Development of a Usutu virus specific real-time reverse transcription PCR assay based on sequenced strains from Africa and Europe.

Usutu virus (USUV) has been isolated in several African and European countries mainly from mosquitoes and birds. However, previous benign and two recent severe cases of human infections point out the need of a tool for the identification of USUV in human samples. A published real-time reverse transcription (RT) PCR assay for the detection of USUV in human blood or cerebrospinal fluid does not take into account the genetic variability of USUV in different geographic regions. Therefore, this article presents a quantitative real-time RT-PCR assay based on sequences from Europe and Africa. Primers and probe were designed in conserved regions among USUV strains that differed from closely related flaviviruses. The specificity of the assay was investigated by testing 16 other flaviviruses circulating in Africa. The sensitivity was determined by testing serial dilutions of virus and RNA standard. Intra- and inter-assay coefficients of variation were evaluated by 10 reactions in a same and in different assays, respectively. The assay provides high analytical specificity for USUV and detection limits of 1.2pfu/reaction for virus dilutions in L-15 medium or human serum and 60 copies/reaction for the RNA standard. The assay needs to be evaluated in a clinical context and integrated in standard diagnosis of flaviviral diseases.

[Aetiology of AFB negative pneumonias in hospitalized HIV patients in Dakar]. / Étiologie des pneumonies BAAR négatives chez les patients infectés par le VIH hospitalisés à Dakar (étude ANRS 1260).

A prospective study was carried out in two tertiary hospitals in Dakar to determine the main causes of sputum acid-fast bacillus (AFB) smear-negative pneumonia in HIV-infected patients. All clinical and microbiological records were reviewed by experts. Seventy patients were finally enrolled. Most of them were hospitalized at an advanced stage of AIDS. The median CD4 cell count was 62/mm(3) and the median body mass index (BMC) was 18 kg/m(2). Thirty-one patients (44 %) were known as seropositive for HIV infection prior to admission. Radiological opacities were localized in 70 % of patients and diffuse in 21 %. Fiberoptic bronchoscopy was performed in 50 patients (71 %). A definite or probable diagnosis was obtained in 55 patients (79 %). Bacterial pneumonia (usually due to Enterobacteriaceae and Pseudomonas aeruginosa), tuberculosis, Pneumocystis pneumoniae and other causes (Kaposi's sarcoma, atypical mycobacteria) were diagnosed in 67 %, 24 %, 5 %, and 13 % of these patients respectively. In conclusion, pneumonia of bacterial origin and tuberculosis can be incriminated in the majority of cases of AFB negative pneumonia observed in HIV patients in Dakar.

[Update on the Mali Pharmacy Code: legislative section]. / Essai de mise au point d'un code de la pharmacie au Mali: partie legislative.

Due to colonization (1895-1960) Mali has been submitted, on the legislative and regulation plan to a corpus of numerous and scattered texts relative to pharmacy. It is essentially the Law AN XI of 21 Germinal or the 11 April 1803 Law; the content of Book V of the Public Health Code relative to pharmacy practice, of which some disposals have been extended to Overseas Territories, Togo and Cameroon in 1953; the 1955 Ministerial Order about the dispatching of the Retail Pharmacy in AOF; the 1960 Law creating the Board of Pharmacists in the Federation of Mali etc. After independence, the new Malian State while renewing the pharmacy legislation issued by the old tutor State which was not opposed to the new fundamental Law, has also set up, according to the political orientations of the moment, some new texts. In the framework of this work, we have collected all the legislative and regulation texts that have been allowed in Mali in the pharmacy area and in that of Studies in Pharmacy. We have examined those texts and proceeded to codify the legislative part. The present Code consists of 189 articles divided in 5 titles layed out in chapters and sections.

[Update on the Mali Pharmacy Code: legislative section]. / Essai de Mise au Point D'un Code de la Pharmacie au Mali : Partie Legislative.

