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Monoclonal antibodies raised against pre-migratory neural crest reveal population heterogeneity during crest development.

In order to address the problem of when heterogeneity arises within premigratory and early migratory neural crest cell populations, mouse monoclonal antibodies were raised against quail premigratory neural crest. Due to the limited availability of immunogen an intrasplenic route for immunization was used. Three monoclonal antibodies (referred to as LH2D4, LH5D3 and LH6C2) were subsequently isolated which recognized subpopulations in 24 h cultures of both quail and chick mesencephalic and trunk neural crest in immunocytochemical studies. Subsequent investigations using a range of six antibodies, including LH2D4, LH5D3 and LH6C2, showed that population heterogeneity (which was not cell cycle related) could be detected as early as 15 h following mesencephalic crest explantation, a stage at which all the neural crest cells were morphologically identical. However, premigratory neural crest from the same axial level of origin was homogeneous, as judged by immunoreactivity patterns with these antibodies. Significant differences were found in the proportion of immunoreactive cells between populations of mesencephalic and trunk neural crest cultures. Double immunofluorescence studies revealed the existence of at least four separate cell populations within individual crest cultures, each identified by their unique antibody reactivity pattern, thus providing some insight into the underlying complexity of subpopulation composition within the neural crest. Immunocytochemical studies on quail embryos from stages 7-22 showed that the epitopes detected by LH2D4, LH5D3 and LH6C2 were not necessarily confined to the neural crest or to cells of crest derivation. All three epitopes displayed a spatiotemporal regulation in their expression during early avian ontogeny. Since the differential epitope expression described in this investigation was detectable as early as 15 h after premigratory neural crest explantation, took place in vitro in the absence of any other cell type and changed progressively with time, we conclude that a certain degree of population heterogeneity can be generated very early in neural crest ontogeny and independently of the tissue interactions that normally ensue in vivo.

Animals

Population heterogeneity in Helicobacter pylori PMSS1 shapes variable mouse infectivity: derivation of the homogeneous reference strain PMSS2.

UNLABELLED: Experimental infection models are widely used to investigate host-microbe interactions, often under the assumption that bacterial populations are genetically uniform. Here, we examined population heterogeneity in the widely used Helicobacter pylori strain PMSS1 and its relationship to variation in mouse infectivity. Single-colony isolates derived from PMSS1 displayed substantial differences in colonization efficiency, indicating that pre-existing variation within the population contributes to infection outcomes. To distinguish the effects of initial population heterogeneity from changes arising during infection, we analyzed PMSS2, a genetically homogeneous reference strain derived from PMSS1 that exhibited consistent infection phenotypes across independently isolated clones. Comparative genomic analysis of isolates recovered from infected mice revealed differences in the extent and patterns of genomic variation between PMSS1- and PMSS2-derived populations. These results demonstrate that variability in infection outcomes can arise from pre-existing heterogeneity within bacterial populations and highlight the importance of considering population composition when interpreting experimental infection studies. IMPORTANCE: Animal infection models are widely used to study how bacterial pathogens cause disease and change during infection. These studies often assume that the bacteria used for infection are genetically uniform. Our study shows that this assumption may not always hold. We found that a commonly used Helicobacter pylori strain contains hidden genetic diversity that leads to large differences in how well bacteria infect mice. By comparing this strain with a genetically uniform derivative, we show how differences present before infection can shape infection outcomes and influence the genetic changes observed during infection. Our findings highlight the importance of considering starting population diversity when interpreting experimental infection studies and are broadly relevant to research on microbial pathogenesis.

Helicobacter pylori

[Sh. flexneri population heterogeneity according to the cellular fatty acid makeup].

The composition of fatty acids in the cells of 100 clones of Sh. flexneri, strain 9054, isolated from a dysentery patient was studied by the method of gas chromatography. Fatty acid composition in 75 clones was similar to the fatty acid composition of the initial culture: 25 clones differed to a variable degree from the prevailing group of clones in the content of unsaturated and cyclopropane fatty acids and, to a considerably lesser extent, in the content of palmitic acid and a number of other saturated fatty acids. After the change of the culture medium (agar and broth) the differences in the fatty acid composition of the clones could still be observed. The data obtained in this study indicate the heterogeneity of Shigella population in the composition of cellular fatty acids.

Chromatography, Gas

Isolation of a heterogeneous population of temperature-sensitive mutants of measles virus from persistently infected human lymphoblastoid cell lines.

Two human lymphoblastoid B-cell lines, WI-L2 and 8866, were infected with the Edmonston strain of measles virus at a multiplicity of infection of 10(-6), and stable persistent infections were established. By immunofluorescence and electron microscopy, the vast majority of cells from both cell lines were expressing viral antigens and releasing virion-like particles. However, very little infectious virus could be detected at 37 degrees C, either by an infectious centers assay or by titration of supernates from persistently infected cultures. When cultures were shifted to 31 degrees C, the cells released a population of virus that was temperature-sensitive. Clonal analysis of supernatant virus at 31 degrees C revealed a highly heterogeneous population of temperature-sensitive mutants, differing in plating efficiency ratios, thermolability, and antigen production at the nonpermissive temperature. Factors such as interferon, defective interfering particles, and extracellular virus do not appear to be important in maintaining the persistent carrier state. These studies have important implications for persistent infections of lymphoid cells in vivo, and the slow neurological diseases associated with measles, subacute sclerosing panencephalitis, and multiple sclerosis.

Antibodies, Viral

Genetics of the HL-A system in a venezuelan heterogeneous population.

