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Seronegativity in Lyme Borreliosis and Other Spirochetal Infections

16 September 2003 

“If false results are to be feared, it is the false negative result 
which holds the greatest peril for the patient.” 


Gestational Lyme borreliosis. Implications for the fetus. 

MacDonald AB. Rheum Dis Clin North Am, 15(4):657-77. 1989. 

Author Year Title Journal 

Borrelia burgdorferi 

1. Dejmkova H; 2002 Seronegative Lyme arthritis caused by Borrelia garinii. Clinical Rheumatology, 21(4):330-4 Hulinska D; Tegzova D;

[From the abstract:]

 “A case of a female patient suffering from Lyme arthritis (LA) without elevated antibody levels to Borrelia burgdorferi sensu lato is reported. Pavelka K; Seronegative Lyme arthritis was diagnosed based on the classic clinical manifestations and DNA-detected Borrelia garinii in blood and synovial fluid of the patient, Gatterova J; after all other possible causes of the disease had been ruled out. The disease was resistant to the first treatment with antibacterial agents. Six months after the therapy, Vavrik P. arthritis still persisted and DNA of Borrelia garinii was repeatedly detected in the synovial fluid and the tissue of the patient. At the same time, antigens or parts of spirochetes were detected by electron microscopy in the synovial fluid, the tissue and the blood of the patient. The patient was then repeatedly treated by antibiotics and synovectomy has been performed.” 

2. Tylewska-2002 Limiation of serologic testing for Lyme borreliosis: evaluation of ELISA and western blot in comparison Wien Klin Wochenschr, 114(13-14):601-5 Wierzbanowska S; with PCR and culture methods. 
Chmielewski T; 
[From the abstract:] 

“No correlation was found between levels of specific B. burgdorferi antibodies detected with a recombinant antigen ELISA and the number of protein fractions developed with these antibodies by immunoblot. Moreover, Lyme borreliosis patients who have live spirochetes in body fluids have low or negative levels of borrelial antibodies in their sera. This indicates that an efficient diagnosis of Lyme borreliosis has to be based on a combination of various techniques such as serology, PCR and 
culture, not solely on serology.” [Testing was performed on samples from 90 patients.] 

3. Breier F; 2001 Isolation and polymerase chain reaction typing of Borrelia afzelii from a skin lesion in a Br J Dermatol, 144(2):387-392 
Khanakah G; seronegative patient with generalized ulcerating bullous lichen sclerosus et atrophicus. 
Stanek G; Kunz G; 
Aberer E; [From the abstract:] “Spirochaetes were isolated from skin cultures obtained from enlarging LSA lesions. These spirochaetes were identified as Borrelia afzelii 
Schmidt B; by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and polymerase chain reaction (PCR) analyses. However, serology for B. burgdorferi 
Tappeiner G. sensu lato was repeatedly negative." 
4. 
Brunner M. 2001 New method for detection of Borrelia burgdorferi antigen complexed to antibody in J Immunol Methods, 249(1-2):185-190 
seronegative Lyme disease. 
[From the abstract:] "...serologic tests for early Lyme disease can be falsely negative due to lack of sensitivity of ELISAs and Western blots. Most routine antibody tests are 
designed to detect free antibodies, and in early, active disease, circulating antibodies may not be free in serum but sequestered in complexes with the antigens which 
originally triggered their production. This difficulty may be overcome by first isolating immune complexes (IC) from the serum and using this fraction for testing. Free Borrelia-
specific antibodies can then be liberated from the immune complexes which may enhance test sensitivity in patients with active disease. We developed a technique that 
captures the antibody component of IC on immunobeads, and subsequently releases the antigen component of IC. Immunoblotting with monoclonal antibody detected at least 
one antigen to be OspA, thus definitively demonstrating a Borrelia-specific antigen in circulating IC in early Lyme disease. This test is also useful in demonstrating Bb antigen 
in otherwise seronegative Lyme disease patients." 

Page1 of 17 


Author Year Title 
Journal 

5. Wang P; 2001 Contribution of HLA alleles in the regulation of antibody production in Lyme disease. Front Biosci, 6:B10-B16 
Hilton E. 
"Of eighteen seronegative LD patients, 14 were OspA PCR positive on mononuclear cells and 5 were positive on CSF. ...The presence of certain HLA alleles with 
seronegativity to disease has been reported in malaria (10), HIV (16,17), rheumatoid arthritis (RA) (18) and spondylarthropathies (SpA) (19). ...Our results provide 
evidence of a correlation between certain HLA genotypes and the ability to mount an antibody response to Bb. In this study, 9 of 22 (40.9%) seropositive LD patients 
and only 1 out of 18 (5.6%) seronegative LD patients had HLA-DR7 alleles. ... 

Our study provides evidence that HLA alleles are involved in antibody responsiveness or non-responsiveness to Bb infection. A low frequency of HLA-DR7 alleles 
and HLA-DR6 alleles and a high frequency of HLA-DR1 alleles may contribute to non-responsiveness of antibody production in LD patients. Thus, genetic 
predisposition may be a critical factor in the regulation of the host immune response and the diagnosis and prognosis of Lyme disease.” 

6. 
Grignolo MC; 2001 Reliability of a polymerase chain reaction (PCR) technique in the diagnosis of Lyme Minerva Med, 92(1):29-33 
Buffrini L; borreliosis. 
Monteforte P; 
Rovetta G. [From the abstract:] "50% of the PCR positive results, obtained with serum and cerebrospinal fluid samples corresponded to patients who were true 
positives at clinical examination but negatives at serologic tests. 62.5% of urine samples positive results belonged to tp patients who had negative serologic 
and serum PCR RESULTS. CONCLUSIONS: The obtained results suggested a good reliability of positive results obtained with the PCR technique used in 
this study and allowed the false negatives of serologic tests to be detected, more specifically when urine samples were used." 

7. 
Klempner MS; 2001 Intralaboratory reliability of serologic and urine testing for Lyme disease. American Journal of Medicine, 110(3):217-19 
Schmid CH; Hu L; 
Steere AC; 
Johnson G; “In the 21 patients with Lyme disease, the results of the initial western blot analysis were positive in 14 cases and negative in 7. ..

McCloud B; Repeat testing of the 7 seronegative samples showed fewer than 5 reactive bands in all samples.
” 
Weinstein A. 


8. 
Honegr K; 2001 [Persistence of Borrelia burgdorferi sensu lato in patients with Lyme borreliosis]. Epidemiol Mikrobiol Imunol, 50(1):10-6 
Hulinska D; 
Dostal V; [From the abstract:] “In 18 patients with Lyme borreliosis the authors proved the persistence of Borrelia burgdorferi sensu lato by detection of the causal agent by 
Gebousky P; immune electron microscopy or of its DNA by PCR in plasma or cerebrospinal fluid after an interval of 4-68 months. ...Examination of antibodies by the ELISA method 
Hankova E; was negative in 7 of 18 patients during the first examination and in 12 of 18 during the second examination. In all negative examinations the specific antibodies were 
et al. assessed by the Western blot or ELISA method after liberation from the immunocomplexes." 
9. Paul A. 2001 [Arthritis, headache, facial paralysis. Despite negative laboratory tests Borrelia can still be MMW Fortschr Med, 143(6):17 
the cause.] 
10. 
Pleyer U; Priem S; 2001 Detection of Borrelia burgdorferi DNA in urine of patients with ocular Lyme borreliosis. Br J Ophthalmol, 85(5):552-5 
Bergmann L;
Burmester G; 
Hartmann C; [From the abstract:] "RESULTS: Only four of six uveitis patients suspected for Lyme borreliosis were ELISA positive, while all six subjects showed a positive western blot. 
Krause A. B burgdorferi PCR was positive in all of these six patients. Whereas two of the 30 controls had a positive Lyme serology, B burgdorferi DNA was not detectable by PCR in any 
sample from these patients. CONCLUSIONS: PCR for the detection of B burgdorferi DNA in urine of uveitis patients is a valuable tool to support the diagnosis of ocular Lyme 
borreliosis. Moreover, these patients often show a weak humoral immune response which may more sensitively be detected by immunoblotting.” 

