Wasmannia Forel ( Hymenoptera : Formicidae : Myrmicinae ) in Argentina : Systematics and Distribution

The ant genus Wasmannia (Formicidae: Myrmicinae) was created by Forel in 1893. Initially, this genus was included in the tribe Ochetomyrmicini because its member species are very similar to those of the genus Ochetomyrmex Mayr (Fernández, 2003). One hundred and ten years later, Bolton (2003) included it in the tribe Blepharidattini, a sister group of the fungus growing ants, Attini s.s. (Shultz & Meier, 1995). However, Ward et al. (2015) recently synonymized the tribe Blepharidattini under Attini. Abstract The ant genus Wasmannia is endemic to the Neotropics, with 10 species occurring within the presumptive native range for the genus from Mexico to Argentina. Only the little fire ant, Wasmannia auropunctata is widely distributed being present from central-eastern Argentina to Bermuda, and has become infamous due to its recent worldwide expansion and status as an invasive pest. The objective of this work was to study the systematics and distribution of Wasmannia in its southern limit of distribution in Argentina. Out of the three species reported so far for Argentina, only W. auropunctata was found to be widely distributed, but abundant only in disturbed habitats mostly in the Northeast. Herein, the distribution of Wasmannia auropunctata is extended and its queen and male castes are redescribed, along with descriptions of gynandromorphs (specimens with left side of the head similar to a male and right side similar to a queen). Wasmannia sulcaticeps and W. williamsoni are much less common and widespread. W. sulcaticepsis mostly found in mountain forests in northwestern Argentina, whereas W. williamsoni is only found in shrublands and grasslands in central eastern Argentina, and most frequently in mountain grasslands. Both species overlap with W. auropunctata, which is more common in the lowlands, between approximately 400 and 1000 m elevation. The queen of W. williamsoni is described and queen and male of W. sulcaticeps are redescribed. A new species, Wasmannia longiseta n. sp. Cuezzo and Calcaterra, recently found in northeastern Argentina, is described based on worker morphology. Wasmannia rochai is recorded for the first time in Misiones, extending its distribution range from São Paulo (Brazil) to Misiones in northeastern Argentina. A key to the worker caste is provided. A cladistic analysis based on discrete and continuous morphological characters is presented as a first attempt to clarify the phylogenetic relationships between the known species of Wasmannia. Sociobiology An international journal on social insects


Introduction
The ant genus Wasmannia (Formicidae: Myrmicinae) was created by Forel in 1893.Initially, this genus was included in the tribe Ochetomyrmicini because its member species are very similar to those of the genus Ochetomyrmex Solenopsidini, Fernández, 2007) is currently inferred as the most closely related genus.All three genera are now included in the tribe Attini (new sense).All species of Wasmannia are small ants that can be differentiated from Blepharidatta andAllomerus by 1) having shallow, always well-developed antennal scrobes, 2) posterior margin of the vertex not pronounced as lobes or teeth (as in Blepharidatta), 3) petiolar node with a distinct anterior and dorsal face, and 4) irregularly striated head, at least in part, in full face view (Longino & Fernandez, 2007).
Besides these characters, the eight known species of Allomerus inhabit internal cavities of plants and have the propodeum unarmed to bidentate.The well-developed spines between the dorsal and posterior face of the propodeum, so typical of Wasmannia, are absent in Allomerus.The body of workers in Allomerus is always smooth and shining, resembling more a small Solenopsis than a worker of Wasmannia.
The genus Wasmannia is mostly endemic to the Neotropics, with ten species occurring from Argentina to Mexico, according to the last taxonomic review (Longino & Fernández, 2007).The little fire ant Wasmannia auropunctata Roger (1863) (LFA) is the most widely distributed species of the genus, being present from central-eastern Argentina north to the Caribbean and Bermuda (Wetterer & Porter, 2003).It has spread fairly recently throughout Pacific and Atlantic islands, and the Mediterranean region (e.g.Israel), and has become a serious pest in Hawaii and the Galapagos, disrupting agricultural practices and threatening wildlife (Foucaud et al., 2010).Kusnezov (1952) hypothesized parthenogenetic reproductionby this species given the high prevalence of colonies containing only females, this was later confirmed by Fournier (et al., 2005).The LFA displays extraordinary reproductive polymorphism with both regular sexual and unusual clonal populations (Foucaud et al., 2007).This rare type of clonality was also recently reported for the invasive longhorn crazy ant Paratrechina longicornis (Latreille) (Pearcy et al., 2011).This unusual reproductive system is postulated to be responsible for unicoloniality, in which individuals from different nests form a large supercolony, an attribute that probably contributes to the success as an invader (Orivel et al., 2009).
