Phylogenetic diversity of Rhizobium species recovered from nodules of common beans (Phaseolus vulgaris L.) in fields in Uganda: R. phaseoli, R. etli, and R. hidalgonense.

A total of 75 bacterial isolates were obtained from nodules of beans cultivated across 10 sites in six agro-ecological zones in Uganda. Using recA gene sequence analysis, 66 isolates were identified as members of the genus Rhizobium, while nine were related to Agrobacterium species. In the recA gene tree, most Rhizobium strains were classified into five recognized species. Phylogenetic analysis based on six concatenated sequences (recA-rpoB-dnaK-glnII-gyrB-atpD) placed 32 representative strains into five distinct Rhizobium species, consistent with the species groups observed in the recA gene tree: R. phaseoli, R. etli, R. hidalgonense, R. ecuadorense, and R. sophoriradicis, with the first three being the predominant. The rhizobial strains grouped into three nodC subclades within the symbiovar phaseoli clade, encompassing strains from distinct phylogenetic groups. This pattern reflects the conservation of symbiotic genes, likely acquired through horizontal gene transfer among diverse rhizobial species. The 32 representative strains formed symbiotic relationships with host beans, while the Agrobacterium strains did not form nodules and lacked symbiotic genes. Multivariate analysis revealed that species distribution was influenced by the environmental factors of the sampling sites, emphasizing the need to consider these factors in future effectiveness studies to identify effective nitrogen-fixing strains for specific locations.

Two new species of Fusarium in the F. incarnatum-equiseti species complex from Oryza sativa in Iran.

Identification of Fusarium species associated with diseases symptoms in plants is an important step toward understanding the ecology of plant-fungus associations. In this study, four Fusarium isolates were obtained from root rot of Oryza sativa L. in Izeh (southwest of Iran) and identified based on phylogenetic analyses combined with morphology. Phylogenetic analyses based on combined translation elongation factor 1-α, calmodulin, RNA polymerase II second largest subunit, and Beta-tubulin (tub2) sequence data delimited two new species, namely F. khuzestanicum and F. oryzicola spp. nov., from previously known species of Fusarium within F. incarnatum-equiseti species complex (FIESC). Morphologically, F. khuzestanicum produces the macroconidia with distinctly notched to foot-shaped basal cells, while basal cells in the macroconidia of F. oryzicola are more extended and distinctly elongated foot shape. Furthermore, these two new species are distinguished by the size of their sporodochial phialides and macroconidia. The results of the present show that the FIESC species complex represent more cryptic species.

Amanita satotamagotake sp. nov., a cryptic species formerly included in Amanita caesareoides.

We evaluated the inclusion of a cryptic species in a Japanese Amanita caesareoides population. We sampled A. caesareoides specimens under various vegetation and climate conditions, and then conducted phylogenetic analyses on sequences from seven loci. The A. caesareoides specimens showed two distinct groups, except when the ITS phylogeny was considered. These two phylogroups showed different distributions: subalpine-cool temperate and temperate-subtropical areas. Although these two phylogroups overlapped in terms of basidiospore size, the latter tended to exhibit smaller basidiospores. In addition, only the former showed mycelial growth on nutrient agar. Based on these phylo-morpho-ecophysiological characteristics, we separated the specimens labeled with the name A. caesareoides into two species. As the lectotype of A. caesareoides showed similarity to the former by DNA analysis, the latter was described as a new species, namely A. satotamagotake. Based on the geographic patterns of the two species, A. satotamagotake may have invaded the natural habit of A. caesareoides because of global warming.

An Extended Application of the Fast Multi-Locus Ridge Regression Algorithm in Genome-Wide Association Studies of Categorical Phenotypes.

