Introduction

Atlantic cod (Gadus morhua) is widely distributed in the North Atlantic and therefore exposed to a wide range of environmental conditions (Brander 2000). These diverse environmental conditions affect all biophysical processes including growth and indeed many studies have demonstrated differences in growth rate among and within cod populations (Svasand et al. 1996; Otterlei et al. 1999; Swain et al. 2003; Brander 2007). However, fish growth is a highly complex process, which is influenced both by the genetic set-up of the organism as well as the environment (Wootton 1998; Imsland and Jónsdóttir 2003; Cardinale et al. 2004). Growth is one of the most frequently studied parameters in fish populations as it is of great importance for the productivity of fish stocks (Jennings et al. 2001). Furthermore, maturation and egg production, and consequently the reproductive output of individuals are known to be affected by growth (Marteinsdottir and Begg 2002).

Studies on Icelandic cod have demonstrated differences in growth rate between at least three cod groups, where cod spawning in inshore waters south of Iceland grew faster than both cod in northern as well as the southern offshore waters (Marteinsdottir et al. 2000; Jónsdóttir et al. 2002, 2006a; Petursdottir et al. 2006). In addition to growth, somatic condition and especially the hepatosomatic index has been observed to vary between cod spawning north and south of Iceland (Pardoe et al. 2008). Recent studies on potential stock structure based on otolith shape (Jónsdóttir et al. 2006a), otolith chemistry (Jónsdóttir et al. 2006b), neutral (microsatellite) and non-neutral (Panthophysin, Pan I locus) genetic markers combined with tagging experiments (Pampoulie et al. 2006) clearly distinguished among cod spawning north and south of Iceland. In terms of the Pan I, the Pan IAA was most abundant north and east of Iceland while Pan IAB and Pan IBB were more abundant west and south of Iceland (Pampoulie et al. 2006). One location south of Iceland was, however, mostly composed of Pan IAA genotype (Pampoulie et al. 2006). It is also of interest to note that, although the genetic pattern was congruent with both types of genetic markers, the level of differentiation was approximately 100-fold higher for the Pan I compared to the microsatellites, a result interpreted as evidence for disruptive selection at the Pan I locus (Pampoulie et al. 2006).

The Pan I locus has been shown to be under natural selection in several other studies (Fevolden and Pogson 1997; Karlsson and Mork 2003; Pogson and Mesa 2004) and is frequently used to distinguish among populations (e.g. Fevolden and Pogson 1997; Karlsson and Mork 2003; Sarvas and Fevolden 2005; Pampoulie et al. 2006). It codes for an integral membrane protein expressed in cytoplasmic transport vesicles but its exact functions in microvesicle trafficking and exocytotic pathways are poorly understood (Windoffer et al. 1999; Brooks et al. 2000). Recently, it has been linked to differences in fish growth rate during the adult (Jónsdóttir et al. 2002; Imsland and Jónsdóttir 2003) and larval stages (Case et al. 2006). Larvae expressing the Pan IAB genotype expressed higher length and dry weight than larvae with the Pan IBB genotype (Case et al. 2006). However, no results were recorded for the Pan IAA genotype, as it was missing from the experiments carried out by Case et al. (2006). In adults, the Pan IAA genotype has been linked to greater size at age in comparison to the other two Pan I genotypes (Jónsdóttir et al. 2002; Imsland and Jónsdóttir 2003). The samples of spawning cod collected by Jónsdóttir et al. (2002) both inshore and offshore exhibited different proportion of Pan I genotypes where high frequency of the Pan IAA genotype was noted for inshore cod while offshore cod exhibited high frequency of Pan IBB genotype (Jónsdóttir et al. 2002). These results were later confirmed by Pampoulie et al. (2006). As data from these two populations were compiled to study the growth properties of each genotype (Jónsdóttir et al. 2002), the recorded difference in growth of individuals carrying the different Pan I genotype could merely reflect distinct populations with different life histories rather than a strict correlation between the Pan I genotypes and size at age.

