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PubMed2002

Comparison of an immortalized human corneal epithelial cell line with Vero cells in the isolation of Herpes simplex virus-1 for the laboratory diagnosis of Herpes simplex keratitis.

BACKGROUND: Herpes simplex keratitis (HSK) is a sight threatening ocular infection often requiring a specific and prompt laboratory diagnosis. Isolation of Herpes simplex virus (HSV-1) in culture provides the most reliable and specific method and is considered as the "Gold Standard" in the laboratory diagnosis of HSK in spite of its low sensitivity. Using "cell lines of corneal origin" for virus isolation may be beneficial under such circumstances, since these cells have been shown to be excellent substrates for the growth of HSV-1 isolated from the cornea. We report a comparative study of a novel human corneal epithelial cell line (HCE) and the Vero cell line in the isolation of HSV-1 from corneal scrapings employing a shell vial assay. METHODS: Corneal scrapings were obtained from 17 patients with a clinical diagnosis of HSK. All the cases were confirmed by virological investigations (PCR and viral antigen detection positive, n = 15, PCR positive, n = 1, Viral antigen positive, n = 1). Scrapings obtained from 10 patients with infectious keratitis of non-viral origin were included as controls. All the scrapings were simultaneously inoculated into shell vials of HCE and Vero cells. Cultures were terminated at 24 h post-infection. Isolation of HSV-1 was confirmed using an indirect immunofluorescence/ immunoperoxidase assay. RESULTS: Virus could be isolated using both or either of the cell lines in 10/17 (58.82%) patients with HSK. HSV-1 was isolated from 10/17 (58.82%) and 4/17(23.52%) specimens in HCE and Vero cells, respectively (P = 0.036). None of the controls yielded HSV-1. While all the 10 (100%) strains were isolated in HCE, Vero yielded only 4/10 (40%) strains in the shell vial culture (P = 0.014). CONCLUSIONS: HCE showed a statistically significant difference in the virus isolation rate with respect to Vero cells. HCE may be an excellent alternative cell line for the isolation of HSV-1, especially from corneal scrapings, for the laboratory diagnosis of HSK.

Animals↗
PubMed2006

Laboratory diagnosis of equine rabies and its implications for human postexposure prophylaxis.

Laboratory diagnosis is essential to confirm suspected cases of equine rabies and to determine the medical care needed for human postexposure antirabies prophylaxis. Equine rabies transmitted by the vampire bat, Desmodus rotundus, has increased gradually in the State of São Paulo. The present study has several objectives, the most important being the evaluation of fluorescent antibody test (FAT) and virus-isolation laboratory tests performed with different equine nervous system tissues (cortical, hippocampus, cerebellar, brainstem and cervical medullar) to determine the tissue for which the two techniques have the highest sensitivity. Analysis by FAT of these five regions of the central nervous system (CNS) from 35 animals showed that there was a greater amount of viral antigen in the brainstem and cervical medullar tissues than in the hippocampus, cortical and cerebellar tissues. While there were no significant differences in the mortality rate of mice inoculated with suspension prepared from the different tissues, a trend towards higher mortality rate was detected with brainstem and cervical medullar tissues. Laboratory diagnosis was not affected by whether the animal had been vaccinated or not, or whether it had died following the natural course of the disease or as a result of euthanasia. Isolation of the rabies virus in equine salivary glands demonstrated the potential risk for humans exposed to infected animals.

Animals↗
PubMed1989

Laboratory diagnosis of Lyme disease.

Different techniques have been developed to aid in laboratory diagnosis of Lyme disease. The detection of serum antibodies, in particular, is relied on heavily by the medical community and is currently the most practical means of confirming B. burgdorferi infections. Although most assays may not detect low amounts of IgM antibody during the initial weeks of infection, application of a capture ELISA method has been reported to improve test sensitivity. Antibodies to Borrelia and Treponema spirochetes will cross-react in conventional assays being used for Lyme disease, but in most cases, these problems can be eliminated by performing other serologic tests or by reviewing clinical and epidemiologic data. Further improvements in the laboratory diagnosis of Lyme disease should be made by standardizing current methods (including commercial test kits), establishing reference laboratories in the United States and Europe, and by developing rapid antigen detection procedures. Finally, serologic determination of B. burgdorferi infections should remain secondary to clinical diagnosis.

