16S RNA Research Articles

Alterations and potential roles of microbial population of pregnant mouse saliva and amniotic fluid.

Prenatal exposure to intrauterine inflammation (IUI) is a crucial event in PTB pathophysiology. However, the relationship between microflora and PTB is not fully elucidated. In this study, we established an intrauterine inflammation mouse model via LPS intrauterine injection. The saliva and amniotic fluid were collected for 16s RNA gene sequencing. The levels of TNF-α and IL-1β in mouse amniotic fluid were determined by ELISA assays. Up to 60% of the operational taxonomic units (OTUs) in the saliva and amniotic fluid of PBS-treated mice were overlapped. LPS treatment-induced changes in the abundance of oral and amniotic fluid microorganisms. Both immune-associated probiotics, salivarius and mastitidis, were still detected in saliva (at significantly increased levels) after LPS-induced intrauterine inflammation and almost no probiotics of any type were detected in amniotic fluid, suggesting that the uterine cavity seems to be more susceptible to LPS compared to the oral cavity. Moreover, the abundance of pathogenic bacteria Escherichia coli was increased in both saliva and amniotic fluid after LPS treatment. The level of TNF-α and IL-1β in amniotic fluid is positively related to the amniotic fluid E. coli abundance. The microbial composition of saliva and amniotic fluid of pregnant mice was similar. LPS-induced intrauterine inflammation decreased the consistency of microbial composition in mouse saliva and amniotic fluid, increased the abundance of E. coli in saliva and amniotic fluid, and decreased the abundance of immune-associated probiotics, especially in amniotic fluid.

Tracking Microbial Diversity and Hygienic-Sanitary Status during Processing of Farmed Rainbow Trout (Oncorhynchus mykiss).

Aquaculture is becoming a strategic sector for many national economies to supply the increasing demand for fish from consumers. Fish culture conditions and processing operations can lead to an increase in microbial contamination of farmed fish that may shorten the shelf-life of fish products and byproducts, and ready-to-eat fishery products. The objective of this study was to evaluate the hygienic-sanitary status of water, environment, and processing of fresh-farmed rainbow trout (Oncorhynchus mykiss) fillets produced in a local fish farm in Andalusia, Spain. To achieve this, a longitudinal study was carried out by collecting environmental (air and food-contact surfaces), water from fish ponds, and rainbow trout samples. Thereby, seven sampling visits were performed between February 2021 and July 2022, where foodborne pathogens and spoilage microorganisms, together with physicochemical parameters, were analysed in the collected samples. Further, microbial identification of microbiota was achieved through a culture-dependent technique using blast analysis of 16S RNA gene sequencing. The results showed that Listeria monocytogenes and Salmonella were not detected in the analysed samples. Regarding the hygienic-sanitary status of the fish farm, the slaughtering bath, the eviscerating machine and the outlet water from fish ponds presented the highest counts of coliforms, Enterobacteriaceae, and Aerobic Mesophilic Bacteria. Staphylococcus aureus and sulphite-reducing Clostridium were identified in the conveyor belts, fish flesh, and viscera. The 16S RNA identification confirmed the presence of viable spoilage bacteria such as Citrobacter gillenii, Macrococcus caseolyticus, Hafnia paralvei, Lactococcus lactis, Lactococcus cremoris, Klebsiella, Escherichia coli, Morganella morganii, and Shewanella. Three of these genera (Citrobacter, Hafnia, and Pseudomonas) were present in all types of samples analysed. The results evidenced potential transmission of microbial contamination from contaminated packaging belts and boxes, evisceration and filleting machines to flesh and viscera samples, thus the establishment of control measures should be implemented in fish farm facilities to extend the shelf-life of farmed fishery products.

Identification of acacia gum fermenting bacteria from pooled human feces using anaerobic enrichment culture.

Commercial acacia gum (AG) used in this study is a premium-grade free-flowing powder. It is a gummy exudate composed of arabinogalactan branched polysaccharide, a biopolymer of arabinose and galactose. Also known as food additive, acacia gum (E414), which is presently marketed as a functional dietary fiber to improve overall human gut health. The health effects may be related to the luminal pH regulation from the short-chain fatty acids (SCFA) production. Studies suggested that amylolytic and butyrogenic pathways are the major factors determining the SCFA outcome of AG in the lower gut. However, the primary bacteria involved in the fermentation have not been studied. This study aimed to investigate the putative primary degraders of acacia gum in the gut ecosystem. Isolation and identification of gum-fermenting bacteria were performed through enrichment culture fermentation. The experiment was conducted in an anaerobic chamber for 144 h in three stages. The study was conducted in triplicate using an anaerobic chamber system. This culture system allows specific responses to support only bacteria that are responsible for gum fermentation among the gut microbiota. Five bacterial strains were isolated and found to be gum-fermenting bacteria. Based on the 16s RNA sequence, the isolates matched to butyrate-producing Escherichia fergusonii, ATCC 35469.

