Protection of Chicken Embryos by Viridans Streptococci Against the Lethal Effect of Staphylococcus aureus

Chicken embryos were used to investigate the mechanism by which viridans streptococci inhibit the growth of pathogenic staphylococci. Ten-day-old embryonated eggs were infected allantoically. At a concentration of 1.8 x 10(2) colony-forming units (CFU) of viridans streptococci, the percentage of fatalities was less than 10%. There was 80% fatality with 8 x 10(1) CFU of Staphylococcus aureus strain 502A and 100% when a 100-fold increase in concentration was used. An inoculum size of 10(2) to 10(3) CFU of viridans streptococci was chosen to protect the embryos against the lethal effect of strain 502A when challenged 24 h later. The survival after challenging at 4 days was 93% in protected eggs and 37% in unprotected eggs. Chicken embryos receiving heat-killed viridans and challenged with strain 502A when examined after 4 days did not demonstrate a protective effect. This protection of embryonated eggs could not be transferred by administration of sterile filtrate of allantoic fluid in which protecting strain was grown. The experimental infection of embryonated eggs has demonstrated that prior allantoic infection with viridans streptococci affords significant protection against subsequent challenge with virulent staphylococci.     

Enhanced saliva-mediated bacterial aggregation and decreased bacterial adhesion in caries-resistant versus caries-susceptible individuals.

A study of saliva-mediated aggregation and adhesion has been carried out in a group of caries-resistant (CR) and caries-susceptible (CS) individuals. The submandibular saliva of the CS group had a much greater potency, as determined by dilution, in promoting adherence to hydroxyapatite beads than did the saliva of CR group. In contrast, the CR group demonstrated a twofold enhancement of saliva-mediated aggregation compared with the CS group. These observations support the hypothesis that saliva-mediated aggregation and adherence are important factors in caries resistance.     

Inactivation of the peroxisomal ABCD2 transporter in the mouse leads to late-onset ataxia involving mitochondria, Golgi and endoplasmic reticulum damage

ATP-binding cassette (ABC) transporters facilitate unidirectional translocation of chemically diverse substances, ranging from peptides to lipids, across cell or organelle membranes. In peroxisomes, a subfamily of four ABC transporters (ABCD1 to ABCD4) has been related to fatty acid transport, because patients with mutations in ABCD1 (ALD gene) suffer from X-linked adrenoleukodystrophy (X-ALD), a disease characterized by an accumulation of very-long-chain fatty acids (VLCFAs). Inactivation in the mouse of the abcd1 gene leads to a late-onset neurodegenerative condition, comparable to the late-onset form of X-ALD [Pujol, A., Hindelang, C., Callizot, N., Bartsch, U., Schachner, M. and Mandel, J.L. (2002) Late onset neurological phenotype of the X-ALD gene inactivation in mice: a mouse model for adrenomyeloneuropathy. Hum. Mol. Genet., 11, 499-505.]. In the present work, we have generated and characterized a mouse deficient for abcd2, the closest paralog to abcd1. The main pathological feature in abcd2-/- mice is a late-onset cerebellar and sensory ataxia, with loss of cerebellar Purkinje cells and dorsal root ganglia cell degeneration, correlating with accumulation of VLCFAs in the latter cellular population. Axonal degeneration was present in dorsal and ventral columns in spinal cord. We have identified mitochondrial, Golgi and endoplasmic reticulum damage as the underlying pathological mechanism, thus providing evidence of a disturbed organelle cross-talk, which may be at the origin of the pathological cascade.     

C7 complement deficiency in an Israeli Arab village

Deficiencies of terminal complement components, particularly the latter ones, are often detected because of increased susceptibility to Neisserial infections. Herein we document the first report of C7 deficiency among a highly inbred Arab population living in the lower Galilee region of Israel. Both biochemical and molecular analysis were performed on samples from infected survivors and parents of children who succumbed to Neisserial infections in a 4-year period. Only the index case who suffered recurrent infections and a sibling who had not suffered an infection during the outbreak were found to be C7-deficient. The mutation was found to be the one previously described to be prevalent among Israeli Jews of Moroccan ancestry (mutation G1135C). The implications of this finding are discussed in the context of family pedigree, the protective effect of complement deficiency, and the clinical outcome.     

