Geographic distribution and regional impacts of Oxyops vitiosa (Coleoptera: Curculionidae) and Boreioglycaspis melaleucae (Hemiptera: Psyllidae), biological control agents of the invasive tree Melaleuca quinquenervia.

The invasive tree Melaleuca quinquenervia (Cav.) Blake is widely distributed throughout peninsular Florida and poses a significant threat to species diversity in the wetland systems of the Everglades. Mitigation of this threat includes the areawide release campaign of the biological control agents Oxyops vitiosa Pascoe and Boreioglycaspis melaleucae Moore. We summarize the results of this release effort and quantify the resulting geographic distribution of the herbivores as well as their regional impact on the target weed. A combined total of 3.3 million individual Melaleuca biological control agents have been redistributed to 407 locations and among 15 Florida counties. Surveys of the invaded area indicate that the geographic distribution of O. vitiosa encompasses 71% of the Melaleuca infestation. Although released 5 yr later, the distribution of B. melaleuca is slightly greater than its predecessor, with a range including 78% of the sampled Melaleuca stands. Melaleuca stands outside both biological control agents' distributions occurred primarily in the northern extremes of the tree's range. Strong positive association between herbivore species was observed, with the same density of both species occurring in 162 stands and no evidence of interspecific competition. Soil type also influenced the incidence of biological control agents and the distribution of their impacts. The odds of encountering O. vitiosa or B. melaleucae in cells dominated by sandy soils were 2.2 and 2.9 times more likely than those predominated by organically rich soils. As a result, a greater level of damage from both herbivores was observed for stands growing on sandy versus organic-rich soils.

Selection on herbivory resistance and growth rate in an invasive plant.

The evolution of increased competitive ability (EICA) hypothesis proposes that invasive species evolve decreased defense and increased competitive ability following natural enemy release. Previous tests of EICA examined the result of evolution by comparing individuals from home and introduced ranges, but no previous study of this hypothesis has examined the process of evolution by analyzing patterns of selection. On the basis of EICA, there should be selection for competitive ability without herbivores and selection for defense with herbivores. Selection on competitive ability should be stronger for genotypes accustomed to herbivores (home range genotypes), and selection on defense should be stronger for genotypes unaccustomed to herbivores (introduced range genotypes). Using a field experiment, we tested these hypotheses for the invasive plant Melaleuca quinquenervia. There was a negative genetic correlation between resistance and growth, indicating a trade-off. However, selection for stem elongation (an indicator of competitive ability) was always positive, and selection on resistance was always negative and did not depend on genotype source or the presence of herbivores. The patterns of selection found in this study contrast with predictions from EICA and accurately predict the lack of evolutionary change in growth and resistance following the introduction of this species from Australia to Florida.

Relevance of 10-min delayed recall in dementia screening.

Within the context of early diagnosis of Alzheimer's disease (AD), there is a growing interest in neuropsychological screening tests. Amongst these tests, we focused on the largely used Memory Impairment Screen (MIS). The objective of the present work was to show that adding a 10-min delayed recall to the MIS, improves the test psychometric characteristics in order to detect dementia in the earliest stages. A prospective study was carried out on a cohort of 270 consecutive elderly ambulatory subjects attending the Broca Hospital Memory Clinic: normal controls (n = 67), mild cognitive impairment subjects (n = 98) and mildly demented patients [n = 105, Mini Mental State Examination (MMSE) = 23 +/- 4]. This study consisted in testing the advantage of the 10-min delayed recall entitled MIS-D compared with the MIS. At a cut-off score of 6, the MIS-D revealed satisfying psychometric characteristics with a sensitivity of 81% and a specificity of 91%, whilst the MIS alone indicated a sensitivity of 60% and a specificity of 88% in detecting dementia. In demented patients with MMSE score > or =26, MIS-D properties still remained satisfying (sensitivity: 75%, specificity: 92%). MIS-D is a more relevant screening test than MIS alone at very early stages of dementia.

Initial impacts and field validation of host range for Boreioglycaspis melaleucae Moore (Hemiptera: Psyllidae), a biological control agent of the invasive tree Melaleuca quinquenervia (Cav.) Blake (Myrtales: Myrtaceae: Leptospermoideae).

