Phenotypic and genotypic study on antibiotic resistance and pathogenic factors of Staphylococcus aureus isolates from small ruminant mastitis milk in South of Italy (Sicily) (original) (raw)
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Staphylococcus aureus is commonly associated with mastitis in dairy herds with potential public health implications. This study was conducted to investigate the existence of S. aureus in mastitic milk and to determine the antimicrobial resistance profiles of the isolated strains as well as the resistance and virulence associated genes. Materials and Methods: Two hundred quarter milk samples were collected from 3 dairy farms at Dakahliya (n = 2) and Damietta (n = 1) Governorates, Egypt from September to December 2016. Conventional culturing and Polymerase Chain Reaction (PCR) assays targeting nuc (thermonuclease) and coa (coagulase) genes were performed. Isolates were tested for its susceptibility against 14 antimicrobial agents using disk diffusion method. All the isolates were screened for the presence of $-lactamases ( blaZ, mec A) and virulence associated ( pvl and tst ) genes by PCR. Results: The S. aureus was detected in 42% (84/200) of the total examined milk samples. Regarding the antibiogram results, S. aureus revealed a high resistance against ampicillin (95.2%) and penicillin (83.3%) and a lower resistance was observed against gentamicin (23.8%), amikacin (16.7%) and ciprofloxacin (14.3%). Multidrug resistances were detected in 83.3% of the isolated S. aureus . Of the 70 penicillin-resistant S. aureus isolates, blaZ gene was identified in 67 (95.7%) isolates. Fifty percent of S. aureus isolates harbored the specific amplicon of mec A gene. Markedly, all mec A positive strains displayed multidrug resistance and were also positive for blaZ gene. The virulence determinants pvl and tst were detected in 7.1 and 11.9% of the isolated S. aureus , respectively. Conclusion: Presence of multidrug resistant and toxin producing S. aureus in dairy farms pose a major risk to public health. Therefore, this study highlighted the importance of developing an efficient control program to inhibit the transmission of S. aureus , particularly multidrug resistant strains to humans.
This study was aimed to examine the recent trends of antibiotic resistance (AR) prevalence in Staphylococcus aureus isolated from milk of animals with clinical mastitis in areas of the Abruzzo and Molise regions in central Italy. Fifty-four S. aureus isolates could be obtained from routine testing for clinical mastitis agents carried out in the author institution in years 2021 and 2022. These were analyzed for phenotypic resistance to eight antibiotics recommended for testing by European norms and belonging to the antibiotic classes used for mastitis treatment in milk producing animals. Moreover, the presence of 14 transferable genetic determinants encoding resistance to the same antibiotics was analyzed by qPCR tests developed in this study. Phenotypic resistance to non-β-lactams was infrequent, with only one 2022 isolate resistant to clindamycin. However, low level resistance to the β-lactam cefoxitin was observed in 59.2% isolates in both years making these isolates classifiable ...
Journal of Applied Veterinary Sciences
The objectives of the study were to detect toxins and antiseptic resistance genes in Staphylococcus aureus isolated from cows with subclinical mastitis in Egypt. A total of 400 quarter milk samples (QMS) were collected from different dairy herds in which quaternary ammonium compounds (QAC) had been used as a disinfectant for more than 3years. The collected samples were subjected to bacterial investigation. S. aureus was successfully isolated confirmed by duplex PCR targeting 16S rRNA and nuc genes. Also determined their antibiogram and sensitivity to disinfectant. Genes of QAC(qacA/B), enterotoxins (Sea, Seb) and exfoliative toxins (ETB) were detected by simplex and multiplex PCR. Results of bacterial investigation revealed 103 (25.75%) S. aureus isolates. Results of antibiogram demonstrate that the most microbial antibiotics resistance were recorded for Penicillin G (85.7%) and Tetracycline (54.2%). While Gentamycin, Neomycin and Amoxicillin+ clavulanic acid show moderate resistance (21.4%, 10% and 7.1%) respectively, although Norfloxacin and Cephradine exhibited seldom resistance with high sensitivity of 95% and 94.3% respectively. Regarding the results of QAC sensitivity, only 8 isolates (7.76%) were resistant to benzalkonium chloride (BC) versus to 13 isolates (12.62%) harbour QAC gene could be detected by PCR with specific amplicon of 220bp corresponding to qacA/B. The results revealed Positive amplification of 102 bp specific for Sea gene in 19(18.44%) isolates and 164bp specific for Seb gene in 13(12.62%) isolates while there is no amplification was detected for etb gene. In conclusion, Antibiogram, as well as the identification of toxigenic and QAC genes in this study, may open another perspective in planning some alternative therapeutic strategies against multi resistances S. aureus mastitis. Monitoring crossresistance between antibiotics and antiseptic should be further investigated.
