Lead toxicity in chicks: interactions with dietary methionine and choline

Two factorial experiments were conducted to study the effects of dietary methionine and choline on lead toxicity in chicks. Dietary variables were 0.3 or 0.63% (experiment 1) or 0.23 or 0.75% methionine (experiment 2); 0 or 1000 ppm lead (as Pb acetate X 3H2O); and 1130 or 3300 mg/kg (experiment 1) or 396 or 1266 mg/kg choline (experiment 2). In both experiments, lead depressed growth while methionine stimulated growth. Growth depression by lead was less with methionine-adequate than with methionine-inadequate diets. There were no differences in growth with the choline-marginal or choline-excess diets. In experiment 2, the methionine x lead interaction for growth was observed with choline-adequate but not with choline-inadequate diets. Lead-induced depression of growth was exacerbated by added choline when methionine-inadequate diets were fed. With methionine-adequate diets, choline level had no effect on the lead-induced depression of growth. Hepatic nonprotein sulfhydryl (NPSH) concentrations were increased by both supplemental methionine and lead with no interaction. Choline levels had no effect on NPSH. Dietary methionine significantly lowered Pb concentration of kidney and muscle but not of bone, liver or blood. Choline had no effect on organ Pb concentrations. Methionine, either dietary or in the dosing solution, had no effect on in situ intestinal absorption or 203PbCl2. These results suggest that lead lowers the chick's choline requirement and that the methyl moiety of methionine does not participate directly in lead detoxication. The amelioration of Pb toxicity by methionine appears to be related to increased excretion of Pb.     

Dietary interrelationships among arginine,methionine, and lysine in young broiler chicks

Since excess dietary lysine (Lys) can increase the chick's arginine (Arg) requirement and excess Arg can increase the chick's methionine (Met) requirement, experiments were conducted to test the hypothesis that responses to dietary Lys and Met are also interrelated. Day-old Ross x Ross chicks were fed a maize-soyabean meal-based diet supplemented with four levels of L-Arg (0, 5, 10 or 20 g/kg), factorially arranged with four levels of supplemental DL-Met (0, 1, 2 or 3 g/kg). Three replicate pens of ten chicks each were randomly assigned to each treatment and fed for 14 d. An increase in Arg in the diet caused growth and feed-intake depression (P=0.0001), but increasing Met in the diet enhanced growth and feed intake (P=0.0001). Arg toxicity was dependent on the Met level of the diet (Arg x Met interaction; P=0.0153). Experiment 2 was conducted to study interrelationships among Arg, Met, and Lys. Eight treatments were factorially combined: two levels of supplemental L-Arg (0 or 10 g/kg), two levels of supplemental DL-Met (0 or 2 g/kg), and two levels of supplemental L-Lys (0 or 6 g/kg). Six replicate pens of eight chicks per treatment were used. A three-way interaction among Arg, Met, and Lys was observed for body-weight gain and feed intake (P<0.023). As expected, kidney arginase activity increased as dietary Lys increased (P=0.0004). No interactions were found for kidney arginase activity. Muscle creatine increased when chicks were fed the higher Arg (25.2 g/kg) diet (P=0.0047). A three-way interaction among Arg, Met, and Lys was found for muscle creatine (P=0.0075). Excess dietary Lys depressed muscle creatinine concentration, but only in the presence of the lower concentrations of Arg and Met. To conclude, an interrelationship among Arg, Met, and Lys was demonstrated, and it was probably related to creatine biosynthesis.     

The effects of dietary amino acid levels upon lysine and methionine requirements for growing rats

The amino acid requirement has been investigated by many laboratories since 1931. Authors considered that the requirement was influenced by the protein level of the basal diet and also by the experimental method. In light of those earlier studies, the present study investigated the effect on lysine and methionine requirements by modifying total nitrogen level or essential amino acid level. The result of our study confirmed that when the amino acid requirement for maximum growth was expressed as the dietary percentage, amino acid to total amino acid ratio, and amino acid to total nitrogen ratio, the requirement had changed according to not only essential amino acids but also to nonessential amino acids. Furthermore, we investigated the relationship between amino acid intake and body protein gain, and found that 6.2 mg lysine and 9.0 mg methionine were required for the maintenance of a rat weighing 80 g and 71.6 mg of lysine and 48.8 mg of methionine were required for one gram of body protein gain. The lysine and methionine intakes required for one gram of protein gain were equivalent to 100% and 124% of amino acid contents of body protein, respectively.     

