Introduction

Eucalyptus L’Hér. (Myrtales: Myrtaceae) is largely planted in Sumatra, Indonesia for production of paper, pulp and viscose goods (PRASETYO et al., 2019; INAIL et al., 2019). These goods are used to supply the local market as well as being imported to countries such as China, India, Malaysia, Singapore, and South Korea (SZULECKA et al., 2016). Eucalyptus plantations in Sumatra are predominantly grown with Eucalyptus pellita F. Muell. and the hybrids of Eucalyptus grandis W. Hill. × E. pellita (BOPHELA et al., 2019; TACHI et al., 2020; TAVARES et al., 2020). Several lepidopteran species have been recorded to infest Eucalyptus in Sumatra, including Ophiusa disjungens (Walker, 1858) (Erebidae) (RAIMON et al., 2020), Polyphagozerra coffeae (Nietner, 1861) (Cossidae) (TAVARES et al., 2020; TACHI et al., 2020), Strepsicrates sp. (Tortricidae) (KKADAN et al., 2020a, 2020b), and Tetracona amathealis (Walker, 1859) (Crambidae) (MELIA et al., 2021).

The Homona spargotis Meyrick, 1910 (Lepidoptera: Tortricidae), of the coffearia group, was known previously as Ericiana spargotis and native to the Northern Territory and Queensland, Australia and currently-apparently distributed in the Australasian realm, where is considered a potential pest on a variety of tropical crops (HERBISON-EVANS & CROSSLEY, 2004; HULCR et al., 2007). The larvae feed primarily on avocado, Persea americana Mill. (Laurales: Lauraceae) and sporadically on coffee, Coffea arabica L. (Gentianales: Rubiaceae), custard-apple, Annona reticulata L. (Magnoliales: Annonaceae), lychee, Litchi chinensis Sonn. (Sapindales: Sapindaceae), macadamia, Macadamia spp. (Proteales: Proteaceae), narra, Pterocarpus indicus Willd. (Fabales: Fabaceae), pariparoba, Piper umbellatum L. (Piperalis: Piperaceae), star fruit, Averrhoa bilimbi L. (Oxalidales: Oxalidaceae), and tea plant, Camellia sinensis (L.) Kuntze (Ericales: Theaceae) (HERBISON-EVANS & CROSSLEY, 2004; HULCR et al., 2007). H. spargotis has not been recorded in Australia from any native host, except Macadamia spp. and P. indicus suggesting its potential as polyphagous on rainforest plants (WHITTLE et al., 1987).

Homona spargotis larvae form a shelter made of rolled leaves held with silk where it lives and feeds. The adult moths are a patchy brown (HERBISON-EVANS & CROSSLEY, 2004), inactive during daytime and found resting in leaf litter and in the canopy (WHITTLE et al., 1987). The wingspan is about 14.0-19.5 mm for males and 20.0-28.0 mm for females (MEYRICK, 1910; WHITTLE et al., 1987). The eggs are yellow and flatted. They are laid on the upper section of the host plant crown in overlapping masses of several hundred (HERBISON-EVANS & CROSSLEY, 2004). The moths have an unusual resting posture, with the hindwings protruding from under the forewings, and the protruded parts folded over (ZBOROWSKI & EDWARDS, 2007).

Not only to identify species, DNA barcoding may also be used to establish species boundary, as recently shown to distinguish multiple species within the Homona genus (HULCR et al., 2007) and Hesperiidae (HEBERT et al., 2004). Genetic distances calculated from the highly conserved mitochondrial cytochrome c oxidase subunit I (mt-COI) gene were shown to form bimodal distribution, clearly separating genetic distance between insects belonging to the same species vs. across species, i.e., intraspecific vs. interspecific genetic distance. Insects within the same species were shown to share similar DNA sequences, corresponding to unique haplotypes associated with morphologically and ecologically distinct caterpillars (HULCR et al., 2007), hence they have a much smaller pairwise genetic distances.

The objectives of the current study were to report the finding of Homona spargotis in Sumatra, Indonesia, confirmed by molecular analysis, and to add a new plant host for this insect.