Isolation and characterization of human apatite-inducing aortic proteolipid

A proteolipid, capable of inducing hydroxyapatite in vitro, can be isolated from human aortic calcified lesions, noncalcified lesions, and nonlesion tissue obtained at autopsy. Analysis of the amino acid composition after acid hydrolysis showed all nucleators to be high in the hydrophobic amino acids glycine and alanine. Estimation of the molecular weight of the nucleator, extracted from calcified lesions, by gel filtration and lipid content showed a minimum molecular weight of 469,000 Da. All nucleators were characterized by the presence of acidic phospholipids which may have a role in the mechanism of calcification.     

Improved DNA markers for efficient analysis of fragile X families

We report the characteristics of two new probes that detect BclI RFLPs useful for analysis of fragile X families. With these two probes and a single blot, 34% of women are heterozygous both for the proximal marker DXS105 (closer to the fragile X locus than the factor IX gene) and for the distal markers DXS52 or the factor VIII gene. Combined with the analysis of previously described polymorphic markers, it is possible to have a majority of families fully informative for flanking markers using a limited number of probes and restriction digests.     

Alternative splicing of exon 14 determines nuclear or cytoplasmic localisation of fmr1 protein isoforms

Impaired expression of the FMR1 gene is responsible for the fragile X mental retardation syndrome. The FMR1 gene encodes a cytoplasmic protein with RNA-binding properties. Its complex alternative splicing leads to several isoforms, whose abundance and specific functions in the cell are not known. We have cloned in expression vectors, cDNAs corresponding to several isoforms. Western blot comparison of the pattern of endogenous FMR1 proteins with these transfected isoforms allowed the tentative identification of the major endogenous isoform as ISO 7 and of a minor band as an isoform lacking exon 14 sequences (ISO 6 or ISO 12), while some other isoforms (ISO 4, ISO 5) were not expressed at detectable levels. Surprisingly, in immunofluorescence studies, the transfected splice variants that exclude exon 14 sequences (and have alternate C-terminal regions) were shown to be nuclear. Such differential localisation was however not seen in subcellular fractionation studies. Analysis of various deletion mutants suggests the presence of a cytoplasmic retention domain encoded in exon 14 and of a nuclear association domain encoded within the first eight exons that appear however to lack a typical nuclear localisation signal.      

Three families with high expression of a fragile site at Xq27.3, lack of anomalies at the FMR-1 CpG island, and no clear phenotypic association.

We report on 3 families where the presence and segregation at high frequency of a fragile Xq27.3 site is not associated with the mutations and methylation anomalies typically seen in the fragile X [Fra(X)] syndrome. In one family, a folate insensitive fragile site was associated with Robin sequence in the propositus. In a second family a fra(X) negative mother has two fra(X) positive sons (one mentally retarded and the other newborn). The third family presents very high expression of a folate sensitive site, unlinked to mental retardation, and was described previously by Voelckel et al. [1989]. The fragile sites in these or similar families recently described must be different from the one associated with the fra(X) syndrome. Their association with a clinical phenotype or with mental retardation is certainly not consistent, and may represent an ascertainment bias. However, the relatively high frequency with which they have been found among previously diagnosed fra(X) families suggests that, at least in some cases, the association with mental impairment may be significant. In two families reported up to now, a male with high expression of such variant fra(X) site failed to transmit it to his daughter, which may reflect an imprinting effect. Previously diagnosed families should be reinvestigated before direct DNA analysis is used for prenatal or carrier diagnosis of the fra(X) syndrome.     

Selection in blood cells from female carriers of the fragile X syndrome: inverse correlation between age and proportion of active X chromosomes carrying the full mutation.

We have studied the patterns of mutation and X inactivation in female carriers of a fragile X mutation, to try to correlate them with various phenotypic features. We used a simple assay, which shows simultaneously the size of the mutation, its methylation status, and DNA fragments that represent the normal active and inactive X chromosomes. We have observed an age dependent process, whereby the 'full' fragile X mutation is found preferentially on the inactive X in leucocytes in adult females, but not in younger ones. This phenomenon was not observed in female carriers of a 'premutation', who have little phenotypic expression. Preliminary data suggest that young females who show preferential presence of a full mutation on the active X in leucocytes may be at increased risk for mental retardation. We have also obtained preliminary evidence for an age dependent decrease in the somatic heterogeneity of full mutations, possibly owing to selection for smaller mutated fragments. If confirmed, the latter phenomenon might account for the known decrease with age of the expression of the fragile site. Our observations suggest that a gene whose expression is affected by the presence of a full mutation (possibly the FMR-1 gene) has a cell autonomous function in leucocytes, leading to a slowly progressive selection for cells where the mutation is on the inactive X chromosome.