Serologic survey for viral and bacterial infections in western populations of Canada lynx (Lynx canadensis)

A serologic survey for exposure to pathogens in Canada lynx (Lynx canadensis) in western North America was conducted. Samples from 215 lynx from six study areas were tested for antibodies to feline parvovirus (FPV), feline coronavirus, canine distemper virus, feline calicivirus, feline herpesvirus, Yersinia pestis, and Francisella tularensis. A subset of samples was tested for feline immunodeficiency virus; all were negative. For all other pathogens, evidence for exposure was found in at least one location. Serologic evidence for FPV was found in all six areas but was more common in southern populations. Also, more males than females showed evidence of exposure to FPV. Overall, prevalences were low and did not exceed 8% for any of the pathogens tested. This suggests that free-ranging lynx rarely encounter common feline pathogens.     

Nuclear RNA turnover

In mammalian cells, significantly more RNA is turned over in the nucleus than in the cytoplasm. However, only recently have we begun to understand the mechanisms and regulation of nuclear RNA decay.     

Asaia sp., an unusual spoilage organism of fruit-flavored bottled water

A gram-negative bacillus was isolated from a batch of fruit-flavored bottled water, which had spoiled as a result of bacterial overgrowth (>10(6) CFU/ml). The spoilage organism was extremely difficult to identify phenotypically and was poorly identified as Pasturella sp. (78.7% identification profile) employing the API 20NE identification scheme, which gave the profile 5040000. Molecular identification through PCR amplification of a partial region of the 16S rRNA gene followed by direct automated sequencing of the PCR amplicon allowed identification of the organism. Due to the sequence identity (100%) between the spoilage organism and a reference strain in GenBank, the spoilage isolate was considered to be an Asaia sp., a recently described genus and member of the acetic acid bacteria. This is the first report of Asaia sp. causing spoilage of a foodstuff and highlights the benefits of molecular identification techniques based on 16S rRNA gene sequences in the identification of unusual spoilage organisms.     

Interacting quantitative trait loci control loss of peripheral tolerance and susceptibility to autoimmune ovarian dysgenesis after day 3 thymectomy in mice

Day 3 thymectomy (D3Tx) results in a loss of peripheral tolerance mediated by CD4(+)CD25(+) T cells and the development of autoimmune ovarian dysgenesis (AOD) in A/J and (C57BL/6J x A/J)F(1) (B6AF(1)) hybrids but not in C57BL/6J mice. Quantitative trait loci (QTL) linkage analysis using a B6AF(1) x C57BL/6J backcross population verified Aod1 and Aod2 that were previously mapped as qualitative traits. Additionally, three new QTL intervals, Aod3, Aod4, and Aod5, on chromosomes 1, 2, and 7, respectively, influencing specific subphenotypes of AOD were identified. QTL linkage analysis using the A x B and B x A recombinant inbred lines verified Aod3 and confirmed linkage to H2. Aod5 colocalized with Mater, an ovarian-specific autoantigen recognized by anti-ovarian autoantibodies in the sera of D3Tx mice. Sequence analysis of Mater identified allelic, strain-specific splice variants between A/J and C57BL/6J mice making it an attractive candidate gene for Aod5. Interaction analysis revealed significant epistatic effects between Aod1-5 and Gasa2, a locus associated with susceptibility to D3Tx-induced autoimmune gastritis, as well as with H2. These results indicate that the QTL controlling D3Tx-induced autoimmune phenomenon are both organ specific and more generalized in their effects with respect to the genesis and activity of the immunoregulatory mechanisms maintaining peripheral tolerance.     

Non-traditional Alu evolution and primate genomic diversity

Alu elements belonging to the previously identified "young" subfamilies are thought to have inserted in the human genome after the divergence of humans from non-human primates and therefore should not be present in non-human primate genomes. Polymerase chain reaction (PCR) based screening of over 500 Alu insertion loci resulted in the recovery of a few "young" Alu elements that also resided at orthologous positions in non-human primate genomes. Sequence analysis demonstrated these "young" Alu insertions represented gene conversion events of pre-existing ancient Alu elements or independent parallel insertions of older Alu elements in the same genomic region. The level of gene conversion between Alu elements suggests that it may have a significant influence on the single nucleotide diversity within the genome. All the instances of multiple independent Alu insertions within the same small genomic regions were recovered from the owl monkey genome, indicating a higher Alu amplification rate in owl monkeys relative to many other primates. This study suggests that the majority of Alu insertions in primate genomes are the products of unique evolutionary events.     

