Protocols:
The following are the main protocols selected by the Diptera TWIG for the present project.
Protocol for Setting Up Malaise Traps and Handling Samples
Protocols for Culicidae (Darlene Judd)
Protocols for Ceratopogonidae (Art Borkent)
Protocols for Dolichopodidae (Daniel Bickel)
Protocols for Asilidae (Eric Fisher)
Protocols for Sarcophagidae (Thomas Pape)
Protocol for Acalyptratae (Stephen Marshall and Matthias Buck)
Protocols for Culicidae (Darlene Judd)
Collecting:
Conservation Areas are visited twice a year, one during the dry season and the other during the rainy season, in order to determine seasonal differences in species composition. Detail information about habitat of immature stages (larval/pupa) is recorded to have a better knowledge in mosquito biology. Adults are collected with entomological reds and ABC traps using light or CO2 emission sources. Females feeding on blood are collected in the field to obtain eggs and progenies. These females are kept into containers having a web cotton layer in the bottom and a piece of cotton soaked with sugar water in the top. Eggs laid by these females on the cotton are transferred into a cup with water by using a brush. Tetra Min baby fish food is used to feed larvae and allow their development. A sample of eggs is preserved in 75% alcohol. Larvae are collected from hollow logs, artificial containers holding water, lagoons, ponds, bromelias, heliconia inflorescences, Aracea petioles, etc. Aquatic reds, Pasteur pipets, aspirators, etc are used for collecting larvae. Collected material is transported to the lab into Whirl-Pak plastic bag or clean containers properly labeled.Larvae rearing:
For identification purposes, it is important that all material reared, larvae and pupae, is slide-mounted. Larval and pupa stages will be associated with the corresponding adults to elaborate taxonomic keys of the three stages. At the lab, larvae are sorted individually o in 2-3 specimens groups and placed into cups with clean water to allow pupation. Some larvae are preserved in 75% alcohol for subsequent mounting. All reared material (larval exuviae, pupae and adults) has the same collecting and rearing code. Exuviae are kept into ¼ dram containers with 75% alcohol.Specimens mounting:
Adults are mounted using triangle system in horizontal position and legs toward the pin. Preferably, the exuviae should be mounted as soon as possible. If it sits around the skins become difficult to mount. Larval and pupal exuviae are put in isopropyl alcohol for 5 minutes, and then mounted in Euparal mounting medium. When mounting larval exuviae, thoracic and abdominal hairs must be extended toward sides, and the siphon has to be slightly put apart from the anal segment. For larva mounting, head has to be dissected and tissues removed with KOH for 5 minutes or with lactic acid for 10-15 minutes. Abdomen is dissected in the seventh segment. From each morphotype, a female and a male are slide-mounted. Antennae, head, wings, legs, and genitalia of these adults are dissected. These body structures are mounted on the same slide and under 10 mm diameter cover slide. Slides are kept in a 45 ºC stove for 3-4 weeks for drying.
Diptera TWIG / Sampling site selection /
Taxa selection / Products / Protocols
