CRISPR mediated transactivation in the human disease vector Aedes aegypti.

As a major insect vector of multiple arboviruses, Aedes aegypti poses a significant global health and economic burden. A number of genetic engineering tools have been exploited to understand its biology with the goal of reducing its impact. For example, current tools have focused on knocking-down RNA transcripts, inducing loss-of-function mutations, or expressing exogenous DNA. However, methods for transactivating endogenous genes have not been developed. To fill this void, here we developed a CRISPR activation (CRISPRa) system in Ae. aegypti to transactivate target gene expression. Gene expression is activated through pairing a catalytically-inactive ('dead') Cas9 (dCas9) with a highly-active tripartite activator, VP64-p65-Rta (VPR) and synthetic guide RNA (sgRNA) complementary to a user defined target-gene promoter region. As a proof of concept, we demonstrate that engineered Ae. aegypti mosquitoes harboring a binary CRISPRa system can be used to effectively overexpress two developmental genes, even-skipped (eve) and hedgehog (hh), resulting in observable morphological phenotypes. We also used this system to overexpress the positive transcriptional regulator of the Toll immune pathway known as AaRel1, which resulted in a significant suppression of dengue virus serotype 2 (DENV2) titers in the mosquito. This system provides a versatile tool for research pathways not previously possible in Ae. aegypti, such as programmed overexpression of endogenous genes, and may aid in gene characterization studies and the development of innovative vector control tools.

Exercise profile and effect on growth traits, carcass yield, meat quality, and tibial strength in Chinese Wannan chickens.

The aim of the study was to understand the dynamic changes in daily step counts (DSC) during the development of chickens and to further explore the effects of exercise on the growth performance, carcass yield, meat quality, and tibial strength of cocks. A total of 600 (half male and half female) 1-day-old Wannan chickens with similar hatching weights were raised under the same rearing conditions. All birds were wing banded and housed in identical cages for from 1 to 8 wk in the experimental poultry house. The dimensions of the cages were 70 × 70 × 40 cm (length × width × height). At the age of 9 to 16 wk, these birds were reared in indoor pens (2 m × 2 m, 1,000 cm2 per bird). In addition, they also had a free-range grass paddock (20 m × 30 m, 1 m2 per bird). The DSC of male and female Wannan chicks were recorded from 70 to 112 d by using a pedometer. At 112 d of age, based on the average DSC, birds were divided into groups representing the highest (HS), medium, and lowest (LS) number of step groups. Fifteen cocks from each group were selected for subsequent experiments. Compared with the LS group, the HS group displayed higher tibial strength (P = 0.025) and lower BW, cooking loss (P = 0.014), shear force (P = 0.023), and drip loss (P = 0.008). The DSC had no effects on the female BW or male carcass parameters. There was no significant change in the DSC of all birds from 70 to 112 d. However, male chickens took more steps than females at 15 (P = 0.025) and 16 (P = 0.012) week of age. In conclusion, the effects of the DSC on the BW of Wannan chickens depend on sex, and enhanced exercise could improve the meat quality and tibial strength of cocks.