Oxacillin Resistance Research Articles

Increasing Incidence of Multidrug Resistance Among Cystic Fibrosis Respiratory Bacterial Isolates

Pseudomonas aeruginosa and Staphylococcus aureus are common pathogens in cystic fibrosis (CF) patients with increasing multidrug resistance (MDR). This study characterized antimicrobial susceptibility trends among organisms isolated from the respiratory tract of CF patients. Microbiological culture and sensitivity results for all CF patients were collected from January 2010 through December 2014. Minimum inhibitory concentrations were obtained using Phoenix® and Etest® methods. Clinical and Laboratory Standards Institute guidelines were used to remove duplicate isolates and develop antimicrobial susceptibility reports. MDR was defined as resistance to one agent in three or more antibiotic classes or oxacillin resistance in S. aureus. Overall, 542 bacterial isolates from 376 cultures were analyzed for trends. P. aeruginosa (41%), S. aureus (40%), and Stenotrophomonas maltophilia (8%) were the most commonly isolated organisms. Multidrug-resistant organism isolation increased from 39% to 49% (r = 0.76, p = 0.13), while representing 47.6% of all isolates. Multidrug-resistant P. aeruginosa incidence increased each year from 26% to 43% (r = 0.89, p = 0.046), while P. aeruginosa isolation decreased from 47% to 38% over the study period (r = -0.93, p = 0.02). MRSA accounted for 62.6% of all S. aureus isolated, while overall multidrug-resistant S. aureus incidence was 73.1% in all cultures. MDR among common pathogens in CF continues to increase. Empiric therapy for CF exacerbations should be targeted to previous antimicrobial susceptibility, and P. aeruginosa and S. aureus should be empirically covered.

Heterogeneous oxacillin-resistant phenotypes and production of PBP2A by oxacillin-susceptible/mecA-positive MRSA strains from Africa

Recent surveillance of MRSA colonizing patients and healthcare workers in two African countries (Angola and São Tomé and Príncipe) reported the frequent recovery of oxacillin-susceptible MRSA (OS-MRSA): Staphylococcus aureus strains that gave positive results with the mecA DNA probe, but had low oxacillin MIC values characteristic of susceptible S. aureus. This apparent dissociation of the drug-resistant phenotype from mecA-the primary genetic determinant of resistance-prompted us to perform a more detailed analysis on nine of the African OS-MRSA strains. Oxacillin MIC values were determined by Etest and population analysis profiles with and without induction of the stringent stress response by mupirocin. Biochemical profiling using SDS-PAGE followed by western blotting was used for the detection of PBP2A protein produced. Cultures of the African MRSA strains (ST88-IVa and ST8-V) showed heterogeneous oxacillin resistance in which the majority of cells exhibited low oxacillin MICs (≤0.75 mg/L), but highly resistant subpopulations were also present with oxacillin MIC values up to several hundred mg/L and with frequencies of 10(-4) to 10(-6). The same strains after induction of the stringent stress response by mupirocin 'converted' the heterogeneous phenotypes into a more homogeneous and higher level resistance. After induction by oxacillin and mupirocin, each of the nine African OS-MRSA strains produced PBP2A-the protein product of mecA. The resistant phenotype of OS-MRSA resembles the phenotypes of historically early MRSA clones. The nature of genetic determinants responsible for the heterogeneous phenotypes of OS-MRSA remains to be determined.

Ocular Pathogens and Antibiotic Sensitivity in Bacterial Keratitis Isolates at King Khaled Eye Specialist Hospital, 2011 to 2014.

