Toxicity and kinetics of spinosad in different developmental stages of the endoparasitoid <Emphasis Type="Italic">Hyposoter didymator</Emphasis> (Hymenoptera: Ichneumonidae) and its host <Emphasis Type="Italic">Spodoptera littoralis</Emphasis> larvae (Lepidoptera: Noctuidae)

The toxicity of spinosad was evaluated using the RaPID Assay^® Spinosad immunosorbent assay in different developmental stages of the parasitoid, Hyposoter didymator, and in its host, fourth-instar larvae of the cotton leafworm Spodoptera littoralis. Spinosad was applied directly to pupae and adults of H. didymator (ingestion or topical application) or to the immature stages of the parasitoid via the host larvae. Low amounts of spinosad were recovered from S. littoralis host larvae after topical treatment, and the compound was mainly retained in the hemolymph. Amounts of spinosad detected in third-instar larvae of H. didymator, pulled out from the hemolymph of parasitized S. littoralis larvae, were 85 pg (3.57 ng a.i./g body weight) in dead larvae, and 82 pg (3.42 ng a.i./g body weight) in alive individuals. After topical treatment of H. didymator cocoons, most of the compound was retained in the silken cocoon, preventing contamination of the pupa. Also in the parasitoid adults, relatively low amounts of spinosad were accumulated in the body overall, but half of all the insecticide recovered was found in the ovaries. The kinetic results obtained help to better understand the toxicity of spinosad in the complex S. littoralis–H. didymator, and to ascertain the compatibility between spinosad and the parasitoid for optimizing the control of lepidopteran pests.     

Compatibility of sulfoxaflor and other modern pesticides with adults of the predatory mite <Emphasis Type="Italic">Amblyseius swirskii</Emphasis>. Residual contact and persistence studies

Amblyseius swirskii Athias-Henriot (Acari: Phytoseiidae) is a very efficient generalist predatory mite of Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) and Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae), worldwide released in horticultural greenhouses. Here, the toxicity of sulfoxaflor and other ten pesticides to A. swirskii adults when applied at their maximum field rate was assessed in the laboratory in terms of mortality and reproductive performance. The duration of the harmful activity when residues were aged under greenhouse was assessed for compounds not classified as harmless in the laboratory, based on the International Organization for Biological Control (IOBC) rules. Sulfoxaflor as well as flonicamid, flubendiamide, metaflumizone, methoxyfenozide, spiromesifen, and spirotetramat were harmless, emamectin was slightly harmful and abamectin, deltamethrin and spinosad were harmful. Emamectin was short-lived and abamectin, deltamethrin and spinosad were slightly persistent under our conditions.     

African Swine Fever Virus Structural Protein pE120R Is Essential for Virus Transport from Assembly Sites to Plasma Membrane but Not for Infectivity

This report examines the role of African swine fever virus (ASFV) structural protein pE120R in virus replication. Immunoelectron microscopy revealed that protein pE120R localizes at the surface of the intracellular virions. Consistent with this, coimmunoprecipitation assays showed that protein pE120R binds to the major capsid protein p72. Moreover, it was found that, in cells infected with an ASFV recombinant that inducibly expresses protein p72, the incorporation of pE120R into the virus particle is dependent on p72 expression. Protein pE120R was also studied using an ASFV recombinant in which E120R gene expression is regulated by the Escherichia coli lac repressor-operator system. In the absence of inducer, pE120R expression was reduced about 100-fold compared to that obtained with the parental virus or the recombinant virus grown under permissive conditions. One-step virus growth curves showed that, under conditions that repress pE120R expression, the titer of intracellular progeny was similar to the total virus yield obtained under permissive conditions, whereas the extracellular virus yield was about 100-fold lower than in control infections. Immunofluorescence and electron microscopy demonstrated that, under restrictive conditions, intracellular mature virions are properly assembled but remain confined to the replication areas. Altogether, these results indicate that pE120R is necessary for virus dissemination but not for virus infectivity. The data also suggest that protein pE120R might be involved in the microtubule-mediated transport of ASFV particles from the viral factories to the plasma membrane.     

Polyprotein processing in African swine fever virus: a novel gene expression strategy for a DNA virus.

This report shows that African swine fever virus (ASFV)--a large DNA-containing virus--synthesizes a polyprotein to produce several of its structural proteins. By immunoprecipitation analysis, we have found that ASFV polyprotein is a 220 kDa myristoylated polypeptide (pp220) which, after proteolytic processing, gives rise to four major structural proteins: p150, p37, p34 and p14. Processing of the ASFV polyprotein takes place at the consensus sequence Gly-Gly-X and occurs through an ordered cascade of proteolytic cleavages. So far, polyprotein processing as a mechanism of gene expression had been found only in positive-strand RNA viruses and retroviruses. According to the results presented here, ASFV is the first example of a DNA virus that synthesizes a polyprotein as a strategy of gene expression.     

