Fusarium Species Associated with Fusarium Head Blight in Hungarian Wheat Fields.

The species composition of the genus Fusarium associated with Fusarium head blight (FHB) in wheat fields of Hungary in the year 2019 was assessed. Symptomatic wheat heads were collected at 20 geographical locations representing different ecosystems. A total of 256 Fusarium strains were isolated and identified by partial sequences of the translation elongation factor 1-alpha gene and, where required, the second-largest subunit of the DNA-directed RNA polymerase gene. Overall, Fusarium graminearum (58.2%) proved to be the dominant species, followed by F. annulatum (formerly F. proliferatum) (17.2%) and F. verticillioides (7.4%). The presence of all other species, including F. culmorum, in the population was less than 5%. F. graminearum was identified as the main species associated with FHB at 14 sampling sites. Fumonisin-producing F. annulatum, primarily known as the pathogen of maize in Hungary, was detected nearly as frequently as F. graminearum at three locations and dominated at two other sites. F. poae was not found during the survey. F. vorosii, a species that is believed to be of Asian origin and was already found in Hungary in 2002, was identified at two locations.

Characterizing genetic diversity of Sclerotium rolfsii isolates by biomapping of mycelial compatibility groupings and multilocus sequence analysis.

Genetic diversity in Sclerotium rolfsii is useful for understanding its population structure, identifying different mycelial compatibility groups (MCGs), and developing targeted strategies for disease management in affected crops. In our study, a comprehensive genetic analysis was conducted on 50 isolates of S. rolfsii, collected from various geographic regions and host plants. Two specific genes, TEF1α and RPB2, were utilized to assess the genetic diversity and relationships among these isolates. Notably, out of 1225 pairings examined, only 154 exhibited a compatible reaction, while the majority displayed antagonistic reactions, resulting in the formation of a barrier zone. The isolates were grouped into 10 distinct MCGs. These MCGs were further characterized using genetic sequencing. TEF1α sequences distinguished the isolates into 17 distinct clusters, and RPB2 sequences classified them into 20 clusters. Some MCGs shared identical gene sequences within each gene, while others exhibited unique sequences. Intriguingly, when both TEF1α and RPB2 sequences were combined, all 10 MCGs were effectively differentiated, even those that appeared identical with single-gene analysis. This combined approach provided a comprehensive understanding of the genetic diversity and relationships among the S. rolfsii isolates, allowing for precise discrimination between different MCGs. The results shed light on the population structure and genetic variability within this plant pathogenic fungus, providing valuable insights for disease management and control strategies. This study highlights the significance of comprehending the varied virulence characteristics within S. rolfsii isolates, categorizing them into specific virulence groups based on disease severity index (DSI) values. The association with MCGs provides additional insights into the genetic underpinnings of virulence in this pathogen. Furthermore, the identification of geographical patterns in virulence implies the influence of region-specific factors, with potential implications for disease control and crop protection strategies.Please confirm if the author names are presented accurately and in the correct sequence (given name, middle name/initial, family name). Author 1 Given name: [G. M. Sandeep] Last name [Kumar]. Author 2 Given name: [Praveen Kumar] Last name [Singh]. Also, kindly confirm the details in the metadata are correct.I confirm that the given names are accurate and presented in the correct sequence.

The RNA polymerase II subunit B (RPB2) functions as a growth regulator in human glioblastoma.

Glioblastoma multiforme (GBM) is the most common and aggressive brain tumor with a poor prognosis. The growth of GBM cells depends on the core transcriptional apparatus, thus rendering RNA polymerase (RNA pol) complex as a candidate therapeutic target. The RNA pol II subunit B (POLR2B) gene encodes the second largest subunit of the RNA pol II (RPB2); however, its genomic status and function in GBM remain unclear. Certain GBM data sets in cBioPortal were used for investigating the genomic status and expression of POLR2B in GBM. The function of RPB2 was analyzed following knockdown of POLR2B expression by shRNA in GBM cells. The cell counting kit-8 assay and PI staining were used for cell proliferation and cell cycle analysis. A xenograft mouse model was established to analyze the function of RPB2 in vivo. RNA sequencing was performed to analyze the RPB2-regulated genes. GO and GSEA analyses were applied to investigate the RPB2-regulated gene function and associated pathways. In the present study, the genomic alteration and overexpression of the POLR2B gene was described in glioblastoma. The data indicated that knockdown of POLR2B expression suppressed tumor cell growth of glioblastoma in vitro and in vivo. The analysis further demonstrated the identification of the RPB2-regulated gene sets and highlighted the DNA damage-inducible transcript 4 gene as the downstream target of the POLR2B gene. The present study provides evidence indicating that RPB2 functions as a growth regulator in glioblastoma and could be used as a potential therapeutic target for the treatment of this disease.

