DNA Barcodes and Species Boundaries of Black Flies (Diptera: Simuliidae)
Black flies play a prominent role in public health and the epidemiology
of parasitic diseases of humans and domesticated and wild animals.
Correct identification is, therefore, vital for identifying species,
understanding their biological attributes, and monitoring pest and
vector species. DNA barcoding is an established molecular tool that
provides rapid and accurate species identification. Our study
strengthens the molecular database for black flies in Malaysia by adding
59 cytochrome c oxidase I sequences for 22 species, of which 14 are
included for the first time. These sequences, combined with those in
public databases, represent a total of 338 sequences for 52 Malaysian
species, nearly 50% of which were collected from type localities. At
the subgeneric level, barcode gap analysis most accurately identified
species in the subgenus Nevermannia (92%), followed by Simulium s. l.
(91%), and Gomphostilbia (81%). The remaining sequences were ambiguous
and could not be distinguished from those of nearest neighbour species
due to an overlap in genetic divergence and low genetic diversity,
especially between insular species. Tree analyses indicate that certain
species had incomplete lineage sorting and low mitochondrial signals.
Possible cryptic species were indicated in the Simulium (Gomphostilbia)
batoense and S. (G.) epistum species groups. Species delimitations were
consistent with morphological identifications except in large species
groups such as the S. (G.) asakoae, S. (G.) batoense, S. (G.) epistum,
and S. (Simulium) melanopus groups. The use of type specimens or
specimens collected from type localities (topotypes) in barcoding is
strongly recommended for reference sequences to increase the reliability
of the molecular database.