Maunakea Invertebrate Threats, Identification, Collection, and Processing Guide

Standard Operating Procedure C Maunakea Invertebrate Threats, Identification, Collection, and Processing Guide Version 1.0, 2/10/2015, Jessica Kirkpatrick & Fritz Klasner Table of Contents 1. Purpose and Scope... 1 Invertebrate Threats... 2 2. List of Common and Targeted Invertebrates... 2 3. Identification of Common and Targeted Invertebrates... 4 4. Arthropod Collection... 14 Collection Supplies... 14 Collecting Potential Threats... 14 Hand Search & Haphazard Collections... 14 Flying Arthropods... 14 Crawling Arthropods... 15 Collection from traps... 15 Wet Traps... 15 Dry Traps... 15 5. Specimen Processing & Identification... 16 Specimen Processing Supplies... 16 Sorting & Identification... 16 Tips for Identification... 16 Presorting... 16 Post sorting... 17 Processing... 17 Data Entry... 17 6. Vial Labels... 19 7. Sorting Datasheet... 20 8. References... 22 9. Revision History Log... 22 10. Recommended Citation... 22 1. Purpose and Scope This standard operating procedure (SOP) guides threat characterization, identification, collection, and processing of invertebrates (arthropods) on Maunakea. The identification guide is not intended to be comprehensive, rather it is representative of common species and will facilitate an entomologist with SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 1 of 22

identification. This SOP should be used under the guidance of a professional entomologist if complete confidence in specimen identification is required. Invertebrate Threats The Wēkiu Bug Working Group, an informal collaboration of University, State, and Federal land managers and entomologists, periodically review potential arthropods threats for risk prioritization on Maunakea. This group identified invertebrates for risk prioritization as those with potential to alter the ecological functioning of the Management Area as well as impacting the integrity of cultural resources or human health. The greatest potential impacts are most likely to be caused by social insects (ants, wasps, bees and termites). These insects live cooperatively and have the greatest potential to establish and spread. For example, the Argentine ant (Linepithema humile) has successfully invaded Haleakalā National Park on Maui and spread to elevations above 9000 ft, reducing abundance of other native invertebrates including pollinators of the Haleakalā silversword (Argyroxiphium sandwicense subsp. macrocephalum). Once established, social insects are very difficult to control or eradicate, often requiring substantial human resources and materials. The following groups of arthropods are high priority target species for both early detection and ongoing monitoring: 1. Ants (Order: Hymenoptera, Suborder: Apocrita, Family: Formicidae) and other taxa that are morphologically similar, i.e. look like ants. 2. Wasps (Order: Hymenoptera, Suborder: Apocrita, Families: Vespidae, Pompilidae, & Mutilidae) and other taxa that are morphologically similar, i.e. look like large wasps. [Excluded are: Suborder Apocrita, Families: Bradynobeanidae, Falsiformicidae, Rhopalosomatidae, Sapygidae, Scoliidae, Sierolomorphidae, Tiphiidae]). 3. Spiders (Order: Araneae) 4. Beetles (Order: Coleoptera) [Excluded are Suborder: Polyphaga, Family: Coccinellidae i.e. ladybugs). 5. Horn & Stable Flies (Order: Diptera, Suborder: Brachycera, Family: Muscidae, Subfamily: Muscinae, Tribe: Stomoxyini). 6. Centipedes (Order: Scolopendromorpha, Family: Scolopendridae, Genus: Scolopendra ). 7. Mollusks (Phylum: Mollusca, Class: Gastropoda). 2. List of Common and Targeted Invertebrates Table 1: The table below is a list of common invertebrates (mostly arthropods) and invertebrate threats that will be targeted for prevention and early detection surveys on Maunakea, sorted by order, family, genus, and species, also specifying common name, and nativity when known. All taxa and species listed are established on UH Managed lands. Nativity of each taxa (whether order, family, genus, or species) is determined from the Hawai i Insect Database; all species within that taxa determines nativity. Taxa is either native, includes both endemic and indigenous species, non-native or introduced, or have both nativities (native and non-native) from identified taxa. This being said, non-native taxa are usually more commonly observed even when both native and non-native species are present within that taxonomic group. SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 2 of 22

