0 Institute of Medicine Final Report on October 2010 Tickborne Disease Workshop

Back in October 2010, the Institute of Medicine (IOM) held a workshop which was broadcast online live (and remains available at TV worldwide), Critical Needs and Gaps in Understanding: Prevention, Amelioration, and Resolution of Lyme and Other Tick-Borne Diseases: The Short-Term and Long-Term Outcomes.

The workshop participants were members of the Institute of Medicine, various researchers, doctors, and members of the Lyme disease patient advocacy community.

A preliminary summary report on the workshop was published by the IOM in April 2011. Now, an official final report has been published and is available on PubMed:

http://www.ncbi.nlm.nih.gov/pubmed/21977545

For a more detailed table of contents, try:

http://www.ncbi.nlm.nih.gov/books/NBK57020/

Editors: Committee on Lyme Disease and Other Tick-Borne Diseases: The State of the Science.

Source: Washington (DC): National Academies Press (US); 2011.
The National Academies Collection: Reports funded by National Institutes of Health.

Excerpt

It was obvious to participants at the workshop that a significant impasse has developed in the world of Lyme disease. There are conflicts within and among the science; policy; politics; medicine; and professional, public, and patient views pertaining to the subject, which have created significant misunderstandings, strong emotions, mistrust, and a game of blaming others who are not aligned with one’s views. Lines in the sand have been drawn, sides have been taken, and frustration prevails. The “walk in the woods” process of conflict resolution or a similar process seems necessary for creating a new environment of trust and a better environment for more constructive dialogue to help focus research needs and achieve better outcomes. Such a process does not imply a compromise of the science but rather is needed to shift to a more positive and productive environment to optimize critical research and promote new collaborations.

---

I'd have to say this is a good report for those who are new Lyme disease and other tickborne illnesses to read in order to get an idea of what issues concern researchers and patients.

In terms of an action item plan and treatment to help patients, though, this report is lacking in either and what is sorely needed at this point in time.

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0 Phage Therapy and Borrelia burgdorferi

EDITED February 27, 2012 to include information on specific phages of B. burgdoferi.

Earlier this week, we discussed the use of phage therapy - the medical use of viruses found in nature that kill bacteria.

Phage therapy has been a part of regular medical treatment in Eastern Europe for over 85 years, but most of the research published has been in the Russian and Georgian languages since the primary former Soviet institution for the research and collection of a huge phage library has been in Tblisi, Georgia.

Eliava Institute of Tblisi, Georgia -
major bacteriophage research center

Those familiar with the Georgian language have stated that detailed documentation for double-blind controls was lacking in research mentioned, so the work as a whole wasn't taken seriously once translated. However, if research came from patient case studies, then documentation wouldn't require blind controls and simply record individual patients' responses. Either way, it is unknown to me how much of the research has been translated or has been made available for translation, given many people do not speak Georgian and because part of the research was written in the era of censorship in Soviet Georgia, some research may not have been published at all - even in a Russian translation.

Tbilisi's Eliava Institute, however, is not the only place in Eastern Europe that has conducted phage research - the Polish Academy of Science has a special institute that is also involved in phage research and therapy. You can learn about their current research here:
The Ludwik Hirszfeld Institute of Immunology and Experimental Therapy (Polish Academy of Science) and read specific research papers in English right here: Evergreen College Guide to Polish Phage Research.

Both of these institutions have had success in treating local patients as well as visitors from abroad. And with growing antibiotic resistance worldwide, one has to wonder why is it phage therapy isn't being used in the west to treat more patients? Why isn't it being used to treat Borrelia burgdorferi, the bacteria which causes Lyme disease?

These are two different questions, one of history and politics, and one of science. To explore them both requires a bit more backstory and examination of the FDA's regulations regarding the adoption of new medical therapies.

In the 1990's, entrepreneurs from the US and Canada traveled to the Eliava Institute to investigate their use of phage therapy and see if they could use the same medical treatment to help patients in the United States. Due to the FDA's regulatory system on all new therapies - especially combination or "cocktail" drug therapies - the use of phage therapy on patients in the United States would be a long way off, and any company investing in phage therapy would be using it for other purposes first.

As a result, in the United States, phage therapy is being used as a spray to protect all kinds of food (the FDA approved of treating cheese first, then other foods) from developing Listeria monocytogenes, bacteria that can lead to severe infection and sometimes even be fatal in vulnerable populations. There have also been treatments developed for veterinary healthcare, such as ear drops for dogs to treat ear infections (otitis media), and the most recent application of phages is using them on surgical equipment and clinic surfaces.

The road to adopting phage therapy for use on treating people in the western hemisphere has been a somewhat rocky one, given that the first entrepeneurs who went to Tblisi and came back to form a phage therapy research startup company had a bit of a falling out: The main financial backer for the company, Canadian Caisey Harlingten, was rumored to have had arguments over who would receive patent rights on work created with the company's new CEO, Richard Honour, and Honour decided to shut down work being done at the Eliava Institute and develop genetically modified phages in the US.

After this, personnel which had been recruited from Tblisi to go work in the United States for Harlingten's company were not happy with this arrangement, jumped ship, and went on to form their own startup, Intralytix. Intralytix - unlike other pioneering phage startups - decided to focus on phage treatments for animals and general products instead of human therapy.

After three years of operating at a loss, Caisey Harlingten resigned from his company, Phage Therapeutics - as did Richard Honour and the chief financial officer.

Last I read, Phage Therapeutics was supposed to have a particular phage that kills 93% of a broad spectrum of over 1,000 of S. aureus and S. epidermidis strains that were isolated from patients in the US, Canada, and South America. This phage was supposed to have been in preclinical trials and was supposed to enter clinical trials against eye infections.

But somewhere along the line, Phage Therapeutics changed hands, their stock devalued, and I discovered that as of February 22, 2008, Phage Therapeutics International Inc. was acquired by Surge Solutions Group, Inc. in a reverse merger. SSGI, Inc., through its subsidiary, Surge Solutions Group, Inc., provides construction and environmental services in Florida. Nothing to do with phage technology. What happened to the above mentioned broad spectrum phage mix?

Where one company falls, others spring up to take their place. There are a growing number of startups in the phage business, but mostly doing business like Novophage, which specializes in using phages to remove biofilms from industrial equipment.

The first clinical trials using phage therapy were conducted in Europe and America. One clinical trial involved a cocktail of eight bacteriophages (five against Pseudomonas aeruginosa, two against Staphlococcus aureus, and one against Escherichia coli) on leg ulcers in 2008 at The Wound Care Center in Lubbock, Texas.  Following that trial, the Southwest Regional Wound Care Center used bacteriophages along with other methods to treat antibiotic-resistant infections under a limited study. Further information on this study has not been published to date.

Bacteriophages are being studied in fighting against E. coli infections in Bangladesh, and phase 2a clinical trials in the UK have been conducted for using phage therapy on chronic inner ear infections caused by Pseudomonas aeruginosa at the Royal National Throat, Nose, and Ear Hosptial in London. Very positive results on clinical and bacteriological efficiency and safety concerns have been reported on this latter trial.

In 2010, a nebulizer treatment using bacteriophages of Burkholderia cepacia complex (full text) to treat cystic fibrosis was developed, and earlier study was completed on the development of an inhaler to treat Staphylococcus aureus or Pseudomonas aeruginosa. So far, the inhalers have yet to be tested on people.

There is an international conference on bacteriophages that is held in Olympia, Washington, and hosted by Evergreen College. Dr. Elizabeth Kutter, professor of microbiology at the college took a keen interest in bacteriophage therapy years earlier, and had traveled to Tbilisi herself to investigate the treatment and their results. Since then, she has been actively pursuing research into bacteriophages and promoting it for use in medicine. The college has its own special phage projects page you can look at to see research conducted on phage therapy around the world.

