Comparison of the diagnosis of malaria by microscopy, immunochromatography and PCR in endemic areas of Venezuela

Whole blood samples (N = 295) were obtained from different locations in Amazonas and Sucre States, in Venezuela. Malaria was diagnosed by microscopy, OptiMAL™ and polymerase chain reaction (PCR), with Plasmodium vivax, P. falciparum, and P. malariae being detected when possible. We identified 93 infections, 66 of which were caused by P. vivax, 26 by P. falciparum, and 1 was a mixed infection. No infection caused by P. malariae was detected. The sensitivity and specificity of each diagnostic method were high: 95.7 and 97.9 percent for microscopy, 87.0 and 97.9 percent for OptiMAL, and 98.0 and 100 percent for PCR, respectively. Most samples (72.2 percent) showed more than 5000 parasites/æL blood. The sensitivity of the diagnosis by microscopy and OptiMAL decreased with lower parasitemia. All samples showing disagreement among the methods were reevaluated, but the first result was used for the calculations. Parasites were detected in the 6 false-negative samples by microscopy after the second examination. The mixed infection was only detected by PCR, while the other methods diagnosed it as P. falciparum (microscopy) or P. vivax (OptiMAL) infection. Most of the false results obtained with the OptiMAL strip were related to the P. falciparum-specific band, including 3 species misdiagnoses, which could be related to the test itself or to genetic variation of the Venezuelan strains. The use of the microscopic method for malaria detection is recommended for its low cost but is very difficult to implement in large scale, population-based studies; thus, we report here more efficient methods suitable for this purpose.

A methodological approach for the construction of a radiation hybrid map of bovine chromosome 5

A bovine 5,000 rad WG-RH panel was used to construct an RH map of bovine chromosome 5 (BTA5). Twenty-one microsatellites and thirteen genes were scored in the panel using PAGE and radioactive labeling. Marker retention ranged from 8.9 percent-25.8 percent and averaged 17.8 percent. Pairwise locus analysis placed all markers in a single syntenic group with a LOD support of 4.0. At a LOD support of 8.0, a centromeric group of 23 syntenic markers was formed. Telomeric groups of 11 and 9 markers were assembled with a LOD support of 6.0 and 8.0, respectively. All markers were ordered by maximum likelihood methods using the program RHMAP. Only 13 markers were ordered with a LOD support of at least 3.0, while 25 and 29 markers were ordered with a support of at least 2.0 and 1.0, respectively. Total length of the comprehensive RH map was 435.9 cR5,000, with an average marker separation of 12.8 cR5,000. The largest gaps in the map were 55.0 and 30.4 cR5,000 in length. The locus orders of markers common to both the RH map and the USDA-MARC linkage map were identical. The relationship between the RH and linkage maps was calculated to be 3.74 cR5,000/cM.

Molecular cytogenetic assignment of genes to bovine chromosome 5

Seven genes were assigned by molecular cytogenetic methods to bovine chromosome 5. To accomplish this, specific primers were either publicly available or were designed from highly conserved regions of the publicly available mammalian gene sequences. The identity of the amplified segments was verified by sequencing and alignment with the published sequences. The optimized primers that amplified the desired bovine genes were used for screening a bovine bacterial artificial chromosome library. The positive clones were localized to a specific band of bovine chromosome 5 by fluorescence in situ hybridization. The genes HOXC4, SP1 and IGFBP6 were localized to band q21, COL2A1 was localized to bands q21-q23, IGF1 was localized to band q26, MB to band q31 and the gene CYP2D6 was localized to band q35. The cytogenetic assignment of SP1, IGFBP6, COL2A1, IGF1, MB and CYP2D6 is first reported here and the assignment of HOXC4 refines the previous assignment of this gene. The identification and localization of these genes further support the development of the human to bovine comparative map through characterizing the homologous segments conserved in the evolution of these species. This information will be useful for the future localization of genes that affect economically important traits in bovines