Genotypic characterization of methicillin-resistant Staphylococcus aureus strains isolated from the anterior nares and catheter of ambulatory hemodialysis patients in Mexico

Methicillin-resistant Staphylococcus aureus (MRSA) is the causal agent of multiple nosocomial infections worldwide, including catheter-associated bacteremia in hemodialysis patients. The purposes of this work were to genetically characterize a group of MRSA isolates from catheter-related infections of ambulatory Mexican hemodialysis patients and to determine whether the strains are the same as those carried by the patients in their anterior nares. Sixteen pairs of MRSA isolates from the catheter (cat) and anterior nares (N) of hemodialysis patients were compared using pulsed-field gel electrophoresis (PFGE), PCR detection of adhesion genes and other virulence markers, and an antibiogram. Three pairs of N/cat MRSA isolates (18.7 %) with identical resistograms also showed the same combination of PCR-detected markers and PFGE pattern; one additional pair showed only an identical electrophoretic PFGE pattern. Of the MRSA isolates, 75 % (n?=?24) were resistant to ≥7 antibiotics, 4 isolates were resistant to 11 antibiotics, and 7 isolates were resistant to the 12 antibiotics tested. The most frequent virulence marker combination found was spa, clfA, clfB, cna, bbp, ebps, map/eap, sdrC, sdrD, sdrE, ica, agr (65.6 %, n?=?21). The SCCmec alleles of the 32 MRSA isolates were IV (n?=?20), I (n?=?7), II (n?=?4), and V (n?=?1), and no SCCmec type III MRSA was found. The genotypic characterization of the MRSA isolates studied in this work will contribute to a better understanding of the virulence gene makeup of catheter-colonizing S. aureus strains and will help to lower the infection risk in these patients.     

Survey of potential factors involved in the low frequency of CP5 and CP8 expression in <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> isolates from mastitis of dairy cattle from Argentina,Chile, and Uruguay

Staphylococcus aureus produces capsular polysaccharides (CPs) both in vivo and under defined culture conditions being serotypes 5 and 8 the most prevalent. S. aureus isolates that fail to produce CP5 or CP8 are defined as non-typeable (NT). Loss of capsule expression, however, may lead to S. aureus persistence in a chronically infected host. The prevalence of NT strains of S. aureus isolated from bovine mastitis varies according to the geographic origin of the strain. The aims of this work were to detect phenotypically and genotypically the capsular profile of 144 S. aureus isolated from bovine mastitis in Argentina, Chile, and Uruguay and explore the factors that are considered to be associated with capsule expression as presence of IS257, IScap, and agr typing of non-related collection. The detection of the IS257, IScap, cap genes, and agr typing was performed using PCR. The detection and quantification of capsular polysaccharide production were performed by ELISA assays. We found that 96% of the S. aureus isolates investigated carried cap5(8) genes but over 75% of strains do not express capsule in the three countries studied. However, only 6 isolates from Argentina carried the IScap element that totally suppressed the expression of the capsule, suggesting that other factors could influence on CP expression. Moreover, the agrI/NT association was statistically significant suggesting that this profile is a phenomenon observed not only in other parts of the world but also in our region.     

Antimicrobial resistance and toxin gene profiles of Staphylococcus aureus strains from Holstein milk

Isolation of Staphylococcus aureus (Staph. aureus) from Holstein milk samples with mastitis and nonmastitis was conducted to estimate its prevalence, antimicrobial resistance and toxin genes. A total of 353 milk samples were collected from three Chinese Holstein herds. Fifty‐three Staph. aureus isolates collected from 29 Staph. aureus‐positive samples were characterized via antimicrobial susceptibility, toxin genes and Pulsed‐field Gel Electrophoresis (PFGE) profiles. The prevalence of Staph. aureus was 4·0–9·5% in mastitic and 7·3–11·5% in nonmastitic samples in the analysed herds. Approximately 61·0% of Staph. aureus strains isolated from mastitis cows were resistant to ≥10 antimicrobials compared with 0% of isolates with nonmastitis. The most frequently observed super antigenic toxin gene was pvl (41·5%) followed by seh + pvl (13·2%). We did not find mecA‐positive methicillin‐resistant Staph. aureus (MRSA) strains, while mecA‐negative MRSA strains were identified in the three herds. PFGE results suggested potential transmission of Staph. aureus strains in different farms. These results open new insights into Staph. aureus transmission and antimicrobial resistance of Holstein dairy cows and into developing strategies for udder health improvement of dairy cattle.     

