Environmental Polhaion (Series A) 25 (1981) 291 304 AN EVALUATION OF BLOOD PLASMA FOR MONITORING DDE IN BIRDS OF PREY CHARLES J. HENNY Patuxent Wildlife Research Center, 480 S W Airport Rd., Corvallis, Oregon 97330, USA & DENNIS L. MEEKER Denver Wildlife Research Center, BMg. 16, Federal Center, Denver, Colorado 80225, USA ABSTRACT Laboratory and field studies show that DDE residues in blood plasma are highly correlated with DDE in the brain, the rate of DDE exposure and the amount of DDE in eggs of free-living birds of prey. A blood sampling approach is proposed to supplement existing environmental contaminant methods available for birds of prey. Residues (EDDT) in plasma provide some direct information; however, a method is proposed to adjust blood plasma residues from four species of birds of prey to the estimated residues in eggs for the purpose of residue interpretation. The blood plasma-egg relationship appears to be species-independent in the four raptors studied. Two predicting equations are presented for estimating egg residues, one for plasma samples collected prior to, or within a few days of, egg laying, and another for the post-laying period. Limitations and advantages of the blood plasma monitoring approach are discussed. The approach may be particularly suitable for endangered species research. INTRODUCTION Monitoring pesticides in birds has traditionally taken the form of either collecting eggs or killing individuals for analysis. A notable exception is the wings from waterfowl which are submitted by duck hunters to the US Fish and Wildlife Service (Heath, 1969). Unfortunately, tertiary consumers, which are likely to be exposed to higher levels of organochlorine pesticide contamination, have no ready-made collection of samples available. Furthermore, these species, some of which are 'endangered', are 291 Environ. Pollut. Ser. A. 0143-1471/81/0025-0291/$02-50 © Applied Science Publishers Ltd, England, 1981 Printed in Great Britain 292 CHARLES J. HENNY, DENNIS L. MEEKER generally not abundant and large numbers could not be killed for pesticide analysis. An alternative approach, preferably a method whereby the potential exists for samples to be collected year after year from the same individuals, is needed. Under sublethal conditions of continuous exposure, residues in the different body tissues are ordinarily directly correlated with each other, as shown by Dindal & Peterle (1968) for DDT and its metabolites (EDDT) in ducks. Based upon this reasoning, a fat biopsy approach was used with peregrine falcons Falco peregrinus in the late 1960s ( Cade et ah, 1968; Enderson & Berger, 1968). Both whole blood and plasma have been used satisfactorily as an indicator of pesticide contamination in man (Radomski et ah, 1971 ; Edmundson et al., 1972, and others). We elected to use plasma in our studies. Radomski et al. (1971) concluded that concentrations of organochlorine pesticides in the blood are apparently in equilibrium with those in the tissue and measurement constitutes a highly useful and readily obtainable means of estimating body burdens and exposure. Recently, Friend et al. (1979) tested the relationship between DDE in blood serum and DDE in the fat of mallards Arias platyrhynchos. The relationship in the ducks was strong and did not appear to fluctuate significantly, even when the birds were under the stress of reduced food intake and surgery. The blood plasma procedure was first field-tested in 1974 to monitor the uptake of EDDT in raptors on a spray area in the Pacific Northwest (Henny, 1977). Residues in plasma paralleled residues in eggs during the American kestrel Falco sparverius field study. More recently, Capen & Leiker (1979) showed that blood serum samples from a wild population of white-faced ibis Plegadis chihi may be used to monitor body burdens of DDE. To test further the DDE relationships between blood plasma, fat, whole body and brain in raptors, twenty-eight captive American kestrels were dosed with seven levels of DDE for 11 days. This paper reports on the laboratory study together with the wild raptor field study. METHODS Twenty American kestrels (eleven males and nine females) were trapped near Boise, Idaho, between the 23rd and the 25th of January 1975 and shipped air freight to the Denver Wildlife