Sibling species in the Llutzomyia longipalpis complex differ in levels of mRNA expression for the salivary peptide, maxadilan.

Maxadilan is a small ( approximately 7 kDa) protein found in the saliva of sand fly species in the Lutzomyia longipalpis complex, vectors of the parasite causing visceral leishmaniasis, Leishmania chagasi. It is a potent vasodilator and also has immunomodulatory affects. Maxadilan recovered from different sibling species of the Lu. longipalpis complex differ in amino acid content by as much as 23%, however all variants possess equivalent vasodilatory activity. Therefore, the dramatic differences in vasodilatory activity of the saliva from different sibling species is probably due to differences in the amounts of maxadilan in their saliva. This is significant because it has been suggested that maxadilan may influence the pathogenesis of leishmanial infections. In this study we measured the amount of maxadilan messenger RNA (mRNA) per pair of salivary glands from individual sand flies by quantitative reverse transcription polymerase chain reaction (RT-PCR) using a competitive method. We report a method using the gene of interest, in this case maxadilan, amplified by the PCR from genomic DNA, as a competitor in the quantitative RT-PCR, taking advantage of differences in the size of these products due to the presence of an intron. Significant differences in amounts of maxadilan mRNA among colonies from Central and South America are described. We found a strong correlation between the amount of maxadilan mRNA detected in salivary glands of different Lu. longipalpis sibling species and previously described differences in the size of erythemas produced by the bite of these species. Therefore, variation in the amount of mRNA suggests that differences in the vasodilatory properties of saliva among the different sibling species are the result of differences in the amount of maxadilan present in the saliva and not differences in the potency of maxadilan peptide variants. The geographical distribution of species with high or low levels of maxadilan gene expression are concordant with the distribution of atypical cutaneous leishmaniasis resulting from infection with Le. chagasi, lending credence to earlier suggestions that maxadilan may be involved with visceralization of this parasite.