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The restriction fragment length polymorphism of the 195 bp repeated DNA sequence of #Trypanosoma cruzi$ was analyzed among 23 #T. cruzi$ stocks giving a reliable picture of the whole phylogenetic variability of the species. The profiles observed with the enzymes Hinf1 and Hae3 were linked together and supported the existence of two groups. Group 1 shows a 195 bp repeated unit (Hinf 1) and high molecular weight DNA (Hae 3), while group 2 presents a ladder profile for each enzyme, which is a characteristic of tandemly repeated DNA. The two groups, respectively, clustered stocks pertaining to the two principal lineages evidenced by izoenzyme and RAPD markers. The congruence among these three independent genomic markers corroborates the existence of two real phylogenetic lineages in #T. cruzi$. The specific monomorphic profiles for each major phylogenetic lineage suggest the existence of ancient sexuality and cryptic biological speciation. (Résumé d'auteur)

The Anopheles genus is probably one of the best studied genera among insects of medical importance. Of more than 500 species currently listed in the world about sixty species are vectors of malaria agents and about thirty species are responsible for most of the transmission [1-3]. The important epidemiological role of anopheline species has motivated many studies of taxonomy and systematics with traditional tools. With the advent of molecular tools, the development of informatics databases and new mathematical concepts on shape, characteriz‐ ing insects has become more and more accurate. Molecular tools based on the nucleotide polymorphism of DNA have allowed the identification of cryptic diversity, confirming and refining previous findings suggesting the existence of many species groups or complexes of sibling species. The development of international genetic sequence database collabora‐ tionshttp://www.insdc.org, http://www.barcodinglife.com/ allowed the use of reference sequences for species identification. Although not well developed, the same need for infor‐ matic databases arose for traditional taxonomy.

BackgroundAnopheles gambiae, a major vector of malaria, is widely distributed throughout sub-Saharan Africa. In an attempt to eliminate infective mosquitoes, researchers are trying to develop transgenic strains that are refractory to the Plasmodium parasite. Before any release of transgenic mosquitoes can be envisaged, we need an accurate picture of the differentiation between the two molecular forms of An. gambiae, termed M and S, which are of uncertain taxonomic status.Methodology/principal findingsInsertion patterns of three transposable elements (TEs) were determined in populations from Benin, Burkina Faso, Cameroon, Ghana, Ivory Coast, Madagascar, Mali, Mozambique, Niger, and Tanzania, using Transposon Display, a TE-anchored strategy based on Amplified Fragment Length Polymorphism. The results reveal a clear differentiation between the M and S forms, whatever their geographical origin, suggesting an incipient speciation process.Conclusions/significanceAny attempt to control the transmission of malaria by An. gambiae using either conventional or novel technologies must take the M/S genetic differentiation into account. In addition, we localized three TE insertion sites that were present either in every individual or at a high frequency in the M molecular form. These sites were found to be located outside the chromosomal regions that are suspected of involvement in the speciation event between the two forms. This suggests that these chromosomal regions are either larger than previously thought, or there are additional differentiated genomic regions interspersed with undifferentiated regions.

Two indigenous bradyrhizobia strains displaying different natural behaviours towards water regime (strain ORS 3257, nodulating more frequently in favourable-water conditions and strain ORS 3260, in limited-water conditions) were studied for their competitivity for nodulation of cowpea (Mouride cultivar) under favourable and limited water conditions in non-sterile soil. The nodule occupancy was studied by PCR-RFLP analysis. Both strains showed good competition with other indigenous rhizobia populations under favourable- and limited-water conditions. Competition between the inoculated strains in the mixture varied between water regimes. In non-limited-water conditions, strain ORS 3257 was the best competitor, whereas in limited-water conditions, strain ORS 3260 was the best competitor. Results indicated that screening of strains according to their environmental origin could ensure successful rhizobia Inoculation.

Cassava bacterial blight, caused by Xanthomonas axonopodis pv. manihotis (Xam), is a widespread disease that affects cassava (Manihot esculenta Crantz). Studies on the pathogen population structure, pathogen diagnosis, identification and expression of plant genes involved in resistance have been carried out. Different molecular techniques were developed to assess the genetic diversity among the Xampopulations. Characterization of Xam population dynamics over time had enable us to determine the different factors that are associated with resistance breakdown and those that influence the genetic structure or virulence phenotypes of the pathogen's population. Methods for detecting the pathogen in vegetative planting materials and true seeds were developed and contributed to reduce the impact of the disease. To better understand the genetics of resistance a quantitative trait loci (QTLs) approach was developed. Using a PCR-based strategy with degenerate primers we isolated two resistance gene candidates in cassava. We also characterized a region of a chromosome rich in R-gene like sequence. In this review we also report the main results obtained by transcript profiling methodologies, cDNA-AFLP and ESTs developed by the authors to characterize the genes involved in disease resistance. All together these techniques allowed the identification of molecular markers either associated to CBB resistance or that may represent putative genes involved in disease resistance. This article reviews current knowledge on the molecular cassava-Xam interactions.