Even though there is interest in bacteriophages, few clinical evaluations have been published on them because the data available are at a very early stage, making it difficult to attract further funding - and as mentioned earlier, the use of phage often involves a "cocktail" of more than one virus to treat a patient and this challenges the FDA's regulatory standpoint on cocktail treatments.

Also, using phage therapy in Eastern Europe focused mainly on treatment for wounds and intestinal infections - conditions which could be treated using phages topically in ointments, sprays, and dressings or capsules and enemas. Intravenous therapy (IV) - while used on occasion - did not make up the majority of treatments given, so little has been known about their effectiveness.

There is some evidence that phage therapy can work in IV therapy, but it was suggested that in this form it is more likely to come with a drawback: just as Lyme disease patients experience a Herxheimer reaction from antibiotic therapy, patients receiving phage therapy can also have a Herxheimer reaction from phage therapy. One veterinary study, though, has shown that no notable negative reactions or effects were noted (Soothill, 2004).

As as a commenter on my previous post mentioned, there are shortcomings as well as benefits to the use of phage. But overall, the risks of using phage therapy seem lower than those of antibiotics so far because the antibiotic resistance issue and risk of C. difficile are gone (someone is even working on phage therapy for C. difficile).

Despite the growing evidence that phage therapy can be safe and effective, there are some challenges that even people who are most unfamiliar with phage therapy have pointed out at least one of them:

• We don't know much about how phages interact with gut flora. Suspicions are most are benign if not helpful because we already have bacteriophages living in our stomachs and intestines all the time.
• Some research has shown one kind of phage - T-even bacteriophage - show inhibition of lysis in low-oxygen environments. 
• Both carbohydrates and bile salts can interfere with bacteriophages ability to replicate in the stomach. 
• If a bacteriophage that was lytic becomes lysogenic, it will integrate with its host, enabling it to transfer bacterial virulence genes into other bacteria. This is why therapeutic phages must be entirely lytic and cannot carry toxic or housekeeping genes associated with lysogeny.

Even though these drawbacks exist, research is underway to find solutions that address them because the risk of not having phage therapy can be worse for some patients with very deadly infections which are becoming increasingly antibiotic resistant.

Can phage therapy work on killing Borrelia burgdorferi?

So far I have not seen any phage therapy research for Borrelia burgdorferi - however, the Phage Therapy Center for patients in Tblisi, Georgia claims they have phage therapy to treat Lyme disease coinfections.

In terms of phage therapy for Lyme disease itself, though - the best answer I can give at this writing is a theoretical maybe someday.

This is based on the idea that there is a phage for every bacteria out there if we were only to look for it and find it. It's also based on the idea that we have the technology available to potentially modify Lyme disease's known phages in order to change its behavior - or perhaps create a delivery system which could lyse Borrelia in a manner that phage does.

But so far - unlike Staphloccocus and other bacteria - few phages which attack and kill Borrelia have been documented. Publications on virulent phages of Borrelia are sparse, and there is only a little more documentation on phages in spirochetes as a whole.

B3-like morphology
phage on spirochete

In 1982, Hayes, Burgdorfer, and Barbour recorded their observations of a phage attacking Borrelia burgdorferi in vivo and took photographs to record the event. The images captured are of a B3-like bacteriophage, described by the researchers as having a "40- to 50-nm elongated head and a tail 50 to 70 nm in length. It appears devoid of collars or kite-tail structure".

There are two aspects of these images below which are  compelling: One is that they give us a rare glimpse of a phage which can actually kill Borrelia burgdorferi. (Wouldn't it be fabulous if we could somehow find a way to harness this as a treatment method, and find phages for all strains of Borrelia?) The second is that we have a photo of gemmae - a form of Borrelia which is not mentioned much in today's genomic oriented Borrelia research.

•  (a) Section profile of a gemma with its attendant membrane bound granules or spherical bodies. Arrows indicate cross-section profiles of bacteriophage heads. (b) Internal attachment of bacteriophage to outer membrane material after plasmolysis of the spirochete. Arrows indicate remnants of plasma membrane.

A passage within the text, "Bacteriophage in the Ixodes dammini Spirochete, Etiological Agent of Lyme Disease", sheds some light on what is known about this phage and its relationship to Borrelia burgdorferi:

"Thus far, only those spirochetes showing left-handed coiling have been found to be phage infected. Figure ld shows phages that are associated with a spirochete with left-handed coiling. Bacteriophage heads in longitudinal and cross-sectional profiles were also observed within granules located within the aneurysmic blebs (Fig. 2a).

Completely assembled phages were more clearly seen in rarely occurring plasmolysed cells (Fig. le and 2b). In negatively stained preparations of spirochetes, they have only been detected internally (Fig. 2c). Bacteriophages previously reported to infect other spirochetes (15-17) are described as polyhedral and tailed (7) or cubic (5) in symmetry."

It appears that only those spirochetes which coil in a counterclockwise direction had phages. Why didn't any spirochetes with a clockwise coil have phages? Is there some inherent difference in their surface which makes it harder for phage to adhere to them?

In 1993, Neubert et al wrote about finding phage which were induced while introducing the antibiotic, ciprofloxacin, to Borrelia spirochetes. These A-1 and B-1 type phages were not virulent phages such as Hayes et al's B3-like phage.

The ultimate Borrelia book, "Borrelia: Molecular Biology, Host Interaction and Pathogenesis", has some passing mention of phages of Borrelia as well as a map of known and possible prophages in its plasmids. It also mentions a more recent discovery than Hayes, Burgdorfer, and Barbour's B3-like phage.

phiBB-1, prophage of
Borrelia burgdorferi

In 2001, Eggers et al published their discovery of a phage of Borrelia burgdorferi (Bb) named phiBB-1 (also written as φBB-1). It is not the best candidate for use in bacteriophage therapy because it is a prophage - also known as a temperate phage or lysogenic phage.

Lysogenic phages remain inactive as viruses when they are prophages, and only replicate together with the host genome unless mobilized. In contrast, virulent phages, having replicated and assembled into complete virions, cause rapid lysis and death of the bacterial cell, with release of 10–100 virions per phage; these virions then find more prey and die out when they cannot find any more bacteria.

Every time Borrelia burgdorferi divides, the viruses internalized in its plasmids divide with it. The viruses are an integral part of the plasmids and contribute to the functionality and antigenic variation of the spirochete - they have become part of the bacteria. In technical terms: The phiBB-1 prophage is capable of transducing a cp32 (circular plasmid) between cells of the same isolate and between different Bb isolates (gene transfer between different Borrelia spirochetes). This means this prophage could play a role in the genetic diversity of different Bb isolates.

Lytic-Lysogenic Phage Cycles
image by Suly12, Wikipedia

See the image to the left. If a bacteriophage is virulent, it will deposit its genes into bacteria so that it replicates and kills the bacteria from inside by lysing its membrane. The viruses then continue in search of more of the same bacteria to feast on. This is called the lytic cycle.

But if a bacteriophage is temperate or lysogenic, though - a prophage - then it will deposit its genes into bacteria so that they mix with the bacteria's own genes and divide with them each time the bacteria divides. This is called the lysogenic cycle.

Borrelia burgdorferi's plasmids contain virus genes which are locked into the lysogenic cycle.


