Groupx+19


 * Lauren Joyce
 * Harpreet Kaur
 * Sarah Cupper
 * Jasmine Tinker
 * Vicki Trimble
 * Ali Al-Ghusein


 * Original Sampling Plan**



We chose to have 6 samples covering the three main habbitats of woodland, shrubland, and grassland. We want to see if height and/or distance has an effect on the type of shells of //Cepaea nemoralis//. We chose to have two samples in each type of environment, but at opposite heights and distances so we can test for the effect of these variables. We expect that habitat will be the variable that exemplies the greatest diversity among this species, as shown by Figure 11.3 below, and would be supported by negligible differences in the height and distance variables within the same habitat.
 * Revised Sampling Plan**

We changed our sampling plan to eliminate the measurement of extraneous variables that would have been measured if we followed our initial plan. We changed our design to eliminate the variable of height by having each sample at the same height. We also have 6 samples, 2 samples for each type of environment to ensure the replication of samples in each type of environment (woodland, shrubland, and grassland). In an attempt to control for the distance variable, we are planning on having each sample about 50 meters away from one another, to keep the distance consistent for each sample collection to eliminate the variable of varied distances. Under this sampling plan, we are planning to focus on the environment variable and how it may have an effect on the phenotypic frequencies of //C. nemoralis.//
 * Methodology/Reasoning:**

// Cepaea nemoralis //, commonly known as grove snails, are known to have a shell polymorphism consisting of colour and banding. The polymorphism for colour is yellow, pink, or brown, but there can be various shades of these colours. The variations in banding ranges from zero to five bands. They are also indigenous to western Europe and have been found living in a variety of habitats, ranging from dunes to wooded areas. //C. nemoralis// is known to be one of the most polymorphic members of the European fauna (Jones et al., 1977).
 * Introduction**

Compared to other animals, snails are known to be smaller and slower and thus the polymorphism changes can be determined by geographic scaling. //C. nemoralis// are often used in polymorphism studies because of their obvious and easily identifiable phenotypic variation. There is currently some evidence that allele frequency is related to population, but the causation for polymorphisms in varying populations of //C. nemoralis// is still under speculation and depends on a variety of factors.

Jones et al. (1977) found that woodland habitats of //C. nemoralis// had predominately brown and pink shells with generally no bands, grassland habitats had yellow shells with bands, and hedgerows had intermediate frequencies. In order to determine if Jones et al.’s findings still hold true, this report aims to determine the polymorphism variation in the woodland, shrubland, and grassland habitats of Pulpit Wood Reserve, near Monks Riseborough, in South-East England. The habitats are similar to that of Jones et al.; the shrubland being the hedgerow equivalent. Similarly to Jones et al whether these variations is caused by genetic drift, gene flow, or selection, such as predation or climate, will be investigated.

The sampling strategy will consist of two separate locations for woodland, shrubland and grassland, with a minimum distance of 50m between each habitat type locations and will also control for variables such as elevation, and ensure replication of each of the three habitats and to minimise the likelihood of failure to find snails or any issues that may be caused by proximity. In addition, large enough samples will be taken from each of the six locations to ensure good replication and representation of the populations in each given loci.

The aim of this report is to determine if there is selection for a specific frequency of shell colour polymorphism dependent on the type of habitat. If there is evidence of spread or loss of certain polymorphisms, then it is likely caused by genetic drift and selection. There are also different locations, so there may be evidence of gene flow in cooperation with genetic drift and/or natural selection. In the absence of natural selection and genetic drift, gene flow will allow for homogeneous frequencies and an equilibrium to be reached. If natural selection and genetic drift are present in the populations observed, gene flow will result in divergence in the populations because it is restricted (Andrews, C. A., 2010). If the populations are diverse and there are different frequencies in shell colour and banding polymorphisms, there may be evidence of natural selection and genetic drift in //C. nemoralis//.

Final Report:

// Cepaea nemoralis, // commonly known as the grove snail, is native to western and central Europe with a habitat ranging from dunes to wooded areas. //C. nemoralis// is a model organism for investigating polymorphism. Polymorphism is the existence of two or more different phenotypes in a randomly mating population of the same species, due to multiple alleles at the locus for a particular trait. //C. nemoralis// exhibit shell polymorphism of both colour and banding; the shell can be varying shades of yellow, pink or brown with a range of 0-5 bands. //C. nemoralis// are comparatively smaller and slower than most other animals so polymorphic changes can be determined by geographic scaling in a relatively small area. There is evidence that allele frequency is related to population, but the causation for polymorphisms in populations of //C. nemoralis// depends on a variety of factors (Jones et al., 1977).
 * Introduction **

This report sets out to determine if there is selection for a specific frequency of shell colour polymorphism depending on the type of habitat. If there is evidence of spread or loss of certain polymorphisms, then it is likely caused by genetic drift or selection. There are also different locations, so there may be evidence of gene flow in cooperation with genetic drift or natural selection. In the absence of natural selection and genetic drift, gene flow will allow for homogeneous frequencies and equilibrium to be reached across areas within the population. If natural selection and genetic drift are present in the populations observed, gene flow will result in divergence in the populations because it is restricted. If there are different frequencies in shell colour and banding polymorphisms, there may be evidence of natural selection and genetic drift in //C. nemoralis//.

