Trypanosoma cruzi: identification of specific epimastigote antigens by human immune sera

Soluble antigens from epimastigotes of Trypanosoma cruzi were analyzed by western blot in terms of their reactivity with sera from patients with Chagas' disease. In addition, sera from patients with visceral (AVL) and tegumentar leishmaniasis (ATL) were also tested in order to identify cross-reactivities with Trypanosoma cruzy antigens. Twenty eight polypeptides with molecular weights ranging from 14 kDa to 113 kDa were identified with sera from Chagas' disease patients. An extensive cross-reactivity was observed when sera from human visceral leishmaniasis were used, while only a slight cross-reaction was observed with sera from tegumentar leishmaniasis. On the other hand, 10 polypeptidesspecifically reacting with sera from Chagas' disease patients were identified. Among them, the antigens with molecular weights of 46 kDa and 25 kDa reacted with all sera teste and may be good candidates for specific immunodiagnosis of Chagas' disease.

Identification and characterization of sex-linked proteins of Schistosoma mansoni

The proteins of adults worms (male and female) of two isolates (BH and RJ) of Shistosoma mansoni were extracted using Triton X-114 phase separation. The SDS-polyacrilamide gel electrophoresis profiles of the three phases (detergent, aqueous and insoluble proteins) obtained were compared after Coomassie blue and silver staining, surface radioiodination and Western blotting. No major differences were detected between the 2 isolates. Of the 25 or more proteins which partitioned into the detergent phase, only about 8 proteins could be surface radiodinated on live adult worms. A comparison was also made between the profiles of mael and females worms, isolated from bisexually infected mice. Two major female-specific and one male-specific band were detected by silver and/or Coomassie staining. The female bands, 32 KDa and 18 KDa, partitioned into the detergent and aqueous phase, respectively. The male-specific band of 42 KDa remained in the insoluble phase. Antigenic differences between male and females protins were detected by Western vlotting using a sera from infected Nectomys squamipes.

Identification of schistosome-infected snails by detecting schistosomal antigens and DNA sequences

Cercarial shedding tests do not provide species identification of the shistosomes concerned and cannot detect prepatent schistosomal infections. We have demonstrated that both immunodetection by ELISA of schistosomal antigens in snail hemophlymph, and dot hybridization of snail extracts by DNA probe representing highly repeated sequences, proved suitable for detecting infected snails during prepatnecy as well as patency. A group-specific monoclonal antibody was found to be suitable for detecting Schistosoma mansoni infection in Biomphalaria sp., but not for positive identification of S. haematobium in Blulinus sp. Comparative evaluation of the diagnostic qualities, and technical aspects and cost of these tests, point to the superiority of the immunodetection approach for large scale detection of snails prepatently infected with S. mansoni. This approach is potentially useful for providing extended information on schistosome-snail epidemiology that may facilitate rapid evaluation of the danger of post-control reinfection, and help make decisions on the time and place of supplementary control measures. In this context the potential usefulness of the immunodetection or DNA probing approach for facilitating catalytic model representation of schistosome-snail epidemiology warrants further evaluation. Specific identification of S. haematobium in Bulinus by either of these approaches may be possible depending on the development of suitable antibodies or DNA probes.

A programme for computer aided identification of Phlebotomine sandflies of the Americas (CIPA): presentation and check-list of American species

The CIPA programme is a collaborative project including two entomologists from France and seven South and Central America countries. Its objective is the development of an expert system for computer aided identification of phlebotomine sandflies from the Americas. It also includes the formation of data bases for bibliographic, taxonomic and biogeographic data. Participant consensus on taxonomic prerequisites, standardization in bibliographic data collections and selection of descriptive variables for the final programme has been established through continous communication among participants and annual meetings. The adopted check-list of American sandflies presented here includes 386 specific taxa, ordered into genera and 28 sub-genera or species groups.

Sequencing and identification of expressed Schistosoma mansoni genes by random selection of cDNA clones from a directional library

We have initiated a gene discovery program in Schistosoma mansoni based on the technique of Expressed Sequence Tags (ESTs), i.e. partial sequences of cDNAs obtained from single passes in automatic DNA sequencers. ESTs can be used to identify genese onf the basis of their homology whith sequences from other species deposited in DNA or protein databases. Trasncripts with sequences without matches in teh databases may represent novel parasite-specific genes. This approach has shown to be very efficient and in less than two years a broad range of novel genes has already been ascertained, more than doubling the number of known S. mansoni genes.

Use of isozyme patterns in the identification of Biomphalaria tenagophila (D'Orbigny, 1835) and B. occidentalis (Paraense, 1981) (Gastropoda: Planorbidae)

Two sibling species of Biomphalaria, B. tenagophila and B. occidentalis were identified using isozyme patterns obtained by horizontal gel electrophoresis. Six diagnostic enzymatic loci were identified in digestive gland homogenates. The results enable us to distinguish the species, calculate the Nei's coefficient of genetic similarity, and provide a basis for making inferences about the pattern of these two planorbid species colonization and distribution.

