Cytochemical demonstration of modification of carbohydrates in the mouse zona pellucida during folliculogenesis

In the present study, lectin-gold cytochemistry and antibodies against ZP2 and ZP3 glycoproteins were used to investigate the oligosaccharide content of mouse ovarian zona pellucida (ZP) during follicular development. The entire thickness of the ZP and several organelles of the oocyte (cortical granules, Golgi apparatus, and vesicular aggregates) were reactive to RCA-I, DSA, AAA, WGA, MAA, and LFA throughout follicular development. HPA labeling was not detected at the earliest stages of follicular folliculogenesis. HPA reactivity was first observed in the ZP, Golgi apparatus, and the vesicles of oocytes at the trilaminar primary follicle stage. HPA labeling in the ZP was always restricted to the inner region of the zona matrix. After neuraminidase treatment, HPA reacted with the entire ZP in ovarian follicles at different stages of development. Immunolabeling with specific antibodies showed that, although ZP2 and ZP3 glycoproteins were uniformly distributed in the zona matrix of ovarian oocytes, there was a progressive increase in thickness of the ZP in parallel with the proliferation of follicular cells. ZP3 glycoprotein was also localized to the Golgi apparatus and vesicular aggregate. The present results suggest: (1) a difference in composition of carbohydrate content between the inner and outer region of the fully developed ZP generated probably by a modification in the biosynthetic pathway of oligosaccharides in the oocyte during folliculogenesis, (2) that newly synthesized ZP glycoproteins displace previously synthesized ZP components in a direction toward the follicular cells and, therefore, no redistribution of the ZP matrix occurs during folliculogenesis, and (3) that the vesicular aggregates in the ooplasm constitute an intermediate step in the secretory pathway of ZP glycoproteins. Accepted: 10 January 2000     

Proteomic analysis of the oxygen-evolving complex of photosystem II under biotec stress: Studies on Nicotiana benthamiana infected with tobamoviruses

We have previously shown that tobamovirus infection induces an inhibition of photosystem II electron transport, disturbing the oxygen-evolving complex (OEC). In the infected plants, the OEC polypeptide pattern was modified when compared to healthy plants, the levels of the PsbP and PsbQ extrinsic proteins being lowered to different extents. In this work we have further investigated by two-dimensional polyacrylamide gel electrophoresis (2-DE) the changes on the OEC protein pattern of thylakoid membranes isolated from Nicotiana benthamiana Domin plants infected with the Spanish strain of pepper mild mottle virus. When the thylakoid membranes from healthy plants were analyzed for the presence of PsbO and PsbP proteins by 2-DE (pI range 4-7) and further immunoassayed by using specific-antisera against these two proteins, it was observed that four polypeptides cross-reacted with each antiserum. These data, along with the N-terminal amino acid sequence determined for the eight polypeptides, indicate that the N. benthamiana PsbO and PsbP proteins correspond to protein families. In the silver-stained 2-DE gels of thylakoid membranes isolated at different days postinoculation from virus-infected plants, it was observed that the content of PsbP polypeptides decreased dramatically with respect to those of PsbO, during the progress of the infection. Interestingly, there was a differential decrease of the different PsbP proteins, indicative of a distinct regulation of their expression.     

Inhibition of photosynthesis by viral infection: Effect on PSII structure and function

