Characterization of New Races of Xanthomonas oryzae pv. oryzae in Mali Informs Resistance Gene Deployment.

Bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae represents a severe threat to rice cultivation in Mali. Characterizing the pathotypic diversity of bacterial populations is key to the management of pathogen-resistant varieties. Forty-one X. oryzae pv. oryzae isolates were collected between 2010 and 2013 in the major rice growing regions in Mali. All isolates were virulent on the susceptible rice variety Azucena; evaluation of the isolates on 12 near isogenic rice lines, each carrying a single resistance gene, identified six new races (A4 to A9) and confirmed race A3 that was previously reported in Mali. Races A5 and A6, isolated in Office du Niger and Sélingué, were the most prevalent races in Mali. Race A9 was the most virulent, circumventing all of the resistance genes tested. Xa3 controlled six of seven races (i.e., 89% of the isolates tested). The expansion of race A9 represents a major risk to rice cultivation and highlights the urgent need to identify a local source of resistance. We selected 14 isolates of X. oryzae pv. oryzae representative of the most prevalent races to evaluate 29 rice varieties grown by farmers in Mali. Six isolates showed a high level of resistance to X. oryzae pv. oryzae and were then screened with a larger collection of isolates. Based on the interactions among the six varieties and the X. oryzae pv. oryzae isolates, we characterized eight different pathotypes (P1 to P8). Two rice varieties, SK20-28 and Gigante, effectively controlled all of the isolates tested. The low association observed among races and pathotypes of X. oryzae pv. oryzae suggests that the resistance observed in the local rice varieties does not simply rely on single known Xa genes. X. oryzae pv. oryzae is pathogenically and geographically diverse. Both the races of X. oryzae pv. oryzae characterized in this study and the identification of sources of resistance in local rice varieties provide useful information to inform the design of effective breeding programs for resistance to bacterial leaf blight in Mali.

Tuberculosis specific responses following therapy for TB: Impact of HIV co-infection.

Characterizing perturbations in the immune response to tuberculosis in HIV can develop insights into the pathogenesis of coinfection. HIV+ TB+ and TB monoinfected (TB+) subjects recruited from clinics in Bamako prior to initiation of TB treatment were evaluated at time-points following initiation of therapy. Flow cytometry assessed CD4+/CD8+ T cell subsets and activation markers CD38/HLA-DR. Antigen specific responses to TB proteins were assessed by intracellular cytokine detection and proliferation. HIV+ TB+ subjects had significantly higher markers of immune activation in the CD4+ and CD8+ T cells compared to TB+ subjects. HIV+ TB+ had lower numbers of TB-specific CD4+ T cells at baseline. Plasma IFNγ levels were similar between HIV+ TB+ and TB+ subjects. No differences were observed in in-vitro proliferative capacity to TB antigens between HIV+ TB+ and TB+ subjects. Subjects with HIV+ TB+ coinfection demonstrate in vivo expansion of TB-specific CD4+ T cells. Immunodeficiency associated with CD4+ T cell depletion may be less significant compared to immunosuppression associated with HIV viremia or untreated TB infection.

Virology. Mutation rate and genotype variation of Ebola virus from Mali case sequences.

The occurrence of Ebola virus (EBOV) in West Africa during 2013-2015 is unprecedented. Early reports suggested that in this outbreak EBOV is mutating twice as fast as previously observed, which indicates the potential for changes in transmissibility and virulence and could render current molecular diagnostics and countermeasures ineffective. We have determined additional full-length sequences from two clusters of imported EBOV infections into Mali, and we show that the nucleotide substitution rate (9.6 × 10(-4) substitutions per site per year) is consistent with rates observed in Central African outbreaks. In addition, overall variation among all genotypes observed remains low. Thus, our data indicate that EBOV is not undergoing rapid evolution in humans during the current outbreak. This finding has important implications for outbreak response and public health decisions and should alleviate several previously raised concerns.

New multilocus variable-number tandem-repeat analysis tool for surveillance and local epidemiology of bacterial leaf blight and bacterial leaf streak of rice caused by Xanthomonas oryzae.

