Epidemiological investigation of Salmonella tilene by pulsed-field gel electrophoresis and polymerase chain reaction.

Pulsed-field gel electrophoresis (PFGE) and DNA fingerprinting by the polymerase chain reaction (PCR) were performed on 11 isolates of Salmonella tilene. Five strains were from a cluster of human patients, six from sugar gliders and pygmy hedgehogs kept as family pets or from local pet retailers, and one isolate from the first North American case of S tilene described in Washington State in 1994. The PFGE restriction patterns showed all isolates to be similar. However, PCR using primers to the 16S and 23S rRNA genes of Escherichia coli demonstrated that the Washington State isolate differed from the rest of the other isolates, which were all similar based upon their DNA fingerprint. This study indicates that reliance on one technique alone may be insufficient to show nuances between strains that are, in many respects, closely related.

Detection of Chlamydia trachomatis antigen by enzyme immunoassay: importance of confirmatory testing.

AIM--To determine when a fluorescence assay for Chlamydia trachomatis elementary bodies in the specimen buffer is of most value as a verification test for genital specimens reactive on screening enzyme immunoassay (EIA). METHOD--Genital swabs from high and medium prevalence populations were tested using EIA. Samples with absorbance values greater than the positive threshold and those within the range of 30% below this value were verified by the MicroTrak direct fluorescence assay (DFA) test. Quotients derived from the threshold value and specimen absorbances were used to establish confidence limits for the EIA. RESULTS--Of 13,283 swabs tested, 474 from the high risk group and 236 from the medium risk group were reactive on EIA and confirmed by DFA. Thirty six (5.9%) patients with confirmed reactive samples would have been missed if the kit criteria alone were followed. When confidence limits were applied to the calculated quotients, only those samples with an EIA quotient of > or = 4.0 were universally confirmed by the DFA. CONCLUSION--A scheme of testing which uses the DFA to verify EIA reactive specimens over a specified range was found to improve the sensitivity and specificity of the EIA screening test.

Screening for antibody to Chlamydia pneumoniae by the complement fixation test.

We performed a retrospective survey for antibody to Chlamydia pneumoniae, by the microimmunofluorescence (MIF) test, on 120 sera that were previously determined to be positive for antibody to ornithosis antigen by the complement fixation (CF) test. The panel of sera comprised 40 paired acute and convalescent sera, and 40 single samples, from 80 patients. Of these patients, 60% were considered to be serologically positive for C. pneumoniae, based on the antibody titers of IgG, IgM, or both. There was no association between the CF titer to ornithosis antigen and the respective IgG or IgM MIF antibody titers. We propose that, in those laboratories routinely using the CF test, sera found to be positive for ornithosis antigen should be further tested by the MIF in order to clarify, both from an epidemiologic and clinical perspective, whether these patients are also serologically positive for C. pneumoniae.

Antibody to hepatitis C virus in selected groups of a Canadian urban population.

In an anonymous survey, 433 sera from Canadian individuals of selected categories were tested for the presence of antibody to hepatitis C virus (HCV) using a recombinant antigen-based immunoassay. About 50% of intravenous drug abusers (IVDA), 10% of transfusion recipients and an overall average of 7.9% of male homosexuals were reactive for antibody to HCV. Individuals with jaundice and negative hepatitis B virus (HBV) serology were not reactive for antibody to HCV compared with 26.7% of those with positive HBV serology. Similarly 58% of male Federal prisoners with positive HBV serology were also HCV-antibody reactive compared with 15% of those with negative HBV serology. A prevalence of 1.2% was recorded for individuals not in any of the above groups. Of 433 sera, 92 were reactive and the discrimination in absorbance values between reactive and not reactive samples was good except for 13 sera, eight of which gave values considerably higher than the average negative value and five which were just above the positive threshold.

Variability in growth of Neisseria polysaccharea on colistin-containing selective media for Neisseria spp.

In a prospective survey of 773 healthy schoolchildren in southern Alberta, Canada, Neisseria polysaccharea was isolated from the pharynxes of only 4 (0.5%) subjects, whereas Neisseria lactamica and Neisseria meningitidis were isolated from 110 (14%) and 15 (2%) children, respectively. These strains of N. polysaccharea, together with three other sporadic isolates from Alberta, Canada, were compared with the type strain from France and strains from Spain and Germany. All strains were phenotypically identical, except that the Canadian and German strains, for which the colistin MICs were 1 mg/liter, failed to grow on Thayer-Martin medium (TMM), whereas the type strain and the Spanish strains, for which the colistin MICs were greater than 7.5 mg/liter, were not inhibited. Multilocus enzyme electrophoresis indicated that six distinct electrophoretic types were present among the seven Canadian strains. Our results show that growth on gonococcal selective media which contain colistin is a variable feature of this taxon.

