KNOWLEDGE SUMMARY

Keywords: CATTLE; COPROANTIGEN; COWS; ELISA; FASCIOLA; FLUKE; SEDIMENTATION; SENSITIVITY

Fasciola hepatica: can the coproantigen ELISA replace the faecal egg sedimentation test?

Jake Collyer, BVMSci^1*

¹ School of Medicine, University of Surrey, Guildford, United Kingdom
* Corresponding author email: jake.collyer@btinternet.com

Vol 9, Issue 4 (2024)
Submitted: 21 August 2023; published: 28 Nov 2024; next review: 25 May 2026
DOI: https://doi.org/10.18849/ve.v9i4.698

PICO question

In adult cattle, is the sensitivity of the coproantigen ELISA test equal or superior to the sensitivity of the faecal egg sedimentation test for the diagnosis of Fasciola hepatica?

Clinical bottom line

Category of research

Diagnosis.

Number and type of study designs reviewed

Three studies were appraised. This included two cross-sectional diagnostic accuracy studies and one case control diagnostic accuracy study.

Strength of evidence

Moderate.

Outcomes reported

The first study reported the findings from 619 tested cattle over 3 sample periods comparing the sensitivity and specificity of the different tests. The sensitivity of the faecal egg sedimentation test varied greatly between the sample periods from 0.81 (95% beta coefficient (BCI) 0.72–0.90) to 0.58 (95% BCI 0.43–0.72) with the coproantigen ELISAs sensitivity remaining consistent at 0.77 (95% BCI 0.64–0.88) throughout.

The second study reported the findings of 200 tested cattle over 2 sampling periods comparing the sensitivity and specificity of the different tests. The mean sensitivity of the coproantigen ELISA was significantly higher than the 4 g/10 g preparations of the faecal egg sedimentation tests at 94% (95% CI 87%–98%) (P < 0.001). 

The third study reported the findings of Coproantigen ELISA testing on 250 bovine faecal samples with 94 confirmed positive for liver fluke via faecal sedimentation testing. The sensitivity of the coproantigen ELISA was 80% and the specificity was 100% compared with 70% and 80% respectively for the faecal egg sedimentation test.

Conclusion

All three studies demonstrated either an increased or equivalent sensitivity of the coproantigen ELISA to the faecal sedimentation test, but only one study reported a statistically significant increase in test sensitivity. Whilst all three studies were diagnostic accuracy validity studies, the systematic sampling strategy of one study was superior to the convenience sampling method of one of the other studies and to the case control method of the other.

Several sources of bias also exist within the included studies. Sampling and selection bias is present in the two of studies due to the animals selected only being sampled over one year. The results of these studies are susceptible to changes in the fluke lifecycle of that year, and the sampled animals are more likely to be fit and well-conditioned as they are presenting for slaughter, and as such are less likely to carry significant/chronic fluke burdens. All three studies are susceptible to validity issues due to an absence of clinical information regarding flukicide treatment and concurrent parasitic diseases which, whilst not impacting the efficacy of diagnostic testing, may cause issues if the studies are to be repeated.

The coproantigen ELISA can be utilised as a suitable adjunctive test to aid in the diagnosis of Fasciola hepatica in adult cattle and has the scope to be used as an early diagnostic test, but whilst the results of the reported studies indicate that the coproantigen ELISA is an accurate and reliable test, it does not provide definitive evidence to warrant the discontinuation of the simple and affordable faecal egg sedimentation test. In order to come to a conclusion regarding the more sensitive test more literature is required that directly compares the coproantigen ELISA to the faecal egg sedimentation test in different clinical scenarios and exploring different diagnostic techniques.

How to apply this evidence in practice

The application of evidence into practice should take into account multiple factors, not limited to: individual clinical expertise, patient’s circumstances and owners’ values, country, location or clinic where you work, the individual case in front of you, the availability of therapies and resources.

Knowledge Summaries are a resource to help reinforce or inform decision making. They do not override the responsibility or judgement of the practitioner to do what is best for the animal in their care.

The evidence

A literature search found two cross-sectional diagnostic accuracy studies (Charlier et al., (2008) and Mazeri et al., (2016)) and one case control diagnostic accuracy study (Palmer et al., 2014). These studies evaluated the diagnostic accuracy and therefore sensitivity and specificity of the different diagnostic tests with the focus in this Knowledge Summary being the comparison of the sensitivity of the faecal sedimentation test (FEST) to the coproantigen enzyme-linked immunosorbent assay (ELISA).

