Screening and exclusion of Zika virus infection in travellers by an NS1-based ELISA and qRT-PCR.

OBJECTIVES: Zika virus (ZIKV) infection during pregnancy may cause neurological abnormalities in the foetus, and therefore fast and accurate laboratory assays are critical for rapid diagnosis. ELISA based on ZIKV NS1 protein has been developed and shown to be sensitive and highly specific; however, its negative and positive predictive values have not been tested. In this study we evaluated the ability of the NS1-based ELISA to exclude ZIKV infection and serve as a first-line screening tool for travellers. METHODS: We tested samples obtained during the peak of ZIKV infection from 1188 symptomatic and asymptomatic Israeli travellers using NS1-based IgG and IgM ELISA, real-time RT-PCR analysis and ZIKV neutralization. The Kaplan-Maier method was used to evaluate the duration of ZIKV RNA in whole blood and urine samples. RESULTS: NS1-based ELISA identified 20 true-positive, five false-positive and four false-negative cases, resulting in sensitivity and specificity of 83.3% (95%CI: 62-94%) and 97.5% (95%CI: 94-99%) respectively, and positive and negative predictive values of 80% (95%CI: 59-92%) and 98% (95%CI: 95-99%) respectively. Based on 14 RT-PCR-positive cases, median time to detect ZIKV RNA in whole blood was 17.5 days (range 5-58 days) and in urine 10 days (range 5-26 days). CONCLUSIONS: The NS1-based ELISA and RT-PCR in whole blood are highly reliable for identification of ZIKV-negative and -positive cases, respectively. Combination of both assays minimizes the risk of false-negative results, and thus allows the exclusion of ZIKV infection in travellers returning from ZIKV-endemic countries, including those who are pregnant or wish for preconception screening.

What mediates tree mortality during drought in the southern Sierra Nevada?

Severe drought has the potential to cause selective mortality within a forest, thereby inducing shifts in forest species composition. The southern Sierra Nevada foothills and mountains of California have experienced extensive forest dieback due to drought stress and insect outbreak. We used high-fidelity imaging spectroscopy (HiFIS) and light detection and ranging (LiDAR) from the Carnegie Airborne Observatory (CAO) to estimate the effect of forest dieback on species composition in response to drought stress in Sequoia National Park. Our aims were (1) to quantify site-specific conditions that mediate tree mortality along an elevation gradient in the southern Sierra Nevada Mountains, (2) to assess where mortality events have a greater probability of occurring, and (3) to estimate which tree species have a greater likelihood of mortality along the elevation gradient. A series of statistical models were generated to classify species composition and identify tree mortality, and the influences of different environmental factors were spatially quantified and analyzed to assess where mortality events have a greater likelihood of occurring. A higher probability of mortality was observed in the lower portion of the elevation gradient, on southwest- and west-facing slopes, in areas with shallow soils, on shallower slopes, and at greater distances from water. All of these factors are related to site water balance throughout the landscape. Our results also suggest that mortality is species-specific along the elevation gradient, mainly affecting Pinus ponderosa and Pinus lambertiana at lower elevations. Selective mortality within the forest may drive long-term shifts in community composition along the elevation gradient.

West Nile virus outbreak in Israel in 2015: phylogenetic and geographic characterization in humans and mosquitoes.

OBJECTIVES: West Nile Virus (WNV) is endemic in Israel and was responsible for several outbreaks in the past 16 years. The aim of the present study was to investigate the spatial distribution of WNV acute infections from an outbreak that occurred in 2015 in Israel and report the molecular and geographic characterization of WNV isolates from human cases and mosquito pools obtained during this outbreak. METHODS: Using a geographical layer comprising 51 continuous areas of Israel, the number of WNV infection cases per 100 000 people in each area and the locations of WNV-infected mosquitoes in 2015 were analysed. Sequencing and phylogenetic analyses followed by geographic localization were performed on 13 WNV human isolates and 19 WNV-infected mosquito pools. RESULTS: Substantial geographical variation in the prevalence of acute WNV in patients in Israel was found and an overall correlation with WNV-infected mosquitoes. All human patients sequenced were infected only with the Mediterranean subtype of WNV Lineage 1 and resided primarily in the coastal regions in central Israel. In contrast, mosquitoes were infected with both the Mediterranean and Eastern European subtypes of WNV lineage 1; however, only the Mediterranean subtype was found in mosquitoes from the coastal region in central Israel. CONCLUSION: These results demonstrate differential geographic dispersion in Israel of the two WNV subtypes and may also point to a differential pattern of human infections. As a geographical bridge between Europe, Asia and Africa, analysis of WNV circulation in humans and mosquitoes in Israel provides information relevant to WNV infections in Eurasia.

