The polymicrobial nature of the oral cavity and claws of cats diagnosed by mass spectrometry and next-generation sequencing.

Close contact between cats and humans increases the risk of transmission of zoonotic pathogens, through bites and scratches due to the complexity of microorganisms in the oral and nail microbiotas of felines. This study investigated the presence of bacteria and fungi in the oral cavity and claws of 100 apparently healthy cats using conventional and selective microbiological culture media, and next-generation sequencing (NGS) and mass spectrometry (MALDI-TOF MS). Furthermore, antimicrobial susceptibility testing of bacteria isolates was performed by disc diffusion method. In total, 671 bacteria and 33 yeasts were identified by MALDI-TOF MS. Neisseria animaloris (10.8%), Staphylococcus felis (8.5%), and Pasteurella multocida (7%) were the most prevalent bacteria in oral cavity samples (n=343), while the most common yeast (n=19) was Candida albicans (68.4%). Staphylococcus pettenkoferi (13.4%), Staphylococcus felis (6.4%), and Staphylococcus simulans (5.8%) were the prevalent bacteria identified in the claw samples (n=328), while Rhodotorula mucilaginosa (57.2%) was the most common yeast (n=14). NGS predominantly identified the genera Moraxella, Neisseria, Pasteurella, and Fusobacterium in oral cavity samples, whereas enterobacteria and staphylococci were prevalent in nail bed samples. In addition, the genera Capnocytophaga and Bartonella were identified, which have been described in serious human infections secondary to feline aggressions. Levofloxacin, marbofloxacin, and amoxicillin/clavulanic acid were the most effective drugs against the main groups of bacteria identified. Multidrug resistance was observed in 17% of the bacterial isolates. Furthermore, three staphylococci harboring the methicillin resistance gene mecA were identified. We highlight the complexity of microorganisms inhabiting the oral/claw microbiotas of cats, the high resistance rate of the isolates to conventional antimicrobial agents, and the zoonotic risk of aggressions caused by bites and scratches from domestic cats.

Antimicrobial resistance patterns of Staphylococcus spp. isolated from clinical specimens of companion animals in Northern Portugal, 2021-2023.

Staphylococcus spp. are growing pathogens in humans and companion animals. The emergence of multidrug-resistant bacterial infections, such as methicillin-resistant Staphylococcus-associated infections, due to zoonotic transmission, is a major public health concern. Domestic animals, such as dogs and cats, are possible reservoirs of multi-resistant bacterial species, which makes it relevant to monitor them due to their proximity to humans. However, there is a lack of information on the real scenario in Europe, especially in Portugal, particularly for animal infections caused by Staphylococcus spp. Therefore, this study aimed to investigate the antimicrobial resistance profile of Staphylococcus spp. isolated from cats and dogs diagnosed with infection in Northern Portugal. During 2021-2023, 96 Staphylococcus isolates from dogs and cats with symptoms of bacterial infection, including animals being treated in veterinary clinics/hospitals and cadavers submitted for necropsy at INIAV were included in the study collection. Of the 96 isolates, 63 were from dogs and 33 were Staphylococcus spp. from cats, most of which were isolated from ear (57% and 18%, respectively), skin (19 % and 27 %, respectively) and respiratory tract infections (6 % and 27 %, respectively). Among all the isolates, 12 different Staphylococcus spp. were identified, with Staphylococcus pseudintermedius being the most identified (61 % from dogs and 30 % from cats). It is noteworthy that 36 % of the isolates were multi-drug resistant and 25 % of the isolates showed a methicillin-resistant phenotype, with the mecA gene having been identified in all these isolates. This study highlights a high occurrence of multidrug-resistant Staphylococcus spp. in companion animals in Northern Portugal. This underlines the potential for cats and dogs to act as reservoirs of antimicrobial resistance, that can be transmitted to humans, posing a serious threat to public health.

Screening of antibiogram, virulence factors, and biofilm production of Staphylococcus aureus and the bio-control role of some probiotics as alternative antibiotics.

