Name: Video: Author Spotlight: Assessing the Olfactory Effects of Airborne Pollutants — Buried Food and Social Odor Tests
Uploaded: 2024-09-13T13:40:00.000+00:00
Duration: 4 min
Description: 605 Views. Qingdao University. This paper provides a detailed description of the buried food test and social odor discrimination experiment to assess the effects of inhaled environmental pollutants exposure on olfactory function in mice.

This work was financially supported by the National Natural Science Foundation of China (82204088, 82273669) and the Natural Science Foundation of Shandong Province, China (ZR2021QH209).

We used male C57BL/6J mice (age: 6-8 weeks; weight: 20-22 g) for all behavioral tests. The mice were subjected to steady conditions (i.e., temperature, 23 &#177; 1 &#176;C; humidity, 55% &#177; 5%, and 12/12 h light-dark cycle with lights on at 7:00). All behavioral tests were performed between 10:00 and 17:00. All animal experiments were approved by the Ethics Committee of the Professional Committee of Animal Experiments of Qingdao University. After a 1-week acclimation period, all mice were exposed to inhalable environmental pollutants.
1. Exposure to pollutants
<ol>
	<li>Intranasal administration by instillation
	<ol>
		<li>Dissolve inhalable environmental pollutants in a 0.9% saline solution. Use 5% zinc sulfate solution once, which has been demonstrated to cause olfactory dysfunction17. For control mice, administer 0.9% normal saline solution.</li>
		<li>Anesthetize the mouse by intraperitoneal injection of 1% sodium pentobarbital (57 mg/kg)18 and assess the depth of anesthesia using the toe-pinch reflex. Apply vet ointment to mouse&#39;s eyes to prevent dryness. Position the mouse on its back on an inclined surface, with the head facing downward.</li>
		<li>Using a pipetting gun, administer 10 &#181;L of the solution in one nostril of the mouse.&#160;Suspended the pipette tip above the nose of the mouse, so that the liquid drips onto the nose,&#160;allowing the mouse to naturally inhale the solution into the nasal cavity.</li>
		<li>To ensure the well-being of the mouse and prevent any potential discomfort, repeat the inhalation for the other nostril after a 10 min interval.</li>
		<li>At 3 days after intranasal administration by instillation in mice, perform the buried food test.</li>
	</ol>
	</li>
</ol>
2.&#160;Buried food test
<ol>
	<li>Carry out food deprivation at 18-24 h before the test by removing all chow pellets from the food hopper of the home cage. Change the bedding materials for mice. Do not remove the water bottle.</li>
	<li>Arrange the operating table as described below.
	<ol>
		<li>At 1 h before the start of the test, take the cage containing the mice to the operating room to rest.</li>
		<li>Arrange the operating room during this period. Use and mark transparent PVC standard mice cages as A as these make a familiar environment. Use and mark the test cage as B, which is a transparent PVC standard squirrel cage. Mark the common cage used to place the mice after the experiment as cage C.</li>
		<li>Cover cages A and B with 3 cm of bedding material and measure them with a ruler. Arrange the cages side by side, with a distance of 0.5 m between each cage (Figure 1).</li>
		<li>Define the experimental area as an area with a radius of 2 m, with the center being the center of the cages. Define the area outside the range of 2 m as the observation area.</li>
		<li>Keep cage C and cages containing untested mice as far away from the experimental area as possible.</li>
	</ol>
	</li>
	<li>Record the time to find food as described below.
	<ol>
		<li>Select a position at random in cage B, bury the food 1 cm below the surface of the bedding, and smooth the surface of the bedding.</li>
		<li>Put the mouse into cage A for 4 min. Transfer the mice to cage B at the end of the timing, turn on the video device, and return to the observation area.</li>
