ITS rDNA Sequences Research Articles

A new species, new host records and life cycle data for lepocreadiids (Digenea) of pomacentrid fishes from the Great Barrier Reef, Australia

A new species of lepocreadiid, Opechonoides opisthoporusn. sp., is described infecting 12 pomacentrid fish species from the Great Barrier Reef, Australia, with Abudefduf whitleyi Allen & Robertson as the type-host. This taxon differs from the only other known member of the genus, Opechonoides gure Yamaguti, 1940, in the sucker width ratio, cirrus-sac length, position of the testes, position of the pore of Laurer’s canal, and relative post-testicular distance. The new species exhibits stenoxenic host-specificity, infecting pomacentrids from seven genera: Abudefduf Forsskål, Amphiprion Bloch & Schneider, Neoglyphidodon Allen, Neopomacentrus Allen, Plectroglyphidodon Fowler & Ball, Pomacentrus Lacépède and Stegastes Jenyns. Phylogenetic analyses of 28S rDNA sequence data demonstrate that O. opisthoporusn. sp. forms a strongly supported clade with Prodistomum orientale (Layman, 1930) Bray & Gibson, 1990. The life cycle of this new species is partly elucidated on the basis of ITS2 rDNA sequence data; intermediate hosts are shown to be three species of Ctenophora. New host records and molecular data are reported for Lepocreadium oyabitcha Machida, 1984 and Lepotrema amblyglyphidodonis Bray, Cutmore & Cribb, 2018, and new molecular data are provided for Lepotrema acanthochromidis Bray, Cutmore & Cribb, 2018 and Lepotrema adlardi (Bray, Cribb & Barker, 1993) Bray & Cribb, 1996. Novel cox1 mtDNA sequence data showed intraspecific geographical structuring between Heron Island and Lizard Island for L. acanthochromidis but not for L. adlardi or O. opisthoporusn. sp.

Coprinopsis alnivora (Psathyrellaceae), a rare species from North America is discovered in Europe

A rare species, Coprinopsis alnivora, was previously known from the type collection in Washington State, USA. Afterward, 11 additional samples were collected from five new host trees at nine localities in Europe (Austria, Croatia, and Slovakia). The species indicated a preference for growth in cavities or wounds of living deciduous trees. Its mycelium and basidiomata were successfully cultivated under laboratory conditions. A detailed morphological description of the basidiomata supplemented with colour photographs and line drawings is provided. Delimitation characters from similar species are discussed. Molecular phylogenetic relationships within the genus Coprinopsis were inferred from ITS rDNA sequences and are presented by a phylogram. Molecular genetic analyses revealed that C. alnivora represents a genetically well-delimited species with six known haplotypes.

Diversity and Ginsenoside Biotransformation Potential of Cultivable Endophytic Fungi Associated With Panax bipinnatifidus var. bipinnatifidus in Qinling Mountains, China.

To obtain novel fungi with potent β-glucosidase for minor ginsenoside production, Panax bipinnatifidus var. bipinnatifidus, which is a traditional medicinal plant containing various ginsenosides, was first employed to isolate endophytic fungi in this study. A total of 93 representative morphotype strains were isolated and identified according to ITS rDNA sequence analyses, and they were grouped into three phyla (Ascomycota, Basidiomycota, and Mucoromycota), five classes (Dothideomycetes, Sordariomycetes, Eurotiomycetes, Agaricomycetes, and Mucoromycetes), and 24 genera. Plectosphaerella (RA, 19.35%) was the most abundant genus, followed by Paraphoma (RA, 11.83%) and Fusarium (RA, 9.70%). The species richness index (S, 34) and the Shannon–Wiener index (H’, 3.004) indicated that P. bipinnatifidus harbored abundant fungal resources. A total of 26 endophytic fungal ethyl acetate extracts exhibited inhibitory activities against at least one pathogenic bacterium or fungus. In total, 11 strains showed strong β-glucosidase activities and also presented with the ability of ginsenoside biotransformation with varied glycoside-hydrolyzing pathways. Excitingly, three genera, namely, Ilyonectria, Sarocladium, and Lecanicillium, and all 11 taxa were first found to have the ability to transform ginsenosides in our study. The results indicated that P. bipinnatifidus could be a new fungi resource with potential novel natural compounds with antimicrobial activity and potent β-glucosidase for varied minor ginsenoside production.

