16S rRNA Gene Sequence Analysis Research Articles

Chitinophaga defluvii sp. nov., a Cyhalofop-Butyl-Degrading Bacterium Isolated from Municipal Sludge.

A yellow-colored, Gram-stain-negative, non-motile, and rod-shaped bacterium, designated strain H8T, was isolated from municipal sludge in Huaibei, China. Strain H8T was able to grow at 15-37°C (optimum at 30°C), pH 6.0-8.0 (optimum at pH 7.0), and 0-2.5% (w/v) NaCl concentration (optimum at 0%). This strain was taxonomically characterized by a polyphasic approach. Based on the 16S rRNA gene sequence analysis, strain H8T represented to the genus Chitinophaga and shared highest sequence similarities with C. arvensicola DSM 3695T (97.5%), C. niastensis JS16-4T (97.3%), C. hostae 2R12T (97.1%), and C. ginsengisegetis Gsoil 040T (96.8%). The 16S rRNA gene similarities with other members of the genus Chitinophaga are less than 96.3%. The only respiratory quinone was a menaquinone with seven isoprene units (MK-7); the major polar lipid was phosphatidylethanolamine; and the predominant fatty acids were iso-C15:0 and C16:1 ω5c. The genome size of strain H8T was 7.6Mb, with 44.3% G + C content. The DNA-DNA relatedness and the average nucleotide identity values among strain H8T and other relatives were all less than 19.7% and 72.6%, respectively, which fall below the threshold value of 70% and 95% for the strain to be considered as novel. The morphological, physiological, chemotaxonomic, and phylogenetic analyses clearly distinguished this strain from its closest phylogenetic neighbors. Thus, strain H8T represents a novel species of the genus Chitinophaga, for which the name Chitinophaga defluvii sp. nov. is proposed. The type strain is H8T (= CCTCC AB 2023228T = KCTC 102174T).

Personalized prediction of glycemic responses to food in women with diet-treated gestational diabetes: the role of the gut microbiota

We developed a prediction model for postprandial glycemic response (PPGR) in pregnant women, including those with diet-treated gestational diabetes mellitus (GDM) and healthy women, and explored the role of gut microbiota in improving prediction accuracy. The study involved 105 pregnant women (77 with GDM, 28 healthy), who underwent continuous glucose monitoring (CGM) for 7 days, provided food diaries, and gave stool samples for microbiome analysis. Machine learning models were created using CGM data, meal content, lifestyle factors, biochemical parameters, and microbiota data (16S rRNA gene sequence analysis). Adding microbiome data increased the explained variance in peak glycemic levels (GLUmax) from 34 to 42% and in incremental area under the glycemic curve (iAUC120) from 50 to 52%. The final model showed better correlation with measured PPGRs than one based only on carbohydrate count (r = 0.72 vs. r = 0.51 for iAUC120). Although microbiome features were important, their contribution to model performance was modest.

Dynamic Spread of Antibiotic Resistance Determinants by Conjugation to a Human-Derived Gut Microbiota in a Transplanted Mouse Model

Background. Antibiotic-resistant (AR) bacteria pose an increasing threat to public health, but the dynamics of antibiotic resistance gene (ARG) spread in complex microbial communities are poorly understood. Conjugation is a predominant direct cell-to-cell mechanism for the horizontal gene transfer (HGT) of ARGs. We hypothesized that commensal Escherichia coli donor strains would mediate the conjugative transfer of ARGs to phylogenetically distinct bacteria without antibiotic selection pressure in gastrointestinal tracts of mice carrying a human-derived microbiota with undetectable levels of E. coli. Our objective was to identify a mouse model to study the factors regulating AR transfer by conjugation in the gut. Methods. Two donor E. coli strains were engineered to carry chromosomally encoded red fluorescent protein, and an ARG- and green fluorescent protein (GFP)-encoding broad host range RP4 conjugative plasmid. Mice were orally gavaged with two donor strains (1) E. coli MG1655 or (2) human-derived mouse-adapted E. coli LM715-1 and their colonization assessed by culture over time. Fluorescence-activated cell sorting (FACS) and 16S rDNA sequencing were performed to trace plasmid spread to the microbiota. Results. E. coli LM715-1 colonized mice for ten days, while E. coli MG1655 was not recovered after 72 h. Bacterial cells from fecal samples on days 1 and 3 post inoculation were sorted by FACS. Samples from mice given donor E. coli LM715-1 showed an increase in cells expressing green but not red fluorescence compared to pre-inoculation samples. 16S rRNA gene sequencing analysis of FACS GFP positive cells showed that bacterial families Lachnospiraceae, Clostridiaceae, Pseudomonadaceae, Rhodanobacteraceae, Erysipelotrichaceae, Oscillospiraceae, and Butyricicoccaceae were the primary recipients of the RP4 plasmid. Conclusions. Results show this ARG-bearing conjugative RP4 plasmid spread to diverse human gut bacterial taxa within a live animal where they persisted. These fluorescent marker strategies and human-derived microbiota transplanted mice provided a tractable model for investigating the dynamic spread of ARGs within gut microbiota and could be applied rigorously to varied microbiotas to understand conditions facilitating their spread.