Biological control of Magnaporthe oryzae using natively isolated Bacillus subtilis G5 from Oryza officinalis roots.

Rice blast, caused by Magnaporthe oryzae, is a major threat to global rice production causing significant crop losses and impacting grain quality. The annual loss of rice production due to this disease ranges from 10% to 30%. The use of biologically controlled strains, instead of chemical pesticides, to control plant diseases has become a research hotspot. In this study, an antagonistic endophytic bacterial strain was isolated from the roots of Oryza officinalis using the traditional isolation and culture methods. A phylogenetic tree based on 16S RNA and whole-genome sequencing identified isolate G5 as a strain of Bacillus subtilis. This isolate displayed strong antagonistic effects against different physiological strains of M. oryzae. After co-culture in LB medium for 7 days, the inhibition rates of the mycelial growth of four strains of M. oryzae, ZB15, WH97, Guy11, and T-39800E were 98.07 ± 0.0034%, 98.59 ± 0.0051%, 99.16 ± 0.0012%, and 98.69 ± 0.0065%, respectively. Isolate G5 significantly inhibited the formation of conidia of M. oryzae, with an inhibition rate of 97% at an OD600 of 2. Isolate G5 was able to provide 66.81% protection against rice blast under potted conditions. Whole-genome sequencing revealed that the genome size of isolate G5 was 4,065,878 bp, including 4,182 coding genes. Using the anti-SMASH software, 14 secondary metabolite synthesis gene clusters were predicted to encode antifungal substances, such as fengycin, surfactin, and bacilysin. The G5 isolate also contained genes related to plant growth promotion. These findings provide a theoretical basis for expounding the biocontrol mechanisms of this strain and suggest further development of biogenic agents that could effectively inhibit rice blast pathogen growth and reduce crop damage, while being environmentally friendly, conducive to ecological development, and a sustainable alternative to chemical pesticides. This study also enriches the relevant research on endophytes of wild rice, which proves that wild rice is a valuable microbial resource bank.

Lung Microbiota in Idiopathic Pulmonary Fibrosis, Hypersensitivity Pneumonitis, and Unclassified Interstitial Lung Diseases: A Preliminary Pilot Study.

(1) Introduction: Although historically, the lung has been considered a sterile organ, recent studies through 16S rRNA gene sequencing have identified a substantial number of microorganisms. The human microbiome has been considered an "essential organ," carrying about 150 times more information (genes) than are found in the entire human genome. The purpose of the present study is to characterize and compare the microbiome in three different interstitial lung diseases: idiopathic pulmonary fibrosis (IPF), hypersensitivity pneumonitis, and nondifferential interstitial lung disease. (2) Material and methods: This was a prospective cohort study where the DNA of 28 patients with ILD was extracted from the lavage and then processed using the standard technique of 16S RNA gene sequencing. In a tertiary teaching hospital in the northern, western part of Romania, samples were collected through bronchoscopy and then processed. (3) Results: The same four species were found in all the patients but in different quantities and compositions: Firmicutes, Actinobacteria, Proteobacteria and Bacteroides. Streptococcus was the most prevalent genus, followed by Staphylococcus and Prevotella. Statistically significant differences in the OUT count for the ten most abundant taxa were found for the genus: Gemella, Actinobacteria, Prevotella, Neisseria, Haemophilus, and Bifidobacterium. The comparative analysis showed a richer microbiota in patients with IPF, as shown by the alpha diversity index. (4) Conclusions: In interstitial lung diseases, the microorganisms normally found in the lung are reduced to a restricted flora dominated by the Firmicutes family. These changes significantly disrupt the continuity of the observed bacterial pattern from the oropharynx to the bronchial tree and lung, possibly impacting the evolution and severity of interstitial lung diseases.

