Human Direct Skin Feeding Versus Membrane Feeding to Assess the Mosquitocidal Efficacy of High-Dose Ivermectin (IVERMAL Trial).

BACKGROUND: Ivermectin is being considered for mass drug administration for malaria, due to its ability to kill mosquitoes feeding on recently treated individuals. In a recent trial, 3-day courses of 300 and 600 mcg/kg/day were shown to kill Anopheles mosquitoes for at least 28 days post-treatment when fed patients' venous blood using membrane feeding assays. Direct skin feeding on humans may lead to higher mosquito mortality, as ivermectin capillary concentrations are higher. We compared mosquito mortality following direct skin and membrane feeding. METHODS: We conducted a mosquito feeding study, nested within a randomized, double-blind, placebo-controlled trial of 141 adults with uncomplicated malaria in Kenya, comparing 3 days of ivermectin 300 mcg/kg/day, ivermectin 600 mcg/kg/day, or placebo, all co-administered with 3 days of dihydroartemisinin-piperaquine. On post-treatment day 7, direct skin and membrane feeding assays were conducted using laboratory-reared Anopheles gambiae sensu stricto. Mosquito survival was assessed daily for 28 days post-feeding. RESULTS: Between July 20, 2015, and May 7, 2016, 69 of 141 patients participated in both direct skin and membrane feeding (placebo, n = 23; 300 mcg/kg/day, n = 24; 600 mcg/kg/day, n = 22). The 14-day post-feeding mortality for mosquitoes fed 7 days post-treatment on blood from pooled patients in both ivermectin arms was similar with direct skin feeding (mosquitoes observed, n = 2941) versus membrane feeding (mosquitoes observed, n = 7380): cumulative mortality (risk ratio 0.99, 95% confidence interval [CI] 0.95-1.03, P = .69) and survival time (hazard ratio 0.96, 95% CI 0.91-1.02, P = .19). Results were consistent by sex, by body mass index, and across the range of ivermectin capillary concentrations studied (0.72-73.9 ng/mL). CONCLUSIONS: Direct skin feeding and membrane feeding on day 7 resulted in similar mosquitocidal effects of ivermectin across a wide range of drug concentrations, suggesting that the mosquitocidal effects seen with membrane feeding accurately reflect those of natural biting. Membrane feeding, which is more patient friendly and ethically acceptable, can likely reliably be used to assess ivermectin's mosquitocidal efficacy. CLINICAL TRIALS REGISTRATION: NCT02511353.

Life history and reproduction of the neotropical caecilian Siphonops annulatus (Amphibia, Gymnophiona, Siphonopidae), with special emphasis on parental care

Due to their mainly fossorial way of life, caecilian amphibians are the least known order of terrestrial vertebrates. Here, we present new observations on the natural history and reproductive biology of the neotropical oviparous, siphonopid caecilian Siphonops annulatus from a long-term study of this species in the field and in captivity. In the studied population, mating occurs between the end of August and beginning of October, and oviposition between November and December, when rainfall peaks. Egg hatching occurs between the end of December and beginning of January. The complete cycle of maternal care, from oviposition to independent, self-sufficient offspring lasts about 3 months. After eclosion, the altricial young feed on the mother's specially modified skin (maternal dermatophagy) and are also supplied by a fluid released from coming from the maternal cloaca. Also presented are observations on the burrows, feeding and social behaviour of S. annulatus.

Membrane feeding of dengue patient's blood as a substitute for direct skin feeding in studying Aedes-dengue virus interaction.

BACKGROUND: Understanding the interaction between Aedes vectors and dengue viruses (DENV) has significant implications in determining the transmission dynamics of dengue. The absence of an animal model and ethical concerns regarding direct feeding of mosquitoes on patients has resulted in most infection studies using blood meals spiked with laboratory-cultured DENV. Data obtained from such studies may not reflect the natural human-mosquito transmission scenario. This study explored the potential of using membrane feeding of dengue patient's blood as a substitute for direct skin feeding. METHODS: Four to six-day old female Ae. aegypti were provided the opportunity to feed via direct exposure to a patient's forearm for 15 min or via exposure to EDTA-treated blood from the same patient through an artificial membrane for 30 min. Mosquitoes from both feeding methods were incubated inside environmental chambers. Mosquitoes were sampled at day 13 post-feeding. Midgut and salivary glands of each mosquito were dissected to determine DENV infection by RT-qPCR and viral titration, respectively. RESULTS: Feeding rates: Direct skin feeding assay (DSFA) consistently showed higher mosquito feeding rates (93.3-100%) when compared with the membrane feeding assay (MFA) (48-98.2%). Midgut infection: Pair-wise comparison between methods showed no significant difference in midgut infection rates between mosquitoes exposed via each method and a strong correlation was observed in midgut infection rates for both feeding methods (r = 0.89, P < 0.0001). Overall midgut viral titers (n = 20) obtained by both methods were comparable (P ≥ 0.06). Salivary gland infection: Pair-wise comparison between both methods revealed no significant difference in salivary gland infection rate. Strong correlation in salivary gland infection was observed between DSFA and MFA (r = 0.81, P < 0.0001). In general, mosquitoes fed directly on dengue patients and those on patients' blood (n = 11) had comparable virus titer (P ≥ 0.09). CONCLUSION: DENV midgut and salivary gland infection rates showed good concordance between DSFA and MFA blood meal exposure methods. Freshly-obtained venous blood in EDTA from dengue patients for MFA can be used as a substitute to DSFA, especially in circumstances where bioethics approval or patient recruitment is difficult to obtain for vector competence studies. Nevertheless, mosquito numbers will need to be increased to compensate for lower feeding rate in MFA.

Metabolic and endocrine changes during the reproductive cycle of dermatophagic caecilians in captivity.

The amphibian order Gymnophiona is poorly known, and studies about their reproduction are mainly comparative and descriptive, focusing on the structure of testes, ovaries and oviducts. However, to understand the reproductive processes, including those of the oviparous dermatophagic species, it is important to know the dynamics of storage and mobilization of energetic substrates to gonads and skin during the reproductive cycle of males and females, as well as the endocrine control associated. For the present study, total lipids and proteins were measured during the annual cycle in the plasma, liver, muscle, testes, ovaries and skin of Siphonops annulatus in captivity. Plasma levels of gonadal steroids (estradiol, testosterone and progesterone) were quantified by radioimmunoassay. Histological analyses of ovaries and testes were performed to classify the maturation stages of the animals during the reproductive cycle. Gonadal maturation in males and females of S. annulatus was accompanied by metabolic changes in reserve tissues, which supported gonadal development and prepared the females' epidermis for skin feeding by the offspring. Even in confinement conditions, females and males synchronized the reproductive period. However, due to the absence of environmental cues in captivity inadequate levels of the hormones responsible for gamete release were triggered, leading to a lack of reproductive success.