Search Results (287)

This review explores the biofilm architecture and drug resistance of Enterococcus faecalis in clinical and environmental settings. The biofilm in E. faecalis is a heterogeneous, three-dimensional, mushroom-like or multilayered structure, characteristically forming diplococci or short chains interspersed with water channels for nutrient exchange and waste removal. Exopolysaccharides, proteins, lipids, and extracellular DNA create a protective matrix. Persister cells within the biofilm contribute to antibiotic resistance and survival. The heterogeneous architecture of the E. faecalis biofilm contains both dense clusters and loosely packed regions that vary in thickness, ranging from 10 to 100 µm, depending on the environmental conditions. The pathogenicity of the E. faecalis biofilm is mediated through complex interactions between genes and virulence factors such as DNA release, cytolysin, pili, secreted antigen A, and microbial surface components that recognize adhesive matrix molecules, often involving a key protein called enterococcal surface protein (Esp). Clinically, it is implicated in a range of nosocomial infections, including urinary tract infections, endocarditis, and surgical wound infections. The biofilm serves as a nidus for bacterial dissemination and as a reservoir for antimicrobial resistance. The effectiveness of first-line antibiotics (ampicillin, vancomycin, and aminoglycosides) is diminished due to reduced penetration, altered metabolism, increased tolerance, and intrinsic and acquired resistance. Alternative strategies for biofilm disruption, such as combination therapy (ampicillin with aminoglycosides), as well as newer approaches, including antimicrobial peptides, quorum-sensing inhibitors, and biofilm-disrupting agents (DNase or dispersin B), are also being explored to improve treatment outcomes. Environmentally, E. faecalis biofilms contribute to contamination in water systems, food production facilities, and healthcare environments. They persist in harsh conditions, facilitating the spread of multidrug-resistant strains and increasing the risk of transmission to humans and animals. Therefore, understanding the biofilm architecture and drug resistance is essential for developing effective strategies to mitigate their clinical and environmental impact. Full article

Background/Objectives: Enterococci, particularly Enterococcus faecalis and Enterococcus faecium, are Gram-positive cocci that can cause severe infections in hospitalized patients. The rise of vancomycin-resistant enterococci (VRE) and vancomycin-variable enterococci (VVE) poses significant challenges in healthcare settings due to their resistance to multiple antibiotics. Methods: We conducted a point prevalence survey (PPS) to assess the prevalence of VRE and VVE colonization in hospitalized patients. Rectal swabs were collected from 160 patients and analyzed using molecular assays (MAs) and culture. Whole-genome sequencing (WGS) and core-genome multilocus sequence typing (cgMLST) were performed to identify the genetic diversity. Results: Of the 160 rectal swabs collected, 54 (33.7%) tested positive for the vanA and/or vanB genes. Culture-based methods identified 47 positive samples (29.3%); of these, 44 isolates were identified as E. faecium and 3 as E. faecalis. Based on the resistance profiles, 35 isolates (74.5%) were classified as VRE, while 12 (25.5%) were classified as VVE. WGS and cgMLST analyses identified seven clusters of E. faecium, with sequence type (ST) 80 being the most prevalent. Various resistance genes and virulence factors were identified, and this study also highlighted intra- and inter-ward transmission of VRE strains. Conclusions: Our findings underscore the potential for virulence and resistance of both the VRE and VVE strains, and they highlight the importance of effective infection control measures to prevent their spread. VVE in particular should be carefully monitored as they often escape detection. Integrating molecular data with clinical information will hopefully enhance our ability to predict and prevent future VRE infections. Full article

