RNA/DNA co-analysis from blood stains--results of a second collaborative EDNAP exercise.

A second collaborative exercise on RNA/DNA co-analysis for body fluid identification and STR profiling was organized by the European DNA Profiling Group (EDNAP). Six human blood stains, two blood dilution series (5-0.001 µl blood) and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by the participating laboratories using a RNA/DNA co-extraction or solely RNA extraction method. Two novel mRNA multiplexes were used for the identification of blood: a highly sensitive duplex (HBA, HBB) and a moderately sensitive pentaplex (ALAS2, CD3G, ANK1, SPTB and PBGD). The laboratories used different chemistries and instrumentation. All of the 18 participating laboratories were able to successfully isolate and detect mRNA in dried blood stains. Thirteen laboratories simultaneously extracted RNA and DNA from individual stains and were able to utilize mRNA profiling to confirm the presence of blood and to obtain autosomal STR profiles from the blood stain donors. The positive identification of blood and good quality DNA profiles were also obtained from old and compromised casework samples. The method proved to be reproducible and sensitive using different analysis strategies. The results of this collaborative exercise involving a RNA/DNA co-extraction strategy support the potential use of an mRNA based system for the identification of blood in forensic casework that is compatible with current DNA analysis methodology.

mRNA profiling for the identification of blood--results of a collaborative EDNAP exercise.

A collaborative exercise on mRNA profiling for the identification of blood was organized by the European DNA Profiling Group (EDNAP). Seven blood samples and one blood dilution series were analyzed by the participating laboratories for the reportedly blood-specific markers HBB, SPTB and PBGD, using different kits, chemistries and instrumentation. The results demonstrate that HBB is expressed abundantly in blood, SPTB moderately and PBGD significantly less. All but one of the 16 participating laboratories were able to successfully isolate and detect RNA from the dried bloodstains even though a majority of the laboratories had no prior experience with RNA. Despite some expected variation in sensitivity between laboratories, the method proved to be reproducible and sensitive using different analysis strategies. The results of this collaborative exercise support the potential use of mRNA profiling as an alternative to conventional serological tests.

Characterization of HSFY, a novel AZFb gene on the Y chromosome with a possible role in human spermatogenesis.

HSFY (heat shock transcription factor, Y chromosome) has been mapped in the AZFb region of the Y chromosome, whose deletion results in severe male infertility. HSFY belongs to the heat shock factor family that has been shown to be implicated in spermatogenesis both in animals and humans. We report the characterization of the genomic structure, the number of copies on the Y chromosome and the expression of the gene. By comparison of data obtained from expression analysis and from GenBank cDNA sequences, seven exons were identified. Alternative splicing generates three different transcripts and proteins, each containing an HSF domain typical of HSF proteins. Two identical and functional full-length copies of HSFY map in palindrome P4 of AZFb, whereas four similar sequences mapping in two clusters in palindrome P1 of AZFc and P3 seem to represent pseudogenes. Sequences similar to few HSFY exons are also located in the short arm of chromosomes Y, X and 22. Expression analysis revealed that the three HSFY transcripts are differentially expressed, transcript 1 being present in many tissues including testis and ejaculated sperm, and transcripts 2 and 3 being testis-specific. These data suggest that HSFY could have an important role in human spermatogenesis.

[Alteration of spermatogenesis and Y chromosome microdelations. Analysis of the DAZ gene family]. / Alterazioni della spermatogenesi e microdelezioni del cromosoma Y. Analisi della famiglia genica DAZ.

The Y chromosome has a fundamental role in sex determination and regulation of spermatogenesis. Three regions (designated as AZFa, b, and c) on the long arm of this chromosome exist, deletions of which result in severe damage to spermatogenesis with azoospermia or severe oligozoospermia. Recent progresses in molecular biology and extraordinary development of assisted reproduction techniques contributed to the research on this chromosome and the genes involved in spermatogenesis. About 10-15% of subjects affected by azoospermia or severe oligozoospermia carry a deletion in one or more AZF regions, 60% of which involves AZFc. The genes responsible for the testicular phenotype observed in these subjects are DBY and USP9Y for AZFa, RBMY1 for AZFb, and DAZ for AZFc. In this article, the current knowledge on biology and genetics of the Y chromosome are reported with particular interest to deletions found in infertile subjects. Furthermore, the more recent advances on DAZ gene and its role in spermatogenesis and male infertility are discussed.

Control of parasitic infections among school children in the peri-urban area of Botucatu, São Paulo, Brazil.