www.springer-ny.com; International audience; In order to promote genome research on coffee trees, one of the most important tropical crops, a bacterial artificial chromosome (BAC) library of the coffee allotetraploid species, Coffea arabica, was constructed. The variety IAPAR 59, which is widely distributed in Latin America and exhibits a fair level of resistance to several pathogens, was chosen. High-efficiency BAC cloning of the high molecular weight genomic DNA partially digested by HindIII was achieved. In total, the library contains 88,813 clones with an average insert size of 130 kb, and represents approximately eight C. arabica dihaploid genome equivalents. One original feature of this library is that it can be divided into four sublibraries with mean insert sizes of 96, 130, 183 and 210 kb. Characterisation of the library showed that less than 4.5% of the clones contained organelle DNA. Furthermore, this library is representative and shows good genome coverage, as established by hybridisation screening of high-density filters using a number of nuclear probes distributed across the allotetraploid genome. This Arabica BAC library, the first large-insert DNA library so far constructed for the genus Coffea, is well-suited for many applications in genome research, including physical mapping, map-based cloning, functional and comparative genomics as well as polyploid genome analyses.

International audience; Cassava suffers from bacterial blight attack in all growing regions. Control by resistance is unstable due to high genotype–environment interactions. Identifying genes for resistance to African strains of Xanthomonas axonopodis pv. manihotis can support breeding efforts. Five F1 cassava genotypes deriving from the male parent ‘CM2177-2’ and the female parent ‘TMS30572’ were used to produce 111 individuals by backcrossing to the female parent. In all, 16 genotypes among the mapping population were resistant to stem inoculation by four strains of X. axonopodis pv. manihotis from different locations in Africa, and 19 groups with differential reactions to the four strains were identified, suggesting that the strains represent different pathotypes. Four genotypes were resistant to leaf inoculation, and three were resistant to both stem and leaf inoculations. Genotypes with susceptible, moderately resistant, and resistant reactions after leaf and stem inoculation partly differed in their reactions on leaves and stems. Based on the genetic map of cassava, single-marker regression analysis of disease severity after stem-puncture inoculation was performed. Eleven markers were identified, explaining between 16 and 33.3% of phenotypic variance of area under disease progress curve. Five markers on three and one linkage groups from the female- and malederived framework of family CM8820, respectively, seem to be weakly associated with resistance to four strains of X. axonopodis pv. manihotis. Based on the segregation of alleles from the female of family CM8873, one marker was significantly associated with resistance to two X. axonopodis pv. manihotis strains, GSPB2506 and GSPB2511, whereas five markers were not linked to any linkage group. The quantitative trait loci (QTL) mapping results also suggest that the four African strains belong to four different pathotypes. The identified pathotypes should be useful for screening for resistance, and the QTL and markers will support breeding for resistance.

The diversity of Bradyrhizobium strains nodulating three cowpea (Vigna unguiculata L. Walp.) cultivars in favorable and water-limited conditions occuring at flowering was analysed. PCR- Restriction Fragment Length Polymorphism (PCR-RFLP) analysis of 16S-23S rDNA intergenic spacer region (IGS) directly applied on 85 crushed nodules distinguished four genetic profiles, IGS types I, II, III and IV. The distribution of these IGS types according to water conditions and cowpea cultivars (B-21, TN 88-63 and Mouride) showed that nodulating strains appeared more diverse in water-limited condition. More than three quarters of prospected nodules presented the IGS type I. They were formed on all three cultivars and in both water conditions. Only a small part of nodules was distributed between the IGS type II, III and IV. Nodules showing the IGS types II and III were found mainly in limited conditions on TN 88-63 and Mouride cultivars, whereas nodules presenting the IGS type IV were collected only from cultivars B-21 and Mouride, in both water conditions. Strains corresponding to the different profiles were isolated. The phylogenetic analysis of 16S rRNA gene sequences showed that they belong to the genus, Bradyrhizobium. The sequence analysis of 16S-23S rDNA IGS revealed that the strains exhibiting IGS types II, III and IV were closely related to some Faidherbia albida isolates from Senegal. IGS type II can be assigned with at least 98% similarity to Bradyrhizobium genospecies IV. IGS types III and IV showed more than 96% similarity with genospecies VII and could belong to the same genospecies. IGS type I, the most frequent, exhibits low IGS similarity with reported sequences in the databases, and could represent a new genospecies. (African Journal of Biotechnology: 2003 2(1): 13-23)

