A new species of Loxosceles Heineken & Lowe (Araneae, Sicariidae), with updated distribution records and biogeographical comments for the species from Mexico, including a new record of Loxoscelesrufescens (Dufour)

Abstract A new species of the spider genus Loxosceles Heineken & Lowe, 1832, Loxoscelesmalintzisp. n., is described from the states of Puebla, Morelos and Guerrero, in the central region of Mexico. The description is based on adult males and females with morphological and ultra-morphological images. Updated distribution maps are provided for the 39 species recorded from the Mexican territory (including the new species). The states with the greatest diversity are Baja California Sur, Baja California and Sonora, with five species each. A total of 441 records for the 39 species, based on arachnological collections, data bases and literature, were used to update the distribution maps. Loxoscelesboneti Gertsch, 1958 is the species with the highest number of records in Mexico, with a total of 58 records from different localities. The states with the most records so far are Guerrero, with 55 records, Morelos, with 35 records, and Baja California Sur, with 30 records. Loxoscelesrufescens (Dufour, 1820), an introduced species, is recorded for the second time in Mexico, from the state of Chihuahua, being the first well-documented record for the country. Mexico has the greatest diversity of species of Loxosceles worldwide, with 39 (two introduced species) of the 134 described species. Additionally, biogeographical comments for the species from Mexico are provided.


Introduction
Spiders of the genus Loxosceles Heineken & Lowe, 1832 are better known in North America as "violin spiders", "recluse spiders", or "brown recluse spiders"; commonly known by the medical community and general public to cause dermonecrotic lesions caused by their poisonous bites and the venom component, Sphingomyelinase D, an enzyme that destroys endothelial cells lining the blood vessels (Vetter and Barger 2002;Vetter and Bush 2002;Vetter et al. 2003Vetter et al. , 2009Wendell 2003;Da Silva et al. 2004;Vetter 2005Vetter , 2008Vetter , 2015Sandidge and Hopwood 2005;Ramos-Rodríguez and Méndez 2008;Manríquez and Silva 2009;Swanson and Vetter 2009). The genus Loxosceles belongs to the spider family Sicariidae Keyserling, 1880, which comprises three genera: Hexophthalma Karsch, 1879, with six species from Africa, Sicarius Walckenaer, 1847, with 21 species distributed in Central and South America, and Loxosceles, with 134 described species worldwide (Magalhães et al. 2017;World Spider Catalog 2018). Recently, Souza and Ferreira (2018) described the first troglomorphic species of Loxosceles from caves of Brazil. According to Binford et al. (2008), species of Loxosceles are classified into eight species groups: reclusa, laeta, amazonica, gaucho, spadicea, rufescens, vonwredei and spinulosa. However, the species group amazonica was merged with the species group rufescens by Duncan et al. (2010) based on molecular data. The reclusa group has the highest diversity, with 50 species, all from North America, primarily Mexico (Gertsch and Ennik 1983). Mexico has the highest diversity of recluse spiders worldwide, with 39 recorded species, of which 37 are native (including the new species herein described) and two are introduced species: Loxosceles reclusa Mulaik, 1940 andLoxosceles rufescens (Dufour, 1820) (Gertsch 1958(Gertsch , 1973Gertsch and Ennik 1983;World Spider Catalog 2018). The first species described from Mexico was Loxosceles yucatana Chamberlin & Ivie, 1938 from the Yucatan Peninsula. The most complete systematic revision for North American species of Loxosceles was published by Gertsch and Ennik (1983), describing 20 new species from Mexico. Thus, this was the last and most complete taxonomic revision for the species that occur in the country. The most recent taxonomic contribution for the species of Loxosceles from Mexico was the description of the male of Loxosceles mulege by Jiménez and Llinas (2005) from Baja California Sur.
