Isoacentric and isocentric chromosomes originating after deletions of human chromosomes

Summary The analysis of a sample of 100 isoacentric (IA) and isocentric (IC) chromosomes, which had originated from spontaneous or radiation-induced deletions in human lymphocytes, is reported. IC and also IA have a strong tendency to be formed after breakage in juxtacentromeric heterochromatin. When euchromatic regions are involved, the breaks are not distributed at random since they frequently occur at places where juxtacentromeric heterochromatin exists in other primate species. It is assumed that intercalary structures conserving some of the properties of heterochromatin exists in human chromosomes in intercalary positions.     

Preferential breakage of sensitive regions of human chromosomes

Summary Whilst studying the chromosomes of the peripheral blood lymphocytes of normal controls and patients with lymphoproliferative disorders, two examples of preferential breakage of a sensitive chromosomal region were found. A patient with lymphocytic lymphoma had a sensitive region in a C9 chromosome coinciding with the secondary constriction. A healthy woman had one A2 chromosome showing an unusually located secondary constriction in which breakage sometimes occurred.     

Plasmodium falciparum: selenium-induced cytotoxicity to P. falciparum

The in vitro antimalarial activity of sodium selenite (NaSe) was investigated and the mechanism of its action was studied. NaSe had antimalarial activity against both the chloroquine-susceptible strain FCR-3 and chloroquine-resistant strain K-1 of Plasmodium falciparum. The shrunken cytoplasm of the parasite was observed in a smear 12 h after treatment with NaSe. Co-treatment with copper sulfate (CuSO(4)) in culture did not affect the antimalarial activity of NaSe, but NaSe cytotoxicity against the mammalian cell line Alexander was decreased significantly. The intracellular reduced glutathione level of parasitized red blood cells was decreased significantly by treatment with NaSe, and the decrease was consistent with their mortality. Treatment with NaSe had a strong inhibitory effect on plasmodial development, and NaSe cytotoxicity to human cells was decreased by co-treatment with CuSO(4). These results suggest that co-treatment with NaSe and CuSO(4) may be useful as a new antimalarial therapy.     

Karyotype comparison between P. chabaudi and P. falciparum: analysis of a P. chabaudi cDNA containing sequences highly repetitive in P. falciparum.

The molecular karyotypes of P. chabaudi and P. falciparum have been compared by pulse field gradient electrophoresis. P. chabaudi has 3 extra chromosomes in the 750-2000 Kb range although the overall number appears to be 14 as is the case for P. falciparum. The chromosomal location of the rRNA genes has been determined for P. chabaudi together with that of a 24 Kd antigen gene. The corresponding cDNA 443 may code for a protein unusually rich in tyrosine and contains sequences highly repetitive in P. falciparum.     

Patterns of meiotic double-strand breakage on native and artificial yeast chromosomes

The preferred positions for meiotic double-strand breakage were mapped on Saccharomyces cerevisiae chromosomes I and VI, and on a number of yeast artificial chromosomes carrying human DNA inserts. Each chromosome had strong and weak double-strand break (DSB) sites. On average one DSB-prone region was detected by pulsed-field gel electrophoresis per 25 kb of DNA, but each chromosome had a unique distribution of DSB sites. There were no preferred meiotic DSB sites near the telomeres. DSB-prone regions were associated with all of the known ”hot spots” for meiotic recombination on chromosomes I, III and VI. Received: 19 March 1996; in revised form: 26 July 1996 / Accepted: 18 August 1996     

Plasmodium falciparum: isolation and characterization of a 55-kDa protease with a cathepsin D-like activity from P. falciparum

Native electrophoresis followed by imprint digest method using hemoglobin as substrate allowed the detection of parasite hemoglobinase activity at acidic pH (3.9 to 5). This protease was inhibited specifically by pepstatin A and insensitive to other protease inhibitors. The molecular weight determination using modified SDS-PAGE followed by imprint digest method, demonstrated a single area of activity at 55-58 kDa, similar to cathepsin D characterized in eucaryotic cells. The parasitic origin has been shown by radiolabeling experiments with [35S]-methionine. The 55-kDa protein was immunoprecipitated by a rabbit anti-cathepsin D serum.     

Altered linkage values in phage P22-mediated transduction caused by distant deletions or insertions in donor chromosomes

Summary The effects of distant deletions or insertions in the Salmonella typhimurium donor strains on P22-mediated cotransducibility of genetic markers was studied. We found that deletions of histidine operon, unit 44 of the chromosome map, changed the linkage of markers purF and aroC (unit 49) and pyrF and trpA (unit 34). They did not change the linkage of more distant markers pyrE and cysE. The effect of three types of insertions was examined. The donor strains carried F factor, Tn10 transposon or pi-his duplication inserted close to histidine operon. These insertions caused alteration of purF-aroC linkage while pyrF-trpA cotransduction values were not affected. These data show that the effect of the chromosome rearrangements extends to at least 5% of S. typhimurium chromosome length and may reach as much as 10% of it. Our results are in agreement with the model of Chelala and Margolin (1974) concerning formation of transduction particles. They indicate that the cotransducibility changes caused by deletions or insertions extent further than it might have been expected from previous reports.     