Coffea canephora# possede l'une des aires de repartition les plus larges du sous-genre #Coffea# : elle s'etend d'ouest en est, de la Guinee au Soudan et, du nord au sud, du Cameroun a l'Angola. La conservation a long terme de cette espece s'effectue en champs a cause du comportement peu orthodoxe de ses graines lors de leur stockage. La premiere analyse de la diversite genetique de cette espece a ete realisee a partir du polymorphisme enzymatique. L'utilisation des marqueurs moleculaires est recente et les resultats presentes dans ce chapitre constituent les premieres donnees sur la variabilite moleculaire de #C. canephora#. L'analyse de la diversite agromorphologique ne permet pas de reveler une forte structuration de l'espece. A l'aide des locus isoenzymatiques discriminant les groupes guineen et congolais, un schema de selection recurrente reciproque de #C. canephora# fonde sur l'utilisation de ces deux groupes a ete propose et son efficacite a ete validee par la suite, ce qui confirme qu'il est important de connaitre la structure de la diversite pour exploiter au mieux les ressources genetiques. Les resultats de l'etude sur les marqueurs moleculaires devraient permettre d'ameliorer ce schema de selection. Un certain nombre de recommandations pour assurer une meilleure gestion #ex situ# des ressources genetiques est avance

L'espèce cultivée asiatique, #Oryza sativa#, a une structure génétique tout à fait particulière, objet des travaux décrits dans ce chapitre. Sa domestication, à partir de l'espèce #O. rufipogon#, est très ancienne. Deux sous-espèces ont été identifiées, #indica# et #japonica# à partir des caractères morphologiques et physiologiques et du comportement en croisement. Les marqueurs moléculaires apportent beaucoup à la compréhension de la diversité génétique du riz. Les isoenzymes, malgré le nombre limité de locus et d'allèles accessibles, ont fourni les premiers éléments de quantification de la différenciation intravariétale. La classification établie sur la base des isoenzymes concorde globalement avec des classifications locales, empiriques, ainsi qu'avec les classifications synthètiques, établies sur des critères multiples associant composantes morphologiques et physiologiques. Des études fondées sur plusieurs types de marqueurs moléculaires, ont permis de construire différentes classifications infraspécifiques. Dans ce chapitre, on compare plusieurs schémas de classification reposant sur différents types de caractères, en concentrant les analyses sur des échantillons largement partagés. Ensuite sont examinées quelques situations particulières permettant d'aborder certains aspects de la dynamique qui conditionne la diversité génétique des riz cultivés.

Microsatellites or simple sequence repeats (SSR) were used to assess polymorphism among 16 Coffea arabica and four Coffea canephora accessions, and to identify DNA introgression fragments from C. canephora in four C. arabica lines. Thirty-one primer pairs allowed for the identification of 92 polymorphic alleles distributed over 37 loci. The C. arabica accessions derived from the genetic bases 'Typica' and 'Bourbon' were grouped separately according to their genetic origin. Two genotypes derived from a spontaneous hybrid (C. arabica x C. canephora) were classified with the C. canephora accessions from Central Africa. Coffea canephora from West Africa were separated from the other accessions studied. Four alleles related to introgression (i.e. present in C. canephora and introgressed lines, and absent in C. arabica) were identified. The SSR markers were used successfully for characterization of a particular cultivar ('Veranero') from Costa Rica, which is known for its late maturity. (Resume d'auteur)

A direct comparison of the genetic linkage maps of sorghum and rice is proposed. It is based on the mapping of a common set of 123 RFLP probes scattered on the genomes of both species. For each species a composite map was established by merging two individual maps comprising many common loci. This enabled us to confirm the global correspondence scheme that had previously been established between the chromosomes of sorghum and rice. It also provided a more detailed insight into the conservation of synteny and colinearity : 69% of the loci mapped on a given rice chromosome mapped to the corresponding homoeologous chromosome in sorghum ; among them, 84% formed a colinear arrangement between the two species. Local inversions and translocations were detected. (Résumé d'auteur)

Eighteen strains of anaerobic thiosulfate-reducing bacteria unable to use sulfate as electron acceptor (TSRnSR) were isolated from four ricefield soils originating from France and the Philippines, using peptides as energy sources, H2 as electron donor, thiosulfate as electron acceptors, and four enrichment methods to vary the selection pressure. They were strict proteolytic asaccharolytic anaerobes producing H2S when grown on thiosulfate + H2. They exhibited the same RFLP profile (11 restriction enzymes tested). Partial sequencing of the 16S rDNA showed that they belonged to the genus Clostridium and were phylogenetically related to C. subterminale. DNA-DNA hybridization of a representative strain with the closest C. subterminale strain (DSM 6970T) yielded a value of 68.9%. Previous counts of TSRnSR in ricefield soils, their identification as Clostridium strains, and the known ubiquity of this genus in such soils indicate that TSRnSR may play a significant role in S cycling in some wetland soils. (Résumé d'auteur)