To investigate prevalence and in vitro susceptibility trends of bacteria isolated from patients with bacterial keratitis from 2011 to 2014 in a tertiary care eye hospital in Saudi Arabia. Retrospective review of bacterial isolates from corneal scraping of eyes with microbial keratitis. The most common isolates and their antibiotic resistance profiles were identified; trend analysis was performed over the study period. A total of 2037 bacterial isolates met inclusion criteria during the study period. Gram positives accounted for 91.4% of isolates, including Staphylococcus epidermidis 962 (27.4%), other coagulase-negative staphylococci 289 (8.2%), Staphylococcus aureus 237 (6.8%), and Streptococcus pneumoniae 159 (4.5%). Pseudomonas aeruginosa was the most common gram-negative isolate (38.4%). All tested Gram positive isolates were sensitive to vancomycin. As a whole, isolates were most sensitive to moxifloxacin and ciprofloxacin with resistance of 3.7% and 3.3%, respectively. Oxacillin resistance was increasingly found in S. aureus (14.8% in 2011, 27.8% in 2014, P = 0.06), but was without significant change in S. epidermidis and other coagulase-negative Staphylococci (range 19.4%-32.0%). There was an increase in moxifloxacin resistance among S. epidermidis, increasing from 0.9% to 12.7%. Using a logistic regression model, the overall change in resistance of bacteria to antibiotics by year was not significant. Gram-positive bacteria represented the majority of bacteria isolated, with a possibly increasing prevalence of oxacillin resistance in S. aureus. Fluoroquinolone resistance is uncommon, and no vancomycin-resistant gram-positive strains were identified. There was no overall significant trend in antibiotics resistance; however, future surveillance studies are recommended.

Therapeutic effects of a combined antibiotic-enzyme treatment on subclinical mastitis in lactating dairy cows

The objective of this study was to evaluate a combined antibiotic-enzyme therapy for Staphylococcus aureus mastitis and biofilm formation. A total of 141 cases of S. aureus chronic mastitis from three farms were divided into two groups: the control group (n = 54) were treated with Nafpenzal® ointment; the enzyme + antibiotic group (n = 87) were treated with Nafpenzal® plus an enzymatic ointment (MastiVeyxym®). Quantitative determination of biofilm formation was determined using a colorimetric microplate assay and the detection of ica genes by PCR. Enzyme + antibiotic therapy did not significantly improve cure rates compared to control (48.3 vs 38.9%). The cure rate for infections caused by biofilm-positive strains was 42.6 and 46.6% for control and enzyme + antibiotic groups, respectively (P > 0.05). Comparison of cure rates between farms showed a relationship with somatic cell count (SCC), parity and oxacillin resistance. 79.4% of the isolates produced biofilm and antibacterial resistance rates for oxacillin, penicillin and streptomycin were 25.5, 71.6 and 95.7%, respectively. These results indicate that while the increase in mastitis cure rate using enzymatic therapy was not significant, this treatment could be useful in some situations.

Emerging Antimicrobial Resistance amongst Common Bacterial Pathogens in Mymensingh Medical College Hospital

Antimicrobial resistance has been coming up as one of the challenging business for treating domiciliary and nosocomial infections. The present study compared rate of resistance towards different commonly used antibiotics for common bacterial pathogens isolated during 2001 to 2003. Regarding Escherichia coli, mentionable increase (p<0.01) of resistance was noted against ceftazidime (47% to 77%) and ceftriaxone (43% to 71%). Imipenem (2% to 1%) and pefloxacin (40% to 17%) showed decreased trend. For Staphylococcus aureus, marked increase in resistance was shown against almost all antibiotics except co-trimoxazole (55% to 57%). Mentionable increase in resistance (p<0.05 and p<0.001, respectively) was noted against ciprofloxacin (17% to 43%) and ceftriaxone (28% to 83%). Although, oxacillin resistance increased from 22% to 42% but no resistance against vancomycin was noted during this period. Strains of Pseudomonas species showed increase (p<0.05) in resistance against ciprofloxacin (47% to 71%), ceftriaxone (50% to 74%) and ceftazidime (39% to 58%). Carbenicillin showed decreased resistance (92% to 50%) and none of the strains was found resistant to imipenem. Emphasis was given towards judicial use of antibiotics by followig local antibiogram.Bangladesh J Med Microbiol 2007; 01 (01): 04-09

Tigecycline antimicrobial activity tested against clinical bacteria from Latin American medical centres: results from SENTRY Antimicrobial Surveillance Program (2011–2014)

