16S rRNA Sequence Analysis Research Articles

Molecular and genetic characterization of LEPTOSPIRA spp. collection strains from the St. Petersburg Pasteur institute based on 16S rRNA gene sequencing data

Leptospirosis is a zoonotic disease found virtually worldwide. Microscopic Agglutination Test with live leptospira (MAT) is the reference method for the serological diagnosis of leptospirosis. MAT is based on assessing serum potential to agglutinate live reference serovar Leptospira maintained at a reference laboratory. At some laboratories having own collections of isolated and reference Leptospira strains applicable for serological diagnosis, those microorganisms are maintained for many years by repeated subculturing, that increases markedly a chance of strain cross-contamination. The lack of adequate quality control for reference strains may affect data of epidemiological studies. Control of Leptospira spp. reference strains purity and stability of their antigenic composition is very important for diagnosis of leptospirosis. The study objective was to compare the 16S rRNA gene nucleotide sequences of some Leptospira strains from the collection of the St. Petersburg Pasteur Institute to with relevant sequences uploaded to GenBank. In this study, 38 Leptospira strains were investigated. Nucleotide sequences of 36 strains were deposited in the international GenBank database, inconsistencies were revealed in two strains. The study found that the control Leptospira strains from the collection of the St. Petersburg Pasteur Institute had minimal dissimilarities from international control strains. The analysis of the resultant 16S rRNA sequences has shown the presence of point mutations, transitions, deletions and insertions, regardless of the strain species. The open leptospira pan-genome demonstrates high genomic variability in species due to the capability of leptospira for lateral gene transfer in order to adapt to changing environmental conditions. The massive acquisition and loss of genes give rise to an increased species diversity. The 16S rRNA gene is suitable for screening diagnostics; however, high level of the fragment similarity and close phylogenetic relationship between different species put bounds to its use in genotyping. The presence of point nucleotide mutations is most likely associated with the evolutionary mechanisms of leptospira, their ability to horizontal gene transfer and crossing-over, including ribosomal genes, but this assumption necessitates additional research. For specimen genotyping it is necessary to select alternative genes with high specificity and sufficient level of nucleotide divergence. The study shows a need for genetic analysis of collection strains in order to control the purity of cultures.

First Report of ‘Candidatus Phytoplasma asteris’ (16SrI-B) Associated with Phyllody Disease of Moth Bean in World

Moth bean (Vigna aconitifolia), a drought and heat-resistant legume from the Fabaceae family, is commonly cultivated in arid and semi-arid regions of the Indian subcontinent In September 2022, phyllody symptoms (Figure 1) were observed on 50-days-old moth bean plants at the ICAR-NBPGR research farm in Jodhpur, Rajasthan, India. The disease incidence ranged from 10 to 25%. To investigate the cause, ten symptomatic VacoJod (1-10) and ten asymptomatic VacoJod (11-20) Vigna aconitifolia plants were collected. Insect populations were also collected from the vicinity using the sweep-net method to examine the role of insect vectors. The leafhopper was identified based on morphological characterization as Empoasca sp. at the Division of Entomology, ICAR-IARI, New Delhi. DNA was extracted from midribs of all collected plants and the Empoasca sp., using Qiagen DNeasy Plant Mini Kit and Blood and Tissue kit, respectively. Nested polymerase chain reaction (Nested-PCR) with universal primers P1/P7 and R16F2n/R16R2 (Deng and Hiruki, 1991; Gundersen and Lee, 1996), and secA gene primers (secAfor1/secArev3 and secAfor2/secArev3) (Hodgetts et al., 2008) were employed to determine phytoplasma species association. Out of the 10 symptomatic plants and 10 leafhopper samples, 6 leafhopper samples and all symptomatic plants produced expected band sizes for the 16S rRNA (approximately 1.25 kb) and secA gene (480 bp). The PCR products were cloned, sequenced, and sequences (two each from moth bean and leafhopper) were submitted to NCBI GenBank with accession numbers OP941130, OP941132, OP941133 and OP941134 for 16S rRNA and OP958868, OP958869, OP958870, and OP958871 for secA gene sequences. Nucleotide BLAST analysis of 16S rRNA sequences revealed a minimum of 99.92% similarity with 'Primula acaulis' yellows phytoplasma (KJ494340) from Czech Republic. All 100% hits corresponded to 16SrI-B group phytoplasmas, for example rapeseed phyllody phytoplasma (CP055264) from Taiwan. Similarly, nucleotide BLAST analysis of secA sequences revealed a minimum of 99.15% sequence similarity with Paulownia witches'-broom phytoplasma (secA) (OP124308) from China. All 100% hits were of 16SrI-B group phytoplasmas, for example Ageratum conyzoides yellowing phytoplasma (MW401697, secA) from India. Phylogenetic analysis using MEGA11 (Tamura et al., 2021) clustered the moth bean and Empoasca sp. phytoplasma strains with 16SrI-B phytoplasma reference strains. iPhyClassifier tool classified the 16S rRNA gene sequences into 16Sr group I, subgroup B, with a similarity coefficient of 1.0 (Figure 2a, 2b). This marks the first report of the association of 'Ca. P. asteris' 16SrI-B related phytoplasma strain with moth bean plants globally. The 16SrI-B phytoplasma strain is prevalent in various crops in India (Singh et al., 2023). This report emphasizes the epidemiological studies and highlights the need for further research and preventive measures to manage the spread of this phytoplasma strain, which could impact crop production and food security in hot and dry regions.

