Helminthology

Author: J BOOMKER

There are various methods the veterinarian can use to diagnose helminthosis and identify the worm species causing the problem. The method used will be based on a number of factors, including the species of animal, the type of parasite, whether the animal is alive or dead, the cost of the individual tests, and the facilities available to the veterinarian.

Faecal examination methods

Faeces provide a useful source of diagnostic material for the identification of the endoparasite infesting an individual animal, or group of animals. Larvae and the eggs of some species can be used to identify the species (see later in chapter) and the number of eggs in a faecal sample can give an indication of the severity and the relevance of an infection. It must be stressed that a faecal egg count merely gives an indication of the presence of reproductively active female worms, and there is no correlation between the egg count and the actual worm burden that the animal carries. For evaluation of worm burdens of species, refer to Table 19 in Addendum B.

Direct microscopic examination

This is a simple and practical method for the veterinarian, in the field, to use. Suspend a small amount of faeces in a few drops of water and place a cover slip over the material. It is possible to detect most eggs or larvae using this method, but due to the small sample this technique will only detect heavy infections.

Flotation methods

These are used to visualise worm eggs or parasites. By suspending the faecal material in a solution with a higher specific gravity than the worm eggs, the latter will float to the surface. The sensitivity of the method used depends on a number of factors – including the specific gravity and viscosity of the solution, the SG of eggs or parasite stages, the concentration of eggs/parasites, and the care and precision with which the sample was processed.

Various methods are used and their advantages and disadvantages are outlined below. Where applicable, details of the individual procedures are given in Addendum B.

Direct flotation method: this qualitative method is suitable for use in private practice for all species and is inexpensive. It is conducted with commercially available devices such as Fecalyzer®, Ovassey® and Ovatector®. The sample used is freshly-collected faeces from the rectum – that can be stored in a refrigerator if there is to be any delay in despatching the specimen to the laboratory. The specimens for submission to the diagnostic laboratory should be placed in a suitable specimen container with a tightly fitting screw cap, and placed in a polystyrene coolbag with an ice pack to prevent development of the eggs. Speedy delivery using an overnight courier is essential. Shredded paper towels or bubble wrap to prevent direct contact between the faecal sample and the frozen ice pack, should be used.

Centrifugal flotation methods: this is a qualitative method used in diagnostic laboratories. The sampling methods are, as for the direct flotation method. For the preparation of flotation fluid, see Addendum B.

McMaster method

This is a quantitative method for counting eggs of gastrointestinal nematodes and the results are expressed as epg (eggs per grams of faeces). This method is used as a monitoring tool for nematode infection in production animals and equids.

It can be used to monitor nematode dynamics and for the Faecal Egg Count Reduction test, which monitors the efficacy of control and can also be used as a screening test to identify the presence of anthelminthic resistance (see under control). Sampling requirements are as for direct flotation. For laboratory instructions for the McMaster method, see Addendum B.

Sedimentation methods

These methods are used for the demonstration of trematodes and lungworm larvae.

Benedek method: this method is suitable for routine diagnostics in a veterinary practice. It provides qualitative and quantitative analysis for trematode eggs, and is inexpensive. The sampling requirements are as for the direct flotation test. See Addendum B for the laboratory method of Benedek’s sedimentation method.

Pitchford-Visser and Bauermann- Wetzel methods: these are confined to use in diagnostic laboratories.

Species-specific tests

Centrifugation-flotation test

This is a specialised, qualitative test used for detecting Spirocerca lupi and Anaplocephala eggs, and must be done by an experienced diagnostician.

Adhesive swab tape test

This cheap, qualitative test can be used to detect cestode eggs in dogs and cats, as well as oxyurid eggs in equids, primates, and humans.

Blood sample examination

Stained thick and thin blood films are used to detect micofilariae in various species such as dogs and cats (Dirofilaria spp). Many filarial infections are detected incidentally in routinely prepared stained thin and thick blood films. However, species identification using this technique is highly unreliable.

