Corresponding author: Shigeki Kobayashi (
Academic editor: E. van Nieukerken
This paper provides new taxonomic and biological data on a complex of gracillariid moths in the endemic genus
Kobayashi S, Johns CA, Lopez-Vaamonde C, Doorenweerd C, Kawakita A, Ohshima I, Lees DC, Hanabergh S, Kawahara AY (2018) Hawaiian
Hawaii constitutes one of the most geographically isolated archipelagos and harbors thousands of unusual, highly threatened endemic species. Phytophagous insects that rely on endemic Hawaiian plants are of special risk as they depend on the survival of their native host plants. The Hawaiian Islands measure just 0.02% of the area of the United States, but account for nearly 70% of the United States’ historically documented plant and animal extinctions (
Larval host plants of
In late April 2016, several of the authors collected numerous blotch mines on leaves of
All adult moths were reared from leaf mining larvae and their pupal cocoons. Leaf mines and cocoons were collected between 2013–2016 in the locations listed in Table
Study sites of
No. | Locality | Island | Collection Longitude and latitude | Elevation (m) | Study Specimens ID | Species name | Host plant |
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1 | Limahuli, Upper Preserve | Kauai |
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900 | AYK-HI10-001, 002 |
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Unknown |
2 | Kokee | Kauai |
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1230 | CJ-433, 442 |
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3 | Kahili | Kauai | No data | 400–500 | CJ-148 |
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4 | Mt. Kaala | Oahu |
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800 | CJ-526 |
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5 | Kamakou | Molokai |
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1170 | CJ-241 |
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6 | Eke | Maui |
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870 | CJ-136, 531 |
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7 | Kauaula* | Maui |
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900 | CJ-381 |
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8 | Waikamoi | Maui |
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1800 | CJ-539 |
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9 | Upper Hamakua Ditch Trail | Hawaii |
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900 | CLV6239 |
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10 | Kohala Watershed Partnership | Hawaii | No data | 700–1500 | CJ-419 |
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11 | Kaumana Trail | Hawaii | 19.45°N, 155.21–155.19°W | 900–1000 | HILO016 |
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12 | Hawai’i Volcanoes National Park† | Hawaii |
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1090 | SKH-5, 10, 13, 15; | ||
HILO053, 054, 059; |
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AYK0001, 0002, CLV6240 |
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Type locality of *
Descriptions focused on the adult stage and leaf mines because of limitations of other material, and because these stages provide a wealth of morphological traits useful for diagnosis. Photographs of leaf mines were taken primarily in the field using Canon EOS 60D and 5D MKIII digital cameras. Some leafmines were scanned using an EPSON Perfection V600 Photo scanner. Observations and measurements were made under a Leica M2 16 dissection microscope at 71–115× and a Leica S6E microscope at 6.3–40× with the aid of a micrometer scale. Images of adults were captured using a Olympus E-330 camera and Moticam 580 5.0 MP. Images were taken at various depths and subsequently stacked using the Helicon Focus 6.22. All images were then edited with Adobe Photoshop Elements 9 into final figures.
For genitalic dissections, the whole abdomen was removed and boiled for 3–4 min in 10% aqueous KOH, and residual scales and soft parts were removed in 70% ethanol. Genitalia were then stained in Chlorazol Black E (1% solution in 70 % ethanol) or acetocarmine for 0.5–1h, dehydrated in a series of 70−100 % ethanol and mounted in Canada balsam on a glass slide.