Background: Antimicrobial resistance (AMR) is increasingly acknowledged as a critical global challenge, posing serious risks to human and animal health and potentially disrupting poultry production systems. Commensal bacteria such as Staphylococcus spp., Enterococcus spp., and Escherichia coli may serve as important reservoirs and vectors of resistance genes. Objectives: This study aimed to assess the AMR profiles of bacterial strains isolated from industrial chicken farms in the Dél-Alföld region of Hungary, providing region-specific insights into resistance dynamics. Methods: A total of 145 isolates, including Staphylococcus spp., Enterococcus spp., and E. coli isolates, were subjected to minimum inhibitory concentration (MIC) testing against 15 antimicrobial agents, following Clinical and Laboratory Standards Institute (CLSI) guidelines. Advanced multivariate statistics, machine learning algorithms, and network-based approaches were employed to analyze resistance patterns and co-resistance associations. Results Multidrug resistance (MDR) was identified in 43.9% of Staphylococcus spp. isolates, 28.8% of Enterococcus spp. isolates, and 75.6% of E. coli isolates. High levels of resistance to florfenicol, enrofloxacin, and potentiated sulfonamides were observed, whereas susceptibility to critical antimicrobials such as imipenem and vancomycin remained largely preserved. Discussion: Our findings underscore the necessity of implementing region-specific AMR monitoring programs and strengthening multidisciplinary collaboration within the “One Health” framework with proper animal hygiene and biosecurity measures to limit the spread of antimicrobial resistance and protect both animal and human health. Full article

Background/Objectives: European hedgehogs (Erinaceus europaeus) are present in areas where there is human activity; therefore, they can be a source of pathogens for other animals and humans. Methods: Eighteen hedgehog carcasses were collected and analyzed for Staphylococcus spp. Isolated strains were typed and analyzed for exfoliative toxins genes and the phenotypic and genotypic characteristics of antimicrobial resistance. Results: A total of 54 strains were isolated and typed as S. aureus, S. xylosus, S. sciuri, S. pseudintermedius, S. simulans, S. chromogenes, S. epidermidis, S. hyicus, and S. lentus. No strains had the eta and etb genes coding for exfoliative toxins. Overall, 39/54 (72.20%) isolates showed phenotypic resistance to at least one antimicrobial and 21/54 (38.80%) showed more than one resistance. The lowest efficacy was observed for erythromycin, with 40/54 (74.08%) strains classified as intermediate and 6/54 (11.11%) classified as resistant. Among the 29 isolates shown to be penicillin-resistant, 11 (37.93%) were oxacillin-resistant, with a minimum inhibitory concentration (MIC). Among the 54 staphylococcal strains, 2 (3.70%) were resistant to vancomycin, both with an MIC value equal to the maximum concentration of the antibiotic tested (256 μg/mL) and 2 (3.70%) had an intermediate resistance profile with an 8 μg/mL MIC value. No strains had the genes vanA and vanB. Two of the 29 (6.90%) penicillin-resistant strains had the blaZ gene; 8 (27.13%) strains had the mecA gene. Overall, 2/54 (3.70%) isolates were classified as extensively drug-resistant (XDR) and 9/54 (16.66%) were classified as multidrug-resistant (MDR). Conclusions: Hedgehogs can harbor antimicrobial-resistant staphylococci and can be sources of these bacteria for other animals and humans. They can also serve as bioindicators of the pathogens and antimicrobial-resistant bacteria circulating in a given habitat. Full article

Cutaneous adverse reactions (CARs) are common complications of epidermal growth factor receptor (EGFR) inhibitor therapy, with papulopustular eruptions and paronychia being the most frequent. Growing scientific evidence implies that Staphylococcus aureus is involved in the pathogenesis of these reactions. This observational prospective study characterized 42 S. aureus strains isolated from CARs, analyzing antibiotic resistance, biofilm formation, soluble virulence factors, and virulence/resistance genes using multiplex polymerase chain reaction (PCR). S. aureus was identified in 90% of lesions; in 33% of cases, nasal and skin isolates were genetically identical. High resistance rates were noted for penicillins (85%) and tetracyclines (57%), while all strains remained susceptible to fluoroquinolones, vancomycin, and rifampicin. All isolates formed biofilms, and DNase/esculinase production significantly correlated with CAR severity. An enzymatic score based on these markers was associated with an 18-fold increased risk of severe reactions. Genotypically, clfA and clfB were prevalent (85.7%), while exotoxin genes were less common. These findings support a key role for S. aureus in exacerbating CARs via antibiotic resistance, biofilm production, and the expression of virulence factor. Additionally, we emphasize the role of routine microbial screening—including nasal swabs—and therapy guided by antibiograms. Furthermore, the enzymatic score may further be validated as a predictive biomarker. Full article

