rDNA Sequences Research Articles

First Report of Pantoea ananatis Causing Bacterial Leaf and Panicle Blight of Rice in Louisiana, USA.

In July 2023, panicle and leaf blight-like symptoms were observed from the rice (Oryza sativa) variety, PVL03, in research field plots in Louisiana (Rayne, LA 70578, USA; 30.21330⁰ N, 92.37309⁰ W). We observed that chlorosis started on the tips and spread throughout the whole leaf. We also observed panicle blight-like symptoms (grain discoloration of rice hulls) on infected rice plants at the heading stage, which ranged from dark brown to black. Disease incidences were approximately 70% in the observed plots. Five plants were sampled from five plots (one plant per plot). Leaf tissues including both symptomatic and healthy parts (~ 1 cm2) were surfaced sterilized and placed on CCNT medium, a semi-selective medium for the rice bacterial panicle blight pathogen Burkholderia glumae (Kawaradani et al., 2000), and on Luria-Bertani (LB) agar supplemented with cycloheximide (50 µg/ml) for non-selective isolation of bacteria. For the panicle samples, the surfaced sterilized rice kernels were dehulled and ground in sterile distilled H2O using a sterile mortar and pestle. The homogenate was spread on the CCNT media and LB agar supplemented with nitrofurantoin (50 µg/ml) and cycloheximide (50 µg/ml). After ~72 h at 41⁰C, round, smooth and yellow-colored bacterial colonies were observed on both media. The two isolates, one from an infected rice leaf (LPL-1) and the other from the infected rice panicles (LPP-1), were initially identified as Pantoea ananatis based on PCR-amplified 16S rDNA sequences (Weisburg et al., 1991). The taxonomic identity of these two isolates was further confirmed through whole genome sequencing (NCBI Accession ID: PRJNA1092676), which was performed through a hybrid sequencing approach using Oxford Nanopore sequencer & Illumina NextSeq2000 system (Plasmidsaurus Inc., South San Francisco, CA, USA). To satisfy Koch's postulates, we inoculated six one-month-old rice plants (cv. Kitaake) with LPL-1 and LPP-1, respectively, by pricking the stems with a sterile toothpick touched to the overnight grown bacterial culture on LB agar (~ 5-8 x 105 bacterial cells). This virulence assay was conducted twice, and rice plants inoculated with sterile distilled water were included as the negative control. By 27 days post inoculation (DPI), the inoculated plants developed symptoms on leaves and panicles, which were like the symptoms originally observed in the field, and additional symptoms such as shot holes and stem necrosis were also observed. From leaf and panicle samples of the artificially inoculated plants, we reisolated bacteria and confirmed the identity of the bacteria using the P. ananatis specific PCR primers PANA_1080 61F and PANA_1080 1009R (Asselin et al., 2016). To the best of our knowledge, this is the first report of bacterial panicle and leaf blight of rice caused by P. ananatis in Louisiana. Since P. ananatis is a recently emerging pathogen of rice worldwide, including the United States (Yu et. al., 2022; Lu et al., 2022; Luna et al., 2023), it is crucial to monitor the occurrence of this pathogen in other rice-growing areas as a potential threat to US rice production.

Identification, Pathogenicity, and Antimicrobial Resistance Analysis of Bacterial Pathogenesis Aeromonas hydrophila from Hybrid Sturgeon (Huso dauricus ♀ × A. schrenckii ♂) in Zhejiang, China

In 2019, a disease outbreak struck a hybrid sturgeon farm (Huso dauricus ♀ × A. schrenckii ♂) in Tiantai, Zhejiang province, leading to the deaths of 8000 sturgeons. The sturgeons exhibited reduced appetite, lethargic and uncoordinated swimming, and physical signs such as reddish petechiae and ulcers on the body and fins. Hemorrhagic spots were observed on the kidneys, spleen, and gonads, alongside reddish intestines with hemorrhagic ascites in the abdominal cavity. ST-1902 was isolated and identified as Aeromonas hydrophila through physiological and biochemical characterization and 16S rDNA sequence analysis. The pathogenicity of ST-1902 was confirmed through a challenge test, with a median lethal dosage (LD50) of 7.9 × 106 CFU/IND. Histopathological examination showed hyperplasia and neoplasm-like changes in the epicedial mesothelial tissues, enlarged and necrosis renal tissue, and serious hemosiderosis in spleen and gills. Virulent genes (Aer, Epa, Alt, Hly, and Act) were detected in ST-1902, corresponding to typical β-hemolysis, extracellular protease, and enterotoxin. Moreover, antimicrobial experiment detection indicated ST-1902 is sensitive to quinolones and phenicols but resistant to sulfamethoxazole, aminoglycoside antibiotics with Sul1, and Intl and Ant (3”)-I. These results suggest that A. hydrophila was the causative agent of the sturgeon disease and highlight the emerging threat it poses to the sturgeon industry.

