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International Journal for Parasitology: Parasites and Wildlife logoLink to International Journal for Parasitology: Parasites and Wildlife
. 2026 Feb 28;29:101214. doi: 10.1016/j.ijppaw.2026.101214

Comparison of wet mount, Mini-FLOTAC and Midi Parasep® coprological techniques for the detection of Crenosoma striatum, Capillaria spp. and Brachylaemus erinacei in European hedgehogs (Erinaceuseuropaeus)

Xiaoran Wu a,⁎, Paul Gilmore a, Anna Sturaro b, Steve Bexton c, Beverley Panto b, Rebecca C Hoyle a, Julian Chantrey d, Yannick Van de Weyer b,d
PMCID: PMC12969349  PMID: 41809673

Abstract

Parasitism, particularly Crenosoma striatum infection, can cause significant morbidity and mortality in European hedgehogs (Erinaceus europaeus). Wet mount is a coprological technique, which despite its relatively low sensitivity, is widely used in practice to detect the presence of endoparasites. The aims of this study were to compare the sensitivities of three coprological techniques and to determine a sensitive, yet practical method that could improve the detection of C. striatum infections in rehabilitating hedgehogs. Faecal samples were collected during autumn from 56 hedgehogs, and each analysed by wet mount, Mini-FLOTAC and Midi Parasep® (A sedimentation by centrifugation technique) to identify C. striatum first stage larvae (L1), Capillaria spp. eggs and Brachylaemus erinacei eggs. Mini-FLOTAC (Se = 98.0%) was significantly more sensitive than both wet mount (Se = 66.7%) and Midi Parasep® (Se = 58.8%) in detecting C. striatum L1 (p < 0.001). Midi Parasep® detected more samples as positive for Capillaria spp. (47/53) and Brachylaemus erinacei eggs (6/53) than Mini-FLOTAC (39/53; 3/53) though is impractical for use in a wildlife rehabilitation setting. The risk of obtaining false negative results when attempting to detect C. striatum L1 and Capillaria spp. eggs in hedgehog faeces could be reduced by using Mini-FLOTAC as an alternative to or in conjunction with wet mount; demonstrating that Mini-FLOTAC is highly sensitive in detection of C. striatum L1 and that it could provide an improved basis for management of crenosomosis in practice.

Keywords: Hedgehog rehabilitation, Crenosomosis, Faecal analyses, Lungworm, Fluke

Graphical abstract

Image 1

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Highlights

  • •

    Mini-FLOTAC is a highly sensitive technique for detecting C. striatum L1.

  • •

    Mini-FLOTAC is more sensitive than wet mount and Mini Parasep for detecting C. striatum.

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    Management of C. striatum can be improved by using Mini-FLOTAC in practice.

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    Mini-FLOTAC maybe less beneficial for management of Capillaria spp. and B. erinacei.

1. Introduction

The European hedgehog (Erinaceus europaeus) is categorised as “Near threatened” by the International Union for Conservation of Nature and the UK population has experienced rapid declines in previous decades (Gazzard and Rasmussen, 2023). They are a common and charismatic wildlife species native to Western Europe; orphaned, injured, or ill animals are frequently presented for rehabilitation (Bexton and Couper, 2019). Parasitism is common in wild hedgehogs, and it can contribute significantly towards morbidity and mortality (Bexton and Couper, 2019; Gaglio et al., 2010; Lehmann et al., 2024). Several endoparasites including nematodes e.g. Capillaria spp., trematodes e.g. Brachylaemus erinacei (B. erinacei), cestodes e.g. Hymenolepis erinaceus, Acanthocephala e.g. Oliganthorhynchus erinacei and protozoa e.g. Cryptosporidium parvum have been identified in hedgehogs (Rasmussen et al., 2021). Those responsible for causing verminous pneumonia such as Crenosoma striatum (C. striatum) are considered among the most detrimental to the animal's health (Lehmann et al., 2024; Rasmussen et al., 2021; Van de Weyer et al., 2023).