Material and methods

COLLECTION OF HOMONA SPARGOTIS

Homona spargotis larvae were collected manually from trees of an E. grandis × E. pellita hybrid in a commercial stand during the first semester of 2020 in Pangkalan Kerinci area (0º 20’ N × 101º 51’ E, 10 m altitude) in Riau, Sumatra, Indonesia. Larvae were placed in one-liter plastic containers and taken to the Entomology Laboratory of the Asia Pacific Resources International Holdings Ltd. (APRIL) of the PT. Riau Andalan Pulp and Paper (RAPP), where they were kept in a room at 26 ± 2º C, 75 ± 15% RH and 14:10 (L:D) h photoperiod. Caterpillars received daily fresh shoots of E. grandis × E. pellita as a food.

MOLECULAR IENTIFICATION OF HOMONA SPARGOTIS

One caterpillar was used for species name confirmation through molecular analysis. DNA extraction and PCR of the mt-COI gene were carried out as previously described (MELIA et al., 2021). The PCR product was then sent to a service provider for Sanger Sequencing. Trimming and consensus sequence building were carried out using the sangeranalyseR package (CHAO et al., 2020) with default settings. We used the consensus sequence to find the most similar sequence in NCBI GenBank using blastn (ALTSCHUL et al., 1990). The top hit, sorted by the highest score, is kept. The consensus sequence, top hits from the blast search as well as several sequences from the Homona sp. Meyrick were aligned using Clustal Omega (SIEVERS et al., 2011), and conserved sequences were identified using Gblocks (TALAVERA & CASTRESANA, 2007). The output was used to build a phylogenetic tree using the maximum likelihood approach (FELSENSTEIN et al., 1981) with a bootstrap value of 1,000 and GTR as the DNA substitution model (TAVARÉ, 1986), which were implemented in the phangorn R package (SCHLIEP et al., 2011).

Species boundary analyses were performed using mt-COI sequences from 65 insects belonging to the Homona genus submitted to NCBI GenBank, which were all available sequences from H. spargotis (13 sequences) and Homona aestivana (Walker, 1866) (12 sequences) as well as a maximum of 20 sequences each from Homona mermerodes Meyrick, 1910 and Homona trachyptera Diakonoff, 1941. These four species were selected following previously published data examining species determination amongst generalist moths (HULCR et al., 2007). Sequences were then aligned to find conserved blocks using the same approach described previously. The resulting aligned sequences were used to calculate genetic distance as implemented in the phangorn R package with default settings. A total of 2080 genetic distances was calculated between all possible pairwise combinations within our dataset, which were then plotted as a histogram.

Results

REPORT OF H. SPARGOTIS IN SUMATRA

This is the first report of H. spargotis in Sumatra (Fig. 1).

Fig. 1.–
Adult Homona spargotis Meyrick, 1910 (Lepidoptera: Tortricidae).

REPORT OF H. SPARGOTIS ON E. GRANDIS × E. PELLITA

The H. spargotis larvae were collected while feeding on an E. grandis × E. pellita hybrid, which represents a new host plant species for this insect.

MOLECULAR ANALYSIS

Species identification through sequencing of the mt-COI gene has become an accepted approach given the difficulty of identification through insect body morphology. Here, we report a leafroller whose closest species is H. spargotis based on molecular analysis (Table 1). This similarity is stable as our sample are clustered together with H. spargotis in 67% of the 1,000 bootsraps (Fig. 2). Despite the sequence resemblance to H. spargotis, our sample is clustered less tightly with the rest of H. spargotis, as compared to other intra-species clustering within the Homona genus (Fig. 2).

Table 1.–

Origin, score, E value, identity, and accession

Fig. 2.–
Phylogenetic tree on Homona spargotis Meyrick, 1910 (Lepidoptera: Tortricidae).