		<li>When the mouse picks up the food block with its forepaw, stop the video recording. In some cases, mice can be seen eating with their heads bent over the food. This behavior also indicates trial success, even if the mouse is not holding the food with its forepaw.</li>
		<li>Record data. Record the time from contact with the mat at the bottom of cage B until food was found for each mouse. If the mice do not find food after 4 min, record the find time as a delay time of 240 s.</li>
		<li>Put the mouse in cage C after the test.</li>
		<li>Remove the feed from cage B and put it in a sealed bag. Replace the food in cage B and test the next mouse after changing the bedding material, as described below.
		<ol>
			<li>For mice housed in the same cage, test with the same set of bedding materials in cage B. For mice in different cages, clean the cage with alcohol and replace the bedding materials.</li>
		</ol>
		</li>
		<li>Add feed and water to mice after the experiment. Perform the social odor discrimination experiment 1 day later. 
		NOTE: Masks need to be worn throughout the process, and transparent gloves need to be changed after each mouse finishes the experiment to avoid odor cross as much as possible. Keep the amplitude of the movements as small as possible to avoid mouse stress.</li>
	</ol>
	</li>
</ol>
3. Social odor discrimination experiment
<ol>
	<li>Urine collection
	<ol>
		<li>Separately collect the urine of sexually mature male mice and female mice and aliquot into tubes in equal volumes19. Pack in 2 mL microcentrifuge tubes with 300 &#181;L of urine in each tube.</li>
		<li>Store the samples at -80 &#176; C until use. Shake tubes to evenly distribute the sample after thawing. Do not thaw and freeze the sample repeatedly.</li>
	</ol>
	</li>
	<li>Arrange the operating table as described below.
	<ol>
		<li>At 1 h before the start of the test, take the cage containing the mice to the operating room to rest. Allow the urine&#160;sample to reach room temperature.</li>
		<li>Arrange the operating room during this period as described in steps 2.2.2-2.2.5.</li>
	</ol>
	</li>
	<li>Record the time to find urine as described below.
	<ol>
		<li>Make a groove with tape around each of the two wide sides of cage B large enough to hold the microcentrifuge tube containing 300 &#181;L of male and female urine. Place the tubes and keep the two tube covers closed for now.</li>
		<li>Put the mouse in cage A and set a countdown for 4 min. Open the two tubes of cage B at the end of the timing.</li>
		<li>Transfer the mouse to the middle position of cage B, at the same distance from the two tubes, then turn on the video recording equipment and gently and slowly retreat to the observation area.</li>
		<li>When the mouse sniffs the tube wall/mouth or even inside the tube, the tube is successfully smelled. At this time, press the stopwatch and record the time as the time of smelling the tube (M s), then continue to record the residence time (X s) until the mouse moves away from the tube.</li>
		<li>Continue timing the time sniffing the other tube. When the mouse sniffs the tube wall/mouth or even inside the tube, the tube is successfully smelled. Press the stopwatch and record the time it took to smell the other tube (N s), then continue to record the residence time (Y s) until the mouse moves away from the tube.</li>
		<li>Transfer the mouse to cage C at the end of the experiment.</li>
		<li>Change the urine tubes and bedding material of cage B after each mouse test. Put the used mouse urine in a clean bag according to sex.</li>
		<li>Change gloves and test the next mouse as described above.</li>
	</ol>
	</li>
	<li>Perform the experiment in a laboratory without special odor, such as strong fragrances or the smell of food, if there are, thoroughly ventilate before experimenting. Avoid personal products that emit strong odors. Wear gloves and masks throughout the procedure to avoid odor crossing as much as possible. Keep the amplitude of the movements as small as possible to avoid mouse stress.</li>
</ol>

