Konservoituneiden lysiinien merkitys Borrelia gariniin DbpA- ja DbpB-adheesioproteiineissa

Tämän syventävien opintojen kirjallisen työn aiheena on Lymen borrelioosi. Lymen borrelioosi on Borrelia burgdorferi bakteerin aiheuttama tauti, joka yleensä leviää ihmiseen puutiaisen pureman välityksellä. Lymen borrelioosia levittäviä puutiaisia esiintyy laajalti pohjoisella pallonpuoliskolla. Lymen borrelioosi voi aiheuttaa monenlaisia oireita; yleisimmin ihoon, niveliin tai hermostoon. Selviytyäkseen ihmiselimistössä borrelia-bakteerit käyttävät pinnallaan esiintyviä proteiineja tarttuakseen isäntäelimistön soluihin. Tämän tutkimuksen aiheena oli tutkia borrelia-bakteerien pinnalla esiintyviä Dbp-proteiineja. Tutkimuksessa pyrittiin selvittämään Dbp-proteiinien aminohapporakenteen merkitystä borrelia-bakteerien kyvylle tarttua ihmissoluihin. Borrelia-bakteerien pintaproteiinien ja niiden sitoutumismekanismien tunteminen voi tulevaisuudessa auttaa kehittämään rokotteita, uusia lääkkeitä tai diagnostisia menetelmiä Lymen borrelioosin hoitoon. Tutkimusta varten tuotettiin bakteerikanta, jonka Dbp-proteiinien rakenne erosi normaalista Dbp-proteiineista tutkittavan aminohapposekvenssin osalta. Tämän mutatoidun borrelia-kannan kykyä sitoutua ihmissoluihin tutkittiin sitoutumiskokein. Sitoutumiskokeissa kannan sitoutumiskykyä verrattiin vastaavien normaaleja Dbp-proteiineja pinnallaan omaavien kantojen kykyyn sitoutua samanlaisten solujen pinnalle. Sitoutumiskokeiden tulokset olivat vaihtelevia. Suuressa osassa sitoutumiskokeen toistoista mutatoidun kannan bakteerit sitoutuivat heikommin ihmissoluihin kuin normaalia proteiinia pinnallaan ilmentävät kannat. Oli siis nähtävissä suuntaus, että mutaatio heikensi bakteerien sitoutumiskykyä ihmissoluihin. Tämä viittaa, että tutkimuksen aiheena oleva aminohapporakenne olisi tärkeä borrelia-bakteerien sitoutumiskyvyssä ihmissoluihin. Eri toistojen suuren vaihtelun ja negatiivisena kontrollina toimineen kannan liian suuren sitoutumiskyvyn vuoksi tutkimuksen tulokset eivät kuitenkaan ole täysin luotettavia ja aiheesta tarvitaan lisää tutkimusta

C6 peptide enzyme immunoassay in Lyme borreliosis serology

The cut-off values used in C6 peptide-based enzyme immunoassay (EIA), a widely used test in Lyme borreliosis (LB) serology, have not been thoroughly analysed. The objective of the study was to examine the performance of the C6 EIA, and to determine optimal cut-off values for the test. The analysed data contained results of 1368 serum samples. C6 EIA index values were compared statistically with the immunoblot (IB) test results. The identified cut-off values were further tested in a well-defined LB patient cohort. Cut-off value 1.6 appeared to be optimal when C6 EIA was used as a stand-alone test. When using C6 EIA as the first-tier test, the optimal cut-off values were 0.9 and 2.4 for negative and positive results. When C6 EIA was used as a second-tier test, samples yielding C6 index values >= 3.0 could be considered positive. The identified cut-off values had also a high sensitivity to identify seropositivity among definite LB patients. The identified cut-off values refine the role of C6 EIA in LB serology. Importantly, the use of C6 EIA leads to a reduction in the number of samples that need to be analysed using an IB, thus also reducing the costs. Two alternative workflows for LB serology including the C6 EIA are suggested.Peer reviewe

C6 peptide enzyme immunoassay in Lyme borreliosis serology

The cut-off values used in C6 peptide-based enzyme immunoassay (EIA), a widely used test in Lyme borreliosis (LB) serology, have not been thoroughly analysed. The objective of the study was to examine the performance of the C6 EIA, and to determine optimal cut-off values for the test. The analysed data contained results of 1368 serum samples. C6 EIA index values were compared statistically with the immunoblot (IB) test results. The identified cut-off values were further tested in a well-defined LB patient cohort. Cut-off value 1.6 appeared to be optimal when C6 EIA was used as a stand-alone test. When using C6 EIA as the first-tier test, the optimal cut-off values were 0.9 and 2.4 for negative and positive results. When C6 EIA was used as a second-tier test, samples yielding C6 index values ≥3.0 could be considered positive. The identified cut-off values had also a high sensitivity to identify seropositivity among definite LB patients. The identified cut-off values refine the role of C6 EIA in LB serology. Importantly, the use of C6 EIA leads to a reduction in the number of samples that need to be analysed using an IB, thus also reducing the costs. Two alternative workflows for LB serology including the C6 EIA are suggested.</p

Conserved lysine residues in decorin binding proteins of Borrelia garinii are critical in adhesion to human brain microvascular endothelial cells

Lyme borreliosis is a tick-borne disease caused by Borrelia burgdorferi sensu lato spirochetes (Lyme borreliae). When the disease affects the central nervous system, it is referred to as neuroborreliosis. In Europe, neuroborreliosis is most often caused by Borrelia garinii. Although it is known that in the host Lyme borreliae spread from the tick bite site to distant tissues via the blood vasculature, the adherence of Lyme borreliae to human brain microvascular endothelial cells has not been studied before. Decorin binding proteins are adhesins expressed on Lyme borreliae. They mediate the adhesion of Lyme borreliae to decorin and biglycan, and the lysine residues located in the binding site of decorin binding proteins are important to the binding activity. In this study, we show that lysine residues located in the canonical binding site can also be found in decorin binding proteins of Borrelia garinii, and that these lysines contribute to biglycan and decorin binding. Most importantly, we show that the lysine residues are crucial for the binding of Lyme borreliae to decorin and biglycan expressing human brain microvascular endothelial cells, which in turn suggests that they are involved in the pathogenesis of neuroborreliosis.</p