Allophlebia, a new genus to accomodate Phlebia ludoviciana (Agaricomycetes, Polyporales)

Allophlebia is proposed as a new genus in Meruliaceae based on morphological characters and molecular data. The genus, so far monotypic, is typified by Peniophora ludoviciana and the new combination A. ludoviciana is proposed. The type species is characterized by a resupinate basidioma, a monomitic hyphal system with clamp connections, two types of cystidia (leptocystidia and metuloids), clavate basidia, and hyaline, thin-walled and ellipsoid basidiospores. A phylogeny for Allophlebia and related taxa was inferred from ITS and nLSU rDNA sequences and new information on the geographic distribution of A. ludoviciana is provided.

Molecular Identification and Virulence of Four Strains of Entomopathogenic Fungi Against the Whitefly, Bemisia tabaci (Hemiptera: Aleyrodidae).

The whitefly, Bemisia tabaci (Gennadius), is a key pest of many economically important crops grown in the field and in greenhouses throughout the world. Because entomopathogenic fungi (EPF) are potential biological control agents for B. tabaci, however, minimal research has been conducted on using fungal strains to control B. tabaci. In this study, four EPF strains were isolated and identified as Lecanicillium attenuatum (Zare & Gams) JL-003, Beauveria bassiana Balsamo (Vuillemin) JL-005, Lecanicillium longisporum (Petch) JL-006, and Akanthomyces lecanii (Zimmerman) JL-007, based on rDNA-ITS sequence analysis. In comparing the virulence of the four fungi against the different life stages (i.e., eggs, 1st-, 2nd-, 3rd-, 4th-instar nymphs, and adults) of B. tabaci the mortality of B. tabaci decreased and LT50 values increased as the conidia concentration decreased in a series of conidia concentrations (1×105, 106, 107, and 108 conidia/mL). The fungal strains L. attenuatum JL-003 (LC50: 1.31×106) and B. bassiana JL-005 (LC50: 0.92×106) were found to be more effective than L. longisporum JL-006 (LC50: 4.97×107) and A. lecanii JL-007 (LC50: 6.46×106). Fourth-instar nymphs, eggs, and adult stages of B. tabaci were less susceptible to all fungal strains compared to 1st-, 2nd-, and 3rd-instar nymphs. The virulence of L. attenuatum, which was tested for the first time on B. tabaci, was found to be more toxic to early-stage nymphs. Our data will be useful in biological control programs that are considering using EPF against B. tabaci.

Morphological, histopathological and secondary structure analysis of second internal transcribed spacer (ITS-2) region of Gigantocotyle explanatum (Trematoda: Paramphistomidae) in buffaloes of Pakistan

Liver infections due to amphistomes in buffaloes cause significant economic losses in livestock sector. The present study investigated the classical morphological features of adult amphistomes combined with histopathology and molecular identity in slaughtered buffaloes from Khyber Pakhtunkhwa, Pakistan. Adult amphistome were collected and morphologically characterized as Gigantocotyle explanatum. Morphometric measurement (n=50) were obtained with 12.05±1.68 mm in length and 5.77±0.81 mm in width, and the values of sagittal section was 6.35±1.03 x 2.98±0.50 mm in size. Severe bile duct pathology was observed with fibrosis of the duct wall with an irregular epithelial border, hyperplasia and inflammatory response with numerous neutrophils and eosinophils. The molecular identity of G. explanatum within Paramphistomidae was confirmed by ITS-2 rDNA sequences phylogenetic analysis based on maximum likelihood method. The genetic data based on ITS-2 secondary structure of G. explanatum consisted of four helix, Helix I, II and IV were conserved as compared with other closely related reference taxa of family Paramphistomidae and Gastrothylacidae. Helix III expressed some variations. The study concluded that rDNA ITS-2 and secondary structure information provides a guide for other researchers to determine the molecular taxonomic position of Paramphistomidae trematodes, data will support future clinical studies and control measures to reduce the amphistomiasis in buffaloes.

