The serum resistance-associated gene as a diagnostic tool for the detection of Trypanosoma brucei rhodesiense.

Abstract

Abstract. In the search for new diagnostic methods that would distinguish Trypanosoma brucei rhodesiense from T. b. brucei and T. b. gambiense, we have developed two polymerase chain reaction (PCR) primer sets. The first primer set was derived from the serum resistance−associated (SRA) gene of T. b. rhodesiense that confers resistance to lysis by normal human serum (NHS). The specificity of the SRA-based PCR was tested on 97 different trypanosome populations originating from various taxonomic groups, host species, and geographic regions. Only one of 25 T. b. rhodesiense samples was negative in this PCR, and none of 72 other samples were positive in this assay. Interestingly, a reference T. brucei strain (TREU927/4) currently used for genome sequencing was negative for the SRA gene; however, this strain was resistant to lysis by NHS. The second primer set was derived from a specific variant surface glycoprotein (VSG) expression site where the SRA gene is expressed (R-ES). This primer set identified the strain as T. b. rhodesiense in 17 of 17 SRA gene-positive strains in which it was tested. These data strongly suggest that expression of the SRA gene is generally involved in resistance to lysis by NHS in T. b. rhodesiense strains.

Human infectivity trait in Trypanosoma brucei: stability, heritability and relationship to sra expression

“…This PCR test is based on the assumption that human serum resistant trypanosomes have the sra gene and non-human infective trypanosomes do not which appears to hold true for the majority of isolates tested from east and southern Africa Welburn et al 2001;Agbo et al 2003), but not for west and central Africa (Berberof et al 2001;Radwanska et al 2002).…”

“…However, while it is clear that the sra gene is responsible for human infectivity in a large number of T. b. rhodesiense isolates Welburn et al 2001), there are exceptions to this in trypanosome lines from East Africa (Rifkin et al 1994;Agbo et al 2003) and, critically, this gene is not present in West African T. b. gambiense isolates (De Greef and Hamers 1994;Radwanska et al 2002). Thus there must be alternative mechanisms determining human infectivity/human serum resistance other than those based on the sra gene.…”

Comparative Detection of Trypanosomal DNA by Loop-Mediated Isothermal Amplification and PCR from Flinders Technology Associates Cards Spotted with Patient Blood

“…These are PCR based, targeting the serum resistanceassociated (SRA) gene that is specific for T. b. rhodesiense (4,6,14) or the T. b. gambiense surface glycoprotein (TgSGP) that is specific for that subspecies (2,15). The need for specialized equipment and a constant supply of electricity has hindered integration of those techniques into the diagnostic algorithm.…”

In Vitro Generation of Human High-Density-Lipoprotein-Resistant Trypanosoma brucei brucei

“…Although compelling evidence supports the role of SRA in human infectivity by T. b. rhodesiense, several investigators have reported the occurrence of human serum-resistant lines of African trypanosomes that lack the SRA gene (2,47,66,67). To investigate the mechanism of SRA-independent resistance, we generated human serum-resistant lines of T. b. brucei 427 (MiTat 1.2) by repeated in vitro exposure to human HDLs.…”

“…Several experiments have shown a correlation between the expression of the SRA gene and resistance to human serum (11,35), and transfection of this gene into T. b. brucei conferred resistance to human serum and TLF (41,74). In addition, the SRA gene has proven to be a reliable marker for T. b. rhodesiense in epidemiological studies throughout East Africa (16,18,39,47,72,73).…”