Antibodies, Bacterial↗
PubMed2001

Laboratory diagnosis of Clostridium difficile disease.

The laboratory diagnosis of Clostridium difficile-associated disease (CDAD) is based on culture and toxin detection in fecal specimens. Culture is performed on a commercially available selective media. C. difficile colony morphology is typical when viewed under a dissecting microscope. Definitive identification is best obtained by gas liquid chromatography. Culture is very sensitive but, when used alone without toxin testing, it leads to low specificity and misdiagnosis of CDAD when high rates of asymptomatic carriage exist. Toxin detection by a tissue culture cytotoxin assay followed by neutralisation with specific antiserum is often considered the standard. However, this approach lacks sensitivity and has not detected up to 30% of patients with confirmed CDAD. Multiple enzyme immunoassays (EIAs) have been introduced by various manufacturers for the detection of toxin A alone or for both toxins A and B. Some of these are designed to give results in less than 1 h. Comparative studies of EIA kits reported that the sensitivity and specificity are slightly lower than cytotoxin assays. Toxigenic culture tests C. difficile isolates for toxin production: colonies isolated on selective media are tested for in-vitro toxin production either by a cytotoxicity assay or by direct EIA. It has higher sensitivity than the cytotoxicity assay and equivalent specificity. In the routine laboratory, culture and toxin detection should be performed on every specimen and, in culture-positive and fecal toxin-negative cases, toxigenic cultures should be performed on isolated colonies.

Antibodies, Bacterial↗
PubMedDate not supplied

Laboratory diagnosis of Chagas' heart disease.

The laboratory diagnosis of Chagas' disease is a complex one. Factors relating to the host immune response and the antigenic variability of T. cruzi must be considered in the final interpretation of tests results. Parasitologic methods for detecting T. cruzi, immunologic methods for detecting T. cruzi antigens in different biological fluids and serologic tests for detection and quantification of different classes of immunoglobulins are well standardized and used in the diagnosis of the acute or chronic phase of the disease. Xenodiagnosis is the most common parasitologic test employed, although it detects only 50% of infections in the chronic phase. Indirect immunofluorescence for detecting IgG and IgM antibodies, hemagglutination and enzyme immunoassay are the serologic tests most frequently employed for diagnosis, to screen blood donors and for seroepidemiologic studies. An important caveat to be remembered is that serologic tests provide only a probable diagnosis, which depends on the prevalence of Chagas disease, as well as on the sensitivity and specificity of the test employed. The use of well defined specific antigens, obtained through recombinant methods or chromatography, opens an important field for the development of very specific tests, without significant loss of sensitivity.

Agglutination Tests↗
PubMedDate not supplied

Quality assessment of malaria laboratory diagnosis in South Africa.

To assess the quality of malaria diagnosis in 115 South African laboratories participating in the National Health Laboratory Service Parasitology External Quality Assessment Programme we reviewed the results from 7 surveys from January 2000 to August 2002. The mean percentage incorrect result rate was 13.8% (95% CI 11.3-16.9%), which is alarmingly high, with about 1 in 7 blood films being incorrectly interpreted. Most participants with incorrect blood film interpretations had acceptable Giemsa staining quality, indicating that there is less of a problem with staining technique than with blood film interpretation. Laboratories in provinces in which malaria is endemic did not necessarily perform better than those in non-endemic areas. The results clearly suggest that malaria laboratory diagnosis throughout South Africa needs strengthening by improving laboratory standardization and auditing, training, quality assurance and referral resources.

Clinical Laboratory Techniques↗
PubMed1995

Haemophilus paraphrophilus infection: a pitfall in laboratory diagnosis.

Many clinical laboratories have difficulty in identifying a group of organisms which are catalase negative, oxidase positive, Gram negative rods. We describe a case of purulent sacroiliitis due to Haemophilus paraphrophilus where the organism was initially misidentified as Eikenella corrodens leading to inappropriate antimicrobial chemotherapy. We review the strains of H. paraphrophilus and E. corrodens that were identified by the National Collection of Type Cultures over the last ten years. Only 21 of 100 strains identified as E. corrodens were submitted as E. corrodens. Seven strains submitted as possible E. corrodens were identified as H. paraphrophilus. Several different species of Gram negative rods may produce pitting on agar and this seems to be poorly recognised. However, further tests are available to facilitate correct identification of these strains.