2,2-Dichloroacetamide exposure induces behavior and memory disorders in mice: Detrimental effects of long-term dietary restriction on neurotoxicity

2, 2-dichloroacetamide (DCAcAm), a nitrogen-containing disinfection byproduct (DBPs), is commonly found in potable water. This study aimed to compare the neurotoxicity of DCAcAm in C57/BL6 mice at both environmentally relevant and higher doses through oral exposure over a 28-day period. Furthermore, the potential effects of dietary restriction (DR) on the cerebral toxicity induced by 20 ppb DCAcAm were examined. The findings indicated that DCAcAm exposure and DR treatment resulted in reduced memory retention and cognitive adaptability in mice. Additionally, higher doses of DCAcAm exposure induced severe brain inflammation and oxidative stress. Metabolic profiling revealed disruptions in fatty acid, energy, and amino acid metabolism in the brain. Remarkably, the negative impacts of 20 ppb DCAcAm on the mice brain were worsened by DR treatment. Analysis of 16S rRNA sequencing revealed notable changes in the composition and structure of intestinal microorganisms after exposure to DCAcAm. This study discovered that DCAcAm has both direct effects on the brain and indirect effects through the microbial-brain-intestinal axis, which collectively result in neurotoxicity and dietary restriction exacerbates these effects. This study provides emerging views on the assessment of the toxicity of nitrogen containing DBPs.

The effects of pks+ Escherichia coli and bile acid in colorectal tumorigenesis among people with cholelithiasis or cholecystectomy.

Patients with cholelithiasis (CL) or cholecystectomy (CE) would have more chances of getting colorectal adenoma (CRA) or cancer (CRC). We aimed to figure out the effects of gut microbiota and bile acid on colorectal neoplasm in CL and CE patients. This was a retrospective observational study that recruited 514 volunteers, including 199 people with normal gallbladders (normal), 152 CL, and 163 CE patients. Discovery cohort was established to explore the difference in gut microbiota through 16S rRNA and metagenomics sequencing. Validation cohort aimed to verify the results through quantitative polymerase chain reaction (qPCR). Significant enrichment of Escherichia coli was found in patients with cholelithiasis or cholecystectomy both in the discovery cohort (16S rRNA sequencing, PNormal-CL=0.013, PNormal-CE=0.042; metagenomics sequencing, PNormal-CE=0.026) and validation cohort (PNormal-CL<0.0001, PNormal-CE<0.0001). Pks+ E.coli was found enriched in CL and CE patients through qPCR (in discovery cohort: PNormal-CE=0.018; in validation cohort: PNormal-CL<0.0001, PNormal-CE<0.0001). The differences in bile acid metabolism were found both through Tax4Fun analysis of 16S rRNA sequencing (Ko00120, primary bile acid biosynthesis, PNormal-CE=0.014; Ko00121, secondary bile acid biosynthesis, PNormal-CE=0.010) and through metagenomics sequencing (map 00121, PNormal-CE=0.026). The elevation of serum total bile acid of CE patients was also found in validation cohort (PNormal-CE<0.0001). The level of serum total bile acid was associated with the relative abundance of pks+ E.coli (r=0.1895, P=0.0012). E.coli, especially pks+ species, was enriched in CL and CE patients. Pks+ E.coli and bile acid metabolism were found associated with CRA and CRC in people after cholecystectomy.

Exploring PLGA-OH-CATH30 Microspheres for Oral Therapy of Escherichia coli-Induced Enteritis.