Protective effects of sulforaphane on inflammation, oxidative stress and intestinal dysbacteriosis induced by triphenyltin in Cyprinus carpio haematopterus

The purpose of this experiment was to study the mitigation effect of sulforaphane (SFN) on fish toxicological damage caused by triphenyltin (TPT) pollution. A total of 320 healthy fish (56.9 ± 0.4g) were randomly placed into four groups, each with four duplicates. The control group was fed the basal diet, the TPT group was exposed to 10 ng/L TPT on the basis of the control group, the SFN group was fed a diet supplemented with 10 mg/kg SFN, the SFN + TPT group was exposed to 10 ng/L TPT on the basis of the SFN group. Each tank had 20 fish and the breeding lasted for 8 weeks. The present study found that the antioxidant enzyme activity in the TPT group was significantly lower than that of the control group (P < 0.05). In addition, compared with the control group, the mRNA expression of pro-inflammatory factors (IL-6, TNF-α) were significantly induced, and the anti-inflammatory factor genes (IL-10, TGF) were significantly inhibited (P < 0.05) in TPT group. SFN relieved the changes of inflammatory factors caused by TPT, ameliorated oxidative stress, improved antioxidant enzyme (include SOD, CAT, GSH, GPx) activities (P < 0.05). 16s RNA analysis indicated that exposure to TPT caused changes in intestinal microflora. The results of the study showed that after exposure to TPT, some beneficial genera of bacteria in the gut of Rhizobiaceae, Bdellovibrio and Candidatus Alysiosphaera were decreased. The bacteria associated with intestinal inflammation including Propionibacterium, Rubrobacter, Anaerorhabdus_furcosa_group, Rikenellaceae and Eubacterium_brachy were upregulated. However, the SFN treatment group significantly down-regulated the above five inflammation-related bacteria. The above results indicated that TPT caused oxidative stress and inflammation in fish intestines, changed the intestinal microflora, and dietary SFN could improve antioxidant status, regulate inflammation and intestinal health. Therefore, SFN is a promising diet additive for improving fish damage caused by TPT contamination.

Comparison of Proline-glutamate-proline-glutamate-polymorphic GC-rich Sequences Family Protein Wag22 (Rv1759c), PE_PGRS31 (Rv1768), PE_PGRS32 (Rv1803), and PE_PGRS33 gene (Rv1818c) in Exponential State and Under In vitro Model of Latency in Same Clinical Isolates of Mycobacterium tuberculosis: Frameshift Mutation in Extensively Drug-resistant and Totally Drug-resistant tuberculosis Bacilli

Abstract Background: Proline-glutamate (PE)/proline-PE (PPE) proteins play an important role in the development of mycobacterial pathogenicity by modulating the host immune system. In the present investigation, the structural changes in PE-polymorphic GC-rich sequences (PGRS) family protein Wag22 (Rv1759c), PE_PGRS31 (Rv1768), PE_PGRS32 (Rv1803), and PE_PGRS33 gene (Rv1818c) were compared and analyzed in exponential state and under in vitro model of latency in same clinical isolates of Mycobacterium tuberculosis (MTB). Methods: MTB strains were isolated from clinically and laboratory-confirmed cases of tuberculosis (TB). The TB isolates were subjected to the Xpert MTB/rifampin test and then, further susceptibility testing using proportional methods was performed on them. The isolates were characterized using both 16S–23S RNA and hsp65 genes spacer polymerase chain reaction-restriction fragment length polymorphism. Selected isolates studied at two experimental set–up at exponential phase OD 600 = 0.05 (5 cfu/mL × 106 cfu/mL) and under zero oxygen and nutrition for 26 months to selected isolates studied at two experimental setup in exponential phase OD600 = 0.05 (5 cfu/mL × 106 cfu/mL) and under zero oxygen and nutrition after 26 months. Whole-genome sequencing was performed on studied isolates and the protein structures were analyzed using a bioinformatics web server. Results: No deletion, insertion, or substation occurred in susceptible, mono-drug and multidrug resistant-TB isolates were observed at PE-PGRS family protein Wag22 (Rv1759c) and PE_PGRS31 (Rv1768) at exponential phase. Although, a large deletion (at Rv1759c; Rv1768) was observed in extensively drug-resistant (XDR) and totally drug-resistant (TDR) TB isolates at the exponential phase. All studied TDR-TB isolates had a common deletion position from amino acid 1 (methionine) to amino acid 83 (glycine) and from amino acid 725 (proline) to amino acid 914 (threonine) at PE-PGRS family protein Wag22 (Rv1759c). At PE_PGRS32 (Rv1803), deletion occurred from amino acid 1 (methionine) to amino acid 212 (glycine) in latent TDR-TB bacilli. No changes in Rv1803 were observed in other studied isolates. In contrast, 66.6% of studied isolates had either insertion, deletion, substitution, or combination of changes at PE_PGRS33 (Rv1818c). However, the majority of changes at Rv1818c occurred in drug-resistant isolates. We also documented the region of deletion and insertion at PE_PGRS33 (Rv1818c) is different in active and latent TDR-TB isolates. Conclusions: Changes in these PE-PGRS family protein was associated with drug susceptibility patterns of individual isolates. Our result showed a total frameshift mutation of protein that had a different length in comparison to the original protein. These changes might disturb the interactions between XDR and TDR-TB isolates and immune responses, which needs further investigation.

