16S rDNA Research Articles

A new species of Diacheopsis (Myxomycetes) and a new habitat for myxomycetes

ABSTRACT We describe a new species, Diacheopsis resinae (Myxomycetes), collected from a microhabitat new for myxomycetes: stem wounds of coniferous trees (Norway spruce) where the resin is overgrown with a community of resinicolous fungi. The 80 known collections come from the Vosges (France), the Black Forest (Germany), Swabian Alp (Germany), and several localities in Denmark and Norway. Observations, but as well as metabarcoding of substrate samples with fungal (ITS [internal transcribed spacer]), bacterial (16S rDNA), and myxomycete (18S nuc rDNA) primers from eight trunks, revealed the new myxomycete to co-occur with resin-degrading ascomycetes (Infundichalara microchona, Lophium arboricola, Zythia resinae). The gram-negative bacterial genera Endobacter and Sphingomonas were found to be abundant in the substrate and may be a food source for the myxomycete. Fruit bodies were found mostly during the more humid winter season, with a peak in January/February. Partial sequences of two independent molecular markers (18S nuc rDNA, EF1α [elongation factor 1-alpha] gene) were obtained for 41 accessions, which form a monophyletic cluster in a two-gene phylogeny of Stemonititidales but do not group with other species of Diacheopsis, thus rendering this genus paraphyletic. The new species, although exclusively developing sessile sporocarps and morphologically undoubtedly falling into the genus Diacheopsis, is most closely related to species of Lamproderma, especially L. album, L. zonatum, and L. zonatopulchellum. Within D. resinae, three groups can be differentiated, which show nearly complete reproductive isolation, as judged from a recombination analysis of the two unlinked markers and the allelic combinations of the EF1α gene.

Morphological and molecular analysis of Propanagrolaimus siweyae sp. n. (Nematoda: Panagrolaimidae) from Molepo dam, Limpopo Province, South Africa, and its relationship with water parameters

AbstractPanagrolaimidae members are bacterivorous with global distribution. There is no information on this species' morphology and molecular characterization in South Africa. During a survey of Molepo dam, Limpopo Province, a population of Propanagrolaimus nematodes belonging to the family Panagrolaimidae was recovered and determined utilizing conventional and molecular characters. Propanagrolaimus siweyae sp. n., was confirmed by morphological and molecular informations. Propanagrolaimus siweyae sp. n. is characterized by a body length 1026–1258 µm (a = 39.4–45.5, b = 5.4–6.5, c = 8.1–10.0, c’ = 6.9–7.8, V = 53–57), post vulval uterine sac 25–53 µm, and tail length 108–158 µm long. The scan electron microscopy (SEM) photographs revealed a tessellated cuticle and six slightly separated lips. The discriminant analysis placed populations of P. siweyae sp. n. close to P. hygrophilus with clear separation from the mentioned species. Partial sequences of the 18S and 28S regions of the ribosomal DNA gene were amplified for P. siweyae sp. n. The phylogenetic analysis grouped P. siweyae sp. n. in a clade with 1.00 and 0.87 posterior probability values together with other Propanagrolaimus and Halicephalobus based on 18S and 28S rDNA, respectively. Principal component analysis revealed no correlation between Propanagrolaimus and water parameters. However, the result showed a high correlation with diatoms (r = 0.937) in Molepo dam (site MD-6). In contrast, The PCA showed salinity, pH, and temperature with no significant effect on nematodes in Molepo dam, Limpopo Province, South Africa. In conclusion, the usefulness of free-living nematodes as bioindicators is an advantage to studying pollution in aquatic systems. Limpopo Province is facing pollution in the river and dam systems. Therefore, finding a bioindicator is critical to examine the ecological role of free-living nematodes such as Propanagrolaimus.

