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The human vaginal microbiota is a critical determinant of vaginal health. These communities live in close association with the vaginal epithelium and rely on host tissues for resources. Although often dominated by lactobacilli, the vaginal microbiota is also frequently composed of a collection of facultative and obligate anaerobes. The prevalence of these communities with a paucity of Lactobacillus species varies among women, and epidemiological studies have associated them with an increased risk of adverse health outcomes. The mechanisms that drive these associations have yet to be described in detail, with few studies establishing causative relationships. Here, we review our current understanding of the vaginal microbiota and its connection with host health. We centre our discussion around the biology of the vaginal microbiota when Lactobacillus species are dominant versus when they are not, including host factors that are implicated in shaping these microbial communities and the resulting adverse health outcomes. We discuss current approaches to modulate the vaginal microbiota, including probiotics and vaginal microbiome transplants, and argue that new model systems of the cervicovaginal environment that incorporate the vaginal microbiota are needed to progress from association to mechanism and this will prove invaluable for future research., (© 2022. Springer Nature Limited.)

Vaginal microbiota composition affects many facets of reproductive health. Lactobacillus iners-dominated microbial communities are associated with poorer outcomes, including higher risk of bacterial vaginosis (BV), compared with vaginal microbiota rich in L. crispatus. Unfortunately, standard-of-care metronidazole therapy for BV typically results in dominance of L. iners, probably contributing to post-treatment relapse. Here we generate an L. iners isolate collection comprising 34 previously unreported isolates from 14 South African women with and without BV and 4 previously unreported isolates from 3 US women. We also report an associated genome catalogue comprising 1,218 vaginal Lactobacillus isolate genomes and metagenome-assembled genomes from >300 women across 4 continents. We show that, unlike L. crispatus, L. iners growth is dependent on L-cysteine in vitro and we trace this phenotype to the absence of canonical cysteine biosynthesis pathways and a restricted repertoire of cysteine-related transport mechanisms. We further show that cysteine concentrations in cervicovaginal lavage samples correlate with Lactobacillus abundance in vivo and that cystine uptake inhibitors selectively inhibit L. iners growth in vitro. Combining an inhibitor with metronidazole promotes L. crispatus dominance of defined BV-like communities in vitro by suppressing L. iners growth. Our findings enable a better understanding of L. iners biology and suggest candidate treatments to modulate the vaginal microbiota to improve reproductive health for women globally., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Evidence for the concept of the "gut-brain axis" (GBA) has risen. Many types of research demonstrated the mechanism of the GBA and the effect of probiotic intake. Although many studies have been reported, most were focused on neurodegenerative disease and, it is still not clear what type of bacterial strains have positive effects. We designed an experiment to discover a strain that positively affects brain function, which can be recognized through changes in cognitive processes using healthy mice. The experimental group consisted of a control group and three probiotic consumption groups, namely, Lactobacillus acidophilus, Lacticaseibacillus paracasei, and Lacticaseibacillus rhamnosus. Three experimental groups fed probiotics showed an improved cognitive ability by cognitive-behavioral tests, and the group fed on L. acidophilus showed the highest score. To provide an understanding of the altered microbial composition effect on the brain, we performed full 16S-23S rRNA sequencing using Nanopore, and operational taxonomic units (OTUs) were identified at species level. In the group fed on L. acidophilus, the intestinal bacterial ratio of Firmicutes and Proteobacteria phyla increased, and the bacterial proportions of 16 species were significantly different from those of the control group. We estimated that the positive results on the cognitive behavioral tests were due to the increased proportion of the L. acidophilus EG004 strain in the subjects' intestines since the strain can produce butyrate and therefore modulate neurotransmitters and neurotrophic factors. We expect that this strain expands the industrial field of L. acidophilus and helps understand the mechanism of the gut-brain axis. IMPORTANCE Recently, the concept of the "gut-brain axis" has risen and suggested that microbes in the GI tract affect the brain by modulating signal molecules. Although many pieces of research were reported in a short period, a signaling mechanism and the effects of a specific bacterial strain are still unclear. Besides, since most of the research was focused on neurodegenerative disease, the study with a healthy animal model is still insufficient. In this study, we show using a healthy animal model that a bacterial strain (Lactobacillus acidophilus EG004) has a positive effect on mouse cognitive ability. We experimentally verified an improved cognitive ability by cognitive behavioral tests. We performed full 16S-23S rRNA sequencing using a Nanopore MinION instrument and provided the gut microbiome composition at the species level. This microbiome composition consisted of candidate microbial groups as a biomarker that shows positive effects on cognitive ability. Therefore, our study suggests a new perspective for probiotic strain use applicable for various industrialization processes.

This study aimed to explore the changes of the vaginal microbiota and enzymes in the women with high-risk human papillomavirus (HR-HPV) infection and cervical lesions. A total of 448 participants were carried out HPV genotyping, cytology tests, and microecology tests, and 28 participants were treated as sub-samples, in which vaginal samples were characterized by sequencing the bacterial 16S V4 ribosomal RNA (rRNA) gene region. The study found the prevalence of HR-HPV was higher in patients with BV (P = 0.036). The HR-HPV infection rate was 72.73% in G. vaginalis women, which was significantly higher than that of women with lactobacillus as the dominant microbiota (44.72%) (P = 0.04). The positive rate of sialidase (SNA) was higher in women with HR-HPV infection (P = 0.004) and women diagnosed with cervical intraepithelial neoplasia (CIN) (P = 0.041). In HPV (+) women, the α-diversity was significantly higher than that in HPV (-) women. The 16S rRNA gene-based amplicon sequencing results showed that Lactobacillus was the dominant bacteria in the normal vaginal microbiota. However, the proportion of Gardnerella and Prevotella were markedly increased in HPV (+) patients. Gardnerella and Prevotella are the most high-risk combination for the development of HPV (+) women. The SNA secreted by Gardnerella and Prevotella may play a significant role in HPV infection progress to cervical lesions., (© 2022. The Author(s).)

Nearly three hundred thousand female patients are diagnosed with ovarian cancer in the world annually, and this number shows an increasing trend. However, characteristic symptoms caused by ovarian cancer are so few that early diagnosis remains challenging, and an effective screening method has not yet been established. Here, we conducted a case-control study in Japan to analyze the association between cervicovaginal microbiome and ovarian cancer, using 16S rRNA amplicon sequencing. Analysis of DNA extracted from cervical smear samples revealed Lactobacillus-dominant and Lactobacillus-deficient, highly-diversified bacterial communities in premenopausal and postmenopausal healthy controls, respectively, as reported for vaginal microbiota previously. We found that cervicovaginal microbiota in ovarian cancer patients, regardless of their menopausal status, were frequently a diversified community and similar to those in healthy subjects at postmenopausal ages. The diverse microbiota was associated with the major histotypes of epithelial ovarian cancer, including serous ovarian cancer and ovarian clear cell cancer. The present study implies the potential of a cervicovaginal microbiome biomarker in screening ovarian cancer in premenopausal women., (Copyright © 2021. Published by Elsevier B.V.)

Introduction: Sialidase activity in the cervicovaginal fluid (CVF) is associated with microscopic findings of bacterial vaginosis (BV). Sequencing of bacterial 16S rRNA gene in vaginal samples has revealed that the majority of microscopic BV cases fit into vaginal community-state type IV (CST IV), which was recently named "molecular-BV." Bacterial vaginosis-associated bacterial species, such as Gardnerella spp., may act as sources of CVF sialidases. These hydrolases lead to impairment of local immunity and enable bacterial adhesion to epithelial and biofilm formation. However, the impact of CVL sialidase on microbiota components and diversity remains unknown., Objective: To assess if CVF sialidase activity is associated with changes in bacterial components of CST IV., Methods: One hundred forty women were cross-sectionally enrolled. The presence of molecular-BV (CST IV) was assessed by V3-V4 16S rRNA sequencing (Illumina). Fluorometric assays were performed using 2-(4-methylumbelliferyl)-α-D-N-acetylneuraminic acid (MUAN) for measuring sialidase activity in CVF samples. Linear discriminant analysis effect size (LEfSe) was performed to identify the differently enriched bacterial taxa in molecular-BV according to the status of CVF sialidase activity., Results: Forty-four participants (31.4%) had molecular-BV, of which 30 (68.2%) had sialidase activity at detectable levels. A total of 24 bacterial taxa were enriched in the presence of sialidase activity, while just two taxa were enriched in sialidase-negative samples., Conclusion: Sialidase activity in molecular-BV is associated with changes in bacterial components of the local microbiome. This association should be further investigated, since it may result in diminished local defenses against pathogens., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Ferreira, Marconi, Parada, Ravel and Silva.)

