Tyrosol-gold nanoparticle functionalized acacia gum-PVA nanofibers for mitigation of Candida biofilm.

Increasing incidences of fungal infections and prevailing antifungal resistance in healthcare settings has given rise to an antifungal crisis on a global scale. The members of the genus Candida, owing to their ability to acquire sessile growth, are primarily associated with superficial to invasive fungal infections, including the implant-associated infections. The present study introduces a novel approach to combat the sessile/biofilm growth of Candida by fabricating nanofibers using a nanoencapsulation approach. This technique involves the synthesis of tyrosol (TYS) functionalized chitosan gold nanocomposite, which is then encapsulated into PVA/AG polymeric matrix using electrospinning. The FESEM, FTIR analysis of prepared TYS-AuNP@PVA/AG NF suggested the successful encapsulation of TYS into the nanofibers. Further, the sustained and long-term stability of TYS in the medium was confirmed by drug release and storage stability studies. The prepared nanomats can absorb the fluid, as evidenced by the swelling index of the nanofibers. The growth and biofilm inhibition, as well as the disintegration studies against Candida, showed 60-70 % biofilm disintegration when 10 mg of TYS-AuNP@PVA/AG NF was used, hence confirming its biological effectiveness. Subsequently, the nanofibers considerably reduced the hydrophobicity index and ergosterol content of the treated cells. Considering the challenges associated with the inhibition/disruption of fungal biofilm, the fabricated nanofibers prove their effectiveness against Candida biofilm. Therefore, nanocomposite-loaded nanofibers have emerged as potential materials that can control fungal colonization and could also promote healing.

Assessing the synergistic potential of bacteriophage endolysins and antimicrobial peptides for eradicating bacterial biofilms.

Phage-encoded endolysins have emerged as a potential substitute to conventional antibiotics due to their exceptional benefits including host specificity, rapid host killing, least risk of resistance. In addition to their antibacterial potency and biofilm eradication properties, endolysins are reported to exhibit synergism with other antimicrobial agents. In this study, the synergistic potency of endolysins was dissected with antimicrobial peptides to enhance their therapeutic effectiveness. Recombinantly expressed and purified bacteriophage endolysin [T7 endolysin (T7L); and T4 endolysin (T4L)] proteins have been used to evaluate the broad-spectrum antibacterial efficacy using different bacterial strains. Antibacterial/biofilm eradication studies were performed in combination with different antimicrobial peptides (AMPs) such as colistin, nisin, and polymyxin B (PMB) to assess the endolysin's antimicrobial efficacy and their synergy with AMPs. In combination with T7L, polymyxin B and colistin effectively eradicated the biofilm of Pseudomonas aeruginosa and exhibited a synergistic effect. Further, a combination of T4L and nisin displayed a synergistic effect against Staphylococcus aureus biofilms. In summary, the obtained results endorse the theme of combinational therapy consisting of endolysins and AMPs as an effective remedy against the drug-resistant bacterial biofilms that are a serious concern in healthcare settings.

Mechanistic insights into antifungal potential of Alexidine dihydrochloride and hexachlorophene in Candida albicans: a drug repurposing approach.

Candida albicans has been listed in the critical priority group by the WHO in 2022 depending upon its contribution in invasive candidiasis and increased resistance to conventional drugs. Drug repurposing offers an efficient, rapid, and cost-effective solution to develop alternative therapeutics against pathogenic microbes. Alexidine dihydrochloride (AXD) and hexachlorophene (HCP) are FDA approved anti-cancer and anti-septic drugs, respectively. In this study, we have shown antifungal properties of AXD and HCP against the wild type (reference strain) and clinical isolates of C. albicans. The minimum inhibitory concentrations (MIC50) of AXD and HCP against C. albicans ranged between 0.34 and 0.69 µM and 19.66-24.58 µM, respectively. The biofilm inhibitory and eradication concentration of AXD was reported comparatively lower than that of HCP for the strains used in the study. Further investigations were performed to understand the antifungal mode of action of AXD and HCP by studying virulence features like cell surface hydrophobicity, adhesion, and yeast to hyphae transition, were also reduced upon exposure to both the drugs. Ergosterol content in cell membrane of the wild type strain was upregulated on exposure to AXD and HCP both. Biochemical analyses of the exposed biofilm indicated reduced contents of carbohydrate, protein, and e-DNA in the extracellular matrix of the biofilm when compared to the untreated control biofilm. AXD exposure downregulated activity of tissue invading enzyme, phospholipase in the reference strain. In wild type strain, ROS level, and activities of antioxidant enzymes were found elevated upon exposure to both drugs. FESEM analysis of the drug treated biofilms revealed degraded biofilm. This study has indicated mode of action of antifungal potential of alexidine dihydrochloride and hexachlorophene in C. albicans.

