Investigating Enterocytozoon bieneusi in pigs farmed in Zhejiang Province, China: Occurrence, genotype identification, evolutionary analysis, and zoonotic risk assessment.

Enterocytozoon bieneusi is a widespread intracellular fungus that can infect both humans and animals, making it a significant zoonotic threat. In the current study, a total of 208 fecal samples were assayed to investigate the prevalence of E. bieneusi in pigs reared in Zhejiang Province, China. Employing polymerase chain reaction (PCR) amplification techniques specifically designed to target the internal transcribed spacer (ITS) region of the small subunit ribosomal RNA (rRNA) gene, the results revealed that 78 samples (37.5 %) tested positive for the presence of E. bieneusi. A total of 19 different genotypes of E. bieneusi were detected. Nine of these genotypes were already known: EbpC (n = 36), KIN-1 (n = 10), PigEbITS7 (n = 8), EbpA (n = 6), Henan III (n = 3), PigEbITS5 (n = 2), Henan-IV (n = 1), EbpD (n = 1), and TypeIV (n = 1), and 10 were novel: ZJP-I to ZJP-X (one each). The present investigation revealed that all the nine known genotypes identified in pigs here, have also been previously discovered in humans. Additionally, the novel genotypes of E. bieneusi discovered here were all classified as belonging to Group 1. These findings suggest the potential for cross-species transmission between humans and pigs.

Phylogenetic Assessment of Understudied Families in Hymenochaetales (Basidiomycota, Fungi)-Reporting Uncovered Species and Reflecting the Recent Taxonomic Updates in the Republic of Korea.

Hymenochaetales Oberw. is an order classified in Basidiomycota of Fungi, and species in this order display notable diversity. They exhibit various fruiting body shapes, including clavarioid, effused-reflexed, and resupinate basidiomes. Few mycorrhizal species have been reported in Hymenochaetales, but wood-decaying species dominate the order. Hymenochaetaceae Imazeki & Toki and Schizoporaceae Jülich are the most species-rich families within Hymenochaetales, and most species in the Republic of Korea belong to these two families. As such, current taxonomic classification and nomenclature are not reflected upon species in the remaining Hymenochaetales families. For this study, a multifaceted morphological and multigenetic marker-based phylogenetic investigation was conducted to, firstly, comprehensively identify understudied Hymenochaetales specimens in Korea and, secondly, reflect the updates on the species classification. Five genetic markers were assessed for the phylogenetic analysis: nuclear small subunit ribosomal DNA (nSSU), internal transcribed spacer (ITS), nuclear large subunit ribosomal DNA (nLSU), RNA polymerase II subunit 2 gene (RPB2), and translation elongation factor 1 gene (TEF1). The results from phylogenetic analysis supported 18 species classified under eight families (excluding Hymenochaetaceae and Schizoporaceae) in Korea. Species formerly placed in Rickenellaceae and Trichaptum sensu lato have been systematically revised based on recent taxonomic reconstructions. In addition, our findings revealed one new species, Rickenella umbelliformis, and identified five formerly nationally unreported species classified under five understudied families. Our findings contribute to a better understanding of Hymenochaetales diversity and highlight the need for continued research.

A set of nematode rRNA cistron databases and a primer assessment tool to enable more flexible and comprehensive metabarcoding.

The ITS-2-rRNA has been particularly useful for nematode metabarcoding but does not resolve all phylogenetic relationships, and reference sequences are not available for many nematode species. This is a particular issue when metabarcoding complex communities such as wildlife parasites or terrestrial and aquatic free-living nematode communities. We have used markerDB to produce four databases of distinct regions of the rRNA cistron: the 18S rRNA gene, the 28S rRNA gene, the ITS-1 intergenic spacer and the region spanning ITS-1_5.8S_ITS-2. These databases comprise 2645, 254, 13,461 and 10,107 unique full-length sequences representing 1391, 204, 1837 and 1322 nematode species, respectively. The comparative analysis illustrates the complementary value but also reveals a better representation of Clade III, IV and V than Clade I and Clade II nematodes in each case. Although the ITS-1 database includes the largest number of unique full-length sequences, the 18S rRNA database provides the widest taxonomic coverage. We also developed PrimerTC, a tool to assess primer sequence conservation across any reference sequence database, and have applied it to evaluate a large number of previously published rRNA cistron primers. We identified sets of primers that currently provide the broadest taxonomic coverage for each rRNA marker across the nematode phylum. These new resources will facilitate more comprehensive metabarcoding of nematode communities using either short-read or long-read sequencing platforms. Further, PrimerTC is available as a simple WebApp to guide or assess PCR primer design for any genetic marker and/or taxonomic group beyond the nematode phylum.