All the environmentally exposed surfaces of the human body harbor ecologically distinct microbial communities with a mutualistic beneficial relationship. Depending on the body sites, microbes may provide metabolic functions, protection against pathogens, and signaling molecules to modulate host physiology and reduce disease susceptibility. Our recent understanding of the vaginal microbiome based on culture-independent 16S rRNA gene sequencing indicates that Lactobacillus-dominated microbial communities of healthy women play an important role in decreasing susceptibility to several urogenital diseases, including bacterial, fungal and viral infections. The findings of shotgun sequencing of the vaginal microbiome suggest that microbial-derived lactic acid, bacteriostatic, bactericidal molecules, and lower vaginal pH mediate such protections and regulations. Bacterial species, the dominant component of the vaginal microbiome, also play a key role in determining the gestation period and birth outcomes of reproductive-age women. The presence of Lactobacillus crispatus species in the vaginal milieu reduces the risk of preterm delivery in women of Asian ancestry. A deeper knowledge of the vaginal microbiota's role in the succession and development of newborn gut bacteria would also be beneficial. The microbiome of the mother changes throughout pregnancy and is linked to the microbiome of the newborn. This chapter highlights updated information and new opportunities for human microbiome research, focusing on the assessment of the risk of preterm birth., (Copyright © 2022 Elsevier Inc. All rights reserved.)

While lactobacilli are not generally regarded as efficient cell factories for heterologous proteins, these food-grade Gram-positive bacteria are attractive as expression hosts for medicinal proteins. Furthermore, tools have been developed not only to secrete the protein of interest, but also to anchor the protein to the cell membrane or the cell wall. Research efforts aimed at the production and surface display of complex vaccine proteins have shown that lactobacilli are capable of producing heterologous proteins that are otherwise difficult to produce in soluble form. Many recent studies on expressing a wide variety of proteins in lactobacilli have employed the pSIP vector system, which offers a wide range of possibilities for inducible expression, including various options for secretion and surface anchoring. The modular nature of the pSIP vectors allows for rapid screening of multiple expression strategies. This chapter describes the pSIP vector system and how it can be used to accomplish protein expression in lactobacilli., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Listeria monocytogenes is an important difficult to control and eradicate foodborne pathogen due to its resistance properties to extreme conditions. Bacteriocins produced by lactic acid bacteria (LAB) can be considered as natural alternatives for safety and quality of foods, since these molecules offer antimicrobial activity against other bacteria., Methods: In this study, Lacticaseibacillus casei, Lactiplantibacillus plantarum, and L. monocytogenes isolates were first characterized by phenotypical tests and 16S rRNA gene using PCR. Then, six types of bacteriocins produced by Lactobacilli strains were identified using molecular tests. The ability of these strains to compete with L. monocytogenes for adhesion and invasion to HT-29 cells was evaluated through colony count and MTT assay. Finally, the level of bacteriocins expression was assessed using qRT-PCR., Results: L. monocytogenes strains were categorized from A1 to A8 based on the source of isolation. In the adhesion assay, L. casei + L. monocytogenes isolated from milk and Lpb plantarum + L. monocytogenes isolated from feces presented the maximum adherence values. Further, Lpb plantarum + L. monocytogenes isolated from blood invaded to HT-29 cell line at the highest level. Eventually, L. casei + Lpb plantarum + L. monocytogenes isolated from placenta revealed more expression levels in comparison with other groups., Conclusion: These results suggest a practical approach to classifying bacteriocins into functional groups that could be used for identifying the best mixture of bacteriocins for usage against L. monocytogenes., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Objective: This study aimed to search for Enterococcus faecalis in children's deep carious dentine and characterize their virulence traits. Material and Methods: Eight isolates from 15 carious molars identified by 16S rDNA species-specific PCR as E. faecalis were included. These eight isolates were subject to identification by MALDI-TOF and characterized regarding: (i) bacterial aggregation and biofilm formation on polystyrene and glass, with/without saliva, as single or dual-species (associated to Streptococcus mutans ); (ii) environmental pH measurement before and after 24 h incubation; (iii) acidogenicity; (iv) gelatinase production; (v) macrophage adherence; and (vi) toxicity towards Caenorhabditis elegans . Statistical analyses were performed using two-way ANOVA/Tukey or Fisher's exact tests. Results: All isolates initially identified as E. faecalis by PCR were correctly identified as Lactobacillus by MALDI-TOF, being designated as Lactobacillus misidentified as Enterococcus (LME). These isolates produced biofilm in the presence of saliva and in the dual-species assays. Bacterial aggregation was only observed in the dual-species model. After 24 h, environmental pH dropped from 7.5 to 4.5 for seven of eight isolates, and to 4.0 in all dual-species models. LME isolates were acidogenic, none of them produced gelatinase or adhered to macrophages, but all presented toxicity towards C. elegans . Conclusions: No E. faecalis were identified in the children's caries lesions. All LME isolates presented important virulence traits, including biofilm formation and high acidogenicity, which cause enamel demineralization, that might increase the risk of dental caries in children carrying LME. Thus, the correct identification and in-depth virulence characterization of microorganisms isolated from dental caries are important to understand the dynamics of this disease.

The bacterial genus Lactobacillus comprises a vast range of strains with varying metabolic and probiotic traits, with genome editing representing an essential tool to probe genotype-phenotype relationships and enhance their beneficial properties. Currently, one of the most effective means of genome editing in bacteria couples low-efficiency recombineering with high-efficiency counterselection by nucleases from CRISPR-Cas systems. In lactobacilli, several CRISPR-based genome editing methods exist that have shown varying success in different strains. Here, we detail a fast and simple approach using two shuttle vectors encoding a recombineering template as well as the Streptococcus pyogenes Cas9, a trans-activating RNA, and a CRISPR array. We provide a step-by-step procedure for cloning the shuttle vectors, sequentially transforming the vectors into lactobacilli, screening for the desired edit, and finally clearing the shuttle vectors from the mutant strain. As CRISPR-based genome editing in bacteria can fail for various reasons, we also lay out instructions for probing mechanisms of escape. Finally, we include practical notes along the way to facilitate each stage of genome editing, and we illustrate the technique using a representative edit in a strain of Lactobacillus plantarum. Overall, this method should serve as a complete guide to performing genome editing in lactobacilli., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Animal microbiota is becoming an object of interest as a source of beneficial bacteria for commercial use. Moreover, the escalating problem of bacterial resistance to antibiotics is threatening animals and humans; therefore, in the last decade intensive search for alternative antimicrobials has been observed. In this study, lactic acid bacteria (LAB) were isolated from suckling and weaned pigs feces (376) and characterized to determine their functional properties and usability as pigs additives. Selection of the most promising LAB was made after each stage of research. Isolates were tested for their antimicrobial activity (376) and susceptibility to antibiotics (71). Selected LAB isolates (41) were tested for the production of organic acids, enzymatic activity, cell surface hydrophobicity and survival in gastrointestinal tract. Isolates selected for feed additive (5) were identified by MALDI-TOF mass spectrometry and partial sequence analysis of 16S rRNA gene, represented by Lentilactobacillus, Lacticaseibacillus (both previously classified as Lactobacillus) and Pediococcus genus. Feed additive prototype demonstrated high viability after lyophilization and during storage at 4 °C and - 20 °C for 30 days. Finally, feed additive was tested for survival in simulated alimentary tract of pigs, showing viability at the sufficient level to colonize the host. Studies are focused on obtaining beneficial strains of LAB with probiotic properties for pigs feed additive., (© 2021. The Author(s).)

Lactobacilli are a promising natural tool against vaginal dysbiosis and infections. However, new local delivery systems and additional knowledge about their distribution and mechanism of action would contribute to the development of effective medicine. This will be facilitated by the introduction of the techniques for effective, inexpensive, and real-time tracking of these probiotics following their release. Here, we engineered three model vaginal lactobacilli ( Lactobacillus crispatus ATCC 33820, Lactobacillus gasseri ATCC 33323, and Lactobacillus jensenii ATCC 25258) and a control Lactobacillus plantarum ATCC 8014 to express fluorescent proteins with different spectral properties, including infrared fluorescent protein (IRFP), green fluorescent protein (GFP), red fluorescent protein (mCherry), and blue fluorescent protein (mTagBFP2). The expression of these fluorescent proteins differed between the Lactobacillus species and enabled quantification and discrimination between lactobacilli, with the longer wavelength fluorescent proteins showing superior resolving power. Each Lactobacillus strain was labeled with an individual fluorescent protein and incorporated into poly (ethylene oxide) nanofibers using electrospinning, as confirmed by fluorescence and scanning electron microscopy. The lactobacilli retained their fluorescence in nanofibers, as well as after nanofiber dissolution. To summarize, vaginal lactobacilli were incorporated into electrospun nanofibers to provide a potential solid vaginal delivery system, and the fluorescent proteins were introduced to distinguish between them and allow their tracking in the future probiotic-delivery studies.