Predicting human immunodeficiency virus type 1-positive sera by using two enzyme immunoassay kits in a parallel testing format.

Two algorithms for screening sera for antibody to human immunodeficiency virus type 1 were compared for their efficiency in identifying a true-positive sample in a population with heterogeneous risk factors, using the criteria of specificity and positive predictive value (PPV). In the first algorithm, all sera were screened by using a single enzyme immunoassay (EIA) kit, and a specificity of 98.6% and a PPV of 69.3% was calculated for true-positive sera. The second algorithm employed two different EIA kits in parallel to screen each sample. In the first instance, a specificity and a PPV of 100% was calculated if a positive sample was defined as reactive by both EIA kits; in the second, a specificity of 99.97% and a PPV of 99.4% was obtained if this criterion was extended to include a combination of one reactive and one equivocal result obtained with the two EIA kits.

Detection of verotoxin in stool specimens.

A total of 132 fecal specimens containing verotoxin (VT) were subjected to counter-current immunoelectrophoresis (CIE). Of these, 113 (85.6%) were found to be positive by CIE. Another 71 stool specimens containing E. coli serogroup O157 but with flagellar antigens other than H7 were tested for verotoxin by CIE. These stool specimens were negative for VT on Vero cell monolayers. Of these 71 stool specimens, 6 (8.5%) gave positive tests for verotoxin by CIE. Forty stool specimen filtrates which were negative for VT (negative controls) were also subjected to CIE. One of these stool specimen filtrates gave a line of precipitation by CIE. The specificity of the CIE test was 93.7%, and the sensitivity was 85.6%. False-positive results may have been due to an antibody component against the somatic antigen (O157) in the antitoxin used; this is a limitation of the CIE test. In a related evaluation, 302 stool specimen filtrates containing VT were retested with Vero cell suspension cultures in microdilution plates. Of these, 281 stool specimen filtrates showed cytotoxic effects within 24 h, while the remaining 21 filtrates showed the effects within 48 h. The use of Vero cell suspension culture is as reliable as the use of Vero cell monolayers and provides detection of verotoxin 24 to 48 h sooner.

Pharyngitis 1987: a survey of physicians' attitudes and practices in southern Alberta.

A questionnaire survey concerning Streptococcal pharyngitis was completed by 85 southern Alberta family physicians. The data revealed a significant trend to overtreat pharyngitis with antibiotics because of the unreliability of clinical diagnosis and the lack of diagnostic manoeuvres with suitable ability to influence management. These data and a further questionnaire survey directed to the appreciation of other bacterial pharyngitides, demonstrate the importance of laboratory reports in biasing treatment, and further suggest that significant overtreatment of non-Streptococcal pharyngitis also occurs.

Inhibition of Escherichia coli serotype O157:H7 by bromthymol blue.

Bromthymol blue, at a concentration of 0.1% in tryptose-glucose broth, inhibited growth of 98.4% of Escherichia coli serotype O157:H7 isolates but only 0.8% of E. coli non-O157:H7 isolates after an overnight incubation at 44.5 degrees C, but not 35 degrees C. The inhibition was dependent on temperature, density of inoculum, bromthymol blue concentration, time of incubation, and composition of the medium. Compared with serologic typing, the inhibition had sensitivity, specificity, predictive values of the positive and negative tests, and overall agreement between the two tests of 98.4, 99.2, 98.4, 99.2, and 98.9%, respectively. The inhibition could be useful as a presumptive test to identify E. coli isolates of serotype O157:H7, especially in laboratories that do not have serotyping capabilities.

Serologic confirmation of Neisseria gonorrhoeae by monoclonal antibody-based coagglutination procedures.

Two commercially available monoclonal antibody coagglutination tests, Phadebact Monoclonal GC OMNI Test (PMGOT; Pharmacia Diagnostics AB, Uppsala, Sweden) and GonoGen (GG; New Horizons Diagnostics, Columbia, Md.), for the confirmation of Neisseria gonorrhoeae were evaluated. The sensitivities of PMGOT and GG were 99.2 and 98.7% and the specificities were 91.5 and 100.0%, respectively. False-positive reactions were observed with Neisseria lactamica and Neisseria meningitidis in PMGOT. A modification of the procedure recommended by the manufacturer for PMGOT was done by substituting Todd-Hewitt broth for 0.9% saline to prepare the suspension of the test organism. This eliminated technical difficulties with the test and resulted in a sensitivity and a specificity of 99.3 and 100.0%, respectively. Advantages offered by the modified PMGOT over GG were the better predictive value of the negative test, the lower cost, the ease of reading of the test, and the lack of noninterpretable results.