Adult↗
PubMed1991

[Neurocysticercosis in childhood. I. Clinical and laboratory diagnosis].

Clinical and laboratory findings among 34 children (age range from 15 months to 13 years) with neurocysticercosis were reviewed. The main symptoms were: intracranial hypertension, 21 cases (62%); epilepsy, 20 cases (59%); hemiplegia, 4 cases (12%). Computed tomography (CT) in 33 children showed typical brain active cysts in 26 patients (79%) and calcifications in 2 (6%). The complement fixation reaction or the indirect immunofluorescent test for cysticercus antibody were positive in CSF in 77% (20 out of 26 patients) and serum in 78% (18 out of 23 patients). CSF pleocytosis was found in 57% of the cases (15/26 patients) with eosinophil cells in 27% (7/26 patients). The most efficient diagnostic test was CT scan of the head, and was confirmed immunologically by measurement of cysticercus antibody titers in CSF and serum.

Adolescent↗
PubMed1987

Studies into the immunopathogenesis and laboratory diagnosis of dengue haemorrhagic fever.

Studies were carried out into the immunopathogenesis and laboratory diagnosis of dengue virus infections. Using an experimental system it was shown that cell-mediated immunity (CMI), as measured by delayed-type hypersensitivity (DTH) was induced in mice infected with dengue virus. The nature of the DTH response satisfies most criteria for a classical DTH reaction. In addition, it was also shown that infection with dengue virus causes a transient immunosuppression as measured by the immune response to other, unrelated antigens. With regard to the laboratory diagnosis of dengue infections, it was found that mosquito cells were a sensitive system for the isolation of dengue viruses and that the success of isolation was related to the antibody content of the serum. A new method for the rapid isolation of dengue viruses was also developed involving the intracerebral inoculation of mosquito larvae. By the use of this method viral antigens can be detected as early as 2-3 days after specimen inoculation. The significance of these findings in relation to the immunopathogenesis, prevention and control of disease syndromes due to dengue viruses is discussed.

Animals↗
PubMed1997

[Current developments in the laboratory diagnosis of rubella].

Thirty years after the introduction of the hemagglutination inhibition assay (HAI), laboratory diagnosis of rubella virus infection has achieved a high reliability. While the HAI remains the reference standard against which newer assays are compared, routine laboratory diagnosis is based mainly on ELISA tests which permit a more rapid and less cumbersome detection of specific IgG and IgM antibody. Although quantification of immunoglobulin G against rubella virus is performed using WHO standards, the correlation between different ELISAs is relatively poor. Despite substantial improvements in virus isolation and nucleic acid amplification techniques, serology remains the mainstay of diagnosis for both acquired and postnatal diagnosis of congenital infection. Differentiation between primary and re-infection is of critical importance during pregnancy and can be achieved relatively reliably by antibody avidity determination or by immunoblot. While current anti-rubella IgM ELISAs are relatively sensitive, their specificity may be limited by cross reactivity with other viruses, i.e. parvovirus B19 and Epstein-Barr virus. Maternal reinfection with congenital rubella syndrome is very rare, however it may be misdiagnosed in the absence of significant IgG antibody titer change and/or IgM antibody.

Enzyme-Linked Immunosorbent Assay↗
PubMed1990

Comparison of a dot immunobinding assay, latex agglutination, and cytotoxin assay for laboratory diagnosis of Clostridium difficile-associated diarrhea.