Hypotheses Of Altering Phages To Lyse Borrelia

In order to put phiBB-1 to work at killing Bb, someone would have to genetically engineer it or introduce some agent which turns it into a virulent phage that kills Bb rather than adding its own DNA to its plasmids. Or, maybe phiBB-1 could be modified in a different way: don't bother changing its prophage nature, just program it to turn off DNA replication and gene expression in the bacteria's plasmids.

Another thing that could be done is to have someone extract the lysing proteins that work with phiBB-1 and find a method of delivery to Bb so those proteins could go to work on killing Bb outside in - maybe attach it to a non-pathogenic adenovirus that is programmed for such an adventure. There are such delivery systems being experimented with in general right now - but nothing yet for Borrelia.

These are wild hypotheses about how an existing phage we know about could be used to kill Bb, but it is not proven this would work. People are thinking of the biotech applications of phiBB-1 - but so far, I have seen only one patent application referring to its use.

The best option, obviously, would be to find naturally occurring phages which lyse Borrelia burgdorferi (as well as other strains) and find a method for using them to treat patients - though there are likely to be technical challenges in applying this as well.


References:
Wired magazine: http://www.intralytix.com/Intral_News_Wired.htm
A controlled clinical trial of a therapeutic bacteriophage preparation in chronic otitis due to antibiotic-resistant Pseudomonas aeruginosa; a preliminary report of efficacy. Wright A, Hawkins CH, Anggård EE, Harper DR. Clin Otolaryngol. 2009 Aug;34(4):349-57.
Viruses Vs. Superbugs: A Solution to the Antibiotics Crisis? By Thomas Häusler
Soothill, J.S. Hawkins, C. Anggard, E.A. & Harper, D.R. (2004) Therapeutic use of bacteriophages. Lancet Inf. Dis. 4, 544-545.
Microbiologist, the magazine of the Society for Applied Microbiology (June 2009, Vol.10 No.2)
Bacteriophage Therapy: Exploiting Smaller Fleas. Stan Deresinski. Clin Infect Dis. (2009) 48 (8): 1096-1101. doi: 10.1086/597405 link: http://cid.oxfordjournals.org/content/48/8/1096.full
Bacteriophage in the Ixodes dammini Spirochete, Etiological Agent of Lyme Disease. Stanley F. Hayers, Willy Burgdorfer, Alan G. Barbour. Journal of Bacteriology, June 1983, p. 1436-1439. link: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC217620/pdf/jbacter00247-0414.pdf
Demonstration of Cotranscription and 1-Methyl-3-Nitroso-Nitroguanidine Induction of a 30-Gene Operon of Borrelia burgdorferi: Evidence that the 32-Kilobase Circular Plasmids Are Prophages. Hongming Zhang and Richard T. Marconi. Journal of Bacteriology. December 2005, Vol. 187, No. 23 p. 7985-7995.
Bacteriophages induced by ciprofloxacin in a Borrelia burgdorferi skin isolate. Neubert U, Schaller M, Januschke E, Stolz W, Schmieger H. Zentralbl Bakteriol. 1993 Aug;279(3):307-15. link: http://www.ncbi.nlm.nih.gov/pubmed/8219501 Bacteriophage-like particles associated with a spirochete. Berthiaume L, Elazhary Y, Alain R, Ackermann HW. Can J Microbiol. 1979 Jan;25(1):114-6.
link: http://www.ncbi.nlm.nih.gov/pubmed/427652
http://en.wikipedia.org/wiki/Lysogenic_cycle

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13 Books: Borrelia. Plus a lesson in terminology.

Borrelia: Molecular Biology, Host Interaction and Pathogenesis. If you have GB £159 or US $310 on hand, and you want to know the state of the science on Borrelia including Borrelia burgdorferi, afzelii, garinii, and its relapsing fever relatives - this is a good book to get.

If you would prefer to review the book before purchasing - or don't want to purchase it at all - check out your local library's interlibrary loan program.


See a detailed chapter outline at this link: http://www.horizonpress.com/borrelia

I have yet to do a review on this book as it is information dense and takes some time to read through - but I pulled some highlights from the chapter outlines (it is pretty robust for an outline, I have to admit)  and provided some vocabulary translation in which some readers may take an interest:

"B. burgdorferi strain B31, the B. burgdorferi type strain, has been studied in the most detail and harbors twelve linear and nine circular plasmids that comprise about 612 kbp. The plasmids are unusual, as compared to most bacterial plasmids, in that they contain many paralogous sequences, a large number of pseudogenes and, in some cases, essential genes. In addition, a number of the plasmids have features suggesting that they are prophages. Some correlations between genome content and pathogenicity have been deduced and comparative whole genome analyses promise future progress in this arena. [CO comment: Refer to Ben Luft's and Steven Norris' research on Bb strains and their ability to cause infection in human hosts.]"

In general, the whole plasmid thing is just weird. Having plasmids that take up one third of your genome is interesting.

Circular plasmids? No problem. You find them in bacteria all the time.

Linear plasmids? That is a bit unusual. Linear plasmids used to be thought of as only in eukaryotic organisms - organisms with a cell nucleus - not prokaryotic organisms like bacteria. Linear plasmids are also found in viruses. But they are found in few bacteria - Borrelia burgdorferi being one of them.

But prophages...  These are very interesting, and what makes some Borrelia strains more harmful to their hosts. That viral genes end up in Borrelia plasmids is pretty fascinating to me, even though this sort of thing happens with other bacteria too - it's fascinating and adds to Borrelia's existing complexity.

Translation for biology/genetics beginners:

(You might want to check this out sometime: http://en.wikipedia.org/wiki/Introduction_to_genetics .)

chromosome = A chromosome is an organized structure of DNA and protein that is found in cells. It is a single piece of coiled DNA containing many genes, regulatory elements and other nucleotide sequences.

plasmid  = Usually described as a double-stranded unit of DNA that replicates within a cell independently of the chromosomal DNA. Three of the plasmids found in all Borrelia burgdorferi isolates have been described as "mini-chromosomes".

Your handy little Bb plasmid cheat sheet:
cp = circular plasmid
lp = linear plasmid
cp26 = needed for viability, encodes OspC.
lp25 and lp28-1 = needed for infectivity in mice; lp28-1 involved in antigenic variation.
cp32 = contains prophage material; encodes a BpaB protein, which appears to play a role in replication and segregation.
lp54 = encodes OspA/B operon and decorin-binding protein operon dpbBA

Other Borrelia plasmids (relapsing fever):
cp32 = described in B. hermsii; does not contain an OspE/F/Elp lipoprotein (unlike Bb)

Plasmids make up about a third of Borrelia burgdorferi's entire genome. (Fraser et al, 1997; Casiens et al, 2000). Linear plasmids are the most genetically diverse part of Bb's genome, and rearrangements and deletions inside them contribute to their uniqueness.

In Bb, there is a division of labor between the chromosome and extrachromosomal elements or “plasmids”. Genes encoded on the chromosome tend to do housekeeping - housekeeping genes are present in single copy - while genes on the plasmids tend to be Borrelia-specific, of unknown function or involved in the infective cycle, and present in multiple, related copies.

Source material:
The Prokaryotes: A Handbook on the Biology of Bacteria (Vol. 7) by Martin Dworkin and Stanley Falkow.
The Linear Hairpin Replicons of Borrelia burgdorferi. Kerri Kobryn. Microbiol Monogr (7) Universite de Sherbooke, Sherbrooke, QC, Canada

genome = the entirety of an organism's hereditary information. It is encoded either in DNA or, for many types of virus, in RNA.

pseudogene = dysfunctional relatives of known genes that have lost their protein-coding ability or are otherwise no longer expressed in the cell. Also known as mutationally damaged or "Junk DNA" - it is not always necessarily "junk" - its function is simply unknown at this time.

essential gene = genes that are indispensable to support cellular life. These genes constitute a minimal gene set required for a living cell - if they don't work, the organism dies.

paralogous = This one takes a little more explaining...