Population Genetic Analysis of Fusarium decemcellulare, a Guaraná Pathogen, Reveals High Genetic Diversity in the Amazonas State, Brazil.

Guaraná is indigenous to the Brazilian Amazon where it has cultural and agroeconomic significance. However, its cultivation is constrained by a disease termed oversprouting of guaraná caused by Fusarium decemcellulare, with yield losses reaching as high as 100%. The disease can affect different parts of the plant, causing floral hypertrophy and hyperplasia, stem galls, and oversprouting of vegetative buds. To date, no study has been conducted characterizing the genetic diversity and population structure of this pathogen. Here, we report genetic diversity and genetic structure among 224 isolates from eight guaraná production areas of Amazonas State, Brazil, that were genotyped using a set of 10 inter-simple-sequence repeat (ISSR) markers. Despite moderate gene diversity (Hexp = 0.21 to 0.32), genotypic diversity was at or near maximum (223 multilocus genotypes among 224 isolates). Population genetic analysis of the 10 ISSR marker fragments with STRUCTURE software identified two populations designated C1 and C2 within the F. decemcellulare collection from the eight sites. Likewise, UPGMA hierarchical clustering and discriminant analysis of principal components of the strains from guaraná resolved these same two groups. Analysis of molecular variance demonstrated that 71% of genetic diversity occurred within the C1 and C2 populations. A pairwise comparison of sampling sites for both genetic populations revealed that 59 of 66 were differentiated from one another (P < 0.05), and high and significant gene flow was detected only between sampling sites assigned to the same genetic population. The presence of MAT1-1 and MAT1-2 strains, in conjunction with the high genotypic diversity and no significant linkage disequilibrium, suggests that each population of F. decemcellulare might be undergoing sexual reproduction. Isolation by distance was not observed (R2 = 0.02885, P > 0.05), which suggests that human-mediated movement of seedlings may have played a role in shaping the F. decemcellulare genetic structure in Amazonas State, Brazil.

Bioactive Polyketides from the Natural Complex of the Sea Urchin-Associated Fungi Penicillium sajarovii KMM 4718 and Aspergillus protuberus KMM 4747.

The marine-derived fungal strains KMM 4718 and KMM 4747 isolated from sea urchin Scaphechinus mirabilis as a natural fungal complex were identified as Penicillium sajarovii and Aspergillus protuberus based on Internal Transcribed Spacer (ITS), partial ß-tubulin (BenA), and calmodulin (CaM) molecular markers as well as an ribosomal polymerase two, subunit two (RPB2) region for KMM 4747. From the ethyl acetate extract of the co-culture, two new polyketides, sajaroketides A (1) and B (2), together with (2'S)-7-hydroxy-2-(2'-hydroxypropyl)-5-methylchromone (3), altechromone A (4), norlichexanthone (5), griseoxanthone C (6), 1,3,5,6-tetrahydroxy-8-methylxanthone (7), griseofulvin (8), 6-O-desmethylgriseofulvin (9), dechlorogriseofulvin (10), and 5,6-dihydro-4-methyl-2H-pyran-2-one (11) were identified. The structures of the compounds were elucidated using spectroscopic analyses. The absolute configurations of the chiral centers of sajaroketides A and B were determined using time-dependent density functional theory (TDDFT)-based calculations of the Electronic Circular Dichroism (ECD) spectra. The inhibitory effects of these compounds on urease activity and the growth of Staphylococcus aureus, Escherichia coli, and Candida albicans were observed. Sajaroketide A, altechromone A, and griseofulvin showed significant cardioprotective effects in an in vitro model of S. aureus-induced infectious myocarditis.

