tetK Genes Research Articles

A One Health Approach Molecular Analysis of Staphylococcus aureus Reveals Distinct Lineages in Isolates from Miranda Donkeys (Equus asinus) and Their Handlers

Donkeys (Equus asinus) are in decline in Europe. Occupational exposure to farm animals has been associated with increased staphylococci carriage. We aimed to isolate S. aureus and coagulase-negative staphylococci (CoNS) from donkeys and handlers and characterize the antimicrobial resistance profiles and genetic lineages of S. aureus strains. Oral and nasal swab samples were collected from 49 Miranda donkeys and 23 handlers from 15 different farms. Staphylococci species were identified by MALDI-TOF MS. The presence of antimicrobial resistance genes and virulence factors was investigated by PCR. Molecular typing was performed in S. aureus isolates. From the 49 donkey samples, 4 S. aureus (8.2%) and 21 CoNS (42.9%) were isolated. Ten handlers (43.5%) were carriers of S. aureus and 4 (17.4%) carried CoNS. The CoNS isolates showed resistance to several classes of antimicrobials encoded by the mecA, aph (3′)-IIIa, ant (4′)-Ia, tetM, tetK, lnuA, ermB, ermC, dfrA and dfrG genes. S. aureus isolates were resistant to penicillin, aminoglicosides and tetracycline harboring the blaZ, aph (3′)-IIIa, tetL, tetM and tetK genes. All S. aureus isolates from donkeys belonged to ST49 and spa-type t208 while the strains isolated from the handlers were ascribed to 3 STs and 7 spa-types. However, human isolates were from different STs than the donkey isolates. Donkeys are mainly colonized by methicillin-resistant S. sciuri. S. aureus transmission between donkeys and their handlers appears not to have occurred since the isolates belonged to different genetic lineages.

Multidrug-Resistant Methicillin-Resistant Coagulase-Negative Staphylococci in Healthy Poultry Slaughtered for Human Consumption.

Coagulase-negative staphylococci are commensals that are known to be prevalent in most environments, and they are also an important reservoir of antimicrobial-resistant genes. Staphylococcal infections in animal husbandry are a high economic burden. Thus, we aimed to determine the prevalence and species diversity of methicillin-resistant coagulase-negative staphylococci (MRCoNS) in poultry slaughtered for human consumption and to study the antimicrobial resistance of the isolates. Swab samples were recovered from 220 commercial chickens, homebred chickens and quails. Species identification was performed using MALDI-TOF. Antimicrobial susceptibility testing was performed by the disc diffusion method against 14 antimicrobials. The presence of antimicrobial-resistant genes was investigated by polymerase chain reaction. Totals of 11 (19.6%), 13 (20.3%), and 51 (51%) MRCoNS were isolated from commercial chickens, homebred chickens and quails, respectively. S. lentus was isolated from all homebred chickens, whereas 11 S. lentus and 2 S. urealyticus were isolated from commercial chickens. As for quails, the most prevalent MRCoNS were S. urealyticus. Almost all isolates had a multidrug-resistant profile and carried the mecA gene. Most isolates showed resistance to erythromycin, clindamycin, penicillin, tetracycline, ciprofloxacin and fusidic acid and harbored the ermA, ermB, ermC, mphC tetK, tetL, tetM and tetO genes. This study showed a frequent occurrence of multidrug resistance in MRCoNS isolated from healthy poultry in Portugal.

Presence of Antibiotic-Resistant in Staphylococcal Subclinical Mastitis in Several Regencies of East Java, Indonesia

Staphylococcal mastitis has been reported as a serious dairy disease in various regions around the world. The occurrence of resistant strains in Staphylococcus species to antibiotics has triggered alternative treatment substituting antibiotic usage on the global scene. This study aimed to investigate the presence of antibiotic-resistant genes in Staphylococcal subclinical-mastitis cases present in several regencies of East Java Province, Indonesia. A total of 592 quarter milk samples were collected from 62 farms in the region with high dairy cattle populations in Lumajang, Banyuwangi, Malang, Sidoarjo, Jember, Pasuruan, Probolinggo, and Mojokerto. Subclinical-mastitis samples were screened using the California mastitis test (CMT). Positive CMT samples were grown on the selective Staphylococcus media and tested for their biochemical properties. The polymerase chain reaction was performed to detect the presence of antibiotic-resistant genes in all isolates ( Staphylococcus sp) using a specific pair-primer for mecA, blaZ, tetK , and tetM genes. The result showed that about 67% of milk samples were subclinical mastitis in several regencies of East Java. About 17.12% of subclinical mastitis was caused by Staphylococcus species ( Staphylococcus aureus , Staphylococcus epidermidis , and other non- aureus Staphylococci (NAS)). The most prevalent region of Staphylococcal subclinical-mastitis was recorded in Jember. However, only NAS species obtained from Mojokerto, Malang, Probolinggo, and Banyuwangi were detected to have a blaZ gene responsible for penicillin resistance. In conclusion, the appearance of the antibiotic-resistant gene in NAS species found in several regencies of East Java can be used as important information to evaluate Staphylococcal subclinical-mastitis treatment.