Current Microbiology, 2010
Staphylococcus aureus is one of the major causes of mastitis in dairy animals and its resistance against multiple antimicrobials always remains crucial concern. Present investigation was carried out to detect the distribution of antibiotic-resistant genes of S. aureus isolates. Isolates (128) of S. aureus from mastitic milk were collected, tested for antibiotics with disc-diffusion method, and resistant genes mecA, linA, msrA msrB, vatA, vatB, vatC ermA, ermC tetK, tetM and aacA-D were detected by PCR. The phenotypic antibiotics resistance percent in S. aureus isolates was classified as tetracycline (36.7), gentamycin (30.5), streptomycin (26.6), kanamycin (25.8) and penicillin G (22.7). All the isolates were susceptible to vancomycin. Among isolates, 10.2% were observed as methicillin-resistant. The distribution of antibiotic-resistant genes was linA (51.6) followed by msrB (46.1), tetK + M (34.4), msrA and aacA-D (26.6%). Different antibiotic-resistant genes combinations (mecA/linA-2; mecA/aacA-D/tetK/linA/msrB-3; mecA/linA/msrA/msrB-3; aacA-D/linA/msrA/msrB-4; aacA-D/linA/msrB-7; linA/msrA/msrB-10; tetK/linA/msrA/msrB-11; aacA/tetK/linA/msrB-12 isolates) were observed. All the isolates lacked amplification of vatA, vatB, ermA and ermC genes. Molecular typing resulted genetic variation in protein A (6–12 repeats) and coagulase genes (A–E patterns) were observed. Coagulase A and D genotypes were more prevalent in antibiotic-resistant isolates, while E, B and C in susceptible ones. The significant observation was the prevalence of methicillin-resistant S. aureus, which were resistant to multiple antibiotics. Findings revealed the status of resistant isolates in herd that might be helpful in treatment, controlling of resistant strains and culling of cows for mastitis reduction.
Genetic Characterization of Resistant S. Aureus Strains Isolated From Cow’s Milk with Mastitis
Anglisticum Journal, 2015
Mastitis therapy is generally unsuccessful due to pathological changes that occur in the breast as a result of the inflammatory response, factors associated with the causes of mastitis, the pharmacokinetic properties of antimicrobial drugs, etc. The main reason for the low effectiveness of antibiotic treatment of mastitis caused by staphylococci is especially, among other things, the resistance of bacteria, where is recognized that Staphylococcus aureus is the first penicillinresistant bacteria. Laboratory techniques based on phenotypic characteristics of resistance have low sensitivity. PCR (Polymerase Chain Reaction) as a method for detecting gene mecA is considered as gold standard test. Detection of mecA gene through PCR is one of the most effective methods for the detection of MRSA. Using this method we identified six methicillin-resistant strains of S. aureus isolated from cow's milk with mastitis. Our study presents the first survey carried out in our country regarding the presence of the mecA gene for antibiotic-resistant strains of S. aureus which is isolated from cow's mastitis. This study has a special importance for food products of animal origin, like milk contaminated with MRSA may constitute a main source of consumer's infection. Mastitis therapy is generally unsuccessful due to pathological changes that occur in the breast as a result of the inflammatory response, factors associated with the causes of mastitis, the pharmacokinetic properties of antimicrobial drugs, etc. The main reason for the low effectiveness of antibiotic treatment of mastitis caused by staphylococci is especially, among other things, the resistance of bacteria, where is recognized that Staphylococcus aureus is the first penicillin-resistant bacteria. Laboratory techniques based on phenotypic characteristics of resistance have low sensitivity. PCR (Polymerase Chain Reaction) as a method for detecting gene mecA is considered as gold standard test (Éva Juhász-Kaszanyitzky 2007).
Antibiotic resistance gene typing in Staphylococcus aureus isolated from bovine mastitis
The study was conducted to determine the antimicrobial resistance pattern of Staphylococcus aureus isolates from bovine mastitis. Milk samples (160) collected aseptically from mastitis affected cows at organized dairy farms in and around Jammu were subjected to microbial culture for the isolation and identification of S. aureus using Baird Parker Agar. Presumptive S. aureus isolates (52) were subjected to molecular confirmation through identification of species specific (nuc) gene. In vitro antimicrobial resistance pattern of the isolates against a panel of 13 selected antibiotics, using disc diffusion technique, revealed that the isolates were mostly sensitive to enrofloxacin followed by vancomycin, ciprofloxacin, tetracycline, chloramphenicol, streptomycin, ceftriaxone, erythromycin and gentamicin whereas maximum resistance was shown towards penicillin G followed by ampicillin, amoxyclav, methicillin, gentamicin, streptomycin, erythromycin, tetracycline and ceftriaxone. Methicillin resistance (MRSA) was recorded in 32.69% S. aureus isolates out of which 41.17% isolates carried mecA gene. Among the gentamicin and tetracycline resistant S. aureus isolates, 61.53% isolates carried aacA-aphD gene and 80% isolates carried tetK gene, respectively. Multidrug resistance (MDR) was observed in 71.15% S. aureus and 82.35% MRSA isolates. In conclusion, S. aureus showed maximum sensitivity to enrofloxacin thereby suggesting the use of this drug for effective treatment of mastitis but the development of resistance against this drug cannot be ruled out in the near future, hence, there is a need for accurate diagnosis of mastitis along with the correct selection of antibiotics to prevent bovine mastitis.
