Carabodes marginatus (Michael, 1884)

Avanzati, Anna Maria, Salomone, Nicola, Baratti, Mariella & Bernini, Fabio, 2004, Genetic diversity in the Carabodes marginatus species group (Acarida, Oribatida, Carabodidae) as inferred from allozymes, Journal of Natural History 38 (15), pp. 1927-1940 : 1933-1938

Are Carabodes marginatus and closely related taxa separating evolving gene pools?

According to our data, these taxonomic entities of soil oribatid mites would represent different evolutionary lineages, as suggested by the observed mean genetic divergence (from D=0.187 to D=0.827). Previous works indicate that the critical genetic distance (D) for distinguishing between morphologically differentiated species in oribatid mites is 0.125 –0.328 (Bernini et al., 1988; Bernini and Avanzati, 1989; Pigliucci et al., 1990), and observed values justify the distinction between the four taxa at the specific level.

Carabodes arduinii is a taxon distantly related to all the studied species. This

result is in agreement with morphological evidence which includes this species in the ‘ coriaceus ’ group.

Two features regarding the species of the ‘ marginatus ’ group are worth noting. First, C. marginatus seems to be more closely related to C. montanus than to the morphologically similar C. affinis . Second, there is a high genetic affinity between C. affinis and C. quadrangulus (separated by a level of divergence D=0.187). Although the D value is sufficient to determine the two taxa at the specific p (1): average frequency of private alleles; N m and N m: quantitative estimates of

w

gene flow based on Slatkin’s (1985a,b) method and F (Nei, 1977) values respectively.

st

n.p.a. Sample size p (1) N m F N m st w

C. marginatus 5 23.68 0.055 2.489 0.382 0.405

level, this finding is surprising because the two species are morphologically well differentiated (Baratti et al., 1995).

It seems that morphological differentiation in these species is not accompanied by a similar degree of genetic differentiation. Data from previous studies have also failed to find any correlation between morphological and genetic variation in oribatid mites (Bernini and Avanzati, 1987, 1989; Bernini et al., 1988; Pigliucci et al., 1990). It is generally accepted that different processes influence molecular and morphological evolution. Molecular rates are mostly determined by the rate of neutral mutations, while rates of morphological evolution are assumed to be controlled by the fluctuating pressure of natural selection (Kimura, 1983). Another possible explanation for the lack of correlation between the two kinds of data is that the genic regulation of some diagnostic characters is unknown and probably not related to protein-coding genes. Clearly, more detailed analyses are necessary to determine the mechanisms governing the many aspects of both molecular and morphological evolution in Carabodes .

Comparisons of inferred levels of gene flow in C. marginatus

Gene flow analysis provides information about the genetic structure of C. marginatus populations and related morphotypes. However, the two different methods of estimating gene flow from allele frequency data (F values and number of private

st alleles p (1)) led to conflicting results (table 5).

Based on F values from this study, the high degree of population subdivision

st indicates very low gene-flow among such geographically grouped oribatid mite populations. Even in sympatric populations, genetic heterogeneity between the two setal forms can be detected. Different evidence was obtained in the quantitative analysis of gene flow by the method of rare alleles. The N m estimate using this approach is much higher, suggesting a considerably higher level of gene exchange between subpopulations.

Similar discrepancies in the two estimates have already been described, as for example in Collembola (Frati et al., 1992) and cave crickets (Caccone and Sbordoni, 1987). It has been suggested (Slatkin and Barton, 1989) that, in analysing ideal data, F and rare-allele methods are equally effective for describing the extent of st

variation in allele frequency among demes. In reality there are practical reasons for preferring F to p (1): the first can be estimated from data from any polymorphic st

loci, whereas p (1) requires that a reasonable number of private alleles be present. Another advantage of F is that it is probably less sensitive to errors in the st

interpretation of electrophoretic data. The results obtained with Slatkin’s method could be misleading due to the low number of detected private alleles. On the other hand, even if F estimates support a genetic discontinuity between the two setal st

forms, in this situation discrepancies in the two methods make it hard to give a precise estimate of gene flow.

Which is the taxonomic status of the morphotypes related to C. marginatus ?

Based on morphological evidence, the only remarkable difference between the seven analysed C. marginatus populations (including both setal forms) is essentially related to the length of the marginal notogastral setae (Baratti et al., in preparation). The presence of both morphs in the same natural environment leads to the exclusion of major ecological factors as possible causes for differentiation. The intra-specific variation of setal length is a well-known feature (McMurthry, 1980; Abou-Setta et al., 1991) and among closely related species intra-specific variation may exceed inter-specific variation (Mayr, 1978). According to morphological and ecological evidence, C. marginatus could be characterized by a high degree of intra-specific variation concerning the shape of the notogastral setae.

Although this conclusion seems plausible, some data suggest an alternative explanation.

The population from the Sibillini Mts. seems to be exclusively characterized by morphotypes with short marginal notogastral setae. No specimens characterized by long marginal setae ( C. marginatus typicus) were found among the approximately 1000 individuals collected at this site and analysed morphologically.

Further evidence contrasting the intra-specific morphological variation hypothesis is provided by allozyme data. Despite an overall low degree of molecular differentiation which distinguishes the tested populations, the UPGMA analysis yields a pattern of relationships that does not follow a geographic scheme, i.e. sympatric populations of the two setal forms are never clustered together. This is an interesting feature, because under the effect of gene exchange, conspecific specimens living in the same area are expected to share the same allelic frequencies. On the contrary, our data clearly indicate different patterns of allelic frequency among local populations. Despite the absence of fixed genetic differences, several loci show alleles with lower frequencies that are never present in both sympatric morphs. This is the case of the PGM locus, where populations of C. marginatus typicus from Fioreta and Mt. Amiata exhibit several allelic variants that are never detected in morphotypes with short marginal setae from the same area. These are both monomorphic at this locus.

The EST-2 locus reflects other important genetic differences. On Mt. Amiata and in Poggibonsi the two sympatric setal forms share the same alleles, but their frequencies are significantly different. It is difficult to explain these important genetic differences between sympatric populations without assuming a lack of gene exchange and significant genetic drift in shaping the distribution of allele frequency. A barrier to gene flow among geographically grouped populations is likely moreover considering the inability of electrophoresis to detect all DNA substitutions. Previous analyses applied to the study of the genetic variability of natural populations demonstrated that allozyme analyses detect genetic differences above the species level, but fail to reveal sufficient polymorphism at the subspecies level due to the absence of change in electrophoretic mobility of many enzymes (Jenczenski et al., 1999; Thomas et al., 1997).

One possible explanation, given the observed low genetic distances, is that these two setal forms diverged very recently, so that much sharing of alleles and lineage sorting is still occurring confounding the phylogenetic delimitation of species boundaries. This would explain why the UPGMA reconstruction partly failed to distinguish the two lineages.

This genetic diversification may have a historical basis in the recent Pleistocene glaciations. Based on molecular data, numerous cases of genetic divergence at subspecific and even specific level have already been reported as a consequence of glacial events in Europe. Interestingly, the highest number of sister species, subspecies and hybrid zone have been described in the southern regions of Italy, Iberia, the Balkans and Greece which acted as refugia (see Hewitt, 1999 for a review). Similarly, climatic changes which determined the fragmentation of former populations of C. marginatus may have created the necessary conditions for divergence in allopatry with secondary intergradation after range expansion from refugia. This would not exclude that occasional gene flow is actually eroding phylogenetic signals in the two taxa.