Dietary selenium requirement of fingerling channel catfish

Two experiments were conducted in aquaria to determine the minimum dietary selenium requirement of fingerling channel catfish (Ictalurus punctatus). Casein-gelatin diets containing graded levels of supplemental selenium (as Na2SeO3) ranging from 0 to 15 mg/kg were fed to catfish for 15 weeks in experiment 1 to broadly define their selenium requirement and toxicity levels. Although growth of catfish was affected by dietary selenium level, significant differences in weight gain were not easily discernible due to variability among the groups of fish. Weight gain data generally indicated that the basal diet containing 0.06 mg Se/kg diet caused growth depression, and a supplemental selenium level of 15 mg/kg also caused a reduced growth response, which indicated selenium toxicity. Selenium concentrations in edible muscle tissue increased almost linearly with increasing dietary selenium levels. Liver and plasma selenium-dependent glutathione peroxidase (Se GSH-Px) activities indicated the selenium requirement of fingerling channel catfish was between 0.1 and 0.5 mg Se/kg diet. In experiment 2, casein-gelatin diets containing incremental levels of supplemental selenium were fed to catfish for 14 weeks to more precisely determine their minimum dietary selenium requirement. Growth data and liver and plasma Se GSH-Px activities indicated that the minimum selenium requirement of fingerling channel catfish fed adequate vitamin E was 0.25 mg Se/kg dry diet. Based on these data, it appears that selenium supplementation of commercial catfish feeds is warranted.     

Quantitative requirements of the hatchling green sea turtle for lysine, tryptophan, and methionine

The quantitative requirement for the amino acids lysine, tryptophan, and methionine was determined for the hatchling green sea turtle (Chelonia mydas). Hatchling green sea turtles were fed synthetic diets of purified substances with the composition of the diet varying in the amount of lysine, tryptophan or methionine present. The lysine requirement was found to be 4.8% of the crude protein (N X 6.25) or 1.7% of the dry diet. The tryptophan requirement was found to be 0.63% of the crude protein or 0.22% of the dry diet. The methionine requirement, in the presence of adequate cystine (3.1% of the crude protein), was found to be 1.5% of the crude protein or 0.49% of the dry diet.     

Dietary choline requirement of juvenile yellow perch (Perca flavescens)

We conducted an 11-wk feeding trial to determine the dietary choline requirement of juvenile yellow perch (Perca flavescens) and to investigate whether dietary phosphatidylcholine (PC) could meet this requirement. Six dietary treatments contained choline concentrations of <0.11, 0.23, 0.34, 0.75, 1.22 or 3.37 g/kg diet. Two additional diets contained 31 g of lecithin/kg diet, with or without supplemental choline chloride (4.0 g choline/kg diet). The total sulfur amino acid concentration was maintained at 1.0 g/100 g diet (methionine/cyst(e)ine, 49:51). Diets were fed to satiation twice daily to triplicate groups of yellow perch initially weighing 16.0 g/fish. Weight gain, feed intake and carcass proximate composition were significantly affected by dietary choline. Weight gains and feed intakes increased as dietary choline increased from 0 to 0.75 g/kg. Both values tended to plateau in fish fed dietary choline levels above 0.75 g/kg. Broken-line analyses of weight gain and feed intake data indicated the dietary choline requirement was 0.598 and 0.634 g/kg diet, respectively. Hepatic lipid concentrations and feed efficiency values were not significantly different. Whole-body fat concentrations increased significantly, whereas ash levels decreased significantly in fish fed increasing levels of dietary choline. Weight gain and feed intake of fish fed diets containing PC were not significantly different from fish fed 0.75 g/kg of dietary choline. However, hepatic lipid concentrations were significantly higher in fish fed the diet containing PC and no choline chloride. Thus, yellow perch require a maximum of 0.598-0.634 g of choline/kg diet for maximum growth and this requirement may potentially be met with 31 g of lecithin/kg diet.     

