Drop-off behaviour of Bt-resistant and Bt-susceptible Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) larvae on Bt-cotton and non-Bt cotton plants.

The highest natural mortality rate of larval Lepidoptera in field populations occurs in the first instar, but it is highly variable. The pattern and degree of survival is not easily predicted but depends on their ability to establish on host plants. Lepidopteran larval dispersal behaviour, known as 'drop-off', happens when the host is unsuitable for larvae to settle and begin feeding. Understanding drop-off behaviour of Helicoverpa armigera (Hübner) with and without physiological resistance to Bt toxins on Bt and non-Bt cotton plants is an important component for resistance management strategies for this insect. We examined the drop-off behaviour of H. armigera to determine: (1) whether they move the same way or differently in response to Bt and non-Bt, and (2) could H. armigera larvae detect Bt toxin levels in cotton plants or did they move independently of toxin levels? In this study, we assessed the drop-off behaviour of Bt-resistant and Bt-susceptible H. armigera neonates on artificial diets and cotton plants with and without Bt toxin during the first 12 h after hatching. Bt-resistant and Bt-susceptible H. armigera neonates behaved differently on Bt and non-Bt substrates. The percentages of Bt-resistant larvae that dropped off Bt and non-Bt cotton plants were not significantly different. In contrast, significantly more Bt-susceptible larvae dropped off Bt cotton than non-Bt cotton plants over time. Although Bt-susceptible larvae could not detect Bt toxin, they showed preference on non-Bt toxin substrates and were more likely to drop off substrates with Bt toxin.

Oviposition site selection and survival of susceptible and resistant larvae of Helicoverpa armigera (Lepidoptera: Noctuidae) on Bt and non-Bt cotton.

In Australia Bt cotton has been planted since 1996, and has greatly improved the control of its key target Helicoverpa armigera (Hübner). There is no strong evidence that genetically modified cotton has been selected for significant physiological resistance to Bt toxin in field populations. There are many possible explanations for the lack of apparent selection that range from high compliance with the resistance management strategy for this technology to a lack of behavioral preference in key traits such as oviposition that could favor survival. To date most experiments that test oviposition of H. armigera on Bt cotton vs. conventional cotton have been done with susceptible moths. We determine the oviposition preference of a field isolated Bt resistant line of H. armigera and a susceptible counterpart when given a choice of non-Bt cotton and Bt-cotton with the same genetic background, and test whether there is any relationship between oviposition site selection (different plant structures) and the survival of the first instar larvae. Within cotton plants, our experiments consistently showed that both resistant and susceptible moths did not choose plants or plant parts that were less toxic in terms of Bt toxin on which to lay eggs. There was one exception in that susceptible moths were more likely to lay eggs on squares of Bt cotton plants than squares of non-Bt cotton. As expected, the mortality of susceptible H. armigera neonates was significantly higher on structures of Bt cotton plants than on those structures of conventional cotton, and survival was greater on flowers than on other structures of Bt cotton. This confirms opportunities for selection for resistance, and demonstrates no advantage in this respect to carrying resistance genes that might overcome the Bt toxins.

Predictive value of anti-cell and anti-human immunodeficiency virus (HIV) humoral responses in HIV-1-exposed seronegative cohorts of European and Asian origin.

Unconventional immune responses have been demonstrated in individuals who, despite repeated exposure to human immunodeficiency virus (HIV) infection, remain seronegative. As environmental exposure to pathogens and genetic background may modulate immune responses differentially, one Italian and two Asian populations of HIV-1-exposed seronegative individuals were studied. In serum samples from each group, IgG to CCR5, IgG to CD4 and IgA to gp41 were measured, which were previously described as markers of unconventional immunity in HIV-exposed seronegative Caucasians. Given the importance of conformational epitopes in virus-cell interactions, IgG to CD4-gp120 complex was also measured. It was found that markers of HIV exposure were present in all populations studied. HIV-specific humoral responses (IgA to gp41 and IgG to CD4-gp120 complex) were extremely significant predictors of HIV exposure (P<0.0001 in both cases), whereas the predictive values of anti-cell antibodies (anti-CCR5 and anti-CD4) varied between populations. Evidence is provided for the correlation of these differences with route of exposure to HIV and level of natural antibodies to cross-reactive microbial antigens. In conclusion, exposed seronegative individuals of ethnically different origins display similar signs of HIV-dependent unconventional immunity. A specific relevance must be attributed to different innate and acquired factors.

