Establishment, Growth kinetics, and Susceptibility to AcMNPV of Heat Tolerant Lepidopteran Cell Lines

Lepidopteran heat-tolerant (ht) cell lines have been obtained with sf-9, sf-21 and several Bombyx cells. They have a distinct karyotype, membrane lipid composition, morphology and growth kinetics from the parental cell lines. In this paper, we report the development of ht cell lines from other insect species and examination of their growth characteristics and virus susceptibility. Adaptation of cell lines sf-9, BTI-TN-5B1-4 (High5) and BTI-TN-MG1 (MG1) to 33℃ and 35℃ was carried out by shifting the culture temperature between 28℃ and higher temperatures by a gradual stepwise increase in temperature. The process of adaption to a higher culture temperature was accomplished over a period of 2 months. The cell lines with the temperature adaption were designated as sf9-ht33, sf9-ht35, High5-ht33, High5-ht35, MG1-ht33, MG1-ht35. These cell lines have been subcultured over 70 passages. Adaption to high temperatures was confirmed by a constant population doubling time with individual cell lines. The population doubling time of heat adapted cell lines were 1-4 h less than these of parental cell lines. Cell shapes did not show obvious change, however, the cell size of sf9-ht cells was enlarged and those of High5 and MG1 ht cells were reduced after heat adaption. When the cell lines were infected with Autographa californica nuclear polyhedrosis virus (AcMNPV) at 28℃, 33℃, 35℃ and 37℃, production of budded virus and occlusion bodies in each cell line was optimum at its own adapted temperature.

Initiation of two ovarian cell lines from Fugu rubripes(Temminck et.Schlegel)

The ovary is an excellent system for studying stem cell renewal and differentiation, which is under the control of ovarian somatic cells. In order to understand oogenesis in Fugu rubripes （Temminck et. Schlegel） as a marine fish model of aquaculture importance, we established cell lines called TSOC 1 and TSOC2 from a juvenile ovary of this organism. TSOC1 is composed of spindle epithelial-like cells, while the other is cobblestone-like ceils. Therefore, TSOC1 and TSOC2 appear to consist of ovarian somatic cells. Growth requirement condition was investigated including temperature, concentration of FBS and pH. Significant fluorescent signals were observed after TSOC1 and TSOC2 cells were transfected with pEGFP-N3 vector, indicating its potential utility for genetic manipulation such as gene function studies. It is shown that these cell lines are effective for infection by the turbot reddish body iridovirus and flounder lymphosystis disease virus as evidenced by the appearance of cytopathic effect and virus propagation in the virus-infected cells, and most convincingly, the observation of viral particles by electron microscopy, demonstrating that TSOC1 and TSOC2 are suitable to study interactions between virus and host cells. It is believed that TSOC1 and TSOC2 will be useful tools to study sex-related events and interactions between primordial germ cells and oogonia cells during oogenesis. Therefore, establishment of ovary cell lines from Fugu rubripes seems to be significant for those research areas.

A novel cell line established from skin tissue of gibel carp(Carassius gibelio)is susceptible to Carassius auratus herpesvirus infection

Frequent outbreaks of emerging infectious diseases in fish,such as Carassius auratus herpesvirus(CaHV)infection has caused great economic losses in China.However,the lack of a sensitive cell culture system has limited studies of CaHV.In the present study,a new cell line(gibel carp skin cell,GiCS)derived from gibel carp(Carassius gibelio)skin tissue was established to create a valuable tool for research of the virus.The GiCS cells consisted mainly of epithelial-like cells,which grew well at 25℃in L-15 medium supplemented with 10–20%fetal bovine serum.Chromosomal analysis revealed that the skin cell line remained amphitriploid,with most chromosome counts being 156(54%).The GiCS cells can be efficiently transfected and expressed exogenous genes.In particular,the GiCS cells showed high susceptibility to CaHV infection,which was confirmed by virus infection tests,detection of viral gene expression,and ultrastructural observation.To our knowledge,it is the first cell line that is highly permissive to CaHV infection.In addition,the cells also showed susceptibility to several aquatic animal viruses from different families including Iridoviridae,Rhabdoviridae,and Reoviridae.In conclusion,these results indicated that the establishment of the GiCS cell line is a significant advance that will be beneficial to future studies of CaHV and other aquatic animal viruses.

Association of HLA Alleles(A, B, DRB1) and HIV-1 Infection in the Han Population of Hubei, China

The HIV susceptibility and resistance alleles in the HLA genes were determined by investigating the distribution characteristics of the HLA alleles（A, B, and DRB1） in HIV-infected individuals of the Han population in Hubei, and by comparing these alleles with HIV-negative individuals from the same area. A cohort of 424 HIV-1 infected individuals were chosen as study subjects, and 836 HIV-negative healthy subjects from the same area served as the control population. HLA-A, B, and DRB1 allele typing was performed using polymerase chain reaction-sequence-specific oligonucleotide probes（PCR-SSOP） and polymerase chain reaction-sequencing based typing（PCR-SBT） techniques. Arlequin ver3.0 was used to analyze the allele and haplotype frequencies of HLA-A, B, and DRB 1, whereas Epi Info 7 and SPSS18.0 was used to analyze the differences in the HLA alleles between the HIV-1 positive and HIV-1 negative groups. A＊02：03, DRB1＊01：01, and DRB1＊15：01 alleles and their haplotypes as well as the HLA_Bw4-Bw6 hybrid showed a protective effect on HIV-1 infection. After adjusting for confounding factors such as age and sex, multivariate logistic regression analysis revealed that B＊15：02G, DRB1＊01：01, and DRB1＊15：01 subtypes were the resistance genes of HIV-1 infection, while B＊13：01 might increase susceptibility to HIV-1 infection. The correlation between A＊02：06 and B＊15：01G subtypes and HIV-1 susceptibility was independent of the age and sex of the host. This study demonstrated the influence of genetic factors in humans such as HLA polymorphism on individuals to resist HIV-1 infection. Association studies of HLA polymorphism, susceptibility/resistance to HIV-1 infection, and hosts＇ genetic background are of significant importance for research on HIV-1 pathogenesis and vaccine design.