Bacterial organisms (n = 13,494) were consecutively collected in 2011–2014 from 21 Latin American medical centres (11 nations). Antimicrobial susceptibility was determined by broth microdilution at a central laboratory. Tigecycline was very active against Gram-positive organisms, with MIC50/90 values of 0.06/0.06 µg/mL for Staphylococcus aureus (n = 2878), 0.06/0.12 µg/mL for coagulase-negative staphylococci (n = 880), 0.06/0.06 µg/mL for enterococci (n = 708) and ≤0.03/≤0.03–0.06 µg/mL for streptococci (n = 1352). All Gram-positive species exhibited 100.0% susceptibility (FDA and/or EUCAST criteria), except for Streptococcus pneumoniae (99.8% susceptible). The S. aureus oxacillin resistance rate varied from 28.0% (Brazil) to 55.0% (Argentina), and the overall vancomycin resistance rate was 15.5% (Enterococcus faecium, 50.3%; and Enterococcus faecalis, 2.3%). The E. faecium vancomycin resistance rate varied from a low (26.3%) in Argentina to a high (71.7%) in Brazil. Against Enterobacteriaceae (n = 4543), tigecycline MIC50/90 values were 0.25/1 µg/mL; 98.3% and 94.2% of strains were considered susceptible according to FDA and EUCAST breakpoints, respectively. Overall, 37.7% and 57.3% of Escherichia coli and Klebsiella pneumoniae exhibited the CLSI ESBL screening phenotype. The highest CLSI ESBL screening phenotype rates among E. coli and Klebsiella spp. strains were observed for isolates collected from Mexico (69.9%) and Chile (69.9%), respectively. Occurrence of carbapenem-resistant Enterobacteriaceae was substantially higher in Brazil (9.0%) and Argentina (6.3%) compared with Chile and Mexico (0.4–0.7%). Tigecycline was also active against Acinetobacter spp. (MIC50/90, 1/2 µg/mL; 92.3/72.1% inhibited at ≤2/≤1 µg/mL) and Stenotrophomonas maltophilia (MIC50/90, 0.5/2 µg/mL; 91.5/83.0% inhibited at ≤2/≤1 µg/mL).

Phenotypic and genotypic detection of methicillin-resistant Staphylococcus aureus in hunting dogs in Maiduguri metropolitan, Borno State, Nigeria.

Aim:To determine the presence of MRSA in hunting dogs in Maiduguri metropolitan.Materials and Methods:Phenotypic methods used includes microscopic technique, colony morphology study, catalase-coagulase tests, and the use of mannitol salt agar test, oxacillin resistance screening agar base, and antibiotic susceptibility testing methods. Genotypic approach was used for deoxyribonucleic acid extraction, and the presence of nuc and mecA gene was detected using polymerase chain reaction (PCR) techniques.Results:Examination of 416 swab samples from nasal and perineal region of dogs revealed a total of 79.5% of S. aureus, where 62.5% of the isolates were MRSA. Molecular analysis revealed that 7nuc genes specific for S. aureus from 20 presumptive MRSA assay were all mecA PCR negative. The isolates were sensitive to gentamicin and ciprofloxacin but proved resistant to cefoxitin and oxacillin.Conclusion:High isolation rate of MRSA was found in hunting dogs. Significant level (p<0.05) of MRSA was isolated in the nasal cavity of hunting dogs than its perineum. Only nuc genes were detected from the MRSA isolates.

Characterization of canine and feline methicillin-resistant Staphylococcus pseudintermedius (MRSP) from Thailand

Methicillin-resistant Staphylococcus pseudintermedius (MRSP) in small animal practice are very difficult to treat due to multi-resistance. In contrast to other countries, little is known about MRSP from Thailand. In particular, information on feline MRSP isolates in general is rare. In total, 39 MRSP isolates from dogs (n=28) and cats (n=11) from Thailand collected from independent clinical cases were used. Oxacillin resistance and presence of the mecA gene was confirmed. Susceptibility to additional 29 antimicrobial agents was tested according to CLSI recommendations. Antimicrobial resistance genes were detected by PCR assays. Molecular typing comprised spa typing, dru typing and macrorestriction analysis with subsequent pulsed-field gel electrophoresis (PFGE). For selected isolates, multi-locus sequence typing (MLST) was performed. All isolates were multi-resistant with resistance to at least six classes of antimicrobial agents. In all cases corresponding resistance genes were detected. In addition to mecA, the genes blaZ, catpC221, aacA/aphD, erm(B), dfrG, tet(M) and tet(K) were identified. Six spa types (t02, t05, t09, t10, t23, t72), eleven dru types (dt8ak, dt10ao, dt10cp, dt10cq, dt11a, dt11bo, dt11cb, dt11cj, dt11v, dt11y, dt11z) and 27 PFGE types (designated as A1-A10, B1-B8, C1-C2, D, E, F, G, H, I, J) were identified. MLST for one isolate of each main PFGE pattern A-J revealed seven types [ST45 (n=3), ST112, ST155, ST282 and the novel types ST432, ST433 (n=2) and ST434]. This study showed that MRSP isolates from clinical cases in individual dogs and cats in Thailand are multi-resistant with similar resistance genes and characteristics as isolates from Europe and North America.

