Carter T. Atkinson, U.S. Geological Survey, Pacific Island Ecosystems Research Center 20250930 tabular digital data https://doi.org/10.5066/P13OSHIG This data publication contains information collected as part of a field and laboratory effort to determine prevalence of avian malaria (Plasmodium relictum) and pox-like lesions (Avipoxvirus spp.) in native and non-native forest birds on Molokaʻi in the Hawaiian Islands. Forest bird banding data, observations of pox-like lesions and blood samples for diagnosis of Plasmodium relictum, the agent responsible for avian malaria in the Hawaiian Islands, were collected in 2003 from wet and mesic habitats on eastern Molokaʻi, including Kalaupapa National Historical Park, Kamakou Preserve, Pelekunu Preserve and Puʻu Aliʻi Natural Area Reserve. These areas were the last locations where Olomaʻo (Myadestes lanaiensis rutha) and Iʻiwi (Drepanis coccinea) were observed in 1980 and 2010, respectively (Smith et al. 2025). Only two species of honeycreepers (Hawaiʻi ʻAmakihi, Chlorodrepanis virens and ʻApapane, Himatione sanguinea) still persist. These data report morphometric data, age, sex, molt status, pox virus-like (Avipoxvirus spp.) lesion occurrence, and diagnostic results for avian malaria from native and non-native forest birds that were captured with mist nets. This XML describes two tabular CSV files: Molokai Banding Table with Diagnostics.CSV and Standard Curve.CSV. The former file contains the analyzed data for Moloka’i, while the latter contains only calibration data Smith, L. K., S. W. Judge, A. S. Genz, L. Berry, L. K. Berthold, H. L. Mounce and R. J. Camp. 2025. Population trends of native and non-native forest birds on the island of Molokai'i, Hawai'i. The Wilson Journal of Ornithology, 137:51-64. These data were collected as part of a larger study funded by the U.S. Geological Survey Ecosystems Mission Area (formerly the National Biological Survey) to investigate the effect of introduced vector-borne avian disease on geographic and altitudinal declines of native Hawaiian forest birds. 20030204 20031106 observed Complete None planned Boundary area of study sites on Moloka"i and Hawaiʻi -157.0383 -154.8665 21.1880 19.4399 ISO 19115 Topic Category biota environment health Keywords avian malaria Hawaiian honeycreepers vector-borne disease Plasmodium relictum parasite Avipoxvirus USGS Metadata Identifier USGS:69aa38a1b66b01b7b32d80a9 USGS Thesarus wildlife disease USGS-EMA-LOW Fish and Wildlife Disease native species Place Keywords Molokaʻi Kamakou Preserve Puʻu Aliʻi Natural Area Reserve Kaulaupapa National Historical Park Pelekunu Preserve Kaunakakai None Plasmodium relictum Avipoxvirus Plasmodiidae Poxviridae Kingdom Protozoa Phylum Apicomplexa TSN: 553099 None. Please see 'Distribution Info' for details. None. These data were collected using funding from the U.S. Government and can be used without additional permissions or fees. Users are advised to read the dataset's metadata thoroughly to understand appropriate use and data limitations. Carter T. Atkinson U.S. Geological Survey, NW-PACIFIC ISLAND REG Volunteer Emeritus physical

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Hawaii National Park HI 96718 USA 808-210-6141 catkinson@usgs.gov This study was supported by the U.S. Geological Survey Ecosystems Mission Area with funding from the Invasive Species and Wildlife and Terrestrial Resources program and the U.S. Geological Survey Natural Resource Protection Program. Samuel Aruch Carter T. Atkinson Amy F. Savage Dennis A. LaPointe 20071001 publication Journal of Wildlife Diseases vol. 43, issue 4 n/a Wildlife Disease Association ppg. 567-575 https://doi.org/10.7589/0090-3558-43.4.567 Laboratory serological analysis of blood samples included positive and negative controls to ensure accuracy of the results. Blood smears were examined by an experienced microscopist with knowledge of Plasmodium morphology, staining characteristics and host cell alterations. Real-time PCR diagnostic assays included no template (water) negative controls and a standardized positive control sample of template DNA from a chronically infected Hawaiʻi ʻAmakihi. The same positive control was included on each assay plate to allow standardization of results. GPS units were not available at the time of the study so GPS coordinates for banding sites are approximated based on map values derived from pocket barometric altimeters with an accuracy of +/- 10 m