C. striatum is a hedgehog specific metastrongylid nematode that effects the lower airways (Lehmann et al., 2024) and its prevalence can be as high as 77% (Schütte et al., 2025). Hedgehogs are infected by ingesting third stage larvae via an infected gastropod intermediate host. Adult worms are found in the bronchi and bronchiolar lumens, producing first stage larvae (L1) which migrate to the lung parenchyma, causing bronchopneumonia with associated clinical signs such as a moist cough, weight loss, dyspnoea, crackly breathing and wheezing (Bexton and Couper, 2019; Lehmann et al., 2024). The L1 are expelled from the lungs, swallowed and shed in the faeces from approximately 21 days post infection (Baruš and Blažek, 1971; Beck, 2007). Therefore, infection can be detected in live animals through the identification of L1 via faecal examination or broncho-alveolar lavage (Cousquer, 2004; Gaglio et al., 2010) though the latter is rarely used due to its invasive nature.

European hedgehogs can be infected with multiple capillarid species including Capillaria aerophila, Capillaria erinacei and Capillaria putorii (Rasmussen et al., 2021; Schütte et al., 2025) with one UK study recording a prevalence of 81% for Capillaria spp. in rehabilitating hedgehogs weighing ≥200 g (Van de Weyer et al., 2023). Another study from Germany recorded a prevalence of 27% for C. aerophila and 68% for C. erinacei and C. putorii (Schütte et al., 2025). Capillaria aerophila contributes towards verminous pneumonia while the latter two affect the gastrointestinal system (Gaglio et al., 2010; Lehmann et al., 2024; Schütte et al., 2025). B. erinacei is a host-specific trematode described as infecting the small intestine and causing clinical signs such as restlessness, weight loss and haemorrhagic enteritis (Beck, 2007; Naem et al., 2015; Vasiliu et al., 2025). Brachylaemus aetechini and Brachylaemus mackoi have been identified in European hedgehogs in Italy (Casanova and Ribas, 2004) however their presence has not been recorded in Great Britain. Both Capillaria spp. and B. erinacei eggs are shed in faeces, enabling infection to be detected through coprological analysis, albeit differential speciation of Capillaria spp. eggs can be challenging based on morphology alone.

The Baermann's migration technique (BT) represents the classical gold standard for detecting Crenosoma L1 (Barutzki and Schaper, 2009). It is a low-cost technique which can produce preparations with minimal background debris and requires little effort and equipment (Tintori et al., 2022). While the BT is frequently used in a clinical context, it relies on the migration of viable larvae meaning ≥12 h are required before the larvae can be obtained and analysed, which can delay treatment.

The wet mount (WM) is frequently used in wildlife practice as it is accessible, low cost and quick (Bexton and Couper, 2019; Van de Weyer et al., 2023). Unfortunately, WM has relatively low sensitivity (Hussein et al., 2017; Nikolay et al., 2014) which can lead to false negative results with infections being missed.

The Midi Parasep® faecal parasite concentrator (Apacor limited UK) (PS) is an analogous derivative of the original centrifugation/sedimentation technique developed by Telemann (1908). This commercially available device follows the principals of the parasite concentration techniques that are routinely used in laboratories for parasite concentration and detection, such as Formalin-Ethyl Acetate Sedimentation and Ridley-Allen formol ether sedimentation (Garcia, 2010; Manser et al., 2016; Ridley and Allen. 1970; Soares et al., 2020). Unlike the BT, PS can detect non-viable larvae as it does not depend on larvae motility and therefore, less effected by the freshness of the sample. Its use and efficacy are well established (Manser et al., 2016) and it has outperformed the Ridley-Allen formol ether sedimentation technique for detecting gastrointestinal helminths (Abdel Aziz et al., 2020). The PS is an enclosed disposable system and therefore has a lower risk of hazard exposure and improved ease of use than the traditional centrifugation/sedimentation techniques (Manser et al., 2016).

The Mini-FLOTAC (MF) is an adaptation of the FLOTAC technique which has been used for detecting C. vulpis in dogs (Rinaldi et al., 2007) and mitigates the need for centrifugation and high-cost laboratory facilities. It has promising applications for use in resource limited, field settings (Barda et al., 2013) and has successfully been used to detect a variety of endoparasites, including gastrointestinal helminths, protozoa, and lungworm (Barda et al., 2013; Cringoli et al., 2017; Ianniello et al., 2020). It has demonstrated high sensitivities in comparison to other faecal analysis methods (Cringoli et al., 2017; Ianniello et al., 2020; Mohammedsalih et al., 2025). We therefore hypothesised that both PS and MF have a higher sensitivity than WM for detecting C. striatum, B. erinacei and Capillaria spp.

