AbstractIn the Esmeraldas focus of onchocerciasis in Ecuador, the larvae of the primary vector Simulium exiguum Roubaud are found in sympatry with the non-vector S. gonzalezi Vargas & Díaz Nájera. Distinguishing the pre-ultimate stage larvae of these two species usually relies on the pattern of markings on the cephalic apotome. These patterns appear either as dark spots against a pale cuticle (positive head pattern) or as pale spots within a darkened area of the cuticle (negative head pattern). Larvae of S. exiguum s.l. usually exhibit positive or no head patterns whilst those of S. gonzalezi usually have negative head patterns. The purpose of this paper was to investigate head pattern variation (scored in one of six categories) in (polytene) chromosomally identified larvae of the two species and cytotypes of S. exiguum from three collection sites in Ecuador (including one in the onchocerciasis focus). The head patterns of a total of 1183 larvae were studied. Chromosomal identification of penultimate instar larvae revealed only the presence of the Cayapa and Quevedo cytotypes of S. exiguum (n=208). Both cytotypes exhibited a wide range of head patterns from positive to negative. The results indicate that negative head patterns cannot be used to reliably distinguish between larvae of S. exiguum s.l. and S. gonzalezi. All except one larva with negative head patterns were females (n=53), suggesting that head pattern in this species complex might be a sex-linked character.