Delta hepatitis virus infection in China.

To assess the prevalence, epidemiological features and prognostic implications of hepatitis D (Delta) in Sichuan Province, The People's Republic of China, 649 sera (515 from HBsAg positive patients and 134 from HBsAg negative subjects) were tested by radioimmunoassay (RIA) for antibody to the hepatitis D virus (anti-HD). Forty-seven sera (7.2%) showed some degree of reactivity. Serial dilutions of these sera indicated that prozoning was not responsible for the equivocal results. Thirty-four of the 47 sera were submitted under code to a second laboratory for independent analysis. According to those results anti-HD antibodies were detected in four of these sera. The overall prevalence of anti-HD in the HBsAg positive patients therefore was 0.8% (4/515). On the basis of clinical, biochemical and histological data 427 HBsAg positive sera were further divided into acute Type B hepatitis, chronic Type B hepatitis, healthy carrier state and hepatocellular carcinoma (HCC) subgroups. Two of 65 (3.1%) anti-HD positive sera belonged to the acute Type B hepatitis group; one of 104 (0.9%), the chronic Type B hepatitis group and one of 246 (0.4%), the healthy carrier group. No antibody was detected in sera from 12 HBsAg positive HCC patients. All HBsAg negative patients were negative for anti-HD antibody. The results of this study indicate that despite a high prevalence of hepatitis B virus infection, positive serology for delta virus is uncommon in Sichuan Province, The People's Republic of China.

Evaluation of the GO Slide (Roche) growth transport system for isolation of Neisseria gonorrhoeae from clinical specimens.

A new growth transport system for the isolation of Neisseria gonorrhoeae from clinical specimens was evaluated. The system, GO Slide (Roche) of F. Hoffmann-La Roche & Co., Basel, Switzerland, showed 88% sensitivity for male urethral specimens and 59% sensitivity for endocervical specimens compared with transport in the Amies transport medium combined with culturing on modified Thayer-Martin medium. The media used appear not to support growth of certain strains of N. gonorrhoeae. recovery and growth-supporting capabilities of this system need to be improved before the system can be used routinely.

The prevalence of antibody to delta virus in western Canada.

To assess the prevalence and pathological role of hepatitis D virus (HDV) infection in western Canada, we tested a total of 310 sera from the province of Alberta, Yukon and Northwest Territories for antibody to HD (anti-HDV) by commercial solid phase radioimmunoassay. Two hundred and forty-five sera were hepatitis B surface antigen (HBsAg) positive. These were classified on the basis of clinical and biochemical data as either acute hepatitis, chronic hepatitis or in the healthy carrier phase of infection Sixty-five HBsAg negative sera from patients with other forms of chronic liver diseases served as controls. Anti-HDV was detected in only four of the HBsAg positive sera (1.6%) and in none of the controls. The prevalence of anti-HDV was significantly higher in patients with chronic hepatitis, three of twenty-two (13.6%) than in patients with acute hepatitis (0%) (p less than 0.05) or healthy carriers (0%) (p less than 0.005). Two of the four anti-HDV positive sera were obtained from patients with a history of parenteral drug abuse. These results indicate that HDV infection is uncommon in western Canada but, when it does occur, is more likely to be associated with chronic inflammatory liver disease. Parenteral drug abuse appears to be the major risk factor for HDV infection in western Canada at this time.

Interaction of L. pneumophilia and a free living amoeba (Acanthamoeba palestinensis).

Co-cultivation of Legionella pneumophila serogroup I and Acanthamoeba palestinensis in Neff's medium at 35 degrees C resulted in the intracellular multiplication of the bacteria as demonstrated by electron microscopy and immunofluorescence. In the closed experimental system used, the number of legionellae rose from 10(7) colony forming units (c.f.u.)/ml initially to a maximum of 10(10) c.f.u./ml on day 5. Legionellae were seen in expelled phagosomes, in some amoebae filling the cytoplasm and in others in which the process of encystment appeared to have commenced. At 20 degrees C the acanthamoebae phagocytosed and digested the legionellae. The bacteria disappeared from the co-cultivation flask by day 2 but reappeared in low numbers (10(2) c.f.u./ml) by day 6 suggesting that even at this temperature some intra-amoebal multiplication occurred.