Due to colonization (1895-1960) Mali has been submitted, on the legislative and regulation plan to a corpus of numerous and scattered texts relative to pharmacy. It is essentially the Law AN XI of 21 Germinal or the 11 April 1803 Law; the content of Book V of the Public Health Code relative to pharmacy practice, of which some disposals have been extended to Overseas Territories, Togo and Cameroon in 1953; the 1955 Ministerial Order about the dispatching of the Retail Pharmacy in AOF; the 1960 Law creating the Board of Pharmacists in the Federation of Mali etc. After independence, the new Malian State while renewing the pharmacy legislation issued by the old tutor State which was not opposed to the new fundamental Law, has also set up, according to the political orientations of the moment, some new texts. In the framework of this work, we have collected all the legislative and regulation texts that have been allowed in Mali in the pharmacy area and in that of Studies in Pharmacy. We have examined those texts and proceeded to codify the legislative part. The present Code consists of 189 articles divided in 5 titles layed out in chapters and sections.

Essai de mise au point d'un code de la pharmacie au Mali : partie legislative

Du fait de la colonisation (1895-1960) le Mali a ete soumis au plan legislatif et reglementaire a un corpus de textes nombreux et epars se rapportant au secteur de la pharmacie. Il s'agit entre autre de la Loi An XI du 21 Germinal ou Loi du 11 avril 1803; du contenu du livre V du Code de la Sante Publique relatif a l'exercice de la pharmacie; dont certaines dispositions ont ete etendues aux territoires d'Outre-Mer; au Togo et au Cameroun en 1953 ; de l'Arrete de 1955 sur la repartition des officines en AOF ; de la Loi de 1960 portant creation de l'Ordre des pharmaciens dans la federation du Mali etc.. Apres l'accession a la souverainete internationale; le nouvel Etat Malien tout en reconduisant la legislation pharmaceutique issue de l'Etat anciennement tuteur qui n'etait pas contraire a la nouvelle loi fondamentale; a egalement mis en place; conformement aux orientations politiques de l'heure; de nouveaux textes. Dans le cadre de ce travail; nous avons recense l'ensemble des textes legislatifs et reglementaires qui ont ete pris au Mali dans le secteur pharmaceutique et celui des etudes en pharmacie. Nous avons fait l'examen critique de ces textes et procede a la codification de la partie legislative. Le present code est presente en 189 articles repartis en 5 titres disposes en chapitres et en sections

[Nosocomial infections: main causative agents]. / Les infections nosocomiales: agents responsables.

INTRODUCTION: Nosocomial Infections acquired within a care unit, may be related to various agents (Bacteria, virus, fungi, parasites...). The study purpose is to give relevent data on the main causative agents. MAIN AGENTS: The authors have shown the prevalent role of bacteria, mainly Enterobacteria, Staphylococcus and Pseudomonas; Legionella is also frequently isolated in some countries. Multiresistant Bacteria could be sometimes associated to nosocomial infections and severe infections. Among Nosocomial Virus: VRS, Rotavirus, Hepatitis B and C Viruses, HIV, Influenza Virus are cited. Fungal Agents (Aspergillus, Candida), Plasmodium, Non Conventional Agents (Prions) are also causative agents of Nosocomial Infection. CONCLUSION: The Isolation of contagious Agent in a hospitalized patient is not the only criteria for Nosocomial Infection typing; Rigorous analysis needed to be done for every case.

Class 2 integron-associated antibiotic resistance in Shigella sonnei isolates in Dakar, Senegal.

Forty-three Shigella sonnei isolates from adult patients with diarrhoea in Dakar were analysed for the presence of integrons. Isolates were resistant to sulphamethoxazole, trimethoprim, tetracycline, streptomycin and spectinomycin. A high prevalence of class 2 integrons (93%) was found. These integrons showed three distinct structures: a class 2 integron, part of the Tn7 family and its derivatives, carrying four cassettes in the order dfrA1-sat-aadA1-orfX; a truncated class 2 integron, without orfX; and a third type ca. 4 kb in size. These class 2 integrons probably play a role in the spread of multiresistance in S. sonnei isolates. To our knowledge, this is the first description of class 2 integrons in S. sonnei isolated in sub-Saharan Africa.

