Phylogenetic diversity of rhizobia associated with horsegram [Macrotyloma uniflorum (Lam.) Verdc.] grown in South India based on glnII, recA and 16S-23S intergenic sequence analyses

Horsegram [Macrotyloma uniflorum (Lam.) Verdc.) is an important grain legume and fodder crop in India. Information on root nodule endosymbionts of this legume in India is limited. In the present study, 69 isolates from naturally occurring root nodules of horsegram collected from two agro-eco-climatic regions of South India was analyzed by generation rate, acid/alkali reaction on YMA medium, restriction fragment length polymorphism analysis of 16S-23S rDNA intergenic spacer region (IGS), and sequence analyses of IGS and housekeeping genes glnII and recA. Based on the rDNA IGS RFLP by means of three restriction enzymes rhizobia were grouped in five clusters (I–V). By sequence analysis of 16S-23S rDNA IGS identified genotypes of horsegram rhizobia were distributed into five divergent lineages of Bradyrhizobium genus which comprised (I) the IGS type IV rhizobia and valid species B. yuanmingense, (II) the strains of IGS type I and Bradyrhizobium sp. ORS 3257 isolated from Vigna sp., (III) the strains of the IGS type II and Bradyrhizobium sp. CIRADAc12 from Acacia sp., (IV) the IGS type V strains and Bradyrhizobium sp. genospecies IV, and (V) comprising genetically distinct IGS type III strains which probably represent an uncharacterized new genomic species. Nearly, 87% of indigenous horsegram isolates (IGS types I, II, III, and V) could not be related to any other species within the genus Bradyrhizobium. Phylogeny based on housekeeping glnII and recA genes confirmed those results found by the analysis of the IGS sequence. All the isolated rhizobia nodulated Macrotyloma sp. and Vigna spp., and only some of them formed nodules on Arachis hypogeae. The isolates within each IGS type varied in their ability to fix nitrogen. Selection for high symbiotic effective strains could reward horsegram production in poor soils of South India where this legume is largely cultivated.     

Species diversity within the Morchella esculenta group (Ascomycota: Morchellaceae) in Germany and France

The distribution of the Morchella esculenta group in Germany and France is examined based on 22 samples, a sample from Montenegro is studied as well. In the recent literature the group was often treated as a single species, M. esculenta sensu lato. Our study, based on the polymorphism of the internal transcribed spacer (ITS) region within the nuclear ribosomal DNA (nrDNA), indicates the presence of three distinct species: M. esculenta (L.) Pers., M. crassipes (Vent.) Pers. : Fr., and M. spongiola Boud. They can be identified easily by restriction fragment length polymorphisms (RFLP) of the ITS region.     

Contrasting responses to ectomycorrhizal inoculation in seedlings of six tropical African tree species

Five caesalpinioid legumes, Afzelia africana, Afzelia bella, Anthonotha macrophylla, Cryptosepalum tetraphylum and Paramacrolobium coeruleum, and one Euphorbiaceae species, Uapaca somon, with a considerable range in seed sizes, exhibited different responses to inoculation by four species of ectomycorrhizal (ECM) fungi, Scleroderma dictyosporum, S. verrucosum, Pisolithus sp. and one thelephoroid sp. in greenhouse conditions. Thelephoroid sp. efficiently colonized seedlings of all of the five caesalpinioid legumes except U. somon, but provided no more growth benefit than the other fungi. Thelephoroid sp. and S. dictyosporum colonized seedlings of U. somon poorly, but stimulated plant growth more than the other fungi. The relative mycorrhizal dependency (RMD) values of the caesalpinioid legumes were never higher than 50%, whilst U. somon had RMD values ranging from 84.6 to 88.6%, irrespective of the fungal species. The RMD values were negatively related to seed mass for all plant species. Potassium concentrations in leaves were more closely related than phosphorus to the stimulation of seedling biomass production by the ECM fungi. Our data support the hypothesis that African caesalpinioid legumes and euphorbe tree species with smaller seeds show higher RMD values than those with the larger seeds.     