The main objective of this study was to examine the relationship between the Pan I locus and three different parameters (fish length, somatic condition and hepatosomatic index) of cod spawning at different areas around Iceland. Specifically, the hypotheses that cod belonging to the same spawning group and expressing different Pan I genotypes exposed differences in growth (length-at-age), somatic condition and hepatosomatic index were tested.

Methods

Sampling

Female and male spawning cod were sampled during the peak of the spawning season in April–May in 2002 (n = 985) and 2003 (n = 1300). Samples were collected from 12 to 13 different spawning locations around Iceland in 2002 and 2003, respectively (Fig. 1; Table 1). Each spawning location was identified with a three digit number, the first digit representing one of the nine regions, the second the depth interval (1 <75 m; 2 75–125 m; 3 >125 m), and the last the station number (Fig. 1). Earlier studies showed differences in growth, otolith shape, otolith chemistry and genetics between cod spawning north and south, as well as at different depth south of Iceland (Jónsdóttir et al. 2002; Jónsdóttir et al. 2006a, b; Pampoulie et al. 2006; Petursdottir et al. 2006). Based on these studies, geographically close spawning locations were grouped together into four groups; shallow southern spawning locations (G1), west of Iceland (G2), northwest, north and northeast of Iceland (G3) and at deep southern locations (G4) (Fig. 1). Sampling was carried out from fishing boats using gillnets, handlines, or Danish seines. A total of 35–121 mature or spawning cod were sampled at each spawning location. At sea the total length of all sampled cod was measured to the nearest centimetre, gutted and ungutted weight of the fish was recorded and sex and maturity stage were determined macroscopically. Sagittal otoliths were carefully removed from each fish, cleaned of adhering tissue and stored dry in paper envelopes until age determination. Gill filaments were preserved in 1 ml of 96% ethanol for genetic analysis. Somatic condition was defined as Fulton’s K:

Fig. 1
figure 1

Sampling locations in spring 2002 (open symbols) and 2003 (filled symbols). Spawning groups divided into four groups: G1 (shallow south: circles), G2 (west; squares), G3 (northwest, north and northeast; up pointing triangles) and G4 (deep south; down pointing triangles). Depth contours at 75, 125 and 500 m

Full size image
Table 1 Number of total individuals of each Pan I genotype (Pan IAA, Pan IAB, and Pan IBB) in each spawning group in 2002 and 2003
Full size table
$$ K = \frac{w}{{l^3 }} \times 100 $$

and the hepatosomatic index was defined as:

$$ {\text{HSI}} = \frac{{{lw}}}{w} \times 100 $$

where w was gutted weight in grams, l the fish length in cm and lw the liver weight in grams.

DNA analysis

DNA was extracted using a Chelex (Biorad 10%) extraction protocol (Walsh et al. 1991) from gill filaments preserved in 1 ml of 96% ethanol. Polymerase chain reactions (PCR) and digested PCR products analyses were performed as described in Pampoulie et al. (2006).

Statistical analysis

Fish length, somatic condition and hepatosomatic index were tested for normality and homogeneity of variance. Hepatosomatic index was standardised by natural-log transformation. Analysis of covariance (ANCOVA) with sex and Pan I genotype as fixed factors and age as a covariate was used to identify which factors influenced fish length. ANCOVA was also used to evaluate differences in hepatosomatic index and somatic condition whereby spawning group and sex were used as fixed factors and Pan I genotype as a covariate. ANCOVA assumes a linear relationship between the dependent and the covariate. Linear relationship was not established between age and either somatic condition or hepatosomatic index. Therefore, age groups were combined for each year. Mean differences in somatic condition and hepatosomatic index between cod carrying the different Pan I genotypes within a spawning group were tested with one-way analysis of variance (ANOVA). Tukey’s HSD was then used to identify the source of significant differences detected by the ANOVA.

