Isolation and partial characterization of bacteria (Pseudoalteromonas sp.) with potential antibacterial activity from a marine costal environment from New Caledonia.

UNLABELLED: Marine bacteria are a rich source of bioactive metabolites. However, the microbial diversity of marine ecosystem still needs to be explored. The aim of this study was to isolate and characterize bacteria with antimicrobial activities from various marine coastal environment of New Caledonia. We obtained 493 marine isolates from various environments and samples of which 63 (12.8%) presented an antibacterial activity against a panel of reference pathogenic strains (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli and Enterococcus faecalis). Ten out of the most promising strains were cultured, fractionated and screened for antibacterial activity. Four of them (NC282, NC412, NC272 and NC120) showed at least an activity against reference and multidrug-resistant pathogenic strains and were found to belong to the genus Pseudoalteromonas, according to the 16S phylogenetic analysis. The NC282 strain does not belong to any described Pseudoalteromonas species and might be of interest for further chemical and biological characterization. These findings suggest that the identified strains may contribute to the discovery for new sources of antimicrobial substances to develop new therapies to treat infections caused by multidrug-resistant bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: With the constant increasing of bacterial resistance against known antibiotics in worldwide public health, it is now necessary to find new sources of antimicrobials. Marine bacteria from New Caledonia were isolated, tested for antibacterial activity and characterized to find new active molecules against multidrug-resistant bacteria. This study illustrates the diversity of the marine ecosystem with potent new bacteria species. Also the potential of marine bacteria as a rich source of bioactive molecule, for example antibiotics, is highlighted.

Partial characterization of an exopolysaccharide secreted by a marine bacterium, Vibrio neocaledonicus sp. nov., from New Caledonia.

AIMS: Exopolysaccharides (EPS) are industrially valuable molecules with numerous useful properties. This study describes the techniques used for the identification of a novel Vibrio bacterium and preliminary characterization of its EPS. METHODS AND RESULTS: Bioprospection in marine intertidal areas of New Caledonia followed by screening for EPS producing brought to selection of the isolate NC470. Phylogenetic analysis (biochemical tests, gene sequencing and DNA-DNA relatedness) permitted to identify NC470 as a new member of the Vibrio genus. The EPS was produced in batch fermentation, purified using the ultrafiltration process and analysed by colorimetry, Fourier Transform Infrared spectroscopy, gas chromatography, Nuclear Magnetic Resonance and HPLC-size exclusion chromatography. This EPS exhibits a high N-acetyl-hexosamines and uronic acid content with a low amount of neutral sugar. The molecular mass was 672 × 10(3)  Da. These data are relevant for possible technological exploitation. CONCLUSIONS: We propose the name Vibrio neocaledonicus sp. nov for this isolate NC470, producing an EPS with an unusual sugar composition. Comparison with other known polymers permitted to select applications for this polymer. SIGNIFICANCE AND IMPACT OF THE STUDY: This study contributes to evaluate the marine biodiversity of New Caledonia. It also highlights the biotechnological potential of New Caledonia marine bacteria.

Outbreak of leptospirosis in New Caledonia: diagnosis issues and burden of disease.

A leptospirosis epidemic affected New Caledonia during the first semester of 2008. A total of 135 cases were diagnosed with a relatively low fatality rate of 3.7%. Heavy rainfalls, related to La Niña, favoured this epidemic. The PCR, routinely used, confirmed 54% of the cases, and the microagglutination test 56%. Epidemiological and economical data on this epidemic are presented and discussed.

Field evaluation of rapid diagnostic tests for meningococcal meningitis in Niger.

