Multicenter Evaluation of the Simplexa VZV Direct Assay for Detection of Varicella-Zoster Virus in Cerebrospinal Fluid and Lesion-Swab Specimens.

Varicella-zoster virus (VZV) is the etiologic agent of varicella (chickenpox) and herpes zoster (shingles) infections commonly involving skin, mucous membranes, and less frequently the central nervous system. Traditional methods for the laboratory diagnosis of these infections are time-consuming, labor-intensive, and often insensitive. As such, these tests are being replaced by more sensitive and rapid molecular methods. This study evaluated the performance of two different molecular assays, the Simplexa VZV Direct and Simplexa VZV Swab Direct, to detect VZV DNA in cerebrospinal fluid (CSF) and lesion-swab specimens, respectively. The Simplexa VZV Direct and Simplexa VZV Swab Direct assays were compared against individual composite reference methods that varied depending on the sample cohort examined. A total of 883 CSF and 452 cutaneous and mucocutaneous prospective, retrospective, and contrived specimens were evaluated in this multicenter study. The results of this study showed that the Simplexa assays demonstrated near perfect agreement (k = 0.98) compared to the composite reference methods for the detection of VZV in CSF and lesion swab specimens. A further comparison between the standard of care molecular assays employed at the site of specimen collection and the Simplexa assays demonstrated excellent agreement (k = 1.0). The Simplexa assays offer rapid and reliable alternatives for the detection of VZV in certain clinical specimens without the need for nucleic acid extraction.

Comparative evaluation of enzyme immunoassay and culture for the laboratory diagnosis of gonorrhea.

A commercially available, solid-phase enzyme immunoassay (EIA), called Gonozyme, was compared to Martin-Lewis medium in Jembec plates for the laboratory diagnosis of gonorrhea. A total of 577 clinical specimens (419 urethral and 158 endocervical) were collected from a high-risk, walk-in patient population attending a sexually transmitted disease clinic. The results showed that EIA was comparable to a conventional cultural procedure for identifying infected and noninfected males. In addition, the system may be used reliably for performing test of cure on urethral samples obtained from this male population. Gonozyme was also comparable to culture in identifying females who had gonococcal infection. However, because of the high incidence of false positive test results, most likely attributable to antigen persistence in endocervical secretions, EIA is not recommended for performing test of cure in the female. Overall, the Gonozyme system is an easily performed, rapid, and reliable system that provides for a noncultural alternative for the laboratory diagnosis of gonorrhea.

Use of modified New York City medium for growth of Mycoplasma pneumoniae. A preliminary report.

The recovery of Mycoplasma pneumoniae from clinical specimens is a time-consuming process that requires highly specialized media. Modified New York City (NYC) medium was evaluated for its ability to support the growth of M. pneumoniae, as compared with conventional culture technics for mycoplasma. In this study, 95 stock strains of M. pneumoniae were inoculated onto modified NYC medium, PPLO methylene blue agar, biphasic medium with methylene blue, and PPLO glucose broth with phenol red to compare each medium's growth-supporting properties. The results demonstrated that the modified NYC formulation was a suitable medium for supporting this growth of stock strains of M. pneumoniae and that growth can be detected a minimum of five to six days sooner than by conventional methods.

Biotypes of Haemophilus influenzae: relationship to clinical source of isolation, serotype, and antibiotic susceptibility.

A total of 94 clinical isolates of Haemophilus influenzae were studied to analyze the relationship of biotype to site of isolation, serotype, and pattern of antimicrobial susceptibility. Systemic infections were caused most commonly by biotype I, and the majority of these isolates possessed type b capsular polysaccharide. Other noncapsulated biotypes of H. influenzae, particularly biotype V, also were associated with invasive disease. Antimicrobial susceptibility testing was performed on all isolates by an agar dilution method against ampicillin, chloramphenicol, cefoxitin, rifampin, and rosoxacin, and all isolates were screened for beta-lactamase activity. Except for 15 isolates that produced beta-lactamase, no other substantial differences in antimicrobial susceptibilities among biotypes of H. influenzae were detected. Encapsulated strains of biotype I had the highest frequency of ampicillin resistance.

The impact of same-day tests versus traditional overnight testing.

Within the last decade, major technologic advances have been made in clinical microbiology that have resulted in the availability of a wide variety of different methods for the rapid reporting of test results. Included among these technologies are rapid methods for producing antimicrobial susceptibility reports that many regard as the most important information generated by the microbiology laboratory. Ideally, the early availability of this important information should favorably affect patient care by enabling the more judicious use of alternative drug therapies that are equally efficacious yet less toxic and less costly to the patient. Clinicians appear to have been reluctant to modify initial empiric therapies, however, despite the availability of the rapid antimicrobial susceptibility report. This article addresses some of the issues responsible for this long-standing problem and discusses and explores various strategies that can be implemented for improving the use and for controlling the cost of antimicrobial agents within the hospital.

Use of the Gen-Probe PACE system for the detection of Neisseria gonorrhoeae in urogenital samples.

