Co-expression of ovine LPS receptor CD14 with Mannheimia haemolytica leukotoxin receptor LFA-1 or Mac-1 does not enhance leukotoxin-induced cytotoxicity

Leukotoxin (Lkt) and LPS are the major virulence determinants of Mannheimia haemolytica that contribute to the pathogenesis of bovine and ovine pneumonic pasteurellosis. We have previously identified bovine and ovine CD18 as the functional receptor for Lkt. LPS complexes with Lkt resulting in increased thermal stability and enhanced cytotoxic activity of Lkt. Cellular recognition of LPS involves several different molecules including CD14. We hypothesized that expression of ovine CD14 together with LFA-1 or Mac-1 would enhance Lkt-induced cytotoxicity. Ovine cDNA for CD14 was amplified by PCR and cloned into mammalian expression vectors. The 1122 bp cDNAs for bighorn sheep (BHS) and domestic sheep (DS) CD14 encode 373 amino acids which exhibit 99% identity with each other. Ovine CD14 plasmids were transfected either into HEK-293 cells, or previous HEK-293 transfectants stably expressing ovine LFA-1 or Mac-1. Flow cytometric analysis of transfectants confirmed the cell surface expression of CD14. The transfectants expressing LFA-1 or Mac-1 and the transfectants co-expressing CD14 with LFA-1 or Mac-1 did not show any significant difference in Lkt-induced cytotoxicity when incubated with LPS complexed Lkt. In contrast, incubation of the LFA-1 or Mac-1 and LFA-1/CD14 or Mac-1/CD14 transfectants with Lkt which lacks LPS, resulted in reduced cytotoxicity. None of the above transfectants showed any difference in [Ca2+](i) elevation when incubated with both types of Lkt preparations. Lkt did not induce any cytotoxicity or [Ca2+](i) elevation in ovine CD14 transfectants or parent HEK-293 cells. Based on these findings, we conclude that expression of CD14 together with LFA-1 or Mac-1 does not enhance Lkt-induced cytotoxicity, whereas LPS enhances cytotoxicity by complexing with Lkt.     

Characterization of Lasiodiplodia species causing leaf blight,stem rot and fruit rot of fig (Ficus carica) in Malaysia

Fig (Ficus carica) is an exotic deciduous plant that is grown worldwide. Fungal diseases pose a major threat to fig plants, affecting their fruit quality and production. This study was conducted to characterize the fungal isolates associated with leaf blight, stem rot and fruit rot of F. carica in Malaysia through morphological analysis, DNA sequencing, multigene phylogenetic analysis and pathogenicity tests. From September 2018 to March 2019, 30 blighted leaves and 30 rotted stems and fruits of F. carica were collected from several nurseries in Malaysia. Thirty fungal isolates that belonged to Lasiodiplodia theobromae (27 isolates) and L. brasiliensis (three isolates) were identified based on morphological characteristics, comparison of DNA sequences and phylogenetic analysis of the internal transcribed spacer (ITS), elongation translation factor 1-α (tef1-α), β-tubulin (tub2) and DNA-directed RNA polymerase II subunit (rpb2). Among the 27 isolates of L. theobromae, nine isolates were obtained from leaves, eight isolates from stems and 10 isolates from fruits, whereas the three isolates of L. brasiliensis were obtained from stems (two isolates) and a leaf (one isolate). The results of pathogenicity tests revealed that L. theobromae and L. brasiliensis isolates were responsible for leaf blight and stem rot of F. carica, whereas fruit rot was caused by L. theobromae isolates. The present study highlighted two different species, L. theobromae and L. brasiliensis, as the causal agents of leaf blight and stem rot of F. carica. Additionally, L. theobromae caused fruit rot of F. carica in Malaysia.     