Mitigating Vibrio-Induced Skin Ulceration in Sea Cucumbers Using Probiotic Strains

Sea cucumbers are valuable in aquaculture, but their cultivation faces challenges from diseases such as skin ulceration syndrome caused by Vibrio alginolyticus (VA). This study aimed to isolate and identify probiotics capable of combating VA and improving sea cucumber’s growth performance. Pathogenic VA was identified, through 16S rDNA sequencing, confirming its high genetic similarity (>99%) to Vibrio alginolyticus. Two Bacillus strains, Bacillus licheniformis YB-1, and Bacillus megaterium YB-2, were isolated as potential probiotics, with identification supported by 16S rDNA phylogenetic analysis and deposition in microbial culture collections. They demonstrated strong antibacterial activity against VA in vitro without exhibiting antagonism when combined. Probiotic tolerance to environmental stressors was observed, while feeding trials revealed significant growth improvements in sea cucumbers, with the highest specific growth rates observed at 1 × 106 CFU/mL for both strains. Immersion challenge tests showed that sea cucumbers treated with probiotics exhibited reduced symptoms of rotten skin syndrome and higher survival rates. The optimal combination of YB-1 and YB-2, with viable bacteria concentrations of 5 × 107 CFU/mL each, achieved a 55% survival rate after a VA challenge, demonstrating their synergistic efficacy. These findings suggest that YB-1 and YB-2 offer promising probiotic solutions for enhancing sea cucumber health and resistance to VA infections in aquaculture.

Limitations of 18S rDNA Sequence in Species-Level Classification of Dictyostelids

Dictyostelid species classification has traditionally relied on morphology, a time-intensive method requiring expert knowledge. This study evaluated the potential and limitations of using the 18S rDNA sequence for species-level classification. 18S rDNA sequences of 16 samples from the Dicty stock center, including 14 samples found in Thailand, were analyzed. Signature sequence analyses confirmed genus-level identification with high accuracy. These sequences were analyzed alongside 309 database entries retrieved from the GenBank database. The analyses confirmed genus-level identification accuracy but highlighted challenges in distinguishing species due to overlapping intraspecific and interspecific variations, negative barcoding gaps, and incorrectly grouped samples to putative taxa by species delimitation analyses. Species delimitation methods, including maximum likelihood (ML) phylogenetic analysis, achieved limited success, with ML showing the highest accuracy but not exceeding 50%. However, species with high barcoding gaps, such as Raperostelium and Rostrostelium, demonstrated potential for accurate classification. These findings support using 18S rDNA for genus-level identification and suggest its possible application for certain species. Expanded sampling is needed to improve species-level classification and to identify more robust DNA markers for dictyostelid diversity studies.

Morphological and Molecular Characterization of Two Species of Ligophorus (Monogenea: Ancyrocephalidae) in Mullets from the Yucatán Peninsula, with Comments on the Geographical Distribution of L. mediterraneus

BackgroundLigophorus Euzet and Suriano, 1977 is a specious genus of ancyrocephalid monogeneans parasitizing mullets around the world, with most species distributed in the western Pacific and the Mediterranean Sea. Only nine out of the 62 species in the genus have been reported from the Americas, and from them, only two have been sequenced.MethodsWe analyzed two species of Mugil (L.) from Northern Yucatán Peninsula. Specimens of Ligophorus were sampled from the gills of their hosts. The morphology of the specimens was examined. In addition, 28S and ITS rDNA sequences were obtained and compared with previous sequences downloaded from GenBank.ResultsWe discovered two species of Ligophorus using morphological and molecular characters, L. mediterraneus, parasitizing the stripped mullet Mugil cephalus off the coast of Celestún, and L. yucatanensis, parasitizing the silver mullet M. curema in four coastal lagoons. Sequence data of the latter species are reported for the first time.ConclusionOur findings showed that two species of Ligophorus occur in mugilids of the Yucatán Peninsula. One represents a widely distributed marine species with records in the Mediterranean Sea and the Yucatán Peninsula, whereas the second one, L. yucatanensis, represents an endemic species restricted to coastal lagoons of the Yucatán Peninsula.