Fructo-oligosaccharide enhanced bioavailability of polyglycosylated anthocyanins from red radish via regulating gut microbiota in mice.

The anthocyanins from red radish (ARR) rich in polyglycosylated pelargonidin glucosides were used as pigment. However, bioavailability of anthocyanins was considered at low level. This work examined the intensive effects of fructo-oligosaccharide (FOS) on ARR bioavailability. Pelargonidin, cyanidin and pelargonidin-3-glucoside showed higher level in serum of mice fed with FOS together with ARR for 8weeks than that fed with only ARR. Co-ingestion of FOS and ARR more effectively elevated the hepatic antioxidant activity by increase in total antioxidant capacity and activities of superoxide dismutase and glutathione peroxidase when compared with intake of ARR. FOS also markedly increased pelargonidin level in cecum of mice. 16S RNA sequencing found that Bacteroides genus play an important role in FOS elevating bioavailability of ARR. Fecal bacteria transplantation verified the positive effects of FOS on ARR bioavailability. These results suggested that combined ingestion of FOS and ARR is effective strategy for bioactivity of ARR.

Characterization of carbapenemase-producing Gram-negative bacilli: first report of blaNDM-1 in Enterobacter cloacae.

The spread of multidrug-resistant bacteria, particularly carbapenem-resistant Gram-negative bacilli (CR-GNB), has become a serious challenge for clinicians due to limited therapeutic options. The aim of the study was to investigate the prevalence of carbapenemase production among clinical isolates recovered from 352 samples collected in Tebessa hospital, Algeria. Bacterial isolates were identified by 16S RNA gene sequencing and susceptibility to antibiotics was determined by disk diffusion method. Carbapenem-resistant isolates were screened for carbapenemase production using modified carba Nordmann-Poirel test, modified Hodge test and imipenem-EDTA combined disc test. Extended-spectrum β-lactamases (ESBL) were detected using double-disk synergy test. Molecular characterization of carbapenemases and ESBL genes was performed by polymerase chain reaction (PCR) and sequencing. A total of 85 Gram-negative bacilli isolates were recovered mainly from urine samples and were identified as: Klebsiella pneumoniae (17.65%), Serratia odorifera (15.29%), Escherichia coli (12.94%), Raoultella ornithinolytica, Enterobacter cloacae (11.76%), Serratia marcescens (10.59%), Morganella morganii (7.06%), Proteus mirabilis (5.88%), Acinetobacter baumannii (4.70%) and Pseudomonas aeruginosa (2.35%). All strains were resistant or intermediate to imipenem and/or ertapenem. ESBL, carbapenemase and metallo-beta-lactamases (MBL) phenotypes were detected in 19 (22.35%), 9 (10.59%) and 2 (2.35%) GNB isolates, respectively. PCR results in nine carbapenemase-producing GNB strains chosen showed the presence of one to four carbapenemase genes (blaGES, blaSME, blaNDM-1, blaVIM, blaGIM, blaSPM, blaOXA-48) in four strains; however, seven strains had at least one ESBL gene (blaTEM-1, blaCTXM-15, blaSHV). In this study, we report the first incidence of blaNDM-1 gene in Enterobacter cloacae isolated from urine sample in Algeria.

Electrochemical DNA Sensor Designed Using the Pencil Graphite Electrode to Detect Listeria monocytogenes.

In the present work, a novel electrochemical DNA sensor was designed to detect L. monocytogenes. Two different gene fragments were selected for the sensor design. One is a 702bp long fragment of the hlyA gene, encoding the synthesis of listeriolysin O toxin, which is unique only to pathogenic strains of L. monocytogenes and is essential for pathogenicity. The other is a 209bp long fragment of the 16S RNA gene found in all species of the Listeria genus. As the working electrode, the pencil graphite electrode was modified in various ways (activated or covered with polypyrrole), and six different combinations were constituted using three types of the modified working electrode and two different gene fragments. The developed system is based on differential pulse voltammetric transduction of guanine oxidation after hybridization between the selected gene fragment (38µg/mL) and the selected fragment-specific inosine-modified probe (1.8 µmol/L) immobilized on a pencil graphite electrode surface. The comparison of all combinations demonstrates that the best results are obtained with the combination formed from a polypyrrole-coated pencil graphite electrode (prepared at 2 scans) and 702bp fragment of the hlyA gene. The analysis time is less than 1 hour, and the necessary DNA concentrations for the analysis have been determined as 8.2 × 10-11M DNA and 2.7 × 10-10M DNA respectively, for the hlyA gene and 16S RNA gene.