Identification, biological characteristics, and fungicide screening of pathogen of black spot in Aconitum carmichaelii

In Hezhang county, Guizhou province, black spot tends to occur to Aconitum carmichaelii in the hot rainy summer, with the incidence up to 50%-70%, seriously impacting the yield and quality of the medicinal material. Thus, this study aims to clarify the pathogen and the occurrence characteristics. To be specific, the pathogen was isolated and identified according to Koch's postulates and the pathogenicity and biological characteristics were determined. In addition, the sensitivity of the pathogen to four microbial fungicides, four botanical fungicides, and five chemical fungicides was determined with the mycelium growth rate method for the purpose of screening out optimal fungicides. The pathogen was identified as Alternaria alternate, as evidenced by the similar colony morphology and microscopic characteristics and 99.55%-100% similarity in sequences of rDNA-ITS, LSU, 18S, and TEF of the two. The optimum growth conditions for A. alternata were 28 ℃, pH 8, and continuous darkness. Bacillus subtilis had strong inhibitory effect on the pathogen, and the inhibition rate was more than 90% when the concentration was 1 mg·L~(-1). In addition, difenoconazole and quinoline copper can also control the pathogen, with median effective concentration(EC_(50)) of 2.92 and 9.02 mg·L~(-1), respectively. This study lays a theoretical basis for the field control of black spot in A. carmichaelii.

Endophytic fungi from Dongxiang wild rice (Oryza rufipogon Griff.) show diverse catalytic potential for converting glycyrrhizin.

Endophytic fungi inhabiting niche environments are novel biocatalyst resources that need to be exploited urgently. In this study, 63 endophytic fungi isolated from Dongxiang wild rice (Oryza rufipogon Griff.) were tested to assess their potentials to transform glycyrrhizin (GL) into glycyrrhetinic acid monoglucuronide (GAMG) or glycyrrhetinic acid (GA), of which 12 strains were shown to have β-d-glucuronidase activity. Based on morphological characteristics and rDNA ITS sequence analysis, the strains S59, L138, L55 and R57 with high GL molar conversion rates (55%, 45%, 65% and 89%) were further identified as Microsphaeropsis arundinis S59, Penicillium rubens L138, Aspergillus flavus L55 and Eupenicillium javanicum R57, respectively. These four strains with four different types of GL conversion processes were identified, i.e., (1) GL → GAMG in M. arundinis S59, (2) GL → GAMG and GA in A. flavus L55, (3) GL → GA in P. rubens L138, and (4) GL → GAMG → GA in E. javanicum R57, in which the bioconversion type (4) is reported for the first time. The study not only provided abundant and diverse β-d-glucuronidase resources that can be used for GL bioconversion, especially for GAMG biosynthesis from endophytic fungi, but also expanded our knowledge of potential roles of endophytes as new biocatalysts in biotransformation.

Revealing hidden diversity in the Cryptomonas erosa clade (Cryptophyceae), with the description of two new species from acidic habitats

ABSTRACT In this study, the cryptomonads of the northern territories of Russia were explored. Four strains were isolated from the different acidic habitats in the Middle Urals and the North of the European part of Russia. Cells of these strains are flattened in the dorsoventral plane, have an asymmetrical outline and are similar in size and shape to Cryptomonas erosa. Phylogenetic relationships inferred using maximum likelihood and Bayesian interference from datasets of nuclear-encoded SSU, LSU, ITS2 rDNA and psbA cpDNA show that the strains form one clade, which was divided into three subclades with high statistical support. One subclade was attributed to Cryptomonas erosa. Two other subclades represent species new to science. Comparison of ITS2 rRNA secondary structures with the compensatory base changes (CBC) approach confirmed the separation between C. erosa and the two new species, C. macilenta sp. nov. and C. parmana sp. nov. Comparison of nuclear V9 SSU and ITS1 rDNA sequences of two new species with metabarcoding data from 218 lakes in Europe showed that C. macilenta and C. parmana have not previously been recorded in Europe.

