Immunity against Moraxella catarrhalis requires guanylate-binding proteins and caspase-11-NLRP3 inflammasomes.

Moraxella catarrhalis is an important human respiratory pathogen and a major causative agent of otitis media and chronic obstructive pulmonary disease. Toll-like receptors contribute to, but cannot fully account for, the complexity of the immune response seen in M. catarrhalis infection. Using primary mouse bone marrow-derived macrophages to examine the host response to M. catarrhalis infection, our global transcriptomic and targeted cytokine analyses revealed activation of immune signalling pathways by both membrane-bound and cytosolic pattern-recognition receptors. We show that M. catarrhalis and its outer membrane vesicles or lipooligosaccharide (LOS) can activate the cytosolic innate immune sensor caspase-4/11, gasdermin-D-dependent pyroptosis, and the NLRP3 inflammasome in human and mouse macrophages. This pathway is initiated by type I interferon signalling and guanylate-binding proteins (GBPs). We also show that inflammasomes and GBPs, particularly GBP2, are required for the host defence against M. catarrhalis in mice. Overall, our results reveal an essential role for the interferon-inflammasome axis in cytosolic recognition and immunity against M. catarrhalis, providing new molecular targets that may be used to mitigate pathological inflammation triggered by this pathogen.

Clostridium perfringens virulence factors are nonredundant activators of the NLRP3 inflammasome.

Inflammasome signaling is a central pillar of innate immunity triggering inflammation and cell death in response to microbes and danger signals. Here, we show that two virulence factors from the human bacterial pathogen Clostridium perfringens are nonredundant activators of the NLRP3 inflammasome in mice and humans. C. perfringens lecithinase (also known as phospolipase C) and C. perfringens perfringolysin O induce distinct mechanisms of activation. Lecithinase enters LAMP1+ vesicular structures and induces lysosomal membrane destabilization. Furthermore, lecithinase induces the release of the inflammasome-dependent cytokines IL-1ß and IL-18, and the induction of cell death independently of the pore-forming proteins gasdermin D, MLKL and the cell death effector protein ninjurin-1 or NINJ1. We also show that lecithinase triggers inflammation via the NLRP3 inflammasome in vivo and that pharmacological blockade of NLRP3 using MCC950 partially prevents lecithinase-induced lethality. Together, these findings reveal that lecithinase activates an alternative pathway to induce inflammation during C. perfringens infection and that this mode of action can be similarly exploited for sensing by a single inflammasome.

Symptomatic responses elicited by electrical stimulation of the cingulate cortex: Study of a cohort of epileptic patients and literature review.

Our understanding of cingulate cortex function is limited. As a method for locating the epileptogenic zone, direct electrical cortical stimulation (ECS) provides an opportunity to understand the functional localization of the cingulate cortex. This study aimed to learn more about the function of the cingulate cortex by analyzing a large body of data from our center and by reviewing existing literature on cortical mapping. We retrospectively analyzed the ECS data of 124 patients with drug-resistant epilepsy who had undergone electrode implantation in the cingulate cortex. The standard stimulation parameters included a biphasic pulse and bipolar stimulation at 50 Hz. Furthermore, we reviewed existing studies on cingulate responses elicited by the ECS and compared them with our results. A total of 329 responses were evoked in 276 contacts using ECS. Of these, 196 were physiological functional responses, which included sensory, affective, autonomic, language, visual, vestibular, and motor responses, along with a few other sensations. Sensory, motor, vestibular, and visual responses were concentrated in the cingulate sulcus visual area (CSv). Furthermore, 133 epilepsy-related responses were evoked, most of which were concentrated in the ventral cingulate cortex. No responses were evoked by 498 contacts. Furthermore, the comparison of our ECS results with those reported in 11 comprehensive reviews revealed that the cingulate cortex is involved in complicated functions. The cingulate cortex is involved in sensory, affective, autonomic, language, visual, vestibular, and motor functions. The CSv is an integrating node of sensory, motor, vestibular, and visual systems.

Unraveling enterohemorrhagic Escherichia coli infection: the promising role of dietary compounds and probiotics in bacterial elimination and host innate immunity boosting.