HIV incidence continues to be unacceptably high in Eastern and Southern Africa, with women disproportionately affected. An increased per-contact risk of HIV acquisition among African, Caribbean, and other Black (ACB) women has been associated with the higher prevalence of bacterial vaginosis (BV) in these communities, wherein the vaginal microbiota is predominated by diverse pro-inflammatory anaerobic bacteria. However, while the vaginal microbiota in BV-free women is typically predominated by one of several different Lactobacillus spp., the degree of HIV protection afforded by a Lactobacillus-predominant vaginal microbiota also varies considerably. Specifically, L. crispatus is associated with an immunoregulatory genital immune environment, exclusion of BV-associated bacteria, and reduced HIV risk. In contrast, less HIV protection or exclusion of BV-associated bacteria and fewer immune benefits have been associated with L. iners-which is unfortunately the most common Lactobacillus species among ACB women. These species-specific clinical differences are underpinned by substantial genomic differences between Lactobacillus species: for instance, the much smaller genome of L. iners lacks the coding sequence for D-lactic acid dehydrogenase and cannot produce the D-lactate isomer that enhances HIV trapping in mucus but encodes for epithelial cell toxins and stress resistance proteins that may enhance bacterial survival in the context of microbiota and environmental fluctuations. While more studies are needed to elucidate whether differences in HIV protection between Lactobacillus species are due to direct genital immune effects or the exclusion of proinflammatory BV-associated bacteria, the current body of work suggests that for BV treatment to succeed as an HIV prevention strategy, it may be necessary to induce a vaginal microbiota that is predominated by specific (non-iners) Lactobacillus species. Video abstract., (© 2021. The Author(s).)

Cruciferous vegetables are rich sources of glucosinolates (GSLs). GSLs are degraded into isothiocyanates, which are potent anticarcinogens, by human gut bacteria. However, the mechanisms and enzymes involved in gut bacteria-mediated GSL metabolism are currently unclear. This study aimed to elucidate the enzymes involved in GSL metabolism in lactic acid bacteria, a type of gut bacteria. Companilactobacillus farciminis KB1089 was selected as a lactic acid bacteria strain model that metabolizes sinigrin, which is a GSL, into allylisothiocyanate. The sinigrin-metabolizing activity of this strain is induced under glucose-absent and sinigrin-present conditions. A quantitative comparative proteomic analysis was conducted and a total of 20 proteins that were specifically expressed in the induced cells were identified. Three candidate proteins, β-glucoside-specific IIB, IIC, IIA phosphotransferase system (PTS) components (CfPttS), 6-phospho-β-glucosidase (CfPbgS) and a hypothetical protein (CfNukS), were suspected to be involved in sinigrin-metabolism and were thus investigated further. We hypothesize a pathway for sinigrin degradation, wherein sinigrin is taken up and phosphorylated by CfPttS, and subsequently, the phosphorylated entity is degraded by CfPbgS. As expression of both pttS and pbgS genes clearly gave Escherichia coli host strain sinigrin converting activity, these genes were suggested to be responsible for sinigrin degradation. Furthermore, heterologous expression analysis using Lactococcus lactis suggested that CfPttS was important for sinigrin degradation and CfPbgS degraded phosphorylated sinigrin., (© 2021. The Author(s).)

Objective: To assess the association between vaginal microbiome (VMB) composition and recurrent early spontaneous preterm birth (sPTB)/preterm prelabour rupture of membranes (PPROM)., Design: Nested case-control study., Setting: UK tertiary referral hospital., Sample: High-risk women with previous sPTB/PPROM <34 +0  weeks' gestation who had a recurrence (n = 22) or delivered at ≥37 +0  weeks without PPROM (n = 87)., Methods: Vaginal swabs collected between 15 and 22 weeks' gestation were analysed by 16S rRNA gene sequencing and 16S quantitative PCR., Main Outcome Measure: Recurrent early sPTB/PPROM., Results: Of the 109 high-risk women, 28 had anaerobic vaginal dysbiosis, with the remainder dominated by lactobacilli (Lactobacillus iners 36/109, Lactobacillus crispatus 23/109, or other 22/109). VMB type and diversity were not associated with recurrence. Women with a recurrence, compared to those without, had a higher median vaginal bacterial load (8.64 versus 7.89 log 10 cells/mcl, adjusted odds ratio [aOR] 1.90, 95% CI 1.01-3.56, P = 0.047) and estimated Lactobacillus concentration (8.59 versus 7.48 log 10 cells/mcl, aOR 2.35, (95% CI 1.20-4.61, P = 0.013). A higher recurrence risk was associated with higher median bacterial loads for each VMB type after stratification, although statistical significance was reached only for L. iners domination (aOR 3.44, 95% CI 1.06-11.15, P = 0.040). Women with anaerobic dysbiosis or L. iners domination had a higher median vaginal bacterial load than women with a VMB dominated by L. crispatus or other lactobacilli (8.54, 7.96, 7.63, and 7.53 log 10 cells/mcl, respectively)., Conclusions: Vaginal bacterial load is associated with early sPTB/PPROM recurrence. Domination by lactobacilli other than L. iners may protect women from developing high bacterial loads. Future PTB studies should quantify vaginal bacteria and yeasts., Tweetable Abstract: Increased vaginal bacterial load in the second trimester may be associated with recurrent early spontaneous preterm birth., (© 2021 John Wiley & Sons Ltd.)

Pregnancy loss is a common obstetric problem. Significant causes of miscarriage include genetic and epigenetic disorders of the embryo, immunological and endocrine factors, uterine malformations, improper embryo selection, and lifestyle. Perhaps a hitherto underappreciated cause of miscarriage may be an abnormal microbiota composition of the female reproductive system. Lactobacillus spp. is the most common bacteria within the reproductive tract. However, the protective role of Lactobacilli in the vagina has been well described in the literature, while it is still unknown what function Lactobacilli may have in the uterus. Moreover, new research shows that Lactobacillus spp. can have a role in miscarriage. However, both molecular and immunological mechanisms of host- Lactobacillus spp. interactions are not fully understood. Understanding these relationships will help address the importance and extent of the protective role of Lactobacillus spp. in miscarriage., Competing Interests: Conflict of interest The authors do not report any financial or personal connections with other persons or organizations, which might negatively affect the contents of this publication and/or claim authorship rights to this publication., (© 2021 Aneta Kiecka et al.)

Objective: To evaluate the impact of vaginal microbiota on pregnancy outcomes in women undergoing assisted reproduction., Design: A prospective cohort study., Setting: A university-based assisted reproductive technology (ART) centre., Population: 223 women undergoing ART treatment., Methods: Prior to embryo transfer, vaginal samples were collected from the posterior fornix. Vaginal microbiota identification was carried out using next-generation sequencing and categorised according to the V3-V4 hypervariable region in the 16S rRNA gene region., Main Outcome Measures: ART clinical outcomes (implantation, clinical pregnancy rates and live birth rates)., Results: The live birth rate in women with community state type (CST)-I (39%) was higher than that in women with CST-III (21.5%) but the difference was not statistically significant (P = 0.052). The relative abundance of Lactobacillus was lower in women who failed to become pregnant (NP group) (67.71%) than in women who became pregnant (PR group) (79.72%). However, this difference was not statistically significant (P = 0.06). In the NP group, the relative abundance of Streptococcus (7.81%) and Gardnerella (9.40%) was higher than that in the PR group (relative abundance of Streptococcus and Gardnerella was 2.28% and 5.56%, respectively). The abundance of Streptococcus was found to be statistically significantly different between the two study groups (P = 0.014). Linear discriminant analysis (LDA) further validated that Streptococcus had the highest contribution (LDA score >4.0) to the difference between these two groups., Conclusions: Streptococcus has the highest contribution to the distinction between the PR and NP groups., Tweetable Abstract: A relatively high abundance of Streptococcus in the vaginal microbiota may be associated with a lower ART success rate., (© 2021 John Wiley & Sons Ltd.)

The potential of the gut microbiome as a driver of individual cognitive differences in natural populations of animals remains unexplored. Here, using metagenomic sequencing of individual bumblebee hindguts, we find a positive correlation between the abundance of Lactobacillus Firm-5 cluster and memory retention on a visual discrimination task. Supplementation with the Firm-5 species Lactobacillus apis, but not other non-Firm-5 bacterial species, enhances bees' memory. Untargeted metabolomics after L. apis supplementation show increased LPA (14:0) glycerophospholipid in the haemolymph. Oral administration of the LPA increases long-term memory significantly. Based on our findings and metagenomic/metabolomic analyses, we propose a molecular pathway for this gut-brain interaction. Our results provide insights into proximate and ultimate causes of cognitive differences in natural bumblebee populations., (© 2021. The Author(s).)