Sea Conch Peptides Hydrolysate Alleviates DSS-Induced Colitis in Mice through Immune Modulation and Gut Microbiota Restoration.

Inflammatory bowel disease (IBD) is a persistent, lifelong inflammation of the digestive system. Dextran sulfate sodium is commonly used to induce colitis in experimental animal models, which causes epithelial damage, intestinal inflammation, mucin depletion, and dysbiosis of the gut microbiota. Various prebiotics, polysaccharides, and polypeptides are used for IBD treatment. In this study, we used a murine model utilizing BALB/c mice, with 10 mice per group, to investigate the treatment effect of sea conch peptide hydrolysate (CPH) on DSS-induced colitis mice. Colitis was induced through the administration of 2.5% DSS in drinking water over a seven-days period. Furthermore, on the eighth day of the experiment, sea conch peptide hydrolysate (CPH) at low (100 mg/kg), medium (200 mg/kg), and high (400 mg/kg) doses, which were continued for 14 days, were assessed for medicinal purposes in DSS-induced colitis mice. Our results showed that CPH treatment significantly alleviated the severity and symptoms of colitis. The epithelial integrity and histological damage were improved. Intestinal inflammation and inflammatory cell infiltration were improved. Furthermore, the expression of pro-inflammatory cytokines was reduced, and intestinal barrier integrity was restored by elevating the tight junction proteins. Moreover, 16s RNA sequencing revealed dysbiosis of the gut microbiota was observed upon DSS treatment, which was reinstated after CPH treatment. An increased level of Firmicutes and Lactobacillus was observed in the treatment groups. Finally, our results suggest that CPH would be recommended as a functional food source and also have the potential to be used as a medicinal product for different gastrointestinal disorders.

B-016 Automated Next Generation Sequencing Fluidic-Based Device for Sample to Library Preparation: A Flexible Solution for Rapid Decentralized Sequencing for Infectious Disease Diagnostics

Abstract Background In addition to the established use of Next Generation Sequencing (NGS), such as drug resistance monitoring in HIV, the recent COVID19 pandemic has shown the crucial need of NGS technologies as tools for identification of clinically relevant variants of the SARS-CoV-2 virus. Typically, in these settings, to be cost-effective most NGS analyses were conducted using larger batch sizes often resulting in long turn-around times of 2–3 weeks the need of trained personnel, and centralized testing facilities. Methods We have developed an easy to use automated fluidic-based benchtop device to bridge a perceived gap in NGS library preparation and provide a solution for decentralized testing that does not require specially trained personnel. With a batch size of 6, smaller hospitals and laboratories can perform NGS library preparation on fresh or archived samples with a fast turn-around-time of two days, facilitating earlier reporting to speed up diagnosis and treatment. The Helaxy NGS fluidic device performs sample nucleic acid extraction and a 2-step PCR in a plastic fluidic card, which is the sole consumable required for the extraction and library prep process.The sample is applied to the fluidic card by manual pipetting followed by loading of the card into the device. Nucleic acids are extracted with a combination of magnetic bead pull-down and centrifugal force in a purification chamber to give an eluate that is spun to a separate PCR chamber that uses the concept of thermal induction and cycling. Barcoding for library preparation follows within the same PCR chamber and the resulting product is transferred into another fluidic card for size selection purification, normalization and pooling to generate a library for input into compatible NGS sequencers, e.g., but not confined to, Illumina instruments. Results In order to demonstrate clinical utility we have shown that the Helaxy NGS fluidic device can successfully extract nucleic acid from at least three sample types: SARS-CoV-2 RNA from nasopharyngeal swabs, HIV RNA from plasma and 16S RNA from stools. Using in-house and commercial assays, we have generated libraries of target amplicons using the fluidic device workflow followed by sequencing with Illumina sequencers iSeq and MiSeq. Sequencing results analysed and reported by the Helaxy NGS Bioinformatics Reporting Pipeline revealed expected variants and mutations in the samples. We correctly identified all the unique mutations such as L211del and G446S in commercially available inactivated Omicron BA1 variant of the SARS-CoV-2 virus as well as characterised drug-resistant mutations such as L90M in the Protease gene and D67N and Y181C in the Reverse Transcriptase gene in commercially available inactivated HIV-1 virus. Conclusion The Helaxy NGS fluidic device can thus effectively provide an easy-to-use automated solution for sample library preparation in decentralized facilities to reduce the turn-around-time from sample collection to result reporting for clinical diagnosis and treatment.