Invasion of south Florida wetlands by the Australian paperbark tree, Melaleuca quinquenervia (Cav.) S.T. Blake (melaleuca), has caused adverse economic and environmental impacts. The tree's biological attributes and favorable ambient biophysical conditions combine to complicate efforts to restore and maintain south Florida ecosystems. Management requires an integrated strategy that deploys multiple biological control agents to forestall reinvasion and to supplement other control methods, thereby lessening recruitment and regeneration after removal of existing trees. This biological control program began during 1997 when an Australian weevil, Oxyops vitiosa (Pascoe), was released. A second Australian insect, the melaleuca psyllid (Boreioglycaspis melaleucae Moore), first introduced during 2002, has also widely established. After inoculation of the psyllid in a field study, only 40% of seedlings survived herbivory treatments compared with 95% survival in controls. The resultant defoliation also reduced growth of the surviving seedlings. A weevil-induced decline at a site comprised mainly of coppicing stumps had slowed after a 70% reduction. Psyllids colonized the site, and 37% of the remaining coppices succumbed within 10 mo. The realized ecological host range of B. melaleucae was restricted to M. quinquenervia; 18 other nontarget plant species predicted to be suboptimal or nonhosts during laboratory host range testing were unaffected when interspersed with psyllid-infested melaleuca trees in a common garden study. Evaluations are ongoing, but B. melaleucae is clearly reducing seedling recruitment and stump regrowth without adversely impacting other plant species. Manifestation of impacts on mature trees will require more time, but initial indications suggest that the psyllid will be an effective supplement to the weevil.

125I derivatives of prostaglandins. A novel approach in prostaglandin analysis by radioimmunoassay.

We report the production of radioactive iodinated (125 I) derivatives of prostaglandins E1, E2, F2alpha and their use in radioimmunological assays. Histamine or tyramine was coupled to the prostaglandins carboxyl group and the iodination was accomplished using the chloramine T method. The high specific radioactivity of these tracers and the resolution of the purification procedure allowed the detection of 0.5 pg of prostaglandins. A comparison with tritiated prostaglandin was made and showed a 10-fold gain in sensitivity. Furthermore in the case of the prostaglandin E1 system using 125I-labelled histamine or tyramine as tracer the cross reaction curves obtained were different from those obtained with [3H]prostaglandin E1; we suggest that the blocking of the carboxyl group alters the prostaglandin E1 structure, modifying its immunoreactivity.

The use of iodinated tracers for a sensitive radioimmunoassay of 13,14-dihydro-15-ketoprostaglandin Falpha.

Radioimmunoassay of 13,14-dihydro-15-ketoprostaglandin Falpha is reported using various 125I-labelled derivatives. The apparent association constants of the antiserum for iodinated tracers are higher than with the homologous hapten. In spite of this, the high specific activities of iodinated tracers (2000 Ci/mmol) allow a 3-fold increase in the sensitivity of the assay when compared with the tritiated derivative. Human plasma levels of 13,14-dihydro-15-ketoprostaglandin Falpha reported (25+/-6 pg/ml) are lower than those previously found by radioimmunoassay, and no sexual difference was found.

Enzyme immunoassay of progesterone at the picogram level using beta-galactosidase as label.

We report here the first sensitive enzyme immunoassay of a hapten. A progesterone beta galactosidase conjugate was prepared using carbodiimide as a bifunctional reagent. Rabbit progesterone antisera were previously obtained. The separation of the bound from the free fraction of the label was performed with the help of polymerized anti rabbit gamma-globulins. The enzyme activity of the bound fraction was determined with O-nitrophenyl-beta-D-galactoside as substrate. Specificity and sensitivity (approximately 15 pg) of this enzyme immunoassay can be successfully compared with radioimmunoassay performances. It thus provides a non radioactive, inexpensive and reliable method of small molecule quantitation.

In vitro prostaglandin synthesis by various rat renal preparations.