Growth response of Afzelia africana Sm. seedlings to ectomycorrhizal inoculation in a nutrient-deficient soil

 The growth and mineral nutrition responses of seedlings of two provenances of Afzelia africana Sm. from Senegal and Burkina Faso, inoculated with four ectomycorrhizal (ECM) fungi (Scleroderma spp. and an unidentified isolate) from the same regions were assessed in a pot experiment in a savanna ECM-propagule-free soil deficient in NPK. There was little variation in the ability of the different fungal species to colonize roots of either provenance of A. africana or to produce external hyphal in soil. Root colonization by ECM fungi and their hyphal development were not related to mineral nutrition or ECM dependency. Differences in P, N, Mg and Ca concentrations in the leaves of inoculated and non-inoculated Afzelia seedlings were not always associated with production of biomass. Only leaf K concentration increased in both provenances after ECM inoculation. However, the Burkina Faso provenance responded better to inoculation with the two fungal isolates than the Senegal provenance in terms of biomass production. This was due to stimulation of root dry weight of the Burkina Faso provenance. Therefore, the hypothesis arises that non-nutritional rather than nutritional effects explain the contribution of ECM inoculation to the growth of A. africana seedlings. Accepted: 27 April 1999     

Genetic polymorphism of ferula mushroom growing on Ferula sinkiangensis

Mating tests, internal transcribed spacer (ITS) sequence analysis, intergenic spacer 1–restriction fragment length polymorphism (IGS1-RFLP), IGS1 sequence analysis, and IGS2-RFLP analysis were carried out on isolates of 17 morphologically different Pleurotus mushrooms collected on Ferula sinkiangensis. The isolates were divided, based on mating tests and ITS sequence analysis, into two groups identical to P. eryngii var. ferulae and P. nebrodensis, respectively. Single spores from these two groups were incompatible, but those from P. eryngii var. ferulae and P. eryngii were compatible and combined to produce 56.25% dikaryon mycelia with clamp connections. The ITS of P. eryngii var. ferulae and P. nebrodensis (GenBank accession no. AY311408) were both 638 bp in size but differed by 3% in sequence. P. eryngii var. ferulae and P. eryngii (GenBank accession no. AY368658) were identical in ITS size and sequence. P. nebrodensis was the dominant population of Pleurotus mushroom growing on F. sinkiangensis. It exhibited genetic diversity. The two species could also be distinguished by IGSI-RFLP, similar to identification by mating tests and ITS sequence analysis. Difference in IGS1-RFLP existed between P. eryngii var. ferulae and P. nebrodensis. The sequence difference reached 2.28%. Both IGS1 size and IGS1-RFLP were similar among the different samples of P. nebrodensis. The 17 isolates were separated into five types based on IGS2 size and IGS2-RFLP, with both interspecies and extraspecies differences. P. nebrodensis exhibited polymorphism and was divided into four types. These results agreed with macroscopic differences. IGS2 might be the effective domain of genetically polymorphic ribosomal DNA in P. nebrodensis mushrooms found in Xinjiang, China.     

Identification of <Emphasis Type="Italic">Ganoderma</Emphasis>, the causal agent of basal stem rot disease in oil palm using a molecular method

