Evaluation of a diet free of animal protein in germfree swine.

BACKGROUND: Two experiments were conducted in which germfree pigs or pigs monoassociated with Lactobacillus paracasei subspecies paracasei were fed either a traditional milk-based diet (Esbilac) or an experimental diet free of animal protein (DFAP). METHODS: Throughout the 16-day study, animals' clinical condition, total weight gain, feed conversion, and bacterial contamination were monitored. At the conclusion of the study the animals were killed, necropsied and tissues sampled for L. paracasei isolation. RESULTS: General pig disposition remained consistent between treatment groups and trials, except for two animals that developed mild diarrhoea during trial 1. All pigs remained viable during the study irrespective the diet fed or probiotic inoculation. Germfree pigs fed the Esbilac diet gained on average a total of 1034 +/- 63.0 g, and had a feed conversion ratio of 0.17 +/- 0.01 g of gain per 1 ml of diet. Germfree pigs fed the experimental diet gained on average a total of 599 +/- 151 g, and had a feed conversion ratio of 0.10 +/- 0.02 g of gain per 1 ml of diet. Monoassociated pigs fed the Esbilac diet gained on average a total of 862 +/- 70.3 g, and had a feed conversion ratio 0.14 +/- 0.01 g of gain per 1 ml of diet. Monoassociated pigs fed the experimental diet gained on average a total of 563 +/- 96.8 g, and had a feed conversion ratio of 0.09 +/- 0.02 g of gain per 1 ml of diet. Lactobacillus paracasei established extensively in pigs fed either the Esbilac or experimental diets. Lactobacillus paracasei had no effect (P >0.05) on piglet growth and did not display any interactions based on the diet fed. Measured growth parameters were statistically different (P <0.05) based on the diet fed and variance seen between trials. CONCLUSION: In conclusion, a DFAP has been developed and has been shown to be capable of sustaining life in piglets up to 16 days of age.

Pre-weaning performance and health of pigs born to cloned (fetal cell derived) swine versus non-cloned swine.

The objective of this study was to compare the pre-weaning performance of pigs derived from cloned versus non-cloned parents. Five cloned gilts and one cloned boar were used to produce five litters of pigs. One of five cloned females and the cloned boar were derived from two genetically unmanipulated fetal fibroblast cell lines. The remaining female clones were derived from a fetal fibroblast cell line in which random insertion of a alpha-1,3-galactosyltransferase gene targeting construct had occurred. Fetal cell lines had similar genetic backgrounds and were derived from three different fetuses in three different litters. Five litters of pigs were also generated from matings between two non-cloned boars and five non-cloned gilts. The mean gestation length, mean litter size, mean birth and weaning weights for male and female pigs were similar for litters derived from cloned parents versus non-cloned parents. The proportions of pigs born live and pigs that survived to weaning were also similar for pigs born to cloned as compared to non-cloned parents. In summary, matings between cloned swine derived from fetal fibroblast cell lines yielded litters of pigs that were similar in the number born, piglet birth weight and perinatal and pre-weaning mortality to litters produced by non-cloned swine.

Recurrent selection for corn earworm (Lepidoptera: Noctuidae) resistance in three closely related corn southern synthetics.

Plant resistance offers a good alternative to pesticides for protecting corn (Zea mays L.) from attack by corn earworm, Helicoverpa zea (Boddie). The GT-CEW-RS8 synthetic is a breeding population with some resistance to corn earworm, but based on variability for this trait, further improvement was possible in this population. S1 recurrent selection for corn earworm resistance was conducted to improve three synthetics derived from GT-CEW-RS8. The objective of this research was to study the effectiveness of these selection programs to increase insect resistance and maintain agronomic performance. Cycles of selection for each population were evaluated under four environments. Significant gains in resistance were realized, but all three synthetics derived from GT-CEW-RS8 showed a negative response for yield. However, yield losses were not very dramatic for 66RM4 and inbreeding depression would not affect the yield performance in hybrid combination. Selection for yield done simultaneously with selection for reducing ear damage by corn earworm and husk tightness could assist in avoiding decreased yield.