Order Family Species Common Name Nativity Acari Multiple Unknown genera Mites Native & Non-native Araneae Lycosidae Lycosa hawaiiensis Lycosa spider Native Araneae Salticidae Unknown genera Jumping spiders Native & Non-native Coleoptera Carabidae Trechus obtusus Ground beetle Non-native Coleoptera Coccinellidae Coccinella septempunctata 7- Spot ladybug Non-native Coleoptera Coccinellidae Hippodamia convergens Converg. ladybug Non-native Collembola Entomobryidae Unknown genera Slender springtail Native & Non-native Dermaptera Forficulidae Forficula auricularia European earwig Non-native Diptera Agromyzidae Multiple genera & unknown Leaf-miner flies Native & Non-native genera Diptera Calliphoridae Multiple genera & unknown Blow flies Native & Non-native genera Diptera Chloropidae Unknown genera Frit flies Non-native Diptera Drosophilidae Multiple genera & unknown Vinegar flies Native & Non-Native genera Diptera Muscidae Multiple genera & unknown House flies Native & Non-native genera Diptera Phoridae Unknown genera Hump-backed fly Native & Non-native Diptera Sarcophagidae Multiple genera & unknown Flesh flies Native & Non-native genera Diptera Sciaridae Multiple genera & unknown Dark-winged Native & Non-native genera fungus gnats Diptera Syrphidae Multiple genera Hover flies Non-Native Hemiptera Aphididae Multiple genera & unknown Aphids Native & Non-native genera Hemiptera Geocoridae Geocoris spp. Big-eyed bug Non-native Hemiptera Lygaeidae Neacoryphus bicrucis Whitecrossed seed Non-Native bug Hemiptera Lygaeidae Nysius palor Seed bug Non-native Hemiptera Lygaeidae Nysius terrestris Seed bug Native Hemiptera Lygaeidae Nysius wekiuicola Wēkiu bug Native Hemiptera Miridae Orthotylus sophoricola Plant bug Native Hemiptera Nabidae Multiple genera Damsel bug Native & Non-native Hemiptera Pseudococcidae Unknown genera Mealybugs Native & Non-native Hemiptera Psyllidae Multiple genera & unknown Jumping plant Native, Non-Native genera louse Hymenoptera Apidae Apis mellifera European honey Non-native bee Hymenoptera Braconidae Multiple genera & unknown Braconid wasp Native, Non-native genera Hymenoptera Colletidae Hylaeus spp. Yellow-masked bee Native & Non-native Hymenoptera Formicidae Cardiocondyla kagustsuchi Common ant Non-native Hymenoptera Ichneumonidae Multiple genera & unknown Ichneumon wasp Native & Non-native genera Lepidoptera Noctuidae Multiple genera & unknown Owlet moths Native & Non-native genera Lepidoptera Pieridae Pieris rapae Cabbage butterfly Non-Native Lithobiomorpha Lithobiidae Lithobuis spp. Stone centipedes Native & Non-native Psocoptera Psocidae Unknown genera Bark lice Native & Non-native Stylommatophora Succineidae Succinea konaensis Amber Snail Native SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 3 of 22

3. Identification of Common and Targeted Invertebrates Table 2. The identification guide of the species listed above are provided below, including a selection of identifying characteristics derived from literature and other text and photographs of common invertebrates and invertebrate threats on Maunakea. Sorted by physical morphological similarities (including body shape, number of legs, wing placement, wing venation, color, etc.), order, family, genus, and species. Order Family Scientific/ Common Name Araneae Lycosidae Lycosa hawaiiensis Identification Characteristics (Source) 3 claws anterior eyes small in a more or less straight row, posterior median eyes very large, posterior lateral eyes smaller positioned well behind the posterior medians Species description uncertain Wolf spider Araneae Salticidae Unknown genera ( C.Triplehorn) Stout bodied with short legs Distinctive eye patterns with anterior median eyes by far the largest Body is often hairy Acari Anystidae Bdellidae Eupodidae Laelaptidae Phytoseiidae Unknown family Jumping spider Multiple genera (C.Triplehorn) Small, less than ½ mm long Abdomen un-segmented Abdomen broadly joined or fused to the anterior region of the body Body usually rounded or oval More than 6 legs Mites Hemiptera Aphididae Multiple genera (J.Castner) Wings present or absent Pair of dorsal tubes (cornicles) project from posterior part of the body Body is pear-shaped and soft Aphids (J.Castner, C.Triplehorn) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 4 of 22