The aim of this study is to determine a more sensitive yet practical larval detection method that can help clinicians improve the management of crenosomosis in hedgehogs. This study compares the sensitivities of three faecal analysis methods: Wet mount, Mini-Flotac and Midi Parasep®. The protocols followed in this study (such as flotation fluid choice) were selected based on suitability towards the detection of C. striatum, the primary target, but the presence of Capillaria spp. eggs and B. erinacei. eggs were also recorded.

2. Material and methods

2.1. Faecal sampling and collection

Faecal samples were obtained from wild hedgehogs in temporary captivity and under veterinary care at either Stapley Grange or East Winch RSPCA wildlife centres between November 11th, 2024 to December 9th, 2024. Inclusion criteria for sampling included new admissions (<48 h), weighing ≥175g, and the ability to collect ≥1g of faeces for transport. Animals with a history of previous anthelmintic administration were excluded from the study. Samples were collected non-invasively by RSPCA staff and analysed using the WM method by the corresponding wildlife veterinarian of the centre at which it was collected. Subsequently, faecal samples were packaged in bubbled envelopes and refrigerated until the end of the day when they were transported via 1st class post to the parasitology laboratory (Leahurst campus, University of Liverpool). Upon receipt, they were stored at 4 °C until analyses by the MF and PS methods were conducted. BT, often suggested as the gold standard technique for lungworm, was not used in this study because of its reliance on live motile larvae, which could not be guaranteed with the study design logistics.

2.2. Wet mount

The wet mount method was performed on-site by the veterinary surgeon at each wildlife centre within 12 h of faecal sampling. Instructions were provided to the veterinarians to place a rice-grain-sized amount of faeces on a glass microscope slide, apply up to 3 drops of saline directly onto the slide with a Pasteur pipette, then mix the faeces with the water to obtain a monolayer. One preparation was made per sample and analysed with brightfield, compound microscopy at 100× magnification for 60 s (to reflect the real-world scenario), whereby 400× magnification was used when more detail was required to make an accurate diagnosis. The findings of the WM (positive/negative for C. striatum, Capillaria spp. and B. erinacei.) were noted on a form, packaged and posted with the corresponding faecal sample, to be recorded by the primary researcher.

2.3. Mini-FLOTAC

The MF method was performed at the Leahurst parasitology laboratory within 7 days of sample collection, having been stored at 4 °C upon receipt. Analysis was conducted based on the method described by Cringoli et al. (2017), using a 1.200 specific gravity zinc sulphate flotation solution as recommended for C. vulpis by the manufacturer. Each sample was first mixed until homogenous. Then the faeces and flotation solution were measured into the fill-FLOTAC at a 1:10 dilution ratio and suspended by rotating and pumping the device 10 times. Due to the relatively small volume of the samples, 0.5g of each faecal sample was measured and suspended with 4.5 ml of flotation solution, maintaining the suggested dilution ratio. The suspension was inverted 5 times before being dispensed into both flotation chambers of the mini-FLOTAC cassette at a 45° angle. A single cassette was prepared for each sample. After 10 min, the key was used to turn the reading disk 90° clockwise, which transferred the floated material from the chambers to the grids on the reading disk. Both grids were analysed under brightfield compound microscopy at 400× magnification.

2.4. Midi Parasep® concentration

Midi Parasep® Faecal Parasite Concentrators produced by © Apacor Limited UK 2023, were used for the PS method which was performed at the Leahurst laboratories following manufacturer guidelines and as previously described (Apacor, 2023; Manser et al., 2016). Due to the small amount of sample obtained per individual, 0.5 (±0.05) g of faeces was used for analysis. All samples were suspended in 10% formalin fixative and-ethyl acetate solvent solutions as soon as possible and at a very maximum of 3 weeks post collection, centrifuged and resuspended in 0.9% saline. One drop (∼0.05 ml) of the suspension was subsequently placed on a glass slide using a Pasteur pipette and covered with a 22 mm × 22 mm glass cover slip for brightfield, compound microscopy.

2.5. Parasite identification

All sample preparations were assessed for the presence of C. striatum L1, Capillaria spp. eggs and B. erinacei eggs. Parasites were identified morphologically based on previous literature (Baruš and Blažek, 1971; Beck, 2007; Mariacher et al., 2021) using brightfield compound microscopy at 100x and 400× magnifications. The presence of other parasite species such as Cryptosporidium parvum were outside the scope of this study and therefore were not recorded.

2.6. Additional calculations and statistical analysis

A power analysis was conducted (Sample size calculator) prior to sampling.