Changes In Antioxidant Enzyme Activity In Turfgrass Cultivars Under Various Saline Water Irrigation Levels To Suit Landscapes Under Arid Regions

Turfgrass cover is an inevitable component in the urban landscapes of the United Arab Emirates. Tolerance to abiotic stress like salt, drought, and high temperature is a potential factor to be considered in the selection of turfgrass for the landscapes in the arid regions. Three Seashore paspalum genotypes, four Bermudagrass cultivars along with tall fescue as a control were screened for enzymatic changes under four different saline irrigation levels of 5,000, 10,000, 15,000, and 20,000 mg L^?1 of salinity. Irrigation with potable water served as the control. Turfgrasses were maintained in a field experiment under factorial randomized block design for a period of two years under sustained saline conditions by working out the leaching fraction to maintain the precise level of salinity in each treatment throughout the experimental period. The activities of antioxidant enzymes such as superoxide dismutase (SOD), ascorbate peroxidase (APX), peroxidase (POD), polyphenol oxidase (PPO), and catalase (CAT) were assayed in each of the saline water treated turfgrass types. The SOD activity was found to be high in Yukon (bermudagrass) and Sea Dwarf paspalum. In the paspalum group, Salam and Sea Dwarf showed the highest SOD activity under saline condition. In Bermuda types, Yukon and SR 9554 showed the highest SOD activity compared to other bermuda types. Compared to other cultivars, Sea Isle 2000 (Paspalum type), SR 9554, and Yukon (Bermuda types) exhibited more APX activity without any significant differences among themselves. There was an elevated activity of POD initially, followed by gradual reduction upon increasing the salinity level. Salam cultivar maintained stable POD activity even at the elevated salinity levels. The paspalum types showed relatively high level of PPO activity compared to other types under salinity. Yukon cultivar showed a significantly higher activity of PPO when the salinity level was raised from 15,000 to 20,000 mg L^?1. A significant increase with respect to CAT activity was exhibited in Sea Dwarf under elevated level of salinity. Saline water irrigation brought about a significant effect on the antioxidant enzyme systems to impart oxidative stress tolerance in turfgrass species.     

Viral interference with MHC class I antigen presentation pathway: the battle continues

CD8+ cytotoxic T lymphocytes (CTLs) play a critical role in the defense against viral infections. In general, CD8+ CTLs recognize antigenic peptides in the context of the major histocompatibility complex (MHC) class I molecule. The MHC class I molecules are expressed on almost all the nucleated cells in the body. The trimolecular complex consisting of the class I heavy chain, beta2-microglobulin and the peptide are generated by the MHC class I antigen presentation pathway. This pathway is designed to sample the intracellular milieu and present the information to the CTLs trafficking the area. This rigorous sampling of intracellular environment enables the CTLs to quickly identify and eliminate the cells that synthesize non-self proteins as a result of a viral infection. Many viruses, including several viruses of veterinary importance, have evolved astounding strategies to interfere with the MHC class I antigen presentation pathway, as a means of evading the CTL response of the host. This review focuses on the diverse mechanisms of viral evasion of the MHC class I antigen presentation pathway with particular emphasis on viruses of veterinary importance.     

Immunoblot assays using recombinant antigens for the detection of Mycoplasma hyopneumoniae antibodies

The 36kDa L-lactate dehydrogenase (LDH) and a 29kDa partial fragment of an ABC transporter ATP-binding protein analogue/multidrug resistance protein homologue (PR2) of Mycoplasma hyopneumoniae were tested for their potential as diagnostic antigens. Recombinant LDH was genetically engineered to contain six histidine residues at its C-terminal end, expressed in Escherichia coli and purified to a high degree using Ni(2+)-chelate affinity chromatography. A partial 262 amino acid segment representing the C-terminal end of the PR2 protein was cloned as a glutathione S-transferase (GST) fusion protein, expressed in E. coli and purified by urea extraction. Purified recombinant LDH-6xHis and PR2-GST were then reacted with pig sera in immunoblot assays. Our immunoblots showed that both proteins detected anti-M. hyopneumoniae antibodies in field and experimentally infected pig sera but not in any of the SPF control sera. The two proteins were specific for M. hyopneumoniae as they did not react with sera of pigs infected with the closely related Mycoplasma flocculare and Mycoplasma hyorhinis which are frequently isolated in pigs but are not of particular concern.     