Multilocus variable-number tandem-repeat analysis (MLVA) is efficient for routine typing and for investigating the genetic structures of natural microbial populations. Two distinct pathovars of Xanthomonas oryzae can cause significant crop losses in tropical and temperate rice-growing countries. Bacterial leaf streak is caused by X. oryzae pv. oryzicola, and bacterial leaf blight is caused by X. oryzae pv. oryzae. For the latter, two genetic lineages have been described in the literature. We developed a universal MLVA typing tool both for the identification of the three X. oryzae genetic lineages and for epidemiological analyses. Sixteen candidate variable-number tandem-repeat (VNTR) loci were selected according to their presence and polymorphism in 10 draft or complete genome sequences of the three X. oryzae lineages and by VNTR sequencing of a subset of loci of interest in 20 strains per lineage. The MLVA-16 scheme was then applied to 338 strains of X. oryzae representing different pathovars and geographical locations. Linkage disequilibrium between MLVA loci was calculated by index association on different scales, and the 16 loci showed linear Mantel correlation with MLSA data on 56 X. oryzae strains, suggesting that they provide a good phylogenetic signal. Furthermore, analyses of sets of strains for different lineages indicated the possibility of using the scheme for deeper epidemiological investigation on small spatial scales.

High level of HIV-1 resistance in patients failing long-term first-line antiretroviral therapy in Mali.

OBJECTIVES: In resource-limited settings, few data are available on virological failure after long-term first-line antiretroviral therapy. This study characterized the genotypic resistance patterns at the time of failure after at least 36 months of a first-line regimen in Mali, West Africa. METHODS: Plasma samples from 84 patients who were receiving first-line antiretroviral treatment and with an HIV-1 RNA viral load (VL) >1000 copies/mL were analysed. Genotypic resistance testing was performed and HIV-1 drug resistance was interpreted according to the latest version of the National Agency for HIV and Hepatitis Research algorithm. RESULTS: At the time of resistance testing, patients had been treated for a median of 60 months (IQR 36-132 months) and had a median CD4 cell count of 292 cells/mm(3) (IQR 6-1319 cells/mm(3)), a median HIV-1 RNA level of 28266 copies/mL (IQR 1000-2 93 495 copies/mL) and a median genotypic susceptibility score of 1 (IQR 1-4). The prevalence of nucleoside reverse transcriptase inhibitor (NRTI) and non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance mutations was 78% and 82%, respectively. Viruses were resistant to at least one drug in 92% of cases. Although etravirine and rilpivirine were not used in the first-line regimens, viruses were resistant to etravirine in 34% of cases and to rilpivirine in 49% of cases. The treatment duration, median number of NRTI and NNRTI mutations and some reverse transcriptase mutations (T215Y/F/N, L210W, L74I, M41L and H221Y) were associated with the VL at virological failure. CONCLUSIONS: This study demonstrated a high level of resistance to NRTIs and NNRTIs, compromising second-generation NNRTIs, for patients who stayed on long-term first-line regimens. It is crucial to expand the accessibility of virological testing in resource-limited settings to limit the expansion of resistance and preserve second-line treatment efficacy.

Analysis of Xanthomonas oryzae pv. oryzicola population in Mali and Burkina Faso reveals a high level of genetic and pathogenic diversity.

Bacterial leaf streak (BLS) caused by Xanthomonas oryzae pv. oryzicola was first reported in Africa in the 1980s. Recently, a substantial reemergence of this disease was observed in West Africa. Samples were collected at various sites in five and three different rice-growing regions of Burkina Faso and Mali, respectively. Sixty-seven X. oryzae pv. oryzicola strains were isolated from cultivated and wild rice varieties and from weeds showing BLS symptoms. X. oryzae pv. oryzicola strains were evaluated for virulence on rice and showed high variation in lesion length on a susceptible cultivar. X. oryzae pv. oryzicola strains were further characterized by multilocus sequence analysis (MLSA) using six housekeeping genes. Inferred dendrograms clearly indicated different groups among X. oryzae pv. oryzicola strains. Restriction fragment length polymorphism analysis using the transcriptional activator like effector avrXa7 as probe resulted in the identification of 18 haplotypes. Polymerase chain reaction-based analyses of two conserved type III effector (T3E) genes (xopAJ and xopW) differentiated the strains into distinct groups, with xopAJ not detected in most African X. oryzae pv. oryzicola strains. XopAJ functionality was confirmed by leaf infiltration on 'Kitaake' rice Rxo1 lines. Sequence analysis of xopW revealed four groups among X. oryzae pv. oryzicola strains. Distribution of 43 T3E genes shows variation in a subset of X. oryzae pv. oryzicola strains. Together, our results show that African X. oryzae pv. oryzicola strains are diverse and rapidly evolving, with a group endemic to Africa and another one that may have evolved from an Asian strain.

