Biomedical subjects

Gerard J Schoone

Publications and source records attributed to Gerard J Schoone.

4 recordsLinked to original sources

PubMed2006

Detection and identification of human Plasmodium species with real-time quantitative nucleic acid sequence-based amplification.

BACKGROUND: Decisions concerning malaria treatment depend on species identification causing disease. Microscopy is most frequently used, but at low parasitaemia (<20 parasites/mul) the technique becomes less sensitive and time consuming. Rapid diagnostic tests based on Plasmodium antigen detection do often not allow for species discrimination as microscopy does, but also become insensitive at <100 parasites/microl. METHODS: This paper reports the development of a sensitive and specific real-time Quantitative Nucleic Acid Sequence Based Amplification (real-time QT-NASBA) assays, based on the small-subunit 18S rRNA gene, to identify the four human Plasmodium species. RESULTS: The lower detection limit of the assay is 100-1000 molecules in vitro RNA for all species, which corresponds to 0.01-0.1 parasite per diagnostic sample (i.e. 50 microl of processed blood). The real-time QT-NASBA was further evaluated using 79 clinical samples from malaria patients: i.e. 11 Plasmodium. falciparum, 37 Plasmodium vivax, seven Plasmodium malariae, four Plasmodium ovale and 20 mixed infections. The initial diagnosis of 69 out of the 79 samples was confirmed with the developed real-time QT-NASBA. Re-analysis of seven available original slides resolved five mismatches. Three of those were initially identified as P. malariae mono-infection, but after re-reading the slides P. falciparum was found, confirming the real-time QT-NASBA result. The other two slides were of poor quality not allowing true species identification. The remaining five discordant results could not be explained by microscopy, but may be due to extreme low numbers of parasites present in the samples. In addition, 12 Plasmodium berghei isolates from mice and 20 blood samples from healthy donors did not show any reaction in the assay. CONCLUSION: Real-time QT-NASBA is a very sensitive and specific technique with a detection limit of 0.1 Plasmodium parasite per diagnostic sample (50 microl of blood) and can be used for the detection, identification and quantitative measurement of low parasitaemia of Plasmodium species, thus making it an effective tool for diagnostic purposes and useful for epidemiological and drug studies.

Animals↗

PubMed2005

Application of direct agglutination test (DAT) and fast agglutination screening test (FAST) for sero-diagnosis of visceral leishmaniasis in endemic area of Minas Gerais, Brazil.

BACKGROUND: The direct agglutination test (DAT) has proved to be a very important sero-diagnostic tool combining high levels of intrinsic validity and ease of performance. Otherwise, fast agglutination screening test (FAST) utilises only one serum dilution making the test very suitable for the screening of large populations. RESULTS: We have tested FAST and DAT for the detection anti-Leishmania antibodies in serum samples from patients with American visceral (AVL) and cutaneous leishmaniases (ACL) in Minas Gerais State, Brazil. The DAT on serum and blood samples of confirmed AVL patients found all samples positive at a serum dilution of > or = 1:800. This dilution was subsequently used as cut off value in the study. The blood and serum samples of these confirmed patients could also be clearly read in FAST using a 1:100 dilution with the same high sensitivity. DAT and FAST were not able to detect significant amounts of antibodies in samples from ACL patients and are not suitable for the diagnosis of this manifestation of the disease. CONCLUSION: We suggest that both DAT and FAST are very practical diagnostic tools for the sero-diagnosis of AVL under rural conditions as both serological tests do not require sophisticated equipment, a cold chain and are very simple to perform.

Journal Article↗

PubMed2005

Quantitative nucleic acid sequence-based assay as a new molecular tool for detection and quantification of Leishmania parasites in skin biopsy samples.

Currently available methods for the diagnosis of cutaneous leishmaniasis (CL) have low sensitivities or are unable to quantify the number of viable parasites. This constitutes a major obstacle for the diagnosis of the disease and for the study of the effectiveness of treatment schedules and urges the development of improved detection methods. In this study, quantitative nucleic acid sequence-based amplification (QT-NASBA) technology was used to detect and quantify Leishmania parasites in skin biopsy samples from CL patients. The assay is based on the detection of a small subunit rRNA (18S rRNA), which may allow for the detection of viable parasites. The QT-NASBA assay was evaluated using in vitro-cultured promastigotes and amastigotes and 2-mm skin biopsy samples from Old and New World CL patients. The study demonstrated that the lower detection limit of the QT-NASBA was two parasites per biopsy sample. Parasites could be quantified in a range of 2 to 11,300,000 parasites per biopsy sample. The QT-NASBA could detect levels of parasites 100-fold lower than those detected by conventional PCR. Test evaluation revealed that the QT-NASBA had a sensitivity of 97.5% and a specificity of 100% in the present study. The QT-NASBA is a highly sensitive and specific method that allows quantification of both Old and New World Leishmania parasites in skin biopsy samples and may provide an important tool for diagnosis as well as for monitoring the therapy of CL patients.

Adolescent↗

PubMed2004

Sero-epidemiological study of canine Leishmania spp. infection in the municipality of Alijó (Alto Douro, Portugal).

Visceral leishmaniosis caused by Leishmania infantum is a zoonotic disease in the Mediterranean basin. We report an epidemiological survey carried out in dogs from the municipality of Alijó in the endemic region of Alto Douro (north Portugal). Performance of the direct agglutination test (DAT) was assessed in 205 matching samples of blood collected on filter paper and serum. A high degree of agreement (97.6%; k = 0.83) was found between the results obtained from both types of samples. DAT was then used to test more blood on filter paper (B-FP) samples from other dogs of the same municipality. The detected sero-prevalence was 18.7% (288/1540), with values ranging from 0.0 to 81.1% in each of the 19 parishes of Alijó. Three distinct geographical zones of mean sero-prevalence could be defined: northwestern (2.5%), intermediate (11.4%) and southern (49.9%). No statistically significant difference was observed between male (19.1%) and female (17.8%) sero-prevalences (P = 0.560). Dogs of 9-11 years of age showed the highest sero-prevalence (28.4%), but all the other age-intervals (0-2, 3-5, 6-8 and 12-17 years) presented values (15.0-22.3%) not significantly different from the mean of the whole study population. Risk factors for canine Leishmania infection were age and geographical zone. Only 5.9% of the sero-positive animals had clinical signs of canine leishmaniosis and the overall prevalence of disease was 1.1%. This study validates the use of B-FP samples and confirms DAT as a simple and sensitive serological test to evaluate the level of canine Leishmania infection in areas of high sero-prevalence.

Age Factors↗