Microbio Exam 3: Microbial Ecology and environmental microbiology

Lecture 13 (Chapters 21-22)

What do animal-associated microbial communities provide for their hosts?

Can provide essential components for hosts.

Bacterial Provisions for hosts examples

1. Bacteria can encode enzymes to synthesize nutrients that are limited in the host’s diet (can allow invasive stinkbugs to colonize wide range of plants)

Population

Group of individuals of one species living in a common location

Ecosystem

Populations of species plus their habitat/environment.

Microbiome

Microbes associated with a host or animal/plant

Metagenome

The sum total of all DNA sequences from an environmental sample.

• Environment can be any natural or human-made environment or the microbiome of a plant or animal

• The DNA sequence obtained directly from a mixture of genomes.

How are environmental taxa’s identified?

SSU rRNA sequencing

Requirement for sampling

First, defining a target community to study

Steps of Samping/Sequencing

1. Separating the microbes from their physical environment

2. Break open the cells

3. Isolate the DNA

Drawback/Benefit of Different DNA Isolation methods

Different methods may lead to different view of a metagenome.

• Single universally applied method can allow for comparison across projects.

  • Multiple extraction methods may maximize coverage for all genomes in a sample

What is the purpose of a preliminary metagenomic screen?

To asses sequece diversity in a microbial community.

What is metabarcoding?

Using NGS to sequence SSU rRNA genes and identify taxa in a microbial community.

What are operational taxonomic units (OTU)’s?

Groups of similar DNA sequences used as a metagenomic approximation of species.

What must be constructed before complete metagenomic sequencing?

A DNA library.

What are the three steps of making a DNA library?

1. Concentrate DNA

2. Fragment DNA by size

3. Add adapters to fragment ends

What is the purpose of adding adapters to DNA fragments?

To prepare DNA fragments for sequencing by allowing them to interact with the sequencing platform.

What happens after DNA sequencing?

Computer analysis assembles overlapping sequences into scaffolds and helps reconstruct genomes.

How are DNA sequences assembled into genomes?

Overlapping DNA fragments (reads) are assembled into contigs, then scaffolds, and eventually patial genomes.

Contigs

Continuous DNA sequences assembled from overlapping reads, with no gaps.

What are scaffolds?

Larger DNA structures made by ordering and connecting contigs, potentially with gaps between them.

What tools are used to assemble and analyze DNA fragments and contigs?

Advanced computational tools (bioinformatics).

What is taxonomic binning?

Grouping DNA sequences based on which organism or taxonomic group they likely belong to.

What is the order of metagenomic sequence assembly?

Reads→ Contigs→Scaffolds→Partial genomes (through taxonomic binning).

What is a computational pipeline in genome assembly?

A series of programs combining mathematical tools and biological assumptions to reconstruct genomes from DNA sequences.

What is a de Bruin graph used for in genome assembly?

To identify connections between overlapping DNA sequences and help reconstruct the original genome.

What are the two types of genome assembly?

1. Reference-based assembly

2. De novo assembly

Reference-based genome assembly

Aligning DNA reads to an existing reference genome to help reconstruct the sequence.

De novo genome assembly

Assembling DNA reads into longer sequences without using an existing referenc

e genome.

MAGs

Metagenome-Assembled Genomes

Matthias Hess

His study yielded 15 partial genomes (MAGs) ranging from 60% to 93% estimated completeness.

• Completeness of each genome was estimated from the number of “core genes” identified.

Core genes

Genes found in nearly all members of a given taxonomic order. (made it to slide 14)