partially or poorly neutralized by current vaccines. IBV is well studied worldwide, nevertheless in African countries epidemiological and scientific data are poor and not updated. This review aims to give a current overview of IBV situation, to establish evolutionary relationship between the African variants and to list some of the potential measures to control IBV in Africa. Three S1 gene hypervariable regions were studied and compared to the reference genotypes/serotypes that found emerging in African regions. This comparison was based on phylogenetic trees, nucleotide and amino-acid sequence analysis. It clearly appears that IBV variants reported in Africa, display a low genetic relationship between them and with the majority of the reference strains emerging in neighboring countries, except the case of variants from Libya and Egypt that show a high relatedness. Also the Massachusetts serotypes were the most prevalent co-circulating with both serotypes, Italy02 type in Morocco and Qx-like genotype in South part of the African continent. In order to control the IBV variants in African regions, an efficient vaccination strategy program should be implemented.

Human papillomavirus genotypes distribution in cervical cancer cases in Gabon.

BACKGROUND: Cervical cancer is a real public health problem in African countries. The relation between HPV and cervical cancer is well established. However, it is known that the distribution of HPV genotypes differ geographically and this may influence the effectiveness of the three available vaccines, which among other HPV genotypes targets the genotypes 16 and 18 that cause about 70 % of cervical cancers cases. The objective of this study was to identify for the first time the HPV genotypes distribution in cervical cancer specimens obtained from Gabonese women. METHODS: A total of 105 cervical samples including 93 formalin-fixed paraffin embedded tissues collected between 2007 and 2013 and 12 fresh biopsies collected in August 2013 were investigated. The presence of HPV DNA was analyzed by nested PCR with primers MY09/11 and GP5+/6+ followed by sequencing for HPV genotyping. RESULTS: Amplification of the housekeeping gene (ß-globin) with PCO4/GH20 primers was successful for 91.4 % (96/105) of the cervical cancer samples and HPV DNA was detected in all the 96 samples. Five different HPV genotypes were identified. HPV 16 [58.3 %; 95 % IC: 48.44-68.16] was the most common genotype followed by HPV 33 [25.0 %; 95 % IC: 16.34-33.66], HPV 18 [8.4 %; 95 % IC: 2.86-13.94], HPV 70 [7.3 %; 95 % IC: 2.1-12.5] and HPV 31 [1.1 %; 95 % IC: -0.986-3.186]. HPV 16 was also the most prevalent in all histological malignant lesions. It was found in 56.6 % of squamous cervical carcinoma and 69.2 % of adenocarcinoma. Concerning the HPV positive adenocarcinoma cases, HPV 18 was identified in 7.7 % (1/13). CONCLUSION: These findings show the predominance of HPV 16 in cervical cancer cases among Gabonese women. However, HPV33 is more prevalent than HPV18. Our study suggests that HPV vaccines may be effective at reducing the burden of cervical cancer in Gabon.

Human papillomavirus genotypes distribution among Gabonese women with normal cytology and cervical abnormalities.