Antibiotic therapy effectively addresses Escherichia coli-induced enteric diseases, but its excessive utilization results in microbial imbalance and heightened resistance. This study evaluates the therapeutic efficacy of orally administered poly (lactic-co-glycolic acid) (PLGA)-loaded antimicrobial peptide OH-CATH30 microspheres in murine bacterial enteritis. Mice were categorized into the healthy control group (CG), untreated model group (MG), OH-CATH30 treatment group (OC), PLGA-OH-CATH30 treatment group (POC), and gentamicin sulfate treatment group (GS). Except for the control group, all other experimental groups underwent Escherichia coli-induced enteritis, followed by a 5-day treatment period. The evaluation encompassed clinical symptoms, intestinal morphology, blood parameters, inflammatory response, and gut microbiota. PLGA-OH-CATH30 microspheres significantly alleviated weight loss and intestinal damage while also reducing the infection-induced increase in spleen index. Furthermore, these microspheres normalized white blood cell count and neutrophil ratio, suppressed inflammatory factors (IL-1β, IL-6, and TNF-α), and elevated the anti-inflammatory factor IL-10. Analysis of 16S rRNA sequencing results demonstrated that microsphere treatment increased the abundance of beneficial bacteria, including Phocaeicola vulgatus, in the intestinal tract while concurrently decreasing the abundance of pathogenic bacteria, such as Escherichia. In conclusion, PLGA-OH-CATH30 microspheres have the potential to ameliorate intestinal damage and modulate the intestinal microbiota, making them a promising alternative to antibiotics for treating enteric diseases induced by Escherichia coli.

Effect of the Combination of Clostridium butyricum and Mycelium of Phellinus igniarius on Intestinal Microbiota and Serum Metabolites in DSS-Induced Colitis.

Clostridium butyricum (CB) and Phellinus igniarius (PI) have anti-inflammatory, immune regulation, anti-tumor, and other functions. This study aimed to explore the therapeutic effect of CB and mycelium of PI (MPI) alone and in combination on colitis mice induced by dextran sodium sulfate (DSS). Mice were randomly assigned to five groups: (1) control (CTRL), (2) DSS, (3) CB, (4) MPI, and (5) CB + MPI (CON). The weight of the mice was recorded daily during the experiment, and the length of the colon was measured on the last day of the experiment. The colons were collected for hematoxylin and eosin staining, colon contents were collected for intestinal flora analysis, and serum was collected for metabolite analysis. The results showed that compared with the DSS group, CB, MPI, and CON treatments inhibited the weight loss and colon length shortening caused by DSS, significantly increased the concentrations of interleukin (IL)-4, IL-10, and superoxide dismutase, and significantly decreased the concentrations of IL-6, tumor necrosis factor-α, and myeloperoxidase. Gene sequence analysis of 16S rRNA showed that CB, MPI, and CON treatments changed the composition and structure of intestinal microorganisms. Metabolome results showed that CB, MPI, and CON treatments changed serum metabolites in DSS-treated mice, including dodecenoylcarnitine, L-urobilinogen, and citric acid. In conclusion, CB, MPI, and CON treatments alleviated DSS-induced colitis in mice by regulating intestinal flora and metabolites, with the CON group having the best effect.

Dietary eugenol ameliorates long-term high-fat diet-induced skeletal muscle atrophy: mechanistic insights from integrated multi-omics.

Eugenol (EU), the major constituent of clove oil, possesses a range of bioactivities. Here, the therapeutic potential of oral EU for mitigating skeletal muscle wasting was investigated in a long-term high-fat diet (HFD)-induced obese mice model. Male C57BL/6J mice, aged six weeks, were assigned to either a chow or a HFD for 10 weeks. Subsequently, the weight-matched HFD-fed mice were allocated into two groups, receiving either 0.2% (w/w) EU supplementation or no supplementation for 14 weeks. Our findings revealed that EU supplementation enhanced grip strength, increased hanging duration, and augmented skeletal muscle mass. RNA sequencing analysis demonstrated that EU modified the gastrocnemius muscle transcriptomic profile, and the differentially expressed genes between HFD and EU groups were mainly involved in the HIF-1 signaling pathway, TCR signaling pathway, and cGMP-PKG signaling pathway, which is well-known to be related to skeletal muscle health. Untargeted metabolomics analysis further showed that EU supplementation significantly altered the nucleotide metabolism in the GAS muscle. Analysis of 16S rRNA sequencing demonstrated that EU supplementation ameliorated the gut dysbiosis caused by HFD. The alterations in gut microbiota induced by EU were significantly correlated with indexes related to skeletal muscle atrophy. The multi-omics analysis presented the robust interaction among the skeletal muscle transcriptome, metabolome, and gut microbiome altered by EU supplementation. Our results highlight the potential of EU in skeletal muscle atrophy intervention as a functional dietary supplement.