The identification of Anopheles (Nyssorhynchus) rondoni (Diptera: Culicidae) in Mato Grosso, Brazil: an analysis of key character variability

A morphological study was made of a population of Anopheles (Nyssorhynchus) rondoni (Neiva and Pinto) from northern Mato Grosso, Brazil. This population usually lacked the primary key character of a dark basal band on hindtarsomere 3, i.e., hindtarsomere 3 was all white as in most other members of the subgenus. It was determined that this species can be recognized instead by the presence of a dark spot on the thorax made up of a large dark prescutellar space that is contiguous with a concolorous central area on the scutellum. A secondary character of a dark area on the costa created by the fusion of the humeral dark, presector dark and sector dark proximal spots is also usually reliable. Regression analyses comparing the lengths and ratios of the dark bands on hindtarsomeres 2 to those on 3 describe a straight line relationship. This suggests that the "atypical" population is at one end of a character gradient. We propose that in the subgenus Nyssorhynchus individuals that have a long basal band on hindtarsomere 2 are more likely to also have a basal band on hindtarsomere 3. The pupal stage of this species has not been previously described. Reared-associated specimens from this study show that the pupa can be easily differentiated from all other Nyssorhynchus by the relatively stout, usually 2 or 3 branched (1-5), setae 1 and 5 on segments IV-VII.

Identification of species related to Anopheles (Nyssorhynchus) albitarsis by random amplified polymorphic DNA-Polymerase chain reaction (Diptera: Culicidae)

Species-specific Random Amplified Polymorphic DNA-Polymerase chain Reaction (RAPD-PCR) markers were used to identify four species related to Anopheles (Nyssorhynchus) albitarsis Lynch-Arribàlzaga from 12 sites in Brazil and 4 in Venezuela. In a previous study (Wilkerson et al. 1995), which included sites in Paraguay and Argentina, these four species were designated "A", "B", "C" and "D". It was hypothesized that species A is An. (Nys.) albitarsis, species B is undescribed, species C is An. (Nys) marajoara Galvão and Damasceno and species D is An. (Nys.) deaneorum Rosa-Freitas. Species D, previously characterized by RAPD-PCR from a small sample from northern Argentina and southern Brazil, is reported here from the type locality of An. (Nys.) deaneorum, Guajará-Mirim, state of Rondônia, Brazil. Species C and D were found by RAPD-PCR to be sympatric at Costa Marques, state of Rondônia, Brazil. Species A and C have yet to be encountered at the same locality. The RAPD markers for species C were found to be conserved over 4,620 km; from Iguape, state of São Paulo, Brazil to rio Socuavo, state of Zulia, Venezuela. RAPD-PCR was determined to be an effective means for the identification of unknown species within this species complex.

Monoclonal antibodies for the identification of New World Leishmania species

Monoclonal antibodies specific for selected species complexes of Leishmania have been employed for the characterization of several representative strains of Leishmania isolated from different hosts and localities in the Americas. In the past 15 years, data have been accumulated concerning (i) the specificities of a number of these monoclonal antibodies and (ii) the antigenic variation (level of the expressed antigenic determinants) occurring among New World Leishmania species or strain variants as recognized by the monoclonal antibodies. This report is an attempt to summarize in brief the data accumulated to date on these points and to indicate the directions for future applications of these specific monoclonal antibodies for identification of leishmanial isolates.

A low stringency polymerase chain reaction approach to the identification of Biomphalaria glabrata and B. tenagophila, intermediate snail hosts of Schistosoma mansoni in Brazil

The low stringency-polymerase chain reaction (LS-PCR) with a pair of specific primers for the amplification of the 18S rRNA gene was evaluated as a means of differentiating between the two Schistosoma mansoni intermediate host species in Brazil: Biomphalaria glabrata and B. tenagophila. Individual snails obtained from different states of Brazil were used and the amplification patterns obtained showed a high degree of genetic variability in these species. Nevertheless, 4 and 3 clearly defined specific diagnostic bands was observed in individuals from B. glabrata and B. tenagophila respectively. The detection of snail specific diagnostic bands suggests the possibility of reliable species differentiation at the DNA level using LS-PCR.

Specific Identification of Biomphalaria tenagophila and Biomphalaria occidentalis Populations by the Low Stringency Polymerase Chain Reaction

Although Biomphalaria occidentalis and B. tenagophila are indistinguishable on the basis of shell morphology and the majority of their genital organs, only the latter is susceptible to infection with Schistosoma mansoni. Thus, the identification of these species is fundamental to epidemiological studies of schistosomiasis. Here we describe a simple and rapid method for differentiating B. tenagophila from B. occidentalis based on low stringency polymerase chain reaction and using a pair of primers specific for the amplification of the 18S rRNA gene. Analysis of the low stringency product profiles of populations of these snails from different geographical regions confirmed this approach as being applicable to the identification of B. tenagophila and B. occidentalis in cases where classical morphology is inconclusive

Comparison of Some Molecular-genetic Techniques for Identification of Leishmania Circulating in Natural Foci of Zoonotic Cutaneous Leishmaniasis in the Central Asia Region

Different molecular-genetic methods were used to identify a cohort of Leishmania strains from natural foci of zoonotic cutaneous leishmaniasis located in Central Asia, on the former USSR territory. The results obtained using isoenzymes, PCR, restriction fragment length polymorphisms of kDNA and molecular hybridization techniques are discussed in terms of their applicability, discrimination power and feasibility for answering questions related to molecular epidemiological research and for detecting mixed Leishmania infections