Pastorex Staph-Plus Research Articles

Staphylococcus aureus small colony variants: A potentially underestimated microbiological challenge in peritoneal dialysis

IntroductionPeritonitis remains the major infectious complication in the setting of peritoneal dialysis (PD). Despite known only moderate pathogenicity, the most frequently detected pathogens in PD-related peritonitis are surprisingly coagulase-negative staphylococci. However, this could be explained, at least in part, by Staphylococcus aureus small colony variants (SCVs) induced by PD fluids (PDFs) and misidentified by routinely used microbiological methods. Material and methodsBacteria were exposed to commonly used PDFs in various regimens designed to simulate daily use as closely as possible. Wild-type isolates and SCVs were subsequently used to determine minimum inhibitory concentrations (MICs), in vitro biofilm formation capacities, and auxotrophies. Underlying genetic alterations were investigated using whole-genome sequencing, and various microbial identification methods were tested to determine their performance for wild-types and SCVs. ResultsStable SCVs could be isolated most successfully after exposure to glucose-containing PDFs alone. The reading of MICs was significantly affected by the reduced growth of SCVs, resulting in lower MIC values in 44% of all tests. Nonsynonymous mutations were found in all but one SCV, while only two isolates showed typical auxotrophic responses. While MALDI-TOF, PCR and Pastorex Staph-Plus correctly identified all S. aureus SCVs, API-Staph and VITEK-2 yielded identification rates of only 40% and 10%, respectively. ConclusionsOverall, the present study has shown that commercially available PDFs induce S. aureus SCVs in vitro, which are difficult to identify and test for antimicrobial susceptibility and can potentially lead to recurrent or persistent infections. Thus, they represent a potentially underappreciated challenge not only for microbiologists, but also for clinicians.

Denture stomatitis associated with small-colony variants of Staphylococcus aureus: a case report

BackgroundThe small-colony variants (SCVs) of Staphylococcus aureus were isolated from persistent and recurrent infections, especially after placement of medical devices having direct contact with human tissues. The emergence of SCVs is a survival strategy of S. aureus which enables them to hide inside host’s cells and induces a less severe immune response than to wild-type S. aureus. However, contrary to other medical devices, dental prosthesis as a surface potentially colonized by SCVs of S. aureus has not been examined thus far. We reported the first case of SCVs - S. aureus infection in denture wearer.Case presentationA 62-year-old woman with a complete removable acrylic denture presented extensive elevated erythematous lesions on the palate, compatible with denture stomatitis. The patient had a history of arterial hypertension, cigarette smoking and wearing denture at night. The fungal colonies, identified as Candida albicans, were cultured on Sabouraud agar. From three swabs (from hard palate mucosa, denture surface and angular cheilitis lesions) were cultured of pinpoint, clear, non-pigmented, and non-haemolytic colonies on Columbia agar. The small colonies turned out to be Gram-positive cocci, catalase-, Pastorex Staph Plus -, and clumping factor-positive, and oxidase-negative. Suspected phenotypically SCVs forms were definitively identified as S. aureus based on PCR amplification of species specific nuc and coa genes. Methicillin-resistance was verified by mecA gene detection. The isolates turned out to be susceptible to methicillin (MSSA) and resistant to gentamicin. The isolate was identified as menadione-auxotrophic variant.ConclusionsThis case demonstrated that oral cavity in denture wearers may be a reservoir of small-colony variants of S. aureus, besides C. albicans. The prevalence of these bacteria and their role in the pathogenesis of oral diseases are not understood. Due to problems with their detection and identification, the true prevalence of oral SCVs may be underestimated.

High prevalence of methicillin resistant staphylococci strains isolated from surgical site infections in Kinshasa.