From comparison of the alignments of the internally transcribed spacers (ITS) of ribosomal DNA from Ganoderma associated with oil palm basal stem rot (BSR) and other Ganoderma species, two specific primer pairs were selected to provide a specific DNA amplification of pathogenic Ganoderma in oil palm. Each primer pair produced a single PCR product of about 450 bp (for primer pair IT1–IT2) and 334 bp (for primer pair IT1–IT3) when oil palm Ganoderma DNA was used. No PCR amplification product was observed when other Ganoderma species DNA was used in PCR amplification with these primer pairs. Three specific restriction enzyme sites were identified in the ITS and intergenic spacer (IGS1) regions. The restriction enzymes MluI, SacI and HinfI were used to digest the ITS-PCR product and restriction enzymes TfiI, ScaI and HincII were used to digest the IGS1-PCR product. Of the three restriction enzymes used in each rDNA region, MluI specifically digested the ITS regions, and TfiI specifically digested the IGS1 region of oil palm Ganoderma. Analysis of the published ITS nucleotide sequences of 31 Ganoderma species showed that the MluI restriction site was not present in other Ganoderma species. The use of both specific primers and restriction enzyme analysis can be applied as a standard protocol to identify pathogenic Ganoderma in oil palm. In this study, the use of specific primers and PCR-RFLP analyses of the rDNA gave consistent results for the characterisation of pathogenic Ganoderma, and indicated that Ganoderma strains associated with BSR disease in oil palms belong to a single species.     

Two new species of Haemaphysalis Koch, 1844 (Acari: Ixodidae) in the H. (Rhipistoma) spinulosa subgroup,parasites of carnivores and hedgehogs in Africa

Haemaphysalis (Rhipistoma) princeps n. sp. (Acari: Ixodidae) and H. (R.) camicasi n. sp. are described based on adults. Adults of H. princeps n. sp. were mostly collected from various carnivores (Carnivora: Canidae, Felidae, Herpestidae, Mustelidae, Viverridae) and hedgehogs (Erinaceomorpha: Erinaceidae); few specimens were found on hare (Lagomorpha: Leporidae), various rodents (Rodentia: Muridae) and an antelope (Artiodactyla: Bovidae) in Benin, Burkina Faso, Cameroon, Chad, Côte d’Ivoire, Ghana, Mali, Nigeria, Senegal, South Sudan and Sudan. Few adults of H. princeps n. sp. were reared from nymphs collected on rodents (Rodentia: Muridae) and shrew (Soricomorpha: Soricidae). Adults of H. camicasi n. sp. were also mostly collected from various carnivores (Carnivora: Canidae, Felidae, Herpestidae, Mustelidae, Viverridae) and hedgehogs (Erinaceomorpha: Erinaceidae); few specimens were found on different rodents (Rodentia: Muridae, Nesomyidae, Sciuridae), hare (Lagomorpha: Leporidae) and bushbaby (Primates: Galagidae) in Benin, Burkina Faso, Cameroon, Central African Republic, Chad, Côte d’Ivoire, Nigeria, Senegal, South Sudan and Togo. Males and females of both species can be differentiated from each other and other H. spinulosa-like ticks by their size, pattern of punctations on conscutum/scutum, size of setae, shape of genital structures, size and shape of spiracular plates, dorsal cornua, posterodorsal and posteroventral spurs on palpal segments II and spur on coxae.

     

Differentiation ofMelampsora rust species on willows in Japan using PCR-RFLP analysis of ITS regions of ribosomal DNA

To develop a reliable method for identifyingMelampsora species parasitic on willows in Japan, we differentiated 10Melampsora species by PCR-RFLP analysis. Internal transcribed, spacer (ITS) regions, including 5.8S ribosomal DNA, of 63 collections of 10Melampsora species and 4 collections of unidentified species were amplified by PCR. The fragments from the 67 collections varied in size (approximately 880 bp, 860 bp and 840 bp). The restriction sites in the amplified DNA fragments were mapped after the RFLP analysis using four restriction enzymes,Dra I,EcoRI,SspI andTaqI. All the collections were divided into 11 RFLP types. In the 6 species,M. caprearum, M. epiphylla, M. kamikotica, M. larici-urbaniana, M. microsora andM. yezoensis, the RFLP type was species-specific. The RFLP type ofM. chelidonii-pierotii andM. coleosporioides was identical. The collections ofM. epitea were separated into three RFLP types. One of these three types was identical with the type ofM. humilis. It is suggested that the PCR-RFLP analysis of ITS regions is a useful and reliable method for species identification ofMelampsora. Contribution No. 131, Laboratories of Plant Pathology and Mycology, Institute of Agriculture and Forestry, University of Tsukuba.     