Response of rats to diets of equal chemical score: effect of lysine or threonine as the limiting amino acid and of an amino acid excess

The effect of lysine or threonine deficiency with or without excesses of all other amino acids was evaluated in a 21-d feeding study with male rats. Four amino acid mixtures were designed to be first limiting for the rat in lysine or threonine and contained either 0 or 50% excess of nonlimiting amino acids. These mixtures were incorporated into purified diets to provide seven levels [20-140% of the National Research Council (NRC) requirement] of the limiting amino acid. Food intake, body weight gain and carcass composition were measured for each rat to determine the effects of the identity of the limiting amino acid and of amino acid excess on the response to dietary chemical score. Significant effects and/or interactions of the identity of the limiting amino acid (i.e., Lys or Thr) and the presence of excess amino acids were seen for each of the measured responses. At equivalent dietary percentages of the NRC requirement, threonine deficiency supported greater body weight gain than did lysine deficiency. At equivalent deficiencies (Lys vs. Thr) threonine-deficient rats were more susceptible to adverse effects of excess amino acids. When the limiting amino acids were incorporated into the diet through incremental addition of the deficient amino acid mixture, rats responded to levels of lysine or threonine in excess of the NRC requirement. These results suggest that the current NRC requirements for these amino acids are too low and that aspects of the dietary amino acid composition other than the percentage deficit of the limiting amino acid can be important determinants of animal response.     

Antagonism of arginine by excess dietary lysine in the growing dog

Experiments were conducted with growing English Pointer puppies to examine the effects of ingesting excess lysine. A purified crystalline amino acid basal diet containing 0.40% L-arginine (the arginine requirement for maximal weight gain) and 0.91% L-lysine was fed in all assays. All diets were kept isonitrogenous by the addition of diammonium citrate, and lysine was supplied as L-lysine acetate. Both weight gain and gain/feed were reduced in the presence of 4% excess dietary lysine. However, 1 and 2% excess supplemental lysine had no effect on performance. In a second experiment, a growth response to supplemental arginine was obtained in the presence, but not in the absence, of a growth-depressing level of lysine (4%). Therefore, lysine appeared to depress growth by antagonizing arginine. The mechanism of the lysine-arginine antagonism was examined in a third experiment. Classic signs of arginine deficiency: orotic aciduria, depressed urea formation, hyperammonemia, a reduction in weight gain, and emesis were observed in puppies consuming excess lysine but not in their pair-fed controls. Excess lysine ingestion neither inhibited nor induced liver arginase, but it did result in a generalized amino aciduria early in the experiment. In addition, lysine did not appear to affect arginine absorption. Therefore, the mechanism behind the lysine-arginine antagonism in the dog remains to be elucidated.     

Excess dietary methionine markedly increases the vitamin B-6 requirement of young chicks

A soy-protein isolate diet that contained essentially no bioavailable vitamin B-6 was used to establish the quantitative effect of excess dietary methionine on the vitamin B-6 requirement of young chicks. When made adequate in vitamin B-6, chicks fed the basal diet required 2 g/kg supplemental DL-methionine to achieve maximal growth, and 10 g/kg additional DL-methionine (total = 12 g/kg) was found to be a tolerable excess level that would not depress voluntary food intake or growth rate. When chicks were fed seven graded doses of supplemental pyridoxine (PN) in diets that contained either adequate (2 g/kg) or excess (12 g/kg) methionine, the vitamin B-6 requirement for maximal growth was found to increase (P: < 0.01) from 0.73 to 1.05 mg/kg, a 44% increase, when 10 g/kg excess methionine was present in the diet. Indeed, this level of excess dietary methionine depressed (P: < 0.01) growth at all PN dose levels < or =1 mg/kg, but not at PN doses of 1.2 or 1.4 mg/kg. Because dietary intakes of both vitamin B-6 and methionine can affect plasma homocysteine levels, dietary methionine (and protein) intake should be considered important factors in setting safe and adequate requirement levels for vitamin B-6.     

Methionine toxicity in the chick: nutritional and metabolic implications.

Several assays with young chicks fed crystalline amino acid diets were conducted to investigate the effects of supplemental glycine, serine, threonine, arginine, or adenine on the growth depression resulting from consumption of excess methionine. Glycine was partially effective in alleviating the growth depression caused by excess methionine. The addition of threonine together with glycine improved performance still further. Efficiency of food utilization for weight gain was greater in birds fed the methionine-imbalanced diet supplemented with glycine and threonine than in those fed the control diet. Supplemental glycine, threonine, or adenine, but not arginine, was effective in ameliorating the hypoglycemia resulting from consumption of excess methionine. The rate of oxidation of a tracer dose of threonine was increased markedly by feeding 1.25% excess methionine. This was reflected in a 20% depression in threonine utilization for weight gain as measured by slope ratio. The data suggest that both threonine and glycine are antagonized by consumption of excess methionine.     