Background: Food safety is a serious challenge in the face of increasing population and diminishing resources. Staphylococcus aureus is a critical foodborne pathogen characterized by its capability to secret a diverse range of heat-resistant enterotoxins. Antibiotic usage in dairy herds resulted in the occurrence of antimicrobial resistance (AMR) patterns among bacterial species, which were consequently transmitted to humans via dairy products. Lactic acid bacteria (LAB) produce bacteriocins, which provide an excellent source of natural antimicrobials with the further advantage of being environmentally friendly and safe. Aim: Detection of multidrug resistance (MDR) S. aureus isolates in concerned samples, molecular characteristics, biofilm production, and the inhibitory role of LAB against it. Methods: Random samples of raw milk and other dairy products were analyzed for S. aureus isolation. Phenotypic and genotypic assessment of AMR was performed, in addition to detection of classical enterotoxin genes of S. aureus. Finally, evaluation of the antimicrobial action of some Lactobacillus strains against S. aureus. Results: Incidence rates of presumptive S. aureus in raw milk, Kariesh cheese, and yogurt samples were 50%, 40%, and 60%, respectively. The highest resistance of S. aureus was to Kanamycin (100%) and Nalidixic acid (89.3%), respectively. (78.66%) of S. aureus were MDR. 11.1% of S. aureus carried mecA gene. In concern with enterotoxins genes, PCR showed that examined isolates harbored sea with a percentage of (22.2%), while sed was found in (11.1%) of isolates. Regarding biofilm production, (88.88%) of S. aureus were biofilm producers. Finally, agar well diffusion showed that Lactobacillus acidophilus had the strongest antimicrobial action against S. aureus with inhibition zone diameter ranging from 18 to 22 mm. Conclusion: There is a widespread prevalence of MDR S. aureus in raw milk and dairy products. Production of staphylococcal enterotoxins, as well as biofilm production are responsible for public health risks. Therefore, installing proper hygienic routines and harsh food safety policies at food chain levels is substantial.

A novel extraction-free dual HiFi-LAMP assay for detection of methicillin-sensitive and methicillin-resistant Staphylococcus aureus.

Staphylococcus aureus (S. aureus) is a leading cause of bacteremia and blood stream infections. Methicillin-resistant S. aureus (MRSA) that first appeared in 1961 often caused hospital-acquired infections (HAIs) and community-acquired infections (CAIs) and was associated with high mortality rate. Accurate and rapid point-of-care testing (POCT) of MRSA is crucial for clinical management and treatment of MRSA infections, as well as the prevention and control of HAIs and CAIs. Here, we reported a novel extraction-free dual HiFi-LAMP assay for discriminative detection of methicillin-susceptible S. aureus and MRSA. The dual HiFi-LAMP assay can detect 30 copies/reaction of nuc and mecA genes with detection limits of 147 and 158 copies per 25 µL reaction, respectively. A retrospective clinical evaluation with 107 clinical S. aureus isolates showed both sensitivity and specificity of 100%. A prospective clinical evaluation with 35 clinical samples revealed a specificity of 100% and a sensitivity of 92.3%. The dual HiFi-LAMP assay can detect almost all S. aureus samples (141/142; 99.3%) within 20 min, implying that the entire HiFi-LAMP assay (including sample process) can be completed within 40 min, extremely significantly shorter than 3-5 days by the traditional clinical microbial culture and antibiotic susceptibility testing. The novel extraction-free dual HiFi-LAMP assay can be used as a robust POCT tool to promote precise diagnosis and treatment of MRSA infections in hospitals and to facilitate surveillance of MRSA at hospital and community settings.IMPORTANCEMethicillin-resistant Staphylococcus aureus (MRSA) was associated with high mortality rate and listed as a "priority pathogen" by the World Health Organization. Accurate and rapid point-of-care testing (POCT) of MRSA is critically required for clinical management and treatment of MRSA infections. Some previous LAMP-based POCT assays for MRSA might be questionable due to their low specificity and the lack of appropriate evaluation directly using clinical samples. Furthermore, they are relatively tedious and time-consuming because they require DNA extraction and lack multiplex detection capacity. Here, we reported a novel extraction-free dual HiFi-LAMP assay for discriminative detection of MRSA and methicillin-susceptible S. aureus. The assay has high specificity and sensitivity and can be completed within 40 min. Clinical evaluation with real clinical samples and clinical isolates showed excellent performance with 100% specificity and 92.3%-100% sensitivity. The novel extraction-free assay may be a robust POCT tool to promote precise diagnosis of MRSA infections and facilitate surveillance of MRSA at hospital and community settings.

Exonuclease-III Assisted the Target Recycling Coupling with Hybridization Chain Reaction for Sensitive mecA Gene Analysis by Using PGM.

