Physiological and morphological changes in bovine mammary glands following intramammary infusion of recombinant interferon-gamma.

Eleven lactating dairy cows were used to evaluate the response of bovine mammary glands to increasing doses of recombinant bovine interferon (rBoIFN)-gamma. Right front and rear quarters were intramammarily infused with eight different doses (10(2) U to 2 x 10(8) U/quarter) of rBoIFN-gamma; each dose was tested in at least two quarters. Left udder halves served as within animal controls in which quarters were injected with a saline placebo or were not infused at all. Milk secretion samples for compositional analysis were collected from each quarter prior to infusion and at 6, 24, 36 and 48 h following infusion. Animals were slaughtered immediately following the 48 h sampling period and mammary tissue was obtained for morphometric analyses. Milk composition was similar between control quarters and those quarters infused with up to 10(5) U of rBo-IFN-gamma during the entire sampling period. Quarters infused with 10(6) U and 10(7) U of rBoIFN-gamma had higher milk somatic cell counts (SCC) following treatment compared with preinfusion values. Changes in the composition of mammary secretion were most dramatic in quarters infused with greater than or equal to 10(8) U of rBoIFN-gamma as indicated by the significant increase in SCC and milk pH with a concomitant decrease in lactose concentration when compared with pre-infusion values or with control quarters. Morphometric analysis of tissue demonstrated an increase in stroma, a decrease in luminal area, and a marked increase in the number of infiltrating leukocytes in those quarters infused with the higher doses of rBoIFN-gamma.(ABSTRACT TRUNCATED AT 250 WORDS)     

Characterization of a wild-type strain of Francisella tularensis isolated from a cat.

Francisella tularensis type A is the primary cause of tularemia in animals and humans in North America. The majority of research on F. tularensis has been done with the attenuated live vaccine strain (LVS), which is a type B, but very few wild-type F. tularensis strains have been characterized. A gram-negative coccobacillus that was isolated in pure culture from the lungs of a cat that died after being lost for 5 days was received for identification at the Virginia-Maryland Regional College of Veterinary Medicine Teaching hospital. The isolate (strain TI0902) was not identified (or was misidentified) by commercial identification systems; however, it was identified as F. tularensis subspecies tularensis (type A) by sequencing a portion of the 16S ribosomal RNA gene. Furthermore, repetitive extragenic palindromic sequences-polymerase chain reaction amplified a 4-kb DNA fragment from TI0902 that was characteristic of F. tularensis type A but not type B. The electrophoretic profile of the lipopolysaccharide of strain TI0902 was identical to that of the LVS by Western blotting with antiserum to LVS. The protein-enriched outer membrane of strain TI0902 contained 6-8 proteins, which were similar in molecular size to those from the LVS. Electron microscopy of negatively stained and alcian blue-stained LVS and TI0902 cells showed that both strains were coccobacillary in shape and may be encapsulated. However, after mouse challenge, the TI0902 strain was clearly more virulent than the LVS strain. Results of this study indicate that the genotype and phenotype of wild-type F. tularensis type A strain TI0902 is similar, but not identical, to that of the LVS strain. Further studies will help determine whether pathogenesis and host-pathogen interactions are also similar between the 2 strains.     

Field‐grown Cotton Exhibits Seasonal Variation in Photosynthetic Heat Tolerance without Exposure to Heat‐stress or Water‐deficit Conditions

Thermotolerance acclimation of photosystem II to heat and drought is well documented, but studies demonstrating developmental impacts on heat tolerance in field‐grown plants are limited. Consequently, climatic variables, estimated canopy temperature, predawn leaf water potential (Ψ_PD), and the temperature responses of maximum quantum yield of photosystem II (F_v/F_m), variable fluorescence (F_v/F₀), quantum yield of electron transport (φ_Eο) and efficiency of PSI electron acceptor reduction (REο/ABS) were characterized for Gossypium hirsutum at three sample times during the growing season (21 June, 2 July and 18 July 2013) under well‐watered conditions. The temperature decreasing a given photosynthetic parameter 15% from the optimum is referred to as T₁₅ and served as a standardized measure of heat tolerance. Ambient and estimated canopy temperatures were well within the optimal range for cotton throughout the sample period, and leaves were verified well watered using Ψ_PD measurements. However, T₁₅ varied with sample date (highest on July 2 for all parameters), being 2 °C (F_v/F₀) to 5.5 °C (φ_Eο) higher on July 2 relative to June 21, despite optimal temperature conditions and predawn leaf water potential on all sample dates. These findings suggest that even under optimum temperature conditions and water availability, heat tolerance could be influenced by plant developmental stage.     

