Revista de Biología Tropical ISSN: 0034-7744 ISSN: 0034-7744 Universidad de Costa Rica Morera, Jessica; Mora-Pineda, Geovanny; Esquivel, Alejandro; Hanson, Paul; Pinto-Tomás, Adrián A. Detection, ultrastructure and phylogeny of Sclerorhabditis neotropicalis (Nematoda: Rhabditidae) nematodes associated with the Azteca ant-Cecropia tree symbiosis Revista de Biología Tropical, vol. 66, no. 1, 2018, pp. 368-380 Universidad de Costa Rica DOI: 10.15517/rbt.v66i1.26919 Available in: http://www.redalyc.org/articulo.oa?id=44955366029 How to cite Complete issue Scientific Information System Redalyc More information about this article Network of Scientific Journals from Latin America and the Caribbean, Spain and Portugal Journal's homepage in redalyc.org Project academic non-profit, developed under the open access initiative Detection, ultrastructure and phylogeny of Sclerorhabditis neotropicalis (Nematoda: Rhabditidae) nematodes associated with the Azteca ant-Cecropia tree symbiosis Jessica Morera1, Geovanny Mora-Pineda2, Alejandro Esquivel3, Paul Hanson2 & Adrián A. Pinto-Tomás4* 1. Pharmaceutical Research Institute (INIFAR), University of Costa Rica, San José, Costa Rica; [email protected] 2. School of Biology, University of Costa Rica, San José, Costa Rica; [email protected], [email protected] 3. Nematology Department, National University of Costa Rica, San José, Costa Rica; [email protected] 4. Center for Research in Microscopic Structures (CIEMic), Center for Research in Cell and Molecular Biology, Biochemistry Department, School of Medicine, University of Costa Rica, San José, Costa Rica; [email protected] * Correspondence Recibido 20-III-2017. Corregido 08-IX-2017. Aceptado 04-X-2017. Abstract: The nematodes involved in ant-plant symbioses have been poorly studied, creating an area of research that is now receiving increasing attention. One example is Sclerorhabditis neotropicalis that inhabit the nests of Azteca ants in Cecropia trees. The goal of the present study is to increase our knowledge about these organisms and their possible symbiotic relationship with the ants and their host plant. The samples were collected during 2011-2013 in different locations in Costa Rica: Parque Nacional Carara (Puntarenas province), San Pedro de Montes de Oca and Highway 27 (San Jose) and La Selva Biological Station (Sarapiquí, Heredia). We examined 576 internodes from 23 plants, comprising four species of Cecropia (including one non-myrmecophyte) and five Azteca species. S. neotropicalis was found in all but one of the Azteca/Cecropia combinations examined. Queen and worker ants were placed in water in order to dislodge nematodes and the results were largely positive, sug- gesting that alate queens carry S. neotropicalis between trees and that workers carry them between internodes. In addition to transport, preliminary observations suggest that the ants provide masses of scraped parenchyma (possibly fertilized with feces) as a substrate for bacteria upon which the nematodes presumably feed. We also found S. neotropicalis associated with Azteca in another myrmecophyte, Cordia alliodora, suggesting that this ant/nematode association is not restricted to Cecropia. SEM photographs of S. neotropicalis were provided to supplement the original description of this species, and molecular phylogenetic analyses employing three differ- ent markers suggest that Sclerorhabditis forms a well-supported clade of bacteria-feeding nematodes associated with Azteca. Indirect evidence suggests that this is a mutualistic association, but the benefits to the ants remain unknown. Rev. Biol. Trop. 66(1): 368-380. Epub 2018 March 01. Key words: nematodes, symbiosis, electron microscopy, Cecropia, Azteca, Costa Rica. One of the best-known ant-plant mutual- principally Azteca (Berg, Franco, & Rosselli, isms occurs between trees in the genus Cecro- 2005). The genus Azteca comprises 84 species, pia (Urticaceae) and certain ant species in the all of which are arboreal, but only 13 of these genus Azteca (Formicidae: Dolichoderinae), are known to be obligate inhabitants of Cecro- both genera being restricted to the Neotropics pia (Longino, 1991b; AntWeb, 2016). (discounting human introductions of Cecro- The Cecropia tree provides a nesting site pia). About 80 % of the 61 species of Cecro- inside the hollow stems and a source of nutri- pia have a mutualistic association with ants, tion for the ants in structures called Müllerian 368 Rev. Biol. Trop. (Int. J. Trop. Biol. ISSN-0034-7744) Vol. 66(1): 368-380, March 2018 bodies located at the base of the petioles. These larvae. As the colony grows the workers chew food bodies contain glycogen and lipids, pro- holes in the septa that separate the internodes viding a valuable food supply for the ants (Longino, 1989). (Rickson, 1971). Other sources of nutrition In internodes containing Azteca larvae, include smaller food bodies located on the it is very common to find large numbers petioles and near the veins on the undersides of of nematodes inhabiting moist, dark-colored young leaves (Rickson, 1976), and honeydew mounds of fine-textured material, which appear from mealybugs (Hemiptera: Pseudococcidae) to consist primarily of parenchyma that the that the ants maintain in their nests (Trimble ants have scraped from the internode walls & Sagers, 2004). Myrmecophytic species of (Longino, 1991a). In Costa Rica, the most Cecropia (those that harbor ants) also possess common nematode appears to be Sclerorhabdi- specialized parenchyma lining the inside sur- tis neotropicalis (Rhabditidae), which probably face of young internodes, though the exact role feeds on bacteria (Esquivel, Abolafia, Hanson, of this tissue in the mutualism is still unclear & Pinto, 2012). The presence of unidentified (Valverde & Hanson, 2011). nematodes has also been reported in young The Azteca ants benefit the plant by cut- saplings recently colonized by Azteca queens ting vegetation that encroaches into the Cecro- (Weng, Nishida, Hanson, & LaPierre, 2007), pia tree and by removing insect herbivores but little else is known about the nematodes (Barnwell, 1967; Janzen, 1969; Schupp, 1986; associated with these ants. Recently, bacteria- Agrawal, 1998), although the level of protec- feeding nematodes (Diploscapter sp. and three tion depends on the species of Azteca (Longino, new species of Sclerorhabditis) were found 1991a; Trimble & Sagers, 2004), the size of associated with another ant-tree mutualism in the ant colony (Rocha & Bergallo, 1992), the Southeast Asia, Crematogaster ants (Mymici- abundance of herbivores (Schupp, 1986), and nae) living in Macaranga trees (Euphorbia- elevation (Trimble & Sagers, 2004). Azteca ceae) (Maschwitz, Fiala, Dumpert, Hashim, & ants also provide Cecropia trees with nitrogen Sudhaus, 2016). The authors of this study sug- (Sagers, Ginger, & Evans, 2000; Dejean, Petit- gested that the nematodes might be cultivated clerc, Roux, Orivel, & Leroy, 2012), and plants by the ants in refuse piles, and that they are colonized by ants grow faster than uncolonized possibly taken into the pospharyngeal gland of plants (Oliveira et al., 2015). worker ants and then fed to the larvae. Another The stems of myrmecophytic Cecropia species in the genus Sclerorhabditis, S. min- species are divided into a series of hollow iata, was recently described by Asghar-Shah internodes that are separated from one another and Vaid (2015), but in this case, the nematodes by solid partitions (septa) at each node. In were not associated with insects, as they were young stems, the upper part of each internode isolated from humus and farmyard manure, has an oval depression (prostoma) where the suggesting that this genus may have a wider wall is thinner and through which the coloniz- distribution than originally thought. ing Azteca queen chews a hole in order to enter In Costa Rica, there are seven species of (Bailey, 1922). Once inside, the queen scrapes Cecropia, five of which harbor Azteca ants plant material over the entrance hole, which (González & Rodríguez, 2015); only the three eventually closes even more tightly as callus most widely distributed species (C. peltata, tissue grows over it. The queen thus remains C. obtusifolia, C. insignis) were included in sealed inside the internode and apparently the present study. The two species lacking relies primarily on her own body reserves to ants include C. pitteri, endemic to the Isla de rear the first brood of workers. Once the first Coco, and C. angustifolia (=C. polyphlebia), workers emerge and reopen the prostoma, they which is restricted to higher elevation cloud begin foraging for Müllerian bodies, storing forests and was included in the present study. them in the internodes, presumably as food for Of the 21 Azteca ant species in the country, Rev. Biol. Trop. (Int. J. Trop. Biol. ISSN-0034-7744) Vol. 66(1): 368-380, March 2018 369 five are restricted to Cecropia (Longino, 2007). literature (Andrassy, 1984; Ahmad, Shah, & The objectives of the present study were: 1) Mahamood, 2007; Esquivel et al., 2012). Determine whether Sclerorhabditis nematodes are associated with the four Cecropia species Examination of individual internodes: examined and all five Azteca; 2) Determine Entire plants mostly 1-2 m in height were col- whether the presence of Sclerorhabditis nema- lected (cut at the base) from seven sites (Table todes in Cecropia is dependent upon the pres- 1). A total of 23 plants from four Cecropia ence of Azteca; 3) Examine queen and worker species, including the non-myrmecophytic