IntroductionSurgical site infections (SSIs) after surgery are usually caused by Staphylococcus aureus and coagulase-negative staphylococci (CNS). In low income countries, methicillin resistant Staphylococcus aureus (MRSA) and methicillin resistant coagulase-negative staphylococci (MR-CNS) surgical site infections are particularly associated with high treatment cost and remain a source of mortality and morbidity. This study aimed to determine the prevalence and the sensitivity to antibiotics of MRSA and MR-CNS isolated from SSIs.MethodsWound swabs were collected from 130 hospitalized surgical patients in two major hospitals of Kinshasa. S. aureus and CNS strains were identified by standard microbiological methods and latex agglutination test (Pastorex Staph-Plus). The antibiotic susceptibility of all staphylococcal strains was carried out using disk-diffusion method.ResultsEighty nine staphylococcal strains were isolated. Out of 74 S. aureus and 15 CNS isolated, 47 (63.5%) and 9 (60%) were identified as MRSA and MR-CNS respectively. Among the MRSA strains, 47 strains (100%) were sensitive to imipenem, 39 strains (89%) to amoxycillin-clavulanic acid and 38 strains (81%) to vancomycin. All MR-CNS were sensitive to imipenem, amoxycillin-clavulanic acid and vancomycin. The isolated MRSA and MR-CNS strains showed multidrug resistance. They were both resistant to ampicillin, cotrimoxazole, erythromycin, clindamycin, ciprofloxacin, cefotaxime and ceftazidime.ConclusionThe results of the present study showed a high prevalence of MRSA and MR-CNS. Imipenem, amoxycillin-clavulanic acid and vancomycin were the most active antibiotics. This study suggests that antibiotic surveillance policy should become national priority as MRSA and MR-CNS were found to be multidrug resistant.

Evaluation of a fourth-generation latex agglutination test for the identification of Staphylococcus aureus

In this study, we evaluated a fourth-generation agglutination assay (Staph Plus; DiaMondiaL[DML]) for the rapid identification of Staphylococcus aureus. First, comparison with three third-generation assays (Slidex Staph Plus, bioMérieux; Staphaurex Plus, Murex Diagnostics; Pastorex Staph-Plus, Sanofi Diagnostics Pasteur) was performed on a predefined strain collection: 265 coagulase-negative staphylococci (CNS), 266 methicillin-resistant S. aureus (MRSA) and 262 methicillin-susceptible S. aureus (MSSA) strains (“strain study”). Second, patient material-derived strains (883 CNS, 847 MSSA and 135 MRSA) were tested concurrently with both the DML and Slidex assays (“daily practice study”). In the strain study, the overall sensitivity and specificity of the DML, Slidex, Staphaurex and Pastorex assays were 99.2% and 100%, 98.1% and 100%, 95.2% and 100%, and 98.2% and 98.8%, respectively. Using the respective tests, the result was indeterminate in 0.0%, 0.6%, 0.4% and 1.5% of the strains. Overall, the sensitivity of the DML and Slidex assays were comparable in both sub-studies. However, in MRSA strains, the sensitivity of the DML assay was significantly lower than the Slidex assay. The specificity of the Slidex assay was significantly higher than the DML assay. However, the percentage of indeterminate results was much higher for the Slidex than the DML assay. In conclusion, the presumptive identification of S. aureus by the DML assay proved to be equal to third-generation latex agglutination assays.

Evaluation of a new Staphylococcus aureus latex agglutination kit, Prolex Staph Xtra, against other third-generation kits

Staphylococcus aureus, including methicillin-resistant strains, continues to be a common cause of infection/colonisation, which necessitates accurate and prompt diagnosis in the laboratory. Several rapid agglutination tests that aid this function are available, and some have been modified to improve their performance. One such kit, Prolex Staph Xtra, has been released recently. This study aims to compare this kit with other improved kits (i.e., Pastorex Staph-Plus, Staphaurex Plus and Staphytect Plus) and investigate their ability to confirm the identity of 100 strains of S. aureus. Results showed that 50 were resistant to methicillin. Specificity was checked against 30 strains of coagulase-negative staphylococci and 20 Enterococcus species isolates. Of the four kits tested, Prolex Staph Xtra and Pastorex Staph-Plus proved superior in terms of sensitivity and speed.