Research Center. Four birds were placed in each of five pens. A blood sample (1 ml) was collected from the jugular vein of each bird for measurement of background contamination. Weights were also recorded. The kestrels were fed live mice (Peromyscus sp.) for the first 2 days, then fed ZU/PREEM ® 'Birds of prey diet' (produced by Hills Div., Riviana Foods, PO Box 148, Topeka, Kansas 66601). The birds in each pen were given 175g of ZU/PREEM® daily (i.e. about 44 g/bird/day) for a 14-day adjustment period. On MONITORING D D E IN BIRD BLOOD 293 the 8th of February D D E was added to the Z U / P R E E M ®. Propylene glycol was the carrier at 1 ~o concentration in the feed. The D D E was mixed at the following rates: 1, 5, 25, 125 and 625 ppm D D E dry weight. The Z U / P R E E M ® contained 60.2 ~o water and the D D E dosage rate on a wet weight basis was therefore 0.40, 2.0, 10, 50 and 250 ppm. The kestrels were maintained on the treated food for 11 days and returned to clean feed on the 19th of February. All live birds were killed on the 3rd of March after being on clean feed for 13 days. In addition to initial weights, weights were recorded at the beginning and end of the dosage period (the 7th of February and the 20th of February) and on the last day of the study (the 3rd of March), at which time the birds were killed and brain, fat and whole body samples analysed. Food not eaten was collected each day and weighed. During the 11-day treatment, birds in pen 1 (0-4 ppm) ate 1,838 g, pen 11 (2 ppm), 1370 g, pen III (10 ppm) 1665 g, pen IV (50 ppm) 1665 g and pen V (250 ppm) I 181 g. The estimated total amount of DDE consumed by the birds in the respective pens was 0-74mg, 2.7mg, 17mg, 83 mg and 290 mg. Eight additional kestrels were trapped in Colorado in October and early November 1975 and brought to the laboratory to study intermediate dosage rates. The procedures used with these birds were essentially the same. The eight birds were divided into two groups and placed on a DDE diet of 200 and 400 ppm DDE on a dry weight basis, or 80 and 160 ppm on a wet weight basis. The treated food was given the birds for 11 days (the 25th of November to the 5th of December), followed by clean food for 56 days during which blood samples (0.5 ml) were taken periodically. All surviving birds were killed on the 56th day, and brain, fat, and whole body samples were analysed. The final blood samples were collected from the heart by means of a disposable 3-cc syringe with a 23-gauge needle. The blood was then placed in a heparinised tube and kept cool until it was centrifuged a few hours later. The plasma was then drawn off and stored frozen. The plasma yield was usually 40 to 50 ~,,. Field samples were collected in a similar manner, except that blood samples (up to 2 ml) from the larger raptors were taken from the brachial vein. Deviations from these procedures (especially storage of plasma) may render comparisons with the data in this paper invalid. All samples were analysed at the Denver Wildlife Research Center using the procedures of Peterson et al. (1976). RESULTS The laboratot 3' study Condition of birds during and after experiment: Initial body weights, beginning dosage, after the 1 l-day dosage period, and weights 2 weeks later ( 13 days in spring study, 14 days in autumn study) are summarised in Table t. Most birds gained 294 C H A R L E S J. H E N N Y , D E N N I S L. MEEKER TABLE 1 THE INITIALAND SUBSEQUENT WEIGHTS AND DDE LEVELS IN THE PLASMA OF THE TWENTY-EIGHT AMERICAN KESTRELS IN THE LABORATORYSTUDY Dosage Bird No. (wet weight) (ppm) Sex Initial Weight (g) After 2 weeks on clean feed Beginning End dosage dosage 0.4 0.4 0-4 0-4 2 2 2 2 10 10 10 10 50 50 50 50 80 80 80 80 160 160 160 160 250 250 250 250 3 4 7 8 1 2 9 10 5 6 ll b 12 17 18 19 20 21 22 23 24 25 26 27 28 13 14 15 16 M F M F F M F M M F M M F M M F M F F M F F F M F M F M 102 102 112 117 114 102 118 117 ll0 100 119 98 19 10 15 16 10 19 17 101 123 112 124 112 122 106 118 111 118 115 121 130 127 110 120 122 117 107 112 104 125 114 122 122 123 141 152 112 129 128 140 113 121 111 121 120 121 124 119 129 128 76 ~ 103 120 121 117 79 a 112 123 120 105 127 I10 135 134 113 122 126 155 88 a 126 81 a lll 85 ~ Plasma DDE (ppm wet weight) Initial After 2 weeks on clean feed 125 120 120 122 