Metarhabditis amsactae: A potential biopesticide isolated from Punjab (India) with potent insecticidal activity and immunomodulatory effects against Galleria mellonella (Lepidoptera: Pyralidae).

A survey was undertaken to isolate entomopathogenic nematodes from Amritsar district of Punjab, India. Out of 20 soil samples collected, two were found positive for the presence of nematodes. 18S and ITS rDNA gene sequencing revealed their identity as Metarhabditis amsactae. To assess its biocontrol potential, Galleria mellonella larvae were treated with concentrations of 20, 40, 80 and 160 IJs/L (infective juveniles/larva) and mortality was recorded from 24 h up to 96 h of nematode exposure. Distilled water without nematodes was used as an untreated control. M. amsactae showed potent larvicidal activity against G. mellonella that was found to be concentration and time dependent. Nematode infection caused 93.33 % larval mortality at 80 IJs/L after 72 h of treatment. 100 % mortality was observed after 96 h. No mortality was observed in control. To evaluate the immunomodulatory effects of M. amsactae, G. mellonella larvae were infected with 100 IJs/L and activities of antioxidant and detoxifying enzymes viz., superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APOX), phenol oxidase (PO), glutathione-S-transferase (GST) and acetylcholine esterase (AChE) were appraised after 12, 24, 36 and 48 h of nematode exposure. Malondialdehyde content was also determined. The results obtained demonstrated a significant elevation in all the enzyme activities at all time intervals in treated larvae when compared with untreated control. MDA levels were also enhanced in response to nematode infection. Thus, the present study revealed high insecticidal potential and immunomodulatory effects of M. amsactae on G. mellonella that should be further explored on other insect pests as well.

Genetic diversity and population structure of Fusarium udum in India and its correlation with pigeonpea wilt incidence.

In a study conducted in India, 50 Fusarium isolates were collected from pigeonpea growing regions and extensively examined for their cultural and morphological characteristics. These isolates exhibited significant variations in traits including growth rate, mycelial growth patterns, color, zonation, pigmentation, spore size, and septation. Subsequently, 30 isolates were chosen for pathogenicity testing on eight pigeonpea genotypes. Results showed distinct reactions, with four genotypes displaying differential responses (ICP8858, ICP8859, ICP8862, and BDN-2), while ICP9174 and ICP8863 consistently exhibited resistance and ICP2376 and BAHAR remained susceptible to wilt disease. To study the interaction between Fusarium isolates and pigeonpea host differentials (HDs), an additive main effects and multiplicative interaction analysis was conducted. The majority of disease incidence variation (75.54%) was attributed to HD effects, while Fusarium isolate effects accounted for only 1.99%. The interaction between Isolates and HDs (I × HD) contributed 21.95% to the total variation, being smaller than HD but larger than I. Based on HD reactions, isolates were classified into nine variants, showing varying distributions across pigeonpea growing states, with variants 2 and 3 being prevalent in several regions. This diversity underscores the need for location-specific wilt-resistant pigeonpea cultivars. Furthermore, genetic analysis of 23 representative isolates, through internal transcribed spacer region of ribosomal DNA and translation elongation factor 1-α gene sequencing, revealed three major clusters: Fusarium udum, Fusarium solani, and Fusarium equiseti. These findings hold potential for developing location-specific wilt-resistant pigeonpea cultivars and enhancing disease management strategies.

Rapid and Direct Detection of the Stubby Root Nematode, Paratrichodorus allius, from Soil DNA Extracts Using Recombinase Polymerase Amplification Assay.

The stubby root nematode, Paratrichodorus allius, is one of the most important plant-parasitic nematodes. Besides root feeding, P. allius also transmits the Tobacco rattle virus in potatoes, which causes corky ringspot disease. Rapid detection of P. allius is key for efficient management. This study was conducted to develop a real-time recombinase polymerase amplification (RPA) assay that is capable of detecting P. allius directly in DNA extracts from soil using a simple portable device in real time. A fluorophore-attached probe was designed to target the internal transcribed spacer (ITS)-rDNA of P. allius and was used along with primers designed previously. The real-time RPA assay had the ability to detect P. allius DNA extracted directly from infested soil with a sensitivity of one-sixteenth portion of a single nematode. This RPA assay was specific, as it did not produce positive signals from non-target nematodes tested. The real-time RPA was found to be rapid as it could even detect P. allius in as little as 7 min. Testing with 15 field soil samples validated the RPA assay developed in this study. This is the first report of P. allius detection directly from soil DNA using real-time RPA and is the fastest method for P. allius detection in soil to date.

