Preliminary observations of a bacteriophage infecting Xenorhabdus luminescens (Enterobacteriaceae)

A bacteriophage infective to Xenorhabdus luminescens, a bacterial symbiont of heterorhabditid nematodes, was recovered from insects that supported poor nematode development. Plaque tests showed the phage particles to be infective only to primary and not secondary colonies of X. luminescens. The phage was not infective to X. nematophilus primaries or secondaries. The bacteriophage particles ranged 80-90 nm in length, with the head ranging from 40 to 50 nm in diameter. Restriction analysis was performed on isolated bacteriophage DNA. This first report of a bacteriophage from Xenorhabdus species has practical implications since it could be detrimental to cultures of Heterorhabditis nematodes that are being produced throughout the world for the biological control of insects.     

Infection of frog tadpoles (Amphibia) by insect parasitic nematodes (Rhabditida)

Summary Infective stage juveniles ofNeoaplectana carpocapsae (Steinernematidae) andHeterohabditis heliothidis (Heterorhabditidae) were able to penetrate through the alimentary tract of young tadpoles ofHyla regilla (Hylidae) andXenopus laevis (Pipidae) and enter the body cavity. Some infectives ofN. carpocapsae were able to release their symbiotic bacterium,Xenorhabdus nematophilus inside the host and in two cases, the nematodes developed into adult females before they perished. Tadpole mortality was associated with foreign bacteria entering the penetration holes made by the invading nematodes. The infective stage juveniles of both nematodes frequently encountered a host defense reaction upon reaching the tadpole's infective stage juveniles of both nematodes frequently encountered a host defense reaction upon reaching the tadpole's coelom.     

Development of malaria parasites in mosquitoes fed with ookinetes suspended in defined media

Information concerning the specific nutritional requirements of malarial parasites developing in the mosquito host has been difficult to obtain, owing primarily to the complex nature of the blood meal that accompanies the parasites and the lack of success in culturing the complete invertebrate cycle ofPlasmodium in vitro. The present report describes a blood-free system for infecting mosquitoes with ookinetes ofPlasmodium berghei and for allowing the latter to develop into infective sporozoites. Ookinetes cultured in vitro were separated from blood proteins, suspended in defined medium, and fed toAnopheles stephensi mosquitoes through a membrane. The mosquitoes were then maintained on the same defined medium plus 5% sucrose. Infectivity of the parasites was demonstrated 17–19 days later by intracardial inoculation of the macerated mosquitoes into hamsters. This system makes it possible to evaluate nutritional factors that affect parasite development in the mosquito host under controlled conditions.This project was supported, in part, by the Public Health Service research grant AI-18345 from the National Institute of Allergy and Infectious Diseases to Prof. K. Maramorosch, and the Charles and Johanna Busch Memorial Fund at Rutgers, The State University of New Jersey.     

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.     

Babesia bovis: the effect of acute inflammation and isoantibody production in the detection of babesial antigens

Summary Immunoblots ofBabesia bovis antigen contain dominant antigens which react not only with antisera toB. bovis but with sera from naive calves recovering from an acute inflammatory reaction. It seems likely these antigens are from the host rather than the parasite.     

Differences in chromosome A arrangement betweenDrosophila madeirensis andDrosophila subobscura

Summary The proximal half of the A (=X) chromosome ofD. madeirensis has a gene arrangement very similar to the A1 or A6 inversions found inD. subobscura. Polytene chromosome analysis of hybrids betweenD. madeirensis and strains ofD. subobscura homozygous for such inversions shows, however thatD. madeirensis has a gene arrangement different from any known forD. subobscura. These results provide evidence for a greater differentiation of the X chromosome in these species than has previously been described; it seems that the X chromosome is the only one that has undergone structural variation during the speciation process.     

Agglutinins from aquatic insects—tumor cell agglutination activity

Agglutinins were identified in whole body extracts of aquatic insects by means of murine tumor cell agglutination, using sarcoma 180 ascites, Ehrlich, and MM-46 cells. Screening revealed agglutinins in 5 of 10 of the larvae tested, and in 2 of 6 of the water-dwelling adult insects;Gerris paludum insularis andGyrinus japonicus. Only the agglutinin from adultG. paludum also agglutinated human erythrocytes. An ascites tumor was converted into a solid form in vivo after administration ofG. paludum agglutinin. The observation that these aquatic insect agglutinins preferentially agglutinate tumor cells has considerable implications in terms of anti-tumor effects such as inhibition of cell proliferation and metastasis.Deceased.     

