A Capsule-Based Model for Immature Hard Tick Stages Infestation on Laboratory Mice

Our technique is suitable for various types of experiments but our mouse model is required for feeding ticks and is necessary to keep the ticks in enclosed area for easy collection and monitoring. The benefit of this simple method is it efficiency, short duration, and the ability to monitor or collect ticks at different time points of an experiment. Our system is suitable for studying tick biology, host-vector pathogen interactions or evaluating different control measure of these medically important arthropods.

This method requires accuracy and attention to detail as immature tick stages are small and feeding them on mice necessitates the enclosed capsule to be be firmly attached to the mouse for several days. Begin by sticking together a two millimeter EVA foam, an adhesive double sticky foam. Use a 20 millimeter diameter leather hole punch to cut a circle from the foam pieces.

Then, use a 12 millimeter hole punch to cut the interior. Peel the protective paper strip from the adhesive double sticky foam and attach a 20 millimeter transparent plastic circle to the foam. If feeding larvae, do not remove the protective paper strip from the adhesive foam and proceed directly with capsule attaching.

Make a one centimeter slit in the transparent plastic then use an entomological pin to create at least 10 small holes for moisture evaporation during the experiment. After the mouse has been anesthetized place it on the manipulation pad and attach a nose cone for continuous isoflurane supply. Ensure that the breathing rate is less than 80 breaths per minute by adjusting the isoflurane level.

Shave the anterior part of the mouse from behind the shoulder blades up to the area just behind the ears creating a shaved area that is greater than the capsule surface. Apply non irritating latex glue to the entire EVA foam site of the capsule and wait for one minute. Place the capsule on the mouse's back and apply constant pressure for three minutes especially on the left and right side of the capsule.

Slightly lift the capsule to visually check its attachment to the skin. To ensure the firm attachment of the capsule apply a slight three minute constant pressure with the fingers especially on the left and right side of the capsule. If non-attached regions are found, apply more glue with a spatula and press for another three minutes.

If a non-attachment region forms during the experiment fix it in the same way. Introduce individual nymphs into the capsule via the one centimeter slit. Slightly squeeze the capsule from two sides to allow the transparent plastic to bend then push the nymph inside the capsule.

Turn the forceps 90 degrees and pull them out. For larvae infestation, remove the paper slip from the attached capsule and place the syringe with the larvae directly inside. Deposit the ticks by pushing the syringe plunger then gently turn the plunger towards the skin to remove the remaining attached larvae.

Once the larvae are deposited onto the skin close the capsule by attaching the transparent plastic circle. Cut a 3.5 millimeter strip of the protective plastic band and apply the protective plastic band around the capsule then return the mouse to the cage. To collect the ticks, create a cross shaped cut in the plastic with a scalpel and use forceps to collect the ticks.

If necessary, reclose the capsule by sticking an adhesive plastic patch on the transparent plastic. the same plastic patch can be used if collection of ticks at multiple time points is required. Alternatively, the mouse can be euthanized and the ticks collected or manually detached after capsule removal.

For successful recovery of the mice, keep them in a cage for one additional week and let the capsules detach naturally. Once the capsule is off, check for abnormal reactions on the skin. Apply any emollient lotion if there is any irritation.

This protocol makes it possible to feed immature hard ticks in EVA foam capsules applied to a mouse's back. The use of the fast drying and non irritating latex glue ensures that the capsule is firmly attached within three minutes and remains attached for at least seven days. Attaching two capsules makes it possible to feed two different groups of ticks on the same animal.

Furthermore, complete recovery of the mice after the experiments, makes it possible to reuse the animals and avoid euthanasia. This system has been successfully used to feed Ixodes ricinus nymphs. A moderate to high engorgement success rate was achieved in two different mouse strains.

In both strains, all nymphs finished the feeding within four to five days while the majority dropped off on the fourth day. When attempting this protocol, it's important to shave a sufficient area of mouse's skin and effectively attach the capsule to avoid it's accidental detachment during the experiment. Besides the study of basic tick biology, this method can be used to explore pathogen transmission from tick to host or host to ticks.

Additionally, the option to reuse the mouse offers a possibility for multiple re-infestations that can be beneficial for immunological studies or investigating different tick-borne pathogen transmissions.