Correction: Managing Climate Change Refugia for Climate Adaptation.

[This corrects the article DOI: 10.1371/journal.pone.0159909.].

Managing Climate Change Refugia for Climate Adaptation.

Refugia have long been studied from paleontological and biogeographical perspectives to understand how populations persisted during past periods of unfavorable climate. Recently, researchers have applied the idea to contemporary landscapes to identify climate change refugia, here defined as areas relatively buffered from contemporary climate change over time that enable persistence of valued physical, ecological, and socio-cultural resources. We differentiate historical and contemporary views, and characterize physical and ecological processes that create and maintain climate change refugia. We then delineate how refugia can fit into existing decision support frameworks for climate adaptation and describe seven steps for managing them. Finally, we identify challenges and opportunities for operationalizing the concept of climate change refugia. Managing climate change refugia can be an important option for conservation in the face of ongoing climate change.

Osteocyte control of bone remodeling: is sclerostin a key molecular coordinator of the balanced bone resorption-formation cycles?

Osteocytes, entrapped within a newly mineralized bone matrix, possess a unique cellular identity due to a specialized morphology and a molecular signature. These features endow them to serve as a bone response mechanism for mechanical stress in their microenvironment. Sclerostin, a primarily osteocyte product, is widely considered as a mechanotranduction key molecule whose expression is suppressed by mechanical loading, or it is induced by unloading. This review presents a model suggesting that sclerostin is major mediator for integrating mechanical, local, and hormonal signals, sensed by the osteocytes, in controlling the remodeling apparatus. This central role is achieved through interplay between two opposing mechanisms: (1) unloading-induced high sclerostin levels, which antagonize Wnt-canonical-ß-catenin signaling in osteocytes and osteoblasts, permitting simultaneously Wnt-noncanonical and/or other pathways in osteocytes and osteoclasts, directed at bone resorption; (2) mechanical loading results in low sclerostin levels, activation of Wnt-canonical signaling, and bone formation. Therefore, adaptive bone remodeling occurring at a distinct bone compartment is orchestrated by altered sclerostin levels, which regulate the expression of the other osteocyte-specific proteins, such as RANKL, OPG, and proteins encoded by "mineralization-related genes" (DMP1, PHEX, and probably FGF23). For example, under specific terms, sclerostin regulates differential RANKL and OPG production, and creates a dynamic RANKL/OPG ratio, leading either to bone formation or resorption. It also controls the expression of PHEX, DMP1, and most likely FGF23, leading to either bone matrix mineralization or its inhibition. Such opposing up- or down-regulation of remodeling phases allows osteocytes to function as an "external unit", ensuring transition from bone resorption to bone formation.Mini Abstract: The osteocyte network plays a central role in directing bone response either to mechanical loading, or to unloading, leading correspondingly to bone formation or resorption. This review shows a key role of the osteocyte-produced sclerostin as a major mediator of the molecular mechanisms involved in the process of adaptive bone remodeling.

Is interaction between age-dependent decline in mechanical stimulation and osteocyte-estrogen receptor levels the culprit for postmenopausal-impaired bone formation?