Mazeri et al. (2016) utilised a systematic sampling strategy to select study participants, Charlier et al. (2008) utilised a convenience sampling strategy to select its study participants, and Palmer et al. (2014) utilised a case control sampling strategy.

Charlier et al. (2008) and Mazeri et al. (2016) focused on comparing the accuracy of multiple diagnostic tests with Mazeri et al. 2016 utilising a Bayesian no universally accepted standard approach for test comparison, and Charlier et al. (2008) using liver necropsy as its standard. Palmer et al. 2014 compared the accuracy of the coproantigen ELISA tests against identification of liver fluke eggs on faecal sedimentation as its standard. The strength of evidence of the included papers is moderate as described in the following sections.

Summary of the evidence

Charlier et al. (2008)

 

Mazeri et al. (2016)

 

Palmer et al. (2014)

 

Appraisal, application and reflection

Liver fluke (F. hepatica) is one of the most important endoparasites affecting livestock within the UK, associated with reduced productivity, increased time to slaughter, liver rejections, and occasional sudden death within herds (Cawdery et al., 1977; Mazeri et al., 2017). The principal concern comes from the apparent increase in cases in recent years as shifting weather patterns lead to alterations in the fluke lifecycle, resulting in unpredictable risk-periods for infection (Skuce and Zadoks, 2013). Therefore, it is more important than ever to have an accurate and reliable test that can provide a definitive diagnosis in order to allocate flukicide treatment/preventative measures appropriately. An evidence review was warranted as no diagnostic test for F. hepatica has been recorded as having 100% sensitivity and specificity (Rapsch et al., 2006), it is important, therefore, to be able to evaluate the accuracy and reliability of the diagnostic tests available in order to advise on which test/combination of tests should be prioritised to obtain a valid diagnosis.

The faecal egg sedimentation test has been a longstanding, affordable and effective test for the detection of F. hepatica for many years. Demonstrating a near perfect test specificity in many studies (Graham-Brown et al., 2019; Reigate et al., 2021), it has historically been the universally accepted test in the diagnosis of liver fluke in farm animal species; however, the faecal egg sedimentation test falls short when considering the test sensitivity as various studies have reported wildly different sensitivities indicating that a negative test result is not definite for being free from infection (Anderson et al., 1999; Arifin et al., 2016; Graham-Brown et al., 2019). Various reasons for this relative insensitivity exist, the foremost of these being the variability with shedding of the Fasciola eggs into the biliary system, which is dependent on the presence of mature flukes within the liver, which itself is dependent on the environmental conditions such as temperature, humidity, and rainfall to propagate the snail intermediate host, and the burdens of Fasciola in affected cattle that influence the number of eggs being shed in faeces at any one time (Beesley et al., 2018; Charlier et al., 2014).

The coproantigen enzyme-linked immunosorbent assay (ELISA) test is another diagnostic test that can be performed on samples of faeces from cattle, with the Bio X Diagnostics (Belgium) F. hepatica test kit being one commonly utilised test. Likewise, multiple studies have reported near perfect test specificity (Kajugu et al., 2012; Kajugu et al., 2015), but data regarding test sensitivity is more variable, similar to the faecal egg sedimentation test. Theoretically the test sensitivity should not be affected by the life stage of the infecting liver fluke or the variability in egg shedding in the faeces, so should be more useful in the detection of early F. hepatica infection (Mezo et al., 2004). This testing methodology does come with several disadvantages, including higher running costs when compared to the faecal egg sedimentation test, a requirement for more advanced laboratory equipment, and diagnostic cut-off values set by the test manufacturer.

This Knowledge Summary compared the sensitivities of the above-mentioned diagnostic techniques to determine if the commonly used faecal egg sedimentation test (FEST) is still an appropriate standard test to perform given the range of other diagnostic tests available. Three papers were identified for inclusion into this Knowledge Summary that addressed the PICO question, two of which reported the results from multiple diagnostic methods on animals designated for slaughter at different slaughterhouses, with the third (Palmer et al., 2014) reporting the findings of coproantigen ELISA testing on known positive faecal egg samples when compared to faecal samples obtained from F. hepatica-free areas.

Whilst all studies reported a coproantigen ELISA test specificity equal to or greater than that of the faecal egg sedimentation test, the sensitivity was significantly increased in only one study (Charlier et al., 2008). Mazeri et al. (2016) exhibited an increased sensitivity in one out of three sampling periods and an equivalent sensitivity in another period, and Palmer et al. (2014) demonstrated an increase in test sensitivity of 80% compared to the reference value of 70%.