Clostridioides difficile is a leading pathogen involved in healthcare-associated diarrhea. With its increasing incidence, mortality, and antibiotic resistance, there is an urgent need for novel therapeutic strategies to address the infection and prevent its recurrence. Gegen Qinlian Decoction (GQD) is a traditional Chinese medicine for the treatment of diarrhea, but its main active ingredient is not known. Therefore, in this study, we evaluated the biological activity of berberine (BER) and baicalein (BAI), key components of GQD, against C. difficile. Time–kill curves and scanning electron microscopy were employed to assess their effects on C. difficile growth, while Enzyme-Linked Immunosorbnent Assay (ELISA) and cytotoxicity assays were used to examine their impact on toxin production. We also employed Quantitative Reverse Transcription PCR (qRT-PCR) to examine how BER and BAI influenced the expression of toxin-associated genes. At sub-inhibitory concentrations, these compounds exerted antibacterial activity against C. difficile by disrupting the integrity of the cell membrane and cell wall. Furthermore, BER and BAI also suppressed toxin production, demonstrating effects comparable to those of vancomycin. This suppression likely resulted from their bactericidal activity and the inhibition of toxin gene expression. This study not only highlights the potential application of GQD in treating C. difficile infections but also offers promising options for developing drugs targeting the growth and virulence of this pathogen. C. difficile infection (CDI) is a leading cause of severe diarrhea, and its treatment remains challenging due to limited drug options and its high recurrence rate. BAI and BER, the main active components of the traditional Chinese medicinal formula GQD, inhibited the growth of C. difficile by disrupting its cellular structure and significantly reduced the production of toxins associated with disease severity. Furthermore, the effects of BAI and BER on C. difficile were comparable to those of conventional antibiotics, suggesting that these compounds could be potential alternative therapies for CDI. This study not only highlights the therapeutic potential of GQD in treating CDI but also provides a replicable research strategy for the development of novel anti-CDI agents. Full article

In this study, we isolated and identified bacteria from the feces of a diarrheal cynomolgus monkey. The results showed that the isolated strain was P. aeruginosa, named PA/CM-101101. Morphological observations indicated that when cultured on Luria–Bertani (LB) nutrient agar at 37 °C for 24 h, the strain formed smooth, slightly elevated colonies with neat and wavy edges. On acetamide agar at the same temperature and duration, the colonies appeared flat with irregular edges and a faint pink periphery, while the medium changed to rose-red; in LB broth at 37 °C for 24 h, the medium became turbid and yellowish-green. Gram staining revealed that it was negative and rod-shaped, without sporulation characteristics. The 16S rRNA gene sequence analysis showed that the sequence identity of the strain shared more than 98.4% similarity with 11 strains of P. aeruginosa from various sources in GenBank. The animal toxicity test showed that it had a strong pathogenic effect on mice. The results of drug sensitivity tests showed that strain PA/CM-101101 was sensitive to amikacin, azithromycin, cefoperazone, ceftazidime, ceftriaxone, ciprofloxacin, gentamicin, imipenem, levofloxacin, meropenem, norfloxacin, ofloxacin, and polymyxin B; however, it displayed resistance to ampicillin, cefadroxil, cefazolin, erythromycin, and vancomycin. The research findings provide valuable insights for diagnosis and treatment strategies for cynomolgus monkeys. It also provides a reference for molecular epidemiological studies. To our knowledge, this is the first time P. aeruginosa isolated from the diarrhea feces of cynomolgus monkey has been reported. Full article

