Preharvest applications of growth regulators and their effect on postharvest quality of table grapes during cold storage

Over 54,600 ha of table grapes (Vitis vinifera), mainly cvs. ‘Thompson Seedless’, ‘Flame Seedless’ and ‘Redglobe’, are planted in Chile. Almost the entire production is exported to the USA, Europe, Asia, or one of several Latin American countries, which typically requires 15–40 d of maritime transportation. During this period, several physical, physiological, and pathological factors cause table grape deterioration. Because berry size is the main quality factor in international markets, farmers often overuse the growth regulators, gibberellic acid (GA₃) and forchlorfenuron (CPPU), in an effort to increase berry size. We examined the effect of preharvest growth regulators on seedless (‘Thompson Seedless’, and ‘Ruby Seedless’) and seeded (‘Redglobe’) table grape cultivars during cold (0 °C) storage plus a shelf life period of 3 d at 20 °C. The overuse of GA₃, eight instead of two GA₃ applications on Thompson Seedless, and the use of one GA₃ application on Redglobe and ‘Ruby Seedless’, increased berry pedicel thickness and lowered cuticle content but induced shatter and predisposed grapes to gray mold caused by Botrytis cinerea. In contrast, CPPU increased berry pedicel thickness and cuticle content but did not increase shatter or gray mold incidence. Clusters that were subjected to overuse of combined GA₃ and CPPU were highly sensitive to shatter, had the thickest pedicel, and developed a high gray mold incidence during cold storage. Hairline, a fine cracking developed during cold storage, was induced on ‘Thompson Seedless’ and ‘Ruby Seedless’ by growth regulators, but no hairline occurred on ‘Redglobe’ table grapes. Therefore, berry quality during cold storage is greatly influenced by growth regulator management in the vineyard.     

Optimizing injection time of GFP plasmid into sheep zygote

Microinjection of exogenous DNA into the cytoplasm of matured oocytes or zygotes is a promising technique to generate transgenic animals. However, the data about the microinjection time and procedure in sheep are limited and have not treated in detail. To obtain more in-depth information, the Sarda sheep oocytes from abattoir-derived ovaries were subjected to IVM and IVF. Then, the GFP plasmid as a reporter gene was injected into the cytoplasm of MII oocytes (n: 95) and zygotes at different post-insemination intervals (6–8 hpi, n: 120; 8–10 hpi, n: 122; 10–12 hpi, n: 110 and 12–14 hpi, n: 96). There were no significant differences in the cleavage rates between the groups. However, blastocyst rate of injected zygotes at all-time intervals was significantly lower than injected MII oocytes and control group (p < 0.05). Interestingly, the proportion of GFP-positive embryos was higher at 8–10 hpi compared with other injected groups (4 % versus 0 %, p  < 0.01). Among these, the proportion of mosaic embryos was high and two of those embryos developed to the blastocyst stage. In conclusion, we settled on the cytoplasmic microinjection of GFP plasmid at 8–10 hpi as an optimized time point for the production of transgenic sheep and subsequent experiments.     

Molecular cloning,purification and immunogenicity of recombinant Brucella abortus 544 malate dehydrogenase protein

The Brucella mdh gene was successfully cloned and expressed in E. coli. The purified recombinant malate dehydrogenase protein (rMDH) was reactive to Brucella-positive bovine serum in the early stage, but not reactive in the middle or late stage, and was reactive to Brucella-positive mouse serum in the late stage, but not in the early or middle stage of infection. In addition, rMDH did not react with Brucella-negative bovine or mouse sera. These results suggest that rMDH has the potential for use as a specific antigen in serological diagnosis for early detection of bovine brucellosis.     

Effects of dietary level of ruminally protected choline on performance and carcass characteristics of finishing beef steers and on growth and serum metabolites in lambs

Ruminally protected choline (RPC) was evaluated in two experiments. In Exp. 1, beef steers (n = 160; average initial BW = 350.9 kg) were fed a 90% concentrate diet with either 0, .25, .5, or 1.0% RPC (DM basis) for 112 to 140 d. Feeding .25% RPC increased ADG 11.6% compared with 0% RPC, but responses diminished with increasing RPC level (cubic response, P < .10). Daily DMI was not affected by RPC level, but feed:gain was improved 6.8% with .25% RPC compared with 0% RPC, and responses diminished with increasing RPC level (cubic response, P < .10). Carcass yield grade increased linearly (P < .10) as RPC level increased, but marbling score was lower for all three RPC-containing diets than for the 0% RPC diet (quadratic response, P < .05). In Exp. 2, 20 Suffolk lambs (initial BW = 29.8 kg) were fed an 80% concentrate diet for 56 d with the same RPC levels as in Exp. 1. Serum triglycerides (TG) and cholesterol (CLSTRL) were measured in weekly blood samples, and intensive blood samples were collected on d 28 and 56 to evaluate serum insulin (INS), GH, and NEFA. For the 56-d feeding period, ADG responded quadratically (P < .10) to RPC level, but DMI and feed:gain were not affected. Serum INS and NEFA concentrations increased linearly (P < .05) and serum GH responded cubically (P < .05) to RPC level on d 28, but no differences were noted on d 56. Serum TG concentrations in weekly samples increased linearly (P < .10) with RPC level, but, averaged over all weeks, serum CLSTRL concentrations did not differ (P > .10) among treatments. Quantities of carcass mesenteric (P < .05) and kidney fat (P < .10) increased linearly, but longissimus muscle and liver fat contents did not differ (P > .10) among RPC levels. Supplementing RPC in high-concentrate diets improved performance, but results were not consistent among RPC levels or between cattle and sheep. Potential effects of RPC might be mediated through alterations in fat metabolism and(or) metabolic hormones related to fat metabolism.     