Identification of species in the genus Pichia by restriction of the internal transcribed spacers (ITS1 and ITS2) and the 5.8S ribosomal DNA gene

In this study, the variability within the ribosomal DNA region spanning the internal transcribed spacers ITS1 and ITS2 and the 5.8S gene (5.8S-ITS rDNA) was used to differentiate species in the genus Pichia. The 5.8S-ITS rDNA region was PCR-amplified and the PCR product digested with the enzymes CfoI, HinfI, and HaeIII. The variability in the size of the amplified product and in the restriction patterns enabled differentiation between species in the genus Pichia, and between Pichia species and yeast species from other genera in the Yeast-id database (). Moreover, the restriction fragment length polymorphism (RFLP) patterns of the 5.8S-ITS enabled misidentified strains to be detected and revealed genetic heterogeneity between strains within the Pichia membranifaciens and Pichia nakazawae species. Ultimately, the RFLP patterns of the 5.8S-ITS rDNA failed to differentiate between some Pichia and Candida species that could be distinguished on the basis of the sequence of the 5.8S-ITS rRNA region or the sequence of the D1/D2 domain of the 26S rDNA gene.     

Molecular analyses of the IGS & ITS regions of rDNA of the psychrophilic yeasts in the genus Mrakia

Species of the genus Mrakia are currently classified as synonyms based on molecular sequence analyses of the large sub-unit ribosomal DNA (LrDNA). Physiological and protein electrophoretic studies, however, reveal possible species differences. To clarify this discrepancy, we undertook molecular sequence analyses of the internal transcribed spacer (ITS) and intergenic spacer (IGS) regions of rDNA from the four psychrophilic Mrakia species and the psychrophilic yeast, Cryptococcus curiosus. Identical ITS sequences were found between C. curiosus, M. nivalis and M. frigida. Although, M. stokesii and M. gelida displayed identical ITS and IGS sequences, their sequences differed from the other three species by 2.3% and 38%, respectively. The results suggest that M. stokesii is a synonym of M. gelida, whereas M. nivalis is a synonym of M. frigida. Sequence differences (1.9%) observed in the IGS region indicates that C. curiosus is a distinct strain of M. frigida.     

Polymorphism in the internal transcribed spacer (ITS) region of the ribosomal DNA among different Fusarium species

Fusarium species causing wilt diseases in different plants were characterised by comparing nonpathogenic and different pathogenic species using rDNA RFLP analysis. The ITS (internal transcribed spacer) region of 12 isolates belonging to the section Elegans, Laseola, Mortiella, Discolor, Gibbosum, Lateritium and Sporotrichiella were amplified by universal ITS primers (ITS-1 and ITS-4) using polymerase chain reaction (PCR). Amplified products, which ranged from 522 to 565 bp were obtained from all 12 Fusarium isolates. The amplified products were digested with seven restriction enzymes, and restriction fragment length polymorphism (RFLP) patterns were analysed. A dendrogram derived from PCR-RFLP analysis of the rDNA region divided the Fusarium isolates into three major groups. Assessment of molecular variability based on rDNA RFLP clearly indicated that Fusarium species are heterogeneous and most of the forma speciales have close evolutionary relationships.     

A new PCR-RFLP-based method for an easier systematic affiliation of European water frogs

We describe a non‐invasive, PCR‐RFLP‐based method that allows reliable determination of the European water frog species Pelophylax lessonae and Pelophylax ridibundus and the hybrid form Pelophylax esculentus. Maximum‐likelihood analysis of ITS2 sequences revealed two robust monophyletic clades corresponding to water frogs of the P. lessonae and P. ridibundus groups. Three restriction enzymes (KpnI, HaeII, and SmaI) were used to digest three conserved ITS2 domains. Taxonomic identification was unambiguous; the three restriction enzymes gave the same results. A French reference sample was identified using allozyme electrophoresis. Our PCR‐RFLP method confirmed circa 83% of identification of the allozyme method. We conclude that the difference between identifications was caused by introgression.     