RNA Post-transcriptional Modifications of an Early-Stage Large-Subunit Ribosomal Intermediate.

Protein production by ribosomes is fundamental to life, and proper assembly of the ribosome is required for protein production. The RNA, which is post-transcriptionally modified, provides the platform for ribosome assembly. Thus, a complete understanding of ribosome assembly requires the determination of the RNA post-transcriptional modifications in all of the ribosome assembly intermediates and on each pathway. There are 26 RNA post-transcriptional modifications in 23S RNA of the mature Escherichia coli (E. coli) large ribosomal subunit. The levels of these modifications have been investigated extensively only for a small number of large subunit intermediates and under a limited number of cellular and environmental conditions. In this study, we determined the level of incorporations of 2-methyl adenosine, 3-methyl pseudouridine, 5-hydroxycytosine, and seven pseudouridines in an early-stage E. coli large-subunit assembly intermediate with a sedimentation coefficient of 27S. The 27S intermediate is one of three large subunit intermediates accumulated in E. coli cells lacking the DEAD-box RNA helicase DbpA and expressing the helicase inactive R331A DbpA construct. The majority of the investigated modifications are incorporated into the 27S large subunit intermediate to similar levels to those in the mature 50S large subunit, indicating that these early modifications or the enzymes that incorporate them play important roles in the initial events of large subunit ribosome assembly.

Molecular docking and antibacterial activity of Sargassum fusiforme extracts against major coral pathogens.

The present study evaluates the antibacterial properties of alkaloids and the crude extracts (ethanol, n-hexane and ethyl acetate) from seaweed Sargassum fusiforme against coral pathogens (Photobacterium galatheae, Vibrio harveyi, Bordetella trematum,and Ochrobactrum pseudogrignonese) isolated from coral Porites lutea. To our knowledge, this is the first in vitro assay for such extracts on Porites lutea coral pathogens. Bacterial pathogens have been identified using 16S RNA and BankIt into gene bank and given the accession numbers (OR401000; OR401001; OR401336, and OR400998 respectively). GC-Mass profiling conducted for n-hexane compounds confirmed the presence of thirty-eight molecules, twelve of which have been previously reported for their bioactivity. The results revealed that alkaloids and n-hexane extract demonstrated eminent antibacterial activity compared to the other extracts against the tested coral pathogenic bacteria. Molecular docking was conducted to evaluate the twelve previously mentioned bioactive molecules to get a full understanding of the interaction of those bioactive molecules on vital bacterial proteins (Hemolysin protein (PDB ID: 1XEZ) and Cytoplasmic proteins (PDB ID: 3TZC)). Docked twelve molecules against hemolysin protein (PDB ID: 1XEZ) came exactly in line with the docked result of the same molecules with cytoplasmic proteins (PDB ID: 3TZC), proving the bioactivity of 6-O-Palmitoyl-L-ascorbic acid, 3TMS derivative; Glycerol monostearate, 2TMS derivative and Eicosanoic acid complexes in antibacterial activity action and score higher than reference ligand. Those three compounds will be investigated separately in future in vitro assay soon. Our conclusions align with the study's antibacterial in vitro assay results. The present study reports the novelty of different extracts of S. fusiforme as an antibacterial agent against coral pathogenic bacteria that trigger diseases in Porites lutea.

C2 (Clinical Case) Complicated Sinusitis in an Adolescent Swimmer in the COVID-19 Pandemic: An Unexpected Microbial Player

Abstract Introduction/Background There are increasing reports of complicated sinusitis following post-pandemic viral surges. Streptococcus anginosus and Staphylococcus aureus (SA) are common causes. However, unusual pathogens may need to be considered in the face of inadequate therapeutic response. Case Description A previously healthy 13-year-old boy presented with fever, facial pain, and headache two weeks following presumed COVID-19 infection based on exposure history and symptoms. While recovering, he had been swimming daily and recalled inhaling pool water. On examination he was febrile with a tender mid-forehead swelling and left periorbital swelling without features of orbital cellulitis. There was no meningism or focal neurological deficits. Investigations revealed leukocytosis (19.1 x109/L), elevated CRP (156.8 mg/L) and sterile blood cultures. Head imaging confirmed sinusitis complicated by Pott’s puffy tumor, a small subdural empyema and sinus venous thrombosis. Ceftriaxone, metronidazole, and vancomycin were initiated with de-escalation of the latter to cloxacillin after sinus cultures grew methicillin-sensitive SA. Despite broadening to meropenem and vancomycin, he progressed to develop cerebritis. Legionella species identified from intracranial sampling using 16s RNA, allowed targeted therapy with levofloxacin and successful resolution after six weeks of therapy. Discussion The development of this patient’s Pott’s puffy tumor and intracranial complications seemed consistent with post-viral sinusitis in terms of onset and growth of SA. However, his inadequate response to broad-spectrum therapy and sterile cultures from intracranial sampling prompted request for 16sRNA testing. Subsequent disclosure of freshwater swimming exposure with aspiration event raised concerns for water-borne pathogens such as Legionella. The aspiration event occurring in presence of persisting congestion, resulted in Legionella species seeding already inflamed sinuses and further tracking intracranially. Without special cultures required to recover Legionella, this pathogen was initially missed, resulting in disease progression and delayed targeted therapy. Fluroquinolones or macrolides are required for treatment of this pathogen. Conclusion Given that post-viral sinusitis may be anticipated during viral surges, practitioners should consider the role of waterborne pathogens in hosts with significant swimming exposure who worsen on usual therapy. Microbiology and infectious diseases involvement is encouraged to optimize detection and management of waterborne infection. Practitioners are reminded that in the face of therapeutic failure, it is critical to revisit the history for unusual exposures, determine immunocompetence, and explore potential bug-drug mismatch.

