Coagulase-positive Staphylococci Isolates Research Articles

Prevalence of Staphylococcus argenteus among food handlers, kitchen utensils, and food samples in Japan.

Staphylococcus argenteus has received increased attention from an aspect of food safety since several food poisoning outbreaks caused by the bacterium were reported in Japan. However, S. argenteus prevalence among food handlers and utensils has not yet been investigated. In this study, we investigated S. argenteus prevalence among a collection of coagulase-positive staphylococci (CPS) that were isolated during food sanitary inspections in Japan. Out of a total of 191 CPS isolates, 14 were identified as S. argenteus. One was isolated from shelled shrimp, nine were isolated from food handlers' hand swabs, and four were isolated from kitchen utensils. Whole-genome sequencing revealed that transmission of S. argenteus from human hands to utensils was possible. Though all 14 isolates were negative for the pvl and tst-1 genes, 6 harbored the seb gene. Only 21.4% of S. argenteus isolates were resistant to antibiotics, while 62.1% of the S. aureus isolates from the same sources were confirmed to be resistant. To the best of our knowledge, this is the first report to demonstrate possible transmission of S. argenteus from food handlers to utensils in food-processing environments.

Molecular differentiation of coagulase-positive staphylococcal isolates carrying mecA- and PVL-encoding genes among healthy males

Molecular differentiation of coagulase-positive staphylococcal isolates carrying mecA- and PVL-encoding genes among healthy males

Prevalence of mecA, mecC and Panton-Valentine-Leukocidin Genes in Clinical Isolates of Coagulase Positive Staphylococci from Dermatological Canine Patients.

Coagulase positive Staphylococci (CoPS) are the leading cause of canine cutaneous and otic infections. Virulence factors associated with Staphylococci include the expression of mec and panton-valentine leukocidin (pvl) genes. Methicillin-resistance (MR) is commonly associated with mecA gene expression, although a recently identified variant, mecC, has been reported. This study aims to evaluate the prevalence of mecA, mecC and pvl genes in 232 clinical isolates of CoPS collected from dogs with pyoderma. A multiplex PCR, and Kirby-Bauer disk diffusion susceptibility test for cefoxitin was performed for all isolates. PBP2a agglutination test was performed on 127 isolates. Standard MRSA isolates were used as positive controls. The mecA gene was identified in 149/232 isolates (64.2%): 116 S. pseudintermedius, 30 S. coagulans and three S. aureus. The pvl gene was present in only 1 isolate of S. pseudintermedius (0.4%), whereas no isolates carried the mecC gene. 34 isolates were resistant to cefoxitin (14.6%) and they were all mecA positive. The results of this study show an MR prevalence of 64.2% confirming concerns about antibiotic resistance in veterinary medicine. In conclusion, this is the first study analyzing the prevalence of mecC and pvl in comparison to mecA, in a large cohort of CoPS clinical isolates from dogs with pyoderma. A multimodal surveillance on the prevalence of mecC and pvl in veterinary medicine is essential to appropriate antimicrobial management.

Enterotoxin- and Antibiotic-Resistance-Encoding Genes Are Present in Both Coagulase-Positive and Coagulase-Negative Foodborne Staphylococcus Strains