IDphy: An International Online Resource for Molecular and Morphological Identification of Phytophthora.

Phytophthora, with 203 species, is a genus of high importance in agriculture worldwide. Here, we present the online resource "IDphy", developed to facilitate the correct identification of species of Phytophthora using the type specimens from the original descriptions wherever possible. IDphy emphasizes species of high economic impact and regulatory concern for the United States. IDphy presents an interactive Lucid key and a tabular key for 161 culturable species described as of May 2018, including 141 ex-types and 20 well-authenticated specimens. IDphy contains standard operating procedures for morphological and molecular characterization, as well as a glossary, image gallery, and numerous links. Each of the 161 factsheets includes access to nomenclature and morphological and molecular features, including sequences of the internal transcribed spacer ribosomal DNA, cytochrome C oxidase subunit I (barcoding genes), YPT1, ß-tubulin, elongation factor 1a, L10, heat shock protein 90, and other genes. IDphy contains an innovative in silico BLAST and phylogenetic sequence analysis using NCBI. The IDphy mobile app, released in August 2021 (free for Android or iOS), allows users to take the Lucid key into the laboratory. IDphy is the first online identification tool based on the ex-types implemented for plant pathogens. In this article, we also include information for 21 new species and one hybrid described after the publication of IDphy, the status of the specimens of the types and ex-types at international herbaria and culture collections, and the status of genomes at the GenBank (currently 153 genome assemblies which correspond to 42 described species, including 16 ex-types). The effectiveness of the IDphy online resource and the content of this article could inspire other researchers to develop additional identification tools for other important groups of plant pathogens.

[A comparison of the costs, reliability and time of result periods of widely used methods, new molecular methods and MALDI TOF-MS in the routine diagnosis of Candida strains]. / Candida tür tayininde rutinde yaygin olarak kullanilan yöntemlerle yeni kullanilmaya baslayan MALDI TOF-MS ve moleküler yöntemlerin sonuç verme süreleri, maliyetleri ve güvenilirliklerinin karsilastirilmasi.

In recent years, the fast and accurate identification of the Candida species is of great importance as the response to antifungal treatment differs among species. Following the treatment of several immunosuppressive diseases, fungal infections can emerge. The aim of this study was to compare the accuracy, costs and time of result periods of the methods used in the identification of the most common human fungal infectious agent, Candida strains. From various clinical samples sent to the Microbiology Laboratory of Karabuk University Training and Research Hospital between July 2016-December 2017, a total of 91 yeast isolates cultivated in blood agar (Becton Dickinson, USA) and/or Sabouraud dextrose agar (SDA-Oxoid, UK), confirmed with colony morphology and microscopic appearance, identified as Candida species with a fully automated identification system (Phoenix™ Yeast ID Panel, Becton Dickinson Diagnostics, USA) were included in the study. All the samples were examined with sequence analysis using ITS1 forward 5'-TCC GTA GGT GAA CCT GCG G-3' and ITS4 reverse 5'-TCC TCC GCT TAT TGA TAT GC-3' primers (Iontek, Turkey) and the matrix-assisted laser desorption-ionisation time of flight mass spectrometry (MALDI TOF-MS) systems. Molecular sequence analysis was accepted as the gold standard method and the results were compared with those of the other methods MALDI TOF-MS and Phoenix™ Yeast ID Panel in respect of the accuracy of the identification of Candida strains. According to the results of the DNA sequence analysis of the 91 Candida isolates included in the study, 24 were identified as Candida albicans, 20 Candida tropicalis, 16 Candida parapsilosis, 13 Candida glabrata, seven Candida kefyr, six Candida krusei, two of each Candida dubliniensis, Candida guilliermondi and one Candida lusitaniae. Compared to the results of the DNA sequence analysis, the accurate identification of the fully automated Phoenix™ system and the MALDI TOF-MS system was found as 92.3% and 97.8%, respectively. In addition to accuracy, costs and time of result periods of the three methods were also compared. Disregarding the cost of the device in the 3 methods, when the comparison was made of the cost per test and the time to results after pure production in SDA agar, the MALDI TOF-MS system was determined to have the lowest costs and provided results in the shortest time. As some of the Candida strains have antifungal resistance, identification of the strains must be a priority in respect of starting early treatment. The MALDI TOF-MS system has high performance in accurate identification, low costs and the system provides the results within minutes, thereby allowing immediate decision to be made for the antifungal treatment to be started. Thus, the morbidity, mortality and cost rates will be reduced. In conclusion, as the MALDI TOF-MS is a rapid, reliable and low cost per test system, it can be considered suitable for routine use in laboratories.