Background: Antimicrobial resistance (AMR) is a growing global concern in poultry production, where antibiotic use can disrupt gut microbiota and enrich antimicrobial resistance genes (ARGs). Objectives: This study aimed to assess the in vivo effects of a veterinary-approved amoxicillin formulation on gut microbiome composition and ARG profiles in broiler chickens. Methods: A total of 120 Ross-308 broiler chickens were randomly allocated into 12 experimental groups (n = 10 per group), with three replicates per treatment. Birds received either full-dose (1×), a subtherapeutic quarter-dose (¼×) of amoxicillin, a placebo (starch), or no treatment. Cloacal swabs were collected on days 0, 14, and 28 for shotgun metagenomic sequencing. One-way ANOVA was used to evaluate treatment effects on body weight, with significant differences observed from day 14 onward (p < 0.0001). Results: The ¼× dose caused a more pronounced microbiome shift than the 1× dose, with a marked reduction in Pseudomonadota and increase in Bacillota and Bacteroidota. ARG abundance declined in the ¼× group (from 1386 to 1012). While TEM-type ESBL genes were ubiquitous, CTX-M-1 emerged only after ¼× treatment. Worryingly, 20 types of vancomycin resistance genes were detected across all samples. Plasmid-borne ARGs and mobile genetic elements decreased in the ¼× group. Conclusions: Even subtherapeutic antibiotic exposure significantly reshapes the gut microbiota composition and ARG landscape, highlighting the need for refined risk assessments and microbiome-conscious antimicrobial policies in poultry farming. Full article

Background:Enterococcus spp. is the third leading cause of healthcare-associated infections in the American continent, often because of the virulence factors that protect the bacterium against host defenses and facilitate tissue attachment and genetic material exchange. In addition, vancomycin, considered a last-resort treatment, has shown reduced efficacy in Enterococcus spp. strains. However, the relationship between bacterial resistance and virulence factors remains unclear. This study intends to evaluate the prevalence of glycopeptide-resistant genotypes and virulence factors in Enterococcus spp. strains. Methods: Over six months, 159 Enterococcus spp. strains causing nosocomial infections were analyzed. Multiplex PCR was performed to identify species, glycopeptide-resistant genotypes, and 12 virulence factors. Results: The most abundant species identified were Enterococcus faecalis and E. faecium. Vancomycin resistance was observed in 10.7% of the isolates, and the vanA genotype was present in 47% of resistant samples. The main virulence factors detected were acm (54%), which is related to cell adhesion; gel E (66%), a metalloproteinase linked to tissue damage; and the sex pheromones cpd (64%) and ccf (84%), which are involved in horizontal gene transfer. A significant association was found between the prevalence of acm, ccf, and cpd in VRE isolates, indicating the potential dissemination of genes to emerging strains via horizontal gene transfer. In addition, a new E. faecium, which displayed five virulence factors and harbored the vanA sequence type, was identified and registered as ST2700. Conclusions:Enterococcus faecalis and E. faecium are clinically critical due to multidrug resistance and virulence factors like acm, which aids host colonization. Genes ccf and cpd promote resistance spread via horizontal transfer, while the emerging ST2700 strain requires urgent monitoring to curb its virulent, drug-resistant spread. Full article

Antimicrobial susceptibility testing (AST) is not routinely performed for C. difficile infection (CDI); however, reports of antimicrobial resistance to various antibiotics have increased. This study aimed to assess the rate of antimicrobial resistance to four antimicrobials (vancomycin, metronidazole, tigecycline, and ciprofloxacin) to assess risk factors for antimicrobial resistance and evaluate MIC variation in patients with recurrence. Data from consecutive patients with CDI admitted to our institution between 1 January 2022 and 30 April 2023 were collected. We performed AST with gradient diffusion and NAAT to evaluate the presumptive presence of R027/NAP1 and toxin production genes. Antimicrobial susceptibility testing was performed on 108 available isolates. We did not find any resistance to vancomycin (median MIC 0.5 μg/mL), metronidazole (median MIC 1 μg/mL), and tigecycline (median MIC 0.016 μg/mL), while resistance to ciprofloxacin was detected in all the samples. Among the recurrent isolates, 37.5% displayed a 2-fold MIC increase for vancomycin, 75% for metronidazole, and 37.5% for tigecycline. After stratifying clinical outcomes according to vancomycin MIC, patients with higher MIC experienced increased 28-day mortality (p value 0.009). Our results were concordant with European surveillance data. MIC increase in all tested antibiotics in patients with CDI warrants further research since decreased susceptibility has been associated with clinical failure. Full article

