Development of a viral biopesticide for the control of the Guatemala potato tuber moth Tecia solanivora.

The Guatemala potato tuber moth Tecia solanivora (Povolny) (Lep. Gelechiidae) is an invasive species from Mesoamerica that has considerably extended its distribution area in recent decades. While this species is considered to be a major potato pest in Venezuela, Colombia, and Ecuador, currently no specific control methods are available for farmers.

Experimental mixtures of Phthorimaea operculella granulovirus isolates provide high biological efficacy on both Phthorimaea operculella and Tecia solanivora (Lepidoptera: Gelechiidae).

The Guatemalan potato moth Tecia solanivora (Povolny) recently invaded part of South America, colonizing zones where Phthorimaea operculella (Zeller), another potato moth species belonging to the same group, was previously established. T. solanivora is now the major insect pest of potato in this area encompassing Venezuela, Colombia and Ecuador.

Genetic and biological analysis of Colombian Phthorimaea operculella granulovirus isolated from Tecia solanivora (Lepidoptera: Gelechiidae).

Tecia solanivora (Lepidoptera: Gelechiidae) is an invasive potato pest of the north of South America that recently colonized zones where Phthorimaea operculella (Lepidoptera: Gelechiidae), a taxonomically related insect, was established. Nowadays, both species can be found in most areas in different proportions. The Phthorimaea operculella granulovirus (PhopGV) was found to efficiently control P.

Cydia pomonella granulovirus genotypes overcome virus resistance in the codling moth and improve virus efficiency by selection against resistant hosts.

Cydia pomonella granulovirus (CpGV) has been used for 15 years as a bioinsecticide in codling moth (Cydia pomonella) control. In 2004, some insect populations with low susceptibility to the virus were detected for the first time in southeast France. RGV, a laboratory colony of codling moths resistant to the CpGV-M isolate used in the field, was established with collection of resistant insects in the field followed by an introgression of the resistant trait into a susceptible colony (Sv).

An isometric virus of the potato tuber moth Tecia solanivora (Povolny) (Lepidoptera: Gelechiidae) has a tri-segmented RNA genome.

A small isometric virus has been isolated from larvae of the Guatemala potato tuber moth, Tecia solanivora (Povolny), collected in Ecuador. It was designated the Anchilibi virus (AnchV). The non-enveloped viral particles have an estimated diameter of 32+/-2 nm.

Characterization and authentication of insect cell lines using RAPD markers.

Random amplified polymorphic DNA (RAPD) analysis was used to characterize 11 insect cell lines, including six from lepidoptera (five species), one from diptera and four from coleoptera (one species: Leptinotarsa decemlineata). Whatever the order and even when comparing two closely related species from the same genus (Spodoptera), the DNA fingerprints are very different from one species or from one primer to the other. On the other hand, two independently isolated cell lines from the lepidopteran Phthorimaea operculella produce nearly identical profiles with only minor differences.

Characterization of cell lines developed from the Colorado potato beetle, Leptinotarsa decemlineata Say (Coleoptera: Chrysomelidae).

In order to isolate new pathogens (viruses, microsporidia, etc.) or to evaluate the efficiency of some pathogens (serovarieties and mutants of Bacillus thuringiensis, fungi, etc.) in the control of Colorado potato beetle, an economically important pest, we established four cell lines from tissues of this insect.

Detection of Phthorimaea operculella granulovirus in vivo and in vitro by a specific DNA probe.

In analyzing populations of non-infected potato tuber moth (PTM) Phthorimaea operculella, using a total DNA probe from Phthorimaea operculella granulovirus (PhopGV), false positive reactions were obtained indicating homology between cellular and viral DNAs. Using a cloned 2.1 kbp fragment of PhopGV DNA, a specific digoxigenin-labeled probe was developed.

DNA content analysis of insect cell lines by flow cytometry.

The DNA content of insect cell lines (6 lepidoptera, 1 coleoptera and 1 diptera) was determined by flow cytometry. The DNA profiles of the 8 cell lines tested were different. They were characterized by the presence of several peaks (2 to 7) corresponding to different ploidy levels, by differences in the fluorescence intensity of each peak and by the proportion of cells in each peak.

Genetic heterogeneity of Phthorimaea operculella granulovirus: restriction analysis of wild-type isolates and clones obtained in vitro.

Genetic heterogeneity of a wild-type granulovirus (Tunisia isolate) of the potato tuber moth Phthorimaea operculella (Phthorimaea operculella granulovirus, Phop GV) has been studied. The heterogeneity was indicated by the presence of several submolar fragments in the profiles obtained by use of several restriction endonucleases. It was also demonstrated by variations in the restriction profile of the wild-type Tunisia isolate that had underwent since 1991 in our laboratory numerous passages in vivo.

Evidence for two small viruses persistently infecting established cell lines of Phthorimaea operculella, deriving from embryos of the potato tuber moth.

Two small viruses were isolated from established cell lines of P. operculella deriving from embryos. The first one probably related to the Nodaviridae family, is a 30 nm in diameter icosahedral virus, with a bisegmented RNA genome and a single polypeptide of 39 kilodaltons.