Declining estrogen levels during menopause are widely considered to be a major cause of age-dependent bone loss, which is primarily manifested by increased bone resorption by osteoclasts. We present accumulating evidence supporting another aspect of metabolic bone loss, suggesting that the combined interaction between age-dependent factors, namely, estrogen deficiency and reduced day-by-day activity/mechanical stimulation, directly leads to a reduction in anabolic processes. Such decreased bone formation results in diminished bone strength and failure to maintain the load-bearing competence of a healthy skeleton and to postmenopausal osteoporosis disorder. Estrogen receptors (ERs), as mediators of estrogenic actions, are essential components of bone osteocyte and osteoblast mechano-adaptive responses. ER expression appears to be upregulated by adequate circulating estrogen levels. ERα signaling pathways participate in the mechanotransduction response through obligatory "non-genomic" actions that occur independently of estrogen binding to ER and by a potentially "genomic", estrogen-dependent mode. The experimental data indicate that cross talk between the ERα-"non-genomic" and Wnt/ß-catenin signaling pathways constitutes the major regulatory mechanism. This interaction uses mechanically and ER-induced prostaglandin E2 as a mediator for the downregulation of osteocyte production of sclerostin. Sclerostin suppression, in turn, is a central prerequisite for load-induced formation and mineralization of the bone matrix. It is therefore plausible that future strategies for preventing and treating postmenopausal osteoporosis may use estrogenic compounds (such as selective estrogen receptor modulators or phytoestrogens) with physical activity, to complement antiresorptive therapy, aimed at stopping further bone loss and possibly even reversing it by stimulation of bone gain.

Vitamin D inhibits captopril-induced cell detachment and apoptosis in keratinocytes.

BACKGROUND: Captopril, an angiotensin I-converting enzyme inhibitor, is a commonly prescribed antihypertensive drug. Its cutaneous side-effects include pemphigus vulgaris acantholysis and bullous pemphigoid-like cell-matrix detachment. This medication also triggers apoptosis in human keratinocytes. Calcitriol, the hormonally active vitamin D metabolite, protects keratinocytes from programmed cell death induced by various noxious stimuli. OBJECTIVES: To examine if calcitriol protects proliferating keratinocytes from the damage inflicted by captopril. METHODS: Autonomously proliferating HaCaT keratinocytes, used as a model for basal layer keratinocytes, were exposed to captopril. Cell detachment was examined visually by light microscopy. Cytotoxicity was assessed by Hoechst 33342 staining and lactate dehydrogenase release. Apoptotic death was assessed by monitoring caspase 3-like activity. RESULTS: Cells exposed to captopril detached and became round. This process was accompanied by programmed cell death. From time-dependent monitoring of cell detachment and apoptosis, and examination of pan-caspase inhibitor effects on cell detachment we concluded that cell death is the consequence of cell detachment from the culture plate and not vice versa. Pretreatment with calcitriol significantly attenuated these events. The effects of calcitriol were already evident at 1 nmol L(-1) concentration of the hormone. CONCLUSIONS: The results of this study show that calcitriol protects keratinocytes from captopril-induced cell detachment and apoptosis.

Upregulation of MMP-9 production by TNFalpha in keratinocytes and its attenuation by vitamin D.

MMP-9, a member of the matrix metalloproteinase family that degrades collagen IV and processes chemokines and cytokines, participates in epidermal remodeling in response to stress and injury. Limited activity of MMP-9 is essential while excessive activity is deleterious to the healing process. Tumor necrosis factor (TNFalpha), a key mediator of cutaneous inflammation, is a powerful inducer of MMP-9. Calcitriol, the hormonally active vitamin D metabolite, and its analogs are known to attenuate epidermal inflammation. We aimed to examine the modulation of MMP-9 by calcitriol in TNFalpha-treated keratinocytes. The immortalized HaCaT keratinocytes were treated with TNFalpha in the absence of exogenous growth factors or active ingredients. MMP-9 production was quantified by gelatin zymography and real-time RT-PCR. Activation of signaling cascades was assessed by western blot analysis and DNA-binding activity of transcription factors was determined by EMSA. Exposure to TNFalpha markedly increased the protein and mRNA levels of MMP-9, while pretreatment with calcitriol dose dependently reduced this effect. Employing specific inhibitors we established that the induction of MMP-9 by TNFalpha was dependent on the activity of the epidermal growth factor receptor, c-Jun-N-terminal kinase (JNK), NFkappaB and extracellular signal-regulated kinase-1/2. The effect of calcitriol was associated with inhibition of JNK activation and reduction of DNA-binding activities of the transcription factors activator protein-1 (AP-1) and NFkappaB following treatment with TNFalpha. By down-regulating MMP-9 levels active vitamin D derivatives may attenuate deleterious effects due to excessive TNFalpha-induced proteolytic activity associated with cutaneous inflammation.

