First record of Tettigettalna mariae Quartau & Boulard, 1995 (Insecta: Hemiptera: Cicadoidea) in Spain

Abstract Tettigettalna mariae Quartau & Boulard 1995 is recorded for the first time in Spain. Thought to be endemic to Portugal (occurring in the southern province of Algarve), the present paper adds its distribution to southern Spain, being an Iberian endemism. The acoustic signals of the new specimens collected were recorded in different localities of Huelva province, in Andalusia during August 2012. According to their present known distribution, specimens of Tettigettalna mariae tend to be sparsely distributed in small range populations in southern Iberian Peninsula, favouring wooded areas with Pinus pinea.

signals (Claridge 1985, Boulard and Mondon 1995, Simões et al. 2000, Quartau et al. 1999. Despite being common and with several thousands of species described worldwide, cicada's biodiversity is still poorly known. Recent work has drawn some concern to the diversity of species in the Iberian Peninsula (Boulard 1982, Boulard 2000, Puissant and Sueur 2010, Sueur et al. 2004, Quartau and Fonseca 1988, showing that this region is an important hotspot for Mediterranean cicada diversity. Particular attention was given recently to smaller species in southern Iberia with nine species assigned to the genus Tettigettalna, seven of which are believed to be endemic to southern Iberia (Puissant and Sueur 2010). Three species (T. mariae, T. josei and T. estrellae) have been described and recorded so far in Portugal only (Sueur et al. 2004) but their distribution range in other parts of the Iberia Peninsula has never been investigated before.
The current knowledge on the distribution boundaries of Tettigettalna species is far from being properly known and extensive field surveys for these cicadas are still missing. Moreover, some of these species are believed to have very restricted distribution ranges as is the case of T. mariae, a new species that was recently described (Quartau and Boulard 1995) and was thought to be endemic to the Algarve, the southern province of Portugal.

Materials and methods
As an outcome of intensive fieldwork in Portugal and Spain for species of the genus Tettigettalna during the summer of 2012, we report here the discovery of a new populations of T. mariae in Andalusia (Southern Spain). Identification was based on the collection of specimens and the recording of male acoustic signals typical of the species.
The field survey was conducted daily from 10:00 h to 19:00 h during sunny weather with temperatures ranging from 29°to 32°C. Searches were conducted by car while driving at a reduced speed, allowing the detection of the songs of calling males. Their acoustic signals were recorded at the collecting site using a Marantz PMD 661 Portable SD recorder (20 Hz -24 kHz) connected to a Telinga Pro 7 Dat-mic microphone (Twin Science) following the procedures given in Simões et al. 2000. Specimens were then captured by hand or by means of a sweeping net.
Geographical coordinates were determined with a GPS (Garmin, Oregon series 550t) for each site where male songs were heard and where specimens were collected. Species confirmation was accomplished with time and frequency analysis of sound recordings using the software Avisoft Sas-Lab Pro (Specht 2012) as in previous analyses (e.g. Quartau et al. 1999, Simões et al. 2000. Acoustic recordings were analyzed with a sampling rate of 44.1 kHz and a resolution of 16 bits. Spectra were computed using FFT with a resolution of 512 points and a Hamming Window. For each male, recordings of about one minute were analysed. Song terminology follows that of Gogala and Trilar (1999) and Gogala and Trilar (2000). The examined material and acoustic recordings are deposited in the general data bank on insect data at the Department of Animal Biology in the Faculty of Sciences, University of Lisbon (FCUL). Collected specimens were stored dry and a front leg was preserved in 100% ethanol for DNA isolation.

Analysis
Our analysis of the morphological and acoustic data confirmed the presence of T. mariae specimens in Spain.
Specimens were collected and recorded in different locations from Huelva province in Andalusia around the following localities: Cartaya, Aljaraque, Moguer, Mazagón, Almonte and Hinojos (Table 1). Records were sparse, even within large patches of suitable habitat, and match its current habitat preference, with T. mariae tending to favour wooded areas of Pinus pinea near the sea in the southern Iberian Peninsula (Figs 1, 2).  Fig. 3) showed the profile of the calling song in agreement with previous studies Boulard 1995, Fonseca 1991, unpublished data). T. mariae specimens have a broad spectrum near 6 -16.5 kHz with maximum energy around 12 kHz. For time domain variables, our results indicated an echeme duration ranging from 0.02 to 0.10s, with an average value of 0.06s. For the echeme period we found a range of 0.16 to 0.54s and average of 0.32s.

Discussion
Previous studies suggested that T. mariae was a Portuguese endemic cicada, seeming to be confined to central Algarve, close to the sea (Sueur et al. 2004) which is an area under increasing human pressure. The coastline of Algarve is heavily urbanized, with many touristic villages and golf courses covering most of Vilamoura, Vale do Lobo and Quinta do Lago. This raises concerns about the conservation of T. mariae given the restricted habitat range of the known populations. The discovery of T. mariae populations in Spain means that the species is not confined to the central wooded area of Algarve, close to the sea, having instead a wider distribution extending to Andalusia. The new populations of T. mariae reported here constitute an important addition to the scarce knowledge of this rare species. However, T. mariae distribution remains heavily fragmented and discontinuous. Consequently the species is still vulnerable to habitat loss caused by changes in land use or forest fires that often jeopardize Pinus pinea woods during the summer, when cicada adult males are active. These threats may cause the decline and eventual extinction of local populations of cicadas and are especially worrying for small range species such as T. mariae (Quartau and Mathias 2010).
With the present data, obtained through our 2012 fieldwork, a new cicada species is listed for Spain and a new endemism for Iberia.  Moreover, the current species list available for the cicadas from Iberian Peninsula are likely to still be incomplete. As the male acoustic signals in cicadas are highly diagnostic for the separation of closely related species (Claridge 1985, Boulard and Mondon 1995, Gogala et al. 2008, Gogala et al. 2011, it is quite possible that to the same specific name may correspond in fact two or more independent sibling species as has happened in other genera, such as Cicadetta (e.g. Gogala et al. 2008, Gogala et al. 2011. All this suggests the presence of a larger number of species in the Iberian Peninsula than those already recorded and calls for further cicada surveys in the area, as well as a better knowledge of cicada biology and ecology, which is the key to the conservation of these interesting insects in the Mediterranean area.