Categorical (either binary or ordinal) quantitative traits are widely observed to measure count and resistance in plants. Unlike continuous traits, categorical traits often provide less detailed insights into genetic variation and possess a more complex underlying genetic architecture, which presents additional challenges for their genome-wide association studies. Meanwhile, methods designed for binary or continuous phenotypes are commonly used to inappropriately analyze ordinal traits, which leads to the loss of original phenotype information and the detection power of quantitative trait nucleotides (QTN). To address these issues, fast multi-locus ridge regression (FastRR), which was originally designed for continuous traits, is used to directly analyze binary or ordinal traits in this study. FastRR includes three stages of continuous transformation, variable reduction, and parameter estimation, and it can computationally handle categorical phenotype data instead of link functions introduced or methods inappropriately used. A series of simulation studies demonstrate that, compared with four other continuous or binary or ordinal approaches, including logistic regression, FarmCPU, FaST-LMM, and POLMM, the FastRR method outperforms in the detection of small-effect QTN, accuracy of estimated effect, and computation speed. We applied FastRR to 14 binary or ordinal phenotypes in the Arabidopsis real dataset and identified 479 significant loci and 76 known genes, at least seven times as many as detected by other algorithms. These findings underscore the potential of FastRR as a very useful tool for genome-wide association studies and novel gene mining of binary and ordinal traits.

Modeling the stress and forces on multi-channel TMS coil arrays in high-field MRI scanners.

Transcranial magnetic stimulation (TMS) is a non-invasive method for stimulating the cortex. Concurrent functional magnetic resonance imaging can show changes in TMS-induced activity in the whole brain, with the potential to inform brain function research and to guide the development of TMS therapy. However, the interaction of the strong current pulses in the TMS coil in the static main magnetic field of the MRI produces high Lorentz forces, which may damage the coil enclosure and compromise the patient's safety. We studied the time-dependent mechanical behavior and durability of two multi-locus TMS (mTMS) coil arrays inside a high-field MRI bore with finite element modeling. In addition, coil arrays were built and tested based on the simulation results. We found that the current pulses produce shock waves and time-dependent stress distribution in the coil plates. The intensity and location of the maximum stress depend on the current waveform, the coil combination, and the transducer orientation relative to the MRI magnetic field. We found that 30% glass-fiber-filled polyamide is the most durable material out of the six options studied. In addition, novel insights for more durable TMS coil designs were obtained. Our study contributes to a comprehensive understanding of the underlying mechanisms responsible for the structural failure of mTMS coil arrays during stimulation within high static magnetic fields. This knowledge is essential for developing mechanically stable and safe mTMS-MRI transducers.

Multi-locus imprinting disturbance (MLID): interim joint statement for clinical and molecular diagnosis.

BACKGROUND: Imprinting disorders are rare diseases resulting from altered expression of imprinted genes, which exhibit parent-of-origin-specific expression patterns regulated through differential DNA methylation. A subgroup of patients with imprinting disorders have DNA methylation changes at multiple imprinted loci, a condition referred to as multi-locus imprinting disturbance (MLID). MLID is recognised in most but not all imprinting disorders and is also found in individuals with atypical clinical features; the presence of MLID often alters the management or prognosis of the affected person. Some cases of MLID are caused by trans-acting genetic variants, frequently not in the patients but their mothers, which have counselling implications. There is currently no consensus on the definition of MLID, clinical indications prompting testing, molecular procedures and methods for epigenetic and genetic diagnosis, recommendations for laboratory reporting, considerations for counselling, and implications for prognosis and management. The purpose of this study is thus to cover this unmet need. METHODS: A comprehensive literature search was conducted resulting in identification of more than 100 articles which formed the basis of discussions by two working groups focusing on clinical diagnosis (n = 12 members) and molecular testing (n = 19 members). Following eight months of preparations and regular online discussions, the experts from 11 countries compiled the preliminary documentation and determined the questions to be addressed during a face-to-face meeting which was held with the attendance of the experts together with four representatives of patient advocacy organisations. RESULTS: In light of available evidence and expert consensus, we formulated 16 propositions and 8 recommendations as interim guidance for the clinical and molecular diagnosis of MLID. CONCLUSIONS: MLID is a molecular designation, and for patients with MLID and atypical phenotypes, we propose the alternative term multi-locus imprinting syndrome. Due to the intrinsic variability of MLID, the guidelines underscore the importance of involving experts from various fields to ensure a confident approach to diagnosis, counselling, and care. The authors advocate for global, collaborative efforts in both basic and translational research to tackle numerous crucial questions that currently lack answers, and suggest reconvening within the next 3-5 years to evaluate the research advancements and update this guidance as needed.