We calculated and plotted the distribution of all possible pairwise genetic distance in four Homona species included in our dendrogram (Figs. 2-3). Interestingly, we confirm the existence of bimodal distribution, separating intra species with inter species genetic distances (Fig. 3). Based on our data, any pairwise genetic distance that exceeds 0.045 are likely to come from samples of different species. The closest genetic distance of our sample to any H. spargotis is 0.053 (to H. spargotis with NCBI accession EF070841.1), which falls within the range of inter species genetic distances. Thus, we concludes our sample is Homona spargotis.

Fig. 3.–
Distribution of pairwise genetic distances amongst 65 Homona Walker (Lepidoptera: Tortricidae) samples.

Discussion

REPORT OF H. SPARGOTIS IN SUMATRA

Homona spargotis has been previously reported in the Australasian realm (HERBISON-EVANS & CROSSLEY, 2004) without however confirmation of the countries with distribution of this insect. Figures of the male aedeagus in DIAKONOFF (1939, 1948) of specimens collected from Java, Indonesia agree with those of H. spargotis in WHITTLE et al., (1987). Other specimens collected beyond Australia to New Guinea (WHITTLE et al., 1987) and in Sri Lanka (KOCHANSKY et al., 1978) are also confirmed as H. spargotis. Specimens collected in Sulawesi are similar to those of H. spargotis described in WHITTLE et al., (1987). The combination of a semicircular costal fold on the fore wing together with dark grey hind wings readily distinguishes the male of H. spargotis from any other species of Homona Walker, 1863 known from Australia (WHITTLE et al., 1987).

REPORT OF H. SPARGOTIS ON E. GRANDIS × E. PELLITA

Homona spargotis, the closest described species from H. spargotis collected in the present study while feeding on a E. grandis × E. pellita clone, has been previously recorded as a predominant pest of P. americana and a sporadic pest of C. arabica, A. reticulata, L. chinensis, Macadamia spp., P. indicus, P. umbellatum, A. bilimbi, and C. sinensis. On P. americana, the infestation of H. spargotis appears to be determined by the presence of growth flushes. Numbers increase quickly at flushing and the moth virtually disappears when no new shoots are present (WHITTLE et al., 1987).

Homona spargotis, in the current study, was seen along with the tea mosquito bug, Helopeltis theivora Waterhouse, 1886 (Hemiptera: Miridae), a sapsucker and other mirid species on the commercial stands of E. grandis × E. pellita clone in Sumatra (KKADAN et al., 2020c). Potential monitoring and control measures of H. spargotis include the use of light traps (HULCR et al., 2007) and sex pheromone traps (for monitoring) (WHITTLE et al., 1987). Homona spargotis is recorded to have two flying activity periods on P. americana in Australia, one from March to June and another from August to December however its moths were collected using light traps throughout the summer months (WHITTLE et al., 1987).

MOLECULAR ANALYSIS

We assessed the possibility of our sample belonging to another species that is similar to H. spargotis following an approach previously used to determine species boundaries in six tortricid moths (HULCR et al., 2007) and Hesperiidae (HEBERT et al., 2004). Both studies showed that genetic distances within species are much closer, as compared to between species, which can be exploited to draw species boundaries.

The confirmation of the existence of bimodal distribution, separating intra species with inter species genetic distances has been previously reported (HULCR et al., 2007; HEBERT et al., 2004).

Conclusions

The used molecular protocol was able to identify the insect species as H. spargotis through analysis of its larvae, which extends the geographical distribution to Sumatra and add a new host plant family (i.e., Myrtaceae) and genus (i.e., Eucalyptus) and a host species (i.e., E. grandis and E. pellita) for this insect.

Acknowledgments

Thanks to Dr. Buck Richardson (LeapFrogOz, Kuranda Kreations, Queensland, Australia) for providing the figure 1. The following Indonesian companies provided financial support to the study: PT. Riau Andalan Pulp and Paper (RAPP) and Asia Pacific Resources International Holdings Ltd. (APRIL).

BIBLIOGRAPHY

ALTSCHUL, S. F., GISH, W., MILLER, W., MYERS, E. W. & LIPMAN, D. J., 1990.– Basic local alignment search tool.– Journal of Molecular Biology, 215(3): 403-410.

