Name: a low-cost method of measuring the in situ primary productivity of periphyton communities of lentic waters
Uploaded: 2022-12-16T13:05:00
Description: Watch this Scientific Journal Video about A Low-Cost Method of Measuring the In Situ Primary Productivity of Periphyton Communities of Lentic Waters at JoVE.com

This study was supported by the Czech Science Foundation (GACR 19-05791S), RVO 67985939, and by the CAS within the program of the Strategy AV 21, Land save and recovery. Many thanks to Ond&#345;ej Sihelsk&#253; for taking the shots in the field - without him, the filming would have been complete hell. The project would not be possible without tight cooperation with companies, Palivov&#253; Kombin&#225;t &#218;st&#237; s.p. and Sokolovsk&#225; Uheln&#225;, who provided access to the studied localities.

NOTE: Before sampling, determine the degree of replications based on the overall project needs, statistical design, or expected amount of sample variability.Five replicate pairs of light and dark incubation bottles are suggested for precise statistical analysis and to account for potential sample loss or breakage. The described floating experimental barge is designed to carry five replicates plus one pair of blank controls; see Figure 1 for a technical drawing of the experimental barge.
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<ol>
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The methodology described in this paper is based on the principle of the light and dark bottle oxygen technique in combination with the noninvasive technique of measuring O2 concentration using optical oxygen sensors. This system allows the parallel measurement of different incubation settings as the optical fiber for measuring O2 can be moved quickly from bottle to bottle. The benthic communities from various depths can differ in taxonomic composition and productivity; simultaneously measuring them...