Two toxigenic Ostreopsis species, O. cf. ovata and O. siamensis (Dinophyceae), from the South China Sea, tropical Western Pacific

In the dinophyte genus Ostreopsis, seven out of 11 described species are known to produce various toxic compounds that were characterized in the palytoxins family. Species in the genus shared identical thecal plate patterns but differed in size, shape, and thecal plate ornamentation. Two species, O. cf. ovata and O. siamensis, have been reported from the Western Pacific, but information on toxin production is scarce. Here, we established nine strains of Ostreopsis from six localities in the South China Sea (SCS), covering the Gulf of Thailand, northern SCS (Hainan Island, Beibu Bay), and southern SCS (Peninsular Malaysia). Their morphology was examined by light and electron microscopy and the molecular phylogeny was inferred based on the LSU rDNA (D1-D3) and ITS rDNA sequences using maximum likelihood and Bayesian inference. Both O. cf. ovata and O. siamensis, albeit morphologically closely related, can be distinguished by a feature of the thecal pores with pronounced ridges in the latter. Molecular data further supported their species identity. Toxin production in the strains was examined by LC-MS/MS. O. cf. ovata strain T5PRBost02 was observed to produce Ovatoxin-k and Ovatoxin-j2 only; while Ostreocin-B and Ostreocin-D was produced by O. siamensis strain T10PRBost04. This is the first report confirming the production of palytoxins analogs in Ostreopsis species from the region.

Morphological variation and phylogeny of Karenia selliformis (Gymnodiniales, Dinophyceae) in an intensive cold-water algal bloom in eastern Hokkaido, Japan

Harmful algal blooms responsible for mass mortalities of marine organisms have been rare in Hokkaido, northern Japan, although fish-killing blooms have been frequently reported from western Japanese coasts. In September–November 2021, a huge and prolonged cold-water bloom occurred along the Pacific coast of eastern Hokkaido, and was associated with intensive mortalities of sea urchin, fish, octopus, shellfish, etc. In this study, morphology and phylogeny of the dominant and co-occurring unarmored dinoflagellates of the Kareniaceae in the bloom were examined by using light microscopy, scanning electron microscopy and molecular phylogeny inferred from ITS and LSU rDNA (D1–D3) sequences. Morphological observation and molecular phylogeny showed that the dominant species was Karenia selliformis, with co-occurrences of other kareniacean dinoflagellates, Kr. longicanalis, Kr. mikimotoi, Karlodinium sp., Takayama cf. acrotrocha, Takayama tuberculata and Takayama sp. The typical cell forms of Kr. selliformis in the bloom were discoid, dorsoventrally flattened, and 35.3–43.6 (39.4 ± 2.1) µm in length, which was larger than the cell sizes in previous reports. Transparent cells of Kr. selliformis, lacking chloroplasts or having a few shrunken chloroplasts and oil droplets, were also found. Cells of Kr. selliformis showed morphological variation, but the species could be distinguished from other co-occurring Karenia species by the nucleus positioned in the hypocone and chloroplasts numerous (46–105) in number and small (2.9–4.6 µm) in diameter. Cell density of Kr. selliformis exceeding 100 cells mL−1 was recorded in the temperature range of 9.8–17.6 °C. The rDNA sequences determined from Kr. selliformis in the blooms of Hokkaido, Japan in 2021 were identical to those from the bloom in Kamchatka, Russia in 2020.

Molecular Characteristics of Regional Chromoblastomycosis in Guangdong, China: Epidemiological, Clinical, Antifungal Susceptibility, and Serum Cytokine Profiles of 45 Cases.