Características microbiológicas y terapéuticas de la sepsis neonatal confirmada en un hospital de Lima, Perú

To describe the microbiological and therapeutic characteristics of confirmed neonatal sepsis, and determineits incidence and mortality, at the National Hospital Arzobispo Loayza (HNAL) from 2011 to 2012, Lima, Peru. Retrospective longitudinal study. It was performed a monitoring to newborns with probable sepsis, and assessed the results from positive blood cultures. The newborns' characteristics were obtained from medical records. The acumulated incidences of confirmed sepsis, mortality and fatality rates were determined. Quantitative variables were reported as men ± standard deviation or median (interquartile range). Qualitative variables were reported as relative frequencies and percentages. The incidence of confirmed sepsis was 4.1 per 1 000 live births (95% confidence interval [CI]: 2.7-5.5). The mortality rate was 0.97 per 1 000 live births (95% CI: 0.29-1.63) and the fatality rate was 23.5% (95% CI: 9-37). The most commonly isolated organism was coagulase-negative Staphylococcus, followed by Staphylococcus aureus, both with high rates of oxacillin resistance (90% and 66.6%, respectively). A 67% of the newborns were treated using ampicillin/amikacin or ampicillin/cefotaxime. Antibiotics were changed in 58% of the newborns because of poor clinical/laboratory evolution or blood culture results. In most cases the seconds antibiotic was vancomycin (42.8%). There is a high incidence of confirmed sepsis and a high fatality rate. The administered treatments were not consistent with the microbiological profiles.

SpoVG Regulates Cell Wall Metabolism and Oxacillin Resistance in Methicillin-Resistant Staphylococcus aureus Strain N315.

Increasing cases of infections caused by methicillin-resistant Staphylococcus aureus (MRSA) strains in healthy individuals have raised concerns worldwide. MRSA strains are resistant to almost the entire family of β-lactam antibiotics due to the acquisition of an extra penicillin-binding protein, PBP2a. Studies have shown that spoVG is involved in oxacillin resistance, while the regulatory mechanism remains elusive. In this study, we have found that SpoVG plays a positive role in oxacillin resistance through promoting cell wall synthesis and inhibiting cell wall degradation in MRSA strain N315. Deletion of spoVG in strain N315 led to a significant decrease in oxacillin resistance and a dramatic increase in Triton X-100-induced autolytic activity simultaneously. Real-time quantitative reverse transcription-PCR revealed that the expression of 8 genes related to cell wall metabolism or oxacillin resistance was altered in the spoVG mutant. Electrophoretic mobility shift assay indicated that SpoVG can directly bind to the putative promoter regions of lytN (murein hydrolase), femA, and lytSR (the two-component system). These findings suggest a molecular mechanism in which SpoVG modulates oxacillin resistance by regulating cell wall metabolism in MRSA.

Clindamycin resistance among Staphylococcus aureus strains in Israel: implications for empirical treatment of skin and soft tissue infections.

The objectives of this study were to characterize isolates of Staphylococcus aureus obtained from skin and soft tissue infections in the community in Israel and to document the sensitivity patterns for commonly used antimicrobial agents. The susceptibilities of S. aureus isolates from skin and soft tissue infections in the community in Israel were reviewed to determine the appropriate empirical therapy for these infections. A total of 7221 isolates were collected during the period 2009-2012; 39% were from children (age 0-18 years). In children, S. aureus oxacillin resistance dropped from 8.4% to 3.8% (p=0.073). While inducible clindamycin resistance increased slightly from 20% to 25%, there was a prominent increase in constitutive clindamycin resistance from 0.1% to 26.8% (p=0.012). In adults, oxacillin resistance increased from 16% to 23% (p<0.001) and constitutive clindamycin resistance increased notably from 5% to 29% (p<0.001). These findings demonstrate a dramatic increase in clindamycin resistance among S. aureus isolates and suggest against the usage of clindamycin as empirical treatment for suspected S. aureus infections in Israel. Beta-lactam anti-staphylococcal agents may be given as empirical treatment for children, but should be considered according to risk factors for adults in Israel.