along established trails, campsites and fence lines. The data are logically consistent. The consistency was verified as part of the quality assurance that occurred during data analysis. For the banding data, measurements and age and sex classifications may be missing for recaptured birds, birds that escaped during the banding process and for errors in recording data. Diagnostic evaluations are missing if blood samples were not collected or if partial blood samples limited type or number of tests. Mist-netting was conducted between February and November 2003 in wet and mesic forest habitats on eastern Molokaʻi. Up to ten mist nets (12 × 2.6 m, 32-mm mesh) were operated 1 - 4 days at each field location. Nets were placed at a height of 1 to 5 m on galvanized metal poles in forest habitat dominated by ʻohiʻa lehua (Metrosideros polymorpha), ranging from 215 to 1096 m. A small number of opportunistic samples were also collected at elevations <50 m in Kalaupapa National Historical Park and the town of Kaunakakai. Nets were operated for ~6 h each day between 0630 and 1400 hours HST, depending on weather conditions. All captured birds were given USGS (Bird Banding Laboratory) aluminum bands and were aged and sexed using a combination of plumage, morphometrics, and breeding characters (Pyle 1997). Breeding condition of captured birds was determined by assessment of the swelling of the cloacal protuberance in males and the development of a brood patch in females (Pyle 1997). Up to 100 μl of whole blood was drawn by jugular venipuncture from each bird into a heparinized syringe with a 28-gauge needle for preparation of blood smears and collection of plasma for serological analysis. When jugular venipuncture was deemed impractical or too difficult, blood was collected into a heparinized capillary tube from the brachial wing vein by swabbing the area with 70% ethanol and then puncturing the vein with a sterile 28 gauge needle. Bleeding was stopped by application of Styptic (anticoagulent) powder and light pressure on the vein with a cotton swab. Blood samples from wild Hawaiʻi ʻAmakihi with chronic, low intensity Plasmodium infections as well as some experimentally infected Hawaiʻi ʻAmakihi with higher intensity infections were included in the study to develop a standard curve for estimating intensity of infection with Plasmodium. Hawaiʻi ʻAmakihi with chronic infections were captured at Nanawale Forest Reserve, Malami Kī Forest Reserve and Bryson's Cinder Cone (Puʻu Kaliʻu) on the Island of Hawaii between 2001 - 2003 as part of a larger project on the ecology and biocomplexity of avian malaria (Atkinson et al. 2025). Hawaiʻi ʻAmakihi used in experimental infections were captured at the Upper Waiākea Forest Reserve in 2001 along Powerline Road and exposed to infective mosquito bites as described by Atkinson et al. (2000, 2013) References Atkinson, C.T., R.J. Dusek, K.L. Woods, and W.M. Iko. 2000. Pathogenicity of avian malaria in experimentally-infected Hawaii Amakihi. Journal of Wildlife Diseases, 36: 197-204. Atkinson, C.T., K.S. Saili, R.B. Utzurrum, and S.I. Jarvi. 2013. Experimental evidence for evolved tolerance to avian malaria in a wild population of low elevation Hawai`i `Amakihi (Hemignathus virens). Ecohealth 10: 366-375 Atkinson, C.T., Woodworth, B., Hart, P., and Jarvi, S., 2025, Island of Hawaii forest bird malaria infection data 2001-2005: U.S. Geological Survey data release, https://doi.org/10.5066/P1475TVO Pyle, P. 1997. Identification Guide to North American Birds, Part I: Columbidae to Ploceidae. Slate Creek Press, Bolinas, California. 20031106 Carter T. Atkinson U.S. Geological Survey, NW-PACIFIC ISLAND REG Volunteer Emeritus physical

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Hawaii National Park HI 96718 USA 808-210-6141 catkinson@usgs.gov A thin blood smear was made immediately after blood was drawn from the jugular or brachial veins, air dried, and fixed with methanol. The remaining blood was spun with a portable field centrifuge to separate plasma from red blood cells. Blood smears were stained for one hour with 6% phosphate-buffered Giemsa, pH 7.2 within two weeks after slide preparation, rinsed with tap water and air dried. Slides were scanned by microscopy at a magnification of 500X to search for intraerythrocytic sexual and asexual stages of Plasmodium relictum. Questionable detections were confirmed by examination with an oil immersion objective at 