A multi-aspect analysis of two analogous aspergillus spp. belonging to section Flavi: aspergillus flavus and aspergillus oryzae.

Microfungal isolates were routinely identified depending on both macro and micro morphological characteristics, sometimes, some fungal isolates appeared to be similar and such cases caused severe confusion for mycologists during the preliminary identification. During our previous studies dealing with isolation of fungi for some biotechnological applications; two mystifying species Aspergillus flavus and Aspergillus oryzae showed similar cultural and macroscopic features. Therefore, the current study aimed to easily distinguish between these two species depending on simple approaches which are routinely followed by a large segment of researchers. Investigation of the macroscopic features was performed to check the fungal growth on four different media (PDA, MEA, YES, and CYA) followed by microscopic examination using an ordinary light microscope, and scanning electron microscope SEM. Also, screening of secondary metabolites for both strains was preliminarily identified to find out the difference between their metabolic profiles. Finally, ITS rDNA was involved to clarify the molecular differences along their partial sequence. Conclusively, the BLAST strategy confirmed the similarity of ITS rDNA segments of both fungal strains that supported our hypothesis. The color of the fungal growth is a very critical factor whereas it is extensively influenced by the type of cultivation media. Accordingly, the YES medium was an inspiring tool assisting in prompt differentiation during the culture investigation step whereas A. oryzae and A. flavus appeared significant mustard yellow and olive green respectively. During the microscopic examination, the CYA medium also had a robust effect on the formation of the conidial chain whereas the knit long chain was observed in A. oryzae while the conidia appeared scattered and not in a chain in the case of A. flavus. Likewise, both two strains possessed different metabolic profiles where A. oryzae is not an Afla toxin producer, unlike A. flavus.

Formation of soil organic carbon pool is regulated by the structure of dissolved organic matter and microbial carbon pump efficacy: A decadal study comparing different carbon management strategies.

To achieve long-term increases in soil organic carbon (SOC) storage, it is essential to understand the effects of carbon management strategies on SOC formation pathways, particularly through changes in microbial necromass carbon (MNC) and dissolved organic carbon (DOC). Using a 14-year field study, we demonstrate that both biochar and maize straw lifted the SOC ceiling, but through different pathways. Biochar, while raising SOC and DOC content, decreased substrate degradability by increasing carbon aromaticity. This resulted in suppressed microbial abundance and enzyme activity, which lowered soil respiration, weakened in vivo turnover and ex vivo modification for MNC production (i.e., low microbial carbon pump "efficacy"), and led to lower efficiency in decomposing MNC, ultimately resulting in the net accumulation of SOC and MNC. In contrast, straw incorporation increased the content and decreased the aromaticity of SOC and DOC. The enhanced SOC degradability and soil nutrient content, such as total nitrogen and total phosphorous, stimulated the microbial population and activity, thereby boosting soil respiration and enhancing microbial carbon pump "efficacy" for MNC production. The total C added to biochar and straw plots were estimated as 27.3-54.5 and 41.4 Mg C ha-1 , respectively. Our results demonstrated that biochar was more efficient in lifting the SOC stock via exogenous stable carbon input and MNC stabilization, although the latter showed low "efficacy". Meanwhile, straw incorporation significantly promoted net MNC accumulation but also stimulated SOC mineralization, resulting in a smaller increase in SOC content (by 50%) compared to biochar (by 53%-102%). The results address the decadal-scale effects of biochar and straw application on the formation of the stable organic carbon pool in soil, and understanding the causal mechanisms can allow field practices to maximize SOC content.

First report of Perkinsus olseni infections in blood cockles Tegillarca granosa on the south coast of Korea.

