Vaccination of lactating dairy cows for the prevention of aflatoxin B1 carry over in milk

The potential of anaflatoxin B(1) (AnAFB(1)) conjugated to keyhole limpet hemocyanin (KLH) as a vaccine (AnAFB(1)-KLH) in controlling the carry over of the aflatoxin B(1) (AFB(1)) metabolite aflatoxin M(1) (AFM(1)) in cow milk is reported. AFB(1) is the most carcinogenic compound in food and foodstuffs amongst aflatoxins (AFs). AnAFB(1) is AFB(1) chemically modified as AFB(1)-1(O-carboxymethyl) oxime. In comparison to AFB(1), AnAFB(1) has proven to be non-toxic in vitro to human hepatocarcinoma cells and non mutagenic to Salmonella typhimurium strains. AnAFB(1)-KLH was used for immunization of cows proving to induce a long lasting titer of anti-AFB(1) IgG antibodies (Abs) which were cross reactive with AFB(1), AFG(1), and AFG(2). The elicited anti-AFB(1) Abs were able to hinder the secretion of AFM(1) into the milk of cows continuously fed with AFB(1). Vaccination of lactating animals with conjugated AnAFB(1) may represent a solution to the public hazard constituted by milk and cheese contaminated with AFs.     

A novel Bacteroidetes symbiont is localized in Scaphoideus titanus, the insect vector of Flavescence dorée in Vitis vinifera

Flavescence dorée (FD) is a grapevine disease that afflicts several wine production areas in Europe, from Portugal to Serbia. FD is caused by a bacterium, "Candidatus Phytoplasma vitis," which is spread throughout the vineyards by a leafhopper, Scaphoideus titanus (Cicadellidae). After collection of S. titanus specimens from FD-contaminated vineyards in three different areas in the Piedmont region of Italy, we performed a survey to characterize the bacterial microflora associated with this insect. Using length heterogeneity PCR with universal primers for bacteria we identified a major peak associated with almost all of the individuals examined (both males and females). Characterization by denaturing gradient gel electrophoresis confirmed the presence of a major band that, after sequencing, showed a 97 to 99% identity with Bacteroidetes symbionts of the "Candidatus Cardinium hertigii" group. In addition, electron microscopy of tissues of S. titanus fed for 3 months on phytoplasma-infected grapevine plants showed bacterial cells with the typical morphology of "Ca. Cardinium hertigii." This endosymbiont, tentatively designated ST1-C, was found in the cytoplasm of previtellogenic and vitellogenic ovarian cells, in the follicle cells, and in the fat body and salivary glands. In addition, cell morphologies resembling those of "Ca. Phytoplasma vitis" were detected in the midgut, and specific PCR assays indicated the presence of the phytoplasma in the gut, fat body and salivary glands. These results indicate that ST1-C and "Ca. Phytoplasma vitis" have a complex life cycle in the body of S. titanus and are colocalized in different organs and tissues.     

Opposing effects of ICOS on graft-versus-host disease mediated by CD4 and CD8 T cells

ICOS, a CD28 family member expressed on activated CD4(+) and CD8(+) T cells, plays important roles in T cell activation and effector function. Here we studied the role of ICOS in graft-vs-host disease (GVHD) mediated by CD4(+) or CD8(+) T cells in allogeneic bone marrow transplantation. In comparison of wild-type and ICOS-deficient T cells, we found that recipients of ICOS(-/-) CD4(+) T cells exhibited significantly less GVHD morbidity and delayed mortality. ICOS(-/-) CD4(+) T cells had no defect in expansion, but expressed significantly less Fas ligand and produced significantly lower levels of IFN-gamma and TNF-alpha. Thus, ICOS(-/-) CD4(+) T cells were impaired in effector functions that lead to GVHD. In contrast, recipients of ICOS(-/-) CD8(+) T cells exhibited significantly enhanced GVHD morbidity and accelerated mortality. In the absence of ICOS signaling, either using ICOS-deficient donors or ICOS ligand-deficient recipients, the levels of expansion and Tc1 cytokine production of CD8(+) T cells were significantly increased. The level of expansion was inversely correlated with the level of apoptosis, suggesting that increased ability of ICOS(-/-) CD8(+) T cells to induce GVHD resulted from the enhanced survival and expansion of those cells. Our findings indicate that ICOS has paradoxical effects on the regulation of alloreactive CD4(+) and CD8(+) T cells in GVHD.     

Pt(II) and Ag(I) complexes with acesulfame: Crystal structure and a study of their antitumoral, antimicrobial and antiviral activities

