Determination of the Size of Maturity of the undatum in English Waters – Defra project MF0231 Andy Lawler Funded by Defra

Contents Determination of the Size of Maturity of the Whelk in English Waters – Defra project MF0231 ...... 0 1. Executive Summary ...... 2 2. Introduction ...... 3 3. Objectives...... 4 4. Methods ...... 4 4.1 Size of maturity ...... 4 4.2 Ageing and growth ...... 8 4.3 Parasitological ...... 9 5. Results ...... 9 5.1 Size of maturity ...... 9 5.1.2. Alternative method of male maturity determination ...... 15 5.2 Ageing and growth ...... 16 5.2.1 Reliability of opercula age determination ...... 16 5.2.2 Growth Model fitting ...... 19 5.2.3. Alternative ageing method ...... 24 5.3 Parasitological ...... 25 6. Discussion ...... 26 6.1. Size of maturity ...... 26 6.2. Ageing and growth ...... 28 6.3. Parasitological ...... 29 7. Conclusions ...... 29 7.1. Implications ...... 30 7.2. Recommendations for future work ...... 30 8. References ...... 31 9. Acknowledgements ...... 33 Appendix 1 ...... 34

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1. Executive Summary

This study has provided estimates of size of maturity (SOM) for in ten important English fisheries using visual observation of the gonad for maturity determination. The potential of using opercula and statolith ring counting methods to age whelks was investigated and variability between alternative readers for the widely used opercula counting (OC) method was summarised. Von Bertalanffy growth models (VBGM) were fitted to length and age data derived by the OC method to provide provisional and plausible growth estimates for each of the ten areas. Estimates of SOM and growth parameters were compared with two sites sampled using identical methodology in a previous study.

Samples of catch were sourced from ten English ports chosen in consideration of the economic value of recent and historic reported landings of whelks. Fishers provided samples from sites typically exploited by the local fishery comprising suitable numbers and size ranges of whelks, enabling precise estimates of SOM. Because of the time consuming nature of the sample processing, only one sample was acquired and analysed from each site. Individual whelks from size and sex stratified sub-samples from each site were measured (shell height), removed from their shell and the maturity status and gender recorded. The (trap door) of each whelk was removed for age analysis. Occurrences of host whelks with atypical reproductive development, or sterility, caused by a parasite were recorded. Statoliths from a small number of whelks were removed after digestion of the body in a caustic solution, polished and their rings counted as an alternative approach to OC.

The probability of a whelk being mature with size was estimated using logistic regression analysis, and the size at which the probability was 0.5 (the definition of SOM) was estimated for both sexes at each site. Results were generally consistent with previous work where corresponding sites had been sampled. However, earlier studies often used different methodologies and differences between results were not necessarily explained by regional variation alone. Estimates of SOM from this study ranged between 44.8 mm and 46.4 mm shell height (female and male, respectively) for a site in the Solent (Portsmouth) and 77.8 mm and 76.2 mm (female and male, respectively) for a site in the (Bridlington). Estimates of SOM generally fell into three groups; those around 70mm and above (Bridlington-North Sea, Exmouth-Western English Channel, Ilfracombe-Celtic Sea and Whitehaven-Irish Sea), those between about 50 to 65mm (Whitstable, Poole, Selsey, Ramsgate and Weymouth in the English Channel and Wells-next-the-Sea – Southern North Sea), and that around 45mm (Portsmouth-Solent). In general estimates of SOM by gender were similar (<5% difference), but at some sites differences were about 10% (e.g. Eastbourne where females had a higher SOM than males and Selsey and Weymouth where SOM for females was lower than for males.

Four scientists independently counted the rings on a sample of approximately 500 whelk opercula, stratified by size group, sex and site. Variability between readers as deviations from the mode were summarised and percentage agreement for each reader calculated using a similar method to that used by colleagues investigating variability in age determination in fin- using otoliths. The highest percentage agreement achieved by one of the readers was close to that typically observed for difficult to age fin-fish such as whiting Merlangius merlangus using the otoliths (79.8% c.f. ~ 80%). The other readers achieved lower percentage agreement scores (63.5-72.8%) highlighting a need for improved methodology or higher levels of expertise. Statoliths isolated from a small sample of whelks already aged by the OC method were polished and rings counted. Despite lack of agreement with the counts from the OC method, rings in the statoliths were on occasion clearly defined and this and uncertainty with the OC method suggests that the statoliths warrant further investigation using improved procedures.

VBGMs fitted to the age and size data by least squares methods provided parameter estimates of L and K generally consistent with earlier work at corresponding sites. Growth models fitted to the data suggest growth for whelks did not vary significantly by gender but varied considerably between sites. Fitted models for whelks

2 from the Portsmouth site exhibited a lower growth rate than other sites (L , 62.62mm and K, 0.41) whilst that for whelks from the Bridlington site was the highest (L , 130.73mm and K, 0.23). These provisional growth estimates suggest that whelks are likely to attain the EU Minimum Landing Size (MLS) of 45mm in as little as 2 years except in the Solent (Portsmouth) where this may take 3 years. When combined with estimates of SOM this suggests that the whelks from Eastbourne and Ramsgate reach sexual maturity in 2 years, those from Portsmouth, Poole, Selsey, Wells-next-the-Sea, Whitstable and Weymouth in about 3 years, whilst those from the Bridlington, Exmouth, Whitehaven and Ilfracombe sites will take about 4 years.

The incidence of observations of atypical gonad development in host whelks caused by digenean parasites was reported as low (<0.5%), but paucity of infected animals and problems with sample fixation prevented positive identification of the species responsible.

The estimates of SOM for all sampled sites except the Solent (Portsmouth) indicated that the current EU MLS of 45mm does virtually nothing to protect spawning stocks. The MLS has a modest conservation value for Eastbourne and Ramsgate and a more significant one for the Solent (Portsmouth).

The implications of the SOM estimates were briefly discussed and recommendations made to evaluate alternative management scenarios, especially a range of potential MLS values. Such an analysis should highlight potential short term losses in yields and the magnitude of potential future gains in yields and the size of the spawning stock and will be needed to convince some members of the fishing industry currently landing large quantities of smaller whelks.

If an increased MLS strategy is shown to be beneficial, clearly a one size fits all approach is not suitable from a biological perspective, but regionally variable MLSs can be even more difficult to implement and enforce. Careful consideration of management options and their implications will therefore be required.

Future work should also include enhancements to determination of age and growth, in particular, improved sampling to provide a more complete range of whelk ages and more robust age determination including validation of the opercula ring counting method with appropriate consideration of individual reader performance.

2. Introduction

Whelk landings in England and were worth over £10million at first sale in 2012, but there are concerns amongst scientists and fisheries managers over the sustainability of the fisheries. Despite these concerns, at present formal stock assessments are not undertaken and the status of English fisheries is unknown and inconsistent with the Marine Strategy Framework Directive and the concept of Good Environmental Status.

The only regulation in most areas is the EU Minimum Landing Size (MLS) of 45mm shell height and current management measures may not be adequate to conserve local whelk stocks, especially if fishing effort were to increase as a result of displacement from other more regulated fisheries. A number of potential policy changes are under consideration, including increasing the MLS, but better selectivity in both fishing gears and on-board sorting devices would also be advantageous. Recently displaced fishing effort from another whelk fishery forced the Kent & Essex Inshore Fisheries Conservation Authority (K&EIFCA) to introduce an emergency byelaw and subsequently to revise there whelk fishery management regime. A permit scheme including pot limitations has been introduced, but alternative technical measures such as increasing the MLS have not. Sussex IFCA regulations include a byelaw which prohibits removal of whelks from the fishery that pass through a sorting device with a 25mm gap (there is a 10% tolerance based on weight).