The innate immune system has developed sophisticated strategies to defense against infections. Host cells utilize the recognition machineries such as toll-like receptors and nucleotide binding and oligomerization domain-like receptors to identify the pathogens and alert immune system. However, some pathogens have developed tactics to evade host defenses, including manipulation of host inflammatory response, interference with cell death pathway, and highjack of phagocytosis signaling for a better survival and colonization in host. Enterohemorrhagic Escherichia coli (EHEC) is a notorious foodborne pathogen that causes severe tissue damages and gastrointestinal diseases, which has been reported to disturb host immune responses. Diverse bioactive compounds such as flavonoids, phenolic acids, alkaloids, saccharides, and terpenoids derived from food varieties and probiotics have been discovered and investigated for their capability of combating bacterial infections. Some of them serve as novel antimicrobial agents and act as immune boosters that harness host immune system. In this review, we will discuss how EHEC, specifically E. coli O157:H7, hijacks the host immune system and interferes with host signaling pathway; and highlight the promising role of food-derived bioactive compounds and probiotics in harnessing host innate immunity and eliminating E. coli O157:H7 infection with multiple strategies.

Emerging Activators and Regulators of Inflammasomes and Pyroptosis.

The inflammasome is a cytosolic immune signaling complex that induces inflammation and pyroptosis. Inflammasome complexes respond to a variety of pathogens, as well as danger or homeostasis-altering signals; they can play critical roles in the development of autoinflammatory conditions and cancer. Studies have now provided additional insights into the activation mechanisms and regulation of established inflammasome complexes, including NLRP1b, NLRP3, NOD-like receptor family apoptosis inhibitory protein (NAIP)-NLRC4, absent in melanoma (AIM)2, caspase-11, and pyrin. New activators and regulators of emerging NLRP6 and NLRP9b inflammasome complexes have also been described. We highlight the latest progress in our understanding of the molecular mechanisms governing inflammasome activation and pyroptosis, including the discovery of the pore-forming protein gasdermin D (GSDMD). We also discuss the importance of inflammasome activators and regulators in health and disease.

Effects of 1-methylcyclopropene and gaseous ozone on Listeria innocua survival and fruit quality of Granny Smith apples during long-term commercial cold storage.

This study evaluated the impact of 1-methylcyclopropene (1-MCP), an ethylene synthesis inhibitor, followed by long-term commercial cold storage with low-dose gaseous ozone on the microbiological safety and quality of fresh apples. Granny Smith apples were inoculated with or without Listeria innocua, treated with or without 1.0 mg/L 1-MCP for 24 h, then subjected to commercial cold storage conditions including refrigerated air (RA, 0.6 °C, control), controlled atmosphere (CA, 2% O2, 1% CO2, 0.6 °C), and CA with 51-87 µg/L ozone gas for up to 36 weeks. RA storage reduced L. innocua on apples by up to 3.6 log10 CFU/apple. CA had no advantage over RA in controlling Listeria. Continuous ozone gas application resulted in an additional ∼2.0 log10 CFU/apple reduction of L. innocua (total reduction up to 5.7 log10 CFU/apple) and suppressed native bacteria and fungi. Treatment with 1-MCP had a minor impact on survival of L. innocua or background microbiota on apples, while it significantly delayed fruit ripening and reduced the incidence of superficial scald and internal browning. In summary, 1-MCP treatment followed by CA storage with low-dose continuous ozone gas can effectively control Listeria on fresh apples and delay fruit ripening.

Integrated multi-omics approaches to understand microbiome assembly in Jiuqu, a mixed-culture starter.

The use of Jiuqu as a saccharifying and fermenting starter in the production of fermented foods is a very old biotechnological process that can be traced back to ancient times. Jiuqu harbors a hub of microbial communities, in which prokaryotes and eukaryotes cohabit, interact, and communicate. However, the spontaneous fermentation based on empirical processing hardly guarantees the stable assembly of the microbiome and a standardized quality of Jiuqu. This review describes the state of the art, limitations, and challenges towards the application of traditional and omics-based technology to study the Jiuqu microbiome and highlights the need for integrating meta-omics data. In addition, we review the varieties of Jiuqu and their production processes, with particular attention to factors shaping the microbiota of Jiuqu. Then, the potentials of integrated omics approaches used in Jiuqu research are examined in order to understand the assembly of the microbiome and improve the quality of the products. A variety of different approaches, including molecular and mass spectrometry-based techniques, have led to scientific advances in the analysis of the complex ecosystem of Jiuqu. To date, the extensive research on Jiuqu has mainly focused on the microbial community diversity, flavor profiles, and biochemical characteristics. An integrative approach to large-scale omics datasets and cultivated microbiota has great potential for understanding the interrelation of the Jiuqu microbiome. Further research on the Jiuqu microbiome may explain the inherent property of compositional stability and stable performance of a complex microbiota coping with environmental perturbations and provide important insights to reconstruct synthetic microbiota and develop modern intelligent manufacturing procedures for Jiuqu.

