Isolation from sewage influent and characterization of novel Staphylococcus aureus bacteriophages with wide host ranges and potent lytic capabilities.

Staphylococcus aureus is a gram-positive pathogen that causes a variety of diseases, including bovine mastitis, which has severe economic consequences. Standard antibiotic treatment results in selection of resistant strains, leading to a need for alternative treatments, such as bacteriophage therapy. Forty-nine S. aureus isolates were obtained from the milk of mastitic cows for use in screening of staphylococcal phages. Fifteen isolates which were positive for both coagulase and hemolysin were assayed by PCR for variation in the X region and the immunoglobulin G-binding region of the protein A gene (spa) and in the carboxy terminus of the coagulase gene (coa) and for the presence of enterotoxin C, G, H, and I genes. The host ranges of 52 phages isolated from sewage influent were determined by performing spot tests with the 15 S. aureus isolates, and two phages were subsequently chosen for further analysis. PhiSA039 had the widest host range, producing clear plaques on 13 of the 15 isolates (87%), while PhiSA012 produced clear plaques on 8 isolates (53%) and was the only phage that produced a clear plaque on a nonmastitic S. aureus strain. Transmission electron microscopy revealed that the phages were similar sizes and belonged to the Myoviridae family. Measurement of optical densities during coculture with S. aureus isolates confirmed the breadth of the PhiSA039 host range and showed that PhiSA012 had potent lytic capability. PhiSA012-resistant bacteria did not appear for three of seven isolates tested (43%) after 65 h of incubation. These two phages are proposed as candidates for phage therapy of bovine mastitis.

Circular dichroism imaging microscopy: application to enantiomorphous twinning in biaxial crystals of 1,8-dihydroxyanthraquinone.

A microscope was constructed for imaging circular dichroism of heterogeneous anisotropic media. To avoid linear biases that are common with electronic circular polarization modulation, we chose a retrogressive solution: mechanical light modulation by rotating a linear polarizer with respect to a quarter wave plate continuously tuned by tilting to the operating wavelength. Our comparatively slow technique succeeds with near-perfect circular input and signal averaging using a CCD camera. We have applied the method to anomalously birefringent crystals of 1,8-dihydroxyanthraquinone that are shown to have intergrown mirror image domains, undetected by X-ray diffraction because the twinning complexity renders differences in anomalous dispersion, already small, unreliable. The origin of the anomalous birefringence and the assignment of the absolute configuration are discussed.

Imaging linear birefringence and dichroism in cerebral amyloid pathologies.

New advances in polarized light microscopy were used to image Congo red-stained cerebral amyloidosis in sharp relief. The rotating-polarizer method was used to separate the optical effects of transmission, linear birefringence, extinction, linear dichroism, and orientation of the electric dipole transition moments and to display them as false-color maps. These effects are typically convolved in an ordinary polarized light microscope. In this way, we show that the amyloid deposits in Alzheimer's disease plaques contain structurally disordered centers, providing clues to mechanisms of crystallization of amyloid in vivo. Comparisons are made with plaques from tissues of subjects having Down's syndrome and a prion disease. In plaques characteristic of each disease, the Congo red molecules are oriented radially. The optical orientation in amyloid deposited in blood vessels from subjects having cerebral amyloid angiopathy was 90 degrees out of phase from that in the plaques, suggesting that the fibrils run tangentially with respect to the circumference of the blood vessels. Our result supports an early model in which Congo red molecules are aligned along the long fiber axis and is in contrast to the most recent binding models that are based on computation. This investigation illustrates that the latest methods for the optical analysis of heterogeneous substances are useful for in situ study of amyloid.