Yield and activity of Autographa californica multicapsid nucleopolyhedrovirus and Phthorimaea operculella granulosis virus in cloned and uncloned cell lines of P. operculella.

Three selected uncloned Pop 2, Pop 3, Pop 4 and two cloned cell lines Pop cl1A and Pop cl2B were derived from the original cell line established from Phthorimaea operculella (ORS-Pop-93). Three new non-selected cell lines ORS-Pop-94A, ORS-Pop-94B and ORS-Pop-95 were also established from embryos of the same insect. Differences in morphology, growth rate and polypeptide profile were determined between these cell lines.

Multiplication of a granulosis virus isolated from the potato tuber moth in a new established cell line of Phthorimaea operculella.

A newly established cell line was obtained from the culture of embryonic cells of the potato tuber moth Phthorimaea operculella in low temperature conditions (19 degrees C) using modified Grace's medium supplemented with 10% fetal bovine serum. The population doubling time was about 80 h when cells were cultivated at 19 degrees C and 38 h at 27 degrees C. The cell line had a relatively homogeneous population consisting of various sized spherical cells.

Multiplication of Spodoptera littoralis granulosis virus in a cell line established from Phthorimaea operculella.

A complete replication of the Spodoptera littoralis granulosis virus (SpliGV) was obtained, in vitro by both virus infection and DNA transfection in the ORS-Pop-95 (Pop-95) cell line established from embryonic cells of the potato tuber moth, Phthorimaea operculella. SpliGV multiplied significantly during several passages in Pop-95 cells at 19 degrees C. When the cells were infected and kept at 19 degrees C for the first 4 hrs and then at 27 degrees C for the rest of the experiment (20 days), the viral multiplication proceeded at the same rate.

A new small RNA virus persistently infecting an established cell line of Galleria mellonella, induced by a heterologous infection.

A persistent infection in a Galleria mellonella cell line was revealed when infected with a maize stem borer picorna-like virus isolated on Sesamia cretica (MSBV). The new virus, completely different from the MSBV, is designated as G. mellonella cell line virus (GmclV), induces spectacular cytopathic effects, and is also considered efficient in vivo.

Establishment of a cell line derived from embryos of the potato tuber moth phthorimaea operculella (Zeller).

A cell line from the main insect pest of potatoes in tropical and subtropical areas, Phthorimaea operculella (Zeller), was obtained from embryoculture. These cells were cultured in Grace's modified medium. The cell line, designated ORS-Pop-93, had a heterogeneous population consisting of spherical and spindle cells with great capacity to adhere and a doubling time of 40 h.

Restriction map of the Casphalia extranea densovirus genome.

A physical map of the Casphalia extranea densovirus genome (CeDNV) was constructed. The size of the intact viral genome was estimated to be 4.9 kilobases or 1.

Expression of epidermal differentiation antigens in cultures of interspecific somatic hybrids (human keratinocytes X mouse fibroblasts 3T3-4E).

Interspecific somatic hybrids have been prepared by fusion of human epidermal cells with mouse fibroblasts 3T3-4E using PEG 4000. Expression of epidermal differentiation antigens (bullous pemphigoid antigens, BP, keratin subsets 55-57 k and 67 k), markers of basal and suprabasal cells, were studied by immunocytochemistry for 10 passages. These markers were detected in the hybrids early after fusion, indicating that cells from both compartments were able to fuse with 3T3-4E cells.

[Interspecies somatic hybrid cultures from human epidermal cells and 3T3-4E mouse fibroblasts].

Interspecific somatic hybrids were obtained by fusion of adult human epidermal cells with Mouse fibroblasts 3T3-4E, deficient in thymidine kinase. These hybrids were identified by their morphology and by the presence of markers from the parental cells. Some characters of keratinocytes such as keratin subunits 50 and 51 K were present in primary cultures and disappeared after serial passages, whereas bullous pemphigoid basement membrane zone antigens persisted for at least 20 passages.

Identification of shared antigenic determinants of different polypeptides from Davis and Towne cytomegalovirus strains with monoclonal antibodies.

Six colonies producing antibodies were obtained by fusing mouse myeloma SP2-O cells with spleen cells from mice immunized with cytomegalovirus Davis strain. Among 88 surviving clones, 29 produced antibodies detectable by immunofluorescence on infected MRC5 cells and 2 others produced neutralizing antibodies against a homologous virus. Supernatants from these 31 positive clones and 4 others which were negative in immunofluorescence or neutralization were tested for their capacity to bind polypeptides from labelled Davis-infected cell extracts.

Antigenic polypeptides of 3 reference strains recognized by sera from cytomegalovirus (CMV) infected patients.

The polypeptides of Cytomegalovirus (CMV) from 3 references strains (Davis, Ad 169 and Towne) were precipitated by immune sera from 5 renal transplanted patients with CMV-re-infection and 2 sera from patients with primary infection (one renal transplant and one child). Two antibody negative sera (tested in complement fixation test and immunofluorescence) were used as controls. The patterns of antigenic late polypeptides depended on the kinetics of in vitro infection.