Research on gut microbiota characteristics of PBC patients at different ALBI grades based on machine learning

BackgroundThe Albumin-Bilirubin (ALBI) score and grade are widely used to stratify patients with primary biliary cholangitis (PBC) into different disease statuses and risk levels. Recent studies have increasingly highlighted the role of gut microbiota in autoimmune liver diseases. This study aimed to investigate the differences in gut microbiota among PBC patients with varying ALBI grades.MethodsClinical data and stool samples were collected from outpatient and inpatient PBC patients between 2019 and 2022. Gut microbiota profiles were obtained using 16S rDNA sequencing of stool samples. We analyzed alpha diversity, beta diversity, LEfSe analysis and pathway function prediction. Additionally, various machine learning methods—including random forest (RF), lasso, gradient boosting machine (GBM) and support vector machine (SVM)—were employed to identify key features and to build and validate predictive models using bootstrap techniques.ResultsClinical characteristics of ALBI grade 1 patients were comparatively better than those of ALBI grade 2 and 3 patients, including multiple laboratory indices. Gut microbiota analysis revealed that species richness and balance were higher in ALBI grade 1 patients. Both the comparison of the most abundant genera and the linear discriminant analysis (LDA) in LEfSe demonstrated that Lachnospira had a higher abundance and better discriminative ability in ALBI grade 1. Pathway function prediction indicated that sulfur metabolism was upregulated in higher ALBI grades. Furthermore, RF identified 10 specific genera, which were then used to build and validate models for discriminating PBC patients according to their ALBI grades. All three models, developed using different machine learning methods, demonstrated good discrimination ability (mean AUC 0.75–0.80).ConclusionThis study highlights significant differences in gut microbiota profiles among PBC patients with different ALBI grades. The increased abundance of Lachnospira and upregulation of sulfur metabolism pathways are notable in patients with lower ALBI grades. The machine learning models developed based on gut microbiota features offer promising tools for discriminating between PBC patients with varying disease severities, which could enhance the precision of treatment strategies.

Isolation and assessment of probiotic potential of Acidipropionibacterium acidipropionici C03B-STR from goat milk with cholesterol-lowering capability.

Elevated levels of cholesterol or triglycerides in the blood are one of the major risk factors for coronary heart disease, which is the foremost leading cause of death across the world. The aims of this study were to isolate and verify the potential probiotic strain and cholesterol-lowering capability from goat milk. The C03B-STR isolate had a broad-spectrum antibacterial property and exhibited remarkable bacteriostatic activity against Klebsiella pneumoniae ATCC 13883 (27.00 ± 1.73mm), Pseudomonas aeruginosa ATCC 27853 (19.33 ± 0.57mm) and Staphylococcus aureus ATCC 10832 (19.00 ± 1.00mm) by agar well diffusion assay. This isolate is closely related to the 16S rDNA sequence of Acidipropionibacterium acidipropionici. Acidipropionibacterium acidipropionici C03B-STR can survive under acidic conditions (60.51 ± 0.02% of survival rate at pH 3.0 for 3h) and was as bile-tolerant strain (> 80% of survival rate at 0.30% bile salts after 3h of incubation). Furthermore, it showed significantly high cell surface hydrophobicity (74.36 ± 0.23%) and aggregation capability (> 83%) (p ≤ 0.05) but displayed low to moderate co-aggregation ability (44-61%). This strain can also be regarded as strongly adhesive (73.69 ± 0.74%) and inhibit pathogen attachment to the Caco-2 cell lines (39.20 ± 0.59-69.01 ± 0.29%). A. acidipropionici C03B-STR appeared to be safe (non-haemolytic) and controllable (sensitive to various antibiotics). Notably, it had the potential to suppress cholesterol uptake in Caco-2 cells (approximately 45%) and also reduced cholesterol level above 84% during the exponential phase under acidic conditions. Thus, A. acidipropionici C03B-STR from goat milk may be a promising novel potential cholesterol-lowering probiotic strain for application in functional foods and biotherapeutics.

Parasitic taxa are key to the vertical stratification and community variation of pelagic ciliates from the surface to the abyssopelagic zone.

An increase in upper-ocean thermal stratification is being observed worldwide due to global warming. However, how ocean stratification affects the vertical profile of plankton communities remains unclear. Understanding this is crucial for assessing the broader implications of ocean stratification. Pelagic ciliates cover multiple functional groups, and thus can serve as a model for studying the vertical distribution and functional strategies of plankton in stratified oceans. We hypothesize that pelagic ciliate communities exhibit vertical stratification caused by shifts in functional strategies, from free-living groups in the photic zone to parasitic groups in deeper waters. 306 samples from the surface to the abyssopelagic zone were collected from 31 stations in the western Pacific and analyzed with environmental DNA (the V4 region of 18S rDNA) metabarcoding of pelagic ciliates. We found a distinct vertical stratification of the entire ciliate communities, with a boundary at a depth of 200m. Significant distance-decay patterns were found in the photic layers of 5m to the deep chlorophyll maximum and in the 2,000m, 3000m and bottom layers, while no significant pattern occurred in the mesopelagic layers of 200m - 1,000m. Below 200m, parasitic Oligohymenophorea and Colpodea became more prevalent. A linear model showed that parasitic taxa were the main groups causing community variation along the water column. With increasing depth below 200m, the ASV and sequence proportions of parasitic taxa increased. Statistical analyses indicated that water temperature shaped the photic communities, while parasitic taxa had a significant influence on the aphotic communities below 200m. This study provides new insights into oceanic vertical distribution, connectivity and stratification from a biological perspective. The observed shift of functional strategies from free-living to parasitic groups at a 200m transition layer improves our understanding of ocean ecosystems in the context of global warming.