Biochemical and metabolomic insights into antifungal mechanism of berberine against Candida glabrata.

An unprecedented expansion of antifungal therapy failure incidences in healthcare settings of Candida glabrata is the matter of global concern that needs to be addressed efficiently and effectively. In this pursuit, the present study has investigated the antifungal mechanism of benzylisoquinoline alkaloid berberine using biochemical, metabolic, and gene expression analysis, with the aim to delineate its therapeutic activity against C. glabrata and differentially fluconazole-responsive clinical isolates. Interestingly, the clinical isolates were found to be highly susceptible to berberine. Berberine was found to control the surface properties like hydrophobicity and charge of the cells. The cell membrane composition was altered by berberine, where the ergosterol and fatty acids were affected. The efflux pump activity was inhibited, and osmotic stress was generated in C. glabrata cells upon berberine exposure. The berberine has also generated oxidative stress and activated antioxidant system in C. glabrata cells. Furthermore, these observations were supported by the transcriptional expression study of C. glabrata cell genes (CDR1, RLM1, SLT2, SUR4, KRE1) and metabolomics analysis. Based on fold change analysis, the study identified 20 differential metabolites upon berberine treatment, which belong to central carbon, amino acids, and nucleotide pathways. The checkerboard analysis revealed the potentiation of some classically used antifungal drugs by berberine, thus suggesting it as a combinatorial nutraceutical adjuvant for the eradication of fungal infections. KEY POINTS: • Berberine exhibited better potency against azole-resistant clinical isolates • Berberine modulated metabolites of different pathways • Berberine generated oxidative stress and blocked efflux pump activity.

Seasonal impact on microbiological quality of drinking water in Solan City of Himachal Pradesh, India.

Water contamination with faecal matter is usually the main cause of microbial waterborne diseases. Such diseases are an alarming situation for small cities in developing countries like India. In this research, to check the microbiological status of drinking water in Solan, Himachal Pradesh (India), water samples were collected from baories/stepwells (n = 14), handpumps (n = 9), and the municipal water distribution system (MWDS) (n = 2) in alternative months of the year (covering three main seasons). In 6 months, 150 samples were collected, and they were all examined for the presence of total coliforms and other bacterial pathogens. The associations between the isolates' ecological and seasonal prevalence were also examined. The coliforms were detected by the Most Probable Number (MPN) method, whose range was noticed from the 2-540/100-ml MPN index. The colony forming unit (CFU) count for different samples at the base log 10 value ranged from 3.03 to 6.19. Different genera isolated and identified were Escherichia coli, Salmonella enteric subsp. enterica, Pseudomonas spp., Klebsiella spp., and Staphylococcus aureus. Overall, 74% of the isolates identified in water samples were from the Enterobacteriaceae family. E. coli was about 42.67% (n = 102), followed by Salmonella enterica subsp. enterica 20.92% (n = 50), Staphylococcus aureus 13.38% (n = 32), Pseudomonas spp. 12.55% (n = 30), and Klebsiella spp. 10.46% (n = 25) amongst the total of 239 isolates. The seasonal impact and the dependency of the occurrence of bacteria on one another were determined to be insignificant in the Spearman correlation test. These results showed that external factors (anthropogenic activities) are mainly responsible for the presence of these bacteria in water resources. The occurrence of bacterial isolates has been noticed in all water samples, irrespective of collecting site or season.

Tetraphenylethene Derivatives Modulate the RNA Hairpin-G-Quadruplex Conformational Equilibria in Proto-oncogenes.

