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Fall armyworm (FAW), Spodoptera frugiperda is a generalist pest known to feed on more than 300 plant species, including major staple crops such as rice, maize and sorghum. Biological control of FAW using a combination of a major indigenous egg parasitoid Telenomus remus and entomopathogenic fungi was explored in this study. Metarhizium anisopliae strains (ICIPE 7, ICIPE 41, and ICIPE 78) and Beauveria bassiana ICIPE 621 which demonstrated effectiveness to combat the pest, were evaluated through direct and indirect fungal infection to assess their pathogenicity and virulence against T. remus adults, S. frugiperda eggs and their effects on T. remus parasitism rates. Metarhizium anisopliae ICIPE 7 and ICIPE 78 exhibited the highest virulence against T. remus adults with LT50 values >2 days. ICIPE 7 induced the highest T. remus mortality rate (81.40 ± 4.17%) following direct infection with dry conidia. Direct fungal infection also had a significant impact on parasitoid emergence, with the highest emergence rate recorded in the M. anisopliae ICIPE 7 treatment (42.50 ± 5.55%), compared to the control ± (83.25 ± 5.94%). In the indirect infection, the highest concentration of 1 x 109 conidia ml-1 of ICIPE 78 induced the highest mortality (100 ± 0.00%) of T. remus adults, and the highest mortality (51.25%) of FAW eggs, whereas the least FAW egg mortality (15.25%) was recorded in the lowest concentration 1 x 105 conidia ml-1 of ICIPE 41. The number of parasitoids that emerged and their sex ratios were not affected by the different fungal strain concentrations except in ICIPE 7 at high dose. This study showed that potential combination of both M. anisopliae and B. bassiana with T. remus parasitoid can effectively suppress FAW populations.

IntroductionMicrobial pathogens invade various human organs, including the oral cavity. Candida albicans (C.a) and Streptococcus mutans (S.m) served respectively as representative oral pathogenic fungi and bacteria to stimulate dental pulp stem cells (DPSCs) and to screen the DPSC subcluster that specifically responded to fungal infection.MethodsDPSCs were obtained from the impacted third molars of six healthy subjects. Then, cells were mixed and divided into three samples, two of which were stimulated with C.a and S.m, respectively; the third sample was exposed to cell medium only (Ctrl). Single-cell mRNA sequencing analysis of treated DPSCs was performed.ResultsDPSCs were composed of four major clusters of which one, DPSC.7, exhibited unique changes compared to those of other subclusters. The DPSC.7 cell percentage of the C.a sample was twice those of the Ctrl and S.m samples. DPSC.7 cells expressed genes associated with the response to reactive oxygen species (ROS) response. DPSC.7 subgroup cells established characteristic aggregation under the stimulation of different pathogens in UMAP. The MAPK/ERK1/2 and NF-κB pathways were up-regulated, DUSP1/5/6 expressions were suppressed, FOS synthesis was activated, the immune-related pathway was induced, and the levels of cytokines, including IL-6 and CCL2, were up-regulated in DPSC.7 cells when stimulated with C.a.ConclusionsOur study analyzed the cellular and molecular properties of DPSCs infected by oral fungi and bacteria with single-cell RNA sequencing. A subcluster of DPSCs responded specifically to infections with different pathogens, activating the MAPK and NF-κB pathways to induce immune responses via the ROS pathway. This suggests novel treatment strategies for fungal infections.

BackgroundAccurate identification of infectious diseases using molecular techniques, such as PCR and NGS, is well-established. This study aims to assess the utility of Bactfast and Fungifast in diagnosing bloodstream infections in ICU settings, comparing them against traditional culture methods. The objectives include evaluating sensitivity and specificity and identifying a wide range of pathogens, including non-culturable species.MethodsWe collected 500 non-duplicate blood samples from ICU patients between January 2023 and December 2023. Specimens underwent traditional culture, MALDI-TOF, VITEK®2 compact system, and NGS-based Bactfast and Fungifast analyses.ResultsOut of the 500 samples, 26.8% (n=134) showed bacterial growth via traditional culture methods, while 4.8% (n=24) were positive for fungal growth. MALDI-TOF and VITEK®2 compact system yielded comparable results, identifying 26.4% (n=132) of specimens with bacterial growth. NGS-based Bactfast detected bacterial presence in 38.2% (n=191) of samples, including non-culturable bacteria missed by traditional methods. However, NGS-based Fungifast showed concordant fungal detection rates with culture methods. Among identified pathogens by culture method included Klebsiella pneumoniae 20.89% (n=28), Enterococcus faecalis 18.65% (n=25), Escherichia coli 15.67% (n=21), Pseudomonas aeruginosa 12.68% (n=17), Acinetobacter baumannii 10.44% (n=14), various Streptococcus species 7.46% (n=10), Mycobacterium tuberculosis 6.71% (n=9), Mycobacterium abscessus 4.47% (n=6), and Salmonella spp 2.98% (n=4). Non-culture-based NGS identified additional (n=33) pathogens, including Klebsiella pneumoniae 27.27% (n=9), Bacteroides fragilis 21.21% (n=7), Aerococcus viridans 15.15% (n=5), Elizabethkingia anopheles 12.12% (n=4), Aeromonas salmonicida 9% (n=3), Clostridium 9% (n=3), and Bacteroides vulgatus 6% (n=2). Candida albicans was reported in 5% (n=24) of samples by both methods.ConclusionNGS-based Bactfast and Fungifast demonstrate high sensitivity in identifying a wide array of bacterial and fungal pathogens in ICU patients, outperforming traditional culture methods in detecting non-culturable organisms. These molecular assays offer rapid and comprehensive diagnostic capabilities, potentially improving clinical outcomes through timely and accurate pathogen identification.

The fission yeast Schizosaccharomyces pombe produces the hydroxamate-type siderophore ferrichrome (Fc). The biosynthesis of Fc requires the Fc synthase Sib1, the ornithine-N5-oxygenase Sib2, and the N5-hydroxyornithine-N5-transacetylase Sib3. In this study, we demonstrate the critical importance of the His248 residue of Sib3 in Fc production. Cells expressing a sib3H248A mutant allele fail to grow in iron-poor media without Fc supplementation. These sib3H248A mutant cells are consistently unable to promote Fc-dependent growth of Saccharomyces cerevisiae cells in cross-feeding experiments. Green fluorescent protein (GFP)-tagged wild-type Sib3 and mutant Sib3H248A exhibit a pancellular distribution. Coimmunoprecipitation assays revealed that both wild-type and Sib3H248A physically interact with Sib2. Further analysis identified a minimal C-terminal region from amino acids 290–334 of Sib3 that is required for interaction with Sib2. Deletion mapping analysis identified two regions of Sib2 as being required for its association with Sib3. The first region encompasses amino acids 1–135, and the second region corresponds to amino acids 281–358 of Sib2. Taken together, these results describe the first example of a physical interaction between an ornithine-N5-oxygenase and an N5-hydroxyornithine-N5-transacetylase controlling the biosynthesis of a hydroxamate-type siderophore.

The massive yield of brewers’ spent grain (BSG) waste inevitably threaten environmental health. Here, solid-state fermentation (SSF) technology featuring multi-strain (MS) inoculation and high-throughput sequencing technology were employed to facilitate the sustainable and clean recycling of BSG waste while revealing the associated underlying microbiological and metabolic mechanisms. MS inoculation displayed a lower pH value (3.91 vs. 4.12) and neutral detergent fiber content (446.24 vs. 476.23 g/kg DM), a higher levels of lactic acid (86.64 vs. 33.07 g/kg DM), acetic acid (6.13 vs. 4.87 g/kg DM), propionic acid (2.78 vs. 2.18 g/kg DM) and crude protein (307.5 vs. 289.15 g/kg DM) than those in the control group. Moreover, MS inoculation inhibited the formation of non-protein-N and ammonia-N, and spoilage microorganism resuscitation, while enhanced substrate preservation. Microbiologically, during the SSF, the group treated with MS inoculation exhibited an increase in the relative abundance of Leuconostoc (0.58%∼6.60%), Weissella (6.22%∼15.42%), Enterococcus (3.15%∼9.08%), Bacillus (17.63%∼31.29%), Lactobacillus (12.89%∼8.29%), Pseudoalteromonas (12.87%∼16.29%), and a decrease in the relative abundance of Acinetobacter (0.79%∼0.02%) and Enterobacteriaceae (0.78%∼0.24%). Metabolically, starch and sucrose metabolism, arginine and proline metabolism, and phenylalanine metabolism significantly influenced the quality of extruded BSG fermented by MS during SSF. The examination of the correlation between the microbiota, metabolites, and fermentation parameters revealed that complex interactions between microbes and the environment factors impact metabolite production. Collectively, inoculating with MS improved fermentation quality and stability, facilitated the clean recycling of BSG, which is linked to complex interactions among microbes, the environment factors and metabolite production.

