Evolutionary divergence of human chromosome 9 as revealed by the position of the ABL protooncogene in higher primates

Attempts to solve the fundamental questions regarding the descent of man are dogged by superstitions and unexamined orthodoxies. The origin of humans, established a decade ago based upon cytological analysis of ape chromosomes, continues to be called into question. Although molecular methods have provided a framework for tracing the paths of human evolution, conclusive evidence remains elusive. We have used a single ABL gene probe derived from human chromosome 9 to assess the direction of change in the equivalent ape chromosomes. This approach has resulted in a few surprises which again challenge the prevailing view of early primate evolution based solely on chromosome banding patterns. The ABL protooncogene is present on human chromosome 9 at band q34. Similar DNA sequences presumed to represent an ABL gene, are present on chromosome 11 in chimpanzee (Pan troglodytes) but at a different relative location, indicating that the mechanism of the origin of human chromosome 9 is far more complex than has previously been suggested. Nevertheless, in gorilla (Gorilla gorilla) and orangutan (Pongo pygmaeus), the equivalent to human chromosome band 9 q34 is apparently located on chromosome 13 at a putative telomeric position and no discernible differences could be established. Despite the presence of the ABL protooncogene on human equivalent ape chromosomes, molecular systematics will continue to generate enigmas in the evolutionary context until the entire genome is sequenced.     

Silver nanoparticles as a potential antimicrobial additive for weaned pigs

Three experiments were carried out in order to determine the potential of silver nanoparticles as an additive in diets for weanling pigs. In Experiment 1, ileal contents of 4 pigs weaned 7 days before were incubated in vitro for 4 h at 37 °C with 0, 25, 50 and 100 μg Ag/g. Metallic silver (in colloidal form) linearly reduced coliforms (P=0.003) and lactobacilli (P=0.041) concentration, but did not affect the lactobacilli proportion compared with the control. In Experiment 2, three groups of 5 weaned pigs were given a diet with 0, 20 or 40 mg Ag/kg. The second week after weaning daily growth of pigs increased linearly (P=0.007) with the dose of silver nanoparticles. A trend (P=0.073) for a linear reduction in the ileal concentration of coliforms was observed by culture counts, but lactobacilli remained unaffected. There were no differences among treatments in the ileal concentration of coliforms or lactobacilli measured by FISH. However, the concentration of total bacteria (P=0.010) and Atopobium (P=0.001) decreased at a decreasing rate. No differences were detected in the other bacterial groups tested, except for a lowest concentration of the Clostridium perfringens/Clostridium histolyticum group in 20 mg Ag/kg (P=0.012). No treatment effect was detected in histological examination of ileal mucosa. In Experiment 3, productive performance and silver retention in tissues with 0, 20 or 40 mg Ag/kg diet were studied with 6 lots of 4 piglets per treatment in five weeks after weaning. Feed intake was highest in treatment with 20 mg Ag/kg (P<0.05). No effect was observed on apparent digestibility coefficients. After 5 weeks, there was no silver retention in skeletal muscles or kidneys, but it was observed, although in minimal proportions, in liver (1.354 and 2.445 μg Ag/g dry liver for 20 and 40 mg Ag/kg). The potential effect of metallic silver as a dietary additive on intake and growth of weaned piglets could be mediated through its antimicrobial properties, either against certain bacterial groups or reducing the microbial load of the small intestine; however, other beneficial effects over the host metabolism cannot be discarded.     

Comparison of the distribution of the repetitive DNA sequences in three variants of Cucumis sativus reveals their phylogenetic relationships

Repetitive DNA sequences with variability in copy number or/and sequence polymorphism can be employed as useful molecular markers to study phylogenetics and identify species/chromosomes when combined with fluorescence in situ hybridization(FISH).Cucumis sativus has three variants,Cucumis sativus L.var.sativus,Cucumis sativus L.var.hardwickii and Cucumis sativus L.var.xishuangbannesis.The phylogenetics among these three variants has not been well explored using cytological landmarks.Here,we concentrate on...     

Evaluation of the environmental specificity of Fluorescence In Situ Hybridization (FISH) using Fluorescence-Activated Cell Sorting (FACS) of probe (PSE1284)-positive cells extracted from rhizosphere soil

We explicitly tested for the first time the ‘environmental specificity’ of traditional 16S rRNA-targeted Fluorescence In Situ Hybridization (FISH) through comparison of the bacterial diversity actually targeted in the environment with the diversity that should be exactly targeted (i.e. without mismatches) according to in silico analysis. To do this, we exploited advances in modern Flow Cytometry that enabled improved detection and therefore sorting of sub-micron-sized particles and used probe PSE1284 (designed to target Pseudomonads) applied to Lolium perenne rhizosphere soil as our test system. The 6-carboxyfluorescein (6-FAM)-PSE1284-hybridized population, defined as displaying enhanced green fluorescence in Flow Cytometry, represented 3.51 ± 1.28% of the total detected population when corrected using a nonsense (NON-EUB338) probe control. Analysis of 16S rRNA gene libraries constructed from Fluorescence Activated Cell Sorted-recovered fluorescent populations (n = 3), revealed that 98.5% (Pseudomonas spp. comprised 68.7% and Burkholderia spp. 29.8%) of the total sorted population was specifically targeted as evidenced by the homology of the 16S rRNA sequences to the probe sequence. In silico evaluation of probe PSE1284 with the use of RDP-10 probeMatch justified the existence of Burkholderia spp. among the sorted cells. The lack of novelty in Pseudomonas spp. sequences uncovered was notable, probably reflecting the well-studied nature of this functionally important genus. To judge the diversity recorded within the FACS-sorted population, rarefaction and DGGE analysis were used to evaluate, respectively, the proportion of Pseudomonas diversity uncovered by the sequencing effort and the representativeness of the Nycodenz^® method for the extraction of bacterial cells from soil.     

