Name: a method for quantifying foliage-dwelling arthropods
Uploaded: 2019-10-20T13:30:00
Description: Watch this Scientific Journal Video about A Method for Quantifying Foliage-Dwelling Arthropods at JoVE.com

The authors would like to thank the U.S. Department of Agriculture Forest Service for funding this project through USFS Agreement 13-CS-11090800-022. We would like to thank J. Suda, W. Holland, and others for laboratory assistance, and R. Richards for field assistance.

1. Building the sampling device prior to going to the field
<ol>
	<li>Using bolt cutters, large wire cutters, or an electric grinding disk, remove the bottom 1/3 of the 30 cm wire tomato cage so that it is approximately 55 cm in length.</li>
	<li>Cut two, 50 cm braces made from aluminum or similarly semi-rigid material to use as attachment rods and braces on each side of the largest end of the tomato cage. At 38 cm from the end, use a table-top vice or large grasping tool such as channel locks to bend the brace to an a...

Two necessities of accurately quantifying arthropod communities are relatively high detection probabilities and known or consistent sampling areas. When sampling for arthropods, less than 100% detection probability can be attributed to either individual arthropods avoiding traps or some individuals that were trapped being undetected during processing. Interceptor traps that intercept flying arthropods (Malaise/window traps, sticky traps, etc.) appear to be the most frequently used approach to enumerate arthropod communit...

