Isolation and sequencing of insecticidal peptides from the primitive hunting spider, Plectreurys tristis (Simon).

About 50 peptide toxins were purified from venom of the primitive hunting spider, Plectreurys tristis. Bioassay by injection into larval Heliothis virescens (tobacco budworm) allowed selection of nine toxins for analysis of amino acid sequences. Total sequences were determined for six of the more insecticidal peptides (46-49 amino acids) and three contained free carboxyl-terminal amino acids. These toxins (plectoxins) are paralytic and/or lethal when injected into insect pests such as the lepidopterous larvae H. virescens, Spodoptera exigua (beet armyworm), and Manduca sexta (tobacco hornworm). We expect these plectoxins to be useful in insect control after delivery by a suitable agent such as a recombinant baculovirus.

Paralytic peptides from hemolymph of the lepidopteran insect Trichoplusia ni Hubner.

1. Two paralytic peptides were isolated and identified from hemolymph of Trichoplusia ni Hubner. 2. These peptides cause rapid, rigid paralysis when injected into the lepidopteran insect, Manduca sexta. 3. The paralytic dose (ED50) of one peptide is 0.05 micrograms/g and the other is 0.9 micrograms/g. 4. Sequences are very similar to paralytic peptides from the hemolymph of other lepidopteran insects.

Identification of insecticidal peptides from venom of the trap-door spider, Aptostichus schlingeri (Ctenizidae).

Nine insecticidal peptides were isolated from the venom of the trap-door spider, Aptostichus schlingeri. Seven of these toxins cause flaccid paralysis of insect larvae within 10 min of injection and all were lethal within 24 hr. The complete amino acid sequences (32-76 residues) of six peptides are presented. The identified peptides contain three or four disulfide bonds and the larger peptides (74-76 residues) are quite similar in sequence with carboxyl termini as free acids, not amidated.

Isolation and identification of paralytic peptides from hemolymph of the lepidopteran insects Manduca sexta, Spodoptera exigua, and Heliothis virescens.

Seven paralytic peptides were isolated and identified from lepidopteran hemolymph. All of these peptides cause rapid, rigid paralysis when injected into Manduca sexta and some other lepidopteran larvae. Each peptide contains 23 amino acid residues including 2 cysteines and the carboxyl termini are acidic. Synthetic peptides in the disulfide or reduced forms, and as carboxyl-terminal acids or amides were equally paralytic. The most potent paralytic peptide, Mas PP I, has the following sequence: H-Glu-Asn-Phe-Ala-Gly-Gly-Cys-Ala-Thr-Gly-Tyr-Leu- Arg-Thr-Ala-Asp-Gly-Arg-Cys-Lys-Pro-Thr-Phe-OH. The two peptides from M. sexta hemolymph are remarkable in that they are autoparalytic (i.e. factors in collected hemolymph that are paralytic when injected into the same larvae).

Paralytic and insecticidal toxins from the funnel web spider, Hololena curta.

One peptide and ten acylpolyamine toxins (curtatoxins) were purified and identified from venom of Hololena curta. The acylpolyamines consist of six different polyamines which are amidated with three different aromatic acids: (3-indolyl)acetic, (4-hydroxy-3-indolyl)acetic and 2.5-dihydroxybenzoic acids. These acylpolyamines instantly paralyze lepidopteran larvae following injection. The most potent insecticidal peptide in H. curta venom contains 38 amino acids and is lethal at 4 micrograms/g when injected into Manduca sexta larvae.

Structures of paralytic acylpolyamines from the spider Agelenopsis aperta.

The structures are given for five paralytic acylpolyamines from the venom of the funnel web spider, Agelenopsis aperta. The acyl moieties are derived from (3-indolyl)acetic acid, (4-hydroxy-3-indolyl)acetic acid, and 4-hydroxybenzoic acid. The polyamine portions of the toxins are novel. Three toxins (AG489, AG505, and AG452) contain 1, 5, 9, 13, 18, 22-hexaazadocosane which is unique as a natural polyamine because of its length and hydroxylation at the 5-aza position. The polyamine portions of two other alpha-agatoxins (AG488 and AG504) are unusual also, containing guanidinooxy moieties.