Colorimetric method for rapid detection of Oxacillin resistance in Staphylococcus aureus and its comparison with PCR for mec A gene.

Rapid and accurate detection of Methicillin Resistant Staphylococcus aureus (MRSA) is an important role of clinical microbiology laboratories to avoid treatment failure. The detection of MRSA is based on phenotypic assays which require at least 24 h to perform. Detection of the mecA gene or of PBP 2a is the “gold standard”, but not always available. The aim of this study was to evaluate a rapid method for detection of MRSA by using 3 (4, 5 dimethyl thiazole -2-yl) -2, 5 diphenyl tetrazolium bromide (MTT). Total 126 isolates of MRSA were collected from tertiary healthcare center and were confirmed by oxacillin screening agar test as per CLSI guidelines. Amplification of mecA gene was performed by using PCR. MTT assay was carried out for all the isolates in 96 well Microtitre plate and compared with standard methods of CLSI. Out of 126 isolates, 98 were found to be mecA positive. MTT method was found to be 98.98% sensitive and 96.43% specific. The MTT based colorimetric method is rapid and simple test for screening of oxacillin resistance in Staphylococcus aureus. It significantly shortens the time to just 7 h required to obtained a drug susceptibility test and could be useful to screen MRSA.

Antimicrobial Resistance, Biofilm Formation and mecA Characterization of Methicillin-Susceptible S. aureus and Non-S. aureus of Beef Meat Origin in Egypt.

Methicillin-resistant Staphylococcus aureus (MRSA) have been found in various farm animal species throughout the world. Yet, methicillin-susceptible S. aureus (MSSA), methicillin-susceptible non-S. aureus (MS-NSA), and methicillin-resistant non-S. aureus (MR-NSA) were not investigated. Therefore, we persued to determine the diversity in their phenotypic virulence assay, phenotypic antimicrobial resistance profile and molecular characterization in one of the food chains in Egypt. Samples were collected during 2013 from beef meat at retail. Twenty seven isolates comprising five species (S. hyicus, S. aureus, S. schleiferi subsp. coagulans, S. intermedius, and S. lentus) were characterized for their antibiotic resistance phenotypic profile and antibiotic resistance genes (mecA, cfr, gyrA, gyrB, and grlA). Out of the 27 Staphylococcus isolates only one isolate was resistant to the 12 antibiotics representing nine classes. Raw beef meat sold across the Great Cairo zone, contains 66.7% of MRS, with highest prevalence was reported in S. aureus (66.7%), while the MRS non-S. aureus strains constituted 66.7% from which S. hyicus (60%), S. intermedius (33.3%), S. schleiferi subsp. coagulans (100%), and S. lentus (100%) were MRS. Seven S. aureus, six S. hyicus, four S. schleiferi subsp. coagulans, three S. intermedius, and one S. lentus isolates although being resistant to oxacillin yet, 11/27 (40.7%) carried the mecA gene. At the same time, the cfr gene was present in 2 of the nine S. aureus isolates, and totally undetectable in S. hyicus, S. schleiferi subsp. coagulans, S. intermedius, and S. lentus. Although, global researches largely focused into MRSA and MR-NSA in animals on pigs, the analysis of our results stipulates, that buffaloes and cattle could be MRSA dispersers and that this theme is not specific to pigs. Detection of MSSA virulence determinants is a must, as although oxacillin resistance may be absent yet, the MSSA may carry the virulence determinants which could be a source of perilous S. aureus for the human community.

Role of the Stringent Stress Response in the Antibiotic Resistance Phenotype of Methicillin-Resistant Staphylococcus aureus.