Emergence of pathogenic bacteria isolates from zea maize extract using 16s rRNA molecular sequencing protocol as a tools for microbial identification and characterization

The purpose of this research work is to determine the molecular identity of bacteria isolated from infected Zea maize using the 16s rRNA molecular sequencing protocol. The samples were obtained from Okeagbe, Akoko north-west local government in Ondo state with latitude and longitude of Okeagbe at 7.6450° N, and 5.7603° E respectively. Preparation of infected maize samples was cultured using the serial dilution method.. Confirmatory characterization of bacteria isolates using 16s rRNA (ribosomal RNA) sequencing procedures (purification, amplication, Sequencing, and DNA extraction) inclusive.The result shows the isolation of the bacteria isolates involved the culturing, inoculation, and plating of the isolate on a plated agar, the identification of the bacteria isolate includes the use of Gram staining, biochemical tests, and characterization using Bergey's manual and antibiotics Susceptibility Test. In Gram staining all bacteria isolates were positive except one, in the biochemical test most bacteria isolate was positive for sugar Fermentation and citrate test and all were negative for the Voges Proskauer test. In antibiotics Susceptibility test few were sensitive, most were susceptible to antibiotics used. With the use of the 16S rRNA and procedures (purification and application of product, Sequencing, and Extraction of DNA) the bacteria isolate were identified and characterized. The phylogenetic analysis and molecular identification of 16S rDNA sequencing revealed that Escherichia coli, Samonella enterica and Staphylococcus aureus were found to infect maize. Molecular characterization based on 16S rRNA Gene sequencing confirms the identity of bacteria. The conventional procedure shows the presence of different arrays of microorganisms in the infected maize, microbes identified are Bacillus subtilis, Bacillus anthracis, Micrococcus luteus, Clostridium sporogenes, Microbacterium lacticum, Clostridium sporogenes, Lactobacillus casei and Micrococcus luteus. The phylogenetic analysis and molecular identification of 16s rRNA sequencing revealed that Escherichia coli, Samonella enterica and Staphylococcus aureus were found to infect maize in Band fragment Base pair 1500bp. In conclusion, the hearsay that maize can only be infected by fungi, it was observed that the possibility of being infected with pathogenic bacteria is imminent. The bottom line is, there should be proper surveillance and food safety in our farm, market and food store, to prevent and totally eradicate emergence of pathogenic organism in our food item.

Efficient nitrite accumulation in partial sulfide autotrophic denitrification (PSAD) system: insights of S/N ratio, pH and temperature

ABSTRACT To provide the necessary nitrite for the Anaerobic Ammonium Oxidation (ANAMMOX) process, the effect of nitrite accumulation in the partial sulfide autotrophic denitrification (PSAD) process was investigated using an SBR reactor. The results revealed that the effectiveness of nitrate removal was unsatisfactory when the S/N ratio (mol/mol) fell below 0.6. The optimal conditions for nitrate removal and nitrite accumulation were achieved within the S/N ratio range of 0.7-0.8, resulting in an average Nitrate Removal Efficiency (NRE) of 95.84%±4.89% and a Nitrite Accumulation Rate (NAR) of 75.31%±6.61%, respectively. It was observed that the nitrate reduction rate was three times faster than that of nitrite reduction during a typical cycle test. Furthermore, batch tests were conducted to assess the influence of pH and temperature conditions. In the pH tests, it became evident that the PSAD process performed more effectively in alkaline environment. The highest levels of nitrate removal and nitrite accumulation were achieved at an initial pH of 8.5, resulting in a NRE of 98.30%±1.93% and a NAR of 85.83%±0.47%, respectively. In the temperature tests, the most favourable outcomes for nitrate removal and nitrite accumulation were observed at 22±1 ℃, with a NRE of 100.00% and a NAR of 81.03%±1.64%, respectively. Moreover, a comparative analysis of 16S rRNA sequencing results between the raw sludge and the sulfide-enriched culture sludge sample showed that Proteobacteria (49.51%) remained the dominant phylum, with Thiobacillus (24.72%), Prosthecobacter (2.55%), Brevundimonas (2.31%) and Ignavibacterium (2.04%) emerging as the dominant genera, assuming the good nitrogen performance of the system.

Therapeutic Effects of Zymomonas mobilis on Experimental DSS-Induced Colitis Mouse Model.

Zymomonas mobilis, a Gram-negative bacteria observed in some popular beverages, is considered safe and has been studied for its potential therapeutic benefits. In this study, we explored its effects on the inflammatory process, tissue integrity, differential gene expression, and microbiota composition in an experimental dextran sulfate sodium (DSS)-induced colitis model in mice. As a result, Z. mobilis alleviated the symptoms caused by DSS administration, as indicated by reduced weight loss, disease activity index, a significant reduction in the colon weight/length ratio, and histopathological improvement. Also, Z. mobilis could restore the mucosal barrier as well as increase the expression of Muc3 and Ocln genes. An analysis of 16S rRNA sequences showed that Z. mobilis alters gut microbiota, increasing Akkermansia muciniphila abundance and decreasing Escherichia coli. Furthermore, Z. mobilis seems to be involved in potentiating a regulatory phenotype by inducing immunomodulatory genes like Tgfb, Il5, Il10, and Foxp3 and reducing the relative mRNA expression of proinflammatory cytokines TNF, Il6, and Il17. Our data suggest that Z. mobilis could alleviate disease progression and be considered a possible probiotic adjuvant for pathologies of the bowel.

