New distribution records, first host plant record and DNA barcoding of the Neotropical plume moth Oidaematophoruspseudotrachyphloeus Gielis (Lepidoptera, Pterophoridae)

Abstract Background Oidaematophoruspseudotrachyphloeus Gielis, 2011 (Lepidoptera, Pterophoridae) is a little-known Neotropical plume moth previously recorded in Ecuador, Peru and Argentina. Its host plants and DNA barcodes are unknown. New information Oidaematophoruspseudotrachyphloeus is recorded for the first time from Chile, based on six specimens from the Azapa Valley (Arica Province) and two from Guayacán (Coquimbo Province). Those from the Azapa Valley were reared from folivorous larvae collected on Ambrosiacumanensis Kunth (Asteraceae), representing the first host plant record for this plume moth. The first DNA barcode sequences of O.pseudotrachyphloeus are provided and used to explore relationships with congenerics.


Introduction
Oidaematophorus Wallengren, 1862 (Lepidoptera, Pterophoridae, Pterophorinae, Oidaematophorini) is a widespread genus of plume moths with 24 species described (Gielis 2003, Matthews 2005, Gielis 2011a, Gielis 2011b, Hernandez et al. 2014). Species of this genus are recognised by forewing venation with R1 absent, R2, R3, R4 and R5 separate, Cu1 from the posterior angle of the discal cell and Cu2 from the discal cell, midleg with scale bristles at base of spur pairs and female genitalia with bell-or widened funnel-shaped antrum (Gielis 2011a). Host plants are mainly Asteraceae, but feeding on representatives of Onagraceae and Plantaginaceae has been also documented (Matthews and Lott 2005).

Materials and methods
Plume moths were reared from larvae collected on Ambrosia cumanensis Kunth (Asteraceae) in the Azapa Valley, Arica Province, northern Chile. Eight larvae were collected in July 2017 and placed in plastic vials with leaves of A. cumanensis and a paper towel at the bottom. Additional leaves were provided until the larvae completed feeding and pupated, about ten days after collection. Before pupation, two larvae were placed in 95% ethanol at -20°C until DNA extraction. Adults emerged about twelve days after pupation in August 2017. A search for additional conspecifics from Chile was performed in the Colección Entomológica de la Universidad de Tarapacá (IDEA), Arica, Chile. The abdomen of each specimen was removed and placed in 10% potassium hydroxide (KOH) for a few minutes for genitalia dissection, stained with Chlorazol black and Eosin and mounted on slides with Euparal. Images of the genitalia were captured with a Sony CyberShot DSC-HX200V digital camera attached to a Leica M125 stereomicroscope. The identification of the specimens was based on comparisons with the original description of O. pseudotrachyphloeus provided by Gielis (2011a) and photographs of the genitalia slide of the male holotype deposited in the Museo de Historia Natural, Universidad Nacional Mayor San Marcos, Lima, Peru (MUSM). The distribution map was generated using SimpleMappr (Shorthouse 2010).
Genomic DNA was extracted from two larvae following the procedures described in Huanca-Mamani et al. (2015). DNA purification, PCR amplification and sequencing of the barcode fragment (Hebert et al. 2003), with the primers LCO-1490and HCO-2198(Folmer et al. 1994, were performed in Macrogen Inc. (Seoul, South Korea), following the PCR programme described in Escobar-Suárez et al. (2017). Additional sequences (Table 1) with species identification and 658 base pair length were downloaded from BOLD (Ratnasingham and Hebert 2007) for analysis, including congenerics and representatives of other genera of the tribe Oidaematophorini: Adaina Tutt, 1905, Emmelina Tutt, 1905and Hellinsia Tutt, 1905. These three genera were used as the outgroup because they are closely related to Oidaematophorus ( Gielis 1993). Unfortunately, sequences of Oidaematophorus currently available in public databases are restricted to Nearctic and Palearctic representatives. The sequences were aligned by the ClustalW method and sequence divergence was estimated by the Kimura 2-Parameter (K2P) method in MEGA X (Kumar et al. 2018). Substitution saturation was estimated with the Xia test (Xia et al. 2003) in DAMBE 7 ( Xia 2018) to assess the presence of phylogenetic information. A phylogenetic tree was inferred through a Maximum Likelihood (ML) analysis using IQ-TREE 1.6.12 (Nguyen et al. 2015) in the web interface W-IQ-TREE (Trifinopoulos et al. 2016). Data were partitioned to codon position and TNe+I, F81+F and HKY+F+G4 were selected as the best-fit models for 1st, 2nd and 3rd partitions, respectively, in ModelFinder (Kalyaanamoorthy et al. 2017). Branch support was calculated with 1,000 replications of ultrafast bootstrap (UFBoot, Hoang et al. 2017). The unrooted tree was visualised in FigTree (Rambaut 2014) to root on the representative of Hellinsia, following Gielis (1993

Taxonomic identification
Six adults (three females, three males) of O. pseudotrachyphloeus (Fig. 1) were obtained from the larvae collected on A. cumanensis in the Azapa Valley. Two additional conspecifics (one female, one male) from Guayacán (Coquimbo Province) were found in the IDEA collection.

DNA barcoding
Two identical sequences of O. pseudotrachyphloeus were obtained (GenBank accessions OK510535, OK510536), which represent the first DNA barcodes for this species. The lowest divergence (9.6% K2P) was with O. balsamorrhizae McDunnough, 1939 andO. cineraceus (Fish, 1881). Oidaematophorus was recovered as a monophyletic group in the ML analysis, but with low UFBoot support (Fig. 2). The relationships of O. pseudotrachyphloeus with congenerics were not well resolved.

Discussion
Based on previous records of O. pseudotrachyphloeus (Gielis 2011a, Ustjuzhanin et al. 2021, the northern limit of its geographic distribution is found in Loja ( Although the ML analysis here presented is based on a single mitochondrial marker, it is promising that sequences of Oidaematophorus were clustered as a monophyletic group in agreement with the most recent morphological definition of the genus (Gielis 2011a). However, the UFBoot support (74%) of this monophylum was low (Minh et al. 2013). Furthermore, as O. pseudotrachyphloeus was the only exclusively Neotropical species included in the alignment, it is not surprising that its relationships were not resolved. A more detailed taxon sampling with emphasis on Neotropical representatives of the genus and the use of additional molecular markers would be needed to understand better the evolutionary relationships of O. pseudotrachyphloeus. Eight species of Oidaematophorus were previously recorded from Chile (Gielis 1991), all of which were later transferred to Hellinsia, based on morphological evidence (Gielis 2011a): H. betsiae Gielis, 1991, H. cinerarius (Philippi, 1864, H. coquimboicus Gielis, 1991, H. grandaevus (Meyrick, 1931, H. hololeucos (Zeller, 1874), H. mallecoicus Gielis, 1991, H. mauleicus Gielis, 1991and H. siskaellus Gielis, 1991. The discovery of O. pseudotrachyphloeus confirms the presence of Oidaematophorus in Chile and highlights the importance of surveys for plume moths in the arid environments of the country as suggested by other recent discoveries (Vargas 2020, Vargas et al. 2020, Ustjuzhanin et al. 2021. Geographic distribution of Oidaematophorus pseudotrachyphloeus Gielis, 2011 in South America. Square (type locality) and triangles indicate previous records, circles indicate new records from Chile.