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The latest outbreak of a coronavirus disease in 2019 (COVID‐19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), evolved into a worldwide pandemic with massive effects on health, quality of life, and economy. Given the short period of time since the outbreak, there are several knowledge gaps on the comparative and zoonotic aspects of this new virus. Within the One Health concept, the current EAACI position paper dwells into the current knowledge on SARS‐CoV‐2’s receptors, symptoms, transmission routes for human and animals living in close vicinity to each other, usefulness of animal models to study this disease and management options to avoid intra‐ and interspecies transmission. Similar pandemics might appear unexpectedly and more frequently in the near future due to climate change, consumption of exotic foods and drinks, globe‐trotter travel possibilities, the growing world population, the decreasing production space, declining room for wildlife and free‐ranging animals, and the changed lifestyle including living very close to animals. Therefore, both the society and the health authorities need to be aware and well prepared for similar future situations, and research needs to focus on prevention and fast development of treatment options (medications, vaccines).

Mineralized polymer scaffolds have proven to be effective biomaterials for inducing osteoinductivity in bone tissue engineering. Sequential mineralization is a promising technique for depositing minerals in three-dimensional (3D) scaffolds. Paper, which is made of cellulose fibers, can be used as a tissue scaffold due to its highly porous structure and flexibility, as well as its excellent ability to wick fluids and support the growth of bone cells. In this study, paper-based, mineralized scaffolds were fabricated using sequential mineralization. We conducted experiments with two groups of scaffolds based on different incubation times in the mineralization solutions (30 min and 24 h). Ten cycles of mineralization were performed for each group. We found that the mineral content increased as the cycle number increased and that the 24-h group scaffolds consistently had more mineralization than did the 30-min group scaffolds when measured at the same cycle number. A quantitative reverse transcription-polymerase chain reaction was performed for two osteogenic differentiation markers of the preosteoblasts that were grown on the mineralized paper scaffolds. The gene expression results for bone-specific markers revealed that the mineralized scaffolds were osteoinductive. Subcutaneous implantation of the scaffolds in rats demonstrated favorable biocompatibility, high vascularization, and non-immunogenicity in vivo. The overall results suggest that the sequentially mineralized paper scaffolds are promising materials for use in bone tissue engineering.

RATIONALE Paper spray (PS) has been developed as a method of choice for point-of-care analysis in many real cases, where its applications can be further expanded with delicate high-throughput design. To achieve this goal, we developed a new PS regime, with the assembly of an induced high voltage into the ion source. Compared with regular DC high voltage, the newly developed setup is capable of high-throughput, simple configuration and rapid switching between individual papers without complicated electric/mechanic design. METHODS A device of high-throughput induced PS (IPS) was designed by using a two-dimensional (2D) rotating platform equipped with a circular glass plate. The paper substrate was placed on the circular glass plate and separated from the electrode. The method avoids physical contact between the electrode and the sample. Charged droplets were generated at the paper tip once an induced high voltage was applied to a wet paper. RESULTS A relatively rapid analytical speed of 2.6 s per sample was achieved via IPS-MS. Rapid quantification of amitriptyline (AMT) in complicated matrices was obtained within 1 min using an isotope internal standard method. Limits of detection for AMt in urine, FBS and blood were calculated to be 1.04, 0.84 and 1.33 ng/mL, respectively. In addition, high-throughput IPS-MS can be used for chemical reaction monitoring. CONCLUSIONS We have demonstrated the versatility of high-throughput IPS-MS in ambient ionization, which successfully simplified the experimental installation and facilitated the experimental operation. Therefore, we believe that high-throughput IPS-MS analysis will be widely used for discovering drugs and screening reactions, and the present design has the potential for applications in paper chip-MS analysis.

A potentially universal approach is presented for enzyme attachment to cellulose that significantly enhances enzyme stability while retaining high activity, and involves no chemical functionalization of cellulose. Bovine serum albumin (BSA) was interlocked in cellulose to form a protein-friendly surface (named BSA-Paper), while also providing COOH and NH2 groups for subsequent attachment of enzymes. The desired enzyme is then mixed with additional BSA and interlocked on BSA-Paper. The second BSA layer dilutes and crosslinks the enzyme for improved stability. Laccase was tested as a model enzyme for interlocking on BSA-Paper, and was found to retain over 100 % activity and was 240 times more stable at 25 °C (half life=180 d) than laccase. This new approach was also tested with a few other enzymes with encouraging results, thus providing a potentially universal method for stabilization of enzymes on cellulose with retention of high activities.

This paper describes enzyme-linked immunosorbent assays (ELISAs) utilizing microfluidic thread/paper-based analytical devices (μTPAD), microfluidic fabric-based analytical devices (μFAD), and microfluidic thread-based analytical devices (μTAD). Here, the quantitative detection of biotinylated goat anti-mouse IgG (system one) and rabbit IgG (system two) antibodies via colorimetric analysis is detailed. In both systems, antibody is spotted on the detection site and subjected to a series of washes, addition of streptavidin-alkaline phosphatase (Strep-ALP) (system 1) or alkaline phosphatase (ALP)-conjugated secondary antibody (system 2), and colorimetric substrate. The devices are scanned and analyzed yielding a correlation between inverse yellow (or purple) intensity. For system one, a linear range of detection at low concentrations of streptavidin-alkaline phosphatase (Strep-ALP) was observed befire the enzyme reached a Vmax . At higher concentrations of Strep-ALP, saturation is achieved for both the μTPAD and μFAD devices. For system two, the IC50 values obtained for the non-trifurcated and trifurcated μTADs were determined to be 180.2 fmol/zone and 133.8 fmol/zone, respectively. The IC50 value was demonstrated to be 1034 fmol/zone and 208.6 fmol/zone for the μTPADs and μFADs, respectively. For all devices the lowest concentration of Strep-ALP or rabbit IgG used in the assay was 3.75 × 10-4 mg/mL and 0.7 fmol/zone, respectively. The development of this technology should further facilitate the use of these platforms for ELISA to detect and quantitate antibodies.