Combined Use of Pastorex Staph-Plus and Either of Two New Chromogenic Agars, MRSA ID and CHROMagar MRSA, for Detection of Methicillin-Resistant Staphylococcus aureus

We describe the search toward a fast and reliable strategy to detect and confirm the presence of methicillin-resistant Staphylococcus aureus (MRSA) in screening samples. First, we evaluated the sensitivities and specificities of oxacillin resistance screening agar (ORSA) with enrichment (tryptic soy broth [TSB] and ORSA [TSB-ORSA]) and without enrichment (ORSA), MRSA ID (MRSA_ID) plates, and CHROMagar MRSA (C_MRSA) plates, all of which were inoculated with equal volumes of a suspension made by emulsifying screening swabs. Whereas the sensitivities after 48 h were similar for all media tested (77% for MRSA_ID and ORSA; 73% for C_MRSA and ORSA after enrichment [TSB-ORSA]), the specificities of MRSA_ID (98% after 24 h and 94% after 48 h) and C_MRSA (98% after 24 h and 90% after 48 h) were superior to the specificities of ORSAs (92% after 24 h and 83% after 48 h) and TSB-ORSA (86% after 24 h and 81% after 48 h). Subsequently, the performance of the Pastorex Staph-Plus agglutination test with presumptive MRSA isolates taken directly from chromogenic agars (direct_Pastorex agglutination) was compared to that of the Pastorex Staph-Plus agglutination test with isolates from blood agar subcultures (conventional_Pastorex agglutination). When the direct_Pastorex agglutination test on MRSA_ID plates was combined with Gram staining, the direct_Pastorex agglutination test with samples from MRSA_ID plates was as reliable as the conventional_Pastorex agglutination test with samples from blood agar subcultures from MRSA_ID plates. In contrast, the direct_Pastorex agglutination test with samples from C_MRSA plates gave false-negative results. Finally, we calculated the processing times of the four different strategies, namely, (i) enrichment in TSB supplemented with NaCl, subsequent culture on ORSA, and the conventional_Pastorex agglutination test; (ii) direct inoculation of ORSA combined with conventional_Pastorex agglutination test; (iii) direct inoculation of MRSA_ID plates combined with Gram staining and the direct_Pastorex agglutination test; and (iv) direct inoculation of C_MRSA plates combined with Gram staining and the direct_Pastorex agglutination test. We concluded that the use of MRSA_ID in combination with Gram staining and the direct_Pastorex agglutination test is faster and more specific than the other strategies tested.

Evaluation of six agglutination tests for Staphylococcus aureus identification depending upon local prevalence of meticillin-resistant S. aureus (MRSA)

Most routine laboratory detection of Staphylococcus aureus isolates is based on rapid agglutination test systems. Failure of agglutination assays to identify meticillin-resistant S. aureus strains (MRSA) has been demonstrated. The aim of this study was to evaluate six commercially available agglutination tests for the detection of meticillin-sensitive S. aureus (MSSA) and mecA-positive MRSA strains. The Dry Spot Staphytect Plus test (Oxoid), the Pastorex Staph Plus test (Bio-Rad), the Slidex Staph-Kit and Slidex Staph Plus test (bioMérieux), the Staphaurex Plus test (Remel) and the Staphylase Test (Oxoid) were used. As determined by pulsed field gel electrophoresis, 52 distinct MRSA strains from five countries, 83 MSSA strains and 150 coagulase-negative staphylococci were included. Species identification and determination of susceptibility patterns were performed using colony morphology, Gram stain, catalase testing, tube coagulase testing, DNase testing, mannitol fermentation, susceptibility testing towards oxacillin by Etest, coagulase gene PCR, fibrinogen receptor gene PCR and PCR of the mecA gene. Sensitivity of the agglutination tests ranged from 82.7 to 100.0 % for MRSA strains and 92.8 to 100.0 % for MSSA strains, respectively. Specificity of the test systems ranged from 91.3 to 99.1 %. None of the six agglutination assays produced correct reactions for all staphylococci tested. Only the Dry Spot Staphytect Plus test correctly identified all 52 MRSA strains. For the other tests kits, sensitivity of MRSA detection was lower than for MSSA isolates. Depending upon the local MRSA prevalence and the parameter of interest (sensitivity or specificity), these test systems may be useful for routine diagnostic purposes.

