Effects of vitamin D3 deficiency on estradiol-17 beta metabolism in the laying hen.

Evidence is presented that in vitamin D deficiency there are aberrations of estrogen metabolism in the laying hen. Five White Leghorn hens, 35 weeks of age, all laying soft shelled eggs as a result of complete withdrawal of supplemental vitamin D3 (D3) from the diet, were found to have a 3-fold higher molar ratio of plasma estradiol-17 beta-3-sulfate (E2 beta-3S) to estradiol-17 beta (E2 beta) compared to five control laying hens fed a D3-supplemented diet. A time course tracer study with [3H]E2 beta administered iv revealed an abnormally high accumulation of [3H]E2 beta-3S in plasma, and concomitantly, a markedly reduced formation of [3H]estradiol-17 alpha-3-sulfate [( 3H]E2 alpha-3S) compared with the control hens. This observation suggests a defect in the conversion of [3H]E2 beta-3S to [3H]E1S, previously shown to be a major pathway for E2 beta metabolism (E2 beta E2 beta-3S in equilibrium E1S E2 alpha-3S). As a result, the MCR of [3H]E2 beta from plasma of the D3-deficient hens was about 15% below normal. The calculated production rate of E2 beta was reduced even more, by 60%. However, the reduced production rate could have been related to the lengthening of the ovulatory cycle due to vitamin D deficiency. It is concluded that vitamin D deficiency affects the biosynthesis and catabolism of E2 beta in the laying hen.

Production, clearance rates and metabolic fate of estradiol-17 beta in the plasma of the laying hen.

A single dose of tritiated estradiol-17 beta (3H-E2 beta) was injected i.v. into 5 high egg producing White Leghorn hens, 31 weeks of age, at 19.2 +/- 2.1 (mean +/- S.D.) hr before oviposition. Blood (2 ml) was sampled at approximately 5 min intervals over 40 min. Whenever possible, metabolites were monitored and identified by the double isotope technique with the addition of the corresponding 14C-labelled standards to plasma prior to analysis. The metabolic half-life and clearance rate of 3H-E2 beta in plasma were 10.9 +/- 1.9 min and 118 +/- 18 ml/min/kg body weight, respectively. The calculated production rate of E2 beta at 19.2 hr before oviposition was 19.5 +/- 5.7 ng/min based on the plasma level (93 +/- 22 pg/ml) measured at that time. The relative concentrations (% of plasma radioactivity) of the major metabolites isolated at 5.7 +/- 0.6 min post injection were, in descending order: estradiol-17 beta-3-sulfate (E2 beta-3S: 14.9 +/- 2.7), estradiol-17 alpha-3-sulfate (E2 alpha-3S; 5.7 +/- 0.3), estrone (E1; 4.6 +/- 0.5), estrone sulfate (E1S; 2.2 +/- 0.5), and estradiol-17 alpha (E2 alpha; 1.2 +/- 0.4). As time proceeded, the relative concentration of E2 alpha-3S gradually increased so that by 43.2 +/- 1.0 min it became the most abundant identifiable metabolite (12.3 +/- 1.1) followed by E2 beta-3S (9.1 +/- 1.7), E1S (1.2 +/- 0.6), E1 (0.7 +/- 0.4) and E2 alpha (0.3 +/- 0.2). These findings are consistent with the view that one of the major pathways of E2 beta metabolism in the circulation of the hen is via E2 beta in equilibrium E2 beta-3S in equilibrium E1S in equilibrium E2 alpha-3S.

Studies on genetic and vaccination-induced resistance of chickens to lymphoid tumor transplants 2. Transmissible lymphoid tumor of Olson.

Transmissible lymphoid tumor (TLT) was inoculated in wing webs of five-week-old chickens of 6 strains. About half of the chickens of each strain had been vaccinated with turkey herpesvirus (HVT) one week before challenge in the wing web with TLT. Tumors which developed at the site of inoculation usually reached maximum size within 2 weeks and then regressed. In some chickens, however, tumors developed in visceral organs and caused death in the 2nd through 5th weeks postinoculation. Comparisons among strains of chickens in Expt. 1 revealed no differences in mortality. Vaccination with HVT reduced mortality and also the incidence of wing-web tumors (WWT) in all strains of chickens. A lymphoid leukosis virus and a Marek's disease (MD) virus of low virulence were detected in preparations of TLT, and it is suggested that the immunity induced by vaccination may have been directed against tumor antigens associated with MD virus.