Telomere length analysis of human mesenchymal stem cells by quantitative PCR

Human mesenchymal stem cells (hMSCs) have attracted much attention for tissue repair and wound healing because of their self-renewal capacity and multipotentiality. In order to mediate an effective therapy, substantial numbers of cells are required, which necessitates extensive sub-culturing and expansion of hMSCs. Throughout ex vivo expansion, the cells undergo telomere shortening, and critically short telomeres can trigger loss of cell viability. Telomeres are nucleoprotein structures that cap the ends of chromosomes, and serve to protect the DNA from the degradation which occurs due to the end-replication problem in all eukaryotes. As hMSCs have only a finite ability for self-renewal like most somatic cells, assaying for telomere length in hMSCs provides critical information on the replicative capacity of the cells, an important criterion in the selection of hMSCs for therapy. Telomere length is generally quantified by Southern blotting and fluorescence in situ hybridization, and more recently by PCR-based methods. Here we describe the quantification of hMSC telomere length by real-time PCR; our results demonstrate the effect of telomere shortening on the proliferation and clonogenicity of hMSCs. Thus, this assay constitutes a useful tool for the determination of relative telomere length in hMSCs.     

Chromosome mapping of ribosomal genes and histone H4 in the genus Radacridium (Romaleidae)

In this study, two species of Romaleidae grasshoppers, Radacridium mariajoseae and R.nordestinum, were analyzed after CMA₃/DA/DAPI sequential staining and fluorescence in situ hybridization (FISH) to determine the location of the 18S and 5S rDNA and histone H4 genes. Both species presented karyotypes composed of 2n = 23, X0 with exclusively acrocentric chromosomes. CMA₃⁺ blocks were detected after CMA₃/DA/DAPI staining in only one medium size autosome bivalent and in the X chromosome in R. mariajoseae. On the other hand, all chromosomes, except the L1 bivalent, of R. nordestinum presented CMA₃⁺ blocks. FISH analysis showed that the 18S genes are restricted to the X chromosome in R. mariajoseae, whereas these genes were located in the L₂, S₉ and S₁₀ autosomes in R. nordestinum. In R. mariajoseae, the 5S rDNA sites were localized in the in L₁ and L₂ bivalents and in the X chromosome. In R. nordestinum, the 5S genes were located in the L₂, L₃, M₄ and M₅ pairs. In both species the histone H4 genes were present in a medium size bivalent. Together, these data evidence a great variability of chromosome markers and show that the 18S and 5S ribosomal genes are dispersed in the Radacridium genome without a significant correlation.     

Combined DNA-RNA Fluorescent In situ Hybridization (FISH) to Study X Chromosome Inactivation in Differentiated Female Mouse Embryonic Stem Cells

Fluorescent in situ hybridization (FISH) is a molecular technique which enables the detection of nucleic acids in cells. DNA FISH is often used in cytogenetics and cancer diagnostics, and can detect aberrations of the genome, which often has important clinical implications. RNA FISH can be used to detect RNA molecules in cells and has provided important insights in regulation of gene expression. Combining DNA and RNA FISH within the same cell is technically challenging, as conditions suitable for DNA FISH might be too harsh for fragile, single stranded RNA molecules. We here present an easily applicable protocol which enables the combined, simultaneous detection of Xist RNA and DNA encoded by the X chromosomes. This combined DNA-RNA FISH protocol can likely be applied to other systems where both RNA and DNA need to be detected.     

Some aspects of the ecology of the Groot Letaba River in the Northern Province,South Africa

The aim of this pilot study was to evaluate the current ecological status of the Groot Letaba River and to compare this information with historical data. The objective was to determine the effects of various impacts on the fish populations of the river. This was done by analysing the water quality and by considering the effect of weirs and dams, as well as various illegal angling activities, on the fish community.

The Groot Letaba River is not highly polluted and the decline in its flow seems to be the greatest threat to the system. During a preliminary study to develop the river's resource potential, it was stated that the annual water allocation from Tzaneen Dam was 103.9 million m³/annum for irrigation, 8.4 million m³/annum for domestic and industrial use and 14.7 million m³/annum for environmental purposes. However, the yield from Tzaneen Dam was only 98 million m³/annum, suggesting that more water had been allocated than was available. As a result only 20% of the simulated natural flow is observed at Letaba Ranch Weir at the lower end of the river.

Over the past few years many weirs and dams, none of which have fishways, have been constructed in the Groot Letaba River, impacting on the flow regime and on the migration potential of many fish species. Tiger fish (Hydrocynus vittatus) and the largescale yellowfish (Barbus marequensis) are two of the more prominent species influenced negatively by these barriers. This problem is aggravated by the illegal netting of fish stranded below these barriers during their spawning migrations.     

Physical mapping of three minisatellite sequences in the Atlantic salmon (Salmo salar) genome

We have determined the localization of three minisatellite loci SsBglIIL.6 , SsBglIIU.20 and SsPstL.26 in the Atlantic salmon ( Salmo salar ) genome by fluorescent in situ hybridization (FISH). FISH analysis of the SsBglIIL.6 and SsBglIIU.20 minisatellites revealed that they are both located in single chromosome pairs allowing their direct identification; in contrast, the SsPstIL.26 probe hybridizes to four different chromosomal pairs. The analysis of chromosomal location of minisatellite sequences could be very useful for studying structural changes that have taken place during chromosome evolution in the karyotype of Salmo salar .