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ASM2012
This group catalogs microblogging notes from talks and events at the 2012 American Society for Microbiology General Meeting. This group is public.
David Paez-Espino - Time Series Metagenomic Analyses of CRISPRs
Ruchira S. Datta liked this
I'm running out of juice. Might lose me mid talk.
- Thomas Sharpton
Describing how CRISPRs work
- Thomas Sharpton
// thanks for all your blogging!
- Ruchira S. Datta
reconstructing crispr loci in silico. i do not understand this method.
- Thomas Sharpton
@Ruchira - anytime!
- Thomas Sharpton
Debbie Lindell - Phages are a Selective Force Driving Cyanobacterial Genome Diversification #ASM2012
Ruchira S. Datta liked this
How do cyanobacteria coexist with virus that infect them?
- Thomas Sharpton
Prochlorococcus has syntenic regions (core) and variable regions
- Thomas Sharpton
T7-like Podoviruses are lytic dsDNA viruses that infect Prochloro
- Thomas Sharpton
Isolated resistant strains and characterized mutations and phenotypes
- Thomas Sharpton
Found mutations in 24 genes, most are cell-surface related
- Thomas Sharpton
viruses had impaired ability to invade the cells
- Thomas Sharpton
// evidence of co-evolution?
- Ruchira S. Datta
all mutations were non-synonymous in predicted coding frames; indels leading to frameshifts. often, multiple different types of mutations in genes. most were non-core genes (83%)
- Thomas Sharpton
@Ruchira - I'm waiting for it, but nothing so far
- Thomas Sharpton
Mutations clustered in genomic islands.
- Thomas Sharpton
Suggests selection against cells carrying attachment genes, selective force enhancing diversity in gene content
- Thomas Sharpton
mutant genes are rare in the environment. however, sequence diversity is higher when present when compared to core background
- Thomas Sharpton
Identify populations with diverse cell-surface genes. many subpopulations with different susceptibility regions.
- Thomas Sharpton
Why aren't all cells resistant? These mutations are associated with a growth rate cost. Also, greater susceptibility to other phages. Avrani et al 2011 Nature 474: 604-608
- Thomas Sharpton
Can the slower growers maintain a decent population size in the natural environment?
- Thomas Sharpton
I missed something or she lost me.
- Thomas Sharpton
Anyhow, summary: Viruses drive diversification of genomes through selection for resistant strains. fitness cost to resistance. Complex dynamic network of interactions.
- Thomas Sharpton
Martin Polz - Ecology and Evolution of Bacterial Populations
Ruchira S. Datta liked this
Microbial diversity is vast: vast in terms of the number of OTUs in the environment, vast in the pan-genome size of closely related strains
- Thomas Sharpton
What are ecologically and evolutionarily cohesive units (populations)?
- Thomas Sharpton
If we are interested in functions of a community, how much diversity do we need to consider?
- Thomas Sharpton
He works with vibrio as a model. Easy to culture. But not very abundant. But pretty large. Very diverse, even at the phenotype level.
- Thomas Sharpton
Obtain isolates from potential microhabitats, fine-scale phylogeny (protein coding genes), modeling: population hypothesis (genomic and physiological properties <-> environmental and environmental manipulation)
- Thomas Sharpton
Ecological population cohesion: association with different types of suspended particles. Builds a tree of sequences obtained from various ecosystems, maps ecological data on the tree, cultures some of the isolates and places them on this tree and makes inferences about evolution ecology/physiological among the group
- Thomas Sharpton
Identified several nested clades of low divergence but clear differences in environmental preference.
- Thomas Sharpton
sequenced genomes from these organisms. protein coding genes very similar, consistent with 16S tree. But, are all loci in genome really consistent with this tree? Has there been recombination among strains?
- Thomas Sharpton
Only identify a few recombination blocs that support ecological splits
- Thomas Sharpton
Shapiro Science 336:48 2012
- Thomas Sharpton
He calls these 'islands of differentiation', which contain genes of potential ecological relevance. Some appear to be involved in biofilm formation.
- Thomas Sharpton
What about recombination within v. between habitats?
- Thomas Sharpton
Finds restricted recombination between habitats compared to within.
- Thomas Sharpton
Also finds restricted gene flow between habitats
- Thomas Sharpton
Proposes a model of speciation. ancestral populations are assumed to occupy same habitat. genes come in from outside and trigger habitat differentiation. this in turn limits recombination between strains, increasing divergence between lineages
- Thomas Sharpton
Rebecca Mueller - Forests, farms and fungi: the impacts of deforestation on soil fungi #ASM2012
Ruchira S. Datta liked this
Yay! A Fungi talk!
- Thomas Sharpton
// fungus are among us
- Ruchira S. Datta
Slide showing that humans impact biodiversity. Largest effect? Land-use change
- Thomas Sharpton
Deforestation in the Brazilian Amazon. Prior work (gibson 2011) finds that the loss of primary forest is significant. Only have evaluated macroscopic organisms at this point.
- Thomas Sharpton
Amazon Rainforest Microbial Observatory is being highlighted. I want to go to there.
- Thomas Sharpton
// too many mosquitoes!
- Ruchira S. Datta
@Ruchria - Fair point. I hear a drier sheet in your back pocket helps
- Thomas Sharpton
// never heard that one before, thanks
- Ruchira S. Datta
Used a spatially-explicit sampling design to study alpha and beta diversity. Used Illumina to target and sequence ITS region of fungal genomes
- Thomas Sharpton
Richness is increased in pastures compared to forests. secondary forests also lower richness relative to pasture site
- Thomas Sharpton
Most of the change seems to be among Ascomycota (up in pasture) and Basidiomycota (down in pasture)
- Thomas Sharpton
Now introducing AM fungi, noting that they affect ecosystem function
- Thomas Sharpton
AMF are the dominant association in most tropical forests
- Thomas Sharpton
Focusing just on the AMF, finds that richness increases in pastures (as above)
- Thomas Sharpton
However, in pastures, there is no significant relationship between geographic distance and community diversity. In forests, there is.
- Thomas Sharpton
Community composition shifts with deforestation
- Thomas Sharpton
Similar response of bacterial communities between sites.
- Thomas Sharpton
Local richness was higher in pastures, but turnover was lower. AMF diversity may decrease over larger scales in response to deforestation.
- Thomas Sharpton
Cheryl Andam - Evolutionary divergence and biogeography of Streptomyces
Ruchira S. Datta liked this
Gram+ Actinbacteria, forms mycelium and dessication resistant spores
- Thomas Sharpton
High amoung of horizontal DNA exchange.
- Thomas Sharpton
Introducing everything is everywhere, habitat filtering, and dispersal limitation and regional diversification hypotheses
- Thomas Sharpton
Isolated 924 strains from 15 sites across the US. 4 soil variables (pH, moisture, temperature, soil order) , rpoB gene analysis
- Thomas Sharpton
Using tool called AdaptML. from the Alm lab: http://almlab.mit.edu/ALM...
- Thomas Sharpton
Regional populations differ in the representation of various strains
- Thomas Sharpton
Therefore, everything is NOT everywhere
- Thomas Sharpton
Phylogenetic clustering of various sites indicates habitat filtering or dispersal limitation and regional diversification
- Thomas Sharpton
Genetic differentiation is positively associated with geographic distance, number of shared sequences between sites decay with distance. This suggests that habitat filtering is not valid.
- Thomas Sharpton
Using haplotype analyses to evaluate the expansion of strains throughout the US and understand their biogeography
- Thomas Sharpton
Hypothesizes that glacial retreat influenced the ecological evolution of soil bacteria, enabling their rapid expansion
- Thomas Sharpton
// I went to a microbial ecology talk just a couple of years ago where the presenter very hesitantly suggested that everything might not be everywhere, as if it were a heresy...
- Ruchira S. Datta
@Ruchira - Funny how things change (quickly). This conference may not have killed baas-becking, but certainly raises doubt about that hypothesis
- Thomas Sharpton
Jed Furhman - Temporal patterns of microbes #ASM2012
Ruchira S. Datta liked this
Interested in understanding temporal patterns of free living marine planktonic bacteria and understanding influence of seasonality
- Thomas Sharpton
// Like Eric Alm?
- Ruchira S. Datta
Gets to study Catalina island!
- Thomas Sharpton
@Ruchira - yes, similar questions :D
- Thomas Sharpton
Finds a strong relationship between temperature and diversity. Notes that huge variation along depth, a few meters down is like many hundreds of meters along surface. Looking mostly at surface here.