Comparative study on inhibitory effects of ferulic acid and p-coumaric acid on Salmonella Enteritidis biofilm formation.

Biofilm cells exhibit higher resistance than their planktonic counterparts to commonly used disinfectants in food industry. Phenolic acids are promising substitute offering less selective pressure than traditional antibiotics. This study aims to evaluate the inhibitory effects of ferulic acid (FA) and p-coumaric acid (p-CA) on Salmonella Enteritidis biofilm formation and explore the underlying inhibitory mechanisms. The minimal inhibitory concentration (MIC) of FA and p-CA were 1.0 and 0.5 mg/ml, respectively. The sub-inhibitory concentration (1/8 MIC) significantly decreased biofilm formation without growth inhibitory effects. The biomass and extracellular polymeric substances (EPS) of S. Enteritidis biofilm as well as the bacterial swimming and chemotaxis abilities were significantly decreased when exposed to sub-MIC concentrations of FA and p-CA. These two phenolic acids showed high affinity to proteins involved in flagella motility and repressed the S. Enteritidis biofilm formation-related gene expressions. Furthermore, these two phenolic acids maintained high antibiofilm efficiency in simulated food processing conditions. This study provided valuable information of multiple phenotypic and molecular responses of S. Enteritidis to these two phenolic acids.

Characteristics of the microbiota and metabolic profile of high-temperature Daqu with different grades.

The superior grade Daqu (S_Daqu) and normal grade Daqu (N_Daqu) have obvious differences in flavor, fracture surface, appearance, etc., which can be accurately grouped by well-trained panel based on their sensory properties. However, the differences in microbial community diversity and metabolites between the S_Daqu and N_Daqu were still unclear. The culture-dependent method, the third generation Pacific Biosciences (PacBio) single-molecule, real-time (SMRT) sequencing technology, and nuclear magnetic resonance (NMR) were combined to show the characteristics in microorganisms and metabolites. Results showed that the fungal counts were higher in N_Daqu while the richness of bacterial communities was higher in S_Daqu (P < 0.05). Lentibacillus, Burkholderia, Saccharopolyspora, Thermoascus, and Rasamsonia were the dominant genera of S_Daqu while Staphylococcus, Scopulibacillus, and Chromocleista were the dominant genera in N_Daqu. The content of differential acids, amino acids, and alcohols including fumarate, glucuronate, glycine, 4-carboxyglutamate, and myo-inositol in S_Daqu was higher than that in N_Daqu by 1H NMR coupled with multivariate statistical analysis. The network analysis regarding microbes and metabolites suggested that Saccharopolyspora showed a strong positive correlation with 4-carboxyglutamate while Thermoascus and Chromocleista were highly negatively correlated with alanine and isobutyrate, respectively. Linear Discriminant Analysis (LDA) Effect Size (LEfSe) revealed that Macrococcus and Caulobacter were regarded as bacterial biomarkers in the S_Daqu while Chromocleista was the key fungal genera in N_Daqu. Functionality prediction indicated that the bacteria in S_Daqu were largely involved in more metabolic activities including biosynthesis, degradation, detoxification, and generation of precursor metabolite and energy.

Genetic interaction of purinergic P2X7 receptor and ER-α polymorphisms in susceptibility to osteoporosis in Chinese postmenopausal women.