RNA G-quadruplex (GQs) sequences in 5'-UTRs of certain proto-oncogenes co-localize with hairpin (Hp) forming sequences resulting in intramolecular Hp-GQ conformational equilibria, which is suggested to regulate cancer development and progression. Thus, regulation of Hp-GQ equilibria with small molecules is an attractive but less explored therapeutic approach. Herein, two tetraphenylethene (TPE) derivatives, TPE-Py and TPE-MePy, were synthesized and their effect on Hp-GQ equilibrium was explored. FRET, CD and molecular docking experiments suggest that cationic TPE-MePy shifts the Hp-GQ equilibrium significantly towards the GQ conformer mainly through π-π stacking and van der Waals interactions. In the presence of TPE-MePy, the observed rate constant values for first and second folding steps were increased up to 14.6 and 2.6-fold, respectively. The FRET melting assay showed a strong stabilizing ability of TPE-MePy (ΔTm=4.36 °C). Notably, the unmethylated derivative TPE-Py did not alter the Hp-GQ equilibrium. Subsequently, luciferase assay analysis demonstrated that the TPE-MePy derivatives suppressed the translation efficiency by ∼5.7-fold by shifting the Hp-GQ equilibrium toward GQ conformers in the 5'-UTR of TRF2. Our data suggests that HpGQ equilibria could be selectively targeted with small molecules to modulate translation for therapy.

Elucidating the lactic acid tolerance mechanism in vaginal clinical isolates of Candida glabrata.

Incidence of vulvovaginal candidiasis are strikingly high and treatment options are limited with nearly 50% Candida glabrata cases left untreated or experience treatment failures. The vaginal microenvironment is rich in lactic acid, and the adaptation of C. glabrata to lactic acid (LA) is the main reason for clinical treatment failure. In the present study, C. glabrata and its vaginal clinical isolates were comprehensively investigated for their growth response, metabolic adaptation and altered cellular pathway to LA using different biochemical techniques, metabolic profiling and transcriptional studies. C. glabrata shown considerable variations in its topological and biochemical features without compromising growth in LA media. Chemical profiling data highlighted involvement of cell wall/membrane, ergosterol and oxidative stress related pathways in mediating adaptative response of C. glabrata towards LA. Further, one dimensional proton (1H) NMR spectroscopy based metabolic profiling revealed significant modulation in 19 metabolites of C. glabrata cells upon growth in LA. Interestingly myo-inositol, xylose, putrescine and betaine which are key metabolites for cell growth and viability were found to be differentially expressed by clinical isolates. These observations were supported by the transcriptional expression study of selected genes evidencing cell wall/membrane re-organisation, altered oxidative stress, and reprogramming of carbon metabolic pathways. Collectively, the study advances our understanding on adaptative response of C. glabrata in vaginal microenvironment to lactic acid for survival and virulence.

In vaginal tract, lactic acid present as a natural carbon source is a potentiating factor for vulvovaginal candidiasis caused by C. glabrata is highest. The present article delineates the lactic acid adaptation in vaginal clinical isolates of C. glabrata using a comprehensive approach of biochemical, metabolic and transcriptional studies.

Geraniol eradicates Candida glabrata biofilm by targeting multiple cellular pathways.

Global burden of fungal infections and associated health risk has accelerated at an incredible pace and needs to be attended at the earliest with an unbeatable therapeutic intervention. Candida glabrata is clinically the most relevant and least drug susceptible Candida species. In the pursuit of mining alternative novel drug candidates, the antifungal activity of a monoterpene phytoactive molecule geraniol (GR) against C. glabrata biofilm was evaluated. Biofilm inhibitory and eradication ability of GR evaluated against C. glabrata along with its clinical isolates. Impact of GR on various cellular pathways was evaluated to delineate its antifungal mode of action. GR has inhibited both planktonic and sessile growth of all the studied C. glabrata strains and eradicated the mature biofilm. GR reduced the carbohydrate and eDNA content, as well as hydrolytic enzyme activity in extracellular matrix of C. glabrata. The chemical profiling, microscopic, and spectroscopic studies revealed that GR targets chitin and ß-glucan in cell wall. Further, results highlighted the reduction of cell membrane ergosterol content, and blocking of ABC drug efflux pump by GR which was also confirmed by RT-PCR where expression of CDR1 and ERG4 was downregulated in GR exposed C. glabrata cells. The fluorescence microscopy and flow cytometry results emphasized the alteration in mitochondrial activity, increased Ca+2 uptake, thus changing the membrane permeability ensuing increased cytochrome C release from mitochondria to cytoplasm. Indeed, GR also has arrested cell cycle in G1/S phase and interfered with DNA replication. These observations suggest GR targets multiple cellular pathways and mediated killing of C. glabrata cells via apoptosis. In conclusion, the present study strengthens the candidacy of GR as novel antifungal therapeutic. Key points • GR inhibits growth and eradicates biofilm of C. glabrata and its clinical isolates. • GR inactivates the hydrolytic enzymes in extracellular matrix. • GR mediates C. glabrata apoptosis by interfering with multiple signaling pathways.

