Mitochondrial DNA from osteoarthritic patients drives functional impairment of mitochondrial activity: a study on transmitochondrial cybrids

With the redefinition of osteoarthritis (OA) and the understanding that the joint behaves as an organ, OA is now considered a systemic illness with a low grade of chronic inflammation. Mitochondrial dysfunction is well documented in OA and has the capacity to alter chondrocyte and synoviocyte function. Transmitochondrial cybrids are suggested as a useful cellular model to study mitochondrial biology in vitro, as they carry different mitochondrial variants with the same nuclear background. The aim of this work was to study mitochondrial and metabolic function of cybrids with mitochondrial DNA from healthy (N) and OA donors. In this work, the authors demonstrate that cybrids from OA patients behave differently from cybrids from N donors in several mitochondrial parameters. Furthermore, OA cybrids behave similarly to OA chondrocytes. These results enhance our understanding of the role of mitochondria in the degeneration process of OA and present cybrids as a useful model to study OA pathogenesis.     

Repair and enterotoxin synthesis by Staphylococcus aureus after thermal shock.

To study repair and enterotoxin synthesis, four staphylococcal strains (FRI-100, FRI-137, FRI-472, and S6) were subjected to sublethal heat treatment, transferred to four liquid repair media (1% powdered skim milk in distilled water, complex medium, M9 minimal salt medium, and saline solution), and then incubated at different temperatures. Powdered skim milk proved to be the most efficient medium for promoting the repair of injured cells, particularly at 37 degrees C. Minimal salt medium also gave good results. Salt tolerance also increased at 4 degrees C, although it did not reach normal values. After 6 h of incubation at 37 degrees C in powdered skim milk, strain FRI-100 synthesized detectable amounts of enterotoxin A. After 10 h of incubation in the same medium at the same temperature, enterotoxins were detected in all of the strains.     

Coupled saccharification and fermentation of pre-treated Eucalyptus wood: a simple kinetic model

Coupled saccharification and fermentation of Eucalyptus globulus wood, pre-treated by acid hydrolysis and sodium hypochlorite, was carried out in two column reactors: one for enzymatic hydrolysis of the substrate at 50°C and the other for fermentation of sugars with calcium alginate-immobilized Saccharomyces cerevisiae at 30°C. A buffered solution containing cellulases at pH 4.8 was recycled through both reactors. The maximum yields were about 0.26 g ethanol per g of substrate. The results were reproduced reasonably well using a simple kinetic model consisting of two successive pseudo-first-order reactions.C. Albornoz and D. M. Ferrari are with the Centro de Investigaciones Tecnológicas (CIT), Administración Nacional de Combustibles, Alcohol y Portland (ANCAP), Pando, Canelones, C.P. 91000, Uruguay. S. Blanco and G. Ellenrieder are with Instituto de Investigaciones para la Industria Química (INIQUI), Universidad Nacional de Salta (UNSa), Buenos Aires 177, 4400 Salta, Argentina.     

Grazing lucerne as fattening management for young bulls: technical and economic performance and diet authentication

Three fattening systems were evaluated from weaning to slaughter in order to find alternatives to grain feeding in young bulls, and to test the reliability of carcass subcutaneous fat colour to discriminate among them. After weaning (224 kg), one group of animals was fed concentrates and straw until they reached the target slaughter weight (450 kg; Feedlot), another group grazed rotationally on lucerne supplemented with 1.8 kg DM/day barley until slaughter (LUC), and the third group had the same management as LUC animals for 3 months (period 1) and thereafter was finished on concentrates and straw until slaughter (period 2; LUC + Feedlot). Animals were weighed weekly and sampled monthly for serum IGF-I and leptin, and plasma non-esterified fatty acids and carotenoid pigment concentration analyses. Carcass characteristics and subcutaneous fat colour were recorded after slaughter. In period 1, Feedlot animals had slightly greater weight gains than their grazing counterparts (P < 0.10), and at the end of period 1 they had 66% greater IGF-I and 35% greater leptin concentration (P < 0.01). Plasma carotenoid pigments were undetectable in Feedlot animals, but increased during grazing in LUC and LUC + Feedlot treatments. In period 2, weight gains were lowest for LUC, intermediate for Feedlot and greatest for LUC + Feedlot animals (P < 0.001), conditioning the time taken to reach slaughter weight (73, 58 and 47 days, respectively; P < 0.05). Leptin and IGF-I concentrations increased in all management systems during period 2. Plasma carotenoid pigment concentration reached its maximum at the end of period 2 in LUC animals, but it decreased sharply in LUC + Feedlot animals in this period. Management did not affect carcass traits except for subcutaneous fat colour. Yellowness, Chroma (C*) and the value of the integral of the translated reflectance spectrum (SUM), estimator of carotenoid pigment content in fat, were higher in LUC than in LUC + Feedlot and Feedlot animals (P < 0.001). Two logistic regressions were obtained to discriminate carcasses from LUC treatment: P (LUC) = (1 + e(18.8-5.6 × lightness-36.9 × redness + 0.3 × SUM + 29.8 × C*))(-1) and LUC + Feedlot treatment: P (LUC + Feedlot)=(1 + e(833.7-11.8 × lightness + 4.7 × redness + 0.2 × SUM-2.5 × C*))(-1). The economic margin, calculated as income achieved minus costs, was greatest for LUC, intermediate for LUC + Feedlot and lowest for Feedlot treatment. Therefore, grazing lucerne supplemented with barley was an interesting alternative for fattening young bulls in these conditions, producing carcasses of similar quality, which could be accurately traced by measuring subcutaneous fat colour.     