Food poisoning by staphylococcal enterotoxins (SE) is a major cause of foodborne illness, often associated with coagulase-positive staphylococci (CPS). The increase in the number of methicillin-resistant Staphylococcus aureus (MRSA) strains is another major problem associated with CPS. However, reports of the association of SE and methicillin-resistant Staphylococcus with coagulase-negative staphylococci (CNS) are beginning to re-emerge. In this context, the aim of this study is to investigate the presence of staphylococcal enterotoxin genes and to characterize the phenotypic and genotypic antimicrobial resistance in 66 isolates of Staphylococcus spp. (47 CNS and 19 CPS) recovered from ready-to-eat (RTE) street food sold in Maputo, Mozambique. Seven virulence genes encoding SE (sea, seb, sec, sed and see) and two toxins (hlb and sak) were screened by multiplex PCR (MPCR). Antimicrobial resistance against 12 antibiotics was evaluated by the disk diffusion method. The presence of genes encoding resistance to penicillin, methicillin, vancomycin and erythromycin (blaZ, mecA, vancA, vancB, ermA, ermB and ermC) were also screened by PCR. At least one of the seven virulence genes assessed in this study was detected in 57.9% and 51% of CPS and CNS isolates, respectively. In CPS isolates, the most frequent gene was hlb (47.4%), followed by sec (15.8%) and sea, seb and sed genes with 5.3% each. In CNS isolates, the most frequent gene was sec (36.2%) followed by sak (17%), hlb (14.9%), sed (12.8%) and seb (6.4%). Five of the twelve CPS in which virulence genes were detected were also antibiotic-resistant. All the CNS isolates harboring virulence genes (n = 27, 57.4%) were antimicrobial-resistant. The prevalence of multidrug resistance was higher (59.6%) in CNS than in CPS (26.3%) isolates. Regarding the presence of antibiotic-resistance genes, blaZ (penicillin-resistant) was the most frequent in both CPS (42.1%) and CNS (87.2%), followed by the mecA (encoding methicillin resistance) and vancA genes (vancomycin-resistant), which represented 36.8% and 31.6% in CPS isolates and 46.8% in CNS isolates, respectively. The prevalence of vancomycin-resistant staphylococci has been increasing worldwide and, to our knowledge, this is the first study to report the occurrence of vancomycin-resistant staphylococci in Mozambique. These results emphasize the need to investigate CNS isolates in parallel with CPS, as both constitute public health hazards, given their potential to produce SE and spread antimicrobial resistance genes.

Prevalence and antimicrobial resistance profiling of Staphylococcus aureus isolated from traditional cheese in Yunnan, China.

The online version contains supplementary material available at 10.1007/s13205-021-03072-4.

A second-order Monte Carlo simulation model to describe coagulase-positive Staphylococci growth in mayonnaise served in Algerian pizzerias

To bridge the data gap on food poisoning caused by coagulase-positive staphylococci (CoPS), especially related to mayonnaise sauce served at Algerian pizzerias, this study aimed to assess the concentration of CoPS in mayonnaise and the probability of exceeding a critical concentration of ≥ 5 log CFU/g. The city of Ain Témouchent in West Algeria was taken as a case study.A probabilistic assessment model was built, taking into account the initial contamination in freshly made mayonnaise and the potential growth before serving. Uncertainty and variability were integrated separately in the model. Uncertainty came from lack of data and model fitting error, variability from natural heterogeneity of biological materials (e.g., microbial strains) and temperature during cold storage.The second-order Monte Carlo procedure was implemented in R using the mc2d package. The following pieces of data were generated to populate the model: CoPS were enumerated and characterized from 57 samples of mayonnaise served at pizzeria in Ain Témouchent city; challenge tests at 23 °C were performed in mayonnaise using three CoPS isolates. The following existing data were also gathered: meteorological data from Ain Témouchent were analysed to build a realistic scenario of storage, while a set of 43 and 35 values of the minimal and maximal growth limits of CoPS, respectively, were collected from the literature and analysed to inform a secondary predictive model describing the growth rate at various storage temperature conditions. A sensitivity analysis was performed to facilitate the interpretation of the results.The results revealed a CoPS prevalence in freshly made mayonnaise of 25% [15%; 37%] with concentrations varying from 0.4 [0.3; 0.9] to 2.9 [2.4; 3.0] log CFU/g. The growth rates at 23 °C, based on challenge tests in mayonnaise, had a median value estimated to be 1.41 [1.17; 1.65] h − 1.Concentration levels according to various scenarios of temperature and serving conditions were calculated. For instance, the median contamination of CoPS in mayonnaise after storage in a refrigerated display counter for 14 h during the hottest months of the year was estimated to be zero. However, the 95th percentile was estimated to be 3.6 [2.9; 4.2] log CFU/g. In this scenario, the probability of exceeding a critical concentration of ≥ 5 log CFU/g was estimated to be 1% [0.3%; 2%], which is low but not negligible.These findings could be used to improve food safety policies and develop a risk management strategy to reduce the food poisoning associated with the consumption of ready-to-use foods in Algerian fast food restaurants.

