The overall goal of this procedure is to determine whether or not parasites are present in laboratory rats or mice. This may be accomplished in a variety of ways, including examination of the fur and skin examination of the feces and examination of the intestinal contents. Ultimately, results can be obtained that give important information on the overall health of the colony through routine examination of these samples.
Visual demonstration of these methods is crucial as if they're not performed thoroughly and carefully. A parasitic infection may be missed. Generally, people new to these methods will struggle because even if techniques are performed correctly, parasites may be misidentified.
The methods we will demonstrate can help answer key questions about the health status of the rats and mice used for your research Prior to taking fur and perianal samples. Prepare at least two glass slides per animal with drops of mineral oil. Lift the mouse of interest from its cage and place on the inside of the overturned cage lid.
Holding it by the tail. Apply a five centimeter length cellophane tape firmly to the mouse's, perineum, and perianal area. If successful, there should be hair stuck to the tape.
Continue restraining the animal and grasp its fur with forceps. Gently pluck fur samples from the scapular area, ventral cervical region, axillary area, inguinal area, and dorsal rump. Place those samples on five centimeter length of cellophane tape.
As each sample is taken, apply the tape to a slide tacky side down. Once all the samples are taken, overlay another drop of mineral oil on each sample and apply glass cover slips. When examining the perianal slides, it is unlikely that eggs of any parasite other than si facia will be found.
For fecal examination, collect two to five pellets from each animal or the cage. If the samples are extremely dry, moisten them with 500 microliters of 0.9%saline. Place the samples in a 10 milliliter pill vial or fecal flotation device.
Place the collection vial in the Petri dish and fill the vial one quarter full of dense flotation medium. Then mash and stir the feces into the solution in the vial until no large pieces remain. Continue to add flotation medium until a meniscus forms above the edge of the vial.
Then place the cover slip on the meniscus and incubate the vial at room temperature for 15 minutes. During the incubation, parasite, eggs and proteasome and oasis rise and adhere to the cover slip, which is then mounted to a slide for further examination. This assay can be performed in something as simple as a pill vial.
If feizer are not available, begin by collecting two to 10 fecal pellets per animal, and if desiccated rehydrate them in 500 microliters, 0.9%saline. Fill a 15 milliliter glass centrifuge tube, one-quarter full with a dense flotation solution and mix the sample. Cover the glass tube with a plastic snap cap, and vortex the mixture for 15 to 30 seconds until it has a homogenous appearance.
Now, add additional flotation solution to each sample tube. To form a slight positive meniscus. Apply a plastic cover slip to each tube and ensure that it makes full contact with the tube blip.
For the best seal, clean the tube with a swab. Next centrifuge, the tubes and cover slips for 10 minutes after the spin. If any cover slips are lost or broken, replace them and gently tip the tubes so that the meniscus touches the new cover slip.
Then remove all of the cover slips and place them on a labeled clean glass microscope slide. An iodine stain will help identification of cysts on an anesthetized or euthanized animal. Trim the dorsal hair coat near the base of the tail and the temporal region of the head or around any skin lesion or other site to be scraped deeply scraped the skin with the scalp blade in the direction opposite of hair growth.
This erodes the epidermis. Wipe the blade off in mineral oil applied to a labeled slide. Add an additional drop of oil if needed.
Before covering the sample with a glass cover slip, place a euthanized animal on a clean dissection board and use forceps to lift the abdominal wall at the genital area. Then using scissors carefully incise the ventral abdominal wall from the genital area to the base of the rib cage, and remove the skin and muscle to expose the intestines. Then collect a portion of the secum and duodenum into the lid of a Petri dish.
Incise each intestinal segment lengthwise. To expose the mucosa, place two drops of 0.9%saline on a single labeled slide. Then using a sterilized inoculating loop, scrape the mucosa, the duodenum, and place the scrapings in one drop of saline.
Repeat the process, transferring a sample from the cecum to the other drop. Remove the remaining intestines. Begin at the duodenum and continue to the descending colon.
Transfer the intestines to the bottom of a 100 milliliter Petri dish in the Petri dish. Use scissors to cut the intestines into smaller sections. Add enough tap water to the dish to barely submerge the collected tissue.
Incubate the sample mixture at 35 to 40 degrees Celsius for a minimum of 10 minutes. This will liberate and expose luminal helmets while the macerated intestinal sample is incubating. Use face contrast microscopy to examine the duodenal and SQL samples for parasites.
After the incubation of the full intestinal sample is completed, examine the dish contents under a dissecting microscope. Using a probe, make a thorough examination. Pinworms will appear as small white hair-like worms.
Tapeworms will appear as segmented flatworms. Collect any worms or suspicious materials with a small forceps and mount on a slide. For further examination under a light microscope, place the euthanized mouse or rat directly on the stage of a dissecting microscope under the 10 x objective, use an applicator stick to part the hair and observe the base of the hair shafts.
Ectoparasites may look like dandruff or a yellow waxy buildup at the base of a hair shaft or directly on the skin. At a minimum search for ectoparasites in the cranial region. Inguinal and axillary regions.
Check carefully between the eyes and the penny between the penny, between the scapula and under the jaw. Using small forceps. Transfer possible ectoparasites and other suspicious materials to a slide.
Mount them in mineral oil for identification. These procedures will find many common parasites. Here are a few examples.
These are SE facia eggs found from perineal tape samples. A fur plug was found to contain the mite myopia musculi from a fecal centrifugation preparation. These are picis tetter eggs.
In this direct examination of intestinal mucosa, a trioni may be seen examination of macerated intestines revealed These adult pinworms Once mastered, this complete examination for parasites can take under an hour. After watching this video, you should have a good understanding of how to perform various examinations for parasites in rats and mice. Don't forget that some of these parasites can be transmitted to humans.
Always follow basic laboratory safety rules.