Pineapple, Ananas comosus (L.) Merr, is the third most important tropical fruit cultivated in all tropical and subtropical countries. Pineapple germplasm includes all seven species of the genus Ananas and the unique species of the related genus Pseudananas. A knowledge of its diversity structure is needed to develop new breeding programs. Restriction fragment length polymorphism (RFLP) was used to study molecular diversity in a set of 301 accessions, most of which were recently collected. This sample was analysed using 18 homologous genomic probes. Dissimilarities were calculated by a Dice index and submitted to Factorial Analysis. The same data were represented as a diversity tree constructed with the score method. Pseudananas sagenarius displayed a high polymorphism and shares 58.7% of its bands with Ananas. Within Ananas, variation appears continuous and was found mostly at the intraspecific level, particularly in the wild species Ananas ananassoides and Ananas parguazensis. As for the cultivated species, Ananas comosus appears relatively homogeneous despite its wide morphological variation and Ananas bracteatus, which is grown as a fence and for fruit, appears still much less variable. By contrast Ananas lucidus, cultivated by the Amerindians for fiber, displays a high polymorphism. This tree displayed a loose assemblage of numerous clusters separated by short distances. Most species were scattered in various clusters, a few of these being monospecific. Some accessions which had not been classified, as they shared morphological traits typical of different species, re-group with one or the other, and sometimes with both species in mixed clusters. No reproductive barrier exists in this germplasm and these data indicate the existence of gene flow, enhancing the role of effective sexual reproduction in a species with largely predominant vegetative mutiplication.

The worldwide threat of tuberculosis to human health emphasizes the need to develop novel approaches to a global epidemiological surveillance. The current standard for #Mycobacterium tuberculosis$ typing based on IS6110 restriction fragment length polymorphism (RFLP) suffers from the difficulty of comparing data between independent laboratories. Here, we propose a high-resolution typing method based on variable number tandem repeats (VNTRs) of genetic elements named mycobacterial interspersed repetitive units (MIRUs) in 12 human minisatellite-like regions of the #M. tuberculosis$ genome. MIRU-VNTR profiles of 72 different #M. tuberculosis$ isolates were established by PCR analysis of all 12 loci. From 2 to 8 MIRU-VNTR alleles were identified in the 12 regions in these strains, which corresponds to a potential of over 16 million different combinations, yielding a resolution power close to that of IS6110-RFLP. All epidemiologically related isolates tested were perfectly clustered by MIRU-VNTR typing, indicating that the stability of these MIR-VNTRs is adequate to track outbreak episodes. The correlation between genetic relationships inferred from MIRU-VNTR and IS6110-RFLP typing was highly significant. Compared with IS6110-RFLP, high-resolution MIRU-VNTR typing has the considerable advantages of being fast, appropriate for all #M. tuberculosis$ isolates, including strains that have a few IS6110 copies, and permitting easy and rapid comparison of results from independent laboratories. This typing method opens the way to the construction of digital global databases for molecular epidemiology studies of #M. tuberculosis$. (Résumé d'auteur)

DOI: 10.1139/gen-44-4-589; International audience

Cassava bacterial blight (CBB), caused by #Xanthomonas axonopodis$ pv. #manihotis$ (#Xam$), is a major disease of cassava (#manihot esculenta$ Crantz) in Africa and South America. Planting resistant varieties is the preferred method of disease control. Recent genetic mapping of an F1 cross (TMS 30572 X CM 2177-2) led to the construction of the first molecular genetic map of cassava. To better understand the genetics of resistance to CBB, we evaluated individuals of the F1 cross for CBB resistance by controlled greenhouse inoculations and visually assessed symptoms on days 7, 15, and 30 days after inoculation, using a scale where 0 = no disease and 5 = maximum susceptibility. Five #Xam$ strains were used : CIO-84, CIO-1, CIO-136, CIO-295 and ORST X-27. Area under the disease progress curve (AUDPC) was used as a quantitative measure of resistance in QTL analysis by single-marker regression. Based on the AUDPC values, eight QTLs (Quantitative trait loci), located on linkage groups B, D, L, N, and X of the female-derived framework map, were found to explain 9-20% of the phenotypic variance of the crop's response to the #Xam$ strains. With the male-derived framework map, four QTLs on linkage groups G and C explained 10.27-27.1% of the variance. A scheme to confirm the usefulness of these markers in evaluating segregating populations for resistance to CBB is proposed. (Résumé d'auteur)