Presence and growth prediction of Staphylococcus spp. and Staphylococcus aureus in Minas Frescal cheese, a soft fresh cheese produced in Brazil

Physical-chemical characteristics of Minas Frescal cheese (MFC) favor the growth of Staphylococcus spp. and allow the production of enterotoxins by specific strains. Here, we aimed to characterize the physical-chemical aspects (pH, storage temperature, and salt content) and the presence of Staphylococcus spp. in MFC samples (n = 50) to support a modeling study for the growth by this microorganism. Coagulase-positive staphylococci isolates were obtained and subjected to PCR assays to identify them as Staphylococcus aureus (nuc) and to detect staphylococcal enterotoxin-related genes (sea, seb, sec, sed, see). Staphylococcus aureus growth kinetics (maximum growth rate, Grmax, and lag time) were predicted based on ComBase model and MFC physical-chemical aspects. Mean counts of Staphylococcus spp. ranged from 3.3 to 6.7 log cfu/g, indicating poor hygiene practices during production. Selected isolates (n = 10) were identified as S. aureus, but none presented classical enterotoxin-related genes. pH, temperature, and salt content ranged from 5.80 to 6.62, 5°C to 12°C, and 0.85% to 1.70%, respectively. The Grmax values ranged from 0.012 to 0.419 log cfu/g per h. Independent of the storage temperature, the lowest Grmax values (0.012 to 0.372 log cfu/h) were obtained at pH 5.80 associated with salt content of 1.7%; independent of the pH and salt content, the best temperature to avoid staphylococcal growth was 7.5°C. Hygienic conditions during MFC production must be adopted to avoid staphylococcal contamination, and storage at temperatures lower than 7.5°C can prevent staphylococcal growth and the potential production of enterotoxins.

DETECTION OF ANTIBIOTICS RESISTANCE GENES IN STAPHYLOCOCCUS AUREUS ISOLATED FROM POULTRY FARMS

Poultry industry in Egypt was subjected to various problems one of them was the early chick mortalities that cause great economic losses and its investigation is the request of many poultry scientists and men. In poultry staphylococci, including Staphylococcus aureus are known to cause various diseases from acute septicemia to chronic osteomyelitis. Two hundred and sixty samples of chickens, ducks and turkeys were collected from different poultry farms. A survey on Staphylococcal infection among chickens, ducks and turkeys was carried out. It was found that out of 260 infected and dead chickens, ducks and turkeys suspected to be infected with Staphylococcosis, 54 cases (20.76%) revealed Staphylococcus micro-organisms. The isolated strains were typed as: Staphylococcus aureus (74.07%), Staphylococcus epidermidis (14.8 %)and Staphylococcus saprophyticus (11%). The results of sensitivity biogram revealed that S. aureus were highly sensitive to Amoxicillin, Ampicillin, Ciprofloxacine, Enrofloxacine and lincomycin and resistant to Erythromycin, Streptomycine and Chloramphenicol. The results of MIC of 6 representative coagulase positive Staphylococci isolates against 6 selected antibiotics commonly used in poultry farms showed that 100% of isolates were sensitive to amoxicillin, enrofloxacin, ciprofloxacin and oxytetracyclin, while 100% of 6 isolates were resistant to streptomycin and lincomycin. The incidence of isolation of Staphylococci from the internal organs of examined birds and from unabsorbed yolk sac, joints, liver, intestinal content and heart blood was 36.3%, 20%, 19.2%, 17.3% and 16.9% respectively. Ten of each detected isolates were examined by cPCR for resistance genes blaZ and aac(6’) aph (2’’). The isolates harbored these resistance genes with percentage of 100% for S. aureus. In experiment, the pathogenicity of the isolated strains of S. aureus for 7 day- old chicks was studied. S. aureus injected subcutaneously, oral and intranasal caused death of 100%, 100% and 26.7% of the used chicks respectively.