OBJECTIVE: To evaluate dipstick rapid diagnostic tests (RDTs) for meningococcal meningitis in basic health facilities. METHODS: Health facility staff received a one-day training. During the meningitis season, they performed RDTs on cerebrospinal fluid (CSF) specimens from suspected cases of meningitis. A frozen aliquot of CSF was later tested using polymerase chain reaction (PCR) to establish the reference diagnosis. RDTs used in health facilities were archived to allow checking the concordance between reported diagnosis and observed results. Reported diagnosis was also compared to PCR diagnosis. A second RDT was performed on each CSF specimen at the reference laboratory. RESULTS: Using RDTs, health facilities reported 382 negative results (73.9%), 114 NmA (22.1%), 12 NmW135 (2.3%) and nine uninterpretable results (1.7%), the latter corresponding to the misuse of a reagent by three agents. The agreement between reported diagnosis and archived dipsticks was excellent (kappa = 0.98). The agreement between PCR diagnosis and reported RDTs results was strong (kappa = 0.82). In health facilities, the sensitivity of RDTs for N. meningitidis A was Se = 0.91. The kappa coefficient measuring the agreement between RDTs operated in the reference laboratory and RDTs operated in health facilities was kappa = 0.78. CONCLUSION: We confirmed that dipstick RDTs to identify N. meningitidis serogroups A, C, W135 and Y can be reliably operated by non-specialized staff in basic health facilities. RDTs proved very useful to recommend vaccination in NmA epidemics, and also to avoid vaccination in epidemics due to serogroups not included in vaccines (NmX).

[ Bancroftian lymphatic filariasis: toward its elimination from the Pacific?]. / Filariose lymphatique de Bancroft: vers l'elimination dans le Pacifique?

The region of the Pacific is historically affected by lymphatic filariasis (LF). Following the World Health Assembly resolution in 1997, the Global Program to Eliminate Lymphatic Filariasis (GPELF) was launched. In the Pacific, the World Health Organization (WHO) has implemented from 1999, the Pacific Program to Eliminate Lymphatic Filariasis (PacELF) bringing together the 22 countries and territories, in a common effort to eliminate the disease. The strategy is based on Mass Drug Administration (MDA); in annual single dose during 5 years of a Diethycarbamazine/Albendazole association distributed to all the population at risk. Among the 22 countries and territories of the Pacific, 16 are endemic and 6 are non endemic. The classification is based according to the filarian antigen prevalence upper or lower than 1%. MDA are decided when the rate of the filarian antigen prevalence is > 1%. The objective of PacELF is to reduce this rate down to < 1%, threshold under which the transmission is supposed to be stopped. From 1999 to 2007, 14 of the 16 endemic countries organized MDA. Eleven of them completed the cycle of 5 treatments and even beyond. But, these MDA reached only 19% of the at risk population, because of logistic difficulties in Papua New Guinea, the most populated country in the Pacific. The investigations carried out in sentinel sites showed a public health impact, by the fall of the number of microfilaria carriers, often down to a rate < 1%. However the rate of circulating antigen prevalence remains often above the required threshold of 1%. Prevalence surveys carried out in 2007, in different endemic countries, revealed the necessity to intensify efforts and to refine strategy for elimination of FL from the Pacific. A lot of progress were obtained, but few problems were identified. Reflexions are imperative and in progress about: the MDA coverage rates while at the same time a certain lassitude appears in the populations and among health staff, the methods to evaluate the effectiveness of MDA, the reliability of the diagnostic tools to decide of the stop of MDA and to certify the absence of the transmission, the relevance of univocal biological criteria for the whole Pacific area, the need for an active surveillance during several years after stopping MDA, particularly in the countries affected by the very efficient vector Aedes polynesiensis. Seven years after its launching, despite undeniable success, the PacELF program did not achieve its very ambitious goal of stopping the transmission. Three years before its term, strong efforts have to be done and additional strategies be implemented. However; it is reasonable to expect the prolongation of the program in order to achieve the final objective. Beyond, in some countries, it will be still necessary to ensure a sustained global drug pressure and an active surveillance to prevent the re-emergence of the disease.

HIV seroprevalence surveys in Nigerien pregnant women: a comparison between 2002 and 2006.