I am very thankful to Kuznetsov for his comments on our recent paper about serial structures published in this journal. I hope this is just the beginning of a much wider, and holistic, discussion on the evolution of serial homologous structures, and of so-called "serial structures" in general, whether they are truly serial homologs or the secondary result of homoplasy. Strangely, Kuznetsov seems to have missed the main point of our paper, what is particularly puzzling as this point is clearly made in the very title of our paper. For instance, he states that "Siomava et al. claim that the serial homologues are false because they are ancestrally anisomeric (dissimilar)' and that" Siomava et al., (Siomava et al., Journal of Morphology, 2020, 281, 1110-1132) expected that if serial homology was true, then the serial homologs would be identical at the start and then only diverge. " However, our paper clearly did not state this. Instead, we stated that (a) serial homology is a real phenomenon, and (b) ancestral dissimilarity is actually likely the norm, and not the exception, within serial homology. In particular, our paper showed that, as clearly stated in its title and abstract, within the evolution of serial homologues these structures "many times display trends toward less similarity while in many others display trends toward more similarity, that is, one cannot say that there is a clear, overall trend to anisomerism." Serial homology is therefore a genuine and much widespread phenomenon within the evolution of life in this planet. It is clearly one of the most important issues-and paradoxically one of the less understood, precisely because of the a priori acceptance of long-standing assumptions that have never been empirically tested, some of them repeated in Kuznetsov's paper-within macroevolution and comparative anatomy.

Silver nanoparticles (AgNPs) are in situ synthesized for the first time on microfibrillated silk (MFS) exfoliated from domesticated Philosamia cynthia ricini (eri) and Bombyx mori (mulberry) silkworm silk fibers. The process is rapid (hours time), does not rely on harmful chemicals, and produces robust and flexible AgNPs coated MFS (MFS-AgNPs) protein papers with excellent handling properties. None of these can be achieved by approaches used in the past to fabricate AgNPs silk systems. MFS bonds the AgNPs strongly, providing good support and stabilization for the NPs, leading to strong wash fastness. The mechanical properties of the MFS-AgNPs papers largely do not change compared to the MFS papers without nanoparticles, except for some higher concentration of AgNPs in the case of mulberry silk. The improved tensile properties of eri silk papers with or without AgNPs compared to mulberry silk papers can be attributed to the higher degree of fibrillation achieved in eri silk and its inherent higher ductility. MFS-AgNPs from eri silk also exhibit strong antibacterial activity. This study provides the basis for the development of smart protein papers based on silk fiber and functional nanomaterials.

Paper-supported cell culture is an unprecedented development for advanced bioassays. This study reports a strategy for in vitro engineering of cell-compatible paper chips that allow for adherent cell culture, quantitative assessment of drug efficiency, and label-free sensing of intracellular molecules via paper spray mass spectrometry. The polycarbonate paper is employed as an excellent alternative bioscaffold for cell distribution, adhesion, and growth, as well as allowing for fluorescence imaging without light scattering. The cell-cultured paper chips are thus amenable to fabricate 3D tissue construction and cocultures by flexible deformation, stacks and assembly by layers of cells. As a result, the successful development of cell-compatible paper chips subsequently offers a uniquely flexible approach for in situ sensing of live cell components by paper spray mass spectrometry, allowing profiling the cellular lipids and quantitative measurement of drug metabolism with minimum sample pretreatment. Consequently, the developed paper chips for adherent cell culture are inexpensive for one-time use, compatible with high throughputs, and amenable to label-free and rapid analysis.

Several eusocial wasps are prominent invaders to remote islands. The paper wasp Polistes chinensis antennalis is native to East Asia, was introduced to New Zealand in 1979 and has expanded its distribution there. This provides an excellent opportunity to examine the impacts of an initial bottleneck and subsequent expansion on genetic structure. We analysed and compared the genetic population structures of the native (Japan and South Korea) and invasive New Zealand populations. Although 94% of individuals had shared haplotypes detected across both populations, the remaining 6% had private haplotypes identified in only one of the three countries. The genetic variation at microsatellite loci was lower in New Zealand than in native countries, and the genetic structure in New Zealand was clearly distinct from that in its native range. Higher frequencies of diploid-male- and triploid-female-producing colonies were detected in New Zealand than in the native countries, showing the reduction in genetic variation via a genetic bottleneck. At least two independent introductions were suggested, and the putative source regions for New Zealand were assigned as Kanto (central island) and Kyushu (south island) in Japan. Serial founder events following the initial introduction were also indicated. The estimated dispersal distance between mother and daughter in New Zealand was twice that in Japan. Thus, the introduction history of P. chinensis antennalis in New Zealand is probably the result of at least two independent introductions, passing through a bottleneck during introduction, followed by population expansion from the point of introduction.

Objective To evaluate endodontic paper point tear test (PPTT) in rabbits and compare changes in corneal touch threshold (CTT) induced by Schirmer tear test (STT) and PPTT. Normal corneal sensitivity recovery time after both tear tests was also measured. Also, mean PPTT and STT values were obtained. Methods Tear production of 20 eyes, from 10 rabbits, was evaluated using STT and the PPTT. Central corneal touch threshold was measured by a Cochet-Bonnet esthesiometer before any tear test was performed (zero time), immediately after the test (1 min), and consecutively at 6, 11, 16, and 26 min. Tests were conducted on three consecutive days: Day 1 – control condition, no tear tests performed only the CTT; Day 2 – CTT before and after PPTT; and Day 3 – CTT before and after STT. CTT values were compared using repeated measures ANOVA. Results Corneal touch threshold was significantly increased for at least 16 min after STT, indicating STT causes corneal discomfort. No difference was found between CTT following PPTT and controls, indicating PPTT caused minimal corneal discomfort. The mean (±SD) value for STT was 5.2 ± 1.0 mm/min and for PPTT was 13.8 ± 1.5 mm/min. Discussion The aqueous fraction of rabbit's tears can be successfully measured by PPTT. This report established reference values for PPTT in rabbits. Additionally, the absence of a significant difference in CTT after PPTT compared with controls shows that PPTT is well tolerated by rabbits. Considering the improved comfort (compared with STT), accuracy, and low cost, PPTT is a bona fide method of measuring aqueous tear production in rabbits.