Microbial safety status of Serro artisanal cheese produced in Brazil

Considering the growing consumption of artisanal foods worldwide, we aimed to evaluate the microbial safety of Serro artisanal cheese (SAC), produced in Minas Gerais State, Brazil. This cheese is produced with raw milk using 1 of 2 natural starter cultures: "pingo" and "rala." A total of 53 SAC samples (pingo = 8; rala = 45) were obtained from different farmers and subjected to conventional and molecular assays to detect and enumerate Listeria monocytogenes, Salmonella spp., coagulase-positive staphylococci (CPS), diarrheagenic Escherichia coli, Mycobacterium tuberculosis, and Brucella abortus. The SAC samples were also subjected to an ELISA to detect classical staphylococcal enterotoxins (CSE: SEA, SEB, SEC, SED, SEE) and to PCR assays to detect staphylococcal enterotoxin-related genes (sea, seb, sec, sed, see). Coagulase-positive staphylococci isolates were obtained and tested by the same assays to detect their potential in CSE production and presence of CSE-related genes. None of the SAC samples showed any of the screened food-borne pathogens and zoonotic agents, and none showed the presence of CSE by phenotypic and genotypic approaches. Despite the absence of microbial hazards, mean counts of CPS in SAC samples were 5.2 log cfu/g (pingo starter) and 4.6 log cfu/g (rala starter), indicating poor hygiene practices during production. None of the tested CPS isolates (n = 116) produced CSE or presented CSE-related genes. Despite the relative microbial safety, hygienic conditions during SAC production must be improved to meet official guidelines established in Brazil.

Staphylococcus aureus and Staphylococcal enterotoxin detection in raw milk and cheese origin coagulase positive isolates

In the present study, a total 110 (60 cheese and 50 cows’ raw milk) samples was analyzed for CPS according to Food Drug Administration (FDA, 2001) previously. After the isolation, to confirmation of the isolates, catalase test, microscopic examination, coagulase test in tube and glucose-mannitol fermentation tests were applied. After the tests, we obtained 97 CPS isolates. The 97 CPS isolates were used as a material. For the confirmation of the isolates being Staphylococci species, 16S rRNA was detected by using PCR assay. For the detection of the CPS isolates being S. aureus, nuc gene detected in the CPS isolates using PCR assay. To determination of the staphylococcal enterotoxin, PCR assay was applied. The 16 S rRNA was detected in a total 97 (35 milk origin and 62 cheese origin) isolates. Therefore, these isolates were evaluated as coagulase-positive staphylococci (CPS). The nuc gene was detected in 50 out of 97 CPS isolates. So, the 50 (18 cheese origin and 32 milk origin) isolates were evaluated as S. aureus. However, none of the staphylococcal enterotoxin genes (SE A,B,C,D,E,G,H,I,J) was detected in 97 CPS or 50 S. aureus isolates.

Extended-spectrum beta-lactamase and Class 1 integrons in multidrug-resistant Escherichia coli isolated from turkeys.