Dietary bile acids supplementation improves the growth performance and alleviates fatty liver in broilers fed a high-fat diet via improving the gut microbiota

This experiment aims to evaluate the effect of bile acids (BAs) in alleviating fatty liver disease induced by a high-fat diet (HFD) in broilers, and the modulation of the gut microbiota involved in this process. A total of 192 one-day-old Arbor Acres (AA) commercial male broilers were randomly divided into 4 groups and treated with the following diet: a basal-fat diet (BFD), a basal-fat diet plus bile acids (BFD + BAs), an HFD, and a high-fat diet plus bile acids (HFD + BAs). Bile acids were supplemented at the early growth stage (3-7 d), middle stage (17-21 d), and late stage (31-35 d). Results showed that BAs treatment had a significant effect on body weight on 14 d and 35 d, and increased the breast muscle weight and its index, but decreased the liver weight and abdominal fat weight on 35 d (P < 0.05). The supplementation of BAs significantly improved the serum lipid profile and decreased the level of triglycerides (TG), total cholesterol (TCHO), and nonesterified fatty acids (NEFA) on 35 d (P < 0.05). Dietary BAs supplementation significantly alleviated the hepatic TG deposition induced by HFD (P < 0.05), which was accompanied by upregulation of peroxisome proliferator-activated receptor gamma (PPARγ) and lipoprotein lipase (LPL) gene expression (P < 0.05). Moreover, the expression levels of hepatic gene adipose triglyceride lipase (ATGL), peroxisome proliferator-activated receptor α (PPARα), and apolipoprotein B (APOB) were greatly increased by BAs treatment. The analysis of 16S rRNA sequencing showed that the microbial diversity of the cecal digesta was increased by BAs in broilers with elevated abundances of Firmicutes, Lactobacillus, Anaerostipes, Sellimonas, and CHKCI002 and decreased abundances of Barnesiella and Akkermansia genus (P < 0.05). Hepatic TG content was positively correlated with the abundance of Oscillospiraceae, but it was negatively correlated with the abundance of Lactobacillus in cecal digesta (P < 0.05). These results indicate that dietary BAs can improve growth performance and alleviate fatty liver disease induced by an HFD via modulating gut microbiota in broilers.

The Atlantic salmon microbiome infected with the sea louse Caligus rogercresseyi reveals tissue-specific functional dysbiosis

The microbiota plays functional roles in almost all living organisms, influencing metabolism, defense, and nutrition. Conventionally, parasitism is defined as a bipartite interaction between host and parasites, where both organisms display molecular responses mainly associated with the immune system. However, the microbial community is another relevant factor that can interact during the parasite infection in fish farming. Therefore, this study aimed to characterize the Atlantic salmon microbiota's dynamics, correlation, and functional potential in skin and gut tissues during the sea louse Caligus rogercresseyi parasitism. The findings showed that the parasitism reduced the alpha and beta diversity in infested salmon's microbiota. This reduction favored the tissue-specific occurrence of opportunistic bacterial pathogens. Notably, Photobacterium was identified primarily on the skin, and Vibrio was predominant in the gut microbiota of infested individuals. Cluster analysis of full-length 16S rRNA sequencing suggested that C. rogercresseyi could be a vector for transmitting Tenacibaculum and Vibrio species. Network analysis evidenced that parasitism also perturbs the correlation of the skin and gut microbiota, revealing fewer associations in infested individuals. Finally, a contrasting effect on the functional potential of the skin and gut microbiome was found. Herein, the parasitism increased the biosynthetic processes and reduced degradation pathways in the skin; meanwhile, the opposite trend was observed in the gut microbiome. Overall, our results evidenced that sea lice parasitism induces tissue-specific functional dysbiosis and provides novel evidence about the harmful effects of sea lice in salmon aquaculture.

Role of an antagonistic bacterium, Bacillus subtilis PE7, in growth promotion of netted melon (Cucumis melo L. var. reticulatus Naud.).