- Thomas Sharpton
Sees similar diversity variation among just Chlorophyll
- Thomas Sharpton
Rare bacterial taxa can be more variable over days at one location
- Thomas Sharpton
This isn't sequence based analyses, but ARISA
- Thomas Sharpton
Finds that most days, things don't change that much. But there are some days where there are sudden transitions in composition
- Thomas Sharpton
Discussing Fuhrman et al PNAS 2006
- Thomas Sharpton
Using a discriminant function finds that you can predict the calender month of the year given the microbial diversity profile with 100% accuracy (!).
- Thomas Sharpton
// ARISA = Automated Ribosomal Intergenic Spacer Analysis
- Ruchira S. Datta
This suggests that that bacteria are not functionally redundant. This challenges the ecological redundancy hypothesis.
- Thomas Sharpton
// Cool!
- Ruchira S. Datta
Majority of taxa fit into function, but it will remove non-predictive taxa. ...overfitting? probably dealt with this.
- Thomas Sharpton
//? is the discriminant function regularized?
- Ruchira S. Datta
Don't know. Good question.
- Thomas Sharpton
What about seasonality of viruses? Chow and Furhman 2012 Environmental Micro
- Thomas Sharpton
Found that viruses are also strongly predictive of calendar month
- Thomas Sharpton
Using tools that consider the entire community to evaluate temporal variation (even non-predictive taxa). Found strong 6 month sine pattern of diversity across a many year study
- Thomas Sharpton
If you go down 15-30 meters, this pattern disappears
- Thomas Sharpton
Looking at T4 viruses finds a similar trends
- Thomas Sharpton
Implies a consistent connection between the core and accessory genomes. But it does not say there is only one accessory genome connected to each core genome time. Also suggests there are not rampant nice-altering mutations, gene losses or gains that sweep most of these populations on time scales of a few years.
- Thomas Sharpton
Looking at association networks to identify organisms that distribute through time together
- Thomas Sharpton
Showing a huge network, looking for highly connected nodes. Looking at SAR11 as an example. Connected to a lot of different types of organisms. Missed the details...
- Thomas Sharpton
Some clusters in network are made up of highly connected sets of nodes.
- Thomas Sharpton
Some of these tight clusters may be driven by seasonality (all up together or down together)
- Thomas Sharpton
// network movie?
- Ruchira S. Datta
@Ruchira Nope. Static images.
- Thomas Sharpton
Are able to identify nodes that represent viruses and identify which phage interact with which bacteria
- Thomas Sharpton
"These are mathematical correlations, not direct interactions". Lots of different things could cause these relationships.
- Thomas Sharpton
He discusses these topics and their implications in Fuhrman, Nature 459:193 (2009)
- Thomas Sharpton
Larry Forney (U Idaho): vaginal microbiota in health and disease #asm2012
Ruchira S. Datta liked this
Vaginal microbiome is remarkable. Bacteria in vagina are directly nutrified by the host-- very different than gut. Identify 5 community "states". Believe that function of these must be conserved. Prospective longitudinal study: have samples before during and after major events.
- Erick Matsen
Menses-- example of microbiome change in response. But trickier than that. Each of the temporal patterns are different from one another. Seems correlated with sexual activity, but responses are very individual. See complete turnover, even in 24hr period.
- Erick Matsen
Profile of community state types. Describe 5 types of transition types. Modeling in terms of hormone cycle, and can see a difference with hormones.
- Erick Matsen
Transcriptome analysis with yeast infections and BV. Clinically diagnosed yeast infection -> look at transcriptome. Species specific changes-- looking at changes in pathways. Future directions include clarifying entry point and exit conditions from disease states.
- Erick Matsen
Their sampling: 135 women daily for 3 weeks.
- Erick Matsen
Janet Jansson (LBNL): Impact of perturbations on the human gut microbiome #asm2012
Ruchira S. Datta liked this
Will be describing perturbations, e.g. antibiotics and disease Describing 2007 clindamycin study, Jernberg et al 2007 ISMEJ. Long term impact of antibiotic use. One clone of bacteriodes that dominates community, resistant to antiboitics. Cites Jernberg et al 2010 microbiology.
- Erick Matsen
Crohn's disease. Swedish twin cohort. Healthy twin pairs, those having UC, and Crohn's. Looking at microbiome, metagenome, meta-transcriptome, meta-proteome, and meta-metabolome. 16s data. See differentiation between UC, Ileal CD, and Colonic CD. A major difference between Faecalibacterium prausnitzii. Metaproteomics. Can also see functions differentiating. Get bacterial proteins and...
more...
- Erick Matsen
Restoration from a perturbed state? Can we restore? C. diff fecal transplant story. Profile her community before and after. Before treatment, very strange community. The recipient very closely matches donor's profile. Cure! Idea of alternate stable states and basins of attraction.
- Erick Matsen
Heidi Kong (NIH): Temporal shifts in the skin microbiome are associated with disease flares and treatment in children with atopic dermititis #asm2012
Atopic dermititis = excema. 15% of kids get it. Disease flares are associated with colonization of Staph aureus. Patients Atopic march: AD in early age -> hay fever -> asthma. Have shown that skin exposure to antigens -> mucosal sensitiization Understanding triggers of AD may allow us to modify the development of AD and other atopic disease. Triad of AD: skin barrier (mutations filaggrin), immune system (elevated IgE, AMP), Microbes (S. aureus) In skin, a lot of hetero WRT location. Protocol-- find AD patients, sample inner elbow, nare.
- Erick Matsen
12 patients, 11 controls. Baseline, flare, and post-flare. Sanger sequence 16s. Inverse correlation between Shannon and the disease severity. In particular, when no treatment and flare, low diversity. Analyze additional flares. Able to predict lower versus higher diversity.
- Erick Matsen
What is accounting for the decrease in diversity? All Staph spp. Speciate staph species. Higher prevalence of S. aureus during flares. Also see increase in S. epi.
- Erick Matsen
Tom Sharpton @tjsharpton (UCSF): Novel bacterial taxa in the human microbiome
Ruchira S. Datta liked this
Collaboration with Weinstock. Have identified 1000s of microbes associate with gut. HMP got a whole bunch of new sequences-- 454 and Illumina. Can we identify novel bacteria? Compare 16s to known 16s sequences. Filter reads with 97% identical match to get putatively novel 16s reads.
- Erick Matsen
Sequencing error a problem. Each putatively novel 454 read has to have coverage with Illumina sequences. If we approach a chimeric junction, we will see a dip in the maximum length of illumina read by base position. Go from 300K reads initially -> 10k putatively novel -> 2k reads covered by illumina -> 141 validated novel reads
- Erick Matsen
Who are these novel organisms? Use his PhylOTU, then place into SILVA tree. Tenercutes, Firmicutes, Bacteroidetes, and Proteobacteria. Firmicutes, Bacteroidetes are the majority. Novel OTUs are closely related to uncultured organisms-- have seen them somewhere before. However, some of them are quite different.
- Erick Matsen
Several novel low-abundance OTUs exist in the human gut. However, see one that is found in lots of samples. This is related to Barnesiella, and abundance on the same order of E. coli. Cool!
- Erick Matsen
Rick Bushman (Penn): Dynamics of the human gut virome #asm2012
Ruchira S. Datta liked this
Looking for viruses. Take stool samples, sequential filtration, cscl banding, nuclease treatment. Yield 10^10-10^11 vlp's per gram of input (!) Only get DNA viruses. Overwhelmingly bacteriophages. 454 0.5M reads (Minot et al 2011, Genome Res) HiSeq 40M reads (Minot et al 2012, PNAS) Lots of variation within the population.
- Erick Matsen
Going to talk about HiSeq dataset. 12 human samples, assemble with SOAPdenovo. Get 1000x coverage for somve viruses some very long. Annotating genes: the usual suspect, RT genes, resistance. Only found one known virus: a papilloma virus.
- Erick Matsen
Great majority completely new. Very little overlap between people. We know that some of the genes look like phage genes. (This is actually a guess). Cmparative metagenomics: viruses are parasites, because bacteria devote most of their coding capacity to replication. Can traverse along genomes and see variation. Once in a while, see region where majority of bases different from one another. These seem to be ~100bp regions. Found 29 such regions. Hypervariable region. Near to RT gene.
- Erick Matsen
Have characterized one of these. Work of Jeff Miller. The major tropism determinant of phage BPP-1. Phage targts a very diverse gene at tip of fiber strand. So this hypervariation is to extend range of binding. Requires RT, repeats. We see a number of other genes that have this sort of system. Convergent evolution with VDJ! Ig superfamily folds, using combinatorics to generate diversity. AAY codons are especially used, avoiding being one jump away from stop codons.