Phylogenetic significance of morphological characters in the tropical Hypotrachyna clade of parmelioid lichens (Parmeliaceae, Ascomycota)

Lichen-forming ascomycetes exhibit often complex morphologies of the vegetative thallus that are usually not found in non-lichenized fungi. This includes the thallus organization and appendical structures associated with the main thallus, such as cilia and rhizines. Such morphological characters are widely employed in the taxonomy of parmelioid lichens, especially at generic level. Within parmelioid lichens, several monophyletic groups can be distinguished, the Hypotrachyna clade being one of them, which includes mostly tropical taxa. In this first molecular study focused specifically on the Hypotrachyna clade, we used maximum parsimony and Bayesian analyses of a combined data set of nuclear ITS and mitochondrial SSU rDNA sequences to (1) test the monophyly of genera presently accepted within the clade and (2) evaluate the phylogenetic value of the morphological characters used to circumscribe genera in parmelioid lichens. Out of the 89 mtSSU and 88 nuITS sequences included in the present study, 121 sequences were newly obtained. Our results show that the taxa within the clade fall into two major groups and that the genus Hypotrachyna is polyphyletic. Everniastrum and Parmelinopsis are nested within Hypotrachyna sensu stricto, the latter being also polyphyletic. Bulbothrix is paraphyletic with Parmelinella nested within and is basal to the second major Hypotrachyna clade. Monophylies of Bulbothrix and Hypotrachyna are significantly rejected. The phylogenetic analysis demonstrates that morphological characters currently used to circumscribe genera in parmelioid lichens, such as cortical anatomy, lobe configuration, cilia, and rhizines have been overestimated and have only minor value in identifying monophyletic groups.     

Identification of T-cell epitopes in nonstructural proteins of foot-and-mouth disease virus

Porcine T-cell recognition of foot-and-mouth disease virus (FMDV) nonstructural proteins (NSP) was tested using in vitro lymphoproliferative responses. Lymphocytes were obtained from outbred pigs experimentally infected with FMDV. Of the different NSP, polypeptides 3A, 3B, and 3C gave the highest stimulations in the in vitro assays. The use of overlapping synthetic peptides allowed the identification of amino acid regions within these proteins that were efficiently recognized by the lymphocytes. The sequences of some of these antigenic peptides were highly conserved among different FMDV serotypes. They elicited major histocompatibility complex-restricted responses with lymphocytes from pigs infected with either a type C virus or reinfected with a heterologous FMDV. A tandem peptide containing the T-cell peptide 3A[21-35] and the B-cell antigenic site VP1[137-156] also efficiently stimulated lymphocytes from infected animals in vitro. Furthermore, this tandem peptide elicited significant levels of serotype-specific antiviral activity, a result consistent with the induction of anti-FMDV antibodies. Thus, inclusion in the peptide formulation of a T-cell epitope derived from the NSP 3A possessing the capacity to induce T helper activity can allow cooperative induction of anti-FMDV antibodies by B cells.     