Effects of Salmonella Typhimurium infection on intestinal flora and intestinal tissue arachidonic acid metabolism in Wenchang chickens

Salmonella infections can lead to intestinal inflammation and metabolic disorders in birds. However, whether arachidonic acid (ARA) metabolism is involved in Salmonella-induced intestinal inflammation remains unclear. This experiment investigated the changes in cecal flora and ARA metabolism in Hainan Wenchang chickens infected with S. Typhimurium using 16s rDNA sequencing and targeted metabolomics. The results showed that the levels of ARA metabolites were increased in the cecum tissue of Wenchang chickens after infection with S. Typhimurium, including prostaglandin E2 (PGE2), prostaglandin F2α (PGF2α), lipoxin A4 (LXA4), ± 8(9)-EET, ± 11(12)-EET, and ± 8,9-DiHETrE. The content of key enzymes for ARA production and metabolism (Phospholipase A2 PLA2 and Cyclooxygenase-2 COX-2) in chicken cecum tissues was increased after S. Typhimurium infection. The relative mRNA levels of inflammatory factors were also increased after infection, including Interferon-γ (IFN-γ), Transforming growth factor-β1 (TGF-β1), Interleukin-4 (IL-4), and Interleukin-6 (IL-6). In HD11 cells, the use of a cyclooxygenase (COX) inhibitor reduced the increased levels of COX-2 and PGF2α induced by S. Typhimurium infection and effectively reduced the inflammatory response. In addition, the number of beneficial genera (e.g., Bifidobacterium, Lactobacillus, and Odorobacterium) in the cecum of Wenchang chickens was significantly reduced after infection with S. Typhimurium. The present study revealed the structure of cecal flora in S. Typhimurium-infected Wenchang chickens. In addition, this study demonstrated that S. Typhimurium activates the ARA cyclooxygenase metabolic pathway, which in turn mediates the development of intestinal inflammation in Wenchang chickens. The results can provide data support and theoretical support for the prevention and control of avian salmonellosis.

Gut microbiota and quantitative traits divergence at different altitude of long-tailed dwarf hamsters, Cricetulus longicaudatus

To investigate the community structure and diversity of gut microflora and their function in body mass regulation, as well as the effects of various locations on gut microbiota and Cricetulus longicaudatus body mass regulation at various elevations. We examined the diversity, abundance, and community structure of the gut microbiota of long-tailed dwarf hamsters from eight regions in Shanxi province during summer using 16S rDNA sequencing technology and analyzed the relationships between these microbiota and environmental variables as well as morphological indicators. The results revealed Firmicutes and Bacteroidetes as the dominant phyla at the phylum level, with Lactobacillus emerging as the predominant genus. We observed differences of gut microflora between different areas, and this diversity is affected by altitude. The high-altitude areas individuals had lower β diversity of gut microbiota than the low-altitude area. Moreover, the body and skull indexes of long-tailed dwarf hamsters also changed with altitude. The result presented in this study indicated that the body size of long-tailed dwarf hamsters conforms to Bergmann's law. And Providencia had significant correlation with body size. Finally, functional analysis of the gut microbiota showed changes in metabolic function that depended on elevation, and collinear network analysis showed how the gut microbiota interacts with each other. All of these results suggest that long-tailed hamsters are different depending on their altitude, with altitude being the main factor affecting both the structure of microbes and the way their metabolism works. This study shows that altitude has a big effect on the gut microbiota and phenotypic traits of long-tailed hamsters. It also shows how well this species can adapt to changes in altitude.

Dietary supplementation with Epimedium contributes to the improvement of hormone levels, gut microbiota, and serum metabolite composition in the Chinese forest musk deer (Moschus berezovskii)

The Chinese forest musk deer (Moschus berezovskii) is a small ruminant animal with special economic value. It is listed as a National Level I key protected species in China. However, these animals are prone to stress responses in captive environments. Epimedium, a traditional Chinese herb with aphrodisiac and anti-stress properties, may have potential benefits for the health of the captive Chinese forest musk deer, though its efficacy requires further investigation. This study aimed to evaluate the effects of dietary supplementation with Epimedium on the hormone levels, gut microbiota composition, and serum metabolism of the Chinese forest musk deer. The fourteen adult male Chinese forest musk deer with similar initial body weights (7.0 ± 0.3 kg) and an average age of 4.5 years were randomly divided into two groups, each containing seven animals. The control group was fed a standard diet without Epimedium, while the Epimedium group received the standard diet supplemented with 15 g Epimedium /kg DM. The results indicated that the inclusion of Epimedium in the diet increased dry matter intake (DMI) and improved the ratio of feed to gain (F/G), with an increase in fecal testosterone levels (p < 0.05). 16S rDNA sequencing analysis revealed that Epimedium enhanced the richness and diversity of the gut microbiota in the Chinese forest musk deer, increasing the relative abundance of beneficial bacteria such as Firmicutes, while reducing the relative abundance of the potentially pathogenic Proteobacteria (p < 0.05). A widely targeted metabolomics analysis identified 25 differential metabolites between the two groups. Significant alterations were observed in key metabolic pathways related to lipid metabolism, hormone regulation, and antioxidation, such as ovarian steroidogenesis, tyrosine metabolism, and glycerophospholipid metabolism. Furthermore, correlation analysis between gut microbiota and serum differential metabolites showed that the relative abundances of Clostridia_vadinBB60_group and UCG-010 were positively correlated with anserine and 7-ketocholesterol, respectively (p < 0.05). In conclusion, Epimedium positively influenced feed intake and hormone levels in the Chinese forest musk deer by modulating gut microbiota composition and serum metabolism.