Chromoblastomycosis (CBM) is a chronic disease caused by several species of dematiaceous fungi. In this study, a regional collection of 45 CBM cases was conducted in Guangdong, China, a hyper-endemic area of CBM. Epidemiology findings indicated that the mean age of cases was 61.38 ± 11.20 years, long duration ranged from 3 months to 30 years, and the gender ratio of male to female was 4.6:1. Thirteen cases (29%) declared underlying diseases. Verrucous form was the most common clinical manifestation (n = 19, 42%). Forty-five corresponding clinical strains were isolated, and 28 of them (62%) were identified as F. monophora; the remaining 17 (38%) were identified as F. nubica through ITS rDNA sequence analysis. Antifungal susceptibility tests in vitro showed low MICs in azoles (PCZ 0.015–0.25 μg/ml, VCZ 0.015–0.5 μg/ml, and ITZ 0.03–0.5 μg/ml) and TRB (0.015–1 μg/ml). Itraconazole combined with terbinafine was the main therapeutic strategy used for 31 of 45 cases, and 68% (n = 21) of them improved or were cured. Cytokine profile assays indicated upregulation of IL-4, IL-7, IL-15, IL-11, and IL-17, while downregulation of IL-1RA, MIP-1β, IL-8, and IL-16 compared to healthy donors (p < 0.05). The abnormal cytokine profiles indicated impaired immune response to eliminate fungus in CBM cases, which probably contributed to the chronic duration of this disease. In conclusion, we investigated the molecular epidemiological, clinical, and laboratory characteristics of CBM in Guangdong, China, which may assist further clinical therapy, as well as fundamental pathogenesis studies of CBM.

First Report of Leaf Spot Disease on Cinnamomum camphora (Camphor Tree) Caused by Epicoccum poaceicola in China.

As an important industrial, pharmaceutical and evergreen shade tree (Singh and Jawaid 2012), the camphor tree (Cinnamomum camphora) has been coppiced in Jiangxi Province, China. From 2017 to 2020, we noticed many camphor trees with leaf spots, with an incidence estimated at 50 to 75%, which could severely inhibit leaf growth and reduce their biomass. A dark-green circle with a watery spot appeared on the infected leaves at the initial stage, and necrosis with forming shot-spots surrounded by yellow halos occurred (Figure 1 A). Five leaves with typical symptoms were sampled and washed with tap water for ca. 15 min. Isolation and morphological analysis were performed following the method of Bao et al. (2019). Among 61 fungal isolates, 49 showed the same culture characters. Colonies on PDA were villose and regular, the reverse was scarlet at the edge of the colony, which was ca. 8.75 cm after 7 days of inoculation (Figure 1 I). Chlamydospores were aseptate, dark brown, smooth, in chains or solitary, ellipsoidal to ovoid, 4.8-9.6 × 4.8-11.1 μm (Figure 1 J). The pycnidia were produced on PDA and varied from 47.4 to 85.8 µm (mean 60.2 µm) × 38.6 to 66.8 μm (mean 49.7 μm) (n = 17) (Figure 1 K). Conidia were hyaline, unicellular, elliptical to ovoid, 4.3-6.4 µm (mean 5.1 µm) × 2.3-3.3 µm (mean 2.8 µm) (n = 52) (Figure 1 L). Pathogenicity tests of isolate XW-9 was carried out in the field. Ten leaves were wounded with a sterilized insect needle and inoculated with mycelium plugs (7-mm diameter). Non-colonized PDA plugs served as the negative controlIn addition, conidial suspensions (105 conidia/mL) of isolate XW-9 were sprayed on surface-sterilized leaves with a further ten leaves being sprayed with sterile water as the control. Symptoms described in this study appeared in 100% of the mycelium-inoculated leaves and more than 80% of the conidium-inoculated leaves after 7 days post-inoculation (Figure 1 B-E). No symptoms were seen in the controls (Figure 1 C). Three days after inoculation, brown spots resembling those observed in the field developed on the inoculated leaves, and some lesions turned into shot holes on the infected leaves (Figure 1 G & H). However, no symptoms were observed on the controls (Figure 1 F). The fungus was re-isolated from the margins of the leaf spots and labelled P-XW-9A. The gene regions for ITS, LSU, tub2, RPB2 and ACT of isolates XW-9 and P-XW-9A were amplified and sequenced. The sequences of rDNA-ITS, LSU, tub2, RPB2 and ACT of XW-9 were GenBank MW142397, MW130844, MW165322, MW446945 and MW165324, respectively and those of P-XW-9A were GenBank MW142398, MW130845, MW165323, MW446946 and MW165325, respectively (Lumbsch, et al. 2000; Aveskamp, et al. 2009; Hou et al. 2020). Phylogenetic analysis using concatenated sequences of ITS, LSU, RPB2, and tub2 showed that isolates XW-9 and P-XW-9A formed a single clade with the reference strain of E. poaceicola CBS 987.95 (Figure 2). Thus, XW-9 was identified as E. poaceicola based on its morphological and molecular characteristics. Significantly, the recovered isolate P-XW-9A also aligned with E. poaceicola fulfilling the criteria for Koch's Postulates. E. poaceicola was only reported as a fungal pathogen of Phyllostachys viridis in China (Liu et al. 2020). To our knowledge, this is the first report of leaf spot disease on camphor trees caused by E. poaceicola in China and our findings will be useful for its management.