Influence of grazing dormant native range or winter wheat pasture on subsequent finishing cattle performance, carcass characteristics, and ruminal metabolism

A winter grazing/feedlot performance experiment repeated over 2 yr (Exp. 1) and a metabolism experiment (Exp. 2) were conducted to evaluate effects of grazing dormant native range or irrigated winter wheat pasture on subsequent intake, feedlot performance, carcass characteristics, total-tract digestion of nutrients, and ruminal digesta kinetics in beef cattle. In Exp. 1, 30 (yr 1) or 67 (yr 2) English crossbred steers that had previously grazed native range (n = 38) or winter wheat (n = 59) for approximately 180 d were allotted randomly within previous treatment to feedlot pens (yr 1 native range = three pens [seven steers/pen], winter wheat = two pens [eight steers/pen]; yr 2 native range = three pens [eight steers/pen], winter wheat = four pens [10 or 11 steers/pen]). As expected, winter wheat steers had greater (P < 0.01) ADG while grazing than did native range steers. In contrast, feedlot ADG and gain efficiency were greater (P < 0.02) for native range steers than for winter wheat steers. Hot carcass weight, longissimus muscle area, and marbling score were greater (P < 0.01) for winter wheat steers than for native range steers. In contrast, 12th-rib fat depth (P < 0.64) and yield grade (P < 0.77) did not differ among treatments. In Exp. 2, eight ruminally cannulated steers that had previously grazed winter wheat (n = 4; initial BW = 407 +/- 12 kg) or native range (n = 4; initial BW = 293 +/- 23 kg) were used to determine intake, digesta kinetics, and total-tract digestion while being adapted to a 90% concentrate diet. The adaptation and diets used in Exp. 2 were consistent with those used in Exp. 1 and consisted of 70, 75, 80, and 85% concentrate diets, each fed for 5 d. As was similar for intact steers, restricted growth of cannulated native range steers during the winter grazing phase resulted in greater (P < 0.001) DMI (% of BW) and ADG (P < 0.04) compared with winter wheat steers. In addition, ruminal fill (P < 0.01) and total-tract OM digestibility (P < 0.02) were greater for native range than for winter wheat steers across the adaptation period. Greater digestibility by native range steers early in the finishing period might account for some of the compensatory gain response. Although greater performance was achieved by native range steers in the feedlot, grazing winter wheat before finishing resulted in fewer days on feed, increased hot carcass weight, and improved carcass merit.     

Detection of challenge virus in fetal tissues by nested PCR as a test of the potency of a porcine parvovirus vaccine

To estimate the potency of a porcine parvovirus (PPV) vaccine, three vaccinated and three non-vaccinated pregnant gilts were infected with PPV and the distribution of the virus was studied in the tissues of their 51 fetuses. Virus detection was attempted using haemagglutination (HA) and immunofluorescence (IF) assays, as well as by standard (single) and nested polymerase chain reactions (PCR). None of the detection methods yielded positive results when used to test for the presence of virus in suspensions of organs from the fetuses from the vaccinated gilts. However, the virus was detected in the fetuses from non-vaccinated gilts as follows: HA was positive in 14 cases out of 23 (60.8%), IF in 16/23 (69.5%), standard PCR in 12/20 (60%), and the nested PCR in 19/23 (82.6%). Although the correlation among the results of various methods of virus detection was rather close (r<0.83), the sensitivity of the nested PCR was the highest, both when testing dilutions of PPV and when analysing the fetal organs. The nested PCR therefore provides a reliable approach for studies of virus distribution in fetal organs, with special reference to potency tests on vaccines.     

Autonomic Innervation of the Equine Urinary Bladder

The distribution and density of intrinsic autonomic nerve fibers and cells were studied in the equine urinary bladder by means of the peroxidase-antiperoxidase immunohistochemical method to localize tyrosine-hydroxylase (TH), and by means of a histochemical technique to detect acetylcholinesterase (AChE) activity. The results suggest that the equine urinary bladder, like that of other mammalian species, possesses a rich autonomic innervation which includes catecholaminergic and acetylcholinesterase positive nerves. At least a part of these nerve fibers have an intrinsic origin from ganglion cell bodies within the bladder wall.     

Characterization of culture supernatant proteins from Brucella abortus and its protection effects against murine brucellosis

In this study, we characterized the secreted proteins of Brucella abortus into the enriched media under the bacterial laboratory growth condition and investigated the pathogenic importance of culture supernatant (CS) proteins to B. abortus infection. CS proteins from stationary phase were concentrated and analyzed using 2D electrophoresis. In MALDI TOF/TOF analysis, more than 27 proteins including CuZn SOD, Dps, Tat, OMPs, Adh, LivF, Tuf, SucC, GroEL and DnaK were identified. Cytotoxic effects of CS proteins were found to increase in a dose-dependent manner in RAW 264.7 cells. Upon B. abortus challenge into phagocytes, however, CS proteins pre-treated cells exhibited lower bacterial uptake and intracellular replication compared to untreated cells. Immunization with CS proteins induced a strong humoral and cell mediated immune responses and exhibited significant higher degree of protection against virulence of B. abortus infection compared to mice immunized with Brucella broth protein (BBP). Taken together, these results indicate that B. abortus secreted a number of soluble immunogenic proteins under laboratory culture condition, which can promote antibody production resulted in enhancing host defense against to subsequently bacterial infection. Moreover, further analysis of CS proteins may help to understand the pathogenic mechanism of B. abortus infection and host–pathogen interaction.