- Erick Matsen
Clearly an important role for RT in gut bacteriophage. Great majority or RTs in their dataset are in a single clade! See very extensive targeted hypervariable regions. Beautiful slide of T4 hoc, a dispensible head protein, ligand for binding. Paper: Fokine et al 2011
- Erick Matsen
Q & A session from microbiome speakers #asm2012
Ruchira S. Datta liked this
Q: We've been studying microbiome for years, but lots of new info. 1. What are the big questions that we can address right now that we couldn't address a couple of years ago? 2. What are the big questions we still need tools for?
- Erick Matsen
A Rob: We just saw that in the talks. A Forney: looking at functions. Conservation of function. A Jansson: we need to have to have more well-annotated genes. We are finding lots of unknown genes. Rob: we really need to bring knowledge together and making knowledge widely available. Earl: we need to do bichem, etc. Huttenhower: we would like to do these sorts of biochem studies on whole communities.
- Erick Matsen
Q: We have a lot of correlation. What about causation? How can we use models?
- Erick Matsen
A: Rick: intervention analysis. Forney: there are things that we can do without understanding mechanisms, so statistical correlations still important. Can use correlations to identify more specific tests. Rob: time series and prospective longitudinal studies are key. Also gnotobiotic animals.
- Erick Matsen
Q: functional pathways seem to be conserved, so how do we go about restoring ecosystems?
- Erick Matsen
A Rob: Guild structure-- what can substitute for what? A Sharpton: building model environments inside of model orgs. A Rob: fine tuned manipulation.
- Erick Matsen
Q: This is a dialog within the host. What comes next for host-microbe interaction research?
- Erick Matsen
A Jansson: can already see it in proteomes. That's not contamination! In addition to inoculating model orgs with microbes, can also give them enzymes and metabolites.
- Erick Matsen
A Huttenhower: can learn a lot from nutritional studies. A Forney: lots of strands-- access to medical treatment, nutrition. Start to think as ecologists. A Kong: Host immune system is different between locations. A Earl: spatial variation.
- Erick Matsen
Q: In the context of large projects, there is an increasing drive towards translation. But pure science is also important! What is the role of these two things.
- Erick Matsen
A Rob: we've just looked at the moon, and there's a lot more to be known. A Bushman: How much does the NIH need to fund translational research, because pharma is very good at marketing things. A Forney: room at the table for all-- not everyone has to do translation.
- Erick Matsen
Ashlee Earl (Broad Inst): The "most wanted" bacteria from human microbiome. #asm2012
It's the reference genomes that give us insight. 5 microeuks, 11 viruses/phage 3 archaea. 1654 bacteria. That's a reasonable fraction of the sequenced genomes. HMP has contributed > 400 unique bacterial species. 70% of all human-associated genomes.
- Erick Matsen
Lots of thought going into what strains important. Allen-Vercoe, Izard, and Dewhirst get "platinum status".
- Erick Matsen
How good is our reference genome collection? Now can recruit 57% of a metagenome. 1. work harder for isolation/single cell. 2. do better finding most wanted.
- Erick Matsen
First definie reference data set. 55M 16s reads. Simplify- cluster with AbundantOTU at 97% to create HMP OTUs. Remove chimeric OTUs with UChime, Characterize with RDP, then align.
- Erick Matsen
For each of these OTUs, how often do we see it, and how different is it from what we already have in DBs. More than 60% of HMP OTUs have less than 97% overlap with a sequenced genome.
- Erick Matsen
119 most wanted orgs. They are in all different body habitats.
- Erick Matsen
Curtis Huttenhower (Harvard): Identifying species, strains, and functional roles in the microbiome #asm2012
Ruchira S. Datta liked this
Nicolas Segata. Start contextualizing some of the new bugs in the environment. Using IMG 3,100 bacterial genomes, as well as other domains. About 10M genes.
- Erick Matsen
1. Identity conserved markers that can be used to infer phylogeny
- Erick Matsen
Using 500 markers that are most phylogenetically informative. We can use the placement of these trees to annotate new trees, however, 33 misannotations.
- Erick Matsen
Says "high confidence", but concatenation well known to lead to high confidence incorrect splits.
- Erick Matsen
MetaPhlAn: Metagenomic phylogenetic analysis. A gene considered to be "core" if in all of the reps of a species. Then look for genes that are markers for a species. We can use this to species.
- Erick Matsen
Very fast, able to run on whole HMP. Major driver is body site. Three different oral communities segregate. He is stating that there are four different species-level vaginal microbiome types.
- Erick Matsen
That's not what we see in gut. Also works great for archaea, euks, and DNA/RNA viruses too. Can run several GB of data in minutes. Don't need cloud.
- Erick Matsen
HUMAnN: quanitfying microbial community function with the HMP unified metabolic analysis network. Identifies pathways.
- Erick Matsen
There are functional differences, certainly, and he has a nice tool for showing differences of abundance. Take home: "most processes are core, and most processes are habitat-specific."
- Erick Matsen
Can do metatranscriptomics as well.
- Erick Matsen
IBD as defined by perturbations. What is causing or correlating with disease? Also can see treatment of effect. Four major metabolic areas that are particularly changed in Crohn's/IBD.
- Erick Matsen
2% organisms significantly different. 12% significantly pathways significantly different.
- Erick Matsen
Lita Proctor (NIH): A brief overview of the human microbiome project #asm2012
Ruchira S. Datta liked this
Lots of sequence data, clinical/phenotype data, reference strain cultures and also cell lines.
- Erick Matsen
Clinically healthy cohort study. 300 m/f. 18-40 yo, "healthy", up to 3 visits in 2 years.
- Erick Matsen
Demo projects: skin, GI, and urogenital.
- Erick Matsen
Rob Knight up first.
- Erick Matsen
Characterize the healthy human microbiome. Has produced a vast dataset.
- Erick Matsen
Interesting conclusion: huge differences in lineages, but consistent function.
- Erick Matsen
HMP was able to confirm this trend across body habitats.
- Erick Matsen
Describing procrustes analysis about mapping metagenomic samples to 16s samples. But the HMP covers only the start of human diversity. Ages, populations, health status.
- Erick Matsen
Showing how rate of gut microbes approaching adult state is consistent across different cultures, although they have different endpoints.
- Erick Matsen
QIIME in the cloud. Antonio Gonzalez has been heading up the effort of data integration.
- Erick Matsen
Are there enterotypes? No support for them in HMP + community data. Clustering is weak, and gradients explain better (ditto with other sites).
- Erick Matsen
Re-examination of original data. Taking the lines an ovals out of their figure 1. No evidence of clustering.
- Erick Matsen
Infant gut microbiota changes radically-- integrate with adults? Take infant time series-- how does this relate to the other gut communities that are in the HMP. That's great work.
- Erick Matsen
Animation with ball moving around showing how things progress to adult state.
- Erick Matsen
Study effects are small compared to the between-site differences. (But that's not a very high bar to clear).
- Erick Matsen
But now showing difference between data based on primer sets. However, still separates out ulcerative colitis from healthy.
- Erick Matsen
Maria G. Dominguez-Bello - The Microbiome of Isolated Peoples #ASM2012
Ruchira S. Datta liked this
Going to discuss the microbiome and vertical transmission
- Thomas Sharpton
Helicobacter pylori will be on stage. Found in stomach, plays role in gastritis and peptic ulcer disease
- Thomas Sharpton
Study (collaboration with Blaser) comparison of HP from East asia, Europe, isolated Venezuela, Caracas.
- Erick Matsen
Distribution of H. pylori across people of various continents finds that Amerindians split between European and Asians. Congruent with Asian origin of Amerindians
- Thomas Sharpton
H. pylori ancestral nucleotides reflect human migrations. Falush 2003 Science
- Thomas Sharpton
She says that HP not a pathogen, rather a commensal.
- Erick Matsen
## Indeed, Blaser says HP prevents esophageal adenocarcinoma
- Ruchira S. Datta
Do we see signals of adaptation and selection in our microbiome?
- Thomas Sharpton
Hugenholtz paper (not cited) illustrates the concordant evolution between primates and their microbial communities. This indicates that vertical transmission occurs in microbiota.
- Thomas Sharpton
As infant mice age, they move from having community profiles that look like mother's vaginas to mother's intestines
- Thomas Sharpton
http://www.plosbiology.org/article... <- there's the citation.
- Erick Matsen
@Erick - Thanks!
- Thomas Sharpton
Implications: mammals are born naturally colonized and vulnerable to colonization (high Tregs). The first colonizers must initiate the education of the immune system. Different pioneer microbiota may have important health implications. She thinks that drinking milk is a very important part of shaping microbiota.
