Rootstock genotype succession influences apple replant disease and root-zone microbial community composition in an orchard soil

Apple replant disease (ARD) is a soil-borne disease complex that affects young apple trees in replanted orchards, resulting in stunted growth and reduced yields. Newly developed rootstock genotypes with tolerance to ARD may help to control this disease. We determined the effects of rootstock genotype rotations during orchard renovation, by investigating root-zone soil microbial consortia and the relative severity of ARD on seven rootstock genotypes (M.9, M.26, G.30, G.41, G.65, G.935, and CG.6210) planted in soil where trees on four of those same rootstocks (M.9, M.26, G.30 and CG.6210) had grown for the previous 15 years. Rootstock genotyping indicated that genetic distances among rootstocks were loosely correlated with their differential responses to ARD. Root-zone fungal and bacterial community composition, assessed by DNA fingerprinting (T-RFLP), differed between M.26 and CG.6210. Soil bacterial communities were influenced most by which rootstock had grown in the soil previously, while fungal communities were influenced more by the current replanted rootstock. In a clone library of bacteria from M.26 and CG.6210 root-zone soil, β-Proteobacteria was the most abundant phylum (25% of sequences). Sequences representing the Burkholderia cepacia complex were obtained only from CG.6210 soil. Rootstock genotypes that were grown in the orchard soil previously affected subsequent ARD severity, but replanting with the same or closely related rootstocks did not necessarily exacerbate this disease problem. Our results suggest that genotype-specific interactions with soil microbial consortia are linked with apple rootstock tolerance or susceptibility to ARD.     

Influence of production process design on inclusion bodies protein: the case of an Antarctic flavohemoglobin

Background  

Protein over-production in Escherichia coli often results in formation of inclusion bodies (IBs). Some recent reports have shown that the aggregation into IBs does not necessarily mean that the target protein is inactivated and that IBs may contain a high proportion of correctly folded protein. This proportion is variable depending on the protein itself, the genetic background of the producing cells and the expression temperature. In this paper we have evaluated the influence of other production process parameters on the quality of an inclusion bodies protein.     

Presence of Leishmania infantum in red foxes (Vulpes vulpes) in southern Italy

Skin, lymph node (popliteal), and bone marrow samples were collected from 50 red foxes (Vulpes vulpes) from May 2004 to May 2005 in southern Italy. Samples were tested for Leishmania infantum by polymerase chain reaction (PCR). The parasite was detected by PCR from 20 of 50 (40%) fox carcasses. All 20 positive cases were PCR-positive from lymph node and bone marrow samples, whereas 17 of 20 positive cases were PCR-positive from skin samples. Infection status was not related to age or sex. This is the first report of leishmaniasis in red foxes in Italy based on PCR results, and these results reinforce the assumption that this wild canid can serve as a reservoir for Leishmania.     

Karyological evolution and systematics of Malagasy microhylid frogs

Microhylid frogs are a group of largely unresolved phylogeny, and diverse data sets are needed to improve the evolutionary understanding of these amphibians. We here report karyological data for 22 species of this family, belonging to the Malagasy genera Anodonthyla, Cophyla, Platypelis, Plethodontohyla, Rhombophryne, and Stumpffia (Cophylinae); Scaphiophryne and Paradoxophyla (Scaphiophryninae); and Dyscophus (Dyscophinae); and the Asian genera Calluella and Ramanella (Microhylinae). All species studied have 2n=26 chromosomes, most of which are metacentric or submetacentric. Chromosome morphology, banding pattern, and position of the nucleolar organizer regions (NORs) provide relevant characters for the understanding of the phylogeny and systematics of these frogs. The species of the Cophylinae are characterized by a subtelocentric or telocentric fourth chromosome pair (submetacentric only in Anodonthyla), which can be seen as a synapomorphy for this subfamily. Shifts in NOR position within the Cophylinae are frequent and agree with recent mitochondrial DNA data, corroborating the non-monophyly of the genus Plethodontohyla. Changes of NOR position and chromosome morphology (i.e., occurrence of subtelocentric and telocentric elements) were also common in this subfamily, possibly being related to their faster mitochondrial substitution rate and high species diversity. The ninth chromosome pair of the examined specimens of Dyscophus guineti, all juveniles, is heteromorphic. In this pair, one of the two chromosomes is longer due to the addition of two heterochromatic segments, raising the possibility that one chromosome of this pair may be a sex chromosome.     

Helper-dependent adenovirus for the gene therapy of proliferative retinopathies: stable gene transfer, regulated gene expression and therapeutic efficacy

BACKGROUND: Ocular neovascular disorders, such as diabetic retinopathy and age-related macular degeneration, are the principal causes of blindness in developed countries. Current treatments are of limited efficacy, whereas a therapy based on intraocular gene transfer of angiostatic factors represents a promising alternative. For the first time we have explored the potential of helper-dependent adenovirus (HD-Ad), the last generation of Ad vectors, in the therapy of retinal neovascularization. METHODS: We first analyzed efficiency and stability of intraretinal gene transfer following intravitreous injection in mice. A HD-Ad vector expressing green fluorescent protein (GFP) under the control of the cytomegalovirus (CMV) promoter (HD-Ad/GFP) was compared with a first-generation (E1/E3-deleted) Ad vector carrying an identical GFP expression cassette (FG-Ad/GFP). We also constructed HD-Ad vectors expressing a soluble form of the VEGF receptor (sFlt-1) in a constitutive (HD-Ad/sFlt-1) or doxycycline (dox)-inducible (HD-Ad/S-M2/sFlt-1) manner and tested their therapeutic efficacy upon intravitreous delivery in a rat model of oxygen-induced retinopathy (OIR). RESULTS: HD-Ad/GFP promoted long-lasting (up to 1 year) transgene expression in retinal Müller cells, in marked contrast with the short-term expression observed with FG-Ad/GFP. Intravitreous injection of HD-Ad vectors expressing sFlt-1 resulted in detectable levels of sFlt-1 and inhibited retinal neovascularization by more than 60% in a rat model of OIR. Notably, the therapeutic efficacy of the inducible vector HD-Ad/S-M2/sFlt-1 was strictly dox-dependent. CONCLUSIONS: HD-Ad vectors enable stable gene transfer and regulated expression of angiostatic factors following intravitreous injection and thus are attractive vehicles for the gene therapy of neovascular diseases of the retina.     

Binding of aflatoxins to the 20S proteasome: effects on enzyme functionality and implications for oxidative stress and apoptosis

Aflatoxins (AF) are contaminants of improperly stored foods; they are potent genotoxic and carcinogenic compounds, exerting their effects through damage to DNA. They can also induce mutations that increase oxidative damage. The goal of this study was to evaluate the possibility that a third mechanism could be involved in the carcinogenic action of aflatoxins, namely, direct binding to key enzymes involved in the regulatory pathways of the cell cycle, thereby modulating enzyme functionality. The 20S constitutive and immunoproteasome peptidase and proteolytic activities were assayed in the presence of aflatoxins B1, G1 and M1. All three toxins activated multiple peptidase activities of the proteasome. Aflatoxin (AF) M1 was the most potent activator of proteasome activity, while the constitutive 20S proteasome was specifically stimulated by AFG1. Furthermore, the effects of AFB1 on cultured hepatoma cells were investigated and the various proteasomal activities determined with cell lysates were differently affected. Taking into account the key role of the proteasome in cellular defense against oxidative stress, the carbonyl group content and the activities of antioxidant enzymes in cell lysates were analyzed. The proapoptotic effect of AFB1 was also investigated by measuring caspase-3 activity and cellular levels of p27 and IkappaBalpha.