The protozoan parasite Perkinsus olseni has become a focus of attention since it has been responsible for mass mortalities and economic losses in a wide range of bivalve hosts globally. The P. olseni host range along the south coast of Korea may extend beyond what was previously understood, and blood cockles in the Family Arcidae are also suggested to be potential hosts of P. olseni. In the present study, we applied histology and molecular techniques to identify Perkinsus sp. infections in the blood cockles Tegillarca granosa, which have been commercially exploited on the south coast of Korea for several decades. Histology and molecular techniques, including genus-specific immunofluorescence assay, species-specific fluorescence in situ hybridization, and phylogeny based on the ribosomal DNA internal transcribed spacer region revealed that T. granosa is infected by P. olseni, although the prevalence was low (0.5%). Histology revealed massive hemocyte infiltrations in the mantle, gill, and digestive gland connective tissues, indicating that the infection exerts negative impacts on the host cockles.

First report of Achlya bisexualis infection in captive-reared Endangered golden mahseer Tor putitora.

Achlya bisexualis is a notorious oomycete pathogen with the potential to cause emerging disease in fish farms. In this study, we report the first isolation of A. bisexualis from captive-reared golden mahseer Tor putitora, an Endangered fish species. The infected fish showed a cotton-like growth of mycelia at the site of infection. The mycelium when cultured on potato dextrose agar produced radially growing white hyphae. The hyphae were non-septate, and some of them carried matured zoosporangium with dense granular cytoplasmic contents. Spherical gemmae with stout stalks were also observed. All the isolates had 100% identity in internal transcribed spacer (ITS)-rDNA sequence and showed highest similarity to that of A. bisexualis. In molecular phylogeny, all the isolates formed a monophyletic group with A. bisexualis which was supported by a bootstrap value of 99%. Based on the molecular and morphological findings, all the isolates were confirmed as A. bisexualis. Further, the anti-oomycete effect of boric acid, a known antifungal agent, against the isolate was evaluated. The minimum inhibitory concentration and minimum fungicidal concentration were found to be 1.25 and >2.5 g l-1, respectively. Isolation of A. bisexualis from a new fish species indicates its possible occurrence in other unreported hosts. Considering its wide infectivity and the potential to cause disease in farmed fishes, its probable prevalence in a new environment and host needs to be closely monitored to prevent the spread of infection, if any, by adopting suitable control measures.

First Morphological and Molecular Characterization of Paratylenchus vandenbrandei (Rhabditida: Tylenchulidae) in Iran.

Paratylenchus vandenbrandei, has been recovered from the rhizospheric soil of Euphrates poplar (Populus euphratica) in the Karkheh protected area of Khuzestan province, southwestern Iran. The species was identified as P. vandenbrandei by the presence of three lines in the lateral fields; conoid rounded lip region; presence of submedian lobes, a stylet 24.0-28.8 µm long; an excretory pore at the level of the anterior part of the pharyngeal bulb; a round-to-oval spermatheca; presence of vulval flaps; and a conoid tail, with a terminus that is rounded or slightly pointed in some specimens. Males have a conoid tail, with a rounded-to-slightly-pointed terminus. The phylogenetic relationships of the species were reconstructed and investigated using partial sequencing of the D2-D3 expansion segments of large subunits, as well as internal transcribed spacer regions (LSU D2-D3 and ITS rDNA) based on Bayesian inference (BI). P. vandenbrandei has formed a clade with P. neonanus, P. minor, P. nainianus, P. chongqinjensis, P. pedrami, P. baldaccii, P. leptos and P. rostrocaudatus with maximal support (BPP = 1.00). To the best of our knowledge, this is the first report of P. vandenbrandei in Iran and the first molecular characterization of the species worldwide.

Characterization of Hoplolaimus seinhorsti and Hoplolaimus pararobustus (Tylenchina: Hoplolaimidae) from banana, with phylogeny and species delineation in the genus Hoplolaimus.

The morphological and molecular characterisations of two lance nematode species isolated from the rhizosphere of banana, Hoplolaimus seinhorsti and H. pararobustus, are provided based on an integrative study that includes light and scanning electron microscopy, phylogenetic analysis and two tree-based molecular species delimitation methods (GMYC and bPTP). Nineteen new sequences were obtained, including 5 partial 18S rRNA, 6 D2-D3 of 28S rRNA, 1 ITS rRNA and 7 COI mtDNA (the first COI sequences of H. seinhorsti and H. pararobustus), and an updated morphological character comparison of 37 Hoplolaimus species is presented. The tree-based molecular species-delimitation approaches employed gave markedly differing results, and also showed remarkable discrepancies among the investigated genes, although the bPTP output was found to agree well with established morphological species delimitations. Both species-delimitation approaches did, however, provide the same output for the COI mtDNA sequences, and the COI mtDNA gene sequence was also found to correspond better to established morphological species. It is therefore recommended by this paper as representing the most suitable barcode marker for Hoplolaimus species identification. This integrative study also resulted in the corrective reassignment of 17 gene sequences that were previously unidentified or incorrectly classified, as well as concluding that H. pararobustus consists of two cryptic species.

