DOI: 10.5061/dryad.n02v6wx94
title: Helminth gut parasites of black grouse (Lyrurus tetrix) in northern England, their impact on productivity and possible sources of infection

description: Black grouse (Lyrurus tetrix) declines continue throughout much of
 temperate Europe, following habitat loss and population isolation,
 associated with reduced productivity. In northern England, only 1,437
 lekking males remained in 2014, genetically isolated from other UK
 populations, and typified by low productivity but high adult survival.
 Their distribution overlaps with that of red grouse (Lagopus scotica) on
 moorland managed for red grouse shooting. Here, quasi-cyclical
 fluctuations in red grouse numbers are driven by the parasite
 Trichostrongylus tenuis. Whilst management for red grouse may benefit
 black grouse survival through reducing predators, we hypothesised that
 parasite transmission from red to black grouse may contribute to low black
 grouse productivity. We measured T. tenuis prevalence and mean intensity
 from 186 black grouse carcasses (40 radio-tagged birds, 30 shot birds, the
 remainder found by chance) and compared them with equivalent measures in
 red grouse. T. tenuis occurred in 63% of black grouse at a mean intensity
 of 390 worms per bird, and in 95% of red grouse at 1,535 worms per bird.
 Seventeen black grouse contained Heterakis spp., and two contained the
 tapeworm Parionella urogalli. T. tenuis intensity in black grouse was 2-4
 times higher in predated birds than in ones that died from other causes.
 Body condition and productivity were both lower when T. tenuis abundance
 in black grouse was higher, supporting the hypothesis that parasites
 contributed to low productivity. Despite T. tenuis abundances varying
 temporally by approximately 15-fold in both grouse species, patterns of
 worm abundance between the species were not correlated, nor did we detect
 any relationships between black grouse breeding success and red grouse
 worm abundance. Despite the lack of direct evidence for parasite
 cross-transmission, we suggest ongoing precautionary surveillance of
 parasitic worms in areas where red and black grouse overlap.
description: Data collection Overall, 186 black grouse
 carcasses were collected between 1998 and 2022. Of these, 40 were
 radio-tagged birds from published studies of population dynamics in the
 North Pennine Dales (Warren and Baines 2002, Baines and Richardson 2007).
 A further 30 birds had been shot and provided by hunters, and the
 remaining 116 were found by staff, gamekeepers, farmers, and reserve
 wardens whilst conducting other tasks and provided for
 examination. Of the 186 carcasses, 160 (86%) were found
 in the North Pennines Dales (109 in Teesdale, 29 in Weardale, 20 in
 Tynedale and one in each of Allendale and Derwentdale), 25 (13%) in the
 Yorkshire Dales, comprising 14 in the Yorkshire Dales National Park (six
 in Wensleydale, five in Swaledale, two in Coverdale, and one in
 Arkengarthdale), nine in the Nidderdale National Landscape, and one from
 an unspecified location in northern England (Fig. 1). Carcasses were
 available in 22 of the 25 years, when they averaged eight per year,
 varying annually from one to 23 per year. The breakdown of the sample by
 age, sex, and region is provided in Table 1. Carcasses
 were aged as either adult or juvenile based on primary feather structure
 and pigmentation (Helminen 1963); sex was confirmed at the time of
 post-mortem examination. Weight (to the nearest 10 g) and wing length
 (flattened to the nearest mm) were measured from 135 fresh and intact
 carcasses. The season of death was recorded as either spring (March to
 May, n=41), summer (June to August, n=37), autumn (September to November,
 n=54), or winter (December to February, n=54). Cause of death was assigned
 for all but one of the 186 carcasses to one of four categories: collision,
 when the carcass was found adjacent to a fence, underneath electric wires,
 or in the immediate proximity of a road, and there was physical damage
 consistent with a collision; predation, when feathers had been either
 plucked or bitten from the carcass consistent with predation by raptors
 and mammals respectively (Thirgood et al. 1998), the carcass partially
 eaten, and, in the case of predation by stoat <em>Mustela
 erminea</em>, small, paired incisor holes were found, usually in the
 neck; shot, when the carcass was provided freshly killed by a hunter, or
 where skinning a carcass revealed pellet holes in the flesh; “other”
 causes were when the above criteria were not met, often where the carcass
 was thin, or the bird was unable to fly. This category probably included
 birds dying from disease. Adult <em>T.
 tenuis</em> abundance was estimated following Wilson (1983), whereby
 worms were collected from one of the two caeca by cutting it open
 lengthways and flushing its contents with water through an 850-μm sieve
 into a 212-μm sieve. The sieved contents were washed with 300 ml of water
 into a beaker and mixed thoroughly. Three 10-ml subsamples were placed
 into separate petri dishes and the total number of adult worms in each
 were counted using a binocular microscope with 25x magnification. Given
 the negligible variation between sub-samples, the sum of these counts was
 multiplied by 10 to estimate the total number of adults worms in a caecum
 and then doubled to give the total number per carcass. Other helminth
 worms observed during this procedure were recorded as present /
 absent. Following Bush et al. (1997), we define
 prevalence, mean intensity, and mean abundance of a parasite species as:
 prevalence = (number of carcasses infected with that species) / (total
 number of carcasses examined), mean intensity = (total number of
 parasites) / (total number of infected carcasses) and mean abundance =
 (total number of parasites) / (total number of carcasses).
