Phylogenetic Research Articles

Genetic characterization of influenza B virus and oseltamivir resistance in pediatric patients with acute respiratory infections: a cross-sectional study.

Influenza virus neuraminidase inhibitors (NAIs) drug usage can result in NAI resistance, especially in children and individuals with weakened immune systems. The aim of the present study was to identify NAI-resistant variants of IBV and to introduce probable novel mutations, phylogenetic study, and its epitope mapping based on NA gene in patients from Shiraz, Iran. A cross-sectional study was conducted between 2017 and 2018 on symptomatic children. A real-time PCR was run for IBV screening. Then, making use of direct sequencing, amplified 1401 bases of NA gene and phylogenetic tree reconstructed. Epitopes were predicted using ABCpred server. From among a total of 235 specimens, 9.7% were identified with IBV infection. Of them, sequence of NA gene for 17 isolates were analyzed. Phylogenetic analysis showed that 15 isolates belonged to Yamagata clade 3 Wisconsin/01-like subclade and 2 were related to Victoria clade 1 Brisbane/60-like subclade (Vic-1A-2). NA gene sequence analysis showed a total of 52 substitutions in which 27 were for BVic and 37 were for BYam isolates and 19 were novel substitutions. Only one substitution (S198N) was found in NA active site and T49M, I120V, N198S, N219K, S295R, D320K N340D, E358K, D384G, and D463N were found as probable resistance variants to NAIs. Epitope mapping showed some major differences in our isolates NA gene. Present study was one of the rare comprehensive studies conducted in Shiraz/Iran on IBV resistant associated variants to NAIs. We reported 11.7% mutation in NA active site and some probable NAIs resistant mutations. Epitope mapping confirmed major changes in NA gene which needs broader studies to confirm.

Genome Wide Association Analysis for Uniform Coleoptiles Emergence and Early Seedling Growth in Rice

Early seedling vigour (ESV) is a complex trait in rice. Detection of QTLs/genes controlling these traits can help us in enhancing the yield potential in rice varieties. Association mapping is a technique based on the principle of linkage disequilibrium that is used to find genes or quantitative trait loci (QTL) underlying the complex traits. In this study of haplotype breeding 281 rice genotypes were taken. ANOVA analysis showed P- value for traits and genotypes was found significant. Similarly, P-value for interaction between the traits and genotypes was also found to be highly significant (1.8663*10-208). Further, mean germination data positively correlated with mean shoot length, mean leaf number, mean culm diameter, mean shoot dry weight, and mean shoot area of 21st day among the 281 genotypes. Among the 281 number of genotypes, 111 genotypes are found to be in PCA1 and 170 genotypes are found to be in PCA2 based on the phenotypic analysis. PCA1 component constituted 29.93% and PCA2 constituted the 13.68% of total variation in the analysis. Besides, whole genome phylogenetic analysis showed three major groups of which Group 1 consists of 215 genotypes, group 2 consists of 38 genotypes and group 3 consists of 28 genotypes respectively. Especially, both the subgroups II and III comprised of the unique genotypes from the indica and aus subpopulations of rice. In this analysis, 16 significant associations (LOD Score >7) for different traits were identified using the three different models (MLM, farmCPU, and blink) for GWAS studies Especially, one major QTL was identified for the mean coleoptiles’ emergence for 10 DAS on 11th chromosome (18983591) which explained 49% of the phenotypic variance. Additionally, another major QTL contributing to the shoot length variation of 29.75% was identified in the Chr02 (32954393) for shoot length trait on 28 DAS. A candidate gene namely Os02g0778400 UMP/CMP kinase A/adenylate kinase (LOC_Os02g53790) was located in the significant SNP region of the GWAS analysis. Further characterization of this gene would assist in elucidation of the mechanism regulating the early seedling length in rice under direct seeded rice.