Molecular epidemiological investigation of Cryptococcus spp. isolated from cats in Japan using multi-locus sequence typing.

Cryptococcosis is an important fungal infection for both humans and cats, but molecular epidemiological studies on strains isolated from cats are limited. We conducted multi-locus sequence typing analysis and antifungal susceptibility testing of 14 Cryptococcus spp. strains from domestic cats in Japan and one strain isolated from a cat in Singapore. All 14 strains from domestic cats in Japan were identified as Cryptococcus neoformans molecular type VNI. The sequence types (STs) included eight cases of ST5, five cases of ST31, and one novel ST. VNI ST5 is the most frequently isolated strain in Japanese patients as well, while there are no records of VNI ST31 being isolated from Japanese patients. The Singaporean cat strain was identified as C. gattii VGIIb (C. deuterogattii), ST7. We compared these results with strains previously reported to have been isolated from cats. This comparison suggested that molecular types of Cryptococcus spp. isolated from cats may differ depending on the country. In the antifungal susceptibility testing of C. neoformans, one strain each exceeded the epidemiological cutoff value (ECV) for amphotericin B and 5-fluorocytosine, while two strains exceeded the ECV for fluconazole. This study reveals the molecular epidemiology of Cryptococcus spp. isolated from cats with cryptococcosis in Japan. It suggests that investigating Cryptococcus spp. carried by cats, which share close living environments with humans, may contribute to the health of both cats and human populations.

Cryptococcosis is an important fungal disease in both humans and cats. We genotyped strains isolated from cats with cryptococcosis in Japan. Our findings revealed that the most common genotype infecting both cats and humans in Japan is identical.

Phylogenetic Insights Reveal New Taxa in Thyridariaceae and Massarinaceae.

Pleosporales is a highly diverse (and the largest) order in Dothideomycetes, and it is widespread in decaying plants in various environments around the world. During a survey of fungal diversity in Sichuan Province, China, specimens of hyphomycetous and Thyridaria-like fungi were collected from dead branches of pine trees and cherry trees. These taxa were initially identified as members of Massarinaceae and Thyridariaceae through morphological examination. Phylogenetic analyses of the Thyridariaceae, combining ITS, SSU, LSU, RPB2, and TEF1 sequence data, indicated a distinct clade sister to Pseudothyridariella and Thyridariella, distinct from any genus in the family. Thus, a new genus, Vaginospora, is proposed to accommodate the type species Vaginospora sichuanensis, which is characterized by semi-immersed globose to oblong ascomata with an ostiolar neck, cylindrical to clavate asci with an ocular chamber, and hyaline to dark brown, fusiform, 3-5-transversely septate ascospores with an inconspicuous mucilaginous sheath. Based on the morphological comparisons and multi-locus phylogenetic analyses (ITS, SSU, LSU, RPB2, and TEF1) of the Massarinaceae, we have identified three collections belonging to the genus Helminthosporium, leading us to propose H. filamentosa sp. nov., H. pini sp. nov., and H. velutinum as a new host record. According to Phylogenetic analysis, H. pini formed an independent clade sister to H. austriacum and H. yunnanense, and H. filamentosa represents the closest sister clade to H. quercinum. Helminthosporium pini is distinct from H. austriacum by the shorter conidiophores and H. yunnanense by the longer and wider conidia. The H. filamentosa differs from H. quercinum in having longer conidiophores and smaller conidia. This study extends our understanding of diversity within Thyridariaceae and Helminthosporium. Our findings underscore the rich biodiversity and potential for discovering novel fungal taxa within these groups.

The genetic diversity and evolution analysis of the Hainan melioidosis outbreak strains.