In the field of neonatal infections nursing, methicillin-resistant Staphylococcus aureus (MRSA) is a major bacterial pathogen. Here, we present a portable biosensor for MRSA detection that is both highly sensitive and portable, owing to its implementation on the personal glucose meter (PGM) platform. The H probe was fixed on the magnetic bead for mecA gene analysis. A blunt 3' terminus appeared in the MBs-H probe when the mecA gene was present. Exonuclease-III (Exo-III) recognized the blunt terminus and cleaved it, freeing the mecA gene and so facilitating target recycling. In the meantime, the remaining H probe-initiated hybridization chain reaction (HCR) led to the desired signal amplification. Portable quantitative detection of mecA gene is possible because PGM can read the quantity of invertase tagged on HCR product. After optimizing several experimental parameters, such as the concentration of Exo-III and incubation time, the constructed sensor is extremely sensitive, with a detection limit of 2 CFU/mL. The results from this sensitive PGM-based sensor are in agreement with those obtained from plate counting methods, suggesting that it can be used to accurately assess the MRSA content in artificial clinical samples. In addition, the PGM sensor can significantly cut down on time spent compared to plate counting techniques. The manufactured sensor provides a promising option for accurate identification of pathogenic bacteria.

Prevalence and antibiogram of Pseudomonas aeruginosa and Staphylococcus aureus clinical isolates from burns and wounds in Duhok City, Iraq.

INTRODUCTION: this study aimed to isolate P. aeruginosa and S. aureus, investigate the antimicrobial resistance of collected isolates, and investigate the distribution of exoU and mecA genes in P. aeruginosa and S. aureus isolates. METHODOLOGY: Out of 150 samples, 32 isolates were identified as P. aeruginosa, 48 isolates were identified as S. aureus. All isolates were checked for AST. Then, a PCR was applied to detect exoU and mecA genes in P. aeruginosa and S. aureus. RESULTS: 12.0% and 29.3% of the samples showed co-isolates and single isolates of studied pathogens, respectively. Regarding burn samples, S. aureus was the most prevalent pathogen (38.0%, 38/100) among males (41.8%, 23/55), followed by P. aeruginosa (27.0%, 27/100) among females (28.9%, 13/45). The highest burn infection rates of S. aureus (50.0%) and P. aeruginosa (32.7%) were recorded among age groups (≥ 50) and (18-49), respectively. Comparatively, wound samples were less infected with these pathogens. P. aeruginosa isolates usually exhibited high resistance to gentamicin, tobramycin, and netilmicin, whereas, imipenem showed low resistance at 46.87%. S. aureus isolates were susceptible to trimethoprim-sulphamethoxazole and rifampin. 56.25% of P. aeruginosa isolates were exoU positive and 37.5% of S. aureus isolates were mecA positive. Results of the cefoxitin inhibition zone with mecA gene amplification, 33.3% isolates were MRSA, 4.2% isolates were nmrMRSA, and 62.5% isolates were MSSA. Most of the resistant isolates of P. aeruginosa carried the exoU gene, 80% resistant isolates to imipenem were exoU positive. CONCLUSIONS: S. aureus was more predominant than P. aeruginosa in burns and wounds infections.

A Study of Community-Acquired Pyodermas with Special Reference to Panton-Valentine Leukocidin (PVL)-Positive Methicillin-Resistant Staphylococcus Aureus.

Background: Community-acquired (CA) pyodermas are one of the most common infections encountered in the dermatology outpatient clinics. A significant number of these conditions are caused by Staphylococcus aureus. CA-methicillin-sensitive Staphylococcus aureus (MSSA) and CA-methicillin-resistant Staphylococcus aureus (MRSA) have specific virulence genes which are associated with these diseases, particularly the Panton-Valentine leukocidin (PVL) genes. The presence of the PVL gene as a virulence factor may be associated with recurrent and severe skin infections. Materials and Methods: A prospective study was conducted with 205 cases of CA pyodermas, of which five were discarded due to mixed isolates. Clinical details were taken and wound exudate was sent for bacteriological examination. Further, the molecular study was performed on all MRSA (7) isolates and 13 randomly selected MSSA isolates using polymerase chain reaction for mecA and PVL genes. Results: Staphylococcus aureus was the most common organism (90%) isolated from primary or secondary CA pyodermas. The prevalence of CA-MRSA among all pyodermas was 3.5% in our community. The PVL gene was not detected in all tested CA-MRSA and CA-MSSA isolates. Conclusion: While pyodermas are common, the prevalence of MRSA is low in the CA pyodermas in our region. PVL does not appear to be a virulence factor among the isolated MRSA. Larger, multicentric, and periodic studies are, however, required to further justify these claims.