The sensitivities of each technique were calculated using the following formula:

Sensitivityofx=numberoftestpositives(x)numberoftestpositives(MFand/orPS)

The total number of positive samples detected by either the MF or PS was considered the ‘gold standard’ in this study. Samples testing positive by WM only, were not included in the prevalence due to its low specificity and interpersonal variation. Contingency tables and McNemar statistical tests were carried out to compare sensitivities of each technique. Statistical analyses were performed using IBM SPSS statistics (29.0.1.0) with a p-value of <0.05 considered as significant.

Ethical approval

Ethical approval (number VREC1533) was granted by the University of Liverpool ethics committee.

3. Results

3.1. Samples collected

A total of 56 faecal samples from different individuals were collected based on the inclusion criteria and were each analysed using all three methods. RSPCA Stapeley Grange and East Winch wildlife centres collected 31 and 25 samples respectively. Two samples were analysed by MF beyond 7 days of collection and another sample lacked WM data, excluding them from further analyses; the final cohort was comprised of 53 samples.

3.2. Parasite identification

All parasites were identified morphologically during microscopic examination. Crenosoma striatum L1 was identified through presence of a sharp, pointed posterior tail (Baruš and Blažek, 1971) (Fig. 1a). Eggs were identified as B. erinacei (Fig. 1b) and Capillaria spp. (Fig. 1c) based primarily on the presence of a thick shell with a single operculum and small aboperculate protuberance or, a lemon-shaped appearance with two asymmetrical bipolar plugs respectively (Beck, 2007; Mariacher et al., 2021).

Fig. 1.

Fig. 1

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Light micrographs of European hedgehog endoparasites detected via Midi Parasep coprological analysis: (a) C. striatum L1 with obvious straight-pointed tail, (b) B. erinacei egg, asymmetrically oval in shape with a thick, smooth, singularly operculated shell, and small aboperculate protuberance, (c) Capillaria spp. egg with two characteristic, asymmetrical bipolar plugs.

3.3. Parasite prevalence

A faecal sample was considered positive based on the identification of at least 1 larva (C. striatum) or 1 egg (B. erinacei/Capillaria spp.) detected by either MF or PS methods. The prevalence of each respective taxon was then determined based on the proportion of positive samples to the total number of processed samples. For all three parasites, samples testing positive by WM method only were not included due to the relatively low sensitivity of the test when performed in a busy wildlife rehabilitation setting, and uncommon occurrence of false positives. A prevalence of 96.2% (51/53) for C. striatum; 94.3% (50/53) for Capillaria spp. and 11.3% (6/53) for B. erinacei was found (Fig. 2).

Fig. 2.

Fig. 2

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Table summarising the number of faecal samples detected as positive for L1 C. striatum, Capillaria spp. eggs and B. erinacei eggs, by different methods (total number of samples N = 53). The ‘Maximum total’ is the number of samples detected as positive by either of the three methods. The prevalence is determined by the number of samples detected as positive in either MF or PS. Samples tested positive by WM only are not included in the prevalence.

3.4. Method sensitivity

The sensitivity of each method varied depending on the parasite taxon, with MF having the highest sensitivity for C. striatum and detecting the most samples as positive overall (Fig. 2, Fig. 3).

Fig. 3.

Fig. 3

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Venn diagram displaying the frequency of samples detected as positive for Crenosoma striatum L1 for the Mini-FLOTAC, Wet mount and Midi Parasep®. Values included in the green outline were included in prevalence.

3.4.1. C. striatum

MF had the best detection rate for C. striatum L1, identifying 50/53 (94.4%) of samples as positive. WM and PS detected C. striatum L1 in 35/53 (66.0%) and 30/53 (56.6%) of samples respectively (Fig. 2). MF (sensitivity = 98.0%) was significantly more sensitive than both the WM (sensitivity = 66.7%) and PS (sensitivity = 58.8%) (p < 0.001) (Fig. 4). However, there was no significant difference between the sensitivity of WM and PS (p = 0.383).

Fig. 4.

Fig. 4

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Table summarising the sensitivities and confidence intervals for the Mini-FLOTAC (MF) Wet mount (WM) and Midi Parasep® (PS) for detecting Crenosoma striatum in European hedgehog (Erinaceus europaeus) faecal samples.

