negative consequences of glacial turbidity for the survival of freshwater planktonic heterotrophic flagellates - black polycarbonate sheet
Whip ell is a key component of the floating food chain in freshwater and marine ecosystems, as they are the main consumers of bacteria.
Although they are ubiquitous in aquatic ecosystems, they are not detectable in numbers in turbid glaciersfed lakes.
Here, we show that glacier particles have a negative effect on the survival and growth of the alien whip flag.
The influence of glacier particles is concentrated.
It is caused by their interference with the absorption of bacteria rather than physical damage.
These results are the first to reveal why the establishment of the divergent whip flag population is hindered in very turbid glacial lakes.
As the glaciers are disappearing around the world, the recently formed turbid melodious Lakes represent an excellent opportunity to learn about the environmental conditions that may shape the lake community at the end of the last glacial action.
Surface water samples obtained from glacier particles were collected with improved Schindler-
Patalas sampler from turbid lakes FAS1 and fas3.
These lakes are located in the Austrian Alps (
47 ° 04 '25-37 "N 10 ° 13' 21-28" E)
Together with the other four systems in this field, it represents the turbid gradient generated by the Ice Age sequence ().
In the dark at 4 °c, place the water sample in 10 liters of carboys for 6 months to allow the particles to settle.
Most of the water is then extracted by suction and the remaining water, including sediment, is dried for 5 days at 45 °c.
In addition, sediment samples were collected from FAS1 with a plastic core and removed and dried in the same way the upper 1mm of sediment.
Then grind the dry material and sift through the metal mesh (Size 63 μm)
, Although no sediment material was retained.
Glacial particles for experimental and further characterizing (see )
Therefore, it is the sum of the settling particles plus the settling particles in the upper sediment.
This parameter is measured by a scatter meter using a handheld turbidity meter (WTW Turb 430u2005T)
Use "white light" from tungsten lamps ".
The method follows the agreement recommended by the US Environmental Protection Agency (180. 1). A three-
Point calibration was carried out with the standard of 0.
02, 10 and 1000 South University.
Three measurements were made for each sample and the mean value was calculated.
In order to obtain a reference value for the abundance of bacteria and whip ellate in Faselfad lakes, water samples were collected from FAS1 (1u2005m depth), FAS3 (
1 µm, 6 µm, 10 µm, 15 µm depth)
And clear FAS4 (
1 M, 6 m, 10 m, 13 m depth)
In the field of summer
Collect water samples using the same type of sample feeder and fix them on site with formaldehyde (
Final concentration 2%)
, And transported by helicopter to the Innsbruck lab in a container that kept cool and dark.
To analyze bacterial abundance, subsamples of 5 min to 15 ml were stained with fluorine chromium 4'6 '-diamidino-2-phenylindole (DAPI)(10u2005μg mL)
Then concentrate on a black polycarbonate membrane filter (pore size 0. 2u2005μm).
Count at least 400 cells under a fluorescence microscope (
Axiophot month, Zeiss)
The magnification is 1250 ×.
Saved sub-samples (20–80u2005mL)
Stained and counted as bacteria, but filtered onto a black polycarbonate membrane filter with a pore size of 0. 8u2005μm.
More water was filtered when HNF was not detected.
In other tests, a small amount of HNF cultures were added to the sample to ensure that they can be detected in the presence of particles.
In order to distinguish between different culture bacteria and self-supporting bacteria, the presence of chlorophyll in individuals was examined.
In an automatic fluorescence plasma, a filter bank with blue excitation is used (
Excitation 450-490 nm, emission filter LP 520 nm).
On July 5, 2011, 5 µl water samples were collected at the FAS4 Lake in the depth of 13 µm (
Usually the depth with the largest abundance of HNF)
And was transported by helicopter to Innsbruck's lab, in a plastic vehicle in the dark near the temperature.
The sample is used to test the effect of two different particle concentrations (14 NTU;
57 mg L and 30 NTU: 113 mg L)
About the natural combination of whip flag.
In order to exclude large organisms, samples were screened through the plankton Network (45 μm mesh size)
It is divided into 100 glass bottles and shaken every day to avoid precipitation.
The experiment was carried out in the dark and at 15 °c and treated and controlled using three repetitions (i. e.
, No particles added), respectively.
In order to exchange the air, the bottle is opened briefly every day.
As mentioned above, the abundance of bacteria and HNF was determined at the beginning of the experiment and after 1 week.
Most of the experiments were done with the HNF species sp. strain JBM10 (4-7 μm in diameter)
It is a common cryptorum genus in freshwater communities.
This species can be classified as small-to medium-sized bacteria.
