Aspirin prevents colorectal cancer by regulating the abundance of Enterococcus cecorum and TIGIT+Treg cells.

Although aspirin can reduce the incidence of colorectal cancer (CRC), there is still uncertainty about its significance as a treatment for CRC, and the mechanism of aspirin in CRC is not well understood. In this study, we used aspirin to prevent AOM/DSS-induced CRC in mice, and the anti-CRC efficacy of aspirin was assessed using haematoxylin and eosin (H&E) staining and by determining the mouse survival rate and tumour size. 16S rDNA sequencing, flow cytometry (FCM), and Western blotting were also conducted to investigate the changes in the gut microbiota, tumour immune microenvironment, and apoptotic proteins, respectively. The results demonstrated that aspirin significantly exerted anti-CRC effects in mice. According to 16S rDNA sequencing, aspirin regulated the composition of the gut microbiota and dramatically reduced the abundance of Enterococcus cecorum. FCM demonstrated that there were more CD155 tumour cells and CD4 + CD25 + Treg cells showed increased TIGIT levels. Moreover, increased TIGIT expression on Treg cells is associated with reduced Treg cell functionality. Importantly, the inhibition of Treg cells is accompanied by the promotion of CD19 + GL-7 + B cells, CD8 + T cells, CD4 + CCR4 + Th2 cells, and CD4 + CCR6 + Th17 cells. Overall, aspirin prevents colorectal cancer by regulating the abundance of Enterococcus cecorum and TIGIT + Treg cells.

Comparative study of the gut microbial community structure of Spodoptera frugiperda and Spodoptera literal (Lepidoptera).

Background: Spodoptera frugiperda, the fall armyworm is a destructive invasive pest, and S. litura the tobacco cutworm, is a native species closely related to S. frugiperda. The gut microbiota plays a vital role in insect growth, development, metabolism and immune system. Research on the competition between invasive species and closely related native species has focused on differences in the adaptability of insects to the environment. Little is known about gut symbiotic microbe composition and its role in influencing competitive differences between these two insects. Methods: We used a culture-independent approach targeting the 16S rRNA gene of gut bacteria of 5th instar larvae of S. frugiperda and S. litura. Larvae were reared continuously on maize leaves for five generations. We analyzed the composition, abundance, diversity, and metabolic function of gut microbiomes of S. frugiperda and S. litura larvae. Results: Firmicutes, Proteobacteria, and Bacteroidetes were the dominant bacterial phyla in both species. Enterococcus, ZOR0006, Escherichia, Bacteroides, and Lactobacillus were the genera with the highest abundance in S. frugiperda. Enterococcus, Erysipelatoclostridium, ZOR0006, Enterobacter, and Bacteroides had the highest abundance in S. litura. According to α-diversity analysis, the gut bacterial diversity of S. frugiperda was significantly higher than that of S. litura. KEGG analysis showed 15 significant differences in metabolic pathways between S. frugiperda and S. litura gut bacteria, including transcription, cell growth and death, excretory system and circulatory system pathways. Conclusion: In the same habitat, the larvae of S. frugiperda and S. litura showed significant differences in gut bacterial diversity and community composition. Regarding the composition and function of gut bacteria, the invasive species S. frugiperda may have a competitive advantage over S. litura. This study provides a foundation for developing control strategies for S. frugiperda and S. litura.

Species prevalence, virulence genes, and antibiotic resistance of enterococci from food-producing animals at a slaughterhouse in Turkey.

