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Concept Map of Research Article: The Potential use of Bacterial Community…

- - - - Generates large amounts of data and obtains sequences for microbial taxa that are new and/or unculturable
    - - Use pyrosequencing to describe temporal changes during carrion decomposition and identify communities which have potential to be useful for PMI estimates
        
        Goal 2: Identify important bacteria taxa from these communities that are useful in estimating physiological time (surrogate of PMI estimations)
        
        Goal 3: Understand how quantifications of the necrobiome and its communities can be useful in forensic investigations
        
        Goal 1: Describe epinecrotic bacterial community composition and succession that occur in vertebrate decomposition
        
        Epinecrotic - organisms residing or moving on the surface of decomposing remains
- - - - Pyrosequencing
        
        Community structure was determined by modified bacterial tagged encoded FLX amplicon pyrosequencing
      - Pyrosequencing Data Analysis
        
        Viable sequences were aligned based on the 16sRNA secondary structure and bacterial taxon richness / relative abundance were determined for each sampling region at each taxonomic rank
      - DNA Extraction
        
        Used a modified chloroform-phenol extraction with DNA products from the three replicate carcasses mixed in equal concentrations and aliquoted for analysis by pyrosequencing
      - Bacterial Community Analyses
        
        Taxon richness was analyzed using linear regression over physiological time - with physiological time calculated by summing average daily temperature
        
        Indicator values were used to identify taxa that best represented communities of each sampling region (0 - no indication, 100 - perfect indication)
        
        Tiered statistical approach was also used to estimate decomposition time using bacterial community composition
- - - - Proteobacteria decreased over time, making Firmicutes the dominant taxon as time progressed (See Fig. 1)
        
        Family level results
        
        1st day: Moraxellaceae was the most dominant (51.3 ± 23.2 %) with Pasteurellaceae, Enterobacteriaceae, and Aerococcaceae representing 8.9 ± 4.9, 7.9 ± 13.8, and 6.8 ± 3.0 % relative abundance
        
        Moraxellaceae reduced after 1st day and wasn't seen on 3rd or 5th sampling day
        
        Aerococcaceae and Enterobacteriacea increased until the 3rd sampling day and then wasn't present on the 5th
        
        Planococcaceae was the most dominant family on the 5th day
        
        There was a significant negative relationship between family taxon richness and time
        
        Significant difference between sampling regions and interaction effect with sampling day
        
        Initial sampling: Bacteroidaceae and Moraxellaceae significant indicators
        
        Sampling day 5: Bacillaceae and Clostridiales Incertae Sedis XI significantly represented communities
- - - - Demonstrates that the understanding of bacterial communities in decomposition can be used to estimate physiological time
        
        Phylum level taxonomic resolution was better to distinguish between days of decomposition
        
        Family level indicator taxa were best at estimating variation in physiological time
  - - - Further analysis is needed to determine how changes in community structure alter insect arrival and colonization patterns
  - - - Samples stores in separate containers at −20 °C until further analysis
        
        Metagenomic sequencing performed once DNA has been extracted and processed: Pyrosequencing / Illumina
        
        Sequences classified and the overall bacterial community assessed by determining taxon richness and relative abundance
        
        Compare to reference data from similar environments & data from nearest weather station can be collected to back calculate the estimated physiological time range