A case of mass occurrence of Sandalodesmus araujoi () in a municipality of São Paulo, Brazil and description of the heretofore unknown female (Polydesmida, Chelodesmidae)

Abstract Somatic characters are shared by many Chelodesmidae groups, and generic placement and species identifications traditionally have been based on gonopodal morphology. Female genitalic characters have been largely neglected and are rarely photographed or illustrated. This is rather unfortunate as the morphology of female genitalia presents important characters and may be decisive for developing a more robust family classification. We describe the heretofore unknown female of Sandalodesmus araujoi (Schubart, 1946), previously known only from the male holotype collected in São Paulo, Brazil in December 1943; discuss the utility of female genitalic characters for species delineation in Sandalodesmus; and report the first case of a mass occurrence in the Chelodesmidae. While an attempt at a formal diagnosis of Sandalodesmus females based on genitalic characters is premature, the vulvar morphology of the three taxa examined in this study suggests that female genitalia are species-specific. Some characters (i.e., asymmetric valves, presence of digitiform projections and reduction of setae on the internal basal portion of the valves) are constant between the species, suggesting utility for generic-level delineation. Mass occurrences of millipedes are typically unpredictable and likely related to variations in environmental conditions and/or anthropogenic modifications of natural habitats. Although the mass occurrence of S. araujoi reported herein was only observed once, the event coincides with the mating period of millipedes during the rainy season in Brazil. On the other hand, the region where the species was found has been the target of intense urban development, including replacement of natural habitats with residential areas, which may have influenced its population dynamics.