Aim:This study aimed to investigate the prevalence and implication of extended-spectrum beta-lactamase (ESBL) producing and Class 1 integrons (int1) gene carriers Escherichia coli isolates that demonstrated multidrug resistance (MDR) phenotypes and was isolated from turkeys that suffered from respiratory manifestations.Materials and Methods:A total of 120 freshly dead turkey poults that suffered from respiratory manifestations, with a history of treatment failure at Hefna, Belbis, Sharqia (Egypt) were sampled. From each bird lung and liver were aseptically collected and transported for laboratory investigations.Results:Examination of samples collected from 120 freshly dead turkey poults revealed the isolation of coagulase-positive staphylococci, coagulase-negative staphylococci, Campylobacter spp., Salmonella spp., Proteus spp., Pseudomonas spp., Klebsiella spp., and E. coli with the prevalence rates of 12/120 (10%), 30/120 (35%), 17/120 (14.2%), 5/120 (4.1%), 17/120 (14.2%), 6/120 (5%), 7/120 (5.8%), and 18/120 (15%), respectively. E. coli isolates were subjected for serotyping and characterization, while the rest of isolates were preserved to be investigated later in further studies. Serogrouping of E. coli isolates revealed the identification of O119, O6, O8, and O169, while 1/18 of isolates was untypable. Studying phenotypic antibiotic susceptibility profiles of isolates revealed that 18/18 (100%) of isolates demonstrated resistance against cefuroxime, tetracycline, and trimethoprim, 16/18 (88.9%) of isolates demonstrated resistance to amoxicillin/clavulanic acid, enrofloxacin, and norfloxacin, 14/18 (77.8%) of isolates demonstrated resistance to doxycycline and ciprofloxacin, and 9/18 (50%) of isolates showed resistance to gentamycin. Double disk synergy test showed that 6/18 (33.3%), 8/18 (44.4%), and 13/18 (72.2%) of isolates demonstrated the phenotypic pattern of ESBL producers with cefepime, cefotaxime, and ceftriaxone, respectively. Genotypic attributes for beta-lactamase TEM gene and int1 gene were studied by reverse transcriptase-polymerase chain reaction and revealed that 17/18 (94.4%) of isolates were positive for both genes. Embryo lethality test indicated that the 18 studied E. coli isolates were considered primary pathogens.Conclusion:The results revealed that 18/18 (100%) of E. coli isolates demonstrated MDR against three or more antibiotic groups, 9/18 (50%) of isolates showed extensive resistance against the nine tested chemotherapeutic agents from seven antibiotic groups. It is recommended to monitor the circulation of MDR and ESBL-producing pathogens in poultry production in a one health approach, as a preventive measure to mitigate the risk imposed on public health.

Prevelence of Methicillin-Resistant Staphylococcus Aureus (MRSA)in Layer Chickens with Reference to its Ability to Form Biofilm and Enterotoxins

This descriptive study was done on 100 samples obtained from layer chicken with symptoms of arthritis and decrease egg production in Assuit Governorate Egypt. A total of 37 Staphylococcal isolates were recovered, of which 21(56.7%) identified as coagulase positive Staphylococcus aureus (SA) and 16 (43.2%) were identified as coagulase negative staphylococci (CoNS). Sensitivity test of S. aureus isolates was performed against 10 common used chemotherapeutic agents, the results showed that all tested isolates were resistant to Amoxicillin , cefoxitin, and methicillin, while all were sensitive to vancomycin and Tylosin. For phenotypic analysis of biofilm formation, Congo Red Agar (CRA) test of 21 coagulase positive S. aureus isolates revealed that 76.2 % of these isolates were positive for biofilm production with variay degrees. A total of 12 representative coagulase positive staphylococci isolates were screened by multiplex PCR targeted the 16S rRNA, nuc and mecA genes to discriminate strains and to detect methicillin resistant strains(MRSA) , another PCR assay for presence of biofilm related genes, icaA, icaD, and PCR assay to detect prevalence of enterotoxin genes (SEs) . The 12 isolates were confirmed as S. aureus .Among the 12 S. aureus strains, 11 were found carrying mecA gene and only one isolate was mecA negative, biofilm related genes were icaD gene detected in 8 of11 S.aureus strains(72.7%), while icaA gene was found in one strain only(9%)and one strain was positive for both icaA and icaD genes. All the examined 11 mecA positive S.aureus strains harbourd enterotoxins genes distributed as follow: Sea found in (81.8%), seb (9%) and seb, sec in (9%) .Our results indicated that poultry S. aureus isolates are likely to be enterotoxigenic and biofilm producing ,and this may constitute a risk for poultry and might pose a public health hazard.

Characterization of Staphylococcus aureus strains and evidence for the involvement of non-classical enterotoxin genes in food poisoning outbreaks.

