Bacterial food web may be key to cystic fibrosis

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[h=2]Bacterial food web may be key to cystic fibrosis[/h]
By
Anne Ju
Cystic fibrosis patients suffer from chronic bacterial infections and thick mucous in their lungs, due largely to a combination of microbial infections and resulting inflammation. A common pathogen, Pseudomonas aeruginosa, which can lay dormant in healthy individuals, becomes virulent in the lungs of cystic fibrosis patients, and Cornell biological engineers think they might know why.
They have shown that P. aeruginosa virulence is “turned on” when it feeds on a particular fermentation product called 2,3 butanediol, demonstrating a direct metabolic relationship between fermenting bacteria and P. aeruginosa. This understanding could lead to more effective treatments for cystic fibrosis patients; rather than the use of antibiotics, disrupting P. aeruginosa’s flow of preferred food could be key to preventing cystic fibrosis-related infections in the lungs.
The research was led by Lars Angenent, associate professor of biological and environmental engineering. A paper, along with a related one by San Diego State University researchers, was published online Jan. 9 in the International Society for Microbial Ecology Journal.
Angenent et al. had previously used bioreactors to show that the presence of 2,3 butanediol promotes cross-feeding between P. aeruginosa and fermenting bacteria, including Enterobacter aerogenes, which makes 2,3 butanediol as a fermentation byproduct.
They are now applying their knowledge of this mutually beneficial microbial relationship to shed light on the microbial environment of lungs affected by cystic fibrosis. In this work, they again used bioreactors to observe the virulence of P. aeruginosa as it fed on both 2,3 butanediol and glucose; the glucose did not cause virulence, while with the 2,3 butanediol, the bacteria became significantly more virulent.
Scientists already knew that P. aeruginosa plays a key role in sickening patients, but the exact microbial interactions and subsequent inflammatory responses that lead to the symptoms have not been well understood.
Because 2,3 butanediol is the substrate, or food, for P. aeruginosa, 2,3 butanediol is the key in the metabolic relationship between different bacteria.
Their relationship results in higher concentrations of phenazines, which are chemicals produced by P. aeruginosa that interact with the fermenter, and are part of the cascade of events that leads to the pathogen’s increased virulence, such as the formation of biofilms that increase lung symptoms in patients. The phenazines combine with oxygen to create free radical compounds, which kill other microbes and cause stress for host cells.
“The signaling is switched on because of the 2,3 butanediol,” Angenent explained. “However, the exact mechanism is still unknown.”
Understanding why P. aeruginosa does well in the lungs of cystic fibrosis patients, and looking at ways to cut off its preferred food supply within that system, could lead to ways to treat the disease, the scientists say.
The paper is called “Metabolite transfer with the fermentation product 2,3-butanediol enhances virulence by Pseudomonas aeruginosa,” and its first author is former graduate student Arvind Venkataraman. Authorship includes former postdoctoral associates Miriam Rosenbaum and Jeffrey Werner, and consultation on quorum sensing, which is the stimulus system through which bacteria communicate, by Stephen Winans, Cornell professor of microbiology. The research was supported by a National Science Foundation CAREER award.

 

[Epona]

This is AWESOME! Thanks for posting this. Where did you find this?

 

[hjmomva]

Assuming this is true, what does that mean in the practice of having CF? What do you eat and/or not eat to have this chemical in your body?

 

[MotherofCFBoy11yrs]

I'm wondering the same. Wikipedia says 2,3 butanediol fermentation is one of the end products of glucose fermentation. One more reason to avoid sugars?