Adverse effects of pulp and paper mill effluent on fish populations have been well documented in many countries over the last two decades. Some of the initial studies were at mills with conventional chlorine bleaching and no secondary effluent treatment. Following installation of secondary treatment, changes in bleaching technology to elemental chlorine-free bleaching, and other process changes, adverse effects on fish were reduced or eliminated at some mills. Because no two mills are exactly alike, it is difficult to predict adverse impacts of any given mill on fish populations. In 1994, a study of female white sucker (Catostomus commersoni) in the Androscoggin River, Maine, USA, showed induction of mixed function oxidase, reductions in gonad size and plasma estradiol, and an increase in plasma testosterone in fish downstream of discharges from three large bleached kraft pulp and paper mills, and host community municipal sewage treatment plants (STP). After all three mills switched to elemental chlorine-free bleaching in the late 1990s, studies from 2001 to 2003 found that the pattern of reproductive impacts on white sucker populations measured in 1994 was not repeated. In addition, population estimates of white sucker from 2002 to 2003 using mark–recapture techniques found that densities and biomass were well within the range of those of a reference population, and of those reported in the literature for unimpacted populations. Detailed studies immediately above and below each mill/sewage treatment plant showed no evidence of reproductive effects. However, a clear pattern of eutrophication was noted, which increased cumulatively downstream below each mill/STP. Environ. Toxicol. Chem. 2011;30:142–153. © 2010 SETAC

Aromatic sensitizers and related compounds (SRCs) originating from thermal recording material as impurities in waste paper have been detected in aquatic environments near cities where the paper industry is flourishing. In the present study, the levels of exposure to such SRCs and the stable isotope ratios (δ13C and δ15N) in different fish species (Konosirus punctatus, Takifugu niphobles, and Tribolodon hakonensis) were analyzed. These fish species were collected from the vicinity of waste paper recycling plants in Japan. Eleven SRCs were identified in fish muscle at total concentration levels (ΣSRCs) of 64 to 818 ng/g wet weight (mean 181 ng/L wet wt). The dominant SRCs in the fish samples were 1,1-di(4-methylphenyl)ethane, 1,2-bis(3-methylphenoxy)ethane, and 1,4-dibenzyloxybenzene. Stable isotope ratios showed that, although three species feed on the same trophic level, K. punctatus and T. niphobles live in waters farther offshore than those in which T. hakonensis live. The contributions of various factors to the accumulation were also assessed. The lipid content (r2 = 0.512, p

Although effluent treatment systems within pulp and paper mills remove many toxicants and improve wastewater quality, there is a need to understand and quantify the effectiveness of the treatment process. At a combined news and kraft pulp and paper mill in northwestern Ontario, Canada, fathead minnow (FHM) reproduction and physiology were examined before, during, and after a short-term (6-d) exposure to 10% (v/v) untreated kraft mill effluent (UTK), 25% (v/v) secondary treated kraft mill effluent (TK), and 100% (v/v) combined mill outfall (CMO). Although UTK exposure significantly decreased egg production, neither TK nor CMO caused any reproductive changes. The expression of six genes responsive to endocrine-disrupting compounds, stress, or metals was then examined in livers of these fish using real-time polymerase chain reaction. In female FHMs, none of the three effluents induced significant expression changes in any genes investigated. By contrast, in males there were significant increases in the mRNA levels of androgen receptor, estrogen receptor (ER) β, and cytochrome P4501A (CYP1A) upon UTK and TK exposure but no changes in ERα or vitellogenin (VTG) gene expression, whereas CMO exposure significantly increased the mRNA levels of ERα, VTG, and CYP1A. Together, these results suggest that kraft effluent before and after biological treatment contained compounds able to induce androgenic effects in FHMs, and that combination of kraft and newsmill effluents eliminated the androgenic compounds while inducing distinct and significant patterns of gene expression changes that were likely due to estrogenic compounds produced by the newsmill. Environ. Toxicol. Chem. 2010;29:430–439. © 2009 SETAC

Paper microfluidics has attracted much attention since its first introduction around one decade ago due to the merits such as low cost, ease of fabrication and operation, portability, and facile integration with other devices. The dominant application for paper microfluidics still lies in point-of-care testing (POCT), which holds great promise to provide diagnostic tools to meet the ASSURED criteria. With micro/nanostructures inside, paper substrates provide a natural 3D scaffold to mimic native cellular microenvironments and create excellent biointerfaces for cell analysis applications, such as long-term 3D cell culture, cell capture/phenotyping, and cell-related biochemical analysis (small molecules, protein DNA, etc.). This review summarizes cell-related applications based on various engineered paper microdevices and provides some perspectives for paper microfluidics-based cell analysis.

We investigated the effects of repairing large tympanic membrane (TM) perforations in rats with a thin silk patch compared with the commonly used paper patch. We performed bilateral myringotomies of 1.8 mm in diameter on 50 adult Sprague-Dawley rats with intact TMs. The perforations in the right ears of 40 rats were treated with a silk patch, and the perforations in the left ears of the same rats were treated with a paper patch. Ten rats acted as controls. The mean healing times of the TM perforations on the silk-patch-treated ears and the paper-patch-treated ears were 7.2+/-1.48 and 9.1+/-1.11 days, respectively (control 10.38+/-1.70 days). The difference between silk-patch- and paper-patch-treated ears was statistically significant, with a mean difference of 1.9 days (0.6-4.5 days). The mean perforation closure times were significantly shorter in silk-patch- and paper-patch-treated ears than in the control animals. The endoscopic and histological findings of this study provide evidence that silk-patch treatment accelerates wound healing and shortens TM perforation closure time. We suggest that the silk patch may prove to be an effective material for repairing TM perforations in human patients in an outpatient clinical setting.