We examined the genotypic diversity of 64 Bradyrhizobium strains isolated from nodules from 27 native leguminous plant species in Senegal (West Africa) belonging to the genera Abrus , Alysicarpus , Bryaspis , Chamaecrista , Cassia , Crotalaria , Desmodium , Eriosema , Indigofera , Moghania , Rhynchosia , Sesbania , Tephrosia , and Zornia , which play an ecological role and have agronomic potential in arid regions. The strains were characterized by intergenic spacer (between 16S and 23S rRNA genes) PCR and restriction fragment length polymorphism (IGS PCR-RFLP) and amplified fragment length polymorphism (AFLP) fingerprinting analyses. Fifty-three reference strains of the different Bradyrhizobium species and described groups were included for comparison. The strains were diverse and formed 27 groups by AFLP and 16 groups by IGS PCR-RFLP. The sizes of the IGS PCR products from the Bradyrhizobium strains that were studied varied from 780 to 1,038 bp and were correlated with the IGS PCR-RFLP results. The grouping of strains was consistent by the three methods AFLP, IGS PCR-RFLP, and previously reported 16S amplified ribosomal DNA restriction analysis. For investigating the whole genome, AFLP was the most discriminative technique, thus being of particular interest for future taxonomic studies in Bradyrhizobium , for which DNA is difficult to obtain in quantity and quality to perform extensive DNA:DNA hybridizations.

An interspecific partial genetic linkage map of #Coffea$ sp. based on 62 backcross hybrids is presented. F1 hybrids were generated by a cross between the wild #C. pseudozanguebariae$ and the anciently cultivated #C. liberica$ var. #dewevrei$ (DEW) ; progeny were then derived from a backcross between F1 hybrid and DEW. The map construction consisted of a two-step strategy using 5.5 and 3.1 LOD scores revealed by simulation file. The map consisted of 181 loci : 167 amplified fragment length polymorphism (AFLP) and 13 radom fragment length polymorphism (RFLP) loci. The markers were assembled into 14 linkage groups, each with 4-31 markers covering 1,144 cM. Segregation distorsion was observed for 30% of all loci, in particular 3:1 and 1:3 ratios equally favouring each of the two parents. The existence of such ratios suggests genetic conversions events. This map also represents an initial step towards the detection of quantitative trait loci. (Résumé d'auteur)

The technique described here is a fast and simple method of extracting chloroplast DNA (cpDNA). It overcomes the need for differential centrifugation using density gradients. The leaves do not have to be kept in the dark and lyophilized before extraction, but lyophilization is still possible. The chloroplasts are specifically lysed in a cell extract of leaves, using a non-ionic detergent. After isolation by centrifugation, the cpDNA is purified by the combined action of proteolytic enzymes and detergents, followed by the elimination of proteins using a mixture of chloroform and isoamyl alcohol. This method provided good quality restriction profiles for all species analyzed.

Deux isolats de la granulose de #S. littoralis$ ont été isolés de larves mortes provenant de champs de cotton non traités dans les governorats Gharbeia et Sharkeia en Egypte. Les 2 isolats ont été caractérisés par leur profils de restriction. Les profils obtenus différent de celui publié précédement en provenance de Côte d'Ivoire en Afrique. Les profils polypeptidiques des capisdes ont été déterminé et comparé avec celui de leur homologue africain. Des tests biologiques ont été entrepris. (Résumé d'auteur)

We isolated 71 slow-growing bacterial strains from nodules of 27 native leguminous plants species in Senegal (West Africa) belonging to the genera #Abrus$, #Alysicarpus$, #Bryaspis$, #Chamaecrista$, #Cassia$, #Crotalaria$, #Desmodium$, #Eriosema$, #Indigofera$, #Moghania$, #Rhynchosia$, #Sesbania$, #Tephrosia$, and #Zornia$ playing an ecological role and having agronomic potential in arid regions. The isolates were charaterised by restriction fragment length polymorphism (RFLP) analysis of PCR-amplified 165 rDNA and comparative SDS-PAGE of whole-cell proteins ; reference strains of the different known rhizobial species and groups were included as references. We conclude that these nodule isolates are diverse, and form several phylogenetic subgroups inside #Bradyrhizobium$. Nodulation tests performed on 5 plant species demonstrated host specificity among the strains studied. (Résumé d'auteur)

La NouvelleCalédonie possède une forte concentration en Casuarinaceae endémiques. On y dénombre huit espèces de #Gymnostoma# et une espèce de #Casuarina : C. collina#. Vingt-six souches de #Frankia# ont été isolées de nodules de #C. collina# et étudiées par diverses techniques incluant l'infectivité, l'effectivité, le spectre d'hôte et la PCR/RFLP de l'ITS 16S-23S. Tous ces #Frankia# présentent les mêmes caractéristiques que celles décrites chez les souches atypiques de #Casuarina# spp. Ce travail a mis en évidence, chez les #Frankia# de #C. collina#, les caractéristiques originales suivantes : (i) ils sont localisés à l'intérieur des nodules ; (ii) ils appartiennent au groupe des #Frankia# infectifs sur #Elaeagnus#;(iii) ils se regroupent dans quatre groupes ITS déjà décrits chez #Gymnostoma#. En fait, il est très probable que les souches atypiques de #C. collina# et les #Frankia# de #Gymnostoma# soient les mêmes. On peut formuler l'hypothèse que les #Frankia# de #Gymnostoma# pourraient utiliser #C. collina# comme alternative en absence de leur hôte naturel ou quand les conditions écologiques sont favorables à #C. collina#.