Healthy cattle, sheep, and goats can be reservoirs for gastrointestinal pathogenic fecal enterococci, some of which could be multidrug-resistant to antimicrobials. The objective of this study was to determine the prevalence and diversity of Enterococcus species in healthy sheep, goat, and cattle carcasses, as well as to analyze the antimicrobial resistance phenotype/genotype and the virulence gene content. During 2019-2020, carcass surface samples were collected from 150 ruminants in a slaughterhouse. A total of 90 enterococci, comprising five species, were obtained. The overall prevalence of enterococci was found to be 60%, out of which 37.7% were identified as Enterococcus (E.) hirae, 33.3% as E. casseliflavus, 15.5% as E. faecium, 12.2% as E. faecalis, and 1.1% as E. gallinarum. Virulence-associated genes of efaA (12.2%) were commonly observed in the Enterococcus isolates, followed by gelE (3.3%), asaI (3.3%), and ace (2.2%). High resistance to quinupristin-dalfopristin (28.8%), tetracycline (21.1%), ampicillin (20%), and rifampin (15.5%) was found in two, four, four, and five of the Enterococcus species group, respectively. The resistance of Enterococcus isolates to 11 antibiotic groups was determined and multidrug resistant (MDR) strains were found in 18.8% of Enterococcus isolates. Characteristic resistance genes were identified by PCR with an incidence of 6.6%, 2.2%, 1.1%, 1.1%, 1.1%, and 1.1% for the tetM, ermB, ermA, aac(6')Ie-aph(2")-la, VanC1, and VanC2 genes in Enterococcus isolates, respectively. Efflux pump genes causing multidrug resistance were detected in Enterococcus isolates (34.4%). The results showed that there were enterococci in the slaughterhouse with a number of genes linked to virulence that could be harmful to human health.

Isolation, identification, and characterization of thermophilic lactic acid bacteria isolated from whey of Kars Kashar cheeses.

Kars Kashar cheese is an artisanal pasta-filata type cheese and geographically marked in Eastern Anatolia of Turkey. The aims of this research were to determine for the first time thermophilic lactic acid bacteria (LAB) of Kars Kashar cheese and characterize the technological properties of obtained isolates. In our research, a number of 15 samples of whey were collected from the different villages in Kars. These samples were incubated at 45 °C and used as the source material for isolating thermophilic LAB. A total of 250 colonies were isolated from thermophilic whey, and 217 of them were determined to be presumptive LAB based on their Gram staining and catalase test. A total of 170 isolates were characterized by their phenotypic properties and identified using the MALDI-TOF mass spectrometry method. Phenotypic identification of isolates displayed that Enterococcus and Lactobacillus were the predominant microbiota. According to MALDI-TOF MS identification, 89 isolates were identified as Enterococcus (52.35%), 57 isolates as Lactobacillus (33.53%), 23 isolates as Streptococcus (13.53%), and one isolate as Lactococcus (0.59%). All thermophilic LAB isolates were successfully identified to the species level and it has been observed that MALDI-TOF MS can be successfully used for the identification of selected LAB. The acidification and proteolytic activities of the isolated thermophilic LAB were examined, and the isolates designated for use as starter cultures were also genotypically defined.

Detection and modeling of Staphylococcus aureus and fecal bacteria in Hawaiian coastal waters and sands.

Microbial pollution of recreational waters leads to millions of skin, respiratory, and gastrointestinal illnesses globally. Fecal indicator bacteria (FIB) are monitored to assess recreational waters but may not reflect the presence of Staphylococcus aureus, a global leader in bacterial fatalities. Since many community-acquired S. aureus skin infections are associated with high recreational water usage, this study measured and modeled S. aureus, methicillin-resistant S. aureus (MRSA), and FIB (Enterococcus spp., Clostridium perfringens) concentrations in seawater and sand at six beaches in Hilo, Hawai'i, USA, over 37 sample dates from July 2016 to February 2019 using culturing techniques. Generalized linear models predicted bacterial concentrations with physicochemical and environmental data. Beach visitors were also surveyed on their preferred activities. S. aureus and FIB concentrations were roughly 6-78 times higher at beaches with freshwater discharge than at those without. Seawater concentrations of Enterococcus spp. were positively associated with MRSA but not S. aureus. Elevated S. aureus was associated with lower tidal heights, higher freshwater discharge, onsite sewage disposal system density, and turbidity. Regular monitoring of beaches with freshwater input, utilizing real-time water quality measurements with robust modeling techniques, and raising awareness among recreational water users may mitigate exposure to S. aureus, MRSA, and FIB. PRACTITIONER POINTS: Staphylococcus aureus and fecal bacteria concentrations were higher in seawater and sand at beaches with freshwater discharge. In seawater, Enterococcus spp. positively correlated with MRSA, but not S. aureus. Freshwater discharge, OSDS density, water turbidity, and tides significantly predicted bacterial concentrations in seawater and sand. Predictive bacterial models based upon physicochemical and environmental data developed in this study are readily available for user-friendly application.