The acceptability, efficacy and safety of quinacrine non-surgical sterilization (QS), tubectomy and vasectomy in 5 provinces in the Red River Delta, Vietnam: a follow-up of 15,190 cases.

OBJECTIVES: To compare the safety, efficacy and acceptability of quinacrine sterilization (QS), tubectomy and vasectomy in Vietnam. METHODS: This study was initiated in January 1998 and completed in February 2000. A sample of 9 districts in 5 provinces, where the prevalence of QS was known to be high, was selected. Every person sterilized in these 9 districts between January 1, 1988 and March 31, 1998 was identified and systematically interviewed by family planning clinicians who had received special training for this project. RESULTS: A total of 15,982 sterilization users were identified and 15,190 were interviewed and examined, including a gynecologic exam, if needed: a follow-up rate of 95%. Of those interviewed, 9,753 used tubectomy, 3,734 used QS and 1,703 used vasectomy. All three methods were found to be safe, although morbidity associated with tubectomy was more serious than with QS or vasectomy. No deaths were reported. After more than 5 years of follow-up, tubectomy had the lowest failure rate: 1.0%, followed by 4.1% with vasectomy. A pregnancy rate of 13.2% was reported with quinacrine, although only a small fraction of these failures were confirmed. A strong preference for QS was found. CONCLUSION: QS has an important role to play in sterilization services in Vietnam.

The rate of ectopic pregnancy for 24,589 quinacrine sterilization (QS) users compared to users of other methods and no method in 4 provinces in Vietnam, 1994-1996.

OBJECTIVE: To determine the rates of ectopic pregnancy with the use of quinacrine sterilization (QS) compared to other methods and no method (non-users). METHODS: Four provinces were selected for their above average numbers of women who had undergone QS: Nam Dinh, Nam Ha, Hai Duong and Hung Yen. Case histories related to surgical treatment of all ectopic pregnancies in these 4 provinces from 1994 through 1996 were collected from all hospitals by researchers from the Ministry of Health in June 1997. Using a questionnaire designed for this study, 120 physicians interviewed every woman in her home who had had an ectopic pregnancy during this period. If deceased, a family member was consulted. All interviews were completed in September 1998. The numbers of users of each method and nonusers were calculated from service statistics and demographic data. RESULTS: Based on 2,551,355 woman-years of exposure, the rate of ectopic pregnancy among users per 1000 woman-years was calculated to be: 0.26 with QS; 0.42 with surgical sterilization (TL) and IUD; 0.45 with the Pill; 0.50 with condoms; 0.78 relying on withdrawal; and 1.18 among non-users. CONCLUSION: Ectopic pregnancy rates for QS, TL, IUD and the Pill were similar and much lower than the rate for non-users of contraception.

The acceptability, efficacy and safety of quinacrine non-surgical sterilization (QS), tubectomy and vasectomy in 5 provinces in the Red River Delta, Vietnam: a follow-up of 15,190 cases.