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Recent collaborative work between the Centre for the environment, fisheries and aquaculture science (Cefas) and Sussex Sea Fisheries Committee (Sussex SFC, now SxIFCA) and funded by the Department for fisheries and rural affairs (Defra), has shown that in the Sussex fisheries, the size at which whelks mature is higher than the current MLS which therefore affords virtually no protection to the spawning stock. As the size at which whelks mature is known to vary regionally this work has been extended with further Defra funding to determine the size of maturity in other English fisheries. In addition to estimates of size at maturity this project investigated age determination methodology with the aim of providing growth rates, as both these data gaps hinder determination of stock status.

3. Objectives Primary aim:

1) To determine the size of maturity of whelks in different regions around the English coast a) Determine appropriate sampling sources based on current and historic fishing patterns and acquire adequate size and sex stratified samples. b) Process whelk samples and compile maturity data and other biometrics into a suitable database. c) Analyse data using logistic regression analysis to determine size of maturity by sex and determine regional variations.

Secondary aims:

2) To investigate the most reliable methodology to age whelks and determine the growth rate in different study regions. a) Investigate utility of opercula ring counting and alternative methods relying on physical characteristics of whelks. b) Apply the most appropriate technique explored in objective 2a, to the size stratified samples (by sex) to provide growth rates for each study region.

3) To isolate and identify the trematode parasites observed in a previous whelk study (collaborative project with Sussex SFC), screen for infestation to determine prevalence and likely impact on the spawning potential of the population. a) Identify species present by histological examination of dissected tissues. b) Screen size stratified samples for infestation and determine incidence of infection rate by region to determine likely impact on the spawning stock.

4. Methods

4.1 Size of maturity

Trends in annual landings of whelks by port summarised from the Fishing Activity Database (FAD, Marine Management Organisation, MMO) were used to inform appropriate sampling areas. Priority was given to high landings in recent periods but consideration was also given to historic data as whelk fisheries can be ephemeral in nature due to market forces and fluctuations in stock sizes. Ten ports were selected as sample sites; Bridlington (Yorkshire,), Wells-next-the-Sea (Norfolk), Whitstable and Ramsgate (Kent), Portsmouth (Hampshire), Poole (Dorset), Weymouth and Exmouth (South Devon), Ilfracombe (North Devon) and Whitehaven (Cumbria) (fig. 4.1.1). The fisheries in Sussex were considered adequately sampled from recent previous collaborative work by Cefas and Sussex SFC and funded by the Fisheries Science Partnership and Defra, which sampled the ports of Selsey and Eastbourne. No additional sampling was carried out for this

4 region but results from this previous work are presented and discussed with consideration of those from this study.

Figure 4.1.1 Map of sample sites, labels show subsequent port of landing.

Volunteer fishing skippers were enlisted with the help of local IFCAs or the MMO, and one sample of whelks was acquired from each of ten sampled ports. Samples were sourced over a two year period, with sampling times being restricted to a two month period in the early part of each year for consistency (16th Jan to 15th March). Fishers were asked to provide a sample of approximately 30kg of whelks based on the total pot content of their baited traps in an area typically fished by the local fleet. Two additional smaller quantities of commercial sized whelks and undersized whelks were requested to ensure adequate coverage of rarer sizes. These sites were where the main grounds were located and not always in close proximity to the landed port (e.g. Bridlington sample was captured off the North Norfolk/Lincolnshire coast, Fig 4.1.1). Samples were deep frozen until required. Size stratified sub samples for each sex were selected from each sample at the Cefas laboratory in Lowestoft.

Whelks were individually weighed and their shell height (from apex to end of , fig. 4.1.2) measured. Opercula were removed and stored in paper packets (otolith packets) for subsequent age analysis.

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Individual whelks were removed from their shells using forceps and a bench vice was used to crush the shell if required.

Figure 4.1.2. Shell height measurement

The development status of the reproductive organs and gender of each whelk was recorded according to table 4.1.1. Sexually immature were categorised as stage I or stage II depending on the absence or presence of gonad tissue on the dorsal edge of the digestive gland (where stage I refers to no gonad tissue and stage II where an immature gonad is beginning to develop but present only as a thin strip of tissue). Sexually mature animals were defined as possessing a significant gonad assumed to be functional. Whelks with atypical gonad or digestive gland tissue as a result of parasite infestation were excluded from the analysis of size of maturity. Photographs of the dissected whelks were taken and archived (examples included in fig. 4.1.3).

Table 4.1.1. Maturity stage key by sex. dg: digestive gland

Maturity stage Male Female I (immature) No differentiation of dg No differentiation of dg II (maturing) Some differentiation of dg Some differentiation of dg restricted to dorsal and anterior restricted to dorsal edge edge III (mature) Presence of a mature testis Presence of a mature ovary (typically brick red in colour and (typically yellow in colour and thicker than 2mm in depth) thicker than 4mm in depth)

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Figure 4.1.3. Photographs of mature (top) and immature (bottom) male (LH) and female (RH) whelks with shells removed

The size of maturity (SOM) is defined by the size at which 50% of the population is mature (equivalent to probability=0.5). Maturity can be considered a binomial process where a whelk is either mature or immature and the probability of a whelk being sexually mature by size and by sex and site was therefore modelled using binomial logistic regression analysis carried out with the glm function within the R statistical modelling framework (R Development Core Team, 2012, Crawley, 2007). This gives the model: p=ea+bx/1+ea+bx where p is the probability of a whelk being mature, x is the shell height of the whelk and a and b are parameters defining the exact shape of the curve.

The resulting S shaped plots of the modelled proportion mature against size are commonly called maturity ogives. Plots of residual and predicted values were used to confirm adequate model fits and data integrity. The SOM and associated 95% confidence intervals were reported. Analyses of deviance were used to test for differences between sexes and sites and the SOM and associated 95% confidence intervals were reported.

For male whelks, standardised penis length has been used as an indicator of sexual maturity in male whelks. This metric was calculated from data collected during this project for comparison with other studies and the gonad based estimates of SOM.

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4.2 Ageing and growth Size and sex stratified sub samples of opercula from whelks from each site were cleaned in fresh water and flattened in small paper envelopes. Various methods were used in an attempt to increase the contrast between striae (growth rings) assumed laid down annually and those formed at more frequent intervals. These were staining (methylene blue), histological clearing agent (Histo-Clear, Fisher Scientific), bleaching (household bleach) and the use of image analysis software on images of the opercula (ImageJ, National Institute of Health). However, additional treatments of the opercula provided no apparent benefit over cleaned and flattened specimens and use of these treatments was therefore abandoned.

Figure 4.2.1. Whelk opercula showing relatively clearly defined growth rings (assumed annual)

To investigate the reliability of the age determination process, four scientists independently counted rings on each operculum. Data for opercula where at least two readers did not agree on the number of rings or where two pairs of readers agreed on different numbers of rings were rejected. Readers recorded the number of observed “annual striae” using a binocular microscope with a magnification of 10 and the specimen illuminated with transmitted light (Santarelli et al, 1985). For each operculum the modal number of rings from the four readings was calculated. Records for opercula where all four readers gave different numbers of rings or where two pairs of readers each agreed on different numbers of rings were reported and excluded from further analyses based on the modal number of rings as they did not have a clear mode. Counts by reader were then summarised by comparing with the modal count assumed to be the true number of rings using methods similar to those used by Cefas and other agencies when assessing the reliability of fin fish age determination using otoliths (Etherton, 2012). The square root of the standardised sum of the squared residuals from the mode (analogous to a standard deviation, “SD”) and this metric further standardised by the mode (analogous to a coefficient of variation, “CV”) were calculated for each modal age.

The age of each whelk was assumed to be: modal number of striae - x in decimal years, where x is the correction in years to allow for sampling earlier in the year than an assumed birth date in April. Von Bertalanffy model (VBGM) growth curves were fitted using the R statistical modelling framework to the observed data by minimising least squares (Sparre et al, 1998, Ricker, 1975, Ogle, 2011). In order to improve the estimation process, the original VBGM parameterisation was used and L0 (the length at age zero) was assumed known, resulting in fewer parameters to determine (Cailliet 2006).