Assessing Olfactory Impairment in Mice Due to Inhaled Environmental Pollutants

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The authors declare no conflicts of interest.

This article introduces two fundamental protocols designed for the swift assessment of olfactory impairment in mice. Varied inhalable environmental pollutants result in distinct levels of olfactory dysfunction in mice. The buried food test is employed to assess the capability to detect volatile odors, while the social odor discrimination experiment evaluates the animal&#39;s capacity to discern and differentiate various social odors. The protocol here serves to evaluate the toxic effects of environmental pollutants on ol...

Olfactory impairment has emerged as a noteworthy public health concern and is independently associated with an increased risk of neurodegenerative diseases. This condition can adversely affect overall well-being, contribute to the development of depressive symptoms, and result in a diminished quality of life. Its impact is prominently observed in the altered perception of food, hindrance in social communication, and heightened negative feelings1. Various factors, including sinonasal disease, upper respiratory tract infection, and traumatic brain injury, have been considered contributors to olfactory impairment in humans2. Notably, inhalable environmental pollutants such as PM2.5, estimated to range from 2% to 16%, enter the body through inspired air, traverse the nasal cavity, and reach specific olfaction-dedicated regions where they are deposited3,4,5,6,7. Recent findings indicate that inhalable environmental pollutants, including PM2.5 and ammonia, can indeed harm olfactory sensory neurons8,9,10. However, further validation is required to ascertain whether such damage directly leads to olfactory dysfunction. Hence, a meticulous evaluation of the effects of inhalable environmental pollutants on olfactory function is of particular importance.
Currently, numerous research laboratories employ mice as an alternative vertebrate model for behavioral experiments aimed at comprehending changes in olfactory function11,12,13,14. Mice were chosen as the preferred model system for investigating vertebrate chemical communication, and it exhibits a remarkable olfactory sensitivity crucial for foraging and social communication15. Furthermore, the continually evolving array of tools for observing and influencing mouse behavior has rendered this species exceptionally appealing for research on olfactory function16.
In this study, we employed the buried food test and the social odor discrimination experiment to assess olfactory impairment in a mouse model exposed to inhalable environmental pollutants. To enhance the precision of evaluation, we opted for the most representative method for assessing olfactory function. We systematically refined this method to ensure simplicity and clarity, allowing us to gauge the extent of olfactory dysfunction induced by inhalable environmental pollutants effectively.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.5-10 &#956;L&#160; adjustable micropipette</td>
			<td>Eppendorf, Germany</td>
			<td>3123000225</td>
			<td>Intranasal instillation</td>
		</tr>
		<tr>
			<td>0.9% saline solution</td>
			<td>Solarbio</td>
			<td>7647-14-5</td>
			<td>Dissolve pollutants</td>
		</tr>
		<tr>
			<td>Anhydrous zinc sulfate</td>
			<td>Macklin</td>
			<td>7733-02-0</td>
			<td>Expose mice</td>
		</tr>
		<tr>
			<td>Centrifuge tube (2 mL)</td>
			<td>Biosharp Incorporated</td>
			<td>BS-20-M</td>
			<td>Place urine</td>
		</tr>
		<tr>
			<td>Electronic balance</td>
			<td>Changzhou Ohaus Co.</td>
			<td>EX125DZH</td>
			<td>Weight anesthetics and pollutants</td>
		</tr>
		<tr>
			<td>GraphPad Prism</td>
			<td>GraphPad Software</td>
			<td>8.0.1</td>
			<td>statistic analysis</td>
		</tr>
		<tr>
			<td>Handheld Dust detector</td>
			<td>TSI&#160;Incorporated</td>
			<td>DuatTrak <img alt="Equation 1" src="/files/ftp_upload/66818/66818eq01.jpg" style="margin: auto;" />8532</td>
			<td>Inhalation-exposed mice</td>
		</tr>
		<tr>
			<td>Video recording equipment</td>
			<td>Apple Inc.</td>
			<td>iPhone 6s Plus</td>
			<td>The activity time of mice was recorded</td>
		</tr>
		<tr>
			<td>Vortex mixer</td>
			<td>Haimen Kylin-Bell Lab Instruments Co.</td>
			<td>Vortex-5&#160;</td>
			<td>Mix solution</td>
		</tr>
	</tbody>
</table>

studying the effects of inhaled environmental pollutants on olfactory function in mice

Olfactory impairment is a significant public health problem and independently predicts the risk of neurodegenerative diseases. Inhaled environmental pollutants exposure may impair olfaction; thereby, there is an urgent need for methods to evaluate the effects of inhaled environmental pollutants exposure on olfaction. Mice are ideal models for olfactory experiments because of their highly developed olfactory system and behavioral characteristics. To assess the effects of inhaled environmental pollutants exposure on olfactory function in mice, a detailed buried food test and social odor discrimination experiment is provided, including the experiment preparation, the selection and construction of experimental facilities, the testing process, and indexes of time. Meanwhile, timekeeping equipment, operational details, and the experimental environment are discussed to ensure the success of the assay. Zinc sulfate is used as the treatment to demonstrate the feasibility of the experimental approach. The protocol provides a simple and clear operational process for assessing the effects of inhaled environmental pollutants on olfactory function in mice.

