Enterococcus Species Research Articles

Genomics of Staphylococcus aureus and Enterococcus faecalis isolated from the ocular surface of dry eye disease sufferers

Ocular surface inflammatory disorders, such as dry eye, are becoming increasingly prevalent. Developing new treatment strategies targeting harmful bacteria could provide significant therapeutic benefits. The purpose of this study was to characterize the common ocular pathogen Staphylococcus aureus and the rarer endophthalmitis-associated species Enterococcus faecalis isolated from the ocular surface of dry eye disease patients in Norway. Together the 7 isolates (5 S. aureus and 2 E. faecalis) comprise the complete set of members of each species isolated in our previous study of the ocular microbiome of 61 dry eye sufferers. We aimed to investigate the pathogenic potential of these isolates in relation to ocular surface health. To this end, we used whole genome sequencing, multiplex PCR directed at virulence genes and antibiotic susceptibility tests encompassing clinically relevant agents. The E. faecalis isolates showed resistance to only gentamicin. S. aureus isolates displayed susceptibility to most of the tested antibiotics, except for two isolates which showed resistance to trimethoprim/sulfamethoxazole and three isolates which were resistant to ampicillin. Susceptibilities included sensitivity to several first-line antibiotics for treatment of ocular infections by these species. Thus, treatment options would be available if required. However, spontaneous resistance development to gentamicin and rifampicin occurred in some S. aureus which could be a cause for concern. Whole genome sequencing of the isolates showed genome sizes ranging from 2.74 to 2.83 Mbp for S. aureus and 2.86 Mbp for E. faecalis, which is typical for these species. Multilocus sequence typing and phylogenetic comparisons with previously published genomes, did not suggest the presence of eye-specific clusters for either species. Genomic analysis indicated a high probability of pathogenicity among all isolates included in the study. Resistome analysis revealed the presence of the beta-lactamase blaZ gene in all S. aureus isolates and the dfrG gene in two of them; while E. faecalis isolates carried the lsa(A) gene which confers intrinsic resistance to lincosamides and streptogramin A in this species. Screening for virulence factors revealed the presence of various pathogenicity associated genes in both S. aureus and E. faecalis isolates. These included genes coding for toxin production and factors associated with evading the host immune system. Some of the identified genes (tst, hylA &hylB) are suggested to be linked to the pathophysiology of dry eye disease. Lastly, the presence of specific S. aureus virulence genes was confirmed through multiplex PCR analysis.

Clinical outcome of ampicillin or ampicillin/sulbactam versus glycopeptides in ampicillin-susceptible Enterococcus faecalis/faecium bacteremia: a 10-year retrospective cohort study

BackgroundGlycopeptides for ampicillin-susceptible Enterococcus faecalis/faecium bacteremia are readily prescribed depending on the severity of the condition. However, there is limited data on the outcomes of glycopeptide use compared to ampicillin-containing regimens for ampicillin-susceptible E. faecalis/faecium bacteremia. From an antibiotic stewardship perspective, it is important to determine whether the use of glycopeptides is associated with improved clinical outcomes in patients with ampicillin-susceptible E. faecalis/faecium bacteremia.MethodsThis retrospective cohort study was conducted at a university-affiliated hospital between January 2010 and September 2019. We collected data from patients with positive blood cultures for Enterococcus species isolates. The clinical data of patients who received ampicillin-containing regimens or glycopeptides as definitive therapy for ampicillin-susceptible E. faecalis/faecium bacteremia were reviewed. Multivariate logistic regression analysis was performed to identify risk factors for 28-day mortality.ResultsAmpicillin-susceptible E. faecalis/faecium accounted for 41.2% (557/1,353) of enterococcal bacteremia cases during the study period. A total of 127 patients who received ampicillin-containing regimens (N = 56) or glycopeptides (N = 71) as definitive therapy were included in the analysis. The 28-day mortality rate was higher in patients treated with glycopeptides (19.7%) than in those treated with ampicillin-containing regimens (3.6%) (p = 0.006). However, in the multivariate model, antibiotic choice was not an independent predictor of 28-day mortality (adjusted OR, 3.7; 95% CI, 0.6–23.6).ConclusionsGlycopeptide use was not associated with improved mortality in patients with ampicillin-susceptible E. faecalis/faecium bacteremia. This study provides insights to reduce the inappropriate use of glycopeptides in ampicillin-susceptible E. faecalis/faecium bacteremia treatment and promote antimicrobial stewardship.