-Kt L(t) = L - (L - L0)e

8 where L is length at time t, L L infinity or asymptotic length is the average maximum length of whelks, K is the Brody growth rate coefficient and L0 (L zero) is the length at time zero (assumed 3mm, Martel et al, 1986). Bootstrapping was used to provide confidence intervals around the model predictions by random sampling of the original observations.

The growth parameter phi prime ( ’), which provides a general indication of growth potential, was calculated by ’ = LogK + 2Log L (Pauly and Munro, 1984). Age data collected for an earlier project carried out off Sussex (Eastbourne and Selsey) and during the same time of the year (January and February) are included for comparison.

In an attempt to validate the opercula ageing method the statoliths were removed from the bodies of a small sample of whelks that had previously been aged by opercula counting. Statoliths are small, hard calcified structures located as a pair in the body of the whelk in an organ called the statocyst and their internal structure is thought to show growth rings similar to fish otoliths (see fig. 5.2.3.1 in results). Using a procedure similar to Richardson (Richardson et al, 2005) the statoliths were glued to microscope slides and polished to reveal the internal ring structure. These were counted under a binocular microscope with the specimens illuminated with transmitted light (magnification x200). The resulting ages were tabulated and compared to the ages obtained from the opercula.

4.3 Parasitological

Whilst whelk samples from the ten sites were being processed for reproductive studies, tissues were visually inspected using an illuminating magnifying lens to detect the presence of parasite infestation. Specimens of infected host tissue were deep frozen or preserved in buffered formaldehyde solution and sent to parasitologists at the Cefas Weymouth laboratory for further examination.

5. Results

5.1 Size of maturity

Just fewer than 4000 whelks were dissected and their reproductive status determined. Results from an additional 500 whelks, a subset of data from the earlier study in the Sussex area and sampled over the same sampling period, were presented and compared alongside those from this project.

A logistic model (maturity v size) fitted to the data for all sites and sexes combined, showed a strong relationship between the probability of a whelk being mature and whelk size. The model explained 33% of the deviance in these data (residual deviance was 3827 on 4416 degrees of freedom).

A second more complex model including site and sex as factors (with interactions) explained 59% of the deviance in these data (Res. Dev. was 2340 on 4370 df). A 2 test on the change in deviance and the computed Akaike information criterion suggested this more complex model was more appropriate than the simpler model and diagnostic plots did not indicate any fitting problems or lack of compliance with model assumptions. Analysis of deviance showed that the maturity ogives differed between some of the sites ( 2, Pr<0.001), and that the relationship between the probability of maturity and size differed between sites (significant size and site interaction, 2, Pr<0.001). The gender of the whelks as a main effect in the model was not significant, but the interaction between sites and sex was ( 2, Pr<0.001). Subsequently, maturity ogives were presented separately for each site and by sex (fig. 5.1.1).

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Figure 5.1.1. Probability of a whelk being mature against whelk size (i.e. maturity ogive) by sex and site. Confidence intervals omitted for clarity

Estimates of SOM with confidence intervals and the observed smallest mature whelk and largest immature whelk are tabulated by port and sex (table 5.1.1). The largest immature whelk occurred in the samples from Whitehaven (88.6mm female) and the smallest mature whelk occurred in the Ramsgate sample (36.6mm male). For comparison estimates of SOM as estimated by Bell and Walker in a previous study are presented in table 5.1.2 (Bell and Walker, 1998). Estimates of SOM with the confidence intervals are also presented in figure 5.1.3.

Table 5.1.1. Estimates of SOM with 95% Confidence intervals, size (shell height mm) of smallest mature whelk and largest immature whelk and sample size (N) by site and sex

SOM 95% CI Smallest Largest Size at 50% mature immature Site Sex N mature Lower Upper observed observed F 121 77.8 75.0 80.6 69.7 86.1 Bridlington M 156 76.2 73. 8 78.75 72.5 86.1 F 114 56.7 54.4 59.0 47.1 65.5 Eastbourne M 129 51.2 48.6 53.7 40.5 70.6 F 110 72.4 70.4 74.4 66.1 80.3 Exmouth M 135 69.2 67.2 71.3 54.1 76.9 Ilfracombe F 189 75.5 73.8 77.2 66.0 86.8

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M 159 75.5 74.0 76.9 55.4 78.6 F 159 63.5 61.5 65.5 56.4 78.8 Poole M 151 66.0 63.3 68.7 51.3 76.0 F 145 44.8 43.4 46.1 38.4 50.9 Portsmouth M 149 46.5 45.3 47.5 38.5 53.1 F 320 52.8 51.0 54.7 36.6 67.2 Ramsgate M 293 49.5 47.7 51.3 37.0 67.3 F 126 59.6 58.0 61.1 53.9 65.1 Selsey M 128 64.6 62.8 66.4 55.8 76.4 F 366 60.6 58.8 62.5 54.2 71.2 Wells M 467 62.5 60.8 64.2 54.9 70.7 F 173 54.7 53.1 56.4 46.8 62.0 Weymouth M 166 59.1 56.9 61.3 50.4 70.2 F 197 69.5 67.2 71.9 57.5 87.7 Whitehaven M 200 74.0 71.6 76.3 52.4 88.6 F 116 60.7 59.1 62.4 51.4 67.8 Whitstable M 148 61.9 60.3 63.6 50.0 74.4 Sum/mean 4417 62.7 60.7 64.7 52.4 73.3

Table 5.1.2. Estimates of SOM with 95% confidence intervals reproduced from Bell and Walker (1998)

Sampling station Size at 50% Lower 95% CI Upper 95% CI mature Scarborough 73.5 67.4 79.6 Ramsgate 60.4 58.8 62.1 Eastbourne 57.4 55.8 59.0 Newhaven 57.1 36.0 78.1 Worthing (chalk) 55.7 54.0 57.3 Worthing () 61.6 59.5 63.7 Selsey 45.2 40.4 49.9 Nabs Tower 55.1 51.8 58.5 Portsmouth 56.5 50.6 62.5 Saundersfoot 70.2 65.0 75.4 Fishguard 67.0 54.6 79.4

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Figure 5.1.2. Estimates of Size of maturity (black line) by site with upper and lower 95% confidence limits (red circles)

Maturity ogives by sex for each site are graphically presented with their confidence intervals along with the observed data summarised to proportion mature by 5mm size group and coerced onto the same horizontal axis (fig 5.1.3 and 5.1.4).

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Figure 5.1.3. Probability of female whelks being mature with whelk size (maturity ogive) by site. With SOM and 95% CIs of model fit. Circles are proportions mature in each 5mm size groupings. Solid reference lines correspond to 50% maturity and SOM. Dashed vertical reference line is the current EU MLS (45mm)

Confidence intervals for SOM estimates were all reasonably narrow (typically within ±2mm), but those for female whelks from the Bridlington site were ±2.8mm. There were marked regional differences between estimates of SOM (range 44.8 to 77.8 mm, females Portsmouth and Bridlington). The difference between estimates of SOM by gender within port of sampling was much smaller (range 0.05mm to 4.4mm for Ilfracombe and Whitehaven, but higher for the samples from Selsey and Eastbourne from an earlier project). The significant interaction term for sex and site in the analysis of deviance is explained in table 5.1.1, with the female estimate of SOM being higher than the males for the Eastbourne site, but lower than the males for the Selsey and Weymouth sites.

Estimates of SOM for Portsmouth (44.8mm female and 46.4mm male) were significantly lower than for the other sites, and those for males from Ramsgate and Eastbourne were also low compared to the other sites. Estimates of SOM for Bridlington were the highest (76.2 and 77.8mm for males and females, respectively) and those from Ilfracombe were similar, as were estimates for male whelks from Whitehaven.