- Thomas Sharpton
## naturally colonized? but per previous talks, sterile?
- Ruchira S. Datta
That's the first time I've seen that Treg comment. Wonder what the evidence is (it's prob there, but don't know).
- Erick Matsen
Rescigno 2009 Biol Rep -> models of immune system fine tuning of the microbiota to regulate homeostasis
- Thomas Sharpton
Thanks!
- Erick Matsen
Western lifestyle impact on the human microbiome: she proposes a model about how our lifestyle has led to microbiome changes that produce chronic diseases
- Thomas Sharpton
Comparing lifestyle differences of living in new york city with indigenous tribes in unimpacted environments
- Thomas Sharpton
looking in amerindians, characterized the oral microbiota of these individuals
- Thomas Sharpton
#? Milk drinking has been under recent positive selection in human subpopulations, so if that's important, is it really comparable?
- Ruchira S. Datta
very important differences between groups (amerindians and nonamerindians), finding organisms that hadn't been previously identified in the mouth as well as taxonomic shifts
- Thomas Sharpton
#? Or maybe she just meant breast milk...
- Ruchira S. Datta
@Ruchira - I think she just meant breast milk postpartum, not long term lactose tolerance
- Thomas Sharpton
@tsharpton Thanks
- Ruchira S. Datta
All amerindians group together and separate from nonamers, but there are two distinct amerindian groups
- Thomas Sharpton
I wonder how much of the lifestyle effect she is finding is simply a result of routine access to medicine
- Thomas Sharpton
## Which amerindians are "isolated" and which not?
- Ruchira S. Datta
Oh, that's a good question
- Thomas Sharpton
Found differences in richness, 64 highly predictive OTUs, most were underrepresentative in US, most were Prevotella
- Thomas Sharpton
Also found differences in genes associated with vitamin metabolism. I don't know which group is enriched.
- Thomas Sharpton
Recently have been characterizing the oral and skin microbiota of Amerindians that have no previous recorded contact. So far have not found antibiotic resistance in E. coli among these individuals.
- Thomas Sharpton
Pretty wild. "I am from outer space. I want your poop."
- Erick Matsen
@Erick - That almost made me laugh out loud during her talk
- Thomas Sharpton
Studying the San bushmen. Interesting. I'm studying the San as well....
- Thomas Sharpton
She's suggesting restoration ecology of human microbiome.
- Erick Matsen
Lu Zhang - Antibiotic Resistance in Non-Food Animals: Impact of Animal Hosts on the Resistance Ecology
Ruchira S. Datta liked this
The rapid emergence of antibiotic resistance (ART) bacteria is a major public health concern
- Thomas Sharpton
Not just used in clinics, but also during food animal production. Government agencies are stepping up efforts to regulate use of antibiotics during food production.
- Thomas Sharpton
Wow. 28.9 million pounds of antibiotics are consumed by animals in the US annually.
- Thomas Sharpton
#!!!
- Ruchira S. Datta
Recent studies suggested that AR persists even in the absence of antibiotic selective pressure.
- Erick Matsen
Exposure to antibiotics contributes to the evolution of ART commensals
- Thomas Sharpton
Can we find ART bacteria in non-food animal's GI tract?
- Thomas Sharpton
i.e. pets and zoo animals.
- Erick Matsen
He notes that there are lots of pets, but I wonder what percentage are treated with antibiotics? I'm guessing is not insubstantial
- Thomas Sharpton
Finds phenotypic resistant population in non-food animal feces.
- Thomas Sharpton
Trying to identify the AR gene carriers in non-food animals. Findings are limited by difficult cultivation conditions
- Thomas Sharpton
This work identifies ART bacteria and gene pools in non-food animals rarely exposed to antibiotics (in agreement with previous studies)
- Thomas Sharpton
Elizabeth Costello - Human Microbiome Development Across Multiple Body Sites in Healthy Infants: Assembly and Re-assembly Following Antibiotic Therapy #ASM2012
Ruchira S. Datta liked this
Postdoc in David Relman's lab at Stanford
- Thomas Sharpton
Neonatal microbial exposure can protect mice against asthma and colitis. Infant derived microbiota protect mice against lethal infection
- Thomas Sharpton
Antibiotic perturbation in the context of early microbiome development: an ongoing study of healthy mother-infant dyads. Routine baseline sampling of infant microbiomes. Antibiotic exposures.
- Thomas Sharpton
Collect infant fecal, saliva and skin swabs on the same days each month. Also, collect maternal fecal, saliva, skin, vaginal and breast milk samples collected at several postpartum months
- Thomas Sharpton
## Interesting to know how maternal microbiome changes from T3
- Ruchira S. Datta
Applying more or less the same data generation process outlined by Rob
- Thomas Sharpton
@Ruchira I agree! but they are only looking postpartum
- Thomas Sharpton
Summarizing the major phyla that are found in their initial samples across ages and sights. Nothing surprising in her assessment
- Thomas Sharpton
Talking about a few specific individuals. Looking at change in taxonomic composition over time. When solid food is added, diversity profile shifts.
- Thomas Sharpton
Saliva and skin have similar development structures
- Thomas Sharpton
Preliminary findings suggests that antibiotics administered for ear infection has an effect on the taxonomic diversity. Some resilience in the system.
- Thomas Sharpton
Stay tuned... (notes were a bit muted because this is all pre-pub).
- Erick Matsen
Rob Knight - Evolution of Mammals and their Gut Microbes
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This is work he's done with Ruth Ley, who couldn't make this year's meeting.
- Thomas Sharpton
His talks are fast, here goes!
- Erick Matsen
How can we develop technologies that allow us to manipulate our microbiota
- Thomas Sharpton
Need to first know how microbes are distributed across our bodies.
- Thomas Sharpton
How can we translate among human and animal models
- Thomas Sharpton
How much can our microbes change?
- Erick Matsen
How much can our microbes change over time (plasticity)?
- Thomas Sharpton
High levels of diversity even from the earliest samples (Eckberg 2005).
- Erick Matsen
Eckburg Science 2005 - High levels of diversity initially seen in the gut has been confirmed with more subjects.
- Thomas Sharpton
MetaHIT shows even "core" species varying in abundance by orders of magnitude in different people.
- Thomas Sharpton
The HMP has shown that there is great diversity across the body as well.
- Thomas Sharpton
HMP has expanded this to lots of other body sites.
- Erick Matsen
How do we grapple with all of this data? Phylogenetics!
- Thomas Sharpton
Showing classic phylogenetic tree in Darwin's origin of species.
- Erick Matsen
Introducing UniFrac. I'm note going to write this out. See the Lozupone and Knight paper from 2005
- Thomas Sharpton
## fortunately the fast talks share content :-)
- Ruchira S. Datta
Used UniFrac to determine that fecal samples are OK for representing the gut. While the stool is a bit different than particular gut locations, see the same co-clustering across samples.
- Thomas Sharpton
UniFrac show clusters by diet, gut type and lineage. Got to go to the zoo to collect samples.
- Thomas Sharpton
Observe clear clustering by diet across various mammals.
- Thomas Sharpton
There are two types of herbivores - the foregut and hindgut fermenters
- Thomas Sharpton
However, they do not see clustering by provenance (zoo v. wild)
- Thomas Sharpton
A small number of phyla dominate these samples across species.
- Thomas Sharpton
Notes that there is convergent evolution in gut morphology in herbivores
- Thomas Sharpton
Co-differentiation of microbes and mammals in recent lineages. Two trees are superimposed on each other. One is the UniFrac community hierarchy and the other is the bear phylogeny. Agreement here is perfect
- Thomas Sharpton
Carefully chosen regions of 16S give same info as full sequence. Used UniFrac to verify that clipped regions of 16S give same clustering patter as full sequence. Some regions are NOT good.
- Thomas Sharpton
This led to development of QIIME, which enables integrative analysis of hundreds of samples
- Thomas Sharpton
He likes to use barcoding of 16S sequences to do a single 454 run across many samples
- Thomas Sharpton
Describing Procrustes, which matches multilevel data (e.g., 16S rRNA v. metagenomic)
- Thomas Sharpton
Although species are distinctive, functions are highly conserved. OTU network shows clear differentiation across samples. KOs, however, don't show separation. Everything shares the KO functions.
- Thomas Sharpton
Even at the genus level, there are very few taxids that are shared across samples, but KO's quite overlapping.
- Erick Matsen
Shows that direction of pathways is crucial. Knowing only what pathways are present may not be enough.
- Thomas Sharpton
^ still wondering about coarseness of the functional classifications.
