The source of coagulase-negative staphylococci in the Endophthalmitis Vitrectomy Study. A comparison of eyelid and intraocular isolates using pulsed-field gel electrophoresis.

OBJECTIVE: To determine the species distribution of coagulase-negative staphylococci (CoNS) in patients with endophthalmitis and to ascertain whether the patient's own flora was a major source of postoperative endophthalmitis following cataract extraction. METHODS: In a 4-year multicenter prospective study, 524 bacterial isolates were submitted from 225 Endophthalmitis Vitrectomy Study patients. From the 524 isolates, 250 represented CoNS cultured from the anterior chamber, the vitreous, or both of the 225 patients. Where possible, paired isolates from an individual patient's eyelid and intraocular compartment(s) were studied by pulsed-field gel electrophoresis, an established molecular strain-typing technique. RESULTS: From all sites the most frequently isolated CoNS were Staphylococcus epidermidis (81.9%) and Staphylococcus lugdunensis (5.9%). Where analysis was possible, eyelid isolates were indistinguishable from intraocular isolates in 71 (67.7%) of 105 comparisons. Non-S epidermidis CoNS caused postoperative endophthalmitis in 5 patients. Four of the 5 had postoperative endophthalmitis caused by S lugdunensis and 1 by Staphylococcus haemolyticus. CONCLUSIONS: Coagulase-negative staphylococci from the patient's periocular skin flora play a significant role in causing intraocular infections, and non-S epidermidis CoNS play a small but significant role. These results reinforce the necessity to follow stringent surgical site preparation prior to eye surgery.

Four-year prospective study of STAPH-IDENT system and conventional method for reference identification of Staphylococcus, Stomatococcus, and Micrococcus spp..

A 4-year prospective study compared the accuracy of the STAPH-IDENT system (bioMérieux Vitek, Inc., Hazelwood, Mo.) with that of the reference procedure of the Centers for Disease Control and Prevention for the identification of Staphylococcus species, Stomatococcus mucilaginosus, and Micrococcus species. The study compared the results from 1,106 cultures (500 eye cultures, 217 strains submitted for reference identification, and 389 known stock strains) representing 21 species of the family Micrococcaceae. The overall agreement of genus and species identifications was 81.1%. The percent agreement for the five most common clinical isolates was as follows: Staphylococcus epidermidis, 97.1% (517 isolates); Staphylococcus hominis, 82.5% (57 isolates); Staphylococcus aureus, 77.2% (162 isolates); Staphylococcus haemolyticus, 75.8% (61 isolates); and Staphylococcus warneri, 64.1% (39 isolates). The lowest percent agreement was with Staphylococcus cohnii (11.1%; (9 isolates). Of the 217 isolates sent to the Centers for Disease Control and Prevention for identification, 60.4% (131) were correctly identified by the STAPH-IDENT system. Of these, S. epidermidis accounted for 23.9%, S. aureus accounted for 15.6%, S. warneri accounted for 6.9%, Staphylococcus lugdunensis accounted for 6.5%, S. haemolyticus accounted for 5.5%, and S. hominis accounted for 4.1%. The STAPH-IDENT system did not perform adequately when dealing with commonly encountered organisms and is unsuitable for identifying uncommon isolates.

Numerical approach to reference identification of Staphylococcus, Stomatococcus, and Micrococcus spp.

A numerical-code system for the reference identification of Staphylococcus species, Stomatococcus mucilaginosus, and Micrococcus species was established by using a selected panel of conventional biochemicals. Results from 824 cultures (289 eye isolate cultures, 147 reference strains, and 388 known control strains) were used to generate a list of 354 identification code numbers. Each six-digit code number was based on results from 18 conventional biochemical reactions. Seven milliliters of purple agar base with 1% sterile carbohydrate solution added was poured into 60-mm-diameter agar plates. All biochemical tests were inoculated with 1 drop of a heavy broth suspension, incubated at 35 degrees C, and read daily for 3 days. All reactions were read and interpreted by the method of Kloos et al. (G. A. Hebert, C. G. Crowder, G. A. Hancock, W. R. Jarvis, and C. Thornsberry, J. Clin. Microbiol. 26:1939-1949, 1988; W. E. Kloos and D. W. Lambe, Jr., P. 222-237, in A. Balows, W. J. Hansler, Jr., K. L. Herrmann, H. D. Isenberg, and H. J. Shadomy, ed., Manual of Clinical Microbiology, 5th ed., 1991). This modified reference identification method was 96 to 98% accurate and could have value in reference and public health laboratory settings.