Osteoporosis (OP) is an increasing public health problem worldwide. Genetic factors are considered to be major contributors to the pathogenesis of OP. The aim of this study was to investigate the association of the purinergic P2X7 receptor (P2X7R) and estrogen receptor-α (ER-α) genes with OP risk, and the effect of the possible interaction between the two genes on predisposition to OP in Chinese postmenopausal women. A total of 596 subjects, including 350 OP patients and 246 controls, were recruited in this case-control study. Five functional single-nucleotide polymorphisms (SNPs) in the P2X7R gene (rs2393799, rs7958311, rs1718119, rs2230911, rs3751143) and two ER-α PvuII and XbaI polymorphisms were genotyped and analyzed. Single-gene variant analysis showed that the carriers of the CC genotype of P2X7R rs3751143 revealed an increased OP risk. Haplotype rs1718119G-rs2230911G-rs3751143C also appeared to be a significant 'risk' haplotype with OP. For the ER-α gene, no evidence of significant association of PvuII or XbaI polymorphism with OP risk was found. Moreover, there was a significant gene-gene interaction between P2X7R rs3751143 and ER-α PvuII; the cross-validation consistency was 10/10 and the testing accuracy was 0.5818 (P = 0.0107). A 1.67-fold-increased risk for OP was detected in individuals carrying the genotypes of AC or CC of rs3751143 and Pp or PP of PvuII compared to subjects with AA of rs3751143 and pp of PvuII. Our findings suggest an important association of the P2X7R rs3751143CC genotype and the rs1718119G-rs2230911G-rs3751143C haplotype with an increased OP risk. Also, the P2X7R rs3751143 and ER-α PvuII two-locus interaction confers a significantly high susceptibility to OP in Chinese postmenopausal women.

Favourable effects of grape seed extract on intestinal epithelial differentiation and barrier function in IL10-deficient mice.

The impairment in the rate of cell proliferation and differentiation leads to a negative consequence on the renewal of the intestinal epithelium, which is the aetiological factor of a number of digestive diseases. Grape seed extract (GSE), a rich source of proanthocyanidins, is known for its beneficial health effects. The present study evaluated the beneficial effects of GSE on colonic cell differentiation and barrier function in IL10-deficient mice. Female mice aged 6 weeks were randomised into two groups and given drinking-water containing 0 or 0.1 % GSE (w/v) for 12 weeks. GSE supplementation decreased serum TNF-α level and intestinal permeability, and increased the colonic goblet cell density that was associated with increased mRNA expression of mucin (Muc)-2. Immunohistochemical analyses showed lower accumulation of ß-catenin in the crypts of colon tissues of the GSE-supplemented mice, which was associated with a decreased mRNA expression of two downstream effectors of Wingless and Int (Wnt)/catenin signalling, myelocytomatosis oncogene protein (Myc) and cyclin D1 (Ccnd1). Consistently, GSE supplementation decreased the number of colonic proliferating cell nuclear antigen-positive cells, a well-known cell proliferation marker, and a weakened extracellular signal-regulated kinases 1 and 2 (ERK1/2) signalling. In summary, these data indicate that supplementation of 0.1 % GSE for 12 weeks improved gut barrier function and colonic cell differentiation in the IL10-deficient mice probably via inhibiting Wnt/ß-catenin pathway.

Host inflammatory response inhibits Escherichia coli O157:H7 adhesion to gut epithelium through augmentation of mucin expression.

Escherichia coli O157:H7, a major Shiga toxin-producing pathogen, has a low infectious dose and causes serious illness in humans. The gastrointestinal tract of cattle is the primary reservoir of E. coli O157:H7, and thus, it is critical to eliminate or reduce E. coli O157:H7 gut colonization. Given that E. coli O157:H7 produces effectors that attenuate inflammatory signaling, we hypothesized that the host inflammatory response acts to perturb E. coli O157:H7 intestinal colonization. Tumor necrosis factor alpha (TNF-α) treatment of HT-29 cells resulted in increased expression of inflammatory cytokine interleukin 1ß (IL-1ß), IL-8, and TNF-α genes and increased IL-8 protein and resulted in decreased adhesion of E. coli O157:H7. Similarly, E. coli O157:H7 adhesion to cattle colonic explants was reduced by TNF-α treatment. Irrespective of the presence of E. coli O157:H7, TNF-α enhanced activation of p65, the key mediator of NF-κB inflammatory signaling, whereas E. coli O157:H7 infection suppressed this pathway by inhibiting p65 activation in HT-29 cells. To further explore the mechanisms linking the inflammatory response to attenuated E. coli O157:H7 adhesion, mucin 2 (MUC2) expression was analyzed, considering that the intestinal mucus layer is the first defense against enteric pathogens and MUC2 is the major secretory mucin in the intestine. MUC2 expression in HT-29 cells was increased by TNF-α treatment and by E. coli O157:H7 infection. However, reducing mucin expression by blocking mitogen-activated protein kinase (MAPK) extracellular signal-regulated protein kinases 1/2 (ERK1/2) and/or phosphatidylinositol 3-kinase (PI3K)/Akt signaling increased E. coli O157:H7 adherence to HT-29 cells. These data suggest that the inflammatory cytokine response acts to protect host epithelial cells against E. coli O157:H7 colonization, at least in part, by promoting mucin production.