P0213 Salivary Veillonella parvula from Crohn’s Disease Patients Exacerbates Intestinal Inflammation

Abstract Background The gut microbiota plays a significant role in the development of inflammatory bowel disease (IBD). There is a close interaction between the oral and gut microbiota. However, the role of the "oral-gut" microbial axis in the development of IBD is not yet clear. Methods 1. Analyzed oral-gut microbiota patterns in 97 IBD patients using 16S rDNA sequencing and bioinformatics methods. 2. A mouse colitis model with DSS was used. It was gavaged with oral microbiota from CD patients to assess the impacts on colitis and gut microbiota. 3. Isolated specific differential bacteria, Veillonella, from CD patient saliva and evaluated their effects on DSS-induced colitis in mice. Results 1. IBD patients had a reduced core gut microbiota, lower diversity (P < 0.05), and increased abundance of potential pathogens such as Escherichia-Shigella. 2. CD patients showed more correlations between saliva and fecal microbiota, with the most significant correlation between Veillonella in saliva and Escherichia-Shigella in fecal (P=0.00018) . 3. The mice which were gavaged with saliva from CD patients showed significant weight loss (P < 0.0001), reduced colon length (P=0.03), increased histopathological scores (P=0.01), decreased expression of colonic mucosal barrier proteins ZO-1 and E-cadherin, and exacerbated gut microbiota dysbiosis . 4. The mice which were gavaged with Veillonella parvula from CD patients showed significant weight loss (P < 0.0001), reduced colon length (P=0.02), increased histopathological scores (P=0.02), decreased expression of colonic mucosal barrier proteins ZO-1 and E-cadherin, and exacerbated gut microbiota dysbiosis. Conclusion The interplay between the oral and gut microbiota potentially influences the progression of IBD, with oral bacteria such as Veillonella possibly intensifying intestinal inflammation in IBD patients by disrupting the gut microbiota balance. Targeted modulation of the gut microbiota and interference with oral-gut microbial interactions may offer a promising therapeutic strategy for managing IBD.

P1311 Mucosal Microbiome Signatures Associated with Postoperative Endoscopic Recurrence in Paediatric Crohn’s Disease

Abstract Background Following ileocecal resection (ICR) for Crohn’s disease (CD), recurrence is frequent even in the biologic era. Gut microbiota is involved in this setting and a few adult studies have suggested that bacterial recolonization after surgery may predict recurrence. Given the inherent risk of postoperative recurrence associated with age and disease duration, studies investigating the role of the gut microbiota in predicting relapse in children undergoing ICR appears critical. We aimed to identify changes in the mucosal microbiome associated with, and predictive of, endoscopic CD recurrence within the first year post-ICR in a paediatric population. Methods Microbiota composition (16S rDNA sequencing) was assessed from ileal and colonic mucosa at the time of ICR and later during endoscopic evaluation from above and below the surgical anastomosis in 33 patients (26 with paired samples) prospectively recruited at a tertiary care paediatric IBD center. Results The study included 33 children (mean age 16±2.7 years) undergoing ICR. At the time of ileo-colonoscopy, 9 (35%) patients had endoscopic recurrence (Rutgeerts’ score≥ i2). ICR strongly modified the gut mucosal microbiota composition with significant changes in β-diversity (R2= 0.03, PERMANOVA test, p=0.01). Bacterial recolonization post-ICR differed according to recurrence status (Figure 1). Post-operatively, recurrence was associated with an increase in Fusobacterium, Dialister genera and in members of the Lachnospiraceae family, and a decrease in Faecalibacterium, Lactobacillus genera and members of the Streptococcaceae family. Microbiota colonizing the neoterminal ileum and subanastomotic colon differed in children with recurrence and showed, compared to remission, elevated and reduced bacterial abundance of the Enterobacter (q=0.1) and Ruminococcaceae (q=0.05) families in colonic samples respectively while the ileal mucosa exhibited profound depletion in Faecalibacterium (q=0.01). Postoperative colonic mucosal relative abundance of Enterobacteriaceae (q=0.06) and Ruminococcaceae (q=0.06) correlated positively and negatively with length of intestinal resection. At the time of surgery, several bacterial taxa able to predict relapse were identified (Figure 2). Conclusion Predicting postoperative CD relapse remains challenging, although several clinical risk factors of recurrence have been identified. Our study demonstrates that bacterial recolonization following ICR might pave the way towards remission or recurrence. Our results highlight, for the first time in a paediatric population, target species that may help clinicians to better define patients at risk of relapse and allow precise modulations of the overall microbial ecosystem towards remission pattern.