11. 
Eldoen G; 2001 [Lyme neuroborreliosis in More and Romsdal]. Tidsskrift for Den Norske Laegeforening, 
Vik IS; Vik E; 121(17):2008-11. 
Midgard R. 
[From the abstract:] "Fourteen of 25 (56%) patients had positive Borrelia burgdorferi-IgM and IgG titres in cerebrospinal fluid despite negative tests in serum." 

Page2 of 17 


Author Year Title 
Journal 

12. 
Brunner M; 2000 Immune complexes from serum of patients with Lyme disease contain Borrelia burgdorferi Journal of Infectious Diseases, 182(2):534-9 
Sigal LH. antigen and antigen-specific antibodies: potential use for improved testing. 
[From the abstract:] "We report sequestration of specific IgM anti-Borrelia burgdorferi (Bb) and Bb antigens within immune complexes (ICs) isolated from serum of patients 
with Lyme disease (LD). ...Immunoblot demonstrated that ICs contained antibodies against specific Bb proteins, whereas reactivity was absent or significantly lessened in 
unprocessed serum." 

13. 
Kaiser R. 2000 False-negative serology in patients with neuroborreliosis and the value of employing of different J Med Microbiol, 49(10):911-5. 
borrelial strains in serological assays. 
[Abstract:] “The risk of obtaining false-negative results in serological assays in serum and CSF specimens with only one strain of Borrelia burgdorferi sensu lato as 
antigen was investigated in 79 patients with neuroborreliosis with specimens obtained at initial presentation. Serum antibodies were assessed by immunoblotting; the 
criteria of Hauser et al. were used to evaluate the test. The intrathecal synthesis of borrelial-specific IgM and IgG antibodies was examined by enzyme immunoassay 
(EIA). Strains of B. burgdorferi sensu stricto (BbZ160), B. garinii (Bbii50) and B. afzelii (PKO) served as sources of antigen in both assays. All patients produced 
either a positive IgM or IgG test in serum with at least one strain of B. burgdorferi sensu lato. Reactivity of IgM or IgG antibodies, or both, with antigens of all three 
strains was demonstrated in 67 (85%) of 79 sera. The correlation of results of immunoblotting with different strains was significantly better for IgG (85%) than for IgM 
antibodies (54%). The variability of positive IgM reactions in 18 specimens was mainly due to the fact that the antibodies were directed to the relevantvariable outer-
surface protein C (p23). Intrathecal synthesis of IgG antibodies was demonstrated in 58 patients (81%) of 72 and of IgM antibodies in 25 of 58 patients. No patient 
had isolated intrathecal synthesis of IgM antibodies. The majority of CSF samples (56 of 58) were assessed as IgG antibody-positive, independent of the borrelial 
strain used as antigen in EIA, whereas only 10 of 25 IgM antibody-positive CSF specimens reacted with all three strains. All patients in the study had intrathecal 
antibody synthesis demonstrable at 6-week follow-up. From this study it is concluded that there is a small, but real, risk of false-negative serological findings at the 
time of initial clinical presentation in patients with typical symptoms of neuroborreliosis. In these patients a negative serological result with one strain should prompt 
the repetition of the test with other strains of B. burgdorferi sensu lato.” 

14. Kmety E. 2000 
[Dynamics of antibodies in Borrelia burgdorferi sensu lato infections.] Bratisl Lek Listy, 101(1):5-7 
[From the abstract:] "...During 1994-1998 at least two serum samples were submitted for serological testing from more than 1200 patients. An 
immunofluorescence test was performed paralelly [sic] with two pools of antigen (B. bg.s.s. + B. afzelii, and two serological different strains of B. garinii, all of local origin)
In 92-96% of patients no change of antibody level was found in repeated tests, about 20% of them being negative (< 1:512)



...Only in 9 cases a rise of the titer appeared during 3 weeks after the first negative sample, at contrary in 7 cases no rise of the titer was seen in that time. 2 patients 
were still after 1 month, 3 after 3 months and 1 even after 7 months (patient with a positive CSF culture) serologically negative.



15. 
Wilke M; 2000 Primarily chronic and cerebrovascular course of Lyme neuroborreliosis: case reports and literature review. Arch Dis Child, 83(1):67-71. 
Eiffert H; 
Christen HJ; "In this context, even the complete absence of specific antibodies has been observed; in a girl diagnosed as having focal vasculitis through CNS biopsy, the presence 
Hanefeld F. of B burgdorferi in CSF was confirmed by polymerase chain reaction. No specific antibodies were detectable. In three other children, B. burgdorferi could be cultured 
from CSF in the absence of specific antibodies in CSF or blood." 

16. 
Sheets JT; Rossi 2000 Evaluation of a commercial enzyme-linked immunosorbent assay for detection of Borrelia J Am Vet Med Assoc, 216(9):1418-22 
CA; Kearney BJ; burgdorferi exposure in dogs. 
Moore GE. 
"The commercial ELISA kit evaluated in this study appeared to lack adequate sensitivity for detecting all potential cases of borreliosis in dogs.” 

Page3 of 17 


Author Year Title 
Journal 

17. 
Wang P; 2000 Detection of Borrelia DNA in circulating monocytes as evidence of persistent Lyme disease. J Spirochetal and Tick-borne Diseases, 
Gartenhaus R; 7(1):16-19 
Sood SK; DeVoti J; 
Singer C; et al. [Abstract:] "We report the detection of Borrelia burgdorferi DNA in circulating monocytes in a 31-year-old female who presented with a flu-like syndrome followed by 
neurological abnormalities after a trip to Southampton, Long Island, New York. ELISA and Western blot were negative. Lymphocyte proliferation assay to Borrelia 
burgdorferi was positive. Borrelia burgdorferi DNA was detected in circulating monocytes using a nested polymerase chain reaction (PCR). Treatment with parenteral 
ceftriaxone resulted in clinical improvement and repeat PCR on monocytes was negative. The use of detecting DNA by PCR from circulating monocytes may be 
useful in evaluating seronegative patients with a high suspicion of Lyme disease." 

18. Brown SL; 1999 Role of serology in the diagnosis of Lyme disease. JAMA, 282(1): 62-65 
Hansen SL; (FDA Medical Bulletin) 
Langone JJ. 
"The Food and Drug Administration (FDA) is concerned about the potential for misdiagnosis of Lyme disease based on the results of commonly marketed tests for 
detecting antibodies to Borrelia burgdorferi, the organism that causes Lyme disease. It is important that clinicians understand that a positive test result does not 
necessarily indicate current infection with B. burgdorferi, and a patient with active Lyme disease may have a negative test result. 

The tests should be used only to support a clinical diagnosis of Lyme disease and should never be the primary basis for making diagnostic or treatment decisions.” 

19. Bertrand E; Szpak 1999 Central nervous system infection caused by Borrelia burgdorferi. Folia Neuropathol, 37(1):43-51 
GM; Pilkowska E; Clinico-pathological correlation of three post-mortem cases. 
Habib N; et al. 
"Case 1: ...Specific borrelia IgM and IgG value in serum and CSF were normal (<250). However, on microscopical examination the spirochete B. 
burgdorferi was demonstrated in serum and CSF. The bacteria were cultured both from blood and from CSF, in CSF they were also identified by PCR." 

20. 
Mikkila H, Karma 1999 [The laboratory diagnosis of ocular Lyme borreliosis.] Graefes Arch Clin Exp Ophthalmol, 
A, Viljanen M, 237(3):225-30 
Seppala I. 
"Seven patients, including two with negative ELISA, had a positive immunoblot. Seven of the 13 patients in whom PCR was examined during clinically active disease 
had a positive PCR result. Immunoblot analysis gave a negative result from the sera of five PCR-positive patients. CONCLUSIONS: For efficient diagnosis of ocular 
Lyme borreliosis, immunoblot analysis and PCR should be used in addition to ELISA." 