Prevalence of adhesin and toxin genes among isolates of <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> obtained from mastitic cattle

Staphylococcus aureus is recognized as a major etiological agent of the most important udder disease; mastitis. The virulence potential of S. aureus isolates is attributed to a combination of extracellular factors and invasive properties that are controlled at the genomic level. In this study molecular variations were studied among the 128 S. aureus isolates obtained from the mastitic crossbred cattle. The polymorphic patterns of protein A, coagulase, and IG genes were studied in the isolates using the PCR and PCR–RFLP methods. In addition, presence of other virulence genes responsible for the expression of adhesins and toxins was also screened. A considerable genetic variation was observed in amplified fragment of SPA, IG and CLF genes. The PCR–RFLP of COA gene yielded five different patterns. Occurrence of EBP, ENO and FNBA was found to be common in both high and low virulence isolates. However, the prevalence of FNBB and toxin genes was higher in clinical than subclinical isolates. The distribution proportions of adhesin genes EBP, FIB, FNBB, CNA, BBP, MAP CAP5, and CAP8 were 60.2, 59.4, 21.9, 6.3, 5.5, 80.5, 56.3, and 22.7%; for AGR I, II, III, and IV were 44.5, 32.8, 12.5, and 10.2%; and for toxins genes HLB, SEB, SEC, SED, SEE, SEG, and SEI were 82.0, 3.1, 5.5, 3.9, 0.8, 16.4, and 55.5%, respectively. The proportions of FNBB, EBP, FIB, HLB, BBP, SEG, and SEI genes were observed more in clinical cases. The significant genetic variations in SPA, COA, IG, and CLF genes were useful to differentiate the isolates that might be valuable for mastitis reduction programme.     

2016-2017年宁夏地区牛源金黄色葡萄球菌的全基因组分析

[目的] 了解宁夏地区奶牛乳腺炎金黄色葡萄球菌（Staphylococcus aureus,SA）代表菌株的基因组序列基本特征,进一步探究其耐药基因型、毒力及进化关系,为兽医临床防治提供理论依据。[方法] 采用纸片法对97株金黄色葡萄球菌临床分离株进行抗菌药物敏感性试验,同时进行葡萄球菌蛋白A（Staphylococcus aureus protein A,spa）分型、多位点序列分型（multilocus sequence typing,MLST）,根据分型结果选取16株代表菌株进行全基因组测序,并对获得的测序序列进行处理分析。[结果] 药敏试验结果显示97株分离株对18种抗菌药物存在不同程度的耐药,其中9株耐甲氧西林金黄色葡萄球菌（methicillin-resistantStaphylococcus aureus,MRSA）对青霉素、氨苄西林、苯唑西林、头孢噻呋、磺胺异噁唑、红霉素、庆大霉素和克林霉素等8种抗菌药物完全耐药,甲氧西林敏感金黄色葡萄球菌（methicillin-sensitiveStaphylococcus aureus,MSSA）菌株对青霉素、氨苄西林、磺胺异噁唑耐药率较高。耐药基因数据库（antibiotic resistance genes database,ARDB）注释分析显示16株代表菌株共携带21种耐药基因,其中norA、tet38、bacA、mepA的携带率较高,与药敏试验结果具有一定的相关性。毒力基因数据库（virulence factors of pathogenic bacteria,VFDB）注释分析显示所有菌株携带多种与粘附、宿主免疫逃逸、分泌、胞外酶编码、铁摄取等疾病相关的毒力基因,MRSA菌株均携带较多毒力因子,MSSA菌株携带毒力因子数目不等。基因岛预测结果显示16株代表菌株存在不同数量的基因岛且MRSA菌株携带基因岛数目及毒力基因岛较多,但耐药基因岛数目与MSSA差异不明显。SNP分析结果显示部分分离株同源性较高,同源性较高的两株MRSA的全基因组基本序列特征差异较小,携带的耐药、毒力基因情况相似。[结论] 宁夏地区牛源SA分离株耐药性情况严重且具有较高的毒力水平,本研究为家畜相关MRSA（livestock-associated MRSA,LA-MRSA）与MSSA基因组序列信息的比较分析及宁夏地区SA感染的临床防控提供参考依据。     