137 -130 117 120 128 -116 132 124 123 131 136(121) c 146(137) 161(160) 121(122) 143(131) 149(140) 159(153) 137 -133 0.18 0-12 0.06 0"04 0.34 0.03 0.39 0.51 0.08 0-55 1-0 0.02 0.29 0.16 0.17 0-88 0-01 <0.0! < 0.01 0-01 0.01 < 0.01 0.01 0.01 0.02 0.01 0.18 0-01 0.04 0.04 0.05 0.08 0"09 -0.19 0.40 0.82 0.47 0-62 2.6 3.4 1.3 5-7 3-3 2.5 1.6 4.3 7.9 9.0 6.7 6-5 -9.5 a Died before completion of study; weight given at time of death. b A crippled bird held in captivity for a long period of time. c Weight after 8 weeks; completion of study. weight during the pre-dosage adjustment period, a notable exception being the only crippled bird (No. 11) used in the study. During the dosage period of the study, more of the survivors tended to gain weight at the lower dosage rates and more tended to lose weight at the higher dosage rates (see summary below). Diet (ppm) Spring low (0-40-10) Spring intermediate (50) Autumn high (80-160) Spring high (250) _+2 g of pre-dosage weight. Weight change Gain NC a Loss 5 2 1 1 4 1 2 0 1 I 4 1 MONITORING D D E IN BIRD BLOOD 295 The 2-week post-period on clean feed yielded weight gains for about half of the birds on the lower dosage rates, but gains were recorded for all birds on the higher rates (50 ppm and higher). The latter birds had generally lost weight during the dosage phase of the study. After 2 weeks on clean feed and under conditions of short treatment and unlimited food, the dosage level of DDE had little or no effect on the lipid content of the fat, brain, or whole body of the birds that survived the study (Table 2). However, the birds dosed in the autumn with 80 and 160ppm DDE showed higher lipid content in all tissues. Since the autumn birds were on clean food for 8 weeks, they are not comparable here for both seasonal and other reasons. The birds that died showed a major weight loss during the last 2 or 3 days of life, much like that reported by Porter & Wiemeyer (1972). The per cent lipid in the brain was similar in the birds that survived (Table 2) and even those that died during the experiment (6.44 _+0.47, n = 5). Bogan & Newton (1977) reported similar findings for European sparrowhawks Accipiter nisus. TABLE 2 LIPID CONTENTOF FAT, BRAINAND WHOLEBODYOF SURVIVINGAMERICANKESTRELSWITH RESPECTTO DOSAGE LEVEL OF DDE (WET WEIGHT) AT TERMINATION OF LABORATORYSTUDY Category 0.4 ppm 2 ppm l Oppm Dosage level 50 ppm 80 ppm 160 ppm 250 ppm Sample size (n) Fat (% lipid) Brain (% lipid) 4 3 3 4 4 3 2 86-8 + 3.77 a 88-0 + 5-29 90.7 ± 3.06 84.0 + 8.29 96.6 i- 1.69 96.4 ± 0.12 95-3 ± 6.08 6 - 5 ± 0 . 5 8 6 . 3 ± 0 - 5 8 6 . 5 ± 0 . 5 0 6 . 8 ± 1 . 5 0 7 . 5 ± 0 - 4 9 7 - 1 ± 0 . 2 5 6.0_+0.00 Whole body ( % lipid) 18.0 _+ 4.24 20.3 +_ 4.04 18-3 ± 1.53 20-8 ± 5.12 25.8 ± 5.06 26.9 ± 8-07 22.0 _+ 2-83 " Mean _+ one standard deviation. Condition of birds that diedduring experiment." Five of fourteen males and none of the fourteen females died during the study. The birds during the last few days of life all showed major weight losses amounting to 27~o, 29~o, 22~o, 29~o and 31 ~o of their weight at the beginning of the dosage experiment. Stickel et al. (1970) correlated brain residues with mortality of experimental cowbirds Molothrus ater. They concluded that residues of 300 ppm and 400 ppm of DDE in the brain (wet weight) can be considered to show increasing likelihood of death from DDE. Three male kestrels in this study (all the males and none of the females on the two highest doses, incidentally) died with residues of 230,223 and 280 ppm of DDE in the brain, suggesting that lethal levels of DDE in male kestrels may be somewhat lower than in male cowbirds, where the lowest observed lethal level was 250 ppm, and 95 ~o of the lethal values were estimated to be between 314 and 793 ppm. In another study in which two of fourteen male kestrels died during a prolonged dietary dosage of 10ppm, brains of birds that died contained 213 and 301 ppm of DDE (Porter & Wiemeyer, 1972). The two birds died at a time of seasonal weight loss and depletion of fat reserves related to reproduction and moult. The results of the present study 296 CHARLES J. HENNY, DENNIS L. MEEKER support the likelihood that DDE was at least the proximal cause of death in the Porter & Wiemeyer (1972) study, even though stress was obviously involved. The