Three New Species of Absidia (Mucoromycota) from China Based on Phylogeny, Morphology and Physiology

Species of Absidia are distributed widely in the environment, while their diversity is insufficiently studied. Three new species, A. frigida, A. gemella and A. longissima, are proposed herein from Xinjiang and Yunnan in China based on phylogenetic, morphological and physiological evidence. According to maximum likelihood (ML), maximum parsimony (MP) and Bayesian inference (BI) analyses, the phylogenetical results suggest that A. frigida, A. gemella and A. longissima are closely related to A. psychrophilia, A. turgida and A. zonata and A. koreana, respectively, based on ITS and LSU rDNA sequences. Absidia frigida is characterized by a lower growth temperature, which does not grow above 24 °C. It differs from A. psychrophilia by sporangiophores, sporangia, columellae, collars and projections. Absidia gemella is distinguished from A. turgida by hypha, sporangiospores, sporangia, projections and sporangiophores. Absidia longissima is discriminated from A. zonata and A. koreana by sporangiophores, columellae and collars. The three new species are described and illustrated in this article.

New report of Biscogniauxia rosacearum as a pathogen on almond trees in Iran

Biscogniauxia species are known as fungal trunk pathogens on various tree species in the world. During a survey of trunk diseases of fruit trees conducted in Iran, a branch dieback was observed on almond trees in Kerman province (in the Southeast of Iran). Evaluation of symptomatic branches revealed wood discoloration in cross-sections and the presence of fruiting bodies of an ascomycete fungus on their bark. A total of 31 fungal isolates were obtained, 13 isolates recovered from necrotic wood tissues and 18 isolates from fruiting bodies. These isolates were subjected to morphological analysis as well as sequencing analysis of the partial ITS-rDNA and beta tubulin gene sequences. These fungal isolates were identified as Biscogniauxia rosacearum. Results of the pathogenicity tests showed that this species is pathogenic on almond shoots. Based on our knowledge, this is the first report of this species on almond trees in Iran and in the world.