To an increasing extent, molecular and genetic characterization is now used to investigate foodborne outbreaks. The aim of this study was to seek molecular links among coagulase-positive staphylococci (CPS) isolated from three recent food poisoning outbreaks in Romania using polymerase chain reaction and pulsed-field gel electrophoresis techniques. The 19 CPS isolates were identified as Staphylococcus aureus by detection of the 23S rDNA gene. Among them, 15 carried at least one staphylococcal enterotoxin-encoding gene. The Calaraşi outbreak strains grouped in pulsotype 2 and were sed/sej/ser-positive, whereas the Arad outbreak strains clustered in pulsotype 17 and were either sed/seg/sei/sej/ser- or seg/sei-positive. The Piteşti outbreak strains clustered in pulsotype 1 and, surprisingly, possessed only one enterotoxin gene, i.e. seh. Similar to other European countries, the seh gene has been identified with increasing frequency in Romanian outbreaks; this highlights the importance of considering the application of methods recommended for staphylococcal enterotoxin regulation in Europe.

Antibiotic resistance of Staphylococcus intermedius strains isolated from 53 natural cases of canine pyoderma

Samples for bacterial cultures on blood agar were obtained from the skin lesions of 53 dogs demonstrating the typical clinical and cytological features of superficial (n=17) or deep (n=36) pyoderma. None of them had received any kind of antimicrobial therapy within the previous month. The Staphylococcus looking colonies were recultured on blood and Chapman agar for staphylococcal species identification and biotyping. All 53 coagulasepositive staphylococcal isolates were eventually identified as S. intermedius. The susceptibility for each of these isolates was tested against 12 different antibiotics with the disc diffusion method. Resistance rates were found to be 0% for amoxicillin/clavulanate, 11.3% for oxacillin, 7.5% for cefalexin, 5.7% for enrofloxacin, 32.1% for erythromycin, 11.3% for tylosin, 20.8% for lincomycin, 18.9% for clindamycin, 32.1% for doxycycline, 5.7% for amicacin, 7.5% for chloramphenicol and 92.5% for trimethoprim/sulfamethoxazole. No correlation between the resistance pattern and either the age, breed, sex, and the animals' lifestyle, or the depth of skin infection, the presence or not of underlying diseases and any past treatment with antibiotics was identified.

Isolation of Coagulase Positive Staphylococci from She-Camel Milk at Eastern Libya and Their Drug Susceptibility Patterns

Isolation of Coagulase Positive Staphylococci from She-Camel Milk at Eastern Libya and Their Drug Susceptibility Patterns

Prevalence of Staphylococcus aureus In Broiler Chickens with Special Reference to Beta-Lactam Resistance Genes in the Isolated Strains

The dramatic increase in the prevalence of antimicrobial-resistant Staphylococcus aureus (S. aureus) is receiving wide spread attention, due to multi-resistant strains, diminishing the usefulness of antibiotics in human and animal medicine and, thereby limiting therapeutic options. In this study, 250 samples of chicken swabs were collected from five different poultry farms (50 samples from each farm) within Upper Egypt area. 157 staphylococci strains were detected with an isolation rate of (62.8%). isolates of S. aureus were confirmed using standard microbiological methods. Eighty one (55.9%) were coagulase positive staphylococci, and 76(48.4%) were coagulase negative staphylococci, phenotypic prediction of β-lactam resistance among the 81 isolated coagulase positive staphylococci was declared using disk diffusion test against, cefoxitin and oxicillin which were used for detection of mecA mediated β-lactam resistance (methicillin resistance) while penicillin and amoxicillin/clavulinic acid (AMC) antibiotic discs were used to monitor blaZ mediated resistance. Out of 81 isolates ,all isolates were 100% resist to oxacillin and 93.8 % were resist to cefoxitin, while 90.1 % were resist to penicillin and 87.6% were resist to AMC. The antibiotic susceptibility results were judged by PCR, the mecA ,and blaZ genes were tested for the evaluation of the susceptibility of coagulase positive Staphylococci against oxacillin and s-lactams, the thirty one reprsentative coagulase positive staphylococci isolates confirmed that all the isolates were Staphylococci by successful amplification of the 228 bp PCR product of the Staphylococcal specific 16S rRNA gene. While , 20 isolates were confirmed to be S.aureus by successful amplification of 279 bp PCR product of the S.aureus specific thermonuclease gene .On using the same multiplex PCR assay, detection of blaZ and/or mecA genes were performed through successful amplification of 517 bp and /or 147 bp specific products, respectively. Out of the 31 reprsentative coagulase positive Staphylococci isolated from broiler chickens, 20 (64.5%) were mecA gene positive while 23 (74%) were β- blaZ gene positive. Also, 15 (48.3%) of the isolates carried both mecA and blaZ genes while only 3 isolates (9.6%) were negative for both genes. Concerning MIC of 12 representative coagulase positive staphylococci isolates against 12 selected antibiotics commonly used in poultry farms, 100% of isolates were resistant to streptomycin, spectinomycin ,ampicillin, Cefotax, Trimethoprim-sulphamethazole, gentamycin, and Methicillin with MIC >128, while 100% of isolates were sensitive to tylosin, lincomycine, enrofloxacin, ciprofloxacin and oxytetracyclin.