Lacrimispora sinapis sp. nov., isolated from pickled potherb mustard (Brassica juncea Coss.).

Strain JR3T was isolated from Chinese pickled potherb mustard (Brassica juncea Coss.) purchased from a local market in Shanghai, China. A polyphasic approach including 16S rRNA gene sequence analysis, average nucleotide identity (ANI) analysis, digital DNA-DNA hybridization (dDDH), determination of G+C content and various phenotypic analyses was employed to characterize strain JR3T. The bacterium was rod-shaped, Gram-stain-positive, terminal round endospore-forming and catalase-positive. The strain could grow at a wide range of temperatures (20-45 °C) and pH values (6.0-8.0). Optimal growth of strain JR3T occurred at 35-40 °C and a pH value of 7.0. The strain exhibited growth at salt (NaCl) concentrations of up to 3% (w/v). The G+C content of the genomic DNA was 44.0 mol%. The major fatty acids were C16 : 0, C19 : 0 c9, 10, summed feature 10 (C18 : 1 c11/t9/t6) and C18 : 1 c9. 16S rRNA gene sequencing revealed that strain JR3T represents a member of the genus Lacrimispora, and it has higher sequence similarity with Lacrimispora amygdalina BR-10T (=DSM 12857T) (98.72%), Lacrimispora saccharolyticum WM1T (98.29%) and Lacrimispora xylanolytica sy1 (98.22%). The dDDH value for strain JR3T and phylogenetically related species within the genus Lacrimispora ranged from 17.7% to 29.9%. The ANI of strain JR3T with its closely related taxa was far lower than the threshold (95%-96%) used for species differentiation. Results of phylogenetic, physiological and phenotypic characterization confirmed that strain JR3T represented a novel species within the genus Lacrimispora, for which the name Lacrimispora sinapis sp. nov. is proposed. The type strain is JR3T=CCTCC AB 2024044T=LMG 33655T.

Microbiome analysis reveals the potential mechanism of herbal sitz bath complementary therapy in accelerating postoperative recovery from perianal abscesses.

The herbal sitz bath formula, as a complementary therapy, effectively alleviates postoperative wound pain and accelerates healing time in patients with perianal abscesses. To investigate its mechanism of action, this study conducted 16S rRNA gene sequencing and bioinformatics analysis on wound exudate samples from patients after perianal abscess surgery. Patients were randomly divided into two groups: one receiving the herbal sitz bath as an adjunctive therapy and the other without this adjunctive therapy. Samples were collected on the first and eighth days after surgery to compare the differences in microbial community composition between the two groups on the eighth day and between the first and eighth days within each group. The study revealed that the herbal sitz bath significantly altered the structure of the microbial community, increasing its diversity and abundance. By reducing Enterococcus and increasing Bifidobacterium, Faecalibacterium, and Ruminococcus, the therapy enhanced the wound's anti-infective capacity and accelerated healing. This study explored the potential mechanism of the herbal sitz bath formula as an adjunctive therapy in promoting postoperative recovery from perianal abscesses, providing valuable data for further research on the role of microorganisms in wound care. These findings contribute to optimising postoperative treatment regimens and facilitating patient recovery.

New insights in the metabolic functions of freshwater sulfate reducing communities during steel corrosion by biophysicochemical, 16S rRNA gene sequence and metaproteomic analysis

New insights in the metabolic functions of freshwater sulfate reducing communities during steel corrosion by biophysicochemical, 16S rRNA gene sequence and metaproteomic analysis

Molecular diagnosis of infective endocarditis from culture-negative valve samples in a tertiary hospital in Iran.