Analyses of DNA polymorphism and virulence variation were used to evaluate the population structure of Xanthomonas axonopodis pv. manihotis, the pathogen causing cassava bacterial blight in Colombia. We collected strains from the major cassava-growing regions which can be grouped into different edaphoclimatic zones (ECZs) according to environmental conditions, production constraints, and economic parameters. DNA polymorphism was assessed by a restriction fragment length polymorphism analysis, using an X. axonopodis pv. manihotis plasmid DNA sequence (pthB) as a probe to evaluate the genetic relatedness among 189 Colombian strains. The sampling intensity permitted the estimation of genetic differentiation within and among ECZs, sites, and fields and even within an individual plant. A multiple correspondence analysis indicated that the Colombian X. axonopodis pv. manihotis population showed a high degree of diversity relative to X. axonopodis pv. manihotis populations studied previously, and the entire collection was grouped into seven clusters. A general correlation was observed between the clusters and the geographical origin of the strains, as each cluster was largely composed of strains from the same ECZ. Representative strains, identified with pthB, were further characterized by ribotyping, hybridization to two repetitive genomic probes (pBS6 and pBS8), and restriction analysis of plasmid contents to evaluate the complementarity of these markers. Virulence variation was observed within the Colombian collection. Strains of different aggressiveness were found in all ecological zones, but no correlation between virulence variation and DNA polymorphism was observed. The genetic and virulence analyses contribute to understanding the X. axonopodis pv. manihotis population structure in Colombia.

RFLP analysis of mitochondrial DNA (mtDNA) was used to study variation within 13 populations of #Clarias batrachus$, sampled respectively in Vietnam, Thailand and in the Indonesian Archipelago. In this study an amplified region corresponding approximately to 2.3 kilobases of the Cytochrome-b and D-loop genes was digested using 8 restriction enzymes (HinfI, Hin6I, MvaI, MspI, HaeIII, BanHI, NdeII, DraI). 12 mtDNA haplotypes were found in 40 specimens. Each sampling location was characterised by one haplotype, except Palembang (Sumatra, Indonesia) and Samarinda (Kalimantan, Indonesia) where 2 and 4 haplotypes were found respectively. The consensus tree calculated from 15 more parsimonious networks showed that mtDNA haplotypes are geographically distributed. Three well differentiated clusters were identified. The first cluster is composed by populations from both Thailand and Vietnam, the second cluster by two populations from West Sumatra between populations which come from highlands (altiude more than 1000 meters in Bukittingui, and 300 meters in Nias Island) and populations located in lowlands. The significant genetic relatedness observed between populations from Sumatra (Jambi, Palembang, Muara Tebo, Teluk Kuantan) and populations from Java indicate a possible common origin which is probably in Kalimantan. This result is supported by the high diversity of haplotypes revealed in Samarinda (Kalimantan) and their intermediate position in the genetic network. The populations from highlands in West Sumatra which share a common haplotype are genetically more related to populations from Thailand and Vietnam than to populations from the rest of Indonesia. These populations seems to be relict populations from a first colonisation event which arise from the continental part of Asia... (D'après résumé d'auteur)

La réaction en chaîne des polymérases/polymorphisme des fragments de restriction (PCR-RFLP) a été utilisée pour séparer des isolats du nématode #Bursaphelenchus$. Les isolats de #B. xylophilus$ examinés provenaient du Japon, des USA, de Chine et du Canada, et ceux de #B. mucronatus$ du Japon, de Chine et de la France. L'ADN ribosomal contenant le gène 5.8S, les segments de transciption interne 1 et 2, et les segments partiels des gènes 18S et 28S ont été amplifiés par PCR. La digestion des produits amplifiés provenant de chaque isolat à l'aide de douze endonucléases de restriction et l'examen des données en RFLP qui en découlent révèlent, par une analyse en grappe, une séparation significative entre #B. xylophilus$ et #B. mucronatus$. Parmi les isolats de #B. xylophilus$ examinés, les isolats pathogènes du Japon, ceux de Chine et des USA étaient tous identiques, tandis que les isloats non pathogènes du Japon étaient légèrement distincts et que ceux du Canada formaient une grappe séparée. Parmi les isolats de #B. mucronatus$, deux isolats provenant du Japon étaient très semblables ; de même un autre isolat du Japon et un isolat de Chine étaient identiques. Les données provenant des séquences d'ADN montrent 98 différences (substitution nucléotidiques ou séparations) dans les 884 paires de bases examinées chez les isolats de #B. xylophilus$ et #B. mucronatus$. Les données provenant des séquences d'ADN chez #Aphelenchus avenae$ et #Aphelenchoides fragariae$ diffèrent non seulement de celles des #Bursaphelenchus$ mais aussi entre elles. Afin de préciser les relations phylogéniques de ces espèces, les données séquentielles du gène 5.8S provenant de l'ADN ribosomal ont été examinées... (D'après résume d'auteur)