The Canadian environmental effects monitoring (EEM) program is a regulated, cyclical, industry-funded program designed to determine whether receiving water impacts exist when a mill is in compliance with its discharge limits. The results from three cycles of the fish monitoring program (1992 to 2004) are available from over 200 surveys of fish compared between sites located upstream and downstream of pulp and paper mill effluent outfalls. Previous meta-analyses have shown a national average response pattern across cycles characterized by an increase in endpoints measuring energy storage and growth and a decrease in a reproductive endpoint, consistent with a response of nutrient enrichment in combination with some form of metabolic disruption. Although the national average pattern of effects was temporally consistent, there was some variability in the magnitude of effects among cycles. Questions were raised as to whether the intercycle variability was due to changes in effluent quality or due, at least in part, to other factors. The present study compares responses over the first three cycles, and shows that the choice of sentinel species is likely to be a major contributing factor to the variability in observed effects. Subset analyses using studies from mills that used the same sentinel species across cycles reveal fairly uniform responses and little evidence of significant improvements in overall fish health from cycles one to three. However, a meta-analysis using 1991 data collected from 10 mills before the implementation of the EEM program and data from the same mills collected during cycles one to three of the program reveal significantly reduced effects on relative liver weight and potential improvements in other endpoints. Environ. Toxicol. Chem. 2010;29:440–452. © 2009 SETAC

Hormonal analysis of urine from free-ranging primates has been limited due to the difficulty of preserving samples under field conditions. Drying urine on filter paper is an alternative for field preservation. This study describes a new laboratory method for eluting steroids from filter paper with methanol, along with a series of experiments used to develop and validate this method. The overall elution recovery of estrone sulfate (ES) from filter paper was 86.4%. Estrone conjugate (E1C) levels in humans and captive orangutans were analyzed by radioimmunoassay (RIA). Values from samples dried on filter paper were significantly correlated with values from matched frozen samples, with elution efficiencies ranging between 97.1% and 102.4%. Creatinine (Cr) measurements from frozen urine compared to urine dried on filter paper were also significantly correlated (r=0.96) with an elution efficiency of 101.7%. After the samples were stored for 2 years, the absolute values of E1C and Cr were significantly lower but were still significantly correlated with frozen urine values. These data demonstrate the effectiveness of filter paper as a medium for preserving urinary steroid samples, and the efficiency of methanol as a solvent for eluting E1C and Cr. This method thus provides a viable alternative to the traditional procedure of freezing urine for field studies, where freezers are not available.

Nerve regeneration was examined in 108 mouse sciatic nerves in a prospective morphometric study in which a new anastomotic technique using nitrocellulose paper was compared with conventional suturing and simple sectioning. Mice were randomly allocated to one of the three arms of the trial and sciatic nerves were examined following surgery at time intervals up to 8 weeks. Serial sections were taken for qualitative and quantitative morphometric analyses. Counts demonstrated that the rate of regeneration was faster in the nitrocellulose group than following nerve section alone, but that the suture group was even better. However, 8 weeks after surgery, there were no significant differences in myelinated axon counts 4mm distal to the site of section between any of the experimental groups. The superlative regenerative capacity of rodent axons may limit the applicability of this model to human nerve tissue.

Paper is used in various applications in biomedical research including diagnostics, separations, and cell cultures. Paper can be conveniently engineered due to its tunable and flexible nature, and is amenable to high-throughput sample preparation and analysis. Paper-based platforms are used to culture primary cells, tumor cells, patient biopsies, stem cells, fibroblasts, osteoblasts, immune cells, bacteria, fungi, and plant cells. These platforms are compatible with standard analytical assays that are typically used to monitor cell behavior. Due to its thickness and porous nature, there are no mass transport limitations to/from the cells in paper scaffolds. It is possible to pattern paper in different scales (micrometer to centimeter), generate modular configurations in 3D, fabricate multicellular and compartmentalized tissue mimetics for clinical applications, and recover cells from the scaffolds for further analysis. 3D paper constructs can provide physiologically relevant tissue models for personalized medicine. Layer-by layer strategies to assemble tissue-like structures from low-cost and biocompatible paper-based materials offer unique opportunities that include understanding fundamental biology, developing disease models, and assembling different tissues for organ-on-paper applications. Paper-based platforms can also be used for origami-inspired tissue engineering. This work provides an overview of recent progress in engineered paper-based biomaterials and platforms to culture and analyze cells.

Pulp and paper mill effluents were examined for in vitro androgenicity using a recombinant yeast-based androgen receptor assay. Low levels of androgenic effects were detected in extracts of some effluents after activated sludge treatment. Yeast cell growth cell was inhibited in some samples, and this interfered with the androgenic response. Solid-phase fractionation revealed androgenicity after partial separation of components in the effluents. Comparison of levels in untreated effluent and in effluent treated in an aerated lagoon showed that this treatment had only a marginal effect on androgenicity. An assay of the fractions eluted with increasing concentrations of methanol showed that androgenic compounds were low to moderately lipophilic. In an attempt to identify these compounds, a number of wood-related compounds (guaiacol, vanillin, β-sitosterol, betulin, pinosylvin-O-methyl ether, and a wood extract enriched in lignans) were examined but were found not to be androgenic. Raw process water was not androgenic, but water from a highly humified lake and process water from the production of pulp from partly decayed wood had low androgenicity. It therefore can be plausibly suggested that the androgens originated in decaying wood. An assay of androgenicity in the bile of juvenile rainbow trout exposed to effluents for 3 weeks showed increased dose-dependent levels of androgens after enzymatic hydrolysis of hormone conjugates. © 2004 Wiley Periodicals, Inc. Environ Toxicol 19: 510–517, 2004.

