Kroeber, Thomas

2013-2, Mramba, Furaha, Oloo, Francis, Byamungu, Mechtilda, Kroeber, Thomas, McMullin, Andrew, Guerin, Patrick, Mihok, Steve

Background: Here we set out to standardize long-lasting, visually-attractive devices for Glossina swynnertoni, a vector of both human and animal trypanosomiasis in open savannah in Tanzania and Kenya, and in neighbouring conservation areas used by pastoralists. The goal was to determine the most practical device/material that would induce the strongest landing response in G. swynnertoni for use in area-wide population suppression of this fly with insecticide-impregnated devices. Methods and Findings: Trials were conducted in wet and dry seasons in the Serengeti and Maasai Mara to measure the performance of traps and targets of different sizes and colours, with and without chemical baits, at different population densities and under different environmental conditions. Adhesive film was used as a simple enumerator at these remote locations to compare trapping efficiencies of devices. Independent of season or presence of chemical baits, targets in phthalogen blue or turquoise blue cloth with adhesive film were the best devices for capturing G. swynnertoni in all situations, catching up to 19 times more flies than pyramidal traps. Baiting with chemicals did not affect the relative performance of devices. Fly landings were two times higher on 1 m2 blue-black targets as on pyramidal traps when equivalent areas of both were covered with adhesive film. Landings on 1 m2 blue-black targets were compared to those on smaller phthalogen blue 0.5 m2 all-blue or blue-black-blue cloth targets, and to landings on all-blue plastic 0.32–0.47 m2 leg panels painted in phthalogen blue. These smaller targets and leg panels captured equivalent numbers of G. swynnertoni per unit area as bigger targets. Conclusions: Leg panels and 0.5 m2 cloth targets show promise as cost effective devices for management of G. swynnertoni as they can be used for both control (insecticide-impregnated cloth) and for sampling (rigid plastic with insect glue or adhesive film) of populations.

2010, Kroeber, Thomas, Kessler, Sébastien, Frei, Jérôme, Bourquin, Martine, Guerin, Patrick

We describe here an in vitro behavioral assay for testing mosquito repellents applied in a dose-based manner to a warm body (34 C) in test cages. The system was used to assess the sensitivity of 4-6-day-old Anopheles gambiae to the insect repellent diethyl methyl benzamide (deet). These tests were made in the absence and presence of additional carbon dioxide (CO(2)) applied as a pulse to activate mosquitoes in the cages. In the absence of the CO(2) pulse the mosquitoes hardly responded to the warm body. Increasing the CO(2) level in the cage by 1,000 parts per million caused a 25-fold increase in the number of landings by mosquitoes on the warm body in 2-min tests. This mosquito activation allowed the measurement of a significant reduction in the number of landings to bite on the warm body with increasing doses of deet (0.4 to 3.8 mu g/cm(2)). An asymptotic nonlinear model fitted to the repellency data in the presence of CO(2) allowed estimation of the effective dose of deet that reduced landings to bite by 50% (ED(50)) at 0.95 mu g/cm(2) (5 nmol/cm(2)) and the corresponding ED(95) at 4.12 mu g/cm(2) (21.5 nmol/cm(2)). This in vitro bioassay has the advantage of permitting a fast throughput of test products under standardized conditions and is suitable for screenings designed for the purpose of discovering lead products with as yet unknown human toxicological and dermatological profiles.

2007, Kroeber, Thomas, Guerin, Patrick

An artificial feeding unit with a reinforced silicone membrane to replace host skin provides ticks with a perch over blood with a tick attachment rate of 75-100%. Some 5 mg of an acaricide like fipronil is sufficient to establish survival curves over different doses down to ppb levels in blood. This in vitro feeding assay for hard ticks is more advantageous than in vivo trials on animals.

2000, Kroeber, Thomas, Kreuzer, M., Senn, M., Langhans, W., Sutter, F.

A 5-week experiment with 24 multiparous early lactating Brown Swiss cows was conducted to investigate the effects of supplementary rumen-protected methionine in conjunction with dietary protein reduction on metabolism and performance after 1 week of control measurement. Three rations containing 175, 150 and 125 g of crude protein/kg feed dry matter were supplemented with methionine. The fourth ration, also only containing 125 g of crude protein/kg dry matter, remained unsupplemented. The four treatment groups had a similar metabolic supply of other essential amino acids, protein and energy, as calculated by various approaches. The two low protein rations were, however, slightly deficient in ruminally degraded protein. Treatment effects remained low on feed intake, forage meal pattern, milk yield and fat as well as lactose content. In contrast, the content and yield of milk protein significantly declined only in the unsupplemented low protein ration relative to the initial value. Compared with this ration, the decline in milk protein yield was clearly delayed in the supplemented low protein ration. Blood plasma methionine tended to be reduced without supplementation and to be increased with additional methionine. Supplementation of methionine reduced other plasma amino acids. Plasma insulin, glucose, lactate, ketone bodies and aspartate amino transferase activity indicated a certain liver stress and a somewhat elevated energy requirement with high and particularly with ION protein content (when unsupplemented). Methionine improved metabolic protein utilization, followed by the lowest plasma, urine and milk urea levels in the supplemented low protein diet. In conclusion, no major adverse effects were assessed under the conditions tested. Supplementation of methionine may nevertheless be useful in rations with particularly low protein content fed to early lactating cows in order co prevent negative long-term effects which were only visible here as trends.

