Genetic characterization and diversity of porcine circovirus type 2 in non‐vaccinated South African swine herds

The porcine circovirus type 2 (PCV2) is a swine infectious viral pathogen of great significance in global swine herds. It was recently detected at another Province of South Africa sequel to the first detection of North American‐like strain (PCV2a) at Gauteng about two decades ago, but there is a dearth of information about the genomic features and diversity of the viral strains in circulation within the country and the entire sub‐Saharan Africa region. To date, only one complete genome of the virus from South Africa is available on global data base. This current effort is therefore geared towards the full‐genome characterization of the circulating PCV2 strains in the pigs of Eastern Cape Province. With the use of conventional polymerase chain reaction method, fifteen complete PCV2 genomes were successfully amplified, sequenced and assembled from field samples obtained from non‐vaccinated pigs in the region. Neighbor Joining and Maximum Likelihood phylogenetic analyses of the ORF2 gene and full genomes unanimously showed that most of the assembled genomes (11) belong to genotype PCV2b. Furthermore, three of the characterized sequences formed clade with other reference mutant PCV2b and PCV2b subtype 1C (i.e. PCV2d) strains from the USA, China and South Korea. The last sequence, however, clustered with other reference strains belonging to PCV2 intermediate clade 2 (PCV2‐IM2), recently identified in a global PCV2 strains phylogenetic analysis. This study reports the first complete genome sequences of PCV2b, PCV2d and PCV2‐IM2 in pigs from South Africa, and it gives a possible insight into the genetic characteristics and variability of the viral strains presently in circulation within the country. It further emphasizes the need for more stringent measures in curtailing the introduction and spread of transboundary swine pathogens in the country and entire Southern African region.     

Golgi study on brain of macular mutant mouse as a model of Menkes kinky hair disease

Summary This study was undertaken to elucidate, using the Golgi method, the neuropathological change in the brain of the macular mutant mouse, whose hemizygote (Ml/y) is considered to be a model of Menkes kinky hair disease (MKHD). The hemizygote mice gradually lost weight after 10 days of age and died with emaciation and seizure around day 15. The normal littermate (+/y) was well developed. In the cerebrum, the arborization of pyramidal neurons in the layer V of the Ml/y was the same as that in the +/y on day 10. However, development of arborization in the Ml/y was delayed in comparison with that in the +/y on days 12 and 14. Purkinje cells with several somal sprouts were observed in the cerebellum in both the Ml/y and +/y on day 7. The somal sprouts in the +/y had regressed gradually by day 12, while they were still in the anterior and middle lobes of the Ml/y on day 14. Additionally, the trunks of Ml/y stem dendrites became thicker and a cactus formation was recognized on the branching portion of the dendrites on day 14. Arborization of these abnormal Purkinje cells was distinctly poor compared with that in the +/y. These results suggest that the growth of the neurons is delayed in the Ml/y and simultaneously their cytoskeletal developments are disturbed, especially in the Purkinje cells. There is a close similarity in many respects to the neuropathological change in MKHD.     

Isolation and characterization of a monoclonal anti-P30 antibody resistant mutant of Toxoplasma gondii

Of the possible iodine-labelled Toxoplasma gondii surface proteins, P30 (apparent Mr 30,000) is the principal one recognized by acute and convalescent anti-toxoplasma sera. This protein which comprises from 3 to 5% of the total parasite protein was used to raise a panel of parasiticidal monoclonal anti-P30 antibodies. One of these monoclonal antibodies was able to select a resistant mutant from a large population of chemically mutagenized wild-type P strain parasites. This mutant retained the wild type sensitivity to other non-P30 parasiticidal monoclonal antibodies as well as polyclonal anti-P30 rabbit sera. Analysis of surface radioiodinated wild type and mutant parasites showed that the mutant had a quantitative reduction in the amount of P30. A comparison of surface biotin labelled wild type and resistant parasites by two dimensional electrophoresis showed that the mutant lacked one and possibly two of several proteins that make up wild type P30. Western blot analysis indicated that the mutant was devoid of antigenically reactive P30. These findings further support the hypothesis that antigenic variants of T. gondii can be induced and may involve the major surface membrane antigens of the parasite.     

HBcAg显性失活突变体质粒的构建及体外表达

目的：探讨HBcAg显性失活突变体质粒pCDNA4-C-GFP的构建及体外表达。方法：选择编码乙肝核心抗原C基因片段及绿色荧光蛋白（green fluorescent protein,GFP)基因片段，利用基因重组技术构建成DNA质粒pCDNA4-C-GFP,并将该质粒转染肝癌细胞株HepG2。结果：通过RT－PCR检测到其RNA的表达，Confocal观察到GFP绿色荧光。结论：HBcAg显性失活突变体质粒pCDNA4-C-GFP的构建成功可以进行对HBV作用的研究。     

Virulence of Candida albicans mutants

Mutant strains of the fungal pathogen Candida albicans blocked in pyrimidine transport and salvage metabolism were tested for virulence in various animal models. The growth rate, germination and proteolytic enzyme production did not correlate with the virulence of the strains. However, a defect in the uridine transport system significantly decreased virulence in murine candidosis, although it had no effect in vaginal candidosis or in a Candida cyst model. It remains unclear whether this is due to the differing host defence mechanisms involved in systemic and superficial mycoses, or to the different requirements of the fungal systems for adherence and tissue invasion in the two types of infection.     