First Report of Didymella glomerata Causing Didymella Leaf Blight on Maize in China.

Maize (Zea mays L.) is a staple food crop worldwide. In July 2021, gray leaf blight was observed on maize leaves in a field located in Panjin (41°7'11.98" N, 122°4'14.57" E), Liaoning Province, China. Nearly 5% of the maize plants were affected in the field. The leaves of the affected plants showed oval to oblong, gray, sunken lesions with yellow or tan margins. The lesions were scattered all over the leaf surface; however, they were absent on the stalks and other parts of the affected plants. To isolate the pathogen, leaf discs (1.25 mm2) excised from the blight lesions were surface-sterilized with 70% ethanol for 30 seconds, followed by 20% NaOCl for 2 minutes and finally rinsed three times with sterilized water. The discs were cultured on potato dextrose agar (PDA) plates supplemented with streptomycin (100 mg/L) and incubated at 25oC under a 12-h photoperiod for 7 days. Six single spore isolates (two per sampled infected leaf) were purified from the PDA culture plates. The fungal colonies of three selected isolates (one per sampled infected leaf; Pj-1, Pj-2, and Pj-3) were dark brown on the PDA plates and devoid of aerial hyphae; all three isolates grew 11 mm/day on the PDA plates. The number of conidia produced by the isolates on the 6-cm PDA plates 7 days after incubation was ranged from 160 x 108 to 208 x 108 (n = 36). Conidia were hyaline, single-celled and ellipsoidal (3.35-3.56 µm [width] x 6.47-6.70 [length] µm; n = 36). To identify the pathogen, four loci, i.e., 28S subunit (large subunit [LSU]) of the nuclear ribosomal (nr) DNA, internal transcribed spacer (ITS) region (ITS1, 5.8S subunit of nrDNA, and ITS2), the second-largest subunit of RNA polymerase II (rpb2) and ß-tubulin (tub2) were amplified using the primer sets described in the study by Chen el al. 2015. BLASTn search against GenBank revealed that the four amplicon sequences originating from Pj-1, Pj-2, and Pj-3 showed 99-100% homology to the type strain CBS 528.66 of D. glomerata. A phylogenetic tree deduced from a maximum likelihood analysis of a concatenated MUSCLE-based alignment of LSU, ITS region, rpb2, and tub2 sequences of 12 isolates/strains showed that the Pj isolates clustered together with CBS 528.66, along with other D. glomerata isolates/strains, with a high bootstrap support value (i.e., 99). Based on both morphological characteristics and molecular phylogeny, Pj-1, Pj-2, and Pj-3 were identified as the D. glomerata isolates. Since the amplicon sequences of the three isolates were identical, only Pj-2 sequences were deposited in GenBank with accession numbers OM372474 (LSU), OK485138 (ITS), OM406188 (rpb2), and OK485135 (tub2). To confirm pathogenicity, 14-day-old plants (V3 growth stage) of a maize cultivar P178 were spray-inoculated with the Pj-2 conidia (1 x 107 conidia/mL) in a growth chamber. The inoculated leaves exhibited typical gray leaf blight lesions (similar to those detected in the maize field) 7 days post-inoculation at 25oC and 95-100% humidity under a 12-h photoperiod, whereas the leaves spray-inoculated with sterilized water remained healthy. The pathogenicity assay was repeated three times; the pathogen was re-isolated from the inoculated leaves each time and confirmed by the morphological characteristics and the molecular phylogeny based on the four loci to be D. glomerata, fulfilling Koch's postulates. This first report of D. glomerata causing Didymella leaf blight on maize will help develop robust disease management strategies against this emerging fungal pathogen.

FUSARIUM-ID v.3.0: An Updated, Downloadable Resource for Fusarium Species Identification.