The aim of this study was to investigate the prevalence of Tropheryma whipplei, Chlamydia psittaci, Chlamydia pneumoniae, Legionella, Brucella, and Francisella tularensis in valve samples from endocarditis patients using the real-time PCR method at a major referral heart hospital in Iran. In this study, 146 paraffin-embedded tissue samples from the heart valves of patients with clinical and pathological evidence of infective endocarditis (IE), who underwent heart valve replacement surgery between 2016 and 2020 at Tehran Heart Center were collected. After DNA extraction from paraffin-embedded valve tissue samples, they were surveyed for the presence of T. whipplei, C. psittaci, C. pneumoniae, Legionella, Brucella, and F. tularensis using quantitative real-time PCR (qPCR). The 16S rRNA gene sequence analysis was used for accurate species identification. Based on the molecular results, T. whipplei, Chlamydia spp., and Legionella spp. were detected in 10 (6.84%), 6 (4.1%), and 3 (2.05%) valve samples, respectively. In addition, one of six positive samples for Chlamydia spp. was identified as C. psittaci. No positive samples for F. tularensis and Brucella were found. In addition, all control valve samples were negative for all investigated pathogens. The findings suggest that specific bacterial species such as T. whipplei, Chlamydia spp., and Legionella spp. are associated with the development of IE. Considering the life-threatening nature of IE, it is critical for healthcare systems to prioritize the identification of its causative agents and develop targeted treatment strategies.IMPORTANCEInfective endocarditis (IE) is a serious and potentially life-threatening condition, and it is associated with significant morbidity, mortality, and complications, making it a major concern in both global and national healthcare systems. Late diagnosis and failure to receive appropriate treatment for patients with culture-negative endocarditis caused by hard-to-grow bacteria can lead to the death of patients. Unfortunately, in Iran, less attention is paid to the role of organisms that are difficult to cultivate in laboratory settings such as Tropheryma whipplei, Chlamydia psittaci, Chlamydia pneumoniae, Legionella, Brucella, and Francisella tularensis in causing culture-negative endocarditis, and these pathogens are overlooked by infectious, cardiologists, and health officials. This study underscores the need for special attention in the diagnosis of the agents of IE.

Neptunitalea lumnitzerae sp. nov. isolated from the phyllosphere of the mangrove Lumnitzera racemosa Willd.

A Gram-stain-negative, rod-shaped, non-motile, aerobic, light-yellow-pigmented bacterium, designated as strain Y10T, was isolated from Lumnitzera racemosa leaf in Iriomote island mangrove forests in Japan. The 16S rRNA gene sequence analysis revealed that the isolate Y10T was affiliated with the family Flavobacteriaceae, and the sequence showed the highest sequence identity to that of Neptunitalea chrysea NBRC 110019T (97.2%)and others with below 96% sequence identity. The draft genome analysis revealed that its genomic size is 3.85 Mbp with G + C content 34.1%. The average nucleotide identity, average amino acid identity, and digital DNA-DNA hybridization values between strain Y10T and Neptunitalea chrysea NBRC 110019T was 79.07%, 76.35% and 19.6%, respectively, which were below the thresholds for prokaryotic species delineation (95-96%, 85%, and 70%, respectively). MK-6 was the only menaquinone, and the major cellular fatty acids (> 10% of the total fatty acids) were iso-C15:0, iso-C17:0 3-OH, and summed feature 3 (comprised of C16:1ω7c-iso and C15:0 2-OH). Based on its distinct phylogenetic position and genotypic and phenotypic characteristics, the strain represents a novel species of the genus Neptunitalea, for which the name Neptunitalea lumnitzerae sp. nov. is proposed. The type strain is Y10T (= DSM 110280T = NBRC 115783T).

Kineococcus halophytocola sp. nov., Isolated from Leaves of Halophyte Sesuvium portulacastrum L.

