7 Did Isabel Diterich Have The Cure For Chronic Lyme Disease?

One researcher whose papers I've been reading recently is Isabel Diterich's. Several years ago she published two papers on Lyme disease which grabbed my attention because they not only revealed a hypothesis of immunosuppression caused by Borrelia burgdorferi spirochetes - but they also revealed a potential cure for chronic Lyme disease.

I say "potential" here with this caveat:

While the treatment did appear turn a man who was disabled into what sounds like the picture of health for at least eight years, he had to take filgrastim for almost two weeks. And filgrastim is an immune modulating drug which can have serious side effects in some people - there have even been a few fatalities.

However, most of the people who have suffered serious side effects from filgrastim were cancer and leukemia patients who already had serious health problems and were at greater risk for being affected by the drug. And most patients - including cancer patients - experience less dramatic effects of fatigue and joint pain from the use of filgrastim - something Lyme disease patients suffer with anyway.

Scary sounding as it is to take a drug which has the risk of serious or even fatal side effects, one has to consider that if better and safer immune modulating drugs could be developed - along with antibiotics - together they might be the cure for chronic Lyme disease.

To quote from Isabel Ditrech's 2003 thesis, "Immunomodulation and new therapeutic strategies in Lyme borreliosis":

"5.3.1 Case report

A 51 year old patient with a history of frequent exposures to tick bites presented with polyarthritis in the fingers and feet. Arthritic destruction of synovial clefts mainly in the metacarpophalangial and in the proximal interphalangial joints of fingers and feet could be demonstrated by X-ray. Low, but clearly positive, serum titers of Borrelia IgG by ELISA and immunoblot (p100 +++) and a negative IgM-ELISA (both MaxPettenkofer-Institute, Munich, Germany) corroborated diagnosis of late stage Borrelia infection.  

A standard two week i.v. treatment with 2 g/day Ceftriaxone (Rocephin,Hoffmann LaRoche, Grenzach-Whylen, Germany) led to transient improvement of symptoms, i.e. subjective decline of arthritis, that lasted for eight weeks. Then, the inflammatory symptoms returned and became progressively worse, indicating that the treatment had probably failed.  

We hypothesized that persistence of Borrelia might be due to a disabled immunocompetence of the patient. Therefore, we tested whether a complete eradication of the pathogen could be achieved by combining immunosupportive treatment with antibiosis. The experimental treatment regimen, applied with the informed consent of the patient, was as follows: First week 2 g Ceftriaxone (Rocephin ) i.v. daily, second week 480 µg s.c. Filgrastim (Neupogen, Amgen, Thousand Oaks, USA) every second day, and third week 2 g Ceftriaxone daily plus 300 µg Filgrastim every second day (Figure 5.1). Neutrophil counts were determined by a Coulter STKS counter (Coulter, Krefeld, Germany)"

So this lays out the background of this individual case report on one patient. What were the results? More quoted from the above thesis:

"5.4.1 Patient case report

The combination therapy of Ceftriaxone plus Filgrastim was well tolerated. Only after the first injection of Filgrastim the patient reported acute but moderate pain in the previously affected joints i.e. the shoulder, fingers and knees. 

Circulating neutrophil counts increased from 1400 to 17000 cells/µl within 24 h after the first Filgrastim injection. Monocyte numbers increased about two-fold, while there was little effect on lymphocytes (Figure 5.2a). The plateau of neutrophil counts at about 17000 cells/µl blood was maintained until one day after the end of treatment.  

The subjective symptoms disappeared during the following six weeks after the treatment. The patient reported that he was able to resume previously abandoned sporting activities including mountain climbing and downhill skiing. Moreover, fine mechanical skills needed for piano playing were restored. 

After three months, the Borrelia IgG titer was negative. The intensity of the immunoblot at this time point was significantly reduced (from +++ to +) and two years later it was negative. Eight years after treatment the patient is still free of arthritic symptoms."

Source:
http://kops.ub.uni-konstanz.de/bitstream/handle/urn:nbn:de:bsz:352-opus-9814/Diss_formated_ENDVERSION.pdf

So it seems like at least for this patient, this method of treatment changed their life so that they could return to all the things they used to do that they loved. I would have liked to know more about this patient and how he is doing today, given it has been years since this study was completed.

And I'd like to know if a similar treatment plan would work for me and everyone else suffering with chronic Lyme disease. To take ceftriaxone and filgrastim for a couple weeks - or something similar, but with fewer side effects - only to be done with this nightmare and get on with my life would be fantastic.

It would mean no more attempts at long term antibiotic treatment and experimentation with alternative medicine. I would just get treatment for three weeks and be done with it... Sounds like a plan to me.

Reflecting on this, over the years there have been anecdotes - stories I've heard passed around Lyme disease support groups - about the occasional chronic Lyme disease patient who went on to discover they had cancer, went through chemotherapy and other supportive treatment for their cancer - only end treatment not only going into remission from cancer  - but saying that they think their chronic Lyme disease is cured, too.

These stories have been around for a while, but I've never personally known anyone who went through this process. It would be great to get a confirmation from their doctors and families that after chemotherapy and supportive treatments, they had a notable and lasting improvement and feel like their old selves again. What if a drug like filgrastim played a role in their recovery?

This isn't the only example of a chronic condition where the cause has been unknown and the symptoms can be debilitating and lead to years of loss of productivity and physical pain... let's consider chronic fatigue syndrome, also known as CFS/ME or CFSIDS.

A study completed last year in Norway showed that rituximab had a profoundly positive effect on people with CFS/ME. In this study, a few people seemed to go into complete remission from their CFS and returned to work and led normal lives. It didn't work for everyone - 40% of study participants did not experience improvement from the drug. It's unknown why. But that it worked so well for the rest of treated patients deserves a closer look because it begins to reveal the mechanisms behind what causes CFS/ME.

While there has been speculation that chronic fatigue syndrome and chronic Lyme disease (CLD) are the same condition, a recent study on the different proteins found in the cerebrospinal fluid (CSF) of both CFS and CLD patients has challenged this notion. At least in terms of objective evidence, the proteins in the CSF of both groups are different. However, what if part of the underlying process behind what causes these conditions is the same?

Quoting the above well-written article from the Phoenix Rising ME web site, let's look at the mechanism behind rituximab and what it does in people with CFS/ME:

"Rituximab is believed to deplete B-cells in two ways; by recruiting other members of the immune system to attack them and by locking on a receptor on the B-cell that tells the cell to kill itself. B-cells are an integral part of the immune response. Until they are activated, B-cells quietly troll the blood, collecting and digesting molecules called antigens that appear to be suspicious. Once they are digested they place bits of them on MHC molecules for T-cells to inspect. If the T-cells decide those molecules came from a pathogen, they turn around and turn the B-cells on – transforming them into antibody producing machines (‘plasma cells’) that can generate from 100s to thousands of antibodies per second.

These antibodies or immunoglobulins are specifically manufactured to attach to a pathogen and physically stop it from locking onto our cells. The antibodies also alert macrophages to come gobble up the pathogen and they turn on other parts of the immune system. B-cells are key players in the immune response but if they go too far; if they get too zealous, they can mistakenly attack our own cells and overactive B-cell activity has been implicated in many auto-immune disorders."

If this sounds familiar to you, then you might have been reading about Viral Genetics' targeted peptide therapy, VGV-L, for treating chronic Lyme disease.

Viral Genetics' patent states the following about treating chronic Lyme disease:

"[0116] It is believed according to the invention that Borrelia burgdorferi also produces a Toll ligand for TLR2. Replacement of the CLIP on the surface of the B cell by treatment with a thymus derived peptide with high affinity for the MHC fingerprint of a particular individual, would result in activation of the important Tregs that can in turn cause reduction in antigen-non-specific B cells. Thus treatment with thymus derived peptides could reactivate specific Tregs and dampen the pathological inflammation that is required for the chronic inflammatory condition characteristic of Lyme Disease. With the appropriate MHC analysis of the subject, a specific thymus derived peptide can be synthesized to treat that subject. Thus individuals with all different types of MHC fingerprints could effectively be treated for Lyme disease."

An easier-to-understand explanation can be found elsewhere - this research report revealed how VGV-L is used to treat HIV. In this instance, just substitute "chronic Lyme Disease" for "HIV" and you can get a picture of what VGV-L does:

"The conventional approach to HIV vaccines, for example, is to develop therapeutic vaccines to stimulate immune system response. The problem with the conventional approach is that the infected cells are camouflaged and not visible to the body’s immune system. The body’s powerful T-cells are unable to seek out and destroy the infected camouflaged cells because they cannot recognize that the cell is infected.