Species within Fusarium are of global agricultural, medical, and food/feed safety concern and have been extensively characterized. However, accurate identification of species is challenging and usually requires DNA sequence data. FUSARIUM-ID (http://isolate.fusariumdb.org/blast.php) is a publicly available database designed to support the identification of Fusarium species using sequences of multiple phylogenetically informative loci, especially the highly informative ∼680-bp 5' portion of the translation elongation factor 1-alpha (TEF1) gene that has been adopted as the primary barcoding locus in the genus. However, FUSARIUM-ID v.1.0 and 2.0 had several limitations, including inconsistent metadata annotation for the archived sequences and poor representation of some species complexes and marker loci. Here, we present FUSARIUM-ID v.3.0, which provides the following improvements: (i) additional and updated annotation of metadata for isolates associated with each sequence, (ii) expanded taxon representation in the TEF1 sequence database, (iii) availability of the sequence database as a downloadable file to enable local BLAST queries, and (iv) a tutorial file for users to perform local BLAST searches using either freely available software, such as SequenceServer, BLAST+ executable in the command line, and Galaxy, or the proprietary Geneious software. FUSARIUM-ID will be updated on a regular basis by archiving sequences of TEF1 and other loci from newly identified species and greater in-depth sampling of currently recognized species.

Weeds Harbor Fusarium Species that Cause Malformation Disease of Economically Important Trees in Western Mexico.

Mango malformation disease (MMD) caused by Fusarium spp. is an important limiting factor in most production areas worldwide. Fusarium mexicanum and F. pseudocircinatum have been reported as causing MMD in Mexico. These two pathogens also cause a similar disease in Swietenia macrophylla (big-leaf mahogany malformation disease) in central western Mexico, and F. pseudocircinatum was recently reported as causing malformation disease in Tabebuia rosea (rosy trumpet) in the same region. These studies suggest that additional plant species, including weeds, might be hosts of these pathogens. The role that weed hosts might have in the disease cycle is unknown. The objectives of this work were to recover Fusarium isolates from understory vegetation in mango orchards with MMD, identify the Fusarium isolates through DNA sequence data, and determine whether F. mexicanum is capable of inducing disease in the weedy legume Senna uniflora (oneleaf senna). Additional objectives in this work were to compare Fusarium isolates recovered from weeds and mango trees in the same orchards by characterizing their phylogenetic relationships, assessing in vitro production of mycotoxins, and identifying their mating type idiomorph. A total of 59 Fusarium isolates from five species complexes were recovered from apical and lateral buds from four weed species. Two of the species within the F. fujikuroi species complex are known to cause MMD in Mexico. Trichothecene production was detected in five isolates, including F. sulawense and F. irregulare in the F. incarnatum-equiseti species complex and F. boothii in the F. sambucinum species complex. Both mating types were present among mango and weed isolates. This is the first report of herbaceous hosts harboring Fusarium species that cause mango malformation in Mexico. The information provided should prove valuable for further study of the epidemiological role of weeds in MMD and help manage the disease.

DNA Sequence-Based Identification of Fusarium: A Work in Progress.

Accurate species-level identification of an etiological agent is crucial for disease diagnosis and management because knowing the agent's identity connects it with what is known about its host range, geographic distribution, and toxin production potential. This is particularly true in publishing peer-reviewed disease reports, where imprecise and/or incorrect identifications weaken the public knowledge base. This can be a daunting task for phytopathologists and other applied biologists that need to identify Fusarium in particular, because published and ongoing multilocus molecular systematic studies have highlighted several confounding issues. Paramount among these are: (i) this agriculturally and clinically important genus is currently estimated to comprise more than 400 phylogenetically distinct species (i.e., phylospecies), with more than 80% of these discovered within the past 25 years; (ii) approximately one-third of the phylospecies have not been formally described; (iii) morphology alone is inadequate to distinguish most of these species from one another; and (iv) the current rapid discovery of novel fusaria from pathogen surveys and accompanying impact on the taxonomic landscape is expected to continue well into the foreseeable future. To address the critical need for accurate pathogen identification, our research groups are focused on populating two web-accessible databases (FUSARIUM-ID v.3.0 and the nonredundant National Center for Biotechnology Information nucleotide collection that includes GenBank) with portions of three phylogenetically informative genes (i.e., TEF1, RPB1, and RPB2) that resolve at or near the species level in every Fusarium species. The objectives of this Special Report, and its companion in this issue (Torres-Cruz et al. 2022), are to provide a progress report on our efforts to populate these databases and to outline a set of best practices for DNA sequence-based identification of fusaria.