Spatio-Temporal Variation in the Exceedance of Enterococci in Lake Burley Griffin: An Analysis of 16 Years' Recreational Water Quality Monitoring Data.

Recreational waterbodies with high levels of faecal indicator bacteria (FIB) pose health risks and are an ongoing challenge for urban-lake managers. Lake Burley Griffin (LBG) in the Australian Capital city of Canberra is a popular site for water-based recreation, but analyses of seasonal and long-term patterns in enterococci that exceed alert levels (>200 CFU per 100 mL, leading to site closures) are lacking. This study analysed enterococci concentrations from seven recreational sites from 2001-2021 to examine spatial and temporal patterns in exceedances during the swimming season (October-April), when exposure is highest. The enterococci concentrations varied significantly across sites and in the summer months. The frequency of the exceedances was higher in the 2009-2015 period than in the 2001-2005 and 2015-2021 periods. The odds of alert-level concentrations were greater in November, December, and February compared to October. The odds of exceedance were higher at the Weston Park East site (swimming beach) and lower at the Ferry Terminal and Weston Park West site compared to the East Basin site. This preliminary examination highlights the need for site-specific assessments of environmental and management-related factors that may impact the public health risks of using the lake, such as inflows, turbidity, and climatic conditions. The insights from this study confirm the need for targeted monitoring efforts during high-risk months and at specific sites. The study also advocates for implementing measures to minimise faecal pollution at its sources.

The risk of viable but non-culturable (VBNC) enterococci and antibiotic resistance transmission during simulated municipal sludge composting.

Sludge composting is a sludge resource utilization method that can reduce pollutants, such as pathogens. Enterococci are regarded as more reliable and conservative indicators of pathogen inactivation than fecal coliforms, which are typically used as indicators of fecal pollution. Non-spore pathogenic bacteria may enter a viable but non-culturable (VBNC) state during composting, leading to residual risk. The VBNC status of bacteria is related to their survival during composting. However, the survival mechanisms of enterococci during sludge composting remain unclear. Therefore, this study aimed to investigate the VBNC state of enterococci in different phases of simulated sludge composting and the fate of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) during the composting process. This study is expected to provide a basis for subsequent exploration of possible methods to completely inactivate enterococci and reduce ARGs during sludge composting. Culturable enterococci were reduced in the thermophilic phase of sludge composting, but the proportion of VBNC subpopulation increased. It was reported for the first time that most VBNC enterococci were killed by extending the cooling phase of sludge compost, and by prolonging the cooling phase the types of ARG were reduced. However, there was a certain quantity (approximately 104/g dry weight) of culturable and VBNC enterococci in the compost products. In addition, MGEs and ARGs exist in both bacteria and compost products, leading to the risk of spreading antibiotic-resistant bacteria and antibiotic resistance when sludge compost products are used.

Both biofilm cytotoxicity and monocytes' adhesion may be used as estimators of enterococcal virulence.

Our study aimed to identify markers of enterococci's virulence potential by evaluating the properties of strains of different sites of isolation. Enterococcal strains were isolated as commensals from faeces and as invasive strains from the urine and blood of patients from the University Clinical Centre, Gdansk, Poland. Changes in monocytes' susceptibility to the cytotoxic activity of isolates of different origins and their adherence to biofilm were evaluated using a flow cytometer. The bacterial protein profile was estimated by matrix assisted laser desorption ionization-time of flight mass spectrometer. The cytotoxicity of biofilm and monocytes' adherence to it were the most accurate factors in predicting the prevalence of the strain in the specific niche. Additionally, a bacterial protein with mass-to-charge ratio (m/z) 5000 was found to be responsible for the increased bacterial cytotoxicity, while monocytes' decreased adherence to biofilm was linked with the presence of proteins either with m/z 3330 or 2435. The results illustrate that monocytes' reaction when exposed to the bacterial biofilm can be used as an estimator of pathogens' virulence potential. The observed differences in monocytes' response are explainable by the bacterial proteins' profile. Additionally, the results indicate that the features of both bacteria and monocytes impact the outcome of the infection.