Clays as dietary supplements for swine: A review

Clays are crystalline, hydrated aluminosilicate molecules composed of alkali and alkaline earth cations along with small amounts of various other elements. The best-known are montmorillonite, smectite, illite, kaolinite, biotite and clinoptilolite. The molecules in these clays are arranged in three-dimensional structures creating internal voids and channels capable of trapping a wide variety of molecules. As a result of this structure, clay minerals are regarded as a simple and effective tool for the prevention of the negative effects of many toxic compounds. Dietary supplementation with clays has been shown to improve weight gain and feed conversion in pigs. Where improvements in performance have been noted, one of the most likely explanations for the improvement is the fact clays increase nutrient digestibility. Clays reduce the speed of passage of feed along the digestive tract which allows more time for digestion. Feeding clays also causes morphological changes in the intestinal mucosa such as an increase in villus height and an increase in the villus height to crypt depth ratio. These changes increase the surface area of the gastrointestinal tract thus increasing nutrient digestibility. Several studies have indicated that feeding clay reduces the incidence, severity and duration of diarrhea in pigs. The mechanism for the reduction in diarrhea is likely due to increases in the numbers of Bifidobacteria and Lactobacillus and decreases in Clostridia and E. coli in the small intestine of pigs fed clays. In addition, the numbers of pigs born alive and weaned, birth weight and weaning weight have been shown to be higher for sows fed clays. Several studies have indicated that clays can help mitigate the effects of mycotoxins. The aim of the present review is to focus on the various clays which have been given attention in recent research and to discuss their potential to improve pig performance.     

Role of Bibersteinia trehalosi, respiratory syncytial virus,and parainfluenza-3 virus in bighorn sheep pneumonia

Pneumonic bighorn sheep (BHS) have been found to be culture- and/or sero-positive for Bibersteinia trehalosi, respiratory syncytial virus (RSV), and parainfluenza-3 virus (PI-3). The objective of this study was to determine whether these pathogens can cause fatal pneumonia in BHS. In the first study, two groups of four BHS each were intra-tracheally administered with leukotoxin-positive (Group I) or leukotoxin-negative (Group II) B. trehalosi. All four animals in Group I developed severe pneumonia, and two of them died within 3 days. The other two animals showed severe pneumonic lesions on euthanasia and necropsy. Animals in Group II neither died nor showed gross pneumonic lesions on necropsy, suggesting that leukotoxin-positive, but not leukotoxin-negative, B. trehalosi can cause fatal pneumonia in BHS.     

Incidence and avoidance of zona fracture in cryopreserved bovine ova and embryos

Ova (n=62), which were collected from slaughterhouse bovine ovaries, and embryos (n=26), which were non-surgically recovered from 11 superovulated crossbred donor cows, were frozen. The frozen ova and embryos were then thawed using two conventional thawing protocols, i.e. at 37 °C for 30 seconds in a water bath and at 25 °C for 2 minutes in air. Some 64.5% of the ova and 53.8% of the embryos thawed in the water bath and 16.1% of the ova and 7.7% of the embryos thawed in ambient air exhibited fractured z:onae pellucidae. The slow thawing protocol had a lower incidence of zona damage in cryopreserved oval and embryos than the fast thawing protocol. A low pregnancy rate (12.5%) was recorded for embryos transferred with zona fracture while embryos transferred with intact zonae had a rate of 35.3%) indicating that embryos with zona damage are less viable.     

Incidence and avoidance of zona fracture in cryopreserved bovine ova and embryos

Ova (n=62), which were collected from slaughterhouse bovine ovaries, and embryos (n=26), which were non-surgically recovered from 11 superovulated crossbred donor cows, were frozen. The frozen ova and embryos were then thawed using two conventional thawing protocols, i.e. at 37 degrees C for 30 seconds in a water bath and at 25 degrees C for 2 minutes in air. Some 64.5% of the ova and 53.8% of the embryos thawed in the water bath and 16.1% of the ova and 7.7% of the embryos thawed in ambient air exhibited fractured zonae pellucidae. The slow thawing protocol had a lower incidence of zona damage in cryopreserved oval and embryos than the fast thawing protocol. A low pregnancy rate (12.5%) was recorded for embryos transferred with zona fracture while embryos transferred with intact zonae had a rate of 35.3%) indicating that embryos with zona damage are less viable.