Melioidosis is a zoonotic disease, with its outbreaks being rare and indicative of an unusual concurrence of extreme climate and natural environmental factors. An outbreak of melioidosis cases emerged in Hainan following Typhoon "Dianmu" from October to December 2021, presenting an opportunity to identify the environmental sources of infection for these cases due to its nature as a well-defined point-source cluster. To investigate the relationship between the occurrence of these melioidosis cases and the environment, we extracted the entire genome of 25 clinical strains and conducted MLST typing, followed by whole genome sequencing and analysis of molecular genetic information for four ST46 genotypes from these strains. Phylogenetic and evolutionary relationships between Hainan sequence types (STs) and those found in other endemic regions were analyzed using IslandPath-DIMO, PHASTER, e-BURST, PHYLOViZ, and the maximum likelihood method. Notably, a total of 25 clinical strains were identified, encompassing 12 STs (ST46, ST1105, ST1991, ST30, ST1992, ST50, ST164, ST55, ST70, ST1993, ST1545, and ST58), with ST1991, ST1992, and ST1993 being newly discovered subtypes. PHYLOViZ clustering analysis divided the strains into two groups (A and B), both closely related to the Asian region. Phylogenetic tree analysis further revealed that most of the strains in this study were closely related to those found in Australia and Thailand. Analysis of patient information and visits to their residences suggested that contaminated water sources might be the primary source of infection during this outbreak. Our findings underscore that extreme weather events, such as typhoons, significantly increase the infection rate of B. pseudomallei, along with its genetic diversity, necessitating additional prevention strategies to control these B. pseudomallei infections.

Average Nucleotide Identity and Digital DNA-DNA Hybridization Analysis Following PromethION Nanopore-Based Whole Genome Sequencing Allows for Accurate Prokaryotic Typing.

Whole-genome sequencing (WGS) is revolutionizing clinical bacteriology. However, bacterial typing remains investigated by reference techniques with inherent limitations. This stresses the need for alternative methods providing robust and accurate sequence type (ST) classification. This study optimized and evaluated a GridION nanopore sequencing protocol, adapted for the PromethION platform. Forty-eight Escherichia coli clinical isolates with diverse STs were sequenced to assess two alternative typing methods and resistance profiling applications. Multi-locus sequence typing (MLST) was used as the reference typing method. Genomic relatedness was assessed using Average Nucleotide Identity (ANI) and digital DNA-DNA Hybridization (DDH), and cut-offs for discriminative strain resolution were evaluated. WGS-based antibiotic resistance prediction was compared to reference Minimum Inhibitory Concentration (MIC) assays. We found ANI and DDH cut-offs of 99.3% and 94.1%, respectively, which correlated well with MLST classifications and demonstrated potentially higher discriminative resolution than MLST. WGS-based antibiotic resistance prediction showed categorical agreements of ≥ 93% with MIC assays for amoxicillin, ceftazidime, amikacin, tobramycin, and trimethoprim-sulfamethoxazole. Performance was suboptimal (68.8-81.3%) for amoxicillin-clavulanic acid, cefepime, aztreonam, and ciprofloxacin. A minimal sequencing coverage of 12× was required to maintain essential genomic features and typing accuracy. Our protocol allows the integration of PromethION technology in clinical laboratories, with ANI and DDH proving to be accurate and robust alternative typing methods, potentially offering superior resolution. WGS-based antibiotic resistance prediction holds promise for specific antibiotic classes.

Integrating multilocus phylogeny and morphological analysis reveals the prevalence of Phytophthora meadii (McRae) associated with abnormal leaf fall disease of Hevea brasiliensis in India.

The Oomycetes fungus Phytophthora spp. which causes Abnormal leaf fall (ALF) disease poses a significant threat as one of the most devastating diseases affecting rubber trees in India. A total of 30 Phytophthora isolates were obtained from ALF-affected samples collected during the Southwest monsoon season of Kerala. The colony morphology of Phytophthora isolates revealed eight different types of growth patterns, with stellate, stellate striated, and petaloid patterns growing rapidly, whereas chrysanthemum pattern grew slowly. Sporangia were papillate to non-papillate in various shapes, and sporangiophores exhibited simple, simple sympodial, or irregularly branching patterns. Highly virulent isolates exhibited petaloid morphology and rapid growth rates. Regardless of their virulence, all isolates showed susceptibility to the fungicide metalaxyl. Under in vitro conditions, the highly virulent isolate (R17) from rubber caused severe infections in chili, brinjal, and tomato with brown water-soaked lesions. Sequence analysis and multi-locus phylogeny of Internal transcribed spacer (ITS), cCytochrome c oxidase 1 (COX 1), Heat shock protein 90 (HSP 90), and Ribosomal protein L10 (RPL 10) confirmed the pathogen as Phytophthora meadii. A comprehensive understanding of both morphological and molecular traits of P. meadii is crucial for precise identification and future genetic variability studies.

