Scanning electron micrograph of feces and eggs of <i>Oncopeltus fasciatus</i> infected with <i>Leptomonas wallacei.</i>

<p>(A) Promastigote forms of <i>L. wallacei</i> in fresh feces showing typical characteristics of live cells and cystic forms (arrow). Bar = 5 µm. (B) High magnification of a promastigote form showing one cystic form (arrow) near to its flagella. Bar = 1 µm. (C) Promastigote form present in the naturally dried feces showing extensive membrane damage and three cystic forms. Note that the cystic forms (arrows) do not show any surface damage. Bar = 1 µm. (D) Low magnification of the surface of the eggs. Bar = 500 µm. (E) High magnification of egg surface showing typical cystic forms of <i>L. wallacei</i> (arrows). Bar = 1 µm.</p

Detection of parasites after experimental transovum transmission of <i>Leptomonas wallacei</i> parasites by <i>Oncopeltus fasciatus</i>.

<p>(A) Detection of parasite infection by PCR. Lane 1- The DNA extracted from an axenic culture of <i>L. wallacei</i> was amplified with primers specific for parasite detection. Lane 2- A pool of DNA samples extracted from gut of parasite-free insects was concomitantly amplified with primers specific for parasite and insect DNA detection. Lanes 3–7- Representative DNA samples extracted from guts of insects that fed on sunflower seeds contaminated with eggshels collected from infected colony were concomitantly amplified with primers specific for parasite and insect DNA detection. On the left, the positions of molecular size markers are shown in base pairs. The figure represents a negative image of the gel. (B) Detection of parasite infection by optical microscopy. The representative micrograph shows Giemsa-stained parasites (arrows) in the gut contents of newly infected insects. The arrowheads indicate the nucleus (N) and kinetoplast (K) of the parasite. Magnification = 400 x.</p

Scanning electron microscopy of <i>Oncopeltus fasciatus</i> guts infected with <i>Leptomonas wallacei</i>.

<p>(A) Midgut of <i>L. wallacei</i>-infected <i>O. fasciatus</i>. The arrows indicate large numbers of parasites near the midgut wall. (B) Hindgut of <i>L. wallacei</i>-infected <i>O. fasciatus</i>. The image shows a massive presence of flagellates. Most of those are attached to the intestinal wall of the hindgut by their flagella, so only their slender bodies can be seen. One of the short-sized flagellates can be seen in the lumen (arrow). Bars = 10 µm.</p

<i>Oncopeltus fasciatus</i> hatched from eggs submitted to surface asepsis were <i>Leptomonas wallacei</i>-free.

<p>(A) Representative gel electrophoresis of PCR-amplified DNA samples extracted from eggs and whole insect guts. Lane 1- The DNA extracted from an axenic culture of <i>L. wallacei</i> was amplified with primers specific for parasite detection. Lane 2- Sample of pooled DNA extracted from eggs collected at the infected colony and submitted to surface asepsis was concomitantly amplified with primers specific for parasite or insect DNA detection. Lanes 3 and 4- Sample of pooled DNA, extracted from 3 pools of five insect guts of insects hatched from eggs submitted to asepsis, was PCR-amplified with primers specific for parasite (lane 3) or insect DNA (lane 4) detection, respectively. On the left, the positions of molecular size markers are shown in base pairs. The figure represents a negative image of the gel. (B and C) Scanning electron microscopy of <i>O. fasciatus</i> midgut and hindgut, respectively. The micrographs show the presence of bacteria (arrows) but absence of parasites. Bars = 10 µm.</p

Horizontal and vertical transmission of <i>Leptomonas wallacei</i> by <i>Oncopeltus fasciatus</i>.

<p>The insects get infected by feeding on feces contaminated with parasites. <i>L. wallacei</i> produces cystic forms, which are present in the insect guts; the feces contaminate the eggshells. Newly hatched nymphs feed on egg remnants and acquire infection. The cartoon represents a model of transmission based upon previous data and the results shown here.</p

Infection rates of insects hatched from eggs laid by <i>Leptomonas wallacei</i>-infected <i>Oncopeltus fasciatus</i> females.

<p>*Groups 1–3 are biological replicates.</p><p>Infection rates of insects hatched from eggs laid by <i>Leptomonas wallacei</i>-infected <i>Oncopeltus fasciatus</i> females.</p