C. diff-CUBE, a dot immunobinding assay (DIA) (Difco Laboratories, Ann Arbor, Mich.) for detection of Clostridium difficile toxin A in stool specimens, was compared with latex agglutination (LA) (Marion Laboratories, Kansas City, Mo.) and cytotoxin assay (CTA) for the laboratory diagnosis of C. difficile-associated diarrhea. A total of 200 stool specimens collected from 169 patients with suspected C. difficile diarrhea were tested. Of the 198 specimens evaluated by all three methods, 36 (18%) from 36 patients were positive by one or more of the tests. Twenty-five, 26, and 23 specimens were positive by CTA, DIA, and LA, respectively; 14 were positive by all three methods. Eight specimens yielded nonspecific LA test results; all eight were negative by CTA, and one was positive by DIA. DIA results agreed with CTA results in 183 (92%) cases and with LA results in 175 (88%) cases. CTA and LA results agreed in 179 (90%) cases. Freezing of the specimen did not appear to adversely affect either the DIA or LA test. These preliminary results suggest that C. diff-CUBE may be useful as a rapid screen for the diagnosis of C. difficile-associated diarrhea. However, for optimum laboratory diagnosis, further testing of all stools that are negative by DIA is warranted.

Bacterial Toxins↗
PubMedDate not supplied

[Congenital ChagaśDisease: epidemiology, laboratorial diagnosis, prognosis and treatment].

The authors review studies about epidemiology, clinical aspects and methods used in laboratorial diagnosis of congenital Chagas'disease, emphasizing the limitations in their specificity and sensibility, and suggest alternative methods to improve the accuracy and the quality of the laboratorial diagnosis of congenital Chagaśdisease, essential to an efficient treatment.

English Abstract↗
PubMed1982

Laboratory diagnosis and oral treatment of CAPD peritonitis.

The laboratory diagnosis of 50 consecutive episodes of peritonitis in patients undergoing continuous ambulatory peritoneal dialysis (CAPD) was studied. The technique which yielded the highest rate (84%) of positive bacteriological diagnoses was inoculation and subculture of thioglycollate broth. Cloudiness of fluid to the naked eye was an accurate predictor of a raised white cell count. A minimum laboratory protocol for the bacteriological diagnosis of CAPD peritonitis was devised and has been in use satisfactorily since the completion of the study. Antibiotic treatment was given orally in the first instance in 43 episodes and was successful in 34.

Administration, Oral↗
PubMed1979

A course in physical, clinical, and laboratory diagnosis for dental education.

A basic knowledge of physical diagnosis techniques and an understanding of those parts of clinical medicine (including laboratory diagnosis) most closely related to dentistry are recognized to be important parts of dental education. The course in physical, clinical, and laboratory diagnosis for dentists pursuing specialty training in oral surgery, endodontics, periodontics, oral diagnosis, prosthodontics, and general practice at The University of Michigan School of Dentistry is described.

Clinical Competence↗
PubMed2002

Laboratory diagnosis of metabolic myopathies.

The metabolic myopathies are a heterogeneous group of disorders inherited by a variety of modes that include gene defects in both the nuclear and mitochondrial genomes. Many factors impact on the expression of the pathogenic mutations that cause these disorders including genetic background, environmental factors, and coexisting disorders. Molecular technology has greatly improved the ability to make definitive diagnoses in many of the metabolic myopathies in the last decade and particularly has demonstrated that the wide diversity in the severity of mutations contributes to understanding genotype-phenotype correlations. In some cases, molecular testing obviates the necessity to perform an invasive muscle biopsy. However, it is also clear that the diagnostic yield from molecular testing is incomplete and particularly low among the mitochondrial myopathies as a group, ranging from approximately 6% to 19% in well-classified high-risk groups. Therefore, it is often essential to combine clinical, biochemical, histopathologic, and molecular data for each patient in order to arrive at a definitive diagnosis. The approach to the laboratory diagnosis of metabolic myopathies is described emphasizing both noninvasive and invasive testing, highlighting the molecular methodologies with the benefits and disadvantages of each technology, and documenting how to determine whether patients have coexisting disorders.

Clinical Laboratory Techniques↗
PubMed1991

Laboratory diagnosis of von Willebrand's disease.

The laboratory diagnosis of von Willebrand's disease (vWD) has become much more difficult because of the identification of numerous variant forms of vWD. The biologic and pathologic variability in individual patients necessitates a comprehensive assessment. Patients with classic type I vWD may be easily identified by using the bleeding time, activated partial thromboplastin time, platelet count, von Willebrand antigen, and ristocetin cofactor tests. In patients with variant forms of vWD, however, multimeric analysis of both platelet and plasma von Willebrand factor may be necessary. Furthermore, more than one assessment may be needed to detect an abnormal result in many of the aforementioned tests.

Bleeding Time↗