Overexpression, purification and biochemical studies of Sortase A from Enterococcus faecalis (Ef) and its inhibition studies with Aloenin

Sortase A (SrtA) is a bacterial transpeptidase that garnishes the bacterial surface by adding various virulent factors or proteins by cleaving the LPXTG-specific motif between T and G amino acids. These virulence factors assist in the attachment of host cells, which are essential for bacterial virulence. Enterococcus species are among the multidrug-resistant bacteria that cause nosocomial infections; they have drawn a lot of attention recently. SrtA from E. faecalis (Ef) plays a critical role in pathogenesis, making it a suitable target for the development of antibacterial agents. Since SrtA is not involved in bacterial growth and is present on the surface of bacteria, the probability of developing antibiotic resistance is minimal. In this work, we have done cloning, expression and purification of Ef-SrtA using IMAC (Immobilised Metal Affinity Chromatography) followed by Gel filtration chromatography. Purified Ef-SrtA showed maximum activity at pH-8 and temperature between 45 and 55 °C. The fluorescent assay for kinetic studies of Ef-SrtA showed Vmax 3.852 µM.min−1 and kcat 7.7 × 10−2s−1 for the hydrolysis of substrate using Abz-LPETG-K(Dnp)-NH2. We have selected fifteen Aloe vera extracted compounds and performed virtual screening and docking experiments to identify potential inhibitors against Ef-SrtA. Among fifteen molecules, Aloenin-a which was bound to the active site with a binding energy of -6.1 kcal/mol, interacted with the active site residues, Arg139, Pro105, Leu39, Ala46, and Cys126. Aloenin-a showed a significant inhibitory effect against Ef-SrtA, with an IC50 value of 20.68 µM. Aloenin-a inhibits biofilm formation at concentrations of 20–250 µg/mL. The fibrinogen assay showed adherence to fibrinogen was reduced in the presence of Aloenin-a for E. faecalis. The results demonstrated that Aloe vera extracts containing Aloenin-a can be a significant antagonist of Ef-SrtA.

A first study of meat-borne enterococci from butcher shops: prevalence, virulence characteristics, antibiotic resistance and clonal relationship.

IntroductionEnterococcus, which used to be thought of as a harmless commensal living in the digestive tract, has now become a highly resistant and highly contagious pathogen that makes nosocomial infections much more common. This study examined enterococci species and their antibiotic resistance phenotypes and genotypes and virulence gene content in Turkish ground beef samples. Methodology A total of 100 ground beef samples were analyzed between May 2020 and May 2021. The isolated strains were identified via matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and confirmed using polymerase chain reaction (PCR) after which they were divided into several species using PCR and tested for antibiotic resistance against 19 antimicrobial agents using the disc diffusion method. The genetic similarity analysis, pulsed-field gel electrophoresis (PFGE) was performed. Results A total of 93 isolates in ground beef were identified, comprised of E. faecalis 72.04%; E. hirae- 11.82%; E. casseliflavous- 7.52%; E. faecium- 5.3%; E. gallinarium- 3.23%. The virulence genes observed in Enterococcus species were distributed as follows: gelE 88.1%, ace 53.7%, efaA 40.8%, asaI 19.3%, esp 6.4%, and cylA 1.07%. A high antibiotic resistance was recorded for tetracycline (43.01%), followed by ampicilin (17.2%), and chloramphenicol (13.9%). 17.2% of Enterococcus isolates were multidrug-resistant. The study determined the high prevalence of antibiotic resistance genes, specifically for tet(L) 10 (10.7%), aac(6')Ie-aph(2")-la 3 (3.2%), and ermB 3 (3.2%). The presence of efflux pump genes were identified in 74.1% of Enterococcus isolates. Genetic characterization of 67 E. faecalis isolates by PFGE revealed 41 pulsed-field gel electrophoresis (PFGE) patterns that were grouped into 15 clusters, which presented more than one strain with 100% similarity. Conclusion Isolates obtained from several areas and butchers had comparable patterns of PFGE, suggesting that the presence of circulating E. faecalis poses a potential public health concern in diverse districts. To mitigate the health hazards associated with the contamination of enterococci from raw to cooked meats, it is necessary to enhance the disinfection of butcheries, promote excellent hand hygiene among butchers, and implement appropriate meat storage and handling methods to prevent bacterial development.