21. 
Oksi J; 1999 Borrelia burgdorferi detected by culture and PCR in clinical relapse of disseminated Lyme Annals of Medicine, 31(3):225-32 
Marjamaki M; borreliosis. 
Nikoskelainen J; 
Viljanen MK. "Three of the 13 patients had only IgM antibodies against B. burgdorferi, and one culture-positive patient was seronegative despite the disseminated stage of the disease. 
The reason for the lack of IgG antibodies, or of both IgM and IgG antibodies, was not restriction of the infection to privileged sites, as all these patients had a multiorgan 
disease. We have previously shown that patients with late LB with live spirochetes or borrelial DNA in their body fluids may have low or negative serum borrelia antibody 
levels." 

22. Hudson BJ; 1998 Culture-positive Lyme borreliosis. Med J Aust, 168(10):500-2 
Stewart M; 
Lennox VA; 
et al. 
[From the abstract:] "We report a case of Lyme borreliosis. Culture of skin biopsy was positive for Borrelia garinii, despite repeated prior treatment with 
antibiotics.' 

"The results of conventional serological and histopathological tests were negative, despite an illness duration of at least two years." 

Page4 of 17 


Author Year Title 
Journal 

23. 
McCaulley, 1998 Guidelines for the clinical diagnosis of Lyme disease. Annals of Internal Medicine, 129(5): 422-423 
Mark E., M.D. 
[Letter to the Editor:] "The position paper on laboratory diagnosis of Lyme disease is based on a widely accepted paradigm that is inconsistent with a growing body 
of medical literature. According to this paradigm, cases of Lyme disease are overwhelmingly seropositive and are unlikely to be associated with persistent symptoms 
after presumed adequate therapy. In addition, any patients remaining persistently symptomatic are presumed to no longer have Lyme disease at all but rather to have 
such conditions as fibromyalgia, depression, or the chronic fatigue syndrome and, as a result, to be unlikely to respond to additional antibiotic therapy. Such 
presumptions are inconsistent with an increasing number of reports. 

A 1994 article reports the increased frequency of multiple symptoms in previously treated patients with Lyme disease compared with controls. Antibodies on ELISA 
were found in less than half of the patients with Lyme disease. Re-treatment was associated with improvement in half of re-treated patients. Had the guidelines been 
followed in a clinical evaluation of these or similar patients, Lyme disease would have been diagnosed in few of them. 

In a 1996 report, Borrelia burgdorferi plasmid DNA was detectable by polymerase chain reaction assay only in a subset of patients with Lyme disease who were 
seronegative. Many case reports have described patients with Lyme disease who remain antigen positive and symptomatic despite intensive antibiotic treatment. 

I suggest the acceptance of a new paradigm that incorporates the above information. Physicians involved in the treatment of Lyme disease should consider that 1) 
Patients with Lyme disease, especially those in late stages of the disease, are frequently seronegative; 2) the persistence of symptoms, which may be vague, is 
common and may respond to additional antibiotic therapy; and 3) there is much to be learned about the optimal treatment of Lyme disease at any stage." 

24. 
Petrovic M; 1998 Lyme borreliosis - a review of the late stages and treatment of four cases. Acta Clinica Belgica, 53(3):178-83 
Vogelaers D; VanRenterghem L;
Carton D; De [From the abstract:] "Difficulties in diagnosis of late stages of Lyme disease include low sensitivity of serological testing and late inclusion of Lyme disease in the 
Reuck J; differential diagnosis. Longer treatment modalities may have to be considered in order to improve clinical outcome of late disease stages...The different clinical cases 
Afschrift M. illustrate several aspects of late borreliosis: false negative serology due to narrow antigen composition of the used ELISA format, the need for prolonged antibiotic 
treatment in chronic or recurrent forms and typical presentations of late Lyme disease, such as lymphocytic meningo-encephalitis and polyradiculoneuritis." 

25. 
American 1997 Lyme encephalopathy may surface despite antibiotic treatment. http://www.medscape.com/CPG/ClinReviews/1997 
Academy of /v07.n06/c0706.cnu/c0706.cnu.html#Lyme 
Neurology 49th 
Annual Meeting "Of the 8 patients with CNS infection, only 2 were seropositive on both the ELISA and Western blot tests. Four had indeterminate ELISA results and a 
April 12-19. negative Western blot, and 2 had negative results on both the ELISA and the Western blot. Neither of the 2 seropositive patients had received antibiotics during the 
first month of infection for early localized or disseminated disease," said the Boston researchers. Of the 6 seronegative patients with CNS infection, however, 5 (84%) 
had received a recommended course of oral or intravenous antibiotics during the first month of infection.'" 

26. Branigan P; Rao J; 
Rao J; Gerard H; 
Hudson A; 
1997 PCR evidence for Borrelia burgdorferi DNA in synovium in absence of positive serology. American College of Rheumatology, 
Vol 40(9), Suppl:S270 
Williams W; 

Arayssi T; Pando "PCR evidence for Borrelia has been identified in synovial biopsies of patients with clinical pictures that had not initially suggested Lyme disease. 
J; Bayer M; All 6 PCR-positive] patients were negative for antibodies to Borrelia and some were PCR positive in synovium despite previous treatment with antibiotics.
Rothfuss S; 
Clayburne G;
Sieck M; 
Schumacher HR. 


Page5 of 17 


Author Year Title 
Journal 

27. Donta ST. 1997 Tetracycline therapy for chronic Lyme disease. 
Clin Infect Dis, Jul;25 Suppl 1:S52-6 
"Treatment outcomes for seronegative patients (20% of all patients) were similar to those for seropositive patients. Western immunoblotting showed reactions to one 
or more Borrelia burgdorferi-specific proteins for 65% of the patients for whom enzyme-linked immunosorbent assays were negative." 

28. Hauser U; 1997 Enzyme-linked immunosorbent assays with recombinant internal flagellin fragments Medical Microbiology & Immunology. 
Wilske B. 
derived from different species of Borrelia burgdorferi sensu lato for the serodiagnosis of 186(2-3):145-51 
of Lyme. 


[From the abstract:] "The serodiagnosis of early Lyme neuroborreliosis is hampered by false negative results and one of the reasons could be the heterogeneity of 
strains of Borrelia burgdorferi sensu lato." 

29. 
Pradella SP; 1997 Acute Borrelia infection. Unilateral papillitis as isolated clinical manifestation. Ophthalmologe, Aug;94(8):591-4 
Krause A; 
Muller A. 
[From the abstract:] "Seronegative values in subjects strongly suspected of having Lyme disease do not necessarily exclude the diagnosis of Lyme disease." 

30. Schumacher HR. 1997 PCR evidence for Borrelia burgdorferi DNA in synovium in absence of positive serology. Abstract ACR 61st National Scientific Meeting 
November 8-12 
31. Aberer E; 1996 Heterogeneity of Borrelia burgdorferi in the skin. American Journal of Dermatopathology, 
Kersten A; Klade 18(6):571-9 
H; Poitschek C; 
Jurecka W. 
"Neuralgias arising 6 months after ECM in spite of antibiotic therapy were evident in a seronegative patient who showed perineural rod-like borrelia structures." 

"The morphological forms of borreliae seen in biopsies were correlated with clinical findings. Seropositive patients showed clumped and agglutinated borreliae 
in tissue, whereas seronegative patients exhibited borreliae colony formation (n=2). ...the behavior of borreliae within collagen fibers is strongly influenced by 
immune recognition by the patient. Borrelia may escape immune surveillance by colony formation and masking within collagen, resulting in seronegativity." 