Prevalence of antibodies to hepatitis E virus in veterinarians working with swine and in normal blood donors in the United States and other countries.

Hepatitis E virus (HEV) is endemic in many developing and some industrialized countries. It has been hypothesized that animals may be the source of infection. The recent identification of swine HEV in U.S. pigs and the demonstration of its ability to infect across species have lent credence to this hypothesis. To assess the potential risk of zoonotic HEV infection, we tested a total of 468 veterinarians working with swine (including 389 U.S. swine veterinarians) and 400 normal U.S. blood donors for immunoglobulin G anti-HEV. Recombinant capsid antigens from a U.S. strain of swine HEV and from a human HEV strain (Sar-55) were each used in an enzyme-linked immunosorbent assay. The anti-HEV prevalence assayed with the swine HEV antigen showed 97% concordance with that obtained with the human HEV antigen (kappa = 92%). Among the 295 swine veterinarians tested from the eight U.S. states (Minnesota, Indiana, Nebraska, Iowa, Illinois, Missouri, North Carolina, and Alabama) from which normal blood donor samples were available, 26% were positive with Sar-55 antigen and 23% were positive with swine HEV antigen. In contrast, 18% of the blood donors from the same eight U.S. states were positive with Sar-55 antigen and 17% were positive with swine HEV antigen. Swine veterinarians in the eight states were 1.51 times more likely when tested with swine HEV antigen (95% confidence interval, 1.03 to 2.20) and 1.46 times more likely when tested with Sar-55 antigen (95% confidence interval, 0.99 to 2.17) to be anti-HEV positive than normal blood donors. We did not find a difference in anti-HEV prevalence between veterinarians who reported having had a needle stick or cut and those who had not or between those who spent more time (> or = 80% of the time) and those who spent less time (< or = 20% of the time) working with pigs. Similarly, we did not find a difference in anti-HEV prevalence according to four job categories (academic, practicing, student, and industry veterinarians). There was a difference in anti-HEV prevalence in both swine veterinarians and blood donors among the eight selected states, with subjects from Minnesota six times more likely to be anti-HEV positive than those from Alabama. Age was not a factor in the observed differences from state to state. Anti-HEV prevalence in swine veterinarians and normal blood donors was age specific and paralleled increasing age. The results suggest that swine veterinarians may be at somewhat higher risk of HEV infection than are normal blood donors.

Conventional resistance of experimental maize lines to corn earworm (Lepidoptera: Noctuidae), fall armyworm (Lepidoptera: Noctuidae), southwestern corn borer (Lepidoptera: Crambidae), and sugarcane borer (Lepidoptera: Crambidae).

Plant resistance is a useful component of integrated pest management for several insects that are economically damaging to maize, Zea mays L. In this study, 15 experimental lines of maize derived from a backcross breeding program were evaluated for resistance to corn earworm, Helicoverpa zea (Boddie); fall armyworm, Spodoptera frugiperda (J. E. Smith); southwestern corn borer, Diatraea grandiosella Dyar; and sugarcane borer, Diatraea saccharalis (F.). Experimental line 100-R-3 was resistant in the field to leaf feeding by fall armyworm and line 116-B-10 was resistant in the field to leaf feeding by fall armyworm and leaf and stalk feeding by southwestern corn borer. When corn earworm larvae were fed field harvested silks from experimental line 81-9-B in the laboratory, their pupal weights were significantly lower than the pupal weights of larvae that were fed silks from the resistant control, Zapalote Chico. Maysin levels lower than those commonly associated with corn earworm resistance were present in the resistant experimental line, 107-8-7, indicating a new basis confers resistance to corn earworm in this line. These resistant experimental lines will provide plant breeders with new sources of resistance to lepidopterous insects for the development of improved maize breeding populations.

Increasing muscle mass in spinal cord injured persons with a functional electrical stimulation exercise program.