PCR identification of Enterococcus Species as a Primary Pathogen in Neonatal Bloodstream Infections at Damascus University Paediatric Hospital

Bacterial septicaemia is one of the most common causes of death in newborns throughout the globe. Recently, gram positive bacteria have been detected as a main responsible of hospital acquired blood infection in developed country especially Enterococcus sp. and Staphylococcus sp. This preliminary study aimed to identify Enterococcus species and Staphylococcus species isolated form acquired blood infection in the department of premature and neonates at paediatric university hospital in Damascus, based on PCR technology. the antibiotic sensitivity these pathogens was also investigated against common antibiotics usually used to treat septicaemia in this hospital. Bacterial gDNA was extracted directly from blood samples of children with acquired Septicemia. The bacterial infection was confirmed in 50 samples based on the amplification of 16srDNA gene. Genus Enterococcus sp. and Staphylococcus species were identified as source of infection in samples using specific primers via PCR technique. The antibiotic sensitivity of isolated bacterial strains was studied using Kirby-Bauer disc method. Our study showed that 47 of the 50 samples of the infection were bacterial. The rate of blood infection caused by Enterococcus species was 24%, and 4% for Staphylococcus sp. the isolated bacteria showed a high sensitivity to Amikacin, medium sensitivity to Vancomycin, and less sensitivity to Ampicillin and Gentamycin. Enterococcus species Showed good sensitivity to Amikacin and Vancomycin and low sensitivity to Gentamycin, while was resistant to Ampicillin. Furthermore, the two isolated stains Enterococcus sp and Staphylococcus species showed a good sensitivity against chlorhexidine. This study confirmed the importance of molecular technologies in diagnosing the acquired septicaemia and identifying the specific infectious agent. The molecular test showed that Enterococcus sp. was an important cause of acquired blood infection in neonates respect to ampicillin resistant staphylococcus species.

Identification of Enterococcus spp. by MALDI-TOF mass spectrometry isolated from clinical mastitis and bulk tank milk samples

BackgroundThroughout a three-year study period, 1,577 bovine clinical mastitis samples and 302 bulk tank samples were analyzed from ten Brazilian dairy herds. Enterococcus spp. was isolated and identified in 93 (5.9%) clinical mastitis samples. In addition, 258 Enterococcus spp. were isolated from the bulk tank samples of the same herds. The identification of Enterococcus spp. isolated from bulk tanks and milk samples of clinical mastitis were accomplished by phenotypic characteristics and confirmed by MALDI-TOF Mass Spectrometry (MS). Fisher test was performed to verify the difference between bulk tanks and mastitis samples.ResultsThe following species were identified from clinical mastitis: E. saccharolyticus (62.4%), E. faecalis (19.4%), E. faecium (15.1%), E. hirae (1.1%), E. mundtii (1.1%), E. durans (1.1%). Furthermore, from 258 bulk tank milk samples, eight enterococci species were isolated: E. faecalis (67.8%), E. hirae (15.1%), E. faecium (4.6%), E. saccharolyticus (4.6%), E. mundtii (3.1%), E. caseliflavus ( 2.7%), E. durans (1.2%), E. galinarum (0.8%).ConclusionsThe difference in species predominance in bulk tank samples (67.8% of E. faecalis) and clinical mastitis (62.4% of E. saccharolyticus) was unexpected and caught our attention. Although Enterococcus spp. are traditionally classified as an environmental mastitis agent, in the present study, E. saccharolyticus behaved as a contagious agent of mastitis, which consequently changed the control patterns to be implemented.