Comparative Mitogenomics Provides Valuable Insights for the Phylogeny and New DNA Barcodes of Ganoderma

Ganoderma is the most important genus in the family Ganodermataceae; many species have attracted much attention and widely cultivated because of their medicinal values, but so far, not a sequenced mitogenome derived from dikaryon strains has been explicitly recorded. Herein, four novel mitogenomes of commonly cultivated Ganoderma (G. leucocontextum H4, G. lucidum G6, G. sinense MZ96 and G. tsugae SS) were de novo assembled and given detail functional annotations. Collinearity analysis revealed that the four mitogenomes shared 82.93–92.02% similarity with their corresponding reference mitogenomes at the nucleotide level. A total of 15 core protein-coding genes (PCGs), along with rrnL and rrnS (mtLSU and mtSSU) were chosen as potential candidates for constructing their individual phylogenetic trees. These trees were compared with those derived from the concatenated sequences of 15 core PCGs. And finally, we found that the atp9 and nad4L were the most reliable markers for the phylogenetic analysis of Ganoderma and chosen as standard sequences to generate new DNA barcodes. This finding was further verified by comparing it against almost all available Ganoderma mitogenomes in the NCBI, with Trametes versicolor (Polyporaceae) and Rigidoporus microporus (Meripilaceae) as two outgroups. A total of 52 mitogenomes from three families were highly conserved, with identical gene lengths for atp9 (222 bp) and nad4L (267 bp). These genes were capable of distinguish distinctly different various species, which are grouped into separate clades within the phylogenetic trees. The closest related clades (I and II), including at least 30 samples of the three classical taxonomic species (G. lingzhi, G. sichuanense and G. lucidum), differed in only one SNP. The single base mutation rate increased with the evolutionary divergence of the phylogenetic clades, from two to three SNPs in earlier clades (e.g., clade IV containing G. leucocontextum) to five to six SNPs in later clades (e.g., clade X containing G. sinense). Despite these variations between species, the atp9 and nad4L genes of Ganoderma mitogenomes consistently encoded the same ATP synthase F0 subunit c (73 aa) and NADH dehydrogenase subunit 4L (88 aa). These two genes have been identified as reliable markers of new DNA barcodes, offering valuable insights and contributing significantly to understanding the evolutionary relationships and phylogeny of the Ganoderma genus and even the Ganodermataceae family.

Amphisphaeria hongheensis sp. nov. (Amphisphaeriaceae, Amphisphaeriales) from Yunnan Province, China

Yunnan Province encompasses the highest fungal diversity in China, and novel fungal species are regularly introduced. Several saprobic fungi were collected during surveys of fungi associated with woody plants in Yunnan Province, Southwestern China. The fungi samples were initially identified as Amphisphaeria. Multi-gene phylogenetic analysis based on combined datasets of LSU, ITS, rpb2, and tub2 regions and morphological characteristics confirmed that the samples represent a previously unidentified fungus belonging to Amphisphaeria, which is named A. hongheensis sp. nov. This paper provides a full description, illustrations, and a phylogenetic tree showing the new species' taxonomic placement.

Updated Taxonomy of Chinese Phlomoides: description of three newly recorded species and their systematic placement

Phlomoides iliensis, P. dshungarica and P. rotala, previously known only from Kazakhstan, are newly recorded in Xinjiang Uygur Autonomous Region, China. Molecular phylogenetic analyses using eight plastid DNA markers (atpB-rbcL, psbA-trnH, rpl16, rpl32-trnL, rps16, trnK, trnL-trnF, trnT-trnL) revealed that these three species are nested within Clade VI, with their relationships to similar or allied species discussed. This study increases the total number of Phlomoides species in China to 64. In addition, diagnostic characters, updated morphological descriptions, detailed illustrations, and a key that encompasses all 16 Phlomoides species found in the Xinjiang Uygur Autonomous Region are provided.

Genomic DNA extraction methods and phylogenetic analysis of Beauveria bassiana from Central Java, Indonesia, and its toxicity against the fall armyworm, Spodoptera frugiperda J.E. Smith (Lepidoptera: Noctuidae)