Prostaglandin synthesis by eight different structures from the rat kidney (while cortex, cortical tubules, glomeruli, outer medulla, papilla, glomerular cultured epithelial and mesangial cells, cultured interstitial medullary cells) was measured in vitro after incubation with [14C] arachidonic acid using high-performance liquid chromatography followed by RIA with four specific anti-prostaglandin antibodies (prostaglandin E2, prostaglandin F2 alpha, 6 keto-prostaglandin F1 alpha, thromboxane B2). Prostaglandin production by the whole cortex and cortical tubules was very low. The order of abundance for isolated glomeruli was thromboxane B2 great than prostaglandin E2 greater than prostaglandin F2 alpha greater than 6 keto-prostaglandin F1 alpha. Mesangial cells synthesized prostaglandin E2 at a markedly high rate, in decreasing order: prostaglandin F2 alpha, thromboxane B2 and 6 keto-prostaglandin F1 alpha. The same order of abundance was observed for epithelial cells. The papilla synthesized essentially prostaglandin E2 and prostaglandin F2 alpha, whereas the main product for the outer medullar was 6 keto-prostaglandin F1 alpha. Cultured interstitial cells synthesized mainly prostaglandin E2 and to a lesser extent prostaglandin F2 alpha. Unidentified peaks eluting between 6 keto-prostaglandin F1 alpha and thromboxane B2 were also observed chiefly with glomeruli but they were absent with the medullary preparations. They disappeared after incubation with indomethacin or aspirin and represented for glomeruli the greatest percentage of conversion of [14C] arachidonic acid. These results show that the prostanoid profile varies markedly with the different regions and cells of the rat kidney.

Active metabolism of arachidonic acid by Kaposi sarcoma cells cultured from lung biopsies (KS-3); identification by HPLC and MS/MS of the predominant metabolite secreted as the 11,12-epoxy-eicosatrienoic acid.

The development of long-term culture of AIDS-KS cells has allowed us to investigate further a possible vascular origin of Kaposi sarcoma. Taking into account the relative specificity of arachidonic acid (AA) metabolism according to cell type, the AA 'cascade' was analyzed in cultured KS-3 cells established from lung biopsies and compared to human umbilical venous endothelial (H-UVE) cells and human myometrial smooth muscle (H-MSM) cells, under basal conditions and after stimulation with vasoactive agents such as histamine or thrombin. Considering strictly the 'prostaglandin' profile given by RIAs, the metabolism of AA was closer, whilst not identical, to H-UVE than to H-MSM cells. However, evaluation of all the eicosanoids released from [3H]AA labeled KS-3 cells revealed that the predominant metabolite was not prostacyclin (PGI2), as suggested from PG RIAs, but an epoxy-eicosatrienoic acid (EET), identified as the 11, 12 isomer by HPLC and MS/MS. The synthesis of this EET is probably cytochrome P-450 monooxygenase dependent. Its potential role in the development of the KS tumor cells is under investigation.

Characterization of monoclonal antibodies against prostaglandin E2: fine specificity and neutralization of biological effects.

The specificity and heterogeneity of the immune response of BALB/c mice immunized with prostaglandin E2 (PGE2) coupled to thyroglobulin was studied. All the animals (n = 50) responded to PGB2, a transformation product of PGE2. However, following repeated injections most of the animals (n = 30) were also able to respond to PGE2. Cellular hybridizations were performed and five anti-PGE2 monoclonal antibodies were isolated and analysed. They are mainly directed against the ring and the omega-chain of PGE2 but their specificity toward the alpha-chain is more limited. The association constants are greater than to 1 X 10(9) M-1. The monoclonal antibody 8E.57.71 (Ka = 1.3 X 10(10) M-1) is particularly convenient for sensitive radioimmunoassays (detection limit 25pg/ml, when iodinated tracer is used). Anti-PGE2 monoclonal antibodies were found to neutralize the specific binding of [3H]PGE2 to rat brain hypothalamic receptors and to inhibit the PGE2 induction of rat fundus muscular contraction.

The biological effects of injected antibodies to estradiol-17 beta and to progesterone in pregnant rats.