A glassy carbon electrode modified with gold decorated iron oxide/ carbon dots for light assisted voltammetric detection of antibiotic resistant microbe Enterococcusfaecalis

Detecting bacteria is essential in managing significant health concerns as it enables timely intervention, reducing complications and improving patient outcomes, particularly in treating common infections that necessitate precise identification for effective symptom management. Enterococcus species represent a notable threat in hospital-acquired infections and urinary tract infections (UTIs), given the increasing prevalence of strains resistant to multiple antibiotics, unresponsive to standard therapies, and carrying various virulence factors. Traditional approaches to identifying Enterococcus faecalis (E. faecalis) have limitations, including prolonged processing times, limited sensitivity, and the potential for false positive results. While Polymerase Chain Reaction (PCR) is a valuable tool, it is susceptible to contamination and variations in DNA concentration. The emerging technique of Photoelectrochemical (PEC) holds promise for enhancing E. faecalis detection by leveraging photogenerated electrons and holes. This study introduces a rapid and precise approach utilizing a light-assisted electrochemical biosensor featuring a glassy carbon electrode modified with a nanocomposite of gold-coated iron oxide and carbon dots (Au@Fe3O4/CDs). The nanocomposite was successfully synthesized and underwent thorough characterization. The investigation has a detection range from 1 to 14 CFU mL−1, along with a notably low limit of detection (LOD: 3 CFU mL−1, LOQ: 10 CFU mL−1). Rigorous examination of real-world samples such as food, water, and soil demonstrated exceptional specificity, reproducibility, and long-term stability of the sensor. The applications of the Au@Fe3O4/CDs nanocomposite in PEC processes underscore the potential of this innovative approach in addressing health concerns associated with bacterial infections and delivering real-time impacts for both healthcare and environmental domains.

Integration of MALDI-TOF MS and machine learning to classify enterococci: A comparative analysis of supervised learning algorithms for species prediction

This research focused on distinguishing distinct matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectral signatures of three Enterococcus species. We evaluated and compared the predictive performance of four supervised machine learning algorithms, K-nearest neighbor (KNN), support vector machine (SVM), and random forest (RF), to accurately classify Enterococcus species. This study involved a comprehensive dataset of 410 strains, generating 1640 individual spectra through on-plate and off-plate protein extraction methods. Although the commercial database correctly identified 76.9% of the strains, machine learning classifiers demonstrated superior performance (accuracy 0.991). In the RF model, top informative peaks played a significant role in the classification. Whole-genome sequencing showed that the most informative peaks are biomarkers connected to proteins, which are essential for understanding bacterial classification and evolution. The integration of MALDI-TOF MS and machine learning provides a rapid and accurate method for identifying Enterococcus species, improving healthcare and food safety.

Identification of vancomycin resistance in enterococcus & determination of minimal inhibitory concentration of vancomycin by E-test

Infections due to Enterococcal species is 2 prevalent cause of Nosocomial Infections in health care settings. They are crucial because to their inherent resistance to penicillins, cephalosporins, and low level aminoglycosides & their tendency to develop higher level of aminoglycoside and vancomycin resistance, which leaves few therapeutic alternatives for therapy. Isolation and identification of Enterococci with their Antibiogram & determination of Minimal Inhibitory Concentration (MIC) of Vancomycin by E-test. The antibiogram generated by Kirby Baur disc diffusion method and standard microbiological procedures were used to identify enterococcal isolates from distinct clinical samples. MIC level for Vancomycin was tested for all the isolates using E-test from BIOMERIEUX. Total of 108 isolates (62 E.faecalis and 46 E. faecium) were tested by E-test for MIC of Vancomycin, of which 40 (37.03%) (22 E.faecalis & 18 E. faecium) were Vancomycin Resistant (VRE) strains, having MIC level >32 μg/ml. Nine strains out of 40 (45%) were found to have MIC >256 μg/ml. The enterococcal isolates were observed to be sensitive to Linezolid (80%), Ofloxacin (74%), Amoxyclav (74%) and Cefaperazone (70%). For the Indian medical community, the rise of antibiotic resistance among Enterococcal species has been a significant concern. So this is need of present era that our laboratories must be well-equipped to report the resistance patterns displayed by enterococci in order to practise evidence-based medicine and reduce hospital acquired infections.