Surface microbiota and associated staphylococci of houseflies (Musca domestica) collected from different environmental sources

Houseflies (Musca domestica) are important mechanical vectors for the transmission of pathogenic microorganisms. In this study, 129 houseflies (69 males and 60 females) were collected from 10 different environmental sources and a laboratory population was used. The surface microbiota of houseflies was identified by Next-Generation Sequencing. Staphylococci from the surfaces of houseflies were selectively isolated and their virulence genes, antibiotic susceptibilities, biofilm formation, and clonal relatedness were determined. Metagenomic analysis results demonstrated that Staphylococcus, Bacillus, and Enterococcus were mostly present on the surface of houseflies at the genus level. Additionally, the isolated 32 staphylococcal strains were identified as Staphylococcus sciuri (n = 11), S. saprophyticus (n = 9), S. arlettae (n = 6), S. xylosus (n = 4), S. epidermidis (n = 1) and S. gallinarum (n = 1). tetK, tetM, tetL, ermC, msrAB, and aad6 genes were found to carry by some of the staphylococcal strains. The strains were mostly resistant to oxacillin, penicillin, and erythromycin and three strains were multi-drug resistant. There was a statistical difference between housefly collection places and antibiotic resistance of isolated staphylococci to penicillin G, gentamicin, and erythromycin (p < 0.05). Biofilm test showed that 17 strains were strong biofilm formers, and it plays important role in the transmission of these bacteria on the surface of houseflies. Staphylococcal strains showed extracellular proteolytic and lipolytic activity in 31 and 12 strains, respectively. Closely related species were found in PFGE analysis from different environmental sources. By this study, surface microbiota and carriage of pathogenic staphylococci on the surfaces of houseflies and their virulence properties were elucidated.

Detection, antibiogram and molecular characterization of MRSA and MSSA isolated from swine

The emergence of bacteria with antimicrobial resistance traits such as methicillin-resistant Staphylococcus aureus (MRSA) in food-producing animals is a significant public health concern. The aim of this study is to determine the prevalence rate, antibiotic susceptibility and the molecular characteristics of Staphylococcus aureus (S. aureus) in swine from selected swine farms in Peninsular Malaysia. The oral and nasal swabs of 200 live swine were collected. Screening and isolation of S. aureus isolates were carried out using phenotypic identification techniques. The identity of the bacteria isolates was confirmed by detection of nuc gene (S. aureus) and mecA gene (MRSA). Antibiogram of S. aureus against 19 antibiotics was developed using Kirby-Bauer test. Molecular detections of antimicrobial resistance and virulence genes as well spa typing of S. aureus was carried out. 25% of swine were carrying S. aureus with 9% being MRSA carriers. S. aureus and MRSA have shown high resistance against clindamycin, tetracycline, chloramphenicol, oxacillin, penicillin, erythromycin, cefoxitin and amoxicillin/clavulanate. Antimicrobial resistance and virulence genes, including tetK, tetL, tetM, ermA, ermB, ermC, msrA and scn genes were detected. 93% of S. aureus were multiple-drug resistant (MDR), suggesting the emergence of MDR S. aureus in swine in Malaysia.

Genotypic and Phenotypic Characterizations of Methicillin-Resistant Staphylococcus aureus (MRSA) on Frequently Touched Sites from Public Hospitals in South Africa.