two other males that died during the study included a crippled bird that could not fly (No. 11) and that had been held in captivity for about 6 months, but began losing weight immediately after its move to Denver- possibly from being forced to compete with three other birds in the pen. DDE (251 ppm) in the brain of this bird did reach the range reported for the three birds that died on high dosage during this study, as well as the range for the two birds that died during Porter & Wiemeyer's (1972) study. The other bird (No. 2) that died contained only 2.6 ppm DDE in the brain--well below the believed lethal range for DDE. DDE residues in body tissues compared with dosage of DDE: The kestrels were killed after being on clean feed for different periods of time in the spring and autumn studies. D D E residues in fifteen of the sixteen kestrels that survived the spring laboratory study (omitting one bird on the high dosage that appeared about to die [165 ppm D D E in the brain]) were correlated with the amount of DDE in the diet (milligrammes per bird). The best correlation between the amount of DDE ingested and a body tissue was with DDE (wet weight) in the brain (y = 0.359 + 0.174(x), r = + 0.970, n = 15). DDE in the plasma was also highly correlated with the amount of DDE ingested (y = 0.179 + 0.132(x), r = + 0.944, n = 15). The relationships were weaker for whole body ( y = 1 3 2 . 7 + 10.9(x), r = +0.649, n = 15) and fat (y = 5607 + 257(x), r = +0.409, n = 15), although all relationships appeared to be linear. The relationship between D D E in the blood plasma (ppm wet weight) after 2 weeks on clean feed and DDE in the diet (ppm wet weight) is shown in Fig. 1 for the combined spring and i~utumn studies (omitting one bird on high dosage that appeared about to die). A strong positive linear relationship (r = +0.933) exists between exposure and DDE in the plasma. DDE in plasma compared with DDE in brain: DDE residues in the brain were significantly correlated with D D E residues in the plasma (Fig. 2). Figure 2 includes both the spring (killed after 2 weeks on clean feed) and autumn (killed after 8 weeks on clean feed) data from the laboratory study plus four birds collected from the wild. The only surviving bird that was excluded was the bird about to die (i.e. 165 ppm in brain). The linear correlation (r = +0-964) from autumn and spring laboratory studies was nearly as strong as that from the spring study alone (r = + 0.984) where all birds were killed after 2 weeks on clean feed. The data indicate that an equilibrium was being maintained over the time periods studied between the blood plasma and the brain. Plasma DDE changes over time: During the autumn study the birds were kept for 56 days following the termination of the dosage phase of the study. A 0.5 ml blood sample was collected at 1 week, 2 weeks, 4 weeks and 8 weeks to evaluate changes in MONITORING D D E 297 IN BIRD BLOOD 10 < oO < J D_ c LLI £3 £3 0 0 ' J • o • 40' ' DDE r =÷ 0933 n:22 8'0 . . 120 . . . in D i e t (ppm 160 wet 20 0 ' 240 ' ! ! 280 weight) Fig. 1. The relationship between DDE in the diet (ppm wet weight) and DDE in blood plasma (ppm wet weight) of American kestrels (values for 0.4 ppm not plotted, but are included in regression). blood plasma DDE levels during the period on clean feed (Table 3). Residues declined during the 8-week period on clean feed. The percentage loss was 16-7 ~o during the second week, averaged 5 % per week during the third and fourth weeks, and averaged 4-4 % per week during the last 4 weeks of the study. The field study Blood plasma samples were collected from wild American kestrels, goshawks, Accipiter gentilis, Cooper's hawks A. cooperii, and sharp-shinned hawks A. striatus in a DDT spray area in the Pacific Northwest (Henny, 1977). The study was designed with two purposes: (1) to evaluate trends and patterns of residue build-up in raptors from a single aerial DDT application and (2) to evaluate the blood plasma technique as a monitoring tool for birds of prey. DDT and its metabolites were combined in the field study because laboratory tests have shown that DDE increased as DDT was mobilised (e.g. Ecobichon & Saschenbrecker, 1968). Eggs (an accepted monitoring approach) were taken, in addition to blood samples, for evaluating the pesticide patterns and trends in the populations. It became readily apparent that residues in plasma paralleled those found in eggs of the American kestrel during the study (Henny, 1977). In fact, a log-log relationship 13 12 11 10 9 uJ a £3 8 7 xu. • 6 Z < n" 5 1.408 X 4 r-- * 0 . 