At present, only three species of Wasmannia have been reported from Argentina: W. auropunctata, W. sulcaticeps Emery (1894), and W. williamsoni Kusnezov (1952).According to Longino & Fernández (2007), W. sulcaticeps and W. williamsoni are related endemic species that occur in the far southern range of the genus and, as stated by Kusnezov (1952), they could be the poorest competitors within the genus, whereas W. auropunctata appears to be the most successful and competitive species of the genus, with a pool of derived attributes and a wide distribution.
The main objectives of this work are to (1) update the distribution of Wasmannia in Argentina, (2) describe a new species for Argentina (Wasmannia longiseta n. sp.), and (3) study the phylogenetic position of the Argentinean species in relation to the rest of the genus.

Materials and methods
This study was conducted by examining specimens of Wasmannia deposited at the Instituto y Fundación Miguel Lillo (IFML) ant collection, Tucumán, Argentina, and new specimens collected during surveys carried out by L. A.C. and L.C. between 2008 and 2013 in northern and central Argentina.The new material examined, including types, was deposited in the IFML, and vouchers were deposited in the Fundación para el Estudio de Especies Invasivas (FuEDEI), Hurlingham, Buenos Aires, Argentina.
General terminology and abbreviations are based on Longino and Fernández (2007), Serna and Mackay (2010) and Serna et al. (2011).Measurements (in mm.) were taken using a Leitz stereoscope at 40x magnification.
HL: Head Length in full face view, measured on a straight line from the anterior margin of the clypeus to the frontovertexal margin of the head.
HW: Head Width in full face view, perpendicular to HL, excluding the compound eyes.
EL: Maximum Length of the compound Eye in dorsal view.SL: Scape Length in dorsal view, excluding the radicule.AD: Maximum Distance between frontal carinae.WL: Weber's Length, measured on a straight line in lateral view of the mesosoma from the most anterior part of the prothorax to the postero-ventral angle of the propodeum, including the propodeal lobes.
PSL: Length of the Propodeal Spine in lateral view measured on an imaginary straight line from its insertion in the propodeum to the apical point of the spine.
PD: Peduncular Length, in lateral view, measured from the petiolar point of insertion in the mesosoma to the anterior face of the petiolar node.
Characters (see Table 1 for a list of the characters and explanations) were obtained from 1) recently collected material, 2) samples deposited in the IFML collection, and 3) literature reports (Wheeler, 1915;Kempf, 1965Kempf, , 1967;;Gonçalves, 1961;Wheeler & Wheeler, 1991;Snelling & Longino, 1992;Longino & Fernández, 2007).A new source of characters, provided by lineal morphometry, normally used to define ant taxa was also included in this phylogenetic approach.In the alpha taxonomic literature there is a vast amount of linear morphometric data that could be useful to infer phylogenetic relationships or, at least, to improve clade support, but they are seldom used.One possible reason are the numerous arguments against the use of continuous characters in phylogenetic reconstructions (Rae, 1998;Wiens, 2001); however, most objections relate to how best to encode those characters (Pimentel & Riggins,1987;Cranston & Humphries, 1988;Stevens,1991), rather than reasons for excluding them from the analysis (Goloboff et al., 2006).
Herein, continuous queen and worker character measurements are expressed as a range of values (minimum to maximum), except in those species whose descriptions are based on only one specimen (see Table 1).The data matrix of continuous characters for gynes and workers (characters 0-11) is shown in Appendix 1, whereas the data matrix of discrete morphological characters for workers (characters 12-45) is shown in Appendix 2.
The resulting character matrix was analyzed with TNT version 1.1 (Goloboff et al., 2008b) under parsimony with implicit enumeration (exact search) using combined data sets for discrete and continuous characters.The discrete character number 44 was considered additive, whereas the rest of the discrete characters were codified and analyzed as non-additive.Continuous characters are automatically considered as additive by TNT.To measure clade support, the original matrix was subjected to symmetric resampling (Goloboff et al., 2003b) with 1000 replicates,(P = 0.25) and cut = 25.Values of symmetric resampling are given as frequency differences (GC values).

Key to workers of Wasmannia
The key of Longino and Fernández (2007) to the worker caste of Wasmannia species was modified to include W. longiseta n. sp.together with all the other species (except W. villosa) as follows: 1-Petiolar node strongly quadrate in lateral view; petiolar peduncle about as long as node; setae on mesosomal dorsum simple and erect, color red brown to orange …..