 

[Melissa75]

Interesting! Here is an excerpt from an article from 2007 - what was known or suspected then about other bacteria/viruses influencing pseudomonas virulence.

http://mic.sgmjournals.org/content/153/4/917.full
[h=3]Community interactions 1: synergism[/h] Coinfections probably result from sequential, rather than simultaneous, colonization, and it appears that some degree of succession (change in community structure over time) occurs within the CF lung (e.g. Hoiby, 1974; Wahab et al., 2004). Molecular profiling studies such as that of Rogers et al. (2004) provide particularly clear evidence for succession within patients. Succession may depend in part on environmental changes resulting from the action of infecting microbes; alginate production by P. aeruginosa, for example, can exacerbate local hypoxia (Worlitzsch et al., 2002), presumably constraining the spectrum of species or strains able to invade. There is also evidence that RSV may render some patients more susceptible to chronic P. aeruginosa infection (Petersen et al., 1981).


Historically, synergism in the CF lung has usually been proposed with respect to Staph. aureus ‘sensitizing’ the lungs to subsequent infection by P. aeruginosa (e.g. Burns et al., 1998; but see also Lyczak et al., 2002). However, there is also evidence for antagonistic interactions between these two species (see below). On the other hand, there is very good evidence that P. aeruginosa can promote further pathogenesis. Firstly, two endobronchially detectable P. aeruginosa quorum-sensing molecules (3-oxo-C[SUB]12[/SUB]-homoserine lactone and 2-heptyl-3-hydroxy-4(1H)-quinolone) have been shown to possess immunomodulatory effects (Hooi et al., 2004). Secondly, Wahab et al. (2004) reported a possible link between pre-existing, chronic colonization by mucoid P. aeruginosa and infection by more rarely observed bacteria such as S. maltophilia, A. xylosoxidans and Mycobacterium abscessus. Moreover, P. aeruginosa has been implicated in promoting BCC pathogenesis via the upregulation of BCC virulence factor expression (Riedel et al., 2001).


Synergism can result in increased disease severity in mixed-species infections. For instance, those patients with coinfections of mucoid P. aeruginosa and rare bacterial species observed by Wahab et al. (2004) had significantly worse lung symptoms than did patients colonized by mucoid P. aeruginosa alone. The normal oropharyngeal microflora (OF) may similarly exacerbate CF lung disease. In CF patients, avirulent species which are normally confined to the upper airways are frequently recovered from the lower airways (Coenye et al., 2002; Santana et al., 2003). It has been shown (Duan et al., 2003) that the presence of OF exacerbates P. aeruginosa-mediated lung damage in infected rats. This resulted from the specific upregulation of P. aeruginosa genes linked with pathogenesis, and appeared to be partially attributable to the P. aeruginosa cells ‘eavesdropping’ on intercellular communication between OF constituents (Duan et al., 2003).


Bacteriophage are present at appreciable concentrations in the CF airways (Ojeniyi et al., 1991), and these may affect the pathology of their target bacteria. P. aeruginosa phage recovered from CF sputum have been shown to be capable of inducing changes in P. aeruginosa serotype and a shift to polyagglutinability – a trait associated with increased patient mortality (Hoiby & Rosendal, 1980).

 

[LouLou]

Isn't this so fascinating?! We all knew that sugar feeds bacteria but this is proof. I know for me I crave sugar badly after having antibiotics the worst (when good gut flora is compromised). As for which foods to avoid, to start it's all processed foods because they mostly have either refined wheat or sugar or both. They both are quickly broken down and create sugar Time to start making healthy snacks from scratch and meals too if you aren't already doing that.

 

[Epona]

Hey friends. This is a great study and I hope that it translates into further research into non-antibiotic treatments for disrupting bacterial biofilm formation.

Although sugar consumption absolutely feeds infection and bacteria proliferation, as some in this thread have mentioned, that is not what this study discusses or concludes. What this study discusses is how certain molecules (2,3 butanediol) created by different pathogenic bacteria (pseudomonas and fermenting bacteria like enterobacter) allow these harmful species to work together to become more virulent and to create biofilms, which are what make bacterial infections chronic and intractable. The whole point of this study and how it can translate into future treatments is that it proves certain bad bacteria work together using specific trigger molecules, so if we develop treatments that target those trigger molecules (blocking them or inactivating them somehow) then we may be able to prevent pseudomonas from becoming a chronic infection, even when a CFer is exposed to it. The study also proves that we need more research into the interactions between the species that make up the CF lung ecosystem. If we can somehow figure out how we can reduce the occurrence of fermenting bacteria in the lungs via lifestyle, diet, or drugs, that may also help decrease the chances of pseudo turning into a chronic infection. At least, this is my interpretation of this study.

