COMPARISON OF A COMMERCIAL ENZYME IMMUNOASSAY WITH PLAQUE REDUCTION NEUTRALIZATION FOR MATERNAL AND INFANT MEASLES ANTIBODY MEASUREMENT

The most practicable assay for measurement of measles IgG (mIgG) in large numbers of sera is an enzyme immunoassay (EIA). To assess how EIA results would agree with those by the gold standard method of plaque reduction neutralization (PRN) we compared the results from the two methods in 43 pairs of maternal and umbilical cord sera, and sera from the corresponding infants when aged 11 - 14 months. In maternal-cord sera, the differences between mean antibody levels by EIA or PRN were not statistically significant, though in individual sera, differences could be large. However, agreement was less good for infants sera, in which levels of mIgG were very low. The conclusions of a study of transplacental transport of mIgG would not be affected by the use of either technique. When studying waning immunity in infants, PRN should be the method of choice, while results from studies using EIA should be interpreted with caution.

TOXOCARIASIS: SEROLOGICAL DIAGNOSIS BY INDIRECT ANTIBODY COMPETITION ELISA

Toxocariasis is caused by infection of man by Toxocara canis and Toxocara cati larvae, the common roundworm of dogs and cats. Because larvae are difficult to detect in tissues, diagnosis is mostly based on serology. Non specific reactions are observed mainly due to cross-reactivity with Ascaris sp antigens. This investigation aimed at developing and evaluating an indirect antibody competition ELISA (IACE) employing a specific rabbit IgG anti-Toxocara canis excretory-secretory antigens as the competition antibody, in order to improve indirect ELISA specificity performed for toxocariasis diagnosis. For that, the rabbit IgG was previously absorbed by Ascaris suum adult antigens. Sensitivity and specificity of IACE were first evaluated in 28 serum samples of mice experimentally infected with T. canis embryonated eggs. Adopting cut-off value established in this population before infection, sensitivity and specificity were 100% after 20 days post-inoculation. For human population IACE was evaluated using sera from 440 patients with clinical signs of toxocariasis and the cut-off value was established with 60 serum samples from apparently healthy individuals. Using as reference test the indirect ELISA performed by Adolfo Lutz Institute, sensitivity was 60.2%, specificity was 98% and concordance was 77.3%. Repeatability of IACE was evaluated by the inter-reactions variation coefficient (2.4%).

Serological diagnosis of toxoplasmosis: usefulness of IgA detection and IgG avidity determination in a patient with a persistent IgM antibody response to Toxoplasma gondii

We report the detection of specific IgA antibodies and the determination of IgG avidity in sequential serum samples from a patient exhibiting significant levels of Toxoplasma-specific IgM antibodies for seven years after the onset of the clinical symptoms of toxoplasmosis. IgM antibodies were detected by an indirect immunofluorescence test and by three commercial enzyme-linked immunosorbent assays (ELISA). Anti-T. gondii IgA was quantified by the a-capture ELISA technique using a commercial kit. As defined by the manufacturer of the IgA ELISA test used, most patients with acute toxoplasmosis have antibody levels > 40 arbitrary units per ml (AU/mL). At this cut-off level, the patient still had a positive ELISA result (45 AU/mL) in a serum sample taken one year after the beginning of clinical manifestations. The IgG avidity-ELISA test was performed with the Falcon assay screening test (F.A.S.T.®) - ELISA system. Avidity indices compatible with a recent Toxoplasma infection were found only in serum samples taken during the first 5 months after the onset of the clinical symptoms of toxoplasmosis. These results show that the interpretation of positive IgM results as indicative of recently acquired toxoplasmosis requires additional laboratory confirmation either by other tests or by the demonstration of a significant rise in the antibody titers in sequential serum samples.

Production of TNF-a by primary cultures of human keratinocytes challenged with Loxosceles gaucho venom

Primary cultures of human keratinocytes were challenged with increasing doses from 10 ng/mL to 2 mg/mL of Loxosceles gaucho venom, responsible for dermonecrotic lesion in humans. TNF-a was investigated by bioassay and ELISA in the supernatant of the cultures challenged with 100 ng/mL, 500 ng/mL, 1 and 2 mg/mL of venom. TNF-a was detected by bioassay in the supernatant of cultures challenged with 100 ng/mL, after 6 h. The cytokine was detected by ELISA in the supernatant of the cells challenged with doses of l mg/mL, after 6 and 12 h. The results point out the capacity of this venom to activate the keratinocytes in primary cultures to produce TNF-a. The production of cytokines could contribute to the local inflammatory process in patients bitten by Loxosceles sp.