Morphological and molecular characterization of the elusive vegetative stage of the diatom Cerataulina bicornis: a new record from Indian Sundarbans

Abstract The resting spore stage of Cerataulina bicornis has been reported sporadically from the Indian Sundarbans, with one sediment core record that dates back to ∼2000 yrs. BP, but its vegetative stage has remained elusive during this entire period. The present report of its vegetative stage is a new record from this estuarine system. A comprehensive morphometric description along with molecular data of rbcL and 18s rDNA of the vegetative stage of C. bicornis is presented. The morphological and morphometric characters observed in the light microscope (LM) and scanning electron microscope (SEM) studies, along with a detailed description of the copulae, establish its distinction from other Cerataulina spp. described from different parts of the world.

Isolation, identification, characterization and in vitro assay of saline tolerant endophytes against groundnut root rot caused by Rhizoctonia bataticola (Taub.) Butler

Groundnut, known as Arachis hypogaea L., is India's significant oil seed crop. Dry root rot, caused by Rhizoctonia bataticola, poses a substantial challenge to cultivating groundnuts. During the roving survey, 60.50% dry root rot disease incidence was recorded in Namakkal district, Tamil Nadu. This study aims to acquire salt-tolerant endophytic bacteria residing in groundnuts with significant antagonistic activity against R. bataticola. A total of 27 bacterial strains were isolated from groundnuts. Among these strains, RMV 3 and RMV 2 are the most effective isolates, exhibiting 60.1% and 50% inhibition zones, respectively. The effective isolates were characterized through morphological, biochemical and phytostimulation activities and 16S rDNA sequencing. Among the isolates, RMV 3 and RMV 2 showed positive results for siderophore, indole acetic acid (IAA) and cellulase test. The strain RMV 3 was identified as Bacillus subtilis through 16S rDNA sequencing. GC-MS analysis identified twenty bioactive compounds produced by B. subtilis RMV 3, such as pyrrolo [12-a] pyrazine-14-dione hexahydro-3 (2-methylpropyl) and hexadecanoic acid methyl ester. The crude metabolite assay demonstrated a 96.6% inhibition of R. bataticola by RMV 3. This study demonstrated that Bacillus subtilis RMV 3, which exhibits a robust antagonistic effect on R. bataticola, can potentially be an effective biocontrol agent for groundnut dry root rot.

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.

Description and phylogenetic position of an unusual genus and species of allocreadiid trematode from a characid freshwater fish in South America

The worldwide distributed family Allocreadiidae Looss, 1902 is predominantly parasitic in freshwater fish and occasionally in amphibians and reptiles. It contains 15 genera, of which only Allocreadium, Creptotrema, Wallinia and Creptotrematina have been reported in Argentina. The records of Allocreadium species still require further verification. The discovery of an allocreadiid digenean parasitizing Pseudocorynopoma doriae led us to erect a new genus for which the name Martorellia n. gen is proposed. It is characterized by poorly developed ventrolateral muscular lobes and a transversely elongated ovary, with the most characteristic feature being the presence of vitelline follicles in two fields (one anterior at level of ventral sucker and one posterior to the testis), which are not confluent in the middle. The phylogenetic tree reconstructed with 28S rDNA sequences placed the new genus as the sister taxa of Creptotrematina. Although both genera are separated by a small genetic distance, it is comparable with those separating other allocreadiid genera. The discovery of Martorellia liciae gen et sp. nov. provides further evidence of the great diversity of Allocreadiidae in South America. The analysis of new samples from different South American hosts with an integrative taxonomic approach as the one used here allows a better understanding of the evolutionary history and biogeography of allocreadiids in the Americas.

