Nasir Rajput

Effect of multiple freeze-thaw cycles on the quality of chicken breast meat.

The objective of this study was to determine the effects of repeated freeze-thaw cycles (0-6) on any physico-chemical changes and lipid and protein oxidation in chicken breast. The results showed that meat colour, a(∗) (redness) and b(∗) (yellowness) values decreased while L(∗) values (lightness) increased with increasing cycle numbers. Increasing freeze-thaw cycles resulted in a greater degree of lipid and protein oxidation, as evidenced by higher contents of malondialdehyde and carbonyl compounds, and lower contents of sulfhydryl groups. Differential scanning calorimetry profiles and SDS-PAGE banding patterns of myofibrillar proteins indicated slight denaturation of myosin and actin with repeated freeze-thaw cycles. The structural changes occurring in proteins caused by oxidation directly affected the ability of muscles to retain water, as confirmed by the nuclear magnetic resonance relaxometery profile. In conclusion, multiple freeze-thaw cycles increased lipid and protein oxidation and decreased water holding capacity and colour stability of broiler chicken breast.

Phytophthora sojae Effector PsCRN70 Suppresses Plant Defenses in Nicotiana benthamiana

Phytophthora sojae, an oomycete pathogen, produces a large number of effector proteins that enter into host cells. The Crinklers (Crinkling and Necrosis, CRN) are cytoplasmic effectors that are conserved in oomycete pathogens and their encoding genes are highly expressed at the infective stages in P. sojae. However, their roles in pathogenesis are largely unknown. Here, we functionally characterized an effector PsCRN70 by transiently and stably overexpressing it in Nicotiana benthamiana. We demonstrated that PsCRN70 was localized to the plant cell nucleus and suppressed cell death elicited by all the tested cell death-inducing proteins, including BAX, PsAvh241, PsCRN63, PsojNIP and R3a/Avr3a. Overexpression of the PsCRN70 gene in N. benthamiana enhanced susceptibility to P. parasitica. The H2O2 accumulation in the PsCRN70-transgenic plants was reduced compared to the GFP-lines. The transcriptional levels of the defense-associated genes, including PR1b, PR2b, ERF1 and LOX, were also down-regulated in the PsCRN70-transgenic lines. Our results suggest that PsCRN70 may function as a universal suppressor of the cell death induced by many elicitors, the host H2O2 accumulation and the expression of defense-associated genes, and therefore promotes pathogen infection.

A Virulence Essential CRN Effector of Phytophthora capsici Suppresses Host Defense and Induces Cell Death in Plant Nucleus

Phytophthora capsici is a soil-borne plant pathogen with a wide range of hosts. The pathogen secretes a large array of effectors during infection of host plants, including Crinkler (CRN) effectors. However, it remains largely unknown on the roles of these effectors in virulence especially in P. capsici. In this study, we identified a cell death-inducing CRN effector PcCRN4 using agroinfiltration approach. Transient expression of PcCRN4 gene induced cell death in N. benthamiana, N. tabacum and Solanum lycopersicum. Overexpression of the gene in N. benthamiana enhanced susceptibility to P. capsici. Subcellular localization results showed that PcCRN4 localized to the plant nucleus, and the localization was required for both of its cell death-inducing activity and virulent function. Silencing PcCRN4 gene in P. capsici significantly reduced pathogen virulence. The expression of the pathogenesis-related gene PR1b in N. benthamiana was significantly induced when plants were inoculated with PcCRN4-silenced P. capsici transformant compared to the wilt-type. Callose deposits were also abundant at sites inoculated with PcCRN4-silenced transformant, indicating that silencing of PcCRN4 in P. capsici reduced the ability of the pathogen to suppress plant defenses. Transcriptions of cell death-related genes were affected when PcCRN4-silenced line were inoculated on Arabidopsis thaliana, suggesting that PcCRN4 may induce cell death by manipulating cell death-related genes. Overall, our results demonstrate that PcCRN4 is a virulence essential effector and it needs target to the plant nucleus to suppress plant immune responses.

A Phytophthora sojae cytoplasmic effector mediates disease resistance and abiotic stress tolerance in Nicotiana benthamiana

Each oomycete pathogen encodes a large number of effectors. Some effectors can be used in crop disease resistance breeding, such as to accelerate R gene cloning and utilisation. Since cytoplasmic effectors may cause acute physiological changes in host cells at very low concentrations, we assume that some of these effectors can serve as functional genes for transgenic plants. Here, we generated transgenic Nicotiana benthamiana plants that express a Phytophthora sojae CRN (crinkling and necrosis) effector, PsCRN115. We showed that its expression did not significantly affect the growth and development of N. benthamiana, but significantly improved disease resistance and tolerance to salt and drought stresses. Furthermore, we found that expression of heat-shock-protein and cytochrome-P450 encoding genes were unregulated in PsCRN115-transgenic N. benthamiana based on digital gene expression profiling analyses, suggesting the increased plant defence may be achieved by upregulation of these stress-related genes in transgenic plants. Thus, PsCRN115 may be used to improve plant tolerance to biotic and abiotic stresses.

