LymeNet Home Page LymeNet Flash Discussion LymeNet Support Group Database LymeNet Literature Library LymeNet Legal Resources LymeNet Medical & Scientific Abstract Database LymeNet Newsletter Home Page LymeNet Recommended Books LymeNet Tick Pictures Search The LymeNet Site LymeNet Links LymeNet Frequently Asked Questions About The Lyme Disease Network LymeNet Newsletter Volume 6 Issue 11 LymeNet Home Page LymeNet Flash Discussion LymeNet Support Group Database LymeNet Literature Library LymeNet Legal Resources LymeNet Medical & Scientific Abstract Database LymeNet Newsletter Home Page LymeNet Recommended Books LymeNet Pictures Search The LymeNet Site LymeNet Links LymeNet Frequently Asked Questions About The Lyme Disease Network LymeNet Home LymeNet Newsletter Library

Volume: 6
Issue: 11
Date: 20-Nov-98


Table of Contents:

I.    LYMENET: ACP Recommendations Opposed by Laboratory Scientists
II.   LYMENET: New German Language LD Web Sites Now Available
III.  EUR J CLIN MICROBIOL INFECT DIS: Analysis of the intrathecal
      immune response in neuroborreliosis to a sonicate antigen and
      three recombinant antigens of Borrelia burgdorferi sensu stricto
IV.   ZENTRALBL BAKTERIOL:  Detection of Borrelia burgdorferi in
      urine specimens from dogs by a nested polymerase chain reaction
V.    ZENTRALBL BAKTERIOL: Expression of outer surface proteins A and
      C of Borrelia afzeliiin Ixodes ricinus ticks and in the skin of
      mice
VI.   ABOUT THE LYMENET NEWSLETTER


Newsletter:

***********************************************************************
*                  The National Lyme Disease Network                  *
*                       http://www.LymeNet.org/                       *
*                         LymeNet Newsletter                          *
***********************************************************************


                   Volume 6 / Number 11 / 20-NOV-98
                                INDEX


I.    LYMENET: ACP Recommendations Opposed by Laboratory Scientists
II.   LYMENET: New German Language LD Web Sites Now Available
III.  EUR J CLIN MICROBIOL INFECT DIS: Analysis of the intrathecal
     immune response in neuroborreliosis to a sonicate antigen and
     three recombinant antigens of Borrelia burgdorferi sensu stricto
IV.   ZENTRALBL BAKTERIOL:  Detection of Borrelia burgdorferi in
     urine specimens from dogs by a nested polymerase chain reaction
V.    ZENTRALBL BAKTERIOL: Expression of outer surface proteins A and
     C of Borrelia afzeliiin Ixodes ricinus ticks and in the skin of
     mice
VI.   ABOUT THE LYMENET NEWSLETTER



=====*=====


I.    LYMENET: ACP Recommendations Opposed by Laboratory Scientists
-------------------------------------------------------------------


Editor's note:  The following letter was submitted to the Annals of
Internal Medicine in response to their position paper on the
laboratory diagnosis of Lyme disease.  The full text of the original
article can be found at:


     http://38.232.17.254/journals/annals/15dec97/pplyme1.htm

--

Letter to the Editor
American College of Physicians
Independence Mall West
6th St. At Race
Philadelphia, PA


To Whom It May Concern,

We have several concerns with the report entitled Guidelines for
Laboratory Evaluation in the Diagnosis of Lyme Disease (Annals Int
Med 127, 1106-1123. 1997)


* The  Centers for Disease Control have developed a set of clinical and
diagnostic criteria for surveillance purposes.  The authors of these
"Guidelines" state, with no substitution, that this criteria is also
applicable to the clinical diagnosis of Lyme disease.  To our
knowledge, no such evidence exists.  It would appear that the published
"Guidelines" have as a basis, a clinical criteria for Lyme disease
diagnosis which has never been tested except for clinical studies
published by the authors themselves. There is no external validation to
support the claim of equivalence between clinical diagnostic criteria
and the CDC surveillance definition.