International audience; The prevalence of allergic diseases such as allergic rhinitis, asthma, food allergy, and atopic dermatitis has increased dramatically during the last decades, which is associated with altered environmental exposures and lifestyle practices. The purpose of this review was to highlight the potential role for dietary fatty acids, in the prevention and management of these disorders. In addition to their nutritive value, fatty acids have important immunoregulatory effects. Fatty acid-associated biological mechanisms, human epidemiology, and intervention studies are summarized in this review. The influence of genetics and the microbiome on fatty acid metabolism is also discussed. Despite critical gaps in our current knowledge, it is increasingly apparent that dietary intake of fatty acids may influence the development of inflammatory and tolerogenic immune responses. However, the lack of standardized formats (ie, food versus supplement) and standardized doses, and frequently a lack of prestudy serum fatty acid level assessments in clinical studies significantly limit our ability to compare allergy outcomes across studies and to provide clear recommendations at this time. Future studies must address these limitations and individualized medical approaches should consider the inclusion of specific dietary factors for the prevention and management of asthma, food allergy, and atopic dermatitis.

To develop an effective hybrid bioartificial liver (BAL) device, the material of the scaffold is very important to support hepatocytes that have both growth ability and hepatic differentiated functions. In this study we used paper (Kimwipe, Kimberly-Clark Corp., Roswell, GA, U.S.A.) as a scaffold. Primary hepatocytes isolated from a normal adult rat liver could proliferate on the paper. The secretion of albumin into culture medium by the cells on the paper increased with time in culture and, compared to the cells on dishes, the amount of 48 h albumin secretion at Day 10 was two times larger. Perpendicular sections of hepatocytes on the paper revealed that the cells fell into cavities made by intersecting fibers, piled up, and formed three to four layers. The piled-up cells changed their shape from flat to cuboidal and enlarged their cytoplasm, which was rich in organelles such as mitochondria and peroxisomes with a nucleoid. In addition, they formed bile canalicular structures between the cells. Their morphological appearance was similar to in vivo hepatocytes. Paper (Kimwipe) may be a good candidate as a scaffold to make a BAL device.

The latitudinal herbivory-defense hypothesis (LHDH) predicts that plants near the equator will be more heavily defended against herbivores than are plants at higher latitudes. Although this idea is widely found in the literature, recent studies have called this biogeographic pattern into question. We sought to evaluate the LHDH in a high-latitude terrestrial ecosystem where fire and mammalian herbivores may contribute to selection for higher levels of defensive chemistry. To address this objective, we collected seeds of Alaska paper birch (Betula neoalaskana) from nine locations along two north-south transects between 55 degrees N and 62 degrees N latitudes in western, interior Canada. The birch seeds were planted in pots in a common garden in Madison, Wisconsin, USA. From the resulting seedlings, we determined levels of chemical defense by assessing the density of resin glands, which have been shown to be negatively correlated with browsing. To assess plant architectural traits such as height, mean individual leaf area, and root-to-shoot ratio, we harvested a subset of the birch seedlings. Further, we used these traits to examine growth-defense trade-offs. Contrary to the LHDH, we found a positive correlation between chemical defense and latitude. Investigating relationships with fire, we found a strong positive correlation between resin gland density and percentage of area annually burned (PAAB) around each collection location and also between PAAB and latitude. Additionally, birch seedlings originating from higher latitudes were shorter, smaller-leaved, and rootier than their lower-latitude counterparts. Growth-defense trade-offs were observed in negative correlations between resin gland density and height and leaf size. Seedlings with higher resin gland densities also allocated less biomass to shoots and more to roots. These results further call into question the LHDH and provide specific information about latitudinal trends in plant defense at high, northern latitudes where fire is a major ecosystem driver and mammals are the main herbivores of deciduous trees such as birches. We propose that these interconnected relationships are the key drivers of the positive correlation between defense and latitude in B. neoalaskana. Understanding patterns of boreal plant defense and growth is especially important because high latitude ecosystems are particularly susceptible to climate change. Key words: Alaska paper birch; Betula neoalaskana; biogeography; chemical defense; grotvth-defense trade-offs; inherent growth rate; latitude; latitudinal herbivory-defense hypothesis; papyr'feric acid; plant ar- chitecture; resin glands; root-to-shoot ratio.

Nonfluorescing protein bands can be detected by the fluorescence optics of the commercial gel electrophoresis apparatus with automated scanning of the migration path (HPGE-1000, LabIntelligence, Belmont CA), taking advantage of the decrease of emission from a fluorescent paper placed below the gel by the absorbance of proteins ("fluorescence reduction"). That decrease of fluorescence gives rise to an inverted protein peak. Nonfluorescent colorless proteins appear to reduce the intensity of light emitted from the fluorescent paper due to absorbance of incident and emitted light. When the absorbance spectrum only slightly overlaps with the excitation and emission spectra of the fluorescent paper, that reduction is weak, and detection sensitivity in that application is consequently only 1/30 of that of fluorescent proteins. By contrast, when the protein is colored so that its absorbance spectrum overlaps widely with the excitation and emission spectra of the fluorescent paper, the sensitivity of "fluorescence reduction" equals 1/4 to 1/5 of that obtained for fluorescent proteins. Bands detected by "fluorescence reduction" provide a quantitative measure of protein load and mobility. The area of the inverted bands is proportional to protein loads up to 16 microg/lane of the gel tray. A theory of "fluorescence reduction" is presented which accounts for the existence of a linear relationship between band area and load.

Rueff F, Przybilla B, Muller U, Mosbech H. The sting challenge test in Hymenoptera venom allergy. Position paper of the Subcommittee on Insect Venom Allergy of the European Academy of Allergology and Clinical Immunology. Allergy 1996: 51: 216–225. © Munksgaard 1996.