Effect of dietary supplementation of marigold pigment on immunity, skin and meat color, and growth performance of broiler chickens

Marigold flower extract, a natural pigment, was used to determine its effect on carcass and skin pigmentation, immunity and growth performance of broiler chickens. Two hundred and forty 1-day-old Arbor Acres broiler chicks were randomly distributed into four treatment groups with six replicates in a randomized block design. Birds were fed basal diet for 42 d with or without supplementation of marigold flower extract at various concentrations, i.e., 0 (MG0, control), 100 (MG100), 150 (MG150) and 200 (MG200) mg/kg of feed, respectively. Feed intake and live body weight were weekly recorded. Carcass and shank color, and antibody titers against Newcastle and Influenza viruses were measured. Results showed that marigold flower extract significantly (p<0.05) improved live body weight and relative thymus weight. However, feed intake, feed conversion ratio (FCR), and spleen and bursa weights were not significantly affected. Yellowness (b*) of breast and thigh muscles increased by the dietary supplementation of marigold flower extract compared with the control diet. However, lightness (L*), redness (a*) and redness to yellowness ratio (a/b) were not influenced by the treatments. Moreover, Roche color fan scores of the shank skin were increased at market age (d 42). The results revealed that marigold extract enhanced antibody titers against Newcastle and influenza viruses. It was possible to conclude that the dietary supplementation with marigold flower extract at the rate of 200 mg/kg of feed enhanced carcass and shank color, antibody titers against ND and AI, and growth performance of broiler chickens.

The effect of dietary supplementation with the natural carotenoids curcumin and lutein on pigmentation, oxidative stability and quality of meat from broiler chickens affected by a coccidiosis challenge.

Abstract 1. An experiment was performed to evaluate the effectiveness of the antioxidants curcumin (CRM) and lutein (LTN) on the quality of meat from coccidiosis-infected broilers. A total of 200 one-day-old Arbor Acre chicks were randomly assigned to a treatment group with 5 replicates. The treatments included a basal diet without carotenoid supplementation (control), with 300 mg/kg CRM, with 300 mg/kg LTN or with a combination (C + L) of 150 mg/kg CRM and 150 mg/kg LTN. All chickens were challenged with Eimeria maxima at 21 d old. 2. The results revealed that the coccidiosis reduced redness of meat, while supplementation with carotenoids improved the fresh meat’s redness (a*) and yellowness (b*) and contributed to colour stability maintenance after storage (1 month at −18°C and 3 d at 4°C). 3. Coccidiosis did not produce lipid and protein oxidation in fresh meat, but after storage for one month, the malondialdehyde levels and carbonyl contents were lower in the CRM and C + L birds and the sulfhydryl contents were higher in C + L birds. 4. The sodium dodecyl sulphate–polyacrylamide gel electrophoresis banding pattern showed equivalent myosin chain fragmentations in all treatment groups, whereas lower intensity actin bands were observed in the control group (CONT). Moreover, myofibril protein denaturation (differential scanning calorimetry) profiles showed a reduction in the CONT myosin and actin peaks. Coccidiosis reduced the meat’s water holding capacity in non-supplemented chicken meat and was improved by natural carotenoid. 5. These results emphasise that coccidiosis did not decrease the eating quality of fresh meat, that natural carotenoids are efficient antioxidants and that CRM (300 mg/kg) fed individually or combined with LTN was the most effective supplemented antioxidant compound.

Construction of an immobilised acetylcholinesterase column and its application in screening insecticidal constituents from Magnolia officinalis

BACKGROUND Application of a matrix-immobilised target enzyme for screening inhibitors is widely used in drug development, but there are few studies in insecticide discovery. In this paper, an economical and effective immobilised acetylcholinesterase (AChE) column was prepared using the sol-gel embedment method, which was further combined with high-performance liquid chromatography for screening the AChE inhibitors and insecticidal compounds from complex natural products. RESULTS AChE inhibitory constituents magnolol and honokiol were isolated from the ethanol extract of Magnolia officinalis, with IC50 values of 0.069 and 0.057 mM respectively. In an in vivo bioassay, magnolol and honokiol showed insecticidal activity against Nilaparvata lugens, with LC50 values of 0.324 and 0.137 mM, which are comparable with that of commonly used insecticide chlorpyrifos (0.233 mM). Moreover, molecular docking was carried out against a homology model of N. lugens AChE. The complexes showed that magnolol and honokiol placed themselves nicely into the active site of the enzyme and exhibited an interaction energy that was in accordance with our activity profile data. CONCLUSION These results demonstrate that magnolol and honokiol have great applied potential to be developed as natural insecticides, and an immobilised AChE column is very useful as a rapid screening tool for target enzymes towards potent inhibitors.