Primary productivity is the only entry of autochthonous carbon into the aquatic systems that form the entire system food web1. Hence, the accurate estimation of primary productivity is an essential step toward understanding the functioning of aquatic ecosystems. Littoral zones are areas of high primary productivity and biodiversity. In addition to phytoplankton, periphyton (hereafter referred to as microbial mats) and macroalgae are assumed to significantly contribute to primary productivity in littoral zones2. Due to their sessile lifestyle and significant spatial heterogeneity, quantification of primary productivity is not trivial. 
Primary productivity is driven mainly by light intensity, temperature, availability of nutrients, and distribution of the ionic species of the carbonate system in the respective depths of euphotic zones3,4. The depth markedly influences the spatial distribution of microbial mats. Microbial communities must cope with the adverse effects of high irradiation and pronounced seasonal temperature variations in shallow depths and with lower light intensity at greater depths. In addition to the depth gradient, dynamic trophic interactions generate multiple and complex spatial patterns at different scales5. This complex system is complicated to simulate in the laboratory. The most accurate way to infer the metabolic activity of individual primary producers from littoral zones is to set up in situ experiments. 
The methodology introduced in this paper is based on the traditional chamber method2,6,7, together with a transportable and easy-to-build low-cost floating barge. This allows the measurement of primary productivity at different depths under the natural light spectrum, temperature, and different distribution of the ionic species of the carbonate system with the depth. The method is based on the principle of light versus dark bottle oxygen, which was first employed to measure phytoplankton photosynthesis6 and is still commonly used6,7. It compares the rate of change in oxygen in bottles kept in the light (which includes the effects of primary productivity and respiration) with those held in the dark (respiration only)8. The method uses oxygen evolution (photosynthesis) as a proxy for primary productivity. The measured variables are net ecosystem productivity (NEP, as a change in O2 concentration over time in light conditions) and ecosystem respiration (RE, as a change in O2 concentration over time in the dark). Gross ecosystem productivity (GEP) is the calculation of the difference between the two (Table 1). The term "ecosystem" is used here to denote that the periphyton is composed of autotrophic and heterotrophic organisms. The most significant improvement of this traditional chamber method is using noninvasive oxygen optical sensors and optimization of this primarily planktonic method for measuring periphytic primary productivity.
The technique is described in the example of measuring microbial mats in the littoral zone of newly emerged post-mining lakes in the Czech Republic-Milada, Most, and Medar. The metabolic activity of microbial mats is determined using direct in situ measurement of O2 fluxes performed directly at specific depths, where the studied communities naturally occur. Heterotrophic and phototrophic activity is measured in closed glass bottles equipped with noninvasive optical oxygen sensors. These sensors detect the partial pressure of oxygen using the fluorescence of light-sensitive dyes. The bottles with microbial mats are suspended and incubated on a floating device at the appropriate depths. The oxygen concentration inside the bottles was continuously measured during the daylight period from the small boat. 
Samples of intact microbial mats are collected and placed in gas-tight incubation bottles at designated depths by scuba divers. Each bottle is equipped with a noninvasive optical oxygen microsensor, which monitors the O2 productivity/consumption over time. All measurements are done in five replicate dark/light pairs in each depth. The temperature and photosynthetically active radiation (PHAR) intensities are measured at respective depths throughout the incubation. After 6 h of in situ incubation (daylight hours), the microbial mats are harvested from the bottles and dried. O2 fluxes are normalized to microbial biomass. As a control, fluxes are corrected for changes in O2 concentration in separate light and dark gas-tight bottles (blank controls) containing lake water without microbial mat biomass. Below are detailed instructions for building the floating barge and performing the whole experiment step-by-step. This paper also presents representative results from the measurements of microbial mats at two depths (1 m and 2 m), with five replicates at each depth. Actual temperature and light intensity were measured during the whole experiment using dataloggers.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Aluminum angle L profile 40 x 40 mm x 3 mm, length 2,000 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Aluminum flat bar 40 x 3 x 350 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Bucket 15 L with concrete infill&#160;</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Carabine hook with screw lock 50 x 5 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>electric tape black</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Extruded polystyrene (XPS) material 500 x 200 x 150 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Fibox 3 LCD trace</td>
			<td>PreSens Precision Sensing GmbH</td>
			<td></td>
			<td>stand-alone fiber optic oxygen meter</td>
		</tr>
		<tr>
			<td>Hondex PS-7 Portable Depth Sounder</td>
			<td>Hondex&#160; - Honda Electronics</td>
			<td></td>
			<td>to measures distances through water - to bottom depth measurement; https://www.honda-el.net/industry/ps-7e</td>
		</tr>
		<tr>
			<td>KORKEN - glass tight-seal jar 0.5 L</td>
			<td>IKEA</td>
			<td></td>
			<td>incubation bottles; https://www.ikea.com/cz/en/p/korken-jar-with-lid-clear-glass-70213545/</td>
		</tr>
		<tr>
			<td>metal hook&#160;</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Oxygen Sensor Spot SP-PSt3-NAU-D5</td>
			<td>PreSens Precision Sensing GmbH</td>
			<td></td>
			<td>non-invasive optical oxygen sensor for measurements under Real Conditions</td>
		</tr>
		<tr>
			<td>SCOUT infantable canoe</td>
			<td>GUMOTEX</td>
			<td></td>
			<td>https://www.gumotexboats.com/en/scout-standard#0000-044667-021-13/11C</td>
		</tr>
		<tr>
			<td>Screw 10 x 170 mm with hexagonal nuts</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Screw 4 x 15 mm with hexagonal nuts</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Screw 4 x 15 mm with wing nuts</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Snap hooks 50 x 5 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Steel Carabine hook 50 x 5 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Steel chain with wire diameter 3 mm, inside link 5.5 x 26 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Steel chain, 5 m</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>toothbrush</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>tweezer</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Washer 10 x 50 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Washer 4 x 10 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Washer 4 x 10 mm</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

a low-cost method of measuring the in situ primary productivity of periphyton communities of lentic waters

Measuring the in situ primary productivity of periphyton during the growing season gradient can elucidate the quantitative effect of environmental drivers (mainly phosphorus concentration and light intensity) and species composition on primary productivity. Primary productivity is mainly driven by light intensity, temperature, availability of nutrients, and distribution of the ionic species of the carbonate system in the respective depths of the euphotic zone. It is a complex system that is very difficult to simulate in the laboratory. This cheap, transportable, and easy-to-build floating barge allows measuring the primary productivity accurately-directly under the actual natural conditions. The methodology is based on measuring the primary productivity in real time using noninvasive oxygen sensors integrated into tightly sealed glass jars, enabling online oxygen flux monitoring and providing new insights into metabolic activities. Detailed seasonal in situ measurements of gross primary productivity of microbial mats (or other benthic organisms) can improve current knowledge of the processes controlling primary productivity dynamics in lentic waters.

<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/64078/64078fig05.jpg" /> 
Figure 5: Net and gross ecosystem productivity of microbial mats during daylight. (A) Light bottle-net ecosystem productivity: time course data of net oxygen productivity of microbial mats from the light bottles. The oxygen concentration change in incubation bottles was measured after 1 h during daylight. Grey circles: bottles with samples of m...