Dual quorum-sensing control of purine biosynthesis drives pathogenic fitness of Enterococcus faecalis.

Enterococcus faecalis is a resident of the human gut, though upon translocation to the blood or body tissues, it can be pathogenic. Here we discover and characterize two peptide-based quorum-sensing systems that transcriptionally modulate de novo purine biosynthesis in E. faecalis. Using a comparative genomic analysis, we find that most enterococcal species do not encode this system; E. moraviensis, E. haemoperoxidus and E. caccae, three species that are closely related to E. faecalis, encode one of the two systems, and only E. faecalis encodes both systems. We show that these systems are important for the intracellular survival of E. faecalis within macrophages and for the fitness of E. faecalis in a murine wound infection model. Taken together, we combine comparative genomics, microbiological, bacterial genetics, transcriptomics, targeted proteomics and animal model experiments to describe a paired quorum sensing mechanism that directly influences central metabolism and impacts the pathogenicity of E. faecalis.

Postbiotic emissaries: a comprehensive review on the bioprospecting and production of bioactive compounds by Enterococcus species

SummaryEnterococcus species have been acknowledged for their diverse metabolic capabilities, resulting in a wide array of bioactive compounds that show promise for therapeutic use. This article presents a thorough examination of the exploration and synthesis of bioactive compounds by Enterococcus species. It encompasses the taxonomy, natural habitat, and significance in the biotechnology of Enterococcus, emphasising the various bioactive compounds it produces, including antibacterial, immunomodulatory, and antioxidant agents. Strategies for exploring potential bioactivities, encompassing isolation methodologies and screening techniques, are discussed. The biotechnological synthesis of bioactive compounds derived from Enterococcus via fermentation processes, optimisation approaches, and scaling methodologies is expounded. Moreover, the utilisation of these bioactive compounds in functional foods, pharmaceuticals, and agriculture, alongside considerations of safety and regulatory aspects, is examined. The review concludes with future perspectives and challenges in the field of Enterococcus‐derived bioactive compounds.

Microbial contamination of spittoons and germicidal effect of irradiation with krypton chloride excimer lamps (Far UV-C 222 nm).

In dentistry, instruments, appliances, and body fluids such as saliva or blood are possible sources of infection. Although conventional antiseptic procedures effectively prevent infection, spittoons cannot be sanitized between each treated patient and are usually washed only with running water. However, there is currently no fast and efficient disinfection method that can be implemented between treatments. An optically filtered krypton chloride excimer lamp using ultraviolet light (Far UV-C) in the 200-230 nm wavelength range (innocuous to humans) has been recently used as a virus- and bacteria-inactivating technology. This study aimed to identify the bioburden of a dental spittoon and examine the susceptibility of two oral Streptococcus and two Enterococci to 222-nm Far UV-C by irradiating the spittoon with 222 nm Far UV-C for 5 min before evaluating the disinfection effect. Bacterial analysis and real-time polymerase-chain reaction testing was used to confirm the spittoon's biological contamination. Bacterial susceptibility to a 222-nm Far UV-C was determined with a graded dose irradiation test. After each treatment, the spittoon was irradiated with 222-nm Far UV-C for 5 min, and the disinfecting effect was evaluated. Microbial analysis of the spittoon's surface was performed using the Silva database. We found that > 97% of the microbes consisted of six bacterial phyla, whereas no viruses were found. Pseudomonas aeruginosa was frequently detected. The 1-log reduction value of two oral-derived Streptococci and two Enterococci species at 222-nm Far UV-C was 4.5-7.3 mJ/cm2. Exposure of the spittoon to 222-nm Far UV-C at 3.6-13.5 mJ/cm2 significantly decreased bacterial counts (p < 0.001). Irradiation with 222-nm Far UV-C at 3.6-13.5 mJ/cm2 significantly eliminates bacteria in spittoons, even when they are only rinsed with water. Hence, 222-nm Far UV-C irradiation may inhibit the risk of bacterial transmission from droplets in sink surfaces.