Phylogeny of the genus Fuscoporia and taxonomic assessment of the F. contigua group.

The genus Fuscoporia of the Hymenochaetaceae is characterized by resupinate to pileate basidiocarps, a dimitic hyphal system with fine crystal aggregates and encrusted generative hyphae in dissepiment edge and tube trama, the presence of hymenial setae, and hyaline, thin-walled, smooth basidiospores. Members of the F. contigua group are easy to distinguish from other species of Fuscoporia because of the moderately large pores, presence of mycelial setae, and large hymenial setae. Here, we explore phylogenetic relationships among 20 species of Fuscoporia based on examination of some 90 collections sampled worldwide. Seven new species are recognized in the F. contigua group-F. americana, F. centroamericana, F. costaricana, F. latispora, F. monticola, F. septiseta, and F. sinica-described from China, Costa Rica, Mexico, and the United States. Phylogenetic analyses inferred from DNA sequences of the nuc rDNA internal transcribed spacer (ITS1-5.8S-ITS2 = ITS), D1-D2 domains of nuc 28S rDNA (28S), and translation elongation factor EF-1 alpha (tef1) support the F. contigua group as one of two major clades within Fuscoporia comprising nine species worldwide.

Nuclear and Wolbachia-based multimarker approach for the rapid and accurate identification of tsetse species.

BACKGROUND: Tsetse flies (Diptera: Glossinidae) are solely responsible for the transmission of African trypanosomes, causative agents of sleeping sickness in humans and nagana in livestock. Due to the lack of efficient vaccines and the emergence of drug resistance, vector control approaches such as the sterile insect technique (SIT), remain the most effective way to control disease. SIT is a species-specific approach and therefore requires accurate identification of natural pest populations at the species level. However, the presence of morphologically similar species (species complexes and sub-species) in tsetse flies challenges the successful implementation of SIT-based population control. RESULTS: In this study, we evaluate different molecular tools that can be applied for the delimitation of different Glossina species using tsetse samples derived from laboratory colonies, natural populations and museum specimens. The use of mitochondrial markers, nuclear markers (including internal transcribed spacer 1 (ITS1) and different microsatellites), and bacterial symbiotic markers (Wolbachia infection status) in combination with relatively inexpensive techniques such as PCR, agarose gel electrophoresis, and to some extent sequencing provided a rapid, cost effective, and accurate identification of several tsetse species. CONCLUSIONS: The effectiveness of SIT benefits from the fine resolution of species limits in nature. The present study supports the quick identification of large samples using simple and cost effective universalized protocols, which can be easily applied by countries/laboratories with limited resources and expertise.

DNA barcoding of the genus Nepenthes (Pitcher plant): a preliminary assessment towards its identification.

BACKGROUND: DNA barcoding is impending towards the generation of universal standards for species discrimination with a standard gene region that can be sequenced accurately and within short span of time. In this study, we were successful in developing efficient barcode locus in the Nepenthes genus. A total of 317 accessions were retrieved from GenBank of NCBI which represent 140 different species Nepenthes and evaluated the efficacy of ITS, rbcl and matK barcode candidates using barcode gap, applied distance similarity, and tree-based methods. RESULT: Our result indicates that single-locus ITS or combined with plastid regions (matK) showed the best species discrimination with distinctive barcoding gaps. Therefore, we tentatively proposed the combination of ITS+matK as a core barcode for Nepenthes genus. CONCLUSION: This study provides a report on DNA barcoding for unique insectivores' Nepenthes genus. As the different species of Nepenthes are higly endemic and endangered, it would be a useful study to understand the evolutionary relationship, sketched in emigration, mislabeling and can be a probable assessment for its biodiversity.

Assessment of Passive Traps Combined with High-Throughput Sequencing To Study Airborne Fungal Communities.