Prevalence of multidrug-resistant Staphylococcus aureus in diabetics clinical samples

Antibiotic resistance in 40 Staphylococcus aureus clinical isolates from 110 diabetic patients (36%) was evaluated. Of these, 32 (80%) of the isolates showed multidrug-resistance to more than eight antibiotics and 35% isolates were found to be methicillin resistant S. aureus (MRSA). All 40 S. aureus strains (100%) screened from diabetic clinical specimens were resistant to penicillin, 63% to ampicillin, 55% to streptomycin, 50% to tetracycline and 50% to gentamicin. Where as low resistance rate was observed to ciprofloxacin (20%) and rifampicin (8%). In contrast, all (100%) S. aureus strains recorded susceptibility to teicoplanin, which was followed by vancomycin (95%). Genotypical examination revealed that 80% of the aminoglycoside resistant S. aureus (ARSA) have aminoglycoside modifying enzyme (AME) coding genes; however, 20% of ARSA which showed non-AME mediated (adaptive) aminoglycoside resistance lacked these genes in their genome. In contrast all MRSA isolates possessed mecA, femA genetic determinants in their genome.

     

Screening of <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> nasal strains isolated from medical students for toxin genes

Three hundred twenty-one students (156 students with no clinical exposure and 165 students with clinical exposure) were screened for nasal colonization by Staphylococcus aureus; 20.9% of students were S. aureus nasal carriers, and 40.3% of S. aureus isolates harbored toxin genes. The most prevalent genes were tst (15.0 %) and sec (13.4 %). Isolates with multiple genes were only found among clinical students (p = 0.045). Six of 11 PFGE clones were positive for toxin genes. Methicillin-resistant (MRSA) isolates were only detected in the clinical students (4.5 %). The exposure of students to the hospital environment neither radically increased S. aureus nasal carriage, nor the frequency of clinically important toxin gene presence, but it could have influenced the positive selection of toxigenic MRSA strains.     

Antimicrobial resistance and presence of <Emphasis Type="Italic">ileS-2</Emphasis> gene encoding mupirocin resistance in clinical isolates of methicillin-resistant <Emphasis Type="Italic">Staphylococcus</Emphasis> sp.

Seventy-eight staphylococcal strains were isolated from surgical-site, blood-stream and other hospital-acquired infections. Eighteen isolates were determined as methicillin (MET)-resistant S. aureus (MRSA), while the remaining were MET-resistant coagulase-negative staphylococci (CoNS). Fifty percent of CoNS strains were multiresistant, while 10 % of isolates were resistant only to β-lactams. Clinical isolates of CoNS were generally more resistant to antimicrobial agents than S. aureus strains. Thirty-nine % of S. aureus strains were resistant only to β-lactams. None of the MRSA strains carried ileS-2 gene; this gene was found in two strains of S. epidermidis.     

Clonal distribution of bone sialoprotein-binding protein gene among Staphylococcus aureus isolates associated with bloodstream infections

Staphylococcus aureus is a leading cause of bloodstream infections (BSI) and diseases that may be caused by hematogenous spread. The staphylococcal adhesin, for which the association with the infections emerging as a complication of septicemia has been well documented, is a bone sialoprotein-binding protein (Bbp). The aim of the study was to assess the prevalence of a bbp gene in S. aureus bloodstream isolates associated with BSI and to investigate to what degree the distribution of this gene is linked to the clonality of the population. Spa typing, used in order to explore the genetic population structure of the isolates, yielded 29 types. Six spa clusters and seven singletons were identified. The most frequent was spa clonal complex CC021 associated with MLST CC30 (38 %). The bbp gene was found in 47 % of isolates. Almost all isolates (95 %) clustered in spa clonal complex CC021 were positive for this gene. All isolates carrying the bbp gene were sensitive to methicillin, and if clustered in the spa CC021, belonged to agr group III. Our study shows that Bbp is not strictly associated with BSI. However, one may conclude that for clonally related S. aureus strains most commonly causing BSI, the risk of Bbp-mediated complications of septicemia is expected to be higher than for other strains.     