BOPHELA, K. N., VENTER, S. N., WINGFIELD, M. J., DURAN, A., TARIGAN, M. & COUTINHO, T. A., 2019.– Xanthomonas perforans: A tomato and pepper pathogen associated with bacterial blight and dieback of Eucalyptus pellita seedlings in Indonesia.– Australasian Plant Pathology, 48(1): 543-551.

CHAO, K. H., BARTON, B., PALMER, S. & LANFEAR, R., 2020.– New results sangeranalyseR: Simple and interactive analysis of Sanger sequencing data in R.– bioRxiv: Available from https://doi.org/10.1101/2020.05.18.102459.

DIAKONOFF, A., 1939.– The genera of Indo-Malayan and Papuan Tortricidae.– Zoologische Mededelingen (Leiden), 21(1): 111-238.

DIAKONOFF, A., 1948.– Records and descriptions of Microlepidoptera.– Treubia, 19(3): 483-524.

FELSENSTEIN, J., 1981.– Evolutionary trees from DNA sequences: A maximum likelihood approach.– Journal of Molecular Evolution, 17(1): 368-376.

HERBISON-EVANS, D. & CROSSLEY, S., 2004.– Homona spargotis Meyrick, 1910.– Australian caterpillars and their butterflies and moths. Available at: http://lepidoptera.butterflyhouse.com.au/tort/spargotis.html (accessed on: 14 September 2020).

HEBERT, P., PENTON, E., BURNS, J., JANZEN, D. & HALLWACHS, W., 2004.– Ten species in one: DNA barcoding reveals cryptic species in the neotropical skipper butterfly Astraptes fulgerator.– Proceedings of the National Academy of Sciences, 101(41): 14812-14817.

HULCR, J., MILLER, S. E., SETLIFF, G. P., DARROW, K., MUELLER, N. D., HEBERT, P. D. N. & WEIBLEN, G. D., 2007.– DNA barcoding confirms polyphagy in a generalist moth, Homona mermerodes (Lepidoptera: Tortricidae).– Molecular Ecology Notes, 7(4): 549-557.

INAIL, M. A., HARDIYANTO, E. B. & MENDHAM, D. S., 2019.– Growth responses of Eucalyptus pellita F. Muell plantations in South Sumatra to macronutrient fertilisers following several rotations of Acacia mangium Willd.– Forests, 10(12): 1054.

KKADAN, S. K., DURAN, A., WIJAYA, R., SINULINGGA, N. G. H. B., TAVARES, W. DE S., TARIGAN, M., WONG, C. Y. & SHARMA, M., 2020b.– Impact of water quality on insecticide efficacy.– International Pest Control, 62(6): 314-317.

KKADAN, S. K., SIRAIT, B. A., ASFA, R., TAVARES, W. DE S., TARIGAN, M., DURAN, A., WONG, C. Y. & SHARMA, M., 2020a.– Evaluation of a spinetoram-based insecticide against lepidopteran and thrips infesting acacia and eucalyptus in Sumatra, Indonesia.– Journal of Entomology and Zoology Studies, 8(2): 1345-1351.

KKADAN, S. K., YESHWANTH, H. M., TAVARES, W. DE S., PASARIBU, I., ABAD, J. I. M., TARIGAN, M., DURAN, A., WONG, C. Y. & SHARMA, M., 2020c.– Mirid pests of Eucalyptus in Indonesia: Notes on damage symptoms, alternate hosts and parasitoid.– Journal of the Kansas Entomological Society, 92(4): 577-588.

KOCHANSKY, J. P., ROELOFS, W. L. & SIVAPALAN, P., 1978.– Sex pheromone of the tea tortrix moth (Homonacoffearia Nietner).– Journal of Chemical Ecology, 4(1): 623-631.