OBJECTIVE: To determine the magnitude of changes in muscle mass and lower extremity body composition that could be induced with a regular regimen of functional electrical stimulation (FES)-induced lower-extremity cycling, as well as the distribution of changes in muscle mass among the thigh muscles in persons with spinal cord injury (SCI). STUDY DESIGN: Thirteen men with neurologically complete motor sensory SCI underwent a 3-phase, FES-induced, ergometry exercise program: phase 1, quadriceps strengthening: phase 2, progressive sequential stimulation to achieve a rhythmic pedaling motion (surface electrodes placed over the quadriceps, hamstrings, and gluteal muscles); phase 3, FES-induced cycling for 30 minutes. Participants moved from one phase to the next when they met the objectives for the current phase. MEASURES: Computed tomography of legs to assess muscle cross-sectional area and proportion of muscle and adipose tissue. Scans were done at baseline (before subjects started the program), at first follow-up, typically after 65.4+/-5.6 (SD) weekly sessions, and at second follow-up, typically after 98.1+/-9.1 sessions. RESULTS: Increases in cross-sectional areas were found in the following muscles: rectus femoris (31%, p<.001). sartorius (22%, p<.025), adductor magnus-hamstrings (26%, p<.001), vastus lateralis (39%, p = .001), vastus medialis-intermedius (31%, p = .025). Cross-sectional area of adductor longus and gracilis muscles did not change. The ratio of muscle to adipose tissue increased significantly in thighs and calves. There was no correlation among the total number of exercise sessions and the magnitude of muscle hypertrophy. CONCLUSIONS: Muscle cross-sectional area and the muscle to adipose tissue ratio of the lower extremities increased during a regular regimen of 2.3 FES-induced lower extremity cycling sessions weekly. The distribution of changes was related to the proximity of muscles to the stimulating electrodes.

Evaluation of transgenic sweet corn hybrids expressing CryIA (b) toxin for resistance to corn earworm and fall armyworm (Lepidoptera: Noctuidae).

Many of the lepidopterous insects which attack sweet corn, Zea mays L., are susceptible to insecticidal proteins produced by Bacillus thuringiensis ssp. kurstaki (Berliner) (Btk). Transgenic sweet corn expressing a synthetic cry gene for production of a Btk-insecticidal protein may provide a more environmentally acceptable means of sweet corn production. Eight transgenic sweet corn hybrids containing a synthetic gene for CryIA(b) protein production (BT11 event) were evaluated for resistance to the corn earworm, Helicoverpa zea (Boddie), and fall armyworm, Spodoptera frugiperda (J. E. Smith). Laboratory tests revealed that all Btk sweet corn hybrids were highly resistant to leaf and silk feeding by neonate 3 and 6 d old corn earworm larvae. Ear damage in the field to the Btk sweet corn hybrids caused by corn earworm was negligible. All Btk sweet corn hybrids, except Btk 95-0901, were moderately resistant to leaf and silk feeding by the fall armyworm. Survival and weight gain were reduced when neonates were fed excised whorl leaves of the Btk plants. Weight gain, but not survival, was reduced when 3- and 6-d-old fall armyworm larvae were fed excised whorl leaves of the Btk plants. Btk sweet corn hybrids appear to be ideal candidates for use in integrated pest management (IPM) programs for both the fresh and processing sweet corn markets, and their use should drastically reduce the quantity of insecticides currently used to control these pests in sweet corn. With appropriate cultural practices, it is highly unlikely that Btk sweet corn will contribute to the development of resistance to Btk proteins in these insects because of the high toxicity of the Cry proteins expressed in these sweet corn hybrids and the harvest of sweet corn ears from fields before larvae can complete development.

Genetic mechanisms underlying apimaysin and maysin synthesis and corn earworm antibiosis in maize (Zea mays L.).

