Encapsulated Staphylococcus aureus strains vary in adhesiveness assessed by atomic force microscopy

Staphylococcus aureus capsular polysaccharides are believed to play a role in adhesion to surfaces and may contribute to their antimicrobial resistance, thereby increasing the rates and severity of associated infections. The purpose of this study was to compare the adhesiveness of distinct S. aureus capsular polysaccharides to determine whether adhesiveness was a general or specific feature across different S. aureus strains. Atomic force microscopy was used to confirm the presence or absence of capsular polysaccharides and to measure adhesive forces on a noncapsulated, serotype 8, and serotype 2 strain of S. aureus. Serotype 8 displayed a larger range of adhesive forces (1-19 nN) than the noncapsulated (0-4 nN) and serotype 2 (0-4 nN) strain. The majority of adhesive forces for serotype 8 were in the 10-15 nN range. Removal of capsular polysaccharides gave a marked decrease in adhesive forces measured for serotype 8 and, to a lesser extent, a decrease for serotype 2. Noncapsulated, serotype 8, and serotype 2 S. aureus had water contact angles of 23.8 (+/-8.9), 34.4 (+/-2.5), and 56.7 (+/-11.2) degrees (mean +/- standard deviation), respectively. For the first time, capsular polysaccharides from serotype 8 (clinically common) and serotype 2 (clinically rare) were demonstrated to have different physical properties, which may account for variations in studies in which clinical isolates are utilized, and the conflict in proposed roles for capsular polysaccharides on S. aureus is explained.     

Comparative evaluation of five agglutination techniques and a new miniaturized system for rapid identification of methicillin-resistant strains of Staphylococcus aureus.

The speciation of methicillin-resistant Staphylococcus aureus (MRSA) poses a significant diagnostic problem when rapid identification methods such as slide agglutination tests, are used, because of the high proportion of false-negative reactions. 150 perfectly identified MRSA strains were tested on 5 commonly used agglutination reagents ("Bacto staph latex test", "Monostaph", "Pastorex staph", "Staphaurex", and "Staphyslide test") in comparison with a new micromethod ("RAPIDEC staph") which detects a type of staphylocoagulase within 2 hours by a fluorescence test. The "RAPIDEC staph" reagent enabled identification of all the MRSA while the agglutination tests gave poorer results: "Monostaph" correctly identified 64.6% of strains, "Staphyslide", 59.3%, "Bacto staph latex test", 44.6%, "Pastorex staph", 38.6% and "Staphaurex", 28.6%. These results show that agglutination slide tests are not reliable enough for the identification of MRSA which are more and more encountered in hospital wards. The authors recommend not to use slide agglutination methods. They suggest the tube test for coagulase which is the reference technique, although it is time-consuming and not well standardized. The results of this evaluation encourage the use of the "RAPIDEC staph" reagent since it is an easy-to-use, reliable technique for the rapid identification of Staphylococcus aureus.     

Staphylococcus aureus strains which are not identified by rapid agglutination methods are of capsular serotype 5.

A total of 183 recent Staphylococcus aureus clinical isolates were tested with three commercially available rapid agglutination methods. The capsular polysaccharide type and resistance to oxacillin of these isolates were also determined. Seven isolates were not identified correctly by agglutination methods. All isolates not identified by the rapid methods were of capsular serotype 5, and of these isolates, six were resistant to oxacillin. The results suggest that these agglutination kits can be improved by the use of antibodies reactive with S. aureus capsular polysaccharide.     

Efficacies of rapid agglutination tests for identification of methicillin-resistant staphylococcal strains as Staphylococcus aureus.

Four commercially available rapid agglutination tests for the identification of Staphylococcus aureus were compared with the tube coagulase test for the identification of 300 methicillin-resistant isolates of staphylococci. Isolates tested included 207 methicillin-resistant S. aureus and 93 coagulase-negative staphylococci, collected from five medical centers. Strain variability was documented by phage typing and antimicrobial susceptibility patterns. Results of rapid identification tests ranged between 82 and 86% sensitivity, significantly poorer than the 98% sensitivity which the tube coagulase test provided.     

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.     

Comparative evaluation of identification systems for testing methicillin-resistant strains of Staphylococcus aureus.