Background/Objectives: Methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Staphylococcus pseudintermedius (MRSP) represent important antimicrobial resistance threats related to companion animals, which can directly or indirectly lead to adverse health effects in humans and animals living in close contact. Characterizing the phenotypic resistance of MRSA and MRSP to a panel of antimicrobials relevant to both veterinary and human medicine is crucial within a “One Health” framework. Methods: In this study, a total of 79 presumptive MRSA isolates (34 from cats, 45 from dogs) and 110 presumptive MRSP isolates (105 from dogs, 5 from cats) from clinical cases were analysed. Real-time PCR was used to detect the presence of mecA and mecC genes, and susceptibility testing was performed using the Sensititre EUST2 panel. Results: Most of the isolates (88.9%, 168/189) were positive for the mecA gene, while a minority (1.1%, 2/189) were mecC-positive (2 MRSA, 1 dog, 1 cat). MRSP isolates exhibited acquired resistance to a broader range of antibiotics compared to MRSA strains. Furthermore, several isolates demonstrated acquired resistance to antibiotics considered critically important for human medicine. Resistance to vancomycin was found in an MRSP isolate from a dog, and resistance to linezolid in an MRSP isolate from a cat. This study reveals that 83.3% (30/36) of MRSA isolates from dogs and 89.3% (25/28) from cats were multidrug-resistant organisms, while MRSP isolates exhibited multidrug resistance in 99% (101/102) of cases for dogs and 100% (4/4) for cats. Conclusions: The extremely high level of multidrug resistance, with some isolates resistant to critically important antibiotics used in human medicine, highlight the importance of monitoring antimicrobial susceptibility in MRSA and MRSP isolates collected from cats and dogs in a One Health perspective. Full article

Daptomycin (DAP) is a therapeutic option for vancomycin-resistant Enterococcus faecium (VRE) infections, but DAP resistance may occur during treatment. Previously, we identified a mutation within the E. faecium lafB gene that induces hypersusceptibility to DAP. The lafB gene encodes a glycosyltransferase involved in lipoteichoic acid anchor synthesis, which makes it a promising target for enhancing DAP efficacy. In this study, we characterized E. faecium LafB protein (EfLafB) biophysical properties, used AlphaFold3 to predict LafB in silico three-dimensional structure, and determined lafB gene mutation’s role in virulence, comparing E. faecium HBSJRP18 (DAP-hypersusceptible) and a lafB revertant, HBSJRP18_2.7, and analyzing bacterial growth kinetics, biofilm formation ability, and virulence in a Galleria mellonella model. After gene cloning and expressing and purifying EfLafB, circular dichroism and SEC-MALS assays revealed its monomeric nature under in vitro conditions, with approximately a 40 kDa molecular mass and a melting temperature of 50 °C. In silico prediction indicated that LafB is an αβ-type protein with two domains conforming to the GT-4 family glycosyltransferases. These results are further supported by the highly conserved amino acids (E257, D91, R184, and K185), likely involved in UDP-Glc binding. The studied lafB gene mutation resulted in a significant decrease in bacterial growth and virulence in the invertebrate model. Full article

Background: The genus Enterococcus includes a diverse group of bacteria that are commonly found in the gastrointestinal tracts of humans and animals, as well as in various environmental habitats. Methods: In this study, Enterococcus lactis RB10, isolated from goat feces, was subjected to comprehensive genomic and functional analysis to assess its safety and potential as a probiotic strain. Results: The genome of E. lactis RB10, with a size of 2,713,772 bp and a GC content of 38.3%, was assembled using Oxford Nanopore Technologies (ONT). Genome annotation revealed 3375 coding sequences (CDSs) and highlighted key metabolic pathways involved in carbohydrate, protein, and amino acid metabolism. The strain was susceptible to important antibiotics, including ampicillin, chloramphenicol, tetracycline, and vancomycin, but exhibited resistance to aminoglycosides, a common trait in Enterococcus species with non-hemolytic activity. Genomic analysis further identified two intrinsic antimicrobial resistance genes (ARGs). The strain also demonstrated antimicrobial activity against Bacillus cereus DMST 11098 and Salmonella Typhi DMST 22842, indicating pathogen-specific effects. Key genes for adhesion, biofilm formation, and stress tolerance were also identified, suggesting that RB10 could potentially colonize the gut and compete with pathogens. Moreover, the presence of bacteriocin and secondary metabolite biosynthetic gene clusters suggests its potential for further evaluation as a biocontrol agent and gut health promoter. Conclusions: However, it is important to note that E. lactis RB10 was isolated from goat feces, a source that may harbor both commensal and opportunistic bacteria, and therefore additional safety assessments are necessary. While further validation is needed, E. lactis RB10 exhibits promising probiotic properties with low pathogenic risk, supporting its potential use in food and health applications. Full article

