Three kinds of long-term feeding experiment with chicks were conducted in a collaborative studies of 5-year project to compare the effectiveness of the type of experiment for assessing not only nutritive value of a novel feed ingredient but also safety of the ingredient to domestic animals and poultry as well as to human being through their product of the meat and the eggs.
Total 585 hens and 138 cocks were fed one of 8 experimental diets for 12 weeks to get total 1, 160 female and 335 male F₁ chicks, One half of the F₁ chicks were raised for 212 weeks feeding the same diet as their parent. The rest of the chicks was raised similarly but for 36 weeks, at which F₂ chicks were obtained. Hatchability of the eggs of the former were tested 8 times, and contents of heavy metals and 3, 4-benzpyrene in the diets, 6 kinds of single cell protein (SCP), the eggs, the meat, and the liver of the hens were analyzed chemically.
Seven kinds of SCP, i.e., 2 kinds of yeast, I and K, grown on methanol, 2 kinds of bacteria, J and L, grown on methanol, one kind of yeast, M, grown on ethanol, and 2 kinds of brewer's yeast, N and O, were tested.
Except average body weight of the hens at 212 weeks of age fed brewer's yeast O, which was significantly lower than that fed the control diet, no significant difference was observed between the responses of the control hens and of the SCP hens in the laying performance during 212 weeks and in the hatchability tests.
Egg production of the SCP hens was significantly higher than that of the control in spring, when reduced egg production in winter was recovered. Small increase in feed intake was observed in spring, but the increase was not large enough to explain the difference in egg production.
No significant difference in contents of Hg, Pb, Cd, As, and 3, 4-benzpyrene was observed among the diets and the liver of the hens fed the diets. Therefore, the data were combined to estimate the mean, standard deviation and 99% fiducial upper limit of the content. Except small amount of Cd in the meat, none of them was detected in the meat and the eggs of the hens fed the diets.

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Three kinds of long-term multi-generation feeding exteriment with chicks were conducted to compare the effectiveness of the types of experiment for assessing not only nutritive value of a novel feed ingredient but also safety of the ingredient to domestic animals and poultry as well as to human beings through their product, the meat and the eggs. Results of 2 kinds of multi-generation feeding experiment out of 3 were described in this paper. The results of the rest were reported in the previous paper1).
In a long-term multi-generation feeding experiment, total 1, 812 female and 505 male chicks, which were F₁, F₂, and F₃ of the parents reported in the previous paper, were fed one of 8 experimental diets, i.e., a control and 7 diets containing of 7 samples of single cell protein, for 68 weeks with 3 hatchability tests. In a short-term multi-generation feeding experiment, total 3, 873 female and 1, 343 male chicks, which were F₁-F₆ of the same parent, were fed one of the 8 experimental diets for 36 weeks with one hatchability test at the end of the life-span of each generation.
Statistical analyses of variance to examine the main effect of diets, A, and an interaction between diets and generations were carried out in the long- and short-term experiments. Statistical analyses of variance were also carried out to compare the performances of the chicks until 36 weeks of age in the long-term experiment and of the chicks of earlier 3 generations in the short-term experiment, and to compare those of the chicks of earlier 3 generations and of later 3 generations.
The responses of the hens to a certain diet were similar regardless of the length of the life-span of one generation, generation and age of the hens. Trends of larger body weight of the hens fed Diet C and K, and of smaller body weight of those fed Diet O, and trends of larger egg weight of the hens fed Diet C, and of smaller egg size of those fed Diet O were observed.
With these findings reported in this, previous and following papers together, a type of feeding experiment was recommended as a method to check the presence of unknown or unexpected injurious factor or factors in a feed ingredient.

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Comparative studies on horizontal polyacrylamide gradient gel electrophoretograms of the plasma and yolk from laying pulletts and the plasma from cockerels revealed the presence of polymorphic proteins in prealbumin-2 (Pa-2) zone either in plasma or yolk of pullets, and lacking of them in plasma of cockerels. The injection of estradiol at 15mg/kg twice dose induced an occurrence of Pa-2 protein bands in the plasma of 17-week-old immature cockerels and pullets.
Analysis of the parentage records on the phenotypes of 87 dams and 283 daughters produced by the matings of 24 sires supports the hypothesis that the three variants (A, AB and B types) in prealbumin-2 (Pa-2) zone either in plasma or yolk of laying pullets are controlled by one autosomal locus with two codominant alleles, Pa-2^A and Pa-2^B (Table 1). The gene frequency of Pa-2^A in White Leghorn and Rhode Island Red pullets is 0.470 and 0.683, respectively.
The prealbumin-2 (Pa-2) proteins is possibly phosvitins, because of showing a similar mobility reported by other researchers (McBEE and COTTERILL3), and that the same type of the variation is found in plasma and yolk obtained from every mature pullet, also found in estrogenized immature chicken plasma, and lacking of them in cockerels as shown in the present study.

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E. coli strains were isolated from wild birds (Japanese tree sparrows (J.S.), Green pheasants (G.P.) and Bamboo partriges (B.P.)) for examining their drug-resistance and detection of conjugative R plasmids. Total E. coli strains isolated (30 strains were colected from each one) were 1380, 810 and 1530 respectively. And the isolation frequency of resistant strains were 87.5% (J.S.), 12.5% (G.P.) and 15.8% (B.P.). Individuals that excreted resistant strains were 46 out of 63 J.S. (73.0%), 13 out of 41 G.P. (31.7%) and 51 out of 76 B.P. (67.1%). The frequency of R⁺ strains among representative resistant strains were 13.9%, 13.5% and 9.2%, respectively. These results indicate that wild birds may also become the carriers of resistant strains and may be responsible for the spread of R plasmids in a wide area.

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Six independent trials were conducted sequentially with semen from the Japanese Long-Tailed Fowl to determine the optimal pre-freeze and freeze procedures for maximum recovery of spermatozoal motility. Highest levels of pre-freeze motility (4.3±0.2) were obtained when semen was diluted 1:3 with Beltsville Poultry Semen Extenter, cooled for 1.0h at 5°C, then 4% dimethylsulfoxide (DMSO) added in bulk, and the sample equilibrated an additional 1.0h at 5°C. Highest levels of post-thaw motility (3.5±0.0) were obtained when semen was first frozen at 1.0°C/min from +5°C to -20°C, then exposed to liquid nitrogen vapor temperatures between -81° and -154°C for 10mim, before plunging into liquid nitrogen.

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[in Japanese]

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