9 6 4 ./ 3 • n = 26 2 1 0 0 I I I I I I I I J I 1 2 3 4 5 6 7 8 9 10 BLOOD Fig. 2. PLASMA (DDE) The relationship between D D E in blood plasma (ppm wet weight) and D D E in the brain (ppm wet weight) of American kestrels. The a u t u m n study ('~tr) and the spring study (O). TABLE 3 DDE RESIDUES ( P P M W E T W E I G H T ) IN AMERICAN KESTREL B L O O D PLASMA, FAT, BRAIN A N D W H O L E B O D Y , F O L L O W I N G A DIETARY I N T A K E OF DDE [FOR 1 l DAYS Diet DDE (ppm wet weight 80 80 80 80 160 160 160 160 Mean Initial" 0.01 <0.01 <0.01 0.01 0.01 <0.01 0-01 0.01 0.01 7 days b 5-4 2.6 3.5 4.7 12 7.8 5.8 . 6"0 Plasma DDE 14 days 28 days . 3.3 2"5 1.6 4.3 7.9 9.0 6.7 . 5.0 . 1-5 2.2 1.1 4.5 9.6 6"5 5.8 . 4.5 56 days Fat 2.0 1.8 0.84 2.3 7.1 6"0 5-6 420 440 220 910 1400 1300 1000 DDE Brain 2.9 3.4 1.7 6.8 11 11 7.5 230 Whole body 100 140 80 180 300 310 370 200 3-7 " Residues in bird at time of capture from wild. b N u m b e r o f days bird was on clean diet, after being on dosage for I 1 days. Birds were sacrificed on the 56th day. c Bird died during study. MONITORING D D E IN BIRD BLOOD 299 between E D D T in eggs and blood plasma collected from the same female kestrels was reported. More data are now available concerning the residue relationship between eggs and blood plasma for kestrels and the three accipiters (Fig. 3). The regressions for the American kestrel and the accipiters were similar (3' = 6"871X 0 " 9 6 4 , T = +0"805, F/ = 34;.t' = 4"855 X°'978, r = 0'641, n = 34); an F-test of the equality of the two regression models (simultaneous equality of the two intercepts and of the two slopes) was not significant (the test statistic value was F 2.6 4 = 1-67: the 5 °/ critical level is 3.10). Therefore, we believe the pooled /o regression (Fig. 3) is best ( y = 6-243 X 1 ' 0 3 3 , r = +0"782, n = 68). Plasma samples 100 , I I I . . . . I ' ' ' '''"I ' ~= 6.243X t°33 r = +0.782 n= 68 ' ' ''-' ° // • I0 fA // o / • IJJ ¢- - iv I 9// o I.O I...- Q/o i ";a. r"., [/q }= &o?9xl°°° r = +0.818 n= 22 0.1 • 0.1~) ' i i , **,,~ A O.t i A i i Ltiil t.O I i 1 i 1 i i I il I0 ~.DDT in Plasma Fig. 3. The log-log relationship between ZDDT (ppm wet weight) in blood plasma and ZDDT (ppm wet weight) in eggs laid by the same female American kestrel (O), goshawk (11), Cooper's hawk (A) and sharp-shinned hawk ( ~ ) during the post-laying period(regression line solid) and during the pre-laying or immediate pre-laying period ((3) (regression l i n e - - ). 300 C H A R L E S J. H E N N Y , D E N N I S L. MEEKER with less than 0.10ppm Y,DDT were excluded. Also, plasma samples collected before, or within 9 days after, egg laying were regressed separately as a unit (see 'Discussion' section and findings below). The fact that the log-log relationship was independent of the species tested greatly enhances the potential value of the approach for evaluating pesticide burdens in birds of prey since relative contamination among species may be determined directly. The similarity of egg residue estimates based on the three different post-laying regression models is shown in Table 4. Timing of blood sampling: The elimination of pesticides from the bodies of birds through egg laying has been shown in many studies. It has been demonstrated in the TABLE 4 A COMPARISON OF ESTIMATED EGG RESIDUES(PPM) BASEDON PLASMA RESIDUES USING THE POST-LAYING REGRESSION MODEL FOR KESTRELS, ACCIPITERS AND POOLED SPECIES Plasma EDDT (ppm wet weight) 0.10 1.0 5.0 Estimated EDDT (ppm) in egg Kestrel Accipiter Pooled 0.51 4.9 24 0-75 6.9 32 0.58 6-2 33 field by Bogan & Newton (1977) that 52~o of the body burden of DDE was eliminated in a clutch of six eggs laid by a European sparrowhawk. Furthermore, renesting information also suggests that a loss of pesticides occurs during egg laying--for example, in herons Ardea cinerea (Prestt, 1970). This loss is probably most pronounced in species in which the weight of the clutch is a high percentage of the body weight. In this study an American kestrel laid a clutch of