The hospital environment acts as a reservoir in the transmission of pathogens, such as MRSA, which may cause hospital-acquired infections. This study aimed to ascertain the prevalence, genetic relatedness, antibiotic resistance, and virulence profile of MRSA on some frequently touched hospital sites in South Africa. A total of 777 swabs were randomly collected from 11 frequently touched sites in the hospital environment of three wards of four public hospitals in the KwaZulu-Natal province of South Africa. Isolation of S. aureus and confirmation were done using genotypic and phenotypic methods. Antibiotic susceptibility testing was performed using the Kirby–Bauer disk-diffusion method. MRSA isolates were determined by the presence of the mecA gene. Virulence and resistance genes were detected using a standard monoplex PCR assay. ERIC-PCR was conducted to evaluate the genetic relatedness. An overall prevalence of 12.7% for S. aureus isolates was obtained. Out of these, 89.9% (89/99) were confirmed to be MRSA. The sites with the highest prevalence were the occupied beds (16.2% (16/99)), unoccupied beds (16.2% (16/99)), patient files (14.1% (14/99)), ward phones (13.1% (13/99)), and nurses' tables (14.1% (14/99)). The virulence genes with the highest observed frequency were hld (87 (87.9%)) and LukS/F-PV (53 (53.5%)). The resistance genes with the highest frequency were the tetM and tetK genes detected in 60 (60.6%) and 57 (57.6%) isolates, respectively. The ERIC-PCR results obtained indicated a high level of genetic diversity; however, intraclonal (within a hospital) and interclonal (between hospitals) clusters of MRSA were observed. The study showed that MRSA can contaminate various surfaces, and this persistence allows for the dissemination of bacteria within the hospital environment. This highlights the need for improved infection prevention and control (IPC) strategies in public hospitals in the country to curb their potential transmission risks.

Molecular characterization of netB and tpeL virulence factors and antimicrobial resistance genes of Clostridium perfringens isolated from herbs and spices.

The present study aimed to determine some virulence-associated genes and antimicrobial multidrug resistance of Clostridium perfringens recovered from herbs and spices widely distributed in the Egyptian market. C. perfringens virulence and resistance factors were determined using PCR targeting the netB, tpeL, ermB, bla and tetK genes. Thirty three out of 392 samples (8.42%) from herbs and spices submitted to our laboratory for bacteriological screening were positive for presence C. perfringens. PCR results for the tpeL gene in isolated C. perfringens revealed 9 out of 33 (27.3 %) of isolates, while netB was not detected. The isolates were resistant to Clindamycin, Vancomycin, tetracycline, and erythromycin with inhibition zones of 6.28 ± 0.63, 8.78 ± 0.41, 9.63 ± 0.63, and 9.84 ± 0.66 mm, respectively. The genes mentioned above were selected to correspond to the ineffective antimicrobials; ermB for erythromycin, tetK for tetracycline, and bla for the remainder. PCR results for antibacterial resistant genes in isolated C. perfringens revealed their presence. From 33 isolates, bla gene was detected in 21 (63.4 %), tetK in 13 (39.4 %) and ermB in only one isolate (3.03 %). Sequencing analysis was done for the bla gene as an example for the detected genes as detected at the highest incidence (63.4%). No cross-relationship was detected upon comparing incidence data of both studied virulence genes and those of antimicrobial resistance. The present findings may explain the resistance of C. perfringens to the examined antibacterials and recommend avoiding the application of them to control the microbe. In addition, the authors recommend following strict hygienic procedures during the industry of herbs and spices to ensure their clearance from Clostridium perfringens before distributing the products as food additives into the markets.

Prevalence and Characteristics of Multidrug-Resistant Livestock-Associated Methicillin-Resistant Staphylococcus aureus (LA-MRSA) CC398 Isolated from Quails (Coturnix Coturnix Japonica) Slaughtered for Human Consumption