Homologous gene sequences = the same, where "homo" means "like, similar" and "hetero" means "different, divergent".

For example, in general, if two or more genes have highly similar DNA sequences, it is likely that they are homologous. "Paralogous" means they are homologous gene sequences which are similar but they occupy different positions or locations in the same genome.

Sequence similarity is often seen in organisms that evolved from the same common ancestor. However, sequence similarity may also arise without common ancestry - short sequences may be similar by chance, and sequences may be similar because both were selected to bind to a particular protein, such as a transcription factor. Such sequences are similar but not homologous.

prophage = Okay... This is where things get weird. A prophage is a phage (viral) genome inserted as part of the linear structure of the DNA chromosome of a bacterium. A temperate phage is integrated into the host chromosome or existing as an extrachromosomal plasmid. This is a latent form of a bacteriophage, in which the viral genes are incorporated into the bacterial chromosomes without causing disruption of the bacterial cell.

In other words, a bacteriophage is a virus that lives inside bacteria, and a prophage is viral genetic material that becomes part of the chromosome in the bacteria (see virus to left, injecting its genetic material inside a non-spirochetal bacteria). So, in this case, at some point in time, a phage known as phiBB-1 (also spelled φBB-1) made its viral genes part of Borrelia burgdorferi's plasmids.

In prophages in general, if the host bacteria is damaged, the prophage is excised from the bacterial chromosome in a process called prophage induction. After induction, viral replication begins via the lytic cycle.

Prophages are important agents of horizontal gene transfer, and are considered part of the mobilome.

In many bacterial species, prophages figure prominently in the biology of these cells, often conferring key phenotypes that can convert a non-pathogenic strain into a pathogen. In other words, while it's residing inside the bacteria, the prophage can turn bacteria that is harmless into harmful bacteria.

Such phenotypic changes can include prophage-encoded toxins, bacterial cell surface alterations, or resistance to the human immune system.

Prophage integration into the host genome can inactivate or alter the expression of host genes. In addition to these direct genetic alterations associated with the addition or inactivation of genes, prophages can also alter the phenotype of bacteria at the population level by facilitating the spread of favorable genes through transduction. (transduction = the process by which DNA is transferred from one bacterium to another by a virus.)

Its significance in Borrelia burgdorferi:

It's not discussed much on Lyme disease patient forums, but some strains of Borrelia burgdorferi have had their plasmids invaded by prophages. The presence of those phages - their viral genetic material - can make Borrelia burgdorferi more pathogenic.

phage particles in Borrelia burgdorferi
from Eggers et al

The phiBB-1 prophage is capable of transducing a cp32 between cells of the same isolate and between different Bb isolates. This means this prophage could play a role in the genetic diversity of different Bb isolates.

The structural proteins for phiBB-1 have not yet been identified, and a number of proteins encoded on cp32 have no homologs in any databases outside of Bb.

cp32 expresses a few outer surface or membrane proteins, of which ospE is the only one of which its function is known. It binds to complement factor H and helps Bb evade the complement system of the host.

A number of different bacteriophages have been observed in association with spirochetes and are being researched now. It may contribute to Lyme disease's ability to persist.

Well-known research on this includes this study:
Eggers et al http://www.ncbi.nlm.nih.gov/pubmed/11466280

Source material:
Wikipedia: http://en.wikipedia.org/wiki/Prophage
Wikipedia: http://en.wikipedia.org/wiki/Introduction_to_genetics
Borrelia: Molecular Biology, Host Interaction and Pathogenesis.
The Prokaryotes: A Handbook on the Biology of Bacteria (Vol. 7) by Martin Dworkin and Stanley Falkow.

pathogenicity = the ability of a pathogen to create infectious disease in an organism.

One of the relevant points to be gleaned from the above is that the genetic makeup of the plasmids are related to the virulence and pathogenicity of Borrelia. (virulence = the degree of pathogenicity within a group or species of microorganisms or viruses as indicated by case fatality rates and/or the ability of the organism to invade the tissues of the host.)

"The highly unusual segmented genomes of Borrelia species can contain over 20 autonomously replicating DNA molecules. Many of the molecules, including the chromosome, are linear with covalently closed hairpin ends. Current knowledge of the replication and maintenance of DNA molecules will be reviewed, including the process of telomere resolution, whereby the covalently closed hairpin ends are generated from replicative intermediates. Finally, the proposal that reverse telomere resolution is the driving force shaping the ongoing rearrangements and telomere exchanges in the linear replicons of Borrelia species will be discussed."

Translation for biology/genetics beginners:

chromosome = A chromosome is an organized structure of DNA and protein that is found in cells. It is a single piece of coiled DNA containing many genes, regulatory elements and other nucleotide sequences.

nucleotide = molecules that when joined together make up the structural units of RNA and DNA. In addition, nucleotides play central roles in metabolism.

covalently closed hairpin ends = strands of genetic material curve back on itself in a hairpin pattern.

telomere = a region of repetitive DNA at the end of a chromosome, which protects the end of the chromosome from deterioration. The telomere shortening mechanism normally limits cells to a fixed number of divisions, so a shorter telomere means there are fewer divisions left for that cell and it will die sooner. Animal studies suggest that this is responsible for aging on the cellular level and sets a limit on lifespans.

telomere resolution = when the replicated linear DNA ends are processed by DNA breakage followed by joining of DNA free ends to the complementary strand of the same DNA molecule.

reverse telomere resolution = when linear DNA cleaves (splits) and joins hairpin telomeres on unrelated (rather than related) DNA molecules. The new linked plasmid structure could fuse to another linear plasmid in the future, and this explains how Borrelia can engage in telomere exchanges. This process may play a major role in the development of weird linear plasmids in Borrelia and their ability to confuse the immune system.

replicon = a DNA molecule or RNA molecule, or a region of DNA or RNA, that replicates from a single origin of replication. For most prokaryotic chromosomes, the replicon is the entire chromosome.

This part, I think most readers will know about the last sentence - but suspect they will be less familiar with the first half:

Borrelia are not Gram-positive...  But... they are not exactly Gram-negative bacteria, either.

"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."

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

Translation for biology/genetics beginners:

Gram-negative = Gram-Negative Bacteria are simply called this because of their detection by the Gram’s Stain test in which they do not retain the crystal violet color (dye) in their cell wall. The Gram-Negative bacteria cell-wall holds the pink or reddish dye once a counterstain chemical is used. This is characteristic of bacteria that have a cell wall composed of a thin layer of a particular substance (called peptidoglycan).

diderm = double-membrane envelope consisting of an inner cytoplasmic membrane and outer membrane found in Gram-negative bacteria.

double-membrane envelopes = See the picture to the left. Monodermic bacteria (mono = one) have one membrane, and didermic bacteria have two (di = two) membranes - the inner membrance (cm) and outer membrane (om).