Detection and differentiation of Leishmania parasites in asymptomatic canine by High-Resolution Melting analysis of microsatellite fragment in ITS gene.

The high prevalence of Leishmania infection was reported in dogs as the main reservoir of CanL in many locations in the old world. Detection and firmly identification of Leishmania species in asymptomatic dogs by reliable method was considered and employed. Non-invasive and non-anesthetized blood sampling in asymptomatic dogs was conducted. Nested, conventional and real-time PCR with HRM technique was performed targeting ITS-rDNA gene. 88 asymptomatic dogs were sampled from three CanL endemic provinces of Iran in 2018-2019. 23 blood samples were Leishmania positive. L. major, L. tropica and L. infantum were accurately identified for the first time with HRM targeting ITS2-microsatellite. Three samples were mixed infections. CLC software TM predictions for microsatellite ITS-rDNA were 86.93 °C: L. major, 85.76 °C: L. tropica and 86.04 °C: L. infantum. Standard strains of Leishmania species were accurately separated with almost one to 2 °C deference (L. major: 86.61 °C, L. infantum: 85.41 °C, L. tropica: 84.82 °C). Each HRM curve represents one species in a sample for helping accurate identification of Leishmania species and even mixed infection when two curves are present. Detecting parasites at primary stages in asymptomatic cases is essential using Real-time HRM. As same as mammalian Leishmania in rodents which is present at early stages and non-pathogenesis, only L. major would exist and other Leishmania disappears. This can conclude also for L. major, L. infantum and L. tropica in dogs. The role of L. major existence in canine blood should be investigated more.

Morphological Characterization of Fresh and 20-Yr-Old Fixed Nematode Specimens of Sauertylenchus Maximus (Allen, 1955) Siddiqi, 2000 Deposited in the USDA Nematode Collection from Arlington National Cemetery, VA, USA.

Sauertylenchus maximus was discovered during a survey conducted at the Arlington National Cemetery, Virginia, for the type specimens of Hoplolaimus galeatus. Besides the fresh material, the fixed specimens of S. maximus were also studied by molecular and morphological means. The morphological and morphometric characteristics of the recovered fresh material were consistent with the original and other description(s) of this species. The fixed specimens used in this study were preserved in a 3% formaldehyde and 2% glycerin solution for over 20 yr. Molecular analyses of the fresh and fixed specimens were performed using internal transcribed spacer, D2-D2 expansion segments of 28S large subunits, and 18S small subunit ribosomal DNA sequences. To our knowledge, this represents the first report of S. maximus from Virginia and the first report of a successful DNA extraction from fixed nematode specimens.

Morphological and Molecular Characterization of Tylenchorhynchus Clarus Allen, 1955 and T. Zeae Sethi & Swarup, 1968 (Rhabditida: Telotylenchidae) from Iraq.

During a survey on the biodiversity of plant-parasitic nematodes in Misan province (southeast Iraq), Tylenchorhynchus clarus and T. zeae were discovered around the rhizosphere of sugarcane and pumpkin, respectively. The morphological and morphometric data were provided for the recovered species. The morphological characters of both populations are in agreement with the type populations and other populations of them. To the best of our knowledge, this is the first report of these two species from Iraq, and a first report of the association of T. zeae with pumpkin. Molecular phylogenetic analyses of the Iraqi populations of T. clarus and T. zeae using the D2-D3 expansion segments of 28S rDNA and internal transcribed spacer (ITS) rDNA sequences using Bayesian inference (BI), showed they form maximally supported clades with other sequences of both species.

Fungal diversity present on rocks from a polar desert in continental Antarctica assessed using DNA metabarcoding.