Lactobacillus iners , first described in 1999, is a prevalent bacterial species of the vaginal microbiome. As L. iners does not easily grow on de Man-Rogosa-Sharpe agar, but can grow anaerobically on blood agar, it has been initially overlooked by traditional culture methods. It was not until the wide application of molecular biology techniques that the function of L. iners in the vaginal microbiome was carefully explored. L. iners has the smallest genome among known Lactobacilli and it has many probiotic characteristics, but is partly different from other major vaginal Lactobacillus species, such as L. crispatus , in contributing to the maintenance of a healthy vaginal microbiome. It is not only commonly present in the healthy vagina but quite often recovered in high numbers in bacterial vaginosis (BV). Increasing evidence suggests that L. iners is a transitional species that colonizes after the vaginal environment is disturbed and offers overall less protection against vaginal dysbiosis and, subsequently, leads to BV, sexually transmitted infections, and adverse pregnancy outcomes. Accordingly, under certain conditions, L. iners is a genuine vaginal symbiont, but it also seems to be an opportunistic pathogen. Further studies are necessary to identify the exact role of this intriguing species in vaginal health and diseases., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zheng, Guo, Wang, Zhou and Ling.)

Lactobacillus paragasseri was identified as a novel sister taxon of L. gasseri in 2018. Since the reclassification of L. paragasseri, there has been hardly any report describing the probiotic properties of this species. In this study, an L. paragasseri strain UBLG-36 was sequenced and analyzed to determine the molecular basis that may confer the bacteria with probiotic potential. UBLG-36 was previously documented as an L. gasseri strain. Average nucleotide identity and phylogenomic analysis allowed accurate taxonomic identification of UBLG-36 as an L. paragasseri strain. Analysis of the draft genome (~1.94 Mb) showed that UBLG-36 contains 5 contigs with an average G+C content of 34.85%. Genes essential for the biosynthesis of bacteriocins, adhesion to host epithelium, stress resistance, host immunomodulation, defense, and carbohydrate metabolism were identified in the genome. Interestingly, L. paragasseri UBLG-36 also harbored genes that code for enzymes involved in oxalate catabolism, such as formyl coenzyme A transferase (frc) and oxalyl coenzyme A decarboxylase (oxc). In vitro oxalate degradation assay showed that UBLG-36 is highly effective in degrading oxalate (averaging more than 45% degradation), a feature that has not been reported before. As a recently identified bacterium, there are limited genomic reports on L. paragasseri, and our draft genome sequence analysis is the first to describe and emphasize the probiotic potential and oxalate degrading ability of this species. With results supporting the probiotic functionalities and oxalate catabolism of UBLG-36, we propose that this strain is likely to have immense biotechnological applications upon appropriate characterization., Competing Interests: The authors have declared that no competing interests exist.

Gut dysbiosis with disrupted enterohepatic bile acid metabolism is commonly associated with non-alcoholic fatty liver disease (NAFLD) and recapitulated in a NAFLD-phenotype elicited by 2,3,7,8-tetrachlorodibenzo- p -dioxin (TCDD) in mice. TCDD induces hepatic fat accumulation and increases levels of secondary bile acids, including taurolithocholic acid and deoxycholic acid (microbial modified bile acids involved in host bile acid regulation signaling pathways). To investigate the effects of TCDD on the gut microbiota, the cecum contents of male C57BL/6 mice orally gavaged with sesame oil vehicle or 0.3, 3, or 30 µg/kg TCDD were examined using shotgun metagenomic sequencing. Taxonomic analysis identified dose-dependent increases in Lactobacillus species (i.e., Lactobacillus reuteri ). Increased species were also associated with dose-dependent increases in bile salt hydrolase sequences, responsible for deconjugation reactions in secondary bile acid metabolism. Increased L. reuteri levels were further associated with mevalonate-dependent isopentenyl diphosphate (IPP) biosynthesis and o-succinylbenzoate synthase, a menaquinone biosynthesis associated gene. Analysis of the gut microbiomes from cirrhosis patients identified an increased abundance of genes from the mevalonate-dependent IPP biosynthesis as well as several other menaquinone biosynthesis genes, including o-succinylbenzoate synthase. These results extend the association of lactobacilli with the AhR/intestinal axis in NAFLD progression and highlight the similarities between TCDD-elicited phenotypes in mice to human NAFLD.

Background: Vaginal microbiota (VMB) composition is altered in women with cervical intra-epithelial neoplasia (CIN) compared to healthy controls and is associated with disease progression. However, the impact of CIN excision on the VMB and innate immunity is not known. This observational study aims to explore the impact of CIN excision on the VMB, antimicrobial peptides (AMP) and proinflammatory cytokines., Methods: We sampled 103 non-pregnant, premenopausal women at the time of excisional treatment for CIN and at their 6-month follow-up visit. A further 39 untreated controls with normal cytology were also sampled. We used metataxonomics to group vaginal swab samples into community state types (CSTs) and ELISA to quantify cytokine and AMP levels in matched vaginal secretions. Analyses were performed to compare the bacterial composition and immune analyte levels before and after CIN excision and in healthy controls., Results: Women with CIN had significantly higher rates of Lactobacillus species depletion pre-treatment compared to healthy controls (CST IV 21/103, 20% vs 1/39, 3%, p = 0.0081). Excision did not change the VMB composition, with CST IV remaining significantly more prevalent after excision compared to untreated, healthy controls (CST IV 19/103, 20% vs 1/39, 3%, p = 0.0142). Prevotella bivia and Sneathia amnii were significantly higher in samples before treatment compared to untreated controls, and Prevotella bivia remained significantly higher amongst the treated, with less Lactobacillus crispatus compared to untreated controls. IL-1β and IL-8 remained significantly elevated pre- (p < 0.0001 and p = 0.0014, respectively) and post-treatment (p < 0.0001 and p = 0.0035, respectively) compared to untreated controls. Levels of human beta-defensin-1 and secretory leukocyte protease inhibitor were both significantly reduced following CIN excision (p < 0.0001); however, their levels remained lower than controls post-treatment., Conclusions: Women with CIN have an increased prevalence of Lactobacillus sp. depletion, high-diversity VMB composition, and higher levels of proinflammatory cytokines and AMPs compared to normal controls. Surgical excision of the disease reduces levels of vaginal AMPs but does not alter VMB composition or cytokine levels. These findings suggest that women with CIN have an inherent predisposition to a high-diversity proinflammatory environment that is not corrected by disease excision. The failure to re-establish a Lactobacillus-enriched CST may explain why women remain at high risk of pre-invasive and invasive disease recurrence., (© 2021. The Author(s).)

Objectives: The aim of this study was to characterize cervical microbiome feature of reproductive-age women in the progression of squamous intraepithelial lesions (SIL) to cervical cancer., Methods: We characterized the 16S rDNA cervical mucus microbiome in 94 participants (age from 18 to 52), including 13 cervical cancer (CA), 31 high-grade SIL (HSIL), 10 low-grade SIL (LSIL), 12 HPV-infected (NH) patients and 28 healthy controls (NN). Alpha (within sample) diversity was examined by Shannon and Simpson index, while Beta (between sample) diversity by principle coordinate analysis (PCoA) of weighted Unifrac distances. Relative abundance of microbial taxa was compared using Linear Discriminant Analysis Effect Size (LEfSe). Co-occurrence analysis was performed to identify correlation among marker genera, and Phylogenetic investigation of communities by reconstruction of unobserved states (PICRUSt) to explore functional features and pathways of cervical microbiota., Results: Alpha diversity(p < 0.05) was higher in severer cervical pathology with lower relative abundance of Lactobacillus as well as higher of anaerobes. Beta diversity (p < 0.01) was significantly different. Marker genera were identified including Porphyromonas, Prevotella and Campylobacter of CA and Sneathia of HSIL. The correlation of differential functional pathways with Prevotella was opposite to that with Lactobacillus., Conclusion: Our study suggests differences in cervical microbiota diversity and relative abundance of reproductive-age females in different stages of cervical carcinogenesis. Marker genera might participate in the lesion progression and will be helpful for diagnosis, prevention and treatment. These findings may lead the way to further study of the cervical microbiome in development of cervical cancer., Competing Interests: Declaration of competing interest The authors report no conflicts of interest, financial or otherwise, related to the subject matter of the article submitted., (Copyright © 2018. Published by Elsevier Inc.)

The composition of the vaginal microbiota is a key element for maintaining gynecological and reproductive health. With the aim of obtaining an accurate overview of the vaginal microbiota of Algerian women, in terms of their age and ethnic group, we conducted a 16S rRNA gene targeted metagenomic analysis of 100 vaginal samples taken from healthy childbearing and menopausal women. These data were used to establish the pattern of the vaginal microbiota during reproductive and postreproductive phases. Hormone levels were correlated to changes in microbial composition for menopausal women. The ethnic comparison revealed a particular microbiota profile for Algerian women, with a dominance of CST III and CST I. A rapid qPCR method developed by the authors was successfully used to identify the vaginal bacterial pattern for a customized gynecological management., (© 2020. Springer Science+Business Media, LLC, part of Springer Nature.)