In the framework of HIV serosurveillance, serosurveys in pregnant women are a good alternative to population-based surveys, which are more difficult to implement. In 2002 and 2006, surveys were conducted in Niger to assess the HIV seroprevalence in pregnant women and to evaluate the trend of the HIV epidemic. The overall seroprevalence was 0.96% (95% CI 0.5-1.7%) and 1.3% (95% CI 0.9-1.8%) in 2002 and 2006, respectively, showing no significant change. In the 2006 overall sample, women living in urban areas were significantly more infected than those in rural environments, with prevalences of 1.9% and 0.7%, respectively (P=0.006). Women with higher school attainment were more often HIV-positive than other women (4.6% vs. 1.7%; P<0.001). The 2006 prevalence, which is among the lowest of the sub-Saharan region, was not significantly different from the national seroprevalence measured in adults in 2002 (0.87%, 95% CI 0.5-1.3%). Close monitoring of the epidemic must be continued.

High sensitivity and specificity of the Pastorex latex agglutination test for Neisseria meningitidis serogroup A during a clinical trial in Niger.

There is a great need for a rapid diagnostic test to guide vaccine choice during outbreaks of meningococcal meningitis in resource-poor countries. During a randomised clinical trial conducted during an epidemic of Neisseria meningitidis serogroup A in Niger in 2003, the sensitivity and specificity of the Pastorex latex agglutination test for this serogroup under optimal field conditions were assessed, using culture and/or PCR as the gold standard. Results from 484 samples showed a sensitivity of 88% (95% CI 85-91%) and a specificity of 93% (95% CI 90-95%). Pastorex could be a good alternative to current methods, as it can be performed in a local laboratory with rapid results and is highly specific. Sensitivity can be improved with prior microscopy where feasible. A study specifically to evaluate the Pastorex test under epidemic conditions, using laboratories with limited resources, is recommended.

[Prevalence of tuberculous meningitis in Niamey's Hospital, Niger]. / Prevalence de la méningite tuberculeuse a l'hôpital de Niamey, Niger.

Tuberculosis is hyperendemic in Niger. In Niamey between June 2002 and May 2004, 996 cerebro-spinal fluids (CSF) collected from meningitis suspected patients have been analysed by PCR for the detection of Neisseria meningitidis, Haemophilus influenzae and Streptococcus pneumoniae: the aetiologic diagnosis was obtained for 208 cases (20.9%). The Mycobacterium tuberculosis PCR assay performed on the negative samples was positive for 4 CSF: 0.4% prevalence among suspected cases of meningitis or 1.9% among confirmed bacterial meningitis.

Evaluation of the Pastorex meningitis kit for the rapid identification of Neisseria meningitidis serogroups A and W135.

The recent emergence of Neisseria meningitidis W135 as a cause of epidemic bacterial meningitis and the availability of a trivalent ACW135 vaccine have created a need for accurate and timely meningococcal serogroup determination for organization of epidemic vaccine response. The sensitivity and specificity of the Pastorex meningitis kit (Bio-Rad) to identify serogroups A and W135 in the African meningitis belt was assessed using PCR testing as the gold standard. The sensitivity and specificity for serogroups A and W135 were 87 and 85%, respectively, while the specificities were 93 and 97%. The positive and negative likelihood ratios for A were 12 and 0.14 and for W135 were 33 and 0.16. The positive and negative predictive values, computed to simulate an epidemic of meningococcal meningitis with an estimated 70% prevalence of N. meningitidis among suspected cases, were 97% and 75% for A and 99% and 73% for W135. In remote locations of the African meningitis belt, latex agglutination is the only currently available test that can rapidly determine meningococcal serogroup. This study showed that latex agglutination performs well and could be used during the epidemic season to determine appropriate vaccine response.

[From genomics to surveillance, prevention and control: new challenges for the African meningitis belt]. / Ceinture africaine de la méningite: de la génomique aux stratégies de surveillance, de lutte et de prévention.