Estradiol-17 beta (E2) and progesterone (P) levels have been effectively reduced in pregnant rats by injecting them with antibodies which bind E2 or P (A-E2- or A-P) with high specificity. Before, and at daily intervals after treatment with A-E2 or A-P, blood was collected sequentially from the tail vein and the levels of circulating A-E2, A-P total E2 (E2t) and total P (Pt) determined. Having established the relationship between A-E2, E2t, bound E2 (E2b) and unbound E2 (E2u), as well as between A-P, Pt, bound P (Pb) and unbound P (Pu), the concentrations of E2u and Pu could be calculated reliably. Treatment of gestating rats with A-E2 and A-P lowered E2u and Pu levels significantly (P less than 0.001). In comparison with the 14 controls, the 17 A-E2 treated (and thus E2u-deficient) animals showed significant increases in placental weight (P less than 0.001). This effect of A-E2 was readily prevented by replacement therapy with diethylstilbestrol (DES), since A-E2 does not bind DES. In 21 pregnant rats, treatment at day 10 of gestation with only 2 ml A-P (total "Binding Capacity" = 6 mug P) critically lowered the Pu levels and provoked abortion. In contrast, the same treatment in 12 rats at day 6 of gestation (when the P levels are higher than at day 10) failed to provoke the appropriate degree of P-withdrawal (Pw) and abortion, illustrating the quantitative nature of the relationship between effective A-P treatment, Pw and the termination of pregnancy.

Inhibitory effect of platelet-activating factor (PAF) on luteinizing hormone-releasing hormone and somatostatin release from rat median eminence in vitro correlated with the characterization of specific PAF receptor sites in rat hypothalamus.

Platelet-activating factor (PAF) exhibits a wide range of biological activities, including the stimulation of secretory processes in various cell types. However, little is known regarding its possible influence on the release of brain neuropeptides. In the present study we have examined the effect of PAF on the release of three hypothalamic releasing hormones in adult male rats, and have characterized the presence of specific PAF binding sites in rat hypothalamic membranes. PAF decreased LHRH and somatostatin (SRIF) release from the median eminence with a maximal inhibition at 10(-14) M for both neuropeptides, whereas GRF release was not significantly altered. Moreover, PAF strongly counteracted the Ca2+ ionophore A 23187-stimulated release of LHRH and SRIF from median eminence and medial basal hypothalamus (greater than 50% inhibition). These results suggest an involvement of Ca2+ dependent events in PAF action. This inhibitory effect was specifically exerted at a hypothalamic site because PAF failed to depress LH and GH release from the anterior pituitary. A specific, reversible and saturable binding of [3H]PAF to membrane preparations of rat hypothalamus was demonstrated and two classes of binding sites were characterized. The affinity (KD) of each binding class was 2.14 +/- 0.32 nM and 61.63 +/- 16.4 nM, respectively, and the corresponding maximal number of each binding class was 25.41 +/- 3.2 fmol/mg protein and 146.2 +/- 47.5 fmol/mg protein. In the same conditions no specific binding was observed using rat pituitary membranes. The specificity of PAF analogs for these binding sites was well correlated to their relative effectiveness in altering LHRH and SRIF release (order of potency: L-652,731, kadsurenone greater than BN 52021 greater than Lyso-PAF). These data suggest that the binding sites identified in the hypothalamus have the characteristics expected of a specific PAF receptor and that PAF effect on neuropeptides release is a receptor-mediated process.

Epoxygenase products of arachidonic acid are endogenous constituents of the hypothalamus involved in D2 receptor-mediated, dopamine-induced release of somatostatin.

The epoxyeicosatrienoic acids (EETs) were discovered as products of a cyclooxygenase/lipoxygenase-independent, cytochrome P-450 catalyzed metabolism of arachidonic acid (AA) termed the "epoxygenase" pathway. The rat hypothalamus is able to synthesize EETs from exogenous AA, and 5,6-EET has been found to release the neuropeptide somatostatin (SRIF) from hypothalamic nerve terminals of the median eminence (ME). In the present study, hypothalami from male rats were examined for the presence of endogenous EETs, using chemical, chromatographic, and mass spectral analysis procedures. The samples were initially separated in a C18 Sepralyte column, fractionated on TLC plates, and purified by reverse phase HPLC. Thereafter, they were esterified (pentafluorobenzyl esters) and subjected to negative ion chemical ionization/gas chromatography (GC)/mass spectral (MS) analysis. The GC retention time and the MS fragmentation patterns revealed the presence of a mixture of 8,9-, 11,12- and 14,15-EETs; instability of 5,6-EET during the isolation protocol precluded its identification. Total hypothalamic EET concentration was estimated to be 120 ng/g wet tissue. The 8,9-regiosomer released SRIF from ME nerve terminals with an ED50 of 5 x 10(-12) M; Dopamine (DA) and the D2 receptor agonist PPHT, but not the D1 receptor agonist SKF-38393, induced SRIF release from the ME. This effect was blocked by clotrimazole and ketoconazole, two inhibitors of microsomal cytochrome P-450 function and AA epoxygenase in particular. In contrast, the inhibitors failed to affect the increase in SRIF release induced by 8,9-EET. These results indicate that: 1) in addition to cyclooxygenase and lipoxygenase products, epoxygenase metabolites of AA are endogenous compounds of the hypothalamus, and 2) EETs may mediate the increase in SRIF release from hypothalamic neurons induced by the interaction of DA with D2 receptors.

