ANIMAL MODELS FOR THE STUDY OF LEISHMANIASIS IMMUNOLOGY

Leishmaniasis remains a major public health problem worldwide and is classified as Category I by the TDR/WHO, mainly due to the absence of control. Many experimental models like rodents, dogs and monkeys have been developed, each with specific features, in order to characterize the immune response to Leishmania species, but none reproduces the pathology observed in human disease. Conflicting data may arise in part because different parasite strains or species are being examined, different tissue targets (mice footpad, ear, or base of tail) are being infected, and different numbers (“low” 1×102 and “high” 1×106) of metacyclic promastigotes have been inoculated. Recently, new approaches have been proposed to provide more meaningful data regarding the host response and pathogenesis that parallels human disease. The use of sand fly saliva and low numbers of parasites in experimental infections has led to mimic natural transmission and find new molecules and immune mechanisms which should be considered when designing vaccines and control strategies. Moreover, the use of wild rodents as experimental models has been proposed as a good alternative for studying the host-pathogen relationships and for testing candidate vaccines. To date, using natural reservoirs to study Leishmania infection has been challenging because immunologic reagents for use in wild rodents are lacking. This review discusses the principal immunological findings against Leishmania infection in different animal models highlighting the importance of using experimental conditions similar to natural transmission and reservoir species as experimental models to study the immunopathology of the disease.

The Office International des Epizooties ad hoc Group on Non Tsetse Animal Trypanosomoses: Its Origin, Scope and Perspectives

A summary of the goals and activities of the ad hoc group on Non Tsetse Transmitted Animal Trypanosomosis belonging to the Office International des Epizooties is presented.

Serological evidence of hepatitis E virus infection in different animal species from the Southeast of Brazil

Serological evidence of hepatitis E virus infection (HEV) has been observed in both humans and different animal species living in non-endemic areas, suggesting that animals could be important reservoir for virus transmission to man. Antibodies to HEV have been detected in some Brazilian population groups. Nevertheless, sporadic cases of acute HEV infection have never been reported. We collected 271 serum samples from several domestic animals and also from pig handlers from Southeast of Brazil in order to investigate the seroprevalence of HEV infection. Anti-HEV IgG was detected in cows (1.42%), dogs (6.97%), chickens (20%), swines (24.3%), and rodents (50%), as well as in pig handlers (6.3%). The recognition of swine HEV infections in pigs in many countries of the world led us to investigate a larger sample of pigs (n = 357) from the same Brazilian region with ages ranging from 1 to > 25 weeks. IgG anti-HEV was detected in 100% of 7-day old pigs. Following a gradual decline between weeks 2 and 8 (probably due to loss of maternal IgG), the prevalence then steady increased until it reached 97.3% of animals older than 25 weeks. Besides the detection of anti-HEV antibodies in different animal species, the results showed that swine HEV infection seems to be almost universal within this Brazilian pig population. This is the first report that shows evidences of HEV circulation in Brazilian animal species and pig handlers.

Shared and non-shared antigens from three different extracts of the metacestode of Echinococcus granulosus

Hydatid cyst fluid (HCF), somatic antigens (S-Ag) and excretory-secretory products (ES-Ag) of Echinococcus granulosus protoscoleces are used as the main antigenic sources for immunodiagnosis of human and dog echinococcosis. In order to determine their non-shared as well as their shared antigenic components, these extracts were studied by ELISA-inhibition and immunoblot-inhibition. Assays were carried out using homologous rabbit polyclonal antisera, human sera from individuals with surgically confirmed hydatidosis, and sera from dogs naturally infected with E. granulosus. High levels of cross-reactivity were observed for all antigenic extracts, but especially for ES-Ag and S-Ag. Canine antibodies evidenced lesser avidity for their specific antigens than antibodies from human origin. The major antigenic components shared by HCF, S-Ag, and ES-Ag have apparent molecular masses of 4-6, 20-24, 52, 80, and 100-104 kDa, including doublets of 41/45, 54/57, and 65/68 kDa. Non-shared polypeptides of each antigenic extract of E. granulosus were identified, having apparent masses of 108 and 78 kDa for HCF, of 124, 94, 83, and 75 kDa for S-Ag, and of 89, 66, 42, 39, 37, and 35 kDa for ES-Ag.