Results

Fish growth

Greater differences were observed in fish length-at-age between spawning groups than between cod carrying the different Pan I genotypes within an area (Fig. 2). Mean length-at-age was greater in G1 (shallow south) than in G3 (northwest, north and northeast) in both 2002 and 2003 (Fig. 2). Cod carrying the Pan IBB genotype had significantly higher length-at-age than cod carrying the Pan IAA and Pan IAB genotypes in G3 (northwest, north and northeast) (Fig. 2; Table 2: ANCOVA, P < 0.001). However, cod carrying the Pan IAA genotype in G1 (shallow south) in 2003 had significantly higher length-at-age than cod carrying the Pan IAB and Pan IBB genotypes (Fig. 2; Table 2: ANCOVA, P = 0.016). Length-at-age for cod carrying the different genotypes in the other spawning groups did not differ significantly (Table 2). The interaction between age and the Pan I genotype was only significant in G1 (shallow south) in 2002 (Table 2: ANCOVA, P < 0.001). Significant differences were detected in length-at-age between sexes in G1 (shallow south) in 2002 and in G1 (shallow south), G2 (west) and G3 (northwest, north and northeast) in 2003 (Table 2: ANCOVA, P < 0.05). The interaction between age and sex was not significant in any spawning group (Table 2). Only two individuals (one at the age of 5 and the other at the age of 6) in G4 (deep south) were carrying the Pan IAA genotype in 2002 but none in 2003.

Fig. 2
figure 2

Mean length-at-age (± 95% CI) for cod in different spawning groups in 2002 and 2003 expressing different Pan I genotype (Pan IAA, Pan IAB or Pan IBB). G1 (shallow south), G2 (west), G3 (northwest, north and northeast) and G4 (deep south)

Full size image
Table 2 Analysis of covariance (ANCOVA) of fish length with Pan I genotype and sex as main factors and age as covariate
Full size table

Condition

The mean somatic condition factor (Fulton’s K) was highest in G1 (shallow south) and G4 (deep south) and lowest for G3 (northwest, north and northeast) (Fig. 3). The mean somatic condition factor was highest for cod carrying the Pan IAA genotype and lowest for cod carrying the Pan IBB genotype in all spawning groups (Fig. 3). The relationship between Pan I genotypes and somatic condition was significantly different between spawning groups (Table 3: ANCOVA, P < 0.001). For all spawning groups (except G4 in 2002) the Pan IAA had significantly higher somatic condition factor than the Pan IAB (Tukey HSD, P < 0.05), which again had significantly higher somatic condition factor than Pan IBB (Tukey HSD, P < 0.05; except in G3 northwest and northeast in both 2002 and 2003). The relationship between the Pan I genotypes and somatic condition was not significantly different between sexes (Table 3). Significant positive relationship was found between fish length and somatic condition (P < 0.001).

Fig. 3
figure 3

Mean somatic condition (± 95% CI) of cod expressing different Pan I genotype (Pan IAA, Pan IAB or Pan IBB) in 2002 and 2003. G1 (shallow south), G2 (west), G3 (northwest, north and northeast) and G4 (deep south)

Full size image
Table 3 Analysis of covariance (ANCOVA) of somatic condition and hepatosomatic index with spawning group and sex as main factors and Pan I genotype as covariate
Full size table

Hepatosomatic index

The mean hepatosomatic index was highest in G2 (west). The mean hepatosomatic index was lowest for G3 (northwest, north and northeast) except for the Pan IBB where the G1 (shallow south) was lowest (Fig. 4). The relationship between Pan I and hepatosomatic index was significantly different between spawning groups (Table 3: ANCOVA, P < 0.001). In contrast to the somatic condition factor, the hepatosomatic index was highest for cod carrying the Pan IBB genotype and lowest for cod carrying the Pan IAA genotype in all spawning groups (except G1 in 2003; Fig. 4). The Pan IAA had significantly lower hepatosomatic index than Pan IAB in all spawning groups except G1 (shallow south) and G2 (west) in 2002 (Tukey HSD, P < 0.05). However, significant differences between the Pan IAB and Pan IBB were only detected in G1 (shallow south) and G3 (northwest, north and northeast) in 2002 and in G2 (west) in 2003 (Tukey HSD, P < 0.05). The relationship between Pan I and hepatosomatic index was significantly different between sexes (Table 3: ANCOVA, P < 0.001). The hepatosomatic index was not significantly related to fish length (P = 0.72).