Studies on genetic and vaccination-induced resistance of chickens to lymphoid tumor transplants. 1. Marek's disease tumor transplant (JMV).

Six strains and 4 inbred lines of chickens that differed in susceptibility to Marek's disease (MD) were inoculated in the wing web with JMV when 5 weeks old. Wing web tumors (WWT) developed in all strains and lines inoculated with low-passage JMV (JMV-L) but were largest in Cornell Strain S (highly susceptible to MD). Of 3 strains inoculated with high-passage JMV (JMV-H), only Strain S had appreciable WWT development. Seventy-five percent of the unvaccinated S strain chickens challenged with JMV-L or JMV-H died during the experiments, and approximately half of this mortality occurred during the second week postinoculation. Inbred Line GC ranked next in susceptibility to Strain S and was more susceptible than other lines and strains, including the strain from which it originated. Vaccination with turkey herpesvirus one week before challenge protected against mortality and suppressed WWT development. The effect on WWT development was less, however, in Strains S and NH than in other strains. The transplantability of the tumor was investigated with the use of sex chromosomes as cell markers. Five to 7 days postinoculation of male Strain S chicks with JMV-L or JMV-H, most cells in metaphase from wing web or visceral tumors were of female origin. By 56 days, only male cells were found in visceral tumors. The interpretation was that early lesions were due to tumor transplantation and later lesions were induced by virus.

Comparison of procedures for collecting semen from ganders and inseminating geese.

Experiments were conducted to compare management of ganders and semen collection procedures with respect to semen and sperm yield, and two frequencies of artificial insemination were tested with respect to fertility. Housing ganders in groups, singly, or singly with the introduction of a female just before collection was the rank order of housing system from least to most successful collection of ejaculates. There were no significant differences among types of housing with respect to semen volume, but ejaculates from ganders housed singly had the greatest (P less than .05) spermatozoal concentrations. Ejaculates collected with an artificial vagina were of greater (P less than .05) volume and total spermatozoal yield but not spermatozoal concentration than those collected by aspiration. Interactions between collector and method were observed for spermatozoal traits. Geese inseminated on 2 consecutive days/wk showed greater (P less than .01) fertility than those inseminated once per week. Therefore, collection of semen with an artificial vagina from ganders housed singly, with insemination weekly but on consecutive days, should result in successful reproduction of geese.

Genetic parameters of plasma very low density lipoproteins, abdominal fat lipase, and protein, fatness, and growth traits of broiler chickens.

Genetic parameters of physiological, growth, and fatness traits were investigated in one control and two selected dam strains of broiler chickens. Feed consumption and efficiency were measured between 28 and 42 days of age but were adjusted to estimate values of population average body weights at these ages. Birds were bled at 45 days of age for assay of plasma very low density lipoproteins (VLDL) and killed at 47 days of age for carcass and fat measurements. Abdominal fat was assayed for lipase activity expressed per milligram of protein (LIP/mgP) or per gram of fat (LIP/gF) and protein content of the enzyme preparation expressed as microgram protein/mL (P/mLEPrep). Absolute values of partial correlations corrected for sex and strain were low between production and physiological traits and between fatness and LIP/mgP but were moderate at .3 between fatness and plasma VLDL, LIP/gF, and P/mLEPrep. Heritabilities were moderate to high (greater than or equal to .32) for growth and fatness traits, moderate (.25) for plasma VLDL, and low (less than or equal to .16) for P/LEPrep, LIP/mgP, and LIP/gF. Genetic correlations involving plasma VLDL were as follows: .49 with body weight at 42 days, -.74 with feed consumption, .64 with feed efficiency, .24 with carcass weight at 47 days, 1.07 with abdominal fat weight, and .97 with abdominal fat percentage. Similarly, absolute values of genetic correlations involving P/mLEPrep tended to be as high or higher but genetic correlations involving LIP/mgP and LIP/gF tended to be lower than those involving plasma VLDL.(ABSTRACT TRUNCATED AT 250 WORDS)

Carbonic anhydrase isozymes of blood and oviducal tissues of Leghorn hens.