Terrestrial arthropods play an important role in our ecosystem. In addition to being of scientific interest arthropods can be both detrimental and beneficial to crops, horticultural plants, and natural vegetation as well as provide an important trophic function in food webs. Thus, understanding the factors that influence arthropod community development and abundance is critical to farmers, pest control managers, plant biologists, entomologists, wildlife ecologists, and conservation biologists that study community dynamics and manage insectivorous organisms. Understanding factors that influence arthropod communities and abundances often requires the capture of individuals. Capture techniques can generally be categorized into qualitative techniques that only detect presence of a species for estimates of species range, richness, and diversity, or semi-quantitative and quantitative techniques that allow for an index or estimate of abundance and density of individuals within a taxonomic group.
Qualitative techniques that only allow inference regarding presence of a species or community structure have an unknown or intrinsically low detection probability or are lacking in providing inference regarding detection probability and size of area sampled. Because detection probability with these techniques is low, variability associated with detection precludes adequate precision for inferring how explanatory variables influence arthropod population metrics. Qualitative techniques used to estimate presence include suction sampling1, light traps2, emergence traps3, feeding patterns on roots4, brine pipes5, baits6, pheromone3, pitfall traps7, Malaise traps8, window traps9, suction traps10, beating trays11, spider webs12, leaf mines, frass13, arthropod galls14, vegetation and root damage15.
Alternatively, semi-quantitative and quantitative techniques allow researchers to estimate or at least consistently sample a specified sample area and estimate probability of detection or assume detection probability is non-directional and adequate as to not obscure the researcher&#39;s ability to detect spatial or temporal variation in abundance. Semi-quantitative and quantitative techniques include sweep nets16, suction or vacuum sampling17, systematic counting of visible arthropods18, sticky traps19, various pot-type traps20, entrance or emergent holes21, chemical knockdown22, sticky and water filled color traps23, and branch bagging and clipping24.
Recent anthropogenic-induced changes to climate and disturbance regimes have led to dramatic changes in plant communities, making interactions between plant-community species composition and arthropod communities an active area of study. Understanding how arthropod communities vary with plant species composition is a critical component for understanding the potential economic and environmental impacts of changes to plant communities. Semi-quantitative or quantitative methods of quantifying arthropod abundance with adequate precision to detect differences among species of plants are needed. In this article, we describe a method for indexing foliage-dwelling arthropods that, with reasonable effort, provided adequate precision to identify differences in individual abundance and biomass, diversity, and richness among 5 taxa of trees commonly found in the southeastern deciduous forests of North America25. This approach provided precision adequate for estimating abundance to allow inference as to how changes in species composition of forest plant communities due to anthropic modified disturbance regimes influence composition of arthropods, potentially influencing abundance and distribution of higher trophic insectivorous birds and mammals. More specifically, by using a modified bagging technique first described by Crossley et al.24, we estimated density of surface, foliage-dwelling arthropods and tested the prediction that we would detect differences in diversity, richness, and abundance of arthropods in the foliage of faster growing more xeric species of trees relative to slower growing more mesic species. The goal of this article is to provide detailed instructions of the technique.
We conducted the study on the Shawnee National Forest (SNF) in southern Illinois. The SNF is a 115,738-ha forest located in the Central Hardwoods region of the Ozarks and Shawnee Hills natural divisions26. The forest comprises a mosaic of 37% oak/hickory, 25% mixed-upland hardwoods, 16% beech/maple, and 10% bottomland hardwoods. The SNF is dominated by second growth oak/hickory in upland xeric areas and sugar maple, American beech, and tulip tree (Liriodendron tulipifera) in sheltered mesic valleys27,28.
Site selection for this method will be dependent on the overarching goals of the study. For example, the primarily goal of our original study was to provide insight into how changes in tree community might influence higher trophic organisms by comparing foliage-dwelling arthropod community metrics between mesic and xeric adapted tree communities. Thus, our primary objective was to quantify the arthropod community on individual trees located within the xeric or mesic tree community. We selected 22 study sites along an oak/hickory (xeric) to beech/maple (mesic) dominated gradient using USFS stand cover maps (allveg2008.shp) in ArcGIS 10.1.1. To prevent potential confounding effects, we selected sites using the following criteria: not located in riparian areas, &#8805;12 ha, and located within contiguous upland-deciduous forest habitat (i.e., elevation above 120 m). All sites contained mature trees &#62;50 years old in hilly terrain, thus comprised similar slopes and aspects. While beech/maple site boundaries were distinguished based on the transition of tree communities, oak/hickory site boundaries were identified artificially using SNF cover maps and ArcGIS 10.1.1. All sites were large forest blocks within un-glaciated terrain; their differences in tree species composition were not due to differences in location on the landscape but were representative of past land usage (e.g., clear cuts or selective harvest). We ground-truthed the maps by uploading discrete polygon shapefiles of each study site to a handheld Global Positioning System (GPS) and verifying tree species composition. We randomly selected sampling points (n = 5) at each site. At each point, we sampled three trees from 0600&#8722;1400 hours during 23 May to 25 June 2014. To locate sample trees, we searched outward to a 30 m radius from vegetation points until mature trees (&#62;20 cm d.b.h.) with branches low enough to sample were found. Typically, the three mature trees that represented three of the five genera (Acer, Carya, Fagus, Liriodendron, and Quercus) of interest and were closest to the center point were sampled.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>13 gallon garbage bags</td>
			<td>Glad</td>
			<td>78374</td>
			<td></td>
		</tr>
		<tr>
			<td>Aluminum rod</td>
			<td>Grainger</td>
			<td>48ku20</td>
			<td></td>
		</tr>
		<tr>
			<td>Pruner</td>
			<td>Bartlet arborist supply</td>
			<td>pp-125b-2stick</td>
			<td></td>
		</tr>
		<tr>
			<td>Telescoping pole</td>
			<td>BES</td>
			<td>TPF620</td>
			<td></td>
		</tr>
		<tr>
			<td>Tomato Cage</td>
			<td>Gilbert and Bennet</td>
			<td>42 inch galvanized</td>
			<td></td>
		</tr>
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</table>

a method for quantifying foliage-dwelling arthropods

Terrestrial arthropods play an important role in our environment. Quantifying arthropods in a way that allows for a precise index or estimates of density requires a method with high detection probability and a known sampling area. While most described methods provide a qualitative or semi-quantitative estimate adequate for describing species presence, richness, and diversity, few provide an adequately consistent detection probability and known or consistent sampling areas to provide an index or estimate with adequate precision to detect differences in abundance across environmental, spatial, or temporal variables. We describe how to quantify leaf-dwelling arthropods by sealing the leaves and end of branches in a bag, clipping and freezing the bagged material, and rinsing the previously frozen material in water to separate arthropods from the substrate and quantify them. As we demonstrate, this method can be used at a landscape scale to quantify leaf-dwelling arthropods with adequate precision to test for and describe how spatial, temporal, environmental, and ecological variables influence arthropod richness and abundance. This method allowed us to detect differences in density, richness, and diversity of leaf-dwelling arthropods among 5 genera of trees commonly found in southeastern deciduous forests.

