John H. Schulz

USING A COMMERCIALLY AVAILABLE RADIOIMMUNOASSAY TO QUANTIFY CORTICOSTERONE IN AVIAN PLASMA

Abstract Using a commercially available corticosterone I125 double-antibody radioimmunoassay, we developed and validated an assay procedure for determining corticosterone levels in small-volume (≤30 μL) avian plasma samples. We evaluated this procedures utility by measuring plasma corticosterone levels in Indigo Buntings (Passerina cyanea), American Goldfinches (Carduelis tristis), Red-eyed Vireos (Vireo olivaceus), and Mourning Doves (Zenaida macroura). Standard biochemical validations (e.g., parallelism, recovery of exogenous corticosterone) demonstrated that the assay accurately and precisely measured corticosterone in avian plasma. We used a stress capture protocol to physiologically validate the assays ability to determine biologically important changes in corticosterone levels. Males and females from four bird species exhibited a significant increase in plasma corticosterone in response to capture, handling, and restraint. Baseline and stress-induced corticosterone levels in our study were similar to reported values for other passerine species using other radioimmunoassay procedures. Our results suggest that this radioimmunoassay procedure is very effective for determining corticosterone levels in small-volume avian plasma samples and is sensitive enough to detect biologically important changes in the adrenocortical activity of birds. Thus, this assay has considerable utility for measuring stress levels and stress responses in small birds (<15 g), from which only small volumes of plasma (≤30 μL) can be collected. Utilización de un Radioinmunoensayo Disponible Comercialmente para la Cuantificación de Corticosterona en el Plasma de Aves Resumen. Desarrollamos y validamos un proceso de ensayo para determinar los niveles de corticosterona en muestras de pequeño volúmen (≤30 μL) de plasma de aves utilizando un radioinmunoensayo para corticosterona I125 de doble anticuerpo disponible comercialmente. Evaluamos este procedimiento midiendo los niveles de corticosterona en Passerina cyanea, Carduelis tristis, Vireo olivaceus y Zenaida macroura. Validaciones bioquímicas estándares (e.g., paralelismo, recuperación de corticoesteroide exógeno) demostraron que el ensayo midió de modo exacto y preciso la corticosterona en el plasma de las aves. Utilizamos un protocolo de captura que producía estrés para validar fisiológicamente la habilidad del ensayo de detectar cambios biológicamente importantes en los niveles de corticosterona. Hembras y machos de las cuatro especies de aves mostraron un incremento significativo en los niveles de corticosterona en el plasma en respuesta a la captura, manipulación y retención. Los niveles basales e inducidos por el estrés de nuestro estudio fueron similares a valores reportados para otras especies paserinas que utilizaron otros procedimientos de inmunoensayo. Nuestros resultados sugieren que este procedimiento de radioinmunoensayo es muy efectivo para determinar los niveles de corticosterona en muestras de pequeño volúmen de plasma de aves y que es suficientemente sensible como para detectar cambios biológicamente importantes en la actividad adenocortical de las aves. De esta manera, este ensayo presenta considerable utilidad para medir los niveles y respuesta al estrés en aves pequeñas (<15 g) de las cuales sólo es posible colectar pequeños volúmenes de plasma (≤30 μL).

Acute Lead Toxicosis in Mourning Doves

Abstract Previous research has suggested that free-ranging mourning doves (Zenaida macroura) may ingest spent lead pellets, succumb to lead toxicosis, and die in a relatively short time (i.e., an acute lead toxicosis hypothesis). We tested this hypothesis by administering 157 captive mourning doves 2–24 lead pellets, monitoring pellet retention and short-term survival, and measuring related physiological characteristics. During the 19- to 21-day posttreatment period, 104 doves that received lead pellets died (deceased doves) and 53 survived (survivors); all 22 birds in a control group survived. Within 24-hr of treatment, blood lead levels increased almost twice as fast for deceased doves compared to survivors (F1,208 = 55.49; P < 0.001). During the first week, heterophil:lymphocyte (H:L) ratios increased twice as fast for deceased doves than with survivors (F1,198 = 23.14, P < 0.001). Posttreatment survival differed (χ2 = 37.4, P < 0.001) among the 5 groups of doves that retained different numbers of pellets, and survival ranged from 0.57 (95% CI: 0.44–0.74) for doves that retained ≤2 lead pellets 2 days posttreatment compared to 0.08 (95% CI: 0.022–0.31) for those doves that retained 13–19 lead pellets on 2 days posttreatment; significant differences existed among the 5 groups. After controlling for dove pretreatment body mass, each additional lead pellet increased the hazard of death by 18.0% (95% CI: 1.132–1.230, P < 0.001) and 25.7% (95% CI: 1.175–1.345, P < 0.001) for males and females, respectively. For each 1-g increase in pretreatment body mass, the hazard of death decreased 2.5% (P = 0.04) for males and 3.8% (P = 0.02) for females. Deceased doves had the highest lead levels in liver (49.20 ± 3.23 ppm) and kidney (258.16 ± 21.85 ppm) tissues, whereas controls showed the lowest levels (liver, 0.08 ± 0.041 ppm; kidney, 0.17 ± 0.10 ppm). For doves dosed with pellets, we observed simultaneous increases in blood lead levels and H:L ratios, whereas packed-cell volume (PCV) values declined. Our results support an acute lead toxicosis hypothesis. Although further research is necessary to investigate the magnitude of lead shot ingestion and toxicosis in mourning doves, we recommend that management agencies initiate development of a long-term strategic plan aimed at implementing a nontoxic shot regulation for mourning dove hunting.