Jones et al. (1977) found there was a correlation between habitat and shell colour in //C. nemoralis//. Snails of woodland habitats had predominately brown and pink shells with no bands, while yellow shells with bands were common in grassland and hedgerows had intermediate frequencies of all colours. This report seeks to explore the polymorphic variation in the woodland, shrubland and grassland habitats of Pulpit Wood Reserve near Monks Riseborough, in South-East England. The habitats are similar to the original study with the shrubland serving as the hedgerow equivalent, whether these variations are caused by genetic drift, gene flow, or selection, such as predation or climate, will remain the aim of the investigation.

Samples will be taken at two separate locations for each habitat type, woodland, shrubland and grassland, with a minimum distance of 50m between samples while controlling for variables such as elevation and ensuring replication of the three habitats. By ensuring a minimum distance of 50m this sampling strategy also hopes to minimise the likelihood of finding insufficient numbers of snails or any other issues that may be caused by proximity; care will be taken to collect large enough samples from each of the six locations in order to ensure good replication and representation of the populations in each given loci.

Word Count: 499


 * Results **
 * Table 1-5 ** . Two-way table of environment in comparison to //C. nemoralis// shell banding and colour. //C. nemoralis// were collected and separated into categories by colour and high (HB) banding (3-5 bands) and low (LB) banding (0-2 bands). Observed and expected ((#)) values were calculated to obtain Chi square values for each comparison.

// X2 // =13.94 // X2 // = 14.62 // X2 // =16.9
 * ** Table 1. ** ||  Brown LB   ||   Brown HB   ||   Yellow LB   ||   Yellow HB   ||   Total   ||
 * Woodland  ||   10 (11.54)   ||   10 (6.07)   ||   7 (11.54)   ||   7 (4.86)   ||   34   ||
 * Grassland  ||   9 (7.46)   ||   0 (3.93)   ||   12 (7.46)   ||   1 (3.14)   ||   22   ||
 * Total  ||   19   ||   10   ||   19   ||   8   ||   56   ||
 * ** Table 2. ** ||  Brown LB   ||   Brown HB   ||   Yellow LB   ||   Yellow HB   ||   Total   ||
 * Grassland  ||   9 (8.03)   ||   0 (3.81)   ||   12 (7.19)   ||   1 (2.96)   ||   22   ||
 * Shrubland  ||   10 (10.97)   ||   9 (5.19)   ||   5 (9.81)   ||   6 (4.04)   ||   30   ||
 * Total  ||   19   ||   9   ||   17   ||   7   ||   52   ||
 * ** Table 3. ** ||  Brown LB   ||   Brown HB   ||   Pink LB   ||   Pink HB   ||   Total   ||
 * Woodland  ||   10 (9.32)   ||   10 (4.91)   ||   6 (10.30)   ||   0 (1.47)   ||   26   ||
 * Grassland  ||   9 (9.68)   ||   0 (5.09)   ||   15 (10.70)   ||   3 (1.53)   ||   27   ||
 * Total  ||   19   ||   10   ||   21   ||   3   ||   53   ||

// X2 // =11.33 // X2 // = 10.8 Calculation for Determining the accepted chi-square value: DF= (#Rows-1)(#Columns-1)= (2-1)(4-1)=3 Df=3, P= 0.05 Accepted //X2//= 7.815
 * ** Table 4. ** ||  Brown LB   ||   Brown HB   ||   Pink LB   ||   Pink HB   ||   Total   ||
 * Grassland  ||   9 (9.16)   ||   0 (4.34)   ||   15 (11.09)   ||   3 (1.45)   ||   27   ||
 * Shrubland  ||   10 (9.84)   ||   9 (4.66)   ||   8 (11.91)   ||   2 (3.55)   ||   29   ||
 * Total  ||   19   ||   9   ||   23   ||   5   ||   56   ||
 * ** Table 5. ** ||  Pink LB   ||   Pink HB   ||   Yellow LB   ||   Yellow HB   ||   Total   ||
 * Woodland  ||   6 (8.24)   ||   0 (2.35)   ||   7 (7.45)   ||   7 (3.14)   ||   20   ||
 * Grassland  ||   15 (12.76)   ||   3 (0.65)   ||   12 (11.55)   ||   1 (4.86)   ||   31   ||
 * Total  ||   21   ||   3   ||   19   ||   8   ||   51   ||
 * Table 6. ** Calculated Chi-square values of habitat and snail shell type comparisons that did not exceed the accepted chi-square value of 7.815.
 * Comparison  ||   Chi-square value   ||
 * Pink v. Yellow in Woodland and Shrubland  ||   2.67   ||
 * Pink v. Yellow in Grassland and Shrubland  ||   7.12   ||
 * Brown v. Pink in Woodland and Shrubland  ||   2.18   ||
 * Brown v. Yellow in Woodland and Shrubland  ||   0.21   ||