Simple SummaryMethicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in both humans and animals worldwide. MRSA associated with livestock is a zoonotic pathogen that has been reported in several animals and, although its infections in humans are rare, this strain is recognized as an occupational hazard for people working in direct contact with livestock. Thus, we aimed to isolate MRSA from quails and to characterize their antimicrobial resistance and genetic lineages. One hundred swab samples were recovered from quails at the slaughterhouse. To investigate the prevalence and antimicrobial resistance of MRSA in poultry, we conducted this study on 100 quails slaughtered for human consumption. The antimicrobial resistance was investigated in all isolates as well as virulence genes and genetic lineages. Twenty-nine MRSA were isolated. The results showed that all MRSA isolates had resistance to multiple antibiotics. All strains were classified as livestock-associated. Most strains belonged to a well-known livestock-associated lineage: CC398.Livestock-associated MRSA (LA-MRSA) is a zoonotic pathogen that has been reported in several animals, and it is often associated with clonal complex (CC) 398. We aimed to isolate MRSA from quails and to characterize their antimicrobial resistance and genetic lineages. One hundred swab samples were recovered from quails at the slaughterhouse. The swabs were inoculated onto CHROMagar™ MRSA agar plates for MRSA isolation. The presence of antimicrobial-resistant genes and virulence factors was investigated by PCR. All strains were typed by MLST, SCCmec-, spa- and agr-typing. From the 100 samples, 29 MRSA were isolated. All strains were resistant to penicillin, cefoxitin, ciprofloxacin, erythromycin and clindamycin and carried the blaZ, mecA, ermB and ermC genes. All strains, except one, showed resistance to tetracycline and harbored the tetM, tetK and tetL genes in different combinations. Twenty strains belonged to ST398 and SCCmec type V, and nine strains belonged to the new ST6831. Twenty-eight out of twenty-nine strains were ascribed to t011 and one to t108. As far as we know, this is the first report of MRSA from quails slaughtered for human consumption. Most strains belonged to ST398-t011, which is the most common LA-MRSA clone found in livestock in Europe.

Virulence Factors in Staphylococcus Associated with Small Ruminant Mastitis: Biofilm Production and Antimicrobial Resistance Genes.

Small ruminant mastitis is a serious problem, mainly caused by Staphylococcus spp. Different virulence factors affect mastitis pathogenesis. The aim of this study was to investigate virulence factors genes for biofilm production and antimicrobial resistance to β-lactams and tetracyclines in 137 staphylococcal isolates from goats (86) and sheep (51). The presence of coa, nuc, bap, icaA, icaD, blaZ, mecA, mecC, tetK, and tetM genes was investigated. The nuc gene was detected in all S. aureus isolates and in some coagulase-negative staphylococci (CNS). None of the S. aureus isolates carried the bap gene, while 8 out of 18 CNS harbored this gene. The icaA gene was detected in S. aureus and S. warneri, while icaD only in S. aureus. None of the isolates carrying the bap gene harbored the ica genes. None of the biofilm-associated genes were detected in 14 isolates (six S. aureus and eight CNS). An association was found between Staphylococcus species and resistance to some antibiotics and between antimicrobial resistance and animal species. Nine penicillin-susceptible isolates exhibited the blaZ gene, questioning the reliability of susceptibility testing. Most S. aureus isolates were susceptible to tetracycline, and no cefazolin or gentamycin resistance was detected. These should replace other currently used antimicrobials.

Draft genome of an extremely drug-resistant st551 Staphylococcus pseudintermedius from an Italian dog with otitis externa

ObjectivesTo determine the draft genome sequence and analyse the genetic features of a Staphylococcus pseudintermedius clinical isolate according to the main typing schemes available, with a special focus on antibiotic resistance. MethodsThe strain was isolated from a case of otitis externa in a dog. Its identity and pattern of antibiotic susceptibility were determined using an automated system. The genome was sequenced using an Illumina platform. MLST, SCCmec typing, resistome, and mobile genetic elements were derived by comparative analysis using available specific databases. ResultsS. pseudintermedius CAM1 isolate has a chromosome size of 2 652 610 bp. It showed a wide pattern of phenotypic resistance, comprising beta-lactams, macrolides and lincosamides, aminoglycosides, fluoroquinolones, tetracyclines, and trimethoprim-sulfamethoxazole. The genetic determinants of the underlying mechanisms were all found by in silico analysis of the genome. The mecA gene for methicillin resistance was harboured by the Vc type of the SCCmec. MLST of the strain was st551. ConclusionBy comparison with the MLST database of S. pseudintermedius and data from published molecular epidemiology studies, CAM1 is the first st551 strain recorded in Italy and, in the context of an already extremely wide antibiotic resistance pattern, it harbours also the tetK gene, the prevalence of which is rare in MDR S. pseudintermedius.