Genetic Characterization, Antibiotic Resistance, and Virulence Genes Profiling of Bacillus cereus Strains from Various Foods in Japan.

Bacillus cereus sensu stricto is a foodborne pathogen that causes food poisoning. Their spore and biofilm-forming abilities persist in various environments and foods. This study investigated the prevalence, virulence, antibiotic resistance, and genetic diversity of B. cereus s. s. strains isolated from various food samples. Of 179 samples, 22.34% were positive for B. cereus s. s., with significantly high detection rates in milk products and raw chicken meat. Forty strains were isolated from positive samples. Matrix-assisted laser desorption ionization/time of flight mass spectrometry analysis revealed nine distinct clusters and multi-locus sequence typing revealed 34 sequence types including 23 novel sequences, demonstrating high genetic diversity among the isolates. PCR analysis revealed that all the strains contained at least one toxin gene, but none contained the cytK gene. Antibiotic resistance tests revealed that all isolates were classified as multidrug-resistant, with high resistance levels, particularly to ß-lactam antibiotics and vancomycin, but were susceptible to gentamicin. All isolates showed variations in biofilm formation. This study highlights the significant public health risk due to B. cereus s. s. and underscores the need for stringent monitoring and control measures in food production to manage antimicrobial resistance and ensure food safety.

A Biological Comparison of Three Colletotrichum Species Associated with Alfalfa Anthracnose in Northern China.

Anthracnose caused by various species of Colletotrichum is one of the most prevalent diseases in alfalfa worldwide that not only reduces forage yields but also severely compromises forage quality. A comprehensive survey was conducted in 2020 in the main production regions of northern China. The survey results showed that alfalfa anthracnose is prevalent in northern China, with the disease incidence ranging from 9% to 45% and the disease index from 5 to 17 (maximum possible score: 100). In total, 24 isolates were collected and identified as three Colletotrichum species (C. trifolii, C. truncatum and C. americae-borealis) based on morphological characteristics and phylogenetic analysis (combined sequences ITS, HIS3, ACT and GAPDH). The three species displayed remarkable environmental adaptability, exhibiting a capacity for growth, sporulation and conidial germination in temperatures ranging from 4 to 35 °C and in different nutrient conditions. Pathogenicity assays showed that C. trifolii was more virulent than the other two species, although the growth vigor (in terms of colony diameter, sporulation and conidial germination) of C. truncatum was the greatest.

Isolation and molecular characterization of a novel relapsing fever group Borrelia from the white-eared opossum Didelphis albiventris in Brazil.

This study aimed to detect, isolate and to characterize by molecular methods a relapsing fever group (RFG) Borrelia in white-eared opossums (Didelphis albiventris) from Brazil. During 2015-2018, when opossums (Didelphis spp.) were captured in six municipalities of the state of São Paulo, Brazil, molecular analyses revealed the presence of a novel RFG Borrelia sp. in the blood of seven opossums (Didelphis albiventris), out of 142 sampled opossums (4.9% infection rate). All seven infected opossums were from a single location (Ribeirão Preto municipality). In a subsequent field study in Ribeirão Preto during 2021, two new opossums (D. albiventris) were captured, of which one contained borrelial DNA in its blood. Macerated tissues from this infected opossum were inoculated into laboratory animals (rodents and rabbits) and two big-eared opossums (Didelphis aurita), which had blood samples examined daily via dark-field microscopy. No spirochetes were visualized in the blood of the laboratory animals. Contrastingly, spirochetes were visualized in the blood of the two D. aurita opossums between 12 and 25 days after inoculation. Blood samples from these opossums were used for a multi-locus sequencing typing (MLST) based on six borrelial loci. Phylogenies inferred from MLST genes positioned the sequenced Borrelia genotype into the RFG borreliae clade basally to borreliae of the Asian-African group, forming a monophyletic group with another Brazilian isolate, "Candidatus B. caatinga". Based on this concatenated phylogenetic analysis, which supports that the new borrelial isolate corresponds to a putative new species, we propose the name "Candidatus Borrelia mimona".