乙型肝炎表面抗原“免疫逃避”突变体在哺乳动物细胞中的表达

利用乙型肝炎病毒ＤＮＡ开放框架上的ＢａｍＨＩ和ＨｐａＩ位点，酶切消化质粒载体ＰＥｃｏｂ６（含双拷贝ＨＢＶＤＮＡ），得到约９００ｂｐ的ＨＢＶ－Ｓ基因片断。将其插入到噬菌粒载体ＰＢｌｕｅｓｃｒｉｐｔｓｋ＋的ＳｍａＩ位点上。然后通过体外寡核苷酸介导的人工定点突变获得一系列（共１２种）Ｓ基因“免疫逃避”突变型。再通过ＥＢ病毒真核表达载体ｐＭＥＰ４上的ＢａｍＨＩ和Ｋｐｎｌ位点将噬菌粒ｐＢｌｕｅｓｃｒｉｐｓｋ＋上的Ｓ基因突变型片断定向克隆到ＰＭＥＰ４上，从而构建了含乙肝Ｓ基因突变型的重组质粒ｐＭＥＰ４ＨＢＳＭ。用其转染人肝癌传代细胞系ＨｅｐＧ２，经潮霉素选择，三周后获得抗性细胞克隆。经用抗ＨＢｓ单克隆抗体（含针对ＨＢｓＡｇ“ａ”抗原决定簇）检测除含变异体１４５（即１４５位上甘氨酸为精氨酸替代）外其余抗变异体ＨＢｓＡｇ均为阳性。经Ｗｅｓｔｅｒｎｂｌｏｔ证实变异体１４５，在分子量约为２３ＫＤ处有一特异ＨＢｓＡｇ蛋白带。     

阿弗菌素产生菌SIPI-AV-99081的选育

阿弗菌素链霉菌SIPI-AV-99081采用UV和NTG进行诱变处理。选育得到一高产菌株和几个典型的突变株。高产菌株AV-h-360的阿弗菌素B1的生产能力较出发菌株提高了3倍，AV-m-486和AV-m-796为阿弗菌素合成阻断突变株，它们都不能产生天然的阿弗菌素。     

Serotonin content is elevated in the dopamine deficient striatum of the weaver mutant mouse

In the present study, we measured the striatal serotonin content of weaver and control mice at different ages. Overall, weaver mutant mice exhibited 50% more striatal serotonin than controls. Neither a rostrocaudal gradient nor an age effect was found for either genotype. An analysis of serotonin content across the dorsoventral extent of the striatum revealed that in the dorsal striatum of the weaver, serotonin content was increased 200%, and in the ventral striatum, the increase amounted to 50% relative to control mice. Serotonin immunocytochemistry also revealed an increase in the dorsal striata of weaver mice. The major increase in striatal serotonin content seen in the weaver striatum occurs in the same region that exhibits the severest dopamine depletion. This observation is consistent with the notion that the increase in serotonin levels may be secondary to the decrease in dopamine content and may play an adaptive or compensatory role.     

Spatial and temporal pattern of purkinje cell degeneration in shaker mutant rats with hereditary cerebellar ataxia

Temporal-spatial patterns of surviving Purkinje cells were studied quantitatively in a rat mutant (shaker) with differential hereditary cerebellar ataxia and Purkinje cell degeneration. Shaker rat mutants are characterized behaviorally as mild if they are ataxic or as strong if they have ataxia and tremor. Purkinje cells degenerate in both mild and strong shaker mutants, but the temporal and spatial patterns of cell death are strikingly different. In mild shaker mutants, Purkinje cell death is temporally restricted, with 31-46% of the Purkinje cells in lobules I-IX dying by 3 months of age. Very few Purkinje cells degenerate after this age. Purkinje cell death is spatially random. In lobules I-IX, every second, third, or fourth Purkinje cell degenerates. Purkinje cells in lobule X do not degenerate. In strong shaker mutants, Purkinje cell degeneration is temporally protracted and spatially restricted. By 3 months of age, most Purkinje cells in lobules I-VIa, -b, and -d, and -d have degenerated. Numerous Purkinje cells in the paravermis of lobules VIIb-VIII have also degenerated. Surviving Purkinje cells in the vermis and lateral hemisphere of lobules VIIb-VIII are aligned in parasagittally oriented stripes or transversely oriented bands. Purkinje cells continue to degenerate in localized areas of the posterior lobe such that, by 18 months of age, surviving Purkinje cells are limited primarily to lobules VIc, VIIa, IXd, and X. Quantitative analysis indicates that none of the Purkinje cells in these lobules degenerate.     

Keratinocyte-derived Cytokines and UVB-Induced Immunosuppression

Ultraviolet radiation (UVB) in sunlight is known to have multiple effects on the immune system. Evidence suggests that UVB-induced immunosuppression is mediated in part by immunosuppressive and immunoregulatory cytokines. Our studies have utilized gene-targeted mutant mice to determine key molecular requirements essential for the development of UVB-induced immunosuppression. Preliminary results from our laboratory suggest that TNF-α plays a regulatory role in contact hypersensitivity, but is not a crucial factor for UVB-induced immunosuppression, and that multiple factors are involved in the induction of UVB mediated immunosuppression.