Several commercial systems are available to distinguish between Staphylococcus aureus and the coagulase-negative species of the Micrococcaceae family. Four latex agglutination systems (Accu-Staph, SeroSTAT, Staphaurex, and Staphylatex) and two hemagglutination systems (Hemastaph and Staphyloslide) were compared for their performance in the rapid identification of 232 isolates of staphylococci, including 114 of methicillin-resistant S. aureus. Accu-Staph, Staphaurex, and Staphyloslide correctly identified 100% of the methicillin-resistant S. aureus isolates; Hemastaph and Staphylatex, 99.1%; and SeroSTAT, 94.7%. Most reactions were easy to interpret, although 15% of the SeroSTAT reactions were weak. Autoagglutination occurred only with isolates of coagulase-negative staphylococci. False-positive reactions were rare and occurred only with systems which did not detect autoagglutination. Five of these six systems appear to be adequate for the rapid identification of S. aureus, including methicillin-resistant isolates.     

Rapid and reliable identification of Staphylococcus aureus by a latex agglutination test

A latex slide agglutination test detecting clumping factor and protein A simultaneously is recommended for rapid and reliable routine identification of Staphylococcus aureus. Strains (836) of staphylococci isolated from clinical specimens were examined, all S. aureus strains identified by conventional methods were correctly differentiated by the latex test, and no false-positive results occurred with other staphylococci. The reagent is easy to prepare since plasma is the coating material.     

Comparison of five agglutination tests for identification of Staphylococcus aureus.

Various commercially produced agglutination kits are widely used for the identification of Staphylococcus aureus. These kits detect the presence of protein A and/or clumping factor on S. aureus. The literature has shown that methicillin-resistant S. aureus (MRSA) isolates which are deficient in both clumping factor and protein A may be misidentified. Two products, Slidex and Staphaurex Plus, utilize specific anti-S. aureus antibodies, potentially giving them greater sensitivity compared to products without these antibodies. We report a prospective study designed to compare the performance characteristics of Fastaph, Slidex, Staphaurex, Staphaurex Plus, Staphyloslide, and the tube coagulase test for the identification of staphylococcal isolates. All discrepant isolates were tested with the Gen-Probe AccuProbe S. aureus test and were identified to the species level with conventional reference biochemicals. A total of 1,193 isolates were tested, including 33 MRSA and 423 methicillin-sensitive S. aureus isolates. The sensitivities and specificities of the tests, respectively, were as follows: Fastaph, 99.1 and 98.9%; Slidex, 99.6 and 96.4%; Staphaurex, 98.9 and 99.9%; Staphaurex Plus, 99.6 and 93.9%; Staphyloslide, 99.1 and 98.9%; and tube coagulase, 99.3 and 100%. Sensitivity was excellent for all of the products tested. The specificities of Fastaph, Staphaurex, and Staphyloslide were excellent, while Staphaurex Plus and Slidex demonstrated less optimal results.     

Comparison of commercial slide agglutination kits with a tube coagulase test for the rapid identification of Staphylococcus aureus from blood culture.

Eighty clinical specimens of BACTEC 9240 blood culture vials, culture positive for staphylococci (38 Staphylococcus aureus and 42 coagulase negative staphylococci), were tested directly for the presence of clumping factor/protein A and free coagulase. Seven commercial slide agglutination kits were compared with a direct-tube coagulase (DTC) method. All tests were performed on blood culture pellets. Sensitivity, specificity, and negative and positive predictive values for the seven commercial kits were extremely variable, whereas a two-hour DTC test was highly predictive of S aureus. There was no significant difference between a two-, six- or 24-hour DTC test. Three (8%) S aureus isolates remained DTC negative even after 24 hours' incubation. Staphylococcal slide agglutination kits should not be used directly on blood culture broths. In contrast, a two-hour DTC test is a useful, rapid screening test for S aureus bacteraemia, provided isolates from DTC negative blood culture broths are re-tested using standard laboratory techniques.     

Failure of rapid agglutination methods to detect oxacillin-resistant Staphylococcus aureus.

Although a latex agglutination test (StaphAurex) and a hemagglutination test (Staphyloslide) correctly identified all strains of Staphylococcus aureus that were susceptible or had intermediate susceptibility to oxacillin, 17 of 73 (23%) and 18 of 73 (25%) strains of oxacillin-resistant S. aureus were not identified by StaphAurex and Staphyloslide, respectively. All strains not detected were resistant to trimethoprim-sulfamethoxazole and rifampin.     

Coagulase testing compared with commercial kits for routinely identifying Staphylococcus aureus.