Inhalable environmental pollutants impair olfactory function in mice. Atmospheric zinc emitted from incinerators and motor vehicles has been demonstrated to be an inhaled pollutant that can result in allergic lung inflammation20. Zinc sulfate is considered one of the typical compounds to cause olfactory dysfunction21. Therefore, we use zinc sulfate as the treatment to expose mice by intranasal instillation and test using the buried food test and social odor discrimination e...

Author Spotlight: Assessing the Olfactory Effects of Airborne Pollutants &#8212; Buried Food and Social Odor Tests

This paper provides a detailed description of the buried food test and social odor discrimination experiment to assess the effects of inhaled environmental pollutants exposure on olfactory function in mice.

Studying the Effects of Inhaled Environmental Pollutants on Olfactory Function in Mice

Olfactory impairment is a significant public health problem and independently predicts the risk of neurodegenerative diseases. Inhaled environmental ...

studying-effects-inhaled-environmental-pollutants-on-olfactory

Universidad San Sebastian

Research

JoVE Journal

Environment

605 Views.  Qingdao University. This paper provides a detailed description of the buried food test and social odor discrimination experiment to assess the effects of inhaled environmental pollutants exposure on olfactory function in mice.

Video: Author Spotlight: Assessing the Olfactory Effects of Airborne Pollutants — Buried Food and Social Odor Tests

Exploring the Effects of Atmospheric Forcings on Evaporation: Experimental Integration of the Atmospheric Boundary Layer and Shallow Subsurface

Evaporation is directly influenced by the interactions between the atmosphere, land surface and soil subsurface. This work aims to experimentally study evaporation under various surface boundary conditions to improve our current understanding and characterization of this multiphase phenomenon as well as to validate numerical heat and mass transfer theories that couple Navier-Stokes flow in the atmosphere and Darcian flow in the porous media. Experimental data were collected using a unique soil tank apparatus interfaced with a small climate controlled wind tunnel. The experimental apparatus was instrumented with a suite of state of the art sensor technologies for the continuous and autonomous collection of soil moisture, soil thermal properties, soil and air temperature, relative humidity, and wind speed. This experimental apparatus can be used to generate data under well controlled boundary conditions, allowing for better control and gathering of accurate data at scales of interest not feasible in the field. Induced airflow at several distinct wind speeds over the soil surface resulted in unique behavior of heat and mass transfer during the different evaporative stages.

A protocol for the design and construction of a soil tank interfaced to a small climate controlled wind tunnel to study the effects of atmospheric forcings on evaporation is presented. Both the soil tank and wind tunnel are instrumented with sensor technologies for the continuous in situ measurement of environmental conditions.

Evaporation is directly influenced by the interactions between the atmosphere, land surface and soil subsurface. This work aims to experimentally study ...

A CO2 Concentration Gradient Facility for Testing CO2 Enrichment and Soil Effects on Grassland Ecosystem Function

Continuing increases in atmospheric carbon dioxide concentrations (CA) mandate techniques for examining impacts on terrestrial ecosystems. Most experiments examine only two or a few levels of CA concentration and a single soil type, but if CA can be varied as a gradient from subambient to superambient concentrations on multiple soils, we can discern whether past ecosystem responses may continue linearly in the future and whether responses may vary across the landscape. The Lysimeter Carbon Dioxide Gradient Facility applies a 250 to 500 &#181;l L-1 CA gradient to Blackland prairie plant communities established on lysimeters containing clay, silty clay, and sandy soils. The gradient is created as photosynthesis by vegetation enclosed in in temperature-controlled chambers progressively depletes carbon dioxide from air flowing directionally through the chambers. Maintaining proper air flow rate, adequate photosynthetic capacity, and temperature control are critical to overcome the main limitations of the system, which are declining photosynthetic rates and increased water stress during summer. The facility is an economical alternative to other techniques of CA enrichment, successfully discerns the shape of ecosystem responses to subambient to superambient CA enrichment, and can be adapted to test for interactions of carbon dioxide with other greenhouse gases such as methane or ozone.

A CO2 Concentration Gradient Facility for Testing CO2 Enrichment and Soil Effects on Grassland Ecosystem Function

The Lysimeter Carbon Dioxide Gradient Facility creates a 250 to 500 &#181;l L-1 linear carbon dioxide gradient in temperature-controlled chambers housing grassland plant communities on clay, silty clay, and sandy soil monoliths. The facility is used to determine how past and future carbon dioxide levels affect grassland carbon cycling.

