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Biomedical subjects

Dieter Klein

Publications and source records attributed to Dieter Klein.

6 recordsLinked to original sources
PubMed2003

Phylogenetic analysis of feline immunodeficiency virus in Central Europe: a prerequisite for vaccination and molecular diagnostics.

Feline immunodeficiency virus (FIV) is a worldwide-occurring lentivirus that severely impairs the immune function of infected domestic cats. Due to structural and biological similarities, FIV represents a promising model for human immunodeficiency virus (HIV) and AIDS. A major obstacle in developing vaccines against lentiviruses is their high mutation rate. Furthermore, mutations in target sequences provide a pitfall for molecular diagnostics. It is therefore important to determine the genetic diversity of lentiviruses in any region where vaccination or implementation of new diagnostic techniques are planned. This study presents a phylogenetic analysis of 30 FIV strains derived from Central Europe. In order to improve the reliability of genotyping, DNA from two different proviral genes was amplified and comparative phylogenetic trees were inferred. The highly coincident results point to the existence of extensive virus variation with the presence of at least two highly divergent subtypes of FIV in Austria and Germany.

Animals↗
PubMed2002

Quantification using real-time PCR technology: applications and limitations.

The introduction of real-time PCR technology has significantly improved and simplified the quantification of nucleic acids, and this technology has become an invaluable tool for many scientists working in different disciplines. Especially in the field of molecular diagnostics, real-time PCR-based assays have gained favour in the recent past. However, the wide use of real-time PCR methods has also highlighted some of the critical points and limitations of these assays. These aspects must be considered to increase the reliability of the obtained data.

Gene Expression Profiling↗
PubMed2002

The murine whey acidic protein promoter directs expression to human mammary tumors after retroviral transduction.

The whey acidic protein (WAP) promoter is known to be active in pregnant and lactating mammary epithelial cells as well as mammary tumors of mice. Here we show that a proximal fragment of the murine WAP promoter, including most elements postulated as being responsible for mammary-specific regulation, confers mammary-specific expression upon a marker gene in transgenic mice even though the distal promoter region, known to be important for rat WAP promoter activity, is lacking. The relatively small size of this fragment allows its insertion into a murine leukemia virus-based retroviral vector in place of the viral promoter. Infection of a number of established human mammary and nonmammary cell lines with such a retroviral vector revealed that the WAP promoter was limited in its activity to mammary tumor cell lines. Expression in tumorigenic mammary cells was even more pronounced when these cells were introduced into the mammary fat pads of mice. This is the first demonstration that the WAP promoter is active in human mammary cells and mammary tumor cells in general, and suggests that the extended proximal WAP promoter may be useful for directing therapeutic gene expression to human mammary tumors.

3T3 Cells↗
PubMed2002

Evolution of replication efficiency following infection with a molecularly cloned feline immunodeficiency virus of low virulence.

The development of an effective vaccine against human immunodeficiency virus is considered to be the most practicable means of controlling the advancing global AIDS epidemic. Studies with the domestic cat have demonstrated that vaccinal immunity to infection can be induced against feline immunodeficiency virus (FIV); however, protection is largely restricted to laboratory strains of FIV and does not extend to primary strains of the virus. We compared the pathogenicity of two prototypic vaccine challenge strains of FIV derived from molecular clones; the laboratory strain PET(F14) and the primary strain GL8(414). PET(F14) established a low viral load and had no effect on CD4(+)- or CD8(+)-lymphocyte subsets. In contrast, GL8(414) established a high viral load and induced a significant reduction in the ratio of CD4(+) to CD8(+) lymphocytes by 15 weeks postinfection, suggesting that PET(F14) may be a low-virulence-challenge virus. However, during long-term monitoring of the PET(F14)-infected cats, we observed the emergence of variant viruses in two of three cats. Concomitant with the appearance of the variant viruses, designated 627(W135) and 628(W135,) we observed an expansion of CD8(+)-lymphocyte subpopulations expressing reduced CD8 beta-chain, a phenotype consistent with activation. The variant viruses both carried mutations that reduced the net charge of the V3 loop (K409Q and K409E), giving rise to a reduced ability of the Env proteins to both induce fusion and to establish productive infection in CXCR4-expressing cells. Further, following subsequent challenge of naïve cats with the mutant viruses, the viruses established higher viral loads and induced more marked alterations in CD8(+)-lymphocyte subpopulations than did the parent F14 strain of virus, suggesting that the E409K mutation in the PET(F14) strain contributes to the attenuation of the virus.

Amino Acid Sequence↗
PubMed2000

Variations in the major envelope glycoprotein GP5 of Czech strains of porcine reproductive and respiratory syndrome virus.

The major envelope glycoprotein genes (ORF5) of seven Czech isolates of porcine reproductive and respiratory syndrome virus (PRRSV) were amplified and their nucleotide sequences were determined. ORF5 displayed nucleotide and amino acid identities of 87.5-100% and 87. 6-100%, respectively, among the isolates. In a phylogenetic tree, all European isolates were grouped in a genotype distinct from that of reference American strains (VR-2332, IAF-Klop). Among the European isolates, two different clades were identified. Two Czech isolates (V-501 and V-503) and Italian strain PRRSV 2156 fell into one clade. The remaining European strains comprised the second clade. Surprisingly, two separately clustered strains (V-501 and V-516) were isolated from the same herd. Additionally, the possible effect of in vitro cultivation on the nucleotide sequence was analysed. Nine point mutations in the ORF5 region resulted from 152 in vitro passages of the V-502 isolate in MARC-145 cells.

Amino Acid Sequence↗