Taxonomic revision of the genus Sarcosphaera (Ascomycota: Pezizales) in Europe and North America revealed unexpected diversity

Sarcosphaera (Ascomycota: Pezizales) is a genus of rare spring fungi, currently encompassing a single species, S. coronaria, which is red-listed in many European countries. The genus is characterized by large bright-colored fruit-bodies and high arsenic content. This study investigates the genetic diversity across Europe and North America. Using integrated molecular techniques, including ITS and 28S rDNA sequencing, we aimed to resolve the taxonomic ambiguities and assess their arsenic hyperaccumulation capabilities, a rare trait among fungi. We identified the presence of two species in the Western Palearctic (S. coronaria, S. crassa) and at least 12 additional lineages in North America, indicating a more complex taxonomic structure than previously recognized. To consolidate the taxonomy and nomenclature of these fungi, we selected reference specimens for S. coronaria and S. crassa and discussed their morphological and ecological differences. Two genera, Caulocarpa (C. montana) and Hydnotryopsis (H. setchellii), a genus comprising hypogeous fungi, are newly placed in the genus Sarcosphaera. Sarcosphaera is thus another genus where the rapid evolution of hypogeous forms from their above-ground ancestors took place. Furthermore, two new Sarcosphaera species, S. columbiana and S. pacifica, are described from the Pacific Northwest. Investigation of arsenic content of Sarcosphaera species revealed that arsenic mass fractions in ascocarps vary significantly between species and even within species across different geographical locations. Our findings have important implications for the taxonomy and conservation of the S. coronaria species complex and also point to future directions for further research in the taxonomy, geomycology, and toxicology of this group of fungi.

Nuclear ribosomal transcription units in Asian Paragonimus species (Paragonimidae: Platyhelminthes): genetic characteristics, polymorphism, and implications for intersuperfamilial phylogeny.

The entire transcribed sequences (from the 5' terminus of 18S to the 3' terminus of 28S rRNA genes) of the ribosomal transcription units (rTU*) of five Asian Paragonimus species were obtained and characterized. The rTU* length was 7661bp for P. heterotremus (LC strain, Vietnam), 7422bp for P. iloktsuenensis (Amami strain, Japan), 6932bp for P. skrjabini miyazakii (OkuST1 strain, Japan), 7422bp for P. ohirai (Kino strain, Japan), and three strains of P. westermani: 8616bp (Megha strain, India), 7292bp (Bogil strain, South Korea), and 7052bp (QT2 strain, Vietnam) without intergenic spacer region (IGS). All seven Asian Paragonimus strains' genetic characteristics were described, including the length of individual genes/regions, repeat polymorphism, base composition, and skewness. To investigate the superfamilial relationships in the Xiphidiata, with a focus on the Troglotrematoidea and its associated superfamilies, we used the PhyML software package to create three comprehensive maximum-likelihood phylogenies. The datasets used were 83 concatenated 28S + 18S, 83 single complete 18S, and 157 single, partial 28S rDNA sequences, respectively, from entire rTUs and/or accessible ribosomal sequences of the same species from the suborders Xiphidiata, Echinostomata, and Haplosplanchnata, with a Schistosoma sequence as an outgroup. Three phylogenetic trees revealed that Echinostomata and Haplosplanchnata are monophyletic, while Xiphidiata is polyphyletic and contains the monophyletic Troglotrematoidea. The concatenated 18S + 28S and single 18S phylogenies revealed well-bootstrap supported seven superfamilies (Troglotrematoidea, Haploporoidea, Gorgoderoidea, Brachycladioidea, Microphalloidea, Plagiorchioidea, and Opecoeloidea) that are monophyletic in the Xiphidiata. The Haploporoidea was a basal superfamily nested close to the Gorgoderoidea and Troglotrematoidea and was not supported as a distinct suborder Haploporata. Six of seven xiphidiatan superfamilies were monophyletic in the partial 28S phylogeny, with the exception of Opecoeloidea, which was separated into two different subclades: Opecoelidae and Stenakridae/Zdzitowieckitrematidae. The monophyletic Haploporoidea/Haploporata was separated from the Gorgoderoidea associates and placed in a marginal group in Xiphidiata. There were two notable clusters in the Paragonimidae: mixed-Paragonimus, which included a fairly compact group of P. heterotremus strains, and P. westermani/siamensis, which was divided into geographical/country strain groups. In conclusion, combined ribosomal rDNA sequences were more effective than single rDNA markers in resolving interfamilial and familial relationships. The ribosomal datasets presented here will be useful for taxonomic reassessment, as well as evolutionary and population genetics research in the Troglotrematoidea and other superfamilies in the Xiphidiata and the class Trematoda.

