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worm burden. We evaluated linear correlations between ascarid and strongylid egg and worm counts in a large data set and found correlation coefficients to be less than 0.30 (Nielsen et al., 2010). There are several possible reasons for this lack of relationship, and these include (1) the effect of the host immune response reducing worm fecundity (Chapman et al., 2002), (2) the mixed species nature of cyathostomin infections and a substantial fecundity variation between species (Kuzmina et al., 2012) and (3) density-dependent egg shedding, where the egg production per worm is decreased, when worm burdens increase (Kotze & Kopp, 2008; Walker et al., 2009). Thus, a faecal egg count does not allow a quantitative assessment of infection intensity, so a horse with a high egg count does not necessarily harbour more worms than one with a low count.
Limitations
The most important limitation of faecal egg counts is that they are not clinical diagnostic tools as they are not useful as a part of a work-up of a clinical case. Parasite eggs in the faeces are a normal finding in healthy horses and do not indicate pathology or disease. For strongyle infections, the larval stages are the pathogenic stages, and these do not produce eggs. Furthermore, as mentioned above, the magnitude of the egg count does not correlate with the worm burden. Thus, there is no value in determining parasite egg counts in horses showing clinical signs suggesting parasitic involvement. Horses are infected by a multitude of different parasite
species and types, but these are not all detectable by faecal egg counts. As outlined above, egg counts are very useful for monitoring strongylid and ascarid infection and treatment efficacy against these. In addition, several modified techniques have been found useful for diagnosing infection with Anoplocephala perfoliata infection (Nielsen 2016). However, it should be noted that a simple McMaster technique has been shown to have a diagnostic sensitivity of less than 10% for this parasite category (Meana et al., 1998; Tomczuk et al., 2014), emphasising that modified techniques are needed. Techniques employing centrifugation-enhanced flotation have been shown to perform with diagnostic sensitivities around 60% for detecting burdens of at least one worm and 90% for detecting infections of 20 worms or more (Kjær et al., 2007; Proudman & Edwards, 1992). Thus, these modified egg counting techniques can detect larger tapeworm burdens with a high level of reliability. Other gastrointestinal parasites are less reliably detected
by faecal egg counts. Eggs of equine stomach worms (Habronema spp.) and threadworms (Strongyloides westeri), can be detected by faecal flotation, but both egg types are relatively thin-walled and may not withstand the osmotic pressure elicited by higher density flotation media. The equine pinworm, Oxyuris equi, does not release its eggs within the intestinal tract, but rather in egg packets on the perianal skin (Reinemeyer & Nielsen, 2014), so the characteristic looking eggs are only occasionally encountered in faecal samples. In addition, botfly larvae (Gasterophilus spp.) are arthropod larvae which do not shed eggs within the intestinal tract.
© 2021 EVJ Ltd.
Methodology principles
The array of different faecal egg counting techniques is enormous, and it can seem challenging to understand their strengths and weaknesses. However,
the
overwhelming majority of techniques is built on the same egg flotation principle, so differences are often relatively minimal. For the flotation-based techniques, two main principles exist: (1) egg counting chamber and (2) test tube with cover slip. The best example of a counting chamber-based
technique is the McMaster (Gordon & Whitlock, 1939), but FECPAK (Presland et al., 2005), FLOTAC (Cringoli et al., 2010) and Mini-FLOTAC (Cringoli et al., 2017) are other examples. Here, a specified amount of faeces is weighed and
suspended in a set volume of flotation medium before it is filtered through cheesecloth or strainers, and a subsample is collected and loaded into a counting chamber. Most of these techniques use passive flotation by leaving the chamber for 5–10 min before counting, but the FLOTAC
technique uses centrifugation of chambers to enhance flotation (Cringoli et al., 2010). Examples of techniques using test tubes with coverslips
are Wisconsin (Cox & Todd, 1962) and Stoll (Stoll, 1930) techniques. Again, a specified amount of faeces is weighed, suspended in a set volume of flotation medium and filtered through cheesecloth or strainers before the entire suspension is transferred to centrifuge tubes. The tubes are filled to the rim creating a slight meniscus and cover slips are placed on top of the tubes before they are centrifuged. Upon centrifugation, the cover slips are removed from the tubes and placed on glass microscope slides and then counted under the microscope. More recently, computerised image analysis technologies
have been developed for automated faecal egg counting and systems now exist for cattle (Elghryani et al., 2020), small animals (Nagamori et al., 2020) and horses (Slusarewicz et al., 2016). These systems have the advantage of their performance being independent of operator training and skill and have been shown to perform with significantly better precision (Cain et al., 2021). Furthermore, processing time is significantly reduced in comparison with manual techniques without compromising diagnostic quality (Slusarewicz et al., 2019). Some of these automated systems have recently been introduced to the veterinary diagnostic market, and more are likely to be added in the years to come. As a result, manual egg counts will likely be replaced by these automated methods in the future.
Multiplication factor
The multiplication factor is used to convert the number of eggs counted to a standardised number of eggs per gram (EPG) of faeces. While the principle behind this calculation is simple and straight-forward, there is substantial confusion and misconception regarding this factor and its implications for diagnostic performance. The multiplication factor is derived from the protocol
for the given egg counting technique. The calculation is based on the amount of faeces processed, the volume of flotation medium in which the faeces are suspended, and the volume of the subsample examined under the microscope. It is important to emphasise that if this
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