Order Family Scientific/ Common Name Hemiptera Psyllidae Multiple genera Identification Characteristics (Source) Small 1.5-5mm long Long antennae with 10 segments Wings held in a tent-like fashion over the body Front wings may be clear and are tough with obvious veins Jumping plant louse Psocoptera Psocidae Unknown genera (C.Triplehorn, G.McGavin) Small (3mm or less) Soft-bodied Wings may be present or absent At rest, the wings are held roof-like over the abdomen Antennae as long as body and threadlike Hump-backed appearance Barklice Collembola Entomobryidae Multiple genera (J.Eiben [pers. comm.], J.Castner, C.Triplehorn) Minute, 3mm or less 4th abdominal segment is large Wings absent Large forked structure (furcula) at the tip of the abdomen Slender springtails Thysanoptera Thripidae Unknown genera Thrips (J.Eiben [pers. comm.], J.Castner, C.Triplehorn) Minute (1-2mm) Body torpedo shaped Wings present or absent Four long slender membranous wings with an extremely long fringe of hairs around the edges Antennae short (J.Castner, J.Eiben [pers. comm.]) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 5 of 22

Order Family Scientific/ Common Name Hemiptera Pseudococcidae Unknown genera Identification Characteristics Elongate, oval body with segments Well developed legs Body soft with white waxy coating Only the males have wings (rarely observed) and look completely different (Source) Mealybugs Lithobiomorpha Lithobiidae Lithobuis spp. (C.Triplehorn, J.Eiben [pers. comm.]) Short legged with 15 pairs of legs Usually brown in color 10-45 mm in length Some legs with strong spines Eyes usually consist of many facets Species are undescribed Stone centipedes Dermaptera Forficulidae Forficula auricularia (C.Triplehorn) Tip of abdomen with large pinchers Body elongate and somewhat flattened Antennae threadlike Brown in color European earwig Coleoptera Carabidae Trechus obtusus (J.Castner) Irridescent elytra Eyes large and bulbous Width of head at eyes narrower than the pronotum Antennae originate between the base of the mandibles and eyes Ground beetle Coleoptera Coccinellidae Hippodamia convergens Convergent ladybug (J.Castner, Liebherr & Takumi) 10 mm or less Body broadly oval to round Usually bright colored Head partially to entirely concealed by pronotum when viewed dorsally Full complement of 13 spots or only a few. White lines converge behind the head (J.Castner, BugGuide.net) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 6 of 22

Order Family Scientific/ Common Name Coleoptera Coccinellidae Coccinella septempunctata 7- Spot ladybug Hemiptera Lygaeidae Neacoryphus bicrucis Identification Characteristics (Source) 10 mm or less Body broadly oval to round Usually bright colored Head partially to entirely concealed by pronotum when viewed dorsally Typically has 7 black spots on elytra One spot next to scutellum that bridges the 2 elytra White patches just above the black bridging scutellar spot (J.Castner, Animal Diversity Web) Fairly large Conspicuous colors (red and black) Whitecrossed seedbug Hemiptera Lygaeidae Nysius wekiuicola (C.Solbreck) Flightless (micropterous) adults 4.5-5.5 mm in length Long legs Dark brown- red in color Females are rounded and much larger Males have a pointed ball at their end Adults are elongate with stripes on their abdomen Nymphs are orange and plump Wēkiu bug Hemiptera Lygaeidae Nysius palor (J.Eiben [pers. comm.]) Dark head Pale shiny wings 2 black spots on wings Seed bug (J.Eiben [pers. comm.]) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 7 of 22

Order Family Scientific/ Common Name Hemiptera Lygaeidae Nysius terrestris Identification Characteristics Brown head Costal margins more strongly expanded than N.palor Patterns on wings (Source) Seed bug Hemiptera Geocoridae Geocoris spp. (J.Eiben [pers. comm.]) Oval bodied with broad head Large bulging eyes Big-eyed bug Hemiptera Miridae Orthotylus sophoricola Plant bug (Cornell University) About 4mm in length General coloration is light bluish to yellowish green Cuneus (triangular shaped region) present on base of forewing Hemiptera Nabidae Multiple genera (D.Polhemus, J.Castner ) Relatively slender and elongate in shape Front femora slightly enlarged Discal cells along fringe of forewings Four-segmented rostrum Damsel bug Diptera Agromyzidae Multiple genera (J.Castner, C.Triplehorn, G.McGavin) Small (1-6mm) Usually black and yellow/ white Head and thorax moderately hairy Leaf-miner flies (C.Triplehorn, G.McGavin) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 8 of 22