32. 
Breier P; Klade H; 1996 Lymphoproliferative responses to Borrelia burgdorferi in circumscribed scleroderma. Br J Dermatol, 134(2):285-91 
Stanek G; 
Poitschek C; 
Kirnbauer R; "These findings show that the pattern of Bb-specific immune responses is more complex than previously thought, and underscore the importance of lymphocyte 
Dorda W; function assays in evaluating the diagnosis of potential Bb infection in seronegative patients." 
Aberer E. 
Page6 of 17 


Author Year Title 
Journal 

33. 
Huppertz HI; 1996 Lymphoproliferative responses to Borrelia burgdorferi in the diagnosis of Lyme arthritis in Eur J Pediatr, 155(4):297-302 
Mosbauer S; children and adolescents. 
Busch DH; 
Karch H. "In one patient with seronegative LA [Lyme arthritis] specific lymphocyte proliferation and polymerase chain reaction for borrelial fla sequences in 
urine were positive." 

34. 
Luft BJ. 1996 Chronic Lyme disease: an evolving syndrome. 9th Annual International Scientific Conference on 
Lyme Disease & Other Tick-Borne Disorders, 
Boston, MA, April 19-20 
[From the abstract:] "In the case of the ticks, environmental factors such as temperature, humidity and source of blood meal may alter the major outer surface proteins 
(Osp) of the spirochete within the tick vector. ...Humans with chronic arthritis are more likely to show an immune response to Osp A." 

[Seronegativity:] "Chronic Lyme disease patients may be seropositive or seronegative with or without a documented history of Lyme disease." 

[Diagnosis:] "Since Lyme disease is a clinical diagnosis, research must continue to improve diagnostic assays using recombinant proteins which are more sensitive 
and specific than the whole organism sonicate used for both ELISA and Western blots." 

35. 
Luft BJ; Dattwyler 1996 Azithromycin compared with amoxicillin in the treatment of erythema migrans. Annals of Internal Medicine, 124(9):785-91 
RJ; Johnson RC; A double-blind, randomized, controlled trial. 
Luger SW; BoslerEM; Rahn DW; 
et al. "Fifty-seven percent of patients who had relapse were seronegative at the time of relapse." 
36. 
Mouritsen CL; 1996 Polymerase chain reaction detection of Lyme disease: correlation with clinical American Journal of Clinical Pathology. 
Wittwer CT; manifestations and serologic responses. 105(5):647-54 
Litwin CM; Yang 
L; Weis JJ; 
Martins TB; [From the abstract:] "...nine serum samples and one synovial fluid from patients with definite clinical features of Lyme disease were found to be negative by EIA and 
Jaskowski TD; Western blot analysis for IgG and IgM antibody, but contained B burgdorferi DNA, as detected by PCR. Polymerase chain reaction analysis of serum and synovial 
Hill HR. fluid may be of significant diagnostic value in Lyme disease, especially in the absence of a serologic response in early, partially treated and seronegative chronic 
disease....This is the first study to report an association between PCR positivity and the absence of a serologic response to Lyme borreliosis." 

37. Mursic VP; 1996 Formation and cultivation of Borrelia burgdorferi spheroplast L-form variants. Infection, 24(3):218-26 
Wanner G; 
Reinhardt S; 
Wilske B; et al. 
This study investigated In vitro morphological variants of B. burgdorferi, in an effort to explain the clinical persistence of active Lyme borreliosis despite antibiotic 
therapy. The authors suggest that these atypical forms may allow Borrelia to survive antibiotic treatment. 

"Penicillin G was the most effective inducer of SL-forms [spheroplast-L-forms). The reversion of this form to the helical parental forms was mostly achieved by 
cultivation of isolated SL-colonies in penicillin G-free medium. The atypical forms isolated from patients treated with antibiotics show similar features. The same 
effect is probably obtained with all other ß-lactam antibiotics." 

"With regard to the polyphasic course of Lyme borreliosis, these forms without cell walls can be a possible reason why Borrelia survive in the organism 
for a long time (probably with all beta-lactam antibiotics) [corrected] and the cell-wall-dependent antibody titers disappear and emerge after reversion." 

38. 
Preac Mursic V; 1996 Kill kinetics of Borrelia burgdorferi and bacterial findings in relation to the treatment of Lyme Infection, 24(1):9-16 
Marget W; Busch borreliosis. 
U; Pleterski 
Rigler D; Hagl S. “The patients had clinical disease with or without diagnostic antibody titers to B. burgdorferi." 
Page7 of 17 


Author Year Title 
Journal 

39. Pachner A. 1995 Early disseminated Lyme disease. 
Am J Med, 98 (suppl 4A):4A-30S-51S – Discussion. 
“The correlation between a positive Western blot and Lyme arthritis is probably the best of almost any Western blot and any Lyme disease manifestation. With neurologic 
disease, I have had a lot of patients who don't have a positive Western blot; they just have not developed a peripheral antibody response, for whatever reason.” 

40. 
Coyle PK; 1995 Detection of Borrelia burgdorferi-specific antigen in antibody negative cerebrospinal fluid in Neurology, 45:2010-2014 
Schutzer SE; neurologic Lyme disease. 
Deng Z; KruppLB; Belman MD; 
Benach JL; [From the abstract:] " RESULTS: Of the 35 of 83 (42%) patients who were positive for OspA antigen in their CSF, 15 (43%) were antigen positive despite being 
Luft BJ. antibody-negative in CSF. Seven of these 15 (47%) had otherwise normal routine CSF analyses. Six of these 15 (40%) patients met strict CDC surveillance criteria 
for Lyme disease: four (27%) patients had seroconversion coincident with new neurologic problems; and three (20%) with characteristic syndromes for Lyme disease 
were seronegative, but had complexed antibody to B. burgdorferi. The final two patients (13%) were seropositive and had unexplained neurologic problems not 
characteristic of Lyme disease. CONCLUSIONS: B. burgdorferi antigen can be detected in CSF that is otherwise normal by conventional methodology, and can be 
present without positive CSF antibody. Since CSF antigen implies intrathecal seeding of the infection, the diagnosis of neurologic infection by B. burgdorferi should 
not be excluded solely on the basis of normal routine CSF or negative CSF antibody analyses." 

[From the article:] "Prompt and precise diagnosis is difficult because basic microbiologic tests such as culture and staining have not been useful, on a broad scale, to 
document the presence of the spirochete in a body fluid. Instead, detection of specific antibodies to B burgdorferi in blood and CSF is commonly used to support 
or refute a clinical suspicion of infection. Many of the commercially available assays have been plagued by lack of sensitivity, specificity, and reproducibility. 
Furthermore, the absence of free antibodies to B burgdorferi components has been documented in well-characterized erythema-migrans-positive cases of Lyme 
disease, including those with prominent neurologic involvement." 

41. 
Karma A; Seppala 1995 Diagnosis and clinical characteristics of ocular Lyme borreliosis. American Journal of Ophthalmology, 
I; Mikkila H; 119(2):127-35 
Kaakkola S; 
Viljanen M;
Tarkkanen A. [From the abstract:] “Results of ELISA disclosed that five patients [out of ten] were seropositive, two patients showed borderline reactivity, and three 
patients were seronegative. Four of the five patients with borderline or negative results by ELISA had a positive result by western blot analysis. ... 
CONCLUSIONS: Late-phase ocular Lyme borreliosis is probably underdiagnosed because of weak seropositivity or seronegativity in ELISA assays." 

42. 
Lawrence C; 1995 Seronegative chronic relapsing neuroborreliosis. European Neurology, 35(2):113-7 
Lipton RB; LowyFD; Coyle PK. 
[From the abstract:] This article reports a Lyme disease patient "who experienced repeated neurologic relapses despite aggressive antibiotic therapy." The patient 
was seronegative. "Although the patient never had detectable free antibodies to B. burgdorferi in serum or spinal fluid, the CSF was positive on multiple occasions for 
complexed anti-B. burgdorferi antibodies, B. burgdorferi nucleic acids and free antigen." 