BackgroundControl techniques using biological control agents such as Beauveria bassiana (Balsamo) Vuillemin have the advantage of not showing any negative impacts on environmental health and safety issues. This study used isolates from B. bassiana collection from Laboratory of Pest Monitoring belonging to Indonesian Ministry of Agriculture (LPHP) in Central Java which showed potential in controlling target pest. The problems that still occur are the lack of facilities and infrastructure and the lack of quality testing of collection isolates in LPHP, so that the isolate identification process is still carried out in simple method, and bioassay testing on the fall armyworm (FAW), Spodoptera frugiperda J.E. Smith (Lepidoptera: Noctuidae) as a target pest, is not commonly conducted. The results of bioassay testing can be used to determine the potential of a biological agent to control target pest.ResultThe two DNA extraction methods showed different results regarding DNA concentration and purity values, but both methods were good and could be used to amplify DNA using PCR. The DNA band was amplified at 500–600 bp using primers ITS 1 and ITS 4. The results of molecular analysis showed that the four isolates of B. bassiana from Central Java were found in the same clade as B. bassiana from South Sumatra, Dhaka, and Oromia, where these isolates showed similar similarities descended from a common ancestor. Genetically, B. bassiana isolates from Central Java show more genetic similarities to B. bassiana isolates from South Sumatra, Indonesia. Quality testing was carried out by calculating the density and germination ability values for LPHP isolates from Sukoharjo (Sukoharjo isolate), Temanggung (Purworejo isolate), and Banyumas (Banyumas and Cilacap isolate), which showed varying results. The bioassay test used three isolates, namely B. bassiana from Sukoharjo, Banyumas, and Cilacap, which were selected based on density values, germination ability, and molecular analysis. The ability to cause death of the three isolates against S. frugiperda showed different results where the isolate from Sukoharjo, Banyumas, and Cilacap caused mortality of 60, 40, and 60%, and the LC50 value of each isolate was 3.3 × 106, 1.3 × 107, and 3.5 × 107 conidia ml−1, respectively.ConclusionMorphological identification by macroscopic, microscopic, and molecular analysis showed that the isolate from the LPHP collection in Central Java, Indonesia, was B. bassiana. Genetically, the four isolates showed similar characteristics to isolates from South Sumatra, Indonesia. B. bassiana isolates from collections from Central Java showed potentials as a biological control agent against S. frugiperda.

Characterization of two Friunavirus phages and their inhibitory effects on biofilms of extremely drug resistant Acinetobacter baumannii in Dakar, Senegal

BackgroundAcinetobacter baumannii is a gram-negative, opportunistic pathogen, that is responsible for a wide variety of infections and is a significant cause of hospital-acquired infections. A. baumannii is listed by the World Health Organization (WHO) as a critical priority pathogen because of its high level of antibiotic resistance and the urgent need for alternative treatment solutions. To address this challenge, bacteriophages have been used to combat bacterial infections for more than a century, and phage research has regained interest in recent years due to antimicrobial resistance (AMR). However, although the vast majority of deaths from the AMR crisis will occur in developing countries in Africa and Asia, few phages’ studies have been conducted in these regions. In this study, we present a comprehensive characterization of the bacteriophages vAbBal23 and vAbAbd25, actives against extremely drug-resistant (XDR) A. baumannii.MethodsPhages were isolated from environmental wastewaters in Dakar, Senegal. The host-range, thermal and pH stabilities, infection kinetics, one step growth assay, antibiofilm activity assay, sequencing, and genomic analysis, were performed to characterize the isolated phages.ResultsComparative genomic and phylogenetic analyses revealed that vAbBal23 and vAbAbd25 belong to the Caudoviricetes class, Autographiviridae family and Friunavirus genus. Both phages demonstrated activity against strains with capsular type KL230. They were stable over a wide pH range (pH 3 to 9) and at temperatures ranging from 25 °C to 40 °C. Additionally, the phages exhibited notable activity against both planktonic and biofilm cells of targeted extremely drug resistant A. baumannii. The results presented here indicate the lytic nature of vAbBal23 and vAbAbd25. This is further supported by the absence of genes encoding toxins, resistance genes and bacterial virulence factors, highlighting their potential for future phage applications.ConclusionPhages vAbBal23 and vAbAbd25 are promising biological agents that can infect A. baumannii, making them suitable candidates for use in phage therapies.

Genome-Based Classification of 'Haloarcula aidinensis' and Description of Three Novel Halophilic Archaea Isolated from an Inland Saline Lake, Coastal Saline Soil, and a Marine Solar Saltern.