A group of hens laying high specific gravity eggs and a group laying low specific gravity eggs were selected from each of 6 Single Comb White Leghorn strains and compared for possible qualitative differences in carbonic anhydrase (CA) isozymes in blood and oviducal tissues. Starch gel electrophoretic patterns of erythrocytes, blood plasma and oviducal tissues of magnum, isthmus and uterus from 173 selected hens disclosed 9 carbonic anhydrase isozymes. In oviducal tissues 5 isozyme zones were routinely observed in gel patterns. Of these zones, the 2 with the slowest migration rate had differing staining intensity depending upon the section of the oviduct examined. Five zones were also observed in patterns of lysed erythrocytes, 4 of them possessing the same electrophoretic mobility as 4 to 5 observed in the oviducal tissues, and 1 zone that was observed only in the erythrocytes. Plasma patterns of hens with active ovaries possessed 2 zones, 1 always and 1 occasionally present in patterns of oviducal tissues. Plasma patterns of hens with inactive ovaries possessed 1 or 2 additional unique CA zones. No qualitative differences were found in enzyme patterns between birds laying high specific gravity eggs and those laying low specific gravity eggs.

Esterase isozymes of the hen's oviduct.

Esterase isozymes of magnum, isthmus and uterus of three strains of Single Comb White Leghorn hens were examined by zone electrophoresis on starch gels. Although three regions (I, II and III) of esterase activity were observed, the electrophoretic system was optimized to characterize the pattern of up to five zones of esterase activity that were identified in Region I. These esterases were classified as aliesterases based on reactions in the presence of various substrates and inhibitors. No genetic polymorphisms were observed for these isozymes. However, two of these isozymes were perceived to have an electrophoretic mobility slightly faster in patterns of the magnum of layers than in the isthmus, uterus, and magnum of non-layers. It was shown that egg albumen was present in relatively high quantities in the magnum of layers and that egg albumen, when added to supernatant preparations of isthmus, uterus and magnum of non-layers, caused the faster electrophoretic mobility of these two esterase isozymes. No relation between specific gravity of eggs laid by hens and presence of various Region I esterase isozymes could be detected.

Effects of vitamin D3 deficiency on adenosine triphosphatase activity of jejunums from white Leghorn hens.

Supplemental vitamin D3 (D3) was removed from the diet given to an experimental group of White Leghorn hens, at 227 days of age, while a control group continued to receive a supplemental diet. By 14 days after D3 withdrawal, egg weight, egg specific gravity, shell weight, percent shell, shell thickness, and plasma calcium were lower (P less than .05) in the experimental compared to the control group. At 30 to 37 days after D3 withdrawal, experimental hens had less (P less than .05) jejunal adenosine triphosphatase (ATPase) activity than the control hens. The study indicated that lack of D3 supplementation in laying diets reduced jejunal ATPase activity as well as egg shell quality.

Optimal dietary level of 1 alpha,25-dihydroxycholecalciferol for eggshell quality in laying hens.