Techniques based on high-throughput sequencing (HTS) of environmental DNA have provided a new way of studying fungal diversity. However, these techniques suffer from a number of methodological biases which may appear at any of the steps involved in a metabarcoding study. Air is one of the most important environments where fungi can be found, because it is the primary medium of dispersal for many species. Looking ahead to future developments, it was decided to test 20 protocols, including different passive spore traps, spore recovery procedures, DNA extraction kits, and barcode loci. HTS was performed with the Illumina MiSeq platform targeting two subloci of the fungal internal transcribed spacer. Multivariate analysis and generalized linear models showed that the type of passive spore trap, the spore recovery procedure, and the barcode all impact the description of fungal communities in terms of richness and diversity when assessed by HTS metabarcoding. In contrast, DNA extraction kits did not significantly impact these results. Although passive traps may be used to describe airborne fungal communities, a study using specific real-time PCR and a mock community showed that these kinds of traps are affected by environmental conditions that may induce losses of biological material, impacting diversity and community composition results.IMPORTANCE The advent of high-throughput sequencing (HTS) methods, such as those offered by next-generation sequencing (NGS) techniques, has opened a new era in the study of fungal diversity in different environmental substrates. In this study, we show that an assessment of the diversity of airborne fungal communities can reliably be achieved by the use of simple and robust passive spore traps. However, a comparison of sample processing protocols showed that several methodological biases may impact the results of fungal diversity when assessed by metabarcoding. Our data suggest that identifying these biases is of paramount importance to enable a correct identification and relative quantification of community members.

DNA barcoding of invasive plants in China: A resource for identifying invasive plants.

Invasive plants have aroused attention globally for causing ecological damage and having a negative impact on the economy and human health. However, it can be extremely challenging to rapidly and accurately identify invasive plants based on morphology because they are an assemblage of many different families and many plant materials lack sufficient diagnostic characteristics during border inspections. It is therefore urgent to evaluate candidate loci and build a reliable genetic library to prevent invasive plants from entering China. In this study, five common single markers (ITS, ITS2, matK, rbcL and trnH-psbA) were evaluated using 634 species (including 469 invasive plant species in China, 10 new records to China, 16 potentially invasive plant species around the world but not introduced into China yet and 139 plant species native to China) based on three different methods. Our results indicated that ITS2 displayed largest intra- and interspecific divergence (1.72% and 91.46%). Based on NJ tree method, ITS2, ITS, matK, rbcL and trnH-psbA provided 76.84%, 76.5%, 63.21%, 52.86% and 50.68% discrimination rates, respectively. The combination of ITS + matK performed best and provided 91.03% discriminatory power, followed by ITS2 + matK (85.78%). For identifying unknown individuals, ITS + matK had 100% correct identification rate based on our database, followed by ITS/ITS2 (both 93.33%) and ITS2 + matK (91.67%). Thus, we propose ITS/ITS2 + matK as the most suitable barcode for invasive plants in China. This study also demonstrated that DNA barcoding is an efficient tool for identifying invasive species.

Phylogenetic assessment of Chromocyphellaceae (Agaricineae, Basidiomycota) and a new lamellate species of Chromocyphella.

Cyphelloid fungi represent a polyphyletic assemblage of reduced agarics, including the brown-spored family Chromocyphellaceae. In order to elucidate the phylogenetic position of Chromocyphellaceae, newly generated sequences of Chromocyphella were included in a multigene alignment of the Agaricineae and phylogenetically analyzed. The current analyses show that the Chromocyphella muscicola specimen used to phylogenetically place Chromocyphellaceae in its original description was misidentified and that the Chromocyphellaceae nests in the Hymenogastraceae, Chromocyphella being sister to Flammula. Chromocyphella is emended, including now a new species with lamellate and stipitate basidiomata, C. lamellata. The name Cymbella crouanii, type species of Chromocyphella, is lecto- and epitypified. Our analyses support a new origin of cyphelloid fungi. The shift to a cyphelloid basidioma from an agaric ancestor is suggested to have occurred in two evolutionary steps in Chromocyphella, an initial reduction in basidioma size and a subsequent loss of lamellae and stipe.

Assessment of species richness in Lake Dal, Kashmir, based on classical approach, physiological approach and rDNA ITS sequences from isolates.