Evaluation of screening media for detection of methicillin-resistant Staphylococcus aureus (MRSA) from surveillance swabs

OBJECTIVES: Highly sensitive and cost-effective screening media that aid in a rapid turnaround time (TAT) are of prime importance in the control of MRSA. Good specificity is also paramount, as false positives significantly increase labor and material costs. METHODS: This study compares selective CHROMagar MRSA™ (BBL, Sparks, Maryland) with 6 ug/mL cefoxitin (Cfox) to commonly used Mannitol Salt Agar with 4 ug/mL Oxacillin (MSox). A total of 2561 MRSA surveillance specimens from 1313 patients (775 nasal swabs, 733 rectal, 266 wound, 682 nasal/axilla/groin/perineum, 105 other) were planted to Cfox and MSox. The order of inoculation was alternated every 200 swabs. Cfox were incubated in the dark at 35°C, and both media were read blinded at 24 and 48 hours. Mauve colonies from the Cfox and yellow from the MSox were tested for capsular antigens 5 and 8 (Pastorex Staph Plus, Sanofi Diagnostics Pasteur, Redmond, Washington), tube coagulase and PBP2a (Denka Seiken Co., Ltd., Tokyo) to identify MRSA directly from the medium when possible. Positive PBP2a reactions were confirmed using the NCCLS 6 ug/mL Oxacillin Screen Agar. RESULTS: From the 2561 swabs, 152 (5.9%) MRSA grew from at least one medium; 133 (87.5%) grew on Cfox and 104 (68.4%) grew on MSox, while 85 (55.9%) grew on both media. Of the total 152 MRSA, 19 (12.5%) failed to grow on Cfox and 48 (31.6%) failed on MSox. The sensitivities for Cfox and MSox were 88.9% and 76%, respectively. Cfox grew 17% of its 133 MRSA at 24 hours, and 83% at 48 hours, while MSox grew 25% of its 104 MRSA at 24 hours and 75% at 48 hours. From the 2407 MRSA-negative specimens, non-<i>S. aureus</i> mauve or yellow colonies (coagulase negatives, enterococci, or Gram-negative bacilli) required work-up on 46 from Cfox and 415 from MSox, resulting in specificities of 98.1% for Cfox and 85.2% for MSox, respectively. CONCLUSION: Compared to MSox, the Cfox plates were both more sensitive and specific (p < 0.0001). This resulted in fewer unnecessary work-ups on Cfox for a saving of ∼$2 per false positive. The impact of the breakthrough rate of 16% associated with MSox in this study represents an extremely significant potential for cost avoidance to laboratories processing large numbers of surveillance screens. However, the impact on overall cost to the laboratory depends on the cost differential between MSox and Cfox.

Evaluation of oxoid mannitol salt agar with cefoxitin for the detection of methicillin-resistant Staphylococcus aureus (MRSA) from surveillance specimens

OBJECTIVES: Challenged by an increasing demand for MRSA surveillance and declining resources, clinical laboratories require sensitive and specific screening media. METHODS: This study compares Mannitol Salt Agar base™ (Difco Laboratories, Detroit, Michigan) containing either 4 ug/mL oxacillin (MSO) or 10 ug/mL cefoxitin (MSF) to Aniline Blue Salt agar with 10 ug/mL cefoxitin (ABF) (all manufactured by Oxoid Inc., Nepean, Ontario). A total of 2561 MRSA screens from 1313 patients (775 nasals, 733 rectal, 266 wound, 682 nasal/axilla/groin/perineum, 105 others) were planted onto MSO, MSF, and ABF. MSO was always planted first, while the order of inoculation for MSF and ABF was varied every 200 specimens. Media were incubated at 35°C and read blinded at 18-24 hours and 48 hours. Yellow colonies from MSO and MSF, and blue colonies from ABF were tested for capsular antigens 5 and 8 (Pastorex Staph Plus, Sanofi Diagnostics Pasteur, Redmond, Washington), tube coagulase (Remel Inc., Lenexa, Kansas), and PBP2a (Denka Seiken Co., Ltd., Tokyo) to identify MRSA. PBP2a reactions were confirmed using the NCCLS 6 ug/mL oxacillin agar. Pulsed-field gel electrophoresis (PFGE) was performed on all discrepant MRSA isolates. RESULTS: Overall, 158 MRSA grew in the 48 hour study period: 136 from MSF, 104 from MSO, 102 from ABF, while 75 grew on all three media. The sensitivity for each media was: MSF (87.8%), MSO (74.5%), and ABF (55.1%). From MSF, 41/136 (30.1%) MRSA grew CONCLUSION: MSF plates were less costly and significantly more sensitive than both MSO and ABF (p

Expression of plasma coagulase among pathogenic Candida species.