Characterization of victorin C,the major host-selective toxin fromCochliobolus victoriae: Structure of degradation products

Summary Several host-selective toxins have been isolated in pure form culture filtrates ofCochliobolus victoriae. Acid hydrolysis of the major toxin, victorin C (apparent mol.wt 796), produced five fragments to which structures1–5 have been assigned. Spectroscopic techniques revealed that the toxin contains an additional subunit corresponding to6; thus all the components of victroin C are accounted for.     

Transgenic and mutant animal models to study mechanism of protection of red cell genetic defects against malaria

Malaria, caused by members of the genusPlasmodia, is still the most prevalent parasitic disease in the world. In an attempt to understand genetic factors conferring resistance to malaria, mouse models of thalassemia, sickle trait, and ankyrin and spectrin deficiency were studied during infection with species of malaria infectious to rodents. Although growth ofP. falciparum is not inhibited in thalassemic erythrocytes in culture, mice carrying a -thalassemia mutation were protected fromPlasmodium chabaudi adami, supporting epidemiologic findings. Transgenic mice expressing ^s hemoglobin were also significantly protected from two species of rodent malaria. Importantly, a significant role for the spleen in protection in the ^s transgenic mice was found. Finally, mice deficient in spectrin and ankyrin were studied with respect to their ability to support the growth of malaria. It was found that spectrin deficient mice were almost completely refractory toP. chabaudi adami andP. berghei. These models will allow further study of host factors in resistance to malaria.     

Long-term memory in territorial grasshoppers

Summary Males ofLigurotettix spp. grasshoppers (Orthoptera, Acrididae, Gomphocerinae), territorial species that defend individual host plant shrubs, home to their territories by memorizing landmarks. Thus, the insects can navigate efficiently toward high quality shrubs which are chemically distinctive and recognizable via gustation, but indistinguishable from a distance.     

Trypanosoma cruzi: metabolic labeling of trypomastigote sialoglycolipids

Summary Trypomastigote forms (infective) ofTrypanosoma cruzi incorporate (³H)-palmitic acid and D-(³H)-galactose into glycolipids. Palmitic acid-labeled acidic glycolipids were partially hydrolyzed with neuraminidase. The labeling of these compounds when the intact cell surface was labeled with galactose oxidase plus NaB³H₄ indicates the membrane location of the sialoglycolipids.This investigation received financial support from SECYT and CONICET to RML; CNPq, FINEP, and UNDP/World Bank/ WHOSSpecial Programme for Research and Training in Tropical Diseases to WC; and FAPESP to BZ and WC. ASC is a fellow from FAPESP.     

Antibacterial effect of 2-hydroxy-N-(3,4-dimethyl-5-isoxazolyl)-1,4-naphthoquinone-4-imine onStaphylococcus aureus

The mechanism by which a new naphthoquinone derivative, the 2-hydroxy-N-(3,4-dimethyl-5-isoxazolyl)-1,4-naphthoquinone-4-imine (INQI-E) has antibacterial effect againstStaphylococcus aureus was studied. The interaction of INQI-E with the bacteria was followed by absorption spectroscopy at 323 and 490 nm. The absorption band of INQI-E at 490 nm undergoes a hypochromic shift with a decrease of intensity. This effect was found to be reversible by oxygenation during the first hours of incubation. The participation of an oxidation-reduction process related to the respiratory chain was demonstrated by oxygen consumption. An increase in O₂ uptake and inhibition ofS. aureus growth was observed. Experiments with three inhibitors of the respiratory chain demonstrated that the pathway induced by INQI-E was antimycin-resistant and KCN- and salicylhydroxamic acid (SHAM)-sensitive, which suggests that INQI-E is capable of diverting the normal electron flow to an alternate superoxide-producing route. On the other hand, experiments with Tiron, a specific scavenger of superoxide, hindered the effect of INQI-E againstS. aureus, indicating that the inhibitory growth effect of this quinone-imine is mainly due to the production of the cytotoxic superoxide radical.     

Comparative aspects of structure and action of molluscan neuropeptides

A number of neuropeptides were isolated from the ganglia and muscles of molluscs, and their actions were examined. Diverse neuropeptides, in addition to several classical neurotransmitters, were suggested to be involved in the regulation of the anterior byssus retractor muscle ofMytilus. A wide structural variety of members of theMytilus inhibitory peptide family was observed in each of the generaMytilus, Achatina andHelix. Gly-Trp-NH₂, the C-terminal dipeptide fragment of the neuropeptide AGPWamide, showed a more potent action than the parent peptide in all of the muscles examined. Peptides related to some molluscan neuropeptides were found to be distributed interphyletically. Some neuropeptides containing ad-amino acid residue were found inAchatina andMytilus. These aspects of molluscan neuropeptides are thought not to be exceptional.     

