Oviposition Activity of Mansonia Species in Areas Along the Madeira River, Brazilian Amazon Rainforest.

Mansonia are aggressive mosquito species that are abundant in aquatic ecosystems where the macrophyte plants occur. These mosquitoes are commonly found across the Amazon/Solimões River basin. However, little is known about the oviposition behavior of these species. In the present study, we registered observations on the oviposition activity of 3 species: Mansonia amazonensis, Ma. humeralis, and Ma. cf. titillans, in 5 species of macrophytes in the vicinities of the Madeira River, Porto Velho, State of Rondônia, Brazil. Overall, 197 egg batches were collected. A greater amount of egg batches was found in Salvinia molesta as compared with other macrophytes sampled. In addition, 2 new oviposition habitats were noted in Ludwigia helmintorrhiza and Limnobium spongia. These findings will be important to understand the reproductive dynamics of these mosquitoes in the Brazilian Amazon basin.

Efficient Monitoring of Adult and Immature Mosquitoes Through Metabarcoding of Bulk Samples: A Case Study for Non-Model Culicids With Unique Ecologies.

The rapid and economical monitoring of mosquitos is imperative to understanding the dynamics of both disease vectors and nuisance species. In light of technological advances in mosquito sampling and DNA sequencing, health agencies can now utilize the full potential of metabarcoding pipelines for rapid and standardizable surveillance. Here, we describe mosquito spatial and temporal variation, with particular focus on Mansonia Blanchard species, in the Madeira (Rondônia State) and the Ribeira (São Paulo) watersheds, Brazil using metabarcoding of the D2 rDNA marker. Sampling and molecular pipelines were used to evaluate the taxonomic contribution of mosquitos in pools of culicids collected en masse from macrophyte-roots (immatures) and from Mosquito Magnet traps and protected human landings (adults). Results for adult captures are comparable to morphological diagnoses and clarify previously unknown temporal and spatial species turnover. Metabarcoding of immature stages also confirmed the extent of the geographical distribution of some species and each taxon's association with macrophyte species. Given the benefits of metabarcoding, such as taxonomic acuity, high throughput processing, and objectivity, we suggest such techniques should be more fully incorporated into culicid monitoring schemes. The metabarcoding protocol described herein paired with standardized field sampling schemes, when used by mosquito monitoring professionals, offers substantial improvements in terms of practicality, speed and cost.

Culicidae-centric metabarcoding through targeted use of D2 ribosomal DNA primers.

A practical limitation to many metabarcoding initiatives is that sampling methods tend to collect many non-target taxa, which become "amplicon noise" that can saturate Next Generation Sequencing results and lead to both financial and resource inefficiencies. An available molecular tool that can significantly decrease these non-target amplicons and decrease the need for pre-DNA-extraction sorting of bycatch is the design of PCR primers tailored to the taxa under investigation. We assessed whether the D2 extension segment of the 28S ribosomal operon can limit this shortcoming within the context of mosquito (Culicidae) monitoring. We designed PCR primers that are fully conserved across mosquitos and exclude from amplification most other taxa likely to be collected with current sampling apparatuses. We show that, given enough sequencing depth, D2 is an effective marker for the detection of mosquito sequences within mock genomic DNA pools. As few as 3,050 quality-filtered Illumina reads were able to recover all 17 species in a bulk pool containing as little as 0.2% of constituent DNA from single taxa. We also mixed these mosquito DNA pools with high concentrations of non-Culicidae bycatch DNA and show that the component mosquito species are generally still recoverable and faithful to their original relative frequencies. Finally, we show that there is little loss of fidelity in abundance parameters when pools from degraded DNA samples were sequenced using the D2 primers.

Trichophoromyia iorlandobaratai (Diptera: Psychodidae), a new phlebotomine species from the Brazilian Amazonia.

Trichophoromyia iorlandobaratai sp. n. Vasconcelos dos Santos, Santos Neto, Sánchez Uzcátegui & Galardo is described and illustrated on the basis of male specimens collected in the municipality of Itaituba, Pará State, Brazilian Amazonia. This species can be differentiated from other morphologically similar species within Trichophoromyia by means of the male characters such as the ratio between the aedeagal ducts/sperm pump, shape and distribution of setae in the gonocoxite, and particularly the distinct shape of paramere.