The suitability of blood collected on filter papers in comparison with corresponding conventional serum samples in the diagnosis of bovine anaplasmosis was studied using the complement fixation test, DOT-ELISA, Western immunoblot and rapid card agglutination test. Dried blood on Whatman filter paper no. 1 was eluted in PBS 0.05% Tween 20 giving an initial dilution of 1:10. The reactivity of the eluted samples in both DOT-ELISA and Western immunoblotting were similar to those obtained with the corresponding straight serum sample dilutions. Filter paper samples gave lower reactivity in the remaining tests when compared with corresponding serum samples. There was no significant difference in the reactivity between the eluates from filter papers stored at temperatures ranging between 15.5 and 24 degrees C and those kept refrigerated. Storage at 15.5 to 24 degrees C did not significantly affect reactivity for up to six months. Eluates from filter papers stored for six months at 15.5 to 24 degrees C continued to give similar reactivity as those from freshly prepared filter papers in both DOT-ELISA and Western blot, and in the rapid card agglutination test. It is concluded that collecting blood on filter papers is a suitable technique for large scale seroepidemiological studies on anaplasmosis and offers many advantages in developing countries where transport and cold chain facilities are a major constraint.

Filter paper was used as a support to absorb micro samples of finger prick blood for detection of antibodies to Entamoeba histolytica (causative organism of amebiasis) by a rapid dipstick dot-EIA technique; 8 μl of blood was sufficient to saturate discs (diameter 6 mm) of Whatman paper No. 3. Conditions of elution of blood from the discs were optimized and the best results were obtained when 0.4 ml of buffer was used for elution for 30 minutes at room temperature. The filter paper technique is extremely useful for field use and for diagnosis of amebic liver abscess on a large scale since it does not involve centrifugation of blood and use of sterile vials because blood dried on paper can be stored in polythene bags at room temperature for up to 3 months and for a week at 42°C without significant loss of antibody activity, thereby eliminating the need of refrigeration of samples for storage or transportation to a reference laboratory. Elution of blood from different discs of the same patient was found to be highly reproducible without appreciable loss of sensitivity or specificity. Twenty-four confirmed ALA, 29 non-ALA, and 25 apparently normal healthy controls, with no previous history of amebiasis, were tested. Sensitivity and specificity of the test was found to be 96% and 92%, respectively. © 1994 Wiley-Liss, Inc.

Animals can influence their social environment by preferentially associating with certain conspecifics. Such preferential association has gained increasing theoretical attention, as it may influence social evolution and population dynamics. However, relatively little empirical work has examined the occurrence of preferential association and its effects on cooperative group formation. Here, we test the factors associated with cooperative group formation in Polistes dominulus nest-founding queen wasps. P. dominulus are a good system to study preferential association, as foundresses can nest alone or in groups and group membership is flexible. We found that both social and environmental factors were associated with partner choice. First, facial patterns were associated with cooperation. Wasps with more similar facial patterns were more likely to cooperate than wasps with less similar facial patterns. This preferential phenotypic association fits the theoretical criteria for the evolution of tag-based cooperation. Season was also associated with cooperation; wasps on early-season nests were more likely to cooperate than wasps on late-season nests. High levels of aggression by nest owners during initial interactions were also correlated with lower probabilities of subsequent cooperation, suggesting that nest owners have some control over group membership. Other factors including body weight, weight similarity and nest productivity were not linked with cooperation. Overall, multiple factors influence cooperation in paper wasps, including facial pattern similarity. The occurrence of preferential phenotypic association in paper wasps is quite interesting and may influence the evolution of cooperation and population divergence in this group.

Wastes from animal production and from the paper industry are often used as amendments to agricultural soils. Although data exist on the impacts of raw animal wastes on N₂O production, little is known regarding the effects of paper wastes and biosolids from treated animal waste. We measured N₂O emissions for two consecutive snow-free seasons (mid-May through the end of October) from poorly drained clayey soils under corn (Zea mays L.). Soils were amended with raw pig slurry (PS) or biosolids (four PS-derived and two paper sludges) and compared with soils with mineral N fertilizer (CaNH₄NO₃) or without N addition (Control). Area-based N₂O emissions from the mineral N fertilizer (average, 8.2 kg N₂O–N ha⁻¹; 4.2% of applied N) were higher (P < 0.001) than emissions from the organic amendments, which ranged from 1.5 to 6.1 kg N₂O–N ha⁻¹ (−0.4 to 2.5% of applied N). The N₂O emissions were positively correlated with mean soil NO₃ availability (calculated as “NO₃ exposure”), which was highest with mineral N fertilizer. In plots treated with organic amendments (i.e., biosolids and raw PS), NO₃ exposure was negatively correlated to the C:N ratio of the amendment. This resulted in lower N₂O emissions from the higher C:N ratio biosolids, especially compared with the low C:N ratio PS. Application of paper sludge or PS-derived biosolids to these fine-textured soils, therefore, reduced N₂O emissions compared with raw PS and/or mineral N fertilizers (P < 0.01).

The aqueous fraction of the tear film and the horizontal palpebral fissure length (HPFL) were measured in exotic and laboratory animals, specifically saffron finches (Sicalis flaveola), chestnut-bellied seed-finches (Sporophila angolensis), red-eared sliders (Trachemys scripta elegans), rats (Rattus norvegicus) and mice (Mus musculus). These species possess small eyes making it difficult to perform the typical Schirmer tear test. Measurement of the aqueous fraction of the tear was performed using the standardized endodontic absorbent paper point tear test (PPTT), accomplished with manual restraint by a single operator. The following results were obtained: saffron finches (n = 42)-HPFL (4.46 ± 0.09 mm) and PPTT (5.10 ± 0.26 mm); chestnut-bellied seed-finches (n = 38)-HPFL (4.77 ± 0.05 mm) and PPTT (4.11 ± 0.34 mm); red-eared sliders (n = 56)-HPFL (8.59 ± 0.08 mm) and PPTT (8.79 ± 0.38 mm); rats (n = 60)-HPFL (6.45 ± 0.09 mm) and PTT (6.18 ± 2.06 mm); and mice (n = 22)-HPFL (3.59 ± 0.27 mm) and PPTT (4.39 ± 1.45 mm).

