A SURVEY OF AVAILABLE MOLECULAR MARKERS FOR VERTEBRATE SPECIES PRESENT IN COMUNIDAD DE MADRID

In this paper I present a survey for molecular markers available for Vertebrate species inhabiting the Comunidad de Madrid, with the aim of being useful to researchers working on the characterization of animal biodiversity in this area.


Introduction
Today, molecular markers are a valuable tool for taxonomic and conservation studies allowing serious inferences in phylogeography, genetic structure of populations, conservation status, and so on (Bruford & Wayne, 1993;Tautz et al., 2003;Avise, 2004).
For this reason, I consider that a survey of available molecular markers for Vertebrate species inhabiting in Comunidad de Madrid in sequence databases could be interesting as a research tool for biodiversity present in this area, giving researchers an overview to molecular knowledge for the species under scrutiny, and contributing to define gaps that can be filled subsequently.
The present survey is realised with this in mind, searching in sequence databases for mitochondrial, nuclear, and microsatellite (frequently repeated, highly polymorphic short sequences) sequences for vertebrate species or genera present in Comunidad de Madrid.
This approach, however, can have relative value only, since new sequences in sequence databases become available almost every day, and could make data shown in this work rapidly obsolete.In this sense, I believe that incorporating these data into a new database in progress in the Museo Nacional de Ciencias Naturales, and their continued review, can increase the value of the work.

Methods
Each species present in Comunidad de Madrid has been searched for number of sequences, using the Taxonomy browser of the WEB page of NCBI (National Center for Biotechnology information USA) (http://www.ncbi.nlm.nih.gov/Taxonomy/taxonomyhome.html).This information has been edited, sorting them into mitochondrial and nuclear sequences and microsatellites when sequence number output is less than 200 sequences.Sequences available for the corresponding genera, with special focus in the presence of microsatellites, have also been analyed.
In cases with a sequence number output greater than 200, I search specifically for the presence of microsatellite using the Entrez option with a query: "Genus name + microsatellite".
In the case of Fish I took the distribution show in the Atlas y Libro Rojo de los Peces continentales de España (Doadrio, 2002) as the basis of the their presence in the region.For Amphibia, I followed Martínez-Solano & González Fernández (2003).In relation to Reptilia, I used the Atlas y Libro Rojo de los Anfibios y Reptiles de España (Pleguezuelos et al., 2002).In the case of Aves, I split the species present in Madrid into sedentary and migrants on the basis of the Anuario Ornitológico de Madrid 2002(De la Puente Nilsson et al., 2002), excluding species cited as accidental.For Mammals, I use the Atlas de los mamíferos terrestres de España (Palomo & Gisbert, 2002).For main synonymies I

FISHES
Fishes species in Comunidad de Madrid are represented by a large number of molecular markers (Table 1).This result is logical if we consider that nine of twenty present species are exotic and with high economic value.Despite having a good number of sequences for Cobitis, no microsatellites has been described.Similarly, the absence of microsatellites for Gobio is remarkable.The taxonomic status of some species has been revised in recent years.In these cases, the search for the different synonymies shows a large number of molecular markers available.Additionally, a recent study (Larno et al., 2005) characterizes new microsatellite markers for Leuciscus that are probably useful in other species (Barinova et al., 2004).

AMPHIBIA
The 19 species present in Madrid are well represented in the GenBank (Table 2) except for Triturus boscai, Triturus pygmaeus and Triturus marmoratus, represented by 6, 1 and 4 sequences respectively and Hyla meridionalis, represented by 11 sequences.However, the Genus Triturus and Hyla are represented by a high number of sequences and among them they are numerous microsatellites that could be useful.All of the genera are well represented, with microsatellites described for Alytes, Bufo, Hyla, Rana, Salamandra and Triturus.In particular, microsatellites are described for Alytes cisternasii, Bufo bufo, Bufo calamita, Hyla arborea, Salamandra salamandra and Triturus alpestris.
Sequences are available from specimens sampled within the Comunidad de Madrid for Alytes cisternasii, Alytes obstetricans, Discoglossus jeanneae and Salamandra salamandra which may help in population studies.