The aim of this study was to determine the plant growth-promoting effect of Bacillus subtilis PE7 on growth of melon plants. B. subtilis PE7 isolated from kimchi was identified based on colonial and microscopic morphology along with analyses of 16S rRNA and pycA gene sequences. Strain PE7 showed different levels of inhibition on phytopathogens and was able to grow at variable temperatures and pH values. Strain PE7 had the ability to produce siderophores, indole-3-acetic acid (IAA), ammonia, exopolysaccharides, and 1-aminocyclopropane-1-carboxylic acid deaminase, as well as solubilize insoluble phosphate and zinc. The IAA secretion of strain PE7 showed a concentration-dependent pattern based on the concentration of l-tryptophan supplemented in the fertilizer-based culture medium. The LC-MS analysis indicates the presence of IAA in the culture filtrate of strain PE7. Treatment of the B. subtilis PE7 culture containing different metabolites, mainly IAA, significantly promoted melon growth in terms of higher growth parameters and greater plant nutrient contents compared to treatments with the culture without IAA, fertilizer, and water. The cells of B. subtilis PE7 attached to and firmly colonized the roots of the bacterized melon plants. Based on our results, B. subtilis PE7 can be utilized as a potential microbial fertilizer to substitute chemical fertilizers in sustainable agriculture.

An ecological explanation for carbon source-associated denitrification performance in wastewater treatment plants

The underlying mechanism associated with the roles of dosed carbon source in denitrification performance remains largely unknown. In this study, three denitrifying consortia (DNC) were constructed via evolutionary top-down enrichment method with well-defined conditions and specific carbon sources (acetate, glucose and their mixture). The reactor operation shows that nearly complete nitrate removal was achieved; however, the glucose feeding resulted in much higher concentrations of biomass and non-settable flocs. The 16S rRNA sequencing suggests that the bacterial diversity of the acetate-fed DNC was significantly higher than those of acetate/glucose-fed and glucose-fed DNCs. The dentrifying population in the acetate-fed DNC was dominated by Propionivibrio (16.1 %) and Thauera (3.4 %); whereas those of acetate/glucose- and glucose-fed DNCs were dominated by Pleomorphomonas (21.5 % and 26.3 %, respectively). Interestingly, the supernatant of acetate-fed DNC contained a high abundance of genera Thauera (averaged at 85.1 %), indicating the free-living nature of Thauera. Both PICURSt2 analysis of 16S rRNA sequencing and metagenomic analysis indicate that the acetate-fed DNC contained higher abundances of denitrifying genes; the acetate/glucose-fed and glucose-fed DNCs, in comparison, enriched genes related to glucose transportation and metabolism. Additionally, the acetate-fed DNC had better network stability than other two groups. This study adds important knowledge regarding the ecological traits of DNC, providing important clues for rational addition of carbon sources in wastewater treatment plants.

Co-infections of Aeromonas dhakensis and Chryseobacterium indologenes in largemouth bass (Micropterus salmoides)

As one of the most important economic fishes, largemouth bass (Micropterus salmoides) is being widely farmed around the world. With the improper expansion and high-density culture, largemouth bass is susceptible to an increasing number of pathogens. Particularly, co-infection of multiple pathogens is ubiquitous, which has become a main obstacle to the development of largemouth bass industry. In the study, a co-infected case caused by Aeromonas dhakensis (A. dhakensis) and Chryseobacterium indologenes (C. indologenes) in largemouth bass was firstly reported. Diseased fish with clinical signs such as exophthalmia, hemorrhage on gill cover, rot or ulceration on gill, skin and fin were collected for further diagnostics. Tissue lesions of liver, spleen and kidney were observed, such as necrosis, hemorrhage, vacuolization and disintegration. Two bacteria strains were isolated from liver, one appeared as a white or yellowish colony, and the other was characterized by golden yellow color. Based on the conjoint analysis of 16S rRNA and gyrB gene sequences, the isolated strains were identified as A. dhakensis and C. indologenes, respectively. Antibiotic sensitivity testing indicated that both A. dhakensis and C. indologenes were sensitive to flumequine, enrofloxacin, ceftazidime and minocycline, and resistant to penicillin, sulfamethoxine sodium, ampicillin and cefamezin. Largemouth bass showed similar clinical signs and tissue lesions to naturally diseased fish following co-infection with the isolated strains, indicating that the isolated strains fulfill Koch's postulates. Besides, a slightly increased pathogenicity was observed following co-infection compared with single infection. The data firstly reveal a co-infected case caused by A. dhakensis and C. indologenes in largemouth bass, not only enriching the relevant information of largemouth bass diseases, but also promoting the diagnosis, prevention and control of A. dhakensis and C. indologenes co-infection.

