In vitro metabolism of the trichothecene 4-monoacetoxyscirpenol by fungus- and non-fungus-feeding insects

Summary The metabolism of the trichothecene 4-monoacetoxyscirpenol by intact gut tissue was determined in the fungus-feeding Nitidulid,Carpophilus hemipterus (L.) and the non-fungus-feeding caterpillars, the fall armyworm,Spodoptera frugiperda (J. E. Smith) and the corn earworm,Heliothis zea (Boddie). The primary metabolite was the hydrolysis product scirpentriol. The amount of metabolism by theC. hemipterus larvae was ca 10 times that of the caterpillars on a per mg protein basis, suggesting metabolic adaptation for feeding on fungi that may contain mycotoxins.Acknowledgment. The authors wish to thank S. Taylor for technical assistance.The mention of firm names or trade products does not imply that they are endorsed or recommended by the U.S. Department of Agriculture over other firms or similar products not mentioned.     

Is ATP synthesized by a vacuolar-ATPase in the extremely halophilic bacteria?

The proton-dependent synthesis of ATP was demonstrated in representative members of the generaHalobacterium, Haloarcula, andHaloferax. In all cases, synthesis was not inhibited by nitrate or N-ethylmaleimide, inhibitors of the vacuolar-like ATPase found in Archaea, but was affected by azide, an inhibitor of F₀F₁-ATP syntheses. These observations extend the earlier observations withHalobacterium saccharovorum and suggest that ATP synthesis in these organisms is brought about by an F₀F₁-APT synthase.     

The evolutionary history ofDrosophila buzzatii. V. Differential survivorship onOpuntia betweenD. buzzatii andD. serido

Summary Survival time ofDrosophila buzzatii adults on anOpuntia (prickly pear) medium was significantly longer than that of its nearest relativeD. serido. A significant difference was also found betweenD. buzzatii adults from two experimental populations, one of them fed onOpuntia rots for more than two years and another one kept on standardDrosophila medium for the same period of time. These results suggest that adult selection may be taking place in cactiphilicDrosophila in their natural habitats and could be responsible for the niche differentiation betweenD. buzzatii andD. serido.This work has been supported by the Comisión Asesora de investigación Científica y Técnica, Spain, by funds to project 4514/79 awarded to the third author.     

Partial sequence of the gene for a serine protease from a halophilic archaeumhaloferax mediterranei R4, and nucleotide sequences of 16S rRNA encoding genes from several halophilic archaea

A part of the gene coding for a halophilic serine protease from a halophilic archaeumHaloferax mediterranei R4 was amplified by PCR and its 672 nucleotide sequence was determined. Tentative translation to the amino acid sequence suggested that the enzyme was quite similar to halolysin produced by another halophilic archaeum strain 172P1. Nucleotide sequences of 16S rRNA encoding genes from 9 halophilic archaea were determined. Alignment of 19 sequences known so far showed that there are more than 20 positions carrying bases or deletions specific for each halobacterial genus:Halobacterium, Haloarcula, Haloferax, andHalococcus.     

A new 9,11-secosterol,stellettasterol from a marine spongeStelletta sp

A new 9,11-secosteroid, stellettasterol (1) was isolated from a Japanese marine sponge,Stelletta sp.; its structure was determined by spectroscopic analysis. All NMR signals of1 were unambiguously assigned by application of various 2D NMR techniques. Stellettasterol exhibited antifungal activity againstMortieralla ramannianus.Part 62 of the Bioactive Marine Metabolites series. Part 61: Fusetani, N., Takahashi, M., and Matsunaga, S., Tetrahedron, in press.     

Intergeneric interactions betweenEimeria separata (Apicomplexa) andNippostrongylus brasiliensis (Nematoda) in the rat

Summary Ova production inNippostrongylus brasiliensis infected rats was significantly greater than in rats singly infected with the helminth whenEimeria separata infections were introduced 4, 6 and 11 days postinoculation withN. brasiliensis. Patent periods were unaltered during concurrent infections. These results suggest that the presence ofE. separata affects helminth fecundity but does not increaseN. brasiliensis longevity as has been shown withE. nieschulzi.This study was supported by NIH MB RS Grant RRO8012-8.     

Fungal competition and mycotoxin production on corn

Summary Aflatoxin and secalonic acid D production in corn in laboratory and field by mixed cultures ofPencillium oxalicum andAspergillus flavus orA. parasiticus was lower than production by the pure cultures. However, mixed culture of these molds withFusarium spp. did not affect mycotoxin production.     