USING FECAL GLUCOCORTICOIDS FOR STRESS ASSESSMENT IN MOURNING DOVES

Abstract Fecal glucocorticoid assays provide a potentially useful, noninvasive means to study physiological responses of wildlife to various stressors. The objective of our study was to validate a method for measuring glucocorticoid metabolites in Mourning Dove (Zenaida macroura) feces. We validated the assay using standard procedures (e.g., parallelism, recovery of exogenous corticosterone) to demonstrate that the assay accurately and precisely measured glucocorticoid metabolites in Mourning Dove fecal extracts. We conducted adrenocorticotropin (ACTH) challenge experiments to validate the assays ability to determine biologically important changes in fecal glucocorticoids. Fecal glucocorticoid levels increased significantly approximately 2–3 hr after administration of ACTH at 50 IU per kg body mass to wild Mourning Doves held in captivity. In contrast, fecal glucocorticoid metabolites did not increase in control birds, birds that received saline injections, or a lower dose of ACTH (1 IU per kg body mass). Variation in overall fecal glucocorticoid metabolite levels may have been influenced by season and the length of time birds were held in captivity. Noninvasive fecal glucocorticoid metabolite analyses, in combination with demographic information, may have considerable utility for monitoring the effects of natural and anthropogenic disturbances on Mourning Dove populations. Uso de Glucocorticoides Fecales Para Evaluar el Estrés en Zenaida macroura Resumen. Las evaluaciones de glucocorticoides fecales representan un medio no invasor potencialmente útil para estudiar las respuestas fisiológicas de los animales silvestres ante agentes causantes de estrés. El objetivo de nuestro estudio fue validar un método para medir metabolitos glucocorticoides en heces de palomas Zenaida macroura. Validamos el método mediante procedimientos estándar (e.g., paralelismo, recuperación de corticosterona exógena) para demostrar que éste mide con exactitud y precisión los metabolitos glucocorticoides en extractos fecales de Z. macroura. Realizamos experimentos de desafío con adrenocorticotropina (ACTH) para validar la habilidad que tenía el método para determinar cambios biológicamente importantes en los glucocorticoides fecales. Los niveles de glucocorticoides fecales aumentaron de forma significativa aproximadamente 2–3 hr después de la administración de ACTH a 50 IU por kg de peso corporal a palomas silvestres mantenidas en cautiverio. En contraste, los metabolitos glucocorticoides fecales no aumentaron en aves control, ni en aves que recibieron inyecciones salinas o una menor dosis de ACTH (1 IU por kg de peso corporal). La variación en los niveles generales de metabolitos glucocorticoides fecales podría haber sido influenciada por la estación y la longitud del período de tiempo en que las aves fueron mantenidas en cautiverio. Los análisis no invasores de metabolitos glucocorticoides, en combinación con información demográfica, podrían ser de considerable utilidad para monitorear los efectos de los disturbios naturales y antropogénicos sobre las poblaciones de Z. macroura.