To understand the issue, think of the Klingon space ship on Star Trek that has its cloaking device activated. The U.S.S. Enterprise has no way of knowing where the enemy is in space. The only hope it has in winning the battle is for the Klingon vessel to be de-cloaked and, once revealed, use their ammunition to destroy it. What’s worse in the case of HIV is that while the infected cell is cloaked, it is also effectively setting off an alarm that triggers the immune system to create inflammation. Why is this important? It turns out that this inflammation is critical for allowing the HIV virus to spread to even more cells.

Many other viruses and bacteria also trigger inflammation but, unlike HIV, the inflammation does not necessarily allow or facilitate the spread of the virus or bacteria itself. However, in these cases, the inflammation itself is harmful because it creates a hostile and inflamed environment that provides the necessary components for a potential autoimmune reaction that can cause the immune system to attack and damage one’s own body. Viral believes that diseases such as Lyme Disease, Multiple Sclerosis and others involve this inflammatory mechanism.

To use the Star Trek metaphor, what Dr. Newell Rogers has developed with TPT is a de-cloaking device for the body’s immune system to use in its pursuit of invaders. Through the development and use of computational biology programs and databases, Dr. Newell Rogers and her team have created a way to remove the camouflage that is cloaking the infected cells, flagging them with custom peptides that allow the body’s immune system to seek out and destroy them.

The key discovery of the TPT platform is that a self-peptide (in other words, one that is naturally produced and a healthy part of one’s normally functioning immune system) called ―CLIP2 that was until now thought only to exist primarily inside certain immune system cells, is sometimes displayed on the outside of cells, thus leading to harmful inflammation. Dr. Newell Rogers discovered that the products of some pathogen invaders such as viruses and bacteria, when picked up on the surface of certain immune system cells, sometimes incorrectly cause those cells to display CLIP externally (i.e. ―ectopically).

Normally, when an invader strikes, this process may promote needed inflammation early in infection, but it is quickly controlled when a more specific, immune response takes over, allowing a highly-targeted immune response to be marshaled against the pathogen. However, when CLIP is improperly displayed, displayed for too long or displayed chronically, the immune system is marshaled to promote a broad and unspecified inflammation without the specific targeting, leaving open the possibility that this inflammation actually turns against one’s own cells. Replacing CLIP is the focus of Viral’s Targeted Peptides because it turns off the harmful alarm."

Read more from the source - including about individual MHC genetic profiles here: http://www.viralgenetics.com/investors/press-releases/Research_2.0_Report_Feb1_2011.pdf

They're using Star Trek metaphors to describe this... I think that's pretty geeky. Awesome.

So, it seems that whether there is current infection or not, VGV-L may be one way to effectively treat chronic Lyme disease and lower inflammation due to runaway immune dysregulation. And if infection is currently present, then it looks like VGV-L will trigger T cells that recognize the infection and summon functional B-cells to fight it.

Now, getting back to Isabel... Remember Isabel, the researcher who used filgrastim and ceftriaxone to treat a patient with chronic Lyme disease about a decade ago? Yes, that Isabel.

Well, she wrote another paper, along with Rauter, Kirshning, and Hartung: "Borrelia burgdorferi-Induced Tolerance as a Model of Persistence via Immunosuppression"

The abstract states:

"If left untreated, infection with Borrelia burgdorferi sensu lato may lead to chronic Lyme borreliosis. It is still unknown how this pathogen manages to persist in the host in the presence of competent immune cells. It was recently reported that Borrelia suppresses the host's immune response, thus perhaps preventing the elimination of the pathogen (I. Diterich, L. Härter, D. Hassler, A. Wendel, and T. Hartung, Infect. Immun. 69:687-694, 2001). Here, we further characterize Borrelia-induced immunomodulation in order to develop a model of this anergy. 

We observed that the different Borrelia preparations that we tested, i.e., live, heat-inactivated, and sonicated Borrelia, could desensitize human blood monocytes, as shown by attenuated cytokine release upon restimulation with any of the different preparations. Next, we investigated whether these Borrelia-specific stimuli render monocytes tolerant, i.e. hyporesponsive, towards another Toll-like receptor 2 (TLR2) agonist, such as lipoteichoic acid from gram-positive bacteria, or towards the TLR4 agonist lipopolysaccharide. Cross-tolerance towards all tested stimuli was induced. Furthermore, using primary bone marrow cells from TLR2-deficient mice and from mice with a nonfunctional TLR4 (strain C3H/HeJ), we demonstrated that the TLR2 was required for tolerance induction by Borrelia, and using neutralizing antibodies, we identified interleukin-10 as the key mediator involved."

Source: http://iai.asm.org/content/71/7/3979.full

Where have I heard something like this before? Oh, Dr. Karen Newell Rogers - that's right - she discussed this at a recent Lyme disease research conference:

"[...]Some researchers would argue that chronic inflammation requires the continuous presence of bacteria, whereas others would suggest that continuous presence of bacteria does not always result in inflammation and that exacerbations of chronic symptoms could result from infection with a different organism--or that chronic symptoms could re-cur from unrelated pro-inflammatory events. Potentially reconciling these seemingly conflicting perspectives on the mechanism of Lyme disease may be the effect of Borrelia burgdoreri’s bacterial by-products on Toll Like Receptors, (TLR)-mediated immune activation. 

TLR appear to be the “gate-keepers” of an inflammatory response. Bacteria, including Borrelia, produce products that, by binding to TLRs on the cell surface, promote leukocyte activation, cytokine production, and acute inflammation. In some genetic backgrounds of mice, acute inflammation is sufficient to fight off infection and resolve disease. In other mouse strains, the pathogens, or in this case the bacteria, get past TLR-induced inflammation and remain symptomatically undetectable in cells and tissues (Barthold, etc); Barthold et al. have found that no matter how severe or mild the disease in any of the genetically inbred strains of mice, there was no more inflammatory disease when the bacteria were eliminated."

And where else have I heard about IL-10 production before? Oh, right - Rituximab, and research on gender differences in antibody response to Borrelia burgdorferi...

From the previously mentioned Phoenix Rising ME article:

"While Rituximab is busy destroying B-cells there is also evidence that it may actually be turning on NK cells – which, of course, habitually underperform in CFS. Rituximab also appears to increase production of IL-10 – a key anti-inflammatory cytokine that may be a protective agent in ME/CFS – and reduces levels of the powerful pro-inflammatory cytokine tumor necrosis factor. A review article suggested that Rituximab was able restore Th1/Th2 balance in the immune system. These results suggest Rituximab could be working as an immunodulator helping to re-balance the immune response by turning down the over-activated parts of it and bumping up the under-active ones."

All this ties together quite nicely, it seems, with other research I have listed here - forming a master hypothesis with different pieces. Does it hold up to scrutiny? Tell me - I'd love to hear your ideas.

But here is the master hypothesis, in its infancy:

1) Host genetics play a role in the ability of mice (and possibly people!) in clearing Borrelia burgdorferi infections. See:

http://campother.blogspot.com/2011/08/immune-infection-hla-dr-alleles.html

The host's genetic background in developing chronic infection is, however, open to debate - and may not play as big a role in disease as Borrelia burgdorferi s.l.'s genetic diversity/VlsE recombination on different plasmids:

http://campother.blogspot.com/2011/08/do-different-genetic-haplotypes-matter.html
http://campother.blogspot.com/2011/08/more-on-genetic-haplotypes-and-lyme.html

2) The genetics of Borrelia burgdorferi strains play a role in how quickly they disseminate into host tissues and also how well they can generate inflammation - which leads to overstimulation of the immune system in production of poor quality plasma b-cells, but also, ironically, immune suppression because of the mechanisms Isabel Diterich and Karen Newell Rogers describe. Refer, also, to Tunev and Barthold et al's research, "Lymphoadenopathy during Lyme Borreliosis Is Caused by Spirochete Migration-Induced Specific B Cell Activation":

http://campother.blogspot.com/2011/06/paper-borrelia-burgdorferi-rst1-ospc.html
http://spirochetesunwound.blogspot.com/2011/07/does-borrelia-burgdorferi-cause.html
http://www.plospathogens.org/article/info%3Adoi%2F10.1371%2Fjournal.ppat.1002066
http://spirochetesunwound.blogspot.com/2010/07/antigen-presentation-in-bloodstream-how.html (refer to other research on relationship between b-cells/plasma cells and T cells)

It could also be that not having enough iNKT cells is an issue:
http://www.pnas.org/content/105/50/19863.full.pdf

2a) The changing pattern of antigenic variation during this time may also be why patients produce an undulating immune response in measured antibodies which echo a more drawn-out response similar to relapsing fever:

http://campother.blogspot.com/2012/02/paper-course-of-antibody-response-in.html
http://campother.blogspot.com/2011/08/antibodies-linked-to-long-term-lyme.html (read comments, too)
http://www.ncbi.nlm.nih.gov/pubmed/9108482
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2772371/
http://www.ncbi.nlm.nih.gov/pubmed/11544329
http://campother.blogspot.com/2011/07/lyme-disease-western-blots-and-antigen.html

It may not be that the tests are lousy for measuring antibodies which are present to Borrelia burgdorferi. It may be that the antibodies are not present because they are tied up in immune complexes.