Furthermore, this study indicates to me that more research needs to be done on how pseudo biofilms form and how we can prevent them. Biofilms are the lynchpin to this entire conversation, including the article you posted, Melissa75. Biofilms are what make CF infections permanent infections, and they allow multiple pathogenic species work together to protect eachother from pesky things our immune system and antibiotics. Antibiotics cannot destroy biofilms, so we need new medical strategies to address this issue, and yet very little research has been done so far. Check out this article on the work of a leader in the field of biofilms, a doc who successfully treats diabetic infections by targeting the biofilms, and not just by throwing antibiotics at them. His work is directly translatable to CF lung disease. I just wish we had a visionary like him in the field of CF research. http://bacteriality.com/2008/04/13/wolcott/

 

[scarecrow]

Could somebody who has more time than me do some research into Lactoferrin and Xylitrol to see if anyone has used them to treat lung infections. If you read the link above, that is what that Dr. is using to treat biofilms on Diabetic wounds that won't heal. those 2 things together seem to break down those biofilms. They might work for us, too.

 

[LittleLab4CF]

Scarecrow,

I will look into Lactoferrin and Xylitol per your request. It should be fun I don't know a thing about either one. Actually, I use Xylitol in my research but not in any way germane to breaking down biofilms. If credible articles are out there, I have subscriptions to a fair number of scientific and medical research periodicals, online databases and such. Hopefully I can digest and explain pertinent information about their use and potential uses. This is important stuff and if breaking down biofilms is a key to ending infection, surprise again. DMSO has been used topically to penetrate biofilms for a long time. Xylitol should be familiar to diabetics. It is an artificial sweetener.


Good old Robert Koch, father of microbiology, was my idol growing up. His discovery of greatness involved one of the most sophisticated biofilms ever devised by a microbe. Many pathogenic bacteria were seen by Koch's staining methods but the TB bacillus wasn't being found because of its unique armored shell. Made from a chitin like material, it is coated form refused staining. Thank vacationing Europeans and sloppy scientists who leave their dirty dishes for a month. Fleming probably copied Koch's technique of discovery by neglect. By Koch leaving his famous agar plates open along with vats of carbolic acid they used for destroying all living things in cleaning lab ware, the acrid carbolic acid fumes etched away the chitin sheathes hiding these deadly killers.


Once Koch's team returned from vacation, continued staining of contaminated samples revealed the beautiful pathogen in shades of Gram's stain along with some partial chitin shells. The idea Dr.Wolcott is discussing is a concept about biofilms formed by different but cooperating forms of bacteria. This isn't what germ theory easily embraces because most bacterial battles are fought against other bacteria and other forms of microbes such as yeasts. Then again, bacteria adapt so beautifully and so quickly, very little would surprise me. A multi-racial cooperative society of bacteria enjoined with a common mission of keeping the bounty just for the group sounds whacky as science gets but with bacteria, I never assume.


I don't question Dr. Wolcott's data and his hypothesis is not one I know to be proven otherwise. A fairly universal anti-biofilm or biofilm solvent is needed. If there are cooperative infections making biofilm, every biofilm will be different, even on the same person. "Biofilm" is a new word for an old and well known issue. A scab is a biofilm, and the great revelation of Fleming’s penicillium mold was the biofilm the mold made like a growing moat around the fungal colony, clearing and consuming everything in its biofilm margin. The biofilm appears two dimensional, but deep in the agar, battles for food and territory rage with chemical warfare in the same three dimensions you find in living tissue.