1000X. To standardize search effort among observers, one hundred 500X fields were examined on each slide. This was equivalent to a search effort of about 30,000 erythrocytes per slide. Birds were scored as positive for Plasmodium relictum if at least one infected erythrocyte was detected during microscopic screening of slides. For estimation of intensity of infection, numbers of parasitized erythrocytes in one hundred, 400X fields of view were counted. A photograph was taken every 10 fields and numbers of erythrocytes were counted with a digital image analysis program as described by Gering and Atkinson (2004). These numbers were averaged and multiplied by 100 to estimate the total number of erythrocytes examined in 100 fields. Intensity of infection was determined by dividing total number of parasitized erythrocytes by the estimated total number of erythrocytes in one hundred 400X microscope fields and then adjusted as the total number of parasitized erythrocytes per 1000 erythrocytes. References Gering, E. and C.T. Atkinson. 2004. A rapid method for counting nucleated erythrocytes on stained blood smears by digital image analysis. Journal of Parasitology, 90:879-881. 20031106 Carter T. Atkinson U.S. Geological Survey, NW-PACIFIC ISLAND REG Volunteer Emeritus physical

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Hawaii National Park HI 96718 USA 808-210-6141 catkinson@usgs.gov After capillary tubes containing whole blood were spun with a portable field centrifuge, hematocrit was measured with a hematocrit reader card to determine packed cell volume. Tubes were then scored with a jewelers file just above the boundary between plasma and packed cells and snapped into two pieces. A micropipette with disposable tips was used to transfer plasma from the upper half of the tube to an empty 0.5 ml screw cap tube. Packed red blood cells were similarly removed from the lower half of the tube after the sealed end was removed and transferred to a 0.5 ml screw cap tube containing 100 ul of DNA lysis buffer (0.1 M Tris-HCL, pH 8.0, 0.1 M sodium ethylenediaminetetraacetic acid, 2% sodium dodecyl sulfate). Tubes were stored on wet ice in the field and then moved to a -70 freezer after return to the laboratory. Plasma from each bird was tested for antibodies to P. relictum with the immunoblotting technique described by Atkinson et al. (1995, 2001). Plasma tested by immunoblotting was considered positive for antibodies to P. relictum if one or more bands characteristic of the immune response to erythrocytic stages of P. relictum were detected on immunoblots of SDS-PAGE gels of crude P. relictum antigen (Atkinson et al. 2005). References Atkinson CT, Woods KL, Dusek RJ, Sileo LS, Iko WM (1995) Wildlife disease and conservation in Hawaii: pathogenicity of avian malaria (Plasmodium relictum) in experimentally infected Iiwi (Vestiaria coccinea). Parasitology 111:S59–S69 Atkinson CT, Dusek RJ, Lease JK (2001) Serological responses and immunity to superinfection with avian malaria in experimentally-infected Hawaii Amakihi. Journal of Wildlife Diseases 37:20–27 Atkinson, C. T., J. K. Lease, R. J. Dusek, AND M. D. Samuel. 2005. Prevalence of pox-like lesions and malaria in forest bird communities on leeward Mauna Loa Volcano, Hawaii. Condor 107: 537–546 20031106 Carter T. Atkinson U.S. Geological Survey, NW-PACIFIC ISLAND REG Volunteer Emeritus physical

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Hawaii National Park HI 96718 808-210-6141 catkinson@usgs.gov Frozen packed cells in lysis buffer were thawed and DNA was purified with Qiagen DNeasy DNA extraction kits (Qiagen, Gaithersburg MD). DNA was extracted with DNeasy blood and tissue kits. DNA concentration and purity was determined with a Nanodrop 1000 spectrophotometer (Thermofisher Scientific, Greenville NC). Malaria diagnostic quantitative PCR (qPCR) reactions were performed with primers 11F (5`-ATTAGCAGAACAAAGAAACTTAACA-3`) and 11R (5`-CATAGAATGAACATATAAACCAG-3`) (Zehtindjiev et al. 2008) with a FAM labeled hydrolysis probe with an internal ZEN quencher in addition to the standard 3' Black Hole quencher to increase sensitivity (5`- /FAM/TGCTTTTGG/ZEN/TGCAAGAGAGTATTCAGT/3IABkFQ/ -3`) (IDT, Corralville IA). Primers and probe were specific for a portion of the Plasmodium relictum lineage GRW4 cytochrome b gene. Each reaction contained 10 ul of iTaq Universal Probe Supermix (Bio-Rad Laboratories, Hercules CA), 100 ng of DNA template, 