Color reddish-yellow to orange brown, variable.Frons between frontal carinae punctate, covered with irregular striae.Only 5-6 striae reach vertexal margin of head.Occipital margin of head with short, curved setae.One curved and long seta present at posterior end of frontal carinae.There are three long hairs on each frontal carina, arranged longitudinally and curved inwards.long, in dorsal view.Gaster weakly punctuate, with long, curved setae, scattered along each segment.
Comments.This species is easily recognized by the strongly square shape of the petiolar node in lateral view, with a sharply defined dorsal and anterior face in worker and queen.As pointed out by Longino and Fernández (2007), there is considerable variation in the development of head sculpture and color.
Gyne Comments.Two sympatric sizes of gynes were described in Costa Rica on the basis of their head size (Longino & Fernández, 2007) and could represent either differences between cryptic species or intraspecific polymorphisms.Gyne variation in head size was not observed in Argentina.Argentinean queens were most similar to the small headed form found in Costa Rica, Jamaica, and Venezuela (Longino & Fernández, 2007).According to Kusnezov (1952), the worker size of populations of W. auropunctata from Misiones province differ from the size of workers found in other populations elsewhere in Argentina; however, we have not observed such variation.
Male Diagnosis.These are unusual sexual caste specimens because the left side of their head and antennae are similar to those of a normal male, but the right side has the characteristics of a queen.In one specimen, both compound eyes are less developed than in a normal male, but larger than a queen´s eyes (EL: 0.325).In the second specimen (Figs.3a-c) the left side of the head, including eyes and antennae, is similar to a male, while the right side is similar to a queen (Figs.3a and  3c).In this specimen both antenna scrobes are well developed.In this last specimen, the left antenna is similar to a male and the right one has the same shape of the queen antenna.In both specimens, meso and metasoma are similar to a normal male (Figs 3b-c), with similar wing venation and a male external genitalia.The only difference noted in the genitalia is referred to the development of the telomere.In both gynandromorphs the telomere is more stout (distal width: 0.1 mm) than in a normal male (Fig 3c).Pygostyle is also longer than in a normal male.
Comments.The two specimens of gynandromorphs were found in a nest of W. auropunctata with putatively sexual castes reproducing clonally (see below), in Colón (32º14´S, 58º08´W), Entre Ríos province, Argentina.This is the first record of gynandromorphs occurring naturally in a LFA colony, however, additional gynandromorphs have been found recently in natural populations of W. auropunctata in São Sebastian, Brazil (LAC, unpublished data) and also found by F. Cuezzo in series of W. auropunctata collected by N. Kusnezov and deposited in IFML.Gynandromorphs previously have been found in clonal LFA colonies that were exposed to temperatures of more than 40°C in the laboratory (Olivier Rey, personal communication).They could have been induced in the laboratory during embryonic development by the effect of the extremely high rearing temperature.Gynandromorphism is common in Hymenoptera (ants: Jones & Phillips, 1985;bees: Wcislo et al., 2004).These sexual mosaics have been reported in more than 40 ant species (Jones &Phillips, 1985) but, to date, their production has only been explained in two cases: sub-lethally high breeding temperature in the pharaoh's ant, Monomorium pharaonis (L.) (Berndt & Kremer, 1982), and Wolbachia infections in the isopod Armadillidium vulgare (Rigaud & Juchault, 1993).Wasmannia longiseta Cuezzo and Calcaterra n.sp.

Diagnosis (worker)
. This species is recognized by the following combination of characters: 1) head in dorsal view reticulate; 2) Malar space and postgena also reticulate without longitudinal carina as in other spp. of Wasmannia, and 3) gaster smooth and shiny, with abundant long (>1 mm), curved and whitish setae.
Etymology.The name of the new species refers to the long whitish setae that are present on its gaster.Comments: No other species of Wasmannia shows the particular set of characters described in the diagnosis.This species is morphologically similar to W. affinis and W. lutzi in several aspects (cephalic sculpture and general shape of the petiole), but differs in the number of setae in the vertexal margin, which is fewer in W. affinis and W. lutzi.These two species have a petiolar peduncle shorter than the length of the petiole in lateral view.