BACKGROUND: Cervical cancer is one of the most common tumors affecting women with a disproportionate mortality occurring in developing countries. Despite the high prevalence of cervical cancer and cervical neoplasia in Gabon, few studies have been performed to evaluate the prevalence and determinants of HPV infection in this country. The aim of this study was to determine the HPV prevalence and distribution in a population of Gabonese women with normal cytology and cervical abnormalities. METHODS: A total of 200 cervical samples collected in the "Departement d'Anatomie et de Cytologie Pathologiques" of the "Faculté de Medecine et des Sciences de la Santé" in Libreville, Gabonwere analyzed. Cytological status was classified according to Bethesda 2001. Nested polymerase chain reaction (PCR) using consensus degenerate PCR primers (MY09/11 and GP5+/6+) was performed for the detection of HPV DNA and HPV typing was done by DNA sequencing. RESULTS: Cytological analysis showed that 87 % of women had normal cytology (n = 174/200). Among the 26 women with cytological abnormalities, predominance (61.5 %; 16/26) of low grade squamous intraepithelial lesion (LSIL) was found and no cervical cancer case was detected. Overall, HPV DNA was detected in 60 % of women (120/200). With respect to the cytological status, HPV DNA was found in 57.5 % of women with normal cervix and 76.9 % of women with abnormal cytology. HPV genotyping was performed on 114 HPV positive cases and revealed the presence of 11 distinct genotypes: 16, 18, 33, 31, 56, 6, 66, 70, 35, 45 and 81. The high risk type HPV 16 was the most common genotype found in all cytological categories. Six HPV positive samples could not be typed by DNA sequencing, probably due to multiple HPV infection. Evaluation of possible risk factors showed that HPV infection was related positively with number of sexual partners (≥3, OR = 2.3; 95 % CI, 1.3-4.3), history of sexually transmitted infection (Chlamydia, OR = 1.9; 95 % CI, 1.01-3.4) and marital status (single, OR = 2.0; 95 % CI, 1.1-3.5). CONCLUSION: The prevalence of HPV infection among Gabonese women is high. Our findings highlight the need to set up a national program to fight cervical cancer, combining Pap smear test and HPV testing, to improve cervical cancer prevention in Gabon.

Comparison of SYBR green I real-time RT-PCR with conventional agarose gel-based RT-PCR for the diagnosis of infectious bronchitis virus infection in chickens in Morocco.

BACKGROUND: A rapid, sensitive, and specific molecular method for the diagnosis of infectious bronchitis virus (IBV) infection is important in curbing infectious bronchitis outbreaks in Morocco and other countries. METHODS: In this study, an easy-to-perform SYBR green I real-time reverse transcriptase polymerase chain reaction (RT-PCR) targeting the nucleocapsid gene of IBV was developed and compared with conventional agarose gel-based RT-PCR for the detection of IBV infection. RESULTS: We found that the SYBR green I real-time RT-PCR was at least 10 times more sensitive than the agarose gel electrophoresis detection method. The assay exhibited high specificity for IBV infection. All negative controls, such as Newcastle disease virus, infectious bursal disease virus, and avian influenza virus, were not detected. CONCLUSION: The SYBR green I real-time RT-PCR test described herein can be used to rapidly distinguish IBV from other respiratory pathogens, which is important for diagnosis and control of infectious bronchitis outbreaks in Morocco. The test is a valuable and useful method as a routine assay for diagnosis of clinical IBV infection in commercial chickens.

Multiple alignment comparison of the non-structural genes of three strains of equine influenza viruses (H3N8) isolated in Morocco.

BACKGROUND: Three equine influenza viruses, A/equine/Nador/1/1997(H3N8), A/equine/Essaouira/2/2004(H3N8), and A/equine/Essaouira/3/2004(H3N8), were isolated from different Equidae during local respiratory disease outbreaks in Morocco in 1997 and 2004. Their non-structural (NS) genes were amplified and sequenced. RESULTS: The results show high homology of NS nucleotide sequences of A/equine/Nador/1/1997 with European strains (i.e., A/equine/newmarket/2/93 and A/equine/Grobois/1/1998) and clustered into the European lineage. However, NS gene of A/equine/Essaouira/2/2004(H3N8) and A/equine/Essaouira/3/2004(H3N8) strains indicated high homology with equine influenza strains that had circulated before 1990 (A/equine/Fontainbleu/1/1979(H3N8), which belonged to a pre-divergent phase Amino acid sequence comparison of the NS1 protein with reference strain A/equine/Miami/1963(H3N8) shows that the A/equine/Nador/1/1997(H3N8) strain has 12 substitutions at the residues D/24/N, R/44/K, S/48/I, R/67/Q, A/86/V, E/139/K, A/112/T, E/186/K, L/185/F, A/223/E, S/213/T and S/228/P. In both A/equine/Essaouira/2/2004(H3N8) and A/equine/Essaouira/3/2004(H3N8) strains, the NS1 sequences present one common mutation at the residue: S/228/P. CONCLUSION: It seems that all of these substitutions are not produced at the key residues of the RNA-binding domain (RBD) and the effector domain (ED). Consequently, we can suppose that they will not affect the potency of inhibition of cellular defences, and the virulence of the Moroccan equine strains will be maintained.