Bacterial dynamics and volatile metabolome changes of vacuum-packaged beef with different pH during chilled storage

This study aimed to assess the growth of spoilage bacteria in Brazilian vacuum-packed beef across different pH ranges (5.4–5.8, 5.8–6.1, ≥6.1) stored at temperatures of 0 °C, 4 °C, and 7 °C. Additionally, the research sought to identify predominant spoilage bacteria at the genus level using 16S rDNA gene sequencing and analyze the principal volatile organic compounds (VOCs) produced by this microbiota through HS-SPME/GC–MS. Lactic acid bacteria (LAB) consistently exhibited counts exceeding 6.0 Log CFU/g, regardless of temperature and pH conditions. The bacterial diversity in the meat samples reflected the influence of slaughterhouse environments, with Pseudomonas and Serratia remaining dominant across different cuts and pH levels. Post-storage, variations in pH and temperature modulated the initial bacterial diversity, leading to a reduction in diversity and an increase in LAB such as Lactobacillus, Lactococcus, Leuconostoc, and Carnobacterium. Notably, these changes were observed within pH ranges of 5.4–5.8 and 5.8–6.1, irrespective of beef cuts and storage temperatures. Based on high throughput sequencing and VOCS, correlation analysis revealed a relationship between the growth of specific spoilage microorganisms under vacuum conditions and the presence of VOCs such as alcohols (e.g., 1-propanol, 2-methyl-) and ketones (e.g., 2-nonanone, 2-octanone, 2-heptanone), identifying them as potential indicators of spoilage bacteria growth.

Dermacentor species (Acari: Ixodidae) in western Canada, with detection of Dermacentor similis.

Numerous tick species are undergoing significant range expansion in Canada, including several Dermacentor spp Koch (Acari: Ixodidae). With the recent description of Dermacentor similis Lado in the western United States, additional research is required to determine the current range of this species. Five hundred ninety-eight Dermacentor spp. were collected from companion animals in the western Canadian provinces of British Columbia, Alberta, and Saskatchewan. Ticks were morphologically identified to species, followed by PCR and gel electrophoresis of the ITS-2 partial gene target (n = 595). Ninety-seven percent (n = 579/595) generated valid banding patterns. The banding pattern for the majority (74%, n = 206/278) of Dermacentor spp. from southern British Columbia was consistent with D. variabilis (Say), while 26% (n = 72/278) was consistent with D. andersoni Stiles. For samples from Alberta, 38% (n = 3/8) had banding patterns consistent with D. variabilis and 63% (n = 5/8) with D. andersoni. All (n = 293) ticks from Saskatchewan had banding patterns consistent with D. variabilis. After the description of D. similis was published, DNA sequencing of mitochondrial (16S rDNA gene, COI gene) and nuclear (ITS-2) markers was used to confirm the identity of 40 samples. Twenty-seven samples that had banding patterns consistent with D. variabilis from British Columbia were confirmed to be D. similis. One sample from Alberta and five from Saskatchewan were confirmed to be D. variabilis and seven samples from British Columbia were D. andersoni. The ITS-2 amplicons were not useful for differentiating between D. variabilis and D. similis. These results provide evidence of D. similis in western Canada and highlight that sequences of the mitochondrial genes are effective for distinguishing D. andersoni, D. variabilis, and D. similis.

Effects of water-insoluble wheat bran-fraction powder on disease activity and caecal microbiota in dextran sodium sulphate-induced inflammatory bowel disease mouse model.