2b) There is also the possibility that Borrelia burgdorferi is occasionally intracellular in nature, though there is not enough in vivo evidence to support this. If so, it would also explain why an undulatory immune response might be present:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3067508/?tool=pubmed
http://campother.blogspot.com/2011/07/fibroblasts-and-lyme-disease-sample.html

Whether or not items #2a and #2b are relevant here remains to be seen - the main point is that Borrelia burgdorferi can lead to both overstimulation of the immune system as well as immune suppression.

Based on this, I surmise that may not be that blood tests are so lousy at detecting antibodies produced by the presence Borrelia burgdorferi. It may be that there is no reliable way to detect the presence of infection by correlating them with the presence of antibody responses (seronegative Lyme disease).

3) What gender you are and your hormone levels and metabolism may play a role in persisting symptoms and prolonged infection as well, so there is ALSO a metabolic cause behind chronic Lyme disease. How well the immune system can respond to initial infection to begin with seems to play a role in developing chronic Lyme disease, as even 10% of acute cases of Lyme disease result in treatment failure.

http://campother.blogspot.com/2012/03/lyme-disease-presents-differently-in.html
http://campother.blogspot.com/2012/01/two-new-hypotheses-for-chronic-lyme.html (read comments, too)
http://www.ncbi.nlm.nih.gov/pubmed/17438273 (this may provide the scientific link for the anecdotes that people who develop chronic Lyme disease generally were under more stress when they contracted the disease)

4) If there are persister cells, this is an additional consideration - throwing more antibiotics at a pathogen which is antibiotic tolerant when it is a persister cell will, at most, keep the infection from getting worse but it won't eliminate it.

http://campother.blogspot.com/2012/01/paper-persistence-of-borrelia.html
http://campother.blogspot.com/2012/02/blog-log-spirochetes-unwound-on.html

See also:
The research of Kim Lewis on persister cells: www.bu.edu/abl/files/killing_persisters.pdf
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3145328/

And it may be that persister cells are more likely to be on the scene earlier, depending on how appropriate a given antibiotic is for treating specific genospecies - refer to item #2 above, but also:

http://campother.blogspot.com/2011/05/abstract-evaluation-of-in-vitro.html

5) Because the host has a sub-optimal immune system, even with long term antibiotics, a subset of the population will have trouble clearing the remaining spirochetes after antibiotics are stopped. Additional antibiotics plus a treatment which eliminates low quality plasma b-cells and promotes the activity of Treg cells which recognize current infection could overturn the dysregulated immune system.

What does this boil down to?

Easy: The argument of "is it a chronic infection or is it an immune disorder, possibly autoimmune" is a false dichotomy and too simplistic.

The circumstances which give rise to chronic Lyme disease are more complex than that, and if people want to solve the chronic Lyme problem, they have to roll up their sleeves and look at more puzzle pieces and how they fit together.

Image credit: 
Original image by Muns on Wikimedia Commons; derived image above by Schlurcher.

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1 Memo: Borrelia Are NOT Gram-Negative Bacteria, And Might Become Resistant

For years, Borrelia have been thought to be Gram-negative because of their diderm and structural appearance upon staining. Borrelia - including Borrelia burgdorferi - are not Gram-negative bacteria, though - even though many people are still referring to them as such. They belong in their own category.

As I was telling Dr. MacDonald this week:

I found information confirming the fact that Borrelia is not Gram-negative or Gram-positive. It's its own special thing:

"Borrelia were thought to be Gram negative because of their double membrane structure, but genetic analysis places them - along with other spirochetes - into a separate eubacterial phylum. Ultrastructural molecular and biochemical studies have emphasized the wide taxonomic gap between spirochetes and Gram-negative bacteria."

- From "The Genus Borrelia" by Melissa Caimano. Prokaryotes (2006) 7:235-293.

"Although Borrelia spirochetes are often, but mistakenly described as Gram-negative bacteria due to their diderm, i.e. double-membrane envelopes, a closer examination reveals significant differences in composition and architecture. Probably most striking is the lack of LPS, the presence of major surface lipoproteins at the host-pathogen interface during transmission, persistence and ensuing pathogenic processes and the additional function of periplasmic flagella in defining cell shape. While surface lipoproteins such as the Osps interact with a variety of ligands in different organ tissues, they are also targets of the immune response and several have emerged as vaccine candidates."

- From Borrelia: Molecular Biology, Host Interaction and Pathogenesis. Edited by D. Scott Samuels and Justin D. Radolf. (2010)

So one could say they are Gram-negative-"like" - but strictly speaking, Borrelia is not Gram-negative bacteria."

He accepted this response, and is now stating that Borrelia burgdorferi is Gram-indeterminate.

There is additional information from the second source cited which indicates that Borrelia spirochetes are different from other bacteria, summarized here:

"Some of the identified periplasmic lipoproteins, i.e. the OppAs, are components of substrate transport complexes. Investigations into integral membrane proteins led to the identification of several Borrelia porins: P13, whose structure and function is unknown, DipA, which is specific for dicarboxylates and P66 (Oms66), which has a dual role as a pore-forming outer membrane protein with an extremely high single channel conductance and an adhesin for β3-integrin. The recently identified Tol homologs BesA, -B and -C appear to form a Type I 'channel' to export exogenous toxic agents such as antibiotics and to maintain infectivity by an unknown mechanism. Initial studies on envelope biogenesis pathways based on diderm proteobacterial model organisms already revealed significant deviations from the norm. This further bolsters the unique status of Borrelia among microbial pathogens."

Damn Borrelia... Why do you have to be such a deviant? Why can't you just conform?

Anyway, the Type I 'channel' they're talking about above which exports or removes antibiotics and helps maintain infectivity is said here to do so using an unknown mechanism. Judging from research out there, this channel relates to transmembrane proteins which span the outer and inner membranes of the bacteria - and it appears to function like an efflux pump (in this case, a resistance-nodulation-division-like pump) which removes antibiotics and is involved in antibiotic resistance. Check out this paper (full text at link):

Source: http://www.plospathogens.org/article/info%3Adoi%2F10.1371%2Fjournal.ppat.1000009

An RND-Type Efflux System in Borrelia burgdorferi Is Involved in Virulence and Resistance to Antimicrobial Compounds

Ignas Bunikis1, Katrin Denker, Yngve Östberg1, Christian Andersen, Roland Benz, Sven Bergström. PLoS Pathog. 2008 Feb 29;4(2):e1000009.

Abstract

Borrelia burgdorferi is remarkable for its ability to thrive in widely different environments due to its ability to infect various organisms. In comparison to enteric Gram-negative bacteria, these spirochetes have only a few transmembrane proteins some of which are thought to play a role in solute and nutrient uptake and excretion of toxic substances. Here, we have identified an outer membrane protein, BesC, which is part of a putative export system comprising the components BesA, BesB and BesC. We show that BesC, a TolC homolog, forms channels in planar lipid bilayers and is involved in antibiotic resistance. A besC knockout was unable to establish infection in mice, signifying the importance of this outer membrane channel in the mammalian host. The biophysical properties of BesC could be explained by a model based on the channel-tunnel structure. We have also generated a structural model of the efflux apparatus showing the putative spatial orientation of BesC with respect to the AcrAB homologs BesAB. We believe that our findings will be helpful in unraveling the pathogenic mechanisms of borreliae as well as in developing novel therapeutic agents aiming to block the function of this secretion apparatus.

---

Obviously, more research in this direction is necessary.

So when people ask, "Camp, why are you so interested in learning about something like TolC? Why don't you want to learn about different kinds of treatment and how they've helped patients?" my answer for them is this:

I do want to learn about different kinds of treatment - it is in my best interest to do so, given I have been dealing with Lyme-related health problems for years. And I make my own decisions about treatment which are largely personal to my situation.