Ectomycorrhizal fungal diversity in orchards of cultivated pecan (Carya illinoinensis; Juglandaceae)

Carya illinoinensis (pecan) belongs to the Juglandaceae (walnut family) and is a major economic nut crop in the southern USA. Although evidence suggests that some species in the Juglandaceae are ectomycorrhizal, investigations on their ectomycorrhizal fungal symbionts are quite limited. Here we assessed the ectomycorrhizal fungal diversity in cultivated orchards of C. illinoinensis. Five pecan orchards in southern Georgia, USA, were studied, three of which were known to fruit the native edible truffle species Tuber lyonii. We sequenced rDNA from single ectomycorrhizal root tips sampled from a total of 50 individual trees. Mycorrhizae were identified by ITS and LSU rDNA sequence-based methods. Forty-four distinct ectomycorrhizal taxa were detected. Sequestrate taxa including Tuber and Scleroderma were particularly abundant. The two most abundant sequence types belonged to T. lyonii (17%) and an undescribed Tuber species (~20%). Because of our interest in the ecology of T. lyonii, we also conducted greenhouse studies to determine whether this species would colonize and form ectomycorrhizae on roots of pecan, oak, or pine species endemic to the region. T. lyonii ectomycorrhizae were formed on pecan and oak seedlings, but not pine, when these were inoculated with spores. That oak and pecan seedling roots were receptive to truffle spores indicates that spore slurry inoculation could be a suitable method for commercial use and that, ecologically, T. lyonii may function as a pioneer ectomycorrhizal species for these hosts.     

Plasmodium falciparum: population genetic analysis by multilocus enzyme electrophoresis and other molecular markers.

Abderrazak, S. B., Oury, B, Lal, A. A., Bosseno, M.-F., Force-Barge, P., Dujardin, J.-P., Fandeur, T., Molez, J.-F., Kjellberg, F., Ayala, F. J., and Tibayrenc, M. 1999. Plasmodium falciparum: Population genetic analysis by multilocus enzyme electrophoresis and other molecular markers. Experimental Parasitology 92, 232-238. The population structure of Plasmodium falciparum, the agent of malignant malaria, is uncertain. We have analyzed multilocus enzyme electrophoresis (MLEE) polymorphisms at 7-12 gene loci in each of four populations (two populations in Burkina Faso, one in Sudan, one in Congo), plus one "cosmopolitan" sample consisting of parasite cultures from 15 distant localities in four different continents. We have also performed random amplified polymorphic DNA analysis (RAPD) and restriction fragment length polymorphism (RFLP) and characterized gene varia tion at four antigen genes in the Congo population. All genetic assays show abundant genetic variability in all populations analyzed. With the isoenzyme assays, strong linkage disequilibrium is apparent in at least two local populations, the Congo population and one population from Burkina Faso, as well as in the cosmopolitan sample, and less definitely in the other Burkina Faso population. However, no linkage disequilibrium is detected in the Congo population with the molecular assays. We failed to detect any nonrandom association between the different kinds of genetic markers; that is, MLEE with RAPD or RFLP, RAPD with RFLP, and so on. Although isoenzyme data show statistical departures from panmictic expectations, these results suggest that in the areas under survey, P. falciparum populations do not undergo predominant clonal evolution and show no clear-cut subdivisions, un like Trypanosoma cruzi, Leishmania sp., and other major parasitic species. We discuss the epidemiological and taxonomical significance of these results.     