Rhizosphere microbiotas play vital roles in resisting environmental stress, transforming soil nutrients, and promoting plant health, growth, and productivity. The effects of rhizosphere microbial community shaping and the characteristics and functions of keystone taxa on blueberries were comprehensively studied by examining the rhizobacteria of healthy old trees (O), young seedlings (OG), and poorly growing seedlings (OB) of O’Neal blueberries. Our results showed that rhizobacterial diversity followed the order OB > > OG > O, and the microbial community of OG was similar to that of O, while that of OB was distinctly different. The predominant rhizobacteria identified included Actinobacteria, Proteobacteria, Firmicutes, Chloroflexi, and Acidobacteria. Firmicutes were highly enriched in healthy blueberries, with Bacillus identified as a key genus that significantly enhanced blueberry growth when inoculated. Bradyrhizobium and Gaiellales were common core bacteria in the blueberry rhizosphere. In contrast, Acidobacteria were the predominant phylum in poorly growing OB, with the specific Vicinamibacterales-related and Latescibacterota-related genera acting as keystone taxa that shaped the microbial community. In addition, bacterial species in Vicinamibacterales might act as a potential pathogen predicted by BugBase. Taken together, these findings provide fundamental insights into the development of the blueberry rhizosphere microbial community and highlight the role of beneficial rhizobacteria, such as Bacillus, in enhancing blueberry growth. This knowledge could contribute to the exploitation of beneficial rhizobacteria and the prevention of pathogens in modern agriculture.

Although aboveground biodiversity has been extensively studied, the impact of nutrient enrichment on soil microbial populations remains unclear. Soil microorganisms serve as important indicators in shaping soil nutrient cycling processes and are typically sensitive to nutrient additions. For this, we employed a factorial combination design to examine the impact of nutrient additions on the composition and function of soil bacteria in a temperate steppe. Nitrogen addition promoted the growth of copiotrophic bacteria (Proteobacteria, Firmicutes, and Bacteroidota) but inhibited the growth of oligotrophic bacteria (Acidobacteria, Chloroflexi, and Verrucomicrobiota). Phosphorus addition alleviated phosphorus deficiency, resulting in a decrease in the abundance of phoD-harboring bacteria (Actinobacteria and Proteobacteria). Significant enhancement of soil bacterial alpha diversity was observed only in treatments with added phosphorus. Changes in NO3−-N, NH4+-N, available phosphorus, and dissolved organic carbon resulting from nutrient addition may have a greater impact on microbial community structure than changes in soil pH caused by nitrogen addition. Moreover, nutrient addition may indirectly impact microbial ecological function by altering nutrient availability in the soil. In conclusion, our study suggests that soil nutrient availability, particularly available phosphorus, affects soil bacterial communities and potentially regulates the biogeochemical cycles of soil ecosystems.

The strain designated as Y139T is a novel Gram-stain-negative, aerobic, and non-motile bacterium, was isolated from a soil sample in McClain County, Oklahoma, United States. The cells of strain Y139T were a rod-shaped, with the width of 0.4–0.7 μm and the length of 1.5–2.0 μm. Growth occurred at 20–37°C (optimum, 30°C), pH 5.5–9.5 (optimum, pH 7.0), and 0–1.0% NaCl (w/v) (optimum, 0%). The polar lipid profiles included phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, phosphatidyldimethylethanolamine, and an unidentified lipid. The major fatty acids included C16:0, iso-C14:0, and C16:1ω9c. Menaquinone-9 (MK-9) was recognized as the only respiratory quinone. Strain Y139T showed the highest 16S rRNA gene sequence similarity to Luteolibacter flavescens MCCC 1K03193T (98.3%). Phylogenetic analysis positioned it within the genus Luteolibacter. The draft genome of strain Y139T consisted of 7,106,054 bp, and contained 5,715 open reading frames (ORFs), including 5,656 coding sequences (CDSs) and 59 RNA genes. The genomic DNA G + C content was found to be 62.5%. Comparing strain Y139T with L. flavescens MCCC 1K03193T and Luteolibacter arcticus CCTCC AB 2014275T, the average nucleotide identity (ANI) values were 80.6 and 82.1%, respectively. Following phylogenetic, physiological, biochemical, and chemotaxonomic analyses, a novel species within the genus Luteolibacter, designated as Luteolibacter soli sp. nov., was proposed for strain Y139T, which was also assigned as the type strain (=KCTC 92644T = MCCC 1H01451T). Further analysis of core genes across 9 Luteolibacter species uncovered significant genomic divergence, particularly in those related to cofactor, vitamin, and energy metabolism. Analysis of biogeographic distribution suggested that lake and soil were the main habitats for the genus Luteolibacter. Additionally, the genus Luteolibacter was sensitive to climate warming and precipitation.

Lyme disease is the most prevalent tick-borne infection caused by Borrelia burgdorferi bacteria in North America. Other Borrelia species are predominately the cause of this disease in Eurasia with some distinct and various overlapping manifestations. Consequently, caution must be exercised when comparing the disease and its manifestations and treatment regimens in North America and Europe. Diagnosis of the early Lyme disease remains difficult using the currently FDA approved serological tests in the absence of a reported tick bite or of erythema migrans in many individuals, non-specific initial symptoms, and the absence of detectable anti-Borrelia antibodies in the prepatent period of infection. Furthermore, it is difficult to distinguish persistence of infection and disease versus reinfection in the endemic regions of Lyme disease by serological assays. If early infection remains untreated, spirochetes can disseminate and could affect various organs in the body with a variety of disease manifestations including arthralgias and musculoskeletal pain, neurologic symptoms and anomalies, and acrodermatitis chronicum atrophicans (ACA) in Europe. Although most patients recover after antibiotic treatment, an estimated ∼10–20% patients in the United States show persistence of symptoms known as post-treatment Lyme disease syndrome (PTLDS). The causes and biomarkers of PTLDS are not well-defined; however, several contributing factors with inconsistent degree of supporting evidence have been suggested. These include antigenic debris, dysregulation of immunological response, bacterial persisters, or combination of these features. This review highlights currently employed treatment approaches describing different antimicrobials used, and vaccine candidates tried to prevent B. burgdorferi infection.

In the Central Plains of China during the Zhou Dynasty (1046-256 BCE), the social hierarchy gradually solidified, accompanied by frequent wars and the phenomena of multicultural and multi-ethnic integration. These social phenomena collectively influenced the population’s genetic structure at that time. However, our understanding of the genetic history of this period remains largely unknown owing to limited ancient DNA studies. In this study, we successfully obtained 11 ancient genomes from the Guanzhuang site during the Zhou Dynasty on the central plain of China. Our findings revealed remarkable genetic continuity with the Neolithic populations of the Yellow River Basin and emphasized genetic diversity through the analysis of uniparental genetic markers. Population structure analysis further confirmed the genetic similarity between the Guanzhuang population and ancient populations of the Yellow River Basin and indicated genetic exchanges with ancient populations from surrounding regions. Intriguingly, signs of inbreeding within the Guanzhuang community cast doubt on the stringent enforcement of the contemporary marital regulations against consanguineous marriages within the same surname or clan. These revelations significantly enhance our insight into the complex interplay of ancient demography and societal organization, concurrently presenting a genetic perspective to view the complex evolution of Chinese civilization’s multiethnic.

Improvements in sequencing quality, availability, speed and costs results in an increased presence of genomics in infectious disease applications. Nevertheless, there are still hurdles in regard to the optimal use of WGS for public health purposes. Here, we discuss the current state (“status quo”) and future directions (“quo vadis”) based on literature regarding the use of genomics in surveillance, hazard characterization and source attribution of foodborne pathogens. The future directions include the application of new techniques, such as machine learning and network approaches that may overcome the current shortcomings. These include the use of fixed genomic distances in cluster delineation, disentangling similarity or lack thereof in source attribution, and difficulties ascertaining function in hazard characterization. Although, the aforementioned methods can relatively easily be applied technically, an overarching challenge is the inference and biological/epidemiological interpretation of these large amounts of high-resolution data. Understanding the context in terms of bacterial isolate and host diversity allows to assess the level of representativeness in regard to sources and isolates in the dataset, which in turn defines the level of certainty associated with defining clusters, sources and risks. This also marks the importance of metadata (clinical, epidemiological, and biological) when using genomics for public health purposes.

BackgroundThe emergence of ESBLs producing cephalosporin-resistant Escherichia coli isolates poses a threat to public health. This study aims to decipher the genetic landscape and gain insights into ESBL-producing E. coli strains belonging to the high-risk clone ST410 from pediatric patients.Methods29 E. coli ST410 isolates were collected from young children and subjected to antimicrobial susceptibility testing, Whole-genome sequencing (WGS), serotype analysis, MLST, ESBL genes, virulence genes, and plasmid profiling.ResultsAntimicrobial susceptibility testing demonstrated a high level of resistance to cephalosporins followed by aminoglycoside, sulfonamide, carbapenem and penicillin group of antibiotics. However, n=20/29 shows MDR phenotype. Phylogenetic group B2 (n=15) dominated, followed by group D (n=7), group A (n=4), and group B1 (n=3). Serotyping analysis identified O1:H7 (n=8), O2:H1 (n=6), O8:H4 (n=5), O16:H5 (n=4), and O25:H4 (n=3). Other serotypes identified included O6:H1, O15:H5, and O18:H7 (n=1 each). The most commonly detected ESBL genes were blaCTX-M, (n=26), followed by blaTEM (n=23), and blaSHV (n=18). Additionally, blaOXA-1 (n=10), blaOXA-48 (n=5), blaKPC-2 (n=3), blaKPC-3 (n=2), blaNDM-1 (n=4), blaNDM-5 (n=1), blaGES-1 (n=2), blaGES-5 (n=1), and blaCYM-1 (n=3). Notable virulence genes identified within the ST410 isolates included fimH (n=29), papC (n=24), hlyA (n=22), and cnf1 (n=18), among others. Diverse plasmids were observed including IncFIS, IncX4, IncFIA, IncCol, IncI2 and IncFIC with transmission frequency ranges from 1.3X10-2 to 2.7X10-3.ConclusionThe ST410 clone exhibited a complex resistance profile, diverse serotypes, the presence of specific resistance genes (ESBL genes), virulence gene repertoire, and diverse plasmids. The blaCTX-M was the most prevalent ESBL gene detected.