As a first description to document the species richness in Dal Lake, a freshwater lake ecosystem in Kashmir valley, an extensive network of sixteen sampling stations with distinguishing features was sampled seasonally for two years. The identification process yielded fifty-one species probably first and new records for this area to date. The taxonomic groups observed were those with species from Ascomycetes (inclusive of yeasts), Basidiomycetes, Blastocladiomycetes, Zygomycetes, and Peronosporomycetes. Each phylum was represented by a single Order, with the exception of the Peronosporomycetes, which was represented by two Orders- Saprolegniales and Pythiales. In the filamentous fungal group, family Trichocomaceae was dominant followed by Saccharomycetaceae, Mucoraceae, Nectriaceae, Tremellaceae and Hypocreaceae. However, in the group of zoosporic & fungal like eukaryotes, family Saprolegniaceae was most dominant followed by Blastocladiaceae and Pythiaceae. A dramatic decrease in fungal load was observed in different seasons with highest colonial load in the summer season and lowest in the winter season. The observed distribution was statistically significant for both the filamentous fungal species (p < 0.01) as well as zoosporic fungi & fungal like eukaryotes (p < 0.05). In order to assess biodiversity patterns of fungi more accurately, it is necessary to repeat such investigations in other areas and to improve the tools for taxonomic identification of these highly diverse but mostly microscopic organisms.

Molecular identification of Candida species isolated from cases of neonatal candidemia using polymerase chain reaction-restriction fragment length polymorphism in a tertiary care hospital.

CONTEXT: Candida spp. is an emerging cause of bloodstream infections worldwide. Delay in speciation of Candida isolates by conventional methods and resistance to antifungal drugs in various Candida species are responsible for the increase in morbidity and mortality due to candidemia. Hence, the rapid identification of Candida isolates is very important for the proper management of patients with candidemia. AIMS: The aim was to re-evaluate the identification of various Candida spp. by polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) and to evaluate the accuracy, speed, and cost of phenotypic methodology versus PCR-RFLP. SETTINGS AND DESIGN: Hospital-based cross-sectional study. MATERIALS AND METHODS: Ninety consecutive clinical isolates of seven Candida species, isolated from blood of neonates and identified by routine phenotypic methods, were re-evaluated using universal primers internal transcribed spacer 1 (ITS1) and ITS4 for PCR amplification and Msp I restriction enzyme for RFLP. STATISTICAL ANALYSIS USED: Kappa test for agreement. RESULTS: The results of PCR-RFLP were 100% in agreement with those obtained using conventional phenotypic methods. Identification could be achieved within 3 work days by both the methods. Our routine methods proved to be cost effective than PCR-RFLP. CONCLUSIONS: We can continue with our routine phenotypic methods and PCR-RFLP can be used for periodic quality control or when conventional methods fail to identify a species.

Molecular identification of Ancylostoma species from dogs and an assessment of zoonotic risk in low-income households, São Paulo State, Brazil.

Hookworm infection stands out for its worldwide distribution and for its veterinary and public health relevance. Based on copromicroscopic examinations and polymerase chain reaction (PCR) amplification of the ITS1-5.8S-ITS2 region, we assessed, respectively, the prevalence of intestinal parasites and the identification of canine hookworm species in faeces recovered from 278 dogs living in households of an inland municipality of São Paulo State, Brazil. Intestinal parasites were found in 67.3% of dogs and hookworm infection was found at the highest prevalence rate (56.6%), followed by Toxocara canis (11.9%), Isospora spp. (11.9%), Giardia spp. (5.8%), Sarcocystis spp. (4.0%), 'Hammondia-like' (1.4%), Dipylidium caninum (1.1%) and Trichuris vulpis (0.7%). Of 158 samples positive for hookworm eggs, 106 (67.1%) were amplified by PCR and, of those, 88 (55.7%) were successfully sequenced for species identification. Single infections with Ancylostoma caninum and Ancylostoma braziliense were recorded in 61.4% and 12.5%, respectively, and mixed infections were found in 26.1%. The nucleotide sequences of both species showed high identity rates (98-100%) when compared with reference sequences. Although A. caninum was the most prevalent hookworm in the dogs assessed, the occurrence of both A. caninum and A. braziliense in single and/or mixed infections poses a potential risk for the local population in a low-income area, especially children, to acquire cutaneous larva migrans (CLM).

Identification of Chinese Caterpillar Medicinal Mushroom, Ophiocordyceps sinensis (Ascomycetes) from Counterfeit Species.