3.4.2. Other parasites

All three detection methods successfully identified both Capillaria spp. and B. erinacei eggs, though to varying degrees of sensitivity. PS detected Capillaria spp. eggs in 47/53 samples and was the most sensitive (47/50, 94%). MF followed with a sensitivity of 78% (39/50), detecting Capillaria spp. eggs in 39/53 samples. Morphological speciation of Capillaria spp. eggs was not attempted. The WM detected Capillaria spp. eggs in 29/53 samples and had a sensitivity of 58% (29/50). WM, MF and PS detected B. erinacei eggs in 12.5% (7/53), 5.4% (3/53) and 10.7% (6/53) of the samples respectively.

4. Discussion

This study highlights MF as a useful tool for detecting C. striatum L1 in European hedgehogs. It produced a higher sensitivity than the frequently used WM method (Bexton and Couper, 2019) and PS: a formol centrifugation/sedimentation technique, used commonly in clinical parasitology laboratories (Abdel Aziz et al., 2020; Manser et al., 2016; Trunant et al., 1981; Young et al., 1979). The high level of sensitivity produced by MF in this study supports previous comparisons of MF with various other faecal analyses methods including flotation, McMaster, centrifugation/sedimentation and direct smears in detecting other metastrongylid and trematode species (Alowanou et al., 2021; Barda et al., 2013; Bosco et al., 2023; Ianniello et al., 2020; Mohammedsalih et al., 2025). Moreover, MF was shown to be particularly sensitive for infections with high parasitic load (Nikolay et al., 2014), which is often the case for C. striatum infections (Schütte et al., 2025). Successful detection of C. striatum L1 and other metastrogylid larvae using MF and FLOTAC has been previously reported (Cringoli et al., 2017; Ianniello et al., 2020; Rinaldi et al., 2007) and this study provides further validation for such use. Our results also demonstrate that MF can detect multiple parasite taxa as well as mixed parasitic infections (Cringoli et al., 2017) in European hedgehogs. The grids within each reading chamber of MF also allow for larval quantification, which can be useful for assessing parasitic load and monitoring treatment efficacy. In contrast to MF, WM and PS are not quantifiable and therefore unsuitable for monitoring larvae per gram (LPG) changes. The McMaster is a quantifiable method that has previously been used to detect C. striatum in hedgehogs (Gaglio et al., 2010; Rasmussen et al., 2021) however its greater time requirements may limit its use in wildlife rehabilitation. Although McMaster and MF were not directly compared in this study, the sensitivity of MF and the prevalence of C. striatum obtained in this study surpassed that of McMaster (78%) as reported by Gaglio et al. (2010) and the prevalence of crenosomosis obtained in other studies (Rasmussen et al., 2021). This suggests superior sensitivity of the technique used in our study for detection of C. striatum, but it should be noted that seasonality (autumn) and selection criteria (e.g. weight of ≥175 g) may have affected the likelihood of detection and resulted in a higher prevalence. MF has also demonstrated a greater sensitivity than McMaster for detecting other helminth species (Alowanou et al., 2021; Mohammedsalih et al., 2025).

BT can also be used to evaluate LPG, is able to produce preparations with minimal debris and is less labour intensive than MF. However, results cannot be obtained for 12-36 h, potentially delaying treatment decisions in practice. Practical challenges such as sample transportation, its consequences on freshness and survival of motile larvae limited our ability to compare MF with BT, which is considered gold standard for detecting metastrongylids such as Crenosoma spp. (Ianniello et al., 2020). Nevertheless, it is possible MF may be preferred, as comparisons by Bauer et al. (2010), Rinaldi et al. (2007) and Schnyder et al. (2011) found BT to be less sensitive than other methods such as McMaster and zinc- or magnesium sulphate based FLOTAC in detecting other species of lungworm larvae. Similarly, the FLOTAC method, upon which MF is based, also outperformed BT in detecting Crenosoma vulpis in dogs (Rinaldi et al., 2007; Schnyder et al., 2011). Moreso, MF obtained higher LPG than BT for A. vasorum, A. abstrusus and Troglostrongylus spp. (Ianniello et al., 2020) however, additional studies specific to C. striatum are required as optimal diagnostic techniques can differ between parasites even of the same genus.