An aerobic, Gram-stain-positive, motile, coccus-shaped actinomycete, designated strain LSe6-4T, was isolated from leaves of sea purslane (Sesuvium portulacastrum L.) in Thailand and subjected to a polyphasic taxonomic studies. Growth of the strain occurred at temperatures between 15 and 38°C, and with NaCl concentrations 0-13%. The 16S rRNA gene sequences analysis indicated that the strain was most closely related to Kineococcus endophyticus KLBMP 1274T (99.2%), and Kineococcus mangrovi L2-1-L1T (99.0%). The genome of strain LSe6-4T was estimated to be 4.22 Mbp in size, with DNA G+C content of 74.57%. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) relatedness values between strain LSe6-4T and the closely related species were less than the threshold values for identifying a novel species. Strain LSe6-4T showed morphological and chemotaxonomic characteristics of the genus Kineocoocus. The whole-cell hydrolysates contained meso-diaminopimelic acid as the diagnostic diamino acid, with arabinose, galactose, and ribose as whole-cell sugars. The predominant menaquinone was MK-9(H2). The polar lipid profile composed of phosphatidylglycerol, diphosphatidylglycerol, an unidentified phospholipid, three unidentified aminophospholipids, and an unidentified aminolipid. Major cellular fatty acids (> 10% of total fatty acids) were anteiso-C15:0 and iso-C14:0. It is evident from the combination of genotypic and phenotypic characteristics that strain LSe6-4T represents a novel species of the genus Kineococcus, for which the name Kineococcus halophytocola sp. nov. is proposed. The type strain is strain LSe6-4T (= TBRC 17798T = NBRC 116401T).

Spatial variations in the microbiota: comparative analysis of microbial composition and predicted functions across different intestinal segments and feces in donkeys.

Donkeys, as single-stomach herbivores, have a complex and diverse microbial community in their digestive tracts. The intestinal bacterial community is crucial for maintaining intestinal homeostasis, as well as the host's overall nutrition and health. However, research on donkey gut microbes is relatively limited, particularly regarding the microbial colonization patterns in different intestinal segments of adult donkeys. Therefore, this study examined the abundance and function of microbiota across various sites of the intestinal tract (duodenum, jejunum, ileum, cecum, colon) and feces of healthy adult Dezhou male donkeys using 16S rRNA gene sequencing and PICRUSt analysis. The results indicate that donkeys have a rich gut microbial diversity and a large microbial population. No significant differences in the indices of alpha diversity were observed among the donkey's duodenum, jejunum, ileum, cecum, colon, and feces. A Venn diagram analysis revealed the presence of both unique (Duodenum: 4645; Jejunum: 3586; Ileum: 4904; Cecum: 4253; Colon: 6135; Feces: 4885) and shared (339) ASVs among the different sections. A principal coordinate analysis (PCoA) revealed significant differences (R2 = 0.2076, p < 0.007) across the six intestinal segments of the donkeys. At the phylum level, Firmicutes (63.64%), Bacteroidetes (20.72%), Verrucomicrobiota (9.16%), Patescibacteria (1.95%), Spirochaetota (1.87%), Actinobacteriota (1.13%), and Proteobacteria (0.42%) were the dominant bacteria in all samples. The Wilcoxon rank-sum test revealed significant differences in the proportions of genera among different intestinal segments. Specific genera were significantly enriched in various segments: Lachnospiraceae_UCG-008 and Sphaerochaeta in the duodenum; Christensenellaceae_R-7_group and Bacillus in the jejunum; NK4A214_group and Alloprevotella in the ileum; UCG-005 and Lactobacillus in the cecum; Clostridium_sensu_stricto_1 and Chlamydia in the colon; and Rikenellaceae_RC9_gut_group and Prevotellaceae_UCG-004 in the feces. A PICRUSt2 functional prediction analysis indicated that carbohydrate metabolism, prokaryotic cellular communities, antimicrobial drug resistance, immune diseases, membrane transport, signal transduction, and transcription exhibited significant differences among the different intestinal segments. This study provided critical primary data on the differences in donkey gut microbiota and the synergistic effects between gut microbiota and host functions. These findings can be used to assess donkey health status, improve breeding, and develop microbial additives.

Nasal microbial diversity is associated with survival in piglets infected by a highly virulent PRRSV-1 strain