Chemical characterization of acylpolyamine toxins from venom of a trap-door spider and two tarantulas.

Two acylpolyamines are identified from venom of the trap-door spider, Hebestatis theveniti. These toxins (paralytic to lepidopteran insect larvae) are amides containing 3-(3-indolyl)lactic acid joined to spermine or 1,13-diamino-4,10-diazatridecane (Het389 and Het403, respectively). Het389 is also abundant in venom from a tarantual from Mozambique (Harpactirella sp.). Two additional acylpolyamines (Apc600 and Apc728) are partially characterized from venom of another tarantula, Aphonopelma chalcodes.

Purification and characterization of two classes of neurotoxins from the funnel web spider, Agelenopsis aperta.

Two classes of paralytic toxins were isolated from the venom of Agelenopsis aperta and their chemical and larvicidal properties characterized. Five acylpolyamine toxins (alpha-agatoxins) of molecular masses 452, 488, 489, 504, and 505 Da produce immediate but reversible paralysis in Manduca sexta following injection. Six insecticidal peptides (mu-agatoxins) produce a gradual but irreversible paralysis. The complete amino acid sequences (36-38 residues) of the mu-agatoxins are presented. These peptides contain eight half-cystines and are quite similar in sequence. At least four of these toxins are amidated at the carboxyl terminus. The secondary structure of one of these toxins (mu-Aga V) was investigated.

O-dephenylation and conjugation with benzoylornithine. New metabolic pathways for 3-phenoxybenzoic acid in chickens.

Metabolism of [benzyl-14C]fluvalinate by chickens produces 3-phenoxybenzoic acid, which is further degraded by two new pathways. The first pathway involves O-dephenylation, not reported previously for related pyrethroids in birds or mammals. O-Dephenylation is a major metabolic route (12% of the applied 14C). In the second pathway, 2% of the applied dose is converted into four conjugates of benzoylornithine (two with 3-hydroxybenzoic and two with 3-phenoxybenzoic acids). The predominant conjugate with benzoylornithine is N2-(3-hydroxybenzoyl)-N5-benzoylornithine.

The fate of dienochlor administered orally and dermally to rats.

Within four days of receiving a single oral dose (1 mg/kg) of [U-ring-14C]dienochlor [bis(pentachloro-2,4-cyclopentadien-1-yl)] female rats excreted 2 and 88% of the applied 14C in urine and feces, respectively. Metabolites could not be identified and the preponderance of the fecal radioactivity consisted of unextractable 14C-labeled residues. Within 1 day virtually all of the dienochlor had been degraded by rats, with only traces of parent dienochlor in excreta and tissues. After four days only 2% of the applied dose remained in tissues (mainly kidney, liver, and gastrointestinal tract). Pharmacokinetic studies with blood plasma and bile showed dienochlor (and/or its metabolites) to be poorly absorbed. Rats were exposed dermally for 24 hr to [14C]dienochlor formulated as Pentac WP miticide both as an aqueous suspension and as an undiluted wettable powder. Half of the dose adhered to the skin and the other half was found in gauze patches used to protect the treated skin. After a 24-hr exposure over 60% of the radiolabel that adhered to skin was removed by washing with an aqueous soap solution and 86% of this rinsing solution was unmetabolized dienochlor. The dienochlor and its metabolites were transported inefficiently from the application site; only 1% of the applied dose was detected in urine plus feces and less than or equal to 0.2% in tissues. With application rates that simulate field exposure by humans, the actual residue of dienochlor and metabolites in skin (i.e., not removable by washing) is about thirteen times higher following exposure to dienochlor as undiluted wettable powder than as an aqueous suspension.