The optimal dietary level of 1 alpha,25-dihydroxycholecalciferol [1,25-(OH)2D3] for eggshell quality was established. White Leghorn hens, 59 wk of age, were fed one of eight diets that contained the same basal ingredients, including 3.1% calcium, but different levels (microgram/kg) or forms of calciferol supplements: no calciferol supplement of any form (56 hens); 27.5 (control) or 55.0 micrograms of cholecalciferol (56 hens each); 3, 5, or 7 micrograms of 1,25-(OH)2D3 (28 hens each); 5 micrograms of 24,25-dihydroxycholecalciferol [24,25-(OH)2D3] with 28 hens; 5 micrograms each of 1,25-(OH)2D3 and 24,25-(OH)2D3 (28 hens). All groups were fed the control diet prior to the 21-wk treatment. The group fed 5 micrograms 1,25-(OH)2D3/kg diet ranked first in specific gravity (SG), e.g., 1.081 versus 1.077 for the control group at Week 21 (P less than .05). The group fed 7 micrograms 1,25-(OH)2D3/kg consumed 30% less feed and laid 20% fewer eggs than the control, but shell quality was not affected. The groups receiving no calciferol supplement or receiving only 24,25-(OH)2D3 laid eggs with significantly lower SG than the control after 2 wk of treatment (1.072 or less versus 1.082 at Week 2). The rest of the treatment groups mentioned were comparable to the control in eggshell quality and egg production. Groups fed the combination of 1,25-(OH)2D3 and 24,25-(OH)2D3 per kilogram of feed, or 1,25-(OH)2D3 alone at 5 micrograms/kg, had significantly higher tibial weights relative to the control group. All groups receiving the diets without cholecalciferol supplementation had markedly reduced hatchability. It was concluded that the optimal dietary level of 1,25-(OH)2D3 for improving eggshell quality without affecting egg production was approximately 5 micrograms/kg and the toxic level was 7 micrograms/kg.

Effects of vitamin D3 metabolites on physiological traits of White Leghorn hens.

This study was conducted to determine the effect of feeding vitamin D3(D3) metabolites on BW of hen, weight of uterus, plasma Ca, jejunal and uterine adenosine triphosphatase (ATPase), and carbonic anhydrase. At 416 days of age each of 7 groups of laying hens was fed the basal ration supplemented with one of 7 concentrations (micrograms per kg) of D3 or its metabolites as treatments: 0 micrograms of D3; 27.5 micrograms of D3; 3, 5, or 7 micrograms of 1,25(OH)2D3; 5 micrograms of 24,25(OH)2D3; and 5 micrograms of 24,25(OH)2D3 plus 5 micrograms of 1,25(OH)2D3. Treatment effects were compared at various periods after the start of the study. Hens fed the unsupplemented ration had lower (P less than .05) values for all traits than hens fed the D3-supplemented ration by 162 days after the start of treatment. In a comparison of all dietary treatments except the one involving 0 micrograms D3, from 154 to 161 days after the start of the experiment, treatment effects were significant (P less than or equal to .05) for BW, uterine ATPase, and carbonic anhydrase; hens fed 5 micrograms of 24,25(OH)2D3 per kg of ration ranked the lowest of all treatment groups for these traits. Hens fed 27.5 micrograms of D3 and those fed 5 micrograms of 1,25(OH)2D3 per kg of ration did not differ (P greater than .05) for any traits studied. The results suggest that 5 micrograms of 1,25(OH)2D3 per kg of ration can replace 27.5 micrograms of D3 per kg of ration but that 5 micrograms of 24,25(OH)2D3 per kg of ration tends to have a negative effect on physiological systems of the hen.

Plasma very low density lipoproteins, abdominal fat lipase, and fatness during rearing in two strains of broiler chickens.

Experimental control Strains 30, representative of commercial broiler dam stocks in the late 1970's, and K, representative of commercial broiler stocks of 20 years earlier, were compared. Development of carcass fatness and related traits and physiological traits from 3 to 17 weeks of age were studied. About 12 chickens of mixed sex of each strain were bled and killed at 2-week intervals beginning at 3 weeks of age for measurement of traits. From 3 to 17 weeks of age, percentage abdominal fat of carcass, carcass fat (ether extract) and plasma very low density lipoproteins (VLDL) increased with age. Percentage carcass nitrogen, ash, and water, lipase activity of abdominal fat expressed per mg protein or per g of fat (LIP/g F), and protein concentration of the abdominal fat enzyme preparation (P/ml EPrep) decreased with age. Strain 30 had a higher percentage of abdominal fat and carcass fat, but less carcass nitrogen, ash, and water than Strain K, although there were interactions with sex and age. For physiological traits, Strain 30 had more LIP/g F and less P/ml EPrep than Strain K. Males had lower percentages of abdominal fat, carcass fat, and plasma VLDL, and higher percentages of carcass nitrogen, ash, and water than females. There were no significant differences between sexes for the other physiological traits. There were significant (P less than .01) partial correlations between plasma VLDL and percentage abdominal fat (.22), and between P/ml EPrep and percentages of abdominal fat (-.25), carcass fat (-.29), carcass nitrogen (.30), and carcass water (.30). These observations in conjunction with multiple regression analyses indicated that, in addition to plasma VLDL, protein concentration of adipose tissue expressed as P/ml EPrep might be a useful predictor of fatness in chickens.