Carbapenemase-producing Klebsiella pneumoniae strains (CP-Kps) have recently been observed to spread rapidly worldwide. New Delhi metallo-β-lactamase (NDM) producing clones of Klebsiella pneumoniae (K. pneumoniae) cause a significant healthcare burden, particularly in Indian sub-continent, where this clone is circulating widely. However, in Italy, data on the incidence of these new clones is limited, and an ST437 NDM-producing K. pneumoniae strain has not been reported to date. A sacral ulcer infection caused by a K. pneumoniae strain was identified in an 85-year-old Italian male patient with several comorbidities. Antimicrobial susceptibility testing revealed an extensive resistance to a wide range of antimicrobials, including novel agents such as cefiderocol and ceftazidime/avibactam. Genomic analysis identified the pathogen as an ST437 K. pneumoniae strain harboring blaNDM-5, blaOXA-232 and blaCTX-M-15 genes. Following the identification of this first case, several infection control measures were implemented in healthcare settings, including direct precautions and reinforcement of standard cross-transmission control measures. The emergence of pathogenic microbial clones carrying new genetic determinants, particularly in a little city, requires prompt diagnosis and therapeutic protocols. An effective infection control system for the early detection and/or control of the transmission of NDM-producing Enterobacteriaceae is also needed. Further investigations are required to better understand the potential transmission routes and evolution of these clones.

ObjectiveIn recent years, the lung-gut axis has received increasing attention. The oxidative stress and systemic hypoxia occurring in chronic obstructive pulmonary disease (COPD) are related to gut dysfunction. That suggests probiotics have a potential therapeutic role in COPD. In this study, we therefore evaluated the ameliorative effects of probiotics on COPD.MethodsSearches were conducted in four electronic databases, including PubMed, Cochrane Library, the NIH clinical registry Clinical Trials. Gov and EMBASE. The data extracted was analyzed statistically in this study using StataMP17 software, with mean difference (MD) chosen as the effect size for continuous variables, and the results expressed as effect sizes and their 95% confidence intervals (CIs). Standardized Mean Difference (SMD) was used if the data units were different.ResultsWe included three randomized, controlled, double-blind clinical trials and five randomized controlled animal studies. The results show that for lung function, probiotics improved %FEV1 in COPD patients (MD = 3.02, 95%CI: 1.10, 4.93). Additionally, in inflammation, probiotics increased IL-10 (SMD = 1.99, 95%CI: 1.02, 2.96) and decreased inflammatory markers such as TNF-α (SMD= -2.64, 95%Cl: -3.38, -1.90), IL-1β (SMD= -3.49, 95%Cl: -4.58, -2.40), and IL-6 (SMD= -6.54, 95%Cl: -8.36, -4.73) in COPD animals, while having no significant effect on C-reactive protein (MD = 0.30, 95%CI: -0.71, 1.32) in COPD patients. For lung structure, probiotics significantly reduced the degree of pulmonary collagen fibers deposition in COPD animals (SMD = -2.25, 95%CI: -3.08, -1.41).ConclusionOverall, probiotics may be an additional approach that can improve COPD. Further clinical trials are needed to evaluate the efficacy, safety, and impact factors of probiotics for COPD.Systematic Review Registrationhttps://inplasy.com/inplasy-2023-4-0023/, identifier INPLASY202340023.

Legionella infection, the causative agent of Legionnaires’ disease, represents a significant threat to human health. The pathogenesis of this infection is intricately linked to the complex interactions between the bacterium and its host, resulting in profound metabolic perturbations. Central to these metabolic shifts is the bacterium’s modulation of lipid metabolism, with changes in lipid synthesis and breakdown modifying membrane composition and function. These alterations can influence cellular signaling and immune responses, further contributing to disease progression. It also disrupts glucose utilization and lipid metabolism, altering cellular energy production and immune responses. Additionally, Legionella infection perturbs amino acid and protein metabolism, affecting protein synthesis and degradation, leading to changes in cellular functions and immune responses. This mini-review underscores the complexity of metabolic perturbations in Legionella infection and their significance in host-pathogen interactions. Understanding these metabolic shifts provides valuable insights into the pathogenesis of Legionnaires’ disease and could lead to the development of novel therapeutic strategies.

Hibernation, an evolved survival trait among animals, enables them to endure frigid temperatures and food scarcity during the winter months, and it is a widespread phenomenon observed in mammals. The gut microbiota, a crucial component of animal nutrition and health, exhibits particularly dynamic interactions in hibernating mammals. This manuscript comprehensively evaluates the impacts of fasting, hypothermia, and hypometabolism on the gut microbiota of hibernating mammals. It suggests that alterations in the gut microbiota may contribute significantly to the maintenance of energy metabolism and intestinal immune function during hibernation, mediated by their metabolites. By delving into these intricacies, we can gain a deeper understanding of how hibernating mammals adapt to their environments and the consequences of dietary modifications on the symbiotic relationship between the gut microbiota and the host. Additionally, this knowledge can inform our comprehension of the protective mechanisms underlying long-term fasting in non-hibernating species, including humans, providing valuable insights into nutritional strategies and health maintenance.

BackgroundMalassezia furfur (M. furfur) is a prevalent dermatophyte that significantly impairs patients’ quality of life. This study aimed to evaluate the synergistic antifungal effects of combined extracts from Rosa rugosa Thunb. (MG) and Coptidis Rhizoma (HL) against M. furfur, both in vitro and in vivo.MethodsHigh-performance liquid chromatography (HPLC) was used to identify the major active compounds present in MG and HL. The antifungal activity of the combined Meilian extract (ML) was assessed using the checkerboard method and time-kill curves. Microstructural alterations in the fungi were observed using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The impact of the extracts on the fungal cell membrane was investigated through propidium iodide staining, protein concentration assays, and ergosterol quantification. Transcriptomic analysis was conducted to elucidate the molecular mechanisms underlying the effects of the extracts. Furthermore, the synergistic antifungal effects of ML were evaluated in a mouse model of seborrheic dermatitis induced by M. furfur.ResultsThe study demonstrated that the combined application of MG and HL significantly affected the integrity of the M. furfur cell membrane and potentially modulated its formation processes. In the M. furfur-induced seborrheic dermatitis model, ML exhibited synergistic antifungal effects and effectively alleviated skin inflammation. These findings provide an important theoretical basis for understanding the antifungal mechanisms of ML and its potential application in dermatological therapy.

IntroductionTraditional fermented foods have long been recognized for their numerous health benefits along with their potential to aid in the treatment of gastrointestinal disorders. These fermented foods have been shown to promote gut health and contribute to a longer, healthier life.MethodsThe high-throughput sequencing using the Illumina MiSeq platform was employed to investigate the microbiome communities of rice-based fermented beverages consumed by ethnic tribes in Southern Assam, namely Zeme Naga, Dimasa Kachari, Hmar, Karbi and Tea tribes.ResultsThe fermented rice-based beverages were highly predominated by Firmicutes, Bacteroides, Proteobacteria, and Actinobacteria exhibiting the highest relative abundance across all tribes. At genus level, significant abundance of pediococcus, lactobacillus, bacillus, leuconostoc, acetobacter, staphylococcus, delftia, erwinia, klebsiella and chrysebacterium were found amongst these ethnic tribes.DiscussionUnderstanding the fermented food microbiome will help to know the relationships between microbial communities and their effect on health of humans amongst the tribes. Furthermore, the use of these fermented products could provide enhanced health benefits to southern Assam region of India.

This study examined the phenol degradation capabilities and oxidative stress responses of Candida tropicalis SHC-03, demonstrating its metabolic superiority and resilience compared to Saccharomyces cerevisiae BY4742 in a culture medium with phenol as the sole carbon source. Through comparative growth, transcriptomic, and metabolomic analyses under different phenol concentrations, this study revealed C. tropicalis SHC-03’s specialized adaptations for thriving in phenol as the sole carbon source environments. These include a strategic shift from carbohydrate metabolism to enhanced phenol degradation pathways, highlighted by the significant upregulation of genes for Phenol 2-monoxygenase and Catechol 1,2-dioxygenase. Despite phenol levels reaching 1.8 g/L, C. tropicalis exhibits a robust oxidative stress response, efficiently managing ROS through antioxidative pathways and the upregulation of genes for peroxisomal proteins like PEX2, PEX13, and PMP34. Concurrently, there was significant upregulation of genes associated with membrane components and transmembrane transporters, enhancing the cell’s capacity for substance exchange and signal transduction. Especially, when the phenol concentration was 1.6 g/L and 1.8 g/L, the degradation rates of C. tropicalis towards it were 99.47 and 95.91%, respectively. Conversely, S. cerevisiae BY4742 shows limited metabolic response, with pronounced growth inhibition and lack of phenol degradation. Therefore, our study not only sheds light on the molecular mechanisms underpinning phenol tolerance and degradation in C. tropicalis but also positions this yeast as a promising candidate for environmental and industrial processes aimed at mitigating phenol pollution.