Distribution of enkephalin in human fetus and infant spinal cord: an immunofluorescence study.

The distribution of enkephalin-like immunoreactivity in the human fetus and infant spinal cord have been studied by indirect immunofluorescence. Enkephalin-like immunoreactive fibers were detectable in the lateral funiculus of fetal spinal cord as early as 10 weeks. At the other fetal ages examined, ranging from 12 to 28 weeks, and in infant, enkephalinlike immunoreactivity was found widely distributed throughout the whole spinal cord. In fetus spinal cord several enkephalin-like immunoreactive cells were sometimes seen scattered in the intermediate gray region. Most of the labeling was, however, represented by thin, varicose, immunofluorescent fibers mainly localized in the intermediate gray regions, in the ventral horn and in the superficial dorsal horn layers where they progressively increased in number. Further, the white matter exhibited enkephalin-like immunoreactive fibers particularly in the lateral funiculus where a dense punctiform immunofluorescence could be seen. On the whole, similar patterns were also visible in infant spinal cord. Thus, the superficial layers of the dorsal horn and the intermediolateral and reticular nuclei areas displayed dense plexuses of immunoreactive fibers. In contrast, the white matter showed only little labeling. In addition, no immunoreactivity was found in fetus and infant dorsal root ganglia. Our results emphasize the wide distribution of the enkephalin-like immunoreactivity in the fetus as in the infant spinal cord and further suggest its first appearance early in fetal life, possibly at the embryonic stage.

Triton X-100 eliminates plasma proteins interference in a radioimmunoassay for luteinizing hormone-releasing hormone (LHRH) and LHRH analogues.

A method is described for the radioimmunoassay of native LHRH and DTrp6-LHRH, an LHRH analogue which does not require extraction of plasma samples. Interference by binding proteins normally present in plasma is removed by addition of Triton X-100 to the binding buffer at a concentration of 1% for LHRH and 0.15% for the LHRH analogue. This approach permits a direct estimation of the peptide level in unextracted plasma with quantitative recoveries for concentrations ranging from 0.015 to 10 ng/ml. Although the antiserum titre is reduced, the affinity of the antibody does not change at the detergent concentrations used in this study. This procedure is recommended for peptide assays in which the non-specific effects of plasma prevent a direct assay.

ELISA for detection of human antibodies to Chlamydiae.

An enzyme-linked immunosorbent assay for the detection of human antibodies to Chlamydiae is described which exploits the cross-react properties between the genus-specific antigen of Chlamydiae and the ReLPS constituent of the outer membrane of a Salmonella minnesota mutant. Of 100 random sera tested by ELISA-ReLPS and immunofluorescence 78% showed an absolute correlation, 15% were positive in immunofluorescence and negative in ELISA and 7% were positive in ELISA and negative in immunofluorescence. Furthermore results obtained by the ELISA-ReLPS on 55 sera from patients with clinical evidence of Chlamydiae infection correlated well with the values obtained by an ELISA using Chlamydia-coated microtitration plates and by two immunofluorescence tests using Chlamydia trachomatis and Chlamydia psittaci as antigens. The method described here is sensitive, simple, reproducible and may be employed for epidemiological and pathogenetic studies of chlamydial infections.

Prostaglandins of the E series inhibit release of noradrenaline in rat hypothalamus by a mechanism unrelated to classical alpha 2 adrenergic presynaptic inhibition.