OBJECTIVES: To compare the safety, efficacy and acceptability of quinacrine sterilization (QS), tubectomy and vasectomy in Vietnam. METHODS: This study was initiated in January 1998 and completed in February 2000. A sample of 9 districts in 5 provinces, where the prevalence of QS was known to be high, was selected. Every person sterilized in these 9 districts between January 1, 1988 and March 31, 1998 was identified and systematically interviewed by family planning clinicians who had received special training for this project. RESULTS: A total of 15,982 sterilization users were identified and 15,190 were interviewed and examined, including a gynecologic exam, if needed: a follow-up rate of 95%. Of those interviewed, 9,753 used tubectomy, 3,734 used QS and 1,703 used vasectomy. All three methods were found to be safe, although morbidity associated with tubectomy was more serious than with QS or vasectomy. No deaths were reported. After more than 5 years of follow-up, tubectomy had the lowest failure rate: 1.0%, followed by 4.1% with vasectomy. A pregnancy rate of 13.2% was reported with quinacrine, although only a small fraction of these failures were confirmed. A strong preference for QS was found. CONCLUSION: QS has an important role to play in sterilization services in Vietnam.

The epidemiology of nematode and fluke infections in cattle in the Red River Delta in Vietnam.

Over a period of 13 months, faecal samples were collected monthly from approximately 45 cattle over 3 months of age. Additionally, 74 calves of 1-2 months were sampled to determine the presence of Toxocara vitulorum eggs. Individual egg counts and infective strongyle larvae from pooled faecal samples were examined. Post-mortem worm counts were carried out on six groups of tracer calves (n=12) that had been kept for 4 weeks on pasture in and around the village studied. The following helminths were identified: T. vitulorum, Cooperia punctata, C. pectinata, C. oncophora, Oesophagostomum radiatum, Trichostrongylus axei, T. colubriformis, Haemonchus spp., Fasciola spp. and Paramphistomum spp. In 8% of the samples collected from young calves, individual egg counts for T. vitulorum were found indicative for pathogenic worm burdens. Strongyle egg counts and worm counts indicated that transmission is low without a distinct seasonality. In animals of 3-9 months old, a strongyle egg count peak can be demonstrated which at a higher age steadily and significantly decreased. In faecal cultures Cooperia spp. were most prominent in all age groups throughout the year with the exception of the period September-November when Haemonchus spp. and Oesophagostomum spp. were most prevalent. Fasciola spp. eggs were found in 22% of the collected faecal samples and the egg counts were low indicating that the intensity of Fasciola spp. infection is mild. Based on the present data, regular anthelmintic treatments seem not to be justified, except for a single treatment at the age of 2 weeks against toxocariosis.

A method for demonstrating gene essentiality in Staphylococcus aureus.

A method for demonstrating whether a gene of Staphylococcus aureus is essential for growth in a rich medium is described. We have used this method to determine whether the murE gene, which encodes the UDP-N-acetylmuramyl tripeptide synthetase required for peptidoglycan synthesis, is essential for growth in S. aureus. In this study, strain CYL368 was constructed from S. aureus RN4220 by placing the murE gene in the chromosome under the control of the spac promoter (a hybrid promoter of the Escherichia coli lac operator and the Bacillus subtilis SPO1 phage promoter). To regulate the murE gene in CYL368, the E. coli lacI gene was expressed from the B. licheniformis penicillinase gene (pcn) promoter in plasmid pMJ8426. Strain CYL368(pMJ8426) grew normally in the presence of isopropyl-beta-d-thiogalactopyranoside but could not grow in the absence of the inducer. These results indicate that the murE gene expressed from the spac promoter in CYL368(pMJ8426) is needed for bacterial growth. We concluded that murE is an essential gene of S. aureus.

The sensitivity and specificity of two methods for detecting Fasciola infections in cattle.

Counts of Fasciola spp. eggs in faeces and measurements of antibody concentration to the excretory/secretory antigens of Fasciola spp. by ELISA were related to the numbers of flukes in the livers of 92 cattle killed in the abattoirs of Hanoi City, Vietnam. In this population, about 22% of the cattle had no flukes, another 22% had between 1 and 10 flukes, 44% between 11 and 100 flukes and 12% had more than 100 flukes in their livers. Of the 14 animals less than 2 years of age, only three were infected. At 2 years of age the mean number of flukes per liver was 10 whereas at 3 years and older, the mean varied between 60 and 80 flukes. Prevalence of infection was 78.3%. No eggs of Fasciola spp. were detected in the faeces of one third of infected cattle and 60% of the counts were less than 100 eggs per gram. The sensitivity of the egg counting method was 66.7% and specificity 100%, overall accuracy was 73.9%. Corresponding values for the ELISA method were 86.1, 70 and 82.6%, respectively. The positive and negative predictive values for the egg counting method were 100 and 45.5% and for the ELISA method were 91.2 and 58.3%, respectively.