Novel insights into perfluorinated compound-induced hepatotoxicity: Chronic dietary restriction exacerbates the effects of PFBS on hepatic lipid metabolism in mice

Perfluorobutane sulfonates (PFBS) have garnered extensive utilization because of their distinctive physicochemical properties. The liver acts as a key target organ for toxicity within the body and is vital for regulating metabolic processes, particularly lipid metabolism. However, there is currently a significant research gap regarding the influences of PFBS on hepatic lipid metabolism, especially in individuals with different dietary statuses. Here, the objective of this research was to examine the effects of PFBS on hepatic function under different dietary conditions. The results suggested that the levels of liver injury biomarkers were significantly upregulated, e.g., transaminase (GPT, GOT), while liver lipid levels were downregulated after exposure to PFBS at concentration of 50 μg/L for 42 days. Moreover, restricted diet further intensified the adverse effects of PFBS on the liver. Metabolomics analysis identified significant alterations in lipid-related metabolites in PFBS-induced hepatotoxicity, PFBS exposure induced a decrease in lysophosphatidylethanolamine and lysophosphatidylcholine. PFBS exposure caused an increase in aldosterone and prostaglandin f2alpha under restricted diet. In PFBS treatment group, histidine metabolism, beta-alanine metabolism, and arginine biosynthesis were the main pathway for PFBS toxicity. Aldosterone-regulated sodium reabsorption as a vital factor in inducing PFBS toxicity in the RD-PFBS treatment group. The analysis of 16S rRNA sequencing revealed that exposure to PFBS resulted in imbalance of gut microbial communities. PFBS exposure induced a decrease in Akkermansia and Lactobacillus, but an increase in Enterococcus. PFBS exposure caused the abundance of Lachnospiraceae_NK4A136_group was significantly elevated under restricted diet. Additionally, disruptions in the expression of genes involved in lipid production and consumption may significantly contribute to lipid imbalance in the liver. This study underscores the importance of recognizing the harmful impact of PFBS on liver function, along with the biotoxicity of contaminant influenced by dietary habits.

SAT046 Canagliflozin Leads To Less Weight Gain, Confers Changes In The Gut Microbiota And Extends Life Span In Male C57BL6 Mice

Abstract Disclosure: E.E. Habeos: None. F. Filippopoulou: None. M. Kanakis: None. G.I. Habeos: None. N. Belmokhtar: None. P. Stathopoulou: None. G. Tsiamis: None. S. Taraviras: None. G. Lagoumintzis: None. D.V. Chartoumpekis: None. Introduction: SGLT2 inhibitors (SGLT2i) are widely prescribed drugs for the treatment of diabetes. Their beneficial health effects appear to be pleiotropic and are not limited to the amelioration of glycemic profile. The SGLT2i-induced glycosuria is suggested to provoke a metabolic shift that mimics the fasting response. Since calorie restriction has been shown to enhance longevity in mice, we hypothesized that treating mice with SGLT2i would also increase their life span. Methods: To this end, 4-months old male C57BL6 mice were fed with a standard chow diet (Control, n=83) or a diet supplemented with 200 mg/kg canagliflozin (Cana, n=92) and were observed up to the natural end of their life. Results: The canagliflozin-treated mice gained less weight over time. For instance, at the age of 10 months, the body weight of control mice was higher (means ± SEM: 33.1 ± 0.26 g) than the body weight of their canagliflozin-treated counterparts (means ± SEM: 31.4± 0.21 g; p&amp;lt;0.01). The development and severity of age-related cataracts were also examined at the age of 14 months, but no statistically significant difference was found in the severity score between the 2 groups. The survival curves of the control and the Cana mice diverged significantly, with the median survival of the control being 107.5 weeks versus 112.5 weeks of the Cana group (p=0.011). As the gut microbiome is known to change with age and be associated with the development of age-related diseases, feces were collected from 17-month-old mice (n=10 per group) and analysis of fecal bacterial 16S rRNA sequences was performed. Differences in beta diversity (diversity of bacterial species across samples) and relative abundance of bacterial species were noted between the Control and the Cana groups. Conclusions-Discussion: Our study highlights the effect of SGLT2i on longevity in a widely used mouse model (C57BL6). The underlying mechanisms leading to the enhancement of longevity are not clear yet. Some microbiota changes associated with the SGLT2i treatment, as described in other mouse models with enhanced longevity, warrant further investigation and analysis. It would be useful to extend these studies employing various SGLT2i with different potencies and selectivity against SGLT so as to determine if the observed effect on longevity is independent of the type of the drug. Presentation: Saturday, June 17, 2023

Genomic evidence of genetic diversity and functional evolution in Flavobacterium columnare.