Influence of dietary zinc on lead toxicity during gestation and lactation in the female rat.

Confirmed pregnant female albino rats received a purified diet containing either 12 or 120 ppm zinc with or without lead (0 to 500 ppm) from day 0 gestation through day 16 of lactation. An 11% reduction in dam weight gain during gestation and a 12% reduction in average pup weight in rats fed the low zinc plus lead diet was markedly different compared to controls not receiving lead and lead supplemented rats receiving supplemental zinc. Milk lead concentration was reduced in rats receiving additional dietary zinc and lead which was reflected in a 15% reduction in pup tibia lead concentration, a 20% reduction in inhibition of pup liver delta-aminolevulinic acid dehydratase activity and a 31% reduction in pup blood porphyrin concentration. Some of the beneficial effects of supplemental zinc on lead toxicity of pups may be explained by a reduction of both liver lead (40%) and blood lead (32%) concentration of dams. However an interaction between lead and zinc at the placental and mammary barriers must also be considered. The results of this study suggest that nutritional status of zinc may be an important factor in modifying lead burden of infants borne by women occupationally and environmentally exposed to lead.     

Essentiality of amino acids for the growing kitten.

The effect of deleting each of the amino acids known to be essential for the young rat was determined in post weanling kittens fed a purified diet containing only L-amino acids as the source of dietary nitrogen. When any one of the 10 amino acids (arginine, lysine, histidine, isoleucine, leucine, methionine, phenylalanine, threonine, tryptophan, valine) were deleted from the diet food intake decreased, the kittens lost weight, and there was a dramatic drop in each corresponding amino acid in the blood plasma; indicating that each of the above amino acids is essential for the kitten. Deletion of all the amino acids except the 10 essential amino acids plus alanine resulted in a decreased weight gain to about 1/3 normal; indicating that although all the other amino acids could be synthesized, one or more of the dispensable amino acids may be required for maximal growth. When any one of the essential amino acids was decreased to one-half that present in the basal diet, there was no decrease in weight gain, indicating that the high protein requirement of the kitten is not the result of an unusually high requirement for the essential amino acids.     

The effect of feeding high levels of low-quality proteins to growing chickens.

1. Three growth trials were done using male broiler chicks. In the first two trials, groundnut meal was used, with and without supplementary methionine and lysine. In the third trial, soya-bean meal was used with and without supplementary methionine. Protein levels ranged in the first trial from 120 to 420 g/kg diet and in the third trial from 120 to 300 g/kg diet. Thus the assumed minimal amino acid requirements of the chick were supplied by high levels of low-quality dietary protein. 2. Diets based on cereals and groundnut meal did not support maximum live-weight gain or maximum efficiency of food utilization at any level of dietary protein. When the principal deficiencies of lysine and methionine were corrected, this protein mixture was capable of supporting the same growth rate as a control diet of cereals and herring meal. 3. Diets based on maize and soya-bean meal did not support quite the same growth rate as similar diets supplemented with methionine, even though the protein level in the unsupplemented diets was sufficient to meet the assumed methionine requirements. 4. These results are interpreted as examples of amino acid imbalance in diets composed of familiar feeding-stuffs. It is concluded that one cannot assume that the poor quality of a protein source can always be offset by increasing the concentration of dietary protein.     

Effect of dietary pH on amino acid utilization and the lysine requirement of fingerling channel catfish.

The pH of amino acid test diets has been shown to be of major importance in dietary amino acid studies in the channel catfish (Ictalurus punctatus). Maximum growth rate and feed conversion was observed when the test diet was adjusted to pH 7. Growth studies, utilizing a 24% crude protein diet containing an amino acid pattern similar to whole egg protein, indicate that the lysine requirement for fingerling channel catfish is about 1.23% of the diet (dry weight basis) or 5.1% of the dietary protein. The dietary requirement was confirmed by serum free lysine analysis. A marked increase in serum free lysine occurred at a dietary lysine level of approximately 1.2% of the diet.     