We collected 626 samples from 323 individual trees composing 5 tree groups. For estimates of total arthropod biomass per meter of branch sampled, the standard error ranged from 12% to 18% of the mean for the 5 tree groups (Table 1). This level of precision was adequate to detect variation among tree groups and a quadratic change in biomass with date25. This technique provided more precision when estimating guild diversity as demonstrated by the sta...

Watch this Scientific Journal Video about A Method for Quantifying Foliage-Dwelling Arthropods at JoVE.com

A Method for Quantifying Foliage-Dwelling Arthropods

We describe how to quantify leaf dwelling arthropods by sealing the leaves and end of branches in a bag, clipping and freezing the bagged material, and rinsing the previously frozen material in water to separate arthropods from the substrate for quantification.

Terrestrial arthropods play an important role in our environment. Quantifying arthropods in a way that allows for a precise index or estimates of density ...

The main advantage of this protocol is it allows researchers to capture both flying and crawling arthropods from the foliage. Although we use this technique on trees it can also be used on bushes and shrubs. Be careful to practice bagging branches quickly without disturbing the foliage so active arthropods do not escape capture.To begin this procedure, use bolt cutters, large wire cutters, or an electric grinding disc to remove the bottom third of a 30 centimeter wire tomato cage so that it is approximately 55 centimeters in length. Then, cut two 50 centimeter braces from aluminum tubing or a similarly semi-rigid material to use as attachment rods and braces on each side of the largest end of the tomato cage. At 38 centimeters from the end use a tabletop vice or larger grasping tools such as channel locks to bend the brace to an approximately 30 degree angle.Attach the long end of each attachment rod to opposite ends of the tomato cage with zip ties. Use either duct tape or electrical tape to support this attachment making sure that the tape is wrapped around at least six centimeters of the cage and rod. Attach the other end of each rod on opposites sides of the end of an extendable pole with zip ties.Use either duct tape or electrical tape to support this attachment making sure to wrap the tape around multiple times and that it overlaps the pole and rods by at least six centimeters. Make sure that the opening of the cage is contact with the end of the telescoping pole when the cage is attached. Then attach the cage directly onto the end of the pole using tape.Attach hook and loop fastener strips at two points to the opening of the cage 90 degrees from the previously attached pole. First, attach two pieces of hook and loop fastener to the outside opening of the bag so that they align with the hook and loop fastener attached to the opening of the cage. Be certain that the hook and loop fastener is aligned so when the bag is inserted and attached the opening to the pull strings of the bag run parallel to the telescoping pole.Insert a 49 liter kitchen garbage bag into the wire tomato cage. Place three gator clips evenly dispersed on each respective side of the bottom of the bag and attach the clips to both the bag and the wire cage so that the bag is held against the cage. Orient the gator clip on the bag opening so that it is attached to the drawstring parallel to the telescoping pole opposite of the location of the pole attachment.Attach paracord to the bag's drawstring closest to the pole. Cut four pieces of plastic or hard rubber tubing in four centimeter sections and attach them at four locations with duct tape or electrical tape. First should be placed on the extending part of the pole about 0.5 meters from the end of the pole nearest the tomato cage.The remaining three should be placed equidistance along the bottom section of extending pole starting about five centimeters from the top of the bottom section. Thread the end of the paracord that is not attached to the bag through the plastic tubing. After this run both paracord strings together through the plastic tubing inserts to hold it in place.For each sample tree, use a random number generator to select a sample height that is within the height of the extension pole when extended at maximum length. Use the random number generator to select a sample distance from the tree trunk and identify a branch that will fit in the bag with minimal disturbance to the foliage. And is the proper height and distance from the trunk based on the numbers generated.Raise the sampling pole to a height parallel with the desired branch. Quickly slide the bag over the branch and then quickly pull the paracord strings that are attached to the drawstrings to seal the bag. Have a second person use the extension pole pruner to clip the branch at the location adjacent to the bag's opening.Then carefully bring the sample bag to the ground and rapidly tie the bag's drawstrings closed. Store the bagged branch in a freezer until ready to conduct the laboratory arthropod analysis. Hold the frozen bag and branch upright and shake the sample branch while it is in the bag to dislodge the arthropods into the bag.Carefully remove the branch from the bag and rinse it in large collection pan to remove any remaining arthropods. Next, empty the material remaining in the bag into the collection pan and remove an non-arthropod debris. Separate the arthropods into the desired taxonomic groups.Noting differences between larvae and adults. Quantify the arthropods as desired. If biomass is of interest either measure the length of arthropods and use published length mass table to estimate biomass.Replace arthropods in small drying pans. Dry in drying oven for 24 hours at 45 degrees Celsius and weigh on an electronic balance. Dry the leaves in a drying oven for 48 hours at 45 degrees Celsius and weigh them on an electronic balance.Or measure the length of all woody branches within the sample. In a representative study, 626 samples are collected from 323 individual trees that composed five tree groups. For estimates of total arthropod biomass per meter of branch sampled the standard error ranges from 12%to 18%of the mean for the five tree groups.This level of precision is adequate to detect variation among tree groups in a quadratic change in biomass by date. This technique provides more precision when estimating guild diversity as demonstrated by the standard error of arthropod guild diversity ranging from 3%to 7%of the mean diversity across the five tree groups. Precision at this level is adequate to detect variation across the five tree groups.Precision of estimates of richness are also very precise as demonstrated by standard errors that ranged from 3%to 7%of the mean richness among the five tree groups. This level of precision is adequate to identify variation among tree groups. A quadratic association with date, decrease in richness with height on the tree, and a positive relationship between arthropod richness and distance from the tree trunk.Be certain that you enclose the branch quickly without disturbing the foliage to ensure active arthropods do not escape capture.

