Development of a PCR assay followed by nonradioactive hybridization using oligonucleotides covalently bound to CovaLink NH microwells for detection of four Plasmodium species in blood samples from humans.

We developed and evaluated a PCR-based assay to detect four Plasmodium species in 79 blood samples from 56 travelers returning from areas where malaria is endemic. DNA amplification targeting a small region of the 18S rRNA gene was performed with Plasmodium genus-specific primers. The biotinylated PCR products were then identified by PCR-colorimetric Covalink NH microwell plate hybridization (CMPH) using species-specific phosphorylated probes covalently bound to a pretreated polystyrene surface. The results from PCR-CMPH showed high specificity, and for 47 of the 56 patients (84%), microscopy and PCR-CMPH results were in agreement. Discordant results were reevaluated with microscopy examination, other molecular methods, and DNA sequencing. Except for one patient, discrepancies were resolved in favor of PCR-CMPH: three mixed infections were detected, four species identification errors were corrected, and two negative results were shown to be positive. Our results indicate that PCR-CMPH is a simple, rapid, and specific method for malaria diagnosis. It employs stable reagents and inexpensive equipment, making it suitable for routine epidemiological use.

Genetic identification of the main opportunistic Mucorales by PCR-restriction fragment length polymorphism.

Mucormycosis is a rare and opportunistic infection caused by fungi belonging to the order Mucorales. Recent reports have demonstrated an increasing incidence of mucormycosis, which is frequently lethal, especially in patients suffering from severe underlying conditions such as immunodeficiency. In addition, even though conventional mycology and histopathology assays allow for the identification of Mucorales, they often fail in offering a species-specific diagnosis. Due to the lack of other laboratory tests, a precise identification of these molds is thus notoriously difficult. In this study we aimed to develop a molecular biology tool to identify the main Mucorales involved in human pathology. A PCR strategy selectively amplifies genomic DNA from molds belonging to the genera Absidia, Mucor, Rhizopus, and Rhizomucor, excluding human DNA and DNA from other filamentous fungi and yeasts. A subsequent digestion step identified the Mucorales at genus and species level. This technique was validated using both fungal cultures and retrospective analyses of clinical samples. By enabling a rapid and precise identification of Mucorales strains in infected patients, this PCR-restriction fragment length polymorphism-based method should help clinicians to decide on the appropriate treatment, consequently decreasing the mortality of mucormycosis.

Laboratory diagnosis of cystic hydatic disease.

The main purpose of this article is to answer the questions about which test to perform for hydatic diagnosis and when. Several techniques for biologic diagnosis and follow-up of human cystic hydatidosis are reviewed. The specificity and sensitivity of immunologic reactions are reported. The differential diagnosis between Echinococcus granulosus and E. multilocularis is examined. The characteristics of the immunologic diagnosis according to the stage and the treatment of hydatidosis disease is discussed. Laboratory diagnosis of cystic hydatic disease is complementary to the clinical data. A judicious association of the usual techniques (indirect immunofluorescence assay, indirect hemagglutination assay, immunoelectrophoresis, co-electrophoresis with antigen 5 identification) confirms the diagnosis in 80% to 94% of hepatic hydatidosis cases and in 65% of pulmonary hydatidosis cases. Special techniques (enzyme-linked immunosorbent assay, Western blot, polymerase chain reaction) must be used for other localizations or when cysts are calcified. A serologic survey is necessary for the follow-up of operated medically treated patients. Despite poor standardization, purified antigens can distinguish between E. granulosus and E. multilocularis infections, although false-positive results are observed during other helminthiases, such as cysticerocosis.

Toxoplasmosis in kidney transplant recipients: report of six cases and review.

Six patients with toxoplasmosis complicating renal transplantation are described, and 25 other reported cases are reviewed. The mean age of the 31 patients was 35.16 years. Most of the recipients (25 of 29) showed signs of toxoplasmosis within 3 months post-transplantation, with fever, neurological disturbances, and pneumonia as the main clinical features. Diagnosis was established at autopsy in 15 cases, by serology in 13 cases, and by direct examination, culture, or polymerase chain reaction of biological samples in 5 cases. Seventeen patients also had concomitant infections. The donor was the likely source of transmission to 10 recipients; reactivation was suspected in two cases. The source of transmission could not be determined for the remaining 19 patients. The mortality rate was 64.5%. Ten of the 11 patients given specific treatment survived, indicating that early diagnosis and therapy are essential.

[Application from chemiluminescence to serological diagnosis of human toxoplasmosis]. / Application de la chimiluminescence au diagnostic sérologique des toxoplasmoses humaines.

Diagnostic Products Corporation has chosen chemiluminescent for the new kit of quantitative measurement of IgG and qualitative detection of IgM antibodies to Toxoplasma gondii, 878 human sera of principal diagnosis situations were tested, and the results obtained with the IMMULITE Toxoplasmosis kit were compared with those of the Parasitology and Mycology Laboratory of the University of Lille. Chemiluminescent allows a sensitive and specific determination of immunity. In the same ways, this method is able to detect earlier specific IgM and IgG during seroconversion. The kit of quantitative measurement of IgG and qualitative detection of IgM is reproducible and sensitive; this confirms the interest for the pediatric diagnosis of congenital toxoplasmosis.

[Biological diagnosis of echinococcosis]. / Diagnostique biologique des échinococcoses.

The authors review the laboratory techniques used for diagnosis of echinococcosis (IFL, IHA, IEP, ES, ELISA) with emphasis on the value and standardization of the antigens utilized. The specificity of these tests makes it possible to differentiate hydatidosis from alveolar hydatid disease.

[Cerebrospinal fluid distribution of metampicillin (author's transl)]. / Cinétique intra-rachidienne de la métampicilline.

UNLABELLED: In order to avoid other antibiotics which interfere with plasma and CSF level determination of a given antibiotic the authors use a chemical method of established specificity. 15 patients presenting with purulent meningitis were given a standard daily dose of metampicillin and divided into 3 groups of 5. On days 2, 4 and 6 after onset of disease plasma and CSF specimens were taken at respectively 1, 2 or 4 hours following metampicillin I.V. injection according to patient group. All patients were cured between days 7 and 21 of therapy. Tolerance was good. RESULTS: --there was no correlation between dose injected and plasma level of drug,--CSF distribution of metampicillin was good and stayed identical throughout the disease period,--peak CSF drug level was delayed as compared to plasma level, occurring more than 4 hours after I.V. injection of metampicillin.