Maternal obesity exacerbates insulitis and type 1 diabetes in non-obese diabetic mice.

Accompanying the dramatic increase in maternal obesity, the incidence of type 1 diabetes (T1D) in children is also rapidly increasing. The objective of this study was to explore the effects of maternal obesity on the incidence of T1D in offspring using non-obese diabetic (NOD) mice, a common model for TID. Four-week-old female NOD mice were fed either a control diet (10% energy from fat, CON) or a high-fat diet (60% energy from fat) for 8 weeks before mating. Mice were maintained in their respective diets during pregnancy and lactation. All offspring mice were fed the CON to 16 weeks. Female offspring (16-week-old) born to obese dams showed more severe islet lymphocyte infiltration (major manifestation of insulitis) (P<0.01), concomitant with elevated nuclear factor kappa-light-chain-enhancer of activated B cells p65 signaling (P<0.01) and tumor necrosis factor alpha protein level (P<0.05) in the pancreas. In addition, maternal obesity resulted in impaired (P<0.05) glucose tolerance and lower (P<0.05) serum insulin levels in offspring. In conclusion, maternal obesity resulted in exacerbated insulitis and inflammation in the pancreas of NOD offspring mice, providing a possible explanation for the increased incidence of T1D in children.

Dietary purple potato supplement attenuates DSS-induced colitis in mice: impact on mitochondrial function.

Inflammatory bowel disease (IBD) is a condition characterized by disrupted intestinal barrier function, abnormal immune response, and mucosal structure loss. This study evaluated the beneficial role of purple potato (PP) supplementation against IBD symptoms using a murine model of dextran sulfate sodium (DSS)-induced colitis, and further explored the underlying mechanisms. Six-week-old C57BL/6J male mice were randomized into two groups and fed a standard rodent diet with or without 10% PP powder for 7 weeks. At the 5th week of dietary supplements, mice in each group were further divided into two subgroups and were either induced with or without 2.5% DSS induction for 7 days, followed by 7 days of recovery. Data showed that PP supplementation ameliorated the disease activity index in DSS-treated mice and reversed the colonic structure loss, mucosal damage, macrophage infiltration, and pro-inflammatory cytokine secretion induced by DSS in the colonic tissue. PP supplementation also restored the levels of tight junction proteins and caudal type homeobox 2 in DSS-treated mice. Furthermore, dietary PP enhanced peroxisome proliferator-activated receptor-γ coactivator-1α signaling pathway, mitochondrial biogenesis, mitochondrial proteostasis, and protein-folding capacity. In summary, dietary PP ameliorated DSS-induced colitis and improved gut structures and barrier function, which was associated with improved mitochondrial function. These results support further investigation of PP as a potential dietary intervention for IBD.

Enhancing biofilm penetration and antibiofilm efficacy with protein nanocarriers against pathogenic biofilms.

Nanocarriers play an important role in enhancing the efficacy of antibiotics against biofilms by improving their penetration and prolonging retention in pathogenic biofilms. Herein, the multifunctional nanocarriers including nanospheres (NS) and nanotubes (NT) with a high biocompatibility and biodegradability were prepared through self-assembly of partially hydrolyzed α-lactalbumin. The effects of these two different shaped nanocarriers on the delivery of antibiotics for biofilm treatment were examined by conducting in vitro antibiofilm experiment and in vivo infected wound model. The strong affinity of NS and NT for the bacterial surface allows antibiotics to be concentrated in the bacteria. Notably, the high permeability of NT into biofilms facilitates deeper penetration and the easier diffusion of loaded antibiotics within the biofilm. Furthermore, the acidic biofilm environment triggers the release of antibiotics from the NT, resulting in the accumulation of high local antibiotic concentrations. Therefore, NT could efficiently clean and inhibit the biofilm formation while also destroying the mature biofilms. In a S. aureus infected wound animal model, treatment with antibiotic-loaded NT demonstrated accelerated healing of S. aureus infected wounds when compared to free antibiotic treatment. These findings indicate that NT nanocarrier strategy is promising for treating bacterial biofilm infections, offering the potential for lower antibiotics dosages and preventing the overuse of antibiotics.