An international conference was held in Niamey, Niger, in November 2005. It aimed at reviewing the current situation in the meningitis belt. This region stretches from Senegal to Ethiopia and is characterized by high levels of seasonal endemicity with large epidemics of meningococcal meningitis occurring cyclically, generally caused by N. meningiditis serogroup A. WHO currently recommends a reactive strategy based on rapid detection of epidemics, intervention with antibiotics to treat cases and mass vaccination with a meningococcal polysaccharide vaccine to halt the outbreak. Epidemiological patterns of the disease in Africa have been changing with the occurrence of outbreaks outside the meningitis belt and with the emergence of serogroup W135, which first caused an epidemic among Hajj pilgrims in 2000 and then a large-scale meningitis outbreak in Burkina Faso in 2002. Consequently enhanced laboratory surveillance and confirmation of the strain responsible for the outbreak are required. New rapid dipstick tests have been developed through a collaboration between Institut Pasteur and CERMES. They are designed for bedside diagnosis and detect meningococcal antigens present in CSF using immunochromatography. The treatment of meningococcal meningitis during epidemics is based on short-course, long-acting oily chloramphenicol. An alternative is the use of ceftriaxone, which is equally effective and can be used in pregnant women and infants. A low-cost, monovalent serogroup A meningococcal conjugate vaccine for large-scale use in Africa is under development. In spite of the emergence of W135 strains in the meningitis belt, N. meningiditis A continues to be the principal strain isolated during the epidemic seasons and elimination of outbreaks of N. meningiditis serogroup A can still be considered as the primary objective of a preventive vaccination strategy.

[Assessment of five rapid tests and two algorithms for the HIV infection diagnosis in Niger]. / Evaluation de cinq tests rapides et de deux algorithmes pour le diagnostic de l'infection par le VIH au Niger.

Niger is a Sahelian country of 1,267,000 km2 and 11 M inhabitants (84% living in rural area). The national seroprevalence of the HIV infection is low (2.08% in urban area and 0. 64% in rural area). Because of limited resources, laboratories and qualified personnel are scarce, so the classical strategy using ELISA methods to diagnose HIV is available only in the capital Niamey. In order to define a strategy suitable to the Nigerian field conditions, we have determined the diagnostic value of five commercialised rapid assays (Génie II, InoCheck, Determine, Capillus and Doublecheck). A panel of plasma, whole blood and dried blood spot from 42 positive and 160 negative control individuals was tested and the sensitivity and specificity of each assay were calculated. The two best algorithms using two tests were selected: a algorithm discriminating HIV1 & 2 combining Determine and Genie II showing an excellent sensitivity and specificity (100%) but with many practical constraints; and a non discriminative algorithm combining Determine and Double-check less specific but workable on capillary blood and fully compatible with the field conditions. Its negative predictive value was 100% and its positive predictive value was > 97% for urban population and > 91% for rural population. Dried blood spots on filter paper proved to be very practical and efficient to use by the reference laboratory for the quality control of the peripheral diagnostic centres.

Molecular epidemiology of meningococci isolated in Niger in 2003 shows serogroup A sequence type (ST)-7 and serogroup W135 ST-11 or ST-2881 strains.

In 2003, in the Zinder and Maradi regions (Niger), epidemics were due to serogroup A:4:P1.9 meningococci belonging to sequence type 7 (ST-7). In Niamey, only sporadic cases were reported: 55% of the meningococcus strains were in serogroup A, and 38% were in serogroup W135 and could be placed in ST-11, identical to the 2002 Burkina Faso epidemic clone, and in ST-2881, a new ST.

Nationwide HIV prevalence survey in general population in Niger.

A national population-based survey was carried out in Niger in 2002 to assess HIV prevalence in the population aged 15-49 years. A two-stage cluster sampling was used and the blood specimens were collected on filter paper and tested according to an algorithm involving up to three diagnostic tests whenever appropriate. Testing was unlinked and anonymous. The refusal rate was 1.1% and 6056 blood samples were available for analysis. The adjusted prevalence of HIV was 0.87% (95% CI, 0.5-1.3%) and the 95% CI of the estimated number of infected individuals was 22 864-59 640. HIV-1 and HIV-2 represented, respectively, 95.6% and 2.9% of infections while dual infections represented 1.5%. HIV positivity rate was 1.0% in women and 0.7% in men. It was significantly higher among urban populations than among rural ones (respectively, 2.1% and 0.6%, P < 10(-6)). Using logistic regression, the variables significantly related to the risk of being tested positive for HIV were urban housing, increasing age and being either widowed or divorced. The estimate from the national survey was lower than the prevalence assessed from antenatal clinic data (2.8% in 2001). In the future, the representativeness of sentinel sites should be improved by increasing the representation of rural areas accounting for more than 80% of the population. Compared with other sub-Saharan countries, the HIV prevalence in Niger is still moderate. This situation represents a strong argument for enhancing prevention programmes and makes realistic the projects promoting an access to potent antiretroviral therapies for the majority.