Continuing increases in atmospheric carbon dioxide concentrations (CA) mandate techniques for examining impacts on terrestrial ecosystems. Most ...

LC-MS Analysis of Human Platelets as a Platform for Studying Mitochondrial Metabolism

Perturbed mitochondrial metabolism has received renewed interest as playing a causative role in a range of diseases. Probing alterations to metabolic pathways requires a model in which external factors can be well controlled, allowing for reproducible and meaningful results. Many studies employ transformed cellular models for these purposes; however, metabolic reprogramming that occurs in many cancer cell lines may introduce confounding variables. For this reason primary cells are desirable, though attaining adequate biomass for metabolic studies can be challenging. Here we show that human platelets can be utilized as a platform to carry out metabolic studies in combination with liquid chromatography-tandem mass spectrometry analysis. This approach is amenable to relative quantification and isotopic labeling to probe the activity of specific metabolic pathways. Availability of platelets from individual donors or from blood banks makes this model system applicable to clinical studies and feasible to scale up. Here we utilize isolated platelets to confirm previously identified compensatory metabolic shifts in response to the complex I inhibitor rotenone. More specifically, a decrease in glycolysis is accompanied by an increase in fatty acid oxidation to maintain acetyl-CoA levels. Our results show that platelets can be used as an easily accessible and medically relevant model to probe the effects of xenobiotics on cellular metabolism.

Here we show isolated human platelets can be used as an accessible ex vivo model to study metabolic adaptations in response to the complex I inhibitor rotenone. This approach employs isotopic tracing and relative quantification by liquid chromatography-mass spectrometry and can be applied to a variety of study designs.

Perturbed mitochondrial metabolism has received renewed interest as playing a causative role in a range of diseases. Probing alterations to metabolic ...

A Flow-through Exposure System for Evaluating Suspended Sediments Effects on Aquatic Life

This paper describes the Fish Larvae and Egg Exposure System (FLEES). The flow-through exposure system is used to investigate the effects of suspended sediment on various aquatic species and life stages in the laboratory by using pumps and automating delivery of sediment and water to simulate suspension of sediment. FLEES data are used to develop exposure-response curves between the effects on aquatic organisms and suspended sediment concentrations at the desired exposure duration. The effects data are used to evaluate management practices used to reduce the interactions between aquatic organisms and anthropogenic causes of suspended sediments. The FLEES is capable of generating total suspended solids (TSS) concentrations as low as 30 to as high as 800 mg/L, making this system an ideal choice for evaluating the effects of TSS resulting from many activities including simulating low ambient levels of TSS to evaluating sources of suspended sediments from dredging operations, vessel traffic, freshets, and storms.

A robust and flexible flow-through exposure system designed to maintain sediment in suspension is presented. The system is used to investigate the effects of suspended sediment on various aquatic species and life stages in the laboratory.

This paper describes the Fish Larvae and Egg Exposure System (FLEES). The flow-through exposure system is used to investigate the effects of suspended ...

Evaluating the Effect of Environmental Chemicals on Honey Bee Development from the Individual to Colony Level

The presence of pesticides in the beekeeping environment is one of the most serious problems that impacts the life of a honey bee. Pesticides can be brought back to the beehive after the bees have foraged on flowers that have been sprayed with pesticides. Pesticide contaminated food can be exchanged between workers which then feed larvae and therefore can potentially affect the development of honey bees. Thus, residual pesticides in the environment can become a chronic damaging factor to honey bee populations and gradually lead to colony collapse. In the presented protocol, honey bee feeding methods are described and applied to either an individual honey bee or to a colony. Here, the insect growth regulator (IGR) pyriproxyfen (PPN), which is widely used to control pest insects and is harmful to the development of honey bee larvae and pupae, is used as the pesticide. The presenting procedure can be applied to other potentially harmful chemicals or honeybee pathogens for further studies.

Herein we present a method to feed pesticide contaminated food to both an individual honey bee and a beehive colony. The procedure evaluates the pesticide effect on individual honey bees by in vivo feeding of basic larval diet and also on the natural condition of beehive colony.