In Vitro Evaluation of Phytobiotic Mixture Antibacterial Potential against Enterococcus spp. Strains Isolated from Broiler Chicken.

Enterococcus spp. are normal intestinal tract microflorae found in poultry. However, the last decades have shown that several species, e.g., Enterococcus cecorum, have become emerging pathogens in broilers and may cause numerous losses in flocks. In this study, two combinations (H1 and H2) of menthol, 1,8-cineol, linalool, methyl salicylate, γ-terpinene, p-cymene, trans-anethole, terpinen-4-ol and thymol were used in an in vitro model, analyzing its effectiveness against the strains E. cecorum, E. faecalis, E. faecium, E. hirae and E. gallinarum isolated from broiler chickens from industrial farms. To identify the isolated strains classical microbiological methods and VITEK 2 GP cards were used. Moreover for E. cecorum a PCR test was used.. Antibiotic sensitivity (MIC) tests were performed for all the strains. For the composition H1, the effective dilution for E. cecorum and E. hirae strains was 1:512, and for E. faecalis, E. faecium and E. gallinarum, 1:1024. The second mixture (H2) showed very similar results with an effectiveness at 1:512 for E. cecorum and E. hirae and 1:1024 for E. faecalis, E. faecium and E. gallinarum. The presented results suggest that the proposed composition is effective against selected strains of Enterococcus in an in vitro model, and its effect is comparable to classical antibiotics used to treat this pathogen in poultry. This may suggest that this product may also be effective in vivo and provide effective support in the management of enterococcosis in broiler chickens.

Enterococcus-derived tyramine hijacks α2A-adrenergic receptor in intestinal stem cells to exacerbate colitis.

Inflammatory bowel disease (IBD) is characterized by dysbiosis of the gut microbiota and dysfunction of intestinal stem cells (ISCs). However, the direct interactions between IBD microbial factors and ISCs are undescribed. Here, we identify α2A-adrenergic receptor (ADRA2A) as a highly expressed GPCR in ISCs. Through PRESTO-Tango screening, we demonstrate that tyramine, primarily produced by Enterococcus via tyrosine decarboxylase (tyrDC), serves as a microbial ligand for ADRA2A. Using an engineered tyrDC-deficient Enterococcus faecalis strain and intestinal epithelial cell-specific Adra2a knockout mice, we show that Enterococcus-derived tyramine suppresses ISC proliferation, thereby impairing epithelial regeneration and exacerbating DSS-induced colitis through ADRA2A. Importantly, blocking the axis with an ADRA2A antagonist, yohimbine, disrupts tyramine-mediated suppression on ISCs and alleviates colitis. Our findings highlight a microbial ligand-GPCR pair in ISCs, revealing a causal link between microbial regulation of ISCs and colitis exacerbation and yielding a targeted therapeutic approach to restore ISC function in colitis.

Genomic epidemiology reveals multiple mechanisms of linezolid resistance in clinical enterococci in China.

BACKGROUND: Infections caused by linezolid-resistant enterococci (LRE) are clinically difficult to treat and threaten patient health. However, there is a lack of studies on long time-span LRE strains in China. For this reason, our study comprehensively revealed the resistance mechanisms of LRE strains collected in a Chinese tertiary care hospital from 2011 to 2022. METHODS: Enterococcal strains were screened and verified after retrospective analysis of microbial data. Subsequently, 65 LRE strains (61 Enterococcus faecalis and 4 Enterococcus faecium, MIC ≥ 8 µg/ml), 1 linezolid-intermediate Enterococcus faecium (MIC = 4 µg/ml) and 1 linezolid-susceptible Enterococcus faecium (MIC = 1.5 µg/ml) were submitted for whole-genome sequencing (WGS) analysis and bioinformatics analysis. RESULTS: The optrA gene was found to be the most common linezolid resistance mechanism in our study. We identified the wild-type OptrA and various OptrA variants in 98.5% of LRE strains (61 Enterococcus faecalis and 3 Enterococcus faecium). We also found one linezolid-resistant Enterococcus faecium strain carried both optrA and cfr(D) gene, while one linezolid-resistant Enterococcus faecium only harbored the poxtA gene. Most optrA genes (55/64) were located on plasmids, with impB-fexA-optrA, impB-fexA-optrA-erm(A), fexA-optrA-erm(A), and fexA-optrA segments. A minority of optrA genes (9/64) were found on chromosomes with the Tn6674-like platform. Besides, other possible linezolid resistance-associated mechanisms (mutations in the rplC and rplD genes) were also found in 26 enterococcal strains. CONCLUSIONS: Our study suggested that multiple mechanisms of linezolid resistance exist among clinical LRE strains in China.