Choline is required by tilapia when methionine is not in excess

Choline is essential in diets fed to most young vertebrates, but previous studies did not confirm the essentiality of choline in diets fed to tilapia. Two experiments were conducted to evaluate the essentiality of dietary choline in such diets. The basal diet used in both experiments contained 32 g crude protein/100 g diet (10.1 g crude protein from casein and gelatin, and 21.9 g from a crystalline L-amino acid mixture). The total sulfur amino acid (TSAA) concentration of the basal diet was 0.28 g/100 g diet, Met:Cys 89:11. In Experiment 1, a 4x2 design was used in which crystalline L-methionine was added to the basal diet resulting in four levels of TSAA (0.28, 0.50, 0.75 or 1.0 g/100 g diet, Met:Cys 89:11, 94:6, 96:4, or 97:3, respectively). At each level of TSAA, diets also contained either 0 or 1 g choline/kg diet supplied as choline chloride. Weight gain, feed efficiency (FE) and serum methionine concentrations were significantly affected by dietary TSAA concentration, but not by dietary choline concentration or the interaction between TSAA and choline. Weight gain, feed efficiency and serum methionine concentrations indicated that the TSAA requirement was 0.5 g/100 g diet. In the second experiment, diets were formulated to contain either 0.28 or 0.5 g TSAA/100 g diet, Met:Cys 89:11 or 94:6, respectively, and graded levels of choline ranging from 1 to 4 g/kg diet in gradations of 1 g/kg. Dietary methionine significantly affected weight gain and FE, whereas dietary choline significantly affected weight gain, FE and survival, and the interaction of methionine and choline significantly affected weight gain. Fish fed diets containing 0.5 g TSAA/100 g diet and 3 g choline chloride/kg diet exhibited the highest weight gain, feed efficiency and survival. On the basis of these data, it seems clear that juvenile tilapia require choline in certain dietary formulations.     

Influence of dietary methionine on the metabolism of selenomethionine in rats

To determine the influence of methionine on selenomethionine (SeMet) metabolism, weanling male rats were fed for 8 wk a basal diet marginally deficient in sulfur amino acids, containing 2.0 micrograms selenium (Se)/g as DL-SeMet and supplemented with 0, 0.3, 0.6 or 1.2% DL-methionine. Increased dietary methionine caused decreased selenium deposition in all tissues examined but increased glutathione peroxidase (GSHPx, EC 1.11.1.9) activity in testes, liver and lungs. A positive correlation was found between dietary methionine and the calculated percentage of selenium associated with GSHPx. In a second experiment, 75SeMet was injected into weanling male rats which had been fed the basal diet containing 2.0 micrograms selenium as DL-SeMet with or without the addition of 1.0% methionine. The selenoamino acid content of tissues and the distribution of 75Se in erythrocyte proteins were determined. In comparison to the rats fed the basal diet without added methionine, significantly more 75Se-selenocysteine was found in liver and muscle, more 75Se was found in erythrocyte GSHPx and less 75Se was found in erythrocyte hemoglobin of rats fed 1.0% methionine. These data suggest that methionine diverts SeMet from incorporation into general proteins and enhances its conversion to selenocysteine for specific selenium-requiring proteins, such as GSHPx.     

Dietary sulfur amino acid requirement of juvenile yellow perch fed the maximum cystine replacement value for methionine