Introduction
With over 5,000 described species, Polydesmida is the most diverse order within the class Diplopoda and includes the two most species-rich families of millipedes, Paradoxosomatidae Daday, 1889 (approx.1,000 described species) and Chelodesmidae Cook, 1895 (750+ described species) (Hoffman 1980, Shelley 2002, Brewer et al. 2012, Nguyen and Sierwald 2013, Enghoff et al. 2015).Chelodesmidae is divided into two subfamilies, Chelodesminae (139 genera) distributed across the Neotropical region, and Prepodesminae (37 genera) known from the Afrotropical and Palearctic regions (Hoffman 1980).Currently, 21 tribes are recognized within the Chelodesmidae, with 19 of them belonging to Chelodesminae, however, more than half of the described genera and species have yet to be assigned to a tribe (Bouzan et al. 2021).
Based chiefly on gonopodal characters, Hoffman (1982) proposed the tribe Sandalodesmini to include the South American Oncoleptodesmus Schubart, 1958 andSandalodesmus Silvestri, 1902.Members of the tribe are small to moderately sized chelodesmids (25-45mm in length) with relatively slender bodies and reduced gonopods, characterized by the presence of a shield-like acropodite which conceals the solenomere in the lateral view, a subspatulate prefemoral process that partially envelops the acropodite, the presence of a spiniform process on the gonocoxa, and a modified, hook-like cannula (Hoffman 1982).Sandalodesmus differs from Oncoleptodesmus by the presence of only two macrosetae on the dorsal side of the gonocoxae and by the absence of spinulations on the lateral margin of the acropodite [except in Sandalodesmus repandus (Schubart, 1985); Silvestri 1902, Schubart 1958, Hoffman 1982, Rojas-Buffet et al. 2022].Sandalodesmus includes 14 species distributed across Argentina, Brazil, Paraguay, and Uruguay and Oncoleptodesmus includes four species only known from the Brazilian state of Rio Grande do Sul (Attems 1931(Attems , 1938;;Schubart 1958;Hoffman 1982;Rojas-Buffet et al. 2022).Sandalodesmus was reviewed by Hoffman (1982) and recently Rojas-Buffet et al. (2022) provided an updated key to the males.
Somatic characters are shared by many genera within the Chelodesmidae and generic placement and species identification have been traditionally based on gonopodal morphology (Hoffman 1971, Pena-Barbosa et al. 2013, Bouzan et al. 2017a).Female genitalic characters have been largely neglected, seldom examined, and rarely photographed or illustrated (Brölemann 1902;Schubart 1946;Hoffman 1990aHoffman , 1990bHoffman , 2000;;Pena-Barbosa et al. 2013;Bouzan et al. 2017bBouzan et al. , 2018aBouzan et al. , 2018b)).Sandalodesmus is no exception, and the females of only six of the 14 known species have been described in some, often very abbreviated, manner.To date, only the females of Sandalodesmus joachimadisi Rojas-Buffet and Bouzan, 2022 have been described in detail, including description of genitalic characters, and illustrated (Rojas-Buffet et al. 2022).This is rather unfortunate as the morphology of the female genitalia presents important characters and may be decisive for developing a more robust classification of the Chelodesmidae (see Hoffman 2012).
Mass occurrences (seemingly sudden increases in local abundance) in the Diplopoda are a result of natural cycles (reproduction swarming or lifecycle periodicity) or unpredictable changes in climate, resource availability, or fragmentation of natural habitat (David 2015, Niijima et al. 2021).Such events have been reported from Africa, Eurasia, and The Americas, and vary in magnitude from localized occurrences of hundreds of individuals to millions of individuals covering extensive areas (Cloudsley-Thompson 1949, Niijima et al. 2021).While small scale swarms can occasionally be a nuisance near human habitations in urban and suburban areas, large outbreaks have been reported to impact livestock, crop production, and even interfere with transportation (Cloudsley-Thompson 1949, Niijima et al. 2021).In an attempt to provide a more robust classification of millipede mass occurrences, Niijima et al. (2021) proposed the separation of these events into swarming (hundreds of individuals), mass emergence (thousands), and outbreak (millions), based on the number of individuals observed.
One of the largest millipede outbreaks ever recorded, occurred in West Virginia, USA in June, 1918 when an estimated 65,340,000 Apheloria virginiensis corrugata (Wood, 1864) covered 75 acres of farmland for over a month (Cloudsley-Thompson 1949).During this event, farmers were reportedly nauseated by the defensive chemicals released by the millipedes and cattle refused to graze in the impacted areas (Cloudsley-Thompson 1949).An equally impressive series of mass outbreaks have been reported for Parafontaria laminata (Attems, 1909) in Japan where mass occurrences of this species have caused multiple road and railroad obstructions between 1920 and 1984 (Niijima and Shinohara 1988, Niijima 1998, Niijima et al. 2021).Interestingly, Parafontaria laminata armigera Verhoeff, 1936 was recently shown to have a lifecycle akin to periodical cicadas with an outbreak event occurring every eight years (Niijima et al. 2021).
In Brazil, millipede population outbreaks are most commonly associated with non-native species including the Asian Oxidus gracilis (C.L. Koch, 1847) and Orthomorpha coarctata (Saussure, 1860) (Polydesmida, Paradoxosomatidae), and more recently the Portuguese millipede, Ommatoiulus moreleti (Lucas, 1860) (Julida, Julidae) (Iniesta et al. 2020(Iniesta et al. , 2021(Iniesta et al. , 2022)).Although all three species reach high densities in urban areas of southeastern Brazil, O. gracilis and O. coarctata have also become established in rural and natural environments, including forests, caves, and islands (Iniesta et al. 2021).There is little published information regarding population outbreaks of native millipedes in Brazil, except for those regarded as agricultural pests.The majority of these accounts concern species of Spirostreptida (Spirostreptidae and Pseudonannolenidae) in cultivated areas where they have been reported to cause damage to seedlings, tubers, and fruits (Schubart 1942;Boock and Lordello 1952;Lordello 1954;Boccardo et al. 1997Boccardo et al. , 2002)).To date, there have been no published accounts of swarming or population outbreaks in the Chelodesmidae.
We report the first case of a mass occurrence in the Chelodesmidae and describe the heretofore unknown female of Sandalodesmus araujoi (Schubart, 1946), previously known only from the male holotype collected in the state of São Paulo, Brazil in 1943.In addition, we discuss the utility of female genitalic characters for species delineation in Sandalodesmus and provide additional information on male characters, including measurements and SEM images.

Materials and Methods
The material examined is deposited in the following collections (curators in parentheses): Instituto Butantan, São Paulo, Brazil (IBSP; A.D. Brescovit), Arachnida and Myriapoda collection of Facultad de Ciencias, Universidad de la República, Montevideo, Uruguay (FCE-Myr; M. Simó).Scanning electron micrographs were taken using a FEI Quanta 250 SEM (FEI, Oregon, USA) with an attached SLR digital camera at Instituto Butantan.Specimens selected for scanning electron imaging were cleaned two times (30 sec.each) ultrasonically, transferred to an ascending series of ethanol dilutions (70, 80, 90, and 100%), bathed for 15 minutes at each step, and critical-point dried.The samples were mounted on aluminum stubs and coated with gold in a sputter coater for 240 seconds.In addition, relevant morphological features were photographed with a Leica DFC 500 digital camera mounted on a Leica MZ16A stereomicroscope and measured to the nearest 0.01 mm with Leica Application Suite ver.2.5.0 (Leica Camera, Wetzler, Germany).Body-rings were measured in dorsal view, and antennomere lengths were measured in lateral view.
Female genitalic characters of S. araujoi (five specimens), S. joachimadisi Rojas-Buffet and Bouzan, 2022 (two specimens) and Sandalodesmus sp.(two specimens from Estação Ecológica Jureia/ Itatins, São Paulo, Brazil) were examined and photographed in order to explore their utility in species delineation and classification.