Application of isotope-selective non-dispersive infrared spectrometry for the evaluation of the 13C-urea breath test: comparison with three concordant methods

The aim of this work was to compare the performance of isotope-selective non-dispersive infrared spectrometry (IRIS) for the 13C-urea breath test with the combination of the 14C-urea breath test (14C-UBT), urease test and histologic examination for the diagnosis of H. pylori (HP) infection. Fifty-three duodenal ulcer patients were studied. All patients were submitted to gastroscopy to detect HP by the urease test, histologic examination and 14C-UBT. To be included in the study the results of the 3 tests had to be concordant. Within one month after admission to the study the patients were submitted to IRIS with breath samples collected before and 30 min after the ingestion of 75 mg 13C-urea dissolved in 200 ml of orange juice. The samples were mailed and analyzed 11.5 (4-21) days after collection. Data were analyzed statistically by the chi-square and Mann-Whitney test and by the Spearman correlation coefficient. Twenty-six patients were HP positive and 27 negative. There was 100% agreement between the IRIS results and the HP status determined by the other three methods. Using a cutoff value of delta-over-baseline (DOB) above 4.0 the IRIS showed a mean value of 19.38 (minimum = 4.2, maximum = 41.3, SD = 10.9) for HP-positive patients and a mean value of 0.88 (minimum = 0.10, maximum = 2.5, SD = 0.71) for negative patients. Using a cutoff value corresponding to 0.800% CO2/weight (kg), the 14C-UBT showed a mean value of 2.78 (minimum = 0.89, maximum = 5.22, SD = 1.18) in HP-positive patients. HP-negative patients showed a mean value of 0.37 (minimum = 0.13, maximum = 0.77, SD = 0.17). IRIS is a low-cost, easy to manage, highly sensitive and specific test for H. pylori detection. Storing and mailing the samples did not interfere with the performance of the test.

Comparison of the efficacy of biodegradable and non-biodegradable scintillation liquids on the counting of tritium- and [14C]-labeled compounds

The widespread use of ³H and 14C in research has generated a large volume of waste mixed with scintillation liquid, requiring an effective control and appropriate storage of liquid radioactive waste. In the present study, we compared the efficacy of three commercially available scintillation liquids, Optiphase HiSafe 3, Ultima-Gold™ AB (biodegradable) and Insta-Gel-XF (non-biodegradable), in terms of [14C]-glucose and [³H]-thymidine counting efficiency. We also analyzed the effect of the relative amount of water (1.6 to 50%), radioisotope concentration (0.1 to 100 nCi/ml), pH (2 to 10) and color of the solutions (samples containing 0.1 to 1.0 mg/ml of Trypan blue) on the counting efficiency in the presence of these scintillation liquids. There were few significant differences in the efficiency of 14C and ³H counting obtained with biodegradable or non-biodegradable scintillation liquids. However, there was an 83 and 94% reduction in the efficiency of 14C and ³H counting, respectively, in samples colored with 1 mg/ml Trypan blue, but not with 0.1 mg/ml, independent of the scintillation liquid used. Considering the low cost of biodegradable scintillation cocktails and their efficacy, these results show that traditional hazardous scintillation fluids may be replaced with the new safe biodegradable fluids without impairment of ³H and 14C counting efficiency. The use of biodegradable scintillation cocktails minimizes both human and environmental exposure to hazardous solvents. In addition, some biodegradable scintillation liquids can be 40% less expensive than the traditional hazardous cocktails.

Isolation of human fungi from soil and identification of two endemic areas of Cryptococcus neoformans and Coccidioides immitis

The present study was carried out in two different areas of Province of Cordoba, Argentina, where there was a suspicious of endemic mycosis. The previous data were the presence of a clinical case of pulmonary cryptococcosis in one area (Alta Gracia) and the previous findings of a high incidence of coccidioidin and cryptococcin reactors in the population of the second one (Villa Dolores). In both areas soil samples for fungi were studied and Cryptococcus neoformans was found in 2/25 samples from Alta Gracia. In Villa Dolores Coccidioides immitis was isolated in 2/40 samples, and C. neoformans in 1/40 samples. Delayed hypersensitivity test with cryptococcin was determined in the population from Alta Gracia and it was found to be 5.3%. Positive cutaneous tests with coccidioidin (33.8%) and cryptococcin (31.9%) in Villa Dolores were obtained. With these findings two endemic areas of systemic mycoses in Cordoba, Argentina were delimited.