The effect of viral infection on photosynthesis was investigated in Nicotiana benthamiana Gray plants infected with different strains of pepper and paprika mild mottle viruses (PMMoV and PaMMoV) and chimeric viral genomes derived from them. In both symptomatic and asymptomatic leaves of virus-infected plants, photosynthetic electron transport in photosystem II (PSII) was reduced. In all cases analyzed, viral infection affected the polypeptide pattern of the oxygen-evolving complex (OEC) in thylakoid membranes. The levels of both the 24 and 16 kDa proteins were reduced to a differing extent when compared with the levels in healthy control. This loss of the OEC extrinsic proteins affected the oxygen evolution rates of thylakoid membranes and leaves from infected plants. Additionally, viral coat protein (CP) was found associated with the chloroplasts and the thylakoid membranes of the infected plants. The CP accumulation level was dependent upon both the post-infection time and the virus analyzed, but independent of the CP itself since hybrid viruses did not behave as their parental viruses with the same CP, with respect to PSII inhibition, CP accumulation rates and OEC protein levels. Modulated chlorophyll (Chl) fluorescence and oxygen evolution measurements carried out in both types of leaves showed that the quantum yield of PSII electron transport was diminished in infected plants with respect to those of control plants. The decrease in electron transport efficiency was mainly caused by a reduction in the fraction of open reaction centers. The infected plants also showed a reduction in the efficiency of excitation capture in PSII by photoprotective thermal dissipation of excess excitation energy.     

Antixenosis resistance to Aphis fabae Scopoli (Hemiptera:Aphididae) in bean cultivars

Aphis fabae Scopoli (Hemiptera: Aphididae) is heteroecious and polyphagous that is harmful on secondary hosts such as many important agricultural products like beet, common bean, faba bean, potato and other products. This aphid is the cause of more than 33 viral transition. One of the mechanisms of plant resistance is antixenosis. This mechanism influences on placement and nutrition of pests that result in less damage. In this study, antixenosis resistance mechanism of 12 varieties of bean was tested. Experiment was on completely randomised design with 12 treatments and 6 replications. Bean varieties include of white bean, kidney bean and wax bean, and each replication includes one pot, and then, pots were placed under the isolated room that were filled with winged adult aphids in circular form. After 24 and 48?h, aphids and level of nymph production were counted. The lowest number of adult aphids was observed on Sayad variety among 12 varieties (during 24?h). The least number of produced nymphs was in Daneshkade variety. In Sayad variety, the frequency of matured insects and produced nymphs was minimum.     

Genetic variation in Malaysian oysters: taxonomic ambiguities and evidence of biological invasion

Genetic diversities in two cultured oyster species, Crassostrea iredalei (Faustino 1932) and Crassostrea belcheri (Sowerby 1871) were assessed using a 581-nucleotide fragment of the mtDNA cytochrome oxidase subunit 1 (CO1) gene. A total of 103 C. iredalei individuals and 120 C. belcheri from 12 populations were sampled along the coast of Malaysia. Trees of unique haplotype samples generated based on Neighbor-Joining (NJ) algorithm revealed that many individuals had been misidentified and did not cluster with their presumed species based on morphological identification. BLAST results of DNA sequences showed presence of previously unreported C. madrasensis in Peninsular Malaysian waters (98% maximum identity). The true identity of the Muar (Crassostrea sp.) and Semporna (Saccostrea sp.) populations were unresolved by two BLAST search and showed less than 88% identity with other species in GenBank. Repeated analysis of these two populations using 487 bp of the mitochondrial 16S gene data showed only a maximum identity less than 97%. Hence, the identity of these specimens remains unclear. Evolutionary divergences within presumed species were 0.001–0.011 and 0.034–0.313 between species. Findings from this study have important implications for aquaculture, management and monitoring of cultured populations as well as conservation of wild oyster species in Malaysia.     

Purification of a new fungal mannose-specific lectin from Penicillium chrysogenum and its aphicidal properties

Several Ascomycetes fungi are commonly used in bio-industries and provide available industrial residues for lectin extraction to be valuable. A lectin from Penicillium chrysogenum, named PeCL, was purified from a fungal culture using gel-filtration chromatography column. PeCL was found to be a mannose-specific lectin by haemagglutination activity towards rabbit erythrocyte cells and was visualised on SDS-PAGE gel. Purified PeCL fraction was delivered via artificial diet to Myzus persicae aphid and was demonstrated to be aphicidal at 0.1?% with higher toxic efficiency than a known mannose-binding lectin Concanavalin A (ConA). A fast and efficient way to purify PeCL and a potential use in pest control is described.     