Fig. 4
figure 4

Mean hepatosomatic index (± 95% CI) of cod expressing different Pan I genotype (Pan IAA, Pan IAB or Pan IBB) in 2002 and 2003. G1 (shallow south), G2 (west), G3 (northwest, north and northeast) and G4 (deep south)

Full size image

Discussion

The results of the present study indicate complicated relationship between growth and condition and the Pan I locus in Atlantic cod. Differences in growth (length-at-age) were greater between cod spawning at different areas around Iceland than between cod carrying the different Pan I genotypes within a spawning area. However, significant differences in growth and condition (somatic and hepatosomatic index) between cod carrying the different Pan I genotypes were observed both north and south of Iceland. Cod carrying the less frequent genotype were found to express the highest length-at-age in each area, e.g. Pan IAA south of Iceland and Pan IBB north of Iceland. Earlier studies have already suggested that the Pan I genotype was related to growth (Jónsdóttir et al. 2002; Case et al. 2006). However, the results of the present study indicate that the relationship between fish growth (and condition) and the Pan I locus is complicated and is likely to be influenced by other factors like size-selective fishing and food supply.

Greater differences were detected in growth (length-at-age) among cod from the different spawning areas than among cod expressing different genotypes at the same spawning area. Cod spawning south of Iceland were found to grow faster than cod spawning north of Iceland (see also Marteinsdottir et al. 2000; Jónsdóttir et al. 2006a). This difference could to some extent be explained by the substantial temperature differences found between these two areas as temperature has been noted to have a direct effect on fish growth (Brander 2000; Björnsson et al. 2001). The recorded temperature above 200 m was 6–10°C south of Iceland while it was between 0 and 7°C north of Iceland in the period from 1990 to 2001 (Malmberg and Valdimarsson 2003).

In the present study, cod carrying the least frequent Pan I genotype in each area (north or south) was the one expressing the highest growth rate. Significant differences between cod carrying the different Pan I genotypes were, however, only detected in two groups (G1 shallow south in 2003 and G3 northwest, north and northeast in 2002 and 2003). In the study of Jónsdóttir et al. (2002) it was suggested that the Pan I locus was related to growth, as cod carrying the Pan IAA grew faster than the Pan IBB. However, this observed pattern was likely to be due to different life histories experienced by two genetically distinguishable populations. Fast growing Pan IAA cod were found close to shore (location 911) while the Pan IBB was more frequent in slower growing cod spawning at greater depth (locations 931, 932, 933). These have been suggested to represent different populations (Jónsdóttir et al. 2002; Pampoulie et al. 2006). The higher growth rate of the Pan IAA cod south of Iceland cannot been explained by this fast growing group mentioned earlier (location 911), as genetic data were not available for this exact location in spring 2003 when the Pan IAA cod were observed to grow faster than the other Pan I genotypes in the shallow southern region. However, in 2002 this fast growing group was present but not found to grow significantly faster than the other Pan I genotypes. Therefore, the observed differences in the present study were likely to be within a population. In another study addressing the relationship between growth and Pan I, faster growth was recorded for larvae expressing the Pan IAB compared to the Pan IBB (Case et al. 2006). The Pan IAA genotype was, however, missing from the study. In cod the Pan IA and Pan IB alleles differ by four amino acid substitutions. Because of that, the alleles could be encoding for different proteins with markedly different physiological properties (Pogson 2001). As environmental conditions vary greatly between the northern and the southern spawning areas, natural selection could be acting on different properties in these two areas. However, it has not been possible to link selection at the Pan I locus with any environmental factor, as little is known about the function of the Pan I locus in fishes (Pogson 2001; Pogson and Mesa 2004).