More smooth-spored species of Inocybe (Agaricales, Ba sidiomycota): type studies and 12 new species from Europe.

Twelve new species of Inocybe (I. adorabilis, I. comis, I. demetris, I. filiana, I. galactica, I. morganae, I. othini, I. ovilla, I. proteica, I. somae, I. suryana and I. venerabilis) are described from Europe on the basis of detailed morphological and molecular investigation. A portrait of the recently described I. ianthinopes is given. All species are smooth-spored and some pruinose only in the apical part of the stipe, and some on entire length. The new species are compared to 24 type specimens (17 characterized by at least partial ITS sequence data), all of which are described and revised here. Epitypes were selected for two species, I. hirtella and I. sindonia. Based on our studies, we confirm that I. kuehneri and I. sindonia on one hand, and I. subalbidodisca and I. ochroalba on the other, are synonyms and furthermore suggest that I. abietis is synonymous with I. catalaunica, I. exilis with I. rufobrunnea, I. hirtellarum with I. mycenoides, I. lapidicola with I. deianae, I. ochraceolutea with I. sindonia, I. stangliana with I. pelargonium, I. subrubens with I. subhirtella and I. sulfovirescens with I. langei. All of the new species are supported by phylogenetic analyses. Among the 16 previously described species accepted here, 10 are represented by types in the phylogenetic analyses and five by own collections corresponding to the type. Two species, I. eutheloides (remaining doubtful) and I. pallidolutea are only treated morphologically. In summary, we describe as new or verify the taxonomic status and provide or corroborate morphological concepts for 37 smooth-spored species of Inocybe. Citation: Bandini D, Oertel B, Eberhardt U. 2022. More smooth-spored species of Inocybe (Agaricales, Basidiomycota): type studies and 12 new species from Europe. Persoonia 48: 91-149. https://doi.org/10.3767/persoonia.2022.48.03.

Malformation Disease in Tabebuia rosea (Rosy Trumpet) Caused by Fusarium pseudocircinatum in Mexico.

Tabebuia rosea (rosy trumpet) is an economically important neotropical tree in Mexico that is highly valued for the quality of its wood, which is used for furniture, crafts, and packing, and for its use as an ornamental and shade tree in parks and gardens. During surveys conducted in the lower Balsas River Basin region in the states of Guerrero and Michoacán, symptoms of floral malformation were detected in T. rosea trees. The main objectives of this study were to describe this new disease, to determine its causal agent, and to identify it using DNA sequence data. A second set of objectives was to analyze the phylogenetic relationship of the causal agent to Fusarium spp. associated with Swietenia macrophylla trees with malformation surveyed in the same region and to compare mycotoxin production and the mating type idiomorphs of fusaria recovered from T. rosea and S. macrophylla. Tabebuia rosea showed malformed inflorescences with multiple tightly curled shoots and shortened internodes. A total of 31 Fusarium isolates recovered from symptomatic T. rosea (n = 20) and S. macrophylla (n = 11) trees were identified by molecular analysis as Fusarium pseudocircinatum. Pathogenicity tests showed that isolates of F. pseudocircinatum recovered from T. rosea induced malformation in inoculated T. rosea seedlings. Eighteen F. pseudocircinatum isolates were tested for their ability to produce mycotoxins and other secondary metabolites. Moniliformin, fusaric acid, bikaverin, beauvericin, aurofusarin. and 8-O-methylbostrycoidin were produced by at least one strain of the 18 isolates tested. A multiplex PCR assay for mating type idiomorph revealed that 22 F. pseudocircinatum isolates were MAT1-1 and that 9 were MAT1-2. Here, we report a new disease of T. rosea in Mexico caused by F. pseudocircinatum.

Updating the taxonomy of Aspergillus in South Africa.