Direct sequencing for rapid detection of multidrug resistant Mycobacterium tuberculosis strains in Morocco.

BACKGROUND: Tuberculosis (TB) is a major public health problem with high mortality and morbidity rates, especially in low-income countries. Disturbingly, the emergence of multidrug resistant (MDR) and extensively drug resistant (XDR) TB cases has worsened the situation, raising concerns of a future epidemic of virtually untreatable TB. Indeed, the rapid diagnosis of MDR TB is a critical issue for TB management. This study is an attempt to establish a rapid diagnosis of MDR TB by sequencing the target fragments of the rpoB gene which linked to resistance against rifampicin and the katG gene and inhA promoter region, which are associated with resistance to isoniazid. METHODS: For this purpose, 133 sputum samples of TB patients from Morocco were enrolled in this study. One hundred samples were collected from new cases, and the remaining 33 were from previously treated patients (drug relapse or failure, chronic cases) and did not respond to anti-TB drugs after a sufficient duration of treatment. All samples were subjected to rpoB, katG and pinhA mutation analysis by polymerase chain reaction and DNA sequencing. RESULTS: Molecular analysis showed that seven strains were isoniazid-monoresistant and 17 were rifampicin-monoresistant. MDR TB strains were identified in nine cases (6.8%). Among them, eight were traditionally diagnosed as critical cases, comprising four chronic and four drug-relapse cases. The last strain was isolated from a new case. The most recorded mutation in the rpoB gene was the substitution TCG > TTG at codon 531 (Ser531 Leu), accounting for 46.15%. Significantly, the only mutation found in the katG gene was at codon 315 (AGC to ACC) with a Ser315Thr amino acid change. Only one sample harbored mutation in the inhA promoter region and was a point mutation at the -15p position (C > T). CONCLUSION: The polymerase chain reaction sequencing approach is an accurate and rapid method for detection of drug-resistant TB in clinical specimens, and could be of great interest in the management of TB in critical cases to adjust the treatment regimen and limit the emergence of MDR and XDR strains.

Evaluation of conventional molecular diagnosis of Mycobacterium tuberculosis in clinical specimens from Morocco.

INTRODUCTION: Tuberculosis is a major public health threat, annually affecting new individuals worldwide, especially those in developing countries. Rapid detection of the agent and effective treatment are two important factors in controlling this disease. METHODOLOGY: The present study aimed to evaluate polymerase chain reaction (PCR) as a rapid and direct molecular method for the diagnosis of Mycobacterium tuberculosis (MTB) in 70 clinical specimens (62 sputum samples, six cerebrospinal fluids, and two biopsies) using heat shock protein (hsp65) as the gene target. Automated sequencing of the same gene was used for the identification of MTB to the species level. RESULTS: The sensitivity of PCR was 81.13%, with specificity of 88.24%; the positive and negative predictive values were 95.56% and 60%, respectively. CONCLUSION: Based on these results, the hsp65 gene sequence can be used to differentiate the members of MTB complex from non-tuberculosis mycobacteria (NTM).

HBV genotypes prevalence, precore and basal core mutants in Morocco.