Resistance to beta-lactam antibiotics in methicillin-resistantStaphylococcus aureus(MRSA) requires the presence of an acquired genetic determinant,mecAormecC, which encode penicillin-binding protein PBP2A or PBP2A', respectively. Although all MRSA strains share a mechanism of resistance, the phenotypic expression of beta-lactam resistance shows considerable strain-to-strain variation. The stringent stress response, a stress response that results from nutrient limitation, was shown to play a key role in determining the resistance level of an MRSA strain. In the present study, we validated the impact of the stringent stress response on transcription and translation ofmecAin the MRSA clinical isolate strain N315, which also carries known regulatory genes (mecI/mecR1/mecR2andblaI/blaR1) formecAtranscription. We showed that the impact of the stringent stress response on the resistance level may be restricted to beta-lactam resistance based on a "foreign" determinant such asmecA, as opposed to resistance based on mutations in the nativeS. aureusdeterminantpbpB(encoding PBP2). Our observations demonstrate that high-level resistance mediated by the stringent stress response follows the current model of beta-lactam resistance in which the native PBP2 protein is also essential for expression of the resistance phenotype. We also show that theStaphylococcus sciuri pbpDgene (also calledmecAI), the putative evolutionary precursor ofmecA, confers oxacillin resistance in anS. aureusstrain, generating a heterogeneous phenotype that can be converted to high and homogenous resistance by induction of the stringent stress response in the bacteria.

Proteomic response of methicillin-resistant S. aureus to a synergistic antibacterial drug combination: a novel erythromycin derivative and oxacillin.

The use of antibacterial drug combinations with synergistic effects is increasingly seen as a critical strategy to combat multi-drug resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA). In this work, the proteome responses in MRSA under the stress of a sub-inhibitory dose of a synergistic drug combination of a novel erythromycin derivative, SIPI-8294, and oxacillin, were studied by label-free quantitative proteomics. Several control treatment groups were designed to isolate proteome responses potentially related to the synergy: (1) the non-synergistic drug combination of erythromycin and oxacillin, (2) SIPI-8294 only, (3) oxacillin only and (4) erythromycin only. Results showed that 200 proteins were differentially expressed in SIPI-8294/oxacillin-treated cells. Among these proteins, the level of penicillin binding protein 2a, the protein mainly responsible for oxacillin resistance in MRSA, was four times lower in the SIPI-8294/oxacillin group than in the erythromycin/oxacillin group, suggesting that SIPI-8294 may interfere with this known oxacillin resistance mechanism. Moreover, hierarchical clustering analysis of differentially expressed proteins under different treatments revealed that SIPI-8294/oxacillin elicits very different responses than the individual drugs or the non-synergistic erythromycin/oxacillin combination. Bioinformatic analysis indicated that the synergistic effect can be further traced to a disruption in oxidation-reduction homeostasis and cell wall biosynthesis.

Inducible-Clindamycin Resistance in &amp;lt;i&amp;gt;Staphylococcus aureus &amp;lt;/i&amp;gt; Isolates in Rivers State, Nigeria