The taxonomy and nomenclature of the genus Aspergillus and its associated sexual (teleomorphic) genera have been greatly stabilised over the last decade. This was in large thanks to the accepted species list published in 2014 and associated metadata such as DNA reference sequences released at the time. It had a great impact on the community and it has never been easier to identify, publish and describe the missing Aspergillus diversity. To further stabilise its taxonomy, it is crucial to not only discover and publish new species but also to capture infraspecies variation in the form of DNA sequences. This data will help to better characterise and distinguish existing species and make future identifications more robust. South Africa has diverse fungal communities but remains largely unexplored in terms of Aspergillus with very few sequences available for local strains. In this paper, we re-identify Aspergillus previously accessioned in the PPRI and MRC culture collections using modern taxonomic approaches. In the process, we re-identify strains to 63 species, describe seven new species and release a large number of new DNA reference sequences.

The phoma-like dilemma.

Species of Didymellaceae have a cosmopolitan distribution and are geographically widespread, occurring in diverse ecosystems. The family includes several important plant pathogenic fungi associated with fruit, leaf, stem and root diseases on a wide variety of hosts, as well as endophytic, saprobic and clinically relevant species. The Didymellaceae was recently revised based on morphological and phylogenetic analyses of ex-type strains subjected to DNA sequencing of partial gene data of the LSU, ITS, rpb2 and tub2 loci. Several poly- and paraphyletic genera, including Ascochyta, Didymella and Phoma were redefined, along with the introduction of new genera. In the present study, a global collection of 1 124 Didymellaceae strains from 92 countries, 121 plant families and 55 other substrates, including air, coral, human tissues, house dust, fungi, insects, soil, and water were examined via multi-locus phylogenetic analyses and detailed morphological comparisons, representing the broadest sampling of Didymellaceae to date. Among these, 97 isolates representing seven new genera, 40 new species and 21 new combinations were newly introduced in Didymellaceae. In addition, six epitypes and six neotypes were designated to stabilise the taxonomy and use of older names. A robust, multi-locus reference phylogenetic tree of Didymellaceae was generated. In addition, rpb2 was revealed as the most effective locus for the identification of Didymellaceae at species level, and is proposed as a secondary DNA marker for the family.

First Report of Aspergillus niger causing Black rot of Grapes in Pakistan.

In June 2015 & 2016, a postharvest survey of table grapes (Vitis vinifera) cv. King's Ruby, was carried out in five different commercial fruit markets of Rawalpindi (33°38'19.2″N, 73°01'45.0″E) district, Punjab Province. Symptoms appeared as brownish lesions with black sporulation on grapes berries. The incidence of these symptoms on bunches ranged from 12 to 17% at all sites. Symptomatic tissue pieces were surface-sterilized with 0.1% sodium hypochlorite (NaOCl) for 30 seconds, rinsed three times with sterile distilled water, dried on filter paper for 45 seconds, and incubated on potato dextrose agar (PDA) at 25°C. After 3 days, dark brown to black mycelium were formed on PDA media. A total of 24 isolates were examined morphologically. The apex of the conidiophore was observed to be radiate. Vesicles were found to be spherical and covered with irregular metulae and phialides. Conidia were globose or subglobose measured (3.14 µm ± 2.24 in averaged diameter: n=50), dark brown to black, with roughened cell walls. The conidiophores were also smooth-walled, hyaline, and became melanized toward the vesicle. These characteristics of the fungus were similar to those described for Aspergillus niger van Tiegh (de Hoog et al. 2000). For molecular identification, the internal transcribed spacer (ITS) region, beta-tubulin (Bt) gene and partial RNA polymerase II largest subunit (RPB2) gene of representative isolate (Asp.n02) was amplified using primers ITS1/ITS4, BT2a/BT2b and RPB2-6F/RPB2-7R respectively (White et al., 1990; Glass & Donaldson, 1995; Liu et al. 1999). Sequences were deposited in GenBank (ITS, MN658871; Bt2, MT117924; and RPB2, MT318289). Based on BLAST analysis, sequences of the ITS region, Bt2 genes, and RPB2 gene showed 99 to 100% similarity of isolate Asp.n02 to Aspergillus niger (Accession Nos. MK307680.1, MN195121.1, MF078661.1 for ITS gene, MN567299.1, MK451029.1, MK451020.1 for Bt2 gene, and MK450788.1, MK450790.1 for RPB2 gene). To complete Koch's postulates, 10-µl aliquots of spore suspensions (106 spores/ml) of isolate: Asp.n 02 was pipetted onto three non-wounded and four wounded (5 mm diam) asymptomatic grape berries cv. King's Ruby (seven berries per isolate), Sterile distilled water was applied to asymptomatic berries similaries to serve as a negative control (Ghuffar et al. 2018; Jayawardena et al. 2018). Berries were incubated at 25 ± 2°C in sterile moisture chambers, and the experiment was conducted twice. Brownish lesions leading to black sporulation similar to the original symptoms were observed on both wounded and non-wounded inoculated berries after 3 days, whereas no symptoms were recorded on the negative control. The morphology of the fungus that was re-isolated from each of the inoculated berries was identical to that of the original cultures. Aspergillus niger was reported previously in Europe and Israel causing mycotoxin (Ochratoxin A) OTA production on Table grapes (Bau et al. 2006). To our knowledge, this is the first report of Aspergillus niger causing black rot of grapes in Pakistan. This finding will help to plan effective disease management strategies against the black rot of grapes in Pakistan.