A simple and sensitive colorimetric approach for mecA gene analysis via exonuclease-III catalyzed signal cascade.

Analysis of mecA gene in Staphylococcus aureus (S. aureus) is essential for controlling infections in intensive care units (ICU) and preventing the use of ineffectual empirical treatments. However, quantitative determination of the mecA gene remains difficult. Herein, we propose a simple and sensitive colorimetric approach by integrating exonuclease-III (Exo-III) assisted signal cascade and G-quadruplex/hemin DNAzymes (G4 DNAzymes) catalyzed 2,2'-azino-bis (3-ethylben-zothiazoline-6-sulfonic acid) (ABTS) based color reaction. In this method, signal amplification does not necessitate the use of complex experimental components, such as multiple enzymes and primer design, while still maintaining a high signal amplifying efficiency. Therefore, the method has a broad mecA gene detection range from 10 fM to 1 nM and a low limit of detection down to 3.4 fM level. Taking the merit of simplicity and high sensitivity, the approach is promising in analyzing mecA gene in S. aureus and diagnosing infections.

Therapeutic potential of natural products and antibiotics against bovine mastitis pathogen of cows and buffaloes.

The present study aims to evaluate the prevalence and antimicrobial sensitivity of Staphylococcus aureus associated with bovine mastitis to selected antibiotics and plant extracts. In the current study, 140 milk samples were collected from cows and buffaloes. Among the 140 samples, 93 samples were positive for sub-clinical mastitis based on the California Mastitis Test (CMT). Out of the total positive samples, 45 were confirmed for S. aureus on a Mannitol salt agar media. The antimicrobial susceptibility test revealed that 44.82% of the isolates were resistant to cefoxitin (oxacillin) confirming methicillin-resistant S. aureus (MRSA) with a higher percentage (51.61%) in the buffalo than in the cow samples. Furthermore, the PCR assay confirmed the presence of the mecA gene in all the MRSA isolates. Among the seven tested antibiotics, sulfamethoxazole + trimethoprim showed high efficacy (71.1%) against methicillin-susceptible S. aureus isolates (MSSA). Oxytetracycline and sulfamethoxazole + trimethoprim showed 20% efficacy against MRSA followed by enrofloxacin (10%). On the other hand, the tested samples from Pistacia chinensis revealed that the ethyl acetate extract of bark showed a maximum zone of inhibition of 21.3 mm against MSSA and MRSA isolates at 3 000 µg/disc. Moreover, the methanol extract of Cotoneaster microphyllus formed a 12.3 mm and 9.1 mm zone of inhibition against the MSSA and MRSA isolates, respectively.

New synergistic antibacterial mechanism of bulky mixed Ti/w hetero-polyoxometalates composed of multi lacunary Keggin structure with oxacillin against vancomycin intermediate-resistant Staphylococcus aureus.

Considering the lack of detailed research on the antibacterial mechanism of polyoxometalates, we examined the synergistic effect of novel bulky mixed Ti/W hetero-polyoxometalates (K9.5H2.5 ï¼»α-Ge2Ti4W20O78］・ 29H2O; αTi4, K9H5 ï¼»α-Ge2Ti6W18O77］・16H2O; αTi6, K23H5ï¼»α-Ge4Ti12W36O154］・39H2O; αTi12, K9H5 ï¼»ß-Ge2Ti6W18O77］・ 45H2O; ßTi6) with the antibiotic oxacillin against vancomycin intermediate-resistant Staphylococcus aureus (VISA) using fractional inhibitory concentration (FIC) index and growth curve in this study. All polyoxometalates used in this study showed remarkable synergistic effects with oxacillin. Its synergistic antibacterial mechanism was examined using reverse transcription PCR (RT-PCR) and penicillin binding protein-2' (PBP2') latex agglutination test. The results suggested that these polyoxometalates did not inhibit mecA gene transcription but resulted in PBP2' protein malfunction. From these results, we concluded that the substances producing resistance in VISA were affected by polyoxometalates depending on their molecular size, facilitating a synergistic antibacterial effect with oxacillin.

