Geminivirus DNA replication and cell cycle interactions

The Geminiviridae family includes a large number of viruses that infect plants and have a unique geminate virion particle, a single-stranded genome of approximately 2.6-3.0 kb, and replicate through a rolling-circle mechanism. Since they encode for just a few proteins (4-6 depending on the members that belong to four different genera), a rich variety of interactions has evolved between viral proteins and host factors to develop the virus replicative cycle. Among them, we have been particularly interested so far: (i). in the interference with cell cycle regulatory proteins of the retinoblastoma-related (RBR)/E2F pathway and (ii). in the interaction with host DNA replication factors necessary for the assembly of a functional replication complex at the viral origin of DNA replication during the rolling-circle stage. Yeast two-hybrid assays revealed that wheat dwarf virus RepA protein, but nor Rep protein, interacts with plant RBR protein. Interestingly, deletion of the C-terminal domain of Rep confers the truncated protein the ability to interact with RBR, suggesting that this domain may hinder the LXCXE RBR-binding motif. Secondary structure predictions support such a possibility.     

Growth and mineral absorption in maize seedlings as affected by increasing NaCl concentrations

The influence of NaCl solutions of decreasing osmotic potentials (¥π = ‐0.44 and ‐0.88 MPa) on seedling growth and on the concentration of the most important macro‐ and micro‐nutrients in the shoots and roots of maize (Zea mays L., cv. Summer II) grown in Hoagland's solution in a growth chamber was studied. Salt stress was imposed on six‐day‐old seedlings for a three day period. Increasing NaCl concentrations induced a reduction in the leaf water potential and a significant decrease in the length and dry weight of the shoots, whereas these two parameters decreased in the roots only at the lowest osmotic potential.

Although the absorption and accumulation of nutrients upon salt stress differed in the two treatments depending on the plant tissue and nutrient, almost all of the macronutrients decreased in the roots and shoots, showing the lowest values at ¥π = ‐0.88 MPa. Sodium and Cl increased continuously, much more in the roots than in the shoots.

A different response was seen in the shoots and roots in terms of micronutrients. In the roots, almost all of the ions reached their maximum concentrations at ¥π = ‐0.88 MPa; whereas in the shoots, they decreased at ¥π = ‐0.44 MPa without undergoing any further decrease at the lowest osmotic potential.     

Efficacy of a combination of 10% imidacloprid/50% permethrin for the prevention of leishmaniasis in kennelled dogs in an endemic area

The efficacy of imidacloprid 10%and permethrin 50% (Advantix; Bayer AG, Germany) in a spot-on formulation was evaluated in the field as a control measure to prevent canine leishmaniasis (CanL) in dogs in an endemic area of southern Italy. In February 2005, out of 845 dogs initially tested for CanL, 631 dogs which tested negative (315 from a kennel in Bari (KB) and 316 from a kennel in Ginosa (KG)) in a serological and a parasitological examination were allocated to one of three groups: Group A-treated with imidacloprid 10% and permethrin 50% once a month; Group B-treated every 2 weeks; and Group C-untreated control animals. All the dogs were examined serologically and parasitologically for CanL prior to the start of the study, in November 2005 (end of the sandfly season) and in March 2006 (end of the study). An initial CanL seroprevalence of 24.7% (209 dogs) was detected in KB and KG. In KB Leishmania infection, inferred by positivity in at least one of the three tests performed at the interim or final follow-up, was found in one animal from Group A and in nine from Group C. No positive animals were detected in Group B, thus giving a final protection efficacy of 88.9% in Group A and 100% in Group B. In KG Leishmania infection was identified in one animal from Groups A and B, respectively, and 11 from Group C (protection efficacy of 90.36% in Group A and 90.73% in Group B). The incidence density rates (IDRs) of infection in both Groups A and B at each kennel were statistically significantly lower than that registered in Group C (KB p<0.05 and KG p<0.01). The results clearly show that a combination of imidacloprid 10% and permethrin 50%, by virtue of its repellent activity against sandflies, is effective under both application regimes in preventing CanL in the field in endemic areas.     

Efficacy of a combination of imidacloprid 10%/permethrin 50% versus fipronil 10%/(S)-methoprene 12%, against ticks in naturally infected dogs

Preventing tick bites is a fundamental step towards reducing the impact of tick-borne protozoal, bacterial and viral diseases (TBDs) in humans and animals. The aim of this study was to evaluate the efficacy of a combination of imidacloprid 10%/permethrin 50% and of fipronil 10%/S-methoprene 12% against ticks in naturally infected dogs and to assess methodological parameters to calculate drug efficacy on tick immature stages.

From July to August 2004, 45 privately owned dogs of various sexes, ages, breeds, coat length and habits were enrolled in a trial carried out in an area (radius approximately 50 km) in Southern Italy. Three homogeneous groups (both for dog population and tick population) were formed: 15 dogs treated with imidacloprid 10% and permethrin 50% spot-on (group A), 15 dogs treated with fipronil 10% and methoprene 12% spot-on (group B) and 15 untreated dogs (group C). The dogs in each group were then sub-grouped according to their age and weight. Two different treatments were administered (time 0 and +28 days) to groups A and B, and the dogs were checked weekly for tick infestation until day +56 post-treatment (p.t.). Twenty-four areas distributed on the whole body surface were examined for ticks at each follow-up, while only at time 0 and at day +56 p.t., ticks were collected from the dogs and identified.