Water-soluble arabinoxylan exerts anti-colitic effect and exhibits ameliorative activity in an inflammatory bowel disease (IBD) mouse model. Water soluble fibre from wheat bran (WB) also exhibits anti-colitic effect. However, arabinoxylan is a primary compound of insoluble polysaccharide (hemicellulose) in WB. This study aimed to clarify the anti-IBD effects of the WB water-soluble (WBS) and water-insoluble (WBI) fractions. WB suspension was autoclaved and fractionated to WBS and WBI. C57BL/6 mice were divided into control (CT), dextran sodium sulphate (DSS), WBI, and WBS groups. They were fed as follows from day 1: CT, standard diet and distilled water; DSS and WBI, 3% (w/v) DSS in drinking water; WBI, 8% (w/w) WBI diet; and WBS, 50% (v/v) WBS and 3% (w/v) DSS in water. DSS group mice showed diarrhoea, body weight reduction, and blood in faeces by day 5 and colon tissue damage by day 6. These inflammatory indices were significantly inhibited by treatment with WBI. Amplicon sequencing of the 16S rDNA (V4) gene of the caecal contents of the CT, DSS, and WBI groups showed that the abundances of Escherichia, Allobaculum, and Bacteroidaceae increased and that of Faecalibaculum decreased in the DSS group. KEGG pathway prediction showed that amino acid metabolism and lipopolysaccharide biosynthesis decreased and increased, respectively, in the DSS group. However, WBI treatment tended to suppress these effects. WBI, rather than WBS, reduces inflammation and maintains the gut microbiota. However, further studies are warranted to elucidate the properties of the WBI active components and efficacy of WBI metabolites on gut microbiota, particularly on Faecalibaculum.

Cloning, Expression, Purification and Enzymatic Characterization of Low-temperature Cholesterol Esterase from Marine Panthenia Agglutinosa

BACKGROUND: The low-temperature cholesterol esterase is primarily used in industries such as papermaking and healthcare. OBJECTIVE: To discover a microorganism with high cholesterol esterase activity and tolerance to low temperatures, leading to the promotion of the sustainable utilization of marine cold-adapted microbial resources and fostering industrial development. MATERIALS AND METHODS: This study isolated a strain producing low-temperature cholesterol esterase from marine samples in the China Bohai Sea. The strain was identified through 16S rDNA sequencing and named Panthenia agglutinosa Y03. The cholesterol esterase gene (PaChe) from P. agglutinosa Y03 was cloned and heterologously expressed in Escherichia coli, and the recombinant enzyme PaChe was purified and characterized. The structure of PaChe was predicted using AlphaFold2, and molecular docking was performed with cholesterol linoleate as the ligand. RESULTS: The enzyme protein has a molecular weight of 56.35 KDa, a theoretical pI of 7.24, lacks a signal peptide, and exhibits structural features of the α/β hydrolase superfamily protein. The concentration of the purified PaChe is 0.5 mg/mL, with a specific activity of 42.7 U/mg. The optimal working temperature is 30 °C, and the enzyme retains activity at 4 °C , demonstrating weaker thermal stability. The optimal pH is 7, and the enzyme maintains over 70% activity at pH 9. Na +, Ca 2+ and Mg 2+ are the primary activators, while Ba 2+, Fe 2+, Mn 2+, Cu 2+ and chemical agents such as SDS as inhibitors, with Cu2+ exhibiting particularly significant inhibitory effects. CONCLUSION: This study establishes the theoretical groundwork for the development and utilization of a novel lowtemperature cholesterol esterase.

The microbiota comparative analysis of the characteristics between colorectal adenomatous polyps and normal mucosal intestinal.

The aim of this study is to systematically examine and compare the characteristics distinguishing colorectal adenomatous polyps from normal mucosal intestinal microbiota. A total of 30 specimens were obtained from patients diagnosed with colorectal adenomatous polyps (adenoma group) who underwent endoscopic removal at Wenzhou People's Hospital between September 2021 and November 2021. Concurrently, 30 normal mucosal specimens were collected from patients without adenomatous polyps (control group). Subsequently, microbiome total DNA extraction was carried out, followed by PCR amplification targeting the V3-V4 region of the 16S rDNA. High-throughput sequencing was conducted using the Illumina MiSeq platform. Subsequent to sequencing, bioinformatics analysis was used to assess the diversity, composition, and functional aspects of the intestinal microbiota in both study groups. A notable dissimilarity in the microbiota structure was identified, specifically within the transverse colon, between these two groups ( P < 0.05). Species composition analysis revealed that Escherichia , Fusobacterium , and Bacteroides were predominant bacteria in both groups, with Escherichia and Enterobacter displaying significant differences at the genera level between the control group and the adenoma group ( P < 0.05). Correlation analysis and functional prediction demonstrated substantial disparities in interactions among dominant intestinal microbial genera within patients from both groups. Additionally, it was discovered that the intestinal microbiomes in patients in the adenoma group exhibited a significantly higher pathogenic potential. Upon conducting a comprehensive analysis, it was discerned that the microbiota present in the transverse colon of the control group exhibited distinctive characteristics that may contribute to the maintenance of intestinal health.