43. 
Millner M. 1995 Neurologic manifestations of Lyme borreliosis in children. Wiener Medizinische 
Wochenschrift,145(7-8):178-82 
"Our own observations in children which suffered from an acute neuroborreliosis (NB) showed the following:... Indeed, there is a seronegative NB also in children." 

Page8 of 17 


Author Year Title 
Journal 

44. Oksi J; Uksila J; 1995 Antibodies against whole sonicated Borrelia burgdorferi spirochetes, 41-Kilodalton flagellin, Journal of Clinical Microbiology, 33(9):2260-4 
Marjamaki M; and P39 protein in patients with PCR- or culture-proven late Lyme borreliosis. 
Nikoskelainen J; 
Viljanen MK. 
[From the abstract:] "These results show that antibodies to B. burgdorferi may be present in low levels or even absent in patients with culture- or PCR-proven late LB 
[Lyme borreliosis]. Therefore, in addition to serological testing, the use of PCR and cultivation is recommended in the diagnosis of LB." 

45. 
Skripnikova IA; 1995 The humoral immunological response of patients with Lyme disease. Ter Arkh, 67(11):53-6 
Anan'eva LP; 
Barskova VG; 
Ushakova MA. "Both acute and chronic borreliosis can be seropositive or seronegative." 
46. Schubert HD; 1994 Cytologically proven seronegative Lyme choroiditis and vitritis. Retina, 14(1):39-42 
Greenebaum E; 
Neu HC. 
[From the abstract:] "RESULTS: Intravitreal spirochetes consistent with Borrelia burgdorferi were found in this seronegative patient. CONCLUSION: Vitreous specimens of 
patients with choroiditis and vitritis of unknown cause should be examined cytologically, particularly when serologic results do not corroborate the clinical findings of Lyme 
disease." 

47. 
Sigal LH. 1994 The polymerase chain reaction assay for Borrelia burgdorferi in the diagnosis of Lyme Annals of Internal Medicine, 120(6):520-521 
disease. 
"Polymerase chain reaction may be more sensitive than antibody detection techniques in human Lyme neuroborreliosis [17,19] and the murine experimental model 

[22] and clearly is more sensitive than current culture techniques. Our experience suggests that a few patients may be positive by PCR despite negative 
immunologic assay results in inflammatory fluid and blood (Sigal LH and Liebling M. Unpublished observation)." 
48. 
Bojic I; 1993 Clinical characteristics of Lyme disease. Vojnosanit Pregl, 50(4):359-64 
Mijuskovic P;
Dokic M; Nozic D; [From the abstract:] "Clinical characteristics of Lyme disease were analysed in 22 patients. Erythema migrans was found in 20 (91%), arthralgia in 18 (81%), 
Lako B; et al. neuralgia in 8 (36%), encephalitis in 3 (13%), carditis in 2 (9%) and arthritis in 2 (9%) patients. The positive antibody titer was found in 14 (63%) patients. 
49. Coyle PK. 1993 
Antigen detection and cerebrospinal fluid studies. In "Lyme Disease," ed. P. Coyle, p.143 
"...spirochetes show a peculiar feature compared to other bacterial neurologic infections: the organisms can be present in CSF without inducing inflammatory 
changes. This is well-documented for neurosyphilis, leptospirosis, and relapsing fever, and appears to be occasionally true for Lyme disease as well. In 
Europe, B. burgdorferi has been cultured from otherwise normal CSF." 

50. 
Häupl T; Hahn G; 1993 Persistence of Borrelia burgdorferi in ligamentous tissue from a patient with chronic Lyme Arthritis & Rheumatism, 36(11):1621-6 
Rittig M; Krause borreliosis. 
A; Schoerner C; 
Schonherr U; et al. 
[From the abstract:] "The initially significant immune system activation was followed by a loss of the specific humoral immune response and a 
decrease in the cellular immune response to B burgdorferi over the course of the disease." [From the article:] "Interestingly, the cellular immune responses were also 
directed against the surface protein OspA during each recurrence of clinical symptoms, even though anti-OspA antibodies were not detectable by immunoblot." 

Page9 of 17 


Author Year Title 
Journal 

51. 
Hulinska D; 1993 Electron microscopy and the polymerase chain reaction of spirochetes from the blood of Central European Journal of Public Health. 
Krausova M; patients with Lyme disease. 1(2):81-5 
Janovska D; 
et al. [From the abstract:] "Results of studies using direct antigen detection suggest that seronegative Lyme borreliosis is not rare and support the hypothesis that Borrelia 
antigens can persist in humans." 

52. 
Kazakoff MA; 1993 Liver function test abnormalities in early Lyme disease. Arch Fam Med, 2(4):409-13 
Sinusas K; 
Macchia C. [From the abstract:] "PATIENTS: Thirty-seven female and 36 male patients with erythema migrans who had not yet been treated with antimicrobial agents. ... 
Only seven patients (9%) had a positive titer in response to the enzyme-linked immunosorbent assay for Lyme disease.” 

53. 
Liegner KB; 1993 Recurrent erythema migrans despite extended antibiotic treatment with minocycline in a Journal of the American Academy of Dermatology, 
Shapiro JR; patient with persisting Borrelia burgdorferi infection. 28(2 Pt 2):312-4 
Ramsay D;
Halperin AJ; 
Hogrefe W; [Abstract:] "Erythema migrans recurred in a patient 6 months after a course of treatment with minocycline for Lyme disease. Polymerase chain reaction on 
Kong L. heparinized peripheral blood at that time demonstrated the presence of Borrelia burgdorferi-specific DNA. The patient was seronegative by Lyme enzyme-linked 
immunosorbent assay but showed suspicious bands on Western blot. Findings of a Warthin-Starry stain of a skin biopsy specimen of the eruption revealed a 
Borrelia-compatible structure. Reinfection was not believed to have occurred. Further treatment with minocycline led to resolution of the erythema migrans." 

54. Schutzer SE. 1993 Seronegative Lyme disease. 
In "Lyme Disease," ed. P. Coyle, p.192 
"The number and percentage of seronegative Lyme disease cases remain controversial. At some academic centers the estimate is 5%, and in certain private settings 
the number may be higher. There is little question that seronegative Lyme disease can exist." 

55. Sigal LH. 1993 Lyme disease: testing and treatment. Who should be tested and treated for Lyme disease Rheum Dis Clin North Am, 19(1):79-93 
[From the abstract:] "LD is not a diagnosis that can be made on the basis of serologic testing. By this is meant that vague symptoms plus a positive serologic test do 
not assure that the patient has LD. On the other hand, a patient with ECM or other manifestations of LD may still be seronegative." 

56. Steere AC. 1993 Seronegative Lyme disease. 
JAMA, (270):1369. 
"The number and percentage of seronegative Lyme disease cases remain controversial. At some academic centers the estimate is 5%, and in certain private settings 
the number may be higher. There is little question that seronegative Lyme disease can exist." 

57. Preac-Mursic V; 1993 First isolation of Borrelia burgdorferi from an iris biopsy. 
J Clin Neuroophthalmology, 13(3):155-61; discussion 162. 
Pfister HW; 
Spiegel H; Burk R; [From the abstract:] “Antibiotic therapy may abrogate the antibody response to the infection as shown by our results. Patients may have subclinical or clinical 
Wilske B; el al. disease without diagnostic antibody titers. Persistence of B. burgdorferi cannot be excluded when the serum is negative for antibodies against it.


