Iophenoxic acid as a serum marker for red deer (Cervus elaphus scoticus)

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Wildlife Research Volume 25, 1998 © CSIRO Australia 1998

A journal for the publication of original scientific research in the biology and management of wild native or feral introduced vertebrates

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Wildlife Research, 1998, 25, 649–654

Iophenoxic acid as a serum marker for red deer (Cervus elaphus scoticus) P. J. Sweetapple and G. Nugent Landcare Research, PO Box 69, Lincoln, New Zealand.

Abstract As part of a study investigating the efficacy of toxic foliage baits for controlling red deer in New Zealand, a pen trial was conducted to determine whether red deer could be marked by oral dosing with the serum marker iophenoxic acid. The doses required to mark red deer, retention times, and the relationship between dose and serum concentration of the marker were also investigated. Three groups of three deer were fed baits containing 0.26, 2.32, and 6.10 mg iophenoxic acid per kilogram of bodyweight. Mean serum iodine concentration was significantly greater in deer fed marked baits for at least 40 days after dosing than in deer fed unmarked baits. The level of marking in each group was proportional to the dose, suggesting that iophenoxic acid may be used to quantify the amount of bait consumed by red deer. Iophenoxic acid is, therefore, an effective serum marker for red deer and could be used in short- and medium-term ecological and bait technology development field trials.

Introduction Introduced wild deer are present throughout most of New Zealand’s forests and are seen as conservation pests by managers of indigenous forests because their selective browsing has modified the floristic structure in these habitats (Challies 1990; McKelvey 1995). In some areas they also pose an animal health risk because they can carry bovine tuberculosis (e.g. Nugent and Lugton 1995) and may be directly or indirectly responsible for infecting and/or maintaining the disease in domestic cattle. To reduce their impacts as pests, deer populations are at present controlled mainly by commercial and recreational hunting and, in some areas, by poisoning with aerially sown carrot or cereal baits. However, these methods do not always reduce deer densities to levels that meet management objectives. As a further step toward development of an alternative cost-effective method of killing deer a study was undertaken to investigate the efficacy of natural foliage bait poisoning (applying a toxic paste to branches of preferred food species) for controlling wild deer populations in indigenous forests. As part of this study an acceptance trial of non-toxic bait was planned for a central North Island, New Zealand, forest containing red deer (Cervus elaphus scoticus). Before this study could be carried out a bait marker was required. Bait markers are non-toxic substances incorporated into baits that visually and/or chemically ‘label’ or ‘mark’ animals that have eaten the marked baits. They have been used in a range of ecological and bait-development studies such as simulated population control or wildlife vaccination studies (e.g. Linhart and Kinnelly 1967; Morgan 1982; Hadidian et al. 1989; Fletcher et al. 1990). Iophenoxic acid (a-ethyl-3-hydroxy-2,4,6-triiodobenzenepropanoic acid), a serum marker, has been identified as a suitable bait marker for a wide range of species including carnivores such as coyotes, raccoons, foxes, dogs, and cats (Larson et al. 1981; Baer et al. 1985; Eason et al. 1994), herbivores such as goats, rabbits and white-tailed deer (Eason and Batcheler 1991; White et al. 1995; King et al. 1998), and omnivores such as the pig (Fletcher et al. 1990). Dosing with iophenoxic acid results in elevated levels of serum iodine. The period of elevated serum iodine concentration following ingestion of iophenoxic acid varies considerably between species. At one extreme, cats (Felis catus) dosed with 1.5 mg iophenoxic acid per kilogram of bodyweight were marked for over 20 weeks (Eason et al. 1994), while at the other,