Nonspecific binding (NSB), a random adsorption of biocomponents such as proteins and bacteria on noncomplementary materials,isoneofthebiggestproblemsinbiological applications including biosensors, protein chips, surgical instruments, drug delivery, and biomedicine. Polyoxyethylene sorbitan laurate (TWEEN), a commercially available chemical with aliphatic ester chains, has shown promise as a medical material and in overcomingproblems associated withNSB. [1‐4] However,stability during solution-based processing and uniformity of the materials that have TWEEN coating on flat substrates or nanomaterials using the selfassembled-monolayer (SAM) method has been an important issue. Further, biocompatible materials with high strength are important for several medical applications including stents, nail implants, and strong invasive instruments. Here, we present the production of a free-standing ‘‘paperlike’’ material composed of TWEEN and reduced graphene oxide (RGO) platelets and obtained by simple filtration of a homogeneous aqueous colloidal suspension of TWEEN/RGO hybrid. The ‘‘TWEEN paper’’ was highly stable in water without leakage of TWEEN and is compliant and sufficiently robust to be handled by hand without breaking. Furthermore, the TWEEN paper was noncytotoxic to three mammalian cell lines and biocompatible, inhibiting nonspecific binding of Gram-positive bacteria. [5] In contrast, RGO paper without TWEEN showed nonspecific bacterial binding. TWEEN is composed ofthree chemical parts (Fig. 1a): aliphatic esterchains that can prevent NSB ofbiomolecules, three-terminal hydroxyl groups that are hydrophilic and can be chemically modified for further applications, and an aliphatic chain that can easily be adsorbed on a hydrophobic surface by noncovalent interaction. Protein microarrays on flat substrates with SAM of TWEEN [4] and highly sensitive biosensors, [1‐3] built using field-effect transistor (FET) behavior of individual carbon nanotube (CNT) strands coated with TWEEN, have demonstrated that TWEEN can be effectively used to overcome NSB.

Iodine deficiency is an important nutritional deficiency, with more than 2 billion people worldwide estimated to be at risk. The developing fetus and young children are particularly at risk. During pregnancy and lactation, iodine requirements increase, whether in iodine-poor or iodine-sufficient countries, making the mother and the developing fetus vulnerable. The American Thyroid Association (ATA) recommends 250 micrograms per day of iodine intake for pregnant and lactating women. The thyroid gland is able to adapt to the changes associated with pregnancy as long as sufficient iodine is present. Dietary intake is the sole source of iodine, which is essential to the synthesis of thyroid hormones. Iodine is found in multiple dietary sources including iodized salt, dairy products, seaweed, and fish. Prenatal vitamins containing iodine are a good source of iodine, but iodine content in multivitamin supplements is highly variable. Congenital hypothyroidism is associated with cretinism. Clinical hypothyroidism has been associated with increased risk of poor perinatal outcome including prematurity, low birth weight, miscarriage, preeclampsia, fetal death, and impaired fetal neurocognitive development. Subclinical hypothyroidism is also associated with poor pregnancy outcomes and potential fetal neurocognitive deficits, but the data are more variable than those for clinical hypothyroidism. We concur with the ATA recommendation that all pregnant and lactating women should ingest (through diet and supplements) 250 micrograms of iodine daily. To achieve this goal, we recommend that all pregnant and lactating women take daily iodine supplementation of 150 micrograms. Birth Defects Research (Part A) 94:677–682, 2012. 2012 Wiley Periodicals, Inc.

The paper gives a brief introduction to canine oncology, including its comparative aspects as basis for recording tumours in the animal kingdom. In an abbreviated presentation of the Norwegian Canine Cancer Project for the years 1990 – 1998, the data (n=14,401) were divided into age groups, each of two years, into different categories of tumours, and into age and gender. As expected, cutaneous histiocytoma was the dominant tumour type in both sexes during the two first years of life. In the age group 2 – 3.99 years histiocytoma was still the largest group in males, but was surpassed by benign epithelial skin tumours in females. After the age of 4 years, benign epithelial skin tumours constituted the greatest circumscribed group in males, and mammary tumours in females, although the summated other tumours, not explained in this survey, dominated overall in males. Maligancies (cancer) were shown in the same way, by corresponding groups of gender and age. While mastocytoma was the most common tumour and non-Hodgkin's lymphoma the second most common during the two first years of life in females, the situation was reversed in males. Later, mammary tumours dominated in females, while different tumour types not further specified in this summarized report dominated in males, until the end of the age registration (above 14 years). Number, sex and location of most common tumours are shown in a tabular outline. Comparative aspects between human and dog tumours are considered: mammary and testicular neoplasia seemed more frequent in dogs than in humans in Norway, while intestinal, pulmonary and prostatic malignancies were less common in dogs. In our study, vascular tumours and tumour-like lesions constituted about 3% of the total data. As benign vascular tumours are incompletely reported to the human Cancer Registry, no dependable comparison may be made, but malignant vascular tumours have been on the rise during the last decades in the Norwegian human population, more so in men then in women. Finally, the article deals briefly with the development of endothelial cells, and the sparse information on causal factors of vascular tumours.

Four pulp and paper mills discharge their effluents in the same section of the Biobio River in central southern Chile. Pulp mill effluents are a very complex mixture with characteristics that depend on the type of raw material, the process technology, and the effluent treatment. To investigate the effect of pulp mill effluent discharges, immature Oncorhynchus mykiss were exposed to river sediments in the laboratory for 29 d. Three sampling areas were defined in a spatial gradient in the river: Preimpact, impact, and postimpact zones relative to the pulp and paper mill discharge areas. Ethoxyresorufin-O-deethylase activities were significantly higher in fish exposed to impact and postimpact sediments when compared to those exposed to preimpact sediments, and higher levels of vitellogenin were observed in the plasma of female fish exposed to impact and postimpact sediments. Histological analysis of the gonadal tissue showed an induction of gonadal maturation in fish exposed to sediment coming from the impact and postimpact zones (oocytes in a vitellogenic state). No site differences were observed in erythrocytes, although differences were noted in the leukocytes in the exposure areas. Finally, the biomarker approach showed evidence that the sediment associated with pulp mill effluent discharges produces some effects in fish under laboratory conditions.