LPS = Lipopolysaccharides (LPS), also known as lipoglycans, are large molecules consisting of a lipid and a polysaccharide joined by a covalent bond; they are found in the outer membrane of Gram-negative bacteria, act as endotoxins and elicit strong immune responses in animals. [CO comment - a question for my readers: If Borrelia burgdorferi do not have LPS, what is producing the endotoxins everyone in the Lyme patient community says are important to detox? Keep reading, the answer is downstream...]

lipid = Lipids are a broad group of naturally occurring molecules which includes fats, waxes, sterols, fat-soluble vitamins (such as vitamins A, D, E and K), monoglycerides, diglycerides, phospholipids, and others. The main biological functions of lipids include energy storage, as structural components of cell membranes, and as important signaling molecules. In this context, lipids make up the inner membrane of Borrelia burgdorferi.

polysaccharide = Polysaccharides are polymeric carbohydrate structures, formed of repeating units (either mono- or di-saccharides) joined together by glycosidic bonds. Examples include storage polysaccharides such as starch and glycogen, and structural polysaccharides such as cellulose and chitin.

lipoproteins = A lipoprotein is a biochemical assembly that contains both proteins and lipids water-bound to the proteins. Many enzymes, transporters, structural proteins, antigens, adhesins and toxins are lipoproteins.

(a) external side view of Borrelia burgdorferi spirochete,
(b)  head-on view of cross-section of Bb, and (c)
side view of cross-section of Bb's diderm membranes.  

periplasmic = The periplasmic space or periplasm is a space between the inner cytoplasmic membrane and external outer membrane of Gram-negative bacteria or the equivalent space outside the inner membrane of Gram-positive bacteria. It may constitute up to 40% of the total cell volume in Gram-negative species, and is drastically smaller in Gram-positive species.

flagella = A flagellum is a tail-like projection that protrudes from the cell body of certain prokaryotic and eukaryotic cells, and functions in locomotion. (Flagella is plural of flagellum.)

Osps = Outer surface proteins. The outer membrane of Borrelia burgdorferi is composed of various unique outer surface proteins (Osp) that have been characterized (OspA through OspF). The Osp proteins are lipoproteins anchored by N-terminally-attached fatty acid molecules to the membrane. They are presumed to play a role in virulence, transmission, or survival in the tick.

ligand = Any substance (e.g. hormone, drug, functional group, etc.) that binds specifically and reversibly to another chemical entity to form a larger complex.

"Several borrelial proteins have been implicated in adherence to host cell surface proteins and extracellular matrix components and are likely to be involved in the homing of Borrelia to histologic compartments within each tissue, penetration of blood vessels and adherence to and migration through endothelial cells and tissue strata at distant sites. Activation of plasmin on the bacterial surface and induction of host proteases are thought to facilitate dissemination and/or inflammation. Most tissue damage appears to result from host inflammatory reactions. Although the mechanisms are not entirely understood, induction of cytokine/chemokine expression by bacterial lipoproteins and the resulting recruitment and activation of lymphocytes, macrophages and granulocytes play a major role in both local histopathology and constitutional symptoms. Despite their relatively low densities in tissues, Borrelia cause neurologic, cardiovascular, arthritic and dermatologic manifestations during the disseminated and persistent stages of infection by mechanisms that remain largely a mystery. Immune evasion mechanisms, including the vls antigenic variation system, complement-regulator acquiring surface proteins (CRASPs), down-regulation of highly antigenic surface proteins (such as OspC) and invasion of protective niches, permit the survival of the pathogens for months to years following infection despite robust antibody and cellular responses."

Translation for biology/immunology/genetics beginners:

host cell surface proteins = proteins on the surface of the host's cell.

extracellular matrix = Any material produced by cells and secreted into the surrounding medium, but usually applied to the noncellular portion of animal tissues. In other words - the space around the outside of cells, but not within cells.

histologic = related to cells and tissue on the microscopic level.

endothelial cells = cells that lines the interior surface of blood vessels, forming an interface between circulating blood in the lumen and the rest of the vessel wall.

plasmin = an enzyme present in blood that breaks down many blood plasma proteins, most notably, it breaks down fibrin clots (blood clots).

protease = an enzyme that breaks down proteins.

cytokine = small cell-signaling protein molecules that are secreted by the glial cells of the nervous system and by numerous cells of the immune system and are a category of signaling molecules used extensively in intercellular communication.

The term "cytokine" has been used to refer to the immunomodulating agents, such as interleukins and interferons. Biochemists disagree as to which molecules should be termed cytokines and which hormones. As we learn more about each, anatomic and structural distinctions between the two are fading.

chemokine = are a family of small cytokines, or proteins secreted by cells. Their name is derived from their ability to induce directed chemotaxis in nearby responsive cells; they are chemotactic cytokines. Chemotaxis is simply the act of cells changing their movement according to certain chemicals in their environment.

lymphocyte = White blood cells. Large lymphocytes include natural killer cells (NK cells). Small lymphocytes consist of T cells and B cells.

macrophage = white blood cells which phagocytose (engulf and then digest) cellular debris and pathogens, either as stationary or as mobile cells. They also stimulate lymphocytes and other immune cells to respond to pathogens.

granulocyte = a category of white blood cells characterized by the presence of granules (meaning "grains") in their cytoplasm (inside the cell membrane). These are thought of as the "phil" brothers of white blood cells: neutrophils, eosinophils, and basophils.

histopathology = the microscopic examination of tissue in order to study the manifestations or signs of disease.

vls = Gene locus in Borrelia burgdorferi which is required for encoding variable surface proteins.

vlsE = VlsE is a lipid-protein conjugate, found on the cell's outer surface during all Borrelia life stages. It is similar to a lipoprotein of the organism that causes African sleeping sickness. Unlike most proteins, VlsE is produced in many forms. It is a complicated protein with several variable regions (VRs), and six invariable regions (IRs).

When synthesizing VlsE, Borrelia periodically replace the VRs with new sequences. This replacement presents fresh surface antigens, and helps Borrelia remain invisible to the immune system. Within four days of being transferred to a mammalian host, VlsE will be produced with more than one VR suite, reducing the strength of the immune response. In ticks, VlsE does not modify the VRs. (Credit to Dr. Albert Burchsted, retired Field Biologist, for this description.)

antigenic variation = the mechanism in which an infectious organism alters its surface proteins in order to evade a host immune response. This change in antigenic profile may occur as the pathogen passes through a host population (also called "antigenic diversity") or may take place in the originally infected host.

The strategy is particularly important for organisms that a.) target long-lived hosts, b.) repeatedly infect a single host, and c.) are easily transmitted. Pathogens that express these characteristics and undergo antigenic variation have a selective advantage over their more genetically stable counterparts.

A number of bacteria use antigenic variation to evade the immune system, but Borrelia burgdorferi's method of antigenic variation is very complex and unusual compared to these.

In the case of Borrelia burgdorferi, the lp28-1 plasmid has been responsible for antigenic variation in vls locus - responsible for vlsE and the changing of Borrelia's outer surface proteins which helps it evade the immune system.

During the past decade, it was thought that removing the lp28-1 plasmid or altering its genetic material would remove Borrelia's infectivity entirely. It reduced it, but removing specific genes near the vlsE locus has done more to reduce Borrelia burgdorferi's infectivity.

The mutation of either of the genes encoding the two subunits of the RuvAB branch migrase blocked transfer of genetic information into vlsE during mouse infections, identifying the first required function for antigenic variation in the Lyme disease spirochete.

But basically, both the cis arrangement of vlsE and the vls silent cassettes in lp28-1 facilitate vlsE gene conversion.

See:
http://www.ncbi.nlm.nih.gov/pubmed/15501789
http://www.ncbi.nlm.nih.gov/pubmed/19266024
http://www.ncbi.nlm.nih.gov/pubmed/19997508

Source material:
The Linear Hairpin Replicons of Borrelia burgdorferi. Kerri Kobryn. Microbiol Monogr (7) Universite de Sherbooke, Sherbrooke, QC, Canada
Borrelia: Molecular Biology, Host Interaction and Pathogenesis.

complement = a small protein which mediates antibody response in the host's body. This is part of a complex immune system known as the complement system.