Cervical cancer is an important health concern worldwide and is one of the leading causes of death in Mexican women. Previous studies have shown changes in the female genital tract microbe community related to human papillomavirus (HPV) infection and cervical cancer; yet, this link remains unexplored in many human populations. This study evaluated the vaginal bacterial community among Mexican women with precancerous squamous intraepithelial lesions (SIL). We sequenced the V3 region of the 16S rRNA gene in cervical samples from 228 Mexican women, including 121 participants with SIL, most of which were HPV positive, and 107 healthy women without HPV infection or SIL. The presence of SIL was associated with changes in composition (beta diversity) and with a higher species richness (Chao1). A comparison of HPV-positive women with and without SIL showed that microbiota changes occurred even in the absence of SIL. Multivariate association with linear models (MaAsLin) analysis yielded independent associations between HPV infection and an increase in the relative abundance of Brachybacterium conglomeratum and Brevibacterium aureum as well as a decrease in two Lactobacillus iners operational taxonomic units (OTUs). We also identified a positive independent association between HPV-16, the most common HPV subtype linked to SIL, and Brachybacterium conglomeratum. Our work indicates that HPV infection leading to SIL is primarily associated with shifts in vaginal microbiota composition, some of which may be specific to this human population. IMPORTANCE Human papillomavirus (HPV) plays a critical role in cervical carcinogenesis but is not sufficient for cervical cancer development, indicating the involvement of other factors. The vaginal microbiota is an important factor in controlling infections caused by HPV, and, depending on its composition, it can modulate the microenvironment in vaginal mucosa against viral infections. Ethnic and sociodemographic factors influence differences in vaginal microbiome composition, which underlies the dysbiotic patterns linked to HPV infection and cervical cancer across different populations of women. Here, we provide evidence for associations between vaginal microbiota patterns and HPV infection linked to ethnic and sociodemographic factors. To our knowledge, this is the first report of the species Brevibacterium aureum and Brachybacterium conglomeratum linked to HPV infection or squamous intraepithelial lesions (SIL).

Egg white peptides (EWPs) can be effectively used to alleviate and treat inflammatory diseases due to their anti-oxidation, anti-inflammation, and microbiota regulation capabilities. A dextran sodium sulfate (DSS)-induced colitis model was used to clarify the regulatory effects of EWPs on colitis. Forty-three peptide sequences were identified from EWPs using LC-MS/MS. The results demonstrated that EWPs decreased the levels of pro-inflammatory cytokines and the extent of crypt damage in a dose-dependent manner. 16S rRNA gene sequencing results indicated that 200 mg/kg EWPs significantly increased the relative abundance of beneficial bacteria Lactobacillus and Candidatus&#95;Saccharimonas, and reduced the relative abundance of pathogenic bacteria Ruminiclostridium and Akkermansia. In addition, the degree of correlation between pro-inflammatory cytokines and microbiota was as follows: interleukin (IL)-1β > IL-8 > IL-6 > tumor necrosis factor-α To summarize, EWPs contributed to the alleviation of colitis symptoms and the intestinal injury through anti-inflammatory effects, repair of intestinal mucosa, and modulation of gut microbiota., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Lactic acid bacteria (LAB) in the natural environment meet multiple stressors such as pH and temperature variations, increased nutrition and metabolite concentrations, harmful chemicals, acidic/oxidative conditions, osmotic pressure, and starvation. However, LAB strains are not subjected to high hydrostatic pressure (HHP) which currently is the most common non-thermal decontamination technology in the food industry. In this context, the LAB response to HHP is more difficult to identify compared to other stress-induced responses, and dnaK , ctsR , and hrcA can serve as essential regulators in this reaction. In the present study, the expression level of dnaK , ctsR , and hrcA mRNAs in 15 LAB strains after the HHP (300 MPa/5') exposure was evaluated. As a result, the HHP-treatment affected the up-regulation of dnaK , ctsR , and hrcA in L. backii KKP 3565, L. backii KKP 3566, L. rhamnosus KKP 3570, L. brevis KKP 3575 strains, whereas, in L. plantarum KKP 3569, L. rhamnosus KKP 3571, L. brevis KKP 3573 all genes were lower expressed. The relative expression level of the dnaK , ctsR , and hrcA either before or after the pressure treatment for L. brevis DSM 6235, L. rhamnosus KKP 3572, L. brevis KKP 3574, L. brevis KKP 3576, L. rossiae KKP 3577, L. curvatus KKP 3578 strains were undetectable. Significant differences in the expression levels were observed, between the control and the HHP treatment strains for dnaK in L. backii KKP 3565, L. backii KKP 3566, L. plantarum KKP 3569, L. rhamnosus KKP 3570, L. rhamnosus KKP 3571, ctsR in, L. backii KKP 3565, L. rhamnosus KKP 3570, L. rhamnosus KKP 3571, and hrcA in L. plantarum KKP 3569, L. rhamnosus KKP 3571. Overall, the studied genes, dnaK , ctsR , and hrcA can be useful markers to indicate the LAB cellular response to HHP. These molecular parameters can help to optimize the desirable LAB growing conditions in industrial processes and to understand the complexity of the stress-related mechanism.

Tibetan kefir grains (TKG) are multi-functional starter cultures used in foods and have been applied in various fermentation systems. This study aimed to investigate the microbial community composition of TKG, the detoxification abilities of TKG and their isolates towards common mycotoxins, and the potential for applying TKG and their associated microbial populations to avoid mycotoxin contamination in dairy products. Cultivation-independent high-throughput sequencing of bacterial and fungal rDNA genes indicated that Lactobacillus kefiranofaciens and Kazachstania turicensis were the most abundant bacterial and fungal taxa, respectively. In addition, 27 total isolates were obtained using cultivation methods. TKG removed more than 90% of the Ochratoxin A (OTA) after 24 h, while the isolate Kazachstania unisporus AC-2 exhibited the highest removal capacity (~46.1%). Further, the isolate exhibited good resistance to acid and bile salts environment. Analysis of the OTA detoxification mechanism revealed that both adsorption and degradation activities were exhibited by TKG, with adsorption playing a major detoxification role. Furthermore, the addition of OTA did not affect the microbial community structure of TKG. These results indicate that TKG-fermented products can naturally remove mycotoxin contamination of milk and could potentially be practically applied as probiotics in fermentation products., (Copyright © 2021. Published by Elsevier Ltd.)

Bacterial genomes often reflect a bias in the usage of codons. These biases are often most notable within highly expressed genes. While deviations in codon usage can be attributed to selection or mutational biases, they can also be functional, for example controlling gene expression or guiding protein structure. Several different metrics have been developed to identify biases in codon usage. Previously we released a database, CBDB: The Codon Bias Database, in which users could retrieve precalculated codon bias data for bacterial RefSeq genomes. With the increase of bacterial genome sequence data since its release a new tool was needed. Here we present the Dynamic Codon Biaser (DCB) tool, a web application that dynamically calculates the codon usage bias statistics of prokaryotic genomes. DCB bases these calculations on 40 different highly expressed genes (HEGs) that are highly conserved across different prokaryotic species. A user can either specify an NCBI accession number or upload their own sequence. DCB returns both the bias statistics and the genome's HEG sequences. These calculations have several downstream applications, such as evolutionary studies and phage-host predictions. The source code is freely available, and the website is hosted at www.cbdb.info.

Introduction: The aim of the study was to investigate prevalence of bacteria most frequently associated with bacterial vaginosis using Amsel's criteria as well as to quantify these bacteria by real-time PCR and to explore the difference in their quantity between healthy and bacterial vaginosis samples., Methodology: For classification of vaginal discharge samples Amsel's criteria have been used. To detect and quantify Gardnerella vaginalis Atopobium vaginae, Lactobacillus spp. and total vaginal microbiome, real-time PCR has been applied., Results: According to results of our study Amsel's criteria matched well with real-time PCR diversification of healthy women and women with BV. Nevertheless, real-time PCR has been more sensitive in diagnosis of bacterial vaginosis. DNA quantification of bacteria demonstrated that mutual abundance of G.vaginalis and A. vaginae was good bacterial vaginosis marker . On the contrary, Lactobacillus spp. was present in high amount in both healthy and bacterial vaginosis samples, but ratio of investigated bacteria was different between them. In fact, G. vaginalis and A. vaginae comprised only 0.1% of total microbiome in healthy, whereas Lactobacillus spp. took 99.3% of it. Nonetheless, in bacterial vaginosis, G. vaginalis and A. vaginae made up 34.4% of total microbiome, while Lactobacillus spp. was 21.6%., Conclusions: According to the results of our study real-time PCR analysis was more sensitive in diagnosis of bacterial vaginosis than Amsel's method, as well as it represented fine tool in making a difference between microbial entities in healthy and bacterial vaginosis samples., Competing Interests: No Conflict of Interest is declared, (Copyright (c) 2021 Nedzib Numanovic, Snezana Ribis, Jelena Cukic, Dane Nenadic, Aleksandar Zivanovic, Predrag Sazdanovic, Violeta Ninkovic, Dejan Baskic.)