Results
The mass occurrence reported herein was brought to the lead author's attention in October 2021 when videos of the event were sent to him by Nicoly Calcagniti (Supplementary Material 1).Subsequently, several male and female specimens were received by the authors and identified as S. araujoi.The mass occurrence took place in the municipality of Pardinho (-23.0825, -48.3852), approx.30 km southeast of the type locality of S. araujoi in the municipality of São Manuel (-22.7423, -48.6191) (Figure 1).According to the residents of the municipality, thousands of individuals were observed around house entrances, gardens, and on pavement in the morning and afternoon hours.
The locality where the mass occurrence took place is part of a transitional zone between the Cerrado and Atlantic Forest biomes in the state of São Paulo (Coutinho 2006).The area has a subhumid temperate climate and is characterized by a patchwork of herbaceous and forested habitats which have resulted from retractions and transgressions during the Quaternary (Coutinho 2006, Alvares et al. 2014, Smith and Mayle 2017).Since the second half of the 20th Century, the region has been severely impacted by human activities, including deforestation, farming practices, livestock production, and accelerated urbanization (Ratter et al. 1997).
Female (IBSP 14278): Coloration in life as in male.Similar to male in general body outline and somatic characters, except sternite of 4 th body ring with less conspicuous and separate projections, all remaining sternites and all legs without modification.Posterior margin of vulvar aperture smooth, without projections.Vulvae (Figures 4D-F 4E-F; 5B, 5D-F).In addition, the vulvae of Sandalodesmus sp. have a pronounced horizontal ridge spanning both valves and delineating the bottom third of each (Figures 5C-D).As compared to the previous two species, the valves of S. joachimadisi are relatively symmetrical, with only a slight enlargement of the external valve, and with fewer and more robust digitiform projections on the internal margin (Figures 5A-B).In addition, the lateral profile of the valves of S. joachimadisi is distinctly subtriangular while those of both S. araujoi and Sandalodesmus sp. are subrectangular, narrower near the operculum and widening towards the apex (Figures 4D, 5A, 5C).

Discussion
Although numerous collection events have targeted the area surrounding the type locality of S. araujoi, until the mass occurrence reported herein, this species was known only from the single male holotype collected in the municipality of São Manuel in December 1943 (Schubart 1946).
Millipede female genitalia undoubtedly contain valuable taxonomic information, however, they have been historically neglected and have been rarely included in species descriptions.Genitalic characters of female Sandalodesmus remain poorly known, and only those of S. joachimadisi have been described and properly illustrated (Rojas-Buffet et al. 2022: figure 3D).While an attempt at a formal diagnosis of females based on genitalic characters is premature, the vulvar morphology of the three taxa examined herein suggests that Sandalodesmus female genitalia are species-specific.In addition, some characters, such as the asymmetric (to different degrees) valves, and the presence of digitiform projections and the reduction or absence of setae on the internal basal portion of the valves, are constant between the species, suggesting utility for generic-level delineation.Additional studies on the morphology of Sandalodesmus female genitalia are needed to assess their systematic importance.
With the exception of the population outbreaks of the Japanese P. laminata armigera, mass occurrences of millipedes are typically unpredictable and likely related to variations in environmental conditions and/or anthropogenic modifications of natural habitats.Although the mass occurrence of S. araujoi reported herein was only observed once, the timing of the event coincides with the mating period of millipedes during the rainy season (October-April) in Brazil (see Alvares et al. 2013).On the other hand, the region where the species was found has been the target of intense urban development, including replacement of natural vegetation with residential areas, which may have influenced its population dynamics.More detailed studies are needed to tease apart the factors contributing to these largely unpredictable events.
The original communities of the Cerrado and the Atlantic Forest in southeastern Brazil, two of the world's premier biodiversity hotspots, have been severely altered by anthropogenic activities, such as medium to large-scale farming practices, livestock production, and accelerated urbanization, and are rapidly disappearing (Fonseca 1985, Durigan et al. 2007).To date, approximately 50% of the Cerrado has been destroyed by human activities (Rodriguez-Zorro et al. 2022), and only 10% of the original vegetation of the Atlantic Forest remains (Colombo and Joly 2010, Joly et al. 2014, Rezende et al. 2018).The region, long known for its vertebrate and invertebrate diversity and high levels of endemicity, was also recently shown to contain the highest species richness and the highest concentration of threatened Spirostreptida in Brazil making its conservation vital for the protection of numerous plant and animal taxa (Iniesta et al. 2023).

Figure 1 .
Figure 1.Known distribution of S. araujoi with the São Manuel type locality shown in yellow and the new record from Pardinho shown in red.