Charlier et al. (2008) conducted a convenience sampling methodology, sampling the first 10–12 cattle that met the inclusion criteria and allowing for up to 3 cattle from the same herd to be sampled. A relatively small sample size was selected for each sampling period, potentially reducing the power of the study and opening the study to error. Similar to the previous study, inclusion criteria were outlined for the sampled cattle and sampling was conducted over multiple periods, allowing for seasonal variation to be accounted for but as this period is only limited to one year it does not account for variations in the lifecycle of F. hepatica.

The sample populations for Mazeri et al. (2016) and Charlier et al. (2008) are conducted using clinically healthy cattle presented for slaughter. These cattle are therefore likely to be well conditioned and unlikely to be carrying significant/chronic fluke burdens. As such they may not be representative of the cattle that would be tested in clinical scenarios, therefore sensitivity of the faecal sedimentation test is likely to be low when compared to the sensitivity of the coproantigen ELISA test as it has been shown to be accurate down to burden of only one fluke.

Palmer et al. (2014) provides a weak body of evidence for this Knowledge Summary and does not entirely address the PICO question. The quality of evidence supplied is reduced by; the case control sampling method, a relatively small positive sample size, failure to include relevant inclusion criteria of submitted faecal samples such as age/grazing status, failure to include confidence intervals when reporting test accuracy, and the modification of the test protocol midway through the study.

Palmer et al. (2014), Mazeri et al. (2016), and Charlier et al. (2008) are open to several sources of validity issues. Namely, they carried an unknown history of flukicide treatment at the time of sampling, potentially affecting the results of each diagnostic test, and an unknown parasite status at the time of sampling. Additionally, geographical location, management, and grazing strategies are not specified in the inclusion criteria of any of the three studies.

All three studies reported thorough faecal collection, storage, and test protocols, including relevant cut-off values for positive results. Additionally, all studies were performed with populations naturally infected with F. hepatica and were therefore more akin to findings identified in clinical practice.

In conclusion, the evidence reviewed provides a moderate argument for utilising the coproantigen ELISA but is not sufficient to be able to recommend a total replacement of the faecal egg sedimentation test for the diagnosis of F. hepatica in cattle just yet, due to the relatively insufficient body of evidence available and limitations of the studies as discussed. This is an economically important disease within the livestock industry and as such the basis of evidence requires significant further development and expansion before any clear conclusions can be drawn due to multiple confounding factors that influence the results of the diagnostic tests. It is; however, important to note the significant role the coproantigen ELISA has to play as an adjunctive test when diagnosing pre-patent infections with F. hepatica, and additional testing should always be considered when faced with cases of high suspicion testing negative.

Methodology

Search strategy

 

Exclusion / inclusion criteria

 

Search outcome

 

ORCiD

Jake Collyer: https://orcid.org/0009-0008-8147-7789

Conflict of interest

The author declares no conflicts of interest.