1.5 ul of each primer at a concentration of 10 uM, 0.5 ul of probe at a concentration of 10 uM and enough PCR grade water to bring the final volume to 20 ul. qPCR reactions were performed in a C1000 Touch Thermocycler with a CFX96 Real-time System (Bio-Rad Laboratories, Hercules CA) with the following thermal profile: 5 min at 95 °C followed by 50 cycles of 5 s at 95 °C, and 30 s at 60 °C. Samples were run in triplicate, along with a negative control containing water and a positive control containing template from a Hawaiʻi ʻAmakihi with a known malarial infection. The same positive control was tested in triplicate on each qPCR plate to allow standardization of plate to plate reactions. Samples were designated positive for P. relictum DNA when there was an exponential increase in fluorescence during the first 45 cycles of PCR amplification. References Zehtindjiev, P., M. Ilieva, H. Westerdahl, B. Hansson, G. Valkiunas and S. Bensch. 2008. Dynamics of parasitemia of malaria parasites in a naturally and experimentally infected migratory songbird, the great reed warbler Acrocephalus arundinaceus. Experimental Parasitology 119:99–110 20231106 Carter T Atkinson USGS - NW-PACIFIC ISLAND REG Volunteer Emeritus physical

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Hawaii National Park HI 96718 USA 808-985-6401 catkinson@usgs.gov The programs LinRegPCR version 2021.2 and Factor-qPCR (Ruitjer et al. 2006, Ruitjer et al. 2015) were used to adjust raw qPCR baseline fluorescence measurements and remove between-run variation that occurs when a quantitative PCR experiment includes multiple plates to accommodate multiple samples and replications. These adjustments in Cq values accounted for plate to plate variation that might be expected when samples were run on different plates, on different days and with different batches of reagents that were prepared the same way. All field samples and all samples that were used for construction of a standard curve for estimating parasitemia were tested with the same positive control and at identical template and reagent concentrations. Both programs are available for download here: https://medischebiologie.nl/files/?main=files&fileName=Factor_qPCR.zip&description=Factor-qPCR:%20removal%20of%20between-run%20variation&sub=Factor_qPCR) References Ruijter, J. M., H. H. Thygesen, J. L. M. Schoneveld, A. Das, B. Berkhout and W. H. Lamers. 2006. Factor correction as a tool to eliminate between-session variation in replicate experiments: application to molecular biology and retrovirology. Retrovirology 3:2 Ruijter, J. M., A. R. Villalba, J. Hellemans, A. Untergasser and M. J. B. van den Hoff. 2015. Removal of between-run variation in a multi-plate qPCR experiment. Biomolecular Detection and Quantification 5: 10-14 20241130 Carter T Atkinson USGS - NW-PACIFIC ISLAND REG Volunteer Emeritus physical

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Hawaii National Park HI 96718 USA 808-985-6401 catkinson@usgs.gov Standard Curve.csv Comma Separated Value (CSV) file containing data. Producer Defined SPECIES Species of bird Producer Defined HAAM Hawaiʻi ʻAmakihi Producer defined BAND U.S. Geological Survey aluminum band number obtained from Bird Banding Laboratory Producer Defined 11951 219124457 TREATMENT How captured birds were handled Producer Defined AVIARY_EXP Bird was removed from the wild and experimentally infected with avian malaria in captivity Producer defined WILD Bird was captured, banded, bled for collection of a blood sample and released into the wild Producer defined ISLAND Island where bird was captured Producer Defined Hawaii Island of Hawaiʻi Producer defined CAPTURE_DATE Date of capture, recorded as mm/dd/yyyy Producer Defined Calendar date when forest birds were captured with mist nets CAPTURE_LOCATION Location where bird was captured Producer Defined Upper Waiākea Forest Reserve Upper Waiākea Forest Reserve, State of Hawaii Producer defined Malama Kī Forest Reserve Malama Kī Forest Reserve, State of Hawaii Producer defined Brysons Study Site, Pu'u Kali'u Bryson's Cinder Cone - Pu'u Kali'u, Kamehameha Schools, Bishop Estate Producer defined Nanawale Forest Reserve Nanawale Forest Reserve, State of Hawaii Producer defined LATITUDE_DD Latitude in decimal degrees, WGS 1984 Producer Defined 19.439940 19.663845 decimal degrees LONGITUDE_DD Longitude in decimal degrees, WGS 1984 Producer Defined -155.369714 -154.866476 decimal degrees ELEVATION_M