[A new case of rhinoscleroma with skin extension]. / Un nouveau cas de rhinosclérome avec atteinte cutanée.

Scleroma is a chronic and specific granulomatous disease of bacterial origin. Klebsialla rhinoscleromatis, a gram-negative bacillus. The majority of cases affect the upper airways, particularly the nose, thus justifying the term of rhinoscleroma. Extension to the palate, the upper lip and the skin is possible. A new case of rhinoscleroma with skin extension is reported. To us, this is the second case in Senegal (Casamance). Epidemiological, clinical, bacteriological and histological aspects of the disease are reviewed. Under suitable antibiotic, evolution is currently favorable.

Prevalence of methicillin-resistant Staphylococcus aureus in eight African hospitals and Malta.

Methicillin-resistant Staphylococcus aureus (MRSA) poses a serious therapeutic problem worldwide, and its frequency in most African countries has not been reported. This study was aimed at determining the prevalence and antibiotic susceptibility patterns of MRSA in eight large hospitals (>500 beds) in Africa and Malta, from 1996 to 1997. Susceptibility to methicillin (oxacillin) and to other drugs was determined by E test (AB Biodisk, Solna, Sweden) on a total of 1440 clinical isolates of S. aureus. Methicillin resistance was detected in 213 (15%) of the 1440 isolates tested. The rate of MRSA was relatively high in Nigeria, Kenya, and Cameroon (21-30%), and below 10% in Tunisia, Malta, and Algeria. All MRSA isolates were sensitive to vancomycin, with MICs 60%) of MRSA strains were multiresistant. There is a need to maintain surveillance and control of MRSA infections in Africa.

[Gastroduodenal peptic ulcer: descriptive study]. / Etude descriptive de la maladie ulcéreuse gastro-duodénale non compliquée.

Our goals were to describe the epidemiological, clinical and endoscopic aspects of gastrointestinal ulcer and to determine the frequency of H. pylori, we included 140 gastrointestinal peptic ulcers from January 1999 to June 2000. Diagnosis of ulcerous disease delt with the presence of an ulcer discovered during a high digestive endoscopy. Gastro-duodenal ulcers bleeding or with stenosis have been excluded as well as patients who were under anticoagulant treatment or antibiotics or under pump of protons inhibitors during the previous month. Data were collected from a unique questionnaire specifying the sociodemographic characteristic, the history of the ulcerous disease, the antecedents, the style of life, and the endoscopic findings. Five biopsies were done using sterile grips and a fast urease test and the direct exam of the smear. The prevalence of gastro-duodenal ulcers in our population of survey was 6.2%. One hundred twenty eight duodenal ulcers (91.4%), and 12 gastric ulcers (8.6%) were found. The average of age was of 37.1 years +/- 15.3. The sex ratio was 2.9. The gastro-duodenal disease had begun for more than 5 years at 40.8%. No difference in the characters of the pain and signs has been found between duodenal and gastric localization of the ulcer. Hp was associated in 91.4% (91.4% when duodenal ulcers, 88.9% when gastric ulcers and all gastric and duodenal ulcers). We conclude that gastrointestinal ulcers occupies an important place in our gastroenterological practice and the infection rate with H. pylori infection is so high during gastro-duodenal ulcers in our country that the eradication of Hp could be proposed in any case of gastro-duodenal ulcer diagnosed by endoscopy.

[Acquired bacteraemias at the intensive care unit]. / Bactériémies acquises dans un service de réanimation.

