San Diego, Calif.) directed at the Mycobacterium tuberculosis complex and Mycobacterium avium-M. intracellulare complex were used to identify acid-fast bacilli directly from specimens grown in BACTEC 12B bottles (Becton Dickinson and Co., Towson, Md.). Clinical specimens were inoculated directly or after decontamination into a BACTEC 12B bottle, Middlebrook 7H11 agar, and Lowenstein-Jensen medium. Conventional media were incubated at 37°C in 5% C02 and examined weekly for 6 weeks. Identification of isolates grown on conventional media by standard biochemicals, morphology, and growth characteristics served as the reference method for identification. BACTEC bottles were incubated at 37°C, and a growth index was taken twice a week. When a growth index of .-100 was reached, 1 ml of BACTEC 12B medium was put into each of three microfuge tubes which were centrifuged for 15 min at 15,000 X g. Pellets were used in hybridization reactions with an M. tuberculosis complex probe, an M. avium probe, and an M. intracellulare probe. The results of the hybridizations of the three probes with the same sample were compared, and the highest percent hybridization was divided by the average of the two lower hybridization values. If this value, the derived patient ratio (DPR), was-3, then the specimen was considered positive for the organism giving the highest percent hybridization. Of the 1,988 specimens cultured, the results of conventional tests for the 190 conventional culture-positive specimens were 64 M. tuberculosis, 61 M. avium, 14 M. intracellulare, 30 other Mycobacterium spp., and 25 non-acid-fast bacilli. There were four cultures that each contained two different Mycobacterium spp. Directly probing the BACTEC 12B sediment, at a DPR of-3 the M. tuberculosis probe identified 83% (53 of 64) of M. tuberculosis isolates, the M. avium probe identified 92% (56 of 61) M. avium isolates, and the M. intracellulare probe identified 86% (12 of 14) of M. intracellulare isolates. There were no false-positive results at this DPR level. The false-negative results from probing the sediment from the BACTEC 12B bottle could not solely be attributed to the number of organisms present, the growth index, or antimicrobial therapy.

Using hemagglutination inhibition (HAI) as a reference method, 292 (40 nonimmune, 252 immune) human serum samples were tested by indirect fluorescent antibody (IFA) and enzyme-linked immunosorbent assay (ELISA) for immune status and quantitation of rubella antibodies. The overall agreement with HAI for immune status was 99.7% (291/292) with IFA and 98.6% (288/292) with ELISA. Two specimens (0.7%, 2/292), negative by HAI, were equivocal by ELISA. Initially a 6.5% (19/292) overall disagreement was obtained for immune status evaluation between HAI and IFA, which was reduced to 0.3% (1/292) upon repeat testing. All of these samples were near the immune/nonimmune cutoff point (95 samples), reflecting an initial disagreement of 20% (19/95) in this category (HAI titers less than 1:20). Likewise, an initial overall disagreement of 4.5% (13/292) was obtained between HAI and ELISA which was reduced to 0.7% (2/292) upon repeated testing. Eleven of the 13 samples were near the immune/nonimmune cutoff point, reflecting an initial disagreement of 11.6% (11/95) with sera having an HAI antibody titer of less than 1:20. Quantitation of rubella antibodies by IFA showed an overall correlation with HAI of 86.6% within less than twofold titer and 99.3% within less than fourfold titers. In testing the ability of ELISA to quantitate antibody, a correlation coefficient (r) of 0.996 was obtained by plotting the measured average optical density (405 nm) of ELISA against the corresponding log of HAI titer. Both IFA and ELISA showed good correlation with HAI for immune status evaluation and for quantitation of rubella antibodies. Technically the HAI was the most cumbersome to perform, whereas IFA was the least technically demanding. Originally, 308 samples were tested; 16 samples (5.2%) could not be evaluated by IFA because of a high level of nonspecific fluorescence. The strict requirement of controlling the temperature range (23 to 24 degrees C) during substrate hydrolysis proved to be a problem with the ELISA test in our laboratory.

Antibody titers to Coccidioides immitis, using coccidioidin antigen, were determined by three methods: the standardized Laboratory Branch complement fixation method (LBCF), a modified version of the Viral and Rickettsial Disease Laboratory complement fixation test (VRDL-CF), and a quantitative immunodiffusion test (QID). Of the 133 samples evaluated, 72 were negative by each method and 57 (42 serum samples, 15 cerebrospinal fluid samples) were positive by all three methods. Four additional specimens (1 serum sample, 3 cerebrospinal fluid samples) were positive by QID alone. All positive patients were diagnosed clinically as having pulmonary or extrapulmonary coccidioidomycosis or both. When titers from two methods were compared, the agreement within ±1 dilution was VRDL-CF/QID, 88.5%; VRDL-CF/LBCF, 85.2%; and LBCF/QID, 82.0%. The agreement of these methods within ±2 dilutions was VRDL-CF/QID, 98.4%; VRDL-CF/LBCF, 96.7%; and LBCF/QID, 93.4%. The VRDL-CF and QID methods are simpler to perform; however, they are yet unrecognized as suitable alternatives to the more cumbersome LBCF. Our data show that they should be considered as options for C. immitis serology.

The detection and identification of intestinal helminths were studied retrospectively by comparing the Formalin concentration technique with the trichrome-stained smear technique. A total of 3,997 stool samples from 1,570 patients were examined by both methods. Of the 3,997 samples, 31% (1,239 of 3,997) contained helminths or protozoans or both. A total of 11% (434 of 3,997) of the samples representing 14% (221 of 1,570) of the patients were positive for one or more helminth species. A total of 570 separate identifications of helminth ova/larvae were made. Among the helminth ova/larvae identified. 14.6% (83 of 570) were detected only in the trichrome-stained smear, representing 6.3% (14 of 221) of the patients. From these data, it can be concluded that unless a diligent search of the stained smear for helminths is made, a significant number of helminth infections may be missed. It is, therefore, recommended that stained stool smears be used to aid the detection of helminth ova/larvae in conjunction with Formalin concentration. The appearance of the most common helminth ova/larvae in trichrome-stained smears is described, along with specific characteristics that may be used for their identification. In addition, the advantages and disadvantages of the trichrome staining technique for helminth identification are discussed.

Sera from 290 hospital patients were tested to compare the sensitivity, specificity, and reproducibility of the hemagglutination treponemal test for syphilis (HATTS) with the fluorescent treponemal antibody absorption test (FTA-ABS). Complete agreement was obtained between the methods when 142 syphilitic sera from patients with various stages of syphilis were tested. By using clinical histories, the specificity with 148 nonsyphilitic sera was determined to be 100% for the HATTS and 96.6% (143 of 148) for the FTA-ABS. Satisfactory reproducibility was obtained with both methods. Compared with the FTA-ABS, the HATTS was more specific, easier, and more economical to perform. We therefore recommend the HATTS as a suitable alternative to the FTA-ABS.