Trans-Anethole Alleviates DSS-Induced Ulcerative Colitis by Remodeling the Intestinal Flora to Regulate Immunity and Bile Acid Metabolism.

Ulcerative colitis (UC) is the most common inflammatory bowel disease (IBD); it is incurable, and the treatment is expensive. Trans-anethole (TA), the main component of fennel, exhibits various biological activities. An increasing number of studies have demonstrated the efficacy of herbal active ingredients in the treatment of UC. This study aimed to investigate the effect and mechanism of TA in UC. In this study, we have experimented on mice with dextran sulfate sodium salt (DSS)-induced UC. The TA group was gavaged with 62.5 mg/kg TA by gavage once daily on days 8-14. To observe the effect of TA on the colon tissue, various investigations were performed, including western blot and immunohistochemistry for intestinal barrier protein expression, TUNEL staining for apoptosis, western blot, and ELISA for inflammation level, flow cytometry for Th17/Treg, LC-MS for blood bile acid content, GC-MS for blood fatty acid content, and 16s RNA for intestinal contents. TA alleviated weight loss in mice with UC; increased colon length; alleviated intestinal mucosal damage; upregulated claudin-1, occludin, and ZO-1 protein expression levels; reduced inflammatory factors in the colon and serum; and alleviated apoptosis. TA reduced fatty acid and bile acid levels by inhibiting colony abundance and reducing Th17/Treg cell differentiation in the colon. We found that TA alleviates DSS-induced UC by remodeling the intestinal flora to regulate immunity and bile acid metabolism.

Parvimonas micra can translocate from the subgingival sulcus of the human oral cavity to colorectal adenocarcinoma.

Oral and intestinal samples from a cohort of 93 colorectal cancer (CRC) patients and 30 healthy controls (non-CRC) were collected for microbiome analysis. Saliva (28 non-CRC and 94 CRC), feces (30 non-CRC and 97 CRC), subgingival fluid (20 CRC), and tumor tissue samples (20 CRC) were used for 16S metabarcoding and/or RNA sequencing (RNAseq) approaches. A differential analysis of the abundance, performed with the ANCOM-BC package, adjusting the P-values by the Holm-Bonferroni method, revealed that Parvimonas was significantly over-represented in feces from CRC patients (P-value < 0.001) compared to healthy controls. A total of 11 Parvimonas micra isolates were obtained from the oral cavity and adenocarcinoma of CRC patients. Genome analysis identified a pair of isolates from the same patient that shared 99.2% identity, demonstrating that P. micra can translocate from the subgingival cavity to the gut. The data suggest that P. micra could migrate in a synergistic consortium with other periodontal bacteria. Metatranscriptomics confirmed that oral bacteria were more active in tumor than in non-neoplastic tissues. We suggest that P. micra could be considered as a CRC biomarker detected in non-invasive samples such as feces.

In vitro screening and characterization of lactic acid bacteria from Lithuanian fermented food with potential probiotic properties.