The effects of eight different prostanoid derivatives (PGs) on the in vitro release of noradrenaline (NA) from rat hypothalamic slices are reported. Prostaglandin E2 (10(-8)-10(-5) M), which does not interfere with the [3H]NA uptake mechanism, inhibited [3H]NA release induced by K+-evoked depolarization. The rank order of inhibition of release of NA for the PGs was: PGE2 greater than PGE1 greater than PGA2 greater than 16, 16-dimethyl-PGE2 greater than 11-epi-PGE2 greater than or equal to 8-iso-PGE2 greater than PGF2 alpha greater than PGD2. It has recently been shown that PGs of the E series specifically bind with a high affinity to membrane preparations of rat hypothalamus. A similar rank order was found for the activity of these PGs in displacing [3H]PGE2 from its binding sites, suggesting that the effect of PGEs on release of NA is mediated by an interaction with PGE2 receptors. Under the same experimental conditions, 10(-6) M clonidine (an alpha 2 adrenoceptor agonist) diminished, and 10(-6) M yohimbine (an alpha adrenoceptor antagonist) increased [3H]NA release, supporting the existence of alpha 2 auto-inhibition. Exposure to 10(-6) M of the alpha 1, alpha 2 adrenergic receptor antagonist phentolamine, a concentration which by itself had no effect on overflow of [3H]NA, blocked the inhibitory effect of clonidine, but failed to antagonize the inhibitory action of PGE2. Moreover, the action of clonidine and yohimbine remained unaffected when PG synthesis was blocked with indomethacin.(ABSTRACT TRUNCATED AT 250 WORDS)

Localization of substance P- and enkephalin-like immunoreactivity in relation to catecholamine-containing cell bodies in the cat dorsolateral pontine tegmentum: an immunofluorescence study.

The distribution of tyrosine hydroxylase-, substance P- and enkephalin-immunoreactive neurons in the cat dorsolateral pons was studied using the indirect immunofluorescence method of Coons. To allow for the visualization of substance P- and enkephalin-immunoreactive cell bodies, colchicine was injected either in the ventricular space or in the cerebral tissue. The distribution of the tyrosine hydroxylase-immunoreactive cell bodies corresponded with the well-known distribution of catecholamine cells in this area of the brain. The observation of adjacent sections treated separately with tyrosine hydroxylase- and enkephalin-antiserum revealed that most catecholaminergic cells contain enkephalin-immunoreactivity. In addition to this catecholamine-enkephalin cell population, a moderate number of substance P-immunoreactive cell bodies was found in dorsolateral pons. The peribrachial nuclei were found to be densely supplied with substance P- and enkephalin-immunoreactive fibers, whereas the medial subdivisions, which contain the majority of the catecholamine cells in the dorsolateral pons, display a moderate number of immunoreactive fibers. These results are suggestive of interactions between peptide-containing and catecholaminergic neurons and also between-peptide-containing and non-catecholamine-containing neurons in the cat dorsolateral pons.

Failure of anti-inflammatory steroids to inhibit prostaglandin production by rat polymorphonuclear leucocytes.

1 Like rabbit polymorphonuclear (PMN) leucocytes, rat peritoneal glycogen-induced PMN leucocytes produced much greater amounts of prostaglandin when incubated with killed bacteria than in the absence of phagocytosable material. 2 Rat PMN leucocytes mainly prostaglandin E2 (PGE2), in amounts up to 17 ng/10(6) cells in 90 min incubation, some 25 times the amount produced by resting cells. 3 Indomethacin and meclofenamic acid inhibited prostaglandin production by resting and phagocytosing cells, the IC50 being of the order of 10(-6) to 10(-7) M for both drugs. 4 Hydrocortisone and dexamethasone at concentrations up to 10(-4) M did not cause significant dose-related inhibition of prostaglandin production in this system. 5 It is suggested that the phagocytosing PMN leucocyte is insensitive to the action of anti-inflammatory steroids with respect to prostaglandin production.

Importance of perinatal testosterone in sexual differentiation in the male rat.

Testosterone secretion in the male rat was high during the late fetal and immediate postnatal periods. It then showed a rapid decrease 3h after birth and remained low until puberty. Male rats from mothers given daily injections of an antibody to testosterone during the week before delivery displayed an LH peak when they were adult, orchidectomized and implanted with oestradiol. However, the amplitude of the peak was far smaller than in female rats from the same mothers treated in the same manner. Thus, the critical period during which testosterone triggers hypothalamic sexual differentiation is very close to birth, possibly starting at the end of the fetal period.