The low copy nuclear region, RPB2 as a novel DNA barcode region for species identification in the rattan genus Calamus (Arecaceae).

Taxonomic complexities, like environmental plasticity and homoplasy, make precise identification challenging in Calamus, the genus of spiny climbing palms of the subfamily Calamoideae (Arecaceae). In the present study, the species discriminatory power of twelve potential DNA barcode regions (rbcL, matK, psbA-trnH, rpoC, rpoB, psbK-psbI, atpF-atpH, psbZ-trnfM, ITS1, ITS2, PRK, and RPB2) were evaluated in 21 species of Calamus from the Western Ghats region of India, using distance, tree, and similarity based statistical methods. Except for the low copy nuclear region, RPB2, none of the tested plastid loci or nuclear loci ITS, either singly or in combinations, could discriminate all the species of Calamus due to low substitution rate of plastid regions and multiple copies of ITS respectively. The RPB2 locus showed highest species resolution with 96% accuracy in similarity based analysis, indicating its potential and efficiency as a barcode locus for the genus. The putative "Calamus gamblei complex" based on overlapping morphology was successfully resolved as six distinct, though closely related, species. The analysis also indicates that C. delessertianus is a morphological variant of C. dransfieldii. In spite of being a low copy nuclear gene region, RPB2 provided an efficient barcode to delineate Calamus species and has the potential to further extend its use as a prospective barcode to other Palm genera.

Species borderlines in Fusarium exemplified by F. circinatum/F. subglutinans.

Fusarium species are known as cross-kingdom pathogens, causing infections in both plants and animals. This ecological variation challenges the species concept of closely similar lineages in the genus. The present paper describes various types of genetic interaction between strains of two neighboring model species with different predilection, F. circinatum and F. subglutinans. Parameters include sequencing of the translation elongation factor 1α (TEF1) and the second largest subunit of RNA polymerase (RPB2), sexual crossing, and vegetative compatibility groups (VCGs). Successful interspecific crosses resulted in either recombination or in homothallic fruiting, the latter being limited to F. subglutinans MAT1 parents. Crossings were skewed, as Fusarium circinatum recombined more often than F. subglutinans. We hypothesize that genetic exchange in Fusarium species is finely regulated with an arsenal of options, which are applied when partners are phylogenetically closely related, leading to fluent species borderlines.

Comparative Evaluation of Etest, EUCAST, and CLSI Methods for Amphotericin B, Voriconazole, and Posaconazole against Clinically Relevant Fusarium Species.