2013, Kroeber, Thomas, Bourquin, Martine, Guerin, Patrick

The threat of transmission of Lyme borelliosis and tick-borne encephalitis by ixodid ticks has resulted in an increasing number of tick repellents coming onto the market. To allow proper evaluation of the efficacy of different types of compounds and their formulations, there is a need for standardised methods for testing ticks repellents. Ticks show a marked negative geotactic response following contact with a potential host, i.e., they climb up in order to locate attachment and feeding sites, whereas exposing ticks to repellents induces positive geotaxis, i.e., ticks walk downwards or drop off the treated host or substrate. We describe here complementary tests that employ these geotactic responses to evaluate repellents: one in vitro on a warm glass plate and the other on the lower human leg (shin). The compounds tested were DEET, EBAAP, icaridin, capric acid, lauric acid, geraniol, citriodiol, citronella essential oil and lavender essential oil, all non-proprietary ingredients of widely distributed tick repellent formulations. In controls on both the warm glass plate and the human leg, the majority of Nodes ricinus nymphs walk upwards. By contrast, in both the in vitro and in vivo tests, effective doses of repellents cause ticks to either walk downwards or fall off the substrates, termed here "affected ticks". The ED75 values for affected ticks on the human leg indicate that the test products can be divided into three groups: (1) icaridin, EBAAP, DEET and capric acid with values between 0.013 and 0.020 mg/cm(2), (2) citriodiol and lauric acid with values between 0.035 and 0.058 mg/cm(2), and (3) geraniol, citronella oil and lavender essential oil with values between 0.131 and 1.58 mg/cm2. The latter three products can be considered as less effective repellents. The tests on the warm glass plate resulted in very similar efficacy rankings for the products tested in vivo, and the ticks' behavioural responses also corresponded closely to those observed on the treated human leg. The ED75 values on the glass plate ranged from half to one sixth needed on the leg. The warm glass plate test thus provides a reliable alternative to human subjects for an initial evaluation of new repellents, and is particularly appropriate for testing products with still to be determined human toxicity and dermatological effects. (C) 2013 Elsevier Inc. All rights reserved.

2010, Schymura, Danuta, Forstner, Maike, Schultze, Anna, Kroeber, Thomas, Swevers, Luc, Iatrou, Kostas, Krieger, Jürgen

Odor-detection in the malaria mosquito Anopheles gambiae involves large families of diverse proteins, including multiple odorant binding proteins (AgOBPs) and olfactory receptors (AgORs). The receptors AgOR1 and AgOR2, as well as the binding protein AgOBP1, have been implicated in the recognition of human host odors. In this study, we have explored the expression of these olfactory proteins, as well as the ubiquitous odorant receptor heteromerization partner AgOR7, in the thirteen flagellomeres (segments) of female and male antenna. Expressing cells were visualized by adapting a whole mount fluorescence in situ hybridization method. In female mosquitoes, AgOR1-expressing olfactory receptor neurons (ORNs) were almost exclusively segregated in segments 3 to 9, whereas AgOR2-expressing ORNs were distributed over flagellomeres 2 to 13. Different individuals comprised a similar number of cells expressing a distinct AgOR type, although their antennal topography and number per flagellomere varied. AgOBP1-expressing support cells were present in segments 3 to 13 of the female antenna, with increasing numbers towards the distal end. In male mosquitoes, total numbers of AgOR- and AgOBP1-expressing cells were much lower. While AgOR2-expressing cells were found on both terminal flagellomeres, AgOR1 cells were restricted to the most distal segment. High densities of AgOBP1-expressing cells were identified in segment 13, whereas segment 12 comprised very few. Altogether, the results demonstrate that both sexes express the two olfactory receptor types as well as the binding protein AgOBP1 but there is a significant sexual dimorphism concerning the number and distribution of these cells. This may suggest gender-specific differences in the ability to detect distinct odorants, specifically human host-derived volatiles.