CRASPS = Borrelia produces complement regulator-acquiring surface proteins (CRASPs) that bind host complement factor H protein. Factor H protein is a negative regulator of the complement cascade (inactivates C3b). Borrelia produces a whole family of CRASP proteins. Different family members are able to interact with different factor H proteins from divergent hosts.

If that sounds too confusing, the thing to remember is that Borrelia has surface proteins which can bind to a host's complement's protein which then inactivates part of the complement system.

down-regulation = the process by which a cell decreases the quantity of a cellular component, such as RNA or protein, in response to an external variable. (Up-regulation is the opposite process.)

So, as I've stated before - researchers do know that the infection can be persistent - the question and controversy has been over whether or not infection persists after one has antibiotic treatment. 

It's my opinion so far that it can persist - especially if one has neuroborreliosis and has been undertreated or untreated. Those who have been suffering from neuroborreliosis and reading this will say it definitely can persist, based on their experience. The question remains as to how one can know whether or not the infection has been eradicated. This is what needs to be known. 

Also, there is persisting, and then there is persisting... One of my nightmare scenarios has been that when the IDSA Lyme disease guidelines panel states that Lyme disease does not respond to long-term antibiotic treatments, what it really means that none of the antibiotics which exist today can effectively eradicate all of it and the immune system isn't always capable of mopping it up after antibiotic treatment.

This is why people began experimenting with a number of alternative treatments: in their experience and opinion, antibiotic treatment either failed to eradicate the infection or stop their symptoms - and sometimes the side effects or infection with C. difficile led to the decision to stop antibiotic treatment.

Onward...

Lyme Disease in Humans - from the last chapter in Borrelia:

"Lyme disease is a rapidly emerging tick-borne, complex, multi-system infectious disorder caused by the spirochetal bacterium Borrelia burgdorferi. The ailment, which affects adults and children alike, is widespread in the Northern Hemisphere and it continues to expand as humans encroach on the sylvatic habitat of the spirochete's mammalian reservoirs."

"Since first identified in the 1970s the incidence of Lyme disease has increased more than 30-fold and it is now considered the most prevalent arthropod-transmitted infection in both the United States and Europe."

This is why I say doctors should look at the possibility of ruling it in, more than ruling it out...

"B. burgdorferi is transmitted by ticks of the Ixodes ricinus complex, including I. scapularis, I. ricinus and I. persulcatus. In North America, B. burgdorferi sensu stricto is the only species proven to be pathogenic for humans [CO note: Relapsing fever Borrelia are pathogenic to humans too, and some species of Borrelia have yet to be determined for their pathogenicity in humans.]. In Europe, both B. afzelii and B. garinii are most commonly associated with human disease.

The spirochete's genomic features, as well as its unique molecular architecture, are considered to have a seminal role not only in how it is transmitted from ticks to humans, but also how it triggers immune responses in afflicted individuals. Inflammatory manifestations associated with the disease result from the host's innate and adaptive immune responses to the bacterium, rather than from toxigenic molecules, which borrelia cannot produce. Indeed, the deposition of spirochetes into human dermal tissue generates a local inflammatory response that becomes manifest as erythema migrans (EM), the hallmark skin lesion of Lyme disease in North America. In Europe, two additional dermatologic disorders, borrelial lymphocytoma and acrodermatitis chronicum atrophicans (ACA) are frequently associated with infection. EM is frequently accompanied by 'flu-like' symptoms, including myalgias, arthralgias and fever, which are generally believed to be cytokine-mediated in response to hematogenous spread of the bacterium. If treated appropriately, the prognosis is excellent; however, if untreated, patients may develop a wide range of inflammatory clinical manifestations, most commonly involving the central nervous system, joints and heart. Within days of treatment, the signs and symptoms associated with the disease typically begin to subside, although in some individuals a complete recovery can take several weeks or even months. A minority of treated patients may go on to develop a poorly defined fibromyalgia-like illness, which is not responsive to prolonged antimicrobial therapy. Below we integrate current knowledge regarding the ecological, epidemiological, microbiological and immunological facets of Lyme disease into a conceptual framework that sheds light on the disorder that healthcare providers encounter."

According to this, Borrelia burgdorferi does not produce toxigenic molecules, and most symptoms are believed to be cytokine-mediated in response to hematogenous spread of the bacterium. (hematogenous = originating in or spread by the blood). They are talking about all Borrelia there.

Hm. That's interesting. Does that mean that what people think is a Herxheimer reaction due to the release of toxins is actually something else, some other process?

I wanted to know what the deal is with this, so I looked it up... Huh, this was interesting:

Borrelia have abundant glycolipids but they do not have an endotoxin-like lipopolysaccharide. And the authors' running hypothesis is that the Herxheimer reaction is due to the action of lipoproteins on toll-like receptors (TLR2) in macrophages and other cells. Apparently, if you infuse antibodies that recognize TNFa (Tumor Necrosis Factor alpha) into someone who is infected with related Borrelia recurrentis before giving them penicillin, it reduced the severity of a Herxheimer reaction (Fekade et al, 1996 - full text at preceding link). When cytokine levels get quite high, this contributes to the reaction. (Borrelia, p. 339)

So, does this mean that detoxing is useless because there is no toxin to remove? The paper goes on to state that "...lipoproteins from two other spirochetes, B. burgdorferi and Treponema pallidum, induce the biosynthesis of tumor necrosis factor in murine macrophages."

So when patients are having a Herxheimer reaction, it is a reaction to the die-off induced by the immune system - not a reaction induced by a toxin.

Regarding the bit in the last section on persistence: Researchers are somewhat divided on this, and even as I write this, studies are published on the use of long-term antibiotic treatment for some of this minority of treated patients with persistent symptoms.

Dattwyler co-wrote this chapter along with Radolf, and one has to wonder if it would have been written somewhat differently and been left open to more uncertainty had Volkman or Barthold co-written it? Something to think about there.

At any rate...the running hypothesis is that infectivity of Borrelia species depends heavily on the evasion from the host response. And this evasion can be based on multiple mechanisms.

You've gotta just love the closing quote on the end of one chapter in the book...

Not.

“If our work with Borrelia has taught us one thing, it is to expect the unexpected. So we should not be surprised to discover that this phylogenetic ancient spirochaetal microorganism has developed several solutions of  its own and that it does not always conform to the dogmatic structure and function of the cell envelope of Gram-negative bacteria. We therefore anticipate a bright future with many challenges and unsolved mysteries for several generations of Borrelia researchers.”

Yeah. Right. Your science project is my life, man. I'm not looking at this as some professional subject of interest. My focus in this is different from yours. By a longshot, baby. By a longshot.

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27 What happened? The Early Days of Lyme Disease

This could end up being part of a series on the history of Lyme disease, but for now I wanted to share something from Polly Murray's book, The Widening Circle.

I really can't recommend this book enough for readers who are either Lyme patients or care about someone who is a patient - it is a well-written account of one woman's journey of struggling with strange symptoms that were plaguing her and her family for years. Her determination to find the cause, and how her actions led to the discovery of Borrelia burgdorferi in ticks led to all that people in the Lyme disease patient community are familiar with now.

Gandhi once said, "Be the change you wish to see in this world," and I think Ms. Murray's life reflects that statement.

Moving on to the question of "What happened?" in regards to how the introduction of Lyme disease back then led to the state of affairs over Lyme disease now (which is something people ask whenever they read the 1993 Senate testimony I recently posted): It's not that easily answered.