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Figure 5.1.4. Probability of male whelks being mature with whelk size (maturity ogive) by site. With SOM and 95% CIs of model fit. Circles are proportions mature in each 5mm size groupings. Solid reference lines correspond to 50% maturity and SOM. Dashed vertical reference line is the current EU MLS (45mm)

Parameters (i.e. slope and inetercept of the linear model) with confidence intervals for each logistic regression equation are presented in table 5.1.3. The low slopes fitted to the regression models for Whitehaven (0.17 female and 0.18 male) and Ramsgate (0.18 both sexes), and to a lesser extent Poole and Eastbourne, showed that whelks from these sites mature over a larger size range than those from the other sites which exhibit steeper gradients in the linear regression model.

Table 5.1.3. Model coefficients (with confidence intervals) from logistic regression analysis by port of sampling and sex.

95% CI 95% CI

Site Sex Intercept Lower Upper slope Lower Upper F -25.77 -15.50 -43.65 0.33 0.20 0.56 Bridlington M -25.18 -16.87 -37.48 0.33 0.22 0.49 F -11.93 -8.02 -16.79 0.21 0.14 0.30 Eastbourne M -9.85 -6.53 -13.92 0.19 0.13 0.27 Exmouth F -23.81 -15.55 -35.64 0.33 0.21 0.49

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M -18.20 -12.60 -25.55 0.26 0.18 0.37 F -37.77 -17.70 -32.65 0.32 0.23 0.43 Ilfracombe M -16.06 -24.94 -57.02 0.50 0.33 0.76 F -12.81 -11.31 -22.28 0.25 0.18 0.35 Poole M -15.51 -9.10 -17.51 0.19 0.14 0.27 F -20.82 -11.00 -21.04 0.35 0.25 0.47 Portsmouth M -9.77 -14.63 -28.76 0.45 0.32 0.62 F -9.77 -7.81 -12.10 0.18 0.15 0.23 Ramsgate M -8.97 -7.11 -11.15 0.18 0.14 0.23 F -24.45 -16.91 -34.62 0.41 0.28 0.58 Selsey M -21.04 -14.50 -29.90 0.33 0.22 0.46 F -20.57 -14.61 -25.93 0.32 0.24 0.43 Wells M -18.61 -15.61 -27.59 0.33 0.25 0.44 F -18.61 -13.57 -25.35 0.34 0.25 0.46 Weymouth M -12.42 -9.13 -16.54 0.21 0.15 0.28 F -13.18 -9.12 -15.60 0.17 0.13 0.22 Whitehaven M -22.32 -9.95 -17.10 0.18 0.13 0.23 F -22.32 -15.18 -32.06 0.37 0.25 0.53 Whitstable M -21.10 -15.11 -29.02 0.34 0.24 0.47

5.1.2. Alternative method of male maturity determination

Figure 5.1.2.1. Relationship between penis length and shell height (top right panel with a log linear transformation, bottom left panel standardised by shell size). Red symbols are those designated as mature by visual inspection of the gonad

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Previous researchers have used a standardised penis length index as an indicator of sexual maturity in male whelks. It has been suggested that a ratio of penis length to shell height of 0.5 or more was indicative of mature whelks (Bell et al, 1998). The relationship between penis length and shell height for mature and immature male whelks can be described by two different log linear regression lines (figure 5.1.2.1). The ratio of penis length to shell height plotted against shell size shows that an index of >=0.5 provides similar, but not identical, results to those where maturity determination was by visual inspection of the gonad.

5.2 Ageing and growth

5.2.1 Reliability of opercula age determination

Plots of deviations from the mode against whelk size for each reader and by site showed a marked level of inconsistency between readers and sites (figure 5.2.1.1 and table 5.2.1.1). Of 508 opercula read, 121 were rejected because the four readers did not produce a clear likely age. This occurred when the ring counts were split between two modes (i.e. two readers agreed on one age and the other two agreed on another) or no readers agreed on a ring count. For Bridlington, Exmouth and Ilfracombe there was a tendency for the deviations to increase with whelk size, but for the other sampling ports the relationship with whelk size was less obvious. The most extreme deviations occurred in the Ilfracombe sample with two readers providing ring counts 4 years higher than the mode for two different whelks. The other sites showed fewer extreme deviations from the mode, with most values within + or -1.

Reader 1 (red) was systematically high over most sites (except Wells, Poole, Portsmouth and Whitehaven). Reader 2 (green) was lower than the mode in most cases (e.g. Bridlington, Eastbourne, Exmouth, Ramsgate, Selsey, Weymouth, Whitehaven and Whitstable) but higher than the mode for the Poole site and variable without bias for Portsmouth and Ilfracombe. Reader 3 (blue) appeared to count high for the Whitehaven site, but low for Eastbourne and Exmouth and with low bias for the other sites. Reader 4 (black) tended to over count in the Eastbourne, Exmouth and Wells sites but under count for Bridlington and Portsmouth. At the other sites reader 4 did not appear to show any obvious bias.

Table 5.2.1.1. Number of opercula read (N1), those with unanimous ageing by all four readers, number where the reading was split, opercula when all readers disagreed and the resulting number of opercula used in the subsequent analysis (N2).

all Split no Modal Site N1 agree decision agree N2 range Bridlington 61 11 10 1 50 1-7 Eastbourne 41 2 13 1 26 1-5 Exmouth 46 2 18 3 25 2-6 Ilfracombe 26 0 6 0 20 3-6 Poole 38 10 5 1 32 1-5 Portsmouth 27 2 6 0 21 2-6 Ramsgate 47 4 10 0 37 1-5 Selsey 36 3 9 0 27 2-5 Wells 56 14 10 0 46 1-6 Weymouth 46 9 8 1 37 1-5 Whitehaven 56 5 12 0 45 1-6 Whitstable 28 4 7 0 21 2-6 508 66 114 7 387

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Figure 5.2.1.1. Deviations from the modal ring count against whelk size for readers (R1-4) by site. A small vertical offset has been introduced to enable visualisation of all observations.

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A summary of the deviances from the mode by reader, for all sites combined, showed that for reader 1 (red), most deviations were due to over counting (figure 5.2.1.2). Reader 2 (green) had a lower incidence of conforming to the mode and deviations were more often due to under counting. Reader 3 (blue) had the highest agreement with the mode with no obvious systematic bias overall. Systematic bias over all sites was also not obvious for reader 4 (black). Percentage agreement with the mode was calculated as 72.8, 63.5, 79.8 and 68.2% for readers 1 to 4, respectively.

Figure 5.2.1.2. Summary of the consistency of performance for each of the four readers (frequency of deviations from the mode with percentage agreement with the mode). All sites and both sexes combined.

The square root of the standardised sum of the squared residuals from the mode (analogous to standard deviation, “SD”) and this metric further standardised by the mode (analogous to a coefficient of variation, “CV”) plotted against modal counts (assumed true age) gives rather ambiguous results (fig. 5.2.1.3). The “SD” metric suggests that reader variability increases with the modal ring count in most sites (except Exmouth, Ilfracombe and Poole). The “CV” metric decreases with increasing modal count for all sites except Whitstable. This indicates that the error in reading tends to increase with increasing age, but at a slower rate than the increase in age. It may also reflect the generally low range and absolute level of ages sampled in this study i.e. no very old whelks were sampled.

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Figure 5.2.1.3. Variation (as two standardised metrics) in ring counts by mode and site (filled circles “SD”, empty circles “CV”)

5.2.2 Growth Model fitting

The original parameterisation of the VBGM fitted to all data by sex fitted moderately well (residual standard error of 9.50 on 385df) although the two observations at the oldest ages and majority of observations in the youngest age group were generally not well described. The fitted models predicted lower mean sizes than observed in the data for ages below 1 and over 6 years old (two points only) (figure 5.2.2.1). Results from ANOVA and comparison of AIC scores show that parameter estimates for both L and K are not significantly different for each gender and subsequent analysis therefore considers models fitted to data for both sexes combined. However, it should also be noted that the data used for fitting growth curves were relatively poor, comprising a relatively narrow age range, with potentially significant error in aging and providing little information on either the curvature in the model or the level of asymptotic length (L .