Two new complexes of platinum(II) and silver(I) with acesulfame were synthesized. Acesulfame is in the anionic form acesulfamate (ace). The structures of both complexes were determined by X-ray crystallography. For K₂[PtCl₂(ace)₂] the platinum atom is coordinated to two Cl⁻ and two N-acesulfamate atoms forming a trans-square planar geometry. Each K⁺ ion interacts with two oxygen atoms of the S(O)₂ group of each acesulfamate. For the polymeric complex [Ag(ace)]_n the water molecule bridges between two crystallographic equivalent Ag1 atoms which are related each other by a twofold symmetry axis. Two Ag1 atoms, related to each other by a symmetry centre, make bond contact with two equivalent oxygen atoms. These bonds give rise to infinite chains along the unit cell diagonal in the ac plane. The in vitro cytotoxic analyses for the platinum complex using HeLa (human cervix cancer) cells show its low activity when compared to the vehicle-treated cells. The Ag(I) complex submitted to in vitro antimycobacterial tests, using the Microplate Alamar Blue (MABA) method, showed a good activity against Mycobacterium tuberculosis, responsible for tuberculosis, with a minimal inhibitory concentration (MIC) value of 11.6 μM. The Ag(I) complex also presented a promising activity against Gram negative (Escherichia coli and Pseudomonas aeruginosa) and Gram positive (Enterococcus faecalis) microorganisms. The complex K₂[PtCl₂(ace)₂] was also evaluated for antiviral properties against dengue virus type 2 (New Guinea C strain) in Vero cells and showed a good inhibition of dengue virus type 2 (New Guinea C strain) replication at 200 μM, when compared to vehicle-treated cells.     

Wild Sicilian Rosemary: Phytochemical and Morphological Screening and Antioxidant Activity Evaluation of Extracts and Essential Oils

To identify the best biotypes, an extensive survey of Sicilian wild rosemary was carried out by collecting 57 samples from various sites, followed by taxonomic characterization from an agronomic perspective. All the biotypes collected were classified as Rosmarinus officinalis L. A cluster analysis based on the morphological characteristics of the plants allowed the division of the biotypes into seven main groups, although the characteristics examined were found to be highly similar and not area‐dependent. Moreover, all samples were analyzed for their phytochemical content, applying an extraction protocol to obtain the nonvolatile components and hydrodistillation to collect the essential oils for the volatile components. The extracts were characterized by LC‐UV‐DAD/ESI‐MS, and the essential oils by GC‐FID and GC/MS analyses. In the nonvolatile fractions, 18 components were identified, namely, 13 flavones, two organic acids, and three diterpenes. In the volatile fractions, a total of 82 components were found, with as predominant components α‐pinene and camphene among the monoterpene hydrocarbons and 1,8‐cineole, camphor, borneol, and verbenone among the oxygenated monoterpenes. Cluster analyses were carried out on both phytochemical profiles, allowing the separation of the rosemary samples into different chemical groups. Finally, the total phenol content and the antioxidant activity of the essential oils and extracts were determined with the Folin–Ciocalteu (FC) colorimetric assay, the UV radiation‐induced peroxidation in liposomal membranes (UV‐IP test), and the scavenging activity of the superoxide radical (O$\rm{{_{2}^{{^\cdot} -}}}$ ). The present study confirmed that the essential oils and organic extracts of the Sicilian rosemary samples analyzed showed a considerable antioxidant/free radical‐scavenging activity.     

Karyopherins in nuclear pore biogenesis: a role for Kap121p in the assembly of Nup53p into nuclear pore complexes

The mechanisms that govern the assembly of nuclear pore complexes (NPCs) remain largely unknown. Here, we have established a role for karyopherins in this process. We show that the yeast karyopherin Kap121p functions in the targeting and assembly of the nucleoporin Nup53p into NPCs by recognizing a nuclear localization signal (NLS) in Nup53p. This karyopherin-mediated function can also be performed by the Kap95p-Kap60p complex if the Kap121p-binding domain of Nup53p is replaced by a classical NLS, suggesting a more general role for karyopherins in NPC assembly. At the NPC, neighboring nucleoporins bind to two regions in Nup53p. One nucleoporin, Nup170p, associates with a region of Nup53p that overlaps with the Kap121p binding site and we show that they compete for binding to Nup53p. We propose that once targeted to the NPC, dissociation of the Kap121p-Nup53p complex is driven by the interaction of Nup53p with Nup170p. At the NPC, Nup53p exists in two separate complexes, one of which is capable of interacting with Kap121p and another that is bound to Nup170p. We propose that fluctuations between these two states drive the binding and release of Kap121p from Nup53p, thus facilitating Kap121p's movement through the NPC.     

Bronchial malondialdehyde DNA adducts, tobacco smoking, and lung cancer

Tobacco smoking is a major risk factor for lung cancer causing, among other effects, oxidative stress and lipid peroxidation. Malondialdehyde (MDA)-DNA adducts can be induced by direct DNA oxidation and by lipid peroxidation. We measured the relationship between bronchial MDA-DNA adducts and tobacco smoking, cancer status, and selected polymorphisms in 43 subjects undergoing a bronchoscopic examination for diagnostic purposes. MDA-DNA adducts were higher in current smokers than in never smokers (frequency ratio (FR) = 1.51, 95% confidence interval (CI) 1.01-2.26). MDA-DNA adducts were also increased in lung cancer cases with respect to controls, but only in smokers (FR = 1.70, 95% CI 1.16-2.51). Subjects with GA and AA cyclin D1 (CCND1) genotypes showed higher levels of MDA-DNA adducts than those with the wild-type genotype (FR = 1.51 (1.04-2.20) and 1.45 (1.02-2.07)). Lung cancer cases with levels of MDA-DNA adducts over the median showed a worse, but not statistically significant, survival, after adjusting for age, gender, and packyears (hazard ratio = 2.48, 95% CI 0.65-9.44). Our findings reinforce the role of smoking in lung carcinogenesis through oxidative stress. Subjects who carry at least one variant allele of the CCND1 gene could accumulate DNA damage for altered cell-cycle control and reduced DNA repair proficiency.