But learning this stuff - these details tucked away in what must appear to most people to be obscure publications in little-known journals outside of researchers within the field? To me, this stuff is what may get us all closer to better treatment for many people.

Do I know this for sure? No. No one does. But with some of the specifics under my belt, I can then at least petition the science world and advocate for the funding to do research on some very particular subjects to get very specific knowledge. Knowledge which may give us a solid idea as to how to treat chronic Lyme disease so that it doesn't become chronic.

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0 Science Articles: Probiotics As Anti-inflammatories, Bacterial Gene Transfer

Probiotics Effective in Combating Antibiotic-Associated Diarrhea, Studies Find; 'Good Bugs' Look Promising as Anti-Inflammatory Agents

In four different studies presented at the American College of Gastroenterology's (ACG) 76th Annual Scientific meeting in Washington, DC, researchers explored the effectiveness of probiotics for antibiotic-associated diarrhea; as an anti-inflammatory agent for patients with ulcerative colitis, psoriasis and chronic fatigue syndrome; and for people with abdominal discomfort and bloating who have not been diagnosed with a functional bowel disorder, such as irritable bowel syndrome (IBS).

Reference:
American College of Gastroenterology

READ MORE at source: http://www.sciencedaily.com/releases/2011/10/111031114951.htm

Bacteria May Readily Swap Beneficial Genes: Microbes Trade Genetic Coding for Antibiotic Resistance and More

Much as people can exchange information instantaneously in the digital age, bacteria associated with humans and their livestock appear to freely and rapidly exchange genetic material related to human disease and antibiotic resistance through a mechanism called horizontal gene transfer (HGT).

Reference:
Chris S. Smillie, Mark B. Smith, Jonathan Friedman, Otto X. Cordero, Lawrence A. David, Eric J. Alm. Ecology drives a global network of gene exchange connecting the human microbiome. Nature, 2011; DOI: 10.1038/nature10571

READ MORE at source: http://www.sciencedaily.com/releases/2011/11/111101125958.htm

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0 News: Antibiotic Shortages Affect Patients

This just in from the Internal Medicine website: Antimicrobial Shortages Affect Patient Quality.

For those who are treating tickborne illnesses, this excerpt may be particularly of importance to you:

"The drugs reported most frequently to be unavailable or in short supply include the intravenous formulation of trimethoprim/sulfamethoxazole (Bactrim), amikacin (Amikin), aztreonam (Azactam), foscarnet (Foscavir), and penicillin G."

Other drugs are listed as well, of which the shortage will affect many people this flu season as one of them is Tamiflu.

(Now would be a good time for more startups to invest in not only antibiotic drug development, but phage therapy as well.)

MORE on how patients have been affected and why this is happening can be found at the link: http://www.internalmedicinenews.com/newsletter/conference-coverage/singleview40841/id-docs-antimicrobial-shortages-threaten-patient-care-quality/802aeb528c.html

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0 Failure of Topical Antibiotics to Prevent Disseminated Borrelia burgdorferi Infection

Last week, I posted an article on how topical azithromycin could help prevent Lyme disease infection. And now, this new abstract has turned up on PubMed...

Failure of Topical Antibiotics to Prevent Disseminated Borrelia burgdorferi Infection Following a Tick Bite in C3H/HeJ Mice. Wormser GP, Daniels TJ, Bittker S, Cooper D, Wang G, Pavia CS. J Infect Dis. 2011 Sep 19. [Epub ahead of print]

Abstract

A prior study in mice has shown that the timely application of topical antibiotics to the skin at the tick bite site could eradicate Borrelia burgdorferi infection. That study, however, did not evaluate antibiotic preparations that are considered suitable for use in humans. In this murine study, topical application of 2% erythromycin and 3% tetracycline preparations that are acceptable for use in humans was found to be ineffective in eliminating B. burgdorferi from the tick bite site or in preventing dissemination to other tissues. Reasons for the discrepant findings are discussed.

---

I find it very odd that they would choose erythromycin as a topical antibiotic for Borrelia burgdorferi - there is research that shows that Borrelia burgdorferi is resistant to erythromycin. I would have never thought to make that a choice for treatment. Tetracycline makes more sense.

How many previous studies of topical antibiotics for treating early Lyme disease have there been, and what have the outcomes been? How many have been done using suitable human antibiotic preparations versus ones only effective in animal subjects?

It seems to me that the researchers who filed this patent might already have a good idea of what might work - is this the same formulation that was used in the study I posted last week?

Composition C
Composition [mg]
Component per unit [1 g]
Azithromycin 150
Dermacryl 79 50
Klucel MF 25
Miglyol 812 50
Ethanol 94% (w/w) ad 1 g

Or is it different?

Results: There is no infection of Lyme disease detectable when composition C is applied to the area where the ticks were allowed to feed. The topical application of Formulation C results in antibody titers, which are not different from naïve mice (KELA values between 10 and 40), whereas the infected mice show KELA values of 160 to 400.

Especially, the tissue and serum samples taken from different parts of the mice 56 days after the tick bite show no Borrelia burgdorferi organisms when cultivated and no specific antigenes are detectable. Tissue probes are taken from heart tissue, bladder, joint and ear. The group of mice topically treated with Formulation C show complete absence of B. burgdorferi , whereas B. burgdorferi is detected in untreated mice, which are exposed to tick-feed.

The recultivation conditions of B. burgdorferi are suitable to detect B. burgdorferi in any morphological form known to date. Surprisingly, Formulation C was able to eradicate B. burgdorferi in a way, that no infectious agents of B. burgdorferi are detected in the target tissues of B. burgdorferi.

How were these tissue probes completed and could they have missed any bacteria? What about the parenchyma of the brain, since this is low passage N40 they're talking about? (Problematic in rodent studies - should be studied in other animals.) I wonder about the methodology...

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24 Do Different Genetic Haplotypes Matter?

It seems people are still holding on to the HLA-DR2 and HLA-DR4 concept of subgroup autoimmunity (subgroups of Lyme disease patients who do not respond to additional antibiotic treatment) in the Lyme disease community.

I get the impression that hypothesis has been dropped by a number of researchers in favor of a new one - except in the case of patients with Lyme arthritis. (I don't think HLA-DR2 is associated with Lyme disease, either, and HLA-DR4 is associated with Lyme arthritis.)

I came across this recently:

National Institute of Allergy and Infectious Diseases, NIH: Impact on Global Health (2009) by Vassil St. Georgiev, PhD

Chapter 22.1 Lyme Disease (Lyme Borreliosis, Lyme Arthritis)
Section 22.1.5.4 The Role of Autoimmune Reactivity in Lyme Disease

"In NIAID-supported clinical studies, case subject patients with PTCLD* were compared with control subjects without such symptoms for the presence of several human leukocyte antigen (HLA) class II (DRB1 and DQB1) genetic markers, some of which are known to be associated with the expression of autoimmune reactivity. The results obtained did not support the involvement of an autoimmune mechanism in PTCLD (24). However, because not all autoimmune diseases are associated with specific HLA haplotypes, these findings do not necessarily exclude that possibility. Definitive proof would clearly involve demonstrating the presence of significant levels of relevant autoimmune antibodies and/or autoreactive T cells in patients with PTCLD but not in treated control subjects without such symptoms. A greater frequency of DRB1*0401, which has been reported to be associated with antibiotic-treatment-resistant arthritis, was noted in the case subject patients; although this finding appeared to be nominally significant (p < 0.05), its biological significance is ambiguous because none of the case subjects considered had symptoms of inflammatory arthritis. (http://www3.niaid.nih.gov/research/topics/lyme/research/autoimmune/)"

* In this NIAID-NIH document, PTCLD stands for Post Treatment Chronic Lyme Disease.

24) Klempner MS, Wormser GH, Wade K, Trevino RP, Tang K, Kaslow R, and Schmid C. (2005) A case-control study to examine HLA haplotype associations in patients with posttreatment chronic Lyme disease. J. Jinfect. Dis. 192(6), 1010-1013.

Now this is an interesting piece of information.

This book from the NIAID-NIH is stating following about patients with PTCLD in NIAID-supported studies:

1) Patients with chronic Lyme disease were compared against control subjects with the same genetic markers and the results did not support an autoimmune mechanism for chronic Lyme disease.

2) Not all autoimmune diseases are associated with specific haplotypes.

3) Those patients with chronic Lyme arthritis have had their condition strongly associated with DRB1*0401.

4) None of the case subjects in trials had inflammatory arthritis. None.