Most antibiotics are nothing more than a particular organism’s biochemical warfare agents harvested and purified for our use. Penicillium fungi excretes a biofilm which we harvest and purify to make penicillin. I see a lot of really sorry legs on diabetics. This is a horror worth investigating and following progress on a potential end to life ending amputation torture.

As for biofilms in CF lungs, the theory is more plausible since mucus is a biofilm in its own right. Mucosal epithelial cells rather than bacteria, but mucus can be feed stock for microbial biofilms. A recent post illustrated different consistencies of lung mucus harbor different bacteria. An agent that dissolves the matrix a bacterial colony produces is opening this biofilm to penetration by the body's defenses and antibiotics. Since many CFers are facing diabetes down along with chronic lung and GI infections, these agents promise help in several areas.

Here’s hoping the good doctor is right!,

LL

 

[Epona]

I see real promise in the development of biofilm-breaking agents such as xylitol and certain enzymes for treatment of chronic CF lung infections. It is actually the only thing that is really going to address this issue - the newest and most expensive antibiotic on the market will only get us so far if the bacterial biofilms of pseudomonas or MRSA are still intact. I am looking forward to the results of the newest trial of inhaled xylitol (http://clinicaltrials.gov/show/NCT01355796). The use of biofilm-breaking enzymes to treat gut infections is now gaining acceptance in the holistic health community (e.g. Interfase Plus), but I am very curious about the possibility of enzymatic biofilm disruption in the lungs. That's close to what pulmozyme is - pulmozyme is an enzyme targeted at breaking up neutrophil elastase, which is what makes up a significant portion of the CF mucus that clogs our airways.

Littlelab - I have a question that you may be able to answer for me. On my sputum culture read outs there is always mention of mucoid and non-mucoid pseudomonas. I asked the docs about the difference, but they are not really very helpful in the education department (what's a doc's first and most important job? Education! Ugh.). Do you know the difference between mucoid and non-mucoid strains of pseudo, and if so could you explain this to me, and how this difference relates to biofilm formation? Thanks.

 

[LittleLab4CF]

Epona, scarecrow, and anybody interested in mucoid and non mucoid P.a.

Mucoid certainly sounds like mucus and thankfully scientific nomenclature still relies on the root language, Latin. Mucoid bacteria, was named so because it glistens when you observe colonies of it on the agar plate or on the microscope slide. In some cases non mucoid verses mucoid bacteria is hydro-inflation, or over filled versions of the same bacteria.

How I wish that was the case for non mucoid and mucoid P.a. (Pseudomonas aeruginosa) in CF patients. Non mucoid P.a. is what most of us catch. If it stays that way, antibiotics have a chance at this MDR or multiple drug resistant bacteria. When bacteria of the same genus i.e. Pseudomonas and maybe the same species i.e. aeruginosa, a growing colony evolves into several species. Think of E. coli, OK, what is E. coli 157 besides a lethal pathogen? It’s the sixth cousin twice removed by a few billion generations.

The biofilm article sited above describes how a colony of bacteria in the body starts out a non mucoidal bacteria infection and can become a mucoidal bacteria infection. As the colony grows the rapid reproduction begets new and slightly different family members. When bacteria swing, their slipping and sliding cause genetic excretions and careless absorbing of genetic excretions, their way of sexual reproduction not the usual asexual reproduction assumed of bacteria. That’s how they are nearly winning all the time.

In the case of P.a. a non mucoid species does not excrete slime called alginate. As it turns out our lungs are perfect for resistant non mucoid P.a. to colonize and make a switch over to a hive cooperative, in theory, of mucoid P.a. or alginate producing and totally resistant P.a. I now understand why Koch’s agar plates conceal the real changes. Without the aid of modern chemical analysis, agar and alginate are identical for all practical purposes. Alginic acid (alginate) is a type of anionic polysaccharide in this case much like agar or kelp derived gelatins.