Molecular strain typing of Mycobacterium tuberculosis complex in Bamako, Mali.

OBJECTIVE: To identify strains of Mycobacterium tuberculosis complex (MTC) circulating in Bamako and to examine the relationship between the strains and their drug susceptibility profiles. METHODS: Between 2006 and 2010, we conducted a cross-sectional study using spoligotyping to identify strains of MTC recovered from 126 tuberculosis (TB) patients under treatment in Bamako, Mali. RESULT: Three members of the MTC were isolated: M. tuberculosis (71.4%), M. africanum (27.8%) and M. bovis (0.8%). Of these, three strains were found to be the most prevalent: M. tuberculosis T1 (MTB T1; 38.9%), M. africanum F2 (MAF2; 26.2%) and M. tuberculosis Latin American and Mediterranean 10 (MTB LAM 10; 10.3%). MAF2 and MTB LAM 10 strains have a lower risk of multidrug resistance (MDR) than MTB T1 (respectively OR 0.1, 95%CI 0.03-0.4 and OR 0.1, 95%CI 0.01-0.8). Age ≥ 32 years (OR 1.4, 95%CI 0.4-3.9), negative human immunodeficiency virus status (OR 0.4, 95%CI 0.1-2.5) and male sex (OR 4, 95%CI 0.9-16.5) were not associated with MDR. The prevalence of MDR among treatment and retreatment failure patients was respectively 25% and 81.8% compared to new patients (2.9%). CONCLUSION: This study indicates a low level of primary drug resistance in Bamako, affirms the importance of using correct drug regimens, and suggests that the MTB T1 strain may be associated with the development of resistance.

A seroepidemiological study of pandemic A/H1N1(2009) influenza in a rural population of Mali.

The swine-origin H1N1 influenza A virus (pH1N1(2009)) started to circulate worldwide in 2009, and cases were notified in a number of sub-Saharan African countries. However, no epidemiological data allowing estimation of the epidemic burden were available in this region, preventing comprehensive comparisons with other parts of the world. The CoPanFlu-Mali programme studied a cohort of 202 individuals living in the rural commune of Dioro (southern central Mali). Pre-pandemic and post-pandemic paired sera (sampled in 2006 and April 2010, respectively) were tested by the haemagglutination inhibition (HI) method. Different estimates of pH1N1(2009) infection during the 2009 first epidemic wave were used (increased prevalence of HI titre of ≥1/40 or ≥1/80, seroconversions) and provided convergent attack rate values (12.4-14.9%), the highest values being observed in the 0-19-year age group (16.0-18.4%). In all age groups, pre-pandemic HI titres of ≥1/40 were associated with complete absence of seroconversion; and geometric mean titres were <15 in individuals who seroconverted and >20 in others. Important variations in seroconversion rate existed among the different villages investigated. Despite limitations resulting from the size and composition of the sample analysed, this study provides strong evidence that the impact of the pH1N1(2009) first wave was more important than previously believed, and that the determinants of the epidemic spread in sub-Saharan populations were quite different from those observed in developed countries.

Confirmation of immunogenic consensus sequence HIV-1 T-cell epitopes in Bamako, Mali and Providence, Rhode Island.

The design of epitope-driven vaccines for HIV has been significantly hampered by concerns about conservation of vaccine epitopes across clades of HIV. In previous work, we have described a computer-driven method for a cross-clade HIV vaccine comprised of overlapping, highly conserved helper T-cell epitopes or "immunogenic consensus sequence epitopes" (ICS epitopes). Here, we evaluated and compared the immunogenicity of 20 ICS HIV epitopes in ELISpot assays performed using peripheral blood monocytes (PBMC) from HIV-infected donors in Providence, Rhode Island, USA and in Bamako, Mali, West Africa. Each core 9-mer HIV sequence contained in a given consensus peptide was conserved in at least 105 to as many as 2,250 individual HIV-1 strains. Nineteen of the 20 ICS epitopes (95%) were confirmed in ELISpot assays using PBMC obtained from 13 healthy, HIV-1 infected subjects in Providence, and thirteen of the epitopes (65%) were confirmed in ELISpot assays using PBMC derived from 42 discarded blood units obtained at the Central Blood Bank in Bamako. Twelve of the epitopes were confirmed in ELISpot assays performed both in Providence and Bamako. These data confirm the utility of bioinformatics tools to select and design novel vaccines containing "immunogenic consensus sequence" T-cell epitopes for a globally relevant vaccine against HIV; a similar approach may also be useful for any pathogen that exhibits high variability (influenza, HCV, or variola for example). An HIV vaccine containing these immunogenic consensus sequences is currently under development.