The Gen-Probe PACE (Probe Assay-Chemiluminescent Enhanced) system for Neisseria gonorrhoeae was compared to Martin-Lewis medium in JEMBEC plates for the direct detection of N. gonorrhoeae in urogenital samples. This 2-hr, nonisotopic chemiluminescent test is based on the use of an acridinium ester-labeled DNA probe that binds with gonococcal target rRNA in a clinical sample. Following the separation of the hybridized probe from the unhybridized probe through the use of magnetic microparticles, the acridinium ester is hydrolyzed from the hybridized probe by the addition of an alkaline hydrogen peroxide solution, resulting in the production of light, which is measured in a luminometer. The amount of light generated is directly proportional to the amount of gonococcal target rRNA present in the sample. A total of 209 urethral and 203 endocervical specimens were collected from a high-risk, clinic population with a gonococcal disease prevalence of 24% during the study period. Statistical analyses of the overall results showed that, compared to culture, the Gen-Probe PACE System had a sensitivity, specificity, and positive and negative predictive values of 90%, 99%, 98%, and 97%, respectively. The comparative results of this study showed that the Gen-Probe PACE System for N. gonorrhoeae is a reasonable, noncultural alternative for detecting gonococci directly in urogenital specimens.

Comparative evaluation of PathoDx Strep A test and culture for the detection of group A streptococci in pharyngeal specimens.

The PathoDx Strep A kit, a 10-min acid extraction and latex agglutination test, was compared with routine culture for the direct detection of group A beta-hemolytic streptococci (BHS) in 414 pharyngeal specimens collected from children with pharyngitis. The results showed that the latex test compared favorably with culture for detecting group A BHS in pharyngeal specimens (sensitivity 96.7%, specificity 97.9%, and positive and negative predictive values of 97.2% and 97.4%, respectively). The comparable number of false-positive (five) and false-negative (six) latex tests along with review of patient histories suggest that these discrepant results were attributable to sampling error during specimen procurement rather than deficiencies in the latex kit. In addition, clear-cut, agglutination reactions were obtained in over 96% of positive latex tests regardless of the amount of group A BHS that was recovered by culture. The PathoDx Strep A test is a rapid, reliable noncultural alternative for the detection of group A BHS in pharyngeal specimens.

Characterization of the A component of Streptococcus zymogenes lysin.

By utilizing conventional techniques of pressure ultrafiltration, gel filtration chromatography, diethylaminoethyl cellulose chromatography, and preparative polyacrylamide electrophoresis, the A component of the group D lysin produced by Streptococcus zymogenes has been purified to a state of apparent homogeneity when determined by the techniques of anionic and cationic disc gel electrophoresis. The A component was found to be a protein possessing a molecular weight of 27,000, a sedimentation coefficient approximating 3.2S, and a net negative charge at physiological pH.

Purification and characterization of the L component of Streptococcus zymogenes lysin.

By utilizing conventional techniques of pressure ultrafiltration, gel filtration chromatography, diethylaminoethyl cellulose chromatography, and preparative polyacrylamide electrophoresis, the L component of the group D lysin produced by Streptococcus zymogenes strains has been purified to a state of homogeneity as determined by the techniques of disc-gel electrophoresis at pH 9.3 and 4.3 and isoelectric focusing. The L component was found to be a protein possessing a molecular weight of 11,000 with a slight net negative charge at physiological pH.

Evaluation of a prototype DNA probe test for the noncultural diagnosis of gonorrhea.

A prototype, nonisotopic, chemiluminescent DNA probe test called the Gen-Probe PACE (Probe Assay-Chemiluminescence Enhanced) system for Neisseria gonorrhoeae (Gen-Probe, San Diego, Calif.) was compared with conventional Martin-Lewis culture medium in JEMBEC plates for the laboratory diagnosis of gonorrhea. This 2-h noncultural assay is based upon the use of an acridinium ester-labeled DNA probe. The rRNA-directed DNA probe hybridizes with the target rRNA, and the hybridized probe is separated from the unhybridized probe through the use of magnetic microparticles. The esterified acridinium is hydrolyzed from the hybridized probe by the addition of an alkaline hydrogen peroxide solution, resulting in the production of visible light which is measured in a luminometer. The amount of light generated is directly proportional to the amount of gonococcal target rRNA present in the sample. A total of 407 clinical specimens (203 urethral and 204 endocervical) were collected from high-risk walk-in patients attending a sexually transmitted disease clinic. Separate patient specimens were collected for culture on Martin-Lewis medium in JEMBEC plates and for DNA probe assay. Statistical analysis of the overall comparative results showed that the DNA probe assay had a sensitivity, specificity, and positive and negative predictive values of 93, 99, 97, and 99%, respectively, in a patient population with a gonococcal disease prevalence of 21%. The results of this comparative study showed that the prototype chemiluminescent DNA probe assay is a rapid and reliable noncultural alternative for the laboratory diagnosis of gonorrhea.

Evaluation of a rapid screening test for detecting group B streptococci in pregnant women.