I find this particularly interesting since the one statement many patients have made is that some researchers have pegged chronic Lyme disease patients as having an autoimmune disorder, when the research in this regard is fuzzy. What indicates to some researchers today that post-treatment chronic Lyme disease is an autoimmune disorder, if it isn't specific haplotypes which are at risk of developing persisting symptoms?  A different molecular mimicry model than that which leads to Lyme arthritis? A different model of persisting symptoms entirely?

I also find it interesting that none of the case subjects had inflammatory arthritis. I don't know how clinicians defined the criteria for inflammatory arthritis and if they did any serology for inflammation, but I can say that I have had joint swelling, muscle pain, and shooting nerve pain during the course of my illness. That none of the case subjects in trials had inflammatory arthritis comes as a surprise to me.

According to research there is a relationship between a specific haplotype and Lyme arthritis. This is just one such example:

From Persistent arthritis in Borrelia burgdorferi-infected HLA-DR4-positive CD28-negative mice post-antibiotic treatment. Iliopoulou BP, Alroy J, Huber BT. Arthritis Rheum. 2008 Dec; 58(12):3892-901.:

"METHODS:We have previously shown that CD28(-/-) mice develop intermittent monarticular Lyme arthritis that is responsive to antibiotics. Since there seems to be a link in humans between persistent arthritic manifestations post-antibiotic treatment and the HLA-DR4 allele, we generated DR4+/+CD28(-/-)MHCII(-/-) mice, infected them with Borrelia burgdorferi, and subsequently treated them with antibiotics.

RESULTS: Thirty-eight percent of the B burgdorferi-infected DR4+/+CD28(-/-)MHCII(-/-) mice, but none of the B burgdorferi-infected CD28(-/-)MHCII(-/-) mice, remained arthritic post-antibiotic treatment. A significant fraction (36%) of these mice, but none of the mice in which arthritis resolved, had serum antibodies to outer surface protein A of B burgdorferi. After abrogation of active B burgdorferi infection, the inflammatory reaction in mice with persistent joint inflammation was restricted to the joints, since their draining lymph nodes were no longer enlarged. Increased CD20 and interferon-gamma messenger RNA expression in the inflamed joints of these mice suggested a possible role of B cells and inflammatory cytokines in the pathogenesis of persistent arthritis post-antibiotic treatment.

Conclusion: The establishment of this murine model allows, for the first time, the elucidation of the immunologic events that lead to persistent Lyme arthritis post-antibiotic therapy in genetically susceptible individuals."

So according to this, little over a third of the mice with a specific allele remained arthritic after antibiotic treatment, and of this third, about a third of them developed antibodies to OspA.

My questions are: How is it that the remaining roughly two thirds of the mice did not remain arthritic? How is it that two thirds of them did not develop antibodies to OspA?

There are many papers written about the association between specific alleles and development of chronic Lyme arthritis:

1993 - Association of treatment-resistant chronic Lyme arthritis with HLA-DR4 and antibody reactivity to OspA and OspB of Borrelia burgdorferi.

2001 - Autoimmune mechanisms in antibiotic treatment-resistant lyme arthritis. (This is where molecular mimicry as a cause of Lyme arthritis is mentioned.The abstract states: "...only one human protein was identified, lymphocyte function associated antigen-1 (hLFA-1), that had sequence homology with OspA (165-173) and predicted binding in the DRB1*0401 molecule.")

2004 - Molecular characterization of the OspA(161-175) T cell epitope associated with treatment-resistant Lyme arthritis: differences among the three pathogenic species of Borrelia burgdorferi sensu lato. (Borrelia that causes Lyme disease in Europe does not usually lead to chronic Lyme arthritis.)

2006 - Antibiotic-refractory Lyme arthritis is associated with HLA-DR molecules that bind a Borrelia burgdorferi peptide (Concluded that binding of a single spirochetal peptide to certain DRB molecules is a marker for antibiotic-refractory Lyme arthritis and might play a role in the pathogenesis of the disease.)

2007 - Clonal Diversification in OspA-specific Antibodies from Peripheral Circulation of a Chronic Lyme Arthritis Patient

A number of these are Steere's papers - not surprising since he has written so much about the rheumatological aspect of Lyme disease. And some of it is not his research, and I could find a few more by others.

I'm looking for more research on chronic Lyme disease's connection to haplotypes and alleles outside of the Lyme arthritis model. The NIAID-NIH book, written in 2009, states there is no connection.

So to sum up:

1) Particular alleles or haplotypes are not associated with chronic Lyme disease as a whole.

2) Particular alleles or haplotypes are associated strongly with chronic Lyme arthritis.

Hm.

Glossary

allele: is one of two or more forms of a gene.Sometimes, different alleles can result in different traits, such as color. Other times, different alleles will have the same result in the expression of a gene. See more info. at: http://en.wikipedia.org/wiki/Allele

haplotype: a combination of alleles (DNA sequences) at different places (loci) on the chromosome that are transmitted together. A haplotype may be one locus, several loci, or an entire chromosome depending on the number of recombination events that have occurred between a given set of loci.See more info. at: http://en.wikipedia.org/wiki/Haplotype

Additional information: http://en.wikipedia.org/wiki/Category:HLA-DR_haplotypes

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0 Superbug Suit: Groups Sue FDA Over Risky Use of Human Antibiotics in Animal Feed

Approximately 70% of all antibiotics used in the United States are given to healthy farm animals at low doses to promote faster growth and compensate for unsanitary living conditions -- a practice that has increased over the past 60 years despite evidence that it breeds antibiotic-resistant bacteria dangerous to humans. The antibiotics, mixed into feed or water for pigs, cows, chicken, and turkeys, are used at levels too low to treat disease, leaving surviving bacteria stronger and resistant to medical treatment.

FDA concluded in 1977 that feeding animals low-doses of certain antibiotics used in human medicine -- namely, penicillin and tetracyclines -- could promote antibiotic-resistant bacteria capable of infecting people. However, despite this conclusion and laws requiring that the agency act on its findings, FDA failed to take any action to protect human health.

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9 Abstract: Evaluation of in-vitro antibiotic susceptibility of Bb

I was recently asked about this study in comments, since there has been much discussion about this research in the Lyme disease patient community online:

Evaluation of in-vitro antibiotic susceptibility of different morphological forms of Borrelia burgdorferi

My opinion of it thus far is one of waiting and seeing - I'm not sure what to think yet and I'm waiting to hear more information... I'm guessing this is an initial publication of findings and a more detailed paper similar to Dr. Sapi's cancer research papers will be published in the near future.

Tinidazole

I have a number of questions and thoughts on it, and prior to this publication, I had the impression that tinidazole has helped a number of Lyme disease patients - but there has been limited research on it (See: Brorsons).

The Brorsons stated in their 2003 paper, An in vitro study of the susceptibility of mobile and cystic forms of Borrelia burgdorferi to tinidazole:

"Acridine orange staining, dark-field microscopy and transmission electron microscopy revealed that, when the concentration of TZ (Tinidazole) was ≥ MBC, the contents of the cysts were partly degraded, core structures did not develop inside the young cysts, and the amount of RNA in these cysts decreased significantly. When cysts were exposed to TZ, both the spirochetal structures and core structures inside the cysts dissolved, and the production of blebs was significantly reduced."

But I digress...

To take each question and thought about Dr. Sapi's latest paper by point:

"Three morphological forms of B. burgdorferi (spirochetes, round bodies, and biofilm-like colonies) were generated using novel culture methods."

Is there a description of these novel culture methods? Have they been through a process of verification and validation? How are they superior to other methods?

If different media is used other than BSK/BSK-H, it would be good to know if that affects the results in some fashion.

"Minimum inhibitory concentration and minimum bactericidal concentration of five antimicrobial agents (doxycycline, amoxicillin, tigecycline, metronidazole, and tinidazole) against spirochetal forms of B. burgdorferi were evaluated using the standard published microdilution technique."

What exactly were the concentrations?

It's important to know, and compare against other studies which have been done in vitro. Of all the literature I've read on this, standardization of these documented concentrations is lacking.

And to note: This was an in vitro test, not in vivo. Given everything that is known about Bb's behavior in vivo, in vitro tests will only give us part of the picture. More on that in a minute, below...

"The susceptibility of spirochetal and round body forms to the antibiotics was then tested using fluorescent microscopy (BacLight™ viability staining) and dark field microscopy (direct cell counting), and these results were compared with the microdilution technique. Qualitative and quantitative effects of the antibiotics against biofilm-like colonies were assessed using fluorescent microscopy and dark field microscopy, respectively."

How do these tests compare to one another relative to results found for each? How do the novel culture methods affect outcome relative to standard culture methods?