Journal of Dairy Science, 2019
Staphylococcus aureus can be associated with bovine mastitis, leading to high financial losses in dairy industry worldwide. In addition, milk products are one of the most common food categories implicated in Staphylococcal Food Poisoning in humans. In this study, we assessed the population structure, phenotypic resistance patterns and virulence and resistance gene profiles of 58 S. aureus strains isolated from bovine mastitis milk in Switzerland. DNA microarray was used to test for virulence and resistance genes, while minimum inhibitory concentrations of various antimicrobial agents were determined by microdilution. We determined clonal complexes and performed multilocus sequence typing and spa typing. The strains were assigned to seven clonal complexes, 10 sequence types and 11 spa types, with CC705 (43%), CC97 (33%), and CC20 (12%) representing the most common lineages and t529 (43%) and t267 (21%) representing the most common spa types. Only one isolate was assigned to CC8, which is linked to high within-herd prevalence of mastitis. A total of 14% (n = 8) of strains was classified as resistant to penicillin and one strain each was classified as oxacillin and pirlimycin resistant. One strain assigned to CC20, ST389 and t2094 exhibited resistance to penicillin, oxacillin, and pirlimycin as well as intermediate susceptibility to erythromycin.
Brazilian Journal of Veterinary Research and Animal Science, 2021
Fifty-two Staphylococcus aureus recovered from papillary ostium and milk samples collected from cows with subclinical mastitis and milking environments in three small dairy herds located in southeastern Brazil were subjected to PCR identification based on the thermonuclease (nuc) gene. All the strains were submitted to in vitro antimicrobial susceptibility testing, and we investigated the sequence types (STs), agr groups (I-IV), virulence genes encoding for Microbial Surface Components Recognizing Adhesive Matrix Molecules (MSCRAMMs), biofilm-associated proteins, bi-component toxins, pyrogenic toxin superantigens, and enterotoxins. Screening for oxacillin resistance (2-6 μg/ml oxacillin), beta-lactamase activity assays, and PCR for the mecA/mecC genes detected 26 methicillin-susceptible S. aureus (MSSA) and 26 mec-independent oxacillin-nonsusceptible S. aureus (MIONSA). While MSSA isolates were found to be susceptible to all antimicrobial agents tested, or only resistant to penicill...
Transboundary and Emerging Diseases, 2010
Staphylococcus aureus is commonly associated with mastitis in dairy herds with potential public health implications. Overall, 303 samples were collected from September 2015 to July 2016 to characterize the phenotypic and genotypic pattern of drug resistance in S. aureus isolated from cases of clinical and sub-clinical bovine mastitis in Central Ethiopia. Milk samples were tested by using California Mastitis Test and positive samples were subjected for bacterial culture, disc diffusion test and polymerase chain reaction (PCR) to detect the presence of antimicrobial resistance. Based on California mastitis test (CMT) result and clinical examination, the prevalence of mastitis was 70.6%. S. aureus was isolated from 36.9% of CMT positive samples. The phenotypic determination of antimicrobial resistance showed that the isolates were most resistant to ampicillin (80%) followed by trimethoprim-sulfamethoxazole (23.3%), tetracycline (15%), streptomycin (10%) and gentamycin (3.3%) and equally to both erythromycin and chloramphenicol (1.6%). Characterization of the antimicrobial resistance gene was done by using PCR. Most of the isolates (56%) contained blaZ gene followed by ermB (33%), ermC (13.3%) and each ermA and msrA appeared only in 2% of the isolates. There was no isolate harboring the methicillin resistance mecA gene. Thirty six percent of the isolates contained more than one antibiotic resistance genes. The highest multidrug resistance (MDR) gene combination was observed by blaZ*ermB (31.25%) genes and the least frequently occurred were blaZ *ermA and msrA*ermB (3.12%) each. This study showed that consumption of raw milk could be considered as a critical source of antibiotic resistant S. aureus.