Prevalence of Staphyloсoссus aureus and Streptococcus agalaсtiae isolated from Raw Milk in Dakahlia Governorate, Egypt

This study was carried out on 200 random milk samples сolleсted from different areas at Dakahlia Governorate for detection of prevalence and antimicrobial resistance of some zoonotic bacteria in milk as Staphylococcus aureus and Streptococci by using biochemical tests, antibiotic sensitivity test, PCR for confirmation and detection of some resistance and virulence genes. Staphyloсoссus aureus (S. aureus) was detected in 63%, 76%, and 42%, while Streptococcus agalaсtiae (St. agalaсtiae) was found in 8%, 10% and 4% of examined individual milk samples, retail milk samples, and milk of Bulk tank, respectively. S. aureus isolates revealed high levels of resistance to ampicillin (100%), nalidxiс acid (90%), oxaсillin (85%), peniсillin (85%), cefoxitin (75%), rifampin (25%), tetraсyсline (20%) and erythromyсin and sulphamethoxazole/ trimethoprim (10%). Whereas, St. agalaсtiae show high resistance to tetracycline (90%), ampiсillin (80%), rifampin (60%), sulphamethoxazole/ trimethoprim (50%), gentamyсin (45%), erythromyсin and nalidxiс aсid (40%), cholorampheniсol (30%), and streptomyсin (25%). PСR results revealed that 4 out of 5 (80%) methicillin-resistant (MRSA) isolates had meсAand 2 (40%) had meсС,while 1 (20%) had Seaenterotoxin. Three isolates (100%) of St. agalaсtiae had sul1 gene and one out of three (33.3%) had tetK genes, while dfrA could not be detected. The main outcome of the current work is that milk can cause severe public health hazards to people because it had a variety of microorganisms. It is important to ensure using good hygienic practices in farms and prevent the haphazard abuse of antibiotics.

Clonal Diversity and Antimicrobial Resistance of Methicillin-Resistant Staphylococcus pseudintermedius Isolated from Canine Pyoderma.

The emergence of methicillin-resistant Staphylococcus pseudintermedius (MRSP) antimicrobial resistance and epidemic genetic lineages is posing a challenge in veterinary medicine due to the limited therapeutical options. MRSP has been identified as an important canine pyoderma pathogen. Thus, we aimed to characterize the antimicrobial resistance and clonal lineages of MRSP isolated from canine cutaneous pyoderma. Thirty-one MRSP isolates recovered from pyoderma were further characterized. The antimicrobial susceptibility testing of the isolates was performed by the Kirby-Bauer disc diffusion method against 14 antimicrobial agents. The presence of antimicrobial and virulence genes was carried out by PCR. Multilocus sequence typing was performed in all isolates. All strains had a multidrug-resistant profile showing resistance mainly to penicillin, macrolides and lincosamides, aminoglycosides, tetracycline and trimethoprim-sulfamethoxazole, which was encoded by the blaZ, ermB, msr(A/B), aac(6′)-Ie-aph(2′′)-Ia, aph(3′)-IIIa, ant(4′)-Ia, tetM, tetK and dfrG genes. All isolates harbored the lukS-I/lukF-I virulence factors. Isolates were ascribed to nine previously described sequence types (STs): ST123, ST339, ST727, ST71, ST537, ST45, ST1029, ST118 and ST1468; and to five STs first described in this study: ST2024, ST2025, ST2026, ST2027 and ST2028. In this study, most isolates belonged to ST123 (n = 16), which belongs to CC71 and is the most common clone in Europe. All isolates were multidrug-resistant, which may impose a serious threat to animal health.

Antagonistic Potential of Dairy Origin Enterococcus faecium Against Multidrug-Resistant Foodborne Pathogens

Probiotic potential of Enterococcus spp. is widely investigated around the globe. The biochemically and molecular characterized E. faecium strains isolated from Dahi (continental yogurt) were evaluated to tolerate simulated gastric environment, bile, sodium chloride, temperature, and pH. The safety was assessed by disc diffusion, broth microdilution, antibiotic resistance genes screening, and hemolytic ability. Enterococci survived simulated gastrointestinal conditions and depicted growth at temperature (15 to ≥42°C), pH (≤2.5 to ≥9.5), 0.3% bile salt and 3% NaCl. All strains were sensitive to ampicillin, vancomycin, kanamycin, gentamicin, streptomycin, tetracycline and ciprofloxacin and harbored vanR, vanX, qnrB2, qnrS, tetK, and tetW resistance genes. E. faecium strains inhibited the E. coli (85%) and S. Typhi (50%) whereas the 10% cell-free culture supernatant (CFCS) of E. faecium halted the growth of E. coli while 15% CFCS completely suppressed S. Typhi. The cell-free culture supernatant retained antibacterial nature after pH and proteinase K treatment, however, it lost activity after heat treatment (≥95°C). The genetic screening revealed that all isolates are capable to produce putrescine biogenic amine. Further assessment of strains for lack of infectivity, cytotoxicity in animals, adhesion to Caco-2 cells and characterization of enterocins is essential to conclude the probiotic potential of these strains.