 <em>T. tenuis</em> intensities in autumn-shot red
 grouse were measured each year at three driven grouse moors in Teesdale,
 the North Pennine Dale from where 59% of black grouse carcasses were also
 recovered. Here, spring densities of red grouse averaged 57 pairs
 km<sup>-2</sup> during the period 2000-22, synchronously
 fluctuating three- to four-fold between a mean low of 27 pairs and a mean
 high of 92 pairs km<sup>-2</sup>. For five consecutive years,
 worms in juvenile red grouse, which have fewer worms than adults (Shaw and
 Moss 1989), were not sampled owing to extremely low worm intensities in
 previous years. Numbers of adult red grouse sampled (both sexes combined)
 varied per year, with a mean of 194 (range 14 - 546). Their worm burdens
 were estimated as described above for black grouse.
 Black grouse breeding success was estimated each August at up to
 three sites in Teesdale using pointing dogs to find and flush females and
 their broods (see Baines et al. 2007). During the period 1998-2022, 890
 females were found at an average of 36 per annum (range 9-74) and occurred
 at a mean annual density of 3.5 km<sup>-2</sup> (range
 1.6-6.0). Counts were combined across sites to give one annual value.
 Three measures of annual breeding success were calculated: the proportion
 of females with broods (number of broods / number of females), brood size
 (number of chicks / number of broods) and chicks reared per female (number
 of chicks / number of females). Statistical
 analysis The 186 carcasses were unlikely to be a random
 sample of the black grouse population. To consider biases amongst
 carcasses, chi-squared contingency tests were used to compare causes of
 death between radio-tagged birds and those non-tagged birds found by
 chance.       Whether <em>T. tenuis</em>
 prevalence and intensity in black grouse carcasses differed in relation to
 region, season, age, sex and cause of death was considered in two parts.
 First, for worm prevalence we used a generalised linear mixed model (GLMM)
 with binomial error and logit link function, with a binary response
 variable (<em>T. tenuis </em>present (1) or absent (0)), and
 region (North Pennines or Yorkshire Dales), season (spring, summer,
 autumn, or winter), age (adult or juvenile) sex (male or female), and
 cause of death (collision, predation, shot or other) as fixed effects.
 Two-way interactions between season, age, and cause of death were also
 included as fixed effects. Second, to consider worm intensity amongst
 infected carcasses, we used a GLMM with Poisson error adjusted for
 overdispersion and a logarithmic link function (Wilson & Grenfell
 1997), with the <em>T. tenuis</em> count per bird as the
 response variable and incorporating the same fixed effects. The year in
 which a black grouse carcass was collected was incorporated as a random
 effect in both models. To determine whether observed
 levels of  <em>T. tenuis </em>infection impacted body
 condition in black grouse, we modelled log<sub>e</sub>(weight)
 as the dependent variable, with log<sub>e</sub>(wing length),
 region, age, sex, season, and log<sub>e</sub>(<em>T.
