Oviposition of Minstrellus grandis (Lepidoptera: Riodinidae) in a harmful ant-plant symbiosis

The oviposition behavior of the rare butterfly Minstrellus grandis (Callaghan, 1999) (Lepidoptera: Riodinidae) is recorded for the first time. Two females laid eggs on the old leaves of an unidentified Triplaris Loefl. ex L. (Polygonaceae), a myrmecophytic plant typically known as ‘Triplaria’ or ‘novice’ tree, inhabited by aggressive ‘taxi’ ants of the genus Pseudomyrmex Lund. 1831 (Hymenoptera: Formicidae). These observations suggest that M. grandis caterpillars live associated with one of the most harmful types of Amazon ant-plant symbiosis.

The systematics of Riodinidae has advanced in recent years, and robust phylogenetic hypotheses supported by fossilcalibrated dating are now available (Seraphim et al. 2018). This backbone is essential to understand the evolutionary history of this remarkable group of butterflies (Kaminski et al. 2013). The main impediment for the advancement on the knowledge on this family, however, is the lack of information on natural history of species in this group, especially for rare and/or restricted species (DeVries 1997). This is the case for members of the subtribe Pachythonina (Nymphidiini), which comprises 33 species in five genera (Callaghan and Lamas 2004;Hall 2007Hall , 2018Dias et al. 2015;Gallard 2017;Gallard and Fernandez 2017). Only recently, the first life cycle information for this lineage was revealed, indicating that their caterpillars are myrmecophilous (i.e., live associated with ants), present armored larval morphology and are carnivorous (Medina 2014;Mota et al. 2020). During a collection expedition to the Serra do Divisor National Park [Parque Nacional da Serra do Divisor (PNSD)], in the western Amazon region (state of Acre, Brazil), we observed, for the first time, the female reproductive behavior of the poorly known riodinid Minstrellus grandis (Callaghan, 1999). This Pachythonina species is rare, known from only five localities in the western Amazon region in Brazil, Bolivia and Peru (Callaghan 1999;Hall 2007). Here we describe its oviposition behavior.
The observations were made in the riparian forest on the right margin of the Moa River, in a frontier-protection deployment area of the Brazilian military forces (Destacamento Especial de Fronteira de São Salvador, 61º Batalhão de Infantaria de Selva) (7°24'47.72"S, 73°12'46.55"W, 200 m), about 8 km east of the border of the PNSD. This region is characterized by a high biological diversity, including riodinid butterflies (Brown and Freitas 2002;Dolibaina et al. 2012Dolibaina et al. , 2016 The females flew slowly and walked along the plant branches, at a height of about three meters from the ground, without attracting the attention of ants (Figure 1a,b). Three oviposition events were observed in both the abaxial and adaxial surfaces of old leaves (Figure 1c). The adult butterflies were collected and are deposited (DZ 51.398 and DZ 51.408) in the collection of the Department of Zoology of Universidade Federal do Paraná, Curitiba, Paraná, Brazil (DZUP). Due to the ant aggressiveness (see Haddad et al. 2009), we were unable to search the Triplaris plant to find possible caterpillars, nor were the eggs and ant vouchers collected. The ants were identified based on behavior, photographs and taxonomist opinion (see Acknowledgments).
Our observations confirm that M. grandis uses the myrmecophytic Triplaris plant as oviposition substrate. Although we cannot confirm whether M. grandis caterpillars are myrmecophilous, nor whether the type of interaction with ants is commensal, mutualistic or parasitic, the observed behavior suggests a unique life cycle, which may represent the first known case of butterfly caterpillars specialized in Pseudomyrmex ants (Pseudomyrmecinae). These large-eyed arboreal ants have stingers and are quite aggressive, especially those that live associated with myrmecophytes, such as those of the Pseudomyrmex triplarinus (Weddell, 1850) group (sensu Chomicki et al. 2015), that are specialized on Triplaris (Benson 1985;Ward 1999;Chomicki et al. 2015;Sanchez 2015). Known associations between myrmecophilous caterpillars and Pseudomyrmex ants are rare and facultative or antagonic (Fiedler 2001;Kaminski et al. 2012).
The Triplaris-Pseudomyrmex ant-plant symbiosis is widely distributed in the Neotropical lowland forests, but the hotspot for these associations is the western part of the Amazon Basin (Sanchez 2015). Interestingly, the four species of Minstrellus Hall, 2007 are also restricted to the western Amazon (Hall 2007). Although some species of Triplaris and Pseudomyrmex are widely distributed, others have restricted distribution ranges (Ward 1999;Sanchez 2015). Thus, the precise identification of Triplaris-Pseudomyrmex associations is essential to understand their evolutionary history and their relationship with the distribution range and apparent rarity This suggests that the occurrence of M. grandis may be seasonal, with adults flying for a short period, as reported for some social parasitic species (Fiedler 1998).
Myrmecophilous caterpillars that exploit ant-plant symbiosis are known to exist (Maschwitz et al. 1984). For instance, Triplaris trees inhabited by Azteca Forel, 1878 (Dolichoderinae) ants are attacked by Theope pieridoides C. Felder & R. Felder, 1865 (Riodinidae) caterpillars (Kaminski et al. 2013). In general, myrmecophilous caterpillars are herbivorous and receive protective benefits from ant partners by inhabiting an enemy-free space (e.g., Kaminski et al. 2010). From herbivorous ancestors, however, some species evolved caterpillars with carnivorous habit and even social parasitism, when caterpillars directly exploit the ants' resources (Pierce et al. 2002). In Pachythonina, it is known that caterpillars of Pachythone xanthe H. Bates, 1868 and Pachythone gigas Godman & Salvin, 1878 are predators of scale insects associated with aggressive Azteca ants (Medina 2014;Mota et al. 2020). In common, the caterpillars have an armored morphology, perforated cupola organs and tentacular nectary organs with possible appeasing function (Mota et al. 2020). These morphological traits can be preadaptations to live with aggressive ants (see Fiedler 1998;Dupont et al. 2016).
Therefore, it would not be unlikely that M. grandis caterpillars are also able to exploit resources from the harmful Triplaris-Pseudomyrmex ant-plant symbiosis. Confirming this hypothesis may add another piece to understand the evolution of carnivory and social parasitism in Pachythonina. Thus, we hope that our report will encourage future fearless efforts to reveal the life cycle of this rare Amazon butterfly.