Resistance to antifolate antimalarial drugs (cycloguanil, a biologically active metabolite of proguanil, and pyrimethamine) is associated with a Ser- to Asn-108 point mutation in the dihydrofolate reductase thymidylate synthase gene of #Plasmodium falciparum$. The frequency of this mutation was studied in 127 clinical isolates obtained in Yaounde, Cameroon using a simple and rapid molecular technique based on the polymerase chain reaction and restriction fragment lengh polymorphism. Of the 127 isolates, pure wild-type Ser-108 codon, pure mutant-type Asn-108 codon, and mixed codons were observed in 66, 55, and six parasites, respectively. The proportion of antifolate-resistant, pure mutant-type codon, with respect to pure wild-type or mixed alleles, was 43% (55 to 127). The results of the molecular assay were compared with those of semimicro isotopic in vitro assay in 34 isolates. All 17 pure Ser-108 isolates and two isolates with mixed alleles were sensitive to both pyrimethamine (50% inhibitory concentration [IC50] < 100 nM) and cycloguanil (IC50 < 50 nM). Fourteen of 15 isolates with the mutant-type Asn-108 codon were resistant to pyrimethamine and cycloguanil. One isolate with Asn-108 showed a slightly elevated pyrimethamine IC50 (78 nM), which was within the sensitive range. This study provides further evidence that antifolate-resistant #P. falciparum$ isolates are already present in Yaounde, Cameroon. (Résumé d'auteur)

DNA polymorphism and variation in virulence of #Xanthomonas axonopodis$ pv. #manihotis$ (Xam), the causal agent of cassava bacterial blight, were studied within a pathogen population from Venezuela. Collections were made in several fields at different sites within an edaphoclimatic zone where cassava is a major crop. DNA polymorphism was assessed by RFLP analysis, using an Xam plasmidic DNA sequence (ptbB) as a probe to determine the relatedness of 91 Venezuelan isolates. A high degree of polymorphism existed among the isolates, whether collected from the same or different fields. Based on a multiple correspondence analysis, the Xam population was distributed into eight clusters and no correlation was observed between genetic diversity and geographic origin. One set of haplotype strains representing the range of variability detected in Venezuela was further characterized by another RFLP analysis using two repetitive genomic probes (pBS6 and pBS8) to establish the usefulness of these probes and their complementarity with the ptbB probe. Variation for virulence was observed in the Xam Venezuelan collection by inoculating a set of cassava cultivars with 28 isolates of the pathogen, each representig a haplotype. Understanding the genetic and pathogenic variation in the pathogen population is useful for designing cassava bacterial blight management strategies. (Résumé d'auteur)

Strains of #Xanthomonas campestris$ pv. #vignicola$ (Xcv), isolated from cowpea leaves with blight or minute pustules and collected from various geographic areas, were selected on the basis of pathological and physiological features. All strains were analyzed for genotypic markers by two methods : ribotyping with EcoRI endonuclease, and RFLP analysis with a plasmid probe (pthB) containing a gene required for pathogenicity from #Xanthomonas campestris$ pv. #manihotis$. Ribotyping revealed a unique pattern for all the strains that corresponded to the previously described ribotype rRNA7. Based on polymorphism detected by pthB among Xcv strains, nine haplotypes were defined. The observed genetic variation was independent of the geographic origin of the strains and of pathogenic variation. Some haplotypes were widely distributed, whereas others were localized. In some cases, we could differentiate strains isolated from blight symptoms and pustules according to haplotypic composition. However, in most cases, no significant differences were observed. Our results and the previous pathogenic and biochemical characterizations suggest that the strains isolated from leaves with blight symptoms or minute pustules belong to the same pathovar. We provide information on pathogen diversity that can be used to identify and characterize resistant germplasm. (Résumé d'auteur)

The biodiversity of nitrogen-fixing organisms is huge. Taxonomic and phylogenetic research is needed to structure this diversity, to facilitate communication among scientists, and to increase our understanding of the evolution and biology of diazotrophs. Molecular tools for taxonomic and biodiversity studies of diazotrophic rhizobia, frankiae, cyanobacteria and bacilli are presented in sections 2 to 5. Sections 6 to 9 focus on problems with genus and species assignment.

Neptunia oleracea is an annual aquatic stem-nodulated legume indigeneous of waterlogged places in Senegal. Bacterial isolates from nodules of N. oleracea can infect Medicago sativa roots but the nodules are not efficient. No information is available on the taxonomy of Neptunia specific bacteria.