Detection of mecA positive staphylococcal species in a wastewater treatment plant in South Africa.

We investigated the prevalence of antibiotic resistant staphylococci and detection of resistant, virulence, and Spa genes in a South African wastewater treatment plant. Species identified were Staphylococcus aureus, S. lentus, S. arlettae, S. cohnii, S. haemolyticus, S. nepalensis, S. sciuri (now Mammaliicoccus sciuri), and S. xylosus. Isolates showed high resistance to methicillin (91%), ampicillin (89%), ciprofloxacin (86%), amoxycillin (80%), ceftazidime (74%), and cloxacillin (71%). Multiple antibiotic resistance (MAR) index for the isolates exceeded 0.2 (0.50-0.70). Among the isolates, 77% were mecA-positive. All S. aureus strains were positive for nuc and 7 Spa gene types. The present study highlights possibility of treated wastewaters being potential reservoir for antibiotic-resistant staphylococci. This is a cause for concern as wastewater effluents are decanted into environmental waters and these are, in many cases, used for various purposes including recreation (full contact), religious (full body submersion), and drinking water for some rural communities and water for livestock.

A double-quenching paperclip ECL biosensing platform for ultrasensitive detection of antibiotic resistance genes (mecA) based on Ti3C2 MXene-Au NPs as a coreactant accelerator.

The global spread of environmental biological pollutants, such as antibiotic-resistant bacteria and their antibiotic resistance genes (ARGs), has emerged as a critical public health concern. It is imperative to address this pressing issue due to its potential implications for public health. Herein, a DNA paperclip probe with double-quenching function of target cyclic cleavage was proposed, and an electrochemiluminescence (ECL) biosensing platform was constructed using Ti3C2 MXene in-situ reduction growth of Au NPs (TCM-Au) as a coreactant accelerator, and applied to the sensitive detection of ARGs. Thanks to the excellent catalytic performance, large surface area and Au-S affinity of TCM-Au, the ECL performance of CdS QDs have been significantly improved. By cleverly utilizing the negative charge of the paperclip nucleic acid probe and its modification group, double-quenching of the ECL signal was achieved. This innovative approach, combined with target cyclic amplification, facilitated specific and sensitive detection of the mecA gene. This biosensing platform manifested highly selective and sensitive determination of mecA genes in the range of 10 fM to 100 nM and a low detection limit of 2.7 fM. The credible detectability and anti-interference were demonstrated in Yangtze river and Aeration tank outlet, indicating its promising application toward pollution monitoring of ARGs.

Dual-Toehold-Probe-Mediated Exonuclease-III-Assisted Signal Recycles Integrated with CHA for Detection of mecA Gene Using a Personal Glucose Meter in Skin and Soft Tissue Infection.

Staphylococcus aureus integrated with mecA gene, which codes for penicillin-binding protein 2a, is resistant to all penicillins and other beta-lactam antibiotics, resulting in poor treatment expectations in skin and soft tissue infections. The development of a simple, sensitive and portable biosensor for mecA gene analysis in S. aureus is urgently needed. Herein, we propose a dual-toehold-probe (sensing probe)-mediated exonuclease-III (Exo-III)-assisted signal recycling for portable detection of the mecA gene in S. aureus. When the target mecA gene is present, it hybridizes with the sensing probe, initiating Exo III-assisted dual signal recycles, which in turn release numerous "3" sequences. The released "3" sequences initiate catalytic hairpin amplification, resulting in the fixation of a sucrase-labeled H2 probe on the surface of magnetic beads (MBs). After magnet-based enrichment of an MB-H1-H2-sucrase complex and removal of a liquid supernatant containing free sucrase, the complex is then used to catalyze sucrose to glucose, which can be quantitatively detected by a personal glucose meter. With a limit of detection of 4.36 fM for mecA gene, the developed strategy exhibits high sensitivity. In addition, good selectivity and anti-interference capability were also attained with this method, making it promising for antibiotic tolerance analysis at the point-of-care.

Microbial Complexity of Oral Cavity of Healthy Dogs Identified by Mass Spectrometry and Next-Generation Sequencing.