Order Family Scientific/ Common Name Diptera Drosophilidae Multiple genera Identification Characteristics (Source) Usually small, yellow, brown or black Shiny clear wings that can have dark markings Light or bright red eyes Anterior edge of wing is thickened and broken at 2 points near where veins join in. Arista of antennae plumose Vinegar flies Diptera Chloropidae Unknown genera (J.Castner, G.McGavin) Small (1-6mm) Can be grey, green, or black with bright yellow markings Body with few obvious hairs and bristles Clear, dark triangular plate on top of the head between the eyes Abdomen is usually broad and tapers towards the hind end Frit flies (G.McGavin) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 9 of 22

Order Family Scientific/ Common Name Diptera Phoridae Unknown genera Identification Characteristics (Source) Small or minute flies easily recognized by the humpback appearance Laterally flattened hind femora Veins short and heavy near the basal half of the anterior edge of the wing Hump-backed flies Diptera Sciaridae Multiple genera (J.Castner C.Triplehorn) Blackish in color The r-m crossvein is in line with and appears as a basal extension of Rs Eyes meet at the base of the antennae Dark-winged fungus gnats Diptera Sarcophagidae Multiple genera (C.Triplehorn) Thorax with a pattern of grey and black longitudinal stripes (never metallic) Abdomen with a grey and black checkered pattern Flesh flies (J.Castner, C.Triplehorn) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 10 of 22

Order Family Scientific/ Common Name Diptera Muscidae Multiple genera Identification Characteristics (Source) Arista (antenna hair)plumose throughout its length Thorax often striped while the sides of the abdomen are pale The anal vein (A2 + CuA2) is short and does not reach the wing margin House flies Diptera Calliphoridae Multiple genera (C.Triplehorn) Body often metallic (sometimes only showing a dark metallic sheen) Large cell nearest wingtip narrows distally Tips of antennae are distinctly feathered Blow flies Diptera Syrphidae Multiple genera (J.Castner, G.McGavin) Many species resemble bees and wasps, frequently yellow and black Spurious (pretend) vein present near middle of wing. Hover flies Hymenoptera Apidae Apis mellifera (J.Castner) Red/brown with black bands and orange yellow rings on abdomen Hair on thorax with less hair on the abdomen Legs are mostly dark brown/black European honey bee (Animal Diversity Web) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 11 of 22

Order Family Scientific/ Common Name Hymenoptera Colletidae Hylaeus spp. Identification Characteristics (Source) Small Black body with yellow markings on the face (not consistent in females) Sparsely hairy Wings with only 2 submarginal cells Wasp-like in appearance Yellow-masked bee Hymenoptera Braconidae Multiple genera (C.Triplehorn, J.Eiben[pers. comm.]) Often dark in color Small-medium (5-15mm) Antennae threadlike and long (greater than or equal to half the body length) Lack of a costal cell in forewings Forewings with two small closed cells below the stigma Braconid wasps Hymenoptera Ichneumonidae Multiple genera (J.Castner, C.Triplehorn) Slender wasps, body laterally compressed Antennae threadlike 16 or more antennal segments Forewings with a large, somewhat trapezoidal- shaped (horse head)closed cell directly below stigma Color and size variable (5-40mm) Ichneumon wasps (J.Castner, C.Triplehorn) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 12 of 22

Order Family Scientific/ Common Name Hymenoptera Formicidae Cardiocondyla kagustsuchi Identification Characteristics Small (2mm) Dark brown to black Wings absent or present Elbowed antennae (Source) Common ant Lepidoptera Noctuidae Multiple genera (J.Castner) Size variable (15-170mm) Antennae threadlike Forewings usually dark with patterns of lines or dots Heavy bodied (for moths) Labial palps are usually long Hair-like antennae Hindwings usually pale with no pattern Owlet moths Lepidoptera Pieridae Pieris rapae Stylommatophora Phylum: Mollusca Succineidae Cabbage butterfly Succinea konaensis (J.Eiben [pers. comm.], J.Castner, C.Triplehorn) White with black at the tips of the forewings Front wings 2 black dots in the central area of each forewing in females, one dot for males. The body is covered with dense hair, colored white in females, and darker in males (University of Florida, Entomology dept.) Thin walled translucent shell Often amber colored Spire is usually very small compared to the huge apertural lip Amber Snail (The living world of Molluscs- website) SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 13 of 22