Development of an RNA anti-parasite based on CRISPR-Cas9 against Chilodonella piscicola

Chilodonella piscicola is a gill and body surface parasite of freshwater fish, affecting the growth of juvenile and adult freshwater fishes. To develop an effective method to manage C.piscicola in aquaculture, we used CRISPR/Cas9 technology to design and validate small guide RNA targeting DNA barcoding ITS-1 and 18S rDNA sequences of C.piscicola to develop a species-specific pesticide. In this study, effective sgRNA targets were found in ITS-1 and 18S rDNA sequences of C.piscicola, respectively. In the soaking experiment, we found that the mixing of Cas9 mRNA and sgRNAs could significantly reduce the survival rate of C.piscicola. The Real-time quantitative PCR results showed that both Cas9 works with the single sgRNA or the mixture of the two sgRNA could destroy the parasite DNA at a specific region. DNA sequencing also verified the effectiveness of single sgRNA or two sgRNAs mixed with cas9 induced mutation on the target fragment of the parasite DNA in the experimental group. The results of this study once again verified the effectiveness of genetic barcoding in the application of anti-parasitic and provided a new idea for anti-parasitic methods.

First report of Fusarium oxysporum causing damping-off of tree bean (Parkia timoriana) seedlings in Northeastern India

Tree bean (Parkia timoriana), is an important multipurpose leguminous tropical deciduous tree species widely grown in the Northeastern states of India. The occurrence of the high incidence of damping-off (86.19%) disease was observed in tree bean nursery grown in Umiam, Meghalaya, a state in Northeastern India. Therefore, a systematic study was undertaken to record the symptoms and characterize the morphological features, the taxonomy of pathogen causing damping-off disease of P. timoriana and confirm it by pathogenicity assay. The infected seeds exhibited typical symptoms of covering with whitish mycelium and cotyledons were completely rotten in later stages of infection and emitted foul odour. Infected seedlings exhibited the symptoms of root rot followed by wilting, dried stem, and complete mortality of the seedlings. The pathogen isolated from the rotten seeds and wilted seedlings consistently showed the association of Fusarium oxysporum and its identity was confirmed based on morphological characteristics and sequences of ITS-rDNA and TEF genomic regions. Five representative isolates (TBSR 1 to 5) exhibited high similarity (>98%) to the known authentic gene sequence of F. oxysporum. The pathogenicity test produced similar to natural symptoms of wilt at 30 days after inoculation, confirming the etiological association of Fusarium oxysporum. To the best of our knowledge, this is the first to report on F. oxysporum causing damping-off disease of P. timoriana in India and worldwide.

Three new species of Helicometroides Yamaguti, 1934 from Japan and Australia, with new molecular evidence of a widespread species.

We report specimens of monorchiids infecting Haemulidae from the waters off Japan and Australia; these specimens represent five species of Helicometroides Yamaguti, 1934, three of which are unambiguously new. Helicometroides murakamii n. sp. infects Diagramma pictum pictum from off Minabe, Japan; Helicometroides gabrieli n. sp. infects Plectorhinchus chrysotaenia from off Lizard Island, Australia; and Helicometroides wardae n. sp. infects Plectorhinchus flavomaculatus and Plectorhinchus multivittatus from off Heron Island, Australia. Helicometroides murakamii n. sp. and H. gabrieli n. sp. conform to the most recent diagnosis of Helicometroides in lacking a terminal organ, but H. wardae n. sp. possesses a terminal organ with distinct, robust spines; despite this morphological distinction, the three form a strongly-supported clade in phylogenetic analyses. We also report specimens morphologically consistent with Helicometroides longicollis Yamaguti, 1934, from D. pictum pictum from off Minabe, Japan, and Diagramma pictum labiosum on the Great Barrier Reef, Australia. Genetic analyses of ITS2 rDNA, 28S rDNA and cox1 mtDNA sequence data for the Japanese specimens reveal the presence of two distinct genotypes. Specimens of the two genotypes were discovered in mixed infections and are morphologically indistinguishable; neither genotype can be associated definitively with H. longicollis as originally described. We thus identify them as H. longicollis lineage 1 and 2, pending study of further fresh material. Genetic analyses of specimens from the Great Barrier Reef are consistent with the presence of only H. longicollis lineage 1. This species thus has a range that incorporates at least Australia and Japan, localities separated by over 7000 km.