First report of the Staphylococcus aureus isolate from subclinical bovine mastitis in the South of Brazil harboring resistance gene dfrG and transposon family Tn916-1545

The aim of this work was to identify at the molecular level the species of coagulase-positive staphylococci isolates from clinical and subclinical bovine mastitis samples in Southern Brazil, and to evaluate the antimicrobial resistance profile, as well as the presence of resistance genes. According to the PCR assay, all 31 isolates were classified as Staphylococcus aureus. The isolates were tested for resistance to penicillin, ampicillin, oxacillin, cefoxitin, cephalothin, ceftiofur, streptomycin, tobramycin, teicoplanin, erythromycin, clindamycin, enrofloxacin, sulfonamide, trimethoprim-sulfamethoxazole, trimethoprim, and tetracycline by the disk diffusion method. Most of the isolates were resistant to sulfonamide (20), followed by ampicillin and clindamycin (16). Twenty isolates were multidrug-resistant. PCR was used for the detection of several antimicrobial resistance genes (ereB, ermB, ermC, tetA, tetB, tetK, tetL, tetM, tetO, Tn916-1545, strA, strB, sul1, sul2, dfrA, dfrG, dfrK, blaZ, mecA, and mecC). The most prevalent antimicrobial resistance genes were tetK and tetL, ereB, followed by tetM, Tn916-1545 and blaZ, detected in 11, nine and four isolates, respectively. For all the tetM gene positive isolates, the presence of conjugative transposons of the Tn916-1545 family was detected. The presence of multidrug-resistant isolates, antimicrobial resistance genes and transposons suggests a potential risk of spreading multi-resistance genes to other bacteria.

Enterotoxin production and antimicrobial susceptibility in Staphylococci isolated from traditional raw milk cheeses in Serbia

ABSTRACTThis study was undertaken to determine the prevalence of coagulase positive staphylococci (CPS) by examining a total of 71 raw milk cheeses. Additionally, enterotoxigenicity, antimicrobial susceptibility and the presence of mecA and mecC genes in the staphylococcal isolates were investigated. The isolation and enumeration procedure of CPS followed the International Organization for Standardization (ISO) standard. The presumptive staphylococci were identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) using the VITEK MS system. VIDAS® Staph enterotoxin II assay was used for the detection of classical enterotoxins. Antimicrobial susceptibility testing (AST) was accomplished performing the disk diffusion method. All suspected methicillin resistant staphylococci were investigated for the presence of mecA and mecC genes by PCR assay. A high prevalence (87.32%) of CPS was detected in the cheeses at contamination levels up to 5.58 log CFU g−1. Among 47 staphylococcal isolates screened for enterotoxin production, only one isolate, identified as S. hyicus, was confirmed as being enterotoxigenic. Resistance to penicillin (63.70%) was the most common resistance among the tested Staphylococcus aureus isolates. The dominant phenotypic resistance patterns in coagulase negative staphylococci (CNS) were resistance to ofloxacin and fusidic acid. All CNS isolates were susceptible to the clinically important antibiotics clindamycin, chloramphenicol, gentamicin, linezolid, rifampicin and trimethoprim-sulfamethoxazole. The mecA and mecC genes were not detected. To the best of our knowledge, this is the first study concerning evaluation of the presence of methicillin resistant staphylococci (MRS) in dairy products in Serbia.