BackgroundPorcine reproductive and respiratory syndrome virus (PRRSV) is a major threat to swine industry worldwide, especially virulent variants arising during the last years, such as Spanish PRRSV-1 Rosalia strain. The role of the nasal microbiota in respiratory viral infections is still to be unveiled but may be promisingly related with the health status of the animals and thus, their susceptibility. The goal of this project was to study the nasal microbiota composition of piglets during a highly virulent PRRSV-1 outbreak comparing animals that died due to the infection with animals that survived it. The microbiota composition was inferred by V3–V4 regions of the 16S rRNA gene sequencing and bioinformatics analysis. To deepen the analysis, we added samples taken from piglets before the outbreak as well as from the sows giving birth to piglets under study.ResultsPiglets that survived the PRRSV-1 outbreak reported a more diverse and different nasal microbiota at three weeks of age compared to piglets dying, which was highly related with the litter of origin and the sow of the piglets. In addition, a high abundance of classical swine nasal colonizers belonging to genera such as Bergeyella, Glaesserella, Neisseria and Moraxella (among others), was related with good outcome. On the other hand, a dysbiotic community dominated by Escherichia and a different clade of Moraxella was found in piglets with bad outcome. Moreover, samples taken before the outbreak showed similar dynamics prior to virulent PRRSV-1 arrival, suggesting that microbiota-related susceptibility was already occurring in the animals and that the increase in mortality seen was related to the new highly virulent strain.ConclusionOur study suggests that the susceptibility to an infection such as PRRSV could be related to the nasal microbiota composition at the moment of infection and may serve as starting point to explore animal resilience. Since the dysbiosis detected as an initial response to infection may be not specific for this virus, further investigations should explore this phenomenon in the context of other viral infections.

Pig nasal and rectal microbiotas are involved in the antibody response to Glaesserella parasuis

Vaccination stands as one of the most sustainable and promising strategies to control infectious diseases in animal production. Nevertheless, the causes for antibody response variation among individuals are poorly understood. The animal microbiota has been shown to be involved in the correct development and function of the host immunity, including the antibody response. Here, we studied the nasal and rectal microbiota composition in association with the antibody response against the pathobiont Glaesserella parasuis. The nasal and rectal microbiotas of 24 piglets were sampled in two farms before vaccination and in one unvaccinated farm (naturally exposed to the pathobiont) at similar time. Microbiota composition was inferred by V3V4 16S rRNA gene sequencing and bioinformatics analysis, and the antibody response was quantified using the variation between the levels before and after vaccination (normalized per farm). Piglets with higher antibody responses showed more diverse nasal and rectal microbial communities compared to piglets with lower responses. Moreover, swine nasal core microbiota colonizers were associated with higher antibody levels, such as several members from Bacteroidales and Clostridiales orders and genera including Moraxella, Staphylococcus, Fusobacterium and Neisseria. Regarding taxa found in the rectal microbiota, associations with antibody responses were detected only at order level, pointing towards a positive role for Clostridiales while negative for Enterobacteriales. Altogether, these results suggest that the microbiota is associated with the antibody response to G. parasuis (and probably to other pathogens) and serves as starting point to understand the factors that contribute to immunization in pigs.

Whole Genome Sequence-Based Classification of Nonomuraea marmarensis sp. nov., Isolated from Island Soil

Actinomycetes are known to produce a vast array of bioactive secondary metabolites with potential therapeutic applications, including antimicrobials, anticancer agents, and enzyme inhibitors. Among these, members of the genus Nonomuraea have received much attention due to their broad ecological importance in nutrient cycling in soil and their ability to produce new bioactive compounds. A novel actinomycetes, designated strain M3C6T, was isolated from soil samples collected on Marmara Island, located in the Istanbul province, aiming to explore the microbial diversity of unexplored habitats, and characterized using a polyphasic approach. The isolate showed chemotaxonomic and morphological features consistent with members of the genus Nonomuraea. The 16S rRNA gene sequence analysis revealed that strain M3C6T shared the highest similarity, at 98.7% sequence identity, to Nonomuraea basaltis 160415T and Nonomuraea turkmeniaca DSM 43926T. However, the ANI and dDDH values between strain M3C6T and these reference strains were fairly low, ranging from 84.0 to 84.6% and 31.8 to 33.7%, respectively, below the generally accepted cutoffs for ANI and DDH that delineate different prokaryotic species. Genomic analysis of strain M3C6T showed that it had a genome size of 10.38 Mbp and a DNA G+C content of 69.5 mol%. Based on these chemotaxonomic, phenotypic, and genomic data, strain M3C6T is classified as a novel species within the genus Nonomuraea, for which the name Nonomuraea marmarensis sp. nov. is proposed. The type strain is M3C6T (= KCTC 49983T = CGMCC 4.8035T). Genomic analyses confirmed the high potential of M3C6T to produce specialized secondary metabolites.

Exploring Dietary Composition in an Invasive Apple Snail From Different Habitats Combining With Intestinal Microbiota and Metabolomics.