Characterization of selachian egg case collagen.

The egg case of the dogfish Scyliorhinus canicula is a remarkable collagenous structure that combines mechanical strength and toughness with high permeability to small molecules and ions. The collagenous lamellae that form over 80% of the thickness of the case wall are secreted by the D-zone of the nidamental (oviducal gland). An acid-soluble collagen extracted from this zone and partially purified ran as a single band on a native gel at pH 4.3. A single band of identical mobility was extracted from egg cases removed from the oviducal gland. SDS-PAGE of both extracts revealed a major component with an apparent molecular weight of 35 kDa and a minor component at 34 kDa. Neither of these components appeared to be glycosylated. Amino acid analysis of the partially purified collagen extracted from the oviducal gland revealed a composition similar to that of the collagenous lamellae of the egg case with glycine accounting for 16% and imino acids for 10% of the total residues. Partial N-terminal and internal sequences were obtained by Edman degradation for peptides extracted from the D-zone of the nidamental gland. Four of the internal sequence fragments showed repeated G-X-Y triplets showing them to be collagenous. These four fragments were novel but showed similarity to the triple-helical domains of mammalian type IV, X, and VI collagens. The noncollagenous N-terminal and pepsin-resistant sequences were unique, showing no significant similarity to known proteins in the database. Several possible N-myristoylation and phosphorylation sites were identified in the noncollagenous sequences.

Modulation of angiotensin II binding affinity by allosteric interaction of polyvinyl sulfate with an intracellular domain of the DuP-753-sensitive angiotensin II receptor of bovine adrenal glomerulosa.

Angiotensin II (AII) is an important regulator of aldosterone secretion by adrenal glomerulosa cells. All interacts with a specific receptor coupled to a guanine nucleotide-binding protein that controls the activity of phospholipase C. Recently, novel All nonpeptide antagonists (DuP-753 and PD-123319) have been shown to discriminate between two subclasses of All receptors in many different tissues. Our studies confirmed that 125I-All specifically labeled two classes of binding sites for All in a membrane preparation of bovine adrenal glomerulosa cells. The first class (DuP-753 sensitive) represented approximately 85% of the total binding sites for All and possessed a high affinity (IC50 of 92.9 +/- 19.5 nM) for DuP-753. PD-123319 did not have any effect on 125I-All binding to this site. The second class of binding sites was more sensitive to PD-123319, with an IC50 of 6.9 +/- 3.7 nM, and had a much lower affinity for DuP-753 (IC50 around 10 microM). The two classes of receptors had different affinities for All. All showed an affinity around 2 nM for All type 1 receptor (AT1)(DuP-753 sensitive) and a higher affinity, around 0.3 nM, for All type 2 receptor (AT2) (PD-123319 sensitive). All-induced steroidogenesis was completely abolished in the presence of 3 microM DuP-753, indicating that this activity was mediated through a DuP-753-sensitive receptor. We also found that polyvinyl sulfate (PVS), a polyanion, could partly inhibit the binding of 125I-All to bovine adrenal glomerulosa cell membranes, with half-maximal efficiency at 17.3 +/- 8.2 nM. The inhibitory effect of PVS was selective for AT1. The inhibitory effect of PVS was due to a change in the affinity state of the receptor. Unexpectedly, PVS had no effect on All-induced steroidogenesis or on All binding to intact bovine adrenal glomerulosa cells. However, the inhibitory effect of PVS on All binding was recovered after permeabilization of cells. Direct interaction of polyanions with AT1 was suggested by the capacity of solubilized photoaffinity-labeled 125I-AT1 to adsorb to heparin-agarose gels. The adsorption of 125I-AT1 to heparin-agarose was inhibited by prior incubation of solubilized receptor with heparin or PVS. These results suggest that All-induced steroidogenesis is mediated by a DuP-753-sensitive receptor and that PVS decreases the affinity of this receptor by interacting with an intracellular domain (possibly the positively charged domain responsible for coupling with guanine nucleotide-binding proteins).