Integrated Gut Microbiome and UHPLC-MS Metabolomics to Reveal the Prevention Mechanism of Pidanjiangtang Granules on IGT Rats

IntroductionPidanjiangtang (PDJT) is a traditional Chinese medicine formula empirically used to treat impaired glucose tolerance (IGT) based on the "Pidan" theory from the classic ancient book Nei Jing. However, the mechanism of PDJT intervention for IGT remains to be studied. ObjectiveThis study aims to explore the mechanism of PDJT granules intervention in IGT by integrating gut microbiome and UHPLC-MS untargeted metabolomics. Materials and methodsThe IGT model was established in 6-week-old male Sprague-Dawley (SD) rats by feeding them a high-fat diet and using an STZ injection. The low, medium, and high doses of PDJT were used for six weeks. Metformin was used as the positive control drug. The efficacy of PDJT was evaluated using fasting blood glucose (FBG), blood glucose maximum (BGmax), blood lipid, and inflammatory factor levels. Finally, 16S rDNA gut microbiome sequencing with metabolomics analysis was used to explore the pharmacological mechanism of PDJT intervention in IGT. ResultsPDJT could reverse the phenotype of IGT rats, reduce blood glucose levels, improve lipid metabolism disorder, and reduce inflammatory response. Gut microbiome analysis found that PDJT can improve gut microbiota composition and abundance of three phyla (Firmicutes, Bacteroidota, Desulfobacterota) and four genera (unclassified_f__Lachnospiraceae, Ruminococcus, Allobaculum, Desulfovibrio), which play an important role in the process of PDJT intervention on glucose metabolism and lipid metabolism in IGT rats. UHPLC-MS untargeted metabolomics showed that PDJT could regulate the levels of 258 metabolites in lipid metabolism pathways, inflammatory response pathways, fat and protein digestion, and absorption. The combined analysis of the two omics showed that improving the body's metabolism by gut microbes may be the possible mechanism of PDJT in treating IGT. Thus, this study provides a new method to integrate gut microbiome and UHPLC-MS untargeted metabolomics to evaluate the pharmacodynamics and mechanism of PDJT intervention in IGT, providing valuable ideas and insights for future research on the treatment of IGT with traditional Chinese medicine.

Short-chain fatty acids enhance muscle mass and function through the activation of mTOR signalling pathways in sarcopenic mice.

Sarcopenia is a prevalent muscle disorder in old people leading to higher fracture rate, mortality, and other adverse clinical outcomes. Evidence indicates that short-chain fatty acids (SCFAs), which are beneficial gut microbial metabolites, were reduced in old people with sarcopenia. This study aimed to determine whether the use of SCFAs as a supplement can be a therapeutic strategy of sarcopenia in a pre-clinical model. Seven-month-old pre-sarcopenic senescent accelerated mouse prone 8 (SAMP8) mice received daily SCFAs cocktail (acetate, butyrate, and propionate) for 3months. Age-matched senescence accelerated mouse resistant 1 (SAMR1) and SAMP8 mice receiving sodium-matched drinking water were control groups. The gut microbiota composition analysis of aged mice with or without sarcopenia was conducted by 16S rDNA sequencing. Gut barrier-related proteins and lipopolysaccharide (LPS) concentration were biomarkers of gut permeability. Colon inflammation levels, circulatory SCFAs concentration, muscle quality, function, and underlying pathways were detected by cell number counting, RT-qPCR, gas chromatography-mass spectrometry, measurements of muscle wet weight and grip strength, ex vivo functional test, treadmill endurance test, transcriptomic sequencing, morphological and immunofluorescent staining, as well as western blot. To investigate the role of mTOR signalling pathways in SCFAs treatment, C2C12 myotubes were treated with rapamycin. Aged SAMP8 mice had different microbiota composition, and lower serum butyric acid compared with SAMR1 mice (P<0.05). SCFAs treatment reversed the increment of colon inflammation (2.8-fold lower of il-1β) and gut barrier permeability (1.7-fold lower of LPS) in SAMP8 mice. Increased muscle mass, myofibre cross-sectional area, grip strength, twitch and tetanic force were found in SCFAs-treated mice compared with control SAMP8 mice (P<0.05). Anti-fatigue capacity (1.6-fold) and muscle glycogen (2-fold) also improved after SCFAs treatment (P<0.05). Transcriptomic analysis showed that AMPK, insulin, and mTOR pathways were involved in SCFAs treatment (P<0.05). Regulation of AKT/mTOR/S6K1 and AMPK/PGC1α pathways were found. SCFAs attenuated fat infiltration and improved mitochondria biogenesis of atrophic muscle. In vitro studies indicated that SCFAs inhibited FoxO3a/Atrogin1 and activated mTOR pathways to improve myotube growth (P<0.05), and rapamycin attenuated the effect of SCFAs through the inhibition of mTOR pathways. This study demonstrated that bacterial metabolites SCFAs could attenuate age-related muscle loss and dysfunction, and protein synthesis-related mTOR signalling pathways were involved both in vivo and in vitro.