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A Simple Method for Automated Solid Phase Extraction of Water Samples for Immunological Analysis of Small Pollutants

A new method for solid phase extraction (SPE) of environmental water samples is proposed. The developed prototype is cost-efficient and user friendly, and enables to perform rapid, automated and simple SPE. The pre-concentrated solution is compatible with analysis by immunoassay, with a low organic solvent content. A method is described for the extraction and pre-concentration of natural hormone 17&#946;-estradiol in 100 ml water samples. Reverse phase SPE is performed with octadecyl-silica sorbent and elution is done with 200 &#181;l of methanol 50% v/v. Eluent is diluted by adding di-water to lower the amount of methanol. After preparing manually the SPE column, the overall procedure is performed automatically within 1 hr. At the end of the process, estradiol concentration is measured by using a commercial enzyme-linked immune-sorbent assay (ELISA). 100-fold pre-concentration is achieved and the methanol content in only 10% v/v. Full recoveries of the molecule are achieved with 1 ng/L spiked de-ionized and synthetic sea water samples.

A protocol for the extraction and pre-concentration of estradiol from water samples by using an automated and miniaturized system is presented.

A new method for solid phase extraction (SPE) of environmental water samples is proposed. The developed prototype is cost-efficient and user friendly, and ...

Administering and Detecting Protein Marks on Arthropods for Dispersal Research

Monitoring arthropod movement is often required to better understand associated population dynamics, dispersal patterns, host plant preferences, and other ecological interactions. Arthropods are usually tracked in nature by tagging them with a unique mark and then re-collecting them over time and space to determine their dispersal capabilities. In addition to actual physical tags, such as colored dust or paint, various types of proteins have proven very effective for marking arthropods for ecological research. Proteins can be administered internally and/or externally. The proteins can then be detected on recaptured arthropods with a protein-specific enzyme-linked immunosorbent assay (ELISA). Here we describe protocols for externally and internally tagging arthropods with protein. Two simple experimental examples are demonstrated: (1) an internal protein mark introduced to an insect by providing a protein-enriched diet and (2) an external protein mark topically applied to an insect using a medical nebulizer. We then relate a step-by-step guide of the sandwich and indirect ELISA methods used to detect protein marks on the insects. In this demonstration, various aspects of the acquisition and detection of protein markers on arthropods for mark-release-recapture, mark-capture, and self-mark-capture types of research are discussed, along with the various ways that the immunomarking procedure has been adapted to suit a wide variety of research objectives.