Clindamycin is indicated in the treatment of skin and soft-tissue infections caused by Staphylococcal species. Treatment of an infection caused by a strain carrying inducible erm gene using clindamycin or any non-inducer macrolide can lead to clinical failure. The present study was aimed to detect inducible-clindamycin resistance (MLSBi) among S. aureus isolates in Port Harcourt, Nigeriaand to study the relationship between clindamycin and methicillin-resistant <i S. aureus (MRSA).Two hundred and five (205) non-duplicate Staphylococcus aureus previously isolated from human sources were randomly collected from three health facilities- University of Port Harcourt Teaching Hospital, Braithwaite Memorial Specialist Hospital and De-Integrated Laboratories-all located in Port Harcourt, Nigeria, for this study from August, 2012 to July, 2013. Isolates were grouped as hospital in-patient (termed hospital- acquired – Nosocomial; n = 76) and out- patient cases (community-acquired; n = 129) Staphylococcus aureus . The isolates collected were reconfirmed following standard laboratory protocols. All confirmed isolates were stored in glycerol at +4°C (later sub-cultured for various phenotypic analyses). Using the disk diffusion method, detection of MRSA was carried out with 1μg of oxacillin (OXOID) placed on Mueller-Hinton agar with 4% NaCl supplementation).Antimicrobial susceptibility testing was performed using Erythromycin (15μg) and Clindamycin (2μg) both obtained from OXOID, UK. All clindamycin-sensitive isolates that were also erythromycin-resistant were subjected to D-Test phenotype (Inducible-clindamycin resistance). Among the 205 S. aureus isolates studied, Forty-four (21.5%) showed resistance to erythromycin, while 38 of these erythromycin-resistant isolates were simultaneously sensitive to clindamycin. Overall, out of 205 isolates, inducible-clindamycin resistance was detected in 23 (11.2%) of the isolates. These 23 (inducible MLSB phenotype) are among 38 erythromycin-resistant S. aureus that were simultaneously sensitive (phenotypically) to clindamycin. Ten (4.9%) of the total (205) study isolates expressed constitutive resistance to clindamycin. Oxacillin Resistance (MRSA) was detected in 25 (12.2%) of the 205 isolates. Among the 38 erythromycin-resistant S. aureus, four were MRSA while 3 (75%) of the 4 erythromycin-resistant MRSA expressed inducible resistance to clindamycin. 20 (58.8%) of 34 erythromycin-resistant MSSA expressed inducible resistance to clindamycin. MRSA phenotype was not significantly correlated (p=0.9430) to inducible-clindamycin resistance. Inducible clindamycin-resistance often leads to treatment failure. The clinical microbiology laboratories in Nigeria should consider routine testing and reporting of inducible clindamycin resistance in S. aureus . There is also the need for sustained surveillance of antimicrobial susceptibilities of S. aureus in this region.

Evaluation of Genotypic and Phenotypic Methods for Detection of Methicillin Resistant Staphylococcus aureus in a Tertiary Care Hospital of Eastern Odisha.

Methicillin resistant Staphylococcus aureus has emerged as an important pathogen in nosocomial and community acquired infections. Accurate and rapid identification of MRSA in clinical specimens is essential for timely decision of effective antimicrobial chemotherapy. The present study was conducted to compare efficacy of four conventional phenotypic methods, with mec- A based polymerase chain reaction (PCR) for MRSA identification. Methicillin resistance was determined in 200 S.aureus isolates by oxacillin disc diffusion, cefoxitin disc diffusion, Oxacillin Resistance Screening Agar and E-test. The results were compared with mec-A based PCR. Among 200 S.aureus isolates 62 (31%) were positive for mec-A gene by PCR. Cefoxitin disc diffusion, Oxacillin Resistance Screening Agar and E-test showed 100% specificity. Oxacillin disc diffusion had lowest sensitivity (82.5%) and specificity (98.5%) among all. The conventional methods take more time than PCR for diagnosing MRSA. Linezolid, Vancomycin & Dalfopristin were the highly sensitive drugs against MRSA isolates. Cefoxitin disc diffusion, is rapid, simple and cheaper, hence can be used routinely as an alternative to PCR for detection of MRSA in resource constraint laboratories.

The Staphylococcus aureus Chaperone PrsA Is a New Auxiliary Factor of Oxacillin Resistance Affecting Penicillin-Binding Protein 2A.

Expression of the methicillin-resistant S. aureus (MRSA) phenotype results from the expression of the extra penicillin-binding protein 2A (PBP2A), which is encoded by mecA and acquired horizontally on part of the SCCmec cassette. PBP2A can catalyze dd-transpeptidation of peptidoglycan (PG) because of its low affinity for β-lactam antibiotics and can functionally cooperate with the PBP2 transglycosylase in the biosynthesis of PG. Here, we focus upon the role of the membrane-bound PrsA foldase protein as a regulator of β-lactam resistance expression. Deletion of prsA altered oxacillin resistance in three different SCCmec backgrounds and, more importantly, caused a decrease in PBP2A membrane amounts without affecting mecA mRNA levels. The N- and C-terminal domains of PrsA were found to be critical features for PBP2A protein membrane levels and oxacillin resistance. We propose that PrsA has a role in posttranscriptional maturation of PBP2A, possibly in the export and/or folding of newly synthesized PBP2A. This additional level of control in the expression of the mecA-dependent MRSA phenotype constitutes an opportunity to expand the strategies to design anti-infective agents.