[Reinfection with Schistosoma haematobium and mansoni despite repeated praziquantel office treatment in Niger, Mali]. / Réinfections par Schistosoma haematobium et mansoni à l'office du Niger au Mali malgré la prise répétée de praziquantel.

The dynamics of reinfection by Schistosoma haematobium and Schistosoma mansoni after repeated treatment with praziquantel (40 mg/kg body weight, single dose) was studied in a cohort of schoolchildren living in an endemic area. A total of 214 urine and 220 stool samples were collected and examined at three different times, i.e., February 1989, July 1989 and February 1990. Mass chemotherapy was administered at the beginning of study (February 89). Treatment was repeated in children with positive tests at each subsequent sampling. Prevalence rates were 55.1 p. 100, 3.7 p. 100, and 35.0 p. 100 for Schistosoma haematobium and 62.7 p. 100, 46.3 p. 100 and 73.1 p. 100 for Schistosoma mansoni in February 1989, July 1989 and February 1990 respectively (p < 0.001). From July 1989 to February 1990, reinfection was observed in 84.5 p. 100 of children by Schistosoma haematobium versus 57.8 p. 100 by Schistosoma mansoni. The risk of reinfection by Schistosoma haematobium was higher in children between the ages of 7 and 10 years than in children between the ages of 11 and 15 years (p < 0.001), The incidence of intense Schistosoma haematobium egg excretion rose from 0 p. 100 in July 1989 to 6.0 p. 100 in February 1990. The incidence of intense Schistosoma mansoni excretion in February 1990 was 4.5 p. 100. The reinfection rate at 7 months was over 50 p. 100 for both parasite species despite repeated treatment. This finding demonstrates that additional measures such as proper sanitation and vector control are needed to control human schistosomiasis in irrigated rice paddies.

Recognition of synthetic 104-mer and 102-mer peptides corresponding to N- and C-terminal nonrepeat regions of the Plasmodium falciparum circumsporozoite protein by sera from human donors.

In the present work, we analyze the recognition of synthetic polypeptides encompassing the aminoterminal (amino acids 22-125) and the carboxy terminal (289-390) regions of the circumsporozoite (CS) protein of Plasmodium falciparum by sera from donors living in endemic area of South America and Africa. Two populations were studied: one on the Colombian Pacific coast, with low endemicity for malaria; and a western African village exposed to a very intense transmission of P. falciparum. Antibodies directed to the two polypeptides were found at high titers in both populations. Furthermore, this response was observed in individuals lacking antibodies to the highly repetitive central sequence of the CS protein (NANP). The epitopes responsible for this recognition were mapped to the region 81-125 and 316-346 of the N- and C-termini, respectively. When the two populations were compared, both showed high titers of antibodies to the two flanking peptides. However, while 95% of the sera from African adults showed antibodies against the repeat region of the CS protein, only 37% of the Colombian adults studied had these antibodies. Furthermore, African donors of various ages exhibited different patterns of recognition of the two polypeptides. In African children less than five years of age, antibodies were found in comparable levels to Colombian adults; however, in older African donors, the response to NANP became dominant. These findings may reflect the skewing effect of the humoral response towards the central repetitive epitope under conditions of frequent exposure to malaria infections. The production of such polypeptides encompassing regions that contain multiple epitopes for antibodies, T helper, and cytotoxic T lymphocyte epitopes would be advantageous in the generation of new and more efficient malaria vaccines.

Analysis of the human antibody response to thrombospondin-related anonymous protein of Plasmodium falciparum.