Type material and additional specimens used in the present study are preserved in the collections of the
Nomenclature adopted in this study for the characterization of forewing pattern in
A total of 16 specimens were DNA barcoded. DNA extraction, PCR amplification and sequencing of the 658 base pair Cytochrome Oxidase 1 (
Sampling information of
Species name | Collection site | Host plant species | Host plant family | Collection ID | BOLD ID | BOLD BIN | GenBank accession no. | Institution of DNA extraction and sequencing of |
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Hawaii |
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West Maui |
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CJ-144 |
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West Maui |
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CJ-064 |
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West Maui |
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CJ-072 |
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Hawaii |
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Hawaii |
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CLV6240 |
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Hawaii |
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IO-322 |
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KPU & Eurofins, Japan |
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Hawaii |
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Hawaii |
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Hawaii |
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Hawaii |
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Hawaii |
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CLV6239 |
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Kauai | Unknown | Unknown | AYK-HI10-002 |
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Kauai | Unknown | Unknown | AYK-HI10-001 |
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Hawaii |
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Hawaii |
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We conducted an ML analysis of the
Intra- and interspecific genetic divergences in DNA barcode sequences among studied
Species |
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[0.88] | |||||
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7.0 | [0.17] | ||||
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6.71 | 5.85 | [0.31] | |||
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8.91 | 7.38 | 8.43 | [0.30] | ||
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11.12 | 11.08 | 10.59 | 13.28 | [1.70] | |
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13.46 | 12.10 | 12.19 | 13.83 | 4.41 | [1.07] |
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13.46 | 15.07 | 14.78 | 15.90 | 13.84 | 14.93 |
Kimura 2-parameter (K2P) distances (%) for barcode DNA sequences of the seven analyzed species in the genus
While single-marker
Institutional voucher numbers are given here for primary type material and museum collections. In the cases of
Abbreviations for collections:
1 | Forewing leaden grey, externally with fuscous brown (Fig. |
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– | Forewing shiny, metallic bronze with bright to dark orange patches |
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2 | A bright orange transverse fascia at 3/4 in middle interrupted with blue patch; an orange medial transverse fascia, narrowing towards dorsum, (Figs |
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– | An orange transverse patch beyond middle to costal 3/4, narrowing towards dorsum, extending to dorsal 2/3, with white costal spot |
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3 | A black patch along costal fold (Figs |
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– | An orange patch along costal fold, fringed with blackish scales (Figs |
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1 | Saccus slender, curved toward dorsum (Fig. |
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– | Saccus broad and straight (Fig. |
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2 | Valva slightly narrowing in middle with terminally rounded dorsal process (Fig. |
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– | Valva with short, pointed dorsal process (Fig. |
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*Male of |
1 | Signa with minute spines (Fig. |
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– | Signa with a pair of larger spines |
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2 | Spines long and slender (Fig. |
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– | Spines on the signa small and rounded (Fig. |
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– | Spines on the signa blunt (Fig. |
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1 | Start of mine spiral-shape (Fig. |
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– | Start of mine linear or serpentine-shape |
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2 | Reddish brown long linear mine following leaf vein (Fig. |
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– | Brown serpentine mines, mature larvae in situ leaves |
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3 | Larvae utilize leaves on larger plants. Mines on |
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– | Larvae utilize leaves on seedlings (Figs |
|
Olinda, Haleakala (Maui).
1 ♂ 7 unsexed, same data and locality as lectotype: 26696|NHMUK010862803; 26661|PARATYPE1/17|BPBM34325; 26667|PARATYPE2/17|
This species was described from 19 specimens: ‘type ♀ (26695); ♂ (28505)’ and 17 ‘paratypes’ from Kauai and Haleakala, Maui. This seems to indicate that Lord Walsingham considered them as holotype, allotype, and paratypes, as indicated on their specimen labels. But as a holotype was not specified in the description, the so-labelled types and paratypes are all to be considered syntypes under the present Code, Article 73.2 (
Adults of
32 (11♂ 15♀ 6 unsexed).