How is something biofilm-like, versus a biofilm?

"Doxycycline reduced spirochetal structures ~90% but increased the number of round body forms about twofold. Amoxicillin reduced spirochetal forms by ~85%–90% and round body forms by ~68%, while treatment with metronidazole led to reduction of spirochetal structures by ~90% and round body forms by ~80%. Tigecycline and tinidazole treatment reduced both spirochetal and round body forms by ~80%–90%."

This is interesting... Some of these results match earlier research findings on specific antibiotics used to treat Bb.

In 2008, Xiaohua Yang, Andrew Nguyen, Dan Qiu, and Ben Luft did some research on the effectiveness of tigecycline and doxycycline on Borrelia burgdorferi in vitro in In vitro activity of tigecycline against multiple strains of Borrelia burgdorferi: http://jac.oxfordjournals.org/content/63/4/709.short

In this abstract it was stated:

"Tigecycline inhibited the growth of and killed the organism more rapidly than doxycycline. Tigecycline was able to kill B. burgdorferi within 24 h at clinically achievable concentrations (> 1 mg/L). In contrast, doxycycline was bacteriostatic and required 48–72 h to achieve its maximal inhibitory effect. The anti-Borrelia activity of the antibiotics was tested against 20 different isolates from three species. Tigecycline was 16- to 1000-fold more active than doxycycline at immobilizing Borrelia for the 20 isolates tested."

The MIC versus the MBC is important to know, and it affects outcomes on a timeline.

Comparing Tigecycline to doxycycline sounds like comparing an axe to a thousand papercuts - the former is just going to be more immediately effective than the latter... And if Bb happens to disseminate rather quickly in an individual patient, there's a chance the doxycycline prescribed may not be enough to adequately treat the patient if you're going by this MIC - especially if one of Bb's strategies is to evade the immune system. (Let's not even get into how ineffective doxycycline is for prophylaxis for now...)

More recently, in 2010, Louis Ates, Christa Hanssen-Hübner, Douglas E. Norris, Dania Richter, Peter Kraiczy and Klaus-Peter Hunfeld conducted the study, Comparison of in vitro activities of tigecycline, doxycycline, and tetracycline against the spirochete Borrelia burgdorferi.

In their abstract they stated:

"The overall rank order of MIC90s was tigecycline (≤0.016 mg/L) > ceftriaxone (0.03 mg/L) > cefotaxime (≤0.125 mg/L) > doxycycline (0.25 mg/L) > tetracycline (0.25 mg/L). The rank order of MBC90s was tigecycline (0.5 mg/L) > ceftriaxone (2 mg/L) > tetracycline (16 mg/L) > doxycycline (16 mg/L) > cefotaxime (>16 mg/L).

High in vitro activity of the new glycylcycline against Borrelia was further substantiated by time-kill experiments performed with B. afzelii isolate EB1. Parallel testing of tigecycline and ceftriaxone demonstrated a bacteriostatic effect for 0.016 mg/L of tigecycline and for 0.03 mg/L for ceftriaxone after 72 h of incubation. Moreover, tigecycline was bactericidal at a concentration of 0.25 mg/L showing a > 3 log₁₀ unit reduction of the initial inoculum, whereas for ceftriaxone a concentration of 2 mg/L was needed."

So as you can see in this earlier research, tigecycline is highly effective in vitro, and doxycycline is ranked much further down the list of effectiveness.

Stepping into our time machine again, and going back to 2004 to Klaus-Peter Hunfeld, Thomas A. Wichelhaus, Rebecca Rödel, Georg Acker, Volker Brade, and Peter Kraiczy's study, Comparison of In Vitro Activities of Ketolides, Macrolides, and an Azalide against the Spirochete Borrelia burgdorferi: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC310164/

Now this was measuring an entirely different group of antibiotics, with the following results:

"The ketolides were the most potent against borrelial isolates on a micrograms-per-milliliter basis. For all agents except cethromycin and telithromycin, the MIC at which 90% of isolates were inhibited (MIC90) and the MBC at which 90% of the isolates were killed were ≥0.01 μg/ml and > 0.25 μg/ml, respectively."

and

"In our study, the rank order of activity by classical macrolides and azalides against borreliae clearly corresponds to the effectiveness of these agents as revealed by in vitro susceptibility studies and clinical treatment trials to date (2, 4, 5, 7, 8, 9, 11, 23, 24), demonstrating higher in vitro effectiveness for azithromycin (MIC90, 0.0156 μg/ml) than for erythromycin (MIC90, 0.0625 μg/ml), roxitromycin (MIC90, 0.0625 μg/ml), and clarithromycin (MIC90, 0.0312 μg/ml). Median MICs of the different substances, however, tended to vary over a 10-fold range between individual strains, with the B. garinii isolate PSth and the B. afzelii isolate EB1 showing the highest MICs for both the classical macrolides and the ketolides."

So in this study, of the antibiotics looked at, ketolides were more effective than macrolides, and azithromycin was more effective than other macrolides.

We also see that MIC's are different for different European Bb isolates. More on that below, in another cited study.

And then they said this about treatment failure:

"Classical macrolides and azalides frequently fail in the therapy of early LD (7, 14, 17, 26), and clinical relapse has been observed following conclusion of treatment (14, 17, 26). Moreover, it has been speculated that resistance may develop in borreliae preexposed to erythromycin owing to resistant subpopulations (25). Based upon our findings, however, the ketolides were superior in vitro on a micrograms-per-milliliter basis when tested alongside classical macrolides under identical test conditions in BSK."

There are a number of studies out there showing that Borrelia burgdorferi can be antibiotic resistant, with some being erythromycin resistant. Because of this, it is critical for doctors to weigh the use of erythromycin in patients - especially pregnant women with Lyme disease - against the risks of another antibiotic which is more effective.

The surprising thing about Dr. Sapi's study, to me, is the part about amoxicillin being that effective at reducing both spirochetal and round body forms. I say this, because I thought earlier research points towards amoxicillin being a less effective treatment than doxycycline.

Did I miss something, though? I thought earlier research did not look at antibiotic impact on round body forms in general, though - other than the Brorsons' metronidazole and tinidazole research.

Image taken from Brosons' research, An in vitro study of the
susceptibility of mobile and cystic forms of Borrelia burgdorferi to tinidazole

But earlier research is out there which confirms the effectiveness of amoxicillin on Borrelia burgdorferi, and found in a 2003 publication, In Vitro Susceptibility Testing of Four Antibiotics against Borrelia burgdorferi: a Comparison of Results for the Three Genospecies Borrelia afzelii, Borrelia garinii, and Borrelia burgdorferi Sensu Stricto.

What's fascinating to note here, again, is that different isolates of Bb respond very differently to different antibiotics!

"In 7 out of 12 comparative evaluations (P  > 0.05), MBCs were significantly different among the three genospecies. B. garinii seemed to be especially susceptible to azithromycin, while amoxicillin had a significantly greater effect on B. burgdorferi sensu stricto compared to the other genospecies. Ceftriaxone had the lowest MBC with B. afzelii and increasingly higher MBCs with B. garinii and B. burgdorferi sensu stricto. Doxycycline did not show any remarkable differences in its effects on the three genospecies."

So amoxicillin apparently doesn't suck when it comes to treating Borrelia burgdorferi, but it's not as effective on the other genospecies. (C'mon, Dr. Luft, please get that test working so we know which Bb we have to treat it the most effectively right off the bat.)

If it is found in vivo that doxycycline creates round bodies that contribute to the spirochete's survival, then doxycycline - what is typically given to patients diagnosed early with Lyme disease - would be contraindicated.

Whether the round bodies are as relevant as a potential "stasis" of metabolism in Borrelia burgdorferi remains to be seen. Either way, in vivo findings are different from in vitro findings.

In vitro, tigecycline was said to be many times more powerful than other antibiotics in killing Borrelia burgdorferi...Yet as we can see from Barthold's experiments on mice, viable spirochetes are found after tigecycline treatment in vivo, and viable enough that they can be picked up by ticks and transmitted to a new host.

Is there any treatment which can be used that would ensure the destruction of these remaining spirochetes, and would their demise lead to the end of persisting symptoms in patients who have them - or would there be an ongoing immune dysregulation which was triggered by their existence which continues after they are all dead?

This is something I'd really like to see studied.

Getting back to the last bit of Dr. Sapi's paper...

"When quantitative effects on biofilm-like colonies were evaluated, the five antibiotics reduced formation of these colonies by only 30%–55%. In terms of qualitative effects, only tinidazole reduced viable organisms by ~90%. Following treatment with the other antibiotics, viable organisms were detected in 70%–85% of the biofilm-like colonies."