Une enquête a été menée concernant les nématodes entomopathogènes dans les cinq régions climatiques du Japon : les îles subtropicales, la région tempérée chaude des côtes du pacifique, la région montagneuse australe, la région tempérée et la région tempérée froide. Plus de 1400 échantillons de sol provenant de 266 sites ont été testés pour la présence de nématodes à l'aide de #Galleria$-pièges. Cent vingt-et-un Steinernematides et 29 Heterorhabditides ont été détectés dans, respectivement, 62 et 64 sites. Huit espèces de Steinernematides ont été reconnues : sept espèces non décrites et #Steinernema kushidai$. Chacune des espèces montre une répartition particulière : trois sont très répandues, une est côtière, une subtropicale et les trois autres montagnardes. En contraste, les #Heterorhabditidae$ ont été identifiés comme #Heterorhabiditis indica$ et #H. megidis$. La première espèce est largement répandue dans la région côtière depuis la zone subtropicale jusqu'à la zone tempérée chaude, tandis que la seconde est surtout rencontrée dans la partie est de la région côtière tempérée. (Résumé d'auteur)

It has been postulated that chloroquine resistance may be associated with a single point mutation at codon 86 of the #Plasmodium falciparum$ multidrug resistance 1 (pfmdr 1) gene. Using a simple and rapid molecular technique involving polymerase chain reaction and restriction fragment length polymorphism, the frequency of the Asn-to-Tyr mutation associated with chloroquine resistance was established among 129 clinical isolates obtained from indigenous patients in Yaoundé, Cameroon. The results showed that 110 of 129 isolates display a mutant codon. The other clinical isolates had either a pure wild-type Asn-86 codon (n = 12) or mixed Asn/Tyr alleles (n = 7). In vitro drug assays were performed to compare the genotype and phenotype in 102 clinical isolates. Of these isolates, 86 displayed pure Tyr-86 mutant codon ; 48 (56%) mutant isolates were chloroquine-resistant (50% inhibitory concentration [IC50] > 100 nM), as expected, but 38 (44%) mutant isolates were chloroquine-sensitive (IC50 < 100 nM). Three chloroquine-resistant isolates and seven chloroquine-sensitive parasites carried a wild-type Asn-86 codon. Mixed alleles were found in six isolates (four chloroquine-sensitive and two chloroquine-resistant isolates). Our results did not confirm previous observations on the possible association between chloroquine resistance phenotype and genotype based on the pfmdr 1 gene. (Résumé d'auteur)

We analysed the genetic differentiation among 17 natural populations of the Nile tilapia Oreochromis niloticus (Linnaeus, 1758) using allozymes and restriction fragment length polymorphism (RFLP) of mitochondrial DNA (mtDNA). The populations studied, from the River Senegal to Lake Tana and from Lake Manzalla to Lake Baringo, represent all subspecies which have been previously described. Sixteen variable nuclear loci showed that these populations can be clustered in three groups: (1) West African populations (Senegal, Niger, Volta and Chad drainages), (2) Ethiopian Rift Valley populations (Lakes Awasa, Ziway, Koka and the Awash River) and (3) Nile drainage (Manzalla, Cairo, Lake Edward) and Kenyan Rift Valley populations (Lakes Turkana, Baringo and River Suguta). Nine different mtDNA haplotypes were found in the RFLP analysis of a 1 kb portion of the D-loop region. The network obtained showed that there are three geographically distinct groups; all West African populations and O. aureus are clustered, the two Ethiopian Rift Valley populations are distinct and between these two groups are the Kenyan and Ugandan Rift Valley populations. Nile populations show affinities both with West African populations and with specimens from Lakes Tana and Turkana. Taxonomic and biogeographical implications of these results are discussed.

Two doubled-haploid rice populations, IR64/Azucena and IRAT177/Apura, were used to identify markers linked to rice yellow mottle virus (RYMV) resistance using core restriction fragment length polymorphism (RFLP) maps. Resistance was measured by mechanical inoculation of 19-day-old seedlings followed by assessment of virus content by enzyme-linked immunosorbent assay tests 15 days after inoculation. IR64/Azucena and IRAT177/Apura populations, 72 and 43 lines, respectively, were evaluated, and resistance was found to be polygenic. Resistance was expressed as a slower virus multiplication, low symptom expression, and limited yield loss when assessed at the field level. Bulked segregant analysis using the IR64/Azucena population identified a single random amplified polymorphic DNA marker that mapped on chromosome 12 and corresponded to a major quantitative trait locus (QTL) evidenced by interval mapping. When pooling RFLP data, integrated mapping of this chromosome revealed that the QTL was common to the two populations and corresponded to a small chromosomal segment known to contain a cluster of major blast resistance genes. This region of the genome also reflected the differentiation observed at the RFLP level between the subspecies #indica$ and #japonica$ of #Oryza sativa$. This is consistent with the observation that most sources of RYMV resistance used in rice breeding are found in upland rice varieties that typically belong to the #japonica$ subspecies. (Résumé d'auteur)