HL-A antigen and gene frequencies of 206 unrelated adults living in the cities of Caracas and Maracaibo have shown that this heterogeneous Venezuelan population sample possess 11 specificities of the first (LA) HL-A series and 15 specificities of the second (Four) series. HL-A2, HL-A9, HL-A5 and W5 showed frequencies higher than those observed in caucasoid populations. The most frequent haplotypes were 2, W5; 2, 5; 9, 12; 2, 12 and 2, X2. No outstandingly high value for gametic association between the alleles of the 2 HL-A series was observed, but haplotypes formed by antigens with dissimilar frequencies in Caucasoids, Negroids and American Indian tribes have shown statistically significant D values. Genetic distance calculated using the HL-A system alone showed that this population is closer to the average Caucasoid and Negroid population tested at the 5th International Histocompatibility Workshop than to 2 American Indian tribes living in the same country.

Female

Population heterogeneity in human sperm DNA content.

The range of variability in the population distribution characteristics of sperm DNA content was surveyed in a selected group of donors. The donors included identical twins, carriers of chromosomal translocations, and oligospermics. The DNA content of sperm was measured individually in an automated cytofluorometer. In contrast to normal donors with high sperm counts and relatively constant modal values of sperm DNA, the balanced carriers of translocations were oligospermic and showed wide dispersion of modal values. This is an unexpected finding, since reciprocal events in the alternate form of segregation, which occurs predominantly in male carriers of balanced translocations, should not alter modal values of sperm DNA nor should they be associated with oligospermia. It appears that sperm DNA content in oligospermia, independent of its association with translocation, varies over a wide range. Thus, it is likely that genetic factor(s) unrelated to chromosome translocation control sperm DNA content.

DNA

[Mouse Lewis carcinoma cell population heterogeneity and sensitivity to cyclophosphamide].

Cell kinetics in the large Lewis carcinoma growing in mice subcutaneously was characterized by the following parameters: at the tumor periphery -tc=11.3 hr, tG2=1.4 hr, tS=8.8hr, tG1=1.1 hr, LI=47%, Pc=61.8%; at the tumour center - 20.7 hr, 2.9 hr, 16.0 hr, 1.8 hr,31.2%, resp. Injection of cyclophosphamide induced degeneration of all the cells in slowly growing central part of the tumour, and only of part of cells on the fast proliferating tumour periphery. It is suggested that in the central part of the tumour there are no stem cells, and that this region regenerates when the transition of cells from the tumour periphery is arrested by cyclophosphamide.

Animals

[Frequency of Australia antigen in a heterogenous population and estimation of 3 various methods of research].

4,000 samples of serum for Australia antigen research were examined by electrosineresis. From these 4,000 serums two groups were formed afterwards: the first formed by 2,466 samples which were examined also by immunodiffusion; the second formed by 1,093 samples which were examined also by latex test; no common serum was in the two groups at the same time. AU frequence in this population sample is within the high limits of the national average. A comparison of the used methods showed the following: immunodiffusion given the smallest number of positive results; electrosineresis given other positive results besides those found out by immunodiffusion; latex test given the largest number of positive results, but greatly discordant from electrosineresis. The achieved results are examined.

Chemical Phenomena

Consanguinity analysis in heterogeneous populations.

Consanguinity analysis can be performed in populations comprising collections of genetic isolates, and the resulting estimates can be valid and useful in phenotypes caused by numerous recessive genes, such as mental retardation and congenital nerve deafness. Maximum likelihood methods are presented for estimating gene frequency and proportions of homozygous cases of morbid phenotypes in such populations.

Consanguinity

[Plasmid heterogeneity in populations of Yersinia pestis strains].

Yersinia pestis strains with the typical plasmid patterns were shown to have the heterogenic populations. Heterogeneity is increased by cultivation passages in artificial nutrient media and is manifested in plasmid elimination within several clones, plasmid integration into the chromosome, appearance of auxiliary plasmids or the ones with increased molecular masses. Passages of strains in experimental animals result in populations homogeneity with the typical plasmid patterns within all clones tested. The clones having changed the plasmid content and selected from heterogenic populations pertain their properties when cultivated in nutrient media and passaged in experimental animals.

Electrophoresis, Agar Gel

[Mechanism of the heterogeneity of a staphylococcal population with respect to methicillin resistance].

The study of the staphylococcal population heterogeneity with respect to methicillin resistance by 2 methods revealed different numbers of the resistant cells in the population. Thus, when the microbial suspension was plated on an agarized medium with methicillin (50 gamma/ml), only 0.0007--0.0005 per cent of the resistant cells were found. When the colonies were replicated from a medium without methicillin to a medium containing methicillin (50 gamma/ml), 84.3--97.3 per cent of the resistant microbial cells were found in the population of the same strains. The main mechanism in the heterogeneity of the staphylococcal population with respect to methicillin resistance was impairement of the phenotype manifestation of the antibiotic resistance under definite conditions.

Culture Media

An expression for the calculation of relative affinities of antibody-ligand interactions.

An expression is derived which allows the calculation of K-a without a value for total binding sites of antibody. The expression yields unambigous K-a values for homogeneous populations of antibody molecules. When heterogeneous populations are used, the value of calculated K-a rises as the fraction of sites bound falls. This expression should be useful in the determination of relative affinity with data obtained from Farr assays of the binding of anti-hapten antibody to hapten. Since values for total binding sites are unnecessary, this equation should be expecially applicable for the study of heterogeneous populations of relatively low affinity antibodies present at low concentrations.

Animals