Ameliorative Effect of Glycyrrhizic Acid on Diosbulbin B-Induced Liver Injury and Its Mechanism.

This study aimed to clarify the protective effect of Glycyrrhizic acid (GL) against Diosbulbin B (DB) - induced liver injury in mice and investigate its mechanisms of action. A liver injury DB was established in mice through the oral administration of DB for 15 days. At the same time, GL was administered to the mice for treatment. After the experiment, the pharmacodynamics and mechanisms of GL in ameliorating DB-induced liver injury were explored using biochemical indexes, non-targeted metabolomics, targeted metabolomics, Western blotting analysis of protein expression, 16S rDNA sequencing, and Spearman correlation analysis. The results show reduced liver function indices and improved DB-induced hepatic pathological changes. It also attenuated DB-induced hepatic inflammation and oxidative stress. Hepatic metabolomics revealed that GL regulated ABC transporters and bile secretion. Targeted bile acid (BA) metabolomics and Western blotting demonstrated that GL improved DB-induced reduction in BA efflux by regulating FXR-mediated efflux transporters. Furthermore, analysis of 16S rDNA gene sequencing revealed that GL effectively restored the relative abundance of beneficial bacteria, reduced the relative abundance of harmful bacteria, and reinstated the structure of the intestinal flora. Additionally, correlation analyses between BA and intestinal flora indicated that Firmicutes, Bacteroidota, TDGA, DGA, UDGA, GDGA, THDGA, and HDGA could serve as major markers for DB-induced liver injury. In conclusion, GL significantly improved DB-induced liver injury by increasing the expression of Nrf2/FXR-BSEP/MRP2/P-gp/UGT1A1, promoting BA efflux, regulating intestinal flora, and alleviating inflammation and oxidative stress.

Tropical fruit-derived Lactiplantibacillus as potential probiotic and antifungal agents against Fusarium oxysporum

Fifty-five lactic acid bacteria (LAB) were isolated from seven selected tropical fruits, with Solanum nigrum exhibiting the highest LAB prevalence and Couroupita guianenis and Musa fruits showing the lowest counts. Two strains isolated from Ficus racemosa demonstrated significant antifungal activity against Fusarium oxysporum. 16S rDNA sequencing identified these strains as Lactiplantibacillus plantarum MYSVCF3 and Lpb. argentoratensis MYSVCF5. The isolates displayed adaptability to a broad range of environmental conditions, including temperatures of 10–45 °C, pH 2–6, and salt up to 7%. The strains tolerated simulated gastrointestinal conditions of acid (pH-2), phenol (0.6%), and bile (0.3%) suggesting potential probiotic attributes. Lpb. argentoratensis MYSVCF5 inhibited F. oxysporum, two ESKAPE group bacteria (P. aeruginosa, S. aureus) plus S. paratyphi and E. coli. The cell-free supernatant (CFS) of Lpb. argentoratensis MYSVCF5 reduced the growth of fungal biomass by 94% and completely inhibited conidial germination, retaining activity even after extended cold storage. LC–MS/MS analysis identified organic acids in the CFS, with citric acid as the most abundant at 34.9 (± 0.3) µg/mL, followed by lactic (8.3 µg/mL) and malic acids (5.2 µg/mL). This study isolated a novel LAB, a potential candidate having probiotics and antifungal properties for application in food and agriculture.

Morphology and phylogenetic position of two novel Dysteria species (Ciliophora: Cyrtophoria) from deep-sea hydrothermal vents on the Central Indian Ridge, Indian Ocean

This study reports the discovery of two novel ciliate species – Dysteria brevicauda sp. nov. and Dysteria parasemilunaris sp. nov. – inhabiting unique environments in recently explored hydrothermal vent fields at the Onnuri and Onbada vents (1978 m and 2523 m depths, respectively) along the Central Indian Ridge in the Indian Ocean. The species were identified using a combination of morphological and molecular approaches. The morphology of D. brevicauda sp. nov. (65–110 × 20–30 μm) is characterized by a protruding dorso-anterior part, a short tail-like structure, converging outer two right kineties at the posterior end, and two frontoventral kineties with a few gaps in the anterior part. D. parasemilunaris sp. nov. (50–70 × 25–35 μm) can be recognized by its elongated oval shape with a dorsoapical collar-like protrusion, a terminally positioned stout podite, and five right kineties, including two frontoventral kineties. D. parasemilunaris sp. nov., which inhabits both the Onnuri and Onbada vents, is morphologically almost identical to Dysteria sp. collected from another hydrothermal vent, Clam Acres, in 1985. Thus, these are considered conspecific species. Phylogenetic analysis based on SSU rDNA sequences places the two new species within Dysteria Group III and V clades, respectively. D. brevicauda sp. nov. is positioned between Spirodysteria kahli (97.3% similarity) and D. subtropica (97.1% similarity). Meanwhile, D. parasemilunaris sp. nov. forms a cluster with D. semilunaris (95.3% similarity) and branches off from D. lanceolata (94.3% similarity). These findings contribute to the understanding of ciliate biodiversity in deep-sea hydrothermal vents. http:zoobank.org/urn:lsid:zoobank.org:pub:FE45985A-C0C1-4BAC-9471-3CC5D6180FEC.