In order to compare it with another swallowing effect, but at the same time, the role of light Nutrition (i. e. , mixotrophic)
Species, we include chickens (Length 12-15 μm)
This is a whip ellate, usually forming colonies, common in freshwater and alpine lakes. sp. and .
Provided by Jens Boenigk. sp.
Growth in inorganic basic media (IBM)
5 mg L of glucose and 40 mg L of protein group.
Training in WCg-medium.
The culture was preserved in an environmental chamber, at 23 °c, in a transparent 50 ml polystyrene tube, with a light time of 8: 16, moving to fresh medium every 2-3.
Two days before the experiment, 1 ml medium and 9 ml fresh medium were transferred to 15 ml test tubes.
At the beginning of the experiment, 100 µμ l cultures and 900 µL fresh medium were transferred to 2 Eppendorf vials.
Treatment and control experiments were performed using five repetitions (i. e.
, No glacier "flour" added "), respectively.
Under the condition of a concentration of 57 mg dry weight L, glacier particles are added to the treatment (14 NTU)
In the case of sp, in the experiment of 7, 14, 30 NTU.
These turbidity values are medium, and may be one order of magnitude higher in other turbid glacial-water-melting lakes.
The experiment was carried out at 15 °c and in the dark. in addition to the test effect in the presence of light, the light was carried out at 23 °c: Dark Light 8: 16The pH-
After adding the glacier "flour", the value in the treatment was examined.
This measurement shows that the addition of glacier "flour" has not changed the pH value extensively (control 7. 09. treatment 7. 05).
To avoid particle deposition, all vials (i. e.
, And controls)
Placed in a spinner (
VWR tube spinner)
The fixed speed is 18 rpm.
The rotation mode is adjusted to 15 min (on)and 45u2005min (off)
Avoid the cycle of potential shear stress on whip flag.
The change of whip ellate abundance over time was evaluated by counting cells with 0 drops.
5-2 u2005 μ l under Olympus BX 50 (
Digital interference contrast field)
A microscope with a magnification of 40-200.
At least 200 cells were counted within a 24-hour time interval.
Count the number of bacteria according to the above time interval.
Grazing rate of Sp.
By using fluorescent labeled bacteria (FLB)as surrogates.
The bacteria are contaminated with yellow.
Green fluorescent dye, 5-(4,6-Nitrogen chloride-2-yl)
Amino fluorescence (DTAF).
Bacteria used to prepare FLB are bacteria that grow together with sp.
It's a mixed culture.
To remove whip ell, filter the media through a polycarbonate filter with a 2 μm aperture.
To estimate the concentration of FLB in the suspension system, cell abundance in small samples (0. 5u2005ml)
Count under a fluorescence microscope at 1250 × magnification.
For the experiment, sp. culture (10u2005ml)
First inoculation into a 390 ml glass bottle with 500 ml IBM medium, 5 days later transfer the dense whip ellate culture to a 50 ml tube for three repeated treatments and controls, respectively.
Estimated volume of FLB to be added (
10-15% of the total number of bacteria in the reference. ), a sample (2u2005ml)
Fixed and stained as described above, as well as bacteria.
Control and treatment tubes are then supplemented with FLB and FLB Glacier particles (57u2005mg L), respectively.
The experiment was carried out in the dark at 16 °c.
Sub-samples of 5 ml were taken after time 0 and 2, 5, 10 and 15 minutes.
Fix the sub-sample with 0 immediately.
25 l alkaline Lugol solution (0. 5%)and 0.
25 ml buffer formaldehyde (40%)
, And stored in the dark at 4 °c until processed within 24 hours.
Then dye the sub-sample with DAPI and concentrate on the black polycarbonate filter (0. 8u2005μm pore-size).
Under the fluorescence microscope, the whip ell was examined with a magnification of 1250 ×.
By switching between filters to evaluate the number of FLB intake per personset for UV (DAPI)
And blue light excitation (DTAF).
At least 60 people have been checked on each filter.
Intake rate (UR)
In FLB cell h, the regression slope between FLB cells and time was obtained by extrapolation for 60 min.
Clearance rate (CR)
In nL cells, h is calculated by dividing UR by the number of FLB used in the experiment.
Experimental results of the influence of glacier "flour" on short time
By comparing the linear regression slope between control and treatment with the Student t-test.
The experimental results of the effects of different particle concentrations on sp were tested.
The experiment of natural community was analyzed. way ANOVA (
Including repeated measurements at different times)with Holm-Sidak post-hoc test.
SigmaStat month in the analysis based on the application software.
Military and StatSoft software statistics.