Research note: avian leukosis retroviral genes are not detected in geese.

Genomic DNA from four strains of geese was analyzed for the presence of endogenous viral elements using a probe that can detect over 20 Rous-associated endogenous viral genes (ev genes) in chickens, as well as a probe and protocol that detects endogenous avian viruses (EAV). Southern blot analysis of genomic DNA did not reveal any ev genes in DNA of 15 geese from Chinese, Synthetic, or two Embden goose strains. Even under low stringency conditions, using a probe that covered most of the polymerase (pol) gene of the Rous-associated virus (RAV) and that revealed EAV elements in a chicken without ev genes, no viral loci were evident in goose DNA.

Scanning electron microscopy of thin and soft shells induced by feeding calcium-deficient or vitamin D-deficient diets to laying hens.

A scanning electron microscopic study was conducted on shells from eggs laid by four groups of hens maintained on different types of diets: a) control, b) vitamin D3-deficient, c) Ca-deficient, and d) vitamin D3-deficient supplemented with 1,25-(OH)2D3. After 1 week for Ca-deficient hens and after 4 weeks for vitamin D3-deficient hens, the thickness of the shell decreased abruptly and numerous thin-shelled and soft-shelled eggs were laid. The study showed that with both Ca-deficient and vitamin D3-deficient diets, the outer layers of the shell (cuticle and spongy) were reduced or absent but the mammillary layer was present even in the thinnest soft-shelled egg. The order in which layers disappeared as treatment progressed was exactly the reverse of the order in which these layers are formed in normal eggs. No eggs were found without mammillary knobs, which suggests that the hens stop laying before Ca concentrations in blood become too low for the formation of the mammillary knobs. Uncalcified portions of the shell organic matrix were never found, suggesting that Ca deposition and matrix formation were inhibited simultaneously. The relationship between fibers of the shell membrane and mammillary knobs was preserved in all cases. The eggshells from hens on 1,25-(OH)2D3-supplemented diets were ultrastructurally indistinguishable from those of hens on diets adequate in vitamin D3.

Divergent selection for incidence of degenerative myopathy of the Musculus supracoracoideus of meat-type chickens.

The response to three generations of selection for incidence of degenerative myopathy of the Musculus supracoracoideus (DMS) in meat-type chickens was studied. From each of two genetic bases named E and F, which were derived from two commercial broiler stocks, a high incidence, low incidence, and a nonselected control strain were initiated. Selection of breeders to produce Generations 1 and 2 was based on the incidence of DMS in their parents after induced wing exercise; mass selection for plasma creatine kinase levels after induced wing exercise was used to produce Generation 3. Little genetic progress in selection occurred in Generations 1 and 2. There was a response in Generation 3, especially in Base E selected for low incidence. Heritability of incidence of DMS was estimated to be .48. Body weights at 8 weeks of age, 18 weeks of age, and breast angle at 18 weeks of age of Generation 3 were greater (P less than .05) for affected than nonaffected birds and positively correlated phenotypically with incidence of DMS. Genetic correlations were extremely variable. The higher incidence of DMS in birds with desirable traits suggests that the breeder must use a multitrait selection program to reduce incidence of DMS as well as improve production traits.

Response to five generations of selection for blood cholesterol levels in White Leghorns.

The response to five generations of selection for high and low plasma cholesterol levels was examined in two lines of Single Comb White Leghorn chickens derived from the same population. An unselected control line was also maintained. Juvenile cholesterol levels of blood plasma were measured in 24,754 birds during the experiment. Males at 9 to 10 weeks of age had higher plasma cholesterol levels than females of the same age. Estimates of the heritability of plasma cholesterol level for males and females from the sire and dam components of variance in unselected population, ranged from .19 to .30. Realized heritabilities from the selected lines were lower and ranged from .14 to .19 with no significant differences between lines or sexes. Differences between the two selected lines in the fifth generation were 37 mg% for males and 33 mg % for females. High cholesterol levels were associated with high mortality. The estimated genetic correlation between plasma cholesterol and egg production was negative. After five generations of selection for plasma cholesterol, yolk cholesterol was 108 mg % lower in the low selected line than in the controls, but there was little difference between the high selected line and controls in this trait.