The persistent problem of lead poisoning in birds from ammunition and fishing tackle

ABSTRACT Lead (Pb) is a metabolic poison that can negatively influence biological processes, leading to illness and mortality across a large spectrum of North American avifauna (>120 species) and other organisms. Pb poisoning can result from numerous sources, including ingestion of bullet fragments and shot pellets left in animal carcasses, spent ammunition left in the field, lost fishing tackle, Pb-based paints, large-scale mining, and Pb smelting activities. Although Pb shot has been banned for waterfowl hunting in the United States (since 1991) and Canada (since 1999), Pb exposure remains a problem for many avian species. Despite a large body of scientific literature on exposure to Pb and its toxicological effects on birds, controversy still exists regarding its impacts at a population level. We explore these issues and highlight areas in need of investigation: (1) variation in sensitivity to Pb exposure among bird species; (2) spatial extent and sources of Pb contamination in habitats in relation to bird exposure in those same locations; and (3) interactions between avian Pb exposure and other landscape-level stressors that synergistically affect bird demography. We explore multiple paths taken to reduce Pb exposure in birds that (1) recognize common ground among a range of affected interests; (2) have been applied at local to national scales; and (3) engage governmental agencies, interest groups, and professional societies to communicate the impacts of Pb ammunition and fishing tackle, and to describe approaches for reducing their availability to birds. As they have in previous times, users of fish and wildlife will play a key role in resolving the Pb poisoning issue.

EFFECTS OF IMPLANTED RADIOTRANSMITTERS ON CAPTIVE MOURNING DOVES

Previous mourning dove (Zenaida macroura) telemetry studies using transmitter glue attachment have found the technique to be relatively short term (<10 weeks), and that transmitter harnesses possibly have deleterious effects on doves. To improve attachment methods, we developed and refined surgical techniques for implanting subcutaneous and intra-abdominal radiotransmitters with external whip antennas in mourning doves, and we determined physiological and pathological responses to the transmitter implants. We used a captive colony of 200 wild-trapped mourning doves to develop and test procedures for subcutaneous implants (SC1), subcutaneous surgeries without implants (SC2), intra-abdominal implants (IA1), intra-abdominal sur- geries without implants (IA2), and a control group without surgeries or implants (CNT); 20 males and 20 females were assigned to each experimental group. Surgeries for IA1 took less time (3.58 t 0.17 min; i? + SE; P < 0.001) than SC1 surgeries (4.36 ? 0.12 min). Heterophil:lymphocyte ratios showed that IA1 and IA2 doves had higher (P = 0.024) posttreatment changes compared with SC1, SC2, or CNT groups. At 4-6 days postsurgery, 153 of 160 (95.6%) doves with surgical treatments showed closed or healed surgical sites with no complications. At 14 days posttreatment, 34 (87%) SC1 implants were located in the thoracic inlet. Gross necropsy findings at 10 weeks postsurgery found that 36 of 39 (92%) SC1 and 36 of 39 (92%) IA1 implants showed little or no tissue response to the implants. Functioning transmitters began failing 2 weeks posttreat- ment, and 85% were not working at 10 weeks posttreatment. Our data suggest subcutaneous implants with external antennas were a better alternative compared to intra-abdominal implants with external antennas, but further testing is needed to compare subcutaneous implants to conventional attachment techniques.

USING A COMMERCIALLY AVAILABLE ENZYME IMMUNOASSAY TO QUANTIFY TESTOSTERONE IN AVIAN PLASMA

Abstract ABSTRACT Using a commercially available testosterone enzyme immunoassay (EIA), we developed and validated an assay procedure for determining testosterone levels in small-volume (20 µL) avian plasma samples. We evaluated this EIAs utility by measuring plasma testosterone levels in Mourning Doves (Zenaida macroura), White-eyed Vireos (Vireo griseus), Red-eyed Vireos (Vireo olivaceus), and Indigo Buntings (Passerina cyanea). Standard biochemical validations (e.g., parallelism, recovery of exogenous testosterone) demonstrated that the assay accurately and precisely measured testosterone in avian plasma. We compared plasma testosterone levels in males and females of all four species and Indigo Buntings in various reproductive stages to physiologically validate the assays ability to determine biologically important changes in testosterone levels. Plasma testosterone levels were higher in males compared to females in three of four species. Prebreeding and breeding male Indigo Buntings had higher circulating testosterone levels than postbreeding males. Testosterone levels in our study were similar to reported values for other passerine species using radioimmunoassay procedures. Our results suggest that this EIA procedure is very effective for determining testosterone levels in small-volume avian plasma samples and is sensitive enough to detect biologically important changes in the gonadal activity of birds. Thus, this assay has considerable utility for measuring testosterone in small birds (<15 g), from which only small volumes of plasma (20 µL) can be collected.

