一种改良的对虾白斑综合征病毒的提纯技术

The envelope proteins of White spot syndrome virus (WSSV) are very fragile and easy to be destroyed during purification. It was difficult to obtain a large quantity of intact virions by routine sucrose gradient centrifugation. After modifying the sucrose gradient by adding citrate sodium, we can obtain a large quantity of intact virions and nucleocapsids. This purified virions and nucleocapsids were subsequently used for analyzing viral structural proteins and DNA extraction. The result showed that this modified techniaue is very efficient for virus purification.     

甜瓜细菌性斑点病拮抗菌P-13的鉴定及其抑菌物质的初步研究

从新疆甜瓜根际土壤中分离到1株甜瓜细菌性斑点病拮抗放线菌P-13, 根据其形态学、生理特征和16S rRNA序列分析, 鉴定该菌株为娄彻氏链霉菌(Streptomyces rochei)。琼脂扩散法生物活性研究表明, 其发酵液对细菌性果腐病菌(Acidovorax avenae subsp. citrull)BFB、细菌性角斑病菌(Pseudomonas syringae pv. Lachrymans)P4的抑菌圈直径分别为19 mm和17 mm以上; 发酵液中抑菌物质主要为胞外代谢物, 不溶于石油醚, 乙醚, 乙酸乙酯等有机溶剂。在100°C处理10 min、pH 6处理6 h或紫外线照射7 h, 该物质抑菌活性不变。纸层析结果表明, 该物质主要为碱性水溶性物质。     

Effects of oral recombinant VP28 expressed in silkworm (Bombyx mori) pupa on immune response and disease resistance of Procambarus clarkii

The envelope protein VP28 of white spot syndrome virus (WSSV) was overexpressed in the silkworm Bombyx mori, which was achieved by using a baculovirus (HyNPV) expression system and by making silkworm pupa as an alternative host, and then it was directly supplemented in diet at a dose of 20 g kg⁻¹ without purification. During a 30 day feeding period, the levels of phenoloxidase (PO) and superoxide dismutase (SOD) in the haemolymph of the tested Procambarus clarkii increased greatly (P < 0.05) when compared to the control crayfish fed with wild-type HyNPV baculovirus-infected silkworms or normal silkworms. Compared with two controls, the crayfish which had been infected for 20 days showed a significantly lower (P < 0.05) mean cumulative mortality (15.6%), which respectively, resulted in relative percent survivals (RPS) of 83.7 and 84.4%. The efficacy to inhibition of viral infection was further studied by in situ hybridization with a WSSV-specific DNA probe. The high levels of PO and SOD might be important for developing resistance against WSSV in these crayfish.     

Analysis of pathogenicity mutants of a bacteriocin producing Xanthomonas perforans

Since the initial discovery of Xanthomonas perforans on tomato in 1991, it has completely displaced Xanthomonas euvesicatoria as the bacterial spot of tomato pathogen in Florida. Previous research has shown that X. perforans produces at least three different bacteriocin-like compounds (BcnA, BcnB, BcnC) antagonistic toward X. euvesicatoria strains. In this study pathogenicity-attenuated, bacteriocin-producing mutants of X. perforans were created to determine their potential as biological control agents for control of X. euvesicatoria. Several candidate genes were chosen based on previous studies in which mutant phenotypes exhibited reduced virulence in either X. perforans (OpgH_Xcv) or the closely related X. euvesicatoria strain 85-10 (hpaB, hpaC, xopA, xopD, avrBs2 and gumD). Each candidate gene in X. perforans was amplified and PCR-assisted deletion mutagenesis was performed in the wild-type (wt) X. perforans strain to create potential attenuation mutants. Each mutant was tested for growth rate, disease severity and antagonism toward X. euvesicatoria strains. Three mutants, XopA, opgH, and gumD were significantly less pathogenic than the wild-type strain with the opgH mutant reaching significantly lower internal populations than all other mutants except hpaC. The opgH-strain was the most affected in its ability to grow internally in plant tissue while inhibiting X. euvesicatoria populations equal to or more than the other mutant strains. This mutant strain could potentially be used as part of an effective biological control strategy.     