Exploring the Effects of Gracilaria lemaneiformis Polysaccharides on the Fecal Microbiota and Fecal Metabolites of Fattening Pigs Based on 16S rDNA and Metabolome Sequencing.

Sixty Duroc × (Landrace × Yorkshire) castrated male finishing pigs were randomly divided into negative control (NC) and gracilaria lemaneiformis polysaccharides (GLP) groups to study the effects of GLP on the fecal microbiota and fecal metabolites of fattening pigs. The NC group was fed a basic diet, and the GLP group was fed a basic diet supplemented with 0.1% GLP. The diversity of the species and the fecal metabolites were analyzed using 16S rDNA sequencing and metabolome sequencing, respectively. The serum immune and antioxidant indices were analyzed using the ELISA assay. The abundances of hazardous bacteria such as Proteobacteria at the phylum level and Shigella at the genus level were extremely significantly decreased (p < 0.01) and those of beneficial bacteria such as Firmicutes at the phylum level (p < 0.01), Clostridium at the genus level (p < 0.01), and Lactobacillus at the genus level (p < 0.05) were significantly increased in the GLP group compared with the NC group. A total of 41 differentially expressed metabolites were identified. The expression of anti-inflammatory and antioxidant active substances, such as methyl cinnamate, protopanaxatriol, and isovanillic acid, was elevated in the GLP group. The ELISA assay showed increased GSH-Px activity (p < 0.01), T-AOC (p < 0.01), IgG (p < 0.01), IgA (p < 0.05), and IgM (p < 0.05) in the GLP group. These results indicate that dietary GLP supplementation can improve the antioxidant ability, anti-inflammatory ability, and immune level of fattening pigs by regulating fecal flora and metabolites and could be used as a functional feed additive.

Therapeutic Potential of Cajanus cajan (L.) Millsp. Leaf Extract in Modulating Gut Microbiota and Immune Response for the Treatment of Inflammatory Bowel Disease.

Background/Objectives: Inflammatory bowel disease (IBD) is a persistent inflammatory condition affecting the gastrointestinal tract, distinguished by the impairment of the intestinal epithelial barrier, dysregulation of the gut microbiota, and abnormal immune responses. Cajanus cajan (L.) Millsp., traditionally used in Chinese herbal medicine for gastrointestinal issues such as bleeding and dysentery, has garnered attention for its potential therapeutic benefits. However, its effects on IBD remain largely unexplored. Methods: In this study, the major compounds from Cajanus cajan leaf extract (CCLE) were initially characterized by LCMS-IT-TOF. The IBD model was developed in C57BL/6 mice by administering continuous 4% (w/v) dextran sodium sulfate (DSS) aqueous solution over a period of seven days. The body weight, colon length, disease activity index (DAI), and histopathological examination using hematoxylin and eosin (H&E) staining were performed in the IBD model. The levels of the main inflammatory factors, specifically TNF-α, IL-1β, IL-6, and myeloperoxidase (MPO), were quantified by employing enzyme-linked immunosorbent assay (ELISA) kits. Additionally, the levels of tight junction proteins (ZO-1, Occludin) and oxidative stress enzymes (iNOS, SOD1, CAT) were investigated by qPCR. Subsequently, flow cytometry was employed to analyze the populations of various immune cells within the spleen, thereby assessing the impact of the CCLE on the systemic immune homeostasis of IBD mice. Finally, 16S rDNA sequencing was conducted to examine the composition and relative abundance of gut microbiota across different experimental groups. In addition, molecular docking analysis was performed to assess the interaction between the principal components of CCLE and the aryl hydrocarbon receptor (AHR). Results: We identified seven bioactive compounds in CCLE: catechin, cajachalcone, 2-hydroxy-4-methoxy-6-(2-phenylcinyl)-benzoic acid, longistylin A, longistylin C, pinostrobin, amorfrutin A, and cajaninstilbene acid. Our results demonstrated that oral administration of CCLE significantly alleviates gastrointestinal symptoms in DSS-induced IBD mice by modulating the balance of gut-derived pro- and anti-inflammatory cytokines. This modulation is associated with a functional correction in M1/M2 macrophage polarization and the Th17/Treg cell balance in splenic immune cells, as well as shifts in the populations of harmful bacteria (Erysipelatoclostridium and Staphylococcus) and beneficial bacteria (Odoribacter, unidentified Oscillospiraceae, Lachnoclostridium, and Oscillibacter) in the gut. Furthermore, cajaninstilbene acid, longistylin A, and longistylin C were identified as potential AhR agonists. Conclusions: The present results suggested that CCLE, comprising stilbenes like cajaninstilbene acid, longistylin A, and longistylin C, protects the epithelial barrier's structure and function against DSS-induced acute IBD by restoring gut microbiota balance and systemic immune response as AhR agonists. Overall, CCLE represents a promising natural product-based therapeutic strategy for treating IBD by restoring gut microbiota balance and modulating systemic immune responses.