4. Arthropod Collection Mollusks will be collected using the same methods described in the Crawling Arthropod section below. Arthropods that are not confidently identified in the field must be collected. Collection types vary amongst hand search and haphazard findings, trap types, and specimen size. Multiple specimens collected in one vial could lead to specimen consumption, and it best to put separate specimens in their own container. Other collection methods may be used. Plastic vials (9 dram) are typically used for field collection, glass vials (1 dram) are typically used for long-term storage and curation. Immediately label all specimens collected and freeze all specimens within 24 hours. Collection Supplies Aspirator Plastic Vials with lids (clean) Forceps Bug / sweep net Glass jar with lid (optional- may come in handy for larger specimens) Gallon Ziploc bags Pre-cut labels Field datasheets with associated survey (usually not needed for hand search and haphazard collections) Pencil Gloves (for putrid specimens) Collecting Potential Threats Arthropods threats, outlined in Section 1, can be collected using the same methods as below, but ensure to take extra precautions to not spread the potential threat. This can be done by simply placing the collected specimen in an extra Ziploc bag. You may also want to take a GPS point at the location in which the specimen was detected, if there is not already a GPS point for the site. Potential new threat species should be reported to within 24 hours. Hand Search & Haphazard Collections This is probably the most common type of collection that will occur. Hand search and haphazard collections includes all specimens that you are searching for, and specimens you just happen to spot and decide to collect. Collection differs between flying and crawling arthropods. Flying Arthropods To capture flying arthropods use a bug net, plastic vial, or glass jar (for larger specimens), pencil, and labels. You do not need to use a net to collect flying arthropods, some can be directly collected in a vial. Collecting arthropods in flight can be very difficult (especially for orders Lepidoptera and Odonata), so it is best to wait for the flying specimen to land, where it can be swept up with a bug net or placed in a vial. Fill out all fields on the label, and put the label in with the specimen. When you make it back to the office put the vial(s), or glass jar, in the freezer. The freezer will kill the specimen, and preserve it until it can be processed. SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 14 of 22

Crawling Arthropods Collection of crawling arthropods typically requires an aspirator, plastic vials, pencil, and labels. Not all crawling arthropods can be aspirated, rather place the vial on the surface near the specimen, and scoop it up. For specimens that can be aspirated, connect an empty vial to the aspirator, point at the specimen you want to aspire, and inhale with the mouth piece. Sometimes specimens get stuck in the aspirator nozzle, if this is the case, use a piece of grass to push the specimen through. Once all specimens have been collected complete a label and put the cap on the vial. When you make it back to the office, put the vial(s), or glass jar in the freezer. The freezer will kill the specimen, and preserve it until it can be processed. Collection from traps Collections from traps will most likely only occur during an survey. Collection methods differ between both wet and dry traps. Wet Traps Wet traps are traps that have liquid when placed to capture and preserve the insects (the liquid is usually propylene glycol or soapy water). You will need plastic vials, forceps, labels, datasheet, empty bottle (for remaining liquid), plastic bag, and a pencil. You may also want paper towels and hand sanitizer. Use your forceps to pick out the arthropods from the liquid. Since the arthropods are dead placing them all in one vial is fine. Be sure to write a label (with appropriate site ID, date, trap type, and collector) and place it in the vial. Pour the remaining liquid in the empty bottle and put the trap in a Ziploc bag. Be sure to hike all supplies and materials out of the field. When you make it back to the office, put the vial(s) in the freezer. The freezer will kill the specimen, and preserve it until it can be processed. Dry Traps Dry traps are placed without liquid. You will need an aspirator, forceps, plastic vials, labels, Ziploc bag, field datasheet, and a pencil. It is very difficult to extract the entire specimen from sticky traps. If you find a specimen of threat or of interest on a sticky trap, record it on the datasheet, and take photos of the specimen. If you are able to, keep the trap in the freezer until an expert can confirm its identification. PBJS stick and vial specimens can be collected using the same methods as Crawling Arthropods under the Hand Search & Haphazard Collections section. A forceps may also be useful for collecting specimens from the PBJS vial. Baited pitfall traps will most likely contain live specimens. If there are any wēkiu bugs in the trap, use your aspirator to suck up the bugs into a vial, and record the sex or life stage, and live or dead as each individual is collected. Release all live Wēkiu bugs and Lycosa hawaiiensis spiders before departing the site. Dead wēkiu bugs, and all other collections can be placed in a vial using the forceps or aspirator. Write a label for the vial and record whether specimens were collected on the field datasheet. The traps may be placed in a Ziploc bag. Be sure to hike all supplies and materials out of the field. When you make it back to the office, put the vial(s) in the freezer. The freezer will kill the specimen, and preserve it until it can be processed. SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 15 of 22