C-glycosyl flavones in maize silks confer resistance (i.e., antibiosis) to corn earworm (Helicoverpa zea [Boddie]) larvae and are distinguished by their B-ring substitutions, with maysin and apimaysin being the di- and monohydroxy B-ring forms, respectively. Herein, we examine the genetic mechanisms underlying the synthesis of maysin and apimaysin and the corresponding effects on corn earworm larval growth. Using an F2 population, we found a quantitative trait locus (QTL), rem1, which accounted for 55.3% of the phenotypic variance for maysin, and a QTL, pr1, which explained 64.7% of the phenotypic variance for apimaysin. The maysin QTL did not affect apimaysin synthesis, and the apimaysin QTL did not affect maysin synthesis, suggesting that the synthesis of these closely related compounds occurs independently. The two QTLs, rem1 and pr1, were involved in a significant epistatic interaction for total flavones, suggesting that a ceiling exists governing the total possible amount of C-glycosyl flavone. The maysin and apimaysin QTLs were significant QTLs for corn earworm antibiosis, accounting for 14. 1% (rem1) and 14.7% (pr1) of the phenotypic variation. An additional QTL, represented by umc85 on the short arm of chromosome 6, affected antibiosis (R2 = 15.2%), but did not affect the synthesis of the C-glycosyl flavones.

Quantitative trait loci and metabolic pathways.

The interpretation of quantitative trait locus (QTL) studies is limited by the lack of information on metabolic pathways leading to most economic traits. Inferences about the roles of the underlying genes with a pathway or the nature of their interaction with other loci are generally not possible. An exception is resistance to the corn earworm Helicoverpa zea (Boddie) in maize (Zea mays L.) because of maysin, a C-glycosyl flavone synthesized in silks via a branch of the well characterized flavonoid pathway. Our results using flavone synthesis as a model QTL system indicate: (i) the importance of regulatory loci as QTLs, (ii) the importance of interconnecting biochemical pathways on product levels, (iii) evidence for "channeling" of intermediates, allowing independent synthesis of related compounds, (iv) the utility of QTL analysis in clarifying the role of specific genes in a biochemical pathway, and (v) identification of a previously unknown locus on chromosome 9S affecting flavone level. A greater understanding of the genetic basis of maysin synthesis and associated corn earworm resistance should lead to improved breeding strategies. More broadly, the insights gained in relating a defined genetic and biochemical pathway affecting a quantitative trait should enhance interpretation of the biological basis of variation for other quantitative traits.

High-intensity Raf signal causes cell cycle arrest mediated by p21Cip1.

Activated Raf has been linked to such opposing cellular responses as the induction of DNA synthesis and the inhibition of proliferation. However, it remains unclear how such a switch in signal specificity is regulated. We have addressed this question with a regulatable Raf-androgen receptor fusion protein in murine fibroblasts. We show that Raf can cause a G1-specific cell cycle arrest through induction of p21Cip1. This in turn leads to inhibition of cyclin D- and cyclin E-dependent kinases and an accumulation of hypophosphorylated Rb. Importantly, this behavior can be observed only in response to a strong Raf signal. In contrast, moderate Raf activity induces DNA synthesis and is sufficient to induce cyclin D expression. Therefore, Raf signal specificity can be determined by modulation of signal strength presumably through the induction of distinct protein expression patterns. Similar to induction of Raf, a strong induction of activated Ras via a tetracycline-dependent promoter also causes inhibition of proliferation and p21Cip1 induction at high expression levels. Thus, p21Cip1 plays a key role in determining cellular responses to Ras and Raf signalling. As predicted by this finding we show that Ras and loss of p21 cooperate to confer a proliferative advantage to mouse embryo fibroblasts.

Quantitative trait loci and metabolic pathways: genetic control of the concentration of maysin, a corn earworm resistance factor, in maize silks.

Interpretation of quantitative trait locus (QTL) studies of agronomic traits is limited by lack of knowledge of biochemical pathways leading to trait expression. To more fully elucidate the biological significance of detected QTL, we chose a trait that is the product of a well-characterized pathway, namely the concentration of maysin, a C-glycosyl flavone, in silks of maize, Zea mays L. Maysin is a host-plant resistance factor against the corn earworm, Helicoverpa zea (Boddie). We determined silk maysin concentrations and restriction fragment length polymorphism genotypes at flavonoid pathway loci or linked markers for 285 F2 plants derived from the cross of lines GT114 and GT119. Single-factor analysis of variance indicated that the p1 region on chromosome 1 accounted for 58.0% of the phenotypic variance and showed additive gene action. The p1 locus is a transcription activator for portions of the flavonoid pathway. A second QTL, represented by marker umc 105a near the brown pericarp1 locus on chromosome 9, accounted for 10.8% of the variance. Gene action of this region was dominant for low maysin, but was only expressed in the presence of a functional p1 allele. The model explaining the greatest proportion of phenotypic variance (75.9%) included p1, umc105a, umc166b (chromosome 1), r1 (chromosome 10), and two epistatic interaction terms, p1 x umc105a and p1 x r1. Our results provide evidence that regulatory loci have a central role and that there is a complex interplay among different branches of the flavonoid pathway in the expression of this trait.