Incidence of degenerative myopathy among turkeys fed corn or wheat based rations.

Turkeys were fed either corn based or wheat based rations and necropsied between 28 and 118 weeks of age. Practically all birds affected with degenerative myopathy were detected among those killed between 77 and 85 weeks of age. The incidence of the disease was 0% among 125 males and 7.9% among 139 females. No significant difference in incidence was observed between birds fed corn based and birds fed wheat based rations.

Calcium-binding proteins in serum of chickens: vitellogenin and albumin.

The Ca-binding proteins of hen serum were resolved by gel filtration in 45Ca buffer and were named CaBP(1) and CaBP(2). The major calcium-binding protein, CaBP(1), had a molecular size of 6.0 X 10(5) daltons and appears to be vitellogenin. Deeley et al. (1975) described vitellogenin as the precursor of lipovitellin and phosvitin, the Ca-binding proteins of egg yolk. The present paper demonstrates that "native" vitellogenin is a calcium-binding protein in the serum of laying chickens. The CaBP(2) co-eluted with and was immunologically identical to chicken serum albumin which also bound 45Ca. Data from elution profiles and phosphorus assays indicated that CaBP(1) and CaBP(2) were different from phosvitin and the Ca-binding proteins of the duodenum and uterus.

Calcium-binding proteins in serum: quantitative differences between thick and thin shell lines of chickens.

Calcium binding by two proteins, vitellogenin and albumin, was measured in serum of a line of hens producing thick (THK) and a line of hens producing thin shells (THN) as well as a line of commercial hens using gel filtration in 45calcium buffer. Vitellogenin was quantitated in serum of THK and THN using a radial immunodiffusion test. Levels of serum calcium were measured and related to the above mentioned parameters. The binding of vitellogenin was significantly greater in 13 THK than 13 THN (672 vs. 508 cpm/ml X 10(-3), but binding by albumin was not significantly different between the two lines (386 vs. 344 cpm/ml X 10(-3). Binding in 11 commercial hens was similar to that for THN. The THK had significantly greater levels of diffusible, non-diffusible, and total serum calcium than THN. Significant positive correlations between total serum calcium and vitellogenin binding were found in THK (.77) and THN (.81) as well as in the commercial hens (.64). Like vitellogenin binding, levels of vitellogenin were significantly greater in THK than THN (4.0 vs. 1.8 mg/ml). These results suggest that in addition to having more diffusible and non-diffusible serum calcium, THK have more calcium binding and more vitellogenin to perform the binding function than THN.

Endogenous viral gene distribution in populations of meat-type chickens.

The present study was designed to document the complexity of endogenous viral (ev) genes and seek evidence for their association with production traits in selected and control strains of meat-type chickens. Three populations were studied, each consisting of a control strain and one to three strains selected for various production traits. The ev genes were revealed by digesting genomic DNA with restriction enzymes and detecting DNA fragments on Southern blots using radioactive probes from nucleotide sequences of the avian leukosis virus genome. A total of 31 polymorphic ev loci were identified in these populations from a SacI digest, with an average of 7.3 ev genes per bird. There were no significant differences in ev genes per bird between strains within populations or between selected and control strains overall. Thirty of 62 comparisons in the three populations indicated ev gene frequency differences (P less than .05). Within populations, 13 of 93 comparisons of ev gene frequencies between control and selected strains and 8 of 62 between three selected strains of a sire population showed such differences (P less than .05). Selection for body weight and feed efficiency had been observed to reduce gene frequencies of the slow-feathering gene, which usually contains the ev21 locus; however, these effects were not detected (.05 less than P less than .06) between strains of the dam population in the current study. Such differences suggested possible associations between ev genes and production traits in meat-type chickens.