REPTILIA
Most species present in Comunidad de Madrid are well represented by sequences in databases (Table 3), except Chalcides bedriagai and Elaphe escalaris, and less so, Blanus cinereus, Coluber hippocrepis, Psammodromus hispanicus and Vipera latastei.However, they are many available sequences for the corresponding Genus except for Blanus.
With respect to microsatellites, a short number has been described for these species and genera.Only for Coronella austriaca and Podarcis muralis do we have microsatellites in GenBank, and at Genus level, microsatellites has been characterized for Elaphe, Lacerta, Natrix, Podarcis and Vipera.I believe that a supplementary effort in order to determine microsatellites for the Reptilia inhabiting the Comunidad de Madrid is necessary.

AVES
They are a large number of molecular markers for Aves, both sedentary (Table 4) and migrants (Table 5), that allow population, biogeographic and kinship studies.However, the number of markers is small for some genera.For this reason, it is necessary to implement the characterization of a larger number of molecular markers.For instance, there are no DNA markers for Melanocorypha among the sedentary group and other genera show a small number of markers (less than 10), including:   It is necessary to remark the absence of microsatellites for Turdus and Ciconia, and the presence of only one microsatellite sequence for Corvus.The case of Philomachus is also interesting.In this genus, the nine sequences available are microsatellites.

MAMMALS
Mammals are well represented by sequences in databases except for the endangered species Galemys pyrenaicus with only four sequences deposited in GenBank.
In respect to the microsatellites, the number of sequences is large, except for Barbastella, Elyomis, Genetta, Hypsugo, Neomys, Pipistrellus, Suncus and Talpa, with no microsatellites characterized.

Conclusions
The species and genera without molecular data deserve a particular effort for the characterization of molecular markers.In particular, I think that is necessary to invest in the obtention of more sequences from the endangered Galemys pyrenaicus, and to obtain sequences for underrepresented avian genera, such as Elanus, Aegypius, Melanocorypha, Lymnocryptes and others.
On the other hand, it is also necessary to characterize microsatellites for genera with no sequences available, considering the current importance of these markers (Bruford & Wayne, 1993), and in particular, a major effort in the characterization of microsatellites in reptiles and fish should be undertaken.

Note in Proof:
Since the submission of this work and the proof corrections, and with respect to Aves, 4 microsatellites have been described for the genus Porphyrio and 16 for the species Nycticorax nycticorax.Many mitochondrial sequences have been published for the genus Scolopax.In the Reptilia, more sequences have been described for the genus Blanus, and 6 microsatellites have been described for Mauremys.Finally, many microsatellites have been described in the mammalian genus Mustela.

Table 1 .
-Sequences in databases for fish present in Comunidad de Madrid.Tabla 1.-Secuencias en las bases de datos para peces presentes en la Comunidad de Madrid.

Table 2 .
-Sequences in databases for Amphibia present in Comunidad de Madrid.Tabla 2.-Secuencias en las bases de datos para anfibios presentes en la Comunidad de Madrid.

Table 3 .
-Sequences in databases for Reptilia present in Comunidad de Madrid.Tabla 3.-Secuencias en las bases de datos para repriles presentes en la Comunidad de Madrid.

Table 4 .
-Sequences in database for sedentary Aves present in Comunidad de Madrid.Tabla 4.-Secuencias en bases de datos para aves sedentarias presentes en la Comunidad de Madrid.

Table 5 .
-Sequences in databases for migrant Aves present in Comunidad de Madrid.Tabla 5.-Secuencias en bases de datos para aves migrantes presentes en la Comunidad de Madrid.

Table 6 .
-Sequences in database for mammals present in Comunidad de Madrid.Tabla 6.-Secuencias en bases de datos para mamíferos presentes en la Comunidad de Madrid.