Nisin inducible production of listeriolysin O in <Emphasis Type="Italic">Lactococcus lactis</Emphasis> NZ9000

Background  

Listeria monocytogenesis a well-characterized food-borne pathogen that infects pregnant women and immunocompromised individuals. Listeriolysin O (LLO) is the major virulence factor of the pathogen and is often used as a diagnostic marker for detection of L. monocytogenes. In addition, LLO represents a potent antigen driving T cell-mediated immunity during infection. In the present work, Lactococcus lactisNZ9000 was used as an expression host to hyper-produce LLO under inducible conditions using the NICE (NIsin Controlled Expression) system. We created a modified pNZ8048 vector encoding a six-His-tagged LLO downstream of the strong inducible PnisA promoter.     

Host plant response and phenotypic plasticity of a galling weevil (Collabismus clitellae: Curculionidae)

The response of a host plant to gall‐inducing insects varies both among and within plants, so that different levels of resources are available to the insects. The weevil Collabismus clitellae Boheman induces galls on the shoots of Solanum lycocarpum St Hil. in south‐east Brazil. Galls are found on a range of parts within an individual plant and are more abundant on smaller plants. In the present study, the host plant response as a possible influence on the performance of C. clitellae both between and within plants was tested. Gall abortion increased with plant height. Within plants, gall size was positively related to shoot diameter and number of chambers within the gall. The increase in gall larval density (number of individuals per gall volume unit) resulted in smaller adults and reduced developmental rates, probably because of resource limitation within the gall. The number of eggs laid by females increased with shoot diameter. Females laid more eggs on thicker shoots, where there are fewer chances to form galls with high larval density. However, this relationship was weak and a large variation was found for adult sizes. The availability of high quality sites is limited to smaller plants and thicker shoots located on the basal region of the plant. The phenotypic plasticity of this insect species in adult size and development time allows individuals growing on low quality sites to reach maturity, thus enhancing exploitation of the host plant.     

Climate‐driven disparities among ecological interactions threaten kelp forest persistence

The combination of ocean warming and acidification brings an uncertain future to kelp forests that occupy the warmest parts of their range. These forests are not only subject to the direct negative effects of ocean climate change, but also to a combination of unknown indirect effects associated with changing ecological landscapes. Here, we used mesocosm experiments to test the direct effects of ocean warming and acidification on kelp biomass and photosynthetic health, as well as climate‐driven disparities in indirect effects involving key consumers (urchins and rock lobsters) and competitors (algal turf). Elevated water temperature directly reduced kelp biomass, while their turf‐forming competitors expanded in response to ocean acidification and declining kelp canopy. Elevated temperatures also increased growth of urchins and, concurrently, the rate at which they thinned kelp canopy. Rock lobsters, which are renowned for keeping urchin populations in check, indirectly intensified negative pressures on kelp by reducing their consumption of urchins in response to elevated temperature. Overall, these results suggest that kelp forests situated towards the low‐latitude margins of their distribution will need to adapt to ocean warming in order to persist in the future. What is less certain is how such adaptation in kelps can occur in the face of intensifying consumptive (via ocean warming) and competitive (via ocean acidification) pressures that affect key ecological interactions associated with their persistence. If such indirect effects counter adaptation to changing climate, they may erode the stability of kelp forests and increase the probability of regime shifts from complex habitat‐forming species to more simple habitats dominated by algal turfs.     

Seed morphology,presence of areoles and water entry at imbibition of <i>Vicia dasycarpa</i> Ten. cv. Tolse F.C.A

The morphological characteristics of the seeds of Vicia dasycarpa Ten. cv. Tolse FCA were studied in relation to the area of imbibition water entry and were considered the presence of areoles. Seeds were analyzed using a stereo, optical and scanning electron microscope (SEM). The determination of the initial water entry area was carried out by immersing the seeds in a solution of tetrazolium (1%). This study showed that this species has seeds with a halo framing the hilum, an inconspicuous dry aril and a deltoid micropyle. The seedcoat pattern is papillose. The tracheid bar is surrounded by a ring of parenchymatous cells, and the tracheids show slight warty and non-vestured pits. It was confirmed the presence of an endospermic radicle pocket that surrounds and protect the radical tip. Two pairs of cotyledonar areoles were identified. It was established that the entry of water during imbibition starts in the area of the lens -having cracks- and moves in the sagittal plane. Both citological characteristic of tracheid bar and areoles presence show an apomorfous state between the Papiplionoids.     