Acquired bacteraemias in intensive care unit (ICU) have some serious consequences in terms of morbidity, mortality and costs. The emergence of multiresistant germs in ICUs, and the therapeutic difficulties that ensue, participate in the aggravation of the prognosis of these infections. The aim of this work was to study the epidemiological aspects of acquired bacteraemias in ICU and the responsible germs sensitivity, to determine strategies of adequate antimicrobial treatment. During the period of study, 31 positive blood cultures collected from 19 patients were considered to be true bacteraemias, giving an incidence rate of 6.3 for 100 admissions. The mean age of our patients was 27.7 years old. There were 16 men and 3 women. Traumatology was the underlying pathology in 52.63% of cases. The origin of the infection was unknown in 84.2%. Flavobacterium spp was the most frequent germ (42%), followed by Pseudomonas aeruginosa (26.2%), and Staphylococcus negative coagulase (10.6%). Only one anaerobic germ was isolated. Flavobacterium spp and Pseudomonas aeruginosa were in general sensitive to most antibiotics used with the exception of aminosides. Staphylococci negative coagulase were methi - resistant. The sensitivity of the anaerobic germ was not tested. The antimicrobial treatment was adapted in 84.2% of cases; the association ciprofloxacine-cefotaxime could be a good alternative in serious infections to Gram negative bacteria. The global death rate among our patients was 42%. The death was directly related to bactereamia in only 15.3% of cases. The existing committee for nosocomial infections control should be more effective in our hospital. The role of this committee is to carry out microbiological surveillance, to recommand and make sure of the application of preventive measures against nosocomial infections, to promote the accessibility of antibiotics such as imipeneme, aztreonam, ceftazidime, vancomycine...., and to propose an appropriate antimicrobial treatment strategy; these measures could reduce notably the morbidity and mortality related to nosocomial infections in general and bactereamias in particular.

[Neonatal bacterial infections at the CUH of Dakar]. / Infections bactériennes néonatales au CHU de Dakar.

OBJECTIVES: It's a retrospective study in order to determine the epidemiology of neonatal bacterial infection and to evaluate the efficiency of the antibiotic protocol in University Teaching Hospital in Dakar. MATERIAL AND METHODES: From January 1st 1997 to December 31st 1998 we have registered 7461 live births, samples of blood are taken from 2312 new-born baby and they received antibiotherapy (beta-lactamine + gentamycin) at the first day based on infections risk evaluated by anamnestic criterias. The treatment is seven to one days long, the antibiotic was adapted according to the antibiogram result. RESULTS: The neonatal infection diagnosis is confirmed in 246 cases, about 33 per 1000 live births or 10.6% of newborn babies having on antibiotherapy. Most current risk factors are premature rupture of membranes (85%) and neonatal suffering (87.8%). Isolated gerras are: Klebsiella pneumoniae (61.5%), Enterobacteria (11.5 Staphylococcus (8.7%), colibacille (6%), Streptococcus (5.5%), Enterococcus (4.1%) and Pseudomonas (2.7%). Most of these germs are resistant to antibiotics currently used in first intention (ampicillin, cefotaxim, gentamycin), in particularly 95% of Klebsiella. Most efficient antibiotics are amikacin, colistin, ceftriaxon and ciprofloxacine. Deaths occurs in 48 cases with 36 in early neonatal period, 79% of mortality rate related to infection by Klebsiella. CONCLUSION: First intention antibiotherapy must be always adapted to the bacterial ecology evolution and must be more selective by using major infections risk factors. We promote early infection diagnosis by using biologic markers which reference is represented by C Reactive Protein.

Two-year surveillance of antibiotic resistance in Streptococcus pneumoniae in four African cities.

Worldwide spread of antibiotic resistance in Streptococcus pneumoniae is a major problem. However, data from West and North African countries are scarce. To study the level of resistance and compare the situations in different cities, a prospective study was conducted in Abidjan (Ivory Coast), Casablanca (Morocco), Dakar (Senegal), and Tunis (Tunisia), from 1996 to 1997. The resistances to eight antibiotics of 375 isolates were studied by E test, and the results were interpreted using the breakpoints recommended by the National Committee for Clinical Laboratory Standards. Overall, 30.4% of the isolates were nonsusceptible to penicillin G (25.6% were intermediate and 4.8% were resistant). Amoxicillin (96.3% were susceptible) and parenteral third-generation cephalosporins (92.7%) were highly active. Resistance to chloramphenicol was detected in 8.6% of the isolates. High levels of resistance were noted for erythromycin (28%), tetracycline (38.3%), and cotrimoxazole (36.4%). Resistance to rifampin was rare (2.1%). There were significant differences in resistance rates between individual countries. Multiple resistance was more frequent in penicillin-nonsusceptible isolates than in penicillin-susceptible isolates. Recommendations for treatment could be generated from these results in each participating country.