We conducted three separate experiments designed to determine the dietary methionine requirement, ability of cyst(e)ine to spare methionine, and the total sulfur amino acid requirement (TSAA) of juvenile yellow perch when fed the maximal amount of cyst(e)ine. The purified basal diet used in each experiment contained 33.6 g of crude protein/100 g diet and 12.0 g of lipid/100 g diet. In the first experiment,;>L-methionine was added to eight diets providing methionine concentrations ranging from 0.37 to 1.77 g/100 g diet in gradations of 0.2 g/100 g diet. Diets were fed for 12 wk to juvenile yellow perch initially weighing 4.7 g/fish. Broken-line analyses of weight gain and feed efficiency data indicated that the dietary methionine requirement was 1.0 g/100 g diet (3.1 g TSAA/100 g dietary protein) and 1.1 g/100 g diet (3.4 g TSAA/100 g dietary protein), respectively. In the second experiment, various ratios of L-cyst(e)ine and L-methionine were added to the basal diet and fed for 12 wk to determine the cyst(e)ine replacement value of yellow perch initially weighing 19.3 g/fish. Weight gain and feed efficiency (FE) data indicated that cyst(e)ine spared up to 51% of the methionine requirement. In the final experiment, graded levels of cyst(e)ine plus methionine in a ratio of 51:49 were added to the basal diet in gradations of 0.1 g/100 g diet (0.5 to 1.2 g TSAA/100 g diet) to determine the dietary total sulfur amino acid requirement. Diets were fed to satiation for 10 wk to fish initially weighing 8. 1 g. Broken-line analyses of weight gain, feed intake and FE data indicated that the dietary TSAA requirement was 0.85, 0.87 and 1.0 g of TSAA/100 g diet (2.5 to 3.0 g of TSAA/100 g of dietary protein), respectively. The majority of dietary TSAA requirements of fish are in the range of 2 to 4 g/100 g of dietary protein and are generally similar to those of both birds and swine, but lower than estimates for rodents.     

米-黄豆饲料添补蛋氨酸对大鼠生长效应的影响

以80%的米与6%黄豆供应约10%蛋白质的饲料,由微生物法分析,其必需氨基酸百分含量如下:精氨酸0.61,组氨酸0.23,异亮氨酸0.53, 亮氨酸0.70,赖氨酸0.29,蛋氨酸0.09,苯丙氨酸0.47,苏氨酸0.36,色氨酸0.11,缬氨酸0.47。 以此基本饲料饲养大鼠,两周平均增加体重29.0g,按Rose推荐的大鼠需要量计算,第一限制氨基酸为蛋氨酸,但添补0.3,0.6,0.9,1.8%DL-蛋氨酸,对大鼠生长均无良好影响。添加1.8%DL-蛋氨酸组体重由对照组的29.0g降至7.0g。 加入1.5%DL-蛋氨酸至基本饲料中引起大鼠生长抑制,添补次限制氨基酸赖氨酸至接近Rose需要量时有部分克服作用,但添补组氨酸、 吡哆醇却未发现相似效应。 过量蛋氨酸和其他各实验组对肝重量的影响与体重有关,各组肝氮、肝脏含水量、血液血红蛋白含量均未见显著性变化。     

Methionine requirement of the growing kitten, in the absence of dietary cystine

This study was conducted to determine the methionine requirement for maximal weight gain and nitrogen retention in growing kittens fed a 24% crystalline amino acid diet containing no cystine. Six male and six female kittens wee fed each of six levels of methionine, 0.45, 0.7, 0.8, 0.9, 1.0 and 1.5% of the diet, for 10 days in a 6 x 6 Latin square design. Diets were made isonitrogenous with alanine and were isocaloric with a calculated metabolizable energy of approximately 5.0 kcal/g. Food intake and body weights were recorded daily, and nitrogen balance for each cat in each 10-day period was determined. On the 7th day of each period, blood was drawn for subsequent analysis of plasma methionine concentrations. From growth response results, the methionine requirement in the absence of dietary cystine was estimated to be 0.07% of the diet for male and female kittens. On the basis of nitrogen balance results, the methionine requirement appeared to be slightly higher, at 0.75% of the diet for both sexes. Plasma methionine concentrations appear to increase exponentially with dietary methionine concentrations, and wee of limited usefulness in estimating the methionine requirement of the kitten in the context of the experimental protocol used here.     

Arginine requirement and apparent absence of a lysine-arginine antagonist in fingerling channel catfish

A series of growth studies, utilizing casein-gelatin based diets supplemented with crystalline amino acids, were conducted to determine the arginine requirement for fingerling channel catfish (Ictalurus punctatus) and to evaluate the effects of excessive levels of dietary lysine and arginine. Weight gain and feed efficiency data indicate the arginine requirement to be 1.03 +/- 0.07% and 1.00 +/- 0.06% of the dry diet, respectively. Based on growth this corresponds to 4.29% of the dietary protein. There was no evidence of an arginine-lysine antagonism when excess lysine was fed in diets adequate or marginal in arginine. Similarly, growth and feed efficiency data suggest the lack of an antagonism when excess arginine is added to diets marginal in lysine. Apparently channel catfish are not as sensitive to disproportionate lysine and arginine levels as are other animals.