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Figure 5.2.2.1. Size against age for all sites combined with fitted lines by gender and both sexes combined

Three models allowing one or both of the fitted parameters to vary between sites and a fourth with common parameter estimates were compared by ANOVA and AIC scores and a model which had different values of L for each site was the most parsimonious, but a model with a variable K gave a similar result. The high negative correlation between the two parameters inherent in the VBGM is demonstrated in figure 5.2.2.2. Subsequent analysis was carried out by fitting the original parameterisation of the VBGM to each site separately.

Figure 5.2.2.2. Example of correlation between the VBGM parameters L and generated from bootstrapping

The model which had different values of L and for each site, fitted well, with residual standard error of 7.51 on 363 df. A plot of residuals against fitted values for the model fitted by site suggests that the errors do not show pronounced heteroscedasticity or other significant problems, but their distribution did exhibit a modest negative skew (figure 5.2.2.3). Given the issues concerning data uncertainty with respect to ageing

20 methodology this model seems adequate and provided plausible parameter estimates for all sites.

Figure 5.2.2.3. Residual against fitted values and distribution of the residuals for the VBGM fitted

The growth curve fitted to the Portsmouth site was distinctly different to the other sites with an obvious lower asymptote (figure 5.2.2.4). The VBGM fitted to the Whitstable data also suggested a lower growth rate over older ages than the other sites. The VBGM for Bridlington and Eastbourne gave the highest and very similar growth rates over most of the age range.

Figure 5.2.2.4. VBGM fitted by site, horizontal reference line at current 45mm EU MLS

21

Individual model fits for each site with associated approximate confidence intervals for the model fit and observations (generated by bootstrapping methods) showed variations in the range of observations, their relative consistency and how successfully they were described by the respective fitted models (figure 5.2.2.5). The range of ages present in the samples was 7 for the Bridlington sample, 6 for Wells and Whitehaven, only 4 for Ilfracombe and Selsey and 5 for the remaining sites (see also table 5.2.1.1). If the age estimates are correct, 0 group whelks were missing from the Exmouth, Whitstable, Portsmouth and Selsey samples and both 0 group and 1 group whelks were not present in the Ilfracombe sample.

Figure 5.2.2.5. The VBGMs fitted by site for both sexes combined. Dashed blue lines are the 95% CIs for the mean length at age and red dashed lines 95% prediction interval (both approximations from bootstrapping). Dashed horizontal reference line is current EU MLS (45mm)

Parameter estimates are presented in table 5.2.2.1 by port of sampling and with those from some earlier studies for comparison. The other studies used the more traditional parameterisation of the VBGM (Beverton

1954; Beverton and Holt 1957) which includes t0 (time at zero size) rather than L0, however the parameters L and K are equivalent so comparison of results is informative. Estimates of L from this study were in the range of 62.62 to 130.73, for the Brody growth coefficient K 0.20 to 0.41, and the growth parameter ’ ranged

22 from 7.39 to 8.25. Where corresponding areas have been studied the estimates from this study are generally consistent with the earlier studies.

Table 5.2.2.1. Estimates of growth parameters from this and other studies

Geographic area Sex L mm K t0 ’ Source This study Bridlington Combined 130.73 0.23 8.25 Cefas Eastbourne 129.79 0.23 8.25 Exmouth 97.13 0.35 8.10 Ilfracombe 129.68 0.23 8.25 Poole 106.01 0.30 8.13 Portsmouth 62.62 0.41 7.39 Ramsgate 121.20 0.20 8.00 Selsey 107.34 0.27 8.03 Wells 116.67 0.27 8.20 Weymouth 102.56 0.27 7.94 Whitstable 107.39 0.30 8.16 Whitehaven 88.38 0.39 8.02 Earlier studies Irish Sea (IOM) 123.7-125 0.2-0.22 Kideys 1991 English Channel 85.5-139.5 0.71-0.18 -1.06 -0.13 Santarelli 1985 SW Irish Sea 106 0.133 -0.959 7.31 Fahy 2000 NW (Cape) 114 0.192 -0.435 7.82 Celtic Sea (Helvic) 106 0.261 1.271 7.98 Shetland F 101.10 0.39 3.19 8.29 Shelmerdine 2007 M 102.04 0.40 3.20 8.33 Shetland (East) F 104.87 0.30 2.17 8.10 M 99.02 0.39 2.66 8.25 Shetland (West) F -185.67 -0.03 -3.65 M 157.52 0.09 -0.32 7.71 South of England 66.05 0.39 0.96 7.44 Cefas Whitstable 64.7-68.7 0.34-1.02 Hancock 1963 English Channel 73 0.221 7.07 Heude-Berthelin 2011

Table 5.2.2.2. 95% Confidence intervals for parameter estimates, number of observations and square root of the estimated variance of the random error (sigma)

L 95% CIs K 95% CIs Site Lower Upper Lower Upper N Sigma Bridlington 109.44 169.52 0.15 0.31 50 8.87 Eastbourne 93.59 263.18 0.09 0.39 26 7.63 Exmouth 83.06 123.88 0.22 0.52 25 8.70 Ilfracombe 105.28 181.96 0.13 0.34 20 6.63 Poole 82.71 171.63 0.15 0.47 32 7.49 Portsmouth 53.80 81.31 0.25 0.67 21 6.54 Ramsgate 93.76 189.71 0.11 0.31 37 6.90 Selsey 91.18 137 0.18 0.36 27 4.53 Wells 100.89 143.06 0.20 0.35 46 6.76

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Weymouth 85.56 136.44 0.17 0.38 37 7.10 Whitehaven 91.48 139.74 0.20 0.42 45 8.90 Whitstable 75.52 113.71 0.25 0.57 21 6.86

5.2.3. Alternative ageing method

Due to the time consuming nature of the extraction and preparation processes statolith removal was attempted on only 13 individual whelks. Of these attempts at least one statolith was successfully removed from 8 individual whelks giving 13 statoliths. Only 8 provided visible rings in the polished statolith (table 5.2.3.1). This highlights the labour intensive nature of this work and a requirement for improved methodology to provide consistent results on a routine basis. In no instance did he estimate of age from the statolith agree with the opercula ring count.

Table 5.2.3.1. Details of 8 whelks where statoliths were successfully removed. Includes whelk size, gender, number of statoliths removed and ring counts on statoliths and opercula. U denotes rings not visible

Ring count Statoliths Shell height mm Sex Opercula removed Statolith Statolith 1 2 75.9 F 2 7 7 5 76.8 M 2 U U 6 57.8 F 1 5 - 3 52.8 M 2 U 4 3 51.9 M 2 3 3 5 42.1 F 2 3 U 2 35.2 M 1 U - 1 34.4 F 1 - 4 2

Following on from this work, Cefas will fund a PhD student based at Bangor University to further investigate the potential of this method. Along with other biological investigations the student will use elemental isotope ratio methods in an attempt to verify both opercula and statolith ring counts. Various images of prepared statoliths are presented in figure 5.2.3.1.

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Figure 5.2.3.1 Images of polished statoliths, lower panels following image enhancement. Approx. magnification x 200

5.3 Parasitological

Photographs of an infected male whelk and that of a normal male whelk of comparable size are presented for comparison (fig. 5.3.1). Only 23 whelks with atypical parasitized gonad and digestive glands were observed in the maturity samples. This equates to less than 0.5% of those animals inspected.