* The authors state that "testing with ELISA is the cornerstone of
laboratory diagnosis for Lyme disease". In fact, it is not. The
commercially available ELISA assays for Lyme Disease do not meet
acceptable criteria according to the group that is responsible for
much of the United States Laboratory proficiency testing program in
Lyme Disease. The data, from a recent study by Bakken et.al. (J Clin
Microbiol 35:537-543,1997), indicated "that" the sensitivity and
specificity of the currently used tests for Lyme Disease are not
adequate to meet the two-tier test approach being recommended by the
CDC/ASTPHLD group". Bakken et. al. (1997) also stated that a screening
test must have >95% sensitivity to adequately screen for Lyme disease
and that the currently available ELISA tests do not perform at that
level.


* The authors have not followed the CDC/ASTPHLD recommendation for
2-tiered testing. That is, all indeterminate and reactive ELISAs
should be reflexed to Western blot (WB), not just indeterminate ELISAs
as the authors of the "Guidelines" suggest. Certainly the authors
realize that reactive Lyme ELISA results may be non-specific because
of a number of cross reacting antibodies (e.g. antibody to the 41 Kda
flagellin protein).


The authors have missed some important studies of Western blotting,
especially those that may be critical of the recommendations of
Dressler et. al. (J. Infect. Dis. 167, 392-400,1993) and the
CDC/ASTPHLD. The report by Engstrom et al (J Clin Microbiol  
33:419-427, 1995) found, for example, that 20% of their Lyme patients
remained seronegative throughout the study and that fewer bands on the
IgG WB could be appropriately used for interpretation. They noted that
the WB was more specific and more sensitive than the ELISA. Their
study also showed that only 19% of patients treated with antibiotics
for 20 days still had a positive ELISA antibody response after one
year, yet almost 60% of their patients continued to be WB positive at
the end of the first year. The study by Aguero- Rosenfeld et al
(J Clin Microbiol 34:1-9, 1996) reported that 89% of patients, with
culture-confirmed erythema migrans (EM), developed specific IgG
antibodies by WB, but only 22% of these patients were positive by the
interpretive criteria proposed by the CDC/ASTPHLD. They further

reported that the duration of the antibody response was related to the
duration of the EM. Tilton et. al. (1997) stated that a highly
sensitive and specific Western blot is desirable for a 2 tiered test
approach or as a primary test. Despite the CDC/ASTPHLD
recommendations, many physicians who treat patients for L.D. do not
believe that an ELISA is an appropriate screening test and consequently
use the Western blot as a primary test.


The authors state that patients not be tested for Lyme Disease unless
the pretest probability of disease is between 0.20 and 0.80. These
recommendations will:


a) rule out any laboratory detection of B. burgdorferi antibodies,
antigens, and/or DNA in non-endemic areas


b) require physicians to screen patients based on epidemiological data
which may not be available to them outside their own local area
require physicians to know the performance characteristics of a wide
variety of Lyme disease tests.


The authors in their comment on non-antibody based tests have chosen to
overlook a number of publications on the utility of direct detection
tests for Lyme disease. A comprehensive review of molecular
techniques for diagnosis of Lyme Disease has recently been published
by Schmidt (Clin Micro Rev 10, 185-201, 1997). They state "evidence
is growing that a positive PCR test can be associated with active
disease after adequate therapy, PCR results are usually negative"
Manak et. al. (J Spirochet Tick Borne Dis 4., 11-20. 1997) in a
well controlled study using the CDC criteria for selection of patients
indicated that PCR on serum, plasma, or buffy coat could be effectively
used to monitor the efficacy of therapy. Similarly, Harris et. al.
(J Spirochet Tick Borne Dis 237, 1995) have validated the Lyme
urinary antigen test (LUAT) in more than 700 LD patients and controls.
The LUAT has a specificity of >95%.  These "Guidelines" only complicate
an already complex disease diagnostic process.


Richard C. Tilton, Ph.D.
Diplomate American Board Medical Microbiology
BBI Clinical Laboratories, Inc.
75 North Mountain Rd.
New Britain, CT 06053
rtilton@bbii.com


Mary N. Sand, Ph.D.
BBI Clinical Laboratories, Inc.
75 North Mountain Rd.
New Britain, CT 06053


Nick S Harris, Ph.D.,
Diplomate American Board Medical Laboratory Immunology
IGeneX Inc. Reference Laboratory
797 San Antonio Road
Palo Alto, CA



=====*=====


II.   LYMENET: New German Language LD Web Sites Now Available
-------------------------------------------------------------
Sender: Marc Gabriel <a229@Lehigh.edu>


Two new web sites dedicated to providing German speaking individuals
with access to Lyme disease information are now available on the web.