Rhizobial Populations in Soils from Natural Acacia senegal and Acacia nilotica Forests in Mauritania and the Senegal River Valley

Eighty-two strains of rhizobia were isolated from soils taken from several sites in Mauritania and Senegal. These soil samples were collected from natural stands of Acacia nilotica and Acacia senegal. The soils from Mauritania were less rich in native rhizobia than the soils from Senegal. The strains were characterized using polymerase chain reaction–restriction fragment length polymorphism and by sequencing the rDNA 16S–23S intergenic spacer region (IGS). They were sorted into seven IGS groups. These groups were not associated with the geographical origin of the strains or with the host-plant species at the site where the soils were collected. Most of the strains were in three of the IGS groups (I, IV, and V). One representative strain from each IGS group was sequenced and showed that the strains were from the genus Mesorhizobium. IGS groups I, IV, and VI were close to the species M. plurifarium (AF34563), IGS groups IIand III were close to the species Mesorhizobium sp. (AF510360), IGS group V was close to the species Mesorhizobium sp. (AF510366), and IGS group VII was close to Mesorhizobium sp. (AF510346).     

Spectrum of plant species foraged by Apis mellifera adansonii Latreille in the North Sudanian phytogeographical region of Burkina Faso

Melissopalynological analysis of 60 honey samples collected from 50 Kenyan Top Bar Hives in the North Sudanian phytogeographical region of Burkina Faso established the plant species foraged by honeybees Apis mellifera adansonii. Forty-three pollen types from 29 families were identified in the honey samples. The most common families were Mimosaceae (9.30%), Asteraceae (6.98%) and Anacardiaceae (6.98%). The most frequently visited plants were Combretum (66.66%), Tridax procumbens (66.66), Acacia seyal-group (50%), Cleome viscosa-group (50%) and Cyperus esculentus (41.67%). The results showed that Apis mellifera adansonii is polylectic with a heterogeneous foraging behaviour. Pollen analysis showed that the honeys from the two main honey flows of Burkina Faso were polyfloral.     

Molecular identification of house dust mites and storage mites

Mites are known causes of allergic diseases. Currently, identification of mites based on morphology is difficult if only one mite is isolated from a (dust) sample, or when only one gender is found, or when the specimen is not intact especially with the loss of the legs. The purpose of this study was to use polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) of the ITS2 gene, to complement the morphological data for the identification of mites to the species level. For this, six species were cultured: Dermatophagoides pteronyssinus, D. farinae, Blomia tropicalis, Tyrophagus putrescentiae, Aleuroglyphus ovatus and Glycycometus malaysiensis. Genomic DNA of the mites was extracted, quantified, amplified and digested individually with restriction enzymes. Hinf I and Ple I differentiated the restriction patterns of D. pteronyssinus and D. farinae. Bfa I and Alu I enzymes differentiated B. tropicalis and G. malaysiensis. Ple I enzyme was useful for the differentiation between T. putrescentiae and A. ovatus. Bfa I was useful for the differentiation of G. malaysiensis from the rest of the species. In conclusion, different species of mites can be differentiated using PCR–RFLP of ITS2 region. With the established PCR–RFLP method in this study, identification of these mites to the species level is possible even if complete and intact adult specimens of both sexes are not available. As no study to date has reported PCR–RFLP method for the identification of domestic mites, the established method should be validated for the identification of other species of mites that were not included in this study.     

Close sequence identity between ribosomal DNA episomes of the non-pathogenicEntamoeba dispar and pathogenicEntamoeba histolytica

