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Phenotype MicroArrays as a complementary tool to next generation sequencing for characterization of tree endophytes.

Blumenstein K, Macaya-Sanz D, Martín JA, Albrectsen BR, Witzell J - Front Microbiol (2015)

Bottom Line: Here, we present detailed descriptions of two different PM protocols used in our recent studies on fungal endophytes of forest trees, and highlight the benefits and limitations of this technique.We found that the PM approach enables effective screening of substrate utilization by endophytes.For the best result, we recommend that the growth conditions for the fungi are carefully standardized.

View Article: PubMed Central - PubMed

Affiliation: Southern Swedish Forest Research Centre, Swedish University of Agricultural Sciences, Alnarp Sweden.

ABSTRACT
There is an increasing need to calibrate microbial community profiles obtained through next generation sequencing (NGS) with relevant taxonomic identities of the microbes, and to further associate these identities with phenotypic attributes. Phenotype MicroArray (PM) techniques provide a semi-high throughput assay for characterization and monitoring the microbial cellular phenotypes. Here, we present detailed descriptions of two different PM protocols used in our recent studies on fungal endophytes of forest trees, and highlight the benefits and limitations of this technique. We found that the PM approach enables effective screening of substrate utilization by endophytes. However, the technical limitations are multifaceted and the interpretation of the PM data challenging. For the best result, we recommend that the growth conditions for the fungi are carefully standardized. In addition, rigorous replication and control strategies should be employed whether using pre-configured, commercial microwell-plates or in-house designed PM plates for targeted substrate analyses. With these precautions, the PM technique is a valuable tool to characterize the metabolic capabilities of individual endophyte isolates, or successional endophyte communities identified by NGS, allowing a functional interpretation of the taxonomic data. Thus, PM approaches can provide valuable complementary information for NGS studies of fungal endophytes in forest trees.

No MeSH data available.


An assortment of nitrogen sources leading to clearly visible differences in optical density (750 nm) and morphology of the fungal mass of Trichoderma harzianum. Measurement at 144 h after inoculation to the plates (bars represent the mean of readings from two plates).
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Figure 7: An assortment of nitrogen sources leading to clearly visible differences in optical density (750 nm) and morphology of the fungal mass of Trichoderma harzianum. Measurement at 144 h after inoculation to the plates (bars represent the mean of readings from two plates).

Mentions: Phenotype MicroArray technique also allowed us to observe the effect of substrates on the morphology of the tested fungi. In particular, nitrogen sources seemed to induce varying morphological responses. For instance, only little fungal mass was produced when T. harzianum grew on cytidine or cytosine (Figure 7). Tyramine and formamide triggered production of in green fungal mass, whereas acetamide resulted in yellow, and adenosine yellow–green, fungal mass. Guanine induced formation of dense, dark green fungal mass in T. harzianum.


Phenotype MicroArrays as a complementary tool to next generation sequencing for characterization of tree endophytes.

Blumenstein K, Macaya-Sanz D, Martín JA, Albrectsen BR, Witzell J - Front Microbiol (2015)

An assortment of nitrogen sources leading to clearly visible differences in optical density (750 nm) and morphology of the fungal mass of Trichoderma harzianum. Measurement at 144 h after inoculation to the plates (bars represent the mean of readings from two plates).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4585013&req=5

Figure 7: An assortment of nitrogen sources leading to clearly visible differences in optical density (750 nm) and morphology of the fungal mass of Trichoderma harzianum. Measurement at 144 h after inoculation to the plates (bars represent the mean of readings from two plates).
Mentions: Phenotype MicroArray technique also allowed us to observe the effect of substrates on the morphology of the tested fungi. In particular, nitrogen sources seemed to induce varying morphological responses. For instance, only little fungal mass was produced when T. harzianum grew on cytidine or cytosine (Figure 7). Tyramine and formamide triggered production of in green fungal mass, whereas acetamide resulted in yellow, and adenosine yellow–green, fungal mass. Guanine induced formation of dense, dark green fungal mass in T. harzianum.

Bottom Line: Here, we present detailed descriptions of two different PM protocols used in our recent studies on fungal endophytes of forest trees, and highlight the benefits and limitations of this technique.We found that the PM approach enables effective screening of substrate utilization by endophytes.For the best result, we recommend that the growth conditions for the fungi are carefully standardized.

View Article: PubMed Central - PubMed

Affiliation: Southern Swedish Forest Research Centre, Swedish University of Agricultural Sciences, Alnarp Sweden.

ABSTRACT
There is an increasing need to calibrate microbial community profiles obtained through next generation sequencing (NGS) with relevant taxonomic identities of the microbes, and to further associate these identities with phenotypic attributes. Phenotype MicroArray (PM) techniques provide a semi-high throughput assay for characterization and monitoring the microbial cellular phenotypes. Here, we present detailed descriptions of two different PM protocols used in our recent studies on fungal endophytes of forest trees, and highlight the benefits and limitations of this technique. We found that the PM approach enables effective screening of substrate utilization by endophytes. However, the technical limitations are multifaceted and the interpretation of the PM data challenging. For the best result, we recommend that the growth conditions for the fungi are carefully standardized. In addition, rigorous replication and control strategies should be employed whether using pre-configured, commercial microwell-plates or in-house designed PM plates for targeted substrate analyses. With these precautions, the PM technique is a valuable tool to characterize the metabolic capabilities of individual endophyte isolates, or successional endophyte communities identified by NGS, allowing a functional interpretation of the taxonomic data. Thus, PM approaches can provide valuable complementary information for NGS studies of fungal endophytes in forest trees.

No MeSH data available.