Tuesday, October 21, 2008

Lyme Lab 101

Unfortunately, practicing physicians are poorly trained regarding laboratory medicine. I have to admit that I am not well versed in the intricacies of IFA, PCR, gene sequencing and other cutting edge lab tests. But I did learn a little something more years ago than I want to admit. Once upon a time I worked in the trenches of the old D.C. General Hospital/Georgetown division. Patients were regularly admitted producing copious sputum. At 3am the rag tattered interns performed gram stains for bacteria and AFB stains for TB. We even spun down blood samples carefully pipetting off the white blood cells, the buffy coat, and gram stained this, looking for bacteria in the blood. If we saw gram positive bacteria in chains we suspected Streptococcal infection (Strep pneumo). If we saw weakly staining gram negative rods we suspected H. flu. We used this preliminary data to guide or choice of antibiotics. We still didn't know for sure which germ the patient had until the culture came back. Fry labs makes a diagnosis based on a blood smear. They don't even have a gram stain. Sorry. You can't distinguish Mycoplasma from Bartonella from Ehrlichia with these stains. You can't get a culture. But now there is a high tech answer. The DNA from the spotted bacteria can be extracted. All bacteria have a segment of DNA called 16S. This segment can be sequenced with technology that would have been unimaginable to the interns at D.C General working at those ungodly hours. The sequencing can be compared to a compendium of the sequencing of all known bacteria. It is like a fingerprint. The bacteria can be definitely identified. No guessing. (Clongen)

Lyme tests: All Western Blots are not equal. State of the Art technology can give results which are extremely accurate.
PCR: I got my first + result from synovial fluid: Supersensitive PCR assay.
I might have to take back things I said about Lyme and PCR in the past.
PCR: New test for Bartonella: 20 strains.
PCR: New test for Babesia, expected soon: 15 strains!
PCR: Mycoplasma: Many positives. First give anti-Mycoplasma antibiotics to chase bacteria from intracellular hiding space.
Ehrilchia: PCRs good: Serology OK and much cheaper.

I am putting together new panels with Clongen which should be very helpful for LLMDS. Clongen has a full service, high tech lab with a huge test menu. I am afraid Tick borne infection profiles will swamp this small lab soon.

9 comments:

dogdoc said...

We weren't trained that badly- they just keep inventing new stuff on us! Not a bad thing. Two ?'s-
1. What are we chasing Mycoplasma out with
2. Have you seen Garth Nicolson's recc's on lab testing for GWS? There are some interesting ideas in this group of lit.

Lyme report: Montgomery County, MD said...

Nicholson is a brave man. I spoke with him in person. He was another bright spot at the ILADS meeting. I will not discus his ideas on line.
Mycomplasma can be killed with tetracycalines, macrolides and quinolones. Take your pick.

Lyme report: Montgomery County, MD said...

PS. Your friend Brian was at the meeting.

dogdoc said...

Doc- I know what kills Mycoplasma (smile). I meant what has been shown to increase PCR yield. Nicholson talks about antibiotic false negatives. Glad to hear Nicholson is a credible source- his work holds some interesting and applicable information I believe. Did he discuss coinfection rates at his presentation? I gotta run see patients. The rest later.

dogdoc said...

Whew- what a morning. Your know its bad when the computer is your quiet time over a quick lunch or after all are tucked in bed. (Actually, it is a nice change of pace). Back to the conference- I hope Bryan fared better there than here. I should have been easier on him. I disagree strongly with presenting to the public that LD is "best" cured by an alternative self help or any other approach. However, I also think that there are folks that are driven into this arena by some of our approaches to LD complex. Sometimes I think we are hitting the tip of the iceberg with a bigger and bigger sledge hammer! Speaking of sledge hammers, did any additional evidence based info come out on antibiotic resistance over the powerpoint lecture set posted at ILADS? Anybody get anything on BLO's? (or I guess in my mind LRO's and RRO's- ie levaquin and rifampin responsive organisms: still have trouble correlating signs on own and in LD ... hmm, resistant mycoplasma and atypical mycobacterium respectively?). I can't help but notice a big picture item- LD complex patients present as an intracellular organism field day. Almost as though we have an "immune biofilm- like" situation- in the body climate of impaired cell mediated immunity, we get multiple players each capable of impairing immunity in a slightly different fashion coming together almost as commensuals helping maintain the favorable immune state. Some probably just tag along for the ride. We have ubiquitous organisms that most are exposed to and harbor longterm kept in check by cell mediated immunity (CMI)until the trigger organism. We have opportunists that the host encounters after CMI is down or as coinfection. Simple LD is one thing- but once CMI breaks down and we get into LDC, all bets are off. I wonder what players we are going to end up with in the end? LD, Babs, bart, erlh/anaplas, mycoplasma (M. fermentans as well as reg ones), chlamydias, neurotopic/persistant viruses HHV, EBV, CMV ect, nontubericle forming mycobacteria, and the list goes on of intracellular and persistant organisms that depend on CMI to keep in check. The key to success with most of our chronic persistant bacterial diseases has been the right combos of drugs for the right length of time (right to overcome resistance & slow replication, and need for return of normal immunity to get the last organisms in check). It will be very interesting to see where we are in 20 years with this one. The more I understand, the less I know. Fascinating puzzle.