Membrane filtration technique

The most sensitive technique to detect patent filarial infections is used for screening dogs and cats for specific infections to satisfy import and export requirements. A sample is taken in EDTA or heparin. The storage and dispatch requirements are as for the ‘direct faecal flotation’ method. This technique should be entrusted only to an experienced diagnostician.

Microfilarial identification by acid phosphatase staining

This is the definitive differential technique for species identification and should only be done by an experienced diagnostician. Sampling is as for the membrane filtration technique.

Detection of encysted larvae of Trichinella spiralis

This method is used to detect the helminths in the musculature of domestic pigs, horses and game animals for human consumption and is conducted by personnel of specialised diagnostic laboratories. This is a requirement to meet import regulations of European and other overseas countries. There are two methods: the compressorium method and enzymatic digestion method.

Larval culture

This entails the recovery and culture of 3rd stage larvae from faecal samples. It is a highly specialised method used in diagnostic and research laboratories by experienced staff.

Serodiagnosis

Immunological tests are not widely used in the diagnosis of helminth infections, partly because the reaction against the intestinal helminths is not specific, and partly because it is easier, faster and cheaper to examine faeces for the presence of eggs. However, important conditions where ova are not produced make it imperative to use serodiagnosis. Examples of such conditions are heartworm (Dirofilaria immitis) in dogs, visceral larval migrans (Toxocara canis), trichinosis (Trichinella spiralis), fasciolosis and Ancylostoma caninum infections in humans. In all these conditions, the ELISA test has been shown to be the most useful serological method. Recently, monoclonal antibodies and antibody-secreting cell probes have been used in the diagnosis of Haemonchus contortus infections in sheep. Western blot techniques are used for the detection of Taenia spp. in dogs and sheep.

There are various commercial test kits available for heartworm antigen detection. The test can detect occult infections and is used for the screening of dogs to meet export requirements to New Zealand. The tests may cause cross reactions with related filarial helminths (Dirofilaria repens). Conducting serological tests and interpreting results should be entrusted only to an experienced diagnostician. These tests are usually very expensive. The sampling technique is as for the membrane filtration technique.

Necropsy

The best method for making a definitive diagnosis of helminthosis, is by doing a systematic necropsy. The effects of the worms can then be seen and in many cases the worms themselves can also be seen and collected.

The technique used for each species involved, is of cardinal importance. The most important points for each system are given below. The details of the necropsy method are given in the Addenda.

Ruminants

Pay special attention to the rumen, the eyes and the first 10 metres of the small intestine. If Trichostrongylus is suspected, gently scrape the mucosa of the small intestines with a glass slide and cover with another glass slide. The glass slides should be held up against the light to see these tiny worms.

Horses

The cranial mesenteric artery must be ligated at the intestines and collected together with the entire aorta. Ligate the thoracic aorta in front of the heart, as well as the iliac arteries caudal to the bifurcation of the aorta. Cut between the ligatures and remove the entire aorta together with the anterior mesenteric artery. Open separately and examine for parasites. Examine the peritoneum visually and collect the larvae of other Strongylus species that may be present. Pay special attention to the large blood vessels of the caecum and the colon, where large migrating larvae of the Strongylus species may also be found.

Dogs

The aorta of dogs must be ligated in the same way as that described for horses. However, it is only necessary to take the thoracic aorta, since Spirocerca lupi occurs here and lesions may be present.

Diagnosis in habitats

This is used mostly for experimental or investigational work, and should be entrusted to a skilled diagnostician.

Collection and counting of infective nematode larvae on herbage

This is a highly specialised and timeconsuming procedure and is only done by personnel of specialised diagnostic laboratories.

Recovery of metacercariae from herbage

This is a highly specialised and timeconsuming procedure and is only conducted by personnel of specialised diagnostic laboratories.

Recovery of snails from habitats

Typical habitats are shallow streams, ponds, dams, marshes, and water troughs – that should be investigated for the presence of the various snails involved in the life cycle of the specific parasite. Personnel from specialised diagnostic laboratories can identify the snails.