Quantification of Plasmodium malariae infection in mosquito vectors.

Plasmodium malariae occurs in various tropical regions throughout the world and causes low, yet significant, levels of morbidity in human populations. One means of studying the ecology and frequency of this parasite is by measuring sporozoite loads in the salivary glands of infected mosquitoes. An effective, species-specific test that can be used to detect the presence of sporozoites in mosquitoes is the circumsporozoite ELISA. The aim of the present study was to standardize the circumsporozoite ELISA for P.malariae, by setting quantification parameters using, as antigen, either a synthetic peptide or extracts of whole sporozoites. The standard quantification curves produced indicated that the assay had a lower threshold of sensitivity of 250 sporozoites in a 50-microl sample, equivalent to about 1250 sporozoites in a mosquito.

Quantitative determination of sporozoites and circumsporozoite antigen in mosquitoes infected with Plasmodium falciparum or P. vivax.

It is essential for malariologists and researchers to have simple and accurate means of assessing the threat of Plasmodium parasites. An attempt was therefore made to re-standardize one of the circumsporozoite (CS) ELISA that can be used to detect and quantify the circumsporozoite antigens of P. falciparum and P. vivax. A two-site, 'sandwich' ELISA based on a monoclonal antibody was used to test for the CS antigen and sporozoites of each Plasmodium species simultaneously. Using the resultant optical-density values, standard curves, that permit the number of sporozoites in an infected mosquito to be estimated from the quantification of the CS antigen, were constructed. Using these plots and the CS ELISA, the presence of just 12.5 sporozoites (i.e. 0.8 pg CS antigen) of P. falciparum, four sporozoites (3.2 pg antigen) of P. vivax-210 or 12.5 sporozoites (32.0 pg antigen) of P. vivax-247 could be demonstrated.

Real-time polymerase chain reaction diagnosis of leishmaniasis in Panama from both fresh and frozen tissue.

Skin biopsies stored in ethanol from 49 patients with suspected cutaneous leishmaniasis (CL) were tested in a real-time polymerase chain reaction (PCR) assay and compared with conventional diagnostic methods. With clinical diagnosis as the gold standard, PCR had a sensitivity of 96% (47/49) vs. 61% (30/49) for histopathology and 33% (16/49) for culture. In addition, DNA was extracted from 70 frozen smears of lesions from suspected cases of CL and tested with the same assay. In these samples, the PCR had a sensitivity of 61% (43/70) vs. 56% (39/70) for histopathology and 41% (29/70) for culture. In this study, real-time PCR offered a rapid diagnosis with an enhanced sensitivity over conventional methods. Although the yield of PCR diagnosis was lower when testing frozen smears, the assay still outperformed existing diagnostic modalities.