Spontaneous resolution or antibiotic effectiveness? Reflection on a case of pediatric urinary tract infections caused by Enterococcus raffinosus.

Enterococcus raffinosus, named by Collins et al. in 1989, is a cocci-shaped bacterium that typically appears in pairs or short chains. As a Gram-positive and non-motile bacterium, it grows at 10°C-45°C, exhibiting negative peroxidase activity [1]. It is a normal flora in the oropharynx and gastrointestinal tract of domestic cats [2] and can also be isolated from human rectal swabs [3], it belongs to the same genus Enterococcus as Enterococcus faecalis and Enterococcus faecium. Enterococcus faecalis and Enterococcus faecium constitute 90% of clinically isolated strains. However, the incidence of other enterococci, excluding E. faecalis and E. faecium, is on the rise [4]. In this case report, a patient with pediatric urinary tract infections caused by E. raffinosus was presented, and a summary of relevant literature was provided.

Antimicrobial resistance pattern of Uro-Pathogens emphasizing non-lactose fermenting gram negative bacilli.

Objectives: To identify various species of non-lactose fermenting gram-negative bacilli involved in urinary tract infections, and to determine their antimicrobial resistance pattern. METHODS: The retrospective, descriptive, cross-sectional study was conducted from January 1 to April 1, 2022, at the Dow University of Health Sciences, Karachi, and comprised data from the institutional diagnostic laboratory that was related to urine samples regardless of age and gender from January 1, 2020, to December 31, 2021. Data was analysed using SPSS version 25. RESULTS: Of the 103,887 urine samples, 41,280(39.7%) were positive, 51,146(49.2%) showed no bacterial growth, 11,000(10.6%) had non-significant bacterial growth and 461(0.4%) had mixed bacterial growth. Of the positive samples, 18359(44.5%) were positive in 2020, and 22,921(55.5%) in 2021. Gram-negative lactose fermenting bacteria included escherichia coli 23,123(22.3%) and klebsiella pneumoniae 2,993(2.9%), gram-negative non-lactose fermenting bacteria included pseudomonas aeruginosa 1,110(1.07%), and gram-positive bacteria included enterococcus 8,008(7.7%). Pseudomonas aeruginosa was most resistant against tobramycin 880(79.3%) and least resistant against piperacillin-tazobactam 146(13%). CONCLUSIONS: Piperacillin-tazobactam was highly sensitive drug against non-lactose fermenting uro-pathogens.

Challenges for heat stress: Intestinal culturable bacteria of Lohmann Brown chickens.

This study aimed to assess how heat stress, specifically within the range of 35-38 °C, affects the populations of culturable intestinal lactobacilli, enterococci, and Escherichia coli, as well as the expression of Heat Shock Proteins (HSP70), in Lohmann Brown chickens. It also explored the influence of the chickens' blood transferrin and ceruloplasmin genotypes on these responses. Thirty chickens underwent eight hours of heat stress, maintained at an average temperature of 37 °C and a relative humidity of 75-80%, with continuous access to food and water. Behavioral monitoring was conducted throughout to prevent excessive heat-related mortality. The Lohmann Brown chickens from the Yerevan "Arax" poultry farm were initially classified based on their blood transferrin and ceruloplasmin genotypes to investigate potential correlations between intestinal bacterial composition and variations in these polymorphisms. A significant correlation was found between heat stress and the abundance of culturable enterococci within the intestinal microbiota, regardless of chicken TfAB, TfBC, CpAB, CpCC and TfAB, TfBC, CpAB, CpCD genotypes. Heat stress led to nearly double the HSP70 levels in chicken blood, along with a reduction in the culturable enterococci population by at least 10,000-fold in the intestinal microbiota. These findings are significant for targeted management strategies to mitigate heat stress in chicken populations.

