Mermithids are the most common parasites of black flies and are associated with host feminization and sterilization in infected hosts. However, information on the species / lineage of black fly mermithids in Southeast Asia, including Malaysia requires further elucidation. In this study, mermithids were obtained from black fly larvae collected from 138 freshwater stream sites across East and West Malaysia. A molecular approach based on nuclear-encoded 18S ribosomal RNA (18S rRNA) gene was used to identify the species identity / lineage of 77 nematodes successfully extracted and sequenced from the specimens collected. Maximum likelihood and neighbor-joining phylogenetic analyses demonstrated five distinct mermithid lineages. Four species delimitation analyses: automated simultaneous analysis phylogenetics (ASAP), maximum likelihood Poisson tree processes with Bayesian inferences (bPTP_ML), generalized mixed yule coalescent (GMYC) and single rate Poisson tree processes (PTP) were applied to delimit the species boundaries of mermithid lineages in this data set along with genetic distance analysis. Data analysis supports five distinct lineages or operational taxonomic units for mermithids in the present study, with two requiring further investigation as they may represent intraspecific variation or closely related taxa. One mermithid lineage was similar to that previously observed in Simulium nigrogilvum from Thailand. Co-infection with two mermithids of different lineages was observed in one larva of Simulium trangense. This study represents an important first step towards exploring other aspects of host - parasite interactions in black fly mermithids.
Sexually anomalous individuals, typically intersexes or gynandromorphs, bear a mixture of male and female traits. Twelve sexually anomalous individuals of the black fly Simulium (Gomphostilbia) trangense Jitklang, Kuvangkadilok, Baimai, Takaoka & Adler were discovered among 49 adults reared from pupae. All 12 sexually anomalous adults were parasitized by mermithid nematodes, although five additional parasitized adults had no overt external anomalies. Sequence analysis of the 18S rRNA gene revealed that the mermithids, possibly representing a new species, are related to Mesomermis spp., with genetic distances of 5.09-6.87%. All 12 anomalous individuals had female phenotypical traits on the head, thorax, forelegs, midlegs, and claws, but male features on the left and right hind basitarsi. One individual had mixed male and female genitalia. The findings are in accord with the trend that mermithid infections are associated with sexually anomalous adult black flies.
Cx. quinquefasciatus is a common nuisance mosquito widely distributed in tropical and subtropical areas. This mosquito is also a vector of urban filariasis. Control with chemicals has been hampered by the development of resistance against chemical insecticides and rising problems of environmental contamination associated with them. Therefore, it is important to adopt more integrated mosquito management approaches that include sustainable, non chemical solutions. The mermithid nematode Romanomermis iyengari is one of several natural control alternatives to synthetic pesticides for mosquito suppression. This study evaluated the effectiveness of the nematode R. iyengari for control of Cx. quinquefasciatus. The nematode R. iyengari was mass-produced, and pre-parasitics (J2) were used for laboratory and field experiments. In laboratory experiments, two concentrations of pre-parasitics (5 and 10 J2 per larva) were tested against L1, L2 and L3 instars larvae of Cx. quinquefasciatus. Infected larvae were observed daily to determine their mortality rate and the number of postparasitic nematodes emerging from dead larvae. In field experiments, 1000, 2000 and 3000 J2/m2 were sprayed in separate natural Cx. quinquefasciatus breeding sites. After treatment, the larval mosquito density in the breeding sites was assessed every 5 days. Laboratory results showed that all tested Cx. quinquefasciatus instars larvae were susceptible to nematode infection. The mortality rates observed for each larval stage indicated that the concentration of 10 J2 kills larvae faster, and that the L1 larvae died earlier than older larvae. The average number of post-parasitic nematodes emerging per larva increases with increasing nematode concentration; also more post-parasitic nematodes emerged from the L2 larvae. Field data showed that, in breeding site treated with 3000 J2 per square meter, larval mosquito reduction reached 97% after nematode application. The dosage of 1000 J2 per square meter did not reduce the larval density. The insect parasitic nematode R. iyengari could be easily used as component of integrated mosquitoes control program in lymphatic filariasis endemic countries.