10.1071/WR97090

1035-3712/98/060649

650

possums (Trichosurus vulpecula) dosed with 10 mg kg–1 were marked for only 1 week (Eason et al. 1994). The length of time that animals are marked must be determined for each species before iophenoxic acid can be used as a bait marker, to be certain that the absence of increased serum iodine concentration is a consequence of non-consumption, rather than low absorption and/or rapid elimination. A pen trial was carried out to determine whether red deer could be marked by oral dosing with iophenoxic acid, the dose required to mark them, the length of time they were marked, and whether iophenoxic acid could be used to estimate the amount of bait eaten. This paper describes the results of that pen trial. Methods Eleven captive red deer at the Lincoln University Deer Farm (Lincoln, New Zealand), were divided into four groups, weighed, and then force-fed a bolus of broadleaf (Griselinia littoralis) leaves containing 0, 20, 200, or 400 mg of iophenoxic acid (Table 1). The iophenoxic acid was custom synthesised by Aldrich Chemical Co., Milwaukee, USA and was applied to the leaves mixed with a carrier that consisted of one part petrolatum grease (soft white paraffin, supplied as ‘Snow White Petrolatum’ by Shell Oil, New Zealand Ltd) and two parts carbopole (a gel of ‘Carbopole 941’, supplied by B.F. Goodrich Chemical Co., and buffered with triethanoline to pH 7). This is the carrier mix we see as most likely to be chosen for future field use in New Zealand. The doses used (0.26–6.1 mg kg–1) covered the range known to have successfully marked large herbivores in previous trials (Eason and Frampton 1992; White et al. 1995). Each treatment group (those animals fed marked baits) consisted of at least one female and one male. All treated animals were either 11 or 23 months old. The control group (those animals receiving unmarked baits) consisted of

Table 1.

Treatments and sample sizes for the four groups of deer dosed with iophenoxic acid

Group

Treatment

Mean Dose (mg kg–1)

Number of animals

1 2 3 4

2.5 g unmarked carrier 20 mg in 0.25 g carrier 200 mg in 2.5 g carrier 400 mg in 5.0 g carrier

0.00 0.26 2.32 6.10

2 3 3 3

one male and one female, both 11 months old. The standard method of measuring iophenoxic acid concentration in animals is measurement of total iodine concentration in blood serum (Mudge et al. 1978; Hadidian et al. 1989). A blood sample (10 mL) was taken from the jugular vein of all the experimental deer, once before dosing (to establish ‘baseline’ iodine concentrations), and then 1, 5, 11, 21, and 40 days after dosing. One animal from each group was also sampled after 182 days. The blood samples were then centrifuged and the resulting serum samples were analysed for total iodine content using the methods described by Hadidian et al. (1989). The mean serum iodine concentrations for each treatment group were compared using repeated measures analysis of covariance using weight of deer as the covariate. Data was loge-transformed to correct for unequal variances between experimental groups. Pairwise comparisons, with Bonferroni adjustments, were used to compare individual groups at any given time. Our studies were conducted in accordance with the guidelines of the New Zealand National Animal Ethics Advisory Committee, and with Landcare Research’s Animal Ethics Committee approval.

Results Mean pre-treatment (baseline) iodine concentration was 6.6 µg per 100 mL serum in the 11 deer used in the trial. Serum iodine concentration in the control group remained stable

651

Serum iodine concentration (µg per 100 mL)

10000

400 mg

1000

200 mg

100

20 mg

10

control

0 0

20

40

182

Time (days) Fig. 1. Semi-log plot of means (plus standard errors) of serum iodine concentration for red deer fed 0, 20, 200, and 400 mg iophenoxic acid. Sample sizes at Day 182 are one deer in each group