Apparent functional independence of the mitochondrial and nuclear transcription systems in cultured human cells.

We have constructed a series of reporter constructs which test the effects of sequence elements from the control region of human mitochondrial DNA on expression in the nucleus, as assayed by transient expression in cultured human cells. The mitochondrial heavy-strand promoter (HSP) was unable to function as a promoter in nuclear DNA. Neither the HSP, nor the binding region for the mitochondrial transcription factor mtTF1 from the light-strand promoter, had any significant or systematic modulatory effects upon transcription from strong or weak RNA polymerase II (pol II) promoters, in three different human cell lines. The same finding held true regardless of orientation with respect to the start site of transcription. Similar results were obtained with a rho 0 derivative of one of these lines, indicating that mitochondrial promoter sequences in the nucleus cannot modulate transcription in response to altered mtDNA copy number. These results support the view that the nuclear and mitochondrial transcription systems in human cells are functionally independent, and do not communicate through factors recognizing shared sequence elements, as suggested by studies in yeast.

Human cord blood contains an IGM antibody to the 41KD flagellar antigen of Borrelia burgdorferi.

Natural antibodies are the IgM products of fetal and neonatal B cells. These are germline encoded low affinity antibodies with multiple specificities to self and exogenous antigens. Lyme borreliosis is the disease resulting from infection with the spirochete, Borrelia burgdorferi. The humoral response to this organism is brisk, directed at multiple proteins, and persistent. Antibody to the 41kd flagellar antigen is found early in disease, but may also be found in non-exposed individuals. These properties suggest that the anti-41kd antibody may be a natural antibody. We report here the finding of an IgM anti-41kd reactivity in 29% of cord blood samples from patients in an area non-endemic for Lyme disease. The results are consistent with the hypothesis that antibody to flagellin may be a germline encoded natural antibody, and could be important in the immunopathogenesis of Lyme arthritis and other arthritides.

Chemical and ultrastructural analysis of corn cuticular lipids and their effect on feeding by fall armyworm larvae.

The cuticular lipid composition of lower and upper leaves of five genotypes of field-grown corn,Zea mays L., was determined by combined gas chromatography-mass spectrometry. Surface lipids of the upper leaves had a higher proportion ofn-alkanes (45-52%) than the lower leaves, while the lower leaves had higher percentages of fatty alcohols (12-18%) than the upper leaves. Scanning electron microscopy showed that the upper leaves of two corn genotypes, MpSWCB-4 and Cacahuacintle X's, had a smooth amorphous appearance, while the lower leaves had a dense array of wax crystals.Spodoptera frugiperda (J.E. Smith) larvae weighed more and developed more rapidly when they were reared on diet containing corn foliage from which the cuticular lipids had been removed than when they were fed untreated foliage. However, growth was not inhibited when larvae were fed diet containing the cuticular lipid extracts or individual cuticular lipid components.

Prenatal diagnosis of thrombocytopenia absent radius syndrome by ultrasound and cordocentesis.

The prenatal diagnosis of thrombocytopenia absent radius (TAR) syndrome, utilizing ultrasound and cordocentesis, is described. To our knowledge, this represents the first prenatal diagnosis of this condition in an index case. The diagnostic evaluation of a fetus with upper extremity limb reduction defects is discussed.

Biochemical activity of centipedegrass against fall armyworm larvae.