We evaluated the fungal diversity associated with carbonate veins and two types of salt encrustation in rocks in a polar desert region of continental Antarctica using DNA a metabarcoding approach. We detected 262,268 reads grouped into 517 amplicon sequence variants (ASVs) assigned to the phyla Ascomycota, Basidiomycota, Mortierellomycota and Mucoromycota. Fourteen ASVs belonging to the genera Trichosporon, Mortierella, Penicillium, Aspergillus, Cladosporium, Coprinellus, Pleurotus and Pseudogymnoascus were assessed to be dominant taxa. The fungal communities of the three habitats sampled displayed high diversity indices when compared with other habitats of Antarctica, although differing in detail, with the highest diversity indices in the gypsum crust type 2. Only 48 of the 517 ASVs (9.28%) were detected in all three habitats, including dominant, intermediate and minor components. In contrast with previous studies of fungal communities living in the ultra-extreme conditions of continental Antarctica, application of metabarcoding revealed the DNA of a rich and complex resident fungal community. The ASVs detected included fungi with different ecological roles, with xerophilic, human- and animal-associated, phytopathogenic, saprotrophic, mutualistic, psychrotolerant and cosmopolitan taxa. This sequence diversity may be the result of deposition of fungal propagules over time driven by air currents, precipitation or human activities in the region.

Species delimitation of Gyrodactylus (Monogenea: Gyrodactylidae) infecting the southernmost cyprinids (Actinopterygii: Cyprinidae) in the New World.

The genus Gyrodactylus von Nordmann, 1832 represents one of the most diverse and widespread taxa within Monogenea, with approximately 500 species described worldwide. Thirty-three species of Gyrodactylus have been recorded in Mexico, and in the last two decades, at least 26 new species have been described mainly from freshwater fish families such as poeciliids, goodeids, profundulids, characids, and cichlids. In this study, we describe two new species of Gyrodactylus infecting freshwater cyprinids based on morphological and molecular characteristics. Gyrodactylus ticuchi n. sp. and Gyrodactylus tobala n. sp. were recovered from Notropis moralesi de Buen and N. imeldae Cortés, respectively, captured in five localities from the State of Oaxaca, Mexico. The new species differ slightly from their congeners in the morphology of the haptoral hard parts and the male copulatory organ. Sequences of the Internal Transcribed Spacers rDNA (ITS1-5.8S-ITS2), cytochrome oxidase subunit I (cox1), and the D2 + D3 domains of the large subunit (28S rDNA) were obtained from multiple specimens and analyzed using Maximum Likelihood (ML) and Bayesian Inference (BI). Phylogenetic hypotheses using ITS rDNA, cox1, and 28S rDNA genes recovered two new species of Gyrodactylus from N. moralesi and N. imeldae; we briefly discuss their phylogenetic relationship with other congeners. These gyrodactylids represent the first species described in species of Notropis from southern Mexico, the cyprinids exhibiting the southernmost distribution in the New World.

Development of Real-Time and Conventional PCR Assays for Identifying a Newly Named Species of Root-Lesion Nematode (Pratylenchus dakotaensis) on Soybean.

A rapid and accurate PCR-based method was developed in this study for detecting and identifying a new species of root-lesion nematode (Pratylenchus dakotaensis) recently discovered in a soybean field in North Dakota, USA. Species-specific primers, targeting the internal transcribed spacer region of ribosomal DNA, were designed to be used in both conventional and quantitative real-time PCR assays for identification of P.dakotaensis. The specificity of the primers was evaluated in silico analysis and laboratory PCR experiments. Results showed that only P.dakotaensis DNA was exclusively amplified in conventional and real-time PCR assays but none of the DNA from other control species were amplified. Detection sensitivity analysis revealed that the conventional PCR was able to detect an equivalent to 1/8 of the DNA of a single nematode whereas real-time PCR detected an equivalent to 1/32 of the DNA of a single nematode. According to the generated standard curve the amplification efficiency of the primers in real-time PCR was 94% with a R2 value of 0.95 between quantification cycle number and log number of P.dakotaensis. To validate the assays to distinguish P.dakotaensis from other Pratylenchus spp. commonly detected in North Dakota soybean fields, 20 soil samples collected from seven counties were tested. The PCR assays amplified the DNA of P.dakotaensis and discriminated it from other Pratylenchus spp. present in North Dakota soybean fields. This is the first report of a species-specific and rapid PCR detection method suitable for use in diagnostic and research laboratories for the detection of P.dakotaensis.

Redescription and synonymization of Oscheius citri Tabassum, Shahina, Nasira and Erum, 2016 (Rhabditida, Rhabditidae) from India and its taxonomical consequences.