5. Specimen Processing & Identification Specimens must be stored in a freezer within 24 hours of collection until they can be sorted and identified in a laboratory. All data is recorded on sorting datasheets. Processing is often done with a partner to facilitate data recording. Specimen Processing Supplies Ethyl alcohol / Ethanol Isopropyl alcohol 91% Forceps Microscope with light source and camera setup Droppers Glass vials with lids and trays (1 dram) Petri dishes Sorting datasheets (Section7) Pre-cut labels (Section 6) Arthropod text books (Study of Insects 7 th edition by C.Triplehorn & N.Johnson, Photographic Atlas of Entomology & Guide to Insect Identification by J.Castner, and Insects by G.McGavin list of identified arthropods (Excel spreadsheet) Print copy of SOP C Sorting & Identification Sorting specimens can be time consuming and tedious. Sorting is a very important process that can be done simultaneously with identification. Sort one plastic (field collection) vial at a time to avoid confusion. Each vial has its own sorting datasheet. Tips for Identification Most collected specimens are common arthropods that can be identified by specific characteristics; many of these can be found in this SOP, Section 3. Commonly Observed Arthropod Identification. Some of these characteristics can be observed and identified easily, while others require a dissecting scope. The person identifying arthropods should be familiar with a microscope, how to set up and use the scope camera, trained in at least basic entomology, and have gone through the identification process with an entomologist. Presorting Specimens should be identified to the lowest taxa possible to determine whether the species is a threat. One of the main things to look for in the presorting process are specimens and taxa that are potential threats. Threats are identified in Section 1. Arthropod Threats. Potential or suspected threats must be reported to OKMM within 24 hours. If the species is still considered a threat after confirmed identification, see the emergency response section of the Maunakea Invasive Species Management Plan. 1) Set up your microscope and workspace to include all supplies, and adequate elbow room to avoid accidents, and stay efficient. 2) Take only specimens for immediate processing out of the freezer. SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 16 of 22

3) Take one plastic vial and put all the specimens in a clean petri dish. 4) Complete the sorting datasheet (1 datasheet per vial) using information from the vial label. 5) Using the dropper, drop some isopropyl alcohol on the specimens, and begin to sort. 6) You will need to use a microscope for this step. Use the forceps and group together specimens that look alike, and to the lowest taxa possible. Use textbooks and the guide in this SOP to help with grouping. Different species from the same family may be grouped together as long as you note the number of morphospecies in the sorting datasheet. When you are able to identify to a lower taxa; the genus or species, group them separately. 7) Record the identification groups (i.e. Braconidae, Muscidae) and individual counts on the datasheet. Place the specimens in glass vials 1/3 full of ethanol (as opposed to isopropyl alcohol). Be sure to make a note on sorting datasheet if any other preservation type was used (i.e. ran out of ethanol, used isopropyl). 8) Write a label in pencil that matches the original plastic vial label and sorting datasheet and write the lowest taxonomic classification known on the back of the label. Place the label in the appropriate glass vial. This will help with further identification and pencil will not bleed or dissolve in alcohol. 9) Specimens and or groups that could not be confidently identified should be placed in separate vials. Each group or specimen goes in its own vial, and placed in a vial tray labeled as unidentified. Don t forget step 8. 10) Repeat step 7 until all specimens in the original plastic vial are sorted with appropriate labels for all glass vials, and data recorded on the sorting datasheet. 11) Repeat steps 2-10 until all collection vials have been sorted. 12) Ensure all glass vials are tightly capped. Place pre-sorted vials in a dry safe place in the office. Place unsorted plastic vials back in the freezer to work on at another lab day. Post sorting Specimens should be identified to the lowest taxa possible. If it is a specimen that you have seen before, go back to those collections and look at what it was identified as before. If it is a brand new specimen, try keying it out using textbooks, online keys, and the HI Insect Database excel sheet (to verify the taxa is in HI). If you still can t figure it out, and you don t have assistance available, take several high quality, high resolution photographs of the specimen and send it to an entomologist. If the entomologist can t confidently identify the specimen from the photo, prepare the specimen to be sent to the experts at the Bishop Museum. Processing Ensure that all vials are appropriately labeled with pencil and are in ethanol-filled glass vials. Vials should be organized in vial trays labeled with the survey type (i.e. Annual Alien Arthropod Survey), and year, with a note of identified or unidentified specimens. Store specimen vials in a dry, safe, pest free location in offices, Bishop Museum, UHH Museum, or elsewhere. Data Entry There are multiple different surveys conducted with associated SOP s. Data (excel sheets) can be found in the associated SOP folders. When entering data, cross-reference sorting datasheet entries with field datasheets (i.e. field datasheet- specimens collected from yellow pan, sorting datasheet- Species A, B, C SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 17 of 22