Genes conferring anti­microbial resistance in cattle with subclinical mastitis

This study was carried out to evaluate the antimicrobial resistance (AMR) as a risk factor associated with some microorganisms isolated from subclinical mastitis (SCM) milk samples from Holstein Friesian dairy animals in Fayoum area, Egypt. The percentage of the SCM in the farm was found to be 41.18% and 63.88% at quarter and cows level respectively, with mean somatic cell count (SCC) of 8.8×105 ± 9.2×103 cells/mL and electrical conductivity (EC) 6.27 ± 0.066 mS/cm for SCM quarter milk samples. Out of the total 444 SCM cow milk samples, the most often isolated microorganisms were Staphylococcus aureus: 296 (66.6%), Enterococcus spp.: 230 (51.80%), Escherichia coli: 210 (47.29%) and Streptococcus agalactiae: 106 (23.87%). AMR was determined by disc diffusion test and the corresponding resistance genes were detected by PCR. Results of the in vitro susceptibility tests performed and the phenotypes indicated that the highest resistance to antibiotics for isolated microorganisms was against penicillin followed by amoxicillin + clavulanic acid, oxacillin and tetracycline, whereas moderate resistance was exhibited to oxytetracycline, ampicillin, sulfamethazole/trimethoprim, cefotaxime and erythromycin. However the most effective antibiotics against most isolates were nitrofurantoin and gentamicin followed by enrofloxacin, norfloxacin and cefoxitin. It was shown that the resistance to tetracyclines was due to the tetK or tetA(A) genes, the resistance to β-lactams (penicillins) – to blaZ and blaTEM genes, to macrolides (erythromycin): to ermB and ermC genes. Methicillin resistance genes were mecA, mec1 and mecC, glycopeptides (vancomycin) resistance gene was vanA, and norfloxacin resistance was attributed to norA gene.

Antimicrobial Susceptibility Pattern in the Bacteria Isolated from Surgical Site Infection: Emphasis on Staphylococcus Aureus; Yasuj City, Southwest Iran.

Surgical site infections (SSIs) in surgical wards remains the most common cause of postoperative complications and realistically is the third most common origin of healthcare-related conditions. Staphylococcus aureus is undoubtedly the most common bacteria causing SSIs. The current study aimed at investigating the antimicrobial susceptibility pattern in bacteria isolated from SSIs, evaluation of tetracycline resistance genes, and SCCmec typing in S. aureus isolates isolated from patients with SSIs from 2018 to 2019 in Yasuj, Kohgiluyeh, and Boyer-Ahmad Province, Iran. This study diligently investigated 240 potential patients. Antimicrobial susceptibility testing was performed properly by the disk diffusion method. For the final confirmation of isolated bacteria, PCR was used. The presence of tet genes and SCCmec typing was carried out by multiplex PCR. The results showed that the most common isolated pathogens included S. aureus, E. coli, P. aeruginosa, Coagulase-negative Staphylococci, and K. pneumonia in 58.8%, 19.8%, 9.2%, 6.8% and 5.4% of cases, respectively. The majority of the Gram positive isolates were resistant against penicillin (86%) and Gram negative were resistant against ciprofloxacin (75.6%). In isolates of Staphylococcus aureus, the mecA gene was detected in 63.6% of isolates. The predominant SCCmec types were type III (59.1%) and type I (18.4%). The tetK and tetM genes were detected in 80.7% and 71.9% of the S. aureus isolates, respectively. There was a statistically significant difference between tet genes (tetK and tetM) from the viewpoint of resistance to tetracycline (p = 0.024). According to the results of the current study, it is recommended to administer vancomycin, amikacin, and imipenem in Yasuj to treat SSIs.

The frequency of tetracycline resistance genes in Escherichia coli strains isolated from healthy and diarrheic pet birds.