Flavobacterium columnare is the causative agent of columnaris disease in freshwater fish. Columnaris disease can cause heavy economic losses in aquaculture. In this study, whole-genome sequencing was used to characterize this pathogen. F. columnare isolate AH-01 had a circular chromosome and plasmid that encoded a total of 3,022 genes. Isolate GX-01 only had a circular chromosome and encoded 2,965 genes. Genomic islands, prophage regions, and CRISPR/Cas systems were identified in both genomes. Both genomes presented evidence of gene variation and horizontal transfer, both of which are the essential components of genetic diversity, genome plasticity, and functional evolution. Single-gene phylogeny and comparative genome analyses were performed to investigate the variation and evolution of this pathogen. Genetic analysis of 16S rRNA and housekeeping gene sequences significantly clustered 55 F. columnare isolates into four clades. The intragroup identity of the 16S rRNA gene exceeded 99%, while the intergroup identity was below the species delineation threshold. We discovered significant translocation, inversion, and rearrangement events that influenced local synteny within each group. Notably, the observed alignments varied considerably among all the studied groups. The core genomes of all strains with available sequences comprised 747 genes, corresponding to approximately 25% of the genome. Core genome multilocus sequence typing, genome-wide orthology and phylogenetic analyses, and average nucleotide identity suggested that the currently existing F. columnare was an assemblage of several distinct species, with levels of divergence at least equivalent to those between recognized bacterial species. The present investigation provided genomic evidence of gene variation and horizontal transfer, which were the basis of genetic diversity, genome plasticity, and functional evolution. The findings supported a proposed new taxonomic perspective on F. columnare.

Pseudomonas benzopyrenica sp. nov., isolated from soil, exhibiting high-efficiency degradation of benzo(a)pyrene.

A Gram-negative, yellow-pigmented, aerobic and rod-shaped bacterium, designated as strain BaP3T, was isolated from the soil. Strain BaP3T grew at 16-37℃ (optimum, 30 °C) and pH 6.0-8.0 (optimum, pH 7.0). Additionally, strain BaP3T could tolerate NaCl concentrations in the range 0-6 % (optimum, 1%). Moreover, strain BaP3T was motile by flagella. The phylogenetic analysis of 16S rRNA sequences showed that strain BaP3T belonged to the genus Pseudomonas, and the sequence was most closely related to Pseudomonas oryzihabitans CGMCC 1.3392T and Pseudomonas psychrotolerans DSM 15758T, with 99.66 % sequence similarity. Pseudomonas rhizoryzae RY24T was the next closely related species, exhibiting 99.38 % 16S rRNA gene sequence similarity. The DNA-DNA hybridization and average nucleotide identity values between strain BaP3T and its closely related types were below 50 and 92 %, respectively. Both results were below the cut-off for species distinction. The genomic DNA G+C content of strain BaP3T was 65.30 mol%. The predominant quinone in strain BaP3T was identified as ubiquinone Q-9. The major cellular fatty acids were summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c), summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c) and C16 : 0. These results indicated that strain BaP3T represents a novel species in the genus Pseudomonas. The type strain is BaP3T (CCTCC AB 2022379T=JCM 35914T), for which the name Pseudomonas benzopyrenica sp. nov. is proposed.

Characterization of rhizosphere Pseudomonas chlororaphis IRHB3 in the reduction of Fusarium root rot and promotion of soybean growth

Rhizosphere beneficial bacteria play an important role in plant health and disease control, and have been widely explored as biocontrol resources and agents for the ecological and sustainable management of soil-borne diseases. In our previous study, a bacterium strain IRHB3 was isolated from soybean rhizosphere under maize-soybean relay strip intercropping. To clarify the characterization of IRHB3 and its biocontrol effects of soybean root rot, we systemically investigated the taxonomy, biological characteristics, bacteriostatic activity, environmental stress tolerance of IRHB3, and also analyzed its effects on the control of root rot as well as growth promotion. Our results showed that IRHB3 as a Gram-negative bacterium was identified as Pseudomonas chlororaphis by multi-locus sequence analysis of 16S rRNA, gyrB, rpoD and gapA. The colony was characterized by yellowish circular and rugby-shaped with rough surface and gullies under scanning electron microscropy.. IRHB3 not only activated kinds of hydrolase such as protease, caseinase, and cellulase, but was also capable of dissolving phosphorus, releasing siderophores and producing IAA. IRHB3 suspension treatment with proper concentration significantly promoted seed germination and seedling growth. Antagonistic assay showed that IRHB3 displayed good inhibition effects on various pathogenic fungi, especially aggressive Fusarium species causing root rot, with the average inhibition rate as high as 69.89%. And meanwhile, introducing IRHB3 into soil effectively alleviated the damage of F. oxysporum to soybean root. Moreover, IRHB3 had strong environmental adaptability of wide pH, low nutrition and salt stress. In conclusion, these results indicate that Pseudomonas chlororaphis IRHB3 from intercropped soybean rhizosphere is one kind of typically plant growth-promoting rhizobacteria (PGPR) and has immense potential and strong environmental adapatability on the control of soil-borne fungal disease.