The high complexity of the oral microbiota of healthy dogs and the close exposure of humans to companion animals represent a risk of the transmission of potential zoonotic microorganisms to humans, especially through dog bites, including multidrug-resistant ones. Nonetheless, a limited number of comprehensive studies have focused on the diversity of the microorganisms that inhabit the oral cavities of healthy dogs, particularly based on modern molecular techniques. We investigated bacterial and fungal organisms in the oral cavities of 100 healthy dogs based on a combination of conventional and selective microbiological culture, mass spectrometry (MALDI-TOF MS), and next-generation sequencing. In addition, in vitro antimicrobial susceptibility patterns of isolates and mecA resistance gene were assessed. A total of 213 bacteria and 20 fungi were isolated. Staphylococcus pseudintermedius (40/100 = 40%), α-hemolytic Streptococcus (37/100 = 37%), and Pasteurella stomatis (22/100 = 22%) were the most prevalent bacteria diagnosed by microbiological culture and MALDI-TOF MS, whereas Aspergillus (10/100 = 10%) was the most common fungi identified. Based on next-generation sequencing of selected 20 sampled dogs, Porphyromonas (32.5%), Moraxella (16.3%), Fusobacterium (12.8%), Conchiformibius (9.5%), Bergeyella (5%), Campylobacter (3.8%), and Capnocytophaga (3.4%) genera were prevalent. A high multidrug resistance rate was observed in Staphylococcus pseudintermedius isolates, particularly to azithromycin (19/19 = 100%), penicillin (15/19 = 78.9%), and sulfamethoxazole/trimethoprim (15/19 = 78.9%). In addition, the mecA resistance gene was detected in 6.1% (3/49) of coagulase-positive staphylococci. Here, we highlight the microbial complexity of the oral mucosa of healthy dogs, including potential zoonotic microorganisms and multidrug-resistant bacteria, contributing with the investigation of the microbiota and antimicrobial resistance patterns of the microorganisms that inhabit the oral cavity of healthy dogs.

Transesophageal Echocardiography vs. Transthoracic Echocardiography for Methicillin-Sensitive Staphylococcus aureus and Methicillin-Resistant Staphylococcus aureus Endocarditis.

Infective endocarditis is an infection of the inner layers of the heart, seen often in intravenous drug users and patients with valvular lesions or prosthetic heart valves. This entity has high mortality and morbidity. The most common causative microorganism is Staphylococcus aureus. In this comprehensive literature review, we focused on both Staphylococcus aureus infections, i.e., methicillin-sensitive Staphylococcus aureus (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA) endocarditis, demographics, use of transthoracic echocardiogram and/or transesophageal echocardiogram for diagnostics, and treatments. Although clinical criteria are relevant, transesophageal echocardiogram plays a vital role in establishing and identifying the presence of infective endocarditis and its local complications, with higher sensitivity in patients with prosthetic valves. The antibiotic selection posed a great challenge for clinicians due to antibiotic resistance and the aggressive nature of Staphylococcus aureus. Early diagnosis of infective endocarditis, when suspected, and effective management by a multispecialty team can improve the outcome for the patients.

Molecular identification, antimicrobial resistance and virulence gene profiling of Staphylococcus spp. associated with bovine sub-clinical mastitis in Bangladesh.

This study was conducted to investigate the diversity and antimicrobial resistance profiling of Staphylococcus species causing sub-clinical mastitis (SCM) in dairy herds in Bangladesh as well as putative risk factors associated with the infections. Individual quarter milk samples were collected from a total of 284 lactating cows from 30 dairy farms were screened by means of California mastitis test; 178 (62.7%) of them had at least of quarter affected by SCM. After conventional microbiological isolation procedures, PCR tests were used for Staphylococcus species identification and detection of antimicrobial resistance and virulence genes. S. chromogenes (65.7%) was the most predominant species followed by, S. epidermidis (20.2%), S. haemolyticus (19.1%), S. aureus (15.7%), and S. sciuri (5.6%). High levels of antimicrobial resistance to ampicillin and amoxicillin/clavulanic acid were observed in S. aureus (82.1% and 75%) and S. sciuri (80% and 70%), while resistance to cefepime was markedly higher in S. chromogenes (95.7%), S. haemolyticus (94.1%), and S. epidermidis (97.2%). Multidrug resistance isolates were identified in all five species. The mecA gene was detected in S. aureus (32.1%) and S. chromogenes (5.98%). In addition, 20% S. sciuri and 17.7% S. haemolyticus carried the cytotoxin (pvl) gene, while 14.3% S. aureus harbored the toxic shock syndrome toxin (tst) gene. Multivariable logistic regression analysis identified "Old aged" (OR [CI]: 3.5 [1-12.4]); "Early stage of lactation" (OR [CI]: 3.4 [1.2-9.7]) and, "Firm udder condition" (OR [CI]: 4.2 [1.2-14.6]) as risk factors associated with SCM caused by S. aureus, S. chromogenes, and S. haemolyticus, respectively. Moreover, "Use of antimicrobials" (OR [CI]: 10.4 [3.4-32.1] and "History of previous clinical mastitis" (OR [CI]: 4.9 [1.2-19.7] for the carriage of methicillin-resistant Staphylococcus spp.