Biochemical characterisation of digestive α-amylase of Red Palm Weevil,Rhynchophorus ferrugineus (Olivier, 1790) (Coleoptera: Curculionidae)

The Red Palm Weevil, Rhynchophorus ferrugineus (Oliver) (Coleoptera: Curculionidae), is a serious pest of a wide range of plant species including coconut, sago, date and oil palms. The α-amylases are the hydrolytic enzymes that are involved in carbohydrate metabolism in insects. So far nothing is done to demonstrate α-amylase activity of R. ferrugineus. Thus, the aim of the current study was to identify and characterise the α-amylase activity to gain a better understanding of digestive physiology of the insect. Thus, the α-amylase in the gut of red palm weevil was isolated and characterised using starch as a substrate. The study showed that the α-amylase is present in the gut of the insect for carbohydrate digestion. The α-amylase has an optimum pH and temperature of 5 and 40°C. The activity of α-amylase was increased by NaCl and KCl and inhibited by other compounds such as MgCl₂, CaCl₂, urea, ethylenediaminetetraacetic acid and sodium dodecylsulfate. Native-PAGE electrophoresis of α-amylase showed two isoenzymes, one major and one minor band showing α-amylase importance in the carbohydrate metabolism of the insect. Understanding of the digestive physiology and α-amylase activity of Red Palm Weevil is important when new management strategies for this economically important pest are devised.     

Seasonal population fluctuations of the diamondback moth,Plutella xylostella (L.) (Lep.: Plutellidae) on different cauliflower cultivars

The diamondback moth, Plutella xylostella (L.) (Lep.: Plutellidae) is one of the most important pests of cruciferous plants throughout the world. In recent years, it has been identified as a serious pest of the cauliflower fields in Tehran province. Resistance of P. xylostella to all main groups of insecticides has been recorded and it is ranked in the 20 most resistant pest species reported until now. According to many researchers, to solve the problem of pest resistance to chemical pesticides, an integrated pest management programme should be used. Despite this condition, it seems that the use of resistant cauliflower cultivars is an appropriate policy for integrated control of the pest in the field. In order to identify the most resistant cultivar in the field, eight cauliflower cultivars in a completely randomised design with five replicates were planted at the Shahed University research field (south of Tehran). Density of eggs, larvae and pupae of P. xylostella were measured every 10 days in these cultivars. The results showed that there is no significant difference between numbers of eggs per plant on different cultivars. But number of larvae and pupae per plant were significantly different among different cultivars. Smilla and Snow mystique cultivars had the highest number of larvae and pupae. On the other hand, Buris and Snow crown cultivars had the lowest number of pupae and Snow crown and SG cultivars had the lowest number of larvae per plant. According to the results, the Buris and Snow crown cultivars had the lowest infestation and had a kind of resistance to pest.     

RF hydrazine plasma modification of chitosan for antibacterial activity and nanofiber applications

Chitosan nano powders were modified using RF hydrazine plasma produced at low pressure (26.66 Pa) with 13.56 MHz frequency at a power of 100 W for 30 min. Characterization and investigation of the properties of plasma-modified chitosan (PMCh) and non-modified chitosan (Ch) were carried out using an optical monochromator, FTIR, florescence analysis, TGA, SEM, and X-ray techniques. FTIR spectra of PMCh indicated a band broadening at 3436 cm⁻¹ that confirmed increasing functional groups based on H-bonding. The number of NH₂ groups was determined from fluorescence analysis. TGA analysis shows that the moisture absorption is three times higher in the PMCh structure. Ch and PMCh in PVA solutions were used to produce nanofibers by the electrospinning method; average fiber diameters were 480 and 280 nm for Ch and PMCh, respectively. It was found that the antibacterial effect of PMCh is better than the Ch for Gram-positive strains.