Lyme report: Montgomery County, MD said...

What evidence? That's the problem.
The Lyme movement started with alternative practitioners and herbalist. That is a major reason for the lack of credibility amongst skeptics. I actually think that the alternative practitioners need to be put in a different basket. They should not be featured at ILADS meetings. We need SCIENCE- or a close facimile.
I am hoping the Clongen technology will help resolve the issues of blood borne bacteria. They keep calling me and telling me that blood samples are loaded. I need to await DNA fingerprinting and hopefully we will have an inkling as to what we are dealing with. I wont be surprised if the DNA fingerprint of these bacteria are not in the catalogue. There are some things I can only tell you in person.

Incidentally, you know as much about the topic as anyone. Scary? Isn't it?

dogdoc said...

Thats a laugh- I read this comment after I wrote the other one. Sounds like grasshopper and master are on the same page. I guess even grasshoppers have to evolve sometimes. Evolution is a humbling process. Oh- many will be in a catalog. Just in bits and peices like a kids collage- pictures recombined and pasted into pretty little pictures. But in the end, the chicken and the egg philosophising will have to lead way to how are we going to clean up the scrambled mess left to burn on the bottom of the frying pan. Can we get it in the sink to soak while we figure out the best tools for getting the crud scraped out of there without peeling of the nonstick coating? By the way, I sincerely hope my minimal level of knowledge is by no means representative of the whole. I am but a country animal doctor and very new to it all at that. And yes- it is scary. And we do desparately need the science. Perhaps it is better to create a new basket than to try to paint the old one? The world has room for many baskets to coexist in peace. I think we need to be careful that the new basket focuses on humanitarian pursuits and takes the high road on the rest. Leave the where and who to someone else and focus on the what and how to deal with it. It may be very difficult to walk that line.

dogdoc said...

PS- for the record, I don't know a darn thing. I don't even want to know. We just are treating a group of patients without typical signs or response to therapy of the wild type diseases. We are treating patients with "onions of disease"- we peel off layers (treat) and we have a new one underneath. We are treating phantom coinfections- ones without tests or typical signs except in this context. We have unusual immunity breakdown patterns. We have a host of previously unheard of syndromes with a chronic related set of endpoints in the body. We are treating Borrelia with drugs for TB and maleria in order to get them to respond for goodness sake. Interesting genetic combinations in bug genomes are popping up independently here and there across the world. We have atypical clusters of disease for their routes of transmission. Either the bacteriophages have been busy or... somebodies been where they should not have gone. Humanity does not have a stellar track record in our time on this earth. We have had the knowledge and technology for an extended period of time. In the end, it matters not who or what. It would be nice if someone would cough up a list of what peices we might be playing with so we could move a bit faster. Some are easier to see in the patterns than others. We need to be able to test for the players, figure out immune support, and get doing the best we can. We really need culture/ sensitivity support for some of these beasties. The thing is- we have the infrastructure to be able to research and develope interventions at this time. And we are being too stupid to set it in motion. Just gotta call them like I see them.

sloborabas said...

Just want to say that this may be the best Lyme-blog I have ever read.

One question to the DOC: one of the primary problems I developed whilst walking around (for years) undiagnosed with Lyme was hyperthyroidism / Graves disease.

I hear it is somewhat unusual to go hyperthyroid, but I am told thyroid problems in general are extremely common with Lyme. Hence I was extremely surprised that thyroid tests (hormones: TSH, T3, T4 and antibodies: Anti-TPO, TgAb) don't appear to be mentioned in your list of diagnostic criteria?