eggs (one egg collected and analysed) that contained 2.42 ppm ZDDT; however, its second clutch contained only 0.90ppm. We also have two instances in the field where blood samples were taken before or during egg laying and again after egg laying (Table 5). In both instances residue levels in the plasma declined after egg laying. Kestrel ENI 10 laid all of her eggs after the 21 st of May, whilst kestrel EI-063 had already laid two eggs by the 6th of May (the initial sampling date), but laid two additional eggs later. The decline in plasma residues after egg laying parallels the body burden decline reported by Bogan & Newton (1977). TABLE 5 WILD FEMALE AMERICAN KESTRELS BLED IN THE FIELD BEFORE AND AFTER CLUTCH COMPLETION Sample number ENI-10 E14)63 Date first bled Plasma ZDDT (ppm) Date last bled Plasma 5-DDT (ppm) 13 M a y 1975 6 M a y 1976 0.21 0.14 14 J u l y 1975 22 M a y 1976 0.09 0.08 MONITORING DDE IN BIRD BLOOD 301 Initial concern about combining plasma samples collected prior to, and within a few days after, egg laying with those collected during the post-laying period was based upon our observations showing a loss of EDDT during egg laying, plus the fact that there was increased lipids in the plasma prior to, and immediately after, egg laying (Table 6; Riddle, 1942). Therefore, a separate regression analysis was made TABLE 6 PLASMA LIPID CYCLE OF FEMALE AMERICAN KESTRELS DURING THE NESTING SEASON Per cent lipid Mean SE Range n - 1 1 to - 2 6 2.40 0.40 2.0-3.2 3 Days before ( - ) or after ( + ) clutch completion -lOtoO + l to 10 + l l to 20 +21to40 4. I 0 0.86 2.(~7-2 5 2.10 0-25 0.2-3.9 19 1.76 0-35 0.4-4.8 14 1.32 0.18 0-2-3.2 18 +41to60 1.88 0.37 0.4-4.0 11 with the twenty-two plasma samples collected before and within 9 days of egg laying (Fig. 3). The somewhat arbitrary break between 9 and I 0 days post-laying was made because seventeen of twenty-two predictions of egg residues using the post-laying regression model showed estimates of residues in eggs that were higher than observed. However, by 10-20 days after laying, the post-laying model showed nearly equal numbers of predicted values above and below the observed. The regression model, based upon samples collected before and immediately after egg laying (.~ = 3.079 x l'°°°, r = +0-818, n = 22), parallels the post-laying model with a nearly identical slope (Fig. 3). A t-test was used to test for a significant difference of the two intercepts, assuming a common slope. The intercepts were significantly different (t = 3.46, 87 degrees of freedom, p < 0.001), indicating that post-laying plasma samples should not be pooled with samples collected prior to, or immediately after, egg laying. Nestling bloodsamph, s."Collecting blood samples from nestlings has not provided much useful information (even in the DDT spray area) except to show that residues are always quite low during this growth phase (usually <0.10ppm). The adult segment of the population provides the most useful information and we have abandoned the sampling of nestlings. DISCUSSION A N D CONCLUSIONS The results from these short-term laboratory experiments with kestrels and the field data from the DDT spray area in the Pacific Northwest show that blood plasma can be used to evaluate exposure to Y,DDT and possibly other organochlorine pesticides. With our approach, we believe raptor populations can be monitored by collecting blood plasma samples. The potential to estimate residue levels in eggs laid from plasma samples improves the usefulness of the approach. Our two equations 302 CHARLES J. HENNY, DENNIS L. MEEKER (in Fig. 3) to adjust the plasma residues to estimated egg residues are best used for plasma samples collected at the corresponding times discussed. The longest period after clutch completion that we collected a sample was 78 days. It is also probable that the adjustment equation can be used with some confidence beyond the 2½ months since Friend et al. (1979) for DDE and Robinson (1968) for H E O D have said that stress (from reduced food intake and surgery) does not appear significantly to influence residue levels in blood. However, Friend et al. (1979) did note that DDE residues in sera of mallards fed intermittently (subject to periodic starvation) were slightly higher than in birds fed continuously (mean in sera, 1.94 compared with 1.82), and slightly higher in birds subjected to the stress of monthly surgery than in those not biopsied (mean in sera, 2-19 compared with 1.67). Thus, birds in a stressed situation may show some elevation in plasma residues. For this reason, general body condition should be noted for each bird sampled. Finally, when applying the technique to free-living birds, it should also be recognised that the possibility exists for an inflated plasma residue to occur if the bird recently ate something heavily contaminated. The procedure by which residues in plasma are adjusted to the estimated residues in the egg provides a method of evaluating, the potential impact of the residues detected in the plasma, since the literature for most birds of prey provides information concerning egg residues that impair reproductive performance or cause significant eggshell thinning. We believe the blood plasma approach can be used to group into at least three categories (low, moderate and high) the E D D T burdens in raptorpopulations, not individuals. The ability to predict accurately egg residues for an individual bird is tenuous because of inherent variability and we therefore recommend that conclusions about population residue burdens be based upon as many plasma samples of adults as possible (preferably at least between fifteen and twenty). Also, if plasma is collected a considerable time before or after the nesting period (i.e. migration or wintering period), we suggest using both regression models to obtain separate estimates and presenting these estimated egg residues as a range. Even if little or no information is known about the significance of pesticide burdens in a species, trends may be established by trapping and collecting blood samples at the same location and time over a period of years. The blood plasma approach to environmental studies is viewed as a supplement to existing procedures (primarily egg analysis and production studies) and not as an end in itself. However, the approach has assets that make it particularly appealing for research with selected species those nesting at low densities, in rugged terrain, and in remote areas where logistical problems predominate; those that are secretive or that nest in extremely dense habitat where nest location is extremely difficult. Possibly blood samples from species in these categories could be collected at concentration sites during another time of the year--during migration, or on wintering areas. The approach has particular assets that make it appealing for endangered species research: (1) birds are not killed, (2) multiple samples are MONITORING DDE IN BIRD BLOOD 303 possible over time (it is suggested that all birds sampled be banded for individual recognition) and (3) discrete populations (sometimes few in number) can be sampled periodically in sufficient numbers to establish pesticide exposure patterns and changes in residue levels. ACKNOWLEDGEMENTS The idea of testing the blood plasma technique under field conditions with birds of prey was prompted from the laboratory studies of Milton Friend. Assistance during the laboratory phase of the study was provided by Harold Baer, Merle Richmond, Rick Hudson, Mike Recktenwald and Earl Schafer. A portion of the kestrels for the laboratory study were trapped by Charles Schwartz and Morlan Nelson. Blood samples from the field investigations were provided by the following biological technicians: Roger Olson, Tracy Fleming, Thomas Smith, Craig Campbell, Timothy Craig, Dale Stahlecker, Kevin Moore, Earl Huff, Frank Renn, Charles Brady, Robert Sheehy and Steven Gray. Kenneth Burnham provided assistance with the statistical analysis. We thank Tony Peterle and Lawrence Blus for reading the manuscript and providing valuable constructive suggestions. REFERENCES BOGAN, J. A. & NEWTON, I. (1977). Redistribution of DDE in sparrowhawks during starvation. Bull. environ. Contam. & Toxicol., 18, 317-21. CADE, T. J., WHI'I~, C. M. & HAUGH, J. R. (1968). Peregrines and pesticides in Alaska. Condor, 70, 170- 8. CAPEN, D. E. & LEIKER,T. J. (I 979). DDE residues in blood and other tissues of white-faced ibis. Environ. Pollut., 19, 163-71. DINDAL, D. L. & PE~RLE, T. J. (1968). 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