Proteins are often used to mark arthropods for dispersal research. Methods for administering internal and external protein marks on arthropods are demonstrated. Protein mark detection techniques, either through indirect or sandwich enzyme-linked immunosorbent assays (ELISAs), are also illustrated.

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Alternative Method of Removing Otoliths from Sturgeon

Extracting the otoliths (ear bones) from fish that have very thick skulls can be difficult and very time consuming. The common practice of making a transverse vertical incision on the top of the skull with a hand or electrical saw may damage the otolith if not performed correctly. Sturgeons (Acipenseridae) are one family in particular that have a very large and thick skull. A new laboratory method entering the brain cavity from the ventral side of the fish to expose the otoliths was easier than other otolith extraction methods found in the literature. Methods reviewed in the literature are designed for the field and are more efficient at processing large quantities of fish quickly. However, this new technique was designed to be more suited for a laboratory setting when time is not pressing and successful extraction from each specimen is critical. The success of finding and removing otoliths using this technique is very high and does not compromise the structure in any manner. This alternative technique is applicable to other similar fish species for extracting the otoliths.

The goal of the protocol is to show an effective method to extract the otoliths from sturgeon carcasses.

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Modification and Application of a Leaf Blower-vac for Field Sampling of Arthropods

Rice fields host a large diversity of arthropods, but investigating their population dynamics and interactions is challenging. Here we describe the modification and application of a leaf blower-vac for suction sampling of arthropod populations in rice. When used in combination with an enclosure, application of this sampling device provides absolute estimates of the populations of arthropods as numbers per standardized sampling area. The sampling efficiency depends critically on the sampling duration. In a mature rice crop, a two-minute sampling in an enclosure of 0.13 m2 yields more than 90% of the arthropod population. The device also allows sampling of arthropods dwelling on the water surface or the soil in rice paddies, but it is not suitable for sampling fast flying insects, such as predatory Odonata or larger hymenopterous parasitoids. The modified blower-vac is simple to construct, and cheaper and easier to handle than traditional suction sampling devices, such as D-vac. The low cost makes the modified blower-vac also accessible to researchers in developing countries.

Assessment of arthropod abundance in crops is critical for investigating population dynamics and species interactions. Here we describe the modification and application of a leaf blower-vac for suction sampling of arthropods in rice.

Rice fields host a large diversity of arthropods, but investigating their population dynamics and interactions is challenging. Here we describe the ...

Reliable Method for Assessing Seed Germination, Dormancy, and Mortality under Field Conditions

We describe techniques for approximating seed bank dynamics over time using Helianthus annuus as an example study species. Strips of permeable polyester fabric and glue can be folded and glued to construct a strip of compartments that house seeds and identifying information, while allowing contact with soil leachate, water, microorganisms, and ambient temperature. Strips may be constructed with a wide range of compartment numbers and sizes and allow the researcher to house a variety of genotypes within a single species, different species, or seeds that have experienced different treatments. As opposed to individual seed packets, strips are more easily retrieved as a unit. While replicate packets can be included within a strip, different strips can act as blocks or can be retrieved at different times for observation of seed behavior over time. We used a high temperature glue gun to delineate compartments and sealed the strips once the seed and tags identifying block and removal times were inserted. The seed strips were then buried in the field at the desired depth, with the location marked for later removal. Burrowing animal predators were effectively excluded by use of a covering of metal mesh hardware cloth on the soil surface. After the selected time interval for burial, strips were dug up and seeds were assessed for germination, dormancy and mortality. While clearly dead seeds can often be distinguished from ungerminated living ones by eye, dormant seeds were conclusively identified using a standard Tetrazolium chloride colorimetric test for seed viability.

Here we present a protocol for assessing seed survivorship, germination and dormancy under field conditions using buried, labeled seed strips and tetrazolium chloride (TZ) viability testing.

We describe techniques for approximating seed bank dynamics over time using Helianthus annuus as an example study species. Strips of permeable polyester ...