58. 
Oksi J; Viljanen 1993 Fatal encephalitis caused by concomitant infection with tick-borne encephalitis virus and Clinical Infectious Diseases, 16(3):392-6 
MK; Kalimo H; Borrelia burgdorferi. 
Peltonen R; 
Marttia R; 
Salomaa P; "Serology for Borrelia was negative. Autopsy revealed necrotizing encephalitis and myelitis with involvement of the dorsal root ganglion. With use of 
et al. polymerase chain reaction tests, segments of two separate genes of B. burgdorferi were amplified from the patient's CSF." 
Page 10 of 17 


Author Year Title 
Journal 

59. 
Keller TL; 1992 PCR detection of Borrelia burgdorferi DNA in cerebrospinal fluid of Lyme neuroborreliosis patients. Neurology, 42(1):32-42 
Halperin JJ;
Whitman M. "PCR detected B burgdorferi DNA in the CSF of seven patients whose blood serology ...failed to demonstrate prior exposure to the organism. The failure of existing 
diagnostic methods to detect patients exhibiting well-recognized manifestations of Lyme disease has been described. This has generally been attributed to abrogation of the 
host immune response by noncurative antimicrobial treatment. This explanation seems unlikely in at least four of the seven patients who had no recollection of having 
received antibiotics. " 

60. Banyas GT. 1992 Difficulties with Lyme serology. 
J Am Optom Assoc, 63(2):135-9 
[From the abstract:] "A major problem has been seronegativity in persons possessing the disease (false negatives). At present, seronegativity in persons strongly 
suspected of having Lyme disease does not necessarily exclude the diagnosis of Lyme disease. The clinician must recognize this in patients who may have Lyme 
disease or a recurrence of the disease." 

61. Dinerman H; 1992 Lyme disease associated with fibromyalgia. Annals of Internal Medicine, 117:281-5 
Steere AC. 
"The small percentage of patients who are seronegative by enzyme-linked immunosorbent assay (ELISA) later in the illness usually have positive 
Western blots or cellular immune responses to borrelial antigens (9,10)." 

62. Fraser DD; Kong 1992 Molecular detection of persistent Borrelia burgdorferi in a man with dermatomyositis. Clinical & Experimental Rheumatology, 
LI; Miller FW. 10(4):387-90. 
[From the abstract:] "Antibiotic therapy was reinstituted after Borrelia burgdorferi was detected in the patient's peripheral blood leukocytes by the polymerase chain 
reaction (PCR). All serologic, T-cell stimulation, and western blot analyses, however, were negative... In addition, this case emphasizes the potential clinical utility of 
PCR technology in evaluating the persistent sero-negative Lyme disease which may occur in immunocompromised individuals." 

63. Keller TL; 1992 PCR detection of Borrelia burgdorferi DNA in cerebrospinal fluid of Lyme neuroborreliosis Neurology, 42(1):32-42 
Halperin JJ; patients. 
Whitman M. 
[Abstract:] "We used the polymerase chain reaction (PCR), a method useful in the detection of Borrelia burgdorferi in vitro, to evaluate CSF in patients thought to have 
neuroborreliosis. Nested pairs of oligonucleotide primers were designed to recognize the C-terminal region of B burgdorferi OspA. CSF samples were obtained from 

(1) patients with immunologic evidence of systemic B burgdorferi infection and clinical manifestations suggestive of CNS dysfunction, (2) seronegative patients with 
clinical disorders consistent with Lyme borreliosis, and (3) patient and contamination controls; all were analyzed in a blinded fashion. PCR detected B burgdorferi 
OspA DNA in CSF of (1) 10 of 11 patients with Lyme encephalopathy, (2) 28 of 37 patients with inflammatory CNS disease, (3) seven of seven seronegative patients 
with Lyme-compatible disorders, and (4) zero of 23 patient controls. Zero of 83 additional contamination controls were PCR-positive” 
64. Faller J; 1991 PCR detection of Borrelia burgdorferi DNA in cerebrospinal fluid of Lyme neuroborreliosis Foot & Ankle, 11(4):236-238. 
Thompson F; patients. 
Hamilton W. 
"Patient 8 had several negative ELISA assays. She then had a lymphocyte reactivity test for cell mediated immune (CMI) response to Borrelia burgdorferi antigen. Her 
peripheral blood lymphocytes were markedly responsive to the spirochete, with an index of 46 (18 is three standard deviations above the controls.)" 

65. Reik L, Jr. 1991 Lyme Disease and the Nervous System. 
New York: Thieme Medical Publishers, Inc. 
“In some cases, specific serum antibody is present but sequestered in immune complexes, and therefore not measurable by routine ELISA.” 
“...test results for serum antibodies are not always positive when neurologic abnormalities develop, especially in stage 2.” 

66. 
Havlik J; 1991 Seronegative Lyme borreliosis. The 5th European Congress of Clinical Microbiol. 
Rohacova H; and Infect. Diseases, Sept. 8-11, Oslo, Norway:1385. 
Hulinska D; et al. 
67. Nields JA; 1991 Tullio phenomenon and seronegative Lyme borreliosis. Lancet, 338(8759):128-9 
Kueton JF. 
68. Steere AC. 1991 Rheumatology research in the 90s. 
Rheumatology News 
Page 11 of 17 


Author Year Title 
Journal 

69. 
Nadelman RB; 1990 Isolation of Borrelia burgdorferi from the blood of seven patients with Lyme disease. American Journal of Medicine, 88:21-6 
Pavia CS; 
Magnarelle LA;
Wormser GP. 
"The absence of significant antibody titers to B. burgdorferi is not uncommon in Lyme disease, especially in early disease. ...Although it was initially believed 
that patients with neurologic Lyme disease generally have antibodies to B. burgdorferi, this may not always be the case. ...We would advise that in an endemic 
area, the differential diagnosis of nonspecific muscle and joint aches without rash should include Lyme disease--even in the absence of antibodies to B. burgdorferi.” 

70. 
Schutzer SE; 1990 Sequestration of antibody to Borrelia burgdorferi in immune complexes in seronegative Lancet, 335(8685):312-5 
Coyle PK; Belman Lyme disease. 
AL; Golightly MG;
Drulle J. 
[From the abstract:] "Apparent B burgdorferi seronegativity in serum immune complexes may thus be due to sequestration of antibody in immune complexes." 

71. 
Bianchi G; 1990 Articular involvement in European patients with Lyme disease. A report of 32 Italian patients. British Journal of Rheumatology, 29(3):178-80 
Rovetta G; 
Monteforte P; 
Fumarola D; 
et al. [From the abstract:] “In addition, interpreting serological tests for antibodies against B. burgdorferi and the real prevalence of arthritis in LD [Lyme disease] is 
complicated by the possible existence of seronegative LD and by the effect of early antibiotic treatment." 

72. Dieterle L; Kubina 1989 Neuro-borreliosis or intervertebral disk prolapse? Dtsch Med Wochenschr, 114(42):1602-6 
FG; Staudacher T; 
Budingen HJ. 
[From the abstract "Between September 1986 and November 1988, 17 patients were hospitalized and treated for neuro-borreliosis. ...Three of 14 patients had no IgG 
antibodies against Borrelia, either in serum or cerebrospinal fluid at the initial examination, two had positive titres in serum only. 

73. Guy EC; 1989 Seronegative neuroborreliosis. Lancet, 1:441 
Turner AM. 
“We wish to report a case with a clinical diagnosis of acute Lyme neuroborreliosis for whom negative serology was reported by a Lyme disease referral centre using 
ELISA. ... 

Western blot analysis of sera revealed both IgM and IgG binding to several B burgdorferi proteins, compatible with acute Lyme disease. Antibody binding to a larger 
number of B burgdorferi proteins was found when the CSF was tested similarly. Our observation of a more diverse antibody response in CSF compared with serum is 
consistent with the findings of a previous study which showed 44% of patients with neurological manifestations of Lyme disease to have positive CSF antibody titres 
but negative serum titres when measured by ELISA. Analysis of CSF for the detection of IgM and IgG binding to B burgdorferi is essential if seronegative cases of 
acute Lyme neuroborreliosis are to be identified.” 