throughout the trial (Fig. 1). Therefore, any animal can be considered marked if it has a serum iodine concentration above 9.6 µg per 100 mL (mean baseline + 2 × standard deviation). For all treatment groups, iophenoxic acid was readily absorbed from the petrolatum/carbopole carrier on the broadleaf leaves by deer, resulting in a serum iodine concentration well above that of the control group (F3,6 = 660, P < 0.001; Fig. 1). Serum iodine concentration was also dependent on bodyweight (F1,3 = 67, P < 0.001), with the largest deer having the lowest concentrations (deer weights were 53–120 kg). All pairwise comparisons of post-dosing mean serum iodine concentrations were significant (P < 0.001) at all times up to and including Day 40, indicating that all three treatment groups were marked for at least 40 days, and that animals receiving high doses were more heavily marked than animals receiving smaller doses (Fig. 1). The dose response appeared to be linear over the range of doses tested (Fig. 2). Using the serum iodine concentration 1 day after dosing as the maximum iodine concentration, serum iodine half-life (estimated by inspection of the data) was approximately 17 days for animals dosed with 400 mg of iophenoxic acid and 11 days for the two other treatment groups (Fig. 1). Despite these half-lives being less than 20 days, all treatment animals still had highly elevated serum iodine concentrations at Day 40. Even 182 days after dosing, the single animals from groups dosed with 400, 200, and 20 mg iophenoxic acid had serum iodine concentrations of 121, 43, and 14 µg per 100 mL respectively (i.e. all were still marked). This reflects an exponential decay rate of the serum iodine, with the rate of elimination of serum iodine slowing with time. Discussion Iophenoxic acid is readily absorbed into the serum of red deer, and is retained for at least 40 days following oral administration of as little as 0.26 mg kg–1. This would provide sufficient

652

Serum iodine concentration (g per 100 mL)

3000

96

2

3

9.

2 )-

=

0.

R

e

os

39

(D

1 .7 y 7 Da I] = [S

2000

1000 0 Day 4 2.089 [SI] =

.4 R

) -14 (Dose

2

6

= 0.9

0 0 20

200 Dose (mg IPH)

400

Fig. 2. Plot of serum iodine concentration against dose of iophenoxic acid (dose-response curve), at Day 1 and Day 40. Simple linear regression models and statistics are given.

time to conduct a wide range of bait acceptance field trials. The very high mean serum iodine concentrations 40 days after animals were dosed with 200 and 400 mg iophenoxic acid, and serum iodine concentrations several times higher than baseline concentrations in two animals tested 182 days after dosing, indicate that red deer are likely to be marked for many months by dosing with 2–6 mg kg–1 iophenoxic acid. White-tailed deer, the only other species of deer for which iophenoxic acid has been tested as a serum marker, were marked for only 3 weeks following dosing with 5 mg kg–1 (White et al. 1995). This shorter period of marking is at least partly due to more rapid elimination of serum iodine than in red deer. Mean elimination rate over 14 days from white-tailed deer with an initial serum iodine concentration of 516 µg per 100 mL was 4.7% per day (White et al. 1995), but was only 1.9% per day over 19 days from red deer with the same initial level of marking (528 µg per 100 mL) in our study. Initial absorption of iophenoxic acid into the blood of the white-tailed deer was not measured by White et al. (1995). The half-life of serum iodine in feral goats (Capra hircus), another mammalian herbivore, was 81 days after dosing with 1.5 mg kg–1 iophenoxic acid (Eason and Frampton 1992), considerably longer than the estimated 11–17 days for the red deer in this study. The considerable variation in retention of iophenoxic acid between these similar species (red deer, white-tailed deer, and goats) and the wide range of absorption and retention of iophenoxic acid in other species (Larson et al. 1981; Baer et al. 1985; Hadidian et al. 1989; Eason et al. 1994) highlights the need to confirm the suitability of this serum marker for each individual species and for each proposed use before undertaking experimental trials.