Elevation above sea level in meters Producer Defined 28 1708 meters SAMPLE_NUMBER Alpha numeric code identifying study site and blood sample number Producer Defined Two or three letter alpha code representing study location followed by 4 digit numeric code representing blood sample number. Alpha codes defined as: AV - Aviary MAL - Malama Kī Forest Reserve BRY - Bryson's Cinder Cone (Pu'u Kali'u) NAN - Nanawale Forest Reserve DNA_LOG Laboratory logbook number for extracted DNA from an individual blood sample Producer Defined 18961 19282 CQ1 Cq value from the first of three qPCR replicates to determine presence of Plasmodium DNA in the blood sample. Reported value is the cycle number at which the fluorescent signal from a sample first rises above a machine determined threshold level. A value of 0 indicates no amplification of parasite DNA Producer Defined 21.806 31.604 cycles CQ2 Cq value from the second of three qPCR replicates to determine presence of Plasmodium DNA in the blood sample. Reported value is the cycle number at which the fluorescent signal from a sample first rises above a machine determined threshold level. A value of 0 indicates no amplification of parasite DNA Producer Defined 21.807 31.755 cycles CQ3 Cq value from the third of three qPCR replicates to determine presence of Plasmodium DNA in the blood sample. Reported value is the cycle number at which the fluorescent signal from a sample first rises above a machine determined threshold level. A value of 0 indicates no amplification of parasite DNA Producer Defined 21.854 31.245 cycles MEAN_CQ Mean Cq value of replicates 1 - 3. Producer Defined 21.822 31.535 cycles ADJUSTED_MEAN_CQ Standardized adjusted mean Cq to account for plate to plate variation among samples that were tested on different plates. Adjustments were based on inclusion of the same positive standard control on each plate and calculated using the programs LinRegPCR and Factor-PCR Producer Defined 21.333 30.828 cycles PARASITEMIA Number of infected red blood cells per 1000 erythrocytes as determined by microscopy Producer Defined 0.019920319 129.007036700 count Molokai Banding Table with Diagnostics.csv Comma Separated Value (CSV) file containing data. Producer Defined SPECIES Four letter alpha numeric code describing each species of forest bird Producer Defined JBWA Japanese Bush Warbler, Horornis diphone Producer defined WAWE Warbling White Eye, Zosterops japonicus Producer defined HAAM Hawaiʻi ʻAmakihi, Chlorodrepanis virens Producer defined APAP ʻApapane, Himatione sanguinea Producer defined RBLE Red-billed Leiothrix, Leiothrix lutea Producer defined NOCA Northern Cardinal, Cardinalis cardinalis Producer defined SBMU Scaley-breasted Munia, Lonchura punctulata Producer defined COMY Common Myna, Acridotheres tristis Producer defined HOFI House Finch, Haemorhous mexicanus Producer defined WRSH White-rumped Shama, Copsychus malabaricus Producer defined HOSP House Sparrow, Passer domesticus Producer defined RCCA Red-crested Cardinal, Paroaria coronata Producer defined DATE Date of capture, recorded as mm/dd/yyyy Producer Defined Calendar date when forest birds were captured with mist nets TIME hr:min:sec, AM or PM Producer Defined NR Not Recorded Producer defined Time of day when bird was captured where AM represents the 12 hour period between midnight and noon and PM represents the 12 hour period from noon to midnight LOCATION Specific location on Molokai where bird was captured Producer Defined Hanalilolilo Trail Kamakou Preserve Producer defined Puuokaeha Puʻuokaʻeha, Kamakou Preserve Producer defined Puukaua Puʻukauā, Kamakou Preserve Producer defined Church Row - Coconut Road near town of Kaunakakai Producer defined Kalawao Kalaupapa National Historical Park Producer defined NPS Office National Park Service Office at Kalaupapa National Historical Park Producer defined USGS Cabin Cabin near Pelekunu Valley stream gauge Producer defined Puuhoi Ridge Puʻuhoi Ridge, located near the head wall of Pelekunu Valley, Pelekunu Preserve Producer defined North Fence North Fence line, Pu'u Ali'i Natural Area Reserve Producer defined SITE Wider region on Molokai where bird was captured Producer Defined Kamakou Preserve Kamakou Preserve, The Nature Conservancy Producer defined Kaunakakai Town of Kaunakakai, State of Hawaii Producer defined Kalaupapa NHP Kalaupapa National Historical Park, National Park Service Producer defined Pelekunu Valley Pelekunu