References

1. Anderson, N., Luong, T.T., Vo, N.G., Bui, K.L., Smooker, P.M. & Spithill, T.W. (1999). The sensitivity and specificity of two methods for detecting Fasciola infections in cattle. Veterinary Parasitology. 83(1), 15–24. DOI: https://doi.org/10.1016/S0304-4017(99)00026-6
2. Arifin, M.I., Höglund, J. & Novobilský, A. (2016). Comparison of molecular and conventional methods for the diagnosis of Fasciola hepatica infection in the field. Veterinary Parasitology. 232, 8–11. DOI: https://doi.org/10.1016/J.VETPAR.2016.11.003
3. Beesley, N.J., Caminade, C., Charlier, J., Flynn, R.J., Hodgkinson, J.E., Martinez-Moreno, A., Martinez-Valladares, M., Perez, J., Rinaldi, L. & Williams, D.J.L. (2018). Fasciola and fasciolosis in ruminants in Europe: Identifying research needs. Transbound and Emerging Diseases. 65(S1), 199–216. DOI: https://doi.org/10.1111/TBED.12682
4. Cawdery, M.J., Strickland, K.L., Conway, A. & Crowe, P.J. (1977). Production Effects of Liver Fluke in Cattle I. the Effects of Infection on Liveweight Gain, Feed Intake and Food Conversion Efficiency in Beef Cattle. British Veterinary Journal. 133(2), 145–159. DOI: https://doi.org/10.1016/S0007-1935(17)34136-2
5. Charlier, J., De Meulemeester, L., Claerebout, E., Williams, D. & Vercruysse, J. (2008). Qualitative and quantitative evaluation of coprological and serological techniques for the diagnosis of fasciolosis in cattle. Veterinary Parasitology. 153(1–2), 44–51. DOI: https://doi.org/10.1016/J.VETPAR.2008.01.035
6. Charlier, J., Soenen, K., De Roeck, E., Hantson, W., Ducheyne, E., Van Coillie, F., De Wulf, R., Hendrickx, G. & Vercruysse, J. (2014). Longitudinal study on the temporal and micro-spatial distribution of Galba truncatula in four farms in Belgium as a base for small-scale risk mapping of Fasciola hepatica. Parasites Vectors. 7(528), 1–8. DOI: https://doi.org/10.1186/S13071-014-0528-0
7. Graham-Brown, J., Williams, D.J.L., Skuce, P., Zadoks, R.N., Dawes, S., Swales, H. & Van Dijk, J. (2019). Composite Fasciola hepatica faecal egg sedimentation test for cattle. Veterinary Record. 184(19), 589–589. DOI: https://doi.org/10.1136/VR.105128
8. Kajugu, P.E., Hanna, R.E.B., Edgar, H.W., Forster, F.I., Malone, F.E., Brennan, G.P., Fairweather, I., 2012. Specificity of a coproantigen ELISA test for fasciolosis: lack of cross-reactivity with Paramphistomum cervi and Taenia hydatigena. Veterinary Record. 171(20), 502–502. DOI: https://doi.org/10.1136/VR.101041
9. Kajugu, P.E., Hanna, R.E.B., Edgar, H.W., McMahon, C., Cooper, M., Gordon, A., Barley, J.P., Malone, F.E., Brennan, G.P. & Fairweather, I. (2015). Fasciola hepatica: Specificity of a coproantigen ELISA test for diagnosis of fasciolosis in faecal samples from cattle and sheep concurrently infected with gastrointestinal nematodes, coccidians and/or rumen flukes (paramphistomes), under field conditions. Veterinary Parasitology. 212(3–4), 181–187. DOI: https://doi.org/10.1016/J.VETPAR.2015.07.018
10. Mazeri, S., Rydevik, G., Handel, I., Bronsvoort, B.M.d. & Sargison, N. (2017). Estimation of the impact of Fasciola hepatica infection on time taken for UK beef cattle to reach slaughter weight. Scientific Reports. 7, 7319, 1–15. DOI: https://doi.org/10.1038/s41598-017-07396-1
11. Mazeri, S., Sargison, N., Kelly, R.F., Bronsvoort, B.M.d.. & Handel, I. (2016). Evaluation of the Performance of Five Diagnostic Tests for Fasciola hepatica Infection in Naturally Infected Cattle Using a Bayesian No Gold Standard Approach. PLOS One. 11(8), e0161621. DOI: https://doi.org/10.1371/JOURNAL.PONE.0161621
12. Mezo, M., González-Warleta, M., Carro, C. & Ubeira, F.M. (2004). An ultrasensitive capture ELISA for detection of Fasciola hepatica coproantigens in sheep and cattle using a new monoclonal antibody (MM3). Journal of Parasitology. 90(4), 845–852. DOI: https://doi.org/10.1645/GE-192R
13. Palmer, D.G., Lyon, J., Palmer, M.A. & Forshaw, D. (2014). Evaluation of a copro-antigen ELISA to detect Fasciola hepatica infection in sheep, cattle and horses. Australian Veterinary Journal. 92(9), 357–361. DOI: https://doi.org/10.1111/AVJ.12224
14. Rapsch, C., Schweizer, G., Grimm, F., Kohler, L., Bauer, C., Deplazes, P., Braun, U. & Torgerson, P.R. (2006). Estimating the true prevalence of Fasciola hepatica in cattle slaughtered in Switzerland in the absence of an absolute diagnostic test. International Journal of Parasitology. 36 (10–11), 1153–1158. DOI: https://doi.org/10.1016/J.IJPARA.2006.06.001
15. Reigate, C., Williams, H.W., Denwood, M.J., Morphew, R.M., Thomas, E.R. & Brophy, P.M. (2021). Evaluation of two Fasciola hepatica faecal egg counting protocols in sheep and cattle. Veterinary Parasitology. 294, 109435. https://doi.org/10.1016/J.VETPAR.2021.109435
16. Skuce, P.J. & Zadoks, R.N. (2013). Liver fluke - A growing threat to UK livestock production. Cattle Practice 21(2), 138–149.