Borreliosis de Lyme: estudio de posibles vectores ixódidos y evaluación de métodos de diagnóstico microbiológico.
Zoonosis caused by bacteria of the genre Borrelia. by the accidental
transmission by infected ticks. The genre Borrelia. concerns to the family
Spirochateaceae, and they are 4 the grounds of Lyme's borreliosis, associated in the
complex broad Borrelia burgdorferi sensu (B. burgdorferi sensu stricto, B. afzelii, B.
garinii and B. Valaisiana) The adherence of 48 hours is the demanded minimum, for
the inoculation of Borrelia burgdorferi sensu lato.
Diagnostic of the Lyme borreliosis: Direcly diasnostic by detection of spirochaeta
with the uses of the clinical specimens (skin biopsies, CSF semple, peripheral blood,
etc.) of patients whith conventional laboratory tintions is relative easy to reconized.
Culture of the organism in BSK medium, molecular characterization of Borrelia isolates
by gene sequences from DNAs amplified (flagellin (flaB), and outer surface protein A
(ospA)) from ticks or patiens confirmed the presence of several Borrelia species.
In the indirect diagnosis use as methods of reference, the
enzimoinmunoanálisis as test of screening and the inmunotransferencia for the
1.- Recolection and caracterization of ixodid ticks from diferent areas of
2.- Investigated la presencia de Borrelia burgdorferi sensu lato in these ixode
3.-To select clinical samples with result in the test of screening positive EIA.
4.-To analyze the above mentioned samples by means of other specific and
confirmatory tests(proofs) as well as to evaluate the presence of cross-reactivity with
other processes nosológicos.
5.-To study the clinical characteristics and evolution of the patients with
serologic results suggestive of borreliosis of Lyme as well as to evaluate his
Materials and methods: Obtaining and characterization of ixodids, one proceeded to
the detection of Borrelia burgdorferi by means of microscopic study, culture and PCR.
The selection of clinical samples, it was realized by means of the method EIA, and the
study of crossed reactivity and his confirmation by means of specific tests. And finally,
there was realized the analysis of signs and clinical symptoms and his relation with
results serológicos. There were realized softened and dissections those who a
microscopic study was practised and culture in BSK II.
In the 2º periode, we made a molecular characterization of B. Burgdorferi from
ticks by gene sequences DNAs amplified (nexteed PCR using 2 primers of gen of outer
surface protein A (OspA), by el Guttman metode et all that permeted the obtention in
the 2ª amplification, a fragment of 345 pairs of bases. By the flagelina, gene, we
using too 2 pairs of primers of the flagelin gene, by the Lebech metode et al. That
allow to obtain in 2 ª amplification, a fragment of 275 bases pair. For the serologic
study, there was in use first a method of sifted EIA IgM-IgG System VIDAS LYT (of
bioMèrieux) and a complementary method IFI Lyme Spot (also of bioMèrieux) that
determines antibodies of class IgM and IgG separately. As confirmatory method, there
was in use the inmunotransferencia B. burgdorferi B31 (of MarDx Diagnostic) and B.
garinii genogrupo 2 (of Gull Laboratories).
The serologic cross reactivity, whith Treponema pallidum, Coxiella burnetii,
Rickettsia conori and Brucella mellitensis, Epstein-Barr virus, Citomegalovirus virus and
AIDS virus. The population of patients corresponded(fitted) with the attended ones in
the area of the HGUV.
1.-The collection and identification of ixodid ticks proceeding from different
areas of the province of Valencia, it has revealed the existence of five genres, with
seven different species. None of them, classified as Ixodes ricinus, recognized principal
vector of Lyme's borreliosis.
2.-En none of the studied ticks has been identified an espiroqueta compatible
with Borrelia burdorferi sl. or another Borrelia's species associated with borreliosis of
3.-We considered that the method most adapted for the microbiological study in
the ixodid, is the ticks dissection with extraction of its glandular structures and medium
intestines. The use of specimens softened for detection for molecular methods is
dissuaded by the probable inhibition of the amplification of the specific ADN
4.-The enzimoinmunoanálisis method of as screening test, a population of
patients selects with clinical very changeable characteristics, but also a group of
patients with typical manifestations and with a concrete clinical evolution, which they
sustain the existence of probable cases of borreliosis of Lyme.
5.-La qualifications of the specific antibodies of class IgM and IgG by means of
inmunofluorescencia and the criteria established for the interpretation of the
confirmatory tests of inmunotransferencia with B. burgdorferi antigens and B. garinii,
prove to be scantily profitable in our way, since they allow neither confirm nor to
discard the existence of the disease in the group of studied patients.
Allows to consider the need of a better epidemiological and clinical study of the
suspicious cases of borreliosis of Lyme, and particularly the development of criteria of
microbiological diagnosis us that they serve to the profile of the disease in our area.
Document Full Text
Advisor:Muñoz Collado, Carlos; Fraile Fariñas, Teresa
School:Universitat de València
Source Type:Master's Thesis
Keywords:microbiologia i ecologia
Date of Publication:05/07/2004