Five commercial Staphylococcus aureus identification kits--Staphaurex (Wellcome), Staphylase (Oxoid), Staphyslide (bioMèrieux), Biostaph (Medlabs) and Bacto Latex (Difco)--were evaluated for the routine identification of S aureus from primary plates in the routine microbiology laboratory. Comparison was made with two methods of tube coagulase testing and five slide methods for detecting clumping factor (slide coagulase testing). Performances were assessed for two groups of organisms, staphylococcal species alone and a combined staphylococcal and non-staphylococcal species group. The effects of growth on selective media and storage of isolates at room temperature and 4 degrees C were investigated. Selective media cannot be recommended, nor can storage of isolates before testing. Ranked according to efficiency value with the combined staphylococcal and non-staphylococcal species group, the kits and coagulase methods performed as follows (the figures in parentheses are the efficiency values for the staphylococcal group alone): tube coagulase reference method 100% (100%), tube coagulase SJH method 99% (99%), Staphaurex 94% (97%), Staphylase 93% (96%), slide coagulase method No 4 93% (94%), slide coagulase method No 5 93% (93%), Bacto Latex 92% (95%), Staphyslide 92% (95%), and Biostaph 87% (91%). It is concluded that a commercial S aureus identification kit should not replace tube coagulase testing for the routine identification of the organism from primary plates and that, even the kits with the best performances, have little advantage over a good slide coagulase test method.     

Comparison of immunoassay kits for detection of staphylococcal enterotoxins produced by Staphylococcus aureus

Four commercial kits, SET-RPLA, RIDASCREEN, SET-EIA, and TECRA, were compared for the efficiency of detecting staphylococcal enterotoxins (SE). There was no non-specific reaction for detection of SE produced by 21 Staphylococcus aureus strains from 5 outbreaks of food poisoning using SET-RPLA, SET-EIA and TECRA kits. The background of the results of RIDASCREEN kit was high and non-specific reactions were present in some strains. In conclusion: (i) TECRA kit is suggested to be used for screening SE producing strains; (ii) SET-RPLA and RIDASCREEN kits are suitable for epidemiological investigation of SE types, but the lack of ability for detecting SEE, long time required for testing with SET-RPLA kit and high background when using RIDASCREEN kit must be overcome; and (iii) because of the complicated test procedures and the lack of ability for detecting SEE, the practicality of SET-EIA kit in screening and epidemiological research purposes is low.     

Twelve aberrant strains of Staphylococcus aureus subsp. aureus from clinical specimens.

A new biovar of Staphylococcus aureus subsp. aureus was isolated from human clinical specimens and described on the basis of studies of 12 isolates that were compared with 11 standard reference strains. Both DNA hybridization experiments and numerical taxonomy analysis demonstrated that these strains were strictly related to S. aureus subsp. aureus; however, they were significantly different from the latter. The atypical strains belonging to the new biovar can be distinguished from typical S. aureus subsp. aureus strains by their alpha-chymotrypsin, alpha-glucosidase, beta-N-acetylglucosaminidase, lipase (C-14), and leucine arylamidase enzymatic activities and novobiocin resistance. Thus, the combination of alpha-glucosidase and beta-N-acetyl-glucosaminidase is more useful for distinguishing these S. aureus strains from the other, typical ones.     

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).     

Evaluation of three commercial agglutination tests for the identification of Staphylococcus aureus.

Three commercially available rapid slide agglutination tests for the identification of Staphylococcus aureus were evaluated with 354 recent clinical isolates (165 strains of S. aureus). The test results of two latex agglutination products, SeroSTAT Staph (Scott Laboratories, Inc.) and Staphylatex (American Micro Scan), and one hemagglutination product, Staphyloslide (BBL Microbiology Systems), were compared with the results of the tube coagulase test, which was read at 4 h (4-h tube coagulase test) and, if negative, again after overnight incubation at room temperature (24-h tube coagulase test). Discrepancies between agglutination and tube coagulase identifications were resolved by use of the thermonuclease, mannitol fermentation, and slide coagulase tests. All sensitivities, specificities, predictive values of a positive result, and predictive values of a negative result for the three agglutination tests were at least 98.8% and comparable with the 4-h tube coagulase test. Best results were obtained with the 24-h tube coagulase test, which yielded one false-negative and no false-positive tests. Agglutination identifications may be performed on organisms taken directly from a primary plate when sufficient growth is present. Kit agglutination procedures yield rapid and reliable identifications and are easy to perform. This study also demonstrates the usefulness of the 24-h tube coagulase test.     

Evaluation of a commercial latex agglutination test for identification of Staphylococcus aureus.

A new, commercially available latex agglutination test (SeroSTAT Staph; Scott Laboratories, Inc., Fiskeville, R.I.) was compared with the tube coagulase and slide coagulase tests as means for identifying Staphylococcus aureus. Of 160 clinical isolates of S. aureus, 159 (99.4%) yielded positive results with the latex agglutination test. Negative latex agglutination test results were obtained with 266 of 267 clinical isolates of Micrococcus spp. and staphylococcal species other than S. aureus (99.6%). The latex agglutination test was found to be a rapid, technically nondemanding method for identifying S. aureus. It was as accurate as the tube coagulase test and more accurate than the slide coagulase test.