Valley, Pelekunu Preserve, The Nature Conservancy Producer defined Puu Alii NARS Pu'u Ali'i Natural Area Reserve, State of Hawaii Producer defined LATITUDE_DD Latitude in decimal degrees, WGS 1984 Producer Defined 21.097139 21.187968 decimal degrees LONGITUDE_DD Longitude in decimal degrees, WGS 1984 Producer Defined -157.038290 -156.876563 decimal degrees ELEVATION_M Elevation above sea level in meters Producer Defined 5 1096 meters CAPTURE_STATUS Code describing whether a bird is newly banded, a recapture from a previous banding session, or escaped from the net or bag prior to full processing at the banding station Producer Defined 1 Newly banded bird Producer defined R Recaptured bird, identified by prior banding Producer defined 0 Bird escaped from net or bag Producer defined BAND_SIZE U.S. Geological Survey aluminum band size based on diameter of tarsus Producer Defined 1 2.39 mm inside diameter Producer defined 0 2.11 mm inside diameter Producer defined 1B 2.77 mm inside diameter Producer defined 1A 3.17 mm inside diameter Producer defined NB Not Banded Producer defined 0A 1.98 mm inside diameter Producer defined 2 3.96 mm inside diameter Producer defined BAND_NUMBER U.S. Geological Survey aluminum band number obtained from Bird Banding Laboratory Producer Defined NB Not banded Producer defined 150037689 805102940 SEX Sex as determined by plumage, breeding condition, morphometrics or some combination of these characteristics Producer Defined ND Not Determined Producer defined M Male Producer defined U Unknown Producer defined F Female Producer defined HOW_SEX Method or methods used to determine sex Producer Defined W Wing Cord Producer defined N Unknown Producer defined A Adult Plumage Producer defined ND Not Determined Producer defined G Graph of morphometric data Producer defined C Cloacal Protuberance Producer defined B Brood Patch Producer defined AGE Age as determined by plumage, breeding condition or morphometric measurements Producer Defined AHY After Hatch Year Producer defined ND Not Determined Producer defined ASY After Second Year Producer defined U Unknown Producer defined SY Second Year Producer defined HY Hatch Year Producer defined HOW_AGE Characteristics that were used to assign Age at capture Producer Defined S Skull Pneumatization Producer defined ND Not Determined Producer defined A Adult Plumage Producer defined N Unknown Producer defined B Brood Patch Producer defined C Cloacal Protuberance Producer defined J Juvenile Plumage Producer defined F Feather Wear Producer defined BP Brood Patch Producer Defined N No Brood Patch Present Producer defined ND Not Determined Producer defined S Smooth - a loss of breast and some abdominal feathers but most of the area is still rather smooth and dark red Producer defined V Vascularized - abdominal skin thickened with increased fluid and vascularization Producer defined W Wrinkled - abdominal skin thinning, wrinkly and scaly Producer defined M Molting - new pin feathers are present on the abdomen Producer defined CP Cloacal Protuberance Producer Defined N None - cloaca not enlarged Producer defined L Large - cloaca larger at tip than at base Producer defined ND Not Determined Producer defined M Medium - cloaca as large at base as at tip Producer defined S Small - cloaca wide at base and narrow at tip Producer defined WEIGHT_BIRD+BAG Weight of cloth bag containing bird Producer Defined ND Not Determined Producer defined 21 55 grams WEIGHT_BAG Weight of cloth bag Producer Defined ND Not Determined Producer defined 9 36.5 grams WEIGHT_BIRD_G Weight of bird Producer Defined ND Not Determined Producer defined 8 42 grams TARSUS_MM Tarsus length in mm Producer Defined ND Not Determined Producer defined 10 37 mm BILL_MM Length of culmen in mm Producer Defined ND Not Determined Producer defined 8.7 21.7 mm WING_MM Wing Chord in mm Producer Defined ND Not Determined Producer defined 49 136 mm TAIL_MM Tail Length in mm Producer Defined ND Not Determined Producer defined 33 141 mm PRIMARY_MOLT Flight Feather Molt Producer Defined ND Not Determined Producer defined N None Producer defined S Full, normal, symmetric molt Producer defined A Adventitious, accidental, asymmetric molt Producer defined TAIL_MOLT Tail Feather Molt Producer Defined ND Not Determined Producer defined N None Producer defined S Full, normal, symmetric molt Producer defined A Adventitious, accidental, asymmetric molt Producer defined BODY_MOLT Body Feather Molt