Prevalence of serotypes of Xanthomonas maltophilia from world-wide sources.

Since its development in 1988, a serologic typing scheme for Xanthomonas maltophilia, based on 31 O antigens, has been successfully used to serotype isolates involved in nosocomial outbreaks in the United States. To determine if this serotyping scheme would be useful in typing X. maltophilia isolates from world-wide sources, we obtained additional isolates from 10 countries; of 900 isolates tested, 795 (88.3%) were typable. In order of predominance, the three most common serotypes were 10, 3 and 19. These three serotypes were most frequently associated with respiratory and blood isolates. This serotyping system is useful as an epidemiologic screening method for universal typing of outbreaks of X. maltophilia infections.

Reevaluation of the API 20E identification system versus conventional biochemicals for identification of members of the family Enterobacteriaceae: a new look at an old product.

The API 20E bacterial identification system has been used for 19 years, often as the standard with which other identification systems are compared. Because the accuracy of this system compared with conventional biochemical tests has not been determined in many years, we evaluated the API 20E linear strip by using 291 typical and atypical strains of the family Enterobacteriaceae taken from a culture collection. At 24 h, the API 20E correctly identified by genus and species 229 of 291 (78.7%) of the strains, using Salmonella and Shigella serotyping where indicated. At 48 h, 95.2% were correctly identified by using additional biochemical tests as recommended by the manufacturer. The API 20E misidentified eight (2.7%) strains; these strains were not limited to any particular genus. When 81 of these Enterobacteriaceae strains were arranged into a weighted assortment correlating to the frequency with which they might be found in a clinical laboratory, the API 20E correctly identified 71 (87.7%) at 24 h and 78 (96.3%) at 48 h. This evaluation concluded that the accuracy of the identification of Enterobacteriaceae strains at 24 h (78.7%) may be significantly lower than that of earlier evaluations. However, there is no significant difference in the ability of the API 20E to correctly identify "challenge" type organisms (229 of 291) versus routine hospital isolates (71 of 81) (P greater than 0.05), but the system is not as accurate as the conventional biochemical method of identification.

Preliminary evaluation of Biolog, a carbon source utilization method for bacterial identification.

The Biolog Identification System (Biolog, Inc., Hayward, Calif.) is a new bacterial identification method that establishes an identification based on the exchange of electrons generated during respiration, leading to a subsequent tetrazolium-based color change. This system tests the ability of a microorganism to oxidize a panel of 95 different carbon sources. We report on a preliminary investigation of the ability of the instrument to identify, using its computer-driven enzyme immunoassay reader, a diverse group of clinically relevant members of the family Enterobacteriaceae and gram-negative non-Enterobacteriaceae. The Biolog reported identifications (correct or incorrect) for 266 of 352 organisms tested (75.6%). Of the 266 identifications reported, 87.3% were correct at the genus level and 75.6% were correct at the species level at 24 h. In the total study of 352 strains, 46.6% were correct to the species level at 4 h and 57.1% were correct to the species level at 24 h. The error rate was 10.4% after 4 h and 9.6% after 24 h. The Biolog performed well with many genera, but problems were encountered with some strains of Klebsiella, Enterobacter, and Serratia. We found the system to be versatile and easy to use.

Agreement between visual and automated UniScept API readings.

The UniScept API system was evaluated for agreement of visual versus automated readings of both its identification panels and its antimicrobial susceptibility panels. The biochemical responses of 340 oxidase-negative and oxidase-positive fermentative bacterial cultures were read both visually and automatically in the UniScept API 20E system. Automated and visual readings agreed with 99.3% of the biochemicals. Of the 45 tests that disagreed, the tests for indole and citrate were most often in disagreement. A total of 470 fermentative and nonfermentative cultures were used in the UniScept MIC system to compare visual and automated readings of susceptibility results with 17 antimicrobial agents. Agreement within +/- 1 dilution occurred with 94.1% of the enteric fermenters and with 91.7% of the other cultures. Comparison of visual and automated readings resulted in very major discrepancies in 0.95% of the readings, with the largest percentage of discrepancies associated with glucose nonfermenters (1.8%). It was felt that an automated reading is an acceptable alternative to a visual reading of the biochemicals but that 0.95% was just within the acceptable range of the 1% allowable very major discrepancies in the automated reading of susceptibilities.