[Feasability of a heart valve bank in Dakar. (Preliminary study of 8 human hearts)]. / Faisabilité d'une banque d'homogreffes valvulaires cardiaque à Dakar. (Etude préliminaire sur 8 prélèvements de coeurs humains)

Authors record a preliminary study led so as to develope techniques of preparation of the cardiac homograft valves. Eight human hearts have been appropriated in the course of medico-legal autopsy. Under sterile hodd, the heart was dissected, aortic and pulmonary valves as well as a mitral valve fragment were collected for a total of 24 valvular levies. After sterlization in an antibiotic solution, valves were preserved at 4 degrees C for the mitral fragment specimen of fresh allograft and to - 196 degrees C in Nitrogen liquidates for the pulmonary and aortic valves. The control of graft quality had consisted in tests of competence during the dissection, to the evaluation of the histological study as well as tests of sterility. Cellular cultures had shown a fibroblast proliferation in 3 cases. It concerned an indeed reliable method but difficult and little sensitivity. Histological tests had shown two types of injurie: the myxoid degeneration (7 times on 8 valves) cryopreserved and a coagulative necrosis whose distribution was identical forthe two modes of conservation. Antibiotic's solution seemed to induce these injuries as do heart ischemia, and the traumatism lihleed to the great cold. The tests of sterility had shown a rate of contamination to 25 % essentially by Pseudomonas. In view of the installation of a behle of allograft bank of quality, we'll need to improve conditions of leavy, to use a freezer adaptation for progressive cryocongelation and the choice of simple method to evaluate valvular viabilty.

[Nasal carriage of Staphylococcus aureus at a hemodialysis unit in Dakar]. / Etude du portage nasal de Staphylococcus aureus dans une Unité d'Hémodialyse à Dakar.

Infectious diseases are the most common complications of the hemodialysis unit patients. Staphylococcus aureus (SA) is the main cause and is supposed to be lodged in the nostrils. The purpose of this study is to assess the importance of the SA presence in the patients and the medical staff nostrils in a senegaleese hemodialysis unit. In this regard, we performed a six-month prospective study on 12 patients on going hemodialysis (11 males and 1 female) and 7 medical agents (4 nurses, 2 physicians, and 1 nurse assistant). On patients we reported the initial nephropathy, the problems of vascular access, the infectious episodes prior to the study, and the duration of dialysis. Three sample series of nostril swabbing were carried out on patients and medical staff at a rate of one series every other month. Blood culture was systematically carried out on patients, with body temperature higher than 38 degrees C, every two months and. Sensibility to several antibiotics was tested for each stain. We found six cases of clinical nephroangiosclerosis, one Wegener granuloma and five unknown causes. All patients had Brescia Cimino arteriovenous fistula vascular and the mean time underdialysis was 36 months (from 7 to 55). Bacteriologically, 124 nostril specimens were performed, 80 on patients, 44 on medical staff. SA was isolated in 64 specimens (52 %) on patients and 34 specimens (24.4 %) on staff. Strains from forty five patients (70.32 %) were sensitive to methicillin versus 23 stains from staff ( 67.6 %). The difference was not statistically significant. Sensibility profil for other antibiotics was the same for patients and staff. All blood culture were negative. Despite nostril carriage noted on patients and staff, there were no infections during this period. However, the presence of permanent infection sites make it mandatory to be on the alert in order to control some of the behavioral changes of the germs, and to put into place an adequate prevention system.