Figure 5.3.1. Photographs of two male whelks (LH with atypical development, RH normal)

Parasitologists at the Cefas Weymouth laboratory were unable to isolate and positively identify the digenean parasite thought to prevent maturation in immature whelks or cause sterility in mature individuals. Inadequate

25 fixation of samples and the paucity of infected material were considered the likely cause. A review of the current understanding of Buccinum parasites is included in Appendix 1.

6. Discussion

6.1. Size of maturity

The common whelk is ubiquitous in UK waters and estimates of SOM have been carried out by various agencies. Studies have utilised different methodologies to determine maturation status, with samples often sourced at different times of the year and with results often restricted to “grey literature”. Earlier studies often show marked variation in the SOM of whelks sourced from different regions (Bell and Walker, 1998, Heude- Berthelin et al, 2011, Henderson and Simpson, 2006). It is unclear how much of the observed regional variation is real or due to different sampling methodologies.

The aim of this current study therefore, was to apply a common methodology to the determination process, for both sexes and to samples sourced from areas relevant to the location of current and recent fisheries.

6.1.1. Seasonal variation of SOM estimates - sampling bias and seasonally varying errors It is likely samples sourced at the same sampling location but during different seasons can give variable results. It is not unreasonable to assume that the seasonal timing of the sampling process can give genuine differences in SOM estimates, but there is also the scope for sampling bias which may vary seasonally. Samples taken by baited traps are reliant on the behavioural activity of the target species and may not be representative of the population. A well known example occurs in the edible crab fishery where mature female crabs are thought not to enter traps after spawning leading to a disproportionate number of immature females to be taken in the catch during the spawning and brooding period. It is not known if whelk traps take representative samples of the immature and mature components of the population over the course of a complete fishing season or during the winter months when samples for this study were collected.

There is the possibility that a spent can be confused with an immature individual and vice versa. Incorrect determination of sexual maturity may also vary seasonally as whelks which are ripe and ready to spawn will be obviously mature due to the presence of large and brightly coloured gonads, whereas the maturation state of spent individuals may not be so obvious.

Although in this study we standardised the timing of sampling in both sampling years to the first quarter to reduce the likelihood of seasonal bias and provide more comparable estimates of SOM, seasonal patterns in the lifecycle of whelks around the UK and between years are not necessarily synchronous.

6.1.2. Alternative age determination The Directorate of Fisheries Research (now Cefas) investigated regional variations in SOM of whelks around England and Wales over 15 years ago (Bell and Walker, 1998). The study used relative penis size to determine sexual maturity in males only, at sites primarily off the South Eastern English coast, with some regional context provided by additional samples from Scarborough and South Wales.

Plots of a penis index (penis length/shell height) with whelk size and by maturation status determined by visual inspection of the gonad from this study, shows that an index of >=0.5 used by the earlier study gives a reasonable indication of male sexual maturity (figure 5.1.2.1). However, results from this latest study show that estimates of SOM can vary by gender and estimates should be determined for each sex.

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The authors of the earlier study had some sampling problems, which led to some unconvincing estimates in some of the study areas. Nonetheless, there are similarities between these earlier results and those presented in this report where corresponding areas were sampled. Perhaps of more interest are the inconsistencies noted in the areas adjacent to the Isle of White. Whereas the lowest estimate of SOM from this study was observed in a sample taken in the Solent and landed into Portsmouth the lowest estimate in the earlier study was from a sample sourced off Selsey approximately 25 miles to the south east. This is evidence of regional variation over a relatively fine spatial scale.

6.1.3. Regional variation Other researchers have described local variations in SOM, growth rates and other biological traits, and most accept there are marked regional differences due to whelk stock structure (Bell and Walker, 1998. Valentinsson et al, 1999. Fahy et al, 2000). We noted there were differences between the shape of whelk shells and the length/weight relationships between sampled sites for this study (not shown). It has been suggested that the observed differences are due to water temperature differences especially summer water temperatures evident along the North South axis of the UK (Bell and Walker, 1998) but other factors such as predation or fishing pressure have also been suggested as possible causes (Fahy et al, 2006). Locally distinct stocks may result from genetic isolation over fine geographic scales and despite the widespread distribution of Buccinum undatum low adult mobility and a lack of dispersive plankton egg or larval stages will limit genetic mixing. A genetic study investigating stock identity and genetic diversity showed differences over large geographic scales, but results presented in this report suggests further genetic studies over smaller spatial scales may provide further useful context (Weetman et al, 2006).

6.1.4. Repeatability of results We would have preferred to take replicate samples in each fishery and over the course of a fishing season to investigate the repeatability of our estimates and fine scale spatial and seasonal variation. Unfortunately the labour intensive nature of the sample processing precluded this; instead we opted for one “good” sample in each area, a strategy compatible with available resources. Here we define “good” as containing suitable numbers of whelks over the required size range to provide an estimate of SOM with relatively tight confidence intervals. From these we were able to produce relatively precise estimates of SOM for each sample. As the fishers who provided our samples were requested to source the sample from within the main grounds within each fishery and to avoid areas where the size structure of the catch appears atypical we hope our samples are representative of the main fishing area in each fishery. Without replication this cannot be confirmed from these data. During the course of this project, Cefas held a workshop designed to share the methodology of sexual maturity determination with local IFCAs. The aim was that local agencies will be able to supplement sampling from this project in some key areas and that such sampling would provide results which are directly comparable with those presented here. Such sampling will therefore be able to determine the repeatability of our results.

6.1.5. Age of maturity Provisional growth rates presented in section 5.2.2 (figure 5.2.2.4) suggest that the age at which whelks achieve SOM varies between sites. Whereas the SOM for whelks sourced from Portsmouth is lower than those sourced from the other sites, they mature at around 3 years old, and comparable with those sourced from Poole, Selsey, Wells, Weymouth and Whitstable. Whelks in the samples landed at Bridlington, Exmouth, Ilfracombe and Whitehaven exhibited higher provisional growth rates, but the higher observed estimate of SOM suggests most mature in their 4th year. Whelks from Eastbourne and Ramsgate may reach sexual maturity in the 2nd year. We remind the reader of the provisional nature of our growth estimates.

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6.2. Ageing and growth

6.2.1. Reliability of opercula age determination The accepted method of age determination for whelks following validation by elemental isotope ratios by Santarelli and Gros, is to count the growth rings on the opercula (Santarelli and Gros, 1985). Whilst age determination of Irish Sea whelks by opercula ring counts were supported by results from length frequency analysis and a mark-recapture experiment they were not consistent with growth observed in aquaria (Kideys, 1996). This author also noted that the isotope ratio work carried out by Santarelli and Gros in 1985 was carried out on only 3 whelks, and they (Kideys) themselves indicated that for the Irish Sea whelks only 16% of opercula had “clear” striae (rings).

We frequently found age determination by the opercula ring counts to be inconsistent between our four readers, which led to a lack of confidence in this method amongst them. One suggested that where growth rings were not clearly defined, there was more agreement between readers, as they were reliant on using the size of the operculum as an indication of the age (not tested). Some authors stated that they adopted exclusion of those opercula where annual growth rings were not clearly defined. Although our methodology of excluding observations where no clear mode between readers was established is effectively the same, we would prefer not to exclude “unclear” opercula because of the possibility of introducing bias.

Our readers had experience in ageing other molluscs and reviews and mini workshops designed to improve the reader’s expertise were carried out before and during this project. We are not aware of any studies that present age data based on opercula ring counts that include an assessment of variability between alternative readers, which is inconsistent with the approach adopted by colleagues responsible for age determination in fin fish by otolith ring counting. Variation between readers of fin-fish otoliths is assessed and the relative reliability of each reader is summarised as a percentage agreement with the modal age (assumed true age, Etherton, 2012. See also ICES age reading reports at http://www.ices.dk/community/Pages/PGCCDBS-doc- repository.aspx#age). For context we note that typical % agreement for difficult to read finfish species, for example whiting (Merlangius merlangus), is 80%; although this is sometimes optimistic (Etherton, pers. Comm.). Our most consistent reader achieved a result comparable with this, but the other readers were less reliable, with the lowest % agreement of only 63.5%. We are prepared to accept that our scientists may have lacked specific whelk ageing expertise, especially at the start of this project, but suggest that this ageing method for whelks is potentially unreliable and that previous studies may not have given consideration to this source of uncertainty.