Selection on the Pan I locus can be linked with numerous other factors. Besides growth, condition is a likely factor. In this study, two different condition factors were tested; the somatic condition and the hepatosomatic index. The somatic condition (defined as Fulton’s K) showed significant relationship with the Pan I locus, where cod carrying the Pan IAA genotype expressed the highest condition in all spawning groups. However, the somatic condition showed significant relationship with fish length and the difference between the genotypes could therefore only be indicating differences in length. Furthermore, in a recent study the somatic condition expressed little variation between spawning areas, sexes or mature/immature individuals (Pardoe et al. 2008). The hepatosomatic index was, however, found to show greater variation between areas, sexes and mature/immature individuals (Pardoe et al. 2008). The results of the present study also showed variation in hepatosomatic index between sexes and a significant relationship with the Pan I locus, but not to fish length. In contrast to the somatic condition, cod carrying the Pan IAA genotype was found to have the lowest hepatosomatic index in all spawning groups (except deep south). A study on cod larvae showed better condition (estimated as the RNA:DNA ratio) for cod larvae expressing the Pan IAB than the Pan IBB genotype (Case et al. 2006). The cod sampled in the present study were spawning and condition (both somatic and hepatosomatic index) has been found to vary among seasons and to be lowest during the time of spawning (Lambert and Dutil 1997; Lloret and Ratz 2000; Mello and Rose 2005). Food supply may also have a great impact on cod condition. Earlier studies on migration behaviour indicated different patterns of cod carrying the different Pan I genotypes, whereas cod carrying the Pan IAA displayed shallow water feeding migration while the Pan IBB displayed deep water feeding migration pattern (Pampoulie et al. 2008). Higher growth rate was furthermore recorded for the shallow water migrating cod indicating greater food supply in shallower waters (Pálsson and Thorsteinsson 2003). The higher somatic condition and lower hepatosomatic index of cod carrying the Pan IAA genotype may possibly be explained by greater food supply, while cod expressing the Pan IBB genotype may experience lower food supply and hence, store greater amount of energy in the liver.

Two alternative explanations for the opposing results shown in the present study are worth mentioning; the trade off between growth and reproduction, and size-selective fishing. Most fish grow continuously throughout life. However, at maturity somatic growth of mature individuals generally declines because energy is reallocated from growth to future reproduction (Wootton 1998). Various factors are known to influence fish growth and reproduction, including food supply (Yoneda and Wright 2005), temperature (Björnsson et al. 2001) and metabolic rate (Wright 1991). These abiotic factors are the most important factors shaping the reproductive ecology of fishes (Wootton 1998). In the present study, the trade-offs between growth and reproduction may have resulted in faster growing cod (e.g. Pan IAA in the south) maturing at earlier age. Different age at maturity has been recorded for faster growing cod spawning south of Iceland compared to slower growing cod north of Iceland, where age at maturity was 5.4–7 years for cod spawning south and north of Iceland, respectively (Marteinsdottir and Begg 2002). Different age at maturity may also well be found for cod expressing different growth rate within the same spawning area. Another factor possibly explaining the results of the present study is size-selective fishing. Fishing is seldom random as most fishing gears are designed to be size selective and fishing often tends to remove the largest, fastest growing individuals in a population (Law 2000; Stokes and Law 2000). A genotype that produces faster growing fish in certain environmental conditions (e.g. Pan IAA in the south) would tend to be at a lower frequency in that region. Less frequent genotype having the highest length-at-age could therefore be explained by the removal of the larger, faster growing fish through selective fishing. It is, however, not possible to answer these issues with the data presented in this study.

The Pan I locus has been suggested to be under positive Darwinian selection (Pogson and Mesa 2004). In Iceland, it has been suggested that disruptive selection acts on the Pan I locus, favouring Pan IAA genotypes in the north and Pan IBB genotypes in the south (Pampoulie et al. 2006). Disruptive selection will typically provide an advantage for reproductive isolation among opposite, extreme phenotypes, well adapted to different environmental conditions (Wallace 1889; Schluter 2001). As mentioned earlier, the less frequent Pan I genotype in each area usually expressed the highest growth rate. It is unlikely that the common genotype in different areas were selected upon without expressing a direct advantage on the phenotype. Hence, it is unlikely that the Pan I locus is directly linked to a single factor like growth or condition and that the relationship might depend on several environmental parameters and be more complex than previously thought.