2007, Kroeber, Thomas, Guerin, Patrick

Prevention of tick bites and transmission of tick-borne pathogens requires the use of molecules that target physiological processes crucial to both tick and pathogen survival. These molecules are best tested in standardized in vitro assays. Because hard ticks require several days to feed to repletion, the development of in vitro feeding assays for these species is challenging. A standard and easily automated feeding assay has been developed for the tick Ixodes ricinus that involves feeding on blood through a membrane that mimics the elasticity of skin. The system can be adapted to feed other hard tick species in vitro. This assay permits, among others, investigations on the role of tick endosymbionts on tick survival, the identification of potential vaccine candidates and drugs, and the application of genomic tools in vitro, including RNA interference experiments.

2010, Biessmann, H., Andronopoulou, E., Biessmann, M. R., Douris, V., Dimitratos, S. D., Eliopoulos, E., Guerin, Patrick, Iatrou, K., Justice, R. W., Kroeber, Thomas, Marinotti, O., Tsitoura, P., Woods, D. F., Walter, M. F.

Haematophagous insects are frequently carriers of parasitic diseases, including malaria. The mosquito Anopheles gambiae is the major vector of malaria in sub-Saharan Africa and is thus responsible for thousands of deaths daily. Although the role of olfaction in A. gambiae host detection has been demonstrated, little is known about the combinations of ligands and odorant binding proteins (OBPs) that can produce specific odor-related responses in vivo. We identified a ligand, indole, for an A. gambiae odorant binding protein, AgamOBP1, modeled the interaction in silico and confirmed the interaction using biochemical assays. RNAi-mediated gene silencing coupled with electrophysiological analyses confirmed that AgamOBP1 binds indole in A. gambiae and that the antennal receptor cells do not respond to indole in the absence of AgamOBP1. This case represents the first documented instance of a specific A. gambiae OBP-ligand pairing combination, demonstrates the significance of OBPs in odor recognition, and can be expanded to the identification of other ligands for OBPs of Anopheles and other medically important insects.

2007, Kroeber, Thomas, Guerin, Patrick

Animal husbandry could not be practised over large areas of the planet without acaricides. The prevention of tick bite and the transmission of diseases requires the use of pesticides, but this contributes to the development of tick resistance against acaricides. This drives the quest for new molecules that target physiological processes crucial to tick survival. In vivo trials involve multiple repetitions because of inherent variations between host animals, requiring large amounts of test products and ticks. An in vitro alternative should permit the testing of the ability of a product to restrict attachment and feeding by ticks at precise doses. In this paper an in vitro feeding system is described where the European tick Ixodes ricinus L. feeds on blood through a cellulose rayon-reinforced silicone membrane. The membrane Shore hardness is modified to imitate the elastic retraction forces of skin that ensure the closing of tick penetration sites on the membrane to prevent bleeding. Tick attachment (75-100%) is achieved by adding chemical and mechanical stimuli to the membrane. Survival curves for different doses of fipronil and ivermectin tested with the method showed highly reproducible acaricide effects within 5 - 7 days. Significant effects are recorded down to ppb levels in blood. Standardised tests can be made with blood from the same donor animal or culture medium under the membrane. (c) 2006 Society of Chemical Industry.

2003, McMahon, Conor, Kroeber, Thomas, Guerin, Patrick

Most in vivo and in vitro tests with repellents or deterrents against ticks have not considered which sensory channel is being targeted. We have recorded the responses of two hard tick species (Acari: Ixodidae) in vitro to determine if such products can disrupt the perception of an attractant in a repellent assay or the perception of an arrestment stimulus in a deterrent assay. Ethyl butylacetylaminopropionate (EBAAP), N,N-diethyl-methyl-benzamide (deet), permethrin and indalone were chosen to test their capacity to inhibit the attraction of Amblyomma variegatum Fabricius to its aggregation-attachment pheromone. Vapours of each test product plus those from a synthetic blend of the pheromone were delivered to the walking tick in an air stream on a locomotion compensator. Neither EBAAP, deet, permethrin nor indalone could inhibit attraction of A. variegatum even when each of the test products was delivered at 10(6) times the pheromone. Indalone did decrease the attraction of A. variegatum to the pheromone and induced repulsion of A. variegatum when presented on its own in the air stream. The effect of permethrin, a sodium channel blocker, was also tested in a deterrent assay measuring the arrestment of Ixodes ricinus (L.) adults on its own faeces and faecal constituents. Permethrin deterred arrestment at doses of 670 fg/cm(2) to 67 ng/cm(2), i.e. at levels five times lower than the dose of chemostimuli present in the arrestment stimulus. This sensitivity to permethrin suggests that it acts via the contact chemoreception channel.