The present work aimed to identify probiotic candidates from Lithuanian homemade fermented food samples. A total of 23 lactic acid bacteria were isolated from different fermented food samples. Among these, only 12 showed resistance to low pH, tolerance to pepsin, bile salts, and pancreatin. The 12 strains also exhibited antimicrobial activity against Staphylococcus aureus ATCC 29213, Salmonella Typhimurium ATCC 14028, Streptococcus pyogenes ATCC 12384, Streptococcus pyogenes ATCC 19615, and Klebsiella pneumoniae ATCC 13883. Cell-free supernatants of isolate 3A and 55w showed the strongest antioxidant activity of 26.37 μg/mL and 26.06 μg/mL, respectively. Isolate 11w exhibited the strongest auto-aggregation ability of 79.96% as well as the strongest adhesion to HCT116 colon cells (25.671 ± 0.43%). The selected strains were tested for their synbiotic relation in the presence of a prebiotic. The selected candidates showed high proliferation in the presence of 4% as compared to 2% galactooligosaccharides. Among the strains tested for tryptophan production ability, isolate 11w produced the highest L-tryptophan levels of 16.63 ± 2.25 μm, exhibiting psychobiotic ability in the presence of a prebiotic. The safety of these strains was studied by ascertaining their antibiotic susceptibility, mucin degradation, gelatin hydrolysis, and hemolytic activity. In all, isolates 40C and 11w demonstrated the most desirable probiotic potentials and were identified by 16S RNA and later confirmed by whole genome sequencing as Lacticaseibacillus paracasei 11w, and Lactiplantibacillus plantarum 40C: following with the harboring plasmid investigation. Out of all the 23 selected strains, only Lacticaseibacillus paracasei 11w showed the potential and desirable probiotic properties.

Lower Airway Dysbiosis Augments Lung Inflammatory Injury in Mild-to-Moderate Chronic Obstructive Pulmonary Disease.

Rationale: Chronic obstructive pulmonary disease (COPD) is associated with high morbidity, mortality, and healthcare costs. Cigarette smoke is a causative factor; however, not all heavy smokers develop COPD. Microbial colonization and infections are contributing factors to disease progression in advanced stages. Objectives: We investigated whether lower airway dysbiosis occurs in mild-to-moderate COPD and analyzed possible mechanistic contributions to COPD pathogenesis. Methods: We recruited 57 patients with a >10 pack-year smoking history: 26 had physiological evidence of COPD, and 31 had normal lung function (smoker control subjects). Bronchoscopy sampled the upper airways, lower airways, and environmental background. Samples were analyzed by 16S rRNA gene sequencing, whole genome, RNA metatranscriptome, and host RNA transcriptome. A preclinical mouse model was used to evaluate the contributions of cigarette smoke and dysbiosis on lower airway inflammatory injury. Measurements and Main Results: Compared with smoker control subjects, microbiome analyses showed that the lower airways of subjects with COPD were enriched with common oral commensals. The lower airway host transcriptomics demonstrated differences in markers of inflammation and tumorigenesis, such as upregulation of IL-17, IL-6, ERK/MAPK, PI3K, MUC1, and MUC4 in mild-to-moderate COPD. Finally, in a preclinical murine model exposed to cigarette smoke, lower airway dysbiosis with common oral commensals augments the inflammatory injury, revealing transcriptomic signatures similar to those observed in human subjects with COPD. Conclusions: Lower airway dysbiosis in the setting of smoke exposure contributes to inflammatory injury early in COPD. Targeting the lower airway microbiome in combination with smoking cessation may be of potential therapeutic relevance.

Multi-Omics Identified THDCA as a Key Contributor to Hyperlipidemia and as a Potential Therapeutic Agent.

In recent years, with the change in human dietary habits, hyperlipidemia (HLP) has become a common chronic disease. Hyperlipidemia is closely related to the incidence of cardiovascular diseases. Due to the increasing incidence and mortality from cardiovascular diseases, it is imperative to develop new medications for reducing lipid levels. With the aim of discovering new treatment options for hyperlipidemia, we conducted a multi-omics analysis of a potential endogenous bile acid compound. Two hyperlipidemia models were established by feeding rats and mice with a high-fat diet. Serum and fecal specimens of rats with hyperlipidemia were collected. Through the combined analysis of lipid metabolism sequencing, 16S RNA intestinal flora sequencing, and bile acid targeted metabolism sequencing, taurohyodeoxycholic acid (THDCA) was found to be a potential lipid-lowering compound. A mouse hyperlipidemia model was developed to verify the anti-hyperlipidemia function of THDCA. Analysis of serum lipid metabolites revealed that the synthesis of bile acid was one of the metabolic pathways that showed significant alterations. 16S RNA sequencing of intestinal flora also found that high-fat diet intake greatly influenced both primary and secondary bile acid biosynthesis. Analysis of bile acid metabolites in the serum and liver tissue found that THDCA in the secondary bile acids is a potential biomarker of hyperlipidemia. Verification experiments in mice confirmed the beneficial function of THDCA in lowering abnormal lipid levels induced by a high-fat diet. THDCA has been identified as a biomarker of hyperlipidemia and has shown potential for the treatment of hyperlipidemia.