Resistance to third and fourth generation cephalosporins is rapidly increasing in bacteria infecting humans. Although many of these problems are linked to human to human transmission and to use of antimicrobials in human medicine, the potential role of community reservoirs such as food producing animals needs to be scrutinized. Resistance to third and fourth generation cephalosporins is emerging in enteric bacteria of food producing animals and also in food of animal origin. The genes encoding resistance to these cephalosporins are transferrable and often linked to other resistance genes. Systemic use of third and fourth cephalosporins selects for resistance, but co-selection by other antimicrobials is also likely to influence prevalence of resistance. Although there are many uncertainties, the potential consequences of a further increase of resistance to this critically important class of antimicrobials in bacteria colonising animals are serious. Measures to counter a further increase and spread of resistance among animals should therefore be considered.

Explaining the maintenance of genetic variation in characters associated with Darwinian fitness is a preoccupation of evolutionary biologists. Spatial or temporal variation in the environment can certainly promote polymorphism, yet even populations of 'model organisms', like fruit flies, kept on invariant protocols for hundreds of generations in the laboratory often show fitness variation that exceeds what would be expected from the input of new mutations alone. Such observations suggest either complexities of selection or of genetic architecture, and offer a powerful tool for the study of mechanisms that promote stable polymorphism. In this issue of Molecular Ecology, Zhang et al. (2013) report examples of nontransitivity in the outcome of postcopulatory sexual selection in the fruit fly, Drosophila, that follow the rules of the popular stalemate-breaking game roshambo - or rock, paper, scissors (RPS). The important feature of RPS is that while each strategy beats one other, it in turn is beaten by the third. Using chromosome extraction lines, the authors confirm earlier findings that the outcome of postcopulatory sexual selection via sperm competition for a male depends, in part, upon the competitor male's genotype. But taking it one step further, they demonstrate the nontransitivities between males required for circular RPS cycles in sperm competition between males, and are able to identify at least four associated loci. Because the postmating phenotype involves hundreds of potentially interacting peptides and receptors, this is an important step to understanding the persistence of variation in a critical component of male fitness.

Caging experiments were conducted using hatchery-reared, immature, female rainbow trout (Oncorhynchus mykiss) in three previously defined areas of the Biobio River (south central Chile) representing a pollution gradient from the pulp and paper mill discharges area: a pre-impact area (upstream area, reference location), an impact area (area directly influenced), and a postimpact area (downstream area, less influenced). No significant changes were observed in the physiological index as represented by condition factor (K) and liver somatic index during different sampling times (after 11, 21, and 30 d of exposure). Ethoxyresorufin-O-deethylase activities were significantly higher in trout caged at the impact and postimpact discharges areas (two- to fourfold) compared with the reference (pre-impact) area, and a strong inhibition of acetylcholinesterase activity, reaching 50%, was observed mainly in fish caged at the impact area. A significant endocrine-disrupting effect (reproductive level) was evidenced by significant increments in gonad somatic index and plasma vitellogenin levels combined with an induction of gonad maturation (presence of vitellogenic oocytes) in trout caged at the impact and postimpact areas. These results, generated by an in situ approach, confirmed our group's findings for trout exposed to sediment in the laboratory: discharges of pulp mill effluent in the Biobio River are associated with the effects evaluated at different biological levels.

It has been proposed that somatic mutations make major contributions to aging. The first paper, based on a gene knock-in mouse, supports a contributory role for mutation in mtDNA1 in aging, but does not support a damaged-mtDNA-producing-more-damaged-mtDNA hypothesis.1 The second paper2 indicates some GC-rich sequences in the nuclear DNA are more sensitive to oxidative damage than mtDNA. As a result, key genes involved in brain function and mitochondrial function are progressively inactivated with age. Failure in these nucleus-encoded mitochondrial genes may be a primary reason for mitochondrial failure in old age. BioEssays 26:1160–1163, 2004. © 2004 Wiley Periodicals, Inc.

Carotenoid-depleted fruit flies, Drosophila melanogaster, were reared on yeast/glucose medium containing lipid-depleted white corn grits and cholesterol. After rearing for more than a year, the yield of flies remained constant and the content of 3-hydroxyretinal in a head was three logarithmic units less than that of normal flies reared on medium containing yellow corn grits. When all-trans retinal was supplied as the sole source of retinoids, the flies formed and accumulated all-trans 3-hydroxyretinal in the dark. To examine the metabolic pathway to produce (3S)-3-hydroxyretinal in Drosophila, all-trans retinal was supplemented for two hours to carotenoid-depleted flies in the dark, and the subsequent changes in the composition of 3-hydroxyretinal enantiomers were analyzed using a chiral column on HPLC. The results indicated initial formation of (3R)-3-hydroxyretinal followed by isomerization into the 3S enantiomer. In another set of experiments, the membrane fraction was obtained from the head homogenate of retinoid-depleted flies and an in vitro assay of 3-hydroxyretinal formation from retinal was performed. The 3-hydroxyretinal produced was the 3R enantiomer, supporting the result obtained from the in vivo experiment whereby (3S)-3-hydroxyretinal is produced from retinal via (3R)-3-hydroxyretinal. Addition of NADPH enhanced 3-hydroxyretinal formation and the presence of carbon monoxide inhibited it, suggesting that hydroxylation at the C3 position of retinal occurred via the monooxygenase activity of cytochrome P-450.