Enamel matrix derivative induces the expression of tissue inhibitor of matrix metalloproteinase-3 in human gingival fibroblasts via extracellular signal-regulated kinase.

BACKGROUND AND OBJECTIVE: Periodontal disease is characterized by increased expression and activity of matrix metalloproteinases (MMPs) and insufficient expression/activity of their inhibitors, tissue inhibitors of matrix metalloproteinases (TIMPs). This altered MMP-TIMP balance results in progressive destruction of gingival and periodontal extracellular matrix. Enamel matrix derivative (EMD), clinically used for periodontal regeneration in a device called Emdogain, has been suggested to enhance gingival healing following periodontal procedures in humans. We previously showed that EMD increases the proliferation of human and rat gingival fibroblasts and protects them from tumor necrosis factor-induced apoptosis. In the present study, the modulation of MMP and TIMP expression by EMD was investigated. MATERIAL AND METHODS: Primary human gingival fibroblasts were treated in vitro with tumor necrosis factor, EMD or both in serum-free conditions, and RNA was analyzed with an extracellular matrix-focused microarray and quantitative real-time polymerase chain reaction. RESULTS: Microarray analysis showed detectable expression of MMP-1, MMP-2, MMP-3, MMP-7 and MMP-13, as well as TIMP-1 and TIMP-3 in untreated cells. There was no apparent regulation of the expression of MMP-2, MMP-7, MMP-13 and TIMP-1 by either tumor necrosis factor or EMD. In contrast, tumor necrosis factor significantly increased MMP-1 expression, and EMD reduced it when both agents were present. Also, EMD significantly induced TIMP-3 expression, an effect which was dependent on activation of extracellular signal-regulated kinase 1/2, since it was totally abolished by a selective extracellular signal-regulated kinase pathway inhibitor. CONCLUSION: These data suggest that EMD may affect gingival health by ways other than cell proliferation/survival, i.e. by stimulation of TIMP-3 production, which could improve the MMP-TIMP balance in gingival tissue and curb extracellular matrix destruction.

Nutrient availability and phytoplankton nutrient limitation across a gradient of atmospheric nitrogen deposition.

Atmospheric nitrogen (N) deposition to lakes and watersheds has been increasing steadily due to various anthropogenic activities. Because such anthropogenic N is widely distributed, even lakes relatively removed from direct human disturbance are potentially impacted. However, the effects of increased atmospheric N deposition on lakes are not well documented. We examined phytoplankton biomass, the absolute and relative abundance of limiting nutrients (N and phosphorus [P]), and phytoplankton nutrient limitation in alpine lakes of the Rocky Mountains of Colorado (USA) receiving elevated (> 6 kg N x ha(-1) x yr(-1)) or low (< 2 kg N x ha(-1) x yr(-1)) levels of atmospheric N deposition. High-deposition lakes had higher NO3-N and total N concentrations and higher total N : total P ratios. Concentrations of chlorophyll and seston carbon (C) were 2-2.5 times higher in high-deposition relative to low-deposition lakes, while high-deposition lakes also had higher seston C:N and C:P (but not N:P) ratios. Short-term enrichment bioassays indicated a qualitative shift in the nature of phytoplankton nutrient limitation due to N deposition, as high-deposition lakes had an increased frequency of primary P limitation and a decreased frequency and magnitude of response to N and to combined N and P enrichment. Thus elevated atmospheric N deposition appears to have shifted nutrient supply from a relatively balanced but predominantly N-deficient regime to a more consistently P-limited regime in Colorado alpine lakes. This adds to accumulating evidence that sustained N deposition may have important effects on lake phytoplankton communities and plankton-based food webs by shifting the quantitative and qualitative nature of nutrient limitation.

Shifts in lake N:P stoichiometry and nutrient limitation driven by atmospheric nitrogen deposition.

