Blog 2: Molecular community analysis of the microbiological composition in different irrigation systems (tree nursery in Schinznach)

Overview

Inside the tree nursery two different irrigation systems are used. The first one is a classical sprinkler which uses water from the pond inside the facility and the second one is a drip irrigation system that uses fresh water. The different usage of water has mainly to do with the water quality. For the drip irrigation system clear water is necessary or otherwise the nozzles would be overgrown with algae and in need for replacement in a short amount of time.

Next to the site, a little stream called “Talbach” is flowing into the river Aare. Possible interactions with the plant nursery and their pond inside the site with the stream should also be investigated.

Goal

This blog entry tries to answer the following main research question:

“Which prerequisites need to be available in order to do a comparison of the different irrigation systems in pot-substrate and their impact for the microbiological community in soil and water?”

In order to do so, a molecular method with its pros and cons are compared against each other. The goal is to find the best analysis technique for the given task and time. To validate if the chosen method is feasible in the project week, an estimated time table is shown.

Additionally, the influence of the used irrigation water, which is stored in the pond on the Talbach, should be assessed.

Possible methods

The irrigation systems and their used water (fresh water or waste water), have a direct impact on the microbiological community in soil. A shift in the microbiological composition, especially in disturbance scenarios e.g. heat, can be observed depending on the used irrigation water. (Frenk et al.)

It is not clear how strong the impact of the pond for the stream Talbach is. Therefor it is necessary to analyse the stream at two different points. Sample point 1 should have no water from the pond in it and is therefore uncontaminated. Sample point 3 is further downstream and came in contact with water from the pond. Sample point 2 is there to have an estimation of the water composition and its quality in the retention basin. 

Figure 1: Overview of the different sample points at the site.

The two separate research questions can be analysed with the following method.

A fast and uncomplicated way to analyse a microbiological community is PCR-DGGE. (Madigan) Firstly, the DNA has to be extracted from the soil or water samples. For that, special kits and procedures are available. (Angel et al.) The isolated DNA can then be further analysed.

For that, a PCR is done with a primer-set for general bacterial community. This could be the primer 341F-GC containing a 40-bp GC clamp for better separation in DGGE or 907R. After amplification the samples are added to a DGGE gel containing 20% to 70% urea with TAE as running buffer (2 M Tris base, 1 M glacial acetic acid, 50 mM EDTA). (Frenk et al.)

Because the goal is to see a shift in the microbiological community and not a detailed phylogenetic tree, a sequencing of the different amplificants is not necessary.

The method is simple and fast to perform but can only give a rough overview of the different bacterial species in the sample. Species which are low in density are most likely not visible on the denaturing gel. 

Time table

To see if the method is feasible in the given time a time table was created.

 

 

References

Angel, Roey, et al. ‘Methanogenic Archaea Are Globally Ubiquitous in Aerated Soils and Become Active under Wet Anoxic Conditions’. The ISME Journal, vol. 6, no. 4, Apr. 2012, pp. 847–62. PubMed Central, doi:10.1038/ismej.2011.141.

Frenk, Sammy, et al. ‘Quality of Irrigation Water Affects Soil Functionality and Bacterial Community Stability in Response to Heat Disturbance’. Appl. Environ. Microbiol., vol. 84, no. 4, Feb. 2018, pp. e02087-17. aem.asm.org, doi:10.1128/AEM.02087-17.

Madigan, Michael T. Brock Biology of Microorganisms. Fourteenth edition, Pearson, 2015.