74. MacDonald AB. 1989 Gestational Lyme borreliosis. Implications for the fetus. 
Rheum Dis Clin North Am, 15(4):657-77 
"From a biologic perspective, most of the fatal cases of LB [Lyme borreliosis] in pregnancy were reactive either in titers in the borderline region or were completely 
nonreactive in serologic tests. The tendency toward seronegativity in pregnancy makes maternal serology a less satisfactory discriminator of maternal infection and 
useless as a practical tool to predict the actual state of the fetus..." 

75. Trock DH; Craft 
JE; Rahn DW. 
1989 Clinical manifestations of Lyme disease in the United States. Connecticut Medicine, Vol 53, No. 6 
"...cases of seronegative Lyme disease have been reported..." 
Page 12 of 17 


Author Year Title 
Journal 

76. 
Dattwyler RJ; 1989 Immunologic aspects of Lyme borreliosis. Rev Infect Dis, II(Suppl 6):S1494-98. 
Volkman DJ; 
Luft BJ. "Three separate groups of investigators have reported individuals who lacked diagnostic levels of specific antibody in their serum, yet had neurologic involvement and 
diagnostic levels of antibody in their CSF. We have confirmed this finding in our laboratory.” 

77. 
Preac-Mursic V; 1989 Survival of Borrelia burgdorferi in antibiotically treated patients with Lyme borreliosis. Infection, 17(6):355-9 
Weber K; Pfister 
HW; Wilske B; 
Gross B; [From the abstract:] "We conclude that early stage of the disease as well as chronic Lyme disease with persistence of B. burgdorferi after antibiotic therapy cannot 
Baumann A; be excluded when the serum is negative for antibodies against B. burgdorferi." 
Prokop J. 
[Seronegativity:] "As shown, negative antibody-titers do not provide evidence for successful therapy; antibody-titers may become negative despite persistence of 

B. burgdorferi." (p.358) 
78. 
Berger BW; 1988 Use of an autologous antigen in the serologic testing of patients with Journal of the American Academy of Dermatology, 
MacDonald AB; erythema migrans of Lyme disease. 18(6):1243-6 
Benach JL. 
[Abstract:] "We attempted to detect an early rise in antibody titers to Borrelia burgdorferi in the serum of patients with erythema migrans of Lyme disease by utilizing 

B. burgdorferi isolates obtained from patients' own skin lesions instead of the B31 reference strain. B. burgdorferi was isolated from nine of 23 skin biopsy 
specimens submitted for culture. Elevated antibody titers were not detected in any of the 23 acute serum samples by immunofluorescence assay. The antigens 
derived from patient isolates were no more effective than the reference strain in detecting antibodies in patients with early Lyme disease." 
79. 
Dattwyler RJ; 1988 Seronegative Lyme disease. Dissociation of specific T- and B-lymphocyte responses to New England Journal of Medicine, 
Volkman DJ; Luft Borrelia burgdorferi. 1;319(22):1441-6 
BJ; Halperin JJ; 
Thomas J; [From the abstract:] "Although these patients had clinically active disease, none had diagnostic levels of antibodies to B. burgdorferi on either a standard enzyme-
Golightly MG. linked immunosorbent assay or immunoflourescence assy. ...We conclude that the presence of chronic Lyme disease cannot be excluded by the absence of 
antibodies against B. burgdorferi and that a specific T-cell blastogenic response to B. burgdorferi is evidence of infection in seronegative patients with clinical 
indications of chronic Lyme disease." 

80. 
Pachner AR. 1988 Borrelia burgdorferi in the nervous system: the new "great imitator". Annals of the New York Academy of Sciences, 
539:56-64 
"The antibody response in serum in CNS Lyme disease seems to be related to the presence of other manifestations; patients who have had both arthritis and CNS 
disease have quite high titers, while those with only CNS disease sometimes do not." 

81. 
Lavoie PE; 1987 Culture positive seronegative transplacental Lyme borreliosis infant mortality. Arthritis Rheum, Vol 30 No 4, 3(Suppl):S50 
Lattner BP; 
Duray PH; 
Barbour AG; “We report a culture positive neonatal death occurring in California, a low endemic region. ...Bb was grown from a frontal cerebral cortex inoculation. The spirochete 
Johnson HC. appeared similar to the original Long Island tick isolate. Silver stain of brain & heart was confirmatory of tissue infection. The mother had been having migratory 
arthralgias and malaise since experiencing horse fly & mosquito bites while camping on the Maine coast in 1971. The family was seronegative for LB by ELISA at 
Yale. Cardiolipin antibodies were also not found." 

82. MacDonald AB. 1987 Lyme disease. A neuro-ophthalmologic view. Journal of Clinical Neuro-Ophthalmology, 
7(4):185-90 
[From the abstract:] "Potential pitfalls in the diagnosis of Lyme disease with an emphasis on false negative serology and currently available 
diagnostic modalities are presented." 

Page 13 of 17 


Author Year Title 
Journal 

83. Hederstedt B; 1986 [Borrelia-diagnos aktuell aret om visar serologisk undersokning av 1985 ars fall.] Lakartidningen, 83:3987-89 
Hovmark A; 
Stiernstedt G; 
Asbring E. 
[According to Guy & Turner, 1989, this study found that 44% of patients with neurological Lyme disease had positive CSF antibody titres but negative serum titres 
when measured using ELISA testing.] 

84. 
Schmidt R; 1985 [Erythema migrans borreliosis in the Federal Republic of Germany. Deutsche Medizinische Wochenschrift, 
Kabatzki J; Epidemiology and clinical aspects.] 110(47):1803-

Hartung S;
Ackermann R. 
[Abstract:] "A positive antibody titre against Ixodes-ricinus-Borrelia (burgdorferi), using indirect immunofluorescence or ELISA, could be detected in serum and (or) 
liquor of 935 (32%) out of a total of 2955 patients between January 1984 and July 1985. In 289 of these cases the typical clinical manifestations were lacking whereas 
a characteristic disease picture enabled a diagnosis to be made in 171 patients with negative or borderline antibody titres. The 1106 cases of infection observed 
covered all regions of the country. A typical clinical syndrome was seen in 817 (74%) of these. Most common were erythema chronicum migrans (n = 458) and 
meningopolyneuritis Garin-Bujadoux-Bannwarth (n = 404); in 42% of the cases meningopolyneuritis was preceded by an erythema. Arthritis (n = 63), acrodermatitis 
chronica atrophicans (n = 72), carditis (n = 13) and lymphadenosis benigna cutis (n = 5) were much less common. Chronic Borrelian encephalomyelitis (n = 45) 
appeared surprisingly often (n = 45). The fact that in 73% of cases the various syndromes appeared alone, were double in 24% and combined only in 3%, illustrates 
the polymorphic nature of this disease." 

Other Spirochetes 

85. 
Stephan C; 2000 [Leptospirosis after a staff outing]. Dtsch Med Wochenschr, 125(20):623-627. 
Hunfeld KP; 
Schonberg A; “[From the abstract:] "The symptoms are non-specific and, moreover, in some cases the laboratory tests are negative, so that clinical diagnosis remains crucial.” 
et al. 
86. 
Jacobs R. 1999 Infectious Diseases: Spirochetal. Syphilis. Current Medical Diagnosis & Treatment 1999. 
38th Edition. Appleton & Lange. Stamford, CT. 
Ed. Tierney LM Jr; McPhee SJ, Papadakis MA. 
“The VDRL titer is usually high (>1:32) in secondary syphilis and tends to be lower (< 1:4) or even negative in late forms of syphilis.” 

87. Vecsei AK, 1999 [Congenital syphilis: late diagnosis in spite of screening]. Wien Klin Wochenschr, 111(10):410-3 
Vecsei PV, 
Dangl-Erlach E, 
[Abstract:] "We report the case of an infant in whom congenital syphilis was diagnosed at the age of 5 weeks. The case is remarkable because of (a) the negative 
venereal disease laboratory test from the cord blood, (b) the incidental diagnosis of the disease in the fifth week of life, (c) pneumonia alba being one of the 
symptoms, (d) the occurrence of a mild Jarisch-Herxheimer reaction after initiation of penicillin therapy and (e) the successful treatment of infection related anaemia 
with recombinant human erythropoietin." 