苯唑西林敏感、mecA基因阳性的猪源金黄色葡萄球菌的流行、分子分型及耐药性研究

[目的] 为了解我国猪源苯唑西林敏感-mecA阳性金黄色葡萄球菌（Oxacillin-susceptible,mecA-positive Staphylococcus aureus,OS-MRSA）的流行情况、菌株分子特征及耐药性,本研究对我国中西部4个省份（甘肃、陕西、河南和广西）的9个规模化养猪场进行鼻腔拭子样本采集。[方法] 运用PCR扩增nuc和mecA基因及苯唑西林耐药性检测对OS-MRSA菌株进行分离鉴定。然后对分离所得的OS-MRSA菌株进行26种毒素编码基因、16种抗生素耐药性以及spa、MLST和SCCmec分型检测。[结果] 结果表明,采集的884份样本中,67份样本7.6%（67/884）分离到金黄色葡萄球菌,包括50株甲氧西林敏感菌株（Methicillin-sensitive Staphylococcus aureus,MSSA）、8株苯唑西林耐受-mecA阳性金黄色葡萄球菌（Oxacillin-resistant mecA-positive,OR-MRSA）和9株OS-MRSA菌株。26种被检毒素编码基因中有9种毒素编码基因被检出,其中hla基因检出率最高,其次为hld、hlb、hlg、sei、sem、seg、sen和seo。此外,67株分离株中仅有16株携带肠毒素编码基因,其中OR-MRSA和OS-MRSA菌株分别占37.5%（6/16）和50.0%（8/16）,且携带毒素编码基因的菌株克隆型均为ST9-t899。16种所测试抗生素中,菌株对12种抗生素表现为耐药,其中MSSA、OR-MRSA和OS-MRSA分离株分别主要对1-8、10-12和7-11种抗生素耐药。所有分离株共有4种克隆型ST398-t571、ST9-t899、ST398-t034和t11241,其中ST9-t899为MRSA菌株唯一克隆型和ST398-t571为MSSA优势克隆型。除4株分离株未检测到SCCmec分型外,IV_b（76.5%,13/17）是MRSA分离株的唯一分型。[结论] 结果表明,我国猪源MRSA分离株对苯唑西林药物敏感性发生了改变,出现了较多的苯唑西林敏感菌株。此外,MSSA和MRSA分离株优势克隆型分别为ST398-t571和ST9-IVb-t899。研究还发现,克隆型与毒素编码基因有显著相关性,携带毒素编码基因的菌株克隆型均为ST9-t899。通过了解我国猪源MSSA、OR-MRSA和OS-MRSA的流行、分子特征和耐药性,可以为我国猪源金黄色葡萄球菌的防控提供数据支持。     

SCC<Emphasis Type="Italic">mec</Emphasis> Type IV,PVL-Negative,Methicillin-Resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> in Cystic Fibrosis Patients from Brazil

Twenty seven S. aureus isolates were obtained from cystic fibrosis (CF) patients at a tertiary care hospital in Brazil. Nineteen (70.4%) were methicillin-susceptible S. aureus (MSSA) and eight (29.6%) methicillin-resistant S. aureus (MRSA). Of the MRSA isolates, four had SCCmec type III and four had SCCmec type IV. PVL genes were not detected in any of the MSSA or MRSA isolates. New studies are necessary to evaluate the exact impact of these different MRSA clones in CF patients.     

Efficacy of Targeted 5-day Combined Parenteral and Intramammary Treatment of Clinical Mastitis Caused by Penicillin-Susceptible or Penicillin-Resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis>

