Serodiagnosis and Immunotherapy in Infectious Disease最新文献

Bordetella pertussis (Bp) and Bordetella parapertussis (Bpp), which are responsible for outbreaks of whooping cough in humans, are closely related, and it is difficult to discriminate between these species immunologically. We developed an immunodiagnostic method using an enzyme-linked immunosorbent assay (ELISA) or immunoblotting, on the basis of the serological differences between lipopolysaccharide (LPS) from Bp and Bpp. In ELISA, the sera from nine out of 11 patients with pertussis (about 80%) possessed high reactivities against whole cells (WC) of Bp in comparison with Bpp-WC, whereas the sera from five patients with the same disease (about 45%) reacted with Bp-LPS more than with Bpp-LPS. High reactivity against Bpp-WC and Bpp-LPS in the sera from 13 out of 16 patients with parapertussis (about 80%) was shown as compared with that against Bp-WC and Bp-LPS, respectively. Immunoblotting showed that all of the sera from pertussis patients reacted more strongly with Bp-LPS than with Bpp-LPS, except those which were unreactive. Almost all of the sera from parapertussis patients reacted with Bpp-LPS more than with Bp-LPS. These results indicated that immunoblotting, rather than ELISA, using LPS from Bp and Bpp is useful for serodiagnosis to distinguish between pertussis and parapertussis.

A study was designed to examine the feasibility of making the specific diagnoses of gonorrhoea and chlamydia infection from a single specimen of urine. Urine specimens and urethral swabs were collected from 212 male attenders at a genitourinary medicine clinic who had evidence of urethritis. Urethral swabs and urine sediments were cultured for Neisseria gonorrhoeae and urine sediments were also tested by Gonozyme and IDEIA enzyme immunoassays for N. gonorrhoeae and Chlamydia trachomatis antigens respectively Sixty-three urethral cultures were positive for N. gonorrhoeae and 83 tests were positive in the routine amplified immunoassay for chlamydia antigen. Nineteen patients had a dual infection of gonorrhoea and chlamydia. The incidence of chlamydia in non-gonococcal urethritis was 43%, this closely agreed with our previous studies. Urine deposit and urethral swab culture for N. gonorrhoeae gave a concordant negative result in 148 patients and urine culture detected 61 out of 63 patients found positive by urethral swab culture, a sensitivity of 96.8%. The immunoassay for gonococcal antigen in urine detected 60 out of 63 urethral culture positive patients. If two persistently equivocal results were taken as reactive then the sensitivity of the test was 98.4% with a specificity of 94%. Our results showed that both N. gonorrhoeae and C. trachomatis can be detected readily using appropriate enzyme immunoassays on a single urine sample from symptomatic males. This approach to sexually-transmitted disease (STD) screening may be applicable to mass populations.

The techniques of immunoblotting and enzyme-linked immunosorbent assay (ELISA) were compared with the Widal test for examining the serum antibody response of patients with clinical typhoid. The Widal test detected antibodies in only four out of 16 patients. By ELISA and immunoblotting six patients were identified. With immunoblotting, patients' sera were found to contain antibodies binding to the 0=12 antigen of lipopolysaccharide (LPS), five of these sera also contained serum antibodies to H=d flagellar antigens. Serum antibodies to Salmonella typhi LPS, as detected by immunoblotting, correlated with high levels of antibodies to LPS as detected with a Salmonella enteritidis LPS ELISA. A rapid immunoblotting procedure was developed using S. enteritidis LPS and flagella prepared from S. meunchen, which could provide a serum test result within 1 day. This immunoblotting procedure might be a useful test to replace the Widal test as a diagnostic test for the serodiagnosis of typhoid.

In this retrospective study a total of 404 stools kept at −70°C were tested for the presence of verotoxin-producing Escherichia coli (VTEC) by the polymerase chain reaction (PCR). Thirteen positive samples from 11 patients were identified by PCR which correlated with previous isolation of E. coli O157:H7. There was no failure to detect VTEC by PCR but PCR did not identify further VTEC isolates. We concluded that (a) the occurrence of VTEC other than serotype O157:H7 is rare in our demographic area, (b) PCR is effective in the identification of E. coli O157:H7, and (c) PCR has the additional advantage over conventional culture methods of identifying VTEC, including sorbitol-fermenting serotypes, which might have been detected if we had extended our sample size.

A sensitive and specific polymerase chain reaction (PCR)-based assay was developed for detection of a single copy of human immunodeficiency virus type 1 (HIV-1) sequence. The different methodologies for preparation of clinical DNA samples were evaluated. The DNA extracted by the Ficoll-Histopaque method gave the best quality for PCR. DNA samples equivalent to 18 μl or less of blood from HIV-1 seropositive individuals were positive by this assay. This procedure should be suitable for early diagnosis of HIV-1 infection in many clinical situations.

The present study was designed to evaluate the utility of two assays, reverse transcription coupled polymerase chain reaction (RT-PCR) and branched DNA (bDNA), to accurately and reproducibly quantitate plasma human immunodeficiency virus (HIV) RNA levels. The bDNA assay quantitated RNA transcripts, prepared from different HIV-1 subtypes (A-E), within 1.5-fold. Similarly, the bDNA assay, standardized to subtype B, was used to quantitate cultured isolates from subtypes A, C-F within 2-fold; however, the RT-PCR assay displayed a 904-fold range. Reproducibility studies demonstrated that the bDNA and RT-PCR assays could be used statistically (P<0.05) to discern less than 2- and 6.8 – 8.1-fold changes in RNA levels, respectively. This study showed that the two assay methods differ in accuracy and reproducibility. These differences need to be considered when choosing specific applications for the methods.

The applicability of the polymerase chain reaction (PCR) for the diagnosis of enteroviral infections is evaluated in this study. A general primer-mediated enterovirus specific PCR was used for the detection of enteroviral RNA in cerebrospinal fluid (CSF) samples from six patients with meningitis, 56 biopsy specimens from 16 patients with congestive heart failure, and two patients with a systemic enteroviral infection. Samples from three patients with meningitis were found positive and in each of eight patients with heart disease, enteroviral RNA was detected in at least one biopsy specimen. From both patients with a systemic enteroviral infection several different organs and tissues were found positive. After reviewing the literature, we concluded that the PCR might become a useful tool for the diagnosis of persistent enteroviral infections and enteroviral-induced meningitis, but that the applicability of PCR for the routine diagnosis of acute enteroviral infections is questionable.

A 141-base pair fragment of human immunodeficiency virus-1 (HIV-1) DNA was amplified using the polymerase chain reaction (PCR). The products were slot-blotted onto a nitrocellulose membrane, revealed with a digoxigenin-labelled probe and quantitated by scanning densitometry. This method for the relative quantitation of HIV-1 DNA achieved reliable results and avoided the use of radioisotopes and the electrophoretic transfer of DNA. Testing of serial dilutions of HIV-1 extracted from infected cells revealed smooth titration curves. A reproducible increase in peripheral blood HIV-1 DNA was documented in a haemophilia patient during disease progression.