A combination of physicians who weren't sure how to treat Lyme disease not long after the disease had been discovered, physicians who came down ill themselves and self-treated for longer if their symptoms relapsed, the media's role in educating the public about Lyme disease leading to fearful patients, politicians in Connecticut who were slow to respond to patient demands for improvements in diagnosing and reporting the disease - all of these contributed to the atmosphere around Lyme disease.

The slow response of politicians and doctors to increasing numbers of patients suffering from Lyme disease led to the creation and growth of Lyme disease patient support groups. Even back then, by 1992 there were over 100 patient support groups for Lyme disease.

I quote an excerpt from Polly Murray's The Widening Circle, pages 266-267:

"Some doctors I have encountered think that Lyme disease is easily treated by a single course of antibiotics in its early stages; if patients fail to overcome the infection after one treatment, their future complaints are considered to be not associated with Lyme disease. These doctors believe that once a patient is treated, he or she no longer has Lyme disease.

In 1987-1988, tremendous media attention was given to the disease; I think this was generally beneficial. Correct information must prevail over ignorance. However, when the public saw the grave outcomes suffered by the patients who appeared on TV, many were scared that if they were bitten by a tick, they would have devastating illness. While a number of people do not respond to treatment or have not had treatment and have had severe complications, a proportionately far greater number are treated and seem to do well. (It is true, however, that some may relapse or enter another stage of the disease, sometimes many years later.)

Some doctors during this period of media attention were inundated by people worried that they might have the disease; they called this anxiety Lyme hysteria and Lyme paranoia. Some patients, as I have mentioned, were told they were "antibiotic junkies". There seemed to be many extremes in attitude, some physicians being unwilling to diagnose Lyme disease even with a classic presentation, and others willing to treat anyone with any vague symptom for Lyme disease. The unreliability of Lyme tests, as we shall see, did not help matters."

So, during this time, according to Polly Murray's account, people who were incredibly sick with Lyme disease were in the media spotlight and it led to fear in many people. The fear was not entirely unfounded - people do suffer greatly from Lyme disease. But if caught early, a significant number of patients go on to do well.

Why this is remains a complicated answer, and something that this blog investigates over time - it won't be limited to a paragraph or two of text.

At any rate, some people were afraid they had Lyme disease, and rather than be compassionate, some of these doctors were making negative statements about such patients. How many, I don't know - Ms. Murray doesn't elaborate on numbers.

It definitely does seem as if there was a greater mix of opinions in the medical profession about how to diagnose and treat Lyme disease. It was less monolithic, and the push from a handful of select professional medical organizations to set the standard of care for all primary care physicians wasn't in place yet.

Ms. Murray continued:

"A number of physicians continued to say that media hype was distorting the true profile of Lyme disease and was scaring people unnecessarily. True, the adverse outcomes were proportionately rare; however, to the rare patient with a devastating outcome, statistics are irrelevant. The fact remains that the more proper information citizens and physicians are armed with, the more likely they will be to protect themselves from Lyme infection and to detect and treat Lyme disease early so that devastating outcomes will be less likely.

As the number of cases continued to rise, some physicians in endemic areas began to see great numbers of patients with Lyme disease. With their growing experience, many of these physicians became convinced that suggested treatment regimens were insufficient to combat the disease in some cases and were calling for longer and more aggressive treatment. They began to encounter patients who clinically seemed to have Lyme yet tested negative, while a number of patients continued to have persistent symptoms and remained chronically ill for years despite treatment. This area of chronic complications is the most controversial and the most disheartening part of the Lyme disease story."

So as the number of cases went up, so did the number of patients who had seronegative Lyme disease and those who seemed to have a condition resistant to treatment.

To get a complete picture of how a disease is going to interact with a population, one can extrapolate what percentage will be infected from a smaller sample size - but it's harder when the presentation is not consistent across the board. Harder when a disease can be seronegative. Harder when it's suspected a disease can be asymptomatic and latent, only to present in its third, most serious stage later.

Part of what fueled the controversy back then was research in 1989 by V. Preac-Mursic and also doctors who were infected with Lyme disease who self-treated themselves longer than what Yale's suggested guidelines were at the time (that 2-4 week treatment length everyone in Lyme world is familiar with).

Ms. Murray wrote,

"Evidence has been found for the persistence of the spirochete in various parts of the body, despite antibiotic treatment and negative tests. A 1990 paper by V. Preac-Mursic and colleagues reported studies of patients who had originally been seropostiive, had been treated with antibiotics, and then had become seronegative. However, spirochetes could be cultured from skin specimens and spinal fluid from these patients, showing a persistence of the infection. In her summary, Dr. Preac-Mursic said, 'We conclude that early stage of the disease as well as chronic Lyme disease with persistence of B. burgdorferi after antibiotic therapy cannot be excluded when the serum is negative for antibodies against B. burgdorferi from CSF [cerebrospinal fluid] and skin biopsy in our patients after antibiotic therapy with normal CSF-values and negative serological tests for B. burgdorferi raises important considerations in the treatment of Lyme borreliosis.' Indeed, this study raises important questions as to both the adequacy of antibiotics in treating the disease and the reliability of the tests in detecting the disease."

The above mentioned study, for the curious, is this one:
PREAC-MURSIC V, WEBER K, PFISTER HW et al.: Survival of Borrelia burgdorferi in antibiotically treated patients with Lyme borreliosis. Infection (1989) 17:355-359.

To see the abstract for this paper (about halfway down the page) and papers with similar focus, I recommend this collection of links found at Lymenet Germany:
http://www.lymenet.de/literatur/niches.htm

There's a lot of science information on the Lymenet Germany web site - much of it in English - so I recommend checking it out some time.

Anyway, knowing this and other research about persistence only fueled the controversy at this time.

By the time 1992 rolled around, and the Fifth International Conference on Lyme Disease was held, the controversy was present at the conference itself.

As Murray reported:

"A number of papers submitted by physicians in endemic areas had been rejected by the conference's program committee. After protests from support groups and patients concerned that important new information on the disease was being excluded, the committee reversed its decision."

Does this sound familiar at all to any of you reading right now? Like maybe how things went down with the October 2010 Institute of Medicine workshop on tickborne illness?

She continued:

"It was my feelings that the patients who attended the conference primarily wanted better research and information on treatment evaluation and the development of more effective therapies and techniques of prevention. They were obviously interested in a cure. Many had found that the prescribed four-week treatment with antibiotics was not sufficient, and that they relapsed if not treated for long enough period of time. They questioned commonly accepted paradigms of the illness and believed that important questions were not being investigated."

And later on,

"After the conference a number of patient representatives wrote to its heads, citing six papers presented at the meeting which endorsed the theory that the spirochete persisted even after treatment and that a patient could be seronegative and yet have Lyme disease. They 'asked for more research on pathogenesis, long-term antibiotics and innovative drug delivery systems' and 'offered their services as participants in an NIH-sponsored effort to find a cure for chronic Lyme disease.'

Those patients sound just like me! That's what I'm asking for, too - more research on pathogenesis, long-term antibiotics, and innovative drug delivery systems.

Whoa.

So, as you can see, the issue of Lyme disease and its treatment being controversial has been with us for many years. The problems and concerns which patients faced twenty years ago are, sadly, the same problems and concerns they face today.

Where do we go next? What can patients do to change this state of affairs without reinventing the wheel?

A lot of awareness has been raised about Lyme disease in recent years - the publishing of Cure Unknown, the making and distribution of the film, Under Our Skin; the increasing online presence of large scale Lyme disease patient social networking sites, and numerous campaigns, fundraising drives, and events have been held to get more attention for Lyme disease.