collected from yellow pan). Double check the data entered from both datasheets and search for errors or inconsistencies. This section will be updated once a database is finalized. SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 18 of 22

Office of Mauna Kea Management 6. Vial Labels SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 19 of 22

7. Sorting Datasheet SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 20 of 22

Sample ID number (GPS code) - GPS - Coordinates - Sample Description Sampling Method Worker Name Date Sorted Collection info HI: Mauna Kea, Date Collected - Site Description Elevation - Collector - Specimens Condition Notes - Order Family Genus Species Morphospecies Adult count Nymphs Notes Order Family Genus Species Morphospecies Adult count Nymphs Notes Order Family Genus Species Morphospecies Adult count Nymphs Notes Order Family Genus Species Morphospecies Adult count Nymphs Notes Order Family Genus Species Morphospecies Adult count Nymphs Notes Order Family Genus Species Morphospecies Adult count Nymphs Notes Order Family Genus Species Morphospecies Adult count Nymphs Notes SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 21 of 22

8. References "Apis Mellifera (honey Bee)." Animal Diversity Web. Web. 10 Oct. 2014. <http://animaldiversity.org/accounts/apis_mellifera/>. Castner, James L. Photographic Atlas of Entomology and Guide to Insect Identification. Gainesville, FL, USA: Feline, 2000. Print. "Geocoris Spp." Geocoris Spp. Web. 10 Oct. 2014. <http://www.biocontrol.entomology.cornell.edu/predators/geocoris.php>. Liebherr, James K., and Raina L. Takumi. "Introduction and Distributional Expansion of Trechus Obtusus (Coleoptera: Carabidae) in Maui, Hawai`i." Pacific Science 56.4 (2002): 365-75. Print. McGavin, George, and Richard Lewington. Insects. New York, N.Y.: Smithmark, 1992. Print. Polhemus, Dan A. "Further Studies On The Genus Orthotylus (Heteroptera: Miridae) In The Hawaiian Islands, With Descriptions Of Thirty-Four New Species." Journal of the New York Entomological Society: 227-333. Print. Solbreck, Christer. "Induction of Diapause in a Migratory Seed Bug, Neacoryphus Bicrucis (Say) (Heteroptera: Lygaeidae)." Oecologia: 41-49. Print. "Species Hippodamia Convergens - Convergent Lady Beetle." Species Hippodamia Convergens. Web. 10 Oct. 2014. <http://bugguide.net/node/view/8374>. Triplehorn, Charles A., and Norman F. Johnson. Borror and DeLong's Introduction to the Study of Insects. 7th ed. Belmont, CA: Thompson Brooks/Cole, 2005. Print. 9. Revision History Log Previous Version # Previous Version Date Previous Version Author Changes Made Reason for Change New Version # Add rows as needed for each change or set of changes associated with each version. 10. Recommended Citation Kirkpatrick, J. and F.Klasner. 2015. Standard Operating Procedure C, Maunakea Invertebrate Threats, Identification, Collection, and Processing Guide. v1.0. 22 pp. In: Vanderwoude, C., F. Klasner, J. Kirkpatrick and S. Kaye. 2015. Maunakea Invasive Species Management Plan. Technical Report No. 191. Pacific Cooperative Studies Unit, University of Hawai i, Honolulu, Hawai i. SOP C, v1.0 2/10/15 Invertebrate Threats, ID Guide, Collection, Page 22 of 22