For the immature stages a semi-quantitative method was adopted and the load of immature stages was evaluated and grouped into four classes up to day +56 p.t. when the mean number of immature ticks (MIT) for each infection class was evaluated.

All the adult ticks collected were identified as brown dog ticks (Rhipicephalus sanguineus). Immature stages were first compared at day +28 p.t.. The efficacy of both products used in groups A and B on adult ticks was high and generally very similar. Conversely, the efficacy of imidacloprid 10% and permethrin 50% against immatures was higher than that of fipronil 10% and methoprene 12% throughout the observation period with statistically significant differences (p < 0.05) at day +28 p.t. (i.e. group A = 98.52%, group B = 72.40%).

On the whole, in analysing the efficacy of both products against adult plus immature ticks, it was found that the combination of imidacloprid 10% and permethrin 50% was more effective than fipronil 10% and methoprene 12%, with the differences being statistically significant at day +28 p.t. (group A = 98.43%, group B = 77.56%).     

Efficacy of moxidectin injectable and pour-on formulations in a pilot control program against bovine hypodermosis in Southern Italy

Bovine hypodermosis is a myiasis caused by Hypoderma bovis and Hypoderma lineatum (Diptera, Oestridae) larvae, which has a severe economic impact on the livestock industry. Though myiasis is widespread throughout Italy, no nationwide eradication program has ever been planned, unlike in other European Countries. With a view to setting up a national control program, a pilot study was carried out in Southern Italy on 9939 cattle bred in an area with a high prevalence of cattle hypodermosis, using moxidectin 0.5% pour-on (Cydectin, Fort Dodge) and 1% injectable (Cydectin, Fort Dodge) formulations. At the recommended dosage, moxidectin displayed efficacy levels of 99.9% in the pour-on and 100% in the injectable formulation, whereas the microdose (1 mg per head regardless of body weight) was less effective (65.7%). This trial contributed to a significant reduction in infestation rates in the study area and represented the first step through which a national program for eradicating warble fly infestation in Italy.     

Molecular characteristics of an immobilization antigen gene of the fish-parasitic protozoan Ichthyophthirius multifiliis strain ARS-6

Ichthyophthirius multifiliis (Ich), a ciliated protozoan parasite of fish, expresses surface antigens (i-antigens), which react with host antibodies that render them immobile. The nucleotide sequence of an i-antigen gene of I. multifiliis strain ARS-6 was deduced. The predicted protein of 47 493 Da is comprised of 460 amino acids (aa's) arranged into five imperfect repeats with periodic cysteine residues with the structure: CX₍₁₉₎₂₀CX₂CX₁₆₋₂₇CX₂CX₂₀₍₂₁₎CX₃. The N-terminal aa's typify a signal peptide motif while a stretch of C-terminal aa's resemble a glycosyl–phosphatidyl–inositol (GPI)-anchor addition site. The degree of deduced i-antigen aa sequence identity of strain ARS-6 (GenBank accession # ACH87654 and # ACH95659) with other I. multifiliis i-antigen sequences present in GenBank ranges from 99% to 36% identity with 52 kDa i-antigens of I. multifiliis strain G5 (accession #s AAK94941 and AAK01661 respectively). Immunoblot analysis of i-antigens following exposure of I. multifiliis theronts to catfish anti- I. multifiliis immune serum did not show any appreciable alteration in i-antigen expression. The mechanism that regulates i-antigen expression in I. multifiliis remains a puzzling question.     

Illegal ingegno fishery and conservation of deep red coral banks in the Sicily Channel (Mediterranean Sea)

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Fish and seafood traceability based on AFLP markers: Elaboration of a species database

Several sociological, health and conservation arguments request a correct labelling of seafood products. Nowadays, molecular genetics is a useful tool for food chain traceability, particularly in regards to species identification. Among the variety of PCR-based molecular markers, AFLPs (Amplified Fragment Length Polymorphisms) have recently been used to investigate genomes of different complexities. This paper assesses the potential use of the AFLP technology to determine fish and seafood species in processed commercial products and domestic stocks. In particular a species database of fish, molluscs and crustaceans has been created with the aim to identify species of origin of seafood products by previously defined AFLP patterns. Different EcoRI and TaqI primer combinations were selected from 20 screened combinations in relation to the total number of detected fragments and polymorphic ones. Most informative combinations were E32/T32, E32/T33, E33/T33, E33/T37, E33/T38, E40/T33, E40/T37, E42/T32, E42/T37. The comparison of informative markers between unknown frozen or fresh products and reference samples has enabled the accurate identification of 32 different species. The taxonomic characterization has been performed either at the species or at the population level depending on the number of available individuals. AFLP variation at the population level is particularly helpful for the stock traceability of domestic strains. Size homoplasy was also investigated in one species to assess the rate of non-homologous comigrating fragments and to detect additional polymorphic markers to be used in stock identification. Results of Band Sharing Index (BSI) and percentage of polymorphic fragments are presented and are discussed in relation to the wide applicability of AFLPs both for fish and seafood safety and authenticity testing in such fields as food traceability and restocking management. The database, available upon request at nonnis@biol.unipr.it, will be continuously updated.