Protocols for Quantifying Transferable Pesticide Residues in Turfgrass Systems

Plant canopies in established turfgrass systems can intercept an appreciable amount of sprayed pesticides, which can be transferred through various routes onto humans. For this reason, transferable pesticide residue experiments are required for registration and re-registration by the United States Environmental Protection Agency (USEPA). Although such experiments are required, limited specificity is required pertaining to experimental approach. Experimental approaches used to assess pesticide transfer to humans including hand wiping with cotton gloves, modified California roller (moving a roller of known mass over cotton cloth) and soccer ball roll (ball wrapped with sorbent strip) over three treated turfgrass species (creeping bentgrass, hybrid bermudagrass and tall fescue maintained at 0.4, 5 and 9 cm, respectively) are presented. The modified California roller is the most extensively utilized approach to date, and is best suited for use at low mowing heights due to its reproducibility and large sampling area. The soccer ball roll is a less aggressive transfer approach; however, it mimics a very common occurrence in the most popular international sport, and has many implications for nondietary pesticide exposure from hand-to-mouth contact. Additionally, this approach may be adjusted for other athletic activities with limited modification. Hand wiping is the best approach to transfer pesticides at higher mowing heights, as roller-based approaches can lay blades over; however, it is more subjective due to more variable sampling pressure. Utility of these methods across turfgrass species is presented, and additional considerations to conduct transferable pesticide residue research in turfgrass systems are discussed.

Transferable pesticide residue experiments in turfgrass systems are integral components of human risk exposure assessments. Experimental approaches to measure transferable residues should be adjusted to the human interaction of interest and turfgrass system dynamics. Three transferable pesticide residue protocols are presented and the suitability across three turfgrass systems is discussed.

Plant canopies in established turfgrass systems can intercept an appreciable amount of sprayed pesticides, which can be transferred through various routes ...

Development of New Methods for Quantifying Fish Density Using Underwater Stereo-video Tools

The use of video camera systems in ecological studies of fish continues to gain traction as a viable, non-extractive method of measuring fish lengths and estimating fish abundance. We developed and implemented a rotating stereo-video camera tool that covers a full 360 degrees of sampling, which maximizes sampling effort compared to stationary camera tools. A variety of studies have detailed the ability of static, stereo-camera systems to obtain highly accurate and precise measurements of fish; the focus here was on the development of methodological approaches to quantify fish density using rotating camera systems. The first approach was to develop a modification of the metric MaxN, which typically is a conservative count of the minimum number of fish observed on a given camera survey. We redefine MaxN to be the maximum number of fish observed in any given rotation of the camera system. When precautions are taken to avoid double counting, this method for MaxN may more accurately reflect true abundance than that obtained from a fixed camera. Secondly, because stereo-video allows fish to be mapped in three-dimensional space, precise estimates of the distance-from-camera can be obtained for each fish. By using the 95% percentile of the observed distance from camera to establish species-specific areas surveyed, we account for differences in detectability among species while avoiding diluting density estimates by using the maximum distance a species was observed. Accounting for this range of detectability is critical to accurately estimate fish abundances. This methodology will facilitate the integration of rotating stereo-video tools in both applied science and management contexts.

We describe a new method for counting fishes, and estimating relative abundance (MaxN) and fish density using rotating stereo-video camera systems. We also demonstrate how to use distance from camera (Z distance) to estimate species-specific detectability.

The use of video camera systems in ecological studies of fish continues to gain traction as a viable, non-extractive method of measuring fish lengths and ...

A Novel Method for the Pentosan Analysis Present in Jute Biomass and Its Conversion into Sugar Monomers Using Acidic Ionic Liquid

Recently, ionic liquids (ILs) are used for biomass valorization into valuable chemicals because of their remarkable properties such as thermal stability, lower vapor pressure, non-flammability, higher heat capacity, and tunable solubility and acidity. Here, we demonstrate a method for the synthesis of C5 sugars (xylose and arabinose) from the pentosan present in jute biomass in a one-pot process by utilizing a catalytic amount of Br&#248;nsted acidic 1-methyl-3-(3-sulfopropyl)-imidazolium hydrogen sulfate IL. The acidic IL is synthesized in the lab and characterized using NMR spectroscopic techniques for understanding its purity. The various properties of BAIL are measured such as acid strength, thermal and hydrothermal stability, which showed that the catalyst is stable at a higher temperature (250 &#176;C) and possesses very high acid strength (Ho 1.57). The acidic IL converts over 90% of pentosan into sugars and furfural. Hence, the presenting method in this study can also be employed for the evaluation of pentosan concentration in other kinds of lignocellulosic biomass.