A series of chi square tables were calculated for varying environments and specific kinds of snail shells pertaining to banding and shell colours. The null hypothesis and alternative hypotheses stated the following: Ho: # of snails in one particular environment= # of snails in other environment HA: # of snails in one particular environment ≠ # of snails in other environment The comparisons that exceeded the accepted chi-square value of 7.815 indicated that the null hypothesis could be rejected and that there may be a significant difference between the environment and snail shell type. The following comparisons exceeded the accepted chi-square value; brown v. yellow in woodland and grassland, brown v. yellow in grassland and shrubland, brown v. pink in woodland and grassland, brown v. pink in grassland and shrubland, and pink v. yellow in woodland and grassland. In the following comparisons the chi-square value did not exceed the accepted chi-square value of 7.815, indicating that the null hypothesis could be accepted and that there was no discrepancy between environments and snail shell types; brown v. yellow snails in woodland and shrubland, brown v. pink snails in woodland and shrubland, pink v. yellow in woodland and shrubland, and pink v. yellow in grassland and shrubland.

The findings of this report indicate that there may be evolutionary selective factors acting on the populations of //C. nemoralis// in the Pulpit Wood Reserve. Jones et al. (1977) found that populations of //C. nemoralis// in woodland habitats had predominately brown and pink shells with no bands, while yellow shells with bands were common in grassland and hedgerows had intermediate frequencies of all colours. In this study, there was no significant difference between brown and pink shells in the woodland and shrubland environments. This may be indicative of both snail shell types being favourable or beneficial to these environments. However, there was also not a difference between pink and yellow or brown and yellow in the woodland and shrubland comparisons. These comparisons indicate that the null hypothesis was accepted and that there was no difference between frequencies of these particular snail shells.
 * Discussion **

Similarly to Jones et al. (1977), the woodland and shrubland polymorphism frequencies were varied and there seemed to be no major selection or drift occurring. One reason may be because these areas allow //C. nemoralis// to be less vulnerable to predators so there are few predator selective pressures. Also the shrubland and woodland areas can vary in colours and density so there may be less of a need for selection and drift and more of a need for a high amount of genetic diversity within the populations.

The chi-square values that exceeded the accepted chi-square value were comparisons between grassland populations to woodland and shrubland populations. As previously mentioned, Jones et al. (1977) found that yellow snails with bands are predominantly found in grassland populations. Unlike Jones et al., the majority of snails found in the grassland populations had low banding. There were no major discrepancies between shell colour, but there were differences between banding. There was a much higher frequency of low banding than high banding. This may be the due to factors such as genetic drift or natural selection. These findings are unlike that of Cameron & Cook (2012) that found that habitat differences in correlation to polymorphism frequencies in //C. nemoralis// are more apparent in colour than banding.

During the sample collection of this investigation, 80% of the snails were dead, 68% of them being dead adults. If factors such as natural selection were acting on the snail populations in Pulpit Wood Reserve, it would be more evident if the samples that were collected were of a greater sample size with a range of subadults and adults to get an understanding of possible evolutionary changes such as drift or selection.

The findings of this report indicate that there are discrepancies polymorphic frequencies, and there may be evolutionary factors acting upon //C. nemoralis// of this reserve such as genetic drift or natural selection. There may also be gene flow between different populations of the same habitat. Although some of the findings differ from Jones et al. (1977), both investigations found differences in polymorphism frequencies that may be due to genetic drift or natural selection.

Word Count: 498

Cameron, R. A. D. & Cook, L. M. (2012) Habitat and shell polymorphism of Cepea Nemoralis (L) : Integrating Evolution Megalab database //J. of Molluscan Studies// 78:179-184.
 * References: **

Jones et al. (1977) Polymorphism in Cepea: A Problem with Too Many Solutions? //Ann. Rev.// // Ecol. Syst. // 8:109-43.