During the course of this project we had hoped to cooperate with Bangor University who have expertise in ageing gastropods using statoliths. Unfortunately this was not achieved during the lifespan of this project. We have trialled the methodology of this technique and it has provided some plausible results and we believe this method warrants additional attention. Recently, Cefas has decided to fund a PhD student based at Bangor University and Cefas Lowestoft who will investigate the potential of other ageing methods as well as other biological issues.

6.2.2. Growth model fitting Exploration of ageing techniques was a secondary aim and we relied on the samples sourced to provide SOM estimates to also provide growth estimates. Whereas the size distributions observed in the samples were suitable for providing information on maturation, a complete range of all ages was not represented in these samples. VBGMs are difficult to fit to data with limited age ranges and the very young and old whelks were missing from the samples. Gear selectivity probably accounted for the lack of young whelks and the rarity of older animals in the catch is probably due to mortality. We would like the opportunity to refine our growth estimates by supplementing the samples with these missing age groups, perhaps by deploying less selective fishing gear and asking fishers to collect very large whelks over a longer fishing period. Improved data would

28 improve the fitting of growth models, potentially removing the need to constrain parameters (i.e. a priori constraint of L0 was applied in this study).

This study made best use of the methods and data available and provisional growth models were fitted to age and size data. They suggest that whelks in our samples reach the current MLS of 45mm shell height in around 2 years, except in the Solent/Portsmouth sample where it was about 3 years. However, perhaps the most important contribution of the project’s age and growth studies is the account of “between reader” variability in age estimates. Future studies may also take into account “within reader” variability by getting the readers to read the same opercula on a number of separate occasions.

6.3. Parasitological

Another secondary aim was to screen for digenean parasites known to cause abnormalities in the reproductive development of Buccinum undatum, and to identify the species responsible. Not only does this parasite have the potential to reduce the reproductive potential of an infested whelk stock, it would also affect the results of this reproductive study if infected whelks were not excluded from the analysis. Including individuals with atypical development and designated as immature, but which without the parasite would have developed normally, would have the affect of increasing the estimate of SOM for the sample. We could argue that such a phenomenon should be accounted for in the estimates of SOM, but we decided that excluding infected whelks would provide a more meaningful result in terms of quantifying maturation rate and that sterility due to parasitism should be quantified, and if need be accounted for , separately. Although we cannot guarantee all infected whelks were removed from the analysis, those individuals with obvious symptoms were excluded (for example the lower than predicted proportions mature presented in the female 75 and 85mm size classes for Poole and Ilfracombe respectively may be due to inclusion of such individuals) (figure 5.1.4).

We were unable to identify the species of digenean fluke infecting the host whelks but we were successful in determining that the incidence of infection was very low and lower than anticipated at the onset of the project. We would recommend taking account of atypical reproductive development in any future studies on reproductive biology.

7. Conclusions

• There was considerable regional variation between estimates of SOM

• Estimates of SOM ranged from 44.8 mm (female) and 46.4 mm (male) taken in the Solent to 77.8 mm (female) and 76.2 mm (male) from the southern North Sea

• Estimates for SOM by gender are less variable but differ in some areas

• The EU Minimum Landing Size of 45mm has limited potential for protecting spawning stocks in all areas except the Solent

• Ageing whelks by counting annual growth rings is not straightforward and there is considerable potential for error and bias

• Counting rings on statoliths is time consuming but worth further investigation for verification or as an alternative to opercula ring counting

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• This project has provided plausible growth parameter estimates for all main English Fisheries but these would benefit from additional sampling over a wider range of size/age classes

• In the samples sourced for this study, the incidence of abnormal reproductive development caused by digenean parasites in host whelks, was very low (<0.5%)

• Data collected during the project would facilitate and inform evaluation of the impacts of EU or regional MLS changes using per recruit methods

7.1. Implications

The estimates of SOM for all sampled sites except the Solent (Portsmouth) indicate that the current EU MLS of 45mm does virtually nothing to protect spawning stocks. There is a modest conservation value of the MLS for Eastbourne and Ramsgate and a more significant one for the Solent (Portsmouth). The potential conservation value to the spawning stock of the current MLS in these three areas will depend on the size structure of the catch and fecundity at size. The contribution of the current MLS in reducing the risk of growth over fishing is not known and will require information on fishing mortality, natural mortality and growth estimates for quantification.

Pragmatically for any MLS to be effective adequate enforcement is required and the large volumes of the whelk landings will always make this problematic. There is also a market for small whelks close to the MLS and use of this technical measure as a management tool, either on its own, or in conjunction with other management measures may not be appropriate. Quantitative modelling (potentially per recruit) could provide a means to evaluate alternative management scenarios, especially a range of potential MLS values. Such an analysis should highlight the short term loss in yields and the magnitude of future gains and the size of the spawning stock and will be needed to convince some members of the fishing industry currently landing large quantities of smaller whelks.

If an increased MLS strategy is shown to be beneficial, clearly a one size fits all approach is not suitable from a biological perspective, but regionally variable MLSs can be even more difficult to implement and enforce. A recent study in the Sussex and Kent & Essex IFCA regions suggests that use of escape holes in whelk traps and the use of onboard sorting devices with appropriate grid spacing will reduce the numbers of undersized whelks in the catch and landings, and these technical measures will compliment appropriate MLS legislation (Lawler et al, 2012).

7.2. Recommendations for future work

Modelling would assess the potential of alternative management measures including different MLS scenarios and stock status monitoring would quantify the urgency of appropriate management measures.

Historically fishing activity and biological data acquisition for most fisheries has generally had lower priority than for quota species, with whelk fisheries being data poor even compared to other shellfish. Gaps in biological data and fishing activity trends need to be addressed before stock status can be determined and models meaningfully parameterised. The restricted number of merchants who trade in whelks compared to some other shellfish may facilitate a targeted, economic and successful biological sampling programme.

The age determination aspect of this project has provided provisional growth rates by area but further work needs to be carried out on the ageing methodology before robust growth rates, suitable for stock assessment

30 work can be estimated. Future work planned for a Cefas funded, Bangor University based, PhD student will provide additional information on age and growth, effective fecundity and other biological parameters.

Recent advancements with genetic markers may facilitate stock determination on a finer geographic scale than earlier work and we would recommend further genetic studies particularly in the sea areas in and adjacent to the Solent.

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9. Acknowledgements

This work was funded by the Department for Environment, Food and Rural Affairs (Defra). We would like to thank staff of the local Inshore Fisheries Conservation and Authorities (IFCA), the Marine Management Organisation (MMO) and members of the fishing industry who contributed to sample collection.

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Appendix 1

Digenean parasites of Buccinum undatum

Report prepared by Matt Longshaw, Population Health and Welfare team, Environment and Animal Health Group, Weymouth for Andy Lawler June 2012

This report has been written to provide an overview of the digenean parasites of whelks following concerns raised regarding their role in castration of whelks around the English coastline. The report briefly covers the lifecycles of typical digenean parasites and provides information on impacts of these parasites to individual and population health as well as zoonotic risks. Information on identification and treatment methods is provided. It provides detailed information on the types of parasites present in whelks including external symptoms of infections as well as information on effects previously reported in the literature. Finally, the report proposes a number of areas of potential future research that could be conducted on these parasites.