Characterisation of methicillin-susceptible Staphylococcus pseudintermedius isolates from canine infections and determination of virulence factors using multiplex PCR

Staphylococcus pseudintermedius is a genuine opportunistic pathogen of the skin, especially in canids. However, characterisation of virulence, antimicrobial resistance and genotypic variability in methicillin-susceptible S. pseudintermedius isolates has not been fully explored. In this study, coagulase-positive staphylococcal isolates collected from dogs of various breeds and ages suffering from dermatitis (n = 70), pyoderma (n = 7), and otitis (n = 7), from districts of Prague (Czech Republic) and surrounding areas, were characterised using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry, and repetitive sequence-based PCR fingerprinting. Susceptibility to antimicrobial agents was determined, virulence factor genes for leukocidin (lukSF-I), exfoliatins (exi, expB, and siet), enterotoxin C (sec<sub>canine</sub>) and enterotoxin-related genes (se-int and sel) were detected using multiplex PCR and the genotypes of S. pseudintermedius isolates were determined using SmaI macrorestriction analysis. The majority of the staphylococcal isolates (n = 84) were identified as S. pseudintermedius (n = 79) and all of them were susceptible to methicillin/oxacillin (MSSP). About half of the strains (n = 41) were resistant to macrolide-lincosamide-streptogramin B antimicrobial agents and resistance was mediated in all but one of the strains by the erm(B) gene. The genes for lukSF-I, siet, se-int, and sel were detected in the majority of the MSSP strains (96.2%, 100%, 100%, and 73.4%, respectively). Investigated canine S. pseudintermedius isolates were highly heterogeneous, which prevented the correlation of any specific lineage to a particular infection, dog breed, or region of origin.

Antimicrobial resistant profiles of Methicillin-resistant Staphylococcus aureus (MRSA) recovered from broiler chicken populations in Egypt

Little is still known about prevalence of methicillin resistant Staphylococcus aureus (MRSA) in broilers, so this study aimed to determine its prevalence, antibiogram patterns and the site of resistant genes using PCR. About 57 staphylococcal isolates of 78 samples (73%) recovered from broiler chicken were classified into coagulase positive staphylococci (CPS) (40.3%) and coagulase negative staphylococci (CNS) (59.6%). The CPS isolates were identified as MRSA (34.8%) and methicillin sensitive Staphylococcus aureus (MSSA) (65.2%) according to oxacillin resistance by disc diffusion method. The CPS isolates showed the highest rate of resistance to trimethoprim/sulphamethoxazole (100% and 93.3% for MRSA and MSSA respectively), erythromycin (100% and 93.3% for MRSA and MSSA respectively), oxacillin and cefoxitin (100%, 87.5% MRSA and 0%, 6.7% MSSA respectively), ceftriaxone (87.5% and 73.3% for MRSA and MSSA respectively) and amoxicillin (75% and 40% for MRSA and MSSA respectively). Moreover, the CNS showed resistance against trimethoprim/sulphamethoxazole with 85.3%, followed by 79.4% erythromycin, 55.9% ceftriaxone, 47% cefoxitin, 47% amoxicillin, and oxacillin (44%). PCR assay revealed that four (80%) of five MRSA isolates were harbored mecA gene. The screening of mecA, dfrA and ermC resistant genes was done on plasmid and chromosomal DNA of four MRSA isolates. All of them were chromosome linked genes, but none of these resistant genes were detected on plasmid DNA. All examined isolates (100%) contained ermC gene. In addition, the dfrA gene was detected in 2 (50%) out of the 4 isolates.