Producer Defined ND Not Determined Producer defined 0 None Producer defined 1 Trace; a few feathers anywhere on body Producer defined 2 Light; a number from any individual tract (fewer than 1/3 overall) or most of head Producer defined 3 Medium, body feathers in several tracts molting Producer defined 4 Heavy: more than half from most tracts (more than 2/3 of body) Producer defined FAT Categorical classification of fat reserves in the furcular hollow Producer Defined ND Not Determined Producer defined 3 Furculum is filled with fat with some covering the clavicle, but area is not bulging Producer defined 2 Furculum is partially (20-40%) filled, mostly along edges Producer defined 1 Trace; fat barely present along sides of furculum Producer defined 4 Furculum is bulging, overflowing with fat Producer defined 0 No fat seen in furculum Producer defined POX_LESIONS Crusty or smooth swelling present on exposed skin of toes, legs, head or wings Producer Defined False Lesion absent Producer defined True Lesion Present Producer defined SKULL Percent Skull Pneumatization Producer Defined ND Not Determined Producer defined 1 Trace pneumatization at very back of skull; 1-5% of total area Producer defined 2 6-33% of skull pneumatization; at back; distinct contrast; whitish dots Producer defined 3 34-66% pneumatization; rear half and some of front, midline and sides Producer defined 4 67-95% pneumatization; two oval red spots on either side of cranium, the rest fully ossified Producer defined 5 96-99% pneumatization; two tiny, pinkish spots Producer defined 6 Complete, fully pneumatized Producer defined 8 Skull examined but degree of pneumatization not determined Producer defined BLOOD Blood sample status Producer Defined True Blood sample collected Producer defined False Blood sample not collected Producer defined PLASMA Plasma sample status Producer Defined True Plasma sample collected Producer defined False Plasma sample not collected Producer defined DNA DNA sample status Producer Defined True DNA sample collected from packed blood cells Producer defined False DNA sample not collected from packed blood cells Producer defined SLIDE Blood smear collection status Producer Defined True Blood smear collected Producer defined False Blood smear not collected Producer defined SAMPLE_NUMBER Alpha numeric code identifying study site and blood sample number Producer Defined AAA999A Three letter alpha code representing study site followed by 3 digit numeric code with or without a subsequent 1 digit alpha code representing blood sample number. Alpha codes defined as: ALI - North Fence, Pu'u Ali'i NAR HOI - Puʻuhoi Ridge, Pelekunu Valley KAI - Kaunakakai KAM - Hanalilolilo Trail, Kamakou Preserve KAU - Puʻukauā, Kamakou Preserve KNP - Kalawao, Kalapapa NHP MES - Puʻuokaʻeha, Kamakou Preserve PEL - USGS cabin, Pelekunu Valley KNP - NPS Office, Kalapapa NHP Producer defined NS No blood sample collected Producer defined HEMATOCRIT Percent packed red blood cells after blood sample spun in a portable hematocrit centrifuge Producer Defined ND Not Determined Producer defined 28 68 relative percentage of blood volume COMMENTS Auxiliary notes made when handling bird. Descriptions of pox-like lesions recorded here. Producer Defined << empty cell >> No Data Producer defined Miscellaneous text describing pox lesions or additional observations IMMUNOBLOT Results from immunoblot assays to detect anti-Plasmodium antibody using the plasma fraction of blood samples Producer Defined ND Not Determined Producer defined P Positive test results. One or more bands characteristic of forest bird malarial infections present on the blot Producer defined N Negative test results. No bands characteristic of forest bird malarial infections present on the blot Producer defined E Equivocal test results. One or more bands of similar size to those present in forest bird malarial infections. Producer defined MICROSCOPY Results from microscopic examination of Giemsa-stained blood smears Producer Defined NS No Slide Producer defined N Negative. Intraerythrocytic parasites not detected after examination of 100, 400X fields Producer defined P Positive. One or more parasitized erythrocytes detected after examination of 100, 400X fields Producer defined U Undetermined, slide was poorly stained or unreadable by microscopy Producer defined qPCR Results of real time PCR assay for detection of rRNA gene of Plasmodium relictum, lineage GRW4 Producer