Delivery by cesarean section (CS) is linked to an increased incidence of food allergies in children and affects early gut microbiota colonization. Furthermore, emerging evidence has connected disordered intestinal microbiota to food allergies. Here, we investigated the impact of CS on a rat model for food allergy to ovalbumin (OVA). Rats delivered by CS were found to be more responsive to OVA sensitization than vaginally born ones, displaying a greater reduction in rectal temperature upon challenge, worse diarrhea, and higher levels of OVA-specific antibodies and histamine. 16S rRNA sequencing of feces revealed reduced levels of Lactobacillus and Bifidobacterium in the CS rats. Preventative supplementation with a probiotic combination containing Lactobacillus and Bifidobacterium could protect CS rats against an allergic response to OVA, indicating that the microbiota dysbiosis contributes to CS-related response. Additionally, probiotic intervention early in life might help to rebuild aberrant Th2 responses and tight junction proteins, both of which have been linked to CS-related high allergic reactions. Taken together, this study shows that disordered intestinal microbiota plays an essential role in the pathogenesis of food allergy mediated by CS. More importantly, interventions that modulate the microbiota composition in early life are therapeutically relevant for CS-related food allergies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Jin, Li, Xia, Li, Zhao, Li, Li, Li, Zhou, Fu, Li and Li.)

The microbiota in the human body play critical roles in many physiological and pathological processes. However, the diversity and dynamics of the female genital tract (FGT) microbiota have not been fully unveiled. In this study, we characterized the microbiome variations in reproductive-aged Chinese women, and we revealed that the cervicovaginal microbiota were dominated by Lactobacillus . Overall, the composition of microbiota in the uterine cavity was more diverse than that in the vagina and cervix. A positive correlation between Lactobacillus iners and Lactobacillus crispatus was observed in both the vagina and the cervix, suggesting that these two species might have a symbiotic relationship in the cervicovaginal microbiota. Moreover, we, for the first time, stratified the reproductive-aged Chinese women into subgroups, based on their microbiome profiles. Furthermore, we identified the bacteria whose abundance changed in the uterine cavity of infertile patients when compared with healthy controls, such as L. iners and L. crispatus . Functionally, the metabolism-related pathways, neurotrophin signaling pathway, and adipocytokine signaling pathway were predominantly dysregulated in the uterine cavity of infertile patients. In conclusion, we characterized a comprehensive microbial landscape in FGT, as well as their functional roles in female infertility of the Chinese population., Competing Interests: Author HY was employed by company SPH Sine Pharmaceutical, Laboratories Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Sun, Ding, Yu, Ji, Xifang, Pang, Wang, Zhang and Li.)

Background: Lactobacillus was described as a keystone bacterial taxon in the human vagina over 100 years ago. Using metagenomics, we and others have characterized lactobacilli and other vaginal taxa across health and disease states, including pregnancy. While shifts in community membership have been resolved at the genus/species level, strain dynamics remain poorly characterized., Methods: We performed a metagenomic analysis of the complex ecology of the vaginal econiche during and after pregnancy in a large U.S. based longitudinal cohort of women who were initially sampled in the third trimester of pregnancy, then validated key findings in a second cohort of women initially sampled in the second trimester of pregnancy., Findings: First, we resolved microbial species and strains, interrogated their co-occurrence patterns, and probed the relationship between keystone species and preterm birth outcomes. Second, to determine the role of human heredity in shaping vaginal microbial ecology in relation to preterm birth, we performed a mtDNA-bacterial species association analysis. Finally, we explored the clinical utility of metagenomics in detection and co-occurrence patterns for the pathobiont Group B Streptococcus (causative bacterium of invasive neonatal sepsis)., Conclusions: Our highly refined resolutions of the vaginal ecology during and post-pregnancy provide insights into not only structural and functional community dynamics, but highlight the capacity of metagenomics to reveal finer aspects of the vaginal microbial ecologic framework., Funding: NIH-NINR R01NR014792, NIH-NICHD R01HD091731, NIH National Children's Study Formative Research, Burroughs Wellcome Fund Preterm Birth Initiative, March of Dimes Preterm Birth Research Initiative, NIH-NIGMS (K12GM084897, T32GM007330, T32GM088129)., Competing Interests: Declaration of Interests The authors declare no competing interests.

The composition of the maternal vaginal microbiome influences the duration of pregnancy, onset of labor, and even neonatal outcomes. Maternal microbiome research in sub-Saharan Africa has focused on non-pregnant and postpartum composition of the vaginal microbiome. Here we aimed to illustrate the relationship between the vaginal microbiome of 99 laboring Ugandan women and intrapartum fever using routine microbiology and 16S ribosomal RNA gene sequencing from two hypervariable regions (V1-V2 and V3-V4). To describe the vaginal microbes associated with vaginal microbial communities, we pursued two approaches: hierarchical clustering methods and a novel Grades of Membership (GoM) modeling approach for vaginal microbiome characterization. Leveraging GoM models, we created a basis composed of a preassigned number of microbial topics whose linear combination optimally represents each patient yielding more comprehensive associations and characterization between maternal clinical features and the microbial communities. Using a random forest model, we showed that by including microbial topic models we improved upon clinical variables to predict maternal fever. Overall, we found a higher prevalence of Granulicatella, Streptococcus, Fusobacterium, Anaerococcus, Sneathia, Clostridium, Gemella, Mobiluncus, and Veillonella genera in febrile mothers, and higher prevalence of Lactobacillus genera (in particular L. crispatus and L. jensenii), Acinobacter, Aerococcus, and Prevotella species in afebrile mothers. By including clinical variables with microbial topics in this model, we observed young maternal age, fever reported earlier in the pregnancy, longer labor duration, and microbial communities with reduced Lactobacillus diversity were associated with intrapartum fever. These results better defined relationships between the presence or absence of intrapartum fever, demographics, peripartum course, and vaginal microbial topics, and expanded our understanding of the impact of the microbiome on maternal and potentially neonatal outcome risk., (© 2021. The Author(s).)

Obesity is a major public health problem related to various chronic health conditions. Lactobacillus species has been reported in obese individuals; however, its role is unknown. We compared the abundance and composition of Lactobacillus species by analyzing feces from 64 healthy control subjects and 88 obese subjects. We isolated one Lactobacillus strain from the feces of a subject with obesity and further analyzed its genetic and molecular features. We found that an increased abundance and higher prevalence of Lactobacillus sakei distinguished the fecal microbiota of the obese group from that of healthy subjects and that it was related to the increased levels of reactive oxygen species (ROS) induced by higher fat intake. The L. sakei ob4.1 strain, isolated from the feces of a subject with obesity, showed high catalase activity, which was regulated by oxidative stress at the gene transcription level. L. sakei ob4.1 maintained colon epithelial cell adhesion ability under ROS stimulation, and treatment with saturated fatty acid increased colon epithelial ROS levels in a dose-dependent manner; however, L. sakei ob4.1 did not change the level of fat-induced colon epithelial ROS. Exposing mice to a high-fat diet revealed that high-fat-diet-induced colon ROS was associated with the increased colonization of L. sakei ob4.1 through catalase activity. Four-week supplementation with this strain in mice fed a high-fat diet did not change their body weights or ROS levels. A high-fat diet induces changes in the colon environment by increasing ROS levels, which provides a colonization benefit to an L. sakei strain with high catalase activity. IMPORTANCE Lactobacillus provides many health benefits; its various species are widely used as probiotics. However, an increased abundance of Lactobacillus has been reported in obesity, and the role of Lactobacillus strains in obesity remains unknown. We found a high abundance of the Lactobacillus sakei species in a group of obese subjects and examined its relationship with a high-fat diet and reactive oxygen species (ROS) in the feces. To find the underlying mechanism, we analyzed and characterized an L. sakei strain isolated from a severely obese individual. We found that higher gut oxidative stress could link high-fat-diet-induced obesity and L. sakei. This translational research identifies the roles of the host gut environment in the colonization and survival of L. sakei .