Some North American synanthropic species of Loxosceles, such as L. reclusa in the United States, have been closely studied for their biological, medical and physiological aspects, analyzing their abundances, distribution and natural history (Vetter and Barger 2002;Vetter and Bush 2002;Vetter et al. 2003Vetter et al. , 2009Wendell 2003;Vetter 2005Vetter , 2008Sandidge and Hopwood 2005;Swanson and Vetter 2009). However, these aspects are poorly known for species from Mexico. It is not yet known whether the introduced synanthropic species collected in houses and buildings may also be collected in natural areas around the houses. In 2017, four collectors collected around 40 Loxosceles misteca in two hours from a house in the state of Tlaxcala, Mexico. However, the species has never been collected in natural areas in the state (Valdez-Mondragón et al. 2018). This has been previously reported by Fischer and Vasconcellos-Neto (2005) with L. laeta and L. intermedia from South America, where these spiders are almost absent from natural areas immediately surrounding the infested buildings where they were collected. Additional research is required for the species from Mexico that have been reported from urban areas.
The primary aim of this paper is to describe a new species of Loxosceles from the central region of Mexico, distributed in the states of Puebla, Morelos and Guerrero. Additionally, we update the distribution records for the Mexican territory providing new records, including that of L. rufescens, an introduced species from the Mediterranean Basin and the Middle East (Nentwig et al. 2017;Tahami et al. 2017). Finally, we discuss the biogeography of the species of Loxosceles from Mexico based on biotic provinces.

Material and methods
The specimens were hand collected and deposited in ethanol (80%) in the Colección Nacional de Arácnidos (CNAN), Institute of Biology, Universidad Nacional Autónoma de México (IBUNAM), Mexico City, and the Laboratorio de Aracnología (LATLAX), Laboratorio Regional de Biodiversidad y Cultivo de Tejidos Vegetales (LBCTV), IBUNAM, Tlaxcala City. The descriptions and observations of the specimens were made using a Zeiss Discovery V8 stereoscope. A Zeiss Axiocam 506 color camera attached to a Zeiss AXIO Zoom V16 stereoscope was used to photograph the specimens. The male palps and female genitalia were dissected in ethanol (80%). The female genitalia were cleaned in potassium hydroxide (KOH-10%) for 5 to 10 minutes. The habitus, female genitalia and palps were submerged in 96% alcohol gel (ethanol) and covered with a thin layer of liquid ethanol (80%) to minimize diffraction during photography (Valdez-Mondragón and Francke 2015). For the electron micrographs, the morphological structures were dissected and cleaned with an ultrasonic cleaner at 20-40 kHz, critical-point dried, and examined at low vacuum in a Hitachi S-2460N scanning electron microscope (SEM). The descriptions were done following Gertsch and Ennik (1983) and Tahami et al. (2017). Morphological nomenclature follows Ramírez (2014), Planas and Rivera (2015) and Magalhães et al. (2017a, b). All measurements are in millimeters (mm). Measurements on electron micrographs are in micrometers (μm). To update the distribution maps, we used literature, databases and networks, mainly of CNAN, LATLAX and the Global Biodiversity Information Facility (GBIF) (http://www.gbif.org). The records of GBIF belong to specimens that where identified by A. Valdez-Mondragón (first author) in 2007 and by W. J. Gertsch (various dates). The specimens were deposited in the CNAN, in Mexico the second representative and diverse biological collection of Loxosceles after LATLAX, which was revised. Nine fieldtrips were made to different states in Mexico to collect additional material of different species: Puebla   Etymology. The species epithet is a noun in apposition and refers to the volcano "La Malinche, Malintzi or Matlalcueye" (meaning "blue skirt" in Nahuatl language), a seismically active volcano (4,420 m) of the Transmexican Volcanic Belt, located in the states of Tlaxcala and Puebla. This last state is where the type locality is located.
Genital area: Seminal receptacles visible by transparency in ventral view (Fig. 31). Seminal receptacles asymmetric, finger-shaped (Fig. 30). Right lobe long and curved, with one small accessory lobe receptacle next to it. Left lobe long, less curved than right one, without accessory receptacles. Base of seminal receptacles wide and strongly sclerotized, directed toward each other in oblique position (Fig. 30). See variation section for more details.
There is little variation in the shape of the male palps, even those from different populations (Figs 55-62). The seminal receptacles of females are asymmetrical and are broadly variable in shape, even in the specimens from the same locality (Figs 63-66). Some specimens have long and wide curved receptacles, finger-shaped (Figs 63, 65), with small accessory lobes receptacles on each side, more visible in some specimens than others (Figs 63, 65). Others have long and thin seminal receptacles (Figs 64,  66). The base of the seminal receptacles is variable; in some specimens wider, rounded and strongly sclerotized, directed towards each other in oblique position, but in other specimens, the base is slightly sclerotized and thinner (Figs 63-67).
