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Немецкие анализы на боррелиоз и сопутствующие инфекции
Современные лабораторные анализы на боррелиоз и ко-инфекции в немецкой лаборатории ArminLabs

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Морфологические и биохимические особенности Боррелии и ее форм

Тема в разделе "Обсуждения публикаций", создана пользователем Niko, 26 ноя 2015.

  1. claraS
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    claraS Уважаемый форумчанин

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    о своём опыте борьбы с бруцеллёзом написала в личку.
     
  2. Julia Göteborg
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    Julia Göteborg Ветеран форума

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    Небольшая зарисовка к ближайшей дискуссии Есть те, которые лечатся травами , брт, бвт и правильным питанием, чтобы не навредить иммунитету
    (и не только к ней, разумеется, и ранее было много подобных, например почти все отсюда Дебаты )

    Извините, но сделать небольшую зарисовку короче у меня не получилось.. :rolleyes: :)

    Resurgence of Persisting Non-Cultivable Borrelia burgdorferi following Antibiotic Treatment in Mice


    Introduction

    The agent of Lyme borreliosis, Borrelia burgdorferi, evades host immunity and establishes persistent infections in its varied mammalian hosts. This persistent biology may pose challenges to effective antibiotic treatment. Experimental studies in dogs, mice, and non-human primates have found persistence of B. burgdorferi DNA following treatment with a variety of antibiotics, but persisting spirochetes are non-cultivable. Persistence of B. burgdorferi DNA has been documented in humans following treatment, but the significance remains unknown. The present study utilized a ceftriaxone treatment regimen in the C3H mouse model that resulted in persistence of non-cultivable B. burgdorferi in order to determine their long-term fate, and to examine their effects on the host. Results confirmed previous studies, in which B. burgdorferi could not be cultured from tissues, but low copy numbers of B. burgdorferi flaB DNA were detectable in tissues at 2, 4 and 8 months after completion of treatment, and the rate of PCR-positive tissues appeared to progressively decline over time. However, there was resurgence of spirochete flaB DNA in multiple tissues at 12 months, with flaB DNA copy levels nearly equivalent to those found in saline-treated mice. Despite the continued non-cultivable state, RNA transcription of multiple B. burgdorferi genes was detected in host tissues, flaB DNA was acquired by xenodiagnostic ticks, and spirochetal forms could be visualized within ticks and mouse tissues by immunofluorescence and immunohistochemistry, respectively. A number of host cytokines were up- or down-regulated in tissues of both saline- and antibiotic-treated mice in the absence of histopathology, indicating host response to the presence of non-cultivable, despite the lack of inflammation in tissues.

    ...
    Non-cultivable B. burgdorferi Resurge in Tissues of Mice at 12 Months Following Antibiotic Treatment

    A number of studies by different research groups have shown that C3H mice infected by either syringe or tick, and treated with doxycycline, tigecycline, or ceftriaxone contained persisting non-cultivable B. burgdorferi in tissues for at least 90 days after completion of treatment [14], [15], [16], [17], [18], [19], [20]. Others [28] have speculated that the persisting non-cultivable spirochetes did not induce inflammation and would eventually die out following treatment. Thus, a ceftriaxone treatment regimen was used to treat C3H mice similar to previous studies that consistently resulted in persistence of non-cultivable B. burgdorferi following treatment [14], [15], [16], [17] to determine the long-term outcome of treatment, including persistence or demise of non-cultivable spirochetes, and returning of cultivability.

    ...
    Discussion

    This study validates a mouse model that can be used for investigation of post-antibiotic persistence of non-cultivable B. burgdorferi. We utilized syringe-inoculation of cultured B. burgdorferi, which is not the natural route of infection, but others [15], [16] have shown persistence of non-cultivable B. burgdorferi with tick-borne infection as well...