Establishing the contextIntestinal dysbiosis is a significant concern among dog owners, and the gut health of pets is an emerging research field. In this context, the Simulator of the Canine Intestinal Microbial Ecosystem (SCIME™) was recently developed and validated with in vivo data.Stating the purpose/introducing the studyThe current study presents a further application of this model by using amoxicillin and clavulanic acid to induce dysbiosis, aiming to provoke changes in microbial community and metabolite production, which are well-known markers of the disease in vivo.Describing methodologyFollowing the induction of dysbiosis, prebiotic supplementation was tested to investigate the potential for microbiota recovery under different dietary conditions.Presenting the resultsThe results showed that antibiotic stimulation in the SCIME™ model can produce significant changes in microbial communities and metabolic activity, including a decrease in microbial richness, a reduction in propionic acid production, and alterations in microbial composition. Additionally, changes in ammonium and butyric acid levels induced by the tested diets were observed.Discussing the findingsThis alteration in microbial community and metabolites production mimicks in vivo canine dysbiosis patterns. A novel dynamic in vitro model simulating canine antibiotic-induced dysbiosis, capable of reproducing microbial and metabolic changes observed in vivo, has been developed and is suitable for testing the effects of nutritional changes.

IntroductionSome cyanobacteria can use far-red light (FRL) to drive oxygenic photosynthesis, a phenomenon known as Far-Red Light Photoacclimation (FaRLiP). It can expand photosynthetically active radiation beyond the visible light (VL) range. Therefore, it holds promise for biotechnological applications and may prove useful for the future human exploration of outer space. Typically, FaRLiP relies on a cluster of ~20 genes, encoding paralogs of the standard photosynthetic machinery. One of them, a highly divergent D1 gene known as chlF (or psbA4), is the synthase responsible for the formation of the FRL-absorbing chlorophyll f (Chl f) that is essential for FaRLiP. The minimum gene set required for this phenotype is unclear. The desert cyanobacterium Chroococcidiopsis sp. CCMEE 010 is unusual in being capable of FaRLiP with a reduced gene cluster (15 genes), and it lacks most of the genes encoding FR-Photosystem I.MethodsHere we investigated whether the reduced gene cluster of Chroococcidiopsis sp. CCMEE 010 is transcriptionally regulated by FRL and characterized the spectral changes that occur during the FaRLiP response of Chroococcidiopsis sp. CCMEE 010. In addition, the heterologous expression of the Chl f synthase from CCMEE 010 was attempted in three closely related desert strains of Chroococcidiopsis.ResultsAll 15 genes of the FaRLiP cluster were preferentially expressed under FRL, accompanied by a progressive red-shift of the photosynthetic absorption spectrum. The Chl f synthase from CCMEE 010 was successfully expressed in two desert strains of Chroococcidiopsis and transformants could be selected in both VL and FRL.DiscussionIn Chroococcidiopsis sp. CCME 010, all the far-red genes of the unusually reduced FaRLiP cluster, are transcriptionally regulated by FRL and two closely related desert strains heterologously expressing the chlF010 gene could grow in FRL. Since the transformation hosts had been reported to survive outer space conditions, such an achievement lays the foundation toward novel cyanobacteria-based technologies to support human space exploration.

BackgroundInvasive pulmonary aspergillosis (IPA) typically occurs in immunocompromised individuals. Severe fever with thrombocytopenia syndrome (SFTS) patients are typically characterized by fever, thrombocytopenia, and leukopenia. These patients typically present with dysregulation of cellular and humoral immunity, which may predispose them to IPA. Our study aimed to identify risk factors for SFTS-associated invasive pulmonary aspergillosis (SAPA) and evaluate its associated prognostic impact.MethodsWe conducted a cohort study between January 2017 and December 2022 in a tertiary hospital in Wuhan City, China. All SFTS patients hospitalized in our department who formally consented were divided into a SAPA group and a non-SAPA group according to whether they were coinfected with aspergillosis or not. The independent risk factors for the SAPA group were determined by multivariate logistic regression. Receiver operating characteristic (ROC) analysis was used to assess the statistical value of parameters to predict SAPA patients. The survival analysis was carried out using the Kaplan–Meier (KM) method.ResultsOf the 269 hospitalized SFTS patients enrolled in the study, 118 (43.87%) cases were diagnosed with SAPA with an average age of 65.71 ± 9.7 years. Multivariate logistic regression analysis revealed that age, neurological complications, serum severe fever with thrombocytopenia syndrome virus (SFTSV) RNA loads, the white blood cell (WBC) count, platelet (PLT) count, albumin (ALB) and globulin (GLB) concentrations, and cardiac troponin I (cTNI) were complementary risk factors for the development of IPA in SFTS patients. The risk score is calculated as 5 times age, plus 6 times neurological complications, plus 10 times RNA (log), plus 5 times WBC, minus 5 times PLT, minus 5 times ALB, plus 5 times GLB, and plus 6 times cTNI. ROC curve analysis showed that the area under the receiver operating characteristic (AUROC) curve represented a risk score of 0.837 (95% CI: 0.789–0.885, p

IntroductionBovine Genital Campylobacteriosis (BGC), caused by Campylobacter fetus subsp. venerealis, is a sexually transmitted bacterium that significantly impacts cattle reproductive performance. However, current detection methods lack consistency and reliability due to the close genetic similarity between C. fetus subsp. venerealis and C. fetus subsp. fetus. Therefore, this study aimed to utilize complete genome analysis to distinguish genetic features between C. fetus subsp. venerealis and other subspecies, thereby enhancing BGC detection for routine screening and epidemiological studies.Methods and resultsThis study reported the complete genomes of four C. fetus subsp. fetus and five C. fetus subsp. venerealis, sequenced using long-read sequencing technologies. Comparative whole-genome analyses (n = 25) were conducted, incorporating an additional 16 complete C. fetus genomes from the NCBI database, to investigate the genomic differences between these two closely related C. fetus subspecies. Pan-genomic analyses revealed a core genome consisting of 1,561 genes and an accessory pangenome of 1,064 genes between the two C. fetus subspecies. However, no unique predicted genes were identified in either subspecies. Nonetheless, whole-genome single nucleotide polymorphisms (SNPs) analysis identified 289 SNPs unique to one or the C. fetus subspecies. After the removal of SNPs located on putative genomic islands, recombination sites, and those causing synonymous amino acid changes, the remaining 184 SNPs were functionally annotated. Candidate SNPs that were annotated with the KEGG “Peptidoglycan Biosynthesis” pathway were recruited for further analysis due to their potential association with the glycine intolerance characteristic of C. fetus subsp. venerealis and its biovar variant. Verification with 58 annotated C. fetus genomes, both complete and incomplete, from RefSeq, successfully classified these seven SNPs into two groups, aligning with their phenotypic identification as CFF (Campylobacter fetus subsp. fetus) or CFV/CFVi (Campylobacter fetus subsp. venerealis and its biovar variant). Furthermore, we demonstrated the application of mraY SNPs for detecting C. fetus subspecies using a quantitative PCR assay.DiscussionOur results highlighted the high genetic stability of C. fetus subspecies. Nevertheless, Campylobacter fetus subsp. venerealis and its biovar variants encoded common SNPs in genes related to glycine intolerance, which differentiates them from C. fetus subsp. fetus. This discovery highlights the potential of employing a multiple-SNP assay for the precise differentiation of C. fetus subspecies.

Horizontal gene transfer (HGT) favors the acquisition and spread of antimicrobial resistance (AMR) genes in Salmonella, making it a major public health concern. We performed a bibliometric analysis to provide the current landscape of HGT in research on Salmonella AMR and identify emerging trends and potential research directions for the future. Data were collected from the Web of Science Core Collection and limited to articles and reviews published between 1999 and 2024 in English. VOSviewer 1.6.19 and CiteSpace 6.2.R1 software were used to conduct bibliometric analysis and visualize co-occurring keywords. A total of 1,467 publications were retrieved for analysis. American researchers contributed the most articles (n = 310). In the meantime, Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement have the highest citation/publication rate of 85.6. Recent studies have focused on the application of whole genome sequencing (WGS), Salmonella quinolone and colistin resistance, and the biocontrol of Salmonella AMR. These findings provide new insights into the role of HGT and help identify new targets for controlling the spread of AMR in Salmonella populations.

Apple is an economically important fruit crop in northern Shaanxi Province in China. In recent years, a new type of apple black spot disease, “bagged apple black spot disease,” has occurred in the main apple production area of Yan’an City, Shaanxi Province, during the apple ripening season. It seriously affects the appearance quality and commercial value of apples. In this study, 120 isolates recovered from symptomatic apples were identified based on morphological characteristics, pathogenicity, and multilocus sequence analyses of the internal transcribed spacer of ribosomal DNA (rDNA-ITS), translation elongation factor 1-α (EF1-α), RNA polymerase II subunit B (RPB2), endopolygalacturonase (endo-PG), and anonymous region OPA1-3. Alternaria alternata was the most abundant species (64%), followed by Fusarium acuminatum (36%). Pathogenicity assays were conducted by inoculating them individually and together on detached apples (Venus Golden and Fuji varieties). The results showed that the two fungal species could infect apples individually and together. Co-infection enhanced the disease severity. F. acuminatum led to increased severity and speed of disease development compared to A. alternata. This is the first report of Fusarium and Alternaria co-infection causing apple black spot disease worldwide, and the first report of F. acuminatum affecting apples. The optimal growth of A. alternata occurred at 25–30°C and pH 7; the optimal growth of F. acuminatum occurred at 25°C and pH 7. The results of this study can provide a theoretical basis for exploring the occurrence and epidemiology of apple black spot disease and strategies for its control.

Over the last few decades, advancements in process safety and quality methods have been significantly improved, yet new challenges continue to emerge in the sustainable food supply chain. This study aimed to investigate some physicochemical and microbiological parameters impacting meat products, particularly cooked sausages, within a sustainable supply chain, focusing on quality, spoilage populations, and syneresis formation under vacuum conditions. A comprehensive analysis was conducted on 355 samples collected over four seasons using high-throughput sequencing (16S/ITS) and microbiological and physicochemical [pH and water activity (aw)] assessments. The microbial growth predictor MicroLab_ShelfLife was employed, and multiple factor analysis (MFA) and agglomerative hierarchical clustering (AHC) were utilized to understand how these variables influence the microbiome resilience of these products. Lactic and acetic acids were correlated with the microbiome of the sausages and the liquid coating covering them using metagenomic analyses. The study highlighted that 52% of the evaluated meat industries in southeastern Brazil are implementing effective protocols for sustainable chain production. The results indicated that the durability of vacuum-packaged cooked sausages was primarily influenced by storage temperature (RV coefficient of 0.906), initial microbial load (0.755), and aw (0.624). Average microbial counts were 4.30 log cfu/g (initial), 4.61 (7°C/4 days), 4.90 (7°C/8 days), 6.06 (36°C/4 days), and 6.79 (36°C/8 days). Seasonal durability analysis revealed that winter had the highest average durability of 45.58 days, while summer had the lowest at 26.33 days. Yeast populations, including Trichosporon sp. and Candida sp., were identified as key genera influencing spoilage dynamics. In addition, Bacillus species emerged as dominant spoilage microorganisms, highlighting the need for new critical controls. This study demonstrates the impact of metagenomic approaches, including ITS and 16S amplicon sequencing, in revealing microbial community dynamics, storage temperature, and aw, which are essential for developing targeted interventions to enhance food safety and quality sustainably.

The stress response of pig herds poses a significant challenge in the pig breeding industry, and investigating strategies to mitigate this stress is of paramount importance. The objective of this study was to investigate the impacts of supplemental feeding of Chinese herbal mixtures to perinatal sows on antioxidant capacity and gut microbiota of sows and their offspring piglets. A total of 60 healthy sows (Large white) at fourth parity were randomly assigned to five treatment groups. The control group received a basal diet, while the TRT1 group received a basal diet supplemented with 2kg/t Bazhen powder (BZP). The TRT2, TRT3, and TRT4 groups were fed a basal diet supplemented with 1kg/t, 2kg/t, and 3kg/t Qi-Zhu- Gui-Shao soothing liver and replenishing blood powder (QZGSP), respectively. The trial lasted for 5weeks, starting from day 100 of gestation until day 21 of delivery. The results demonstrated that the inclusion of 2kg/t and 3kg/t QZGSP significantly enhanced the antioxidant capacity of sows and their offspring piglets to different degrees, thereby effectively alleviating oxidative stress. Analysis of gut microbiota revealed that QZGSP influenced the composition of gut microbiota in both sows and their offspring piglets. Specifically, at the genus level, the abundance of Christensenellaceae_R-7_group in the gut microbiota of sows in the TRT4 group was significantly lower than that in the TRT1 group (p < 0.05), while the relative abundance of Lactobacillus in the gut microbiota of sows in the TRT4 group was significantly higher than that in the CON group (p < 0.05). Furthermore, at the genus level, compared to those in the TRT1 group, piglets from the TRT4 group exhibited a significant decrease in relative abundance of Escherichia-Shigella, Parabacteroides, and Methanobrevivacter (p < 0.05), but a significant increase in Phascolarctobacterium (p < 0.05). Spearman correlation analysis indicated a positive correlation between relative abundance of Christensenellaceae_R-7_group and serum contents of T-AOC and CAT (p < 0.05), as well as a negative correlation with serum concentration MDA (p < 0.05). Additionally, there was a positive correlation between relative abundance Lactobacillus and serum levels SOD (p < 0.01) and GSH-Px (p < 0.05). Therefore, supplementation of 3kg/t QZGSP in the periparturient sow diet significantly augmented antioxidant capacity in both sows and offspring piglets, while concurrently modulating the composition and structure of their intestinal microflora. The findings from this study demonstrate that QZGSP represents a beneficial feed additive for perinatal sows.

P32 protein serves as a crucial structural component of Goat pox virus (GTPV), which causes a highly virulent infectious disease in sheep and goats. Despite the fact that P32 has been widely expressed in the previous studies, it is difficult to obtain recombinant P32 efficiently. This study aimed to achieve soluble expression of P32 recombinant protein and to develop its specific monoclonal antibody. The gene fragment of P32Δ (GP32Δ) was synthesized by optimizing the coding sequence of amino acids 1-246 of the known goatpox P32 protein. Subsequently, GP32Δ was cloned into a prokaryotic expression vector for expression and purification, resulting in the successful production of soluble recombinant protein rP32Δ. Utilizing rP32Δ, an indirect ELISA method was established by immunizing 6-week-old BALB/c mice with inactivated GTPV as the antigen. Through hybridoma technology, three monoclonal antibody hybridoma cell lines secreting anti-goat pox virus rP32Δ were screened, designated as 2F3, 3E8, and 4H5, respectively. These monoclonal antibodies, classified as IgG1, IgG2a, and IgG2b, respectively, with κappa light chains, were characterized following ascites preparation and purification. Indirect ELISA results demonstrated that the ELISA potency of the three monoclonal antibodies exceeded 1:12800. Furthermore, Western blot analysis revealed specific reactivity of both 3E8 and 4H5 with rP32Δ, while immunofluorescence assays confirmed 3E8's ability to specifically recognize GTPV in cells. The preceding findings demonstrate the successful acquisition of the soluble expressed recombinant P32 protein and its specific monoclonal antibody 3E8 in this study, thereby laying a foundational material basis for the establishment of a GTPV detection method.

BackgroundCurrently, Sodium-glucose cotransporter 2 (SGLT2) inhibitors demonstrate additional effects beyond glucose control on the gut microbiota and circulating metabolites. The gut microbiota and metabolites have been found to be useful in elucidating potential biological mechanisms of pulmonary diseases. Therefore, our study aims to investigate the effects of gut microbiota and metabolites mediating SGLT2 inhibition in 10 pulmonary diseases through Mendelian randomization (MR) research.MethodsWe conducted a two-sample, two-step MR study to assess the association between SGLT2 inhibition and 10 pulmonary diseases and to investigate the mediating effects of gut microbiota and metabolite. Gene-fine mapping and annotation of mediators by FUMA and Magma analyses were performed, and causal associations of mapped genes with diseases were assessed by muti-omics MR analyses. Possible side effects of SGLT2 inhibition were assessed by PheWAS analysis.ResultsSGLT2 inhibition was linked to a reduced risk of T2DM, Interstitial lung disease (ILD), Pneumoconiosis, Pulmonary tuberculosis, and Asthma(OR=0.457, 0.054, 0.002, 0.280, 0.706). The family Enterobacteriaceae and order Enterobacteriales were associated with SGLT2 inhibition and ILD(95% CI:0.079–0.138). The family Alcaligenaceae and X-12719 were linked to pneumoconiosis (95% CI: 0.042–0.120, 0.050–0.099). The genus Phascolarctobacterium was connected to pulmonary tuberculosis (95% CI: 0.236–0.703).The degree of unsaturation (Fatty Acids), ratio of docosahexaenoic acid to total fatty acids, and 4-androsten-3beta,17beta-diol disulfate 2, were associated with asthma(95% CI: 0.042–0.119, 0.039–0.101, 0.181–0.473). Furthermore, Fuma and Magma analyses identified target genes for the four diseases, and proteomic MR analysis revealed six overlapping target genes in asthma. PheWAS analysis also highlighted potential side effects of SGLT2 inhibition.ConclusionsThis comprehensive study strongly supports a multi-omics association between SGLT2 inhibition and reduced risk of interstitial lung disease, tuberculosis, pneumoconiosis, and asthma. Four identified gut microbiota, four metabolites, sixteen metabolic pathways, and six target genes appear to play a potential role in this association. The results of the comprehensive phenome-wide association analysis also identified the full effect of SGLT2 inhibitors.

The indiscriminate use of antibiotics has resulted in a growing resistance to drugs in Pseudomonas aeruginosa. The identification of antibiotic resistance genes holds considerable clinical significance for prompt diagnosis. In this study, we established and optimized a Recombinase-Aided Amplification (RAA) assay to detect two genes associated with drug resistance, oprD and arr, in 101 clinically collected P. aeruginosa isolates. Through screening for the detection or absence of oprD and arr, the results showed that there were 52 Imipenem-resistant P. aeruginosa (IRPA) strains and 23 Rifampin-resistant P. aeruginosa (RRPA) strains. This method demonstrated excellent detection performance even when the sample concentration is 10 copies/μL at isothermal conditions and the results could be obtained within 20 minutes. The detection results were in accordance with the results of conventional PCR and Real-time PCR. The detection outcomes of the arr gene were consistently with the resistance spectrum. However, the antimicrobial susceptibility results revealed that 65 strains were resistant to imipenem, while 49 strains sensitive to imipenem with oprD were identified. This discrepancy could be attributed to genetic mutations. In summary, the RAA has higher sensitivity, shorter time, and lower-cost instrument requirements than traditional detection methods. In addition, to analyze the epidemiological characteristics of the aforementioned drug-resistant strains, we conducted Multilocus Sequence Typing (MLST), virulence gene, and antimicrobial susceptibility testing. MLST analysis showed a strong correlation between the sequence types ST-1639, ST-639, ST-184 and IRPA, while ST-261 was the main subtype of RRPA. It was observed that these drug-resistant strains all possess five or more virulence genes, among which exoS and exoU do not coexist, and they are all multidrug-resistant strains. The non-coexistence of exoU and exoS in P.aeruginosa is related to various factors including bacterial regulatory mechanisms and pathogenic mechanisms. This indicates that the relationship between the presence of virulence genes and the severity of patient infection is worthy of attention. In conclusion, we have developed a rapid and efficient RAA (Recombinase-Aided Amplification) detection method that offers significant advantages in terms of speed, simplicity, and cost-effectiveness (especially in time and equipment aspect). This novel approach is designed to meet the demands of clinical diagnostics.

The McMurdo Dry Valleys (MDVs) of Antarctica are a mosaic of extreme habitats which are dominated by microbial life. The MDVs include glacial melt holes, streams, lakes, and soils, which are interconnected through the transfer of energy and flux of inorganic and organic material via wind and hydrology. For the first time, we provide new data on the viral community structure and function in the MDVs through metagenomics of the planktonic and benthic mat communities of Lakes Bonney and Fryxell. Viral taxonomic diversity was compared across lakes and ecological function was investigated by characterizing auxiliary metabolic genes (AMGs) and predicting viral hosts. Our data suggest that viral communities differed between the lakes and among sites: these differences were connected to microbial host communities. AMGs were associated with the potential augmentation of multiple biogeochemical processes in host, most notably with phosphorus acquisition, organic nitrogen acquisition, sulfur oxidation, and photosynthesis. Viral genome abundances containing AMGs differed between the lakes and microbial mats, indicating site specialization. Using procrustes analysis, we also identified significant coupling between viral and bacterial communities (p = 0.001). Finally, host predictions indicate viral host preference among the assembled viromes. Collectively, our data show that: (i) viruses are uniquely distributed through the McMurdo Dry Valley lakes, (ii) their AMGs can contribute to overcoming host nutrient limitation and, (iii) viral and bacterial MDV communities are tightly coupled.

IntroductionWeak aerobic stability is a notable challenge for whole-plant corn silage, particularly in hot and humid regions. Acetobacter is commonly regarded as an indicator of aerobic deterioration in silage, yet its precise role in fermentation and during aerobic exposure, as well as the factors that promote its growth, remain insufficiently understood.MethodsIn this study, whole-plant corn silage was prepared using a bagged method with controlled dry matter (DM) content at 20%, 25%, and 30%, and initial concentrations of A. pasteurianus at 40%, 50%, and 60%. The silage was stored for 60 days under varying temperatures (20°C, 30°C, and 40°C). Following the anaerobic storage phase, the silage was exposed to air at room temperature (20-25°C) for 7 days, both with and without A. pasteurianus inoculation.ResultsThe results demonstrated that A. pasteurianus did not impact the nutritional value of the silage during anaerobic fermentation, maintaining a low pH (< 3.80). However, during aerobic exposure, the presence of A. pasteurianus significantly reduced the aerobic stability of the silage. The microbial community shifted from primarily Klebsiella species initially to Lactobacillus and Acetobacter species post-ensiling. During the aerobic exposure phase, A. pasteurianus and A. fabarum became the dominant species. Response Surface Methodology (RSM) analysis identified optimal conditions for the proliferation of A. pasteurianus during the aerobic phase, which occurred at 28°C, 25% DM, and 52% initial concentration at 3 ml/kg.DiscussionThese findings confirm that A. pasteurianus plays a critical role in reducing the aerobic stability of whole-plant corn silage. Additionally, the study identifies the optimal conditions that favor the proliferation of A. pasteurianus, offering valuable insights for the development of strategies to prevent and control this bacterium, thereby improving the aerobic stability of silage in hot and humid regions.

BackgroundResearch on the effects of intestinal microbiota transplantation (IMT) on chronic HBV infection (CHB) progression associated liver disease (HBV-CLD) and alterations in the microbiota post-IMT are quite limited for the moment.MethodsBy integrating microbiome with metabolome analyses, we aimed to the function of IMT and the alterations of gut microbiota in patients with HBV-CLD. First, this study included 20 patients with HBV-CLD and ten healthy controls. Then, 16 patients with CHB were given IMT with donor feces (heterologous) via oral capsule. Fecal samples from CHB patients were obtained before and after IMT, as well as healthy controls, for 16S rDNA sequencing and untargeted metabolomics analysis.ResultsThe proalbuminemia were significantly increased after IMT, and the HBsAg and TBA showed a significant decrease after IMT in the HBV-CLD patients. There was statistical difference in the Chaol indexes between between CHB patients and healthy controls, suggesting a lower abundance of the gut microbiota in HBV-CLD patients. In addition, there was statistical difference in the Shannon and Simpson indexes between prior to IMT and post-IMT, indicating that the impaired abundance of the gut microbiota had been improved after IMT. The host-microbiota-metabolite interplay, amino acid metabolism, nicotinate and nicotinamide metabolism, starch and sucrose metabolism, steroid biosynthesis, and vitamins metabolism, were significantly lower in HBV-CLD patients than healthy controls.ConclusionIMT may improve the therapeutic effects on patients HBV-CLD. Furthermore, IMT appears to improve amino acid metabolism by impaired abundance of the gut microbiota and therefore improve liver prealbumin synthesis.

PurposeWe aimed to retrospectively investigate an outbreak of linezolid-resistant Staphylococcus epidermidis (LRSE), at Tours University Hospital between 2017 and 2021.MethodsTwenty of the 34 LRSE isolates were included in the study. Antimicrobial susceptibility testing was performed using the disk diffusion method and MICs of last-resort antibiotics were determined using broth microdilution or Etest®. Seventeen of the 20 resistant strains were sent to the French National Reference Centre for Staphylococci to determine the mechanism of resistance to linezolid. The clonal relationship between LRSE strains was assessed by PFGE and the sequence type determined by MLST. We retrospectively evaluated a new typing tool, IR-Biotyper®, and compared its results to PFGE to evaluate its relevance for S. epidermidis typing. Medical records were reviewed, and antibiotic consumption was determined. Search for a cross transmission was performed.ResultsAll LRSE strains showed high levels of resistance to linezolid (MICs ≥ 256 mg/L) and were multi-drug resistant. Linezolid resistance was associated with the 23S rRNA G2576T mutation and none of the 17 strains analyzed carried the cfr gene. Ninety-five percent of the 20 LRSE studied strains were genetically related and belonged to sequence-type ST2. The dendrogram obtained from IR-Biotyper® showed 87% congruence with the PFGE analysis. Prior to isolation of the LRSE strain, 70% of patients received linezolid. No patients stayed successively in the same room.ConclusionLinezolid exposure may promote the survival and spread of LRSE strains. At Tours University Hospital, acquisition of the resistant clone may also have been triggered by hand-to-hand transmission by healthcare workers. In addition, IR-Biotyper® is a promising typing tool for the study of clonal outbreaks due to its low cost and short turnaround time, although further studies are needed to assess the optimal analytical parameters for routine use.

IntroductionEnterococci are commensals of the gastrointestinal tract of humans and animals that evolved into opportunistic pathogens with high antimicrobial resistance and virulence. Multidrug-resistant Enterococcus is a major cause of hospital-acquired infections worldwide. For this reason, the characterization of non-clinical reservoirs of Enterococci and their epidemiological link to resistant hospital isolates is crucial for controlling their spread.MethodsA total of 295 samples collected from livestock (pigs and cows, n = 135) and environment (public buses, passengers hands, and urban environments, n = 160) were screened for Enterococcus spp. E. faecium antimicrobial resistance profiles, virulence potential, and clonal population were further characterized.ResultsEnterococci were detected in 90.5% (n = 267) of the samples, with a higher prevalence in livestock (100%) than the environment (82.5%, p

ObjectiveThe purpose of this study was to investigate the epidemiological and genomic characteristics of Vibrio parahaemolyticus (V. parahaemolyticus) isolates from outbreaks in Huzhou, China.MethodsThis study aims to analyze the epidemiological data on V. parahaemolyticus outbreaks reported in Huzhou from 2019 to 2023. A total of 70 V. parahaemolyticus outbreak isolates were collected. The antibiotic resistance, serotypes, molecular typing, and genomic characteristics of these isolates were analyzed.ResultsMost outbreaks of V. parahaemolyticus infection occurred in the summer, and the majority of outbreaks occurred in restaurants and rural banquets. High resistance rates were observed for ampicillin (AMP, 24.29%), followed by tetracycline (TET, 15.71%) and trimethoprim-sulfamethoxazole (SXT, 15.71%). The newly emerged serotype O10:K4 became dominant from 2021 to 2023, with most isolates belonging to ST3. The resistance gene blaCARB was frequently detected among these isolates. The pulsed-field gel electrophoresis (PFGE) and whole-genome single-nucleotide polymorphisms (wgSNPs) effectively differentiated the nine outbreaks.ConclusionThe newly emerged serotype O10:K4 became dominant from 2021 to 2023, with most isolates being ST3. PFGE and WGS technologies provided reliable methods for typing and identifying V. parahaemolyticus for outbreaks.

The global prevalence of obesity is rising year by year, which has become a public health problem worldwide. Many animal and clinical studies have shown that Lactiplantibacillus plantarum is considered an ideal probiotic and potential supplement for the treatment of obesity. In this study, we aimed to complete the genome sequence of L. plantarum HOM2217, which was isolated from human milk, and study its physiological characteristics and anti-obesity effects in 3T3-L1 cells and rats fed a high-fat diet (HFD) to determine its potential as a starter for functional food products. Whole-genome analysis demonstrated that HOM2217 contained a single circular chromosome of 3,267,529 bp with a GC content of 44.5% and one plasmid (62,350 bp) with a GC content of 38.5%. Compared to the reference strains, HOM2217 demonstrated superior tolerance to gastrointestinal conditions, higher adhesion to intestinal epithelial cell lines, potent antimicrobial activity against Enterobacter cloacae ATCC 13047, and effective cholesterol removal ability in vitro. Treatment with heat-killed HOM2217 significantly reduced lipid accumulation and intracellular triglyceride production in 3T3-L1 adipocytes. Daily treatment of HFD-fed rats with HOM2217 for 7 weeks decreased body weight, body weight gain, and body fat without changes in food intake. HOM2217 also significantly increased the serum high-density lipoprotein cholesterol (HDL-C) level, decreased the serum tumor necrosis factor (TNF-α) and increased short-chain fatty acid (SCFA) (formic acid, acetic acid, and butyric acid) levels in the cecum. Thus, HOM2217 could potentially prevent obesity in rats by inhibiting inflammatory responses and regulating lipid metabolism and SCFAs expression. Therefore, HOM2217 has potential as an alternative treatment for obesity.

IntroductionApplying biochar (BC) to reduce toxic substance accumulation in soil, either through direct adsorption or modulation of the microbial community, has received considerable attention. However, a knowledge gap exists regarding how BC regulates microbial community structure and functions to mitigate toxic substance accumulation.MethodsWe previously identified p-coumaric acid (p-CA) as a representative autotoxin in tobacco rhizosphere soil. On this basis, this study simulated a soil environment with p-CA accumulation to investigate the impacts of BC on p-CA, soil physicochemical properties, and microbial community structure and function.ResultsThe results showed that p-CA could be directly adsorbed onto BC, which followed the pseudo-second-order kinetic model (R2 = 0.996). A pot experiment revealed that BC significantly reduced soil p-CA, altered soil microbial composition, and enhanced bacterial community diversity. A weighted correlation network analysis showed a close association between taxon 1 in the microbial network and p-CA, suggesting a pivotal role for this taxon in reducing p-CA, with Devosia and Nocardioides identified as potential key contributors to this process. The prediction of possible keystone taxa functions showed that BC increased the relative abundances of aromatic compound degraders. Mantel tests indicated that soil organic matter exerted the greatest influence on keystone taxa functions and hub genera.DiscussionThese findings suggest that BC may either directly chemisorb p-CA or indirectly facilitate p-CA degradation by regulating the functioning of keystone taxa. The results of this study provide a novel perspective for further investigation of the mechanisms through which BC reduces the accumulation of toxic substances in soil.

High lead (Pb) levels in agricultural soil and wastewater threaten ecosystems and organism health. Microbial remediation is a cost-effective, efficient, and eco-friendly alternative to traditional physical or chemical methods for Pb remediation. Previous research indicates that micro-organisms employ various strategies to combat Pb pollution, including biosorption, bioprecipitation, biomineralization, and bioaccumulation. This study delves into recent advancements in Pb-remediation techniques utilizing bacteria, fungi, and microalgae, elucidating their detoxification pathways and the factors that influence Pb removal through specific case studies. It investigates how bacteria immobilize Pb by generating nanoparticles that convert dissolved lead (Pb-II) into less harmful forms to mitigate its adverse impacts. Furthermore, the current review explores the molecular-level mechanisms and genetic engineering techniques through which microbes develop resistance to Pb. We outline the challenges and potential avenues for research in microbial remediation of Pb-polluted habitats, exploring the interplay between Pb and micro-organisms and their potential in Pb removal.

BackgroundThe gut microbiota has been demonstrated to have a significant role in the pathogenesis and progression of a variety of diseases, including prostate cancer, prostatitis, and benign prostatic hyperplasia. Potential links between prostate diseases, immune cells and the gut microbiota have not been adequately investigated.MethodsMR studies were conducted to estimate the effects of instrumental variables obtained from genome-wide association studies (GWASs) of 196 gut microbial taxa and 731 immune cells on the risk of prostate diseases. The primary method for analysing causal relationships was inverse variance-weighted (IVW) analysis, and the MR results were validated through various sensitivity analyses.ResultsMR analysis revealed that 28 gut microbiome taxa and 75 immune cell types were significantly associated with prostate diseases. Furthermore, reverse MR analysis did not support a causal relationship between prostate diseases and the intestinal microbiota or immune cells. Finally, the results of the mediation analysis indicated that Secreting Treg % CD4 Treg, Activated & resting Treg % CD4 Treg, and Mo MDSC AC inhibited the role of the class Mollicutes in reducing the risk of PCa. In prostatitis, CD8+ T cells on EM CD8br hinder the increased risk associated with the genus Eubacterium nodatum group. Interestingly, in BPH, CD28- CD25++CD8br AC and CD16-CD56 on HLA DR+ NK promoted the role of the genus Dorea in reducing the risk of BPH.ConclusionThis study highlights the complex relationships among the gut microbiota, immune cells and prostate diseases. The involvement of the gut microbiota in regulating immune cells to impact prostate diseases could provide novel methods and concepts for its therapy and management.

Porcine reproductive and respiratory syndrome virus (PRRSV) has caused severe economic losses to the global swine industry. In recent years, the incidence of PRRSV-1 has been gradually increasing in China, but there are still few studies on it. In this study, clinical samples for PRRS virus isolation were collected from a pig farm in South China in 2022. We effectively isolated a strain of PRRSV utilizing PAM cells and demonstrated its consistent transmission capability on Marc-145 cells. The isolated strain was confirmed as PRRSV-1 by RT-qPCR, IFA, electron microscopy, etiolated spot purification and whole genome sequencing, the strain was named GD2022. The length of GD2022 genome is 15058nt; Based on the genome-wide genetic evolutionary analysis of GD2022, the strain was classified as PRRSV-1. Further genetic evolutionary analysis of its ORF5 gene showed that GD2022 belonged to PRRSV-1 subtype 1 and formed an independent branch in the evolutionary tree. Compared with the sequence of the classical PRRSV-1 strain (LV strain), GD2022 has several amino acid site mutations in the antigenic region from GP3 to GP5, these mutations are different from those of other PRRSV-1 strains in China. Recombination analysis showed no recombination events with GD2022. In addition, piglets infected with GD2022 displayed clinical respiratory symptoms and typical pathological changes. In this study, a strain of the PRRSV-1 virus was isolated using both PAM cells and Marc-145 and proved to be pathogenic to piglets, providing an important reference for the identification, prevention, and control of PRRSV-1.

Ferroptosis is a novel form of programmed cell death, which is different from apoptosis, pyroptosis and autophagy in morphology and biochemistry. Ferroptosis is characterized by condensed mitochondrial membrane densities, vanished of mitochondria crista and outer membrane rupture in morphology, and the accumulation of intracellular iron, lipid peroxidation (LPO), decrease of GSH and inhibition of GPX4 in biochemistry. Japanese encephalitis virus (JEV) and Herpes simplex virus (HSV) are both common neurotropic viruses that can cause neurological disorders, such as severe encephalitis. JEV and HSV have been demonstrated to be able to induce ferroptosis. This process is closely related to the inhibition of the GSH-GPX4 system, ACSL4 phosphorylation, and Nrf2 ubiquitination. In this review, we summarized the mechanisms by which JEV and HSV induced ferroptosis in the current study. In addition, we found a strong relationship between endoplasmic reticulum (ER) stress and ferroptosis, and we therefore speculated that sustained ER stress might be a prerequisite for ferroptosis in JEV and HSV-induced diseases.

BackgroundEarly childhood caries (ECC) are a prevalent chronic disease in young children. However, there has been limited research on the microbiota in different tissue levels of the same tooth in children with ECC. This study aimed to investigate the dynamic changes in bacterial diversity during the progression of Severe Early Childhood Caries (S-ECC) within the same tooth, from the tooth surface to the root canal, by collecting tissue samples from different areas of the affected tooth.MethodsTwenty primary teeth with periapical periodontitis were selected from 20 children aged 3–5 years, with 100 samples collected from the different layers: uncavitated buccal enamel surface without white spot lesion (surface), the outermost layer of the dentin carious lesion (superficial), the inner layer of carious dentin (deep), necrotic pulp tissue (pulp), and root exudate (exudate). The taxonomy of each OTU representative sequence was analyzed against the 16S rRNA database. Comparisons of alpha diversity between groups were performed. The number of shared and unique genera between groups counted. Beta diversity was contrasted to evaluate differences in bacterial community composition, and the relationships between the microbiota and samples were analyzed. The heatmap analysis of the 30 most abundant genera was used, which highlighted their relative distribution and abundance. The significantly abundant taxa (phylum to genera) of bacteria among the different groups were identified. The differences of relative abundance between bacterial genera among the five groups were analyzed. Significant Spearman correlations were noted, and visualization of the co-occurrence network was conducted.ResultsBacterial 16S rRNA gene sequencing showed that most genera were present in all layers, with the number of shared genera increasing as the disease advanced. The bacterial communities and core genera in the co-occurrence network changed with progression to severe ECC.ConclusionAn increase in both the quantity and complexity of bacterial interactions was observed. This study emphasized the importance of paying attention to the relationship between microbial species rather than just checking changes in bacterial species structure when investigating the role of bacteria in disease progression.