Detection of Toxoplasma gondii soluble antigen, SAG-1(p30), antibody and immune complex in the cerebrospinal fluid of HIV positive or negative individuals

Active infection by T. gondii was evaluated by immunoassay for soluble SAG-1 (p30), the major surface antigen from T. gondii, specific antibodies and immune complexes in human cerebrospinal fluid (CSF) samples. A total of 263 samples of CSF were collected from hospitalized patients presenting neurological disorders and analyzed for antibodies to HIV. Patients were divided into two groups: HIV positive (n = 96) or HIV negative (n =167). The results of the assays showed that 45% of all samples were positive for soluble SAG-1. Toxoplasma Ag/Ab immune complexes were detected in 19% of the CSF samples and 62% were positive for T. gondii- specific IgG. A combination of these assays in the presence of clinical findings consistent with active Toxoplasma infection may predict the presence of toxoplasmic encephalitis. Moreover, detection of soluble SAG-1 in the CSF of these individuals appears consistent with active infection.

Antibody response in cattle after vaccination with inactivated and attenuated rabies vaccines

Despite the absence of current official reports showing the number of cattle infected by rabies, it is estimated that nearly 30,000 bovines are lost each year in Brazil. In order to minimize the important economic losses, control of the disease is achieved by eliminating bat colonies and by herd vaccination. In this study, we compare the antibody response in cattle elicited by vaccination with an attenuated ERA vaccine (AEvac) and an inactivated-adjuvanted PV (IPVvac) vaccine. The antibody titers were appraised by cell-culture neutralization test and ELISA, and the percentage of seropositivity was ascertained for a period of 180 days. IPVvac elicited complete seropositivity rates from day 30 to day 150, and even on day 180, 87% of the sera showed virus-neutralizing antibody titers (VNA) higher than 0.5IU/ml. There were no significant differences between the VNA titers and seropositivity rates obtained with IPVvac in the two methods tested. AEvac, however, elicited significantly lower titers than those observed in the group receiving inactivated vaccine. In addition, the profiles of antirabies IgG antibodies, evaluated by ELISA, and VNA, appraised by cell-culture neutralization test, were slightly different, when both vaccines were compared.

Central nervous system paracoccidioidomycosis. Report of a case successfully treated with Itraconazol

Paracoccidioidomycosis (PCM) is a primary pulmonary infection that often disseminates to other organs and systems. Involvement of the central nervous system (CNS) is rare and due to the fact that both clinical alertness and establishment of the diagnosis are delayed, the disease progresses causing serious problems. We report here a case of neuroparacoccidioidomycosis (NPCM), observed in a 55 year-old male, who consulted due to neurological symptoms (left hemiparesis, paresthesias, right palpebral ptosis, headache, vomiting and tonic clonic seizures) of a month duration. Upon physical examination, an ulcerated granulomatous lesion was observed in the abdomen. To confirm the diagnosis a stereotactic biopsy was taken; additionally, mycological tests from the ulcerated lesion and a bronchoalveolar lavage were performed. In the latter specimens, P. brasiliensis yeast cells were visualized and later on, the brain biopsy revealed the presence of the fungus. Treatment with itraconazole (ITZ) was initiated but clinical improvement was unremarkable; due to the fact that the patient was taking sodium valproate for seizure control, drug interactions were suspected and confirmed by absence of ITZ plasma levels. The latter medication was changed to clonazepam and after several weeks, clinical improvement began to be noticed and was accompanied by diminishing P. brasiliensis antigen and antibody titers. In the PCM endemic areas, CNS involvement should be considered more often and the efficacy of itraconazole therapy should also be taken into consideration.

Significance of isolated hepatitis B core antibody in blood donors from São Paulo

The clinical significance of isolated anti-HBc is still a challenge. To elucidate the real importance of this finding in our blood donors, an investigation algorithm was tested. One hundred and twelve isolated anti-HBc seropositive blood donors underwent clinical evaluation and retesting of HBV markers. Those who presented repeatedly reactive isolated anti-HBc, received a single dose of hepatitis B recombinant vaccine to verify anti-HBs early response. A HBV-DNA determination by PCR was done for those who did not test positive to anti-HBs after vaccine. The level of anti-HBc was recorded as a ratio of the sample-to-cut-off values (S:C ratio) in 57 candidates at donation. Comparing true and false-positive anti-HBc results, the different S:C ratios of them were statistically significant and when less than 2, implying in a false-positive result probability over 80%. A high percent of false-positive results (16.07%) was verified after anti-HBc retesting. HBV immunity was characterized in 49.11%, either by anti-HBs detection in retesting (15.18%), or after a single dose HBV vaccination (33.93%). HBV-DNA was negative in all tested donors. In conclusion, this algorithm was useful to clarify the meaning of isolated anti-HBc in most of our blood donors.