Page 14 of 17 


Author Year Title 
Journal 

88. 
Uribe CS; 1998 [Neurosyphilis and the prozone effect]. Rev Neurol, 27(160):970-2 
Garcia FA. 
[Abstract:] "INTRODUCTION: Neurosyphilis (NS) is an entity which still frequently presents to our Neurology Department. The prozone phenomenon occurs in 
approximately 2% of all cases of late primary syphilis or secondary syphilis; we have found no cases described of prozone and neurosyphilis occurring together. 
CLINICAL CASE: We present the unusual case of a 44 year old patient with NS and dementia PGP (progressive general paralysis). Initially serum VDRL was 
negative, but in CSF reacted at dilutions of 1:32. When serum VDRL was repeated using dilutions, it was reactive 1:128 and serum FTA was also reactive. The patient 
was treated with i.v. crystalline penicillin, after which his condition improved. CONCLUSIONS: We wish to draw attention to the possibility that patients with a 
dementia syndrome and negative serum VDRL may have the prozone phenomenon, and the laboratory should therefore be asked to do serial dilutions." 

89. 
Tikjob G; Russel 1991 Seronegative secondary syphilis in a patient with AIDS: identification of Treponema pallidum Journal of the American Academy of Dermatology, 
M; Petersen CS; in biopsy specimen. 24(3):506-8 
Gerstoft J; 
Kobayasi T. 
90. 
Berkowitz K; 1990 False-negative syphilis screening: the prozone phenomenon, nonimmune hydrops, and Am J Obstet Gynecol, 163(3):975-7 
Baxi L; Fox HE. diagnosis of syphilis during pregnancy. 
[From the abstract:] "Recently we encountered four cases of false-negative syphilis serologic results in women who gave birth to infants with congenital syphilis. The 
false-negative results were caused by the prozone phenomenon. The prozone phenomenon, seen during primary and secondary syphilis, occurs because a higher 
than optimal amount of antibody in the tested sera prevents the flocculation reaction typifying a positive result in reagin tests. Serum dilution is necessary to make the 
correct diagnosis. We recommend that for any pregnant woman with apparently negative syphilis serologic results in whom fetal compromise of unknown etiology 
exists, particularly nonimmune hydrops, nontreponemal testing should be repeated using serum dilutions to prevent a missed diagnosis of syphilis. We further 
recommend serum dilution as a routine procedure for all pregnant women in areas of high syphilis prevalence." 

91. 
Borisenko KK; 1989 [Characteristics of the course of latent forms of syphilis]. Vestn Dermatol Venerol, (11):25-9 
Vinokurov IN; 
Toporovskii LM. 
[Abstract:] "Seven patients with latent syphilis are described, in whom the routine serologic tests (RST) were negative during the first examination and over the course 
of therapy, and the specific tests (T. pallidum immobilization and immunofluorescence) were repeatedly positive before therapy. Early latent seropositive recurrent 
forms of syphilis were detected in the majority of these patients' sexual partners. The patients were not administered antisyphilis therapy before. The diagnosis of 
latent seronegative early syphilis negative in the RST is epidemiologically significant, for it helps timely carry out the necessary treatment and prophylaxis measures 
to prevent the disease dissemination." 

92. Ovcinnikov NM. 1981 [Important problems in the serodiagnosis of syphilis.]

Vestn Dermatol Venerol, 8:22-26 
[According to Mattman L., 1993: "It is thought [by Ovcinnikov] that false negative serological tests for syphilis may be explained because cystic and granule stages of the 
treponeme have not stimulated antibody reactive with the spirochetal stage."] 

93. Il'in II. 1981 [Seronegative forms of latent syphilis]. Vestnik Dermatologii i Venerologii. (1):66-9 
Pakhomova LV. 
Page 15 of 17 


Author Year Title 
Journal 

94. Sparling PF. 1971 Diagnosis and treatment of syphilis. 
New England Journal of Medicine, 284: 642-653 
Some infected patients had negative or equivocal serologic tests for syphilis. “These studies [reviewed] emphasize the fact that late syphilis can occur 
even if all serologic tests are negative.” 

Includes a review of recent [as of 1971] evidence indicating that penicillin treatment is not always curative in patients with late syphilis. "Penicillin therapy of 

neurosyphilis has not been as effective [as in early syphilis]. Several studies have reported relapses... Clinical progression of symptomatic neurosyphilis is relatively 

common despite antibiotics." (p.650) 

95. 
Ch'ien L. 1970 Seronegative dementia paralytica: report of a case. Journal of Neurology, Neurosurgery & Psychiatry. 
Hathaway BM. 33(3):376-80 
Israel CW. 
96. Hallock J; 1968 Congenital syphilis in an infant of a seronegative mother. Obstet Gynecol, 32(3):336-8 
Tunnessen WW. 
97. Roitburd MF. 1968 [2 cases of seronegativity in secondary syphilis]

Vestn Dermatol Venerol, 42(8):82-3 
98. Smith JL. 1968 Spirochetes in late seronegative syphilis, despite penicillin therapy. 
Med Times, 96(6):611-23 
99. 
Smith JL; 1967 Spirochetes in the aqueous humor in seronegative ocular syphilis. Persistence after Archives of Ophthalmology, 44:474-477 
Israel CW. penicillin therapy. 
"The purpose of this communication is to report the presence of spirochetes in the aqueous humor both before and after the intramuscular administration of 
9,200,000 units of long-acting penicillin for seronegative ocular syphilis. The treponemes were found by the flourescein antibody technique, in which the spirochetes 
stained with flourescein tagged anti-Treponema pallidum globulin when viewed with ultraviolet microscopy. ... 

Spirochetes have now been found in aqueous humor, cerebrospinal fluid, liver, and lymph nodes in several patients with late seronegative syphilis." 

100. Smith JL; 1967 The presence of spirochetes in late seronegative syphilis. JAMA, 199(13):980-984 
Israel CW. 
[Abstract:] "Late seronegative syphilis refers to clinical signs of ocular or neurosyphilis in a patient whose routine blood (reagin) test is nonreactive, but in whom a 
specific treponemal test is reactive. This report documents the presence of spirochetes in aqueous humor, cerebrospinal fluid, and at liver biopsy in such patients. 
Spirochetes have been found in the aqueous humor with no biomicroscopic abnormality and in cerebrospinal fluids which had normal cell counts, protein levels, and 
reagin and colloidal gold test results. Identification of the organisms depends on use of the flourescein antibody technique, in that spirochetes stain with 
flourescein-tagged anti-Treponema pallidum globulin on ultraviolet microscopy." 

[From the article:] "The finding of motile spirochetes in the aqueous humor of animal eyes which showed no clinical signs of inflammation at the time of paracentesis 
led to the initial clinical studies reported here. It must be emphasized that several patients were found to have negative findings in darkfield examinations of aqueous 
humor and cerebrospinal fluid, in which later study with the fluorescent antibody technique revealed morphologically typical spirochetes which stained with anti-T 
pallidum globulin. Darkfield examination of CSF requires that the fluid be centrifuged and examined within ten minutes after lumbar puncture in order to see motile 
treponemes." 

Page 16 of 17 


Author Year Title Journal 
101.Smith JL; Singer 
JA; Moore MB, Jr; 
1965 Sero-negative ocular and neuro-syphilis. American Journal of Ophthalmology, 59:753-762 
Yobs AR. 

102. Taylor WH; Smith 1965 Experimental seronegative syphilis. American Journal of Ophthalmology, 
JL; Singer JA. 60:1093-1098 
103. McCord. 1929 Study of two hundred autopsies made on syphilitic foetuses. Am J Obst & Gynec, 18:597 


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