Torus shaped bands at the 2R telomere region and at the region 68 of the salivary gland chromosomes ofDrosophila auraria

Summary The telomere of the 2R arm of the salivary gland chromosomes ofD. auraria exhibits a definite toroidal structure in routine squashed preparations, stained either by propionic orcein-carmine or by fluorescent dyes. There is evidence that a band (or bands) of region 68 (possibly homologous to that ofD. melanogaster) of the 3L chromosome arm also exhibits a toroidal structure. These toroids are associated with heterochromatin, but it is not certain that they are themselves heterochromatic.Acknowledgments. This work was supported by a grant from Volkswagenwerk-Stiftung to C.D.K. The outstanding technical assistance of Ms G. Karamanlidis is acknowledged.     

On the specific attraction of the males of the six-toothed spruce bark beetle,Pityogenes chalcographus (L.) to a mixture of synthetic pheromones of the eight-toothed spruce bark beetle,Ips typographus (L.) (Coleoptera,Scolytidae)

Summary Traps containing a mixture of attractants forIps typographus also caught other species of bark beetles. The numbers ofPityogenes chalcographus, Pityogenes conjunctus, andTrypodendron lineatum were too high to be interpreted as accidental captures. The males of the polygamousP. chalcographus were specifically attracted, indicating that theI. typographus pheromone or one of its components acts as a kairomone and primary attractant for the pioneering males ofP. chalcographus.     

C-band differentiation between the chromosomes of two subspecies of the chironomid midgeChironomus thummi

Summary The metaphase chromosomes ofChironomus th. thummi contain approximately 17% more pericentric C-band heterochromatin than the chromosomes ofChironomus th. piger with 11% heterochromatin. InCh. th. thummi, the proportion of heterochromatin appeared to be much larger in metaphase chromosomes than in polytene chromosomes. This discrepancy is interpreted as being due to the specific chromosome organization and not as the result of an underreplication of heterochromatin during polytenization.     

An extremely cadmium-sensitive strain ofChlorella

Summary Growth of 14 strains from fiveChlorella species is rather insensitive towards cadmium. One strain (211-1a) ofC. saccharophila, however, was found to have a sensitivity towards this toxic heavy metal about 100 times higher than that of the other strains ofC. saccharophila.Acknowledgment. For excellent technical assistance I am indebted to Mrs U. Knoch and Mrs E. Weitemeyer.     

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.     

Photodecomposition of orellanine and orellinine,the fungal toxins ofCortinarius orellanus Fries andCortinarius speciossimus

Summary The photosensitivity of orellanine, the main toxin ofCortinarius orellanus Fries mushrooms, and its transformation to orelline via orellinine is discussed. All three substances were found in methanolic extracts ofCortinarius orellanus andCortinarius speciossimus mushrooms. The problem of homogeneity of orellanine is also discussed.Acknowledgment. The authors wish to express their gratitude to Prof. Dr W. Steglich from the Institut für Organische Chemie und Biochemie der Universität Bonn (Federal Republic of Germany) for having provided them with a sample of dried material ofC. speciossisimus.     

Alternariol,a dibenzopyrone mycotoxin ofAlternaria spp., is a new photosensitizing and DNA cross-linking agent

Summary The mycotoxin alternariol (3,4,5-trihydroxy-6-methyldibenzo [a] pyrone) but not alternariol monomethyl ether (3,4-dihydroxy-5-methoxy-6-methyldibenzo [a] pyrone) is phototoxic toEscherichia coli in the presence of near UV light (320–400 nm). The phototoxicity bioassays with a DNA repair-deficient mutant ofE. coli suggested that DNA may be the molecular target for photo-induced toxicity of alternariol. Interactions between alternariol and double-stranded, supercoiled DNA suggest that alternariol interacts with DNA by intercalation. No DNA breakage was detected in this system; however, alternariol forms a complex and cross-links double-stranded DNA in near UV light. These results suggest that alternariol is a new phototoxic, DNA-intercalating agent and is a DNA cross-linking mycotoxin in near UV light.Acknowledgment. Dr Albert Stoessl (Agriculture Canada, London, Ontario, Canada) generously provided a mixture of alternariol and alternariol monomethyl ether, and made many helpful suggestions. Dr Ashwood-Smith (University of Victoria, Victoria, British Columbia, Canada) kindly supplied the microorganisms through Dr G.H.N. Towers (University of British Columbia, Vancouver). We gratefully acknowledge the gifts. It is a pleasure to acknowledge the able assistance of Mr S. Tallevi.