A population of a nematode species belonging to the genus Oscheius was isolated in western Uttar Pradesh, India. Morphological and morphometrical studies on this species showed its high similarity with six species described previously from Pakistan (Oscheius citri, O. cobbi, O. cynodonti, O. esculentus, O. punctatus and O. sacchari). The molecular analysis of the ITS1-5.8S-ITS2 rDNA sequences of the Indian population and the six species described from Pakistan showed that all the sequences are almost identical. Thus, based on morphological and molecular characteristics, all of the six above-mentioned Pakistani species and Indian strain do not differ from each other, hence can be considered synonyms. The correct name for this taxon is the first described species O. citri. Additionally, the phylogenetic analysis of the 18S rDNA and the 28S rDNA sequences showed that Oscheius citri is sister to the clade formed by O. chongmingensis and O. rugaoensis from China. The high similarity of morphological and morphometric characteristics of O. citri and other species, O. maqbooli, O. nadarajani, O. niazii, O. shamimi and O. siddiqii, suggest their conspecificity; however, lack of molecular data for these species does not allow this hypothesis to be tested.

Morphological and molecular characterization of Bitylenchus hispaniensis (Nematoda: Telotylenchidae) from Iran.

During a survey on the biodiversity of plant-parasitic nematodes in Khuzestan province (southwest Iran), Bitylenchus hispaniensis was discovered around the rhizosphere of the euphrates poplar tree. The morphological and morphometric data were provided for the recovered species. To the best of our knowledge, this is the first report of B. hispaniensis from Iran and for the first time in association with euphrates poplar worldwide. Molecular phylogenetic analyses of the Iranian population of B. hispaniensis using the D2-D3 expansion segments of 28S rDNA and internal transcribed spacer (ITS rDNA) sequences using Bayesian inference (BI), showed a maximally supported clade with other sequences of the species.

A new collar rot disease of cowpea (Vigna unguiculata) caused by Aplosporella hesperidica in India.

Cowpea is an important pulse crop cultivated in arid and semi-arid regions of the world. During field survey, a characteristic wilt was observed in around 45 ha of cowpea fields with incidence 17-25%. Infection was seen in pre-flowering stage and infected plants showed quick wilt symptoms with tan lesions near the stem-soil interface. Fungal pathogens associated were isolated on PDA, which produced dark to grey olivaceous colonies in the centre, and aerial mycelia were appressed with floccose and white to smoke-grey. Conidia are aseptate, initially hyaline, smooth-walled, broadly ellipsoidal with rounded ends becoming dark brown. Based on these morphological features, the fungal pathogen was identified as Aplosporella sp. The ITS-rDNA region was amplified using ITS1/ITS4 primers and sequenced. The nBLAST and phylogenetic analysis confirmed the pathogen as Aplosporella hesperidica. The Koch's postulates were performed on 45-days-old cowpea plants with mycelial disc of A. hesperidica. Development of typical necrotic lesions was observed after 28 days of post-inoculation and the pathogen's identity was confirmed based on re-isolation. Efficacy of fungicides evaluated in vitro showed that the pathogen is highly sensitive to systemic fungicides rather than the contact fungicides. The cowpea production was severely affected owing to the causative agent A. hesperidica. The collar rot disease of cowpea by A. hesperidica is the first report in India. SIGNIFICANCE AND IMPACT OF THE STUDY: A new collar rot disease of cowpea recorded from India has been investigated. The necrotic lesions were enlarged and eventually quick wilt and death of the host plant was observed with incidence ranged from 17 to 25%. Associated fungal pathogen was isolated and identified as Aplosporella hesperidica based on morphology and ITS-rDNA sequence analysis. Koch's postulates were performed under greenhouse conditions and in vitro evaluation of fungicides shows that the pathogen is sensitive to systemic fungicides. This is the first report of A. hesperidica causing collar rot disease of cowpea in India.

Fusarium fujikuroi causing Fusarium wilt of Lactuca serriola in Korea.