Phylogeny and taxonomy of two new species in Dictyosporiaceae (Pleosporales, Dothideomycetes) from Guizhou, China.

Two novel species within the family Dictyosporiaceae are described and illustrated from terrestrial habitats on dead culms of bamboo and an unidentified plant, respectively. Through morphological comparisons and the multi-locus phylogenetic analyses of combined LSU, ITS, SSU, and tef1-α sequence dataset, two species, Gregaritheciumbambusicola, Pseudocoleophomaparaphysoidea are identified. Phylogenetically, both species clustered into a monophyletic clade with strong bootstrap support. Gregaritheciumbambusicola sp. nov. can be distinguished from other species within the genus based on its almost straight ascospores. Pseudocoleophomaparaphysoidea sp. nov. differs from other species in its conidiogenous cells intermixed with paraphyses, longer conidiogenous cells and larger conidia. The identification of this lineage contributes to our understanding of the classification of Dictyosporiaceae.

Temporal evolution of bacterial species and their antimicrobial resistance characteristics in wound infections of war-related injuries in Ukraine from 2014 to 2023.

BACKGROUND: This study continues surveillance of antimicrobial resistance associated with combat injuries in Ukraine. AIM: To compare species composition, antibiotic resistance profiles, and emergence of new resistance genes between 2014-2020 and 2022-2023. METHODS: This was a retrospective multi-centre microbiological survey in Ukrainian hospitals. Antibiotic susceptibility, whole-genome sequencing and multi-locus sequence typing were conducted on 154 organisms obtained from 125 casualties between 2022 and 2023. FINDINGS: The data revealed a predominance of Gram-negative bacteria, particularly Acinetobacter baumannii (35.7%), Pseudomonas aeruginosa (14.9%) and Klebsiella pneumoniae (20.7%). High levels of carbapenem resistance were observed among A. baumannii {meropenem 72.2% [39/54, 95% confidence interval (CI) 58.4-83.5]; imipenem 66.7% (36/54, 95% CI 52.5-78.9)}, K. pneumoniae [meropenem 90.6% (29/32, 95% CI 75.0-98.0); imipenem 81.2% (26/32, 95% CI 63.6-92.8)] and P. aeruginosa [meropenem 47.8% (11/23, 95% CI 26.8-69.4); imipenem 60.8% (14/23, 95% CI 38.5-80.3)] strains. A. baumannii sequence type (ST)-78 and ST-400 were prevalent from 2014 to 2020, while five strains of ST-1077 were newly identified in 2022-2023. P. aeruginosa strains showed diversity across 16 STs, with ST-773 increasing in frequency and new STs emerging, but lacking carbapenemase genes. K. pneumoniae exhibited increased genetic diversity over time, with three STs from 2014 to 2020 and six new STs, including blaNDM-1, blaOXA-48 and blaKPC2 carriers, in 2022-2023. CONCLUSION: The prevalence of multi-drug-resistant isolates with STs associated with a high risk of global dissemination is increasing.

Discovering New QTNs and Candidate Genes Associated with Rice-Grain-Related Traits within a Collection of Northeast Core Set and Rice Landraces.

Grain-related traits are pivotal in rice cultivation, influencing yield and consumer preference. The complex inheritance of these traits, involving multiple alleles contributing to their expression, poses challenges in breeding. To address these challenges, a multi-locus genome-wide association study (ML-GWAS) utilizing 35,286 high-quality single-nucleotide polymorphisms (SNPs) was conducted. Our study utilized an association panel comprising 483 rice genotypes sourced from a northeast core set and a landraces set collected from various regions in India. Forty quantitative trait nucleotides (QTNs) were identified, associated with four grain-related traits: grain length (GL), grain width (GW), grain aroma (Aro), and length-width ratio (LWR). Notably, 16 QTNs were simultaneously identified using two ML-GWAS methods, distributed across multiple chromosomes. Nearly 258 genes were found near the 16 significant QTNs. Gene annotation study revealed that sixty of these genes exhibited elevated expression levels in specific tissues and were implicated in pathways influencing grain quality. Gene ontology (GO), trait ontology (TO), and enrichment analysis pinpointed 60 candidate genes (CGs) enriched in relevant GO terms. Among them, LOC_Os05g06470, LOC_Os06g06080, LOC_Os08g43470, and LOC_Os03g53110 were confirmed as key contributors to GL, GW, Aro, and LWR. Insights from QTNs and CGs illuminate rice trait regulation and genetic connections, offering potential targets for future studies.