Evaluation of the updated QUANTUM II system for the identification of gram-negative bacilli.

The QUANTUM II system (Abbott Diagnostics, Irving, Tex.) was evaluated with 65 species of gram-negative bacilli from various culture collections at the Centers for Disease Control. The QUANTUM II system accurately identified 92.5% of 335 isolates tested, as follows: 92.6% of 258 members of the family Enterobacteriaceae, 92.7% of 55 nonfermenters, and 91% of 22 oxidase-positive fermenters. These results were obtained by using the additional biochemical and serologic tests recommended by the manufacturers of the QUANTUM II system. The 25 misidentified cultures generally belonged to newly recognized genera, atypical strains, or slower-growing strains of more widely known genera. The system identified the most commonly encountered organisms at an accuracy of greater than or equal to 95%. The system is efficient, accurate, and rapid.

Serological classification of Xanthomonas maltophilia (Pseudomonas maltophilia) based on heat-stable O antigens.

Twenty-six serotypes of Xanthomonas maltophilia were defined by using 15 antisera described by Hugh and Ryschenkow (R. Hugh and E. Ryschenkow, J. Gen. Microbiol. 26:123-132, 1961) and 11 new antisera. The antisera were prepared by immunizing rabbits with bacterial strains heated at 100 degrees C for 2 h. Twelve antisera required adsorptions with cross-reacting heterologous immunizing strains. We tested 275 clinical and environmental strains of X. maltophilia with 26 antisera by the slide agglutination technique. A total of 259 (94.2%) strains were typeable, with 137 (49.8%) strains agglutinating in three antisera.

Evaluation of PASCO MIC-ID system for identifying gram-negative bacilli.

A production model of the semi-automated PASCO MIC-ID system (PASCO Laboratories, Wheat Ridge, Colo.) was evaluated with 122 groups or species of gram-negative bacilli, which included typical (499 cultures) and atypical (37 cultures) strains of fermenters and nonfermenters. The PASCO system identified 90.9% of 536 cultures accurately; these included 90.8% of 152 nonfermenters, 93.8% of 308 enteric fermenters, and 78.9% of 76 oxidase-positive fermenters. These results were obtained with the aid of serologic tests and a few additional biochemical tests, when recommended by the PASCO system. Of the 14 misidentified nonfermenters, 3 were Pseudomonas paucimobilis, 3 were Weeksella virosa (Centers for Disease Control group IIf), 2 were Xanthomonas (Pseudomonas) maltophilia, and 6 were randomly distributed among the other groups and species tested. The 19 enteric fermenters that were misidentified were randomly distributed among the groups and species tested. Of the 16 misidentified oxidase-positive fermenters, 4 were Pasteurella ureae, and 12 were randomly distributed among the other groups and species. The system identified the most commonly encountered organisms at a rate of 95% or better. The PASCO system is easy to inoculate and read. A slightly improved data base should remedy most of the identification problems.

Evaluation of the MicroScan antimicrobial susceptibility system with the autoSCAN-4 automated reader.

The American MicroScan (American MicroScan, Mahwah, N.J.) identification and antimicrobial susceptibility system consists in part of an automated reading system (autoSCAN-4) with data management capabilities. We evaluated the system with 404 gram-negative and 170 gram-positive facultative anaerobic and aerobic bacteria. We compared MicroScan results read automatically and visually with each other and with the results obtained by the reference method (read visually). The overall agreement within +/- 1 log2 dilution was 94.3% when the MicroScan system (read automatically) was compared with the reference method (read visually), 96.4% when MicroScan panels (read visually) were compared with reference panels, and 97.4% when the autoSCAN-4 automated reading was compared with the visual reading of the MicroScan panels. Total discrepancies (susceptibility interpretation category changes) for the MicroScan system compared with the reference method were 7%, with 6.2% considered a minor discrepancy. The autoSCAN-4 and the complete MicroScan system yielded accurate results compared with the reference method.

autoSCAN-4 system for identification of gram-negative bacilli.