Watch this Scientific Journal Video about A Low-Cost Method of Measuring the In Situ Primary Productivity of Periphyton Communities of Lentic Waters at JoVE.com

A Low-Cost Method of Measuring the In Situ Primary Productivity of Periphyton Communities of Lentic Waters

Presented here is a cost-effective and transportable method/facility for measuring the primary productivity of microbial mats under actual in situ environmental temperature and light conditions. The experimental setup is based on widely available materials and can be used under various conditions while offering the advantages of laboratory-based models.

A Low-Cost Method of Measuring the In Situ Primary Productivity of Periphyton Communities of Lentic Waters

Measuring the in situ primary productivity of periphyton during the growing season gradient can elucidate the quantitative effect of environmental drivers ...

Detailed in-situ measurement of the gross productivity of periphyton or any water microorganism in a suitable site can improve current knowledge of the processes controlling primary productivity dynamic in lentic waters. The major improvements in the proposed methods are non-invasiveness, cost-effectiveness, and the possibility of simultaneous measurements in various location. It also enables the collection of large data assets without additional expenses.Replacing the barge in conducting the experiments, use an inflatable kayak as it's easily transportable. Select a place with an ideal depth to anchor the float. Next, attach the assembled barge behind the stern of the boat and carefully lower the anchor along the side of the boat.Align the anchor by hanging it slightly below the water surface so that the float can be easily towed with the anchor to the spot with the required depth. After reaching the spot, untie the anchor from the boat and lower it to the bottom, then secure the barge to the anchor chain and the chains for attaching the incubation bottles to the barge. To prepare the incubation bottles, attach oxygen optical sensor spots to the inner wall of transparent, wide-neck 0.5-liter bottles with gas-tight seals, then make an opaque layer for the dark treatment bottles by wrapping them with black electrical tape.Cut a tiny hole in the spot of the placed optical sensor and ensure that the cut hole is slightly smaller than the diameter of the sensor to prevent light from entering the bottle. Place the incubation bottles in the portable box and dive with the box to the respective depth without disturbing the sediment in the surrounding water. Next, using long tweezers, carefully fill the samples into the incubation bottles, taking care not to disturb the biomass of the sample too much.If the microbial mats grow on a solid surface such as a small stone, carefully transfer the whole stone with intact biomass into the bottle. Fill one pair of light and dark bottles with clean water from the respective depths without any sediments to serve as blank controls. After ensuring that the water in all incubation bottles is clean and contains no disturbing sediment, bring the closed bottles to the boat anchored to the floating barge.Attach the first two pairs of incubation bottles to the snap hooks on the first chain, then measure the initial oxygen concentration in each bottle using the fiber-optic oxygen meter. Attach the optical cable of the meter to the oxygen sensor mounted inside the bottle and within a few seconds, read out the oxygen concentration in the meter and record the measured value. Immediately after measurement, carefully lower the chain with the attached bottles back into the water, ensuring that the incubation bottles are placed at the same depth from which the biomass placed in them was sampled.After one hour, measure the oxygen concentration again by carefully pulling each chain with the bottles into the boat. Reading the oxygen value was demonstrated previously and lowering the samples into the water again. After completing all the measurements, take the samples out from the incubation bottles and scrub the microbial mat grown on the surface of a hard substance like stone with a toothbrush or small knife, then transfer the contents to plastic flasks.An increase in the oxygen concentration over time is apparent in both the control and the sample bottles exposed to light indicating net ecosystem productivity. Nevertheless, the slope of increase is significantly higher in bottles with microbial mat samples. The change in the oxygen concentration over time in the dark bottles is the sum of autotrophic and heterotrophic respiration.In this case, the slope of oxygen concentration in the control bottle is not significantly different from zero. Subtracting the respiration rates from net ecosystem productivity yields the rates of gross ecosystem productivity and the data complies well with the definition. A practical application of this method was achieved in the fields of three post-mining lakes that show the gross ecosystem productivity of the periphytic community during the vegetation season.During the biomass sampling process underwater, it is important to ensure that no air bubbles remain trapped inside the incubation bottles once they are closed after adding the samples. Using this procedure, it is possible to quantify oxygen exchange between the water body and any organism or community of organisms of a suitable size. This method allows studying the year-round primary productivity of selected organism in situ by making the natural condition as much as possible, thus giving a better idea of its relative importance for whole-life carbon metabolism.

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