Supplementary files

Fig. S1. Maximum Likelihood phylogenetic tree of the genus Chloridium based on analysis of a concatenated data set (ITS, LSU) treated with Gblocks. Species names given in bold and marked in blue colour are Chloridium species described by Wu &amp; Diao (2022). Branch support of nodes ≥ 75 % ML is indicated above or below branches.&lt;br /&gt;Fig. S2. Maximum Likelihood phylogenetic tree of the family Chaetosphaeriaceae based on analysis of a concatenated data set (ITS, LSU) treated with Gblocks. Species names given in bold and marked in blue colour are Chloridium species described by Wu &amp; Diao (2022). Branch support of nodes ≥ 75 % ML is indicated above or below branches.&lt;br /&gt;Fig. S3. Maximum Likelihood phylogenetic tree of genera Adautomilanezia, Chloridium and Sporoschisma based on analysis of a concatenated data set (ITS, LSU) treated with Gblocks. Species names given in bold and marked in blue colour are Chloridium species described by Wu &amp; Diao (2022). Branch support of nodes ≥ 75 % ML is indicated above or below branches.&lt;br /&gt;Fig. S4. Geographical distribution of Chloridium spp. based on data from the GlobalFungi database.&lt;br /&gt;Table S1. Estimates of evolutionary divergence between ITS rDNA sequences. The number of base differences per site from between sequences are shown. Analyses were conducted using the Kimura 2-parameter model [1]. This analysis involved 116 nucleotide sequences. All ambiguous positions were removed for each sequence pair (pairwise deletion option). There were a total of 562 positions in the final data set. Evolutionary analyses were conducted in MEGA11 [2].&lt;br /&gt;Table S2. Biogeography, substrate and habitat affinity of Chloridium inferred from the GlobalFungi database. The overview of all samples in the database (Sheet 1), summary across all Chloridium species (Table S3), and detailed metadata and primary data for all Chloridium species are presented.&lt;br /&gt;Table S3. Summary of biogeography, substrate and habitat affinity of Chloridium inferred from the GlobalFungi database.&lt;br /&gt;Table S4. Growth on different temperatures (5–37 °C) after 14 d. The average of three measurements is presented. The values of average Mean Annual Temperature (MAT), average Mean Annual Precipitation (MAP) for each species are presented. MAT and MAP were obtained from GlobalFungi database and the primary data are presented in Table S2.

Identification and pathogenicity ofPratylenchus scribnerion tomato in Sichuan Province of People's Republic of China

AbstractRoot-lesion nematodes (Pratylenchusspp.) are a group of economically important pathogens that have caused serious economic losses in many crops. In 2019, root-lesion nematodes were recovered from tomato (Solanum lycopersicum) root samples collected from Sichuan Province, People's Republic of China (PRC). Extracted nematodes were disinfected, and one individual female was cultured on a carrot disc for propagation at 25 °C by parthenogenesis and designated the SC isolate. Afterwards, the isolate was identified on the basis of morphometric and molecular markers. Both morphometric characters and molecular analysis of the internal transcribed spacer region gene (ITS) of ribosomal DNA, the D2-D3 expansion region of the 28S rDNA gene and the mitochondrial cytochrome oxidase I (mtDNA-COI) gene revealed that the species of root-lesion nematode wasPratylenchus scribneri. The Bayesian tree inferred from the ITS rDNA, 28S rDNA and mtDNA-COI gene sequences also showed that this isolate formed a highly supported clade with otherP. scribneriisolates. The pathogenicity of the root-lesion nematode SC isolate on tomato was assessed, showing that tomato was a suitable host forP. scribneri. To the best of our knowledge, this is the first report ofP. scribnerion tomato in Sichuan Province, PRC. These are also the first molecular data obtained fromP. scribnerion tomato in the PRC, and the pathogenicity ofP. scribnerito tomato was studied for the first time. This study provides scientific data for the detection, identification and control of tomato root-lesion nematode disease.