The presence of pesticides in the beekeeping environment is one of the most serious problems that impacts the life of a honey bee. Pesticides can be ...

Production and Measurement of Organic Particulate Matter in the Harvard Environmental Chamber

The production and the evolution of atmospheric organic particulate matter (PM) are insufficiently understood for accurate simulations of atmospheric chemistry and climate. The complex production mechanisms and reaction pathways make this a challenging research topic. To address these issues, an environmental chamber, providing enough residence time and close-to-ambient concentrations of precursors for secondary organic materials, is needed. The Harvard Environmental Chamber (HEC) was built to serve this need, simulating the production of gas and particle phase species from volatile organic compounds (VOCs). The HEC has a volume of 4.7 m3 and a mean residence time of 3.4 h under typical operating conditions. It is operated as a completely mixed flow reactor (CMFR), providing the possibility of indefinite steady-state operation across days for sample collection and data analysis. The operation procedures are described in detail in this article. Several types of instrumentation are used to characterize the produced gas and particles. A High-Resolution Time-of-Fight Aerosol Mass Spectrometer (HR-ToF-AMS) is used to characterize particles. A Proton-Transfer-Reaction Mass Spectrometer (PTR-MS) is used for gaseous analysis. Example results are presented to show the use of the environmental chamber in a wide variety of applications related to the physicochemical properties and reaction mechanisms of organic atmospheric particulate matter.

This paper describes operation procedures for the Harvard Environmental Chamber (HEC) and related instrumentation for measuring gaseous and particle species. The environmental chamber is used to produce and study secondary organic species produced from the organic precursors, especially related to atmospheric organic particulate matter.

The production and the evolution of atmospheric organic particulate matter (PM) are insufficiently understood for accurate simulations of atmospheric ...

Preparation of DMMTAV and DMDTAV Using DMAV for Environmental Applications: Synthesis, Purification, and Confirmation

Dimethylated thioarsenicals such as dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV), which are produced by the metabolic pathway of dimethylarsinic acid (DMAV) thiolation, have been recently found in the environment as well as human organs. DMMTAV and DMDTAV can be quantified to determine the ecological effects of dimethylated thioarsenicals and their stability in environmental media. The synthesis method for these compounds is unstandardized, making replicating previous studies challenging. Furthermore, there is a lack of information about storage techniques, including storage of compounds without species transformation. Moreover, because only limited information about synthesis methods is available, there may be experimental difficulties in synthesizing standard chemicals and performing quantitative analysis. The protocol presented herein provides a practically modified synthesis method for the dimethylated thioarsenicals, DMMTAV and DMDTAV, and will help in the quantification of species separation analysis using high performance liquid chromatography in conjunction with inductively coupled plasma mass spectrometry (HPLC-ICP-MS). The experimental steps of this procedure were modified by focusing on the preparation of chemical reagents, filtration methods, and storage.

Preparation of DMMTAV and DMDTAV Using DMAV for Environmental Applications: Synthesis, Purification, and Confirmation

This article presents modified experimental protocols for dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV) synthesis, inducing dimethylarsinic acid (DMAV) thiolation through mixing of DMAV, Na2S, and H2SO4. The modified protocol provides an experimental guideline, thereby overcoming limitations of the synthesis steps that could have caused experimental failures in quantitative analysis.

Dimethylated thioarsenicals such as dimethylmonothioarsinic acid (DMMTAV) and dimethyldithioarsinic acid (DMDTAV), which are produced by the metabolic ...

An Experimental Protocol for Studying Mineral Effects on Organic Hydrothermal Transformations

Organic-mineral interactions are widely occurring in hydrothermal environments, such as hot springs, geysers on land, and the hydrothermal vents in the deep ocean. Roles of minerals are critical in many hydrothermal organic geochemical processes. Traditional hydrothermal methodology, which includes using reactors made of gold, titanium, platinum, or stainless-steel, is usually associated with the high cost or undesired metal catalytic effects. Recently, there is a growing tendency for using the cost-effective and inert quartz or fused silica glass tubes in hydrothermal experiments. Here, we provide a protocol for carrying out organic-mineral hydrothermal experiments in silica tubes, and we describe the essential steps in the sample preparation, experimental setup, products separation, and quantitative analysis. We also demonstrate an experiment using a model organic compound, nitrobenzene, to show the effect of an iron-containing mineral, magnetite, on its degradation under a specific hydrothermal condition. This technique can be applied to study complex organic-mineral hydrothermal interactions in a relatively simple laboratory system.

