Usefulness of Counterimmunoelectrophoresis for detecting toxin of Clostridium difficile.
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Biomedical subjects
Publications and source records attributed to R W Ryan.
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Thirty-two human isolates of enterococci were tested for antibiotic synergy by using penicillin and one of six aminoglycosides. Three methods were used: synergy screen, microdilution checkerboard, and time-kill curves. The synergy screen accurately predicted synergy for gentamicin-penicillin combinations, and this synergy was later confirmed by time-kill curves. The microdilution checkerboard method suffered from inherent variation, and agreement with time-kill curves ranged from 92% (twofold reduction in minimum inhibitory concentration) to 4.2% (fourfold reduction in minimum inhibitory concentration). We suggest that enterococci be screened for synergy (i.e., presence or absence of high-level resistance) by using the criterion of growth or no growth in the presence of 2,000 microgram of an aminoglycoside per ml. The microdilution checkerboard test for synergy is not recommended.
A strain of Clostridium perfringens, type A, has been isolated from the intestine of a dog which died from parvovirus infection. This Clostridium strain produces a toxin which can be detected by counterimmunoelectrophoresis, using C. difficile antitoxin, and produces cytotoxicity in WI-38 cell culture. Cytopathology can be blocked by C. difficile antitoxin. Its role in canine parvovirus infection is unknown.
Fifty fecal specimens were tested by three methods, bacterial isolation, counterimmunoelectrophoresis, and tissue culture, for Clostridium difficile and its toxin. Ten specimens (20%) were positive by all three methods. An additional eight specimens were toxin positive only by counterimmunoelectrophoresis. Although counterimmunoelectrophoresis and tissue culture are of equivalent sensitivity, the additional dilution necessary for tissue culture assay may be critical when only small concentrations of toxin are present.
Whole cells of Pseudomonas aeruginosa possess rhodanese activity. The enzyme can be released by rapidly resuspending the cells in cold Tris--HCl buffer. Approximately 95% of the rhodanese activity is released by cold shock. Release of the enzyme can be inhibited either by preincubating the cells with Mg2+ or by incorporating Mg2+ into the shocking buffer. The effect of Mg2+ can be reversed by washing the cells twice with buffer prior to cold shock. While rhodanese can be released from P. aeruginosa by cold shock, lactic dehydrogenase, a cytoplasmic enzyme, remains within the cell. Diazo-7-amino-1,3-napthalenedisulfonic acid, a compound which does not penetrate the cytoplasmic membrane, completely inactivated rhodanese and alkaline phosphatase, a periplasmic enzyme, whereas lactic dehydrogenase retained its full activity. These data suggest that rhodanese in P. aeruginosa, like alkaline phosphatase, is located distal to the cytoplasmic membrane in the periplasmic space. Electron micrographs also show that portions of the lipopolysaccharide outer membrane are shed from the cell during cold shock, while cells preincubated with Mg2+ did not release segments of their outer membrane.
Members of the genus Pseudomonas are widely implicated in human disease. Although most isolates are P. aeruginosa, there have been serious outbreaks of infection with other members of the genus. The susceptibility patterns of 5) Pseudomonas isolates are reported. It was notable that of the four antibiotics tested, uniform susceptibility was observed with minocycline.
An automated agar plate streaker was evaluated. The Autostreaker mechanizes the agar plate streaking process by providing storage for plates, labeling and streaking one or more plates for either isolation or quantitation, and stacking in one of several racks for subsequent incubation. Results showed the Autostreaker to produce agar plates with well-separated colonies and accurate colony counts. A total of 1,930 clinical specimens were processed either in parallel with manual methods or solely by the Autostreaker. Technologist acceptance of machine-streaked plates was outstanding.
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A micro broth-dilution method was developed for the antibiotic susceptibility testing of Bacteroides frgilis. Eighty strains of B. fragilis were tested against six antibiotics using the agar dilution test as the reference method. The microdilution test yielded results in 24 hours, and agreement with the reference test was satisfactory. Certain subspecies-specific patterns of antibiotic susceptibility were observed. However, larger numbers of subspecies should be evaluated before subspeciation of B. fragilis can be used as a reliable predictor of antibiotic susceptibility.