I'd like to see analysis of how each of the five antibiotics fared relative to one another within biofilm-like colonies, rather than a range. It'd be good to do a direct comparison of each antibiotic against each form in vitro including biofilm-like colonies.

So... On the whole, I'd say take note of the study, with the message that independent confirmation and reproducibility of the methods chosen and these findings are important - and it's good to make note of these findings in relationship to other research already completed.

If anyone heads to the University of New Haven on the 21st to see the presentation on this, I'd love to get a report from you in comments about what was said.

In the end, I'll leave you with this closing thought from a paper from 2005, In Vitro Susceptibility Testing of Borrelia burgdorferi Sensu Lato Isolates Cultured from Patients with Erythema Migrans before and after Antimicrobial Chemotherapy :

"... similar to failures of chemotherapy for Treponema pallidum in syphilis (24), clinical treatment failures have been reported to occur in early LB cases for almost every suitable antimicrobial agent (10, 12, 28, 38, 42). Furthermore, the currently available diagnostic techniques do not reliably discriminate among possible reinfection, true endogenous relapse, and coinfection with other tick-borne pathogens (12). These drawbacks together with the phenomenon of resistance to therapy in individual patients undoubtedly contribute to the inconsistencies surrounding the optimal treatment regimens for LB and are often misinterpreted and misused to support prolonged antibiotic treatment regimens. However, relatively few cases of culture-proven treatment failure have been published (19, 22, 28, 29, 37, 38, 39), and the underlying mechanisms of antimicrobial resistance in B. burgdorferi sensu lato remain unresolved."

And there you have it. This pretty much characterizes the scientific reasons contributing to the ongoing controversy, five years later: Yes, there are treatment failures; yes, they are hard to diagnose and distinguish from coinfection and reinfection; yes, there is antimicrobial resistance; yes, scientists state these issues contribute to what is viewed as a misuse of prolonged antibiotic treatment.

But if treatment is necessary - whether it is a relapse or a new infection - then treatment is necessary.

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1 The Friday Four

In this week's Friday Four, we'll look at how some bacteria avoid antibiotics by shutting down and hiding until it's safe to come out again,  students who go bacteriophage hunting,  disrupting bacteria's communication or quorum sensing in future antibacterial treatments,  tests which use bacteria's scent to detect not only their presence but species, strain, and their antibiotic resistance profile.

CO message to readers: The Friday Four postings will be on hiatus for at least the month of May during Lyme Awareness Month.

1) 'Going off the grid' helps some bacteria hide from antibiotics

Link: http://www.sciencedaily.com/releases/2011/04/110425153611.htm

ScienceDaily (2011-04-25) -- Call them the Jason Bournes of the bacteria world. Going "off the grid," like rogue secret agents, some bacteria avoid antibiotic treatments by essentially shutting down and hiding until it's safe to come out again.

Comments:

I want to keep this one short and sweet: What if those few Borrelia burgdorferi left behind in collagen that some researchers say are not viable or non-dividing are just basically in stasis instead? What if they have shut down their metabolic processes and only look mostly dead? (This is starting to remind me of the scene in that movie, The Princess Bride, where Westley is... Oh, never mind, if you haven't seen it, I don't want to spoiler it for you. It's a fun movie. I will tell you the Bourne series is one of the best action series in my opinion - along those lines, I like Memento too...)

Source Reference:
Xiaoxue Wang, Younghoon Kim, Seok Hoon Hong, Qun Ma, Breann L Brown, Mingming Pu, Aaron M Tarone, Michael J Benedik, Wolfgang Peti, Rebecca Page, Thomas K Wood. Antitoxin MqsA helps mediate the bacterial general stress response. Nature Chemical Biology, 2011; DOI: 10.1038/nchembio.560

2) Phage hunting students find new bacteriophages in soils of St. Louis suburbs

Link: http://www.sciencedaily.com/releases/2011/04/110425135645.htm

ScienceDaily (2011-04-25) -- Twelve students who had participated in an unusual biology course as freshmen have found two bacteriophages, viruses that prey exclusively on bacteria, in the soil of two suburbs of St. Louis, Missouri. As the finders, they had the naming rights; the new phages are called Angelica and Uncle Howie.

Comments:

This is as awesome as being an amateur astronomer. If you're an amateur astronomer, if you find an object in the sky no one has discovered before, it can be named after you or you can decide what you want to name it. Here, students are discovering their own bacteriophages in the dirt and naming them anything they want.

I posted this mainly because I think it's cool, and I wish I had gotten the opportunity to do this in school, too. Well, who knows... maybe I'll go back to school someday, just to be able to take a course like this and name my own bacteriophage Camp Other. If I did, though, I'd try to find one that consumed Borrelia burgdorferi.

Source Reference:
Pope WH, Jacobs-Sera D, Russell DA, Peebles CL, Al-Atrache Z, et al. Expanding the Diversity of Mycobacteriophages: Insights into Genome Architecture and Evolution. PLoS ONE, 2011; 6 (1): e16329 DOI: 10.1371/journal.pone.0016329

3) Bacteria interrupted: Disabling coordinated behavior and virulence gene expression

Link: http://www.sciencedaily.com/releases/2011/04/110421122329.htm

ScienceDaily (2011-04-22) -- New research reveals a strategy for disrupting the ability of bacteria to communicate and coordinate the expression of virulence factors. The study may lead to the development of new antibacterial therapeutics.

Comments:

Bonnie Bassler is up to it again. I love her presentation on TED, and if you haven't seen it, you really should set aside 18 minutes of your time to watch her video on how to get bacteria to talk and how to get them to shut up.

And recently she was on a team that did more research on how to stop bacterial infections by shutting up them up.  Four points in turn outlined their strategy for how one could stop bacterial infection by stopping quorum sensing:

1. Quorum-sensing (QS) antagonists represent potential antibacterial therapeutics
2. They can bind LuxR-family transcription factors in competition with autoinducers
3. The antagonists stabilize a closed conformation incapable of binding operator DNA
4. This inhibition strategy may be generalizable to other multidomain receptors

Which means that there are antagonists which can bind to certain factors that normally autoinducers would bind to - the antagonists are competition for them,  much like Saccharomyces bouldarii can be competition for other yeasts and C. difficle. When the antagonists bind to the factors, they will not bind to operator DNA.

So to sum up: If you can stop autoinducers, you can stop the bacteria from communicating. You can shut it up. If you shut it up, you can tell it to stop having sex and the immune system police will evict it, much like a loud annoying neighbor.

You think I'm kidding, and making this story up? I'm not - I'm merely telling the story to illustrate a point: In order for gene transcription to be activated in the bacteria, the cell must encounter autoinducers secreted by other cells in its environment.

Here's a basic diagram of how Gram-negative bacteria engages in quorum sensing (noting that Borrelia burgdorferi is not exactly Gram-negative or Gram-positive here, it is somewhat closer to Gram-negative so I include that model here):

What you need to imagine here is that this oval represents a bacterium, and that initially a small number of bacteria are doing this all at the same time in their host, whether that be human or not.

Here the LuxI protein makes the autoinducers (green pentagons) which then diffuse freely outside. Each bacterium doing the same, the concentration of external autoinducer is a measure of the size of the population (quorum).

When the autoinducer concentration is high (meaning the bacteria has reproduced to a certain population)  the autoinducer binds to a cognate receptor LuxR (cognate means having the same form and ad hoc characteristics to bind specifically to the molecule it receives).

This is quorum sensing.

The complex auto inducer-Lux R then binds at target gene promoters and activate their effect (transcription) which has behavioral consequences.

In other words, once the bacteria reaches a certain threshold, the level of autoinducers is very high, and the number of bacteria goes up. The high autoinducer level means more bacteria, and more bacteria means more autoinducers. It's a self-perpetuating feedback loop. If you can prevent the loop from even getting started, bacterial numbers will remain low.

So, you're probably wondering, does Borrelia burgdorferi engage in quorum sensing, and if so, can we get it to shut up also?

This has actually been somewhat under debate. Some research has stated that Borrelia burgdorferi has an autoinducing cognate receptor called LuxS, but it doesn't have the necessary autoinducer to bind to it, which in this case would be AI-2.

More recent research has shown that there might be a more complicated method for Borrelia burgdorferi involved for synthesizing its own autoinducers... Might.