Relationship of bursal anti-steroidogenic peptide (BASP) and histone H1

Previous in vitro research from our laboratory has demonstrated the existence of a protein purified from the chicken bursa of Fabricius, with potent antisteroidogenic and antiproliferative action on granulose cells and lymphocytes, respectively called Bursal anti-steroidogenic peptide (BASP). This protein is heat-labile, basic, and amino- and carboxy-terminus blocked. In highly purified form, the protein presents as a doublet on SDS-PAGE electrophoresis with an apparent MW of approximately 29 and approximately 32 kDa. Recently, Nanoflow Q-TOF Mass Spectrometry amino acid sequencing allowed determination of a convincing partial amino acid sequence, strongly suggesting a probable relationship of BASP with histone H1. Bursal cDNA expression library screening, using an antibody produced against BASP, also identified a clone with a sequence matching histone H1. Presently, we have demonstrated that SDS-PAGE electrophoresis of highly purified and bioactive BASP, and commercially-available calf thymus derived histone H1, produced similar doublets at approximately the same apparent MW, and that the electrophoretic profile of these 2 preparations were strikingly similar following 2 dimensional gel electrophoresis. The BASP doublet produced on SDS-PAGE was recognized by a commercially available monoclonal antibody recognizing a highly conserved region of histone H1. Furthermore, calf thymus histone H1 was found to suppress mitogen-stimulated chicken B-cell proliferation in a concentration-related manner, similar to the action of BASP. These data indicate that BASP shares substantial structural homology with, and may be identical to, histone H1.     

A phenotype-sensitizing Apoe-deficient genetic background reveals novel atherosclerosis predisposition loci in the mouse

Therapeutic intervention for atherosclerosis has predominantly concentrated on regulating cholesterol levels; however, these therapeutics are not efficacious for all patients, suggesting that other factors are involved. This study was initiated to identify mechanisms that regulate atherosclerosis predisposition in mice other than cholesterol level regulation. To do so we performed quantitative trait locus analysis using two inbred strains that each carry the atherosclerosis phenotype-sensitizing Apoe deficiency and that have been shown to have widely disparate predilection to atherosclerotic lesion formation. One highly significant locus on chromosome 10 (LOD = 7.8) accounted for 19% of the variance in lesion area independent of cholesterol. Two additional suggestive loci were identified on chromosomes 14 (LOD = 3.2) and 19 (LOD = 3.2), each accounting for 7-8% of the lesion variance. In all, five statistically significant and suggestive loci affecting lesion size but not lipoprotein levels were identified. Many of these were recapitulated in an independent confirmatory cross. In summary, two independently performed crosses between C57BL/6 and FVB/N Apoe-deficient mice have revealed several previously unreported atherosclerosis susceptibility loci that are distinct from loci linked to lipoprotein levels.     

Iron detoxification properties of Escherichia coli bacterioferritin. Attenuation of oxyradical chemistry

Bacterioferritin (EcBFR) of Escherichia coli is an iron-mineralizing hemoprotein composed of 24 identical subunits, each containing a dinuclear metal-binding site known as the "ferroxidase center." The chemistry of Fe(II) binding and oxidation and Fe(III) hydrolysis using H(2)O(2) as oxidant was studied by electrode oximetry, pH-stat, UV-visible spectrophotometry, and electron paramagnetic resonance spin trapping experiments. Absorption spectroscopy data demonstrate the oxidation of two Fe(II) per H(2)O(2) at the ferroxidase center, thus avoiding hydroxyl radical production via Fenton chemistry. The oxidation reaction with H(2)O(2) corresponds to [Fe(II)(2)-P](Z) + H(2)O(2) --> [Fe(III)(2)O-P](Z) + H(2)O, where [Fe(II)(2)-P](Z) represents a diferrous ferroxidase center complex of the protein P with net charge Z and [Fe(III)(2)O-P](Z) a micro-oxo-bridged diferric ferroxidase complex. The mineralization reaction is given by 2Fe(2+) + H(2)O(2) + 2H(2)O --> 2FeOOH((core)) + 4H(+), where two Fe(II) are again oxidized by one H(2)O(2). Hydrogen peroxide is shown to be an intermediate product of dioxygen reduction when O(2) is used as the oxidant in both the ferroxidation and mineralization reactions. Most of the H(2)O(2) produced from O(2) is rapidly consumed in a subsequent ferroxidase reaction with Fe(II) to produce H(2)O. EPR spin trapping experiments show that the presence of EcBFR greatly attenuates the production of hydroxyl radical during Fe(II) oxidation by H(2)O(2), consistent with the ability of the bacterioferritin to facilitate the pairwise oxidation of Fe(II) by H(2)O(2), thus avoiding odd electron reduction products of oxygen and therefore oxidative damage to the protein and cellular components through oxygen radical chemistry.