Presence of mecA-positive multidrug-resistant Staphylococcus epidermidis in bovine milk samples in Brazil

Bacteria of the genus Staphylococcus are one of the major pathogens causing bovine mastitis. In recent decades, resistance of this genus to oxacillin (methicillin) has been a matter of concern due to the possibility of reducing the effectiveness of mastitis treatments and the transfer of resistance determinants to other bacteria. Oxacillin resistance was studied in 170 staphylococci from bovine milk samples, including 79 Staphylococcus aureus and 91 coagulase-negative staphylococci (CNS). The susceptibility profile of 10 antimicrobial agents used in veterinary practice was determined by the Etest method. In addition to the Etest, the phenotypic characterization of oxacillin resistance was tested using the cefoxitin disk diffusion test. All isolates were screened by PCR to detect the mecA gene in 2 different regions of the gene. The isolates with an oxacillin minimum inhibitory concentration ≥0.5µg/mL or resistant to cefoxitin were identified by sequencing a 536-bp fragment of the 16S rRNA gene. This group of isolates was also evaluated for the presence of blaZ and mecC genes. Molecular analysis of the mecA gene was carried out by typing of the staphylococcal cassette chromosome mec (SCCmec). The relatedness of the mecA-positive isolates was evaluated by macrorestriction of chromosomal DNA followed by pulsed-field gel electrophoresis. With the exception of penicillin and oxacillin, 86% of the isolates showed susceptibility to cephalothin, gentamicin, erythromycin, sulfonamide, trimethoprim-sulfamethoxazole, and tetracycline. All S. aureus isolates were susceptible to oxacillin, whereas 47% (n=43) of the CNS isolates were resistant. The CNS isolates showed a higher resistance to cephalothin, erythromycin, tetracycline, and gentamicin in comparison with S. aureus. The mecA gene was only detected in 10 CNS isolates, identified as Staphylococcus epidermidis, and classified into 3 pulsotypes (A, B, and C) and 4 subtypes (A1, B1, B2, and B3). Among the isolates with an oxacillin resistance phenotype, 12 were positive for the blaZ gene, and 9 of them were mecA-positive. Two of the oxacillin-resistant isolates amplified the mecA homolog gene of Staphylococcus sciuri and none amplified mecC. Three SCCmec types, I, IV, and V, were found. Our results suggest that Staphylococcus epidermidis can be a reservoir for mecA for other Staphylococcus species. Studies investigating the molecular and phenotypic profile of antimicrobial resistance in staphylococcal species should be performed for controlling the spread of resistance and the selection of appropriate therapeutic measures.

Species distribution and in vitro antimicrobial susceptibility of coagulase-negative staphylococci isolated from bovine mastitic milk.

BackgroundCoagulase-negative staphylococci (CoNS) are the most common bovine mastitis causing bacteria in many countries. It is known that resistance for antimicrobials is in general more common in CoNS than in Staphylococcus aureus but little is known about the antimicrobial resistance of specific CoNS species. In this study, 400 CoNS isolates from bovine mastitic milk samples were identified to species level using ribotyping and MALDI-TOF MS, and their antimicrobial susceptibility was determined using a commercially available microdilution system. The results were interpreted according to the epidemiological cut-off values by the European Committee on Antimicrobial Susceptibility testing.ResultsThe most common CoNS species were S. simulans, S. epidermidis, S. chromogenes and S. haemolyticus. Penicillin resistance was the most common type of antimicrobial resistance. Staphylococcus epidermidis was the most resistant among the four major species. Almost one-third of our S. epidermidis isolates were resistant to >2 antimicrobials and close to 7 % were multidrug resistant. The majority of S. epidermidis isolates were resistant to benzylpenicillin. On the contrary, only few S. simulans isolates were penicillin-resistant. Phenotypic oxacillin resistance was found in all four main species, and 34 % of the isolates were oxacillin resistant. However, only 21 isolates (5 %) were positive for the mecA gene. Of these, 20 were S. epidermidis and one S. sciuri. mecC positive isolates were not found.Conclusion Staphylococcus epidermidis differed from the three other major CoNS species as resistance to the tested antimicrobials was common, several isolates were multidrug resistant, and 19 % of the isolates carried the mecA gene encoding methicillin resistance.Electronic supplementary materialThe online version of this article (doi:10.1186/s13028-016-0193-8) contains supplementary material, which is available to authorized users.