The Occurrence of Colistin Resistance in Potential Lactic Acid Bacteria of Food-Producing Animals in India.

The overuse of colistin, the last-resort antibiotic, has led to the emergence of colistin-resistant bacteria, which is a major concern. Lactic acid bacteria which are generally regarded as safe are known to be reservoirs of antibiotic resistance that possibly pose a threat to human and animal health. Therefore, this study assessed the prevalence of colistin antimicrobial resistance in livestock in India, that is lactic acid bacteria in healthy chickens, sheep, beef, and swine of Mysore. Diverse phenotypic and genotypic colistin resistance were examined among the lactic acid bacterial species (n = 84) isolated from chicken (n = 44), sheep (n = 16), beef (n = 14), and swine (n = 10). Hi-comb, double-disk diffusion tests, Minimum Inhibitory Concentration (MIC), and biofilm formation were assessed for phenotypic colistin resistance. Specific primers for colistin-resistant genes were used for the determination of genotypic colistin resistance. Around 20%, 18%, and 1% were colistin-resistant Lactobacillus, Enterococcus, and Pediococcus species, respectively. Among these, 66.67% exhibited MDR phenotypes, including colistin antibiotic. The identified resistant isolates are Levilactobacillus brevis LBA and LBB (2), Limosilactobacillus fermentum LBF (1), and Pediococcus acidilactici CHBI (1). The mcr-1 and mcr-3 genes were detected in Levilactobacillus brevis LBA, LBB, and Pediococcus acidilactici CHBI isolated from chicken and sheep intestines respectively. The study identified colistin resistance determinants in lactobacilli from food animals, emphasizing the need for enhanced surveillance and monitoring of resistance spread. These findings underscore colistin resistance as a significant medical concern and should be integrated into India's ongoing antimicrobial resistance monitoring programs.

Isolation and characterization of enterococci from poultry reveals high incidence of Enterococcus thailandicus in Victoria, Australia.

Antibiotic resistance is a global health crisis. Roughly two-thirds of all antibiotics used are in production animals, which have the potential to impact the development of antibiotic resistance in bacterial pathogens of humans. There is little visibility on the extent of antibiotic resistance in the Australian food chain. This study sought to establish the incidence of antibiotic resistance among enterococci from poultry in Victoria. In 2016, poultry from a Victorian processing facility were swabbed immediately post-slaughter and cultured for Enterococcus species. All isolates recovered were speciated and tested for antibiotic susceptibility to 12 antibiotics following the Clinical Laboratory Standards Institute guidelines. A total of 6 farms and 207 birds were sampled and from these 285 isolates of Enterococcus were recovered. Eight different enterococcal species were identified as follows: E. faecalis (n=122; 43%), E. faecium (n=92; 32%), E. durans (n=35; 12%), E. thailandicus (n=23; 8%), E. hirae (n=10; 3%), and a single each of E. avium, E. gallinarum, and E. mundtii. Reduced susceptibility to older classes of antibiotics was common, in particular: erythromycin (73%), rifampin (49%), nitrofurantoin (40%), and ciprofloxacin (39%). Two vancomycin-intermediate isolates were recovered, but no resistance was detected to either linezolid or gentamicin. The relatively high numbers of a recently described species, E. thailandicus, suggest this species might be well adapted to colonize poultry. The incidence of antibiotic resistance is lower in isolates from poultry than in human medicine in Australia. These results suggest that poultry may serve as a reservoir for older antibiotic resistance genes but is not driving the emergence of antimicrobial resistance in human bacterial pathogens. This is supported by the absence of resistance to linezolid and gentamicin.