'Haloarcula aidinensis' was described by a pioneer Chinese scientist focused on halophilic archaea thirty years ago, and the type strain of 'Haloarcula aidinensis' was recently classified based on phylogenetic, phylogenomic, and comparative genomic analyses. Other three novel halophilic archaeal strains, CK38T, DT43T, and SYNS111T, isolated from diverse saline environments in China, were simultaneously subjected to polyphasic classification. 'Haloarcula aidinensis' A5 was found to be related to Haloarcula amylolytica, while strains CK38T, DT43T, and SYNS111T represented three novel species of Haloarcula based on phylogenetic, phylogenomic, and comparative genomic analyses. These strains can be distinguished from other species within the genus Haloarcula based on multiple phenotypic characteristics. The major phospholipids, phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, and phosphatidylglycerol sulfate can be detected in these strains, while the glycolipid profiles of these strains are diverse. Strains A5 and DT43T contained glucosyl mannosyl glucosyl diether and a diglycosyl diether, while other strains CK38T and SYNS111T had sulfated mannosyl glucosyl diether and mannosyl glucosyl diether. Thus, strain A5 should be a reference strain of Har. amylolytica and three novel species of Haloarcula, Haloarcula sediminis sp. nov., Haloarcula brevis sp. nov., and Haloarcula regularis sp. nov. are proposed to accommodate strains CK38T (= CGMCC 1.62732T = JCM 36675T), DT43T (= CGMCC 1.18924T = JCM 36146T), and SYNS111T (= CGMCC 1.62601T = JCM 36149T).

Whole-Genome Sequencing Reveals the Population Structure and Genetic Diversity of Salmonella Typhimurium ST34 and ST19 Lineages.

Salmonella Typhimurium is an invasive gastrointestinal pathogen for both humans and animals. To investigate the genetic framework and diversity of S. Typhimurium, a total of 194 S. Typhimurium isolates were collected from patients in a tertiary hospital between 2020 and 2021. Antimicrobial susceptibility testing was used to confirm the resistance phenotype. Whole-genome sequencing and bioinformatics analysis were performed to determine the sequence type, phylogenetic relationships, resistance gene profiles, Salmonella pathogenicity island (SPI) and the diversity of the core and pan genome. The result showed that 57.22% of S. Typhimurium isolates were multidrug resistant and resistance of total isolates to the first-line drug ciprofloxacin was identified in 60.82%. The population structure of S. Typhimurium was categorized into three lineages: ST19 (20.10%, 39/194), ST34-1 (47.42%, 92/194) and ST34-2 (40.65%, 63/194), with the population size exhibiting increasing trends. All lineages harbored variety of fimbrial operons, prophages, SPIs and effectors that contributed to the virulence and long-term infections of S. Typhimurium. Importantly, ST34-1 lineage might potentially be more invasive due to the possession of SPI1-effector gene sopE which was essential for the proliferation, internalization and intracellular presence of S. Typhimurium in hosts. Multiple antimicrobial resistance genes were characteristically distributed across three lineages, especially carbapenem genes only detected in ST34-1&2 lineages. The distinct functional categories of pan genome among three lineages were observed in metabolism, signaling and gene information processing. This study provides a theoretical foundation for the evolved adaptation and genetic diversity of S. Typhimurium ST19 and ST34, among which ST34 lineages with multidrug resistance and potential hypervirulence need to pay more attention to epidemiological surveillance.

Genomic Characterization and Comparative Analysis of Streptococcus zhangguiae sp. nov. Isolated from the Respiratory Tract of Marmota Himalayana.

Two Gram-stain-positive, oxidase-negative, non-motile, facultative anaerobic, α-hemolytic, coccus-shaped bacteria (zg-86T and zg-70) were isolated from the respiratory tracts of marmots (Marmota Himalayana) on the Qinghai-Tibet Plateau of China. Phylogenetic analysis of the 16S rRNA gene and 545 core genes revealed that these two strains belong to the Streptococcus genus. These strains were most closely related to Streptococcus respiraculi HTS25T, Streptococcus cuniculi CCUG 65085T, and Streptococcus marmotae HTS5T. The average nucleotide identity (ANI) and digital DNA‒DNA hybridization (dDDH) were below the threshold for species delineation. The predominant cellular fatty acids (CFAs) in this novel species were C16:0, C18:0, and C18:1ω9c, whereas the primary polar lipids were phosphatidylglycerol (PG), phosphatidylethanolamine (PE) and an unknown phosphoglycolipid (PGL). The optimal growth conditions for the strains were 37°C, pH 7.0, and 0.5% (w/v) NaCl on brain-heart infusion (BHI) agar supplemented with 5% defibrinated sheep blood. Comparative genomics analyses revealed the potential pathogenicity of strain zg-86T through comparisons with suis subclade strains in terms of virulence factors, pathogen-host interactions (PHIs) and mobile genetic factors (MGEs). Based on the phenotypic characteristics and phylogenetic analyses, we propose that these two isolates represent novel species in the genus Streptococcus, for which the names Streptococcus zhangguiae sp. nov. (the type strain zg-86T=GDMCC 1.1758T=JCM 34273T) is proposed.