The prevalence of intestinal parasitosis was investigated in a primary school located in Rubião Júnior, a peri-urban district of Botucatu, São Paulo state, Brazil, in order to assess the effect of treatment and practical measures of prophylaxis in the control of parasitic infections among 7-to-18-year-old school children of a low socio-economic status. The first series of parasitological examinations included 219 school children, of which 123 (56.1%) were found to be infected with one or more parasite species. Eighty-four children carrying pathogenic parasites were submitted to various anti-parasitic treatment schedules. We re-evaluated 75 (89%) students after 4 to 6 months post-chemotherapy. The results indicate that the combination of treatment with prophylactic measures has been successful in the control of parasitic infections, since reinfection rates were generally low (< or = 5.3%), except for Giardia lamblia infections (18.6%), and a marked reduction on the prevalence rates was observed with a significant percentage of cure (> or = 73.1%) in children infected with most parasite species. The reasons for the apparent failure in the control of infections caused by Hymenolepis nana and Strongyloides stercoralis are discussed.

[Effect of Momordica charantia L. in mice infected with Plasmodium berghei]. / Efeito de Momordica charantia L.em camundongos infectados por Plasmodium berghei.

According to the World Health Organization malaria is one of the major public health problems in Brazil and all over developing countries, where 80% of the population use traditional medicine to solve their primary medical problems. Both treatment and control of this parasitosis have become difficult, because of parasite strains that are resistant to conventional drugs, such as chloroquine. That makes the search for new antimalarial drugs not only important but urgent. We aimed therefore at evaluating the effects of Momordica charantia L. (Cucurbitaceae) in mice infected with Plasmodium berghei. We used aquose and ethanotic extracts in a dose of 1000 mg/kg of body weight, orally, for five consecutive days (i.e. from day 2 to day 6 postinfection). We then followed up the parasitaemia during the course of infection. Although the population use this plant as an antimalarial, in our experimental conditions, M. charantia extracts have not shown such activity.

[Serologic study of experimental infections by Trypanosoma evansi, in guinea pigs]. / Estudo sorológico de infecções experimentais por Trypanosoma evansi, em cobaias.

Sera of 20 guinea-pigs experimentally infected with Trypanosoma evansi were obtained in order to compare the efficacy of gel diffusion, indirect immunofluorescence and agglutination tests, to detect antibodies to T. evansi. The fluorescent antibody test was positive in six (6) animals and the antibody titres were very low (1:4 to 1:16). The agglutination test detected trypanosomal antibodies in sera one (1) week after infection. After two (2) weeks all animals were positive with high titres (1:8.000 to 1:250.000). Agglutination was inhibited when sera were treated with 2-Mercapto-ethanol. This fact suggests that IgM is the principal class of antibodies in sera of infected guinea-pigs. Precipitating antibodies were not detected during the course of infection.

Malaria no municipio de Humaita, Estado do Amazonas. II - Alguns aspectos epidemiologicos comparativos entre 1976 e 1979. / Malaria in the municipality of Humaita, State of Amazonas. II - Some comparative epidemiological aspects of 1976 and 1979

Em 1976 foram estudados 409 e em 1979 foram observados 330 individuos, habitantes das diversas regioes do Municipio de Humaita em povoados situados ao longo das estradas, em localidades na calha do Rio Madeira e na zona urbana.Foram feitas observacoes clinicas completas de todos os individuos estudados, incluindo os antecedentes epidemiologicos. O estudo epidemiologico compreendeu os seguintes elementos: historia pregressa de surtos de malaria, presenca de febre, esplenomegalia e indice esplenico. No seu conjunto os resultados indicam ser a malaria mais frequente, grave e persistente nas estradas, ao contrario das localidades situadas ao longo do Rio Madeira onde ela mostrou tendencia a diminuir no periodo de 1976 a 1979 e sugerem ainda sua ausencia na zona urbana

Alteracoes do sodio, potassio e ureia sericos em camundongos experimentalmente infectados com T. cruzi (Cepa Y). / Serum changes of sodium, potassium and urea in mice experimentally infected with T. cruzi (Strain Y)

Camundongos de 25 dias de idade foram inoculados intraperitonialmente com 200 tripomastigotas sanguineos de T. cruzi, cepa Y. Semanalmente 15 camundongos foram sacrificados para verificacao dos niveis sericos de sodio, potassio e ureia. Os niveis sericos de sodio, potassio e ureia foram encontrados alterados apos 4 a 6 semenas de infeccao. Nossos achados mostram que, mesmo pequenos inoculos, acarretam alteracoes bioquimicas em camundongos em fase cronica da doenca de Chagas