A production model of the autoSCAN-4 system (American MicroScan, Inc., Mahwah, N.J.) was tested with not more than 11 strains each of 73 groups or species of gram-negative bacilli from various Centers for Disease Control culture collections. The strains included typical and atypical strains of enteric fermenters, nonenteric fermenters, and nonfermenters. The autoSCAN-4 system identified 95.3% of all 405 cultures accurately: 95.4% of 307 members of the family Enterobacteriaceae, 96.6% of 29 nonenteric fermenters, and 94.2% of 69 nonfermenters. Manual readings of the same trays provided essentially the same results, with a maximum change of only +1.6% identification accuracy of members of the Enterobacteriaceae. These data were obtained by all required additional tests, including serology and computer consultation when indicated. Only 19 of the cultures tested were misidentified. These were distributed randomly throughout the various groups and species except that Edwardsiella tarda was usually missed because of poor H2S reactions in the test medium. Of six Yersinia enterocolitica isolates, two were not identified. Only one nonenteric fermenter, a Pasteurella sp., and four nonfermenters (three Pseudomonas sp. and one Centers for Disease Control group Ve-2) were misidentified.

Evaluation of the Rapid Strep system for the identification of clinical isolates of Streptococcus species.

A total of 247 strains of streptococci isolated from humans were tested for identification in the Rapid Strep system. The identification rates and identification levels were different for each Streptococcus species. Our data indicate that the Rapid Strep system will identify nearly all the beta-hemolytic Streptococcus species if serological procedures are used in conjunction with the rapid physiological procedures. Of the group D streptococci, 98% of the enterococci and 95% of the non-enterococci were correctly identified. Of the commonly occurring viridans species, 85% were correctly identified, but only 10% of the less frequently occurring viridans species were identified. A total of 90% of the Streptococcus pneumoniae and 60% of the Aerococcus strains were correctly identified.

Evaluation of two Salmonella typhi strains with reduced virulence for use in teaching and proficiency testing.

A total of 21 cases of laboratory-acquired typhoid fever associated with teaching and proficiency tests occurred in the United States during a 33-month period, prompting a search for less virulent strains of S. typhi which would be suitable for teaching purposes. Two strains were evaluated which are reported to have reduced virulence for mice. Strain Ty21a is a genetically constructed mutant that lacks the enzyme UDP-glucose-4-epimerase. This strain has reduced virulence for humans if grown under special laboratory conditions (in the presence of 0.1% d-galactose) and has been evaluated as a candidate for use as a live, oral vaccine. Strain H901 was originally isolated in Russia in 1918. It has not been tested in humans, but its nonmotile variant, O901, has been found to be somewhat less virulent for humans; however, it can cause infection with doses of 10(7) organisms. In teaching exercises, all strains should be treated as though they are fully virulent. Ty21a and H901 were satisfactory, but not ideal, for teaching purposes. Biochemically, they could be identified by conventional tests and by commercially available diagnostic systems, although Ty21a was H(2)S negative. Serologically, both strains posed problems. Both Ty21a and H901 were Vi antigen negative, and Ty21a was rough and grew poorly. Both strains were susceptible to antibiotics, including chloramphenicol, ampicillin, and trimethoprim-sulfameth-oxazole. When Ty21a and H901 were mixed with Escherichia coli and plated, Hektoen and salmonella-shigella agars were most useful for their recovery. The appearance of Ty21a and H901 on differential plating media was typical, although Ty21a had smaller colonies. The plating efficiency on MacConkey agar for Ty21a was 0.6 compared with 1 for H901. Neither strain can be recommended unequivocally for teaching purposes; instead, the advantages and disadvantages of each must be considered. Both strains have been deposited in the American Type Culture Collection (Ty21a = ATCC 33459 = CDC 2861-79; H901 = ATCC 33458 = CDC 2862-79).

Evaluation of the Enteric-Tek system for identifying Enterobacteriaceae.