Unveiling the dynamics of hepatitisC virus transmission among injection drug users and men who have sex with men: A comprehensive study in Japan.

In Japan, despite low nationwide hepatitisC (HCV) incidence, new infections among people who inject drugs (PWID) and men who have sex with men (MSM) hinder HCV elimination. We explored HCV transmission dynamics and screened HCV recombination within these populations. This cross-sectional study recruited HCV-infected patients from Osaka National Hospital, Osaka, Japan, from January 2010 to September 2023. Data from questionnaires and medical records were analyzed. Serum samples collected before anti-HCV treatment underwent HCV RNA extraction, and sequencing of full core (576bp) and NS5B (267bp) regions using the Sanger method. Genotype distribution was determined by phylogenetic analysis, and recombinant screening was conducted. A total of 115 patients were categorized into non-MSM PWID (31), MSM PWID (15), MSM non-PWID (25), and non-MSM non-PWID (44). Positive amplification rates were 99.1% (114/115) for the full-core region, and 96.5% (111/115) for NS5B. No intergenotypic recombination was detected. The predominant genotype in non-MSM PWID was 2a (58%), whereas genotype 1b was most common in MSM PWID, MSM non-PWID, and non-MSM non-PWID groups (79%, 64%, and 68%, respectively). Nucleotide sequence similarity of 94.75%-100% was found in HCV strains from MSM PWID and MSM non-PWID in both full-core and NS5B regions, whereas strains from non-MSM PWID and non-MSM non-PWID were distinct. The findings suggest that the transmission route in PWID is determined by MSM status, whereas MSM groups showed the same transmission route regardless of PWID. HCV control measures should be focused not only on PWID, but also on MSM to achieve HCV elimination in Japan.

Enterobacter-infecting phages in nitrogen-deficient paddy soil impact nitrogen-fixation capacity and rice growth by shaping the soil microbiome

Bacteriophages (“phage”) play important roles in nutrient cycling and ecology in environments by regulating soil microbial community structure. Here, metagenomic sequencing showed that a low relative abundance of nitrogen-fixing bacteria but high abundance of Enterobacter-infecting phages in paddy soil where rice plants showed nitrogen deficiency. From soil in the same field, we also isolated and identified a novel virulent phage (named here as Apdecimavirus NJ2) that infects several species of Enterobacter and characterized its impact on nitrogen fixation in the soil and in plants. It has the morphology of the Autographiviridae family, with a dsDNA genome of 39,605 bp, 47 predicted open reading frames and 52.64 % GC content. Based on genomic characteristics, comparative genomics and phylogenetic analysis, Apdecimavirus NJ2 should be a novel species in the genus Apdecimavirus, subfamily Studiervirinae. After natural or sterilized field soil was potted and inoculated with the phage, soil nitrogen-fixation capacity and rice growth were impaired, the abundance of Enterobacter decreased, along with the bacterial community composition and biodiversity changed compared with that of the unadded control paddy soil. Our work provides strong evidence that phages can affect the soil nitrogen cycle by changing the bacterial community. Controlling phages in the soil could be a useful strategy for improving soil nitrogen fixation.