Entamoeba dispar andEntamoeba histolytica are now recognized as two distinct species-the former being nonpathogenic to humans. We had earlier studied the organization of ribosomal RNA genes inE. histolytica. Here we report the analysis of ribosomal RNA genes inE. dispar. The rRNA genes ofE. dispar, like their counterpart inE. histolytica are located on a circular rDNA molecule. From restriction map analysis, the size ofE. dispar rDNA circle was estimated to be 24·4 kb. The size was also confirmed by linearizing the circle withBsaHI, and by limited DNAseI digestion. The restriction map of theE. dispar rDNA circle showed close similarity to EhR1, the rDNA circle ofE. histolytica strain HM-1:IMSS which has two rDNA units per circle. The various families of short tandem repeats found in the upstream and downstream intergenic spacers (IGS) of EhR1 were also present inE. dispar. Partial sequencing of the cloned fragments ofE. dispar rDNA and comparison with EhR1 revealed only 2·6% to 3·8% sequence divergence in the IGS. The region Tr and the adjoiningPvuI repeats in the IGS of EhR1, which are missing in thoseE. histolytica strains that have one rDNA unit per circle, were present in theE. dispar rDNA circle. Such close similarity in the overall organization and sequence of the IGS of rDNAs of two different species is uncommon. In fact the spacer sequences were only slightly more divergent than the 18S rRNA gene sequence which differs by 1·6% in the two species. The most divergent sequence betweenE. histolytica andE. dispar was the internal transcribed spacer, ITS2. Therefore, it was concluded that probes derived from the ITS1 and ITS 2 sequences would be more reliable and reproducible than probes from the IGS regions used earlier for identifying these species.     

Genetic Diversity of Fusarium oxysporum Populations Isolated from Tomato Plants in Tunisia

Fusarium crown and root rot of tomato (Lycopersicon esculentum) caused by Fusarium oxysporum f. sp. radicis‐lycopersici is a new devastative disease of tomato greenhouse crops in Tunisia. Nothing is known neither about the population of this pathogen in this region, nor about the population of F. oxysporum f. sp. lycopersici the causal agent of Fusarium wilt of tomato. In order to examine the genetic relatedness among the F. oxysporum isolates by intergenic spacer restriction fragment length polymorphism (IGS‐RFLP) analysis and to elucidate the origin of the formae specialesradicis‐lycopersici in Tunisia by looking for genetic similarity of Tunisians isolates with isolates from a foreign source, the genetic diversity among F. oxysporum f. sp. radicis‐lycopersici and F. oxysporum f. sp. lycopersici populations was investigated. A total of 62 isolates of F. oxysporum, obtained from symptomless tomato plants, were characterized using IGS typing and pathogenicity tests on tomato plants. All Fusarium isolates were highly pathogenic on tomato. Fusarium oxysporum f. sp. radicis‐lycopersici isolates were separated into five IGS types. From the 53 F. oxysporum f. sp. radicis‐lycopersici isolates, 34 isolates have the same IGS types (IGS type 25), and the remaining 19 isolates were distributed into four IGS types. However, the only nine isolates of F. oxysporum f. sp. lycopersici have six different IGS types. This difference of diversity between the two formae speciales suggests that F. oxysporum f. sp. radicis‐lycopersici isolates have a foreign origin and may have been accidentally introduced into Tunisia.     

Molecular identification of co-occurring Cortinarius and Dermocybe species from southeastern Australian sclerophyll forests

 The ability of restriction fragment length polymorphism (RFLP) analysis of the rDNA internal transcribed spacer (ITS) region to discriminate 10 co-occurring Cortinarius and Dermocybe species at a southeastern Australian sclerophyll forest site was assessed. Using the basidiomycete-specific primers ITS1F and ITS4B, some taxa were separated on the basis of individual RFLP patterns derived using the restriction endonucleases Hae III or Hinf I. Combined data from both endonucleases were, however, required to separate all taxa [Dermocybe austro-veneta Clel. (Moser & Horak), C. rotundisporus Clel. & Cheel, C. archeri Berk., C. sinapicolor Clel., C. violaceus (L.: Fr.) S.F.Gray, C. radicatus Clel. and four morphologically-distinct, but unidentified Cortinarius spp.]. ITS sequence comparisons confirmed that D. austro-veneta belongs in Dermocybe, that C. rotundisporus is correctly placed in subgenus Phlegmacium, and suggest that Australian C. violaceus collections are not conspecific with northern hemisphere C. violaceus. Accepted: 4 March 1999