Pomacea canaliculata is recognized as a globally invasive aquatic species. Analyses of intestinal microbiota, dietary composition, and metabolism of invasive species can enhance our understanding of their feeding strategies and physiological adaptation strategies to the environment. Intestinal content samples were collected from P. canaliculata inhabiting three distinct environments including a pond, a river, and a ditch. These samples were subjected to 16S rRNA gene sequencing analysis and multiple metabarcoding analyses, including eukaryotic 18S rRNA, mitochondrial cytochrome c oxidase I (COI), and chloroplast rbcL genes. In addition, metabolomics analysis was conducted on the intestinal content samples to investigate metabolic change. The highest dietary diversity in P. canaliculata was observed in the ditch, and females exhibited a higher dietary diversity than males in the pond. The 18S rRNA gene has a high potential for identifying the dietary components of omnivorous species. The intestinal microbiota of P. canaliculata from different habitats displayed significant variations, attributed to differences in food resources and other environmental factors. Bacteria in the aquatic environment had minimal impact on the intestinal microbiota of P. canaliculata. Overall, P. canaliculata exhibited adaptive changes in physiological characteristics across different habitats, including alterations in diet, which, in turn, influence microbiota and metabolic pathways such as amino acid biosynthesis in the intestine. The present study investigated the physiological mechanisms that enable P. canaliculata to adapt to diverse habitats, considering various factors including diet, which is important for comprehending its invasive potential and the subsequent threats it poses to aquatic ecosystems.

Integrating 16S rRNA Gene Sequencing and Metabolomics Analysis to Reveal the Mechanism of L-Proline in Preventing Autism-like Behavior in Mice.

Autism spectrum disorder (ASD) is characterized by impaired social interaction and repetitive stereotyped behavior. Effective interventions for the core autistic symptoms are currently limited. This study employed a valproic acid (VPA)-induced mouse model of ASD to assess the preventative effects of L-proline supplementation on ASD-like behaviors. The method of 16S rRNA sequencing and untargeted metabolomics analyses were conducted to investigate the modulation of gut microbiota and gut metabolites by L-proline. The results indicated that L-proline supplementation significantly prevented ASD-like behavioral disorders, including alleviating social communication deficits and reducing repetitive behavior in the ASD mice. The 16S rRNA sequencing analysis revealed that L-proline regulated the composition and structure of gut microbiota. L-Proline supplementation enhances the abundance of the Verrucomicrobia at the phylum level and the Akkermansia at the genus level, while concurrently reducing the abundance of the Patescibacteria at the phylum level, as well as the Ileibacterium, Candidatus_Saccharimonas, and Lachnospiraceae_UCG-006 at the genus level in the VPA-induced mouse model for ASD. Additionally, the untargeted metabolomics results indicated that L-proline also modified the gut metabolite profiles. Functional analysis of the gut microbiota and KEGG pathway enrichment analysis of differential metabolites between the L-proline-supplemented and VPA groups corroborated that L-proline decreased pathways related to nucleotide metabolism, taurine and hypotaurine metabolism, and pyruvate metabolism, while increasing pathways involved in alpha-linolenic acid metabolism and phenylalanine, tyrosine, and tryptophan biosynthesis. The integrative metabolomic and microbiome analyses showed strong connections between the gut metabolites and gut microbiota affected by L-proline. These findings suggest that the modulatory effects of L-proline on gut microbiota and its metabolites may play a crucial role in preventing autism in mice. These findings suggest that dietary L-proline may represent a viable, effective option for preventing the physiological and behavioral deficits associated with ASD in mice.

Unveiling Microbiota Profiles in Saliva and Pancreatic Tissues of Patients with Pancreatic Cancer.

The pancreas, previously considered a sterile organ, has recently been shown to harbor its own microbiota that may influence tumor biology and patient outcomes. Despite increasing interest in the impact of the microbiome on cancer, the relationship between pancreatic tissue and oral microbiomes in pancreatic ductal adenocarcinoma (PDAC) remains limited. In this study, the oral and pancreas tissue microbiomes of patients with PDAC were compared to patients with other periampullary cancers (DC/AC) and a healthy control group using 16S rRNA gene sequence analysis. The results showed a significant reduction in microbial diversity in the saliva of cancer patients compared to healthy controls, while the PDAC patients exhibited a distinct microbial profile in their pancreatic tissues, consisting predominantly of Firmicutes, Proteobacteria, and Actinobacter, after filtering the microbiome of the indoor environment. Notably, the presence of oral bacteria such as Anoxybacillus, Clostridium, and Bacillus in pancreatic tissues suggests potential translocation from the oral cavity. This study emphasizes the importance of understanding the role of body fluid and tissue microbiota in pancreatic cancer, proposing that oral dysbiosis may contribute to disease progression. Moreover, the results suggest that the microbiome of the indoor environment in which samples are collected and analyzed is also important in microbiota analysis studies.