Genome-wide analysis of the PYL gene family and identification of PYL genes that respond to cold stress in Triticum monococcum L. Subsp. Aegilopoides

Abscisic acid (ABA) is a key plant hormone that regulates plant growth and response to stress . Pyrabactin resistance 1-like (PYR/PYL) proteins are ABA receptors involved in the initial steps of ABA signaling. Triticum monococcum L. subsp. aegilopoides is an important germplasm resource for wheat. In this study, we identified 15 PYL genes from T. monococcum L. subsp. aegilopoides and found that they were distributed across five chromosomes. Based on phylogenetic analysis, we classified these genes into three subfamilies. Members of each subfamily have similar gene structures and contain a common motif. Further analysis revealed that the promoters have multiple hormone-related elements. We found 7, 33, and 49 collinear gene pairs in three different ploidy wheat species (T. urartu, T. turgidum and T. aestivum), indicating that PYL genes are relatively conserved during the process of wheat polyploidization. Additionally, interaction networks and miRNA targets were predicted, revealing interactions between PYL proteins and key components of the abscisic acid signaling network. miR9666b-3p may serve as a central factor in PYL involvement in the abscisic acid network. Through RNA-seq analysis and qPCR validation, three genes (TbPYL2, TbPYL5, and TbPYL12) were found to potentially play a role in cold stress. These findings lay the groundwork for further research on PYL genes in T. monococcum L. subsp. aegilopoides.

Transcriptomic Analysis of Cell Stress Response in Wickerhamomyces anomalus H4 Under Octanoic Acid Stress

The purified yeast strain H4, identified as W. anomalus through morphological, genetic, and phylogenetic analyses, was characterized and compared to a commercial Saccharomyces cerevisiae strain X16. W. anomalus H4 exhibited distinct morphological features. It demonstrated notable tolerance to 11% ethanol, 220 g/L glucose, and 200 mg/L octanoic acid, similar to X16, except for having a lower tolerance to SO2. Survival analysis under various stress conditions revealed that ethanol and octanoic acid had the most detrimental effects, with 56% cell mortality at 13% ethanol and 400 mg/L octanoic acid. Transcriptomic analysis under octanoic acid stress showed that at 200 mg/L, 3369 differentially expressed genes (DEGs) were induced, with 1609 being upregulated and 1760 downregulated, indicating broad transcriptional reprogramming. At 400 mg/L, only 130 DEGs were detected, suggesting a more limited response. KEGG pathway analysis indicated that most DEGs at 200 mg/L were associated with the “ribosome” and “proteasome” pathways, reflecting disruptions in protein synthesis and turnover. At 400 mg/L, the DEGs were primarily related to “DNA replication” and “pyruvate metabolism”. These findings highlight the adaptive mechanisms of W. anomalus H4 to environmental stresses, particularly octanoic acid, and its potential for use in brewing and fermentation processes.

Identification and characterization of a novel Pavlovskyella (Acari: Argasidae) from Chile, parasite of the culpeo fox (Lycalopex culpaeus).

Soft ticks (Argasidae) of the subgenus Pavlovskyella Pospelova-Shtrom are worldwide distributed parasites of medical importance. However, the systematics of the subgenus are currently under debate because genetic data shows that the group is paraphyletic. Meanwhile, species of Pavlovskyella continue to be discovered. In this study a novel species of the subgenus is described from specimens collected on a fox in central Chile. The larva of this new species differentiates from other Pavlovskyella spp. by having the following combination of characters: subpyriform dorsal plate; 15 pairs of setae, 7 anterolateral, 3 central, and 5 posterolateral, and hypostome with denticles in the distal third. Nymphs and adults of the species lack cheeks, eyes or bulging structures on the flank, but exhibit dorsoventral grooves, and humps on tarsi I, II, and III. Moreover, a patch of glabrous integument appears on the distal portion of coxal folds. A phylogenetic analysis using the mitogenome indicates a monophyletic group composed by Ornithodoros (Pavlovskyella) brasiliensis Aragão, Ornithodoros (Pavlovskyella) furcosus Neumann, Ornithodoros (Pavlovskyella) improvisus Muñoz-Leal & Venzal, and Ornithodoros (Pavlovskyella) rostratus Aragão. Interestingly, a phylogeny using 18S-28S rDNA sequences shows that South American Pavlovskyella spp. are paraphyletic, as depicted in previous studies. Adding species of the subgenus from the Neotropical region to phylogenetic analyses could aid to solve this paraphyly. Furthermore, this is the fifth species of Pavlovskyella described in South America, and the second in Chile.