Community context influences the conjugation efficiency of Escherichia coli.

In urinary tract infections (UTIs), different bacteria can live in a polymicrobial community consisting of different species. It is unknown how community members affect the conjugation efficiency of uropathogenic Escherichia coli. We investigated the influence of individual species often coisolated from urinary infections (UTI) on the conjugation efficiency of E. coli isolates in artificial urine medium. Pairwise conjugation rate experiments were conducted between a donor E. coli strain containing the pOXA-48 plasmid and six uropathogenic E. coli isolates, in the presence and absence of five different species commonly coisolated in polymicrobial UTIs to elucidate their effect on the conjugation efficiency of E. coli. We found that the basal conjugation rates of pOXA-48, in the absence of other species, are dependent on the bacterial host genetic background. Additionally, we found that bacterial interactions have an overall positive effect on the conjugation rate of pOXA-48. Particularly, Gram-positive enterococcal species were found to enhance the conjugation rates towards uropathogenic E. coli isolates. We hypothesize that the nature of the coculture and physical interactions are important for these increased conjugation rates in an artificial urine medium environment.

Guarding the walls: the multifaceted roles of Bce modules in cell envelope stress sensing and antimicrobial resistance.

Bacteria have developed diverse strategies for defending their cell envelopes from external threats. In Firmicutes, one widespread strategy is to use Bce modules-membrane protein complexes that unite a peptide-detoxifying ABC transporter with a stress response coordinating two-component system. These modules provide specific, front-line defense for a wide variety of antimicrobial peptides and small molecule antibiotics as well as coordinate responses for heat, acid, and oxidative stress. Because of these abilities, Bce modules play important roles in virulence and the development of antibiotic resistance in a variety of pathogens, including Staphylococcus, Streptococcus, and Enterococcus species. Despite their importance, Bce modules are still poorly understood, with scattered functional data in only a small number of species. In this review, we will discuss Bce module structure in light of recent cryo-electron microscopy structures of the B. subtilis BceABRS module and explore the common threads and variations-on-a-theme in Bce module mechanisms across species. We also highlight the many remaining questions about Bce module function. Understanding these multifunctional membrane complexes will enhance our understanding of bacterial stress sensing and may point toward new therapeutic targets for highly resistant pathogens.

Antimicrobial Evaluation of Two Polycyclic Polyprenylated Acylphloroglucinol Compounds: PPAP23 and PPAP53.

Polycyclic polyprenylated acylphloroglucinols (PPAPs) comprise a large group of compounds of mostly plant origin. The best-known compound is hyperforin from St. John's wort with its antidepressant, antitumor and antimicrobial properties. The chemical synthesis of PPAP variants allows the generation of compounds with improved activity and compatibility. Here, we studied the antimicrobial activity of two synthetic PPAP-derivatives, the water-insoluble PPAP23 and the water-soluble sodium salt PPAP53. In vitro, both compounds exhibited good activity against methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium. Both compounds had no adverse effects on Galleria mellonella wax moth larvae. However, they were unable to protect the larvae from infection with S. aureus because components of the larval coelom neutralized the antimicrobial activity; a similar effect was also seen with serum albumin. In silico docking studies with PPAP53 revealed that it binds to the F1 pocket of human serum albumin with a binding energy of -7.5 kcal/mol. In an infection model of septic arthritis, PPAP23 decreased the formation of abscesses and S. aureus load in kidneys; in a mouse skin abscess model, topical treatment with PPAP53 reduced S. aureus counts. Both PPAPs were active against anaerobic Gram-positive gut bacteria such as neurotransmitter-producing Clostridium, Enterococcus or Ruminococcus species. Based on these results, we foresee possible applications in the decolonization of pathogens.

CRISPR-Cas System, Antimicrobial Resistance, and Enterococcus Genus-A Complicated Relationship.