A case of human rabies in the Urban Area of Ribeirão Preto, SP, Brazil

A 39-year old male patient was admitted to the University Hospital of the Faculty of Medicine of Ribeirão Preto with signs and symptoms of sudden dyspnea, generalized myalgia and behavioral disorders. The initial suspicion was alcohol abstinence syndrome and the patient was referred for psychiatric and neurologic care. The evolution of the patient with a worsening of signs and symptoms, presence of crises of tachypnea, agitation, difficulty to swallow, irritability and hydrophobia, and his report of having been bitten by a suspected dog raised the hypothesis of rabies. The diagnosis was confirmed by examination of a corneal impression, biological tests in the cerebrospinal fluid (CSF) and saliva and visualization of Negri bodies in nervous tissue (direct immunofluorescence). The patient evolved with agitation, aggressiveness, and worsening tachypnea intercalating with apnea, and died on the 4th day after admission

Prevalence of human papillomavirus infection in rural villages of the Bolivian Amazon

Cervical cancer constitutes a major health problem in developing countries like Bolivia. The roles of certain genotypes of human papillomaviruses (HPVs) in the pathogenesis of cervical cancer is well established. The prevalence of HPV infection among sexually active women varies greatly. Information regarding HPV infection in Bolivia is very much scarce, specially in regions like the Amazonian lowland. We studied 135 healthy women living in four rural localities of the Bolivian Amazon. Presence of HPV in DNA extracted from cervical swabs was analyzed using a reverse line hybridization assay. The estimated overall HPV infection prevalence among the studied rural localities was 5.9% (ranging from 0-16.6%). These values were unexpectedly low considering Bolivia has a high incidence of cervical cancer. The fact that Amazonian people seem to be less exposed to HPV, makes it likely that some other risk factors including host lifestyle behaviors and genetic background may be involved in the development of cervical cancer in this population.

Human herpesvirus-7 as a cause of exanthematous ilnesses in Belém, Pará, Brazil

We screened sera from 370 patients suffering from exanthematous illnesses in Belém, North Brazil, for the presence of human herpesvirus-7 (HHV-7) IgM and IgG antibodies. Samples were obtained from January 1996 to December 2002 and were processed by a HHV-7-specific indirect immunofluorescence assay (IFA). HHV-7-specific IgM and/or IgG antibodies were found in 190 (51.4%) of these patients, with similar prevalence rates (IgM+ and IgG+ subgroups taken together) for female and male subjects: 52.5% and 50.3%, respectively. Serological status as defined by IgG was identified in 135 (36.5%) patients. In 55 (14.9%) of the patients HHV-7 IgM antibodies were detected. HHV-7 IgM- and- IgG antibody rates were similar (p > 0.05) when male and female subjects are compared: 14.4% versus 15.3% and 38.1% versus 35.0%, respectively. Statistically significant difference (p = 0.003) was noted when HHV-7-IgM-positive female and male patients aged 5-8 months are compared. Prevalence rates ranging from 4.6% (female, 5-8 months of age) to 93.3% (female, > 10 years of age) and 12.2% (male, 5-8 months) to 80.0% (male, 8-10 years of age) were noted in the IgG- positive subgroups. A subgroup (n = 131) of patients with IgM or IgG HHV-7 antibodies were examined for the presence of DNA using a polymerase chain reaction/nested PCR. Recent/active HHV-7 infection occurred at a rate of 11.0% (6/55) among patients whose samples presented IgM+ specific antibodies. In a subgroup (n = 76) of patients with high HHV-7-IgG antibody levels (titre > 1:160) DNA could not be detected in sera examined by PCR/nested PCR. Of the six recent/active infections, four subjects with less than 1 year and two with 3 and 6 years of age, presented typical exanthem subitum (E.S), as defined by higher fever (> 38.0 ºC) with duration of 24 to 72 hours, followed by a maculopapular skin rash. Our results underscore the need for searching HHV-7 infection in patients with exanthematous diseases, particularly those presenting with typical E.S. HHV-7 appears therefore to emerge as a newly recognized pathogen of exanthem in our region.