The Enteric-Tek wheel (Flow Laboratories), consisting of 14 different biochemical parameters for rapidly identifying Enterobacteriaceae, was evaluated and compared with the conventional method for completely identifying 301 enteric cultures, representing 36 species. The Enteric-Tek system correctly identified 264 (97.8%) of the 270 common or typical strains and 26 (83.9%) of the 31 unusual or atypical strains tested, demonstrating an overall identification accuracy rate of 96.3%. There were 80 (26.6%) correctly identified strains requiring additional tests. Of the 11 (3.6%) misidentifications, 5 (3 Klebsiella and 2 Salmonella strains) were correctly identified at the genus level. When 4,228 individual tests in the Enteric-Tek wheel were compared with the conventional tubed media, 96.4% of the tests agreed; urease, citrate, adonitol, and lactose agreed less than 97%. The Enteric-Tek system was found to be reliable and accurate in producing identifications at the genus and species level within 18 to 24 h.

Evaluation of the modified Micro-ID system for identification of Enterobacteriaceae.

Micro-ID is a system designed to identify the Enterobacteriaceae by utilizing reagent-impregnated disks for 15 biochemical tests. Since its initial evaluations, the system has undergone modification in formulation and in its computer data base. In a dual-center evaluation, 306 isolates of Enterobacteriaceae were tested: 145 common and typical isolates at the Mayo Clinic and 161 unusual or atypical isolates at the Center for Disease Control. Each laboratory also exchanged 50 cultures to test the system's reproducibility. Micro-ID correctly identified 142 (98%) of the common clinical isolates and 123 (76%) of the unusual or atypical organisms. However, in this latter group, three species tested were not in the system's data base. When these organisms were deleted from the analysis, 138 of 146 (95%) of the unusual or atypical isolates were correctly identified. Analysis of the 100 isolates identified in duplicate revealed 93% reproducibility of genus and species identification and 62% reproducibility of octal code numbers. Of the 31 strains with the same identification but different code numbers, 74% differed in only one biochemical test.

Interference by Neisseria gonorrhoeae growth by other bacterial species.

Growth of Neisseria gonorrhoeae from clinical specimens has been enhanced by the use of selective media that inhibit the simultaneous growth of other microorganisms. One explanation for this enhancement could be that certain other bacteria inhibit gonococcal growth. This hypothesis was examined by testing 167 bacterial isolates for in vitro gonococcal inhibition; 34.1% of the isolates failed to inhibit the gonococcus, but 12.0% produced weak inhibition and 53.9% strongly inhibited N. gonorrhoeae. The pattern of in vitro gonococcal inhibition was consistently the same for all the individual isolates within some species, but individual isolates within other bacterial species varied in their ability to inhibit the gonococcus. Consistently strong in vitro N. gonorrhoeae inhibitors were Citrobacter diversus, Enterobacter cloacae, Serratia marcescens, and Pseudomonas. The in vivo significance of gonococcal interference was demonstrated in the subcutaneous chamber model of N. gonorrhoeae infection.

Evaluation of the pathotec Rapid I-D system for identification of Enterobacteriaceae.

The PathoTec Rapid I-D System for identifying Enterobacteriaceae was evaluated with 471 cultures. In 4,910 individual test comparisons, 95.5% of the results agreed, with results of only two test strips, those for esculin hydrolysis and urease production, agreeing with conventional tests in less than 94% of the trials. The PathoTec system exhibited 94.3% accuracy in identifying these cultures in a double-blind study with conventional media and procedures as the alternate system. Two newly developed test strips, for 0-nitrophenyl-beta-D-galactopyranoside and ornithine decarboxylase, were found to be highly reliable.

Evaluation of the Bactrol Disks, a set of quality control cultures.

A set of commercially available quality control cultures was evaluated. These cultures were found to be an acceptable alternate to lyophilized or continuously subcultured cultures for a quality control program.

r-b Expanders: their use in identifying routinely and unusually reacting members of Enterobacteriaceae.

This cooperative study between a large clinical laboratory and a reference laboratory evaluated the performance of the expanded r/b system for identifying Enterobacteriaceae. The 2,200 cultures isolated in the normal hospital routine presented no problem of identification to the r/b system. About 250 "atypical" cultures which were exchanged between the collaborating laboratories were also identified accurately. The expanded r/b system was found to perform as well as most biochemical-physiological identification systems, and when used appropriately was highly satisfactory as a system for identification of Enterobacteriaceae.