Methicillin-Resistant Staphylococcusaureus Strains Isolated from Burned Patients in a Tunisian Hospital: Molecular Typing, Virulence Genes, and Antimicrobial Resistance.

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the major causes of a variety of infections in hospitals and the community. Their spread poses a serious public health problem worldwide. Nevertheless, in Tunisia and other African countries, very little molecular typing data on MRSA strains is currently available. In our study, a total of 64 MRSA isolates were isolated from clinical samples collected from burned patients hospitalized in the Traumatology and Burns Center of Ben Arous in Tunisia. The identification of the collection was based on conventional methods (phenotypic and molecular characterization). The characterization of the genetic support for methicillin resistance was performed by amplification of the mecA gene by polymerase chain reaction (PCR), which revealed that 78.12% of S. aureus harbors the gene. The resistance of all the collection to different antibiotic families was studied. Indeed, the analysis of strain antibiotic susceptibility confirmed their multi-resistant phenotype, with high resistance to ciprofloxacin, gentamicin, penicillin, erythromycin, and tetracycline. The resistance to the last three antibiotics was conferred by the blaZ gene (73.43%), the erm(C) gene (1.56%), the msr(A) gene (6.25%), and tet(M) gene (7.81%), respectively. The clonal diversity of these strains was studied by molecular typing of the accessory gene regulator (agr) system, characterization of the SCCmec type, and spa-typing. The results revealed the prevalence of agr types II and III groups, the SCCmec type III and II cassettes, and the dominance of spa type t233. The characterization of the eight enterotoxins genes, the Panton-Valentine leukocidin and the toxic shock syndrome toxin, was determined by PCR. The percentage of virulence genes detected was for enterotoxins (55%), tst (71.88%), leukocidin E/D (79.69%), and pvl (1.56%) factors. Furthermore, our results revealed that the majority of the strains harbor IEC complex genes (94%) with different types. Our findings highlighted the emergence of MRSA strains with a wide variety of toxins, leukocidin associated with resistance genes, and specific genetic determinants, which could constitute a risk of their spread in hospitals and the environment and complicate infection treatment.

Effectiveness of raw bacteriocin produced from lactic acid bacteria on biofilm of methicillin-resistant Staphylococcus aureus.

Background and Aim: Probiotics are proven beneficial to health since they enhance immunity against dangerous pathogens and increase resistance to illness. Bacteriocin produced by lactic acid bacteria (LAB), demonstrates a broad inhibitory spectrum and therapeutic potential. This study aimed to isolate LAB-producing bacteriocin and investigate the effect of crude bacteriocin on biofilm from methicillin-resistant Staphylococcus aureus (MRSA). Materials and Methods: This study used randomly collected 80 white soft local cheeses (40 each from cows and sheep) from different supermarkets in Basrah Province. The obtained samples were cultured and the bacterial suspension of S. aureus was prepared at 1.5 × 108 cells/mL. The crude bacteriocin extracted from LAB was obtained, and the tube was dried and inverted to detect the biofilm loss at the bottom. Results: There were 67 (83.75%) LAB isolates. Among 40 milk samples collected directly and indirectly, there were 36 (83.33%). Staphylococcus aureus isolates based on conventional bacteriological analysis and biochemical tests. Molecular testing was conducted to identify LAB and MRSA. Depending on genotypic results, the effect of white soft local cheese (cows and sheep) and the amplification results of the 16S rRNA gene were detected in 46 LAB isolates from white soft local cheese from cows and sheep. Based on the molecular identification of the mecA, results on Staphylococcus determined that only 2 of 36 isolates of S. aureus carried the mecA. Moreover, there were 26 (86.66%) isolates (MRSA) from samples of raw milk from local markets and subclinical mastitis in cows. The ability of LAB isolates was tested. The effects of bacteriocin production on preventing biofilm growth and formation were investigated. Results demonstrated that bacteriocin has high activity. Microtiter plates applied to investigate the ability of S. aureus to produce biofilms revealed that all isolates were either weak or moderate biofilm producers, with neither non-biofilm nor strong biofilm producers found among the tested isolates. Conclusion: Lactic acid bacteria demonstrate a high ability to produce bacteriocin. Crude bacteriocin from LAB has a restrictive effect on biofilms produced by MRSA; thus, it can be used to reduce the pathogenicity of this bacterium.