- Erick Matsen
On to microbiome stability. Obesity has a relatively subtle effect, although we can distinguish lean from obese with 90% accurately. Discussing Dan Knights' machine learning work (random forests) on this subject.
- Thomas Sharpton
One striking physiological change is during pregnancy. Shows that gut microbial communities alter dramatically between T1 and T3. This one hits close to home.
- Thomas Sharpton
91 women in cohort.
- Erick Matsen
Comparing to background individuals (not pregnant) finds that T1 women look more like background, while T3 looks very different.
- Thomas Sharpton
T3 has more opportunistic pathogens???
- Thomas Sharpton
More inflammation in T3 stool (IL levels increased)
- Thomas Sharpton
3rd trimester women have gut communities unlike anything previously seen in HMP
- Erick Matsen
Transfer of pregnant microbiota to germ-free mice. Results in physiological changes, including increase adiposity in recipients of T3 microbiota!
- Thomas Sharpton
We can understand how the microbiome develops in infants.
- Thomas Sharpton
Live demo time. I've seen him to this before; very brave
- Thomas Sharpton
Looking at infants across various mammals, finds that we start off similar to bear microbiota and then diverge away from them
- Thomas Sharpton
This is reproducible across multiple replicates and cultural settings.
- Erick Matsen
Also looking across cultures, though I missed the conclusion here
- Thomas Sharpton
US microbiota very different from amerindian microbiota.
- Thomas Sharpton
#? More similar to bear microbiota than some other mammals, or bear was just an example?
- Ruchira S. Datta
Erick is asking - I missed that detail if he said it during the talk.
- Thomas Sharpton
## thanks!
- Ruchira S. Datta
Rob says that they had most of the major mammals represented in the study, humans were closest to bears.
- Thomas Sharpton
## Thanks, that's kind of cool!
- Ruchira S. Datta
## I think bears are omnivorous
- Ruchira S. Datta
@Ruchira - You are correct. They are.
- Thomas Sharpton
Laura Cox - Early life microbiota shapes adult body composition #ASM2012
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All organisms born sterile, successive colonization of GI tract. Intestinal microbial can influence host physiology. For example, body fat accumulation.
- Thomas Sharpton
Sub-therapeutic antibiotic treatment (STAT) promotes growth of livestock
- Thomas Sharpton
STAT does not work with antifungals.
- Erick Matsen
Effect is strongest when given very early.
- Erick Matsen
Hypothesis: STAT changes early life microbiota and influences weight gain
- Thomas Sharpton
Mouse study finds an elevated early life growth rate in animals given STAT from birth, esp. in males
- Thomas Sharpton
This suggests that these four weeks are especially important
- Thomas Sharpton
Next question: is early life STAT sufficient?
- Erick Matsen
Yes. Just giving for first 4 weeks changes long-term fat composition.
- Erick Matsen
I think she assembled on PCR amplified 16S variable regions sequenced via Illumina. Chimeras?
- Thomas Sharpton
Maybe that doesn't matter: PCA analysis shows strong differentiation between high fat diet, normal chow and low fat diet (?)
- Thomas Sharpton
^ ahem, PCoA.
- Erick Matsen
Effect of STAT - antibiotics differentiate community composition from controls. Remove antibiotics from these mice results in a shift back to control
- Thomas Sharpton
@Erick duly noted. I know better.
- Thomas Sharpton
Does appear to be a continuum based on how long they are on microbes.
- Erick Matsen
Find that switch to high fat diet results in change in taxonomic composition (at phylum level). See same things with initiation of STAT and, as before, a rebounding of composition once STAT is stopped.
- Thomas Sharpton
These mice were obese.
- Erick Matsen
Can see similar trends at all ranks.
- Erick Matsen
Is it the antibiotics or the microbes? Is the STAT phenotype transferrable?
- Erick Matsen
Conducted microbiota transfer experiment (oral gavage) - mice that received STAT microbes gain more weight over time. This shows it's the microbes, not the antibiotics, that produce the effect.
- Thomas Sharpton
The donor and transfer samples look very similar.
- Thomas Sharpton
#? So the antibiotics kill off some microbes, permitting overgrowth of the STAT microbes?
- Ruchira S. Datta
Ruchira-- I don't actually know. I'll see if I can ask her.
- Erick Matsen
Alejandro Reyes - A Gnotobiotic Mouse Model for Characterizing Phage Bacterial Host Dynamics in the Human Gut #ASM2012
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Not going to talk too much about human gut microbiota. But, it is dominated by bacteria, few phyla, many species, great number of strains. Marked by interpersonal variation. Do phages play a role in modulating the structure of gut communities?
- Thomas Sharpton
Time course study of 12 individuals (4 MZ female twin pairs and mothers, sampled 3 times)
- Thomas Sharpton
Metagenomic analysis of fecal viromes from healthy humans suggests a temperate viral lifestyle
- Thomas Sharpton
80% unmapped reads!
- Erick Matsen
Most viral diversity not described previously. Virome dominated by phage. High abundance of prophages and temperate phages.
- Thomas Sharpton
What are the benefits of a temperate lifestyle in the gut?
- Thomas Sharpton
Benefits of a "temperate" lifestyle-- integrate into genome. Only comes out when stress signals.
- Erick Matsen
Horizontal gene transfer of genes, superinfection protection, host niche expansion, etc. Also, lysis can yield constant diversity dynamics
- Thomas Sharpton
But, we need a defined model of the human gut ecosystem to study phage-host dynamics
- Thomas Sharpton
Start with 15 prominent sequenced members of human gut microbiota into groups of 5 adult germ-gree mice.
- Erick Matsen
Introduced 15 prominant sequenced members of the human gut microbiota into groups of 5 adult germ-free C57BI/6 mice
- Thomas Sharpton
Once model bacteria assemble, add pool of previously characterized purified VLPs from 5 healthy adults
- Thomas Sharpton
Three sets of mice. Some get VLPs, some get heat killed VLP, some get no microbes + VLPs.
- Erick Matsen
Mice remained germ-free.
- Erick Matsen
Once assembled, add exogenous phage to model community. Assembly is robust, w/in and between treatment lines
- Thomas Sharpton
They identify novel viruses in this analysis.
- Thomas Sharpton
Viruses from stages VLP attack of model human gut microbiota change community strucure. Showing relative abundance plot of B. caccae. A strong decline in abundance as abundance of phage spikes
- Thomas Sharpton
Can see specificity between viruses and bacteria. Bacteria caccae and phage "C1c1".
- Erick Matsen
Assembled the viral genome, found terminases, helicases, DNA polymerases, carbo binding proteins and bacterial stress response transcription factors
- Thomas Sharpton
B. cellulosilyticus changes before VLP gavage. This drop is driven by prophage induction.
- Thomas Sharpton
They are able to induce prophages with diet switches! Between BK and Western diet.
- Erick Matsen
Induction of prophage observed with diet switches
- Thomas Sharpton
Seems like a powerful model and more complex than some of the polyassociation models I've seen in the past.
- Thomas Sharpton
Itai Sharon - Microbial species and strain variations during infant gut colonization #ASM2012
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Itai is from the Banfield lab at UC Berkeley. How many talks have we seen from that group so far? Impressive.
- Thomas Sharpton
Hrm...the sound needs to be checked on that mic.
- Thomas Sharpton
Discussing strain variation: shared core genome, varying strain-specific regions. Prochlorococcus is very abundant in oceans; the genome does not uniformly recruit reads from ocean metagenome.
- Thomas Sharpton
Some genomic islands have significantly decreased recruitment.
- Erick Matsen
Iwase et al Nature 2010 - Staphylococcus epidermidis inhibits Staphylococcus aures biofilm formation: some strains inhibit growth, while others don't. Strain level variation is important to understand.
- Thomas Sharpton
Metaphor of palm tree, trunk is core, leaves are the strains. Referencing Ley et al 2005, Ley et al 2006, Xu et al 2007
- Erick Matsen
What is the role of strain variation? May provide genomic flexibility to enable niche adaptation in reponse to selective pressures (Ley 2005)
- Thomas Sharpton
It is hard to study; culturing, 16S and traditional metagenomics are all limited here
- Thomas Sharpton
Genome extraction from the environment: metagenomic sequencing where they do an initial assembly followed by binning and larger assembly. Results in large contigs, though not always perfect.
- Thomas Sharpton
Now discussing the infant gut microbiome: a simple community that rapidly shifts in response to changing conditions
- Thomas Sharpton
citing Morovitz et al PNAS 2010. Shifts in community structure with respect with nutrition.
- Erick Matsen
Found that strain variation is present from early stages of gut colonization. Used the above approach on four metagenomic samples.
