title: Comparisons of methods for estimating parasitaemia in trypanosomiasis
Creators: name: Singh-Kelley, Satpal K.
Publisher: name: The University of Edinburgh
Year: 1981
Subjects: subject: Annexe MSc Digitisation Project 2022 Block 47
subject: Trypanosomiasis
subject: Parasitaemia
subject: Haemocytometer
subject: Thin Smear
subject: DEAE/cytospin technique

description: "Matching", Haemocytometer and. Thin Smear techniques were compared, to quantify infections with T. congolense and. T. brucei in mice. Preliminary investigations on blood, with high T. congolense infections (containing approximately 32 trypanosomes per wet blood, film) showed, that all three techniques gave consistently reproducible results with replicate samples of blood taken on a single occasion. Estimations of the numbers of trypanosomes in mice infected with T. congolense at concentrations of organisms less than 2-3 trypanosomes per wet blood film, were statistically similar. However, there were significant differences between the "Matching" technique and Haemocytometer and Thin Smears (p &lt; 0.05) at para-sitaemia levels of 9-15 organisms per field. T. congolense infections also showed significant differences between tail and heart blood numbers (p &lt; 0.05) and the significance of these differences was greater at higher levels of parasitaemias (9-15 organisms per field; p &lt; 0.01). Comparable results were produced between the "Matching" technique and Haemocytometer and Thin smears, when T. brucei infections were quantified at infection levels of 2-3 organisms per wet blood film; however results from Haemocytometer and Thin smears differed significantly (p &lt; 0.05). Estimations of infections with higher numbers of organisms (9-15 trypanosomes per wet blood film) produced results that were statistically in agreement. Numbers of trypanosomes in tail blood when compared to those in heart blood were significantly different (p &lt; 0.05) when counts were made on Thin smears. Counts from peripheral blood were consistently lower than those from heart blood with T. brucei infections. Diluents for counting T. congolense in a haemocytometer were compared, and it was found that 1% glutaraldehyde and 1% methylene blue produced significantly lower counts than 0.9% NaCl or carbol fuschin (p &lt; 0.05). However, there were no statistical differences in results between methylene blue diluent and gluter-aldehyde and carbol fuschin diluent and 0.9% NaCl. A new technique, the DEAE/cytospin was developed for quantifying low chronic infections of T. congolense in rabbits and T. evansi in sheep. Trypanosomes were first eluted in mini-columns (Lumsden, Kimber and Strange, 1977) and the eluate further concentrated by cytospinning onto glass slides, which were then stained in 10% Giemsa. This technique was compared to the Measured Volume Smear and it was found that a large percent of trypanosomes were lost during the process, therefore producing highly significant differences in counts (for T. evansi infections) between the two techniques (p &lt; 0.001). When similar techniques were applied to chronic T. congolense infections in rabbits, results obtained were too variable to be analysed statistically.