1. http://www.LymeNet.de/
  This site mirrors much of the information on the LymeNet web site.
German translation has been provided by a German physician.  Marianne
Steenken is responsible for this site.


2. http://www.ariplex.com/lyme/lyme_top.htm
  Authored by Aribert Deckers, this site offers a wealth of
information, much of it translated from English.  A extensive list
of links is also provided.


Both sites offer a translation of the Burrascano treatment protocol.


=====*=====


III.  EUR J CLIN MICROBIOL INFECT DIS: Analysis of the intrathecal
     immune response in neuroborreliosis to a sonicate antigen and
     three recombinant antigens of Borrelia burgdorferi sensu stricto
----------------------------------------------------------------------
AUTHORS: Kaiser R, Rauer S
ORGANIZATION: Neurologische Klinik und Poliklinik der Albert-Ludwigs-
             Universitat Freiburg, Germany.
REFERENCE: Eur J Clin Microbiol Infect Dis 1998 Mar;17(3):159-66
ABSTRACT:
The intrathecal synthesis of borrelial-specific IgM- and IgG-antibodies
was studied in 67 patients with neuroborreliosis and in 14 patients
with neurosyphilis (controls). Antibody concentrations in serum and in
the cerebrospinal fluid were determined by an enzyme immunoassay (EIA)
using, as antigens, a sonicate of Borrelia burgdorferi, the recombinant
14 kDa flagellin fragment, the outer surface protein C (22 kDa), and
the high molecular mass protein p83 (83 kDa). In the sonicate EIA,
IgG- and/or IgM-antibodies to Borrelia burgdorferi in serum were

detected in all patients with neuroborreliosis and in 71% of patients
with neurosyphilis. Intrathecal synthesis of borrelial-specific IgG-
and/or IgM-antibodies was demonstrated in 82% of patients with
neuroborreliosis and in 71% of patients with neurosyphilis.
Immunoglobulin G- and/or IgM-antibodies in serum against any of the
recombinant antigens were detected in 92% of patients with
neuroborreliosis and in none of those with neurosyphilis. Intrathecal
synthesis of IgG- and/or IgM-antibodies to individual recombinant
antigens was demonstrated in 67% of patients with neuroborreliosis
and in none of those with neurosyphilis. The sensitivity of the
recombinant antigens in serum was almost equal to that of the
sonicate EIA, whereas the recombinant antigens were clearly less
sensitive in the estimation of the intrathecal specific immune
response. It was concluded that in suspected cases of neuroborreliosis,
the estimation of high specific antibodies in the recombinant EIA
will be helpful in confirming the diagnosis.



=====*=====


IV.   ZENTRALBL BAKTERIOL:  Detection of Borrelia burgdorferi in
     urine specimens from dogs by a nested polymerase chain reaction
---------------------------------------------------------------------
AUTHORS: Bauerfeind R, Kreis U, Weiss R, Wieler LH, Baljer G
ORGANIZATION: Institut fur Hygiene und Infektionskrankheiten der Tiere,
             Justus-Liebig-Universitat Giessen, Germany.
REFERENCE: Zentralbl Bakteriol 1998 May;287(4):347-61
ABSTRACT:
A nested PCR (nested flagellin PCR) carrying an internal E.
coli DNA control was established and compared with an in-vitro culture
method for the detection of Borrelia burgdorferi in urine specimens
of dogs. The predicted specific amplicon of the flagellin gene fla
was generated from all cultured strains of B. burgdorferi tested
(comprising three European genospecies). In contrast, all 13 strains
of seven other flagellated bacterial species were negative. The PCR
detection limit yielded 20 cells of B. burgdorferi per ml of
double-distilled water and approx. 250 bacteria per ml of dog urine.