The role of magnetoencephalography in preoperative localization and postoperative outcome prediction in patients with posterior cortical epilepsy.

OBJECTIVE: We aimed to explore the value of magnetoencephalography in the presurgical evaluation of patients with posterior cortex epilepsy. METHODS: A total of 39 patients with posterior cortex epilepsy (PCE) and intact magnetoencephalography (MEG) images were reviewed from August 2019 to July 2022. MEG dipole clusters were classified into single clusters, multiple clusters, and scatter dipoles based on tightness criteria. The association of the surgical outcome with MEG dipole classifications was evaluated using Fisher's exact tests. RESULTS: Among the 39 cases, there were 24 cases of single clusters (61.5%), nine cases of multiple clusters (23.1%), and six cases of scattered dipoles (15.4%). Patients with single dipole clusters were more likely to become seizure-free. Among single dipole cluster cases (n = 24), complete MEG dipole resection yielded a more favorable surgical outcome than incomplete resection (83.3% vs. 16.7%, p = 0.007). Patients with concordant MRI and MEG findings achieved a significantly more favorable surgical outcome than discordant patients (66.7% vs. 33.3%, p = 0.044), especially in single dipole cluster patients (87.5% vs. 25.0%, p = 0.005). SIGNIFICANCE: MEG can provide additional valuable information regarding surgical candidate selection, epileptogenic zone localization, electrode implantation schedule, and final surgical planning in patients with posterior cortex epilepsy.

Impact of Metacognition on Health-Related Behavior: A Mediation Model Study.

Objective: The study aims to explore the correlation mechanism among metacognition, attitude toward physical exercise, and health-related behavior in high school students. Methods: A total of 869 students (17 ± 1.70) from Anhui, Zhejiang, Shandong, and Fujian provinces were selected by stratified sampling to complete the Metacognition Questionnaire, Health-Related Behavior Self-Rating Scale, Attitude Toward Physical Exercise Scale, and Depression-Anxiety-Stress Scale (Simplified Chinese version, DASS-21). Results: (1) Metacognition was negatively predictive of attitude toward physical exercise and health-related behavior (ß = -0.236, P < 0.01; ß = -0.239, P < 0.01) but positively predictive of negative emotion (ß = 0.496, P < 0.01); (2) attitude toward physical exercise was positively predictive of health-related behavior (ß = 0.533, P < 0.01) but negatively predictive of negative emotion (ß = -0.336, P < 0.01); and (3) negative emotion was negatively predictive of health-related behavior (ß = -0.389, P < 0.01). Conclusions: Metacognition not only has a directly predictive effect on health-related behavior but also predicts it through attitude toward physical exercise. Negative emotion also mediates the relationship between metacognition and attitude toward physical exercise.

Engineering conjugative CRISPR-Cas9 systems for the targeted control of enteric pathogens and antibiotic resistance.

Pathogenic Escherichia coli and Salmonella enterica pose serious public health threats due to their ability to cause severe gastroenteritis and life-threatening sequela, particularly in young children. Moreover, the emergence and dissemination of antibiotic resistance in these bacteria have complicated control of infections. Alternative strategies that effectively target these enteric pathogens and negate or reduce the need of antibiotics are urgently needed. Such an alternative is the CRISPR-Cas9 system because it can generate sequence-specific lethal double stranded DNA breaks. In this study, two self-transmissible broad host range conjugative plasmids, pRK24 and pBP136, were engineered to deliver multiplexed CRSIPR-Cas9 systems that specifically target Enterohemorrhagic and Enteropathogenic strains of E. coli (EHEC and EPEC), S. enterica, and blaCMY-2 antibiotic resistance plasmids. Using in vitro mating assays, we show that the conjugative delivery of pRK24-CRISPR-Cas9 carrying guide RNAs to the EPEC/EHEC eae (intimin) gene can selectively kill enterohemorrhagic E. coli O157 eae+ cells (3 log kill at 6 h) but does not kill the isogenic Δeae mutant (P<0.001). Similar results were also obtained with a pBP136 derivative, pTF16, carrying multiplexed guide RNAs targeting E. coli eae and the S. enterica ssaN gene coding for the type III secretion ATPase. Another pBP136 derivative, TF18, carries guide RNAs targeting S. enterica ssaN and the antibiotic resistance gene, blaCMY-2, carried on the multi-drug resistant pAR06302. Introduction of pTF18 into bacteria harboring pAR06302 showed plasmids were cured at an efficiency of 53% (P<0.05). Using a murine neonate EPEC infection model, pTF16 was delivered by a murine derived E. coli strain to EPEC infected mice and showed significant reductions of intestinal EPEC (P<0.05). These results suggest that establishing conjugative CRISPR-Cas9 antimicrobials in the intestinal microbiome may provide protection from enteric pathogens and reduce antibiotic resistance without disrupting the normal microbiota.