Synergistic Antimicrobial Activity of Ceftriaxone and Polyalthia longifolia Methanol (MEPL) Leaf Extract against Methicillin-Resistant Staphylococcus aureus and Modulation of mecA Gene Presence.

Medicinal plants are an essential source of traditional curatives for numerous skin diseases. Polyalthia longifolia (Sonn.) Thwaites (Annonaceae family) is a medicinal plant used to cure skin illnesses. P. longifolia is usually applied in folkloric therapeutical systems to treat skin diseases. The methicillin-resistant Staphylococcus aureus (MRSA) bacteria is among the essential bacteria contributing to skin diseases. Hence, to verify the traditional medicinal claim of P. longifolia usage in skin disease treatment, the current research was performed to study the synergistic antibacterial activity of standardized Polyalthia longifolia methanol leaf extract (MEPL) against MRSA bacteria. The synergistic antimicrobial activity result of ceftriaxone, when mixed with MEPL, against MRSA was investigated by the disc diffusion method, broth microdilution method, checkerboard dilution test, and modulation of mecA gene expression by multiplex polymerase chain reaction (multiplex PCR). The MEPL extract exhibited good synergistic antimicrobial activity against MRSA. Using the checkerboard method, we confirmed the synergistic effect of MEPL from P. longifolia and ceftriaxone (2:1) for MRSA with a marked reduction of the MIC value of the ceftriaxone from 8000 µg/mL to 1000 µg/mL. Moreover, the combination of MEPL with ceftriaxone significantly (p < 0.05) inhibited the presence of the resistant mecA gene in the tested strain. The LC-ESI-MS/MS analysis identified compounds that were reported to exhibit antimicrobial activity. Conclusively, the MEPL extract, an important etiological agent for skin diseases, showed worthy synergistic antimicrobial action against MRSA bacteria, thus supporting the traditional use of P. longifolia.

Antibiotic resistance and mecA characterization of Staphylococcus hominis from filarial lymphedema patients in the Ahanta West District, Ghana: A cross-sectional study.

Background and Aim: Filarial infections affect over 150 million people in the tropics. One of the major forms of filarial pathologies is lymphedema; a condition where the immune response is significantly altered, resulting in changes in the normal flora. Staphylococcus hominis, a human skin commensal, can also be pathogenic in immunocompromised individuals. Therefore, there is the possibility that S. hominis could assume a different behavior in filarial lymphedema patients. To this end, we investigated the levels of antibiotic resistance and extent of mecA gene carriage in S. hominis among individuals presenting with filarial lymphedema in rural Ghana. Method: We recruited 160 individuals with stages I-VII lymphedema, in a cross-sectional study in the Ahanta West District of the Western Region of Ghana. Swabs from lymphedematous limb ulcers, pus, and cutaneous surfaces were cultured using standard culture-based techniques. The culture isolates were subjected to Matrix-Assisted Laser Desorption/Ionization Time of Flight (MALDI-TOF) mass spectrometry for bacterial identification. Antimicrobial susceptibility testing (AST) was performed using the Kirby-Bauer method. mecA genes were targeted by polymerase chain reaction for strains that were cefoxitin resistant. Results: In all, 112 S. hominis were isolated. The AST results showed resistance to chloramphenicol (87.5%), tetracycline (83.3%), penicillin (79.2%), and trimethoprim/sulphamethoxazole (45.8%). Of the 112 strains of S. hominis, 51 (45.5%) were resistant to cefoxitin, and 37 (72.5%) of the cefoxitin-resistant S. hominis haboured the mecA gene. Conclusion: This study indicates a heightened level of methicillin-resistant S. hominis isolated among filarial lymphedema patients. As a result, opportunistic infections of S. hominis among the already burdened filarial lymphedema patients in rural Ghana may have reduced treatment success with antibiotics.