Mechanisms of Reproductive Thermotolerance in Gossypium hirsutum: The Effect of Genotype and Exogenous Calcium Application

Although photosynthetic thermotolerance has been investigated extensively in cotton leaves, reports on the biochemical influence of the pistil in promoting fertilization thermostability are limited. To evaluate the effect of temperature, genotype, and exogenous calcium application on fertilization and pistil biochemistry in cotton, thermosensitive (cv. ST4554 B2RF) and thermotolerant (cv. VH260) plants were grown under control (30/20 °C) or high‐temperature (38/20 °C) conditions during flowering, and exogenous CaCl₂ was applied to flowers 1 day prior to anthesis. Measured pistil parameters included fertilization efficiency; protein concentration; glutathione reductase, superoxide dismutase (SOD) and NADPH oxidase activities; and ATP and calcium levels. Exogenous calcium had no effect on reproductive thermostability due to poor calcium uptake under high temperature. High temperature resulted in a 19.2 % decline in fertilization efficiency for ST4554 but no change in fertilization efficiency for VH260 relative to optimal temperature conditions. Pre‐stress glutathione reductase and SOD activities were higher in VH260 than ST4554 pistils, and calcium and ATP content were higher in VH260 than ST4554 pistils. It was concluded that pre‐stress antioxidant enzyme activity, ATP and calcium content of the pistil may be associated with reproductive thermotolerance in cotton. These findings should be confirmed in future experiments with a larger number of cultivars.     

Seasonal Effect on Zebu Embryo Quality as Determined by Their Degree of Apoptosis and Resistance to Cryopreservation

In order to optimize the production of embryos under tropical conditions and to test a possible seasonal effect on embryo quality, 40 Zebu cows were superovulated during the dry season (April to May) and during the rainy season (July to August). A total of 116 (average 2.7/cow) and 83 embryos (3.5 average/cow) were obtained during the respective seasons. After classification as good, fair or poor quality, embryos were tested based on their ultrastructural differences (n = 53 dry season 16 good, 20 fair and 17 poor and n = 61 rainy season 21 good, 20 fair and 20 poor) and their degree of apoptosis using the TUNEL technique (n = 30 during the dry season and n = 55 in the rainy season). Structural characteristics determining embryo quality varied between good and fair quality embryos. No difference, however, was observed between good, fair and poor quality embryos from the two seasons. The number of TUNEL-positive cells was different among embryos (p < 0.001), being lower in labelled cells of good quality embryos regardless of the season. Fewer apoptotic cells were observed in embryos assigned in all three quality levels during the rainy season (p < 0.001). Ultrastructural evaluations confirmed the results obtained by TUNEL. Cryopreserved embryos of good (n = 25 in each season) and fair quality (n = 11 dry season; n = 17 rainy season) showed a significant decrease of TUNEL-positive cells during the rainy season (p < 0.05). Results suggest that embryos collected in the dry season have more cellular damage in contrast; embryos cryopreserved in the rainy season appeared morphologically better equipped to result in a pregnancy following transfer.     

Natural bovine lentivirus type 1 infection in Holstein dairy cattle. II. Lymphoid tissue lesions

Bovine immunodeficiency virus (BIV) in Holstein cows was associated with morphologic evidence of lymphoid organ deficiency. Cows were subjected to normal management practices including parturition and lactation without adverse environmental stresses. During the clinical disease process there was marked weight loss and wasting with frequent and severe concurrent infections. Lymphoid follicular hyperplasia and dysplasia in lymph nodes, and hypertrophy and hyperplasia in hemal lymph nodes were characteristics of the lymphoid tissues. Atrophy of lymphoid cell compartments with depletion of lymphocytes and a lymphocytic lymphoid folliculitis were components of the lymphoid system pathology. The nodal tissue lesions resembled those observed in feline, simian, and human lentiviral disease. A functional correlation with immune system deficiency was the development of multiple bacterial infections which failed to resolve after appropriate therapy. The BIV-associated disease syndrome in dairy cows may be useful as a model system for investigation of the pathogenesis of the lymphoid organ changes that occur in humans and animals with lentiviral infection.     

Plasma pharmacokinetics of selamectin after a single topical administration in the American bullfrog (Rana catesbeiana).

Parasitism is common in wild and captive amphibians; however, pharmacologic data are lacking for anthelmintic drugs. This study was developed to determine the plasma pharmacokinetics of selamectin after topical administration in bullfrogs. Thirty-two adult American bullfrogs (Rana catesbeiana) were randomly assigned into eight groups of four with each group representing a different collection time point. Seven groups received selamectin (6 mg/ kg) topically and the remaining group served as the untreated control group. One group of frogs was euthanized and blood samples immediately collected on days 0 (control), 1, 5, 10, 15, 20, 25, and 30. Plasma was analyzed for selamectin using high performance liquid chromatography with fluorescence detection. Individual samples were analyzed, then data were reported as the mean of the four frogs at each time point. A histologic evaluation of the lung, liver, kidney, and skin tissues was performed and none of the frogs showed histologic evidence of toxicity due to selamectin administration. The mean peak plasma concentration was 162.5 +/- 42.3 ng/ml, area under the curve was 2,856 ng day/ml, mean residence time was 12.2 days, and disappearance half-life was 1.87 days. Based on the plasma pharmacokinetics, bullfrogs appear to absorb selamectin very efficiently, concentrations reach high levels in the plasma, and there were no apparent histologic effects from single dose administration.     