- Thomas Sharpton
The gut is a very complex community. I wonder what kind of fact checking they are doing on their assemblies to eliminate chimeras.
- Thomas Sharpton
Now discussing a time-series Illumina-based metagenomic study of infant gut colonization: 11 samples, 15-24 days after birth, found 6 complete, 2 near complete, 11 partial genomes
- Thomas Sharpton
Does complete mean "no gaps?"
- Thomas Sharpton
"Carrol study" (fake name). 11 samples, collected on days 15-24 after birth. Illumina HiSeq. 6 complete, 2 near complete and 11 partial microbial genomes as well as 4 complete viral genomes. Also detailed description of community dynamics.
- Erick Matsen
Doing strain-level binning using time series data.
- Erick Matsen
Data analysis: new approaches for metagenomic assembly, QC and binning. Conduct strain level binning based on scaffold abundance patterns. They call this ESOM (a Self Organizing Map procedure). This produces maps where dots represent DNA segments and dark regions show cluster boundaries.
- Thomas Sharpton
ESOM self organizing maps as previously described by Banfield. Sharon et al, submitted <- looking forward to reading!
- Erick Matsen
@Erick agreed!
- Thomas Sharpton
Finds that the community composition changes over time in their samples
- Thomas Sharpton
Child was by c-section. Can see lots of skin microbiome representation (P. acnes, etc.).
- Erick Matsen
Also, binning reveals multiple S. epidermidis strains. Total length of the core genome is ~2.2Mbp (~85% of genome size)
- Thomas Sharpton
S. epidermindis: two dominant strains, three infecting phages. Phages 013 and 014 infect strain 1, pahve 4-6 infects strain 3 (!!)
- Erick Matsen
Find 2 dominant strains, three infecting phages. Differential representation of each strain over time
- Thomas Sharpton
Strain representation correlate with various strains, suggesting interactions between particular phage and strains
- Thomas Sharpton
Identify several strain-specific genes, including phage resistance, drug resistance, biofilm formation, transporters
- Thomas Sharpton
Propionibacterium: two species, one becomes abundant while the other disappears over time.
- Thomas Sharpton
The skin bacterium P. acnes comes up again here
- Thomas Sharpton
New propi species!
- Erick Matsen
Shows that strain variation is present in the gut from early stages and that metagenomics can reveal it. Proliferation/decline of certain strains is probably due to a small number of genes.
- Thomas Sharpton
This species also appears to be metabolically different than its closest relative.
- Erick Matsen
Talk by Jeff Weiser (U Penn): ... and Immunity shapes microbes #asm2012
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Strep penumoniae. It is simultaneously an invasive pathogen and a commensal organism.
- Erick Matsen
Streptococcus pneumoniae is an invasive pathogen that colonizes the upper respiratory tract. Also a commensal organism.
- Thomas Sharpton
Leading cause of pneumonai, otits media, and meningitis. Has a big fat capsule, that keeps away the complement system.
- Erick Matsen
Developed a genomic screen for bacterial factors involved in resistance to OPH killing. Looking for mutants that are killed more in the presence of neutrophils
- Thomas Sharpton
They do see mutants that have to do with capsule. But there are some other mutants whose mechanism is not clear.
- Erick Matsen
A common theme among their mutants is that they all form streptococci of increased chain length
- Thomas Sharpton
Idea-- strep with a small size is a virulence determinant in Step pneu because it allows this pathogen to evade opsonophagocytotic killing.
- Erick Matsen
Using chemistry to manipulate length, and indeed longer -> increased killing.
- Erick Matsen
Or can go back to mutants and see the amount of C3 deposited and see that longer -> more complement.
- Erick Matsen
The longer the strep chains, the increased chance of the chain having complement on their surface. The amount of complement affects the propensity of immune killing.
- Thomas Sharpton
Found that decreased chain length promotes a competitive advantage in vivo. The longer the chain of the mutant, the less effective it was at invading a mouse host. In complement deficient mouse, this effect goes away.
- Thomas Sharpton
There may be size selection on pathogenic cells given this observation. Indeed, most pathogenic microbes are very small.
- Thomas Sharpton
Typically seen in clinical specimens in diplococcus. (Double-bacteria).
- Erick Matsen
Does the host exploit the fact that larger bacterial parcticles are more easily killed?
- Erick Matsen
Other side might be that smaller chains is better for immune system so it can agglutinate them with antibodies.
- Erick Matsen
Assessed impact of agglutination on OPH killing. Found it promotes complement-dependent but Fc-independent OPH killing
- Thomas Sharpton
Is this true of other organisms as well? Tested in H. influenzae and found that gllutination enhances complement mediated lysis here too
- Thomas Sharpton
Does bacteria does anything to counter this agglutination?
- Erick Matsen
Do bacteria counter host's antibody agglutination? Discusses Immunoglobulin A1 and how it has a hinge region that is targeted by proteases of successful pathogens
- Thomas Sharpton
Passive immunization experiment: type specific human IgA1 is not protective because pneumo expresses a protease to cleave.
- Erick Matsen
Finally, host has an antibody against this protease.
- Erick Matsen
Grow pneumo is human nasal surface airway fluid -> long forms, compared to in nutrient medium.
- Erick Matsen
Interestingly, finds that the long forms of pathogens increases adherence to host mucosal sites
- Thomas Sharpton
These longer chain forms have a relatively small advantage in colonization.
- Thomas Sharpton
#? Dynamic equilibrium -- compute optimal length of chain?
- Ruchira S. Datta
Charles Bevins - Nets and Harpoons Innate Immune mediators of Host-Microve Interaction in the Small Intestine #ASM2012
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Talking about defensins, evolutionary old family of effector molecules, wide range of activity. They form dimers, which allows one side to associate with microbial membrane
- Thomas Sharpton
Focusing on alpha-defensin 5 and 6, expressed by paneth cells in the intestinal crypts
- Thomas Sharpton
Paneth cells have been implicated in IBD pathogenesis, esp. Crohn's disease
- Thomas Sharpton
They express NOD2 and ATG16L1, susceptibility genes for Crohn's disease
- Thomas Sharpton
Cartoon model: sensing of noxious microbes -> secretion of defensins by patheth cells.
- Erick Matsen
Defensins help defend from pathogens and shape the microbiota
- Thomas Sharpton
Study these molecules in mice that don't express HD5
- Thomas Sharpton
HD5 protects from Salmonella typhimurium
- Thomas Sharpton
Add HD5 to mice protects against infection with Salmonella typhimurium.
- Erick Matsen
Another idea is that these defensins shape the microbiota.
- Erick Matsen
Salzman Nat Immun 2012 - comparing WT mice to HD5 mice finds that there is a relative increase in Bacteroidetes and a reciprocal decrease in Firmicutes
- Thomas Sharpton
Found complete absence of Segmented Filamentous Bacteria in HD5 mice. Candidatus arthromitus, colonize intestines of numerous species, typically adheere tightly to epithelium in ileum
- Thomas Sharpton
Th17 cells increased in mice with SFB
- Thomas Sharpton
Now on to HD6
- Thomas Sharpton
HD6 is, surprisingly, not antimicrobial! At least not Salmonella.
- Thomas Sharpton
HD6 is abundantly expressed and its protein sequence is highly conserved across the homonids, which differs from HD5
- Thomas Sharpton
100% conservation to chimps and gorilla!
- Erick Matsen
Suggests that HD6 plays an important role, though this role is unknown.
- Thomas Sharpton
Raises various hypotheses, and all were turned down by experimental data
- Thomas Sharpton
Story of graduate student working on this project. Tough times.
- Erick Matsen
HD6 mouse model challenged with Salmonella found that none of the HD6 mice died and after a few days, there were differences in the number of bacteria colonizing the host in the pyers patches and spleen
- Thomas Sharpton
This suggests that HD6 inhibits invasion
- Thomas Sharpton
Found that invasion of Salmonella and Yersina rates decreased as a function of HD6. These organisms invade in very different ways, suggesting that this is a general mechanism.
- Thomas Sharpton
Actually making "nets" to prevent invasion. How?
- Erick Matsen
HD6 forms serious multimerization.
- Erick Matsen
The targets for HD6 are flagella and fimbriae. They can form these nanonets ex vivo.
- Erick Matsen
So, in summary, HD5 = harpoons, HD6 = nets. Wow.
- Erick Matsen
Sam Miller (U Washington): Using bacteria to probe innate immune diversity #asm2012
Central hyp: pathogents hav driven evolution of the human genome. Bacteria are unique probes about mammalian processes because they have co-evolved.