Centipedegrass,Eremochloa ophiuroides (Munro) Hack, severely inhibits growth of the fall armyworm larva,Spodoptera frugiperda (J.E. Smith). Fresh centipedegrass extracts and extract fractions were deposited on Celufil, incorporated into meridic-based diets and bioassayed against neonate larvae of the fall armyworm in the laboratory. The methanol extract (F1) caused the greatest reduction in larval weight. When F1, was partitioned between méthylene chloride and water, the activity was transferred to the water-soluble fraction (F5), which, when further fractionated using preparative C-18 reverse-phase chromatography, yielded active F7 and F8 fractions. Gas chromatograph-mass spectrometry and high-pressure liquid chromatography (HPLC) showed F7 to be 95% caffeoylquinic acids with chlorogenic acid as the major constituent. HPLC analysis of F8 revealed maysin [2″-O- α-L-rhamnosyl-6-C-(6-deoxy-Xylo-hexos-4-ulosyl)luteolin] and other luteolin derivatives. Chlorogenic acid and other caffeoylquinic acids, maysin, and other luteolin derivatives are the major factors responsible for the antibiotic resistance of centipedegrass to larvae of the fall armyworm.

Transitory changes of hormones in the plasma of parturient pony mares.

Frequent blood samples were collected from 8 pony mares before, during and after labour, parturition and placental expulsion and assayed for progesterone, oestradiol, androstenedione and LH concentrations by radioimmunoassay. A significant (P less than 0.05) decrease in progesterone, oestradiol and in the progesterone:oestradiol ratio was not detected until 0.5 h after foaling. Androstenedione concentrations rose before and peaked at parturition and then declined. A significant (P less than 0.05) rise in LH was detected 0.5 h after parturition. This LH peak was not detected in one mare and she was the only mare that did not ovulate within the first 20 days post partum. These results suggest that: (1) the foal may be an important factor in the production of progesterone and oestradiol by the feto-placental unit; (2) the pituitary is capable of releasing LH immediately after parturition; (3) the parturient rise in LH may be due to removal of negative feedback inhibition by progesterone and/or oestradiol; and (4) the parturient rise in LH at parturition, combined with already elevated concentrations of FSH, may be involved in the rapid growth of follicles post partum.

Absence of sexual dimorphism in the goat: Induction of luteinizing hormone discharge in the castrated male and female and in the intersex with estradiol benzoate.

Gonadectomized male (n = 5) and female (n = 5) and intact intersex goats (n = 2) were injected i.m. with 50 mug 17(beta)-estradiol benzoate (EB). After treatment, there was a transient 6- to 9-hr decrease in circulating levels of LH followed by a preovulatory-like discharge of LH in all goats. Release peaked at 12 to 18 hr after EB treatment. The magnitude of discharge and the time from treatment until peak of release were not influenced by the goat's sex. These findings suggested that the positive feedback effects of estrogen on LH release were not sexually differentiated in the goat. Since tonic concentrations of LH prior to EB treatment were not different among the groups, the studies also suggested that the intersex goats lacked the inhibitory gonadal influences on gonadotropin release that characterize intact animals.

Identification of a gonadotropin-releasing hormone-like factor in the rabbit fetal placenta.

The biological and immunological gonadotropin-releasing hormone (GnRH)-like activities in rabbit fetal placentas collected at Day 18 of gestation were investigated. Both crude and partially purified acid extracts of placental tissue were tested. A similarly prepared liver extract served as a control. Immunological GnRH-like activity, determined through a GnRH radioimmunoassay was 1.3-2.0 pg/mg protein for the crude placental extract, 7.1-9.2 pg/mg protein for the partially purified placental extract and was nondetectable for liver extract. Both the crude and partially purified placental extracts increased (P less than 0.01) luteinizing hormone (LH) release by dispersed rabbit pituitary cells, whereas the liver extract had no effect. The (Ac-D-p-Cl-Phe1,2, D-Trp3, D-Arg6, D-Ala10)-GnRH antagonist was used to determine whether the biological GnRH-like activity in the placental extract was mediated through GnRH receptors. All three doses of antagonist (10, 100 and 1000 ng) suppressed the biological GnRH-like activity in the placental extracts. Molecular sieve chromatography of the partially purified placental extract showed that the immunoreactive GnRH-like factor eluted in the same fractions as the GnRH standard. These data indicate that the rabbit fetal placenta has both immunological and biological GnRH-like activity.