Effect of intermittent water flow on biodegradation of organic micropollutants in the hyporheic zone.

Water scarcity in the Mediterranean area has increased the number of intermittent rivers. Recently, hyporheic zones (HZ) of intermittent rivers have gained attention since a substantial part of the stream's natural purification capacity is located within these zones. Thus, understanding the flow dynamics in HZs is crucial for gaining insights into the degradation of organic micropollutants. A lab-scale study using column experiments was conducted in an attempt to elucidate the environmental processes accounting for the biodegradation capacity of the HZ under flow intermittency. A mixture of six compounds including pesticides (chloranthraniliprole, fluopyram and trifloxystrobin) and pharmaceuticals (venlafaxine, amisulpride and paroxetine) spiked at 1μg/L level was used for degradation kinetic studies and at 1mg/L for transformation products identification using suspect/non-target liquid chromatography high-resolution mass spectrometry approaches. The experiments lasted 60 days, divided into two 14-day phases: one before and one after a 5-week desiccation period. Bacterial community was charaterized by high-throughput DNA sequencing. The results suggested that intermittent flows stimulated the biodegradation of three compounds namely fluopyram, trifloxystrobin and venlafaxine, showing a large range of biodegradation profiles in batch water/sediment testing system according to OECD 308 tests. Biodegradation rate enhancement was ascribed to the occurrence of additional transformation routes after the desiccation period of river sediment, with the formation of new transformation products reported for the first time in the present work. 16S rDNA sequencing revealed that the desiccation period favored the growth of nitrifying and denitrifying bacteria which could partially explain the emergence of the new transformation pathways and most specifically those leading to N-oxide derivatives. Identification of transformation products also revealed that reductive transformation routes were relevant for this study, being dehydrogenation, dehalogenation, ether bond cleavage and sulfone reduction into sulphide important reactions. These results suggest that the intermittent flow conditions can influence the HZ biodegradation capacity.

Structural basis of nucleosome recognition by the conserved Dsup and HMGN nucleosome-binding motif.

The tardigrade Dsup and vertebrate high mobility group N (HMGN) proteins bind specifically to nucleosomes via a conserved motif whose structure has not been experimentally determined. Here we used cryo-EM to show that both proteins bind to the nucleosome acidic patch via analogous arginine anchors with one molecule bound to each face of the nucleosome. We additionally employed the natural promoter-containing 5S rDNA sequence for structural analysis of the nucleosome. These structures of an ancient nucleosome-binding motif suggest that there is an untapped realm of proteins with a related mode of binding to chromatin.

Molecular Phylogenetic Relationships and Cultural Conditions of Calocybe indica

Calocybe indica known as a milky white, indigenous and medicinally important edible mushroom. The cultural conditions, molecular identification, and phylogenetic relationships of selected strains of Calocybe indica were examined. The optimal temperature for mycelial development was found to be 30°C, whereas the lowest growth occurred at 15°C. The most favorable growth occurs at a pH of 5. Based on the growth characteristics of various culture media, potato dextrose agar proved to be the most advantageous, while Hennerberg and Czapek Dox media were less suitable. Molecular identification and the phylogenetic relationships of the chosen IUM strains of C. indica were examined through their ITS and RAPD analyses. The selected strains of C. indica had total sequence lengths of ITS1, ITS2, and 5.8S rDNA ranging from 570 to 620 bp. The sizes of the ITS1 and ITS2 regions vary among strains, with ITS2 showing greater variety than ITS1. All strains contained identical 5.8S rDNA sequences, according to sequencing analysis. Eleven out of the twenty arbitrary 10-base oligonucleotide primers utilized to amplify genomic DNA segments for the chosen C. indica strains were effective. RAPD-PCR resulted in distinct multiple products, demonstrating significant variability among the strains assessed. The quantity of amplified bands differed based on the primers utilized or the strains evaluated. The DNA polymorphisms displayed consistent traits in the replicated trials. Thus, it can be concluded that ITS sequencing and RAPD methods were effective in identifying the genetic diversity of the chosen strains of Calocybe indica. Plant Tissue Cult. &amp; Biotech. 34(2): 205-216, 2024 (December)

The microbial communities and metabolic profiles of follicular fluid in patients with premature ovarian insufficiency.