Candida coagulase production was assessed by the classical tube test. All Candida krusei strains were coagulase negative, but most C. albicans and C. tropicalis strains can produce coagulase. Some strains agglutinated the Pastorex Staph-Plus reagent, probably because of antigen similarities to coagulase produced by Staphylococcus aureus that may cause mistakes in clinical laboratories.

Cost-effectiveness of different MRSA screening methods

We describe a model to examine the cost-effectiveness of various laboratory-screening approaches to detect methicillin-resistant Staphylococcus aureus (MRSA). A critical literature review was used to derive relevant data on the sensitivity (X), specificity (S) and time to result (T) of different tests. Additional cost information was provided by a hospital. Tests were considered in four interactive groups based on a hierarchy of procedures used in laboratories. X, S and Ts of screening tests were then used in formulae to calculate effectiveness for the various tests. The model was developed to explore the effects on MRSA infection acquisition of differing X, S and T for the different tests in detecting MRSA colonized patients admitted to a high-risk unit such as an intensive care unit. It was concluded that taking a sample from the nose alone and inoculating directly on to Ciprofloxacin Baird-Parker agar without broth incubation and confirmation by a Pastorex Staph-Plus test without any methicillin resistance confirmation test was the most cost-effective approach. The complexity of designing this apparently simple scenario is apparent, and we describe many other factors that would need to be considered to refine this model further. However, this and other models should aid the debate and development of more cost-effective screening strategies given the lack of standardization or agreement concerning so many of the variables within the UK and elsewhere.

Staphylococcal Colonization in Atopic Dermatitis Treatment with Topical Tacrolimus (Fk506)

Staphylococcal Colonization in Atopic Dermatitis Treatment with Topical Tacrolimus (Fk506)

Reliability of Rapid Kits forStaphylococcus aureusIdentification

We assessed the performance of four rapid Staphylococcus aureus detection kits: Slidex Staph-Kit (bioMerieux, France), Avistaph (Omega Diagnostics, England), Staphylase (Oxoid, Unipath, England), Pastorex Staph-Plus (Sanofi Diagnostics Pasteur, France). We tested 244 coagulase-positive ( S aureus ) and 127 coagulase-negative staphylococci (CNS) clinical isolates. The sensitivity and negative predictive values were 96.6% and 99%, respectively, for all kits. The specificity and positive predictive values, however, were different: 91% and 96%, respectively, for Slidex Staph-Kit; 67% and 86%, respectively, for Avistaph; and 94% and 97%, respectively, for both Staphylase and Pastorex Staph-Plus. The percentage of uninterpretable results among kits for testing S aureus and CNS isolates, respectively, were: 0 and 4.7% for Slidex Staph-Kit; 4% and 7% for Avistaph; 4% and 7.8% for Staphylase; and 0 and 0.8% for Pastorex Staph-Plus. The reliability and ranking of these kits were 98% for Staph-Plus, 98% for Staphylase, 97% for Slidex Staph-Kit, and 89% for Avistaph. Although these kits are rapid and useful, they are subject to variations and limitations in their performance, and caution is warranted when interpreting their results.

Evaluation of new agglutination test for identification of oxacillin-susceptible and oxacillin-resistant Staphylococcus aureus

A new agglutination test (Monostaph +; Bionor, Skien, Norway) has been developed. This new agglutination test has been compared with two other agglutination tests for the identification of 128 isolates of Staphylococcus aureus and 82 coagulase-negative staphylococci. The sensitivities of both Monostaph + and Pastorex Staph-Plus were excellent (98.7 and 97.4%, respectively) in detection of oxacillin-resistant Staphylococcus aureus. The specificity was 96.4% (two Staphylococcus epidermidis isolates and one Staphylococcus hominis isolate were false positive).