Daqu, a brick-shaped product spontaneously fermented under an open environment, has been regarded as the starter of fermentation, raw enzyme preparation and raw materials for baijiu production. However, its contribution in baijiu fermentation has not been fully elaborated yet. Here, the effects of daqu microbiota on baijiu fermentation were investigated under both field-scale and lab-scale conditions. In field-scale baijiu fermentation, the dominant daqu microbes (average relative abundance>10.0%), including unclassified&#95;Leuconostocaceae, Thermoascus, and Thermomyces, tended to dominate the early stage (0-7 d). However, the rare daqu microbes (average relative abundance <0.1%, e.g., Kazachstania) tended to dominate the middle and late stages (11-40 d). In addition, some genera showed differences in species diversity between daqu and fermented grains. The average relative abundance of Lactobacillus was over 75% during baijiu fermentation, and most of them were affiliated with Lactobacillus acetotolerans, while Lactobacillus crustorum dominated the Lactobacillus OTUs in daqu. The similar patterns were also observed during lab-scale baijiu fermentation. The results of function prediction showed the enriched metabolic pathways were associated with glycolysis and long-chain fatty acid esters in baijiu fermentation. These results improved the understanding of daqu microbiota function during baijiu fermentation and provided a basic theory to support the regulation of baijiu production., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Fructansucrases (FSs), including inulosucrase (IS) and levansucrase (LS), are the members of the Glycoside Hydrolase family 68 (GH68) enzymes. IS and LS catalyze the polymerization of the fructosyl moiety from sucrose to inulin- and levan-type fructans, respectively. Lactobacillus-derived FSs have relatively extended N- and C-terminal sequences. However, the functional roles of these sequences in their enzymatic properties and fructan biosynthesis remain largely unknown. Limosilactobacillus reuteri (basionym: Lactobacillus reuteri) 121 could produce both IS and LS, abbreviated as Lare121-IS and Lare121-LS, respectively. In this study, it was found that the terminal truncation displayed an obvious effect on their activities and the N-terminal truncated variants, Lare121-ISΔ177-701 and Lare121-LSΔ154-686, displayed the highest activities. Melting temperature (T m ) and the thermostability at 50 °C were measured to evaluate the stability of various truncated versions, revealing the different effects of N-terminal on the stability. The average molecular weight and polymerization degree of the fructans produced by different truncated variants did not change considerably, indicating that N-terminal truncation had low influence on fructan biosynthesis. In addition, it was found that N-terminal truncation could also improve the activity of other reported FSs from Lactobacillus species., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Clinical data exist that support the utility of topical probiotics for certain dermatological diseases such as atopic dermatitis, acne, and psoriasis. However, there is paucity of data on the use of live lactobacilli to control axillary malodor. The objective of this study was to determine whether application of topical oil-based cream containing live Lactobacilli could decrease malodor-producing bacteria in the axilla of healthy subjects., Aims: To determine the effects of topical cream with live lactobacilli on malodor-producing bacteria in the axilla of healthy subjects., Patients/methods: Twenty-five adult volunteers comprising 12 males and 13 females provided informed consent. Axillary skin swabs were collected before and after 14 days application of topical cream containing live Lactobacillus pentosus KCA1. Bacterial DNA was extracted, and V4 region of the 16S rRNA was amplified and sequenced in a pair-end configuration on the Illumina MiSeq platform rendering 2 × 150 bp sequences. Microbial taxonomy to species level was generated using the Greengenes database. Linear discriminant analysis (LDA) effect size (LEfSe) was used to identify biologically and statistically significant differences in relative abundance., Results: Actinobacteria decreased from 70% to 24%, and Firmicutes increased from 26.6% to 73.9% among the female participants. In males, Actinobacteria decreased from 65% to 38%, while Firmicutes increased from 24% to 57%. Corynebacterium decreased from 62.91% to 36.63%, while Lactobacillus increased from 0.06% to 23.11%. In males, unliked females, there were reduction of Staphylococcus species associated with malodor, notably Staphylococcus hominis, Staphylococcus hemolyticus, and Staphylococcus lugdunensis. Bacterial functional gene- Pyridoxal protein dependent enzymes involved in biotransformation of malodor precursor to volatile thioalcohols were down-regulated., Conclusions: Application of Lactobacillus pentosus KCA1 cream led to a significant decrease in the relative abundance of odor-producing Corynebacterium species in both female and male subjects. Some species associated with malodor especially Corynebacterium striatum, Corynebacterium jeikeium, Corynebacterium tuberculostearicum, and Staphylococcus hominis decreased by 96%, 73%, 7%, and 20.8%, respectively, in males., (© 2021 Wiley Periodicals LLC.)

Cervical microbiota (CM) are considered an important factor affecting the progression of cervical intraepithelial neoplasia (CIN) and are implicated in the persistence of human papillomavirus (HPV). Collection of liquid-based cytology (LBC) samples is routine for cervical cancer screening and HPV genotyping and can be used for long-term cytological biobanking. We sought to determine whether it is possible to access microbial DNA from LBC specimens, and compared the performance of four different extraction protocols: (ZymoBIOMICS DNA Miniprep Kit; QIAamp PowerFecal Pro DNA Kit; QIAamp DNA Mini Kit; and IndiSpin Pathogen Kit) and their ability to capture the diversity of CM from LBC specimens. LBC specimens from 20 patients (stored for 716 ± 105 days) with CIN values of 2 or 3 were each aliquoted for each of the four kits. Loss of microbial diversity due to long-term LBC storage could not be assessed due to lack of fresh LBC samples. Comparisons with other types of cervical sampling were not performed. We observed that all DNA extraction kits provided equivalent accessibility to the cervical microbial DNA within stored LBC samples. Approximately 80% microbial genera were shared among all DNA extraction protocols. Potential kit contaminants were observed as well. Variation between individuals was a significantly greater influence on the observed microbial composition than was the method of DNA extraction. We also observed that HPV16 was significantly associated with community types that were not dominated by Lactobacillus iners., Competing Interests: The authors would like to declare the following patents/patent applications associated with this research: M.N. is one of the inventors named in the patents and patent applications for the HPV therapeutic vaccine PepCan. Patent issued: Human Papilloma Virus Therapeutic Vaccine Nakagawa, M. and Chang, B.S. Patent No. 9,974,849 issued on 5/22/2018 Patent application: Human Papilloma Virus Therapeutic Vaccine Nakagawa, M. and Chang, B.S. International Application (PCT/US14/60198) filed on 10/11/2014. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Refractory responses to standard-of-care oral metronidazole among recurrent bacterial vaginosis (BV) patients is not rare, and recurrence within a year is common. A better understanding of the bacterial determinants of these outcomes is essential. In this study we ask whether changes in specific species of Gardnerella are associated with poor short or long term clinical outcomes, and if and how resurgence of Lactobacillus species affects these outcomes. We quantify Lactobacillus isolates as a proportion of total vaginal bacteria using the LbRC5 qPCR assay, and 5 prevalent species of Gardnerella using primers that target species-specific polymorphisms within the cpn60 gene. The study includes 43 BV patients: 18 refractory, 16 recurrent, and 11 remission patients, sampled daily for up to two weeks post-treatment; clinical outcomes were tracked for up to 9 months. Persistently high titers of Gardnerella Gsp07 were associated with refractory responses, and persistently low abundance of Gardnerella Gsp07 and G. swidsinskii / G. leopoldii were associated with remission. Lactobacillus species abundance rose in 4-14 days after initiation of treatment in most but not all recurrent and remission patients, although increases were more sustained among remission patients. The findings suggest that Gardnerella Gsp07 and G. swidsinskii / G. leopoldii are markers of poor clinical outcome or may directly or indirectly suppress recovery of Lactobacillus species, thereby interfering with clinical recovery. Therapies that target these strains may improve patient outcome., Competing Interests: The authors have declared that no competing interests exist.

In order to explore the correlation between the production of propionic acid (PA) and the succession of bacterial community during the fermentation of gray sufu, high-throughput sequencing and HPLC (High Performance Liquid Chromatography) were used to monitor the changes of bacterial community and metabolite content. The abundance and metabolite concentration of Propionibacterium increased rapidly in the early stage of fermentation. In the middle stage, the abundance of Lactobacillus began to increase, while the pH decreased rapidly. In the late stage, the concentration of PA began to decrease, but it remained at a high level at the end of fermentation. Correlation analysis showed that Lactobacillus and Bacillus had a strong negative correlation with PA and its precursor. The results showed that Fusobacterium, Providencia, Lactobacillus and Bacillus could be the key factors to reduce the PA content. This study provides a new idea for the quality control of traditional fermented food., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Human beta defensins (hBDs) may play an important role in the progression of lichen sclerosus (LS), due to their ability to induce excessive stimulation of extracellular matrix synthesis and fibroblast activation. The genetic ability of the individual to produce defensins, the presence of microbes influencing defensin production, and the sensitivity of microbes to defensins together regulate the formation of an ever-changing balance between defensin levels and microbiome composition. We investigated the potential differences in postmenopausal vaginal microbiome composition and vaginal hBD levels in LS patients compared to non-LS controls. LS patients exhibited significantly lower levels of hBD1 (p = 0.0003), and significantly higher levels of hBD2 (p = 0.0359) and hBD3 (p = 0.0002), compared to the control group. The microbiome of the LS patients was dominated by possibly harmful bacteria including Lactobacillus iners, Streptococcus anginosus or Gardnerella vaginalis known to initiate direct or indirect damage by increasing defensin level production. Our observations highlight that correcting the composition of the microbiome may be applicable in supplementary LS therapy by targeting the restoration of the beneficial flora that does not increase hBD2-3 production., (© 2021. The Author(s).)

Adhesion to host surfaces for bacterial survival and colonization involves a variety of molecular mechanisms. Ligilactobacillus ruminis, a strict anaerobe and gut autochthonous (indigenous) commensal, relies on sortase-dependent pili (LrpCBA) for adherence to the intestinal inner walls, thereby withstanding luminal content flow. Here, the LrpCBA pilus is a promiscuous binder to gut collagen, fibronectin and epithelial cells. Structurally, the LrpCBA pilus displays a representative hetero-oligomeric arrangement and consists of three types of pilin subunit, each with its own location and function, i.e. tip LrpC for adhesion, basal LrpB for anchoring and backbone LrpA for length. To provide further structural insights into the assembly, anchoring and functional mechanisms of sortase-dependent pili, each of the L. ruminis pilus proteins was produced recombinantly for crystallization and X-ray diffraction analysis. Crystals of LrpC, LrpB, LrpA and truncated LrpA generated by limited proteolysis were obtained and diffracted to resolutions of 3.0, 1.5, 2.2 and 1.4 Å, respectively. Anomalous data were also collected from crystals of selenomethionine-substituted LrpC and an iodide derivative of truncated LrpA. Successful strategies for protein production, crystallization and derivatization are reported.

Lactic acid bacteria (LAB) are a heterogeneous group of bacteria which are Gram-positive, facultative anaerobes and non-motile, non-spore forming, with varied shapes from cocci to coccobacilli and bacilli. Lactobacillus is the largest and most widely used bacterial species amongst LAB in fermented foods and beverages. The genus is a common member of human gut microbiome. Several species are known to provide benefits to the human gut via synergistic interactions with the gut microbiome and their ability to survive the gut environment. This ability to confer positive health effects provide them a status of generally recognized as safe (GRAS) microorganisms. Due to their various beneficial characteristics, other factors such as their resistance acquisition were overlooked. Overuse of antibiotics has made certain bacteria develop resistance against these drugs. Antibiotic resistance was found to be acquired mainly through conjugation which is a type of lateral gene transfer. Several in vitro methods of conjugation have been discussed previously depending on their success to transfer resistance. In this review, we have addressed methods that are employed to study the transfer of resistance genes using the conjugation phenomenon in lactobacilli., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Typical traditional probiotics lactobacilli and bifidobacteria are gaining great interest to be developed as living diagnostics and therapeutics for improving human health. However, the mechanistic basis underlying their inherent health beneficial property remain incompletely understood which can slow down the translational pipeline in the functional food and pharmaceutical field. Efficient genome editing will advance the understanding of the molecular mechanism of the probiotics' physiological properties and their interaction with the host and the host microbiota, thereby further promote the development of next-generation designer probiotics with improved robustness and tailored functionalities. With the expansion of genome editing strategies such as CRISPR-Cas-based tools and IPSD assisted genome engineering as well as other synthetic biology technologies, the research and application of these health-promoting bacteria for the food and pharmaceutical industry will be further enhanced., (Copyright © 2020 The Author. Published by Elsevier Ltd.. All rights reserved.)

Approximately 15 % of clinically recognized pregnancies end in miscarriage. To explore the vaginal microbiota profile in women diagnosed with early pregnancy miscarriage (including missed miscarriage [M] or empty-sac miscarriage [E]), the microbial community structure in vaginal fluid was evaluated by Illumina MiSeq sequencing of the 16S rRNA gene V4 region and compared with that in women with normal pregnancy (P). Taxa identified in samples from the P, E, and M groups formed distinct clusters. The M group had the highest bacterial species richness and diversity, with lower Lactobacillus levels and higher Bacteroides, Halomonas, Miscellaneous-Crenarchaeota, Bacillus, Staphylococcus, Escherichia/Shigella, and Acetobacter levels than in the other two groups. The vaginal community-state types differed significantly among the three groups (P = 0.02) but were similar between the P and E groups (P = 0.21). Moreover, we identified an optimal marker set composed of 12 operational taxonomic units based on a random forest model that distinguished the M and P groups, with areas under the receiver-operating characteristic curve of 86.76 % and 93.33 % in the training and test groups, respectively. In conclusion, this study highlights that patients with early pregnancy miscarriage had a significantly different vaginal microbiota profile. Microbial markers analyzed by RT-qPCR may be applied for the etiological diagnosis of miscarriage. Further studies should be performed to elucidate the possible mechanism of special strains affecting miscarriage during early pregnancy., (Copyright © 2021. Published by Elsevier Ltd.)

The features of the vaginal microbiota (VM) community can reflect health status, and they could become new biomarkers for disease diagnosis. During pregnancy, domination of bacteria of the genus Lactobacillus in the VM community is regarded as a keystone because they stabilize the VM by producing antimicrobial compounds and competing adhesion. An altered VM composition provides a marker for adverse pregnancy outcomes. This nested case-control study aimed to characterize the VM in women with a tubal pregnancy (TP) presenting with pain and/or uterine bleeding in early pregnancy. Chinese women with a symptomatic early pregnancy of unknown location were the study cohort. 16S rDNA gene-sequencing of V3-V4 variable regions was done to assess the diversity, structures, taxonomic biomarkers, and classification of the VM community. The primary outcome was the location of the early pregnancy. The VM community in women with a TP showed higher diversity (PD-whole-tree, median: 8.26 vs . 7.08, P = 0.047; Shannon Diversity Index, median: 1.43 vs 0.99, P = 0.03) and showed different structures to those in women with an intrauterine pregnancy (IUP) ( R = 0.23, P < 0.01). Bacteria of the genus Lactobacillus were significantly enriched in the IUP group, whereas bacteria of the genera Gardnerella and Prevotella were significantly enriched in the TP group. Lactobacillus abundance could be used to classify the pregnancy location (AUC = 0.81). Non- Lactobacillus -dominated microbiota (≤ 0.85% Lactobacillus ) was significantly associated with a TP (adjusted odds ratio: 4.42, 95% confidence interval: 1.33 to 14.71, P = 0.02). In conclusion, among women with a symptomatic early pregnancy, a higher diversity and lower abundance of Lactobacillus in the VM is associated with a TP., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Ruan, Zhang, Chen, Liu, Zhu, Huang, Liu, Luo, Deng and Gao.)

Background: In recent years, researchers are interested in the discovery of active compounds from traditional remedies and natural sources, as they reveal higher therapeutic efficacies and improved toxicological profiles. Among the various traditional treatments that have been widely studied and explored for their potential therapeutic benefits, kefir, a fermented beverage, demonstrates a broad spectrum of pharmacological properties, including antioxidant, anti-inflammation, and healing activities. These health-promoting properties of kefir vary among the kefir cultures found at the different part of the world as different media and culture conditions are used for kefir maintenance and fermentation., Methods: This study investigated the microbial composition and readily found bioactive compounds in water kefir fermented in Malaysia using 16S rRNA microbiome and UHPLC sequencing approaches. The toxicity effects of the kefir water administration in BALB/c mice were analysed based on the mice survival, body weight index, biochemistry profile, and histopathological changes. The antioxidant activities were evaluated using SOD, FRAP, and NO assays., Results: The 16S rRNA amplicon sequencing revealed the most abundant species found in the water kefir was Lactobacillus hilgardii followed by Lactobacillus harbinensis, Acetobacter lovaniensis, Lactobacillus satsumensis, Acetobacter tropicalis, Lactobacillus zeae, and Oenococcus oeni. The UHPLC screening showed flavonoid and phenolic acid derivatives as the most important bioactive compounds present in kefir water which has been responsible for its antioxidant activities. Subchronic toxicity study showed no toxicological signs, behavioural changes, or adverse effects by administrating 10 mL/kg/day and 2.5 mL/kg/day kefir water to the mice. Antioxidants assays demonstrated enhanced SOD and FRAP activities and reduced NO level, especially in the brain and kidney samples., Conclusions: This study will help to intensify the knowledge on the water kefir microbial composition, available phytochemicals and its toxicological and antioxidant effects on BALB/c mice since there are very limited studies on the water kefir grain fermented in Malaysia.

The objective of this study was to investigate the effect of polysaccharide extracts from persimmon (PPE) on the proliferation of Lactobacillus and the gut microbiota of mice. Lactobacillus strains were cultured in medium containing PPE, and differential gene expression was evaluated using transcriptomics. In addition, 16S rDNA was employed to analyze the abundance and diversity of fecal colonies in mice, and the influence of PPE on the intestinal flora in mice was further examined. The results showed that Lactobacillus acidophilus NCFM and Lactobacillus acidophilus CICC 6075 could proliferate in PPE medium. Gene ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomics (KEGG) pathway analysis indicated that glucose metabolism-related genes, such as phosphoyruvate hydratase (eno) and PTS mannose transporter subunit IIAB (manX), were up-regulated. The metabolic pathways of fructose and mannose were also significantly up-regulated. After gavage of mice with PPE, 16S rDNA sequencing of mouse feces indicated that the beneficial bacteria in the intestines proliferated and the abundance of harmful bacteria was reduced. PPE can maintain the balance of intestinal microorganisms in mice. Therefore, PPE has a significant positive effect on both Lactobacillus proliferation and gut microbiota of mice., (Copyright © 2021 Elsevier B.V. All rights reserved.)