MELIA, T., SINULINGGA, N. G. H. B., MARETHA, M. V., WIJAYA, R., EFENDI, R. R., OLIVEIRA, L. S. S., KKADAN, S. K., TAVARES, W. DE S., TARIGAN, M. & DURAN, A., 2021.– First report of Tetracona (Agrotera) amathealis (Walker, 1859) on a Eucalyptus grandis W. Hill × Eucalyptuspellita F. Muell. (Myrtaceae) hybrid in Indonesia (Lepidoptera: Crambidae, Spilomelinae).– SHILAP Revista de lepidopterología, 49(195): 407-412.

MEYRICK, E., 1910.– Revision of Australian Tortricina.– Proceedings of the Linnean Society of New South Wales, 35(145): 215.

PRASETYO, A., AISO-SANADA, H., ISHIGURI, F., WAHYUDI, I., WIJAYA, I. P. G., OHSHIMA, J. & YOKOTA, S., 2019.– Variations in anatomical characteristics and predicted paper quality of three Eucalyptus species planted in Indonesia.– Wood Science and Technology, 53(1): 1409-1423.

RAIMON, LAKSAMANA, A., SINULINGGA, N. G. H. B., FERLIANDA, I. A., SAMOSIR, M. N., PANE, S. B., KKADAN, S. K., TAVARES, W. DE S., TARIGAN, M. & DURAN, A., 2020.– First report of Ophiusa disjungens(Walker, 1858) (Lepidoptera: Erebidae) on Acacia mangium (Fabaceae), and damage and notes of its biology on Eucalyptus (Myrtaceae) commercial plantations in Sumatra, Indonesia.– SHILAP Revista de lepidopterología, 48(191): 439-447.

SCHLIEP, K. P., 2011.– Phangorn: Phylogenetic analysis in R.– Bioinformatics, 27(4): 592-593.

SIEVERS, F., WILM, A., DINEEN, D., GIBSON, T. J., KARPLUS, K., LI, W., LOPEZ, R., MCWILLIAM, H., REMMERT, M., SODING, J., THOMPSON, J. D. & HIGGINS, D. G., 2011.– Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega.– Molecular Systems Biology, 7: 539.

SZULECKA, J., OBIDZINSKI, K. & DERMAWAN, A., 2016.– Corporate–society engagement in plantation forestry in Indonesia: Evolving approaches and their implications.– Forest Policy and Economics, 62(1): 19-29.

TACHI, T., SHIMA, H., TAVARES, W. DE S. & TARIGAN, M., 2020.– A new species of Cossidophaga Baranov from Indonesia (Diptera: Tachinidae), a parasitoid of the carpenter moth, Polyphagozerra coffeae (Nietner) (Lepidoptera: Cossidae).– Oriental Insects, 54(4): 545-555.

TALAVERA, G. & CASTRESANA, J., 2007.– Improvement of phylogenies after removing divergent and ambiguously aligned blocks from protein sequence alignments.– Systematic Biology, 56(4): 564-577.

TAVARÉ, S., 1986.– Some probabilistic and statistical problems in the analysis of DNA sequences.– Lectures on Mathematics in the Life Sciences, 17(2): 57-86.

TAVARES, W. DE S., KKADAN, S. K., HENDRIK, A. M., TARIGAN, M., ASFA, R., YAKOVLEV, R. V., TACHI, T., DURAN, A., WONG, C. Y. & SHARMA, M., 2020.– Notes on the biology and natural enemies of Polyphagozerra coffeae (Nietner, 1861) infesting Eucalyptus pellita F. Muell. (Myrtaceae) trees in Riau, Indonesia (Lepidoptera: Cossidae, Zeuzerinae).– SHILAP Revista de lepidopterología, 48(190): 333-349.

WHITTLE, C. P., BELLAS, T. E., HORAK, M. & PINESE, B., 1987.– The sex pheromone and taxonomic status of Homona spargotis Meyrick sp. rev., an Australian pest species of the Coffearia group (Lepidoptera: Tortricidae: Torticinae).– Australian Journal of Entomology, 26(2): 169-179.

ZBOROWSKI, P. & EDWARDS, T., 2007.– A Guide to Australian Moths: 103 pp. CSIRO Publishing, Clayton South.

Notas de autor

Autor para la correspondencia / Corresponding authorWagner_Tavares@aprilasia.com