[Improvement of tuberculosis diagnosis by the Mycobacteria Growth Indicator Tube (MGIT) in a developing country laboratory]. / Amélioration du diagnostic de la tuberculose par le Mycobacteria Growth Indicator tube (MGIT) dans un laboratoire de pays en développement.

In order to improve tuberculosis diagnosis in a developing country (Senegal), we evaluated a new liquid-based medium and nonradioactive system, Mycobacteria Growth Indicator Tube (MGIT), with individual clinical specimens collected in Dakar. The main purpose was to compare the time to detection and the rate of recovery of Mycobacterium tuberculosis complex and to determine its importance with respect to Lowenstein-Jensen (LJ), a liquid-based-medium for isolation of M. tuberculosis complex. 531 specimens were processed with Mycoprep kit containing NaOH-N-acetyl L-cystein and inoculated on both LJ and MGIT and incubated at 37 degrees C for 60 days. For each medium, the recovery rate and the time to detection were recorded. Among the 531 specimens, of which 121 smears were positive, 32.5% (173/531) grew the M. tuberculosis complex. Of these, 103 were smear positive (S+) and 70 smear negative (S-). LJ recovered 54.9% (95/173) and MGIT recovered 91.9% (159/173). Disagreements were observed with 92 isolates, LJ failed to recover 78 while MGIT failed to recover 14. The overall mean time to detection was 20.1 days for LJ and 10.5 days for MGIT. MGIT has shown a better sensitivity in isolation with significant reduction in reporting culture for M. tuberculosis complex. As a simple and a nonradiometric system, it could be used in conjunction with egg-based media in developing countries laboratories.

Rapid susceptibility testing of Mycobacterium tuberculosis by the Mycobacteria Growth Indicator Tube (MGIT AST SIRE).

OBJECTIVE: To evaluate the reliability of the Mycobacteria Growth Indicator Tube (MGIT AST) for susceptibility testing of Mycobacterium tuberculosis. METHODS: Seventy strains of M. tuberculosis were tested for susceptibility to streptomycin, isoniazid, rifampicin and ethambutol by comparing MGIT AST results to those obtained by the method of proportion (MOP) on Lowenstein-Jensen (LJ) and Middlebrook 7H10 media. The 7H10 MOP was considered the method of reference. RESULTS: The turnaround time for MGIT AST was 6.2 days (5-10 days) and for MOP it was 18-21 days. With rifampicin, MGIT AST agreed for all isolates with both MOP. For streptomycin, MGIT AST and 7H10 MOP agreed for 64 isolates (91.4%); 61 were susceptible and three resistant. LJ MOP and 7H10 MOP agreed for 64 isolates (92.2%); 62 were susceptible and three resistant. With isoniazid, both MOP agreed for all isolates, while MGIT AST and 7H10 MOP had two discrepancies. For ethambutol, MGIT AST and 7H10 MOP were concordant for 66 isolates; 65 were susceptible and one resistant. Both MOP were concordant for 67 isolates; 66 were susceptible and one resistant. CONCLUSIONS: Based on these results, MGIT AST is a time-saving method and can be used as an alternative to the BACTEC System. MGIT AST is reliable as far as rifampicin and isoniazid are concerned; however, additional studies are needed for streptomycin and ethambutol.

[Micromethod for identification of enterobacteria]. / Microméthodes d'identification des Enterobactéries.

The aim of this study was to set accurate and reliable methods in the identification of Enterobacteriaceae. In Micro CSB Entero each of a strip with 20 cupules containing dehydrated substrates for biochemical identification of bacterial species. Strips were inoculated with 100 microliters per cupule and incubated. After 18 hours, reagents were added. Baye's theorem was used to validate tests. Reactions from Micro CSB were cleared and easily read. 102 strains of Enterobacteriaceae were identified by MicroCSB--89.3% were correctly identified. 0.9% were identified with other tests 9.8% were incorrectly identified MicroCSB gives many advantages: This method save time, space and is less expensive than the other methods and it gives security for identifying species belonging to the different genera.