Using Statistical Population Reconstruction to Estimate Demographic Trends in Small Game Populations

Abstract Statistical population reconstruction offers a robust approach to demographic assessment for harvested populations, but current methods are restricted to big-game species with multiple age classes. We extended this approach to small game and analyzed 14 years of age-at-harvest data for greater sage-grouse (Centrocercus urophasianus) in Oregon, USA, in conjunction with radiotelemetry data to reconstruct annual abundance levels, recruitment, and natural survival probabilities. Abundance estimates ranged from a low of 26,236 in 1995 to a high of 39,492 in 2004. Annual abundance estimates for adult males were correlated with a spring lek count index (r  =  0.849, P < 0.029). We estimated the average annual harvest mortality for the population to be 0.028, ranging from 0.021 to 0.031 across years. We estimated the probability of natural survival of adult females to be 0.818 (), somewhat higher than that of adult males (S ˆ  =  0.609, ). Our precision in reconstructing the population was hampered by low harvest rates and the few birds tagged in the radiotelemetry investigations. Despite these issues, our analysis illustrates how modern statistical reconstruction procedures offer a flexible framework for demographic assessment using commonly collected data. This approach offers a useful alternative to small-game indices and would be most appropriate for species with 5 or more years of age-at-harvest data and moderate-to-heavy harvest rates.

Monitoring Presence and Annual Variation of Trichomoniasis in Mourning Doves

Abstract Information about the annual variation of trichomoniasis in mourning doves (Zenaida macroura) may be important in understanding mechanisms affecting mourning dove populations. The objectives of this study were to monitor the presence and annual variation of Trichomonas gallinae for 6 yr in a local mourning dove population using hunter-killed doves. During 1998–2003, 4052 hunter-killed doves were sampled for the presence of T. gallinae; 226 (5.6%) tested positive (4.4%–10.6% range). Results of the monitoring effort were relatively consistent during the 6-yr period, with the presence of T. gallinae being within the range of previously reported estimates for mourning doves. Asymptomatic carriers in one segment of the dove population may provide a mechanism for spreading the disease to other segments of the mourning dove population.

Accuracy of Techniques Used to Assign Mourning Dove Age and Gender

Sex ratios of mourning doves (Zenaida macroura) reported in banding studies have differed markedly between states and years suggesting that techniques used to assign gender may be biased. We tested this hypothesis with experienced mourning dove biologists (n = 4) who assigned age, gender, primary molt progression, and certainty of classification, using external characteristics of harvested doves (n = 468) in Missouri during September 1993. The probability of correctly classifying adult mourning doves, verified with gonadal examination, was 0.904-0.985 and 0.957-0.990 for immatures. Assessment of age and gender combined was 0.944-0.990 for adult females and 0.533-0.993 for adult males. Error probabilities of observer ability to correctly assign age and gender was 0.0-0.045 for adult females and 0.007-0.451 for adult males. Among individual mourning doves examined, only 1 adult and 6 immature doves were incorrectly assigned age and gender by all 4 participants. Error in assigning gender to adult doves could have contributed to disparate sex ratios reported in previous mourning dove banding studies. We recommend that standard training procedures be developed and implemented across states and regions in future banding studies of mourning doves.

Experimental Lead Pellet Ingestion In Mourning Doves (Zenaida macroura)

ABSTRACT Because the relationship between lead pellet availability and ingestion by mourning doves (Zenaida macroura) remains uncertain, we conducted an experiment to determine if doves held in captivity freely ingest lead shotgun pellets, investigate the relationship between pellet density and ingestion and monitor physiological impacts of doves ingesting pellets. We conducted two trials of the experiment with <60 doves per trial. We randomly assigned 10 doves to one of six groups per trial; 10, 25, 50, 100, 200 pellets mixed with food and a control group with no pellets. We monitored ingestion by examining x-rays of doves 1-d post-treatment and monitored the effects of lead ingestion by measuring heterophil∶lymphocyte (H∶L) ratios, packed-cell volume (PCV), blood lead, liver lead and kidney lead. Pooled data from both trials showed 6 of 117 (5.1%) doves ingested lead pellets. Two mourning doves ingested multiple lead pellets in each of the treatments containing a mixture of 25, 100 and 200 lead pellets and food. Doves ingesting lead pellets had higher blood lead levels than before treatment (P  =  0.031). Post-treatment H∶L ratios, however, were not different compared to pre-treatment values (P  =  0.109). Although post-treatment PCV decreased for four of six doves ingesting lead pellets, overall they were not lower than their pre-treatment values (P  =  0.344). Liver (P < 0.0001) and kidney (P  =  0.0012) lead levels for doves ingesting pellets were higher than doves without ingested pellets. Our lead pellet ingestion rates were similar to previously reported ingestion rates from hunter-killed doves and our physiological measurements confirm earlier reports of a rapid and acute lead toxicosis. Similar to previous field research, we did not observe a relationship between pellet density in the food and ad libitum pellet ingestion. Although one approach would be to ban lead shot for mourning dove hunting on managed public hunting areas, further research is necessary to ensure that policy development and implementation have a consensus among stakeholders.