Wild gorillas as a potential reservoir of Leishmania major

Abstract

Vector-borne parasites of the genus Leishmania are responsible for severe human diseases. Cutaneous leishmaniasis, a common form of the disease, is most often caused by the transmission of Leishmania major to humans by female phlebotomine sand flies. Apes are increasingly being seen as a source of zoonotic diseases, including malaria and rickettsiosis. To examine whether gorillas harbor Leishmania species, we screened fecal samples from wild western lowland gorillas (Gorilla gorilla gorilla) in Cameroon for the presence of these pathogens. Of 91 wild gorilla fecal samples, 12 contained Leishmania parasites, and 4 contained phlebotomine sand fly vectors. The molecular identity was determined by running 3 different polymerase chain reaction tests for detection of L. major. Next, fluorescence in situ hybridization was performed to visualize L. major parasites in fecal samples from the gorillas. Both promastigote and amastigote forms of the parasite were found. This work strongly suggests that wild gorillas carry pathogenic Leishmania parasites.

Keywords: FISH; Leishmania major; PCR; detection; feces; gorilla.

Figure 1.

Overview of the analysis of the 91 gorilla stool samples tested for the presence of Leishmania species by real-time polymerase chain reaction (PCR); sequencing of 18S ribosomal RNA, internal transcribed spacer (ITS), and Cytb PCR products; fluorescence in situ hybridization; and real-time PCR to detect the sand fly vector. ^aSamples collected from the same gorilla; ^bNo reliable results were obtained from the microsatellite analyses; ^cMicrosatellite analyses were not performed in these samples. Abbreviation: rDNA, ribosomal DNA.

Figure 2.

Phylogeny of the Leishmania species sequences amplified from wild western lowland gorillas stool samples, based on the 18S ribosomal RNA (rRNA) gene (500 bp; A), the internal transcribed spacer (ITS) region (430 bp; B), and Cytb (820 bp; C).

Figure 3.

Confocal microscopy images of stool samples stained with the FAM-labeled 18S ribosomal RNA Leishmania-specific probe and the nuclear dye DAPI. A, Images of axenic Leishmania major promastigotes double stained by fluorescence in situ hybridization. B–E, Images of stool sample number 23 (40× original magnification [B, D] and 100× original magnification [C, E]), showing the presence of L. major promastigote parasites (B, C) and L. major amastigote forms (D, E). F, Images of stool sample number 31 (Leishmania-free sample; 100× original magnification).