IntroductionThis study investigates the dynamic shifts in soil bacterial communities within a Salix matsudana afforested ecosystem transitioning from agricultural land. Understanding the temporal variability in bacterial diversity and community structures is crucial for informing forest management and conservation strategies, particularly in regions undergoing afforestation.MethodsWe employed high-throughput sequencing across three distinct months (August, September, and October) to analyze the temporal variability in bacterial community composition and diversity. Network analysis was utilized to identify keystone species and assess community stability under varying environmental conditions, including fluctuations in temperature and precipitation.ResultsWe uncover significant temporal variability in bacterial diversity and community structures, which are closely tied to fluctuations in temperature and precipitation. Our findings reveal the abundance of the dominant bacterial phyla, such as Actinobacteria and Proteobacteria, which did not change overall, highlighting the stability and resilience of the microbial community across seasonal transitions. Notably, the increasing similarity in community composition from August to October indicates a reduction in species turnover, likely driven by more homogeneous environmental conditions. Through comprehensive network analysis, we identify the pivotal role of keystone species, particularly the human pathogen Nocardia, in maintaining community stability under reduced soil moisture. The observed variations in community connectivity underscore the microbial community’s resilience and adaptability to seasonal shifts, with higher stability in August and October contrasting with the instability observed in September.DiscussionThese results underscore the complex interplay between stochastic and deterministic processes in bacterial community assembly, significantly shaped by prevailing environmental conditions. The insights gained from this research have far-reaching implications for forestry management and conservation strategies, particularly in regions undergoing similar afforestation efforts.

Selenium (Se) is a vital trace element integral to numerous biological processes in both plants and animals, with significant impacts on soil health and ecosystem stability. This review explores how soil microorganisms facilitate Se transformations through reduction, oxidation, methylation, and demethylation processes, thereby influencing the bioavailability and ecological functions of Se. The microbial reduction of Se compounds, particularly the conversion of selenate and selenite to elemental Se nanoparticles (SeNPs), enhances Se assimilation by plants and impacts soil productivity. Key microbial taxa, including bacteria such as Pseudomonas and Bacillus, exhibit diverse mechanisms for Se reduction and play a substantial role in the global Se cycle. Understanding these microbial processes is essential for advancing soil management practices and improving ecosystem health. This review underscores the intricate interactions between Se and soil microorganisms, emphasizing their significance in maintaining ecological balance and promoting sustainable agricultural practices.

BackgroundWheat germ is known for its antioxidant, anti-inflammatory, and disease resistance properties in animals. However, its effect on the gut of Sichuan white geese remains unclear.MethodIn this study, thirty 250-day-old geese were divided into three equal groups, the control group, LWG group (21.8% wheat germ) and HWG group (43.6% wheat germ), the experiment lasted 12 weeks. We assessed various aspects of geese intestinal health, including barrier function, digestibility, antioxidant capacity, immunity, microbiota, and metabolism.ResultsThe study revealed a significant increase in villus height (VH), villus height-to-crypt depth (VH/CD) ratio, amylase, and lipase activities in the duodenum and ileum, increased putrescine levels in the duodenum and jejunum, as well as spermidine levels in the jejunum (P < 0.05). LWG increased the total antioxidant capacity (T-AOC) in the duodenum, while decreasing levels of intestinal malondialdehyde (MDA), serum lipopolysaccharide (LPS), interleukin-6 (IL-6), and diamine oxidase (DAO) activity (P < 0.05). Furthermore, LWG increased the relative abundance of Oscillospiraceae_unclassified, Ligilactobacillus, and Roseburia, as well as increased levels of acetic acid, butyric acid, and valeric acid, while decreasing the relative abundance of Subdoligranulum, Flavonifractor, and Klebsiella. Additionally, we observed 17 up-regulated genes and 25 down-regulated genes in the jejunum, which are associated with the cell cycle and immunity. These genes play roles in pathways such as the p53 signaling pathway, cell cycle regulation, and pathways associated with immune modulation. On the other hand, HWG increased intestinal VH and spermidine levels, as well as amylase and lipase activities in the duodenum (P < 0.05). It also elevated ileal T-AOC and sIgA levels (P < 0.05), while reducing intestinal MDA content, serum LPS levels, DAO activity, and propionic acid in cecum contents (P < 0.05). Moreover, HWG increased the relative abundance of Ligilactobacillus, Oscillospiraceae_unclassified, and Roseburia (P < 0.05).ConclusionOverall, wheat germ diets, particularly the LWG diet demonstrated the ability to enhance antioxidant capacity, digestibility, immunity, and barrier properties of the intestinal tract, while modulating the gut microbiota and metabolism. Therefore, wheat germ diets hold promise in improving intestinal health by preserving barrier function and regulating flora structure.

BackgroundThe association between preterm birth and Mycoplasma species such as Mycoplasma hominis and Ureaplasma urealyticum has been extensively investigated. In a clinical setting, conventional diagnostic methods for them involve culture methods for Mycoplasma spp. and Ureaplasma spp., along with PCR tests. However, the clinical utility of these tests remains controversial, highlighting the necessity for more robust and reliable methods for identifying and understanding Mycoplasma infections.ObjectiveThis study aimed to assess the distribution of microbiota in pregnant women with Mycoplasma hominis and Ureaplasma urealyticum infection by the comparison of conventional diagnostic methods with vaginal microbial community analysis.Study DesignThis prospective case–control study involved 228 Korean pregnant women and utilized vaginal microbial community analysis, Ureaplasma/Mycoplasma culture, and 12-multiplex PCR for sexually transmitted diseases. Cross-correlation analysis in SPSS 27 compared the results of two conventional methods with vaginal microbial community analysis. R software generated box plots depicting the relative abundance of microorganisms. Network analysis was conducted using Cytoscape.ResultsPositive Ureaplasma urealyticum culture findings were observed in 60.2% of patients, with 76.4% positive for Ureaplasma parvum PCR and 13.2% positive for Ureaplasma urealyticum PCR. Mycoplasma hominis culture was positive only in two patients, while Mycoplasma hominis PCR was positive in eight women. Vaginal microbial community analysis identified significant differences in relative abundances of Gardnerella species type I and Fannyhessea vaginae between the Ureaplasma urealyticum PCR positive and negative groups. Mycoplasma hominis PCR positive patients exhibited significant differences in 11 bacterial species, including Gardnerella species I and Fannyhessea vaginae.ConclusionThis study suggests that STD-PCR may be more accurate than Ureaplasma/Mycoplasma culture for the diagnosis of Mycoplasma hominis and Ureaplasma urealyticum infection. Also, the presence of Gardnerella species I and Fannyhessea vaginae implies their potential influences on Ureaplasma urealyticum and Mycoplasma hominis infections based on results of vaginal microbial community analysis. Therefore, vaginal microbial community analysis may give the more information of their pathophysiology.

IntroductionWe report head-to-head comparison of the bivalent and quadrivalent HPV vaccine efficacies against immediate precursors of cervical cancer from 15 years’ country-wide cancer registry follow-up of phase III trial cohorts and an age-aligned cohort of unvaccinated women.MethodsThese individually and/or clusterrandomized cohorts of HPV6/11/16/18- and HPV16/18-vaccinated and unvaccinated women were enrolled, respectively, in 2002, 2004, and 2003/2005. The trial cohorts comprised initially 16- to 17-year-old HPV6/11/16/18-vaccinated FUTURE II (NCT00092534) participants (866) and HPV16/18-vaccinated PATRICIA (NCT00122681) and 012 trial (NCT00169494) participants (2,465), and 16,526 initially 16- to 19-year-old unvaccinated controls. After active 4-year clinical follow-up, passive, country-wide Finnish Cancer Registry (FCR) follow-up for cervical intraepithelial neoplasia grade 3 (CIN3) and adenocarcinoma in situ (AIS) was based on consented use of unique personal identifiers and started 6 months after the end of the FUTURE II and PATRICIA trials in 2007 and 2009, and ended at the end of 2019. The follow-up with altogether 229,020 follow-up years was age-aligned to ensure that similarly aged cohorts were passively followed up for 15 years post=vaccination for the intention-to-treat analyses of vaccine efficacy.ResultsOverall, we identified 5 and 16 CIN3 (no AIS) cases in the HPV6/11/16/18 and HPV16/18 cohorts, respectively, during the FCR-based follow-up. In the unvaccinated cohort, we identified 281 CIN3 cases, 20 AIS cases, and 13 cases with invasive cervical cancer. Vaccine efficacies against CIN3+ were 68.4% and 64.5% for the quadrivalent and the bivalent vaccines, respectively, with overlapping confidence intervals.DiscussionLong-term follow-up of randomized, initially adolescent HPV-vaccinated and unvaccinated cohorts shows, in this head-to-head setting, that the bivalent and quadrivalent HPV vaccines are equally effective against immediate precursors of cervical cancer.

IntroductionThus far, the impact of kaolin mining activities on the surrounding native plants and rhizosphere microecology has not been fully understood.MethodsIn this study, we used 16S rRNA high-throughput sequencing to examine the impact of kaolin mining on the rhizosphere bacterial communities and functions of three local plant species: Conyza bonariensis, Artemisia annua, and Dodonaea viscosa.ResultsThe results showed that kaolin mining significantly reduced the diversity of rhizosphere bacteria in these plants, as indicated by the Shannon, Simpson, Chao1, and observed species indices (p