Isolating large nested deletions in bacterial and P1 artificial chromosomes by in vivo P1 packaging of products of Cre-catalysed recombination between the endogenous and a transposed loxP site.

A general approach for isolating large nested deletions in P1 artificial chromosomes (PACs) and bacterial artificial chromosomes (BACs) by retrofitting with a loxP site-containing Tn10 mini-transposon is described. Cre-mediated recombination between the loxP site existing in these clones and one introduced by transposition leads to deletions and inversions of the DNA between these sites. Large deletions are selectively recovered by transducing the retrofitted PAC or BAC clones with P1 phage. The requirement that both loxP sites in the cointegrate be packaged into a P1 head ensures that only large deletions are rescued. PCR analyses identified these deletions as products of legitimate recombination between loxP sites mediated by Cre protein. BACs produce deletions much more efficiently than PACs although the former cannot be induced to greater than unit copy in cells. Mammalian cell-responsive antibiotic resistance markers are introduced as part of the transposon into genomic clone deletions for subsequent functional analysis. Most importantly, the loxP site retrofitting and P1 transduction can be performed in the same bacterial host containing these clones directly isolated from PAC or BAC libraries. These procedures should facilitate physical and functional mapping of genes and regulatory elements in these large plasmids.     

P. falciparum msp1 and msp2 genetic diversity in P. falciparum single and mixed infection with P. malariae among the asymptomatic population in Southern Benin

Plasmodium falciparum and Plasmodium malariae infections are prevalent in malaria-endemic countries. However, very little is known about their interactions especially the effect of P. malariae on P. falciparum genetic diversity. This study aimed to assess P. falciparum genetic diversity in P. falciparum and mixed infection P. falciparum/P. malariae isolates among the asymptomatic populations in Southern Benin. Two hundred and fifty blood samples (125 of P. falciparum and 125 P. falciparum/P. malariae isolates) were analysed by a nested PCR amplification of msp1 and msp2 genes. The R033 allelic family was the most represented for the msp1 gene in mono and mixed infection isolates (99.2% vs 86.4%), while the K1 family had the lowest frequency (38.3% vs 20.4%). However, with the msp2 gene, the two allelic families displayed similar frequencies in P. falciparum isolates while the 3D7 allelic family was more represented in P. falciparum/P. malariae isolates (88.7%). Polyclonal infections were also lower (62.9%) in P. falciparum/P. malariae isolates (p < 0.05). Overall, 96 individual alleles were identified (47 for msp1 and 49 for msp2) in P. falciparum isolates while a total of 50 individual alleles were identified (23 for msp1 and 27 for msp2) in P. falciparum/P. malariae isolates. The Multiplicity of Infection (MOI) was lower in P. falciparum/P. malariae isolates (p < 0.05). This study revealed a lower genetic diversity of P. falciparum in P. falciparum/P. malariae isolates using msp1 and msp2 genes among the asymptomatic population in Southern Benin.     

Heritability of P. falciparum and P. vivax Malaria in a Karen Population in Thailand

The majority of studies concerning malaria host genetics have focused on individual genes that confer protection against rather than susceptibility to malaria. Establishing the relative impact of genetic versus non-genetic factors on malaria infection and disease is essential to focus effort on key determinant factors. This relative contribution has rarely been evaluated for Plasmodium falciparum and almost never for Plasmodium vivax. We conducted a longitudinal cohort study in a Karen population of 3,484 individuals in a region of mesoendemic malaria, Thailand from 1998 to 2005. The number of P. falciparum and P. vivax clinical cases and the parasite density per person were determined. Statistical analyses were performed to account for the influence of environmental factors and the genetic heritability of the phenotypes was calculated using the pedigree-based variance components model. The genetic contribution to the number of clinical episodes resulting from P. falciparum and P. vivax were 10% and 19% respectively. There was also moderate genetic contribution to the maximum and overall parasite trophozoite density phenotypes for both P. falciparum (16%&16%) and P. vivax (15%&13%). These values, for P. falciparum, were similar to those previously observed in a region of much higher transmission intensity in Senegal, West Africa. Although environmental factors play an important role in acquiring an infection, genetics plays a determinant role in the outcome of an infection with either malaria parasite species prior to the development of immunity.     

恶性疟原虫多抗原表位基因的融合与表达

本研究以霍乱毒素B亚基(CT-B)基因为载体,构建了含不同抗原表位的恶性疟原虫的融合基因CTB/ATE和CTB/AWTE。前者除含有恶性疟原虫裂殖子表面主要抗原表位杂合多肽基因SPf66外,还含有很强的T辅助细胞表位CST3和Tc细胞表位;后者在此基础上将我国发现的B细胞表位NKNDD基因经8次串联后融合其中、两种形式的融合基因经测序正确后转入大肠杆菌TK1046中,产量分别为10mg/L及5mg/L。表达产物CTB/AWTE经亲和层析纯化的双抗夹心ELISA测定表明,该融合蛋白在保留了与抗CTB抗体结合的同时,与抗NKNDD单抗的结合效价达1∶8000。