We present a protocol for the synthesis of C5 sugars (xylose and arabinose) from a renewable non-edible lignocellulosic biomass (i.e., jute) with the presence of Br&#248;nsted acidic ionic liquids (BAILs) as the catalyst in water. The BAILs catalyst exhibited better catalytic performance than conventional mineral acid catalysts (H2SO4 and HCl).

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Elemental data are commonly used to infer plant quality as a resource to herbivores. However, the ubiquity of carbon in biomolecules, the presence of nitrogen-containing plant defensive compounds, and variation in species-specific correlations between nitrogen and plant protein content all limit the accuracy of these inferences. Additionally, research focused on plant and/or herbivore physiology require a level of accuracy that is not achieved using generalized correlations. The methods presented here offer researchers a clear and rapid protocol for directly measuring plant soluble proteins and digestible carbohydrates, the two plant macronutrients most closely tied to animal physiological performance. The protocols combine well characterized colorimetric assays with optimized plant-specific digestion steps to provide precise and reproducible results. Our analyses of different sweet corn tissues show that these assays have the sensitivity to detect variation in plant soluble protein and digestible carbohydrate content across multiple spatial scales. These include between-plant differences across growing regions and plant species or varieties, as well as within-plant differences in tissue type and even positional differences within the same tissue. Combining soluble protein and digestible carbohydrate content with elemental data also has the potential to provide new opportunities in plant biology to connect plant mineral nutrition with plant physiological processes. These analyses also help generate the soluble protein and digestible carbohydrate data needed to study nutritional ecology, plant-herbivore interactions and food-web dynamics, which will in turn enhance physiology and ecological research.

Quantifying Plant Soluble Protein and Digestible Carbohydrate Content, Using Corn Zea mays As an Exemplar

The protocols described herein provide a clear and approachable methodology for measuring soluble protein and digestible (non-structural) carbohydrate content in plant tissues. The ability to quantify these two plant macronutrients has significant implications for advancing the fields of plant physiology, nutritional ecology, plant-herbivore interactions and food-web ecology.

Elemental data are commonly used to infer plant quality as a resource to herbivores. However, the ubiquity of carbon in biomolecules, the presence of ...

Design and Use of an Apparatus for Quantifying Bivalve Suspension Feeding at Sea

As shellfish aquaculture moves from coastal embayments and estuaries to offshore locations, the need to quantify ecosystem interactions of farmed bivalves (i.e., mussels, oysters, and clams) presents new challenges. Quantitative data on the feeding behavior of suspension-feeding mollusks is necessary to determine important ecosystem interactions of offshore shellfish farms, including their carrying capacity, the competition with the zooplankton community, the availability of trophic resources at different depths, and the deposition to the benthos. The biodeposition method is used to quantify feeding variables in suspension-feeding bivalves in a natural setting and represents a more realistic proxy than laboratory experiments. This method, however, relies upon a stable platform to satisfy the requirements that water flow rates supplied to the shellfish remain constant and the bivalves are undisturbed. A flow-through device and process for using the biodeposition method to quantify the feeding of bivalve mollusks were modified from a land-based format for shipboard use by building a two-dimensional gimbal table around the device. Planimeter data reveal a minimal pitch and yaw of the chambers containing the test shellfish despite boat motion, the flow rates within the chambers remain constant, and operators are able to collect the biodeposits (feces and pseudofeces) with sufficient consistency to obtain accurate measurements of the bivalve clearance, filtration, selection, ingestion, rejection, and absorption at offshore shellfish aquaculture sites.

A flow-through device for using the biodeposition method to quantify filtration and feeding behavior of bivalve mollusks was modified for shipboard use. A two-dimensional gimbal table built around the device isolates the apparatus from boat motion, thereby allowing the accurate quantification of bivalve filtration variables at offshore shellfish aquaculture sites.

As shellfish aquaculture moves from coastal embayments and estuaries to offshore locations, the need to quantify ecosystem interactions of farmed bivalves ...