Adults: Oahu Is.: 2♀, Mt. Kaala, 18.ii.1923, Swezey coll., host: “
Molokai Is.: 1♀, Kainalu [Kainalu Forest, South East Molokai Forest Reserve], 27?vii.1927,
Lanai Is.: 2♂, 2750 ft, Munro Trail, 2.x.1976, K. & E. Sattler BM1976-605, BMNH(E)1621676 and BMNH(E)1621677,
Maui Is., in
Hawaii Is., host:
Larvae: 2 unsexed, Kokee, Kauai Is., 16&26.vi.2015 (stored), C.A. Johns leg., host:
This species is very similar to
Diagnostic features of four
Species name |
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|
|
|
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Forewing | Shiny, metallic bronze with bright orange-ochreous | Similar to |
Shiny, metallic bronze with dark brownish orange | Leaden grey, externally with brownish fuscous |
Basal patch | Black, triangular-shape | Absent, orange transverse fascia from costal fold to dorsal 1/4 | Brownish orange with black ground color, sometimes black | Brownish fuscous |
Apical orange transverse fascia | Absent | Present | Absent | Absent |
Apical portion | Fuscous, sometimes orangish encroaches on the apex | Fuscous | Fusocus with dark orange scales | Leaden gray |
Genitalia | ||||
Valva | Broad | Unknown | Rather long and narrowing in the middle | having rather shorter and pointed dorsal process |
Vinculum | Small, inflexed on the ventral side | Unknown | Large, inflexed on the ventral side | Small, inflexed on the ventral side |
Saccus | Slender and long, curved toward dorsal side | Unknown | Broad and straight | Broad and straight |
Spine on signum | Long and slender | Rather smaller and rounded | Rather blunt | Minute |
Distributiona,b |
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Host plant speciesa,b |
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Larval habit type | Leaf dropper | Unknown (probably non leaf dropper) | Non leaf dropper | Non leaf dropper |
Mining form | Long, linear, along leaf vein | At first spiral, later blotch | Serpentine | Serpentine |
Mine color | Red | Brown | Brown | Brown |
a As indicated by published data (
Forewing shiny, metallic bronze with bright orange-ochreous patches: a black triangular basal patch along the costal fold (Figs
Adults of
Distribution of
Forewing coloration and pattern of
Male genitalia of
Genitalia of
Kauai, Oahu and Lanai: new record, Maui (Walsingham 1907), Molokai and Hawaii (Big Island) (
(Figs
Resting posture of adult
Biology of
BIN
We identified two adult moths (Coll ID CJ-144 / GenBank accession no. ID
Kauaula (Maui).
Holotype ♀, Kauaula, Maui, 18.viii.2014 (stored in 99% ethanol), C.A. Johns leg., host:
2 unsexed (CJ-064, CJ-072), entirely sacrificed for molecular analysis and belonging to BIN
The forewing pattern of this species is similar to that of
Biology of
Maui.
(Figs
BIN
The specific epithet is derived from the type locality, Kaua`ula (pronounced ‘cow-wa-u-la’) Valley, an important site for Hawaiian endemic plants and culturally and spiritually for Native Hawaiians.
mountains, 2000 ft near Honolulu (Oahu).
Holotype ♂, Mts. 2000 ft near Honolulu, Oahu, 25.x.1892, Perkins. 25857|BM slide no. 472|Walsingham Collection. 1910–427.|NHMUK010305330| in
22 (8♂ 11♀ 3 unsexed)
Kauai Is: 1♂, Mt. Kahili, 18.vi.2013 (stored), N. Tangalin leg., Nat Collection, host:
Oahu Is: 1♂, Kahana, 1.i.1928, O.H. Swezey Collector, “
Molokai Is, in
Hawaii Is., Hawai’i Volcanoes National Park, host:
This species is very similar to
Biology of
Kauai: new record, Oahu (Walsingham 1907), Molokai (
(Figs
BIN
We collected
Hawai’i Volcanoes National Park (Big Island).
Holotype ♂, Hawai’i Volcanoes National Park, Hawaii (Big Island), 25.iv.2016, A. Kawakita leg., host:
Paratypes, in
Among
Hawaii (Big Island).
(Figs
Biology of
Biology of
Mine forms and characters of
Maximum likelihood tree of
BIN BOLD:ADF137. The five specimens sequenced for
The specific epithet,
We obtained DNA barcode data for 16 individual specimens (
Hawaiian
Swezey collected
On Maui,
We collected larvae of
In addition to providing morphological and molecular evidence to delimit species limits among the Hawaiian
It is likely that detailed molecular work among islands will reveal further cryptic species but native hostplants and habitats are under great threat.
We thank Cynthia King (Hawaii Department of Land and Natural Resources, Division of Forestry and Wildlife) for collecting and forest access permits, Rhonda Loa (Hawaii Volcanoes National Park) and Pat Bily (The Nature Conservancy) for land access, permitting assistance, logistical and field support, Daniel Rubinoff (University of Hawai‘i at Mänoa) for conception of the project, and the participants and organizers of the leaf miner workshop conducted on Puna, HI in 2016. Klaus Sattler is thanked for extensive discussion about material in the