Characterization and Enzymatic Assay of Cellulase-Producing Probiotic Bacteria Isolated from Traditional Fermented Bamboo of Bangladesh

Fermented bamboo (also known as Ngwopo) is a traditional food item among the indigenous population living in the south-east of Bangladesh. Though fermented foods harbour a significant number of probiotics and enzyme-producing bacteria, there is a lack of research on isolating beneficial bacteria from native fermented food products in Bangladesh. Thus, considering the extensive utilisation of cellulase enzymes and probiotics, our study was conducted with the objective of isolating and characterizing native cellulase-producing bacteria from fermented bamboo and performing crude enzyme activity assay. The sample was collected from Khagrachari hill tracts district of Bangladesh. Three isolated bacteria, namely FB-1, FB-2, and FB-3, screened based on the carboxymethyl cellulose medium, were tested for biochemical and probiotic properties, and the best isolate was genetically identified by molecular characterization through 16S rRNA gene sequencing and evolutionary analysis. All three gram-positive isolates were observed to be positive for starch hydrolysis and fermentation of sucrose, suggesting their ability to breakdown carbohydrates while testing negative for gelatinase, indole, and H2S test. The isolates demonstrated moderate bile salt and pH tolerance and low to medium autoaggregation. FB-2 had 75% susceptibility to the tested antibiotics, while other isolates were susceptible to 50% antibiotics. All of the bacteria were non-pathogenic, showing γ-haemolysis. FB-2 strain showed an enzyme production capacity of 1.56 units/ml. The best performing isolate was identified as Bacillus cereus. Therefore, the study discovered a native bacteria potentially suitable as probiotic and as a source of cellulase for industrial processes, highlighting the enriched indigenous cuisines of Bangladesh.

Azotobacter biodiversity in Egypt using microbiological, biochemical, and molecular-biology multidisciplinary approach.

The presence of Azotobacter bacteria in the soil plays an important role in increasing its fertility and enhancing plant health. Azotobacter diversity depends on several environmental factors, particularly soil texture, pH, and nutrient content. The current study investigated the diversity of Azotobacter in various soil samples collected from 10 different governorates along the river Nile valley and its delta, Northern Mediterranean shore, Sinai, and Upper Egypt regions. The sampling sites spanned different environmental and ecological conditions of the Egyptian land either cultivated (agricultural land) or uncultivated (desert land). Fifty Azotobacter isolates were isolated and characterized based on cell morphology, culture properties, physiological, biochemical, and molecular characteristics. In addition, the alginate production capacity of the isolates was investigated. The results indicated that Egyptian soils are rich in Azotobacter diversity. The isolates were Gram-negative short rods, appearing either as single cells or in diploid structures. The isolates showed high variability in alginate production where two isolates (BH3 and AST4) were the highest alginate producers (3.12 and 4.22g alginate L- 1), respectively. 16S-rDNA sequencing and 16S-rDNA RFLP analyses indicated that despite the presence of Azotobacter salinestris and Azotobacter vinelandii in the Egyptian soil, Azotobacter chroococcum was the predominant species. In addition, sequence analysis of the gene coding for the transcription factor AlgU confirmed the results of 16S-rRNA gene sequence analysis. RAPD-REP and BOX-PCR were used to study the polymorphism among the isolates. High levels of microbial diversity were found using these DNA primers as 6-9 fingerprinting profiles were retrieved.

Pseudomonas retamae sp. nov., a novel endophytic bacterium with plant growth-promoting potential, isolated from root nodules of Retama monosperma in Northwestern Algeria.