Assessment of the Intensity of Behavioural Traits and Ovulation between Synchronized and Non-synchronized Cows

Thirty cyclic, non-suckled Brahman cows were divided into three groups, all of which were synchronized sequentially with CIDR-B and observed continuously for 100 h to determine different behavioural oestrus signs. Twenty-four hours after implant withdrawal, all synchronized cows in the group, together with all other cows displaying oestrus, were subjected to intensive ultrasonographic observations (every 6 h for 120 h) to pinpoint the moment of ovulation. In the first group, oestrus and ovulation response was 60% (6/10), in the second 44% (4/9) showed oestrus and six ovulated, and in the third group oestrus and ovulation were 80% (8/10). Significant differences were observed between the second and third groups (p < 0.05). No differences were observed in the duration of oestrus, time when oestrus was displayed after implant withdrawal, time of ovulation and onset of oestrus, end of oestrus to ovulation, and intensity of oestrus on a point scale. The relationship between duration of oestrus and time of ovulation was r(2) = 0.16. Ovulation, on average, was 32.1 +/- 14.5 h after the onset of oestrus, 22.3 +/- 16.5 h after the end of oestrus, and 91.8 +/- 16.7 after implant withdrawal, although no significant differences were observed. One non-synchronized animal showed oestrous activity in the second group but failed to ovulate. In the third group, 8 animals showed oestrus, 4 with high concentrations of progesterone. Of the other four one ovulated. In conclusion, oestrous behaviour is not necessarily the best marker to predict the time when ovulation takes place due to variation in the length of the oestrous period and the possible integration of non-ovulatory animals into sexually active groups.     

Is the Production of Embryos in Small‐Scale Farming an Economically Feasible Enterprise?

The present assay attempts to evaluate the feasibility of using embryo transfer in small community farmers by in vivo study and by modelling the results obtained. From the total of 59 donor cows, 62.7% responded to treatment, with a significant difference (p = 0.002) in the percentage of the response between breeds, being 90.5% (19/21) in Holstein and 47.4% (18/38) in Brahman. A total of 283 embryos were graded as transferable, while 141 as non‐transferable, without difference in the percentage of transferable embryo by breed (p = 0.18). The mean of transferable embryos graded as class I and II was not different between Holstein and Brahman (p = 0.96 and p = 0.92, respectively); besides, no differences were observed in the other grades (non‐transferable). The highest difference in costs, regardless of its quality by breed, was seen in the lower levels of probable fertility of the embryo transferred, even reaching several hundred dollars. When modelling the expected costs for embryo produced and transferred, values can reach nearly $2000.00 when the probable fertility is only 10%. However, when the probable fertility was 60%, embryo cost was close to $300.00. This technology seems to be viable on average or high‐scale systems, having a superovulatory response between 60 and 80% with 4–6 transferrable embryos. Yet, in small‐scale farming, due to the reduced number of donors and/or recipients, the costs surpass the economical feasibility of the technique.     

Relationship of Progesterone,Bovine Pregnancy-Associated Glycoprotein-1 and Nitric Oxide with Late Embryonic and Early Fetal Mortalities in Dairy Cows

The aim of the present study was to determine the relationship of progesterone (P4), bovine pregnancy-associated glycoprotein-1 (bPAG-1) and nitric oxide (NO) levels with late embryonic (LEM; day 28 to day 42) and early fetal mortalities (EFM; > day 42 to day 56) in dairy cows. Transrectal ultrasonography (6–8 MHz) was performed in 100 Holstein-Friesian cows at days 28, 42 and 56 after artificial insemination (AI; day 0) to diagnose pregnancy and to monitor the fate of the embryo. After ultrasound scanning of each cow, a milk sample was collected for assessment of P4 by an ELISA test and a blood sample was collected for assessment of bPAG-1, by using a double-antibody radioimmunoassay, and serum NO metabolites (nitrate + nitrite). Based on ultrasonographic examinations and bPAG-1-RIA, 41 of 100 inseminated cows were confirmed pregnant at day 28 after AI. Nine cows suffered of LEM, and 6 cows suffered of EFM and the overall pregnancy loss rate was 36.6% (15/41) between days 28 and 56 of pregnancy. By logistic regression analysis, there were no significant relationships between the level of P4 and bPAG-1 at day 28 after AI and the occurrence of LEM and EFM. Also, there were no significant relationships between the levels of P4 and bPAG-1 at day 42 and the occurrence of EFM. On the other hand, a significant relationship (P<0.05) was found between NO level at day 28 and the occurrence of LEM. In conclusion, measurement of the serum NO concentration at day 28 of pregnancy might help to predict the outcome of pregnancy by day 42 in dairy cows but further studies are needed to confirm this.