An analysis of culture-based methods used for the detection and isolation of Salmonella spp., Escherichia coli, and Enterococcus spp. from surface water: A systematic review.

Identification of methods for the standardized assessment of bacterial pathogens and antimicrobial resistance (AMR) in environmental water can improve the quality of monitoring and data collected, support global surveillance efforts, and enhance the understanding of environmental water sources. We conducted a systematic review to assemble and synthesize available literature that identified methods for assessment of prevalence and abundance of bacterial fecal indicators and pathogens in water for the purposes of monitoring bacterial pathogens and AMR. After screening for quality, 175 unique publications were identified from 15 databases, and data were extracted for analysis. This review identifies the most common and robust methods, and media used to isolate target organisms from surface water sources, summarizes methodological trends, and recognizes knowledge gaps. The information presented in this review will be useful when establishing standardized methods for monitoring bacterial pathogens and AMR in water in the United States and globally.

Dissemination, virulence characteristic, antibiotic resistance determinants of emerging linezolid and vancomycin-resistant Enterococcus spp. in fish and crustacean.

Enterococci are emerging nosocomial pathogens. Their widespread distribution causes them to be food contaminants. Furthermore, Enterococci can colonize various ecological niches and diffuse into the food chain via contaminated animals and foods because of their remarkable tolerance to unfavorable environmental circumstances. Due to their potential dissemination to humans, antimicrobial-resistant Enterococci in fish are a worldwide health issue. This study characterized AMR, ARGs, VAGs, gelatinase activity, and biofilm formation in Enterococcus spp. recovered from fish and seafood and evaluated potential correlations. 54 Enterococcus spp. strains(32.73 %)were isolated from 165 samples (75 Oreochromis niloticus, 30 Argyrosomus regius, and 60 Shrimp), comprising 30 Enterococcus faecalis (55.6 %) and 24 Enterococcus faecium (44.4 %) with total 32.73 % (54/165), The maximum prevalence rate of Enterococcus spp. was observed in Nile tilapia (34/54; 63 %), followed by shrimp (14/54; 25.9 %) and Argyrosomus regius (6/54; 11.1 %). The maximum prevalence rate of E. faecalis was observed in Nile tilapia (22/30; 73.3 %), followed by shrimp (8/30; 26.7 %) with significant differences. The prevalence rate of E. faecium was observed in Nile tilapia (12/24; 50 %), followed by shrimp (6/24,25 %). E. faecium is only isolated from Argyrosomus regius (6/24,25 %). Isolates exhibited high resistance against both tetracycline (90.7 %) and erythromycin(88.9 %), followed by gentamycin (77.8 %), ciprofloxacin (74.1 %), levofloxacin (72.2 %), penicillin (44.4 %), vancomycin (37 %), and linezolid (20.4 %). 50 strains (92.6 %) exhibited resistance to more than two antibiotics, 5 strains (10 %) were XDR, and the remaining 45 strains (90 %) were classified as MDR. 92.6 % of the isolates had MARindices >0.2, indicating they originated in settings with a high risk of contamination. Additionally, ten ARGs were identified, with tet(M) 92.6 %, followed by erm(B) (88.9 %), aac(6')-Ie-aph(2″)-Ia(77.8 %), tet(K) (75.9 %), gyrA (74.1 %), blaZ (48.1 %), vanA (37 %), vanB (31.5 %), optrA (20.4 %), and catA(3.7 %). Biofilm formation and gelatinase activity were observed in 85.2 %, and 61.1 % of the isolates, respectively. A total of 11 VAGs were detected, with gelE as the most prevalent (83.3 %) followed by agg(79.6 %), pil (74.1 %), both sprE and asa1 (72.2 %), hyl (70.4 %), eps(68.5 %), EF3314 (57.4 %), ace (50 %), and cylA (35.2 %) with no detection of cylB. In conclusion, the emergence of linezolid-resistant -vancomycin-resistant enterococci recovered from Egyptian fish and shrimp, suggests that fish and seafood might participate a fundamental part in the emergence of antimicrobial resistance among humans.