(1) Background: The rise in antibiotic resistant bacteria poses a significant threat to public health worldwide, necessitating innovative solutions. This study explores the role of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) in the context of antibiotic resistance among different species from the Enterococcus genus. (2) Methods: The genomes of Enterococcus included in the study were analyzed using CRISPRCasFinder to distinguish between CRISPR-positive (level 4 CRISPR) and CRISPR-negative genomes. Antibiotic resistance genes were identified, and a comparative analysis explored potential associations between CRISPR presence and antibiotic resistance profiles in Enterococcus species. (3) Results: Out of ten antibiotic resistance genes found in Enterococcus species, only one, the efmA gene, showed a strong association with CRISPR-negative isolates, while the others did not significantly differ between CRISPR-positive and CRISPR-negative Enterococcus genomes. (4) Conclusion: These findings indicate that the efmA gene may be more prevalent in CRISPR-negative Enterococcus genomes, and they may contribute to a better understanding of the molecular mechanisms underlying the acquisition of antibiotic resistance genes in Enterococcus species.

Is It Useful to Repeat Blood Cultures in Endocarditis Patients? A Critical Appraisal.

Previous guidelines for endocarditis have suggested repeating blood cultures until they become negative, with limited evidence. Literature reviews were conducted (1) on the incidence of persistent bacteremia and association with outcome and (2) on timing of valve culture negativization to examine the claim for prolongation of antibiotic therapy starting from negative blood cultures. Persistent bacteremia and fever may be present in the first 3 days of endocarditis, despite treatment, and are more common in Staphylococcus (especially MRSA) and Enterococcus species. Persistent bacteremia (48-72 h), persistent infection (day 7), and new onset septic shock are related and predict in-hospital mortality. It is, however, persistent infection at day 7 and septic shock that primarily determine the infectious course of endocarditis, and not persistent bacteremia. Valve cultures at surgery become negative in most cases (>85-90%) after 14-21 days of antibiotic therapy, with no calculated benefit for prolonging therapy after 21 days. Persistent infection at 7 days after appropriate antibiotic therapy is a better key event for prognosis then positive or negative blood cultures at 48-72 h. Therapy prolongation from the day of negative blood cultures is not reasonable. There is no need to survey blood cultures in endocarditis patients after starting therapy.

Antimicrobial Resistance of Enterococcus Isolates from Poultry Farms in the Republic of Serbia (Autonomous Province of Vojvodina).

Enterococcus species are significant intestinal commensals of animals, including poultry. However, they have emerged as important opportunistic infective agents in both veterinary and human medicine as well as major nosocomial pathogens, owing to their increasing antimicrobial resistance. This research aimed to investigate the prevalence and antimicrobial resistance patterns of Enterococcus spp. isolated from poultry farms in the north of Serbia. A total of 40 samples of overshoes or feces were collected from 40 poultry farms and analyzed for the presence of Enterococcus spp. using PCR or MALDI-TOF mass spectrometry for their identification. The number of isolates was 40 and included 11 isolates from laying hens, 2 isolates from turkeys, 3 from broiler breeders, and 24 from broilers. The Kirby-Bauer disk diffusion method was used to test for antibiotic susceptibility in accordance with the Clinical and Laboratory Standards Institute and EUCAST guidelines. The results showed that Enterococcus faecalis was isolated from 37.5% farms, and E. faecium from 42.5%. E. hirae was identified in 15% of poultry establishments, and E. durans and E. thialandicus on 2.5%. Notably, resistance to erythromycin, streptomycin, fluoroquinolones, and tetracyclines among the frequently used antibiotics was found. Furthermore, 35% of the isolates had multidrug resistance (MDR). In order to prevent the spread of antibiotic resistance in chicken farming and protect the health of the public and animals alike, our findings highlight the critical need for improved surveillance and control measures. To effectively establish a containment strategy for Enterococcus spp. isolated from poultry farms, more research into the processes behind their antibiotic resistance is required.