To draw from this Polish research paper from 2009 (http://www.aaem.pl/pdf/16001.pdf):

"...the studies of von Lackum et al.[62] demonstrated that B. burgdorferi encodes functional Pfs and LuxS enzymes for the breakdown of toxic products of methylation reactions. According to these observations, B. burgdorferi was shown to synthesize the final product, 4,5-dihydroxy-2,3-pentanedione (DPD) during laboratory cultivation. DPD undergoes spontaneous rearrangements to produce a class of pheromones collectively named autoinducer 2 (AI-2). The addition of in vitro-synthesized DPD to the culture of B. burgdorferi manifested in differential expression of a distinct subset of proteins, including the outer surface lipoprotein VlsE. Although many bacteria for regeneration of methionine can utilize the other LuxS product, homocysteine, B. burgdorferi did not show such an ability. It is hypothesized that B. burgdorferi produces LuxS for the express purpose of synthesizing DPD, and utilizes a form of that molecule as an AI-2 pheromone to control gene expression [4]."

Those cited papers are:

[62] Von Lackum K, Babb K, Riley SP, Wattier RL, Bykowski T, Stevenson B: Functionality of Borrelia burgdorferi LuxS: the Lyme disease spirochete produces and responds to the pheromone autoinducer-2 and lacks a complete activated-methyl cycle. Int J Med Microbiol 2006, 296, 92-102 -and-
[4] Babb K, von Lackum K, Wattier RL, Riley SP, Stevenson B: Synthesis of autoinducer 2 by the lyme disease spirochete, Borrelia burgdorferi. J Bacteriol 2005, 187, 3079-3087

I need to read more about it, at this point the above is currently hypothetical and an in vitro test, so the answer to your question is (unless you know something I don't): the jury is still out on this one.
.
Source Reference:
Guozhou Chen, Lee R. Swem, Danielle L. Swem, Devin L. Stauff, Colleen T. O'Loughlin, Philip D. Jeffrey, Bonnie L. Bassler, Frederick M. Hughson. A Strategy for Antagonizing Quorum Sensing. Molecular Cell, Volume 42, Issue 2, 199-209, 22 April 2011 DOI: 10.1016/j.molcel.2011.04.003

4) Get a whiff of this: Low-cost sensor can diagnose bacterial infections

Link: http://www.sciencedaily.com/releases/2011/04/110427171636.htm

Colorimetric sensor array
overlaid on petri dish

ScienceDaily (2011-04-28) -- Bacterial infections really stink. And that could be the key to a fast diagnosis. Researchers have demonstrated a quick, simple method to identify infectious bacteria by smell using a low-cost array of printed pigments as a chemical sensor. In only a few hours, the array not only confirms the presence of bacteria, but identifies a specific species and strain. It even can recognize antibiotic resistance -- a key factor in treatment decisions.

Comments: So the abstract for this paper is as follows:

"Rapid identification of both species and even specific strains of human pathogenic bacteria grown on standard agar has been achieved from the volatiles they produce using a disposable colorimetric sensor array in a Petri dish imaged with an inexpensive scanner. All 10 strains of bacteria tested, including Enterococcus faecalis and Staphylococcus aureus and their antibiotic-resistant forms, were identified with 98.8% accuracy within 10 h, a clinically important time frame. Furthermore, the colorimetric sensor arrays also proved useful as a simple research tool for the study of bacterial metabolism and as an easy method for the optimization of bacterial production of fine chemicals or other fermentation processes."

The full text requires paid access, however, just looking at what is known here between the article and abstract, I have to wonder how accurate a test this could be to detect Borrelia burgdorferi. I could see this rapid strain identification being useful for identifying bacteria for bacteriophage treatments and also for detecting the presence of bacteria on specific surfaces in hospitals or from open wounds. This wouldn't work well for something that is deeply embedded in collagen, but it might work from a synovial fluid sample better than current detection tests for Bb there.

Source Reference:
James R. Carey, Kenneth S. Suslick, Keren I. Hulkower, James A. Imlay, Karin R. C. Imlay, Crystal K. Ingison, Jennifer B. Ponder, Avijit Sen, Aaron E. Wittrig. Rapid Identification of Bacteria with a Disposable Colorimetric Sensing Array.Journal of the American Chemical Society, 2011; : 110427110353066 DOI: 10.1021/ja201634d

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5 One way to treat Borrelia naturally?

Many Lyme disease patients have used antibiotics for treating Lyme disease and other tickborne coinfections. They have years of scientific study behind them and many reports of patient improvement come from doctors and specialists - patients have had a lot of success with them.

But sometimes antibiotic use leads to various side effects, digestive problems, and potentially, undesirable secondary infection with C. difficile. Using probiotics can often help with digestive problems and prevent C. difficile, but it is not guaranteed.

In some cases - due to allergies or intolerance of the side effects - patients have to stop antibiotic treatment. Because of this, patients have opted at some point in their treatment to stop taking antibiotics after a while and switch to alternative treatments such as herbs.

Whether a patient decides to use antibiotics or herbs, one thing on the horizon seems certain: Eventually antibiotic resistance will lead to more restrictive use of antibiotics, and antibiotic resistance may challenge patients' ability to treat some of their own infections.

However, there is one completely natural possibility that might treat Borrelia and some other tickborne infections in the future which is rarely mentioned in the west other than as a curiosity - yet everyone in the world is surrounded by this abundant and prosperous source of healing from nature all the time.

Much as there are different probiotic bacteria are found in yogurt and probiotic supplements that Lyme disease patients take -- there are viruses in our environment that are helpful to us.

A lot people think of a few things when they hear the word "virus": they think of H1N1 or the swine flu, colds, herpes, HIV, and meningitis, for a start. Not good things. But like the probiotic bacteria that we consume in yogurt all the time, viruses are also present in our environment - in our food, our soil, our drinking water, and our own digestive systems.

Like adding probiotic mixes to your yogurt, these helper viruses have been approved by the FDA to be sprayed on the surface of cheese across the US in order to prevent the development of the bacteria, Listeria monocytogenes, from causing serious disease in pregnant women,  immunocompromised people such as cancer patients, and those with immuno-deficiences. Thousands of people can be severely sickened by Listeria and in some cases even die. So the use of these viruses in food such as cheese is beneficial.

In addition to providing protection from harmful bacteria in food, these helpful viruses have also been used to help save baby calves from dying of diseases which cause severe diarrhea and prevent salmonella from colonizing chickens.

The method for treating these cases was find out which bacterial strains the animals were infected with in order to find the viruses which would eat them. Then use these viruses just as they are found in nature, with no genetic engineering required - put the viral material in pills, injections, or lotions in order to treat the infection.

So this leads one to wonder if this all-natural, non-GMO treatment which is low-cost compared to antibiotics and so abundant in nature can kill off bacterial infections in animals - why can't they kill off infections in people too?

Well, they can.

Watch the next two videos, paying special attention to the first video.


The first video is a 48 minute BBC documentary on the use of viruses to kill bacteria, also known as "bacteriophage therapy" in the former Soviet republic of Georgia, in the Eliava Institute of Tblisi.

Note that if the institute seems run down, filming was done after the collapse of the Soviet Union and the hospital just came out of a civil war - thus buildings had poor maintenance, but the technology to use bacteriophage therapy was in place and used. (After a period of economic instability and social problems - followed by the Rose Revolution - Georgia and Tblisi have been doing much better in the past several years.)

So this documentary is a little dated but general principles remain the same - it explains very well what bacteriophage therapy is and how it has been used in Europe for over 60 years through the 1990's (it continues to be used today - more on recent research using phages will be posted this week).

Youtube (3 parts)

BBC Horizon - 1997 - The Virus That Cures

This second video is from Canadian television as well as CBS news and is more recent - it contains two clips back to back about two people who were treated with phage therapy and their results. Don't miss it - the results are amazing when you realize the initial prognosis each patient was given.

Case studies on phage treatment plus Evergreen College, 

Washington State phage research - [Time: 9:26 minutes]

Is bacteriophage therapy this effective? Does it have any pitfalls? Why don't we hear more about it here yet, given the rising number of cases of antibiotic resistance to deadly bacteria such as MRSA? What can it treat so far? How can this treatment help Lyme disease patients in the future? Here's just one more video just to get a different angle on it from Australian news (Channel 7 and Channel 9). It talks more about history, plus business investments and projections for human trials...

The Forgotten Cure - on Sunday Sunrise, Channel 7 - 

plus a short clip on phages from Channel 9

More on this later this week - for now, check out the videos and let me know what you think, including your own questions and concerns about this kind of medical treatment. [CO note: Continue reading part two of this series, "Phage Therapy and Borrelia burgdorferi".]

This work by Camp Other is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported License.

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