Study on the stereoselectivity of inositol 1,4,5-trisphosphate recognition sites of bovine adrenal cortex.

Inositol 1,4,5-trisphosphate (InsP3) is an intracellular messenger generated from the hydrolysis of phosphatidylinositol 4,5-bisphosphate by phospholipase C in response to Ca(2+)-mobilizing stimuli. InsP3 interacts with a specific receptor responsible for the release of sequestered Ca2+ from an intracellular store. The purpose of the present study was to evaluate the relative affinities of the naturally occurring D-isomer of InsP3 and that of its L-stereoisomer for the InsP3 receptor and the InsP3 metabolizing enzymes from bovine adrenal cortex. The InsP3 receptor recognized D- and L-isomers with respective affinities of 4.8 nM and 7.3 microM. This high degree of selectivity was also reflected in the capacity of both isomers to mobilize Ca2+ from the microsomal preparation. The partially purified InsP3 kinase preparation was also able to discriminate between the two stereoisomers. The activity of the kinase was half-maximally inhibited in the presence of 11 microM L-InsP3, a value much higher than the Km of the kinase for D-InsP3 (0.4 microM). Both stereoisomers exhibited equipotent affinities (around 17 microM) for the particulate preparation of InsP3 phosphatase. The enzyme, however, appeared to hydrolyze L-InsP3 at a much slower rate. These results demonstrated that the different recognition sites for InsP3 were expressing distinct levels of stereoselectivity. This property, which is an important aspect of ligand-receptor interaction, could be exploited for the design of new selective drugs interfering with InsP3 action and metabolism.

Interaction of polyanions with the recognition sites for inositol 1,4,5-trisphosphate in the bovine adrenal cortex.

Inositol 1,4,5-trisphosphate (InsP3) serves as a second messenger for Ca2+ mobilization in a wide variety of cells. InsP3 activates a specific receptor/channel located on an internal Ca2+ store. Because heparin has already been shown to block the action of InsP3, we have looked at the influence of other polyanions (dextran sulfate and polyvinyl sulfate) on the action and metabolism of InsP3 in the bovine adrenal cortex. Polyvinyl sulfate blocked InsP3 binding to adrenal cortex microsomes with a half-maximal efficiency of 250 nM. Scatchard analyses revealed that this effect was not competitive. The Ca2+ releasing activity of InsP3 on the same microsomal preparation was monitored with the fluorescent indicator, fura-2. Polyvinyl sulfate blocked this activity with a half-maximal efficiency of 80 nM. The effect of polyvinyl sulfate could not be overcome by supramaximal doses of InsP3, suggesting a non-competitive inhibitory effect. The activity of InsP3 phosphatase from bovine adrenal cortex microsomes was also studied. Polyvinyl sulfate inhibited the activity of the phosphatase with a half-maximal efficiency of 5 microM. Lineweaver-Burk plots revealed that this effect was not competitive. Polyvinyl sulfate was able to inhibit the activity of InsP3 kinase from bovine adrenal cortex cytosol. The half-maximal dose was 15 nM and the Lineweaver-Burk analysis showed that the inhibition was not competitive. The effect of dextran sulfate 5000 (DS-5000) on these activities was also studied. DS-5000 inhibited in a competitive manner the binding of InsP3 to its receptor (IC50 of 34 microM), the release of Ca2+ induced by InsP3 (IC50 of 6.5 microM) and the activity of InsP3 phosphatase (IC50 of 57 microM).(ABSTRACT TRUNCATED AT 250 WORDS)