The dysregulation of biliary tract microflora is closely related to primary choledocholithiasis: a multicenter study.

Bile microecology changes play an important role in the occurrence and development of choledocholithiasis. At present, there is no clear report on the difference of bile microecology between asymptomatic patients with gallbladder polyps and choledocholithiasis. This study compared bile microecology between gallbladder polyp patients and patients with choledocholithiasis to identify risk factors for primary choledocholithiasis. This study was conducted in 3 hospitals in different regions of China. Bile samples from 26 patients with gallbladder polyps and 31 patients with choledocholithiasis were collected by laparoscopic cholecystectomy and endoscopic retrograde choledocholithiasis cholangiography (ERCP), respectively. The collected samples were used for 16S ribosomal RNA sequencing and liquid chromatography mass spectrometry analysis. The α-diversity of bile microecological colonies was similar between gallbladder polyp and choledocholithiasis, but the ß-diversity was different. Firmicutes, Proteobacteri, Bacteroidota and Actinobacteriota are the most common phyla in the gallbladder polyp group and choledocholithiasis group. However, compared with the gallbladder polyp patients, the abundance of Actinobacteriota has significantly lower in the choledocholithiasis group. At the genera level, the abundance of a variety of bacteria varies between the two groups, and Enterococcus was significantly elevated in choledocholithiasis group. In addition, bile biofilm formation-Pseudomonas aeruginosa was more metabolically active in the choledocholithiasis group, which was closely related to stone formation. The analysis of metabolites showed that a variety of metabolites decreased in the choledocholithiasis group, and the concentration of beta-muricholic acid decreased most significantly. For the first time, our study compared the bile of gallbladder polyp patients with patients with choledocholithiasis, and suggested that the change in the abundance of Actinobacteriota and Enterococcus were closely related to choledocholithiasis. The role of Pseudomonas aeruginosa biofilm in the formation of choledocholithiasis was discovered for the first time, and some prevention schemes for choledocholithiasis were discussed, which has important biological and medical significance.

Identification and biological characteristics of Enterococcus casseliflavus TN-47 isolated from Monochamus alternatus.

With the widespread use of antibiotics, the incidence of antibiotic resistance in microorganisms has increased. Monochamus alternatus is a trunk borer of pine trees. This study aimed to investigate the in vitro antimicrobial and biological characteristics of Enterococcus casseliflavus TN-47 (PP411196), isolated from the gastrointestinal tract of M. alternatus in Jilin Province, PR China. Among 13 isolates obtained from the insects, five were preliminarily screened for antimicrobial activity. E. casseliflavus TN-47, which exhibited the strongest antimicrobial activity, was identified. E. casseliflavus TN-47 possessed antimicrobial activity against Staphylococcus aureus USA300 and Salmonella enterica serovar Pullorum ATCC 19945. Furthermore, E. casseliflavus TN-47 was sensitive to tetracyclines, penicillins (ampicillin, carbenicillin, and piperacillin), quinolones and nitrofuran antibiotics, and resistant to certain beta-lactam antibiotics (oxacillin, cefradine and cephalexin), macrolide antibiotics, sulfonamides and aminoglycosides. E. casseliflavus TN-47 could tolerate low pH and pepsin-rich conditions in the stomach and grow in the presence of bile acids. E. casseliflavus TN-47 retained its strong auto-aggregating ability and hydrophobicity. This strain did not exhibit any haemolytic activity. These results indicate that E. casseliflavus TN-47 has potential as a probiotic. This study provides a theoretical foundation for the future applications of E. casseliflavus TN-47 and its secondary metabolites in animal nutrition and feed.

Investigation of some changes and clonal relationship in enterococci isolates due to relocation of a hospital.

