Identification of glycinin and beta-conglycinin subunits that contribute to the increased protein content of high-protein soybean lines

Seed protein concentration of commercial soybean cultivars calculated on a dry weight basis ranges from approximately 37 to 42% depending on genotype and location. A concerted research effort is ongoing to further increase protein concentration. Several soybean plant introductions (PI) are known to contain greater than 50% protein. These PIs are exploited by breeders to incorporate the high-protein trait into commercial North American cultivars. Currently, limited information is available on the biochemical and genetic mechanisms that regulate high-proteins. In this study, we have carried out proteomic and molecular analysis of seed proteins of LG00-13260 and its parental high-protein lines PI 427138 and BARC-6. Sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis revealed that the high-protein lines accumulated increased amounts of beta-conglycinin and glycinins, when compared with Williams 82. High-resolution two-dimensional electrophoresis utilizing pH 4-7 and pH 6-11 ampholytes enabled improved resolution of soybean seed proteins. A total of 38 protein spots, representing the different subunits of beta-conglycinin and glycinin, were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. High-protein was correlated with an increase in the accumulation of most of the subunits representing beta-conglycinin and glycinin. Comparisons of the amino acid profiles of high-protein soybean lines revealed that the concentration of sulfur amino acids, a reflection of protein quality, was not influenced by the protein concentration. Southern blot analysis showed the presence of genotypic variation at the DNA level between PI 427138 and BARC-6 for the genes encoding group1 glycinin, beta-conglycinin, Bowman-Birk inhibitor (BBI), and the Kunitz trypsin inhibitor (KTI). LG00-13260 inherited the allelic variants of the parental line PI 427138 for glycinin, beta-conglycinin, and KTI, while BBI was inherited from the parental line BARC-6. The results of our study indicate that high-seed protein concentration is attributed to greater accumulation of specific components of beta-conglycinin and glycinin subunits presumably mediated by preferential expression of these genes during seed development.     

Regional seroprevalence of bluetongue virus in cattle in Illinois and western Indiana

OBJECTIVE: To estimate seroprevalence of bluetongue virus (BTV) and the geographic distribution of seropositive cattle herds in Illinois and western Indiana. SAMPLE POPULATION: 10,585 serum samples obtained from cattle in 60 herds during 3 transmission seasons (2000 through 2002). PROCEDURES: In a longitudinal study, serum samples were tested for BTV antibodies by use of a competitive ELISA. Four geographic zones were created by use of mean minimum January temperature. A multivariable mixed-effects logistic regression model with a random effect for herd was used to estimate seropositive risk for zone, age of cattle, herd type, and transmission season. RESULTS: Overall, BTV antibodies were detected in 156 (1.5%) samples. Estimated seroprevalence in 2000, 2001, and 2002 was 1.49%, 0.97%, and 2.18%, respectively. Risk of being seropositive for BTV was associated with geographic zone and age. Seroprevalence increased progressively from northern to southern zones, with no evidence of BTV infection in the northernmost zone. In the southernmost zone, annual seroprevalence ranged from 8.65% to 11.00%. Adult cattle were 2.35 times as likely as juvenile cattle to be seropositive. CONCLUSIONS AND CLINICAL RELEVANCE: Overall seroprevalence was lower than has been reported for Illinois cattle. Bluetongue virus antibodies were distributed heterogeneously in this region. Only in the southernmost zone was seroprevalence consistently > 2%. Regionalization of BTV risk based on state borders does not account for such variability. Serologic data could be combined with landscape, climate, and vector data to develop predictive models of BTV risk within transitional regions of the United States.     

Antitheilerial activity of BW720C (buparvaquone): a comparison with parvaquone

A series of hydroxynaphthoquinones, all derivatives of the antitheilerial hydroxynaphthoquinone parvaquone (993C, Clexon; Wellcome) was tested for antitheilerial activity against Theileria parva (Muguga) in vitro. BW720C (buparvaquone) was 20 times more active than parvaquone. When tested in vivo BW720C cured all 13 cattle infected with T parva and all six infected with T annulata treated at a dose rate of 2.5 mg (kg bodyweight)-1 while parvaquone at 20 mg kg-1 cured nine of 10 cattle. All 16 untreated control cattle died of theileriosis.     