A simple method for studying whole sections of rice grain

We developed a new and simple method to collect sections of a whole brown rice kernel for investigation of histological properties. A single kernel of rice was dehydrated through a graded ethanol series, transferred to xylene, and embedded in paraffin. During sectioning of the blocks using a rotary microtome, we used a special adhesive tape to collect and place the sections on slides so they remained flat. The use of the adhesive tape technique combined with autofluorescence characteristics allowed us to visualize cell walls throughout an entire longitudinal or transverse section of a whole rice kernel. We obtained scanning electron microscopy images of the sections to determine section quality.     

Staphylococcus aureus and Wegener's granulomatosis

Wegener's granulomatosis (WG) is a form of systemic vasculitis. It is characterized by granulomatous inflammation in the upper and lower airways, vasculitis and necrotizing glomerulonephritis, and is strongly associated with antineutrophil cytoplasmic antibodies against proteinase 3. Since the etiology of the disease is not clear, treatment, consisting of corticosteroids and immunosuppressives, is nonspecific and associated with severe side effects. Pinpointing the trigger(s) of the disease would highly improve treatment. Clinical evidence shows that an infectious agent, the bacterium Staphylococcus aureus, is a risk factor for disease relapse, suggesting its involvement in the pathogenesis of WG. Here we review both clinical and experimental data that either indicate or support a role for S. aureus in WG.     

Autoantibodies directed to novel components of the PM/Scl complex, the human exosome

The autoantigenic polymyositis/scleroderma (PM/Scl) complex was recently shown to be the human homologue of the yeast exosome, which is an RNA-processing complex. Our aim was to assess whether, in addition to targeting the known autoantigens PM/Scl-100 and PM/Scl-75, autoantibodies also target recently identified components of the PM/Scl complex. The prevalence of autoantibodies directed to six novel human exosome components (hRrp4p, hRrp40p, hRrp41p, hRrp42p, hRrp46p, hCsl4p) was determined in sera from patients with idiopathic inflammatory myopathy (n = 48), scleroderma (n = 11), or the PM/Scl overlap syndrome (n = 10). The sera were analyzed by enzyme-linked immunosorbent assays and western blotting using the affinity-purified recombinant proteins. Our results show that each human exosome component is recognized by autoantibodies. The hRrp4p and hRrp42p components were most frequently targeted. The presence of autoantibodies directed to the novel components of the human exosome was correlated with the presence of the anti-PM/Scl-100 autoantibody in the sera of patients with idiopathic inflammatory myopathy (IIM), as was previously found for the anti-PM/Scl-75 autoantibody. Other clear associations between autoantibody activities were not found. These results further support the conception that the autoimmune response may initially be directed to PM/Scl-100, whereas intermolecular epitope spreading may have caused the autoantibody response directed to the associated components.     

Habitat loss and mammalian extinction patterns: are the reserves in the Quadrilátero Ferrífero, southeastern Brazil, effective in conserving mammals?

Habitat loss is considered to be the principal cause of the local extinction of mammals worldwide. We assessed the extinction pattern of medium- and large-sized mammals caused by the effects of habitat loss in reserves in the Quadrilátero Ferrífero, southeastern Brazil, and discussed the effectiveness of these natural remnants for conserving mammals. A literature review and field collections were conducted from 2006 to 2011 to estimate the composition and richness of mammals in nine remnants of different sizes, including reserves and non-protected areas. A species–area relation and a nested subset analysis were performed, and a degree of sensitivity to habitat loss was obtained for each species according to its frequency of occurrence. Forty-five species of mammals were recorded. There was a strong species–area relation involving the legal size of reserves. High species richness was associated with large reserves, and the z value was within the range of very isolated continental remnants. The mammalian community exhibited a nested occurrence pattern, suggesting that most species were part of a more continuous ecosystem and that non-random extinction caused by habitat loss occurred in southeastern Brazil. The negative relation found between species frequencies and body weights suggested that selective species loss is associated with decreases in the size of the reserves. The estimated viable size required to conserve all of the sensitive species is greater than the size of the largest reserve inventoried. We recommend the aggregation of neighboring natural remnants and the creation of new reserves to reduce extinction risks.     