Development of DNA probes for cytotoxin and enterotoxin genes in enteric bacteria

Summary DNA probes to identify the genes encoding toxins in enteric bacteria have been developed. Use of these probes reduces the number of animals required for toxicity testing, as suspect bacteria can be directly tested for the presence of toxin. We have augmented the gene probes available by developing probes against theEscherichia coli enterotoxin LTII and shiga toxin fromShigella dysenteriae 1.The LTII gene fromE. coli 357900 was identified and characterised and a suitable internal probe was obtained. The LTII gene was found not to be common among enterobacteriae from various geographical locations. Isolates predominately of animal origin from Nigeria and Thailand hybridized with the probe.The shiga toxin gene was isolated fromS. dysenteriae 1 by a combination of in vivo and in vitro methods. An internal probe was identified and used against different serogroups ofShigella andE. coli isolated. The probe was found to hybridize withS. dysenteriae 1 isolates and also someS. flexneri andS. sonnei strains. Representatives were tested for toxin production and found to produce toxin at low levels.     

Suppression of phytohemagglutinin induced splenocyte proliferation during concurrent infection withEimeria nieschulzi andNippostrongylus brasiliensis

Summary Results suggest that infection withEimeria nieschulzi (Protozoa) interferes with splenocyte proliferation induced by infection withNippostrongylus brasiliensis (Nematoda).This study was supported by NIH MBRS Grant RR08012-15.     

Studies on the fungal phytotoxin victorin: Structures of three novel amino acids from the acid hydrolyzate

Summary The host-selective phytotoxin victorin, produced by the fungusCochliobolus victoriae, was found to be at least partially peptidic in nature, and did not contain victoxinine. The exact mass of the M-H ion was measured by FABMS as 795.1877. Derivatives of three major acid hydrolysis products were isolated. The structures of the corresponding amino acids were assigned as 2S,3R-3-hydroxyleucine, 5,5-dichloroleucine, and 3-hydroxylysine. A into victorin by the fungus in vivo.     

Endocytosis and inositol hexakisphosphate levels inras transformants ofDictyostelium discoideum amoebae

Summary Fluid-phase pinocytosis kinetics and lysosomal enzyme secretion parameters were measured inDictyostelium discoideum amoebae constructed from strain AX3 by transformation with a multicopy plasmid carrying either a normalras gene (ras-Gly12), a mutatedras gene (ras-Thr12) or by the vector carrying the geneticin resistance gene only (pDNEO2). It was found that the pinocytosis rate and extent as well as the lysosomal enzyme secretion were slightly different in the three strains. These changes, however, were related to minor modifications of the cellular volumes. The overall concentration of inositol hexakisphosphate was similar in the three strains.This work was supported in part by a grant from the Ligue Nationale Française contre le Cancer (n^o 880258) to MS, and by a grant from the Fonds National Suisse de la Recherche Scientifique (No. 3.623-087) to CR.     

Fungitoxicity ofm-fluorophenylalanine-containing peptides towardsPythium ultimum

Summary The tripeptide L-m-fluorophenylalanyl-L-alanyl-L-alanine was much more fungitoxic towardsPythium ultimum than the dipeptide L-m-fluorophenylalanyl-L-alanine orm-fluorophenylalanine. The fungitoxicity of the tripeptide was reduced by L-alanyl peptides and phenylalanine, but not by other amino acids. In contrast, the fungitoxicity ofm-fluorophenylalanine was unaffected by peptides, and was antagonized by several amino acids. These results suggest the effective delivery ofm-fluorophenylalanine into the cell by a tripeptide carrier.     

Competitiveness and communication for effective inoculation byRhizobium,Bradyrhizobium and vesicular-arbuscular mycorrhiza fungi

After a short summary on the ecology and rhizosphere biology of symbiotic bacteria and vesicular-arbuscular (VA) mycorrhiza fungi and their application as microbial inocula, results on competitiveness and communication are summarized. Stress factors such as high temperature, low soil pH, aluminium concentrations and phytoalexins produced by the host plants were studied withRhizobium leguminosarum bv.phaseoli andRhizobium tropici onPhaseolus beans. Quantitative data for competitiveness were obtained by usinggus ⁺ (glucoronidase) labelled strains, which produce blue-coloured nodules. ForPhaseolus-nodulating rhizobia, a group specific DNA probe was also developed, which did not hybridize with more than 20 other common soil and rhizosphere bacteria. Results from several laboratories contributing to knowledge of signal exchange and communication in theRhizobium/Bradyrhizobium legume system are summarized in a new scheme, including also defense reactions at the early stages of legume nodule initiation. Stimulating effects of flavonoids on germination and growth of VA mycorrhiza fungi were also found. A constitutive antifungal compound in pea roots, -isoxazolinonyl-alanine, was characterized.     

Spasmin-like proteins in various ciliates revealed by antibody to purified spasmins ofCarchesium polypinum

Summary It was found that some ciliates,Stentor, Spirostomum andBlepharisma, which can contract rapidly like the stalks of Vorticellidae, have Ca²⁺-binding proteins that are very similar to spasmins, in the immunological sense. The presence of spasmins in other Protozoa and in some Metazoa was also investigated.     