How much this helps towards getting the research that is so badly needed remains to be seen. The funding and development of the Columbia University Lyme and Tickborne Diseases Research Center is definitely one of the bright spots in the Lyme patient community, but more such spots are needed and were needed years ago.

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

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0 Books: Mini Review on "Bull's Eye"

I've been reading bits and pieces from Bull's Eye: Unraveling The Medical Mystery Of Lyme Disease.

Someone told me it may be a frustrating read, so I was prepared for that - but it hasn't been that difficult to get through, actually, once you get past the fact that the author can write about Alan Steere more matter-of-factly than anyone I know.

That's one of the main criticisms I've heard other Lyme patients who have read it have about the book - but after reading a huge chunk of it, I'd have to say that it is more balanced about discussing the controversy than I thought it would be and that much of it focuses on things other than Steere: the history of Lyme disease in Connecticut, history of Borrelia in Europe, general research on Borrelia burgdorferi, epidemiology, difficulties in diagnosing Lyme disease, problems with serological testing, the Dearborn criteria, coinfections, shortcomings in the IDSA view of Lyme disease, the Ed Masters story, experimental treatments, genomes, vaccines, and legal issues.

And it's an easy read, for what it's worth - for the most part, people who aren't knowledgeable about Lyme disease or its controversy can jump right in and begin getting an idea of the big picture pretty quickly.

One of the things that makes it readable is the author, Jonathan Edlow, MD,  can outline procedures using metaphors and analogies in a basic way so that those reading this kind of material for the first time can 'get it', and I've found  the Ambiguity in the Lab chapter to be one of more interesting reads because of how it was written.

In the book, the author describes for the reader how Western blot tests are done for Lyme disease:

"Like the ELISA, the Western blot tests for antibodies, but it allows the laboratory to find precisely which antigens a patient's blood contains antibodies to - not just whether there are any kinds of anti-borrelial antibodies. 

First, the B. burgdorferi is put into a detergent to break up all its proteins. Each of these proteins has different sizes and each has a slight electrical charge. These proteins (antigens) are placed on a gel to which an electrical current is applied. The proteins then migrate across the gel, which is full of nooks and crannies, rather like an English muffin. Because the proteins have different sizes, they move through the gel at different rates, the larger ones moving more slowly than the smaller ones. After a certain amount of time migrating with the electrical current across the gel, the proteins from the B. burgdorferi have traveled different distances. 

Imagine a massive jungle gym with evenly spaced bars going every which way. A group of people who are four to seven feet tall are instructed to start at one end of the jungle gym and travel through the latticework as far as they can in five minutes toward the other side. In this group of people, ten are exactly four feet tall; ten are exactly four feet, six inches tall; ten are five feet tall, and so on - such that there are seven groups of ten people who are all the same height. 

Since the spaces between the bars of the jungle gym are uniform, the smaller people will be able to travel faster across this jungle than the larger, heavier people. When the five minutes are up, the larger people will be closer to the start and the smaller closer to the finish. Assuming that the participants are equal in their abilities, at the end of the five minutes, the seven groups will settle out in seven distinct regions of the jungle gym. If one were to take a picture of the apparatus at the end of the five minutes, it would show seven bands - clusters of people of the same height - interspersed with bare areas of the jungle gym devoid of anyone.

This is what happens to the proteins from borrelia in the Western blot. They migrate across the gel, through the network of obstructions, at different rates on the basis of their molecular weights. After a specified period of time, the proteins are clustered on the gel at specific regions. They are then transferred (blotted) onto a special kind of white membrane. Specific antibodies (attached to a dye and so that they can be seen) will bind to specific borrelial proteins, and colored bands will appear on the white membrane."

Edlow explains other processes and definitions by using metaphor and analogy throughout the book, making it easier to visualize them. He uses the idea of a machine to sort different kinds of fruit to explain what sensitivity and specificity mean in terms of blood tests detecting infection, and uses different models and makes of cars to help describe cross-reactivity in testing. The way he puts things makes it easier for people with no prior knowledge of Lyme disease and testing to understand certain concepts that they can later build on.

There are other reasons I like this book, and a few reasons I don't, but I wanted to take time aside to share this one aspect.

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1 Reading list

So, remember those books that someone ordered for me a few weeks ago? Well, they're here, and I now own them.

It also turns out that today was the day books I wanted to borrow showed up... I may be spending more time reading than posting at this rate. I really didn't expect all these titles to show up the same time:


Bull's Eye: Unraveling the Medical Mystery of Lyme Disease by Jonathan Edlow, MD

The Widening Circle by Polly Murray

The Neurological Manifestations of Pediatric Infectious Diseases and Immunodeficiency Syndromes by Leslie L. Barton and Neil R. Friedman, MBChB

Medical Microbiology by Patrick R. Murray, PhD, Ken S. Rosenthal, PhD, and Michael A. Pfaller, MD

Just what I need - a little light reading.

I don't know that I'm going to be able to do a review of any one of the ones I'm borrowing now - I'm mainly using them to do research.

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0 Books: Tickborne Diseases and Disease Vectors

I've just been informed I'm getting two books in the mail in about a week or so, and when they get here, I'll be sure to let everyone know and offer my own review of them once I've read a significant portion of them...

Tick-Borne Diseases of Humans edited by Jesse L. Goodman, David T. Dennis, and Daniel E. Sonenshine.

Amazon product description: A ready resource, this book covers key information that will be highly useful to students and professionals in the fields of human and veterinary medicine, public health, medical entomology, acarology, and ecology. Written by experts with specialized field knowledge, "Tick-Borne Diseases of Humans" presents state-of-the-art information on disease epidemiology, transmission, and ecology.

The book is divided into three sections, each of which can be used independently or in concert with the remaining two sections.

 Section I integrates divergent information relevant to the full spectrum of tick-borne diseases, incorporating tick biology and identification, distribution of the diseases ticks transmit, and various strategies for tick control. In addition, this section comprehensively reviews the clinical approach to a patient with a possible tick-borne affliction.

Section II is devoted to in-depth profiles of specific diseases, including information on disease history, biology, epidemiology, ecology, transmission, clinical manifestations, diagnosis, treatment and prevention. And, Section III examines the geographical distribution of tick-borne diseases and their vectors.

This book examines the striking increase in incidence and our subsequent awareness of a broad array of tick-borne diseases.

It addresses both vector and disease perspectives, including state-of-the-art information on disease epidemiology, transmission, and ecology; clinical and laboratory findings; diagnosis; and treatment and prevention.

It includes a useful full-color insert, with maps of vector and disease distribution, an atlas of clinical and pathologic images, and illustrations of diagnostically important skin lesions and blood smears; introduces public health practitioners, research scientists, and students to the field and also provides references for information beyond traditional areas of expertise; and also presents accessible information to an informed public on disease transmission, clinical laboratory diagnosis and treatment, and history of infections.      

The Biology of Disease Vectors by Barry J. Beatty and William C. Marquardt. (This is the first edition, and I'll be getting the second edition shortly thereafter to make a comparison.)

This comment is stated on the second edition on Elsevier: "Biology of Disease Vectors presents a comprehensive and advanced discussion of disease vectors and what the future may hold for their control. This edition examines the control of disease vectors through topics such as general biological requirements of vectors, epidemiology, physiology and molecular biology, genetics, principles of control and insecticide resistance. Methods of maintaining vectors in the laboratory are also described in detail. No other single volume includes both basic information on vectors, as well as chapters on cutting-edge topics, authored by the leading experts in the field. The first edition of Biology of Disease Vectors was a landmark text, and this edition promises to have even more impact as a reference for current thought and techniques in vector biology."

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