Karyotype diversity of Polybia (gr. occidentalis) species complex (Hymenoptera: Vespidae): Taxonomic and evolutionary implications

AbstractPolybia is a common wasp genus found in most of South America. Polybia (gr. occidentalis) encompasses several species that are difficult to identify because of their similar morphologies. Our goal was to cytogenetically characterize three species belonging to Polybia (gr. occidentalis) and discuss the importance of cytogenetic data for integrative taxonomy. Polybia colonies were sampled from different regions of Brazil to perform classical and molecular cytogenetic analyses. Polybia paulista and Polybia sp. 1 showed 2n = 34 and 18S rDNA loci on two chromosomes, whereas Polybia sp. 2 showed 2n = 40 and 18S rDNA clusters on four chromosomes. The microsatellites GA(15), GAG(10), CAA(10), TTAGG(6), and TCAGG(6) showed similar distributions among the species, forming blocks in the euchromatic regions of the chromosomes, whereas CGG(10) did not yield any positive markings. In contrast, TAT(10) hybridized on the centromeric heterochromatin, showing differences in the number of marked chromosomes among the species. Therefore, it may potentially be a species‐specific cytotaxonomic marker in this group of wasps, but this feature needs further investigation. Fluorochromes evidenced that AT‐rich DAPI+ sequence distribution was coincident with heterochromatin, while the distribution of CG‐rich CMA3+ sequences was coincident with the 18S rDNA region. The present study revealed differences in diploid number, heterochromatin content, 18S rDNA sites, and microsatellite patterns between morphologically similar species, demonstrating the usefulness of cytotaxonomy in studying species complexes.

Molecular phylogeny of Boninia (Platyhelminthes: Polycladida), with description of a new species from the Pacific coasts of Panama.

Mesopsammic polyclad members in the family Boniniidae have attracted attention in terms of their evolutionary shifts of microhabitat and their unique morphology such as a pair of pointed tentacles extending from the anterolateral margins and prostatoid organs harbouring stylets. Here, we establish a new species of this family as Boninia panamensis sp. nov. from the Pacific coasts of Panama, based on its morphological characteristics of (i) four cerebral and 61-80 marginal eyespots, (ii) two prostatoid organs located anterior and posterior to the penis papilla, and (iii) two uterine canals departing from the anterior part of the Lang's vesicle. We also report Boninia cf. uru from Hawai'i, USA, based on its morphological identity with B. uru from Okinawa, Japan, along with their genetic distances for the partial cytochrome c oxidase subunit I (COI ) sequences, which were beyond the range of intraspecific differences observed in congeners in this study. Boninia oaxaquensis is also reported from Panama as a new locality for the species. Involving the above-mentioned three species sequenced herein, we reconstructed molecular phylogenetic trees of Boninia based on the four gene markers (18S rDNA, 28S rDNA, 16S rDNA and COI ). Our phylogenetic trees indicated the synapomorphy within the genus Boninia of the small numbers of stylets (2-4) and the connection route of the uterine canals to the Lang's vesicle. The results also showed a characteristic distribution pattern in which pairs of species in distinct lineages occurred sympatrically with different microhabitats, as observed in Boninia uru and Boninia yambarensis in Okinawa and B. panamensis sp. nov. and B. oaxaquensis in Panama. In addition, we discuss possible speciation pathways in this genus based on the tree topology. ZooBank: urn:lsid:zoobank.org:pub:D414BACD-C14A-4B34-8AF9-7ACBA28F46D0.