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05/05/2011 09:10 PM

EVA SAPI article-BIOFILMS/ANTBIOTICS, may program!

Bettyg
 
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Eva Sapi Article Biofilms and antibiotics

Eva Sapi will be giving a presentation of this material in May

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

Original Research (518) Article views

Authors: Sapi E, Kaur N, Anyanwu S, Luecke DF, Datar A, Patel S, Rossi M, Stricker RB

Published Date May 2011 , Volume 2011:4 Pages 97 - 113 DOI 10.2147/IDR.S19201

Eva Sapi1, Navroop Kaur1, Samuel Anyanwu1, David F Luecke1, Akshita Datar1, Seema Patel1, Michael Rossi1, Raphael B Stricker2

1Lyme Disease Research Group, Department of Biology and Environmental Sciences, University of New Haven, New Haven, CT, USA;

2International Lyme and Associated Diseases Society, Bethesda, MD, USA

Background: Lyme disease is a tick-borne illness caused by the spirochete Borrelia burgdorferi.

Although antibiotic therapy is usually effective early in the disease, relapse may occur when administration of antibiotics is discontinued.

Studies have suggested that resistance and recurrence of Lyme disease might be due to formation of different morphological forms of B. burgdorferi, namely round bodies (cysts) and biofilm-like colonies.

Better understanding of the effect of antibiotics on all morphological forms of B. burgdorferi is therefore crucial to provide effective therapy for Lyme disease.

Methods:

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

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.

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.

Results:

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%.

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.

Conclusion:

Antibiotics have varying effects on the different morphological forms of B. burgdorferi.

Persistence of viable organisms in round body forms and biofilm-like colonies may explain treatment failure and persistent symptoms following antibiotic therapy of Lyme disease.

i received this from email just now.

bettyg, iowa leader

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07/16/2011 04:30 PM
Bettyg
 
Posts: 32241
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Dr. Sapi and Teams Honorable Research

from PINELADY

7-16-11

http://www.ncbi.nlm.nih.gov/pubmed/21753890

Infect Drug Resist. 2011;4:97-113. Epub 2011 May 3.

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

Sapi E,

Kaur N,

Anyanwu S,

Luecke DF,

Datar A,

Patel S,

Rossi M,

Stricker RB.

Source

Lyme Disease Research Group, Department of Biology and Environmental Sciences, University of New Haven, New Haven, CT, USA;

Abstract

BACKGROUND:

Lyme disease is a tick-borne illness caused by the spirochete Borrelia burgdorferi.

Although antibiotic therapy is usually effective early in the disease, relapse may occur when administration of antibiotics is discontinued.

Studies have suggested that resistance and recurrence of Lyme disease might be due to formation of different morphological forms of B. burgdorferi,

namely round bodies (cysts) and biofilm-like colonies.

Better understanding of the effect of antibiotics on all morphological forms of B. burgdorferi is therefore crucial to provide effective therapy for Lyme disease.

METHODS:

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

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.

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.

RESULTS:

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%.

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.

CONCLUSION:

Antibiotics have varying effects on the different morphological forms of B. burgdorferi.

[b]Persistence of viable organisms in round body forms and biofilm-like colonies may explain treatment failure and persistent symptoms following antibiotic therapy of Lyme disease.

PMID:

21753890

[PubMed][/b]


07/27/2011 11:40 PM
Bettyg
 
Posts: 32241
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Everything you thought you knew about cyst busters is wrong

Sunday, July 24, 2011

Eva Sapi's recent research calls into question everything we thought we knew about Lyme "cysts." In fact it destroys the old thinking.

We have heard about cell wall antibiotics, intracellular antibiotics and cyst-busters. Think again.

She investigated the effect of various antibiotics on Lyme spirochetes and round body forms - also know as cystic forms.

Doxycycline worked according to plan.

Doxy inhibits protein synthesis - it kills bacteria, including Lyme, by action within the cytoplasm, inhibiting the manufacture of proteins required for the bacteria's survival.

Doxy and others are commonly referred to as intracellular antibiotics.

Spirochete loads decreased by about 90% while cyst levels increased by 200% - just as expected.

Then amoxicillin data was presented. Amoxicillin inhibits the formation of bacterial cell walls.

Amox and similar drugs should then only be effective in killing spirochetes with an intact cell wall. This is where the results start deviating from the plotted course.

Amoxicillin killed 90% of spirochete forms - OK, but -- it also killed 68% of the cystic forms!

Amoxicillin and other cell wall drugs are not cyst busters - only specific anti-parasite drugs kill cysts - or so we thought.

Well lets think again for a second: what are cysts?

Are they balled up forms of spirochetes with a different kind of membrane - or blebs (also described) expressed through the spirochete membrane?

Maybe the former retain much of the cell wall from the original spirochete - maybe that is why amoxicillin works here.

This would seem to clear up a nagging question raised by others. Are cysts and L-forms really the same thing?

These results show that cysts cannot represent a version of L-forms or spheroplasts which result when gram negatives shed their cell walls.

If this were the case, a cell wall drug would be ineffective.

Cysts and L-forms are distinct and different forms. (There may be a hole in this reasoning. I will explain later).

OK So we have learned something new: cell wall antibiotics can also kill some cyst forms which are not L-forms.

Let's look at some more data.

Tigecyline is a not a cyst drug either. Wrong. Tigecycline kills 90% of spirochetes, good so far, but it also kills 90% of cysts!

Tigecycline is an intracellular antibiotic similar to doxycycline! Another fly in the ointment.

