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PubMed2002

DNA-based methods for the detection and the identification of phytoplasmas in insect vector extracts.

DNA extraction and storage methods have been evaluated with laboratory-reared leafhoppers and/or field-collected leafhoppers and psyllids. Detection of four different phytopathogenic phytoplasmas, belonging to three taxonomic groups, has been achieved by several direct or nested polymerase chain reaction (PCR) methods with such DNA extracts. Reactions differed in both the 16/23S ribosomal primer pairs used and the specific assay and cycling conditions. Merits and possible hindrances of the various primer pairs, in relation to insect DNA extracts, are discussed. However, identification of the phytoplasma(s) necessarily relied on comparison of the polymorphism in length of the amplified DNA fragments obtained by restriction with appropriate endonucleases. Endonuclease digestion is crucial for determining the identity (subgroup affiliation) of phytoplasmas of the same groups that can be carried by an individual vector.

Animals↗
PubMed1999

Neolignans inhibit Trypanosoma cruzi infection of its triatomine insect vector, Rhodnius prolixus.

Two neolignans, burchellin and nordihydroguaiaretic acid (NDGA), were toxic only to Trypanosoma cruzi clone Dm28c maintained in brain heart infusion (BHI) medium at a concentration of 100 microg/ml, not 10 microg/ml. When Rhodnius prolixus was fed with epimastigotes of T. cruzi and treated simultaneously with a single dose of burchellin or NDGA at 10 pg/ml of blood meal the number of parasites in the gut decreased. Whereas burchellin was only partially active, NDGA drastically reduced the number of epimastigotes and metacyclic trypomastigotes of T. cruzi in the excreta (urine plus feces). When the insect larvae were pretreated with burchellin or NDGA at 20 days before the infection with T. cruzi a significant reduction in the number of parasites in the gut occurred. However, when both compounds were applied at 20 days after the establishment of T. cruzi infection, although burchellin significantly reduced the gut infection, neither compound could abolish the infection entirely within the subsequent 15 days.

Animals↗
PubMedDate not supplied

Chagas' disease and its insect vector. Effect of azadirachtin A on the interaction of a triatomine host (Rhodnius prolixus) and its parasite (Trypanosoma cruzi).

The ED50 for moulting inhibition by injected azadirachtin A is for fourth instar larvae of all the triatomines, Triatoma vitticepes, T. pseudomaculata, T. maculata, T. brasiliensis, T. lecticularis, T. matogrossensis, T. infestans, Rhodnius prolixus. R. neglectus, R. robustus, Panstrongylus megistus, and P. herrera in the range of 10-25 ng/larva. In Rhodnius prolixus, the survival of T. cruzi was studied after treatment with the drug. If the trypomastigotes were fed in presence of 1.0 microgram azadirachtin A/ml blood, the number of parasites decreased near to the limit of detection within 30 days. If the drug was applied 20 days after T. cruzi infection, it still completely abolished the parasite in the host's gut within the subsequent 20 days. The same holds true if the insect larvae were pretreated with azadirachtin A 20 days before the subsequent infection with T. cruzi. Azadirachtin A. if applied at the dose of 1 microgram/ml, did not affect the hemolytic activity of the crop contents or the proteinase content of the intestine. A parallel between azadirachtin effects on the hormone balance of the host and growth inhibition of the parasite is discussed on the basis of the present results.

Animals↗
PubMed1981

Absence of detectable alteration in the kinetoplast DNA of a Trypanosoma brucei clone following loss of ability to infect the insect vector (Glossina morsitans).

A monomorphic bloodstream population of Trypanosoma brucei EATRO 1244 was derived from a cloned pleomorphic parental population by 77 rapid passages through mice. Loss of pleomorphism was accompanied by increased virulence of trypanosomes towards the mammal, by loss of ability to infect the tsetse fly, Glossina morsitans, loss of ability to transform to the procyclic stage in vitro at 26 degrees C, and by loss of oligomycin-sensitive ATPase activity in trypanosome homogenates. No differences in the maxicircle component of the kinetoplast DNAs (kDNA) of the two populations were detected by electron microscopy of kDNA network spreads or by electrophoretic analysis of restriction endonuclease digests. It appears, therefore, that loss of transmissibility and associated ability of the trypanosomes to activate the mitochondrion need not necessarily be the result of deletions in the mitochondrial (maxicircle) genome. We suggest that point mutations in critical mitochondrial genes, undetectable using out methods, or mutations of nuclear genes coding for important mitochondrial enzymes, may account for the observed changes in phenotype.

Animals↗
PubMed1996

P2 protein encoded by genome segment S2 of rice dwarf phytoreovirus is essential for virus infection.