Staphylococcus aureus is a significant bacterial pathogen responsible for a wide range of diseases in both humans and animals. This study aimed to investigate nucleotide sequence variations, gene expression patterns, and serum biomarkers, including acute phase proteins (APPs), hormonal fluctuations, and iron profile parameters in sheep affected by pneumonia. Additionally, the study focused on the isolation and characterization of S. aureus from pneumonic sheep, with particular emphasis on the prevalence of methicillin-resistant S. aureus (MRSA) strains. Blood samples were collected from both healthy and pneumonic sheep for gene expression and biochemical analyses, while nasal swabs from pneumonic sheep were used for bacterial isolation and identification. Out of 100 nasal swabs analyzed, 44% tested positive for Staphylococcus spp., and 61.4% of these were confirmed as S. aureus by PCR. The mecA gene, a key marker of methicillin resistance, was identified in 17 isolates (38.6% of the S. aureus-positive samples). MRSA isolates showed complete resistance to amoxicillin, cloxacillin, and erythromycin, and high resistance to penicillin, amoxicillin, and tetracycline; however, all MRSA strains remained fully susceptible to vancomycin. Gene expression analysis revealed that TLR2, CLEC4E, PTX3, CXCL8, and IL15RA were significantly upregulated (p < 0.05) in pneumonic ewes, while SOCS3 expression was markedly downregulated. Sequence analysis of immune-related genes revealed notable nucleotide differences between healthy and affected animals. Furthermore, the pneumonic group exhibited significantly elevated levels of APPs, cortisol, and growth hormone, along with reduced levels of insulin, T3, and T4. These findings underscore the zoonotic risk posed by MRSA and emphasize the need for robust surveillance and antibiotic stewardship to control its spread. The study also highlights the importance of molecular diagnostics in accurately identifying MRSA and elucidating resistance mechanisms, thereby facilitating targeted treatment and informed management strategies. Full article

Staphylococcus aureus is an important cause of food intoxication, which has the potential to induce diverse infections, toxinoses and life-threatening diseases among humans and animals. This study investigated the prevalence, antimicrobial resistance, and genetic diversity of S. aureus and methicillin-resistant S. aureus (MRSA) in retail raw meat from Shandong (March 2021–October 2022). The distribution of virulence genes, antimicrobial susceptibility, and genetic diversity of these isolates were analyzed. From a total of 442 samples, 87 (19.7%) S. aureus and 11 (2.5%) MRSA were isolated. According to the antimicrobial susceptibility testing, it was found that all the S. aureus isolates were resistant to at least one antimicrobial. Most isolates (95.9%) were resistant to penicillin, with high resistance to ampicillin (82.7%) and multidrug resistance in 76.5% of cases. One isolate could simultaneously resist eleven antimicrobials (ERY-CLI-GEN-SMZ-FFC-PEN-PRL-AMC-CIP-TET-AMP). In contrast, all the isolates showed sensitivity to vancomycin. The most prevalent virulence gene was sed, accounting for 10.2%, followed by sec (8.2%). Regarding genetic polymorphism, these isolates were divided into 21 different sequence types (STs) using multilocus sequence typing (MLST) and 33 staphylococcal protein A (spa) types using spaTyper 1.0 tool. The most prevalent sequence types were ST398 (22.4%), followed by ST7 (20.4%), while ST59, ST1, ST188, ST9, ST398, and ST7 were observed in MRSA isolates. The most prevalent spa types were t034 (15.3%), followed by t899 (10.2%), while t441, t127, t184, t899, t034, and t091 were observed in MRSA isolates. In conclusion, our study highlights the high prevalence of S. aureus and MRSA in different retail raw meats in Shandong. This poses a potential threat to food safety and underscores the need for enhanced surveillance and stricter antibiotic control measures. Full article