Diversity of Multi-Drug Resistant Avian Pathogenic Escherichia coli (APEC) Causing Outbreaks of Colibacillosis in Broilers during 2012 in Spain

Avian pathogenic Escherichia coli (APEC) are the major cause of colibacillosis in poultry production. In this study, a total of 22 E. coli isolated from colibacillosis field cases and 10 avian faecal E. coli (AFEC) were analysed. All strains were characterised phenotypically by susceptibility testing and molecular typing methods such as pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST). The presence of 29 virulence genes associated to APEC and human extraintestinal pathogenic E. coli (ExPEC) was also evaluated. For cephalosporin resistant isolates, cephalosporin resistance genes, plasmid location and replicon typing was assessed. Avian isolates belonged to 26 O:H serotypes and 24 sequence types. Out of 22 APEC isolates, 91% contained the virulence genes predictors of APEC; iutA, hlyF, iss, iroN and ompT. Of all strains, 34% were considered ExPEC. PFGE analysis demonstrated a high degree of genetic polymorphism. All strains were multi-resistant, including those isolated from healthy animals. Eleven strains were resistant to cephalosporins; six contained bla _CTX-M-14, two bla _SHV-12, two bla _CMY-2 and one bla _SHV-2. Two strains harboured qnrA, and two qnrA together with aac(6’)-Ib-cr. Additionally, the emergent clone O25b:H4-B2-ST131 was isolated from a healthy animal which harboured bla _CMY-2 and qnrS genes. Cephalosporin resistant genes were mainly associated to the presence of IncK replicons. This study demonstrates a very diverse population of multi-drug resistant E. coli containing a high number of virulent genes. The E. coli population among broilers is a reservoir of resistance and virulence-associated genes that could be transmitted into the community through the food chain. More epidemiological studies are necessary to identify clonal groups and resistance mechanisms with potential relevance to public health.     

HIV-1 requires Arf6-mediated membrane dynamics to efficiently enter and infect T lymphocytes

As the initial barrier to viral entry, the plasma membrane along with the membrane trafficking machinery and cytoskeleton are of fundamental importance in the viral cycle. However, little is known about the contribution of plasma membrane dynamics during early human immunodeficiency virus type 1 (HIV-1) infection. Considering that ADP ribosylation factor 6 (Arf6) regulates cellular invasion via several microorganisms by coordinating membrane trafficking, our aim was to study the function of Arf6-mediated membrane dynamics on HIV-1 entry and infection of T lymphocytes. We observed that an alteration of the Arf6-guanosine 5'-diphosphate/guanosine 5'-triphosphate (GTP/GDP) cycle, by GDP-bound or GTP-bound inactive mutants or by specific Arf6 silencing, inhibited HIV-1 envelope-induced membrane fusion, entry, and infection of T lymphocytes and permissive cells, regardless of viral tropism. Furthermore, cell-to-cell HIV-1 transmission of primary human CD4(+) T lymphocytes was inhibited by Arf6 knockdown. Total internal reflection fluorescence microscopy showed that Arf6 mutants provoked the accumulation of phosphatidylinositol-(4,5)-biphosphate-associated structures on the plasma membrane of permissive cells, without affecting CD4-viral attachment but impeding CD4-dependent HIV-1 entry. Arf6 silencing or its mutants did not affect fusion, entry, and infection of vesicular stomatitis virus G-pseudotyped viruses or ligand-induced CXCR4 or CCR5 endocytosis, both clathrin-dependent processes. Therefore we propose that efficient early HIV-1 infection of CD4(+) T lymphocytes requires Arf6-coordinated plasma membrane dynamics that promote viral fusion and entry.     

All clinically-relevant blood components transmit prion disease following a single blood transfusion: a sheep model of vCJD

Variant CJD (vCJD) is an incurable, infectious human disease, likely arising from the consumption of BSE-contaminated meat products. Whilst the epidemic appears to be waning, there is much concern that vCJD infection may be perpetuated in humans by the transfusion of contaminated blood products. Since 2004, several cases of transfusion-associated vCJD transmission have been reported and linked to blood collected from pre-clinically affected donors. Using an animal model in which the disease manifested resembles that of humans affected with vCJD, we examined which blood components used in human medicine are likely to pose the greatest risk of transmitting vCJD via transfusion. We collected two full units of blood from BSE-infected donor animals during the pre-clinical phase of infection. Using methods employed by transfusion services we prepared red cell concentrates, plasma and platelets units (including leucoreduced equivalents). Following transfusion, we showed that all components contain sufficient levels of infectivity to cause disease following only a single transfusion and also that leucoreduction did not prevent disease transmission. These data suggest that all blood components are vectors for prion disease transmission, and highlight the importance of multiple control measures to minimise the risk of human to human transmission of vCJD by blood transfusion.