Trends of fumonisin contamination and animal intoxication through monitoring 1991 to 1997 corn crop in the State of Paraná, Brazil

Eleven feed samples associated with six animal (horse and poultry) intoxication outbreaks (1991) in the state of Paraná, Brazil, were evaluated for fungal and fumonisin contamination. In order to estimate the␣trend of livestock intoxication, fumonisin contamination was monitored in corn produced both at the commercial level (1991, 1995 crop), and in an experimental field at a local Agronomy Institute (1997 crop). The total mould count in the feed samples ranged from 2.9 × 10³ to 1.9 × 10⁷ CFU/g, with Fusarium verticillioides as the predominant species, at a high count of 2.4 × 10⁴–6.5 × 10⁵ CFU/g. Fumonisins (FB₁ + FB₂) were detected in all corn-based feed samples at levels ranging from 2.89 to 14.54 μg/g. All 27 Northern corn samples (1991 crop) were contaminated with fumonisins at levels ranging from 2.32 to 16.64 μg/g. Twenty-six (96.3%) out of 27 corn samples from the Central-Southern region (1995 crop) were positive for fumonisins (FB₁+FB₂), with the range of 0.07–3.66 μg/g, while all 37 Northern samples (1995 crop) were contaminated with fumonisins ranging from 0.57 to 9.97 μg/g. Twenty-one out of 37 corn samples from the Northern region (1997 crop) were positive for fumonisins, but at low level (range of 0.05–2.67 μg/g). The results showed a decreasing trend in fumonisin contamination over the years. Nowadays animal intoxication outbreaks rarely occur in this State, as both animal producers and feed industries have become conscious about monitoring of corn and other raw materials at the quality control level.     

Production of heterologous protein by Methylobacterium extorquens in high cell density fermentation

The green fluorescent protein (GFP) was used as a model protein to study the recombinant protein production by the strain Methylobacterium extorquens ATCC 55366. Scale-up from shake flasks to 20 l fed-batch fermentation was achieved using methanol as a sole carbon and energy source and a completely minimal culture medium. Two different expression vectors were used to express GFP. Clone PCM-GFP containing the vector pCM110 with native promoter of the methanol dehydrogenase PmxaF produced approximately 100-fold more GFP than the clone PRK-GFP containing the vector pRK310 with the heterogeneous promoter Plac. Several fed-batch fermentations with and without selective pressure (tetracycline) were run in a 20 l stirred tank fermenter using the two different clones of M. extorquens. The methanol concentration was monitored with an on-line semiconductor gas sensor in the culture broth. It was maintained at a non-toxic level of 1.4 g l(-1) with an adaptative control which regulates the methanol feed rate. The same growth profile was achieved in all fermentations. The maximum growth rate (micro(max)) was 0.18 h(-1) with an overall yield (Y(X/S)) of 0.3 g g(-1) methanol. With this high cell density fermentation process, we obtained high levels (up to 4 g l(-1)) of GFP with the clone PCM-GFP. The maximum specific GFP production (Y(GFP/X)) with this clone was 80 mg g(-1) representing approximately 16% of the total cell protein. Additional feeding of pure oxygen to the fermenter permitted a longer phase of exponential growth but had no effect on the total yields of biomass and GFP. The specific GFP production of clone PCM-GFP remained unaffected in the presence or absence of selective pressure (tetracycline), within the initial 50 h of the fermentation culture. These results suggest that M. extorquens ATCC 55366 could be an interesting candidate for overexpression of recombinant proteins.