- Erick Matsen
Pathogens have driven evolution of the human genome. Signatures of selection in the human genome are more common in genes involved in immune response.
- Thomas Sharpton
Epidemiological and genetic studies indicate that genotype influence risk of death from infectious disease
- Thomas Sharpton
Sorensen et al NEJM 1988.
- Erick Matsen
Investigate cellular infection phenotypes.
- Erick Matsen
"Cellular GWAS" in vitro. Reduces envornmental variation due to differences in dose, medical treatment, etc. Accessible to small labs.
- Erick Matsen
1. Measure phenotypic variation in susceptibility to infection
- Erick Matsen
Uses the HapMap to asses the functional genomics of variation in host response. These are parent-offspring trios that let us look at heritability of phenotype
- Thomas Sharpton
Remarkably, can use these cells in vitro and get same impact as in vivo.
- Erick Matsen
Hi-HOST: high throughput human in vitro susceptibility testing. studied ~400 human cell lines. Invade cells with GFP labelled bacteria, measure cytokines from these cells, stain, fix, sort and analyze for bacteria replication and phenotypes
- Thomas Sharpton
First application, Caspase-1 Salmonella-induced pyropotosis.
- Erick Matsen
Started with a quantitative read out for Caspase-1: Salmonella induced pyrotosis. Caspase 1 activations has been implicated in numerous inflammatory diseases
- Thomas Sharpton
This phenotype shows diversity in response. Shows a very small but consistent (?) heritable phenotype.
- Erick Matsen
First question is whether phenotype is well distributed (is it diverse). Showing a correlation between parent and offspring phenotype intensity. Not sure I believe that there is a correlation...
- Thomas Sharpton
Look for SNPs associated with cell death. 9 SNPs pass functional criteria.
- Erick Matsen
Genome-wide association of Caspase-1 inflammatory response: 1.6 M snps, 475 associated with cell death, 9 pass functional criteria, 7 pass initial validation (RNAi against nearby genes)
- Thomas Sharpton
7 SNPs pass initial validation-- RNAi against nearby gene induces cell death.
- Erick Matsen
Variation in Salmonella-induced cell death: CARD8. Shown to bind Caspase1 and inhibit its activation when overexpressed in 293s or THP1s
- Thomas Sharpton
This is Razmara et al 2002.
- Erick Matsen
CARD8 allele is an important and functional. Important for response to Salmonella flagellin.
- Erick Matsen
Functional consequences of common CARD8 dysfunction allele includes increased inflammatory response, severity of rheumatoid arthritis and risk of SARS
- Thomas Sharpton
A SNP near the APIP gene is associate with pyropoptis. APIP= APAF1-interacting protein.
- Erick Matsen
One SNP identified is associated with APIP, a pyroptosis gene.
- Thomas Sharpton
pyroptosis is rapid and inflammatory, unlike apoptosis
- Thomas Sharpton
Model for APIP SNP action is proposed. Too quick for my fingers.
- Thomas Sharpton
APIP SNP also associated with increase apoptosis in response to caboplatin (cancer drug).
- Erick Matsen
Finds evidence that APIP inhibits pyroptosis when conducting an RNAi knockdown of the gene
- Thomas Sharpton
Shows that APIP inhibits cell death, but it's not clear if it is direct or indirect inhibition
- Thomas Sharpton
APIP has homology to the 3rd enzyme in the methionine salvage pathway, characterized in microbes.
- Erick Matsen
Methionie salvage pathway has only been described in Bacillus and yeast (not sure what yeast he is referring to). Found that APIP is required to utilize MTA as a methionine source in human cells.
- Thomas Sharpton
I wonder if screens of metagenomes associated with these samples could identify the MTA pathway and, possibly, the organism responsible for this activity?
- Thomas Sharpton
This suggests that APIP might deplete MTA and control cell death rates. Suggests that there may be a nutritional switch during infection. Early, maximize bacterial clearance (deplete Methionie), late infection...missed the rest.
- Thomas Sharpton
The APIP high pyroptosis allele is associated with decreased culture positivity for SIRS
- Thomas Sharpton
Asks if there is an optimal inflammatory response in humans
- Thomas Sharpton
What would be the natural selection for this response and is there evidence for it? As our population density increased, we accumulated 'crowd diseases', which would result in polygenic selection for decreased CARD8 response (did I get this right? was fast on the end).
- Thomas Sharpton
Finds frequency of allele is relatively low in hunters and gatherer populations
- Thomas Sharpton
Sarkis Mazmanian - Learning to tolerate our microbial self
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Dysbiosis during inflammatory bowel disease (IBD). Germ-free animals do not develop colitis and immune reactivity to commensal microbes due to defects in regulation. Observe alterations in the microbiota of IBD patients. Two major phylotypes are found in the healthy gut (Bacteriodetes and Firmicutes), but many phylotypes show up in diseased individuals.
- Thomas Sharpton
reference Lee and Mazmanian, Science 2010
- Erick Matsen
These extra phylotypes are Proteobacteria and Firmicutes.
- Erick Matsen
Bacteroides fragilis is a model organism that they study. gram negative, obligate anaerobe, prominent commensal in human gut, synthesizes at least 8 unique capsular polysaccharide complexes from distinct genomic loci. at least two of which have a novel zwitteroinc structure (PSA and PSB).
- Thomas Sharpton
PSA protects from chemically induced intestinal inflammation
- Erick Matsen
Use TNBS to induce inflammation, PSA rescues gut towards an untreated condition
- Thomas Sharpton
TNBS produce IL-17
- Erick Matsen
Colitis is driven by pro-inflammatory T-helper cells and elevated IL-17/IL-23 levels
- Thomas Sharpton
Is PSA increasing activity of Tregs?
- Erick Matsen
Finds that PSA mediates expansion of functionally suppressive CD4+CD25+Foxp3+ Tregs during TNBS colitis.
- Thomas Sharpton
Cells w/ increased Foxp3 expression are immunosuppressive
- Thomas Sharpton
... so better able to control inflammation in vivo.
- Erick Matsen
Round et al PNAS 2012
- Thomas Sharpton
Wanted to identify what sensors detect PSA, queried Toll-like receptor 2 via knockout mouse lines, found protection from TNBS colitis by PSA is required by this receptor.
- Thomas Sharpton
It comes from change of IL-17 levels.
- Erick Matsen
PSA is packaged into outer membrane vesicles.
- Erick Matsen
Found that PSA is packaged into outer membrane vesicles (OMVs) using electron microscopy and indigo blots. PSB is also in outer membrane, but PSG is not.
- Thomas Sharpton
Are these OMVs protective from colitis?
- Erick Matsen
OMVs protect mice from colitis in a PSA-dependent manner. Some amazing differences in the physiology of the intestines of various mice in this experiment
- Thomas Sharpton
Again, this is associated with a change of cytokine levels.
- Erick Matsen
GAdd45alpha expression isrequired for PSA-mediated protection from colitis.
- Erick Matsen
Looked at Bone Marrow DC in WT and Gadd45a-/- lines. Found IL-10 levels drop in WG-OMV relative to deltaPSA-OMV
- Thomas Sharpton
Model: PSA of B. fragilis prevents pro-inflammatory immune responses in the gut to protect from colitis.
- Thomas Sharpton
Idea: can we mimic this pathway theraputically?
- Erick Matsen
Q: How did this evolve-- is there a role for PSA during homeostasis?
- Erick Matsen
Compared germ-free and wildtype microbiota and find that germ free mice are missing Th17 in their intestine. Additional experiments find that PSA restrains Th17 responses to B. fragilis
- Thomas Sharpton
Finds that TLR2 expression on CD4+ T cells is required for in vitro IL-10 production by PSA
- Thomas Sharpton
B. fragilis seems to occupy the mucus layer. So it can impact immune system.
- Erick Matsen
Beautiful confocal microscopy images reveals interactions between B. fragilis and the host. In the absense of PSA, observe a deficiency of microbes that associate with the mucosal sites in the gut
- Thomas Sharpton
but this can be recovered by adding PSA orally.
- Erick Matsen
PSA signals through TLR2 on Tregs to suppress TH17 cell responses during mucosal colonization by B. fragilis.
- Thomas Sharpton
In particular, when TLR2 deleted or PSA not there, fragilis gets kicked out.
- Erick Matsen
Hypotheses regarding potential causes of dysbiosis due to western lifestyles and its role in intestinal health and disease: host genetics, lifestyle (diet, stress), early colonization, medical practices (vaccination use, antibiotics, hygiene)
- Thomas Sharpton
Shows slide showing inverse correlation between infectious diseases and human inflammatory disorders. Maybe how we are treating acute diseases yields the emergence of chronic diseases.