Objectives: To investigate the isolation rates, antimicrobial resistance rates, minimum inhibitory concentration values of antimicrobial agents, and clonal relationships of Enterococcus faecalis and Enterococcus faeciumdue to the relocation of a hospital to a newly constructed building. METHODS: The comparative, prospective study was conducted at adult general intensive care units of the Mus State Hospital, Mus, Turkey, in two phases; before the relocation from January 25 to December 1, 2014, and after the relocation from February 10 to May 24, 2015. Rectal swab samples were collected 72 hours post-hospitalisation. Identification of Enterococcus faecalis and Enterococcus faeciumisolates was determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and antimicrobial resistance with minimum inhibitory concentration values was detected with Vitek 2 system. The clonal relatedness among the strains was investigated by pulsed-field gel electrophoresis. Data was analysed using SPSS 23. RESULTS: Of the 69 patients, 37(53.62%) were related to pre-relocation phase; 20(54.1%) females and 17(45.9%) males with mean age 62.81±21.71 years. There were 32(46.37%) patients in the post-relocation phase; 13(40.6%) females and 19(59.4%) males with mean age 62.69±21.35 years (p>0.05). Of the 84 enterococci strains isolated, 51(60.7%) were Enterococcus faecium; 28(55%) before relocation and 23(45%) after relocation (p=0.77). The remaining 33(39.3%) isolates were Enterococcus faecalis; 16(48.5%) before relocation and 17(51.5%) after relocation (p=0.73). Multiple strains were located in 7(18.9%) patients before relocation and in 7(21.9%) after relocation. In 1(3.1%) patient after relocation, 2(8.7%) Enterococcus faecium isolates with different resistance and pulsed-field gel electrophoresis patterns were detected. There were no significant differences between the isolation and antibiotic resistance rates before and after relocation (p>0.05), and a clonal relation between the isolates was not detected (p>0.05). Decreased minimum inhibitory concentration values were noted for some antibiotics. CONCLUSIONS: Clonal relationship between the isolates and change in the rates of isolation and antimicrobial resistance of Enterococcus faecalis and Enterococcus faecium was not detected due to relocation. Minimum inhibitory concentration values could be used to reveal relocation-related changes in isolates obtained from patients hospitalised in intensive care units.

Comparative analysis of proteomic adaptations in Enterococcus faecalis and Enterococcus faecium after long term bile acid exposure.

BACKGROUND: All gastrointestinal pathogens, including Enterococcus faecalis and Enterococcus faecium, undergo adaptation processes during colonization and infection. In this study, we investigated by data-independent acquisition mass spectrometry (DIA-MS) two crucial adaptations of these two Enterococcus species at the proteome level. Firstly, we examined the adjustments to cope with bile acid concentrations at 0.05% that the pathogens encounter during a potential gallbladder infection. Therefore, we chose the primary bile acids cholic acid (CA) and chenodeoxycholic acid (CDCA) as well as the secondary bile acid deoxycholic acid (DCA), as these are the most prominent bile acids. Secondly, we investigated the adaptations from an aerobic to a microaerophilic environment, as encountered after oral-fecal infection, in the absence and presence of deoxycholic acid (DCA). RESULTS: Our findings showed similarities, but also species-specific variations in the response to the different bile acids. Both Enterococcus species showed an IC50 in the range of 0.01- 0.023% for DCA and CDCA in growth experiments and both species were resistant towards 0.05% CA. DCA and CDCA had a strong effect on down-expression of proteins involved in translation, transcription and replication in E. faecalis (424 down-expressed proteins with DCA, 376 down-expressed proteins with CDCA) and in E. faecium (362 down-expressed proteins with DCA, 391 down-expressed proteins with CDCA). Proteins commonly significantly altered in their expression in all bile acid treated samples were identified for both species and represent a "general bile acid response". Among these, various subunits of a V-type ATPase, different ABC-transporters, multi-drug transporters and proteins related to cell wall biogenesis were up-expressed in both species and thus seem to play an essential role in bile acid resistance. Most of the differentially expressed proteins were also identified when E. faecalis was incubated with low levels of DCA at microaerophilic conditions instead of aerobic conditions, indicating that adaptations to bile acids and to a microaerophilic atmosphere can occur simultaneously. CONCLUSIONS: Overall, these findings provide a detailed insight into the proteomic stress response of two Enterococcus species and help to understand the resistance potential and the stress-coping mechanisms of these important gastrointestinal bacteria.