Digenean parasites (also known as trematodes or ) are common parasites of a number of vertebrate and hosts. Lifecycles are complex and usually involve at least three hosts - invertebrates (including molluscs and ) and vertebrates (fish, birds, mammals). In brief, are released from adult digeneans (which are hermaphroditic) and following a short period of further development in water, hatch to release a free swimming ciliated miracidium stage. This penetrates into a molluscan host and develops into a sac-like structure known as a mother sporocyst. Depending on the species of digenean, this then further develops into a daughter sporocyst or into a redia. Cercaria develop within the redia or daughter sporocyst. Cercaria are released from the molluscan host and actively swim to find a new host. In a small number of species the cercaria directly infect a final host and become adults. However, in most cases cercaria infect a second intermediate host and encyst as metacercaria within host tissues. Transmission to the final host usually occurs following ingestion of the second intermediate host.

A typical digenean lifecycle is shown below: Almost all organs in the host can be infected and in some cases, infections can lead to pathological changes, alterations in behaviour, loss of function of affected organ and death (Alda et al., 2010; Bass et al., 2007; Johnson and Hartson, 2009; Jonsson and André, 1992; Karvonen et al., 2010; Lafferty and Kuris, 2009; Malek, 2001). Some digeneans are recognised as important zoonotic agents, leading to wide spread issues in those areas where there is a predilection for consuming raw or undercooked food (Chai et al., 2005; De Liberato et al., 2011; Dreyfuss and Rondelaud, 2011; Fried et al., 2004; Lima dos Santos and Howgate, 2011; Toledo et al., 2011). of the digeneans is based primarily on morphological features with nomenclature based predominately on adult stages of the parasite. More recently the use of molecular techniques has been applied to assist with the linking of different life stages, in descriptions of new species and for furthering our understanding of cryptic species complexes (Aiken et al., 2007; Anderson and Barker, 1998; Bartoli et al., 2000; Blasco-Costa et al., 2010; Cavaleiro et al., 2012; Hall et al., 1999; Oliva et al., 2010; Valdivia et al., 2010).

Whilst there has been a reasonably large body of work conducted on treatment approaches for digeneans in vertebrate hosts (Shirakashi et al., 2012; Singh and Das, 2001), little has been achieved in development of methods to control such infections in molluscan host, in part due to the relatively low value of molluscs, the limited amount of farming and the mode of action of selected drugs which may interfere with the same pathways in the target pathogen and its molluscan host. It seems unlikely that chemotherapy will be applied in the control of molluscan pathogens at any point in the near future. However, understanding and knowledge of parasite lifecycles can be used to eradicate alternate hosts which contain infective stages thus breaking the lifecycle and reducing or eliminating stages in the whelk. Such intervention is only likely to be useful under farmed conditions.

Despite its relatively wide distribution and its economic and ecological importance, few parasites or pathogens of B. undatum have been recorded. It is not clear if this is due to a real lack of parasites in the host or due to a

34 lack of studies having been conducted on the species. However, in e.g. spp., at least 11 species of trematodes have been reported along with a number of other infections (Arakelova et al., 2004; Avdeev et al., 1986; Berry, 1962; Clausen et al., 2008; Curtis, 2002; Galaktionov and Skirnisson, 2000; James, 1968; Lauckner, 1984; Rothschild, 1941) thus demonstrating that gastropods can be infected by a large number of parasites. In addition to a small number of digeneans, Buccinum undatum has been recorded as a host for two protistans – Merocystis kathae, a coccidian occurring in the kidney and Piridium sociabile occurring near the ventral surface of the foot (Patten, 1935; 1936; Valentinsson et al., 1999); neither appears to be particularly detrimental to host survival. In addition, the turbellarian Graffilla buccinicola has been recorded in the kidneys and digestive glands of whelks (Jennings and Phillips, 1978; Valentinsson et al., 1999), again with no apparent effect on host survival.

To date seven digeneans have been reported from B. undatum occurring in the digestive gland, gonad, stomach and kidney – prevalence in any given population is generally less than 25%. Steringophorus furciger occurs as an adult within the stomach (Køie, 1969; Tétreault et al., 2000). However, although it attains the same size in whelks as it does in a fish host (normally a flatfish), it does not appear to produce eggs in whelks (Køie, 1969) and thus it is unclear if the whelk stage is truly adult or if whelks are an incorrect host. A second species capable of undergoing progenesis, precocious egg production in the cercarial or metacercarial stage (Lefebvre and Poulin, 2005), is Proctoeces maculatus (=P. buccini) occurring in the kidney (Loos-Frank, 1969). It was recorded at low prevalence but high intensity in whelks from the North Sea by Loos-Frank (1969) who incorrectly considered that the stages noted were true adults with the parasite being able to complete its lifecycle within the whelk host.

Whilst it appears that many of the parasites occurring in whelks are mainly harmless, those associated with the digestive gland and/or gonads are considered to be detrimental to host survival and have been implicated in castration of both males and females. Typically these parasites lead to destruction of the gonads and a marked reduction in penis size and mass (Køie, 1969). Renicola sp. has been recorded in the digestive gland of whelks from Danish waters and from the eastern seaboard of Canada (Køie, 1969; Tétreault et al., 2000). It forms small yellow bodies within the tubules of the digestive gland; its impact on whelks appears to be minimal (Tétreault et al., 2000) although Køie (1969) did suggest that it led to a reduction in the size of the penis of male whelks. Anomalotrema koiae (=Cercaria buccini in part) has been recorded as sporocysts in the digestive gland and gonads of whelks from the UK and in Danish waters (Køie, 1969). In contrast to Køie (1969), Lebour (1912) suggested that the parasite did not castrate its host although it was only recorded in 4 individuals during a survey by Lebour (1912). The sporocysts are generally colourless (Lebour, 1912) and no external colour change in the animal is noted as a result (Køie, 1969). In contrast, the digestive gland of whelks infected with Zoogonoides viviparus appears pinkish-yellow in colour (Lebour, 1918); the parasite also affects the gonads leading to its destruction (Valentinsson et al., 1999). Early stages are found throughout the visceral mass but with a predilection for the tissues between the digestive gland tubules and the gonad (Køie, 1987). Older sporocysts replace the gonadal tissue and digestive gland and although there is some host reaction, this is generally minimal (Køie, 1987). Siddall and Des Clers (1994) and Siddall et al. (1993) demonstrated that there was a negative correlation with Z. viviparus prevalence in whelks associated with sewage dumping at sea. They considered that the low prevalence of infection close to the sewage dump site was due to the toxic effects of trace metals in the dump site on the infective miracidium stage. Externally, infections by Neophasis anarrhichae (=Neophasis lageniformis =Neophasis pusilla =Acanthopsolus lageniformis) and (=Cercaria neptuni =Cercaria neptunae) are manifested as a “sickly grey” colouration to the digestive gland (Lebour, 1918; Rochette et al., 2001). Both have been shown to lead to destruction of gonads and a reduction in penis size (Køie, 1969; Køie, 1971; Rochette et al., 2001). Neophasis anarrhichae infects sexually mature individuals with infection prevalence being positively correlated with whelk size (Tétreault et al., 2000). Tétreault et al. (2000) suggested that in the wild the parasite may lead to mortality although empirical data obtained under laboratory conditions did not support the view, possibly as a result of a lack of predators and an abundance of food. In addition to occurring in the digestive gland and gonads, N. anarrhichae also occurs in the albumin gland of females and the seminal vesicles of males. Consideration could be given to the following as future studies: Confirmation of the taxonomy of the digeneans occurring within British whelks through the use of morphological and molecular techniques. Obtain an understanding of the geographical distribution of these parasites. Consider the use of parasites as biological indicators of stock structure and as pollution indicators (Siddall and Des Clers, 1994; Siddall et al., 1993). Elucidate the lifecycle of whelk parasites in British waters

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Consider the interactions between gonadal maturation, parasitism and contaminants through fields surveys and experimental challenges. Elucidate the impact of infections on survival, fecundity and other biological characteristics of the host through integrated field sampling and experimental challenges.

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