杭州西湖风景区外来杂草的调查研究

通过野外调查和文献资料的整理分析,确定杭州西湖风景区有外来杂草56种,隶属于21科44属。对西湖风景区外来杂草的种类组成、来源、分布特点进行分析,结果表明:(1)种类较多的科是菊科、苋科、豆科;(2)原产地以来源于北美洲的最多;(3)容易入侵地表裸露较大的生境;(4)发生具有季节差异;(5)种类、分布上呈动态变化。并就外来杂草近二十至三十年的变化进行了讨论。     

Modulatory effects of Tabebuia impetiginosa (Lamiales, Bignoniaceae) on doxorubicin-induced somatic mutation and recombination in Drosophila melanogaster

The wing Somatic Mutation and Recombination Test (SMART) in D. melanogaster was used to study genotoxicity of the medicinal plant Tabebuia impetiginosa. Lapachol (naphthoquinone) and β-lapachone (quinone) are the two main chemical constituents of T. impetiginosa. These compounds have several biological properties. They induce apoptosis by generating oxygen-reactive species, thereby inhibiting topoisomerases (I and II) or inducing other enzymes dependent on NAD(P)H:quinone oxidoreductase 1, thus affecting cell cycle checkpoints. The SMART was used in the standard (ST) version, which has normal levels of cytochrome P450 (CYP) enzymes, to check the direct action of this compound, and in the high bioactivation (HB) version, which has a high constitutive level of CYP enzymes, to check for indirect action in three different T. impetiginosa concentrations (10%, 20% or 40% w/w). It was observed that T. impetiginosa alone did not modify the spontaneous frequencies of mutant spots in either cross. The negative results observed prompted us to study this phytotherapeuticum in association with the reference mutagen doxorubicin (DXR). In co-treated series, T. impetiginosa was toxic in both crosses at higher concentration, whereas in the HB cross, it induced a considerable potentiating effect (from ~24.0 to ~95.0%) on DXR genotoxity. Therefore, further research is needed to determine the possible risks associated with the exposure of living organisms to this complex mixture.     

Analysis of eighteen deletion breakpoints in the parkin gene

Parkin mutations are responsible for the pathogenesis of autosomal-recessive juvenile parkinsonism (AR-JP). On initial screening of Japanese patients with AR-JP, we had found that approximately half of the parkin mutations are deletions occurring between exons 2 and 5, forming a deletion hot spot. In this study, we investigated the deletion breakpoints of the parkin mutations in 22 families with AR-JP and examined the possible association between these deletion events and meiotic recombinations. We identified 18 deletion breakpoints at the DNA nucleotide sequence level. Almost all these deletions were different, indicating that the deletion hot spot was generated by recurrent but independent events. We found no association between the deletions and specific DNA elements. Recent copy number variation (CNV) data from various ethnic groups showed that the deletion hot spot is overlapped by a highly polymorphic CNV region, indicating that the recurrent deletion mutation or CNV is observable worldwide. By comparing Marshfield and deCODE linkage maps, we found that the parkin deletion hot spot may be associated with a meiotic recombination hot spot, although such association was not found on comparison with recent high-resolution genetic maps generated from the International HapMap project. Here, we discuss the possible mechanisms for deletion hot spot formation and its effects on human genomes.     

Pathological changes in Fenneropenaeus indicus experimentally infected with white spot virus and virus morphogenesis

To demonstrate pathological changes due to white spot virus infection in Fenneropenaeus indicus, a batch of hatchery bred quarantined animals was experimentally infected with the virus. Organs such as gills, foregut, mid-gut, hindgut, nerve, eye, heart, ovary and integument were examined by light and electron microscopy. Histopathological analyses revealed changes hitherto not reported in F. indicus such as lesions to the internal folding of gut resulted in syncytial mass sloughed off into lumen, thickening of hepatopancreatic connective tissue with vacuolization of tubules and necrosis of rectal pads in hindgut. Virus replication was seen in the crystalline tract region of the compound eye and eosinophilic granules infiltrated from its base. In the gill arch, dilation and disintegration of median blood vessel was observed. In the nervous tissues, encapsulation and subsequent atrophy of hypertrophied nuclei of the neurosecretory cells were found. Transmission electron microscopy showed viral replication and morphogenesis in cells of infected tissue. De novo formed vesicles covered the capsid forming a bilayered envelop opened at one end inside the virogenic stroma. Circular vesicles containing nuclear material was found fused with the envelop. Subsequent thickening of the envelop resulted in the fully formed virus. In this study, a correlation was observed between the stages of viral multiplication and the corresponding pathological changes in the cells during the WSV infection. Accordingly, gill and foregut tissues were found highly infected during the onset of clinical signs itself, and are proposed to be used as the tissues for routine disease diagnosis.