Impact of Bacillus licheniformis SV1 Derived Glycolipid on Candida glabrata Biofilm.

In the present investigation, we have evaluated the antibiofilm potential of Bacillus licheniformis SV1 derived glycolipid against C. glabrata biofilm. Impact of isolated glycolipid on the viability of C. glabrata and on inhibiting as well as eradicating ability of its biofilm were studied. Further, morphological alterations, reactive oxygen species generation (ROS) production and transcriptional expression of selected genes (RT-PCR) of C. glabrata in response with isolated glycolipid were studied. The isolated glycolipid (1.0 mg ml-1) inhibited and eradicated C. glabrata biofilm approximately 80% and 60%, respectively. FE-SEM images revealed glycolipid exposure results in architectural alteration and eradication of C. glabrata biofilm and ROS generation. Transcriptional studies of selected genes showed that the expression of AUS1, FKS1 and KRE1 were down-regulated, while that of ergosterol biosynthesis pathway and multidrug transporter increased, in the presence of glycolipid.

Functional characterization of Candida glabrata ORF, CAGL0M02233g for its role in stress tolerance and virulence.

Present investigation is aimed to analyze the role of an uncharacterized ORF of Candida glabrata (CBS138), CAGL0M02233g (an ortholog of RAD53, a key DNA checkpoint effector in Saccharomyces cerevisiae) in tolerance of various stresses and in biofilm formation. The CAGL0M02233g was cloned in p416TEF shuttle vector for constitutive expression under TEF1 promoter in BG14 strain (ura3 auxotrophic C. glabrata), and upregulated expression of the cloned ORF was confirmed by immunoblotting. The constitutive expression of CAGL0M02233g rendered cells resistant to the DNA damage stressor (MMS), replication stressor (HU) and hypoxia mimetic (CoCl2) in plate spot and growth curve assays. Hypoxia (a low oxygen condition) is an imperative host factor that influences Candida pathogenesis. Biofilm formation by the BG14 cells transformed with p416TEF-CAGL0M02233g (REX cell) was reduced to approximately 50% under hypoxia. It is notable that biofilm formation by the REX cells was significantly lower than that of BG14 cells transformed with p416TEF vector (VC cell) under hypoxia. The biofilm of the REX cells has shown higher susceptibility to fluconazole than that of VC cells under hypoxia and REX cells at normoxia. This is the first report on the function of CAGL0M02233g in tolerance of various stressors and in modulation of the biofilm under hypoxia.

Fluoxetine triggers selective apoptosis in inflammation-induced proliferating (Ki-67high ) thymocytes.

Inappropriate functioning of the immune system is observed during sustained systemic inflammation, which might lead to immune deficiencies, autoimmune disorders and cancer. Primary lymphoid organs may progress to a deregulated proliferative state in response to inflammatory signals in order to intensify host defense mechanisms and exacerbate an inflammatory niche. Fluoxetine, a selective serotonin reuptake inhibitor, has recently been projected as an anti-inflammatory agent. This study had been designed to evaluate the potential novel role of fluoxetine in reversing inflammation-induced immune dysfunction. Lipopolysaccharide (LPS) administration in Swiss albino mice potentiated a systemic inflammatory response, along with increased proliferation of thymocytes and peripheral blood mononuclear cells, as evident from increased Ki-67 expression. The proliferative changes in the immune system were mainly associated with increased phosphorylation of PI3k, AKT and IκB along with elevated NFκB-p65 nuclear translocation. The Ki-67high thymocytes obtained from LPS administered mice demonstrated significantly low p53 nuclear activity, which was established to be mediated by NFκB through reduced nuclear translocation of p53 during LPS-induced proliferative conditions, thereby blocking p53-dependent apoptosis. Fluoxetine supplementation not only reversed the proinflammatory condition, but also induced selective apoptosis in the proliferation-dictated Ki-67high thymocytes possibly by modulating the hypothalamus-pituitary-adrenal axis and inducing glucocorticoid receptor activation and apoptosis in these proliferation-biased immune cells, authenticating a novel antiproliferative role of an established drug.

Structured clinical documentation in the electronic medical record to improve quality and to support practice-based research in epilepsy.