Color reddish-yellow to orange.Frons, between frontal carina, punctuate with irregular striae weakly marked, reaching the vertexal margin of the head.Occipital margin of the head with short, curved setae.Frontal carina with four long hairs arranged longitudinally along an imaginary line that runs along the anterior edges and curves.Antenna with 11 segments.Antennal scrobes shallow with sculpture similar to rest of head but without striae.Punctuate sculpture more defined laterally on head in full face view.Ventral margin of scrobe weakly developed.Disc of clypeus with several weakly developed striae anteriorly divergent.Masticatory margin of mandible with five teeth, no denticles and basal margin without teeth or denticles.Compound eye well developed, protruding from lateral margin of headin full face view.Malar space with 5-7 longitudinal irregular carinae.Vertexal margin straight with median notch.Promesonotum with four pairs of long, clavate, and curved setae (length approx.0.1 mm).Humeral angle well developed with one long and curved hair.Mesosomal dorsum rugose with 6-8 longitudinal carinae strongly developed on anterior half of pronotum.Propodeum with one pair of curved setae shorter than those of promesonotum.Short propodeal spines, divergent in dorsal view.In lateral view, propodeal spines shorter than length of petiole and posteriorly directed, with wide base.Petiole triangular, with 1 pair of long, curved setae, similar in longitud to those of promesonotum, anterior margin well differentiated, joining rest of profile in a curve, profile without defined ridge or angle.Mesosoma, petiole, and postpetiole in lateral view, strongly spotted.Metapleural gland strongly developed, bulky.Propodeal lobe rounded and well developed.Petiolar peduncle shorter than petiole in lateral view.Long, acute spine ispresent on anterior ventral margin of peduncle.In dorsal view petiole with rounded anterior edge.In dorsal view postpetiole square and wider than long with four long and curved setae disposed in a middle line.Gaster feebly punctuate, with long curved setae, scattered along each segment.

Male unknown
Comments.The main characters to easily recognize workers of W. rochai, and separate this species from other of Wasmannia, are the presence of curved and clavate hairs on the dorsum of the mesosoma, short propodeal spines with a wide base, and a strong and well developed spine in the anteroventral part of the petiole.This species has been recorded from Costa Rica to São Paulo State, Brazil (Longino & Fernández, 2007).We extend its distribution north to Mexico and south to the seccional Yacuí (25º41´S, 54º26´W, 243m) of the Parque Nacional Iguazú, Misiones, Argentina.This species co-occurred with W. longiseta n. sp. in a secondary rainy subtropical forest belonging to the Paranaense phytogeographical province (Atlantic Forest ecoregion).This species is not as behavioral aggressive as W. auropuntata but is considered as a pest, particularly in the cocoa plantations of the southeast and southwest regions of Bahia State, Brazil (Souza et al., 2009).Color variable from dark brown to yellowish brown in the workers of the same nest.Area between frontal carinaerugoreticulatewith some regular carinae (8-10) reaching the frontovertexal margin.Malar space with four well developed longitudinal carinae.Antenna with 11 segments.Scrobe shallow and foveate.The ventral margin of scrobe formed by preocular carina which reaches frontovertexal margin.Clypeus with eight welldeveloped not bifurcated longitudinal carinae, the four central carinae reach posterior margin of clypeal disc and continue anteriorly.Sculpture between carinae foveate, bothonclypeal disc andfrons.Masticatory margin of mandible with 5 teeth and no denticles, basal margin smooth and without teeth.Vertexal margin weakly concave to straight, with six erect setae curved anteriorly, the two outer hairswith length similar to innermost and located on frontovertexal corner.Compound eye well developed, protruding from lateral margin of headin full face view.Promesonotum with eight longitudinal and irregular carinae.Propodeal spines long and straight, posteriorly directed.Propodeal declivity foveate.Petiole with two pairs of long setae weakly spatulatehairs; anterior margin distinct, forming a curve with the rest of the petiole.Anterior face not separated from dorsum nor defined by an angle.Petiole and postpetiole stronglyfoveate.Petiolelonger than wide in dorsal view; postpetiolewider than long and with rounded corners.Petiolar peduncle shorter than length of petiole in lateral view, with small anteroventral spine.First tergum of gasterfinely striate, with abundant, whitish, long and curved setae,thickenedatits apex.

Worker
Gyne Compound eye well-developed, bulky.Antennae with 11 segments, terminal club with two antenomeres.Dorsal surface of mandible with longitudinal thin striae.Masticatory margin of mandible with five teeth.Malar space rugo-reticulate.Disc of clypeus longitudinally striate, well developed, similar to striae between frontal carinae.Pronotum poorly developed antero-dorsally, so that scutum in dorsal view encompasses more than half of mesosoma.Humeral angle well developed, angulate, not rounded.Mesonotal dorsumwith more than 20 strong, longitudinal and very regular striae.Axilla well developed, almost triangular and medially continuous by a narrow strip of integument.Scutellum strongly striate.Anapleural sulcus completely separates anepisternum from katepisternum, but sulcus weakly impressed.Anepisternum, katepisternum and metakatepisternum finely striate.Propodeal spine short and thick at base.Propodeal lobe acute.Cinctus 1 and 2 well-developed.Petiolar peduncle about same length as petiolar node in lateral view.Sterno-postpetiolar process shaped as blunt and short spine.Metasoma 3 weakly punctate and covered with abundant and subdecumbent, thin setae.