Combined parenteral and intramammary treatment of mastitis caused by Staphylococcus aureus was compared to parenteral treatment only. Cows with clinical mastitis (166 mastitic quarters) caused by S. aureus treated by veterinarians of the Ambulatory Clinic of the Faculty of Veterinary Medicine during routine farm calls were included. Treatment was based on in vitro susceptibility testing of the bacterial isolate. Procaine penicillin G (86 cases due to β -lactamase negative strains) or amoxycillin-clavulanic acid (24 cases due to β -lactamase positive strains) was administered parenterally and intramammarily for 5 days. Efficacy of treatments was assessed 2 and 4 weeks later by physical examination, bacteriological culture, determination of CMT, somatic cell count and NAGase activity in milk. Quarters with growth of S. aureus in at least one post-treatment sample were classified as non-cured. As controls we used 41 clinical mastitis cases caused by penicillin-susceptible S. aureus isolates treated with procaine penicillin G parenterally for 5 days and 15 cases due to penicillin-resistant isolates treated with spiramycin parenterally for 5 days from the same practice area. Bacteriological cure rate after the combination treatment was 75.6% for quarters infected with penicillin-susceptible S. aureus isolates, and 29.2% for quarters infected with penicillin-resistant isolates. Cure rate for quarters treated only parenterally with procaine penicillin G was 56.1% and that for quarters treated with spiramycin 33.3%. The difference in cure rates between mastitis due to penicillin-susceptible and penicillin-resistant S. aureus was highly significant. Combined treatment was superior over systemic treatment only in the β -lactamase negative group.     

Isolation of methicillin-resistant Staphylococcus aureus from caprine mastitis cases

There is no report on isolation of methicillin-resistant Staphylococcus aureus (MRSA) strains from mastitis infection in goats. This study reports two MRSA strains that were isolated from caprine mastitis. A total of 42 Staphylococcus aureus (S. aureus) strains collected from caprine mastitis cases between 2008 and 2009 were examined. Two (4.8%) out of 42 S. aureus strains were identified as MRSA by Kirby–Bauer disc diffusion and mecA polymerase chain reaction (PCR) methods. Based on the coa gene polymorphism, the goat strains were grouped into 6 types. By using rapid amplified polymorphic DNA (RAPD) assay, 10 different patterns were obtained from 42 S. aureus strains, and strains were located in 6 sub-groups. A total of 71% (n = 30) of the strains were clustered in one main group and placed 4 sub-groups by RAPD assay. The two MRSA strains produced identical patterns and distinguished from other S. aureus strains by RAPD method. This paper is the first report of MRSA isolation from caprine clinical mastitis cases.     

Spa typing of Staphylococcus aureus Isolated from Clinical Specimens from Outpatients in Iraq

Methicillin-resistant Staphylococcus aureus (MRSA) is notorious as a hospital superbug and a problematic pathogen among communities. The incidence of MRSA has substantially increased over time in Iraq. The aim of this study was to determine the prevalence and spa types of MRSA isolates from outpatients or patients upon admission into hospitals. Various biochemical tests identified S. aureus isolates, and then this identification was confirmed by PCR using species-specific 16S rRNA primer pairs. Antibiotic susceptibility was determined against methicillin, oxacillin, and vancomycin using the disk diffusion method. Vancomycin MIC was detected by VITEK 2 compact system. All the identified isolates were screened for the presence of mecA and lukS-PV-lukF-PV genes; 36 of them were subjected to spa typing-based PCR. Out of 290 clinical samples, 65 (22.4%) were S. aureus, of which 62 (95.4%) strains were resistant to oxacillin and methicillin. Except for two isolates, all MRSA isolates were mecA positive. One of the three MSSA isolates was mecA positive. Five strains were resistant to vancomycin. Fourteen (21.5%) isolates were positive for the presence of lukS-PV-lukF-PV genes. Spa typing of 36 S. aureus isolates revealed eleven different spa types, t304 (30.3%), t307 (19.4%), t346 (8.3%), t044 (8.3%), t15595 (8.3%), t386 (5.5%), t5475 (5.5%), t17928 (2.8%), t14870 (2.8%), t021 (2.8%), and t024 (2.8%). These findings could be useful for assessing the genetic relatedness of strains in the region for epidemiological and monitoring purposes, which would be essential to limiting the spread of MRSA.     

Molecular characterization of methicillin-resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> nasal isolates from Turkey