A thorough polyphasic taxonomic study, integrating genome-based taxonomic approaches, was carried out to characterize the RB5T strain isolated from root nodules of Retama monosperma growing on the coastal dunes of Bousfer Beach (Oran, Algeria). The 16S rRNA gene sequence analysis revealed that strain RB5T had the highest similarity to Pseudomonas granadensis LMG27940T (98.94%) and Pseudomonas gozinkensis IzPS32dT (98.73%). Phylogenetic studies, including both 16S rRNA gene sequence and multilocus sequence analysis using 16S rRNA, gyrB and rpoD housekeeping genes, positioned RB5T in a distinct branch alongside its closest relative, P. granadensis LMG27940T. Phylogenomic analysis using the Bac120 marker set and Type (Strain) Genome Server confirmed the unique position of RB5T and its close relationship with P. granadensis LMG27940T. Similarly, genomic comparisons using average nucleotide identity based on blast (ANIb) and digital DNA-DNA hybridization (dDDH) revealed values of 92.85 and 59.3%, respectively, when compared with its closest relative, P. granadensis LMG27940T. Both values fall below the established species delimitation thresholds of 95-96% for ANIb and 70% for dDDH, providing strong genomic evidence that strain RB5T represents a novel species. Further average nucleotide identity comparisons with unclassified Pseudomonas spp. (384 genomes) and metagenomic-derived genomes from the Genome Taxonomy Database (GTDB) showed values between 84.27 and 89.2%, indicating that strain RB5T belongs to a unique evolutionary line. The genome of RB5T, with a size of 6 311 310 bp and a G+C content of 60%, harbours several key genes associated with plant growth-promoting traits, making it a promising candidate for sustainable agriculture. Phenotypically, RB5T strain is an aerobic, rod-shaped, Gram-negative, non-spore-forming bacterium that is motile with a single polar flagellum. It grows under a wide range of temperature (4-42 °C) and pH (5-10) conditions and tolerates up to 6% (w/v) NaCl. The main cellular fatty acid composition of RB5T includes C16:0, C17:0 cyclo and the summed features 3 consisting of C16:1 ω7c/C16:1 ω6c. Based on the phylogenetic, phenotypic, chemotaxonomic and genome comparison analyses, strain RB5T was identified as a novel species of the genus Pseudomonas, for which the name Pseudomonas retamae sp. nov. is proposed. The type strain is RB5T (=DSM 117471T=LMG 33633T=CIP 112482T).

Phylogenetic characterization of Bifidobacterium kimbladii sp. nov., a novel species from the honey stomach of the honeybee Apis mellifera.

Six novel Bifidobacterium strains H1HS16NT, Bin2N, Hma3N, H6bp22N, H1HS10N, and H6bp9N, were isolated from the honey stomach of Apis mellifera. Cells are Gram-positive, non-motile, non-sporulating, facultatively anaerobic, and fructose 6-phosphate phosphoketolase-positive. Optimal growth conditions occur at 37°C in anaerobiosis in MRS medium added with 2% fructose and 0.1% L-cysteine. The 16S rRNA gene sequences analysis revealed clustering with Bifidobacterium species found in honeybees. Strains Hma3N, H6bp22N, and H1HS16NT showed significant similarity to Bifidobacterium polysaccharolyticum JCM 34588T, with an average similarity of 99.63%. In contrast, strains Bin2N, H1HS10N, and H6bp9N were closely related to Bifidobacterium apousia JCM 34587T, with an average similarity of 99.22%. Moreover, strains Hma3N and H6bp22N exhibited ANI values of 96.65% and 96.53% when compared to Bifidobacterium polysaccharolyticum JCM 34588T, while strains H1HS16NT, Bin2N, H6bp9N, and H1HS10N revealed ANI values of 94.18%, 94.33%, 94.22%, and 95.50% respectively when compared to B. apousia JCM 34587T. dDDH analysis confirmed that strains Hma3N and H6bp22N belong to B. polysaccharolyticum, whereas strains H1HS16NT, Bin2N, H6bp9N, and H1HS10N represent a novel species. The peptidoglycan of the novel species is of the A4α type (L-Lys-D-Asp). The main cellular fatty acids of the type strain H1HS16NT are C16:0, C14:0, C19:0 cyclo ω9c, and C18:1 ω9c. The DNA G+C content of the type strain is 60.8mol%. Genome analyses of the strains were also conducted to determine their biosynthesis-related gene clusters, probiotic features, and ecological distribution patterns. Phenotypic and genotypic characterization show that strain H1HS16NT is distinct from the type strains of other recognized Bifidobacterium species. Thus, Bifidobacterium kimbladii sp. nov. (H1HS16NT=DSM 115187T=CCUG 76695T) is proposed as a novel Bifidobacterium species.