Sequence and characterization of the major early phosphoprotein p32 of African swine fever virus.

The gene encoding protein p32, the most abundant and immunogenic protein induced by African swine fever virus at early times of infection, has been mapped in the EcoRI C' fragment of the genome of the Vero cell-adapted virus strain BA71V. Sequencing analysis has shown the existence of an open reading frame, named C'204L, encoding 204 amino acids. The protein is phosphorylated in serine residues located in the 115 N-terminal amino acids and was phosphorylated when expressed in cells infected with a vaccinia virus recombinant. Protein p32 is not glycosylated in spite of the presence of two putative N-glycosylation sites in the deduced amino acid sequence of the polypeptide. Immunofluorescence experiments have shown that the protein is localized in the cytoplasm of infected cells and not in the plasma membrane. In addition, the protein has been found in the soluble fraction and not in microsomes from BA71V-infected Vero cells. Low levels of the protein have been detected in the medium from infected swine macrophages, which probably corresponds to nonspecific release of cytoplasmic proteins. The protein encoded by other virus isolates shows different electrophoretic mobilities, indicating variability of p32.     

Nest Defence of Nesting Chinstrap Penguins (Pygoscelis antarctica) against Intruders

We tested some predictions of parental investment theory by studying the aggressive behaviour of colonial nesting chinstrap penguins (Pygoscelis antarctica) against human intruders into their nesting territories. We tested for differences in the aggressive behaviour of penguins according to offspring age (eggs vs. chicks), offspring number, nest location in the colonies (central vs. peripheral) and sex. Offspring age was the main factor influencing nest defence, although nest location and sex were also important. Chicks were defended more strongly than eggs, in accordance with changes in the reproductive value of offspring, and this increase in aggressiveness was not related to revisitation of the same individuals. The level of aggression of penguins breeding in central sites was higher than that of peripheral birds, a difference that could be due to the lower residual reproductive value of central-nesting, probably older, birds. The stronger aggressiveness of males could be due to a combination of factors related to sexual selection and life-history traits. Offspring number did not affect the level of nest defence.     

Comparison of the sequence of the gene encoding African swine fever virus attachment protein p12 from field virus isolates and viruses passaged in tissue culture.

Comparison of the amino acid sequence of the African swine fever virus attachment protein p12 from different field virus isolates, deduced from the nucleotide sequence of the gene, revealed a high degree of conservation. No mutations were found after adaptation to Vero cells, and a polypeptide with similar characteristics was present in an IBRS2-adapted virus. The sequence of the 5' flanking region was conserved among the isolates, whereas sequences downstream of the gene were highly variable in length and contained direct repeats in tandem that may account for the deletions found in different isolates. Protein p12 was synthesized in swine macrophages infected with all of the viruses tested.     

Genetic analysis of bacteriophage phi 29 of Bacillus subtilis: integration and mapping of reference mutants of two collections.

Reference mutants of Bacillus subtilis phage phi 29 of the Madrid and Minneapolis collections were employed to construct a genetic map. Suppressor-sensitive and temperature-sensitive mutants were assigned to 17 cistrons by quantitative complementation. Three-factor crosses were used to assign an unambiguous order for the 17 cistrons. Recombination frequencies determined by two-factor crosses were used to construct a linear genetic map of 24.4 recombination units. The genes were numbered sequentially from left to right (1 to 17) according to their relative map position.     

Mapping of temperature sensitive mutants of bacteriophage phi 29

Summary Temperature-sensitive mutants of eleven complementation groups of phage 29 have been mapped by means of two-factor crosses. The results show the existence of a single non-circular linkage map. Cistrons expressed early after infection are clustered at the left end of the map.     

Characterization of a protein covalently linked to the 5' termini of the DNA of Bacillus subtilis phage phi29.

We have isolated a covalent DNA-protein complex from bacteriophage φ29 particles. Polyacrylamide gel electrophoresis and tryptic peptide analysis showed that the protein present in the complex is very similar or identical to p3, an early induced protein essential for viral DNA replication.When the DNA-protein complex is treated with the restriction endonuclease EcoRI, the protein is specifically associated to the two terminal fragments, A and C. The protein is probably linked to the 5′ termini of the DNA since proteinase K-treated DNA is resistant to phosphorylation with polynucleotide kinase, even after treatment with alkaline phosphatase, while it is sensitive to exonuclease III. By electron microscopy the protein is visualized as a dot located at the ends of unit length DNA molecules.Mixed infection of Bacillus subtilis, at 42 °C, with ts² mutants in cistrons 2 and 3 only produces ts 2 progeny. This finding suggests that an inactive protein p3 bound to the DNA of the ts 3 mutant is not replaced by a functional protein and, as a consequence, replication of the ts 3 DNA does not occur.