Remarks. Gertsch (1958) and Gertsch and Ennik (1983) reported Loxosceles zapoteca Gertsch, 1958 (female specimen) and Loxosceles boneti Gertsch, 1958 (immature specimen) from the state of Puebla, with Loxosceles malintzi sp. n. being the third species from the state (Figs 75, 78). However, in the case of L. zapoteca, males from Puebla are unknown, so we cannot corroborate the accurate identity of the species. In the collected material of L. malintzi sp. n. from localities near Acatlán de Osorio where L. zapoteca was reported (Fig. 75), only males of the new species were collected but no males of L. zapoteca. Also, although there is high variation in the seminal receptacles in L. malintzi (Figs 63-67), the seminal receptacles are completely different from those of L. zapoteca (Gertsch and Ennik 1983: figs 48-52). Also, the male palp and female genitalia are different in both species (Gertsch and Ennik 1983: figs 32-35, 48-51). The record of L. boneti from Puebla is also doubtful: the specimen is an immature, and the type locality of L. boneti is Acapulco, Guerrero, 250 km from Puebla (Fig. 75).
Natural history. The specimens of Loxosceles malintzi sp. n. were collected in a tropical deciduous forest (Figs 11-16). The micro habitat where the specimens were collected was under and among large rocks, and inside of rotten and dry cactus of the genus Opuntia and Pachycereus (arrows,Figs 13,14,17). At the type locality, the specimens were collected close together on a live large cactus (Pachycereus). They were collected at night when males are more active. These specimens were collected at 1.5-2.0 m high in the live cactus where their webs where located. In addition, the new species has anthropogenic habits: the specimens from San Pablo Anicano, Puebla were collected inside a house, under a concrete laundry sink and among concrete blocks in a yard. Even an adult male was collected at night walking on the kitchen floor of the house.

Distribution. Loxosceles rufescens
Remarks. In Mexico, L. rufescens is only known from two records, from the states of Tamaulipas and Chihuahua (Fig. 73). Chickering (1937) reported L. rufescens from San Carlos Mountains, Tamaulipas; however, he never described or illustrated any specimen, which makes his record questionable (Fig. 74).
Updated distribution records for the 39 species of Loxosceles from Mexico. A total of 461 records of the 39 species of Loxosceles distributed in Mexico were reviewed. Twenty records were discarded for not having complete localities or having doubtful georeferences. Thus, a total of 441 records were used to make the distribution maps (Figs 73-76). The states with the most records are Guerrero with 55, Mo- relos with 35, and Baja California Sur with 30 (Fig. 75). The state of Tabasco only has a single record (Appendix 1, Fig. 75). The most diverse states are Baja California Sur, Baja California, Sonora (with five species each), Guerrero, Tamaulipas (with four species each), and Oaxaca, Puebla, Hidalgo, Coahuila, San Luis Potosí, Nuevo León (with three species each) (Figs 73,74). The least diverse states are Durango, Zacatecas, Michoacán, Querétaro, Chihuahua and Sinaloa (with two species each); Jalisco, Guanajuato, Quintana Roo, Colima, Chiapas, Yucatan, Campeche, Tabasco, Veracruz, Mexico City, Nayarit, Aguascalientes, Tlaxcala and state of Mexico (with a single species each) (Figs 73-76).
Regarding the number of total records per species of Loxosceles, the species with the most records are L. boneti with 59 and L. colima with 57 (Fig. 75). The species with the least number of records are L. barbara, L. carmena, L. francisca, L. insula, L. luteola and L. rufescens with a single record each (Fig. 74). A new record of L. misteca was found for Tlaxcala (Figs 75). A third record of L. reclusa was found for Tamaulipas (Fig. 74). The record of L. rufescens from Ciudad Juárez, Chihuahua represents the second record for the country of this introduced species and the first well-documented and illustrated record from Mexico (Figs 68-72, 73, Appendix 1).