Male (n = 1): HL: 0.52; HW: 0.47; EL: 0.22; WL: 0.92.Dark brown, with yellowish mandibles, antennae and legs.Head rectangular in full face view.Mandible triangular with five teeth.Antenna with 13 segments, last antennomere of funicle longest.Scape not reaching frontovertexal margin of head.Compound eye well developed, covering half of lateral side of head.Three ocelli well developed; lateral ocellus reaching frontovertexal margin of head.Area between ocelli with 4-5 strongly impressed rugae.Malar space reduced, crossed by two strong longitudinal carinae.Rest of head in full face view punctuate and crossed by longitudinal and thin striae.Clypeal disc with several parallel and longitudinal carinae.Fore wing with three closed cells, plus open radial cell.Pterostigma present.Hind wing with one basal closed cell.Hamuli with 4 hooks.Peduncle short.Petiolar node triangular in profile with blunt apex and no dorsal margin.Abdominal sternum IX (=subgenital plate) medially projecting as triangle and distally rounded.Pygostyle well developed, one segmented.Telomere short and thick.Digitus and cuspis not observed.
Comment: Because of the scarcity of material (only one male and one dealate gyne could be studied) the descriptions of alates of W. sulcaticeps must be considered as provisional.
Geographical distribution.Wasmannia sulcaticeps was mostly found in mountain forests in northwestern Argentina (Yungas ecoregion), overlapping with W. auropunctata generally between 400-700 meters of altitude in secondary forests, such as the Parque Nacional Calilegua (Jujuy) and Parque Provincial La Florida (Tucumán) (Fig 4).Additional individuals of this species were also found for the first time in secondary forest in the Catamarca province and in lowlands in a secondary gallery forest in the Reserva Natural Iberá, in the Corrientes province (Fig 4).
Reddish-brown with gaster slightly darker than rest of body, sometimes slightly darker than W. auropunctata.Head in dorsal view with erect, short (<0.05 mm) and abundant setae.
Head between frontal carinae, from posterior margin of the clypeal disc to the frontovertexal margin covered with very regular longitudinal carinae (approx.10-12).Carinae present in malar space.Disc of clypeus bears more than 6 longitudinal and irregular carinae.Antennal scrobe with a short medial longitudinal carina that barely exceeds posterior margin of compound eye.Ventral margin of antennal scrobe defined by preocular carina that reaches frontovertexal margin.Antennal scape fails to reach frontovertexal margin.Antenna with 11 segments.Compound eye well developed, protruding from lateral margin of head in full face view.Frontovertexal corner bears long, curved seta at each side.Humeral angle acute, strongly developed.Pro-and mesonotum with 8-10 longitudinal carinae.Propleura with four longitudinal carinae and foveolate, sometimes weakly developed, meso and metapleura foveolate, without carinae.Rounded epicnemial process well developed.Dorsal and lateral sides of propodeum foveolate.In lateral view propodeal spine with wide base.Propodeal spine length less than petiole in lateral view.In dorsal view propodeal spine joined by curved line.Metapleural gland well developed and bulky, protruding beyond mesosomal profile.Propodeal lobe well developed, foveolate and almost rectangular with round angles.Short peduncle.Petiole in dorsal view slightly narrower than postpetiole, both heavily foveolate.In lateral view petiolar node rounded.Sternopostpetiolar process well developed with short spine.Metasomal tergite 3 (fourth abdominal segment) weakly reticulate.Gaster covered with long, weakly curved and sparse hairs.