We compared EUCAST and CLSI methods versus Etest for antifungal susceptibility testing of 20 clinically relevant Fusarium species against amphotericin B, posaconazole, and voriconazole. The median Etest amphotericin B and posaconazole MICs were 1 dilution higher than the median EUCAST and the CLSI MICs. The essential agreement (within ±1/±2 dilutions) was 60/90%, 80/95%, and 70/85% between the Etest and EUCAST methods and 80/95%, 75/95%, and 45/100% between the Etest and CLSI methods for amphotericin B, voriconazole, and posaconazole, respectively. The categorical agreement was >85%. Etest can be used for antifungal susceptibility testing of Fusarium species.

Multidrug-resistant Fusarium in keratitis: a clinico-mycological study of keratitis infections in Chennai, India.

In this study, we aimed to present the first molecular epidemiological data from Chennai, India, analyse keratitis cases that have been monitored in a university hospital during 2 years, identify the responsible Fusarium species and determine antifungal susceptibilities. A total of 10 cases of keratitis were included in the study. Fusarium isolates were identified using the second largest subunit of the RNA polymerase gene (RPB2) and the translation elongation factor 1 alpha (TEF1). Antifungal susceptibility was tested by the broth microdilution method according to the Clinical and Laboratory Standards Institute (CLSI) methodology. The aetiological agents belonged to Fusarium solani species complex (FSSC) (n = 9) and Fusarium sambucinum species complex (FSAMSC) (n = 1), and the identified species were Fusarium keratoplasticum (n = 7), Fusarium falciforme (n = 2) and Fusarium sporotrichioides (n = 1). All strains showed multidrug resistance to azoles and caspofungin but exhibited lower minimum inhibitory concentration (MIC) to natamycin and amphotericin B. Fusarium keratoplasticum and Fusarium falciforme belonging to the Fusarium solani species complex were the major aetiological agents of Fusarium keratitis in this study. Early presentation and 5% topical natamycin was associated with better patient outcome. Preventative measures and monitoring of local epidemiological data play an important role in clinical practice.

Plant diversity and plant identity influence Fusarium communities in soil.

Fusarium communities play important functional roles in soil and in plants as pathogens, endophytes, and saprotrophs. This study tests how rhizosphere Fusarium communities may vary with plant species, changes in the diversity of the surrounding plant community, and soil physiochemical characteristics. Fusarium communities in soil associated with the roots of two perennial prairie plant species maintained as monocultures or growing within polyculture plant communities were characterized using targeted metagenomics. Amplicon libraries targeting the RPB2 locus were generated from rhizosphere soil DNAs and sequenced using pyrosequencing. Sequences were clustered into operational taxonomic units (OTUs) and assigned a taxonomy using the Evolutionary Placement Algorithm. Fusarium community composition was differentiated between monoculture and polyculture plant communities, and by plant species in monoculture, but not in polyculture. Taxonomic classification of the Fusarium OTUs showed a predominance of F. tricinctum and F. oxysporum as well of the presence of a clade previously only found in the Southern Hemisphere. Total Fusarium richness was not affected by changes in plant community richness or correlated with soil physiochemical characteristics. However, OTU richness within two predominant phylogenetic lineages within the genus was positively or negatively correlated with soil physiochemical characteristics among samples within each lineage. This work shows that plant species, plant community richness, and soil physiochemical characteristics may all influence the composition and richness of Fusarium communities in soil.

Fusarium praegraminearum sp. nov., a novel nivalenol mycotoxin-producing pathogen from New Zealand can induce head blight on wheat.

We report on the molecular and morphological characterization of a novel type B trichothecene toxin-producing species (i.e. B clade) recovered from litter in a maize field near Wellington, New Zealand, which is described as Fusarium praegraminearum sp. nov. This species was initially identified as F. acuminatum based on morphological characters. However, it differs from this species by producing longer, slightly asymmetrically curved macroconidia in which the apical cell is not as pointed and by its much faster colony growth rate on agar. Molecular phylogenetic analyses of portions of 13 genes resolved F. praegraminearum as the most basal species within the B clade. Mycotoxin analyses demonstrated that it was able to produce 4-acetylnivalenol and 4,15-diacetylnivalenol trichothecenes, the nontrichothecene sesquiterpenes culmorin and hydroxy-culmorins, and the estrogen zearalenone in vitro. Results of a pathogenicity experiment revealed that F. praegraminearum induced moderate head blight on wheat.