Discussion
All 32 states of the Mexican Republic, including Mexico City, have records of some native or introduced species of Loxosceles (Figs 73-77). Regarding the distribution of species of Loxosceles in Mexico, although the highest diversity of species is in the north-west, more field work is necessary to collect additional material, mainly from the Baja California Peninsula where the species have been described based on one specimen of one sex (male or female) or few specimens (Fig. 73) (Gertsch and Ennik 1983).
Regarding the introduced species in Mexico, the record of L. rufescens from Ciudad Juárez, Chihuahua (Fig. 73), a widely distributed species throughout the Mediterranean Basin and the Middle East (Nentwig et al. 2017;Tahami et al. 2017), is the first well documented and illustrated record of this species from Mexico (Figs 68-72). The third record of L. reclusa from Mexico was found for the state of Tamaulipas. The first two records were recorded by Gertsch and Ennik (1983) (Fig. 74). Loxosceles reclusa is an introduced species. The natural distribution is from the south-central United States, from southern Illinois south to Texas and from eastern Tennessee west to Kansas (Saupe et al. 2011: fig. 2A, B). The records of L. arizonica from Coahuila state are doubtful (Fig.  74) because the natural distribution of the species is from Arizona, USA. For the central region of Mexico, some of the records are introduced species in temperate climates and mainly in urban zones. Such is the case of L. misteca in Mexico City and Tlaxcala (reported for the first time) (Fig. 75). The type locality of L. misteca is from Taxco de Alarcón, in Guerrero state; it is a common species in tourist caves of the state such as Grutas de Cacahuamilpa, Grutas del Mogote, Pozo Melendez, and Cave of Carlos Pacheco. These caves are located in a tropical deciduous forest, a habitat preferred by many species of Loxosceles from Mexico, mainly from the Pacific region (Figs 11-17). Regarding the species of Loxosceles from Mexico City, Durán-Barrón and Ayala-Islas (2007) and Durán-Barrón et al. (2009) reported two species: L. misteca and one undetermined species (Loxosceles sp.), probably a immature specimen of L. misteca. Also, Gertsch (1958) recorded L. nahuana for Mexico City based on an adult female; however, Gertsch and Ennik (1983) only cited specimens of L. nahuana from Hidalgo state where this species is distributed (Fig. 75), which makes the record this species in Mexico City questionable.
According to the biogeographical scheme for Mexico by Morrone (2004Morrone ( , 2005, all biogeographical provinces have recorded species of Loxosceles (Fig. 77). The highest diversity of species of Loxosceles from Mexico is towards the north, and the diversity tends to decrease towards the south of the country (Figs 73-76). The records of Loxosceles from Mexico are located mainly in biogeographical provinces of lowlands and in dry and tropical forests, including tropical deciduous forests, and also deserts, such as Baja California, Del Cabo, Sonorense, North Altiplano, Pacific Coast, Sierra Madre del Sur and Depression of the Balsas provinces (where L. malintzi sp. n. is distributed, Fig. 78) (Fig. 77). Although most of the species of Loxosceles from Mexico are distributed in tropical deciduous forest (Figs 11-17), species such as L. chinateca and L. yucatana are distributed in tropical rain forests. Loxosceles chinateca is from the states of Oaxaca and Veracruz (Gulf of Mexico and Oaxaca provinces), whereas L. yucatana is from the states of Chiapas, Tabasco and Yucatan Peninsula (Gulf of Mexico, Peten and Yucatan provinces) (Fig. 76). The records of Loxosceles in biogeographical provinces with mountains at high elevations (> 2000 m.a.s.l.), temperate climates, and with pine, oak or oak-pine forest are scarce. Such is the case of the Sierra Madre Occidental, highlands of North Altiplano, South Altiplano, Transmexican Volcanic Belt and Los Altos de Chiapas provinces, where some records of Loxosceles might be those of introduced species (Fig. 77). This idea is supported by ecological niche modeling for the species of Loxosceles from Mexico (in press). In the case of the Sierra Madre Oriental province, composed of high mountains and temperate and mountain mesophyll forests, the records of Loxosceles are mainly from the east of the province where the elevations are lower and the climate is more tropical (Fig. 77). Many of these records are from karstic caves, one of the preferred microhabitats of some species from Mexico (e.g. L. misteca, L. boneti, L. chinateca, L. tehuana, L. tenango and L. yucatana).