Human activities have more than doubled the amount of nitrogen (N) circulating in the biosphere. One major pathway of this anthropogenic N input into ecosystems has been increased regional deposition from the atmosphere. Here we show that atmospheric N deposition increased the stoichiometric ratio of N and phosphorus (P) in lakes in Norway, Sweden, and Colorado, United States, and, as a result, patterns of ecological nutrient limitation were shifted. Under low N deposition, phytoplankton growth is generally N-limited; however, in high-N deposition lakes, phytoplankton growth is consistently P-limited. Continued anthropogenic amplification of the global N cycle will further alter ecological processes, such as biogeochemical cycling, trophic dynamics, and biological diversity, in the world's lakes, even in lakes far from direct human disturbance.

Vitamin D protects keratinocytes from deleterious effects of ionizing radiation.

BACKGROUND: Radiotherapy can induce severe skin responses that may limit the clinically acceptable radiation dose. The responses include erythema, dry and moist desquamation, erosions and dermal-epidermal blister formation. These effects reflect injury to, and reproductive failure of, epidermal cells and may also be due to dysregulation of the tissue remodelling process caused by excessive proteolytic activity. Calcitriol, the hormonally active vitamin D metabolite, protects keratinocytes from programmed cell death induced by various noxious stimuli. OBJECTIVE: To examine whether calcitriol protects proliferating keratinocytes from the damage inflicted by ionizing radiation under conditions similar to those employed during radiotherapy. METHODS: Autonomously proliferating HaCaT keratinocytes, used as a model for basal layer keratinocytes, were irradiated using a linear accelerator. Cell death was monitored by vital staining, executioner caspase activation, lactic dehydrogenase release and colony formation assay. Induction of matrix metalloproteinase-9 was assessed by gelatinase activity assay and mRNA determination. Levels of specific proteins were determined by immunoblotting. RESULTS: Treatment with calcitriol inhibited both caspase-dependent and -independent programmed cell death occurring within 48 h of irradiation and increased the colony formation capacity of irradiated cells. These effects may be attributable to inhibition of the c-Jun NH(2)-terminal kinase cascade and to upregulation of the truncated antiapoptotic isoform of p63. Treatment with the hormone also attenuated radiation-induced increase in matrix metalloproteinase-9 protein and mRNA levels. CONCLUSIONS: The results of this study suggest that active vitamin D derivatives may attenuate cell death and excessive proteolytic activity in the epidermis due to exposure to ionizing radiation in the course of radiotherapy.

EGFR in Enamel Matrix Derivative-induced gingival fibroblast mitogenesis.

We previously reported that EMD (Enamel Matrix Derivative) induces proliferation of human gingival fibroblasts via activation of Extracellular Regulated Kinase (ERK), and this study assessed the possible mediatory role of EGFR (Epidermal Growth Factor Receptor) in this effect. Treatment of gingival fibroblasts with EMD resulted in tyrosine phosphorylation of the EGFR, as assessed by immunoblotting and ELISA, while EMD-induced ERK activation and thymidine incorporation were markedly inhibited (approximately 40-50%) by a specific EGFR tyrosine kinase inhibitor. Using appropriate inhibitors, we established that EMD-induced EGFR activation is largely due to shedding of HB-EGF (Heparin-binding EGF) from the cell membrane via a metalloproteinase-mediated process. Finally, the addition of PP1, a Src family inhibitor, abrogated both EGFR phosphorylation and ERK activation. Taken together, these results indicate that, at least in human gingival fibroblasts, EMD-induced ERK activation and proliferation are partially due to a Src-dependent, metalloproteinase-mediated transactivation of EGFR.

Enamel matrix derivative protects human gingival fibroblasts from TNF-induced apoptosis by inhibiting caspase activation.