Earth-abundant minerals play important roles in the natural hydrothermal systems. Here, we describe a reliable and cost-effective method for the experimental investigation of organic-mineral interactions under hydrothermal conditions.

Organic-mineral interactions are widely occurring in hydrothermal environments, such as hot springs, geysers on land, and the hydrothermal vents in the ...

Using Caenorhabditis elegans for Studying Trans- and Multi-Generational Effects of Toxicants

Information about toxicities of chemicals are essential in their application and waste management. For chemicals at low concentrations, the long-term effects are very important in judging their consequences in the environment and on human health. In demonstrating long-term influences, effects of chemicals over generations in recent studies provide new insight. Here, we describe protocols for studying effects of chemicals over multiple generations using free-living nematode Caenorhabditis elegans. Two aspects are presented: (1) trans-generational (TG) and (2) multi-generational effect studies, the latter of which is separated to multi-generational exposure (MGE) and multi-generational residual (MGR) effect studies. The TG effect study is robust with a simple purpose to determine whether chemical exposure to parents can result in any residual consequences on offspring. After the effects are measured on parents, sodium hypochlorite solutions are used to kill the parents and keep the offspring so as to facilitate effect measurement on the offspring. The TG effect study is used to determine whether the offspring are affected when their parent is exposed to the pollutants. The MGE and MGR effect study is systematical used to determine whether continuous generational exposure can result in adaptive responses in offspring over generations. Careful pick-up and transfer are used to distinguish generations to facilitate effect measurement on each generation. We also combined protocols to measure locomotion behavior, reproduction, lifespan, biochemical and gene expression changes. Some example experiments are also presented to illustrate the trans- and multi-generational effect studies.

Using Caenorhabditis elegans for Studying Trans- and Multi-Generational Effects of Toxicants

Trans- and multi-generational effects of persistent chemicals are essential in judging their long-term consequences in the environment and on the human health. We provide novel detailed methods for studying trans- and multi-generational effects using free-living nematode Caenorhabditis elegans.

Information about toxicities of chemicals are essential in their application and waste management. For chemicals at low concentrations, the long-term ...

Data Collection on Marine Litter Ingestion in Sea Turtles and Thresholds for Good Environmental Status

The following protocol is intended to respond to the requirements set by the European Union&#39;s Marine Strategy Framework Directives (MSFD) for the D10C3 Criteria reported in the Commission Decision (EU), related to the amount of litter ingested by marine animals. Standardized methodologies for extracting litter items ingested from dead sea turtles along with guidelines on data analysis are provided. The protocol starts with the collection of dead sea turtles and classification of samples according to the decomposition status. Turtle necropsy must be performed in authorized centers and the protocol described here explains the best procedure for gastrointestinal (GI) tract isolation. The three parts of the GI (esophagus, stomach, intestine) should be separated, opened lengthways and contents filtered using a 1 mm mesh sieve. The article describes the classification and quantification of ingested litter, classifying GI contents into seven different categories of marine litter and two categories of natural remains. The quantity of ingested litter should be reported as total dry mass (weight in grams, with two decimal places) and abundance (number of items). The protocol proposes two possible scenarios to achieve the Good Environmental Status (GES). First: &#34;There should be less than X% of sea turtles having Y g or more plastic in the GI in samples of 50-100 dead turtles from each sub-region&#34;, where Y is the average weight of plastic ingested and X% is the percentage of sea turtles with more weight (in grams) of plastic than Y. The second one, which considers the food remain versus plastic as a proxy of individual health, is: &#34;There should be less than X% of sea turtles having more weight of plastic (in grams) than food remains in the GI in samples of 50-100 dead turtles from each sub-region&#34;.

The protocol focuses on the collection of sea turtle samples, describing all the steps from the animal recovery and necropsy to the classification and quantification of ingested marine litter. Moreover, the representative results show how to use the collected data to elaborate the possible thresholds for Good Environmental Status.

The following protocol is intended to respond to the requirements set by the European Union&#39;s Marine Strategy Framework Directives (MSFD) for the ...