Using the bacterial culture method, urine specimens collected from
216 dogs in Germany were all diagnosed negative for spirochetes by
in-vitro culture and dark-field microscopy. In contrast, DNA of B.
burgdorferi was detected in 32 specimens (14.8%) by PCR. 31 urine
specimens (14.4%) showed inhibitory activity in the PCR assay.
However, 94 (44%) were inhibitory in the culture assay. The majority
of the PCR-positive dogs exhibited major clinical symptoms which have
not been reported in the course of B. burgdorferi infection previously,
e.g. cystitis (14/32 dogs) or prostatitis (5/32 dogs). Our results
indicate that the analysis of urine specimens by the nested flagellin
PCR is a highly valuable procedure for the diagnosis of B.
burgdorferi infections in dogs.



=====*=====


V.    ZENTRALBL BAKTERIOL: Expression of outer surface proteins A and
     C of Borrelia afzeliiin Ixodes ricinus ticks and in the skin of
     mice
---------------------------------------------------------------------
AUTHORS: Leuba-Garcia S, Martinez R, Gern L
ORGANIZATION: Institut de Zoologie, University of Neuchatel,
             Switzerland.
REFERENCE: Zentralbl Bakteriol 1998 May;287(4):475-84
ABSTRACT:
Several studies have described changes in the expression of proteins,
especially of OspA and OspC, of B. burgdorferi sensu stricto during
tick feeding. In this study, the expression of OspA and OspC of B.
afzelii in unfed and feeding I. ricinus nymphs and in the subsequent
adults was followed by means of the immunofluorescence test.
Spirochaetes expressing OspA and OspC were observed in 70% and 80%,
respectively of the unfed nymphs. In feeding and in fully engorged
ticks, spirochaetes expressed OspC, while OspA disappeared 24 hours
after the beginning of the blood meal. Spirochaetes expressing OspC

in salivary glands were observed in one engorged tick. After molting,
in unfed adults spirochaetes again expressed OspA and OspC but did so
less frequently (6% and 13%, respectively). The mouse strain (AKR/N or
BALB/C) on which ticks had their infectious blood meal influenced OspC
expression in the following tick stage. In the skin of AKR/N mice, at
the tick feeding site, B. afzelii expressed OspC only, as was shown
by immunostaining.



=====*=====


VI.   ABOUT THE LYMENET NEWSLETTER
-----------------------------------------------------------------------
For the most current information on LymeNet subscriptions,
contributions, and other sources of information on Lyme disease,
please refer to the LymeNet Home Page at:
                  http://www.lymenet.org
-----------------------------------------------------------------------
To unsubscribe from the LymeNet newsletter, send a message to:
                   listserv@lehigh.edu
On the first line of the message, write:  unsub lymenet-l
-----------------------------------------------------------------------
LymeNet - The Internet Lyme Disease Information Source
-----------------------------------------------------------------------
Editor-in-Chief: Marc C. Gabriel <a229@Lehigh.EDU>
           FAX (for contributions ONLY): 908-789-0028
Contributing Editors: Carl Brenner <cbrenner@ldeo.columbia.edu>
                     John Setel O'Donnell <jod@Equator.com>
                     Frank Demarest <76116.2065@CompuServe.com>

Advisors: Carol-Jane Stolow, Director <carol_stolow@lymenet.org>
         William S. Stolow, President <bill_stolow@lymenet.org>
         The Lyme Disease Network of New Jersey
-----------------------------------------------------------------------
WHEN COMMENTS ARE PRESENTED WITH AN ATTRIBUTION, THEY DO NOT
NECESSARILY REPRESENT THE OPINIONS/ANALYSES OF THE EDITORS.
-----------------------------------------------------------------------
THIS NEWSLETTER MAY BE REPRODUCED AND/OR POSTED ON BULLETIN BOARDS
FREELY AS LONG AS IT IS NOT MODIFIED OR ABRIDGED IN ANY WAY.
-----------------------------------------------------------------------
SEND ALL BUG REPORTS TO a229@Lehigh.EDU
-----------------------------------------------------------------------


Home | Flash Discussion | Support Groups | On-Line Library
Legal Resources | Medical Abstracts | Newsletter | Books
Pictures | Site Search | Links | Help/Questions
About LymeNet


© 1994-1999 The Lyme Disease Network of New Jersey, Inc.
All Rights Reserved.
Use of the LymeNet Site is subject to the Terms of Use.