Pet birds have close contact to human and resistant bacteria can transfer from birds to intestinal flora of human. This study was carried out to determine the tetracycline resistance genes in Escherichia coli strains associated with enteric problem in pet birds. Totally, 295 cloacal swabs were collected from 195 healthy and 100 diarrheic pet birds in Isfahan province, Iran. The presence of E. coli was identified by conventional bacteriological, biochemical, and molecular examinations. The presence of tetracycline resistance genes (tetA, tetB, tetC, tetD, tetE, tetG, tetK, tetL, tetM, tetO, and tetS genes) were examined using three multiplex PCR. The results showed that 18.9% and 43% of cloacal samples of healthy and diarrheic pet birds contained E. coli, respectively. The mean percentage of E. coli isolated from cloacal samples of diarrheic birds was significantly higher than the healthy birds (46.6 vs 23.1%). In healthy birds, out of 37 E. coli isolates, 10 isolates were resistant to tetracycline, harboring tetA and tetB genes (3 tetA vs 7 tetB), but in the diarrheic birds, of 26 resistance E. coli, 11, 12, and 3 strains contained tetA (42.3%), tetB (46.15), and tetA+tetB (11.53%) genes. The percentage of tet genes were significantly higher in diarrheic birds than healthy birds (58.9 vs 24.0%). Both resistant genes of tetA and tetB were detected in E. coli isolates that are related with efflux pump activity. These genes can be transferred between Gram-negative bacteria and they have the potential ability to be transferred to the environment and human flora.

Molecular studies on antimicrobial resistant genes in predominant staphylococci and E. coli isolates from bovine mastitis in different regions of Karnataka

Bovine mastitis is a persistent and major problem in the dairy industry globally. Staphylococci and Escherichia coli are predominantly responsible for this condition. The aim of this study was to look for the genes associated with antimicrobial resistance in these predominant bacteria isolated from bovine mastitis in different regions in Karnataka. In all, 211 bacterial isolates were recovered from bovine mastitis cases in Karnataka. Of these, 85, 23 and 47 were Staphylococcus aureus, S. epidermidis and E. coli, respectively, as confirmed by PCR. Among S. aureus, blaZ was detected in 61.17% (52/85), mecA and tetK detected in 3.52% (3/85), aacA-aphD, ermC and strA genes detected in 4.7% (4/85), tetM was detected in 16.4% (14/85) and strB in 7.05% (6/85) isolates. None of the isolates contained tetO and ermA genes and 30.58% (26/85) isolates did not reveal any of the genes. Among S. epidermidis, blaZ was detected 43.47% (10/23), mecA and tetM detected in 30.43% (7/23), aacA-aphD and ermC detected in 13.04% (3/23), tetK detected in 8.69% (2/23), strA detected in 4.34% (1/23) and strB detected in 21.73% (5/23) isolates. None of them had tetO and ermA genes and 13.04% (3/23) isolates did not reveal any of the genes screened. Among E. coli, blaTEM and tetA genes were detected in 100% (47/47), blaSHV and strA were detected in 31.9% (15/47), blaOXA 30 detected in 12.76% (6/47), tetM and tetB detected in 2.12% (1/47), and strB was detected in 51.06% (24/47). None of the isolates contained aacA-aphD, mecA, tetO, tetK, ermA and ermC genes. This genetic profile of prevalence of antimicrobial genes for varied antibiotics/antimicrobials indicates the possible wider antimicrobial resistance among the predominant species of bacteria causing bovine mastitis. Further, phenotypic characterisation of these isolates with respect to their antimicrobial resistance would be of significance.

Comparative characterisation of human and ovine non-aureus staphylococci isolated in Sardinia (Italy) for antimicrobial susceptibility profiles and resistance genes.