OBJECTIVE: Using the electronic medical record (EMR) to capture structured clinical data at the point of care would be a practical way to support quality improvement and practice-based research in epilepsy. METHODS: We describe our stepwise process for building structured clinical documentation support tools in the EMR that define best practices in epilepsy, and we describe how we incorporated these toolkits into our clinical workflow. RESULTS: These tools write notes and capture hundreds of fields of data including several score tests: Generalized Anxiety Disorder-7 items, Neurological Disorders Depression Inventory for Epilepsy, Epworth Sleepiness Scale, Quality of Life in Epilepsy-10 items, Montreal Cognitive Assessment/Short Test of Mental Status, and Medical Research Council Prognostic Index. The tools summarize brain imaging, blood laboratory, and electroencephalography results, and document neuromodulation treatments. The tools provide Best Practices Advisories and other clinical decision support when appropriate. The tools prompt enrollment in a DNA biobanking study. We have thus far enrolled 231 patients for initial visits and are starting our first annual follow-up visits and provide a brief description of our cohort. SIGNIFICANCE: We are sharing these EMR tools and captured data with other epilepsy clinics as part of a Neurology Practice Based Research Network, and are using the tools to conduct pragmatic trials using subgroup-based adaptive designs.

Nanoformulations for dismantling fungal biofilms: The latest arsenals of antifungal therapy.

Globally, fungal infections have evolved as a strenuous challenge for clinicians, particularly in patients with compromised immunity in intensive care units. Fungal co-infection in Covid-19 patients has made the situation more formidable for healthcare practitioners. Surface adhered fungal population known as biofilm often develop at the diseased site to elicit antifungal tolerance and recalcitrant traits. Thus, an innovative strategy is required to impede/eradicate developed biofilm and avoid the formation of new colonies. The development of nanocomposite-based antibiofilm solutions is the most appropriate way to withstand and dismantle biofilm structures. Nanocomposites can be utilized as a drug delivery medium and for fabrication of anti-biofilm surfaces capable to resist fungal colonization. In this context, the present review comprehensively described different forms of nanocomposites and mode of their action against fungal biofilms. Amongst various nanocomposites, efficacy of metal/organic nanoparticles and nanofibers are particularly emphasized to highlight their role in the pursuit of antibiofilm strategies. Further, the inevitable concern of nanotoxicology has also been introduced and discussed with the exigent need of addressing it while developing nano-based therapies. Further, a list of FDA-approved nano-based antifungal formulations for therapeutic usage available to date has been described. Collectively, the review highlights the potential, scope, and future of nanocomposite-based antibiofilm therapeutics to address the fungal biofilm management issue.

The Significance of Systemic Inflammatory Markers in Prognosis of Head and Neck Squamous Cell Cancers.

This present study aimed to assess the predictive significance of two systemic inflammatory markers, the neutrophilic to lymphocytic ratio (NLR) and platelet to lymphocytic ratio (PLR), in evaluating the prognosis of individuals. The research involved 47 patients diagnosed with head and neck squamous cell carcinoma, all of whom were histo-pathologically confirmed and aged over 18 years. The patients were monitored every 6 months for a period of 18 months. The average age of the study participants was 57.66 ± 13.5 years, with 42 (89.36%) being male and 5 (10.64%) female. After 6 months, the mean PLR in patients with residual/recurrence was 161.5 ± 8.5, which was significantly, exceeded that of patients without residual/recurrence (109.07 ± 36.29; p value < 0.0001). However, no significant correlation was seen between the NLR (p value = 0.822) and residual/recurrence after 6 months. After 12 months, the mean NLR in patients with recurrence was 4.89 ± 0.69, which was significantly higher compared to patients without recurrence (3.48 ± 1.01; p value = 0.025). Conversely, no significant association was found between the PLR (p value = 0.751) and recurrence after 12 months. Notably, there were no significant associations observed in NLR and PLR at the 18-month mark. Elevated levels of the NLR and PLR can serve as indicators of poor prognosis and the presence of residual/recurrent disease in head and neck malignancies.

Intra-tumor ROS amplification by melatonin interferes in the apoptosis-autophagy-inflammation-EMT collusion in the breast tumor microenvironment.