Although this study utilised MF in a laboratory setting, evaluations by Barda et al. (2013), Boelow et al. (2022) and Cringoli et al. (2017) have identified it as a practical tool for resource and time limited settings, supporting its use in wildlife centres. MF does not require centrifugation, a fume hood nor fresh faeces, having still been effective after 7 days post sample collection (Bauer et al., 2010). This could provide users with more flexibility, which may be useful in a busy wildlife hospital setting. Despite being more labour intensive and costly than the wet mount, MF may be superior due to its significantly higher sensitivity and ability to quantify parasitic load (Cringoli et al., 2017). For example, it can be applied to clinically ill animals with a strong suspicion of lungworm infection but negative WM results, or to investigate treatment-resistant infections. While the specificity of each method was not evaluated in this study, both MF and PS are likely to have a specificity near 100% as the parasites could be directly visualised and morphologically assessed. While PS produced preparations with the least debris, the final microscopic preparation obtained by MF may have less background contamination than WM, due to its 90° rotation mechanism (Mohammedsalih et al., 2025). This could reduce the risk of misidentification and produce more specific results than WM.

PS follows similar principals and has comparable sensitivity to other techniques such as the Ridley-Allen and formol-ether concentration methods (Abdel Aziz et al., 2020; Manser et al., 2016). Its compatibility with non-motile larvae made it the preferred method for use in this study. Traditional formalin centrifugation/sedimentation methods (on which PS is based) are highly effective and produced higher sensitivities than other methods such as WM for detecting intestinal helminths and protozoa (Agarwal et al., 2025; Hussein et al., 2017; Soares et al., 2020; Young et al., 1979). However, this has not been investigated for detection of C. striatum with PS specifically and its relatively low sensitivity obtained in this study is unexpected as PS utilises centrifugation which should concentrate and improve the recovery of parasites from the samples (Garcia, 2010; Manser et al., 2016; Trunant et al., 1981). One explanation could be the longer interval between faecal sampling and PS analyses compared to the other techniques meaning that structural integrity and detectability of the larvae may have been affected (Schnyder et al., 2011). Albeit no morphological evidence of larval disintegration was observed, simultaneous analyses of each technique would have been ideal but this was hampered by logistical challenges. PS is more complex and prone to human error compared to WM and MF techniques which are considered easy to perform with relatively low risk of technical errors (Garcia, 2010; Young et al., 1979). Both sedimentation and flotation techniques are procedures for concentrating parasites in faecal samples (Garcia, 2010), and previous studies have recorded poorer sensitivities in the former (Bosco et al., 2023; Cringoli et al., 2017). While PS has a reduced risk of hazard exposure and is less labour intensive than traditional formol centrifugation/sedimentation techniques, a centrifuge is required and a fume cupboard is recommended (Manser et al., 2016) limiting its use in a wildlife rehabilitation setting. Moreover, costs and the low sensitivity demonstrated in this study do not support the use of PS for the detection of C. striatum.

Despite being less sensitive than MF (Agarwal et al., 2025; Barda et al., 2013; Van de Weyer et al., 2023), WM is cheap and quick and may therefore remain a useful tool in practice; particularly for when MF is unavailable or unaffordable. This study's sensitivity of WM for diagnosing crenosomosis (66.7%) was still relatively good and improvements in its sensitivity can be achieved through repeated sampling of individual animals (Bexton and Couper, 2019) or by taking more time to assess each sample. It should be noted that not all samples detected as positive by PS and WM were identified as positive by MF. These false negatives may be because the larval load was too low to be detected by MF and other methods may be more suitable for levels of infection below 5 LPG (Ianniello et al., 2020). The BT may also be a useful tool as it is affordable, requires low effort, and produces preparations with minimal debris. However, the sensitivity of this method was not investigated in this study, due to practical limitations (i.e. wildlife veterinarians didn't have the time to perform and interpret the technique on-site).

All methods produced positive results in samples that tested negative in the alternatives used in this study (Fig. 3) showing that each can be valuable for detecting C. striatum L1. The use of molecular detection methods such as PCR analysis may have been valuable, providing an alternative gold standard which would have allowed better evaluation of both sensitivity and specificity. This would have also enabled for respiratory and gastrointestinal Capillaria spp. To be distinguished from each other (Schütte et al., 2025). However, this was outside the budget of the present study.