A new species of Leptobrachella Smith 1925 (Anura, Megophryidae) from Lai Chau Province, Vietnam

The genus Leptobrachella (Anura, Megophryidae) was originally described, based on the type species from Sarawak (Malaysia), Leptobrachella mjöbergi Smith. The taxa in the group were previously classified into different genera, i.e, Paramegophrys Liu; Leptolalax Dubois; Lalax Delorme, Dubois, Grosjean & Ohler; and Lalos Dubois, Grosjean, Ohler, Adler & Zhao. However, Yuan et al. synonymised Leptolalax with Leptobrachella in 2017. Members of Leptobrachella inhabit the forest floor and rocky streams in hilly evergreen forests. They are widely distributed from southern China and Myanmar through mainland Indochina to Peninsular Malaysia and the island of Borneo. However, the species diversity of the genus was indicated to be underestimated by phylogenetic analyses and a series of new species have been discovered recently. In Vietnam, 34 species of Leptobrachella are currently known and 75% (or 24 species) have been described or newly recorded from the country since 2010. We describe a new species, Leptobrachella huynhi sp. nov., from Sin Ho District, Lai Chau Province. The new species is distinguished from its congeners by genetic divergences ranging from 3.62 to 18.51% (16S rRNA gene) and morphological differences: size medium (SVL 37.8–40.2 mm in adult females); head longer than wide; tympanum distinct; skin on entire dorsum shagreened; toes without webbing and with narrow lateral fringes; supratympanic ridge slightly rough with few nodules; dorsum grey-brown with indistinct dark brown markings; an interorbital region with a stacking double Y-shaped marking; centre of belly creamy-white, outer edges of belly brown with small whitish spots; iris copper. The new species is the 35th species of the genus Leptobrachella known from Vietnam.

Molecular analysis of vector-borne pathogens in Eurasian badgers (Meles meles) from continental Europe

BackgroundVector-borne pathogens (VBPs) are increasing in significance in veterinary medicine and public health settings, with wildlife playing a potentially crucial role in their transmission. Eurasian badgers (Meles meles) are widely distributed across Europe. However, information currently available on the prevalence of VBPs in badgers is limited. The objective of the current study was to investigate the occurrence of Anaplasmataceae, Bartonella spp., Mycoplasma spp., Rickettsia spp., Piroplasmida, Trypanosomatida and Filarioidea in badgers and subsequently, based on the results, assess the potential risk to domestic animals, other wildlife and humans.MethodsBetween 2017 and 2021, blood or spleen samples from 220 badgers were collected in nine continental European countries: Austria (n = 7), Bosnia and Herzegovina (n = 2), Croatia (n = 22), France (n = 44), Germany (n = 16), Hungary (n = 7), Italy (n = 16), Romania (n = 80) and Serbia (n = 26). VBPs were identified by performing PCR analysis on the samples, followed by Sanger sequencing. Additionally, to distinguish between different Babesia lineages we performed restriction fragment length polymorphism (RFLP) analysis on piroplasm-positive samples, using HinfI as restriction enzyme. A phylogenetic analysis was performed on Mycoplasma spp.ResultsThe pathogens identified were Babesia sp. badger type A (54%), B (23%), and C (37%); Trypanosoma pestanai (56%); Mycoplasma sp. (34%); Candidatus Mycoplasma haematomelis (8%); Candidatus Mycoplasma haematominutum (0.5%); and Ehrlichia spp. (2%). Rickettsia spp., Bartonella spp. and filarioid nematodes were not detected among the tested samples.ConclusionsThe large sample size and diverse study populations in this study provide valuable insights into the distribution and epidemiology of the analyzed pathogens. Some of the VBPs identified in our study show high similarity to those found in domestic animals, such as dogs. This finding suggests that badgers, as potential reservoirs for these pathogens, may pose a threat not only to other wildlife but also to domestic animals in close vicinity. Continuous surveillance is essential to monitor VBPs in wildlife as a means to enable the assessment of their impact on other wildlife species, domestic animals and human health.Graphical

Morphological, Biochemical and Molecular Characterization of Plant Growth Promoting Rhizobacteria with Synergistic Effect against Fusarium oxysporum

