Dear researchers,
My current research focuses on the bacterial interactions in activated sludge of wastewater treatment system. I write to consult a few questions about my confusions in investigating bacterial interactions of complex biofilm communities. To estimate microbial interactions, it is important to move beyond macroscale analysis and focus on micron-scale heterogeneity and spatial associations with enough throughput and statistical associations. Now I’m looking for approaches to sampling and sequencing micron-scale biofilm samples, but there were still some questions puzzling me. The questions are as follows:
(1) Many studies have found that fine-scale heterogeneity in microbial communities is a common characteristic of biofilm samples. Our data also confirmed that the bacterial population in activated sludge had significant spatial heterogeneity at the micron scale. I believe that the micron-scale heterogeneity is an inherent property of the biofilm community that has nothing to do with measurement. But I'm not sure if that's correct. For example, I pulverized the biofilm samples and used flow cytometry sorting to produce 1000 single clusters (80 μm), after which I sequenced each single cluster and calculated micron-scale heterogeneity. Will these procedures (e.g., pulverizing) result in any additional heterogeneity? or Does this approach capture true micron-scale heterogeneity?
(2) When sampling at centimeter to micrometer scales (e.g., 1cm, 0.1cm, 0.01cm, 1mm, 0.1mm, 10 μm), we may see various spatial heterogeneity of biofilm communities. Are there criteria for selecting the optimal length scale at micron-scale to estimate the spatial heterogeneity of biofilm communities? How can we select the optimal length scale for studying spatial heterogeneity and interspecies interactions in complex biofilm communities?
Would it be possible for you to explain these questions to me?
Regards,
Thanks in advance.
This is an interesting problem. I'm answering principally to make sure I get to see the other answers.
I think your approach of fragmenting the biofilm then sequencing the fragments is a good one. I think the answers would be still more interesting if you did some form of metabarcoding in order to characterise all the species in the fragments; this would reveal microscale associations within the biofilm.
Thanks for your reply and attention. Our team has sequenced 200 single micron-scale clusters of anammox consortia. We did discover several interesting results that differed from the bulk-scale researches. Coinciding with your suggestion, we are now looking for some metabarcoding methods to enlarge the sampling throughput.
- Badges
- Science topic