Methicillin-resistant Staphylococcus aureus (MRSA) colonize most frequently in the anterior nares of the nose and cause serious infections all over the world. The aim of this study was to determine the nasal carriage rate of S. aureus and MRSA strains in Turkish elementary school children. We also analyzed molecular characterizations of MRSA strains by using pulse field gel electrophoresis (PFGE), multi locus sequence typing (MLST), staphylococcal chromosomal cassette mec (SCCmec) typing, and detection of the Panton-valentine leucocidin (PVL) gene. The nasal swabs were obtained from 4,050 children during a 4 month period in Ankara. In vitro antimicrobial susceptibility testing to 1 μg oxacillin and 30 μg cefoxitin was determined by a disk diffusion method. We found that the 1,001 of 4,050 (24.7%) children were colonized with S. aureus. Three S. aureus strains were resistant to oxacillin and cefoxitin. The rate of MRSA among all children was 0.07%. The MRSA strains revealed three different PFGE pattern. All MRSA isolates by harbored the SCCmec type IV element, but not the PVL gene. The two MRSA isolate belonged to sequence type (ST) 30, whereas the other one was a unique type. The results of this study demonstrated that S. aureus nasal carriage rate was consistent with previous studies. However, MRSA carriage rate was low. This study also indicated that the ST30-type IV without PVL gene MRSA clone may be expected to spread in Turkish community.     

Characterization of Virulence Factors and Genetic Background of Staphylococcus aureus Isolated from Peking University People’s Hospital Between 2005 and 2009

The aim of this study was to investigate both the genetic features of MRSA strains and the occurrence of virulence factors produced by Staphylococcus aureus strains isolated from Peking University People’s Hospital in Beijing, China, between 2005 and 2009. A total of 179 S. aureus strains were isolated, 139 of which were MRSA. The MRSA strains were characterized epidemiologically by SCCmec typing, spa typing and agr typing, then were classified into different genetic groups. The prevalence of genes coding for 14 exotoxins and eight adhesion factors among the S. aureus samples was assessed via polymerase chain reaction. Cluster analysis based on virulence factors-encoding gene content was performed to divide the MRSA isolates into valid clusters. Correspondence analysis was done to analyze the correlation between virulence factors clusters and genetic groups. JCSC1716-agrI-t030 (67.6%), SCCmec-IIIA-agrI-t030 (14.4%), SCCmec-IIIA-agrI-t037 (8.6%) and SCCmecII-agrII-t002 (2.2%) were four predominant MRSA clones. PVL was positive only in MSSA strains, there were at least three superantigenic toxins in our HA-MRSA clones, the prevalence of 16 virulence factors genes (sea, seb, sec, sed, seg, sei, sej, pvl, lukE-lukD, eta, bbp, can, ebp, clfA, fib, fnbB) in MRSA and MSSA was found to be significantly different from MSSA (P < 0.05). Results of correspondence analysis among clusters based on virulence factors genes and groups based on genetic typing illustrated not only the correspondence relationship between groups and clusters overall (P < 0.001), but also the genetic diversity of MRSA strains with respect to virulence factors genes.     

Detection of Malaysian methicillin-resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> (MRSA) clinical isolates using simplex and duplex real-time PCR

The aim of this study was to develop a methicillin-resistant Staphylococcus aureus (MRSA) detection method based on the melting temperature analysis profiling of S. aureus clinical isolates from three different hospitals in Malaysia. Simplex and duplex real-time PCR assay was used for the simultaneous detection of nuc (species-specific) and mecA (methicillin-resistance) genes in a single SYBR Green I real-time PCR tube assay. Evaluations were based on the melting temperature (T _m) analysis of the amplicons using 23 S. aureus clinical isolates including three ATCC S. aureus standard strains. Real-time PCR amplification products with melting peaks at 78.39 ± 0.4°C and 74.41 ± 0.6°C were detected for nuc and mecA genes, respectively. Each real-time PCR assay was completed within two hours. This rapid genotypic method is useful for the detection of resistant determinant (mecA) and identification of S. aureus (nuc) clinical isolates, thus benefiting patient therapy in hospitals.     

Antibacterial Effects of Green Tea Polyphenols on Clinical Isolates of Methicillin-Resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis>

The antibacterial effects of tea polyphenols (TPP) extracted from Korean green tea (Camellia sinensis) against clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) were evaluated. Characterization of the minimal inhibitory concentration (MIC) of oxacillin for 30 S. aureus strains isolated from patients treated with oxacillin identified 13 strains with an oxacillin MIC ≥ 4 μg/mL as methicillin-resistant Staphylococcus aureus (MRSA) (range: 8 to 512 μg/mL), while 17 strains were methicillin-susceptible Staphylococcus aureus (MSSA) (range: 0.25–0.5 μg/mL). The MICs of TPP ranged from 50 to 180 μg/mL for both the MSSA and the MRSA strains. The MICs of oxacillin for each of the 13 MRSA strains were reduced between 8- and 128-fold when these strains were coincubated with sub-MIC (≤0.5× MIC) levels of TPP, demonstrating that the combination of TPP plus oxacillin was synergistic for all of the clinical MRSA isolates. Two-dimensional polyacrylamide gel electrophoresis identified 14 extracellular proteins of MRSA-13 down-regulated and 3 proteins up-regulated by exposure to TPP. These studies demonstrate that TPP can differentially stimulate the expression of various proteins in these bacteria and synergize the bactericidal activity of oxacillin for MRSA.     