- Thomas Sharpton
Inverse correlation between infectionus diseases and human inflammatory disorders. Increase in MS, Crohns, T1 diabetes, and asthma. All of these diseases seem to have a Treg component.
- Erick Matsen
Wow, quite a remarkable talk.
- Erick Matsen
Akiko Iwasaki - From innate virus recognition to adaptive immunity
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Winner of the 2012 Eli Lilly and Company Research Award
- Thomas Sharpton
Question: how are viruses recognized by the immune system? Which of the various viral sensing pathways lead to adaptive immunity?
- Thomas Sharpton
Hierarchy of innate signals for adaptive immunity: ranges from homeostasis (bottom) to harm(top), bottom is PAMP (pathogen associated molecular pattern), then Vita-PAMP, then PICD (pathogen induced cellular damage), including virulence, damage, pathogenic strategies, top is immunogenicity.
- Thomas Sharpton
Plasmacytoid DCs (pDCs) are sentinel of virus infection. potent producers of type I IFNs in response to a diverse set of viruses. present in blood, lumpoid tissues and can be recruited to the sites of infection.
- Thomas Sharpton
Notes that distinct sensors detect viruses. Pichlmair and Reis e Sousa in Immunity 2007 have a nice review figure on this topic. pDCs use a distinct pathway compared to other sensor cell types. PAMP plays an important role in inducing the pDC pathway.
- Thomas Sharpton
Only live infection by VSV and Sendai virus induces high levels of IFN-alpha
- Thomas Sharpton
Autophagy may be the mechanisms that differentiates live from dead virus. Generating autophagy deficient pDCs in mice fail to recognize VSV infection.
- Thomas Sharpton
How is cell damage inflicted by virus replication strategies recognized by the host cell?
- Thomas Sharpton
General mechanism of NLRP3 inflammasome activation slide is being shown: Ichinohe and Iwasaki Uirusu 2009
- Thomas Sharpton
Notes that incluenza virus encodes an ion channel (M2) which might be recognized by the host cell. Used ion channel deletion mutant of virus and found that the channel is required for inflammasome activation
- Thomas Sharpton
But, M2 expression on incoming virion is not sufficient for inflammasome activation.
- Thomas Sharpton
Influenza is recognized by 3 pathways: TLR7, Ion channel recognition triggering inflammasome, missed the third. Thinks that other viruses are likely recognized by these pathways as well.
- Thomas Sharpton
IL-1R but not PAMP, drive CD8 T cell immunity against influenza virus
- Thomas Sharpton
Influenza-specific T cell responses depend on inflammasome activitation
- Thomas Sharpton
Inflammasomes are required for survival of mice following flu infection.
- Thomas Sharpton
looking at the role of commensal bacteria, find they are required for virus-specific Th1 and CTL responses.
- Thomas Sharpton
Finds that commensal microbiota primes signal 1 in vivo at steady state (tested comparing water to antibiotic diet)
- Thomas Sharpton
Interrectal injection of LPS in antibiotic treated mice restore virus-specific immune responses
- Thomas Sharpton
Intact microbiota and inflammasomes are required for DC migration from lung to the draining lymph nodes
- Thomas Sharpton
She finds that there may be a hierarchy of innate signals for adaptive immunity
- Thomas Sharpton
Julia Oh - How primary immunity shapes bacterial communities
Ruchira S. Datta liked this
Various factors can shape the structure of commensal communities. These include environmental factors and transient members, host phyiology, host genotype, and host immune system
- Thomas Sharpton
She's in the Segre Lab at the NIH
- Erick Matsen
Atopic dermatitis (AD) is marked by staphylococcal colonization and decreased community diversity
- Thomas Sharpton
Found that monogenic mutations can cause shared phenotypes characterized by primary immunodeficiency
- Thomas Sharpton
... even though the sources of this immunodeficicency can be from different sources.
- Erick Matsen
Want to characterize the composition of the microbiome among these patients to unravel how the host genotype, immune system, microbiome influence disease
- Thomas Sharpton
Collected communities from various locations on the body. Conducted 16S sequences.
- Thomas Sharpton
Challenge is that these immunodeficient patients are on treatment, but they controlled by looking at non-immunodeficient cases of AD.
- Erick Matsen
Curious if permissivity of the skin yields changes in diversity, changes in community structure or taxonomic composition
- Thomas Sharpton
Found that community diversity is similar in skin unaffected by dermatitis, but decreases in skin affected with dermatitis
- Thomas Sharpton
Using "theta index" to compare stability of microbiomes.
- Erick Matsen
Found less site-specificity in community structure in primary immunodeficiency patients
- Thomas Sharpton
Identified taxa that uniquely colonized immune deficient patients
- Thomas Sharpton
In particular, Staph is the dominant in controls.
- Erick Matsen
Staphylococcus species dominate AD patients.
- Thomas Sharpton
Also found that immunodeficient skin is uniquely colonized by Serratia
- Thomas Sharpton
However, immunodeficient individuals have lots of Serratia, but not much difference between the affected and unaffecteds. So probably not pathogenic.
- Erick Matsen
non-aureus staphylococci predominated the diseased HIES microbiome
- Thomas Sharpton
Staph is important, so speciated them. Staph aureus dominated immunocompetent AD.
- Erick Matsen
Correlating taxa to clinical markers. Corynebacteria associated with a return to healthy state.
- Erick Matsen
P.acnes, and Lacto, Str. oralis, all bad for recovery.
- Erick Matsen
Wrap up: primary immunodifeiciency may increase permissiveness of the skin. Unique traits were observed in the skin disease in primary immunodeficiency patients.
- Erick Matsen
Yingfei Ma: DNA Virome in the Human Distal Esophagus #asm2012
Ruchira S. Datta liked this
Esophageal brushing samples -> Illumina -> bioinfo. More than fifteen viral families detected in the esophagus. Five detected in 2 more more samples. Cyco-like viruses identified in 13/50 samples-- non enveloped sequence. All metagenomce sequences branch out separate from NCBI sequences (mostly from stools).
- Erick Matsen
Torque teno virus identified in 15/50 samples. First found in patients with Hep. Well mixed with DB sequences.
- Erick Matsen
HPV- identified in 3/50 samples. Human adenovirus in 23/50 samples, and HHV 4 in 15/50 samples.
- Erick Matsen
HPV is only found in cancer patients, but everything else does not distinguish. It does appear that heavy viral load does correspond with negative outcomes.
- Erick Matsen
They think that lots of viruses can either cause cell death or transition to cancer.
- Erick Matsen
Talk: Crawling holobionts and microbial mats: Forest Rohwer (SDSU). #asm2012
Ruchira S. Datta liked this
Holobiont idea: from Lynne Margulis. All these bits that fits together. Coral holobiont: protist, virus bacteria, and archaea. Mix and match to get different behavior.
- Erick Matsen
Metagenomics. Coral- associated bacteria are predominantly heterotrophs. They eat carbs. Corlas live in low N2. Crenarchaeota, to nitrify, Fungi to Ammonification. Coral forms the strucutre, CCA- corraline crustose alga, and macro algaes and turf algaes.
- Erick Matsen
MANY species. Fish eat turf alga. Then all the way up to the top. Almost no nutrients go back to the system. If overfishing, then lots more turf. A whole lot of dissolved organic carbon, which then causes coral disease. As algae covers more area, less grazing pressure. Then fewer fish.
- Erick Matsen
Doing transect survey. Is coral winning or losing vs algae? Corals lose to turf when the ilsands are inhabited.
- Erick Matsen
Dissolved compounds from algae kill corals, and antibiotics protect corals from algae.
- Erick Matsen
Algal derived compounds increase microbial grown and hypoxia on corals.
- Erick Matsen
Relative oxygen content: Oxic on alga and coral, but hypoxic on interface.
- Erick Matsen
What's going on at interface? Raise algae and corals in aquaria, then pyrosequencing, QIIME and Unifrac, heatmap. Difficult to simulate actual interface community.
- Erick Matsen
These mats crawl across reefs. Coral reefs are designed to work in a low nutrient environment, and die when eutrophied. Example of Line Islands-- very iron limited. But when a ship runs into it, then will kill a large area around it. Shipwreck from 20 years ago. Look at rubble samples near shipwreck, can see elevated iron. A little bit of iron doesn't change coral itself. But if you incubate corals with iron, all corals die, unless you put in ampicillin.
- Erick Matsen
Black reefs are a crawling holobiont, moving over coral. Algae come in, increase DOC, then bacteria can move in.
- Erick Matsen
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