Thrombospondin-related anonymous protein (TRAP) of the malaria parasite Plasmodium falciparum shares two sequence motifs with other proteins which possess adhesive properties. Recently, findings indicate that TRAP is an antigen which contributes to antisporozoite immunity. We have cloned and expressed the TRAP coding sequences in Escherichia coli to investigate the human humoral immune response against this protein in a region of malaria endemicity of West Africa characterized by a seasonal transmission. Our results show that antibodies against TRAP are present in infected individuals. The anti-TRAP antibodies were analyzed in both a longitudinal and a prospective study. The longitudinal analysis shows seasonal fluctuations of the levels of specific antibodies as well as age-dependent quantitative differences. The immune response is long-lived in most of the adults and some of the older children but short-lived in young children. More importantly, the prospective analysis suggests that the presence of anti-TRAP antibodies in older children before the beginning of malaria transmission correlates with the subsequent control of parasite densities.

A prospective study of the association between the human humoral immune response to Plasmodium falciparum blood stage antigen gp190 and control of malarial infections.

The human humoral immune response to the Plasmodium falciparum merozoite surface antigen gp190 was analyzed to determine the rate of reinfection by the parasite and the ability to control parasite density. The prospective study was carried out in a West African village where malaria is hyperendemic. No correlation between the antibody titers and protection against infection was observed within the group of children. Positive and negative associations of antibody specificities with protection against and/or control of parasitemia were, however, found for adolescents and adults, respectively. Thus, in adolescents, the presence of antibodies to gp190 fragment M6 correlates with a 50% reduced risk of P. falciparum infection and an increased ability to control parasitemia, whereas in adults, the humoral response to some of the polymorphic regions of gp190 associates with an increased risk of infection.

[Incidence of malaria and S hemoglobinopathy in the pediatric hospital milieu in Bamako, Mali]. / Incidence du paludisme et hémoglobinose S en milieu hospitalier pédiatrique bamakois au Mali.

A study on the incidence and gravity of malaria among children suffering of sickle cell anemia was carried out in the pediatric hospital of Bamako-Mali. On the 236 cases of fever studied, 54.2% were malaria infected and 21.2% were carriers of the sicklemic trait. There was a significant difference between malaria affecting AA children and HbS children, specially concerning parasites load.

Human antibody response to the major merozoite surface antigen of Plasmodium falciparum is strain specific and short-lived.

The precursor of the major merozoite antigen of Plasmodium falciparum, gp190, is considered a candidate for inclusion in a malaria vaccine. This protein, which consists of conserved, dimorphic, and polymorphic sequences, is very immunogenic in humans. In a longitudinal study carried out with 94 inhabitants of a rural community in Mali, West Africa, we show that in this endemic area naturally acquired gp190-specific antibodies are predominantly directed against the dimorphic parts of one of the main alleles of gp190. The presence of antibodies against these dimorphic regions correlates with the prevalence of the corresponding antigen in the infecting parasite population. Moreover, qualitative as well as quantitative differences were found in the time course of the humoral immune response to the dimorphic regions in adults and children, who differ in their susceptibility to malaria infection.

[Impact of curtains and blankets impregnated with permethrin on the malarial indicators and the number of malarial attacks per child in a village in an area hyperendemic for malaria on the Malian savannah (preliminary results of the first year study)]. / Impact des rideaux et couvertures imprégnés de perméthrine sur les indices paludométriques et le nombre d'accès palustres par enfant dans un village d'hyperendémie palustre de savane Malienne. Résultats préliminaires de la 1re année d'étude.

From May 1989 to June 1990 an experimental study has been undertaken in two village's malarial hyperendemy of the Malian savannah. This study's objective was to test the effectiveness, on paludometric parameters and the openness of the population to the impregnated curtains and blankets, of permethrin as strategy of controlling malaria. The experimental village (Tiénéguébougou) and the control one (Kambila) are located in the same ecosystem and as at least 3 km any from each other. The impregnated curtains were installed in all the experimental village's houses and all the 216 residents have received impregnated blankets. The entomological, parasitical and clinical parameters were measured monthly from may 1989 to november 1989. The impregnated curtains were accepted by the population. However the impregnated blankets were little successful. Their impact on the entomological parameters is remarkable with a reduction of more than 98% of ma and the rate of entomological inoculation (REI) in the houses. The reduction of parasitical index is much less perceptible. The effect on the parasitical pressure was manifested by a highly significant difference of splenic index between the two villages (lower in the control village). The main vectors belong to complex: A. gambiae s.l. which rate of parity was not affected by the impregnated material. Very efficient against the harmfulness of in house hematophagic insects, the curtains impregnated by pyrethroids are not sufficient for interrupting the transmission of malaria. Actually, the study is continued for 2 years with a best evaluation of the malarial morbidity.