Unveiling genetic signatures associated with resilience to neonatal diarrhea in lambs through two GWAS approaches.

Neonatal diarrhea presents a significant global challenge due to its multifactorial etiology, resulting in high morbidity and mortality rates, and substantial economic losses. While molecular-level studies on genetic resilience/susceptibility to neonatal diarrhea in farm animals are scarce, prior observations indicate promising research directions. Thus, the present study utilizes two genome-wide association approaches, pKWmEB and MLM, to explore potential links between genetic variations in innate immunity and neonatal diarrhea in Karacabey Merino lambs. Analyzing 707 lambs, including 180 cases and 527 controls, revealed an overall prevalence rate of 25.5%. The pKWmEB analysis identified 13 significant SNPs exceeding the threshold of ≥ LOD 3. Moreover, MLM detected one SNP (s61781.1) in the SLC22A8 gene (p-value, 1.85eE-7), which was co-detected by both methods. A McNemar's test was conducted as the final assessment to identify whether there are any major effective markers among the detected SNPs. Results indicate that four markers-oar3_OAR1_122352257, OAR17_77709936.1, oar3_OAR18_17278638, and s61781.1-have a substantial impact on neonatal diarrhea prevalence (odds ratio: 2.03 to 3.10; statistical power: 0.88 to 0.99). Therefore, we propose the annotated genes harboring three of the associated markers, TIAM1, YDJC, and SLC22A8, as candidate major genes for selective breeding against neonatal diarrhea.

Rapid genotyping of Toxoplasma gondii isolates via Nanopore-based multi-locus sequencing.

Toxoplasma gondii is an obligate intracellular parasite associated with severe disease, especially in the immunosuppressed. It is also a cause of congenital malformation and abortion in both animals and humans and is considered one of the most important foodborne pathogens worldwide with different strains showing variable distribution and differing pathogenicity. Thus, strain-level differentiation of T. gondii isolates is an essential asset in the understanding of parasite's diversity, geographical distribution, epidemiology and health risk. Here, we designed and implemented an Oxford Nanopore MinION protocol to analyse genomic sequence variation including single nucleotide polymorphisms (SNPs) and insertion/deletion polymorphisms (InDel's) of four different genomic loci, part of protein coding genes SAG2, SAG3, ROP17 and ROP21. This method provided results with the sequencing depth necessary for accurate differentiation of T. gondii strains and represents a rapid approach compared to conventional techniques which we further validated against environmental samples isolated from wild wood mice. In summary, multi-locus sequence typing (MLST) of both highly conserved and more polymorphic areas of the genome, provided robust data for strain classification in a platform ready for further adaption for other strains and pathogens.

Molecular phylogeny reveals Varroa mites are not a separate family but a subfamily of Laelapidae.

Varroa mites, notorious for parasitizing honeybees, are generally classified as Varroidae. Their extremely modified morphologies and behaviors have led to debates regarding their phylogenetic position and classification as an independent family. In this study, two different datasets were employed to reconstruct the phylogenies of Varroa mites and related Laelapidae species: (1) 9257 bp from the whole 13 mitochondrial protein-coding genes of 24 taxa, (2) 3158 bp from 113 taxa using Sanger sequencing of four nuclear loci. Both mitochondrial and nuclear analyses consistently place Varroa mites within the Laelapidae. Here we propose to place Varroa mites in the subfamily Varroinae stat. nov., which represents a highly morphologically adapted group within the Laelapidae. Ancestral state reconstructions reveal that bee-associated lifestyles evolved independently at least three times within Laelapidae, with most phoretic traits originating from free-living ancestors. Our revised classification and evolutionary analyses will provide new insight into understanding the Varroa mites.