Defined ND Not Determined Producer defined N Negative. No amplification of parasite rRNA genes detected Producer defined P Positive. Amplification of parasite rRNA genes detected Producer defined QPCR_PROCEDURE Laboratory procedure number for determination of qPCR test results Producer Defined ND Not determined, qPCR test not performed Producer defined Numbers represent a particular laboratory notebook record designated as year test was performed (YY) followed by laboratory notebook number (XXXX) DNA_LOG Laboratory logbook number for extracted DNA from an individual blood sample Producer Defined ND Not Determined, DNA sample was not available or not recovered from an extraction Producer defined 18556 20484 count NG_REACTION Nanograms of template DNA used in a qPCR reaction. Samples with values less than 100 were from extractions with low concentrations of template DNA. Producer Defined ND Not Determined, template DNA not available for testing Producer defined 12 100 ng CQ1 Cq value from the first of three qPCR replicates to determine presence of Plasmodium DNA in the blood sample. Reported value is the cycle number at which the fluorescent signal from a sample first rises above a machine determined threshold level. A value of 0 indicates no amplification of parasite DNA Producer Defined ND Not Determined Producer defined 0 45.08 cycles CQ2 Cq value from the second of three qPCR replicates to determine presence of Plasmodium DNA in the blood sample. Reported value is the cycle number at which the fluorescent signal from a sample first rises above a machine determined threshold level. A value of 0 indicates no amplification of parasite DNA Producer Defined ND Not Determined Producer defined 0 40.92 cycles CQ3 Cq value from the third of three qPCR replicates to determine presence of Plasmodium DNA in the blood sample. Reported value is the cycle number at which the fluorescent signal from a sample first rises above a machine determined threshold level. A value of 0 indicates no amplification of parasite DNA Producer Defined 0 44.15 cycles MEAN_CQ Mean Cq value of replicates 1 - 3. A value of 0 indicates no amplification of parasite DNA Producer Defined ND Not Determine Producer defined 0 45.08 cycles ADJUSTED_MEAN_CQ Standardized adjusted mean Cq to account for plate to plate variation among samples that were tested on different plates. Adjustments were based on inclusion of the same positive standard control on each plate and calculated using the programs LinRegPCR and Factor-PCR Producer Defined ND Not Determined Producer defined 0 43.53 cycles This section of the XML includes two ‘Detailed’ tabs with an associated CSV file named Molokai Banding Table with Diagnostics.CSV and an associated Standard Curve.CSV. The Molokai Banding Table with Diagnostics CSV file combines forest bird morphological, physiological, malarial, and pox-like lesion occurrence for forest bird captures at multiple sites on Molokaʻi between February and November, 2002. The Standard Curve CSV file contains parasitemia data (number of infected cells/1000 red blood cells) for a sample of chronically infected wild Hawaiʻi ʻAmakihi and a sample of captive Hawaiʻi ʻAmakihi that were experimentally infected with Plasmodium relictum. None GS ScienceBase U.S. Geological Survey mailing address

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Denver CO 80225 United States 1-888-275-8747 sciencebase@usgs.gov Unless otherwise stated, all data, metadata and related materials are considered to satisfy the quality standards relative to the purpose for which the data were collected. Although these data and associated metadata have been reviewed for accuracy and completeness and approved for release by the U.S. Geological Survey (USGS), no warranty expressed or implied is made regarding the display or utility of the data on any other system or for general or scientific purposes, nor shall the act of distribution constitute any such warranty. Any use of trade, firm, or product names is for descriptive purposes only and does not imply endorsement by the U.S. Government. Digital Data https://doi.org/10.5066/P13OSHIG None 20260504 U.S. Geological Survey, NW-PACIFIC ISLAND REG Data Steward physical

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Hawaii National Park HI 96718 USA 808-210-6141 pierc-datasteward@usgs.gov FGDC Biological Data Profile of the Content Standard for Digital Geospatial Metadata FGDC-STD-001.1-1999