Epidemiological as well as experimental studies have established that the pineal hormone melatonin has inhibitory effects on different types of cancers. Several mechanisms have been proposed for the anticancer activities of melatonin, but the fundamental molecular pathways still require clarity. We developed a mouse model of breast cancer using Ehrlich's ascites carcinoma (injected in the 4th mammary fat pad of female Swiss albino mice) and investigated the possibility of targeting the autophagy-inflammation-EMT colloquy to restrict breast tumor progression using melatonin as intervention. Contrary to its conventional antioxidant role, melatonin was shown to augment intracellular ROS and initiate ROS-dependent apoptosis in our system, by modulating the p53/JNK & NF-κB/pJNK expressions/interactions. Melatonin-induced ROS promoted SIRT1 activity. Interplay between SIRT1 and NF-κB/p65 is known to play a pivotal role in regulating the crosstalk between autophagy and inflammation. Persistent inflammation in the tumor microenvironment and subsequent activation of the IL-6/STAT3/NF-κB feedback loop promoted EMT and suppression of autophagy through activation of PI3K/Akt/mTOR signaling pathway. Melatonin disrupted NF-κB/SIRT1 interactions blocking IL-6/STAT3/NF-κB pathway. This led to reversal of pro-inflammatory bias in the breast tumor microenvironment and augmented autophagic responses. The interactions between p62/Twist1, NF-κB/Beclin1 and NF-κB/Slug were altered by melatonin to strike a balance between autophagy, inflammation and EMT, leading to tumor regression. This study provides critical insights into how melatonin could be utilized in treating breast cancer via inhibition of the PI3K/Akt/mTOR signaling and differential modulation of SIRT1 and NF-κB proteins, leading to the establishment of apoptotic and autophagic fates in breast cancer cells.

16S rRNA female reproductive microbiome investigation reveals Dalfopristin, Clorgyline, and Hydrazine as potential therapeutics for the treatment of bacterial vaginosis.

Bacterial vaginosis (BV) is a prevalent vaginal illness resulting from a disruption in the vaginal microbial equilibrium. The vaginal microbiota has been shown to have a substantial impact on the development and continuation of BV. This work utilized 16S rRNA sequence analysis of vaginal microbiome samples (Control vs BV samples) utilizing Parallel-Meta 3 to investigate the variations in microbial composition. The unique genes identified were used to determine prospective therapeutic targets and their corresponding inhibitory ligands. Further, molecular docking was conducted and then MD simulations were carried out to confirm the docking outcomes. In the BV samples, we detected several anaerobic bacteria recognized for their ability to generate biofilms, namely Acetohalobium, Anaerolineaceae, Desulfobacteraceae, and others. Furthermore, we identified Dalfopristin, Clorgyline, and Hydrazine as potential therapeutic options for the management of BV. This research provides new insights into the causes of BV and shows the potential effectiveness of novel pharmacological treatments.

Biomarker Identification for Preterm Birth Susceptibility: Vaginal Microbiome Meta-Analysis Using Systems Biology and Machine Learning Approaches.

PROBLEM: The vaginal microbiome has a substantial role in the occurrence of preterm birth (PTB), which contributes substantially to neonatal mortality worldwide. However, current bioinformatics approaches mostly concentrate on the taxonomic classification and functional profiling of the microbiome, limiting their abilities to elucidate the complex factors that contribute to PTB. METHOD OF STUDY: A total of 3757 vaginal microbiome 16S rRNA samples were obtained from five publicly available datasets. The samples were divided into two categories based on pregnancy outcome: preterm birth (PTB) (N = 966) and term birth (N = 2791). Additionally, the samples were further categorized based on the participants' race and trimester. The 16S rRNA reads were subjected to taxonomic classification and functional profiling using the Parallel-META 3 software in Ubuntu environment. The obtained abundances were analyzed using an integrated systems biology and machine learning approach to determine the key microbes, pathways, and genes that contribute to PTB. The resulting features were further subjected to statistical analysis to identify the top nine features with the greatest effect sizes. RESULTS: We identified nine significant features, namely Shuttleworthia, Megasphaera, Sneathia, proximal tubule bicarbonate reclamation pathway, systemic lupus erythematosus pathway, transcription machinery pathway, lepA gene, pepX gene, and rpoD gene. Their abundance variations were observed through the trimesters. CONCLUSIONS: Vaginal infections caused by Shuttleworthia, Megasphaera, and Sneathia and altered small metabolite biosynthesis pathways such as lipopolysaccharide folate and retinal may increase the susceptibility to PTB. The identified organisms, genes, pathways, and their networks may be specifically targeted for the treatment of bacterial infections that increase PTB risk.