Currently, plant growth-promoting rhizosphere bacteria (PGPR) are heavily exploited as microbial inoculants in agricultural production among which Pseudomonas sp. and Bacillus sp. are widely used in plant growth promotion and disease control as excellent inoculum strains. Plant growth-promoting rhizobacteria has the potential to be used in agriculture to promote plant growth and health through various mechanisms. Therefore, identifying novel strains tailored to specific agricultural requirements continues to be a key area of research. In the present research work, the strains of PGPR isolated from soybean rhizosphere at Department of Plant Pathology and characterized at morphological, biochemical and molecular level. These strains are now used in ‘Biomix’ formulation a microbial consortium product developed at the Department of Plant Pathology VNMKV, Parbhani to benefit the farmers. Studying the molecular, biochemical, and morphological traits of these microbial strains which are now involved in VNMKV produced ‘Biomix formulation is crucial to maintain its consistency, purity and optimize its formulation. Molecular characterization and phylogenetic analysis will ensure accurate identification and classification of the isolates, enhancing the biofertilizer's effectiveness in agriculture. All three isolates have been identified as per Bergey’s Manual for the determination of bacteriology. All isolates tested positive for starch hydrolysis, citrate utilization, catalase activity, indole production, gelatine hydrolysis, and tetrazolium reduction. Additionally, the 16S rDNA gene sequences of the various potential isolates were validated as P. fluorescens VNMKV1, P. putida VNMKV1, and B. subtilis VNMKV1. The potential of these novel strains of plant growth-promoting rhizobacteria as biocontrol agents was explored by screening against soil-borne pathogenic fungi highlighting their potential to be used as bioinoculant agents.

Genomic Insights into Pig Domestication and Adaptation: An Integrated Approach Using Genome-Wide Selection Analysis and Multiple Public Datasets

As a global focus of animal husbandry, pigs provide essential meat resources for humans. Therefore, analyzing the genetic basis of adaptability, domestication, and artificial selection in pigs will contribute to further breeding. This study performed a genome-wide selection sweep analysis to identify candidate genes related to domestication and adaptive selection via data from 2413 public genotypes. Two complementary statistical analyses, FST (fixation index) and XP–EHH (cross–population extended haplotype homozygosity) were applied. The results revealed that numerous candidate genes were associated with high-altitude adaptability (e.g., SIRPA, FRS2, and GRIN2B) and habitat temperature adaptability (e.g., MITF, PI3KC2A, and FRS2). In addition, candidate genes related to the domestic genetic imprint of indigenous pigs (e.g., TNR, NOCT, and SPATA5) and strong artificial selection pressure in commercial breeds (e.g., ITPR2, HSD17B12, and UGP2) were identified in this study. Specifically, some MHC–related genes (e.g., ZRTB12, TRIM26, and C7H6orf15) were also under selection during domestication and artificial selection. Additionally, a phylogenetic comparative analysis revealed that the genetic divergence between populations does not fully follow the geographical distribution and management history in the major histocompatibility complex region/major histocompatibility complex II haplotypes, unlike that of the genome-wide genotypes. Furthermore, the higher heterozygosity and haplotype alleles of MHC reduce the differences between populations. Briefly, this study not only helps promote the relative theoretical understanding of environmental adaptive selection and domestication but also provides a theoretical reference for disease-resistant breeding in pigs.

Specialization restricts the evolutionary paths available to yeast sugar transporters.

Functional innovation at the protein level is a key source of evolutionary novelties. The constraints on functional innovations are likely to be highly specific in different proteins, which are shaped by their unique histories and the extent of global epistasis that arises from their structures and biochemistries. These contextual nuances in the sequence-function relationship have implications both for a basic understanding of the evolutionary process and for engineering proteins with desirable properties. Here, we have investigated the molecular basis of novel function in a model member of an ancient, conserved, and biotechnologically relevant protein family. These Major Facilitator Superfamily sugar porters are a functionally diverse group of proteins that are thought to be highly plastic and evolvable. By dissecting a recent evolutionary innovation in an α-glucoside transporter from the yeast Saccharomyces eubayanus, we show that the ability to transport a novel substrate requires high-order interactions between many protein regions and numerous specific residues proximal to the transport channel. To reconcile the functional diversity of this family with the constrained evolution of this model protein, we generated new, state-of-the-art genome annotations for 332 Saccharomycotina yeast species spanning approximately 400 million years of evolution. By integrating phylogenetic and phenotypic analyses across these species, we show that the model yeast α-glucoside transporters likely evolved from a multifunctional ancestor and became subfunctionalized. The accumulation of additive and epistatic substitutions likely entrenched this subfunction, which made the simultaneous acquisition of multiple interacting substitutions the only reasonably accessible path to novelty.