The interfacial strength of secondary osteons from the diaphysis of the Thoroughbred equine third metacarpal was evaluated using the fiber pushout test. The pushout was performed on 300-500 microm sections of 4x4x15 mm bone blocks machined from four anatomic regions of the cortex. Pushout strength was evaluated from proximal to distal location within the diaphysis on four osteon types classified under polarized light on adjacent histologic sections from each block. The shear strength of the interfaces were estimated from shear lag theory. Differences were found in the interfacial strength of osteons based on appearance under polarized light with bright field having the highest interfacial strength (40.3 MPa). The lowest strength was found in the dark field osteons (22.8 MPa). The dorsal region had the highest shear strength and toughness compared to all other regions. The cement line and interlamellar interfaces are similar in strength, but exhibit regional dependence--specifically, the palmar region strength is less (17.5 MPa) than the osteon interlamellar interfaces (30.4 MPa) and osteon type dependent (alternating significantly weaker than other types). Histomorphometry revealed significant regional differences (p<0.0001) in osteon area fraction among the four osteon types as well as differences in the osteon diameter (p=0.01), with dorsal regions having larger osteons (170 microm) than the palmar region (151 microm). Fatigue life and fracture toughness of Haversian bone are reported in the literature to be regionally dependent and are known to be associated with osteon pullout--an osteon interfacial phenomenon. Therefore, the results presented in this study are important to further the understanding of the mechanisms of fragility and damage accumulation in cortical bone.

Escherichia coli have evolved adaptive systems to resist strongly acidic habitats in part through the production of 2 biochemically identical isoforms of glutamate decarboxylase (GAD), encoded by the gadA and gadB genes. These genes occur in E. coli and other members of the genospecies (e.g., Shigella spp.) and originated as part of a genomic fitness island acquired early in Escherichia evolution. The present duplicated gad loci are widely spaced on the E. coli chromosome, and the 2 genes are 97% similar in sequence. Comparison of the nucleotide sequences of the gadA and gadB in 16 strains of pathogenic E. coli revealed 3.8% and 5.0% polymorphism in the 2 genes, respectively. Alignment of the homologous genes identified a total of 120 variable sites, including 21 fixed nucleotide differences between the loci within the first 82 codons of the genes. Twenty-three phylogenetically informative sites were polymorphic for the same nucleotides in both genes suggesting recent gene conversions or intergenic recombination. Phylogenetic analysis based on the synonymous substitutions per synonymous site indicated 2 cases in which specific gadA and gadB alleles were more closely related to one another than to other alleles at the corresponding locus. The results indicate that at least 3 gene conversion events have occurred after the gad gene duplication in the evolution of E. coli. Despite multiple gene conversion events, the upstream regulatory regions and the 5' end of each gene remains distinct, suggesting that maintaining functionally different gad genes is important in this acid-resistance mechanism in pathogenic E. coli.

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