Premature ovarian insufficiency (POI) is a condition characterized by ovarian dysfunction occurring before the age of 40, and its etiology is multifactorial, including genetic, immunological, infectious, environmental, and iatrogenic factors, with over half of the cases remaining unexplained. Whether the microbial communities and metabolites in follicular fluid, which is the direct microenvironment for oocyte survival, are related to POI has not been reported. In this study, Follicular fluid samples of 26 patients with POI and 27 controls with a normal ovarian reserve were collected and analyzed using 16S rDNA sequencing and untargeted metabolomics. Conjoint analysis was performed to identify key microbial communities and metabolites that might be involved in POI. Patients with POI exhibited significant alterations in microbial richness and diversity and metabolic profile in their follicular fluid. The downregulation of ABC transporters and upregulation of the citrate cycle (TCA cycle) might be critical for the development and progression of POI. G-Rhodopseudomonas and g-Caulobacter were identified as key microbial genera, while L-aspartic acid, citrate, isoleucine, and cytidine were identified as key metabolites. These findings offer novel insights into the pathogenesis of POI and might pave the way for improved clinical outcomes for individuals with POI.

Integrating 16S rDNA and metabolomics to uncover the therapeutic mechanism of electroacupuncture in type 2 diabetic rats.

This study aimed to investigate the impact of electroacupuncture (EA) on blood glucose levels, gut microbiota, short-chain fatty acids (SCFAs), and glucagon-like peptide-1 (GLP-1) in a rat model of type 2 diabetes mellitus (T2DM). Forty Sprague-Dawley (SD) rats were randomly assigned to five groups (n = 8/group) using a random number table: normal control, T2DM model, electroacupuncture (EA), EA + antibiotics (EA + A), and antibiotics (A). The normal rats received a standard diet and saline gavage, while the other groups were fed a high-fat diet and emulsion. The EA + A and A groups received additional antibiotic solution gavage. The normal, model, and A groups were immobilized and restrained for 30 min, six times per week, for 4 weeks. The EA and EA + A groups received EA treatment at specific acupoints for 30 min, six times per week, for 4 weeks. EA parameters were continuous waves at 10 Hz and 1-2 mA. During the intervention, water and food consumption, body weight, fasting blood glucose (FBG), and oral glucose tolerance test (OGTT) were monitored. Pancreatic tissue was examined using hematoxylin and eosin (H&E) staining. Fecal microbial communities were analyzed by 16S rDNA sequencing, and short-chain fatty acids (SCFAs) were measured using gas chromatography-mass spectrometry (GC-MS). Serum levels of fasting insulin (FINS), glycated hemoglobin (HbA1c), and glucagon-like peptide-1 (GLP-1) were determined using enzyme-linked immunosorbent assay (ELISA). EA significantly improved daily water intake, food consumption, and body weight in T2DM rats (p < 0.01). EA also reduced FBG, the area under the curve of the OGTT, FINS, and homeostasis model assessment of insulin resistance (HOMA-IR) in T2DM rats (p < 0.05). The ELISA results showed a lower concentration of HbA1c in the EA group (p < 0.05). EA improved the overall morphology and area of pancreatic islets, increased the number of β-cell nuclei, and alleviated β-cell hypertrophy. The abundance of operational taxonomic units (OTUs) in the EA group increased than the model group (p < 0.05), and EA upregulated the Shannon, Chao1, and Ace indices (p < 0.05). EA increased the concentrations of acetic acid, butyric acid, and GLP-1 (p < 0.05). Correlation analysis revealed negative associations between Lactobacillaceae (R = -0.81, p = 0.015) and Lactobacillus (R = -0.759, p = 0.029) with FBG. Peptostreptococcaceae and Romboutsia were negatively correlated with HbA1c (R = -0.81, p = 0.015), while Enterobacteriaceae was positively correlated with OGTT (R = 0.762, p = 0.028). GLP-1 was positively correlated with acetic acid (R = 0.487, p = 0.001), butyric acid (R = 0.586, p = 0.000), isovaleric acid (R = 0.374, p = 0.017), valeric acid (R = 0.535, p = 0.000), and caproic acid (R = 0.371, p = 0.018). Antibiotics disrupted the intestinal microbiota structure and weakened the therapeutic effects of EA. EA effectively improved glucose metabolism in T2DM rats. The hypoglycemic effects of EA were associated with the regulation of gut microbiota, SCFAs, and GLP-1.