Comments: This species was only known from its type locality in Castex (Espinal ecoregion), La Pampa, in central Argentina.A new survey was conducted during 2011 to search forits presence in Buenos Aires and La Pampa provinces.However, its presence could only be confirmed 59 years after its discovery by a small colony found under a stone next to a Caldén tree (Prosopis caldenia) scrubland in Castex.The collecting site was located in the ecotone between the Pampeana and Espinal ecoregions.Unfortunately, gynes or males were not found.In 2012, a few workers of this species were captured by C. Ramos in three pitfall traps placed in Olavarría and Coronel Suárez in southern Buenos Aires and Macachín in western La Pampa province, and by S. Santoandre in six pitfall traps placed between 436-1025 m elevation in the Parque Provincial Ernesto Tornquist, next to the locality of Sierra de la Ventana, also in southern Buenos Aires.A new survey was conducted in these sites in 2013 to try to collect sexuals of this species.Fortunately, two small colonies (one of them containing a queen) were manually found under small stones at 412 and 536 m elevation in the park Tornquist.This protected area is located in the Austral district of the Pampeana ecoregions (Cabrera & Willink,1980).The climate in this mountainous region is temperate, with higher humidity in the inter-mountains areas.Annual rainfall ranges between 500 -800 mm (De Fina, 1992).The mean temperature in summer and winter is of 20.5 and 8°C, respectively; the absolute maximum temperature in summer is of 40°C, while in winter frosts are frequent and it occasionally snows.

Phylogeny of Wasmannia
Only one tree was obtained under implicit enumeration (Fig 9).L = 93.648,CI=0.718, and RI = 0.613, which showed a low level of homoplasy.The cladistic analysis treating each dataset (continuous or discrete characters) separately could not resolve all the relationships between the species of Wasmannia, resulting in a polytomous tree.
The analysis shows that the clade [Blepharidatta + Allomerus + Wasmannia] proposed by Ward et al. (2015) based on molecular data is extremely well supported (GC= 91) by morphological data.Wasmannia is as a monophyletic group, defined by the following synapomorphies: worker: antennal scrobe with sculpture (character 19) and postpeciolar shape (character 34).
Although the cladogram shows high resolution for the genus, group support was relatively low for some clades within the genus.

Discussion
The only species widely distributed in Argentina is the little fire ant, W. auropunctata, while the other four recorded species are rare and/or inconspicuous.The distribution of the W. auropunctata known from previous studies by Kusnezov (1952), Kempf (1972), Cuezzo (1998), and Fuentes et al.(1998) (Buenos Aires, Entre Ríos, Córdoba, Santa Fe, Corrientes, Chaco, Formosa, Tucumán, Jujuy, Salta and Misiones) was extended in this work to the provinces of Santiago del Estero, Catamarca, and La Rioja.W. auropunctata was not found in natural/native habitats of the Monte ecoregion, nor in the Patagonian ecoregion (L.A.C., unpublished data).The finding of W. auropunctata in Lozano (southern Buenos Aires province, 34º51´S) extends the known range 100 km further south than previously recorded in northeastern Buenos Aires (Reserva Natural Otamendi; 34º13'S) (Fuentes et al., 1998).
The wide distribution and relatively high abundance of W. auropunctata could be explained by the fact it has the largest and most fecund queen within the genus (Kusnezov, 1952;Longino & Fernández, 2007).However, the higher abundance of W. auropunctata in disturbed habitats, mostly in central-eastern Argentina, seems to be more related with the fact that sexual castes are produced almost exclusively by clonal reproduction (clonal populations) in this region (Rey et al. 2012, LAC, unpublished data).
All individuals collected in Argentina morphologically identified as W. auropunctata were genetically grouped in clade B within W. auropunctata (as defined by Mikheyev & Mueller, 2007), which shares genetic similarities with several populations from the invasive range (e.g.Israel, New Caledonia, and Western Africa) (Rey et al. 2012, LAC, unpublished data).
Interestingly, molecular evidence revealed that clonal populations present in the floodplain of the Paraná River in the locality of Zárate (Buenos Aires, Fig 4) are the putative source population of the W. auropunctata introduction in Israel.Clonal queens from Israel display a multilocus microsatellite genotype similar to that found in queens from Zárate; the Israeli clonal males also display a multilocus haploid genotype remarkably similar to the clonal males found in Zárate.As in the Israeli population, this Argentinean clonal population seems to be more adapted to low temperatures than those clonal populations present in tropical regions (Rey et al., 2012).
The finding of clonal populations of W. auropunctata up to 2125 m elevation in the locality of Tumbaya, Jujuy, was surprising because it had been previously found only up to approximately 1070 m elevation (Wetterer & Porter, 2003).The discovery of W. auropunctata in the Puna ecosystem (Tumbaya, Jujuy) under extremely cold and dry conditions (annual rainfall 179 mm and mean and minimum temperatures of 8.1ºC and -8ºC in July, the coldest month, De Fina, 1992) was unexpected.According to Kusnezov (1952), W. auropucntata was not able to resist prolonged drought.However, our finding supports laboratory evidence that clonal populations present in Argentina are strongly adapted to very low temperatures and probably also to low levels of humidity in environments such as the Puna.This record represents both the highest altitude and the most severe natural environmental conditions reported so far for this species.