Ophiocordyceps sinensis is a valuable traditional Chinese medicine with a high market price. In this study, a polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP) method based on 2 enzymes was developed to distinguish O. sinensis from 6 common counterfeit species. To verify the applicability of this method, we experimentally tested O. sinensis organisms, tablet preparations made from O. sinensis, and cultured mycelia isolated from O. sinensis. To validate the results from this PCR-RFLP method, all real samples were identified by internal transcribed spacer sequencing. This is, to our knowledge, the first time the PCR-RFLP method has been applied to identify O. sinensis. The selection of 2 restrictive enzymes for identification dramatically improved the accuracy and efficiency of this method. It is the great advantage of this method that sampling from either of 2 parts of O. sinensis-the fruiting body or the caterpillar body-would not cause any difference in the final experimental results. Therefore, this method is not only feasible for testing crude drugs of O. sinensis but it is also useful when the crude drugs are broken down into powder or made into tablets, demonstrating the promising prospect of application in quality control.

Phylogenetic assessment of global Suillus ITS sequences supports morphologically defined species and reveals synonymous and undescribed taxa.

The genus Suillus represents one of the most recognizable groups of mushrooms in conifer forests throughout the Northern Hemisphere. Although for decades the genus has been relatively well defined morphologically, previous molecular phylogenetic assessments have provided important yet preliminary insights into its evolutionary history. We present the first large-scale phylogenetic study of the boundaries of each species in the genus Suillus based on the most current internal transcribed spacer (ITS) barcode sequences available inPUBLIC databases, as well as sequencing of 224 vouchered specimens and cultures, 15 of which were type specimens from North America. We found that species boundaries delimited by morphological data are broadly congruent with those based on ITS sequences. However, some species appear to have been described several times under different names, several species groups cannot be resolved by ITS sequences alone, and undescribed taxa are apparent, especially in Asia. Therefore, we elevated S. tomentosus var. discolor to S. discolor; proposed synonymies of S. neoalbidipes with S. glandulosipes, S. borealis with S. brunnescens, Boletus serotinus and B. solidipes with Suillus elbensis, S. lactifluus with S. granulatus, S. himalayensis with S. americanus; and proposed usage of the names S. clintonianus in the place of the North American S. grevillei, S. weaverae for North American S. granulatus, S. ampliporus in the place of the North American S. cavipes, and S. elbensis in place of the North American S. viscidus. We showed that the majority of Suillus species have strong affinities for particular host genera. Although deep node support was low, geographic differentiation was apparent, with species from North America, Eurasia, and Asia often forming their own clades. Collectively, this comprehensive genus-level phylogenetic integration of currently available Suillus ITS molecular data and metadata will aid future taxonomic and ecological work on an important group of ectomycorrhizal fungi.

Yucatán in black and red: Linking edaphic analysis and pyrosequencing-based assessment of bacterial and fungal community structures in the two main kinds of soil of Yucatán State.

Yucatán State is dominated by two kinds of soil, named "Black Leptosol" and "Red Leptosol", which are interwoven across the State. In this work, we analyzed the relation between the edaphic characteristics and the bacterial and fungal community structures in these two kinds of Leptosol. The results revealed that Black Leptosol (BlaS) had a higher content of calcium carbonates, organic matter, nitrogen, and phosphorus than Red Leptosol (RedS). The most outstanding difference in the bacterial community structure between BlaS and RedS was that while in BlaS Actinobacteria was the most abundant phylum (43.7%), followed by Acidobacteria (26.9%) and Proteobacteria (23.6%), in RedS the bacterial community was strongly dominated by Acidobacteria (83%). Two fungal phyla were identified in both kinds of soil; Ascomycota, with 77% in BlaS and 56% in RedS, and Basidiomycota, with 22% in RedS and only 0.67% in BlaS. The most relevant difference between the two fungal communities was that excepting for Fusarium sp., all the species they had were different. Thus, in contrast with bacterial communities, where most of the major OTUs were present in both kinds of soil, fungal communities appeared to be unique to each kind of Leptosol.

Human fascioliasis endemic areas in Argentina: multigene characterisation of the lymnaeid vectors and climatic-environmental assessment of the transmission pattern.