A potent attractant of zoospores ofAphanomyces cochlioides isolated from its host,Spinacia oleracea

A highly potent attractant of zoospores ofAphanomyces cochlioides, a causal fungus of the root rot disease of spinach (Spinacia oleracea), was isolated from spinach roots, and its structure was determined by spectroscopic evidence and chemical synthesis as cochliophilin A (5-hydroxy-6,7-methylenedioxyflavone,1). A chromosorb particle prepared by soaking in solution of1 showed a potent attracting activity toward the zoospores using concentrations of1 above 10^–9 or 10^–10 M.     

Cryopreservation of parasites

Summary In this review, advances in cryopreservation of helminth parasites are reported. Our own studies demonstrate that metacestodes ofEchinococcus multilocularis can be maintained in a viable state for at least 1–2 years by appropriate deep-freezing and storage in liquid nitrogen. Infective larvae of the nematodeToxocara canis cryopreserved for 1 week in liquid nitrogen were maintained after thawing in vitro in a chemically defined medium for 35 weeks. Although motility of previously deep-frozen larvae was reduced they produced secretory/excretory antigens of similar immunodiagnostic quality as those from unfrozen larvae. Whereas infective larvae of several species of trichostrongylids can be easily cryopreserved, the infective larvae of the cattle lungworm,Dictyocaulus viviparus, and muscle larvae ofTrichinella spiralis are more sensitive to damage by subzero temperatures. Therefore, survival rates after cryopreservation are low, but improvement of the cooling schedules appears to be feasible. It is concluded that cryopreservation of certain stages of helminth and protozoan parasites is a useful technique for long-term storage of defined isolates, which can contribute considerably to reducing the number of experimental animals usually required for serial passages.     

Endocrine-dependent expression of circadian clock genes in insects

Current models state that insect peripheral oscillators are directly responsive to light, while mammalian peripheral clock genes are coordinated by a master clock in the brain via intermediate factors, possibly hormonal. We show that the expression levels of two circadian clock genes, period (per) and Par Domain Protein 1 (Pdp1) in the peripheral tissue of an insect model species, the linden bug Pyrrhocoris apterus, are inversely affected by contrasting photoperiods. The effect of photoperiod on per and Pdp1 mRNA levels was found to be mediated by the corpus allatum, an endocrine gland producing juvenile hormone. Our results provide the first experimental evidence for the effect of an endocrine gland on circadian clock gene expression in insects. Received 31 October 2007; received after revision 7 January 2008; accepted 9 January 2008 D. Dolezel, L. Zdechovanova: These authors contributed equally to this work.     

Aberrant,canavanyl protein formation and the ability to tolerate or utilize L-canavanine

Summary L-Canavanine, 2-amino-4-(guanidinooxy)butyric acid, and L-arginine incorporation into de novo synthesized proteins was compared in six organisms. Utilizing L-[guanidinooxy¹⁴C]canavanine and L-[guanidino¹⁴C]arginine at substrate saturation, the canavanine to arginine incorporation ratio was determined in de, novo synthesized proteins.Caryedes brasiliensis andSternechus tuberculatus, canavanine utilizing insects;Canavalia ensiformis, a canavanine storing plant; and to a lesser extentHeliothis virescens, a canavanine resistant insect, failed to accumulate significant canavanyl proteins. By contrast,Manduca sexta, a canavanine-sensitive insect, andGlycine max, a canavanine free plant, readily incorporated canavanine into newly synthesized proteins. This study supports the contention that the incorporation of canavanine into proteins in place of arginine contributes significantly to canavanine's antimetabolic properties.     

Preferential destruction of chloroplast nucleoids in zygotes in green algaeDictyosphaeria cavernosa andAcetabularia calyculus

Summary The preferential destruction of chloroplast nucleoids in young zygotes in the coenocytic algaDictyosphaeria cavernosa and the giant unicellular algaAcetabularia calyculus was studied by high resolution epifluorescent microscopy. The chloroplast nucleoids (DNA) in the chloroplast from one of the parents were preferentially destroyed soon after the mating of male and female gametes.     

Enhanced production of aggregation pheromones in four stored-product coleopterans feeding on methoprene-treated oats

Summary Production of aggregation pheromones by maleOryzaephilus surinamensis, O. mercator, Cryptolestes ferrugineus, andTribolium castaneum was enhanced by feeding on methoprene-treated oats, implicating juvenile hormone in control of pheromone production. Methoprene application to control insects in stored food products may cause enhanced pheromone production by these insects, thus drawing additional beetles into the treated product.Acknowledgments. We thank Dr. G. B. Staal of Zoëcon Corp. for a generous gift of methoprene. Particular thanks are due to J. Dodic for technical assistance. Research funded by the Natural Sciences and Engineering Research Council of Canada and the Science Council of British Columbia.