The time elapsed between sampling and faecal analysis for the MF and especially the PS techniques is a limitation of this study that may have affected the reliability of parasite detection. For example, detection of live Dictyocaulus viviparous larvae from bovine faeces was reduced to 74% after 3 weeks of storage at 3-6 °C (Rose, 1956). Another study of small ruminant faeces stored at 4 °C noted a marked reduction in nematode egg counts only after the third week of storage (Drimtzia and Papadopoulos, 2018). Therefore, some uncertainly remains as there may be differences between parasitic species. Apart from postage, hedgehog faecal samples were kept refrigerated at 4 °C. The short period (often ≤36 h) without refrigeration during transport may have affected larval integrity and consequently detectability, though low ambient temperatures (average of 7.3 °C) during the study period minimise the likelihood of significant degradation. Moreover, our applied techniques do not require larvae to be alive, detection of a single larvae or egg is sufficient for a positive result and morphological evidence of larval or egg disintegration during microscopy was not observed for either technique. Hedgehogs are often affected by high Crenosoma larval burdens (Schütte et al., 2025) and therefore mild to moderate reduction of larval counts due to storage are more likely to impact the reliability of quantitative results but could still enable a reliable qualitative result (positive/negative) in our study. More samples may have been detected as positive if additional time was allocated for WM examination. However, quantification was not required for WM, time in clinical practice is often limited and the number of hedgehog patients in autumn is high. As such the 60 s period was determined to reflect the real-world scenario. Due to the subjective nature of faecal analyses and microscopy, the results may be affected by interpersonal variation and user experience. Nevertheless, each faecal analyst was experienced, received the same protocols and identification criteria thereby reducing these effects.

As other fluke species have not been described in UK hedgehogs, B. erinacei was the most likely species to be detected, and detailed comparisons of species-specific fluke morphology was not routinely performed resulting in some degree of presumption in its diagnosis. Our results suggest that the MF is not optimal for detecting B. erinacei as both WM and PS identified more samples as positive for this trematode. WM identified most samples as positive for B. erinacei. However, WM is associated with abundant background material which, combined with the small size of B. erinacei eggs, may uncommonly result in misidentification, especially when interpreted by inexperienced rehabilitators. Sedimentation methods have been suggested for diagnosing B. erinacei. (Beck, 2007). Contrastingly, MF has successfully detected other trematode species, producing better sensitivity than sedimentation methods (Bosco et al., 2023) albeit a higher specific gravity flotation solution was utilised than in this present study. It is possible that the low sensitivity for fluke seen in this study was because the chosen flotation solution (determined for Crenosoma spp.) was not of sufficient specific gravity to adequately float B. erinacei eggs. The same could be posited regarding Capillaria spp. eggs. Further research investigating different flotation solutions may be useful to find the optimal solutions for detection of Capillaria spp and B. erinacei.

5. Conclusions

This study demonstrates MF is a sensitive yet practical coprological method that can help improve the management of hedgehog crenosomosis in wildlife rehabilitation centres. All three techniques tested in this study successfully detected C. striatum L1, Capillaria spp. eggs and B. erinacei eggs, though to varying degrees. Further parasitic stages were not detected, and other parasitic species were not recorded. BT, the recommended gold standard for detecting metastrongylid larvae, was not evaluated due to logistical challenges but it may nevertheless be a useful tool in practice. However, MF achieved high sensitivity (98%) for the detection of C. striatum and may be considered equally suitable or even superior than BT considering that it is not affected by larval viability. MF was significantly more sensitive for detecting C. striatum L1 than WM and PS while there was no significant difference between the latter two. Other methods may be more sensitive for detecting B. erinacei than MF, but further studies are required. MF can be a valuable tool for when animals presenting with clinical signs compatible with verminous pneumonia test negative for C. striatum on WM, to more reliably rule out C. striatum infections and inform vets of treatment efficacy. When used alongside clinical examinations and responses to supportive therapy, MF can guide treatment decisions and promote responsible anthelmintic use.

CRediT authorship contribution statement

Xiaoran Wu: Writing – review & editing, Writing – original draft, Software, Methodology, Investigation, Formal analysis, Data curation, Conceptualization. Paul Gilmore: Methodology, Conceptualization. Anna Sturaro: Investigation. Steve Bexton: Writing – review & editing, Investigation. Beverley Panto: Investigation. Rebecca C. Hoyle: Supervision, Resources. Julian Chantrey: Supervision, Resources, Methodology, Investigation, Conceptualization. Yannick Van de Weyer: Writing – review & editing, Methodology, Investigation, Conceptualization.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

We sincerely thank the Stapeley Grange and East Winch RSPCA wildlife centres for their collaboration and aid in sample collection.

This research was funded by the budget allocated for the intercalated BSc in Veterinary Conservation Medicine (XW) at the University of Liverpool. There was no external funding.

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