The other four species found in Argentina (W.sulcaticeps, W. rochai, W. williamsoni, and W. longiseta n. sp.) were much less common than W. auropunctata, and they were mostly present in natural and/or disturbed habitats.W. sulcaticeps was recorded for the first time for Corrientes and Catamarca provinces; it was previously known only from Buenos Aires, Santa Fé, Córdoba, Tucumán, Salta, and Jujuy provinces (Cuezzo, 1998;Vittar & Cuezzo, 2008).Although intensive surveys were conducted at multiple sites and in different biogeographic regions of Argentina, W. williamsoni was only found in central-eastern Argentina, suggesting it may be a relict endemic species.W. williamsoni was more common in the Parque Provincial Ernest Tornquist.This park protects several rare and endemic species of the Ventania mountainous system (Sellés-Martínez, 2001), which originates from the Tertiary period (around 22 million years ago).This could be the case of W. williamsoni, which is mostly restricted to this region and seems to show very small populations confined mainly to a specific habitat type.According to Longino & Fernández (2007), W. williamsoni and W. sulcaticeps are two related species that occur at the far southern limit of distribution of the genus and, as stated by Kusnezov (1952), could be the most primitive members of the genus, acting in the present as relicts.It is important to note that both species overlapped with W. auropunctata (more commonly in the lowlands) between approximately 400 and 1000 m elevation.
The cladistic analysis shows that the genus Wasmannia is monophyletic and sister to the genus Allomerus.Only four clades within the genus Wasmannia were strongly supported.One of them showed that the new species found in northeastern Argentina (W.longiseta n. sp.) is the sister species of W. affinis, while W. lutzi is the sister species of this latter clade.The clade [W.lutzi +W.longiseta + W. affinis] is characterized by sharing a unique development of the antennal scrobe.Wasmannia affinis and W. lutzi are, according to Longino and Fernádez (2007), related species distributed from São Paulo to Paraná states in southeastern Brazil, the neighboring region to where W. longiseta n. sp.occurs.
Another well supported clade was [W.rochai + W. sigmoidea].These are two allopatric species; W. rochai is known to occur from Guatemala to São Paulo state in Brazil, now extending its southern limit of distribution to northeastern Argentina, whereas the current reported range of W. sigmoidea is smaller and apparently restricted to the Caribbean region (Guiana, Antilles St. Vincent, Grenada, Puerto Rico, Costa Rica, and Venezuela) (Longino & Fernández 2007).In St. Vincent, W. sigmoidea can co-occur with W. auropunctata (Kuznezov, 1963).However, this species was recently reported from several surveys conducted by Brazilian researchers in northeastern Brazil (e.g. in the state of Bahia), and thus both species can co-occur at least in this region as well.
W. sulcaticeps and W. williamsoni were resolved as sister species with relative high support, which is concordant with Longino and Fernández (2007) who suggested that these are two closely related species.The size of workers of both species (W.sulcaticeps and W. williamsoni ) is similar and much larger than workers of other known species of Wasmannia.The hypothesis presented by Kusnezov (1952), which mentioned that these species may be primitive members of the genus, is partially supported by this analysis.
The position of W. auropunctata in relation to the other species of the genus could not be well clarified.Although the most parsimonious tree places W. auropunctata as a sister group of [W.longiseta + W. affinis + W. lutzi], this grouping is not well supported by symmetric resampling.New studies based in further sources of characters (such as molecular) will be useful to clarify the relationships that are not well supported in the analysis here presented.
The morphometric characters used in this work, rarely used by myrmecologists as a source of phylogenetic information, have proved to be useful in resolving the relationships of species within the genus and in adding support to several clades.
