Isolation and characterization of myrmexins, six isoforms of venom proteins with anti-inflammatory activity from the tropical ant, Pseudomyrmex triplarinus.

Venom from the tropical ant, Pseudomyrmex triplarinus, has anti-inflammatory properties demonstrated by the carrageenin-induced edema animal mode. A multi-protein complex that inhibits edema was isolated from the venom and was further characterized by high performance liquid chromatography (HPLC), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) and amino acid sequencing. Although the complex exhibited a single band in SDS-PAGE electrophoresis, six proteins (isoforms) were resolved and purified to homogeneity and were designated myrmexin I-VI.

Partial biochemical characterization of venom from the ant, Pseudomyrmex triplarinus.

Venom from the ant, Pseudomyrmex triplarinus, contains 12 proteins with mol. wts of > 100,000-4200, and they constitute 41.5% of the dry weight.

Evaluation of a single oral dose of lufenuron to control flea infestations in dogs.

A single dose of lufenuron was administered to dogs to test its efficacy in controlling cat flea (Ctenocephalides felis) infestations for at least 30 days. Efficacy measurements revealed marked differences in the reproduction capability of fleas collected from dogs in the treatment vs the control group. Essentially, all of the eggs collected from dogs treated with lufenuron were unable to develop into normal adult fleas.

Toxicity of the venom from Nasonia vitripennis (Hymenoptera: Pteromalidae) toward fly hosts, nontarget insects, different developmental stages, and cultured insect cells.

A venom preparation from Nasonia vitripennis, a wasp ectoparasitoid of fly pupae, was assayed for lethality in different stages of insects representing ten different orders and in cultured insect cells. In most cases, the motor activity of the injected insects remained completely normal for 1-2 days after the injection and displayed none of the symptoms of paralysis commonly reported for venoms of the Hymenoptera. A natural host, the flesh fly Sarcophaga bullata, was highly sensitive in the pupal stage (LD50 = 5.

High-level expression using baculovirus, purification, and characterization of a monomeric form of type II calmodulin-dependent protein kinase.

The type II calmodulin-dependent protein kinase is an oligomer existing in multiple isozymic forms. To facilitate investigations of the regulatory mechanisms of this complex enzyme, we have constructed a truncated, calmodulin-dependent monomer of the alpha subunit. The N-terminal enzyme fragment (alpha 315) was expressed at high levels in a baculovirus/insect cell expression system.

A serum-free medium for the culture of insect cells and production of recombinant proteins.

A low protein aqueous lipid supplement (Ex-Cyte VLE), in combination with pluronic polyol, is an effective replacement for fetal bovine serum for insect Sf-9 cells. Serum-free medium with lipid supplement and pluronic (SFM-LP) supported higher cell viability and maximum cell populations than serum-supplemented medium. No adaptation procedures are required when switching cells from serum-containing medium to SFM-LP, and growth rates remain constant during continued passages in SFM-LP.

Effect of an experimental systemic compound, CGA-184699, on life stages of the cat flea (Siphonaptera: Pulicidae).

The experimental drug CGA-184699 (N-[2,5-dichloro-4-(1,1,2,3,3,3-hexafluoropropoxy)-phenylaminoc arbonyl]-2,6-difluorobenzamide) was evaluated for efficacy against cat fleas, Ctenocephalides felis (Bouché), held in flea cages on cats. When administered orally, the compound acts systemically and prevents development of the next generation of fleas. There was no effect on adults, but eggs from adults that fed on treated cats had reduced viability.

Expression of three recombinant proteins using baculovirus vectors in 23 insect cell lines.

Recombinant Autographa california baculoviruses expressing genes for pseudorabies virus glycoprotein (gp50T), human plasminogen (HPg), and beta-galactosidase (beta-gal) were used to infect 23 cell lines or strains. The objectives were to compare amounts of recombinant proteins expressed in the cell lines, compare yields from clones and parent lines, investigate the effects of long-term culture in serum-free medium on production, and determine if some lines yield gp50T with different glycosylation patterns. For HPg, IZD-MB0503 had the highest yield and four other lines (IPLB-TN-R2, IPLB-SF-1254, IPLB-LdEIta, and CM-1) had levels above that of SF-9 cells.

Protective effect of methylcellulose and other polymers on insect cells subjected to laminar shear stress.

The relative sensitivity of two insect cell lines to laminar shear stress was determined, and the protective effect of polymers added to the growth media of two insect cell lines, Trichoplusia ni (TN-368) and Spodoptera frugiperda (SF-9), was evaluated. TN-368 and SF-9 cells were found to be equally sensitive to laminar shear stress. Methylcellulose [0.

Physical mapping and field inversion gel electrophoresis of Amsacta moorei entomopoxvirus DNA.

Agarose in situ digestion was used to prepare intact Amsacta moorei entomopoxvirus (AmEPV) DNA from embedded occlusion bodies (OBs). Direct dissolution of OBs in agarose eliminated the necessity for separate purification of virions. A physical map of AmEPV DNA was constructed for five restriction enzymes (BamHI, EcoRI, HindIII, PstI, and XhoI) using single and multiple digests, and isolated fragment digestions.

Inhibition of human platelet aggregation and secretion by ant venom and a compound isolated from venom.

Venom from the tropical ant, Pseudomyrmex triplarinus, has activity against rheumatoid arthritis. Since platelets are involved in inflammatory responses, they were employed to study the effects of venom on prostaglandin-dependent human platelet aggregation and secretion. The assay is very sensitive and uses microliter volumes, which makes it useful as a screen during isolation and characterization of venom components.

Primary culture of ant venom gland cells.

Venom from the ant Pseudomyrmex triplarinus reduces the symptoms and swelling of rheumatoid arthritis. The cells that produce the venom were dissected from larval and pupal ants and culture conditions studied. Cell dissociation, with minimal amount of damage, was done with 0.

The isolation and characterization of the MP and FP plaque variants of Galleria mellonella nuclear polyhedrosis virus.

Two plaque morphologies were detected in assays of Galleria mellonella nuclear polyhedrosis virus preparations which had been serially passaged in the TN-368 cell line. The FP (few polyhedra) plaques were larger (0.7-mm average diameter) and the infected cells contained less than 10 polyhedra each.

Cell cycle analysis and synchronization of the TN-368 insect cell line.

A cell cycle analysis of the Trichoplusia ni (TN-368) insect cell line is described. By means of autoradiography and percent labeled metaphase data, the cell cycle parameters were determined to be as follows: S, 4.5 hr; G2, 8.

Altered phenotypic expression of ultraviolet-irradiated nuclear polyhedrosis virus.

UV (254 nm) irradiation of nuclear polyhedrosis virus results in formation of small plaques. The small-plaque effect is reversed by photoreactivation. Analysis of plaque formation kinetics indicates a slower formation and subsequent enlargement of plaques from UV-irradiated virus in comparison to non-irradiated virus.

A comparison of survival and repair of UV-induced DNA damage in cultured insect versus mammalian cells.

Survival and unscheduled DNA synthesis (UDS) were measured in a cultured insect cell line, TN-368, and a cultured mammalian cell line, V-79-4, following exposure to several fluences of ultraviolet light. TN-368 cells were approximately seven times more resistant to the lethal effects of UV than V-79 cells, as determined by colony formation. The amount of UDS per unit amount of DNA is about the same in both cell types 4 hr after 10-50 J/m2 UV irradiations.