 tenuis</em> abundance + 1) as fixed effects and year as a random
 effect in a GLMM with a normal distribution and identity link function
 (cf. Packard & Boardman 1988). The model was equivalent to
 investigating factors affecting a body condition index defined as
 log<sub>e</sub>(weight/(wing
 length)<em><sup>b</sup></em>) where the exponent
 <em>b</em> accounted for the allometric relationship between
 weight and wing length. One obvious outlier (a juvenile male with long
 wing and low weight, not yet fully grown) was omitted from the
 analysis. Annual <em>T. tenuis
 </em>intensities in black grouse carcasses collected throughout the
 calendar year in the North Pennine Dales, where 86% of the sample was
 collected, were compared with those in red grouse shot in the autumn at
 three moors in Teesdale, one of the North Pennine Dales. Data were not
 collected from all moors in all years. To overcome missing moor-years, we
 fitted a GLM with logarithmic link function and overdispersed Poisson
 error, using total worms on a moor as the dependent variable and
 log<sub>e</sub>(number of grouse sampled from the same moor)
 as an offset, and moor and year as explanatory variables. We used the
 predictions from the model for year, setting offset = 0, to give annual
 values for the mean abundance of <em>T. tenuis</em> in  shot
 red grouse in Teesdale. Annual mean <em>T. tenuis
 </em>abundances
 (log<sub>e</sub>(<em>x</em>+1)-transformed) for
 each grouse species were compared in a general linear model (GLM), with
 worms in black grouse as the response variable, weighted by the number of
 carcasses in each year, and <em>T. tenuis</em> in red grouse
 as a covariate, first in the same year, then in the previous year, to help
 account for differences in timing of data collection between grouse
 species and any lag in response. Measures of annual
 black grouse breeding success in Teesdale were related to annual measures
 of worms in black grouse in all of the North Pennine Dales for 22 years
 during the period 1998-2022. The proportion of females with broods was
 modelled in a GLM with a binomial error and logit link function, with
 broods found per annum as the response variable and females observed per
 annum as the binomial total. Both brood size and chicks reared per hen
 were modelled with a Poisson error and logarithmic link function,
 log<sub>e</sub>(broods) and
 log<sub>e</sub>(females) respectively as offsets, and black
 grouse annual worm mean abundance (log<sub>e</sub>-transformed
 to linearize the relationship), weighted by the number of carcasses, as
 the explanatory variable. Insufficient black grouse carcasses and brood
 counts were available from the Yorkshire Dales to enable a similar
 comparison. The analysis was then repeated using red grouse
 log<sub>e</sub>(annual worm mean abundance) in the same year
 and in the previous one as an explanatory variable to consider whether
 worm abundance in red grouse influenced black grouse breeding
 success. For GLM and GLMM models, we reviewed
 standardised residuals for normality and bias using a Shapiro-Wilk test
 (Shapiro & Wilk 1965) coupled with visual examination of their
 frequency distribution, Normal plots, half-Normal plots and residuals
 versus fitted values. We adjusted Poisson and binomial errors for
 overdispersion where the residual deviance was significant when treated as
 a chi-square value with degrees of freedom equal to the residual degrees
 of freedom. The significance of main effects and interactions were
 assessed using Wald statistics, obtaining significance levels from a
 chi-square distribution if standardised residuals were normally
 distributed, and from random permutation tests (Edgington and Onghena
 2007) if not. Interactions were dropped if not significant at
 P&lt;0.05. Analyses were conducted using Genstat 24th Edition (VSN
 International, Hemel Hempstead, UK).
description: # Helminth gut parasites of black grouse (Lyrurus tetrix) in northern
 England, their impact on productivity and possible sources of infection
 Dataset DOI: [10.5061/dryad.n02v6wx94](10.5061/dryad.n02v6wx94) ##
 Description of the data and file structure To measure the relationship
 between black grouse (host) body condition and breeding success, and the
 intensity and abundance of the parasitic worm they and red grouse hosted. 
 ### Files and variables #### File: BLACK_GROUSE_STRONGYLE_WORMS.csv
 **Description:** Measures of age, sex, wing length, and weight of black
 grouse caracasses, together with the region where they were found, the
 likely cause of death, and the number of parasites they contained. Missing
 values are indicated as empty cells. ##### Variables * Season: that the
 carcass was found; 1=Spring, 2=Summer, 3=Autumn, 4=Winter * Month: that
 the carcass was found * Year: that the carcass was found * Sex: male or
 female * Age: adult or juvenile * Region: North Pennines or Yorkshire
 Dales, one blank where region was unknown * Cause: cause of death
 summarised into Collision, Predation, Shot or Other * Cause of death:
 original cause of death * Weight (g) * Wing length (mm) * HETworms: number
 of Heterakis worms * Ttworms: number of Trichostrongylus tenuis worms ####
 File: Adult_red_grouse_sampled.csv **Description:** Number of *T. tenuis*
 parasitic worms in shot red grouse in relation to grouse age and year. 
 ##### Variables * year: Year in which sample was collected * age: age of
 red grouse, either adult or young * worms: Number of *T. tenuis* worms in
 each grouse #### File: Black_grouse_breeding_success_and_parasites.csv
 **Description:** Measures of black grouse annual breeding success in
 relation to measures of *T. tenuis* worms in both adult red grouse and all
 sampled black grouse in the North Pennines, together with the number of
 birds sampled. Missing values are indicated as empty cells. * year: year
 in which the measures were made * BKHENS: the number of female black
 grouse from which estimates of breeding success were made in each year *
 BKBROODS: the number of broods of black grouse observed from within the
 sample of female black grouse found in each year * BKCHICKS: the total
 number of chicks of black grouse from the broods found in each year *
 RGTTEGGRAB: the natural log transformed mean number of *T. tenuis* worms
 in shot adult red grouse across three moors in Teesdale, North Pennines in
 each year * RGCORPS: the annual number of adult red grouse sampled to give
 the value in RGTTEGGRAB * RGTTPRED: the predicted annual mean number of
 *T. tenuis* worms in adult red grouse * LOGBKWORMS: the natural log
 transformed annual mean number of *T. tenuis* worms in all black grouse
 sampled within the North Pennines * BKCORPSENP: the number of sampled
 black grouse upon which the value in LOGBKWORMS is based