Bartonella species detection in captive, stranded and free-ranging cetaceans

We present prevalence of Bartonella spp. for multiple cohorts of wild and captive cetaceans. One hundred and six cetaceans including 86 bottlenose dolphins (71 free-ranging, 14 captive in a facility with a dolphin experiencing debility of unknown origin, 1 stranded), 11 striped dolphins, 4 harbor porpoises, 3 Risso's dolphins, 1 dwarf sperm whale and 1 pygmy sperm whale (all stranded) were sampled. Whole blood (n = 95 live animals) and tissues (n = 15 freshly dead animals) were screened by PCR (n = 106 animals), PCR of enrichment cultures (n = 50 animals), and subcultures (n = 50 animals). Bartonella spp. were detected from 17 cetaceans, including 12 by direct extraction PCR of blood or tissues, 6 by PCR of enrichment cultures, and 4 by subculture isolation. Bartonella spp. were more commonly detected from the captive (6/14, 43%) than from free-ranging (2/71, 2.8%) bottlenose dolphins, and were commonly detected from the stranded animals (9/21, 43%; 3/11 striped dolphins, 3/4 harbor porpoises, 2/3 Risso's dolphins, 1/1 pygmy sperm whale, 0/1 dwarf sperm whale, 0/1 bottlenose dolphin). Sequencing identified a Bartonella spp. most similar to B. henselae San Antonio 2 in eight cases (4 bottlenose dolphins, 2 striped dolphins, 2 harbor porpoises), B. henselae Houston 1 in three cases (2 Risso's dolphins, 1 harbor porpoise), and untyped in six cases (4 bottlenose dolphins, 1 striped dolphin, 1 pygmy sperm whale). Although disease causation has not been established, Bartonella species were detected more commonly from cetaceans that were overtly debilitated or were cohabiting in captivity with a debilitated animal than from free-ranging animals. The detection of Bartonella spp. from cetaceans may be of pathophysiological concern.     

Regional risk of exporting cattle seropositive for bluetongue virus from the United States

OBJECTIVE: To create a stochastic model to quantify the risk that shipments of cattle from regions within the United States would contain animals seropositive for bluetongue virus and to determine shipment-level accuracy of serologic testing by use of a competitive ELISA (c-ELISA). SAMPLE POPULATION: 19,216 shipments containing 528,918 cattle and calves. PROCEDURE: Data were obtained on number of animals and state of origin of cattle in export shipments originating within the United States between January 1994 and March 2002. Probability distributions for size of export shipments were determined for all states within the United States, and distributions for agar gel immunodiffusion and c-ELISA accuracy (sensitivity and specificity) were determined from expert opinion and review of the literature. The model simulated selection of a shipment and then determined the probability that a threshold number or percentage of cattle within that shipment would have a positive c-ELISA result. Shipment-level sensitivity, specificity, positive-predictive value, and negative-predictive value were calculated. RESULTS: Substantial differences were evident in the regional probability of a shipment being declared positive, with shipments from northeastern states having the lowest probability and shipments from southwestern states having the highest probability. The c-ELISA had variable predictive values at the shipment level, depending on the threshold used and the prevalence of antibody-positive cattle within the region. CONCLUSIONS AND CLINICAL RELEVANCE: Results from this study will aid importers in making scientifically based decisions regarding risk of importing antibody-positive cattle.     

Effect of whole-body vibration in the vertical axis on cortisol and adrenocorticotropic hormone levels in piglets

Vibration, being a consequence of motion during transport, may impair the welfare of pigs. Therefore, the primary objectives of this study were 1) to evaluate during transport simulation the use of ACTH and cortisol plasma levels, which are part of a basic adaptation mechanism of pigs and 2) to define comfort conditions for pigs related to the frequency and acceleration of vibration. Pigs with a body weight between 20 and 25 kg were vibrated in the vertical direction for 2 h at 2, 4, 8, and 18 Hz, in combination with root mean square acceleration magnitudes of 1 or 3 m/s2. Blood was sampled at regular intervals before, during, and after vibration as the pig's behaviors were recorded. Data on ACTH, cortisol, and behavior could be collected from 104 vibrated pigs and 21 controls. In addition, eight animals (3 controls, 5 vibrated) were treated with 0.1 mg of dexamethasone/kg BW, eight animals (3 controls, 8 vibrated) with 0.1 mg naloxone/kg BW, and six (2 controls, 4 vibrated) with a physiological salt solution. Blood samples were taken and products were administrated via an intravenous catheter. The pigs spent less time lying during both hours of vibration treatment than during control conditions. Compared with 2 and 4 Hz, time spent lying was 10 times shorter at 8 Hz and 18 times shorter at 18 Hz. At 1030, ACTH levels were significantly higher than basal levels in animals vibrated at 2 (P < 0.0001), 4 (P < 0.002), and 18 Hz (P < 0.0006). After 1 h, levels returned to basal values. Cortisol levels increased very rapidly after the beginning of vibration (P < 0.0001) and remained higher until 1 h after cessation of vibration (P < 0.003). An inferrence of the lines of equal responses for ACTH and cortisol indicated that, in the beginning of vibration exposure, pigs were extremely susceptible to vibrations at lower frequencies (2 and 4 Hz), whereas at the end of vibration exposure the responses were higher at 18 Hz. The application of dexamethasone and naloxone underpinned the emotional component of the response strategy of pigs to vibration. Hence, vibration during transport should be minimized in order to enhance the adaptive capacities of pigs.