Loss of infectivity to insect vector cell monolayers of rice dwarf phytoreovirus (RDV) after CCl4 treatment was associated with the removal of one of the viral proteins from the virus particles. This protein, encoded by genome segment S2 and thus named P2 protein, was located at the outer capsid of the virus particle. When RDV was treated with CCl4 for various times, the reduction in the amount of P2 protein and the loss of viral infectivity to vector cell monolayers were proportional to treatment time. RDV purified using CCl4 treatment thus lacking P2 protein, lost the ability to infect vector insects through feeding and consequently failed to be transmitted to plants. These results suggest that P2 protein is essential for RDV infection of the insect cells and may be related to transmission of the virus by the vector insect.

Amino Acid Sequence↗
PubMed1997

The minor outer capsid protein P2 of rice gall dwarf virus has a primary structure conserved with, yet is chemically dissimilar to, rice dwarf virus P2, a protein associated with virus infectivity.

The nucleotide sequence of the genome segment 2 (S2) of rice gall dwarf virus (RGDV), a phytoreovirus, when compared with the amino acid sequence of a component protein of the virus, showed that S2 potentially encoded a 127K minor outer capsid protein. This 127K protein designated as P2 and the 127K minor outer capsid protein (also termed P2) of rice dwarf virus (RDV) are similar in size, located in the outer capsid, and have well-conserved predicted polypeptide sequences, suggesting similar functions. Infectivity to insect vector cell monolayers of RGDV was maintained and the P2 protein was retained irrespective of carbon tetrachloride (CCl4) treatment. This is in contrast to the infectivity of RDV which is removed along with P2 protein following CCl4 treatment. RGDV with P2 was acquired by vector insects and transmitted to host plants, although RDV lacking P2 could not be transmitted to plants as previously published. These results imply that RDV and RGDV require P2 proteins for virus infectivity to vector insects.

Amino Acid Sequence↗
PubMed2003

Seroprevalence and risk factors for Trypanosoma cruzi infection in the Amazon region of Ecuador.

Trypanosoma cruzi infection in the Ecuadorian Amazon region has recently been reported. A seroepidemiologic survey conducted in four provinces in this region indicates a seroprevalence rate of 2.4% among the 6,866 samples collected in 162 communities. Among children < OR = 10 years of age, 1.2% were seropositive. Risk factors for T. cruzi seropositivity were having been born and remaining in the Ecuadorian Amazon provinces, age, living in a house with a thatch roof and open or mixed wall construction, recognizing the vector insects, and reporting being bitten by a triatomine bug. These data suggest active transmission of Chagas' disease in the Ecuadorian Amazon region is associated with poor housing conditions, and highlight the need for further studies aimed at understanding the biology of the insect vectors, reservoir species, and the clinical impact of T. cruzi infection as the basis for future educational and control programs in this region.

Adolescent↗
PubMed2004

Does Tetranychus urticae (Acari: Tetranychidae) use flying insects as vectors for phoretic dispersal?

Whether the spider mite Tetranychus urticae uses flying insects as vectors for phoretic dispersal was experimentally tested. Two bean plants were placed in a microcosm, and a mite population was introduced onto one of the plants. Either Phaenicia cuprina Wiedemann (Diptera: Calliphoridae) or Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) was then introduced into the microcosms as a hypothetical vector insect. T. urticae populations on the second bean plant were monitored to detect any evidence of phoretic dispersal. Instances of dispersal were detected at extremely low frequency, suggesting that phoretic dispersal of T. urticae mediated by winged insects is probably rare in the wild.

Animals↗
PubMed1998

Expression and purification of biologically active porcine follicle-stimulating hormone in insect cells bearing a baculovirus vector.

Biologically active recombinant porcine FSH (rec-pFSH) free from the cognate pituitary glycoprotein hormone LH was produced. It was synthesized by a baculovirus vector-insect cell system using two cDNAs encoding the glycoprotein alpha and FSH beta subunits. Its antigenicity was the same as that of pFSH prepared from the pituitary. Glycosylation of rec-pFSH was shown by tunicamycin treatment but the molecular mass of each subunit was lower than that of pituitary-derived FSH, because of the absence of trimming of terminal sugars in insect cells. Rec-pFSH was secreted into the culture medium at about 1 mg/l and purified in six fractions, because of the heterogeneity of the sugar group, by S-Sepharose and concanavalin A-Sepharose column chromatography. The biological activity of rec-pFSH was examined by measuring its effect on progesterone secretion from porcine granulosa cells and germinal vesicle breakdown (GVBD) of porcine oocytes. It showed adequate activity with respect to progesterone secretion, although some fractions rich in the sugar group showed lower activity than that of pituitary-derived FSH. It exhibited higher GVBD activity than that of pituitary-derived FSH at concentrations as low as 1 ng/ml. These results demonstrate that the baculovirus vector-insect cell system can provide biologically active rec-pFSH.

Animals↗