Emdogain, a formulation of enamel matrix derivative (EMD), is used clinically for regeneration of the periodontium (tooth supporting tissues), but the molecular mechanisms of its action have not been elucidated. Several clinical studies suggested that EMD may also improve gingival healing after periodontal surgery and thus affect the fate of gingival fibroblasts (GFs). Since these cells are targets for local inflammatory mediators such as TNF, a pro-apoptotic cytokine, during the course of periodontal disease, we tested whether EMD protects human GFs (hGFs) from TNF-induced cytotoxicity. Quiescent primary hGFs were challenged with TNF (10-100 ng/ml) with or without EMD (100 microg/ml) pretreatment. Cell viability was assessed by neutral red staining, cell death by LDH release and apoptosis by caspase activity. Signaling pathways were evaluated by Western blotting and pharmacological inhibitors. TNF induced classical signs of apoptosis in hGFs, including typical cellular morphology and increased caspase activity. TNF-induced cytotoxicity was entirely caspase-dependent. Pretreatment (4-24 h) with EMD dramatically inhibited the activation of initiator and executioner caspases and enhanced hGF survival. Although TNF induced the activation of p38 MAPK, JNK, ERK and PI-3K signaling, these pathways were not crucial for EMD protection of hGFs. However, EMD increased the levels of c-FLIP(L), an anti-apoptotic protein located upstream of caspase activation. These data demonstrate, for the first time, that EMD protects hGFs from inflammatory cytokines and, together with our recent reports that EMD stimulates rat and human GF proliferation, could help explain the mechanisms whereby in vivo use of EMD promotes gingival healing.

Enamel matrix derivative stimulates human gingival fibroblast proliferation via ERK.

Emdogain, a formulation of Enamel Matrix Proteins, is used clinically for periodontal regeneration to stimulate PDL (periodontal ligament), cementum, and bone formation. Its effects on gingival fibroblasts and tissue have not been thoroughly studied. Therefore, we investigated the mechanisms by which Emdogain affects the cell cycle of human gingival fibroblasts. Without serum, Emdogain (50 microg/mL) induced human gingival fibroblast entry into the S phase and DNA synthesis, but not completion of the cell cycle. With low serum concentrations (0.2-0.5%), Emdogain synergistically induced completion of the cell cycle, resulting in increased cell numbers. The mitogenic response to Emdogain depended on Extracellular Regulated Kinase (ERK) activation, which occurred in two waves, peaking after 15 min and 4 to 6 hrs, since it was abolished by U0126, a specific MAPK inhibitor. Inhibition of the second wave was sufficient to abrogate mitogenesis. This study characterized the mitogenic effect of Emdogain on primary human gingival fibroblasts, its cooperation with serum growth factors, and the key mediatory role of the ERK cascade.

Programmed cell death of stressed keratinocytes and its inhibition by vitamin D: the role of death and survival signaling pathways.

The epidermis is confronted with multiple environmental and pathophysiological stresses. This study shows that TNFalpha, oxidative stress, hyperosmotic and heat shock induced both caspase-dependent and independent cell death in human HaCaT keratinocytes. The hormonal form of vitamin D, 1,25(OH)2D3, which is an autocrine hormone in the epidermis, protected the cells from all the examined stresses and pathways leading to cell death. We aimed to define the signaling pathways that determine the life-death balance of stressed keratinocytes and participate in their protection by 1,25(OH)2D3. As assessed by employing specific inhibitors, the survival pathways mediated by the EGF receptor, ERK, PI-3K or Src kinase, or basal transcriptional activity are important for unstressed cell survival. However, only the EGF receptor, PI-3K and the Src kinase pathways mediate the survival of stressed cells in a stimulus-specific manner. Inhibition of the p38 and/or the JNK death pathways reduced caspase activation induced by oxidative stress, hyperosmotic shock and TNFalpha. The protective effect of 1,25(OH)2D3 was not mediated by the examined survival pathways. 1,25(OH)2D3 inhibited the stress-induced activation of p38 and JNK. Since mimicking this effect by pharmacological inhibition resulted in the attenuation of caspase activation, we infer that these pathways are involved in keratinocyte protection by 1,25(OH)2D3.

13C-urea breath test for success of Helicobacter pylori eradication: study of 5885 Israeli patients.