A biofilm of water bound up with alginic acid makes a three dimensional living space for bacteria in the form of a syrup or gel like non Newtonian fluids. It is a nutrient common for all species of P.a. in the colony. More like a bio-medium, everything the bacteria need easily transports through this viscous artificial mucus.

Greater virulence is credited to mucoid P.a. and though it wasn’t termed a “biofilm” its manner of protecting the bacterial colony meets a biofilm definition. Alginate is edible by people, you can get some in your eye and it won’t do more than be in your eye. It is truly advanced warfare when pathogens live in a selective cocoon that they can set back and consume us and we can’t do a thing about it.

So we’ve gone from a metabolite found in mucoid P.a. that may be tied to food. That may be of diagnostic help or it may eventually be part of an eradication protocol. Two agents, xylitol and Lactoferrin make more sense now as well. Polysaccharides are nature’s plastic feed stock or building blocks for an entire family of bio-polymers. Like oils being soluble in thinner oils like alcohol, sugars are broken down by thin sugars like xylitol or even acetone. Lactoferrin may be acting differently but P.a. metabolizes lacto-proteins. I have my homework to do on Lactoferrin will report back.

The disparate contributions on this topic post are strangely interconnected. I won’t go so far to say we have a solution but hopefully Epona’s loaded with bait question clears the fog and connects all the parts.

How many other biofilm protected bacteria work the same way? How many other types of biofilms need to be decoded?

LL

 

[Epona]

Thanks so much for this LL. If I may attempt to distill your explanation into a few words, is it that when pseudo is in a pre-biofilm state (i.e. "planktonic"), it is non-mucoid. Once Pa moves into a biofilm-producing state (mediated by alginate secretion), it becomes mucoid. Is that a fair summary? That's facinating how use of agar plates may not be correctly noting the presence of alginate (i.e. biofilm) on a Pa culture due to the fact that alginate looks like agar!

I came across an incredibly useful text from the U. of Copenhagen on the role of biofilms in chronic infection, and lucky for us the main example of study used throughout the text is pseudo infection in CF lungs! It's so long I admit I haven't gotten through the whole thing, but what I did read was very helpful. Here's the link: http://www.ncbi.nlm.nih.gov/pubmed/23635385

Other question: do you do, or do you know of anyone who does genetic sequencing of CF mucus samples to determine what's actually down there in a more complete picture? When we get the regular old sputum cultures at our CF centers, what we're really getting is just a picture of the species that can grow on agar, but leaves out many many bacteria, viruses, molds, etc. that may not be culturable using that procedure. So essentially, we are never getting a whole picture of the microbes that are down there unless we get a genetic snapshot. I know that labs are now doing this more regularly for stool samples - any news of use of this tool for sputum samples? In an affordable way, i.e. a way that insurance will pay for it?

Thanks!

 

[curiousme]

Hi,
I'm new to cf.com, though not new to CF or to biofilms. Biofilms are what make bacterial (& other) infections impervious to antimicrobials, & research in recent years indicates that a key - possibly the key - to busting up biofilms is to deprive the pathogens of iron.

I already have done a lot of research on biofilms (of Pseudomonas a & many other pathogens), as well as on infection & iron more generally; & I've looked into lactoferrin & Xylitol, used separately & together, as treatments for Pseudomonas a & other infections. But I haven't written it all up yet in a way that I find sufficiently precise, exhaustive or eloquent - & I'm not sure when I'll be able to do so. However, in the meantime I'd be happy to walk someone else through my notes, if anyone is interested.

BTW, LittleLab4CF, I have found your posts very illuminating, interesting & helpful so far. I'd love to see or hear anything you have to say about the genetics of CF, & genetics in general, & about anything else!

Best wishes to all,
CuriousME
(My name is Mary Ellen, so that's what the ME stands for in "CuriousME." Does this moniker make me look like an egotistical ass? Pray, do tell, anyone, so I can change it sooner rather than later.)