Nasal Staphylococcus aureus and S. pseudintermedius carriage in healthy dogs and cats: a systematic review of their antibiotic resistance,virulence and genetic lineages of zoonotic relevance

The molecular ecology of Staphylococcus aureus, Staphylococcus pseudintermedius and their methicillin-resistant strains in healthy dogs and cats could serve as good models to understand the concept of bacterial zoonosis due to animal companionship. This study aims to provide insights into pooled prevalence, genetic lineages, virulence and antimicrobial resistance (AMR) among healthy dogs and cats. Original research and brief communication articles published from 2001 to 2021 that reported the nasal detection of S. aureus and S. pseudintermedius in healthy dogs and cats in the community, homes and outside veterinary clinics were examined and analysed. Forty-nine studies were eligible and included in this systematic review. The pooled prevalence of nasal carriage of S. aureus/methicillin-resistant S. aureus (MRSA) in healthy dogs and cats were 10.9% (95% CI: 10.1–11.9)/2.8% (95% CI: 2.4–3.2) and 3.2% (95% CI: 1.9–4.8)/0.5% (95% CI: 0.0–1.1), respectively. Conversely, the pooled prevalence of S. pseudintermedius/methicillin-resistant S. pseudintermedius (MRSP) in healthy dogs and cats were 18.3% (95% CI: 17.1–19.7)/3.1% (95% CI: 2.5–3.7) and 1.3% (95% CI: 0.6–2.4)/1.2% (95% CI: 0.6–2.3), respectively. Although highly diverse genetic lineages of S. aureus were detected in healthy dogs and cats, MSSA-CC1/CC5/CC22/CC45/CC121/CC398 and MRSA-CC5/CC93/CC22/CC30 were mostly reported in dogs; and MSSA-CC5/CC8/CC15/CC48 and MRSA-CC22/CC30/CC80 in cats. Of note, MSSA-CC398 isolates (spa-types t034 and t5883) were detected in dogs. Genetic lineages often associated with MSSP/MRSP were ST20/ST71, highlighting the frequent detection of the epidemic European MRSP-ST71 clone in dogs. S. aureus isolates carrying the luk-S/F-PV, tst, eta, etb and etd genes were seldomly detected in dogs, and luk-S/F-PV was the unique virulence factor reported in isolates of cats. S. pseudintermedius isolates harbouring the luk-S/F-I, seint and expA genes were frequently found, especially in dogs. High and diverse rates of AMR were noted, especially among MRSA/MRSP isolates. There is a need for additional studies on the molecular characterization of isolates from countries with under-studied nasal staphylococci isolates.     

Development and application of a triplex-PCR assay for rapid detection of methicillin-resistant Staphylococcus aureus from pigs

A triplex-PCR assay was developed and evaluated for rapid detection of methicillin-resistant Staphylococcus aureus (MRSA) recovered from various biological samples of pig. Three sets of primers were designed to target mecA, 16S rRNA and nuc genes of MRSA. The specific amplification generated three bands on agarose gel, with sizes 280 bp for mecA, 654 bp for 16S rRNA and 481 bp for nuc, respectively. A potential advantage of the PCR assay is its sensitivity with a detection limit of 10² CFU per ml of bacteria. In all, 79 MRSA isolates recovered from various samples of pigs were subjected to the amplification by the triplex-PCR assay and all the isolates yielded three bands corresponding to the three genes under this study. No false-positive amplification was observed, indicating the high specificity of the developed triplex-PCR assay. This assay will be a useful and powerful method for differentiation of MRSA from methicillin-sensitive S. aureus, coagulase-negative methicillin-resistant staphylococci and coagulase-negative methicillin-sensitive staphylococci.