Marker rescue transformation by linear plasmid DNA in Bacillus subtilis

Although plasmid-free Bacillus subtilis cannot be transformed for markers carried by linear or nicked plasmid DNA, a resident plasmid can rescue a marker on such damaged DNA under certain conditions. Linearized chimeric plasmid DNA has been used to transform cultures carrying a resident plasmid which is homologous with a portion of the donor. This system has revealed the following properties of the marker rescue process: (1) It is recE dependent. (2) It requires the presence in the resident plasmid of sequences which are homologous to the donor. (3) When the selected marker is on a nonhomologous segment it must be flanked by segments which are homologous to the resident plasmid. (4) The efficiency of rescue varies in a regular way with the position of the linearizing cut. (5) Marker rescue is first order with respect to DNA concentration. These properties and other data are interpreted as providing a strong indication that marker rescue occurs by recombination, although an alternative explanation involving recE-dependent recircularization of the donor plasmid has not been eliminated. Our results also suggest that if the major pathway of marker rescue is by recombination, an average of 0.15 Mdal (single strand) must be removed from each donor DNA molecule or otherwise rendered unavailable for recombination and that the exchange frequency during transformational recombination is approximately 0.2 to 0.5 Mdal⁻¹.     

Fish assemblage structure in an estuary of the Atlantic Forest biodiversity hotspot (southern Brazil)

We described the fish assemblage in the estuary of the Guaraguaçu River (one of the largest tributaries of the Paranaguá Bay Estuary, located within Brazil’s Atlantic Forest Biosphere Reserve) from June 2005 to May 2006, and assessed the seasonal and spatial effects of abiotic environmental attributes on the fish assemblage structure. Despite some oscillations in salinity, the upper and lower estuaries had year-round persistent oligohaline and polyhaline conditions, respectively. Despite high species richness (55 species), the Guaraguaçu River Estuary fish community contains a few dominant taxa; 11% of the richness accounts for >60% of its density and biomass. The most abundant species (in terms of both biomass and density) was Atherinella brasiliensis. Species whose densities were most strongly associated with the upper estuary were Centropomus parallelus, Ctenogobius schufeldti, Eucinostomus melanopterus, Platanichthys platana, Trinectes paulistanus, and Eugerres brasilianus. Those whose densities were most strongly associated with the lower estuary were A. brasiliensis, Sphoeroides greeleyi, Eucinostomus argenteus, Sphoeroides testudineus, Diapterus rhombeus, and Harengula clupeola. Throughout the year, canonical correspondence analysis identified: (1) the pattern of horizontal stratification of salinity along the river as being the most important variable for explaining most of the fish fauna structure; and (2) a strong relationship between the fish fauna and the salinity gradient along the estuary. Analysis of similarity further confirmed that each estuarine zone supports a year-round persistent and relatively homogeneous fish species assemblage. Total mean density and biomass remained constant over time in each estuarine habitat, but density shifted in the most abundant species, which appears related to recruitment patterns. Such species and abundance persistence likely occurs because seasonal rainfall-induced changes in river discharge are not sufficient to significantly shift runoff and salinity and thus fish assemblage structure (species composition, density and biomass) along the estuary. Such a lack of seasonal fish fauna movement as a response to changes in river flow contrasts with other estuarine systems around the world.     

Molecular investigations of a locally acquired case of melioidosis in Southern AZ, USA

Melioidosis is caused by Burkholderia pseudomallei, a Gram-negative bacillus, primarily found in soils in Southeast Asia and northern Australia. A recent case of melioidosis in non-endemic Arizona was determined to be the result of locally acquired infection, as the patient had no travel history to endemic regions and no previous history of disease. Diagnosis of the case was confirmed through multiple microbiologic and molecular techniques. To enhance the epidemiological analysis, we conducted several molecular genotyping procedures, including multi-locus sequence typing, SNP-profiling, and whole genome sequence typing. Each technique has different molecular epidemiologic advantages, all of which provided evidence that the infecting strain was most similar to those found in Southeast Asia, possibly originating in, or around, Malaysia. Advancements in new typing technologies provide genotyping resolution not previously available to public health investigators, allowing for more accurate source identification.     

Histopathological effects of cisplatin,doxorubicin and 5-flurouracil (5-FU) on the liver of male albino rats

Cisplatin, doxorubicin and fluorouracil (5-FU), drugs belonging to different chemical classes, have been extensively used for chemotherapy of various cancers. Despite extensive investigations into their hepatotoxicity, there is very limited information on their effects on the structure and ultra-structure of liver cells in vivo. Here, we demonstrate for the first time, the effects of these three anticancer drugs on rat liver toxicity using both light and electron microscopy. Light microscopic observations revealed that higher doses of cisplatin and doxorubicin caused massive hepatotoxicity compared to 5-FU treatment, including dissolution of hepatic cords, focal inflammation and necrotic tissues. Interestingly, low doses also exhibited abnormal changes, including periportal fibrosis, degeneration of hepatic cords and increased apoptosis. These changes were confirmed at ultrastructural level, including vesiculated rough endoplasmic reticulum and atrophied mitochondria with ill-differentiated cisternae, dense collection of macrophages and lymphocytes as well as fibrocytes with collagenous fibrils manifesting early sign of fibrosis, especially in response to cisplatin and doxorubicin -treatment. Our results provide in vivo evidence, at ultrastructural level, of direct hepatotoxicity caused by cisplatin, doxorubicin and 5-FU at both light and electron microscopi. These results can guide the design of appropriate treatment regimen to reduce the hepatotoxic effects of these anticancer drugs.     

The Ras guanine nucleotide exchange factor RasGRF1 promotes matrix metalloproteinase-3 production in rheumatoid arthritis synovial tissue

Introduction  

Fibroblast-like synoviocytes (FLS) from rheumatoid arthritis (RA) patients share many similarities with transformed cancer cells, including spontaneous production of matrix metalloproteinases (MMPs). Altered or chronic activation of proto-oncogenic Ras family GTPases is thought to contribute to inflammation and joint destruction in RA, and abrogation of Ras family signaling is therapeutic in animal models of RA. Recently, expression and post-translational modification of Ras guanine nucleotide releasing factor 1 (RasGRF1) was found to contribute to spontaneous MMP production in melanoma cancer cells. Here, we examine the potential relationship between RasGRF1 expression and MMP production in RA, reactive arthritis, and inflammatory osteoarthritis synovial tissue and FLS.