OK. Cysts with their lower metabolic rate, still need ribosomal proteins to survive, just not at the levels of intact spirochetes.

Tigecyline is a more powerful drug, higher levels are delivered into the cytoplasm of the cysts. This makes sense.

Cyst forms are still essentially a pleomorphic version of Lyme bacteria with somewhat different features. In this scenario, cysts could be L-forms.

But we have already shown that this is not true because amoxicillin can kill them. Right?

Amoxil is a cell wall drug. I thought so.

Kersten, (antimicrobial agents and chemotherapy, May 1995, p. 1127-1133) states that Beta-lactam antibiotics, which include amox, penicillin and Rocephin, have been shown to cause a specific loss of total intracellular RNA in the absence of cell wall hydrolysis.

In other words, amoxil could possibly work in part as an intracellular agent. If this is right, cyst forms of Lyme could still be L-forms.

So perhaps we have not shown that L-forms and cyst forms are different after all.

The question remains unanswered.

Let's get to the Cyst-busters.

It takes antiparasitic drugs, so we thought, to kill the cysts. Cyst-busters, anti-parasite drugs, kill parasites (and Lyme cysts) not bacteria.

The so called cyst-busters were heretofore used in combination or cycled with other antibiotics.

Previous thinking was that typical antibiotics would kill spirochetes and/or L-forms and that cyst busters would disrupt only the cystic forms.

Cyst-busters do not kill intact spirochetes - so we are told. Very wrong this time.

I cannot cover the whole Sapi study.

The most exciting finding is that Tindamax (tinidazole) - our premier Cyst-buster, is the most effective drug overall.

This "cyst-buster" kills 90% of cysts and spirochetes: by far the best drug.

We don't know it's effect on L-forms, but we can guess. Tindamax probably works by an intracellular mechanism.

If this is true, it should be equally effective against L-forms.

It gets even better. Tindamax is the only drug which does a great job on biofilm colonies as well!(not to be discussed now). More on biofilms later.

Tindamax passes the blood brain barrier and penetrates well into most tissues. It has been effective in my patients with neurocognitive deficits - neuroborreliosis.

Recently I tried it on another sort of patient. This patient has had intractable Lyme arthritis of his knees.

This young athlete had been extensively treated with IV Rocephin followed by a year of typical oral antibiotics. Knee effusions have persisted - until I prescribed Tindamax.

Now, after two months, the fluid in his knees has evaporated. His knees are dry and painless for the first time in over one year.

This raises the question: should Tindamax be used as mono-therapy?

Well, I cannot endorse blanket use at this time.

Tindamax has a black box warning.

It has been associated with cancer in some laboratory animals. Perhaps there are more compelling reasons to use Tindamax, but this will have to wait for another post.

My nagging question:

Why does penicillin kill Lyme? It shouldn't.

Lyme is a gram negative bacteria. While certain Beta-lactam antibiotics can kill gram negative bacteria, penicillin cannot.

Penicillin is only active against gram positive bacteria.

Maybe this other mechanism alluded to above, the alternative intracellular RNA mechanism is significant and explains why penicillin kills Lyme spirochetes. Maybe not.

We need to continually reevaluate things which we have assumed to be true, because many of them are not.

Posted by Lyme report: Montgomery County, MD at 6:31 PM

4 comments:

la quinta kid said...

Awesome post. Keep up the good work. Out of all the information out there on lyme, I find your blog to be the most level headed and logical.

July 24, 2011 9:25 PM

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Brittany Adams said...

I really love this post!!!

Speaking as one of your patients, I must say that when you switched me to Tindamax a few months ago, it made all the difference in the world and i'm starting to feel better than I have in years.

And I give you full credit for my progress with treating me for chronic lyme disease. Keep up the good work and awesome posts! Smile

July 24, 2011 9:42 PM

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T9im said...

Hi Doc:

I went to Dr. Sapi's conference and she also presented combinations and Tindamax with Doxy worked the best.

It was great to see the presentations and while the bacteria load only went down to 10% the duration was not for a long period (I remember 72 hours).

Our daughter was on tindamax for 7 months and it helped tremendously with the neuro symptoms (she is 11 and has Lyme and Babesia) but we stopped due to some adverse reactions

(alergic - itching which was triggered when we increased the dossage and anorexia - fortunately after appx. 6 weeks her appetite picked up but we have a ways to go to increase body weight).

One of Dr. Sapi's research students indicated it can take up to 6 weeks for a cyst to break and release spirochetes.

July 25, 2011 4:53 AM

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ShowMeMissouri said...

Thank you for the informative post. Son, 12, just started Tindamax and Bactrim. He is sleeping a lot at the moment, but I hope it is a healing sleep! Thank you again for all of your posts!

July 25, 2011 8:03 AM

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http://lymemd.blogspot.com/2011/07/everything-you-thought- you-knew-about.html

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06/05/2012 04:59 AM
Bettyg
 
Posts: 32241
VIP Member
I'm an Advocate

Eva Sapi Biofilms = 5-22-12

Eva Sapi Lyme Disease Biofilms 5-22-2012 Western CT State University

VIDEO 34 MINUTES ... you can NOT see what she is discussing.

the microphones cover up most of her face.

GREAT ONE TO WORK ON HERE AND LISTEN IN THE BACKGROUND since you can't see any charts she discusses!

http://www.youtube.com/watch?v=l5acJNoP3gQ&feature=youtu.be

bettyg, iowa activist

Post edited by: Bettyg, at: 06/05/2012 05:06 AM

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