Insights into the bacterial, fungal, and phage communities and volatile profiles in different types of Daqu.

Daqu has a rich and diverse microbiota, giving them a suitable biotope for phages. However, the absolute abundances of bacteria and fungi, as well as the phage community characteristics in Daqu, remain unclear. In this study, the microbiota absolute abundance, indigenous phage composition and function, and volatile compound profiles of high-temperature Daqu (HTDQ), medium-temperature Daqu (MTDQ), and low-temperature Daqu (LTDQ) were investigated. Absolute microbiota quantification revealed that there were significant variations in microbial composition and absolute abundance across three types of Daqu. The absolute abundances of the top 30 bacterial genera in LTDQ, MTDQ, and HTDQ were 6.0 × 105, 5.3 × 104, 1.4 × 105 copies/ng DNA, while the top 30 fungal genera had 8.5 × 107, 2.1 × 106, and 6.2 × 105 copies/ng DNA, respectively. LTDQ were enriched in Pantoea, Staphylococcus, and Saccharomycopsis; MTDQ were dominated by Saccharopolyspora, Staphylococcus, Saccharomycopsis, and Aspergillus; HTDQ were dominated by Saccharopolyspora, Bacillus, Byssochlamys, and Saccharomycopsis. Volatile profile analysis revealed that LTDQ, MTDQ, and HTDQ comprised 68, 68, and 75 compounds, respectively, with 39 compounds shared by the three types. Fourteen volatile compounds were identified as highly discriminative features among three types of Daqu, which were closely related to Saccharopolyspora, Thermoactinomyces, Kroppenstedtia, Byssochlamys, and Thermomyces. Metaviromics indicated that Genomoviridae, Mimiviridae and Siphoviridae, and Parvoviridae were the dominant viruses in LTDQ, MTDQ, and HTDQ, respectively. The hosts of phages in Daqu mainly included Lactobacillus, Enterobacter, Pantoea, Bacillus, Pediococcus, and Staphylococcus. Phages may prey on numerous microbes living on Daqu via pathways such as genetic information processing, signaling and cellular processes, and replication and repair. This study highlights the use of absolute quantification to avoid misjudgment of differential taxa in comparative microbiome analysis and provides new insights into the phage community and function in Daqu.

Planktonic Growth and Biofilm Formation by Providencia rettgeri and Subsequent Effect of Tannic Acid Treatment under Food-Related Environmental Stress Conditions.

ABSTRACT: Providencia rettgeri is an opportunistic foodborne pathogen with a strong biofilm-forming ability in low-nutrition environments. However, information regarding the impact of simulated food processing conditions on P. rettgeri planktonic growth and biofilm formation is limited. Using response surface methodology (RSM), the combined effects of temperature (19 to 37°C), pH (5 to 9), and sodium chloride (NaCl) concentration (0.50 to 2.0%, w/v) were applied to construct planktonic growth and biofilm formation models for P. rettgeri. For both RSM models, an increase in NaCl concentration restricted P. rettgeri growth. Planktonic growth and biofilm formation were maximum at 27.83 and 25.41°C, respectively. Tannic acid (TA) is a highly effective antibacterial agent that inhibited planktonic and biofilm P. rettgeri under optimal growth conditions. The viability of P. rettgeri cells was decreased by TA treatment, which caused destruction of the cell membrane and production of endogenous reactive oxygen species. TA significantly inactivated P. rettgeri biofilms, as verified by observation. The obtained models in this study may be useful for describing the impact of temperature, pH, and NaCl concentration on the growth by P. rettgeri in the food processing environment and better understanding the impacts of food-related conditions on bacterial planktonic growth and biofilm formation. These results obtained for P. rettgeri planktonic cells and biofilms can provide a framework for removal strategies for other foodborne pathogens.