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To understand better the relationships between blood-group antigens and bacterial constituents, examples of 23 gram-negative bacteria (representing the 10 genera Citrobacter, Edwardsiella, Enterobacter, Escherichia, Klebsiella, Proteus, Pseudomonas, Salmonella, Serratia, and Shigella) were tested for the presence of Kl-like antigens by hemagglutination-inhibition (HAI) assays against both IgG and IgM anti-Kl. Saline-suspended whole organisms, cell-free culture media, and disrupted organisms were used to test for such antigens in, on, and secreted by the microorganisms examined. Disrupted organisms of an isolate of Shigella sonnei nonspecifically inhibited IgG anti-Kl as well as IgG antibodies of the specificities Kpb, Fya, S, and c. However, only Escherichia coli 0125:B15, subtype 12808, had specific K1-like activity (no activity with other IgG [(k, Kpb, Jka, Fya, S, c] and IgM [A, B, M, P1] antibodies). Disrupted organisms inhibited IgM but not IgG anti-K1 in the HAI assay. A second subtype, E. coli 0125:B15, subtype 12809, exhibited no K1-like activity. These findings support the report of K1 activity in cell-free broth cultures of E. coli 0125:B15 (subtype unspecified). Thus, although not all E. coli 0125:B15 possesses K1-like activity, the finding of such activity in at least one E. coli subtype confirms the idea that bacterial components may play a role in the production of naturally occurring antibodies directed against non-ABO red cell antigens.
BACKGROUND AND OBJECTIVES: Single dose regimens have advantages in the treatment of STD. Azithromycin has unique pharmacokinetics that may make single dose regimens feasible. Treatment with a single 1 g dose of azithromycin was compared to 100 mg doxycycline twice daily for seven days. STUDY DESIGN: This was a randomized third-party blinded study on 183 male patients, 176 of whom could be evaluated for efficacy. RESULTS: Chlamydia trachomatis was cultured from 148 patients, 79 receiving azithromycin and 69 receiving doxycycline. Six patients receiving azithromycin had positive cultures on follow-up, four were known to have had sexual intercourse with infected partners. Fifty-one patients had gonorrhea; 28 were treated with azithromycin and 23 with doxycycline. Neisseria gonorrhoeae was eradicated from all patients except one receiving azithromycin. He denied sexual exposure during follow-up. Sixty patients were infected with Ureaplasma urealyticum, 35 were treated with azithromycin and 25 with doxycycline. Five patients in each group had positive cultures on follow up. Three patients receiving azithromycin and two receiving doxycycline were known to have had sexual exposure during follow-up. CONCLUSION: A single dose of azithromycin showed similar effectiveness as a 7-day regimen of doxycycline.
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Among 270 CT scans of the thorax obtained over a 7-month period, four patients (1.5%) with calcified herniated thoracic disks were identified. Each of these patients presented with abnormal chest radiographs and had a CT examination for evaluation of suspected malignancy. None showed any signs or symptoms of thoracic spinal cord compression. The clinical significance of incidental thoracic disk protrusions is unknown. It may be that these patients are at risk for the later development of symptomatic disk disease.
Currently, the method of choice for the laboratory diagnosis of Clostridium difficile disease is the detection of cytotoxin in stool filtrates by tissue culture. Since many hospital laboratories do not have tissue culture facilities, there is a need for a rapid test which is both sensitive and specific to diagnose C. difficile disease. A commercial latex agglutination was compared with the conventional cytotoxin tissue culture assay for the detection of C. difficile or its toxin(s) in fecal specimens. Of the 574 specimens evaluated, 111 were cytotoxin positive while 97 were positive by the latex agglutination test. There were 17 specimens positive by latex agglutination but negative by tissue culture assay. The overall sensitivity and specificity of the CDT latex test was 86.1 percent and 95.3 percent respectively. This rapid latex test can serve as an excellent screening procedure for the presence of C. difficile. Those specimens positive by the latex test should be further evaluated for the presence of cytotoxin by tissue culture.
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