Evaluation of hemostasis disorders and anticardiolipin antibody in patients with severe leptospirosis

A prospective study was designed to evaluate disorders of hemostasis and levels of anticardiolipin antibodies (ACL) in 30 patients with severe leptospirosis and acute renal failure (ARF) (ARF was defined as serum creatinine > or = 1.5 mg/dL). The patients had been admitted to the Walter Cantídio University Hospital, São José Infectious Diseases Hospital and General Hospital of Fortaleza, Ceará, from August 1999 to July 2001. They all were male, with a mean age of 32 ± 14 years and with clinical and laboratory diagnoses of ARF leptospirosis. The time elapsed between onset of symptoms and the first hemorrhagic manifestation was 9 ± 4 days. Bleeding was observed in 86% of the patients. Laboratory tests showed significantly high levels of urea (181 ±95 mg/dl), fibrinogen, (515 ± 220 mg/dl), prothrombin time (13.3 ± 0.9 seconds) and low platelet counts (69 ± 65x10³/mm³) on admission. There was no elevation in activated partial thromboplastin time or thrombin time. Levels of IgM and IgG ACL concentrations were significantly increased (p < 0.05) in leptospirosis patients when compared to control patients (28.5 ± 32.4 vs. 11.5 ± 7.9MPL U/ml and 36.7 ± 36.1 vs. 6.5 ± 2.5 GPL U/ml), respectively. Vasculitis, thrombocytopenia and uremia should be considered important factors for the pathogenesis of hemorrhagic disturbances and the main cause of death in severe leptospirosis.

Primary cutaneous cryptococcosis due to Cryptococcus neoformans var. gattii serotype B, in an immunocompetent patient

The authors report a male patient, a seller with no detected immunosuppression, with an extensive ulcerated skin lesion localized on the left forearm, caused by Cryptococcus neoformans var. gattii serotype B. Oral treatment with fluconazole was successful. A review of the literature showed the rarity of this localization in HIV-negative patients. In contrast, skin lesions frequently occurs in HIV-positive patients, with Cryptococcus neoformans var. neoformans serotype A predominating as the etiological agent. In this paper, the pathogenicity of C. neoformans to skin lesions in patients immunocompromised or not, is discussed, showing the efficacy of fluconazole for the treatment of these processes.

A simple and cheaper in house varicella zoster virus antibody indirect ELISA

We have developed a cheaper an simple in house indirect ELISA that uses the live attenuated VZV vaccine as a coating antigen. The alternative ELISA had an agreement of 94% when compared with a commercial VZV ELISA kit. Moreover, our ELISA proved to be more reliable than the kit when assessing true negative samples. By adding a standard serum, we were able to produce results in international units per millilitre. Also, the addition of an extra step with 8M urea allowed the assessment of VZV IgG avidity without excessive costs. The cost per sample to test VZV IgG was 2.7 times cheaper with our ELISA, allowing the testing of many samples without the burden of production of VZV antigen in the laboratory.

Primary pulmonary AIDS-related lymphoma

Extranodal involvement is common in lymphomas associated with human immunodeficiency virus infection (HIV) and acquired immunodeficiency syndrome (AIDS). However, primary pulmonary AIDS-related non-Hodgkin's lymphoma is very rare and only few reports were published in the medical literature. Clinical presentation is nonspecific, with "B" and respiratory symptoms. Also, patients were with advanced immunodeficiency at the time of diagnosis. Generally, chest radiography showed peripheral nodules or cavitary masses. Primary pulmonary lymphoma associated with AIDS is generally a high-grade B-cell non-Hodgkin lymphoma and Epstein-Barr virus is strongly associated with the pathogenesis of these tumors. We report a patient with AIDS and primary pulmonary lymphoma which clinical presentation was a total atelectasis of the left lung.

Primary liver AIDS-related lympoma

Non-Hodgkin's lymphomas (NHL) are the second most frequent malignancies in AIDS patients. The majority of NHL associated with AIDS involves extranodal sites, especially the digestive tract and the central nervous system. Primary liver lymphoma (PLL) is an uncommon neoplasm among these patients. Ultrasonography and computed tomography scans may be helpful in the diagnosis of focal hepatic lymphoma. Image-guided fine-needle biopsy with histopathology of the liver lesions is the gold standard for the diagnosis of hepatic lymphoma. We report a case of PLL as the initial manifestation of AIDS in a patient without any previous infection by hepatitis C or B virus, presented as multiple and large hepatic masses.

Seroconvertion to hepatitis B vaccine after weight reduction in obese non-responder

Decreased responses to hepatitis B vaccine have been associated with some host conditions including obesity. Susceptible non-responders to a primary three-dose vaccine series should be revaccinated. Those who maintain a non-responder condition after revaccination with three vaccine doses are unlikely to develop protection using more doses. This is a description of an obese woman who received six doses of hepatitis B vaccine and persisted as a non-responder. She was submitted to a vertical banded gastroplasty Roux-en-Y gastric bypass Capellas's technique. After weight reduction, she received three additional doses of vaccine and seroconverted. Further studies should help clarify the need to evaluate antibody levels and eventually revaccinate the increasing population of individuals who undergo weight reduction.