653

The proportional increases in serum iodine concentration with increases in the amount of iophenoxic acid given to the trial deer indicates that this serum marker could be used to quantify the amount of bait eaten by individual red deer. Such quantitative information is useful for calculating toxic loadings in bait-development studies. The linear nature of the dose-response curve over the dose range of 20–400 mg means that the amount of bait consumed can be easily calculated over a useful range of consumption. However, the relatively rapid initial elimination of serum iodine means that the time that baits were eaten needs to be known (i.e. baits must be made available for only a few days) before this can be done. In studies that use natural foliage baits, this will usually be practical only in trials with captive animals or in small-scale field trials. Serum iodine concentration following ingestion of baits marked with iophenoxic acid has previously been used to quantify the amount of bait eaten by goats (Parkes 1991) and foxes (Saunders et al. 1993). In summary, iophenoxic acid is an effective serum marker for red deer and could be used in short- and medium-term ecological and bait-development field trials. Acknowledgments Thanks to D. Batcheler for conducting the serum iodine assays, to M. Keeley (Lincoln University Deer Farm) for the use of the deer and assistance with dosing and blood sampling, to R. Barker for statistical analysis of the results, to C. Eason for advice and comments on this manuscript, to T. Pearson for drafting the figures and to B. Warburton and M. Ogle-Mannering for reviewing and editing this manuscript. This work was funded by the Animal Health Board of New Zealand. References Baer, G. M., Shaddock, J. H., Hayes, D. J., and Savarie, P. (1985). Iophenoxic acid as a serum marker in carnivores. Journal of Wildlife Management 49, 49–51. Challies, C. N. (1990). Red deer. In ‘The Handbook of New Zealand Mammals’. (Ed. C. M. King.) pp. 436–457. (Oxford University Press: Auckland.) Eason, C. T., and Batcheler, D. (1991). Iophenoxic and iopanoic acid as bait markers for feral goats. Wildlife Research 18, 85–90. Eason, C. T., and Frampton, C. M. (1992). The comparative plasma pharmacokinetics of iopanoic and iophenoxic acid in the goat. Xenobiotica 22, 185–189. Eason, C. T. , Batcheler, D., and Frampton, C. M. (1994). Comparative pharmacokinetics of iophenoxic acid in cats and brushtail possums. Wildlife Research 21, 377–380. Fletcher, W. O., Creekmore, T. E., Smith, M. S., and Nettles, V. F. (1990). A field trial to determine the feasibility of delivering oral vaccines to wild swine. Journal of Wildlife Diseases 26, 502–510. Hadidian, J., Jenkins, S. R., Johnston, D. H., Savarie, P. T., Nettles, V. F., Maiski, D., and Baer, G. M. (1989). Acceptance of simulated oral rabies vaccine baits in urban raccoons. Journal of Wildlife Diseases 25, 1–9. King, D. R., Robinson, M. H., Eason, C. T. and Batcheler, D. (1998). Iophenoxic acid as a biomarker for rabbits (Oryctolagus cuniculus). Wildlife Research 25, 65–68. Larson, G. E., Savarie, P. J., and Okuno, I. (1981). Iophenoxic acid and mirex for marking wild, baitconsuming animals. Journal of Wildlife Management 45, 1073–1077. Linhart, S. B., and Kinnelly, J. T. (1967). Fluorescent bone labelling of coyotes with dimethychlortetracycline. Journal of Wildlife Management 31, 317–321. Morgan, D. R. (1982). Field acceptance of non-toxic and toxic baits by populations of the brushtail possum (Trichosurus vulpecula Kerr). New Zealand Journal of Ecology 5, 36–43. Mudge, G. H., Desbiens, N. I., and Stibitz, G. R. (1978). Binding of iophenoxate and iopanoate to human serum albumin. Drug Metabolism and Disposition 6, 432–439. McKelvey, P. (1995). ‘Steepland Forests.’ (Canterbury University Press.) 295 pp. Nugent, G., and Lugton, I. (1995). Prevalence of bovine tuberculosis in wild deer in the Hauhungaroa range, North Island, New Zealand. In ‘Tuberculosis in Wildlife and Domestic Animals’. (Ed. F. Griffin, and G. de Lisle.) pp. 273–275. Otago Conference Series 3. (University of Otago Press: Dunedin.)

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Parkes, J. P. (1991). Phytotoxicity, poison retention, palatability, and acceptance of carriers used in compound-1080–foliage baits for control of feral goats. Wildlife Research 18, 687–694. Saunders, G., Harris, S., and Eason, C. T. (1993). Iophenoxic acid as a quantitative bait marker for foxes. Wildlife Research 20, 297–302. White, L. M., Huetter, D. P., Linhart, S. B., Savarie, P. J., and Van Brackle, M. D. (1995). Iophenoxic acid as an oral biomarker in white-tailed deer. Wildlife Society Bulletin 23, 194–197.

Manuscript received 23 June 1997; accepted 9 June 1998

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