(n = 17): HL: 0.69-0.74;HW: 0.77-0.84;EL: 0.21-0.26;WL: 1.47-1.57.Color and pilosity similar to worker.Head wider behind compound eyes.Scape barely reaches vertexal margin.Frontal carina separated by 10-12 longitudinal, poorly developed and irregular striae.Antennal scrobe deep and reticulate, with longitudinal carina that starts atventral margin of antennal torulus and almost reaches posterior margin of compound eye.Preocular carina runs along ventral margin of scrobe almost to occipital angle of head.Compound eye well-developed, located close to anterior margin of head, and protruding.Antena with 11 segments; apical club with two antennomeres.Mandibular dorsum with longitudinal, thin striae.Masticatory margin of mandible with five teeth.Malar space with three to four longitudinal striae.Disc of clypeus with more than 10 well developed, longitudinal striae.Pronotum poorly developed anterodorsally scutum encompasses more than half of mesosoma in dorsal view.Humeral angle rounded.Mesonotum dorsally striate, with thin, irregular striae and poorly developed.Axilla well developed, almost triangular and continuous in midline by narrow strip of integument.Middle area of scutellum with a deep, longitudinal groove.Anapleural sulcus completely divides anepisternum from the katepisternum.Anepisternum, katepisternum and metakatepisternum finely striate.Propodeal spine thick at its base.Propodeal lobe rounded.Fore and hind wings infuscate.Fore wing with three closed cells: costal, radial (=basal) and cubital (=subbasal).No closed discal cell.Hind wing with a closed radial cell and cubital cell, almost entirely delimited by veins.Hamuli composed by five hooks.Cinctus 1 and 2 well-developed.Petiolar peduncle longer than petiolar node in lateral view.Sterno-postpetiolar process sharp and short.Metasoma III weakly punctuate, covered with sparse, subdecumbent and thin setae.

Fig 4 .
Fig 4. Main distribution regions of the five species of Wasmannia found in Argentina, including several sites mentioned in the text.
WorkerHolotype:HL: 0.50; HW: 0.42; EL: 0.10; SL: 0.40; AD: 0.25;PSL: 0.175;WL: 0.52; PD: 0.10; PTL: 0.10; PPTL: 0.12; PTW: 0.12; PPTW: 0.15; CI: 0.85; OI: 0.20.Paratype workers (n=2): HL: 0.50-0.52;HW: 0.42-0.45;EL: 0.10; SL: 0.40; AD: 0.25; PSL: 0.175;WL: 0.47-0.5;PD: 0.10-0.12;PTL: 0.10-0.17;PPTL: 0.12; PTW: 0.10-0.12;PPTW: 0.15; CI: 0.85-0.86;OI: 0.20.Reddish-yellow head, mesosoma and anterior metasoma, with posterior half of first gastral segment and posteriorly dark brown.Frons between frontal carinae rugo-reticulate with irregular striae, 5-6 striae reach vertexal margin of head.Three long simple setae present on margin of each frontal carina separated by distance similar to or greater than length of hairs.Frontal carinae with three erect hairs, shorter than previous ones, arranged longitudinally along an imaginary line that runs along the anterior edges.The vertexal margin has six simple setae curved towards the frons (Fig5a).Antenna with 11 segments.Antennal scrobe shallow with weak sculpture similar to rest of the head.Preocular carina runs along ventral margin of scrobe, reaching posterior margin of compound eye.Disc of clypeus with four longitudinal and well developed carinae.Each carina forked toward posterior margin of clypeus and with four simple setae with a similar length to those of frontal carina.Reticulate sculpture between frontalcarinae, both in clypeus and frons.Masticatory margin of mandible with five teeth, no denticlesand basal margin without teeth or denticles.Compound eye well developed, protruding from lateral margin of headin full face view.Malar space with 4-5 longitudinal and irregular carinae.Vertexal margin slightly concave medially.Promesonotum with 3-4 pairs of long, weakly spatulate and curved setae (length approx.0.1 mm).Humeral angle strongly developed with long hair on angle.Mesosomal dorsum with three longitudinal carinae that intersect at irregular intervals forming weak grid that doesnot obscure stippling of integument.Propodeum with two pairs of curved setae shorter than promesonotal setae.Long propodeal spines, with length similar to t hairs of mesosomal dorsum.In lateral view, propodeal spines longer than length of petiole and posteriorly directed.In dorsal view propodeal spines diverge from one another.Metapleural gland strongly developed, bulky.Posteropropodeal lobe rounded and well developed.Petiole with three pairs of long setae (0.1-0.15 mm), anterior margin well differentiated, forming gentle curve with rest of petiolar profile.Mesosoma, petiole, and postpetiole in lateral view, strongly spotted.Petiolar peduncle longer than length of petiole in lateral view.Short acute spine present on anterior ventral margin of peduncle.In dorsal view petiole long with a rounded anterior edge, tapering forward; in lateral view, node of petiole with distinct anterior face not forming acute angle with dorsal margin (Fig5b), posterior face of petiolar node weakly developed.In dorsal view, postpetiole ovaland barely wider than long.Gaster smooth and shiny, with abundant long (>1 mm), curved and whitish setae.