The QUIDEL Group B Strep Test is an enzyme immunoassay (EIA) that was compared with culture for the rapid detection of moderate to high levels of group B streptococci (GBS) colonization in pregnant women. A total of 331 pregnant women were included in the study protocol, and GBS were cultured from 19 of these patients in moderate or greater amounts (incidence of 5.7%). Compared with culture, the EIA had a sensitivity, specificity, and positive and negative predictive values of 89, 99, 89, and 99%, respectively. With a sensitivity of 89%, the 95% confidence interval for this assay is 88 to 90%. The QUIDEL EIA test can be performed in less than 10 min and appears to be a reliable method for detecting moderate or greater amounts of GBS in vaginal or cervical specimens.

A new medium for the presumptive identification of gram-positive uropathogens.

A significant percentage of urinary tract infections are caused by gram-positive microorganisms. The rapid identification of these uropathogens is important in determining the appropriate treatment of such infections. A selective medium containing colistin sulfate and nalidixic acid (Columbia CNA Agar) was modified by the addition of esculin, ferric ammonium citrate, mannitol and phenol red. The new medium (Esculin-Mannitol Agar) was extensively evaluated as a primary plating medium for urine specimens. Isolates were presumptively identified solely by colonial morphology and reaction of the medium. Presumptive identification of the isolates was confirmed by conventional tests. The accuracy of the presumptive identification indicated that Esculin-Mannitol Agar was useful in the primary plating of urine specimens when employed together with an appropriate medium for the recovery of gram-negative organisms.

Bicomponent nature of lysin from Streptococcus zymogenes.

Nonlytic mutants of Streptococcus zymogenes X-14 were isolated after exposure to nitrosoguanidine. Cross-streaking of certain of these mutants on brain Heart Infusion (BHI) blood agar plates resulted in formation of spur-shaped zones of hemolysis at the junction of the two streaks. Two types of mutants were recognized. Both of these excreted into the medium substances which are nonlytic but which together produce lytic activity. These substances behaved as an enzyme with an activator. Hence, one mutant type appeared to produce an activator and the other the catalytic molecule. Active complexes were temperature-sensitive and were inhibited by some teichoic acids as is the parental type of lysin.

Comparison of the in vitro activities of teichomycin A2 and vancomycin against staphylococci and enterococci.

The in vitro activity of teichomycin A2 was compared with that of vancomycin by standardized agar dilution testing against clinical isolates of staphylococci and enterococci. Both antibiotics had similar activities against staphylococci. However, teichomycin A2 was significantly more active against enterococci.

In vitro comparative activity of moxalactam, GR 20263, and N-formimidoyl thienamycin to other beta-lactam antibiotics and tobramycin against enterobacteriaceae and staphylococci.

The in vitro activities of moxalactam, GR 20263, and N-formimidoyl thienamycin were compared to those of cephalothin, cefazolin, cefamandole, cefoxitin, and tobramycin against 152 strains of Enterobacteriaceae and staphylococci. The results showed that moxalactam, GR 20263, and N-formimidoyl thienamycin each had significantly improved activity toward the gram-negative organisms tested compared to the other beta-lactams and tobramycin. N-formimidoyl thienamycin was particularly impressive with respect to its activity toward Staphylococcus aureus and S. epidermidis as compared to moxalactam, GR 20263 and the older beta-lactam drugs.

Meningitis caused by maltose-negative variant of Neisseria meningitidis.

A maltose-negative variant of Neisseria meningitidis, Slaterus Y, was recovered from a patient with meningitis. A report of the case is presented and the medical-legal significance of such an isolate is briefly discussed.

Comparison of modified New York City medium with Martin-Lewis Medium for recovery of Neisseria gonorrhoeae from clinical specimens.

A modified formulation of New York City medium was comparatively evaluated with Martin-Lewis medium for the recovery of Neisseria gonorrhoeae from clinical specimens. A total of 240 strains of gonococci were recovered from 1,250 specimens collected from walk-in patients attending a sexually transmitted disease clinic. N. gonorrhoeae was cultivated on both of these media from 182 clinical specimens with an additional 58 gonococcal strains isolated on either of the media. Of these discrepant gonococcal isolates, 27 strains were recovered on only modified New York City medium, whereas the remaining 31 strains were recovered on only Martin-Lewis agar. The differences in these isolation rates were not statistically significant. The overall results showed that modified New York City and Martin-Lewis media were comparable in their ability to grow gonococci from clinical material. Since modified New York City medium is capable of supporting the growth of N. gonorrhoeae, Mycoplasma pneumoniae, and urogenital mycoplasmas and inhibiting the growth of commensal microorganisms, it is possible that it may have considerable application as a multifunctional plating medium within the clinical laboratory.

Recovery and identification of anaerobes: a system suitable for the routine clinical laboratory.

A system for the isolation of anaerobes based upon the use of reducible solid media is described. Plates of reducible media prepared and stored aerobically were reduced before use by incubation in a GasPak jar for 24 h. Clinical specimens for culture were carefully selected. The value of Amies transport medium was confirmed. Selective and nonselective formulations of reducible media were used for primary isolation. Abbreviated identification schemes based in part on gas-liquid chromatography are presented. The suitability of this system for improving the recovery and identification of anaerobes in a routine clinical laboratory is documented.