Use of dipsticks for rapid diagnosis of cholera caused by Vibrio cholerae O1 and O139 from rectal swabs.

We evaluated the recently developed dipsticks for the rapid detection of Vibrio cholerae serotypes O1 and O139 from rectal swabs of hospitalized diarrheal patients after enrichment for 4 h in alkaline peptone water. The sensitivity and specificity of the dipsticks were above 92 and 91%, respectively. The dipsticks represent the first rapid test which has been successfully used to diagnose cholera from rectal swabs, and this would immensely improve surveillance for cholera, especially in remote settings.

Serodiagnosis of human plague by a combination of immunomagnetic separation and flow cytometry.

BACKGROUND: Plague is a severe, highly communicable bacterial disease caused by Yersinia pestis. It is still endemic in more than 20 countries worldwide. Although known as a devastating disease for centuries, laboratory confirmation of clinical suspected cases is still problematic. No standardized and internationally approved test system is commercially available. The aim of this study was the introduction and evaluation of a combination of immunomagnetic separation and flow cytometry for the serodiagnosis of human plague. METHODS: Paramagnetic polystyrene beads were coated with purified F1 capsular antigen (F1 CA) and reacted with sera from plague patients, from 26 laboratory personnel vaccinated against plague and from 102 healthy blood donors (HBD). After incubation with fluorescein isothiocyanate-conjugated anti-human rabbit IgG, particle-associated fluorescence was detected by flow cytometry. RESULTS: Anti-F1 CA antibodies could be demonstrated in all patients with bacteriologically confirmed plague and in 22 sera (84.6%) from vaccinees. Only one serum in the HBD group showed a weakly positive reaction. The total assay time was less than 2 h. CONCLUSIONS: Compared with a recently published combination of an anti-F1 CA enzyme-linked immunosorbent assay (ELISA) and immunoblot, the new assay showed the same sensitivity as the ELISA and almost the same specificity (99.0 versus 100%) as the immunoblot. Allowing a rapid, reliable, and quantitative analysis, immunomagnetic separation combined with flow cytometry might replace both conventional immunoassays.

One-step immunochromatographic dipstick tests for rapid detection of Vibrio cholerae O1 and O139 in stool samples.

We describe the development and evaluation of a rapid diagnostic test for Vibrio cholerae O1 and O139 based on lipopolysaccharide detection using gold particles. The specificity ranged between 84 and 100%. The sensitivity of the dipsticks ranged from 94.2 to 100% when evaluated with stool samples obtained in Madagascar and Bangladesh. The dipstick can provide a simple tool for epidemiological surveys.

Bacteriological follow-up of tuberculosis treatment: a comparative study of smear microscopy and culture results at the second month of treatment.

SETTING: Significance of a positive bacillary examination of sputum at 2 months of treatment in relation to the viability of the bacilli and the final treatment result. OBJECTIVE: To compare the results of smear microscopy and sputum culture at the second month of tuberculosis treatment and to follow the progress of the patients. METHODS: Follow-up of 297 patients with smear-positive pulmonary tuberculosis in Madagascar, 152 of whom were smear-positive at 2 months of treatment and 145 smear-negative. The number of bacilli was recorded, as were the culture results and the final outcome of treatment. RESULTS: Among the 152 patients who were smear-positive at the second month, 77 (51%) were culture-negative; there were 12 (8%) treatment failures and four relapses (4.6%). Among the 145 smear-negative patients, 22 (15%) were culture-positive, of which one was a treatment failure (1%). CONCLUSION: The majority of failures and relapses were observed in the group of smear-positive patients. It is important to reinforce the surveillance of these patients in order to reduce the number lost to follow-up. Furthermore, a positive smear microscopy at the end of the second month is not sufficiently specific for early identification of treatment failures. It is preferable to wait until the fifth month, as the great majority of patients who are positive at 2 months achieve cure. The treatment strategy currently recommended in Madagascar is satisfactory.