We present the comparative characterisation of 195 non-aureus staphylococci (NAS) isolates obtained from sheep (n = 125) and humans (n = 70) in Sardinia, Italy, identified at the species level by gap gene polymerase chain reaction (PCR) followed by restriction fragment length polymorphism analysis with AluI. Isolates were tested phenotypically with a disc diffusion method and genotypically by PCR, for resistance to 11 antimicrobial agents including cationic antiseptic agents. Among the ovine isolates, Staphylococcus epidermidis (n = 57), S. chromogenes (n = 29), S. haemolyticus (n = 17), S. simulans (n = 8) and S. caprae (n = 6) were the most prevalent species, while among human isolates, S. haemolyticus (n = 28) and S. epidermidis (n = 26) were predominant, followed by S. lugdunensis and S. hominis (n = 4). Of the 125 ovine isolates, 79 (63.2%) did not carry any of the resistance genes tested, while the remainder carried resistance genes for at least one antibiotic. The highest resistance rates among ovine isolates were recorded against tetracycline (20.8%), and penicillin (15.2%); none was resistant to methicillin and two exhibited multidrug resistance (MDR); one of which was positive for the antiseptic resistance smr gene. By contrast, most human isolates (59/70, 84.3%) were resistant to ⩾1 antimicrobials, and 41 (58.6%) were MDR. All 52 (74.3%) penicillin-resistant isolates possessed the blaZ gene, and 33 of 70 (47.1%) harboured the mec gene; of these, seven were characterised by the Staphylococcal Chromosomal Cassette (SCCmec) type IV, 6 the type V, 5 of type III and one representative each of type I and type II. The majority (57.1%) was erythromycin-resistant and 17 isolates carried only the efflux msrA gene, 11 the methylase ermC gene and an equal number harboured both of the latter genes. Moreover, 23 (32.8%) were tetracycline-resistant and all but one possessed only the efflux tetK gene. qacA/B and smr genes were detected in 27 (38.6%) and 18 (25.7%) human NAS, respectively. These results underline a marked difference in species distribution and antimicrobial resistance between ovine and human-derived NAS.

Genotypic and Phenotypic Characterization, Antibiotic Resistance and Virulence Patterns of Staphylococcus aureus Isolated form Goat Mastitis

Objective: To determine the phenotypic and genotypic characterization of the virulence and antimicrobial susceptibility of S. aureus. Design: Descriptive study Samples:175 raw goat milk samples Procedures: A total of 87 S. aureus strains was isolated from goat with mastitis in north-west cost of Egypt. PCR technique was used to detect genes coding some virulence and antimicrobial properties while phenotypic antimicrobial susceptibility was determined by the disc diffusion method. Results: Most S. aureus isolates were resistant to amoxycillin (89.7%), penicillin G (88.5%), and ampicillin (86%) while were less resistant to other members of β-lactam group as cefoxitin (17.2%) and ceftriaxone (10.3%).The lowest resistant rates were toward fluroquinolone group members; ciprofloxacin, levofloxacin and ofloxacin in rates 1.2%, 2.3% and 4.5%, respectively. Tetracycline showed the highest level of phenotypic resistance (34.6%) and prevalence of a tetK gene (39.1%). The virulence factors, spa and coa exhibited prevalence rate of 6.9% and 8%, respectively with significant relationship between them. Conclusion and Clinical relevance: The problem of antimicrobial resistance needs strict measures to limit the indiscriminate use of antibiotics and concerted research efforts to limit the spread of resistant bacteria, which will negatively affect human health and control of the infectious diseases in both humans and animals.

Molecular characterization of antimicrobial resistance genes on farms and in commercial milk with emphasis on the effect of currently practiced heat treatments on viable but nonculturable formation

Despite the considerable advances that have been made to improve dairy food safety, there is rising concern that pasteurization is not sufficient for the destruction of plasmid-mediated antimicrobial resistance (AMR) genes of resistant bacteria and could stimulate bacteria to enter into a viable but nonculturable (VBNC) state. In the current study, we surveyed the prevalence of 1 genomic and 9 plasmid-mediated AMR genes in 100 samples (bulk tank milk and milk filter socks) at the farm level and 152 commercial milk samples (pasteurized and UHT milks) and assessed the VBNC state in dairy bacteria. Results revealed that sul2 was the most prevalent plasmid-mediated gene in milk filter socks (96%), bulk tank milk (48%), pasteurized milk (68%), and UHT (43%) milk; in contrast, mecA was not detected in any sample. Additionally, commercial pasteurization (as currently practiced) failed to decrease the prevalence of the blaTEM-B1 (43%), tetK (30%), and tetA (55%) plasmid-mediated AMR genes; thus, commercial pasteurization may be one of the factors creating the VBNC state in some dairy bacteria. Continued research is necessary to identify bacterial species entering the VBNC state after pasteurization, to assess their potential hazard level and shed more light on the expression and possibility of horizontal gene transfer of those plasmid-mediated AMR genes.