Helicobacter pylori (Hp) infection is highly prevalent in many countries and may cause gastritis, peptic ulcer disease, gastric cancer, and lymphoma. Successful eradication depends on the specific treatment used, patient compliance, and Hp antibiotic resistance. The primary aim was to characterize groups of patients with one or more failures of Hp eradication treatment. The secondary aim was to evaluate the factors that influence eradication failure. Between April 1, 1998, and December 31, 2001, 5885 patients were studied for the success of Hp eradication with the 13C-urea breath test (13C-UBT): 5442 after one course of treatment (Group I), 380 after two courses (Group II), and 63 after three courses (Group III). The 13C-UBT was positive in 27.8%, 37.4%, and 47.6% of patients in Groups I, II, and III, respectively (P(I-II) = 0.000, P(II-III) = 0.126). A combination of omeprazole, amoxicillin, and clarithromycin (OAC) was used in 31.3%, 27.4%, and 7.9% of Groups I, II, and III, respectively, and a combination of omeprazole, amoxicillin, and metronidazole (OAM) in 15.2%, 28.9%, and 28.6%, respectively. Regimens that contained clarithromycin were used in decreasing order in Groups I, II, and III, and regimens containing metronidazole, bismuth, or tetracycline, in increasing order. The only good prognostic factor for successful eradication was Israeli origin, while European-American and Asian-African origin, recurrence of symptoms, a history of duodenal ulcer, and chronic proton pump inhibitor (PPI) use did not favor successful eradication. Our results suggest that origin, history of peptic disease, and chronic PPI use are predictors of eradication failure.

An ancillary tool for the diagnosis of amyloid A amyloidosis in a variety of domestic and wild animals.

Immunohistochemistry, the standard method for diagnosing amyloid A (AA) amyloidosis, is limited in animals because it requires a large array of animal-specific anti-AA antibodies, not commercially available. The Shtrasburg method (SH method) is a highly specific and sensitive technique, helping in the diagnosis and determination of AA amyloidosis in humans. The aim of this study is to determine whether the SH method is applicable in the diagnosis of AA amyloidosis in a variety of animals. Tissue samples were obtained from animals suffering from spontaneous or experimentally induced AA amyloidosis (mice, hamsters, guinea pigs, cheetahs, cats, cows, ducks, a dog, a goose, a chicken, and a turaco). Detection of the amyloid and quantitative evaluation were performed using Congo red staining, and specific AA typing was performed by the potassium permanganate technique. The studied tissues were subjected to the SH method, which confirmed the AA nature of the amyloid deposit, by displaying in polyacrylamide gel electrophoresis protein bands consistent with the molecular weight of the species-specific AA, in all the animals examined, except mice, hamsters, and guinea pigs. N-terminal analysis of these bands corroborated their AA origin. We conclude that the SH method may be used as an ancillary simple tool for the diagnosis of AA amyloidosis in a large number of domestic and wild animals. Moreover, our findings further increase the feasibility of applying this method in humans.

Trichloroacetic acid cycling in Sitka spruce saplings and effects on sapling health following long term exposure.

Trichloroacetic acid (TCA, CCl(3)COOH) has been associated with forest damage but the source of TCA to trees is poorly characterised. To investigate the routes and effects of TCA uptake in conifers, 120 Sitka spruce (Picea sitchensis (Bong.) Carr) saplings were exposed to control, 10 or 100 microg l(-1) solutions of TCA applied twice weekly to foliage only or soil only over two consecutive 5-month growing seasons. At the end of each growing season similar elevated TCA concentrations (approximate range 200-300 ng g(-1) dwt) were detected in both foliage and soil-dosed saplings exposed to 100 microg l(-1) TCA solutions showing that TCA uptake can occur from both exposure routes. Higher TCA concentrations in branchwood of foliage-dosed saplings suggest that atmospheric TCA in solution is taken up indirectly into conifer needles via branch and stemwood. TCA concentrations in needles declined slowly by only 25-30% over 6 months of winter without dosing. No effect of TCA exposure on sapling growth was measured during the experiment. However at the end of the first growing season needles of saplings exposed to 10 or 100 microg l(-1) foliage-applied TCA showed significantly more visible damage, higher activities of some detoxifying enzymes, lower protein contents and poorer water control than needles of saplings dosed with the same TCA concentrations to the soil. At the end of each growing season the combined TCA storage in needles, stemwood, branchwood and soil of each sapling was <6% of TCA applied. Even with an estimated half-life of tens of days for within-sapling elimination of TCA during the growing season, this indicates that TCA is eliminated rapidly before uptake or accumulates in another compartment. Although TCA stored in sapling needles accounted for only a small proportion of TCA stored in the sapling/soil system it appears to significantly affect some measures of sapling health.