The study of hepatitis B virus (HBV) genomic heterogeneity has become a major issue in investigations aimed at understanding the relationship between HBV mutants and the wide spectrum of clinical and pathological conditions associated with HBV infection. The objective of the current study was to find out the pattern of HBV genotypes circulating in Morocco and to investigate the precore (PC) and basal core promoter (BCP) mutants' status in Moroccan chronic hepatitis B patients. Viral genotypes were determined in 221 chronic carriers using INNO-LiPA HBV assay and hemi-nested PCR. Phylogenetic analysis was performed in 70 samples, and multiplex PCR method was used to confirm some genotyping results. PC and CP mutants were determined using Inno-Lipa. All isolates were successfully genotyped. The genotype distribution was D in 90.45% of cases, A (5.9%), E (1 case), and mixed genotypes (5 A/D and 2 D/F) in 3.17% patients. HBV carried in the HBV/D samples could be assigned to D7 (63.3%), D1 (32.7%) and 2% of strains to each D4 and D5, all HBV/A belonged to A2 subgenotype and HBV/E strain could not be sub-genotyped. In 70 studied strains, HBV mutants were detected in 88.6% of cases; PC mutants were detected in (40%) of patients and 21.5% present a mixture of wild type and G1896A mutation. BCP mutants were observed in 65.7% of cases, 22.9% were found to have the T1762/1764A double mutation, 18.6% had A1762/1764T mutation and 22.9% of patients showed the A1762T/G1764A double mutation with either A1762T/G1764T mutation. Co-infection by PC and BCP mutants was detected in 52.9% of cases. Movement from place to place most likely shapes the observed genotype distribution and consequent prevalence of genotypes other than A2 or D7 in this population. High circulation of PC and BCP mutants is common in chronic hepatitis B infection in Morocco.

Aberrant promoter methylation and expression of UTF1 during cervical carcinogenesis.

Promoter methylation profiles are proposed as potential prognosis and/or diagnosis biomarkers in cervical cancer. Up to now, little is known about the promoter methylation profile and expression pattern of stem cell (SC) markers during tumor development. In this study, we were interested to identify SC genes methylation profiles during cervical carcinogenesis. A genome-wide promoter methylation screening revealed a strong hypermethylation of Undifferentiated cell Transcription Factor 1 (UTF1) promoter in cervical cancer in comparison with normal ectocervix. By direct bisulfite pyrosequencing of DNA isolated from liquid-based cytological samples, we showed that UTF1 promoter methylation increases with lesion severity, the highest level of methylation being found in carcinoma. This hypermethylation was associated with increased UTF1 mRNA and protein expression. By using quantitative RT-PCR and Western Blot, we showed that both UTF1 mRNA and protein are present in epithelial cancer cell lines, even in the absence of its two main described regulators Oct4A and Sox2. Moreover, by immunofluorescence, we confirmed the nuclear localisation of UTF1 in cell lines. Surprisingly, direct bisulfite pyrosequencing revealed that the inhibition of DNA methyltransferase by 5-aza-2'-deoxycytidine was associated with decreased UTF1 gene methylation and expression in two cervical cancer cell lines of the four tested. These findings strongly suggest that UTF1 promoter methylation profile might be a useful biomarker for cervical cancer diagnosis and raise the questions of its role during epithelial carcinogenesis and of the mechanisms regulating its expression.

Characterization and antibiotic susceptibility of Listeria monocytogenes isolated from poultry and red meat in Morocco.

This study was carried out on 426 samples of raw meats collected from butcheries and supermarkets in Casablanca, Morocco. The samples were examined for the occurrence of Listeria species. Strains of Listeria monocytogenes were characterized by several biochemical tests and confirmed by polymerase chain reaction (PCR). ß-hemolytic cultures and nonhemolytic isolates were tested for biochemical properties with the Listeria API test. Among the 43 Listeria species isolates; we identified 10 strains for L. monocytogenes (23.3%), 31 strains for L. innocua (72.1%) and 2 strains for L. welshimeri (4.6%). Strains of L. monocytogenes were separated by multiplex PCR; two serogroups IIb and IVb were thus differentiated. Antibiotic susceptibility of L. monocytogenes to 21 antibiotics was determined by the disk diffusion method. All isolates were susceptible to a wide range of the tested antibiotics with the exception of nalidixic acid, colistine and cephalosporins second and third generation for which they were all resistant.