BACKGROUND: In South America, fascioliasis stands out due to the human endemic areas in many countries. In Argentina, human endemic areas have recently been detected. Lymnaeid vectors were studied in two human endemic localities of Catamarca province: Locality A beside Taton and Rio Grande villages; Locality B close to Recreo town. METHODS: Lymnaeids were characterised by the complete sequences of rDNA ITS-2 and ITS-1 and fragments of the mtDNA 16S and cox1. Shell morphometry was studied with the aid of a computer image analysis system. Climate analyses were made by nearest neighbour interpolation from FAO data. Koeppen & Budyko climate classifications were used. De Martonne aridity index and Gorczynski continentality index were obtained. Lymnaeid distribution was assessed in environmental studies. RESULTS: DNA sequences demonstrated the presence of Lymnaea neotropica and L. viator in Locality A and of L. neotropica in Locality B. Two and four new haplotypes were found in L. neotropica and L. viator, respectively. For interspecific differentiation, ITS-1 and 16S showed the highest and lowest resolution, respectively. For intraspecific analyses, cox1 was the best marker and ITS-1 the worst. Shell intraspecific variability overlapped in both species, except maximum length which was greater in L. viator. The desertic-arid conditions surrounding Locality A, the semiaridity-aridity surrounding Locality B, and the very low yearly precipitation in both localities, are very different from the typical fascioliasis transmission foci. Lymnaeids are confined to lateral river side floodings and small man-made irrigation systems. Water availability only depends on the rivers flowing from neighbouring mountains. All disease transmission factors are concentrated in small areas where humans and animals go for water supply, vegetable cultures and livestock farming. CONCLUSIONS: The unusually high number of DNA haplotypes and the extreme climate unsuitable for F. hepatica and lymnaeid development, demonstrate that the transmission foci are isolated. Seasonal transmission may depend on the timely overlap of appropriate temperature and river water availability. Lymnaeids and F. hepatica have probably reached these localities by livestock introduction. DNA differences regarding other populations of L. neotropica and L. viator in Argentina suggest an introduction independent from the spreading movements which allowed these two lymnaeids to expand throughout the country.

Molecular inference, multivariate morphometrics and ecological assessment are applied in concert to delimit species in the Russula clavipes complex.

Species of Russula subsect. Xerampelinae are notoriously difficult to identify and name and have not been subject to molecular study. A group of species, referred to here as the R. clavipes complex, growing in association with Salix, Betula and Populus as well as coniferous tree species from temperate to arctic and alpine habitats, were examined. Analyses of the nuc rDNA internal transcribed spacer (ITS) region and a numerical analysis of morphological characters were used. The R. clavipes complex is a monophyletic group within Russula subsect. Xerampelinae, according to molecular results. The complex includes three species: R. nuoljae is a phylogenetically and morphologically well-supported species while the other two, R. clavipes and R. pascua, are similar based on ITS data and morphology but separate based on their ecology. Russula pseudoolivascens is conspecific with R. clavipes Several combinations of characters traditionally used in the taxonomy of R. subsect. Xerampelinae are inappropriate for species delimitation in this group and the adequacy of the ITS for species identification in this group is discussed. Detailed microscopic observations on the type collection of R. nuoljae are presented and illustrated, along with a key to the European members of R. subsect. Xerampelinae.

Comparative analysis of microsatellites in five different antagonistic Trichoderma species for diversity assessment.

Microsatellites provide an ideal molecular markers system to screen, characterize and evaluate genetic diversity of several fungal species. Currently, there is very limited information on the genetic diversity of antagonistic Trichoderma species as determined using a range of molecular markers. In this study, expressed and whole genome sequences available in public database were used to investigate the occurrence, relative abundance and relative density of SSRs in five different antagonistic Trichoderma species: Trichoderma atroviride, T. harzianum, T. reesei, T. virens and T. asperellum. Fifteen SSRs loci were used to evaluate genetic diversity of twenty isolates of Trichoderma spp. from different geographical regions of India. Results indicated that relative abundance and relative density of SSRs were higher in T. asperellum followed by T. reesei and T. atroviride. Tri-nucleotide repeats (80.2%) were invariably the most abundant in all species. The abundance and relative density of SSRs were not influenced by the genome sizes and GC content. Out of eighteen primer sets, only 15 primer pairs showed successful amplification in all the test species. A total of 24 alleles were detected and five loci were highly informative with polymorphism information content values greater than 0.40, these markers provide useful information on genetic diversity and population genetic structure, which, in turn, can exploit for establishing conservation strategy for antagonistic Trichoderma isolates.