Lactuca serriola L. (syn. L. scariola L.) is an annual Asteraceae plant, native to Europe, and accidentally introduced to Korea in the late 1970s (Yim and Jeon, 1981). The Korean Ministry of Environment designated this weed as a harmful plant, which may disturb the balance of ecosystems (Kim et al., 2013). In July 2019, wilting symptoms of prickly lettuce were found among a roadside in Sangju (36°26'15'' N, 128°07'35'' E), Korea, with a disease incidence of 60%. Initial symptoms appeared pale to dark brown lesions on the basal stem and leaves of the plant, and over time the lesions expanded to the upper parts of the plant, resulting in extensive rot. Ultimately, the plants wilted and died. Symptomatic vascular tissues (5 x 5 mm2) of two diseased plants were surface sterilized in 2% NaClO solution for 1 min, followed by 70% ethanol for 1min, and rinsed two times in sterile distilled water. The pieces were placed on potato dextrose agar (PDA) and incubated at 25°C in the dark for a week. Each single spore isolate was obtained from the hypha tip growing on PDA and examined for morphological and molecular analysis. A representative isolate has been deposited at the Korean Agricultural Culture Collection (KACC49588). Macroconidia were slender, straight, and measured 20.4 to 59.6 × 2.5 to 3.9 µm (n=50), with three to five septa. Microconidia were clavate and measured 6.1 to 13 × 2.5 to 3.3 µm (n=50). Chlamydospores were absent. The colonies developed white aerial mycelium and turned pale purple after a week on PDA. Both morphological and cultural characteristics of the Korean isolate were close to Fusarium fujikuroi (Leslie and Summerell, 2006). Also, DNA sequence-based identification was carried out using primer sets of ITS1-F/ITS4 for the internal transcribed spacer (ITS) rDNA region, Btu-F-F01/Btu-F-R01 for ß-tubulin (TUB) gene (Watanabe et al., 2011), and EF-1/EF-2 for translation elongation factor 1-alpha (TEF) gene. The resulting sequences were deposited in GenBank with accession numbers MT102938 for ITS, MT182734 for TUB, and MT625962 for TEF. On a BLASTn search, the Korean isolate revealed 100% sequence identity with the verified sequences of F. fujikuroi MF984413.1 for ITS, 99.79% (1 out of 368 bp is different) with U34415.1 for TUB, and 99.85% (1 out of 675 bp) with MN193860.1 for TEF. Pathogenicity was tested by dipping the roots of five healthy prickly lettuce seedlings in the spore suspension (1 × 106 conidia/ml) for 1 hour. Inoculated plants were transplanted into pots and maintained in a growth chamber at 90% relative humidity and 20°C. Five non-inoculated plants served as controls. After four weeks, wilt symptoms accompanied by expanding brown spots were observed on the basal stem and leaves of all inoculated seedlings, whereas the control plants remained symptomless. The fungus present on the inoculated plants was identical to one of the original infections, fulfilling Koch's postulates. Based on the morphological characteristics, sequencing data, and pathogenicity test, the pathogen was identified as F. fujikuroi. Bakanae (foolish seedling) disease caused by F. fujikuroi is one of the most serious rice diseases in Asia, but also this pathogen has been recorded on Lactuca sativa in Thailand (Farr and Rossman 2020). To our knowledge, this is the first report of F. fujikuroi causing Fusarium wilt on L. serriola in Korea.

A new species of Creptotrematina (Trematoda: Allocreadiidae) from characid fishes of Brazil: morphological and molecular data.

A new species of Creptotrematina Yamaguti, 1954 was collected from characid fishes, Astyanax fasciatus (Cuvier, 1819) and Astyanax lacustris Lucerna & Soares, 2016 from the Batalha River in the State of Sao Paulo, Brazil. The new species most closely resembles Creptotrematina aguirrepequenoi, but differs by the elongated shape of vitelline follicles, the extension of these follicles in the posterior end of body and the fact that they are not confluent. The morphological differences were confirmed through molecular data. Three specimens were sequenced, and molecular analyses were based on the internal transcribed spacers 2 and D1-D3 domains of the 28S ribosomal RNA gene. The obtained topologies showed the new species as a sister taxon of C. aguirrepequenoi, a species originally described from Astyanax mexicanus in Mexico, and later found in Astyanax aeneus in Costa Rica. Isolates of the new species are reciprocally monophyletic, and genetic distance values are similar to those observed in other species pairs within Allocreadiidae. These findings corroborate that the genus Creptotrematina is mostly a parasite of characids, and widely extended across the Americas, with representative species occurring between Argentina and northern Mexico.