    ..Furthermore, the short serum half-life of ceftriaxone in mice as an explanation for persistence of non-cultivable B. burgdorferi was negated with a study that utilized tigecycline [14], which has a half-life of 11.6 hours in mice and achieved a serum maximum level 70 times greater than the B. burgdorferi N40 MBC [43]. Treatment with ceftriaxone, and high or low doses of tigecycline resulted in the same outcome: persistence of non-cultivable B. burgdorferi in tissues of treated mice at 90 days after completion of treatment [14].

    Results of this study demonstrated not only persistence, but also resurgence of non-cultivable B. burgdorferi in tissues of mice at up to 12 months following antibiotic treatment, despite the continued inability to culture spirochetes from tissues. In the absence of the ability to culture spirochetes from tissues of treated mice, this study endeavored to determine the physiologic state of persisting non-cultivable B. burgdorferi. Non-cultivable B. burgdorferi DNA acquisition was demonstrated by xenodiagnosis (including transstadial persistence from larva to nymph), B. burgdorferi spirochetal forms were visualized by immunofluorescence in xenodiagnostic ticks, RNA transcription of multiple B. burgdorferi-specific genes was detected in the tissues of antibiotic-treated mice at 12 months after antibiotic treatment, and spirochetal forms were visualized in the tissue of an antibiotic-treated mouse by immunohistochemistry. It has been speculated that following antibiotic treatment, persisting non-cultivable spirochetes were non-viable and would eventually be eliminated [15], [28], [44]. Results of the current study suggested a similar trend if the experiment were to be stopped at 8 months, as the prevalence of PCR-positive tissue sites declined with time through 8 months. However, there was resurgence of B. burgdorferi DNA in tissues at 12 months to levels nearly equivalent to those in untreated persistently infected mice. These findings underscore the difficulty of detecting non-cultivable B. burgdorferi following antibiotic treatment, as interpretation of xenodiagnosis depends upon timing and number of ticks tested, and the outcome of infection must be measured by analysis of multiple tissues at appropriate intervals. Non-cultivability of persisting B. burgdorferi is likely a reflection of their limited replication kinetics. In a recent study involving antibiotic-treated macaques, small numbers of spirochetes were found in culture after several weeks of incubation [22], and others have demonstrated slow outgrowth of spirochetes in cultures from antibiotic-treated humans after prolonged incubation of up to 10.5 months [45]. In the present study, we attempted to incubate cultures for several weeks, as in the macaque study, but were unable to grow viable spirochetes.

    The mechanism for resurgence of non-cultivable B. burgdorferi at 12 months after treatment remains to be determined, but may be related to declining antibody response. Antibody is critical for resolution of arthritis and carditis, as well as global reduction of spirochete burdens in tissues [40], [46], [47]. Disease resolution can be maintained and reduced tissue burdens can be sustained in infected severe combined immunodeficient mice by passive immunization with immune serum from persistently infected immunocompetent mice, but disease recurs and resurgence of spirochete burdens occurs as the passive antibody decays [48]. The antigenic target for maintaining reduced tissue burdens has not been determined, so cause and effect in the current study could not be confirmed. Antibody titers against several in vivo-expressed N40 recombinant proteins (DbpA, OspC, Arp, BmpA) [49], similar to antibody against B. burgdorferi lysates, were also diminished at all intervals following antibiotic treatment (data not shown). In a related study, recrudescence of non-cultivable B. burgdorferi following antibiotic treatment was attempted by transient immunosuppression of the mice with corticosteroids without apparent consequence [15], but that outcome would be expected when spirochete burdens in tissues are under control of antibody.

    The current study builds upon similar evidence of non-cultivable B. burgdorferi persistence in studies involving dogs, mice and macaques. While it has been concluded that non-cultivable B. burgdorferi is not infectious [16], we previously demonstrated transmission of B. burgdorferi DNA by ticks that fed upon antibiotic-treated mice to naïve mice, as well as transmission of B. burgdorferi DNA by transplantation of B. burgdorferi DNA-positive heart base and tibiotarsus tissue allografts from treated mice into recipient mice, with dissemination of the DNA in the recipient mice [14]. Transtadial transmission of B. burgdorferi DNA from larvae that fed upon treated mice to nymphs and then to adults was also demonstrated [14]. RNA transcription of a limited number of B. burgdorferi genes by persisting non-cultivable spirochetes was shown after antibiotic treatment of both mice [14] and macaques [22], but analyses were limited to only a few target genes. In the current study, the LDA approach allowed simultaneous analysis of multiple gene targets, indicating more global transcription activity of persisting non-cultivable B. burgdorferi at 12 months. Added to earlier studies that demonstrated persistence of B. burgdorferi DNA in tissues of antibiotic-treated dogs [9], [21], [50], the collective conclusion is persistence of non-cultivable spirochetes following treatment of mice, macaques and dogs with various antibiotics. The limited infectivity of persisting non-cultivable B. burgdorferi has no biological significance for the natural enzootic cycle of B. burgdorferi, but it remains to be determined if they have significance in the post-treatment host. Thus, validation of a reproducible animal model for study of this phenomenon is important.

    It has been suggested that persisting non-cultivable B. burgdorferi may be irrelevant, in that they are non-pathogenic and do not cause disease [28]. This is difficult to prove, since in the normal course of events, fully virulent B. burgdorferi infects multiple tissues without inflammation. Experimental animal studies have shown that arthritis and carditis are more severe in animals that are genetically susceptible, immature or immunodeficient, or in animals inoculated with high doses of spirochetes that can replicate to high enough titer to colonize specific tissue sites before immune-mediated disease remission [30]. Inflammation is related to colonization of specific tissue sites that are prone to inflammation, including synovium and vascular media, resulting in acute arthritis (synovitis), arteritis and carditis. The host antibody response eliminates or prevents colonization of these sites, so that the normal course of infection in adult, immunocompetent animals is minimal or none. The lack of inflammation, therefore, in tissues of antibiotic-treated mice at 12 months, with resurgence of spirochete DNA burdens equivalent to those found in saline-treated mice, does not prove the argument one way or the other, since there is no or minimal inflammation in tissues of infected saline-treated mice at 12 months with similar or greater spirochete burdens. The only histologic evidence of host response in both groups of mice was segmental lymphoplasmacytic infiltrates in plantar artery adventitia and sparse infiltrates in the epicardium, albeit less obvious in the antibiotic-treated mice.

    The lack of morphological evidence of inflammation under normal circumstances, as well as after antibiotic treatment, prompted us to evaluate host response through LDA analysis of host cytokines. Avoiding over-interpretation of specific cytokine responses, our results indicated that mice were responding to the presence of persisting non-cultivable B. burgdorferi. The LDA approach allows simultaneous analysis of multiple host cytokines relative to age-matched uninfected mouse tissues, but does not measure absolute values. It is therefore useful for visualizing a “fingerprint” of host response, indicating a pro-inflammatory cytokine state in both persistently infected, saline-treated and antibiotic-treated mice at 12 months after treatment. Recently, it has been shown that spirochete antigens persist in tissues of antibiotic-treated mice, which may be responsible for cytokine activation [16]. The presence of persisting, non-cultivable B. burgdorferi that transcribe RNA and express antigens in tissues may contribute to this phenomenon.
    ...
    Resurgence of Persisting Non-Cultivable Borrelia burgdorferi following Antibiotic Treatment in Mice

     
  3. Julia Göteborg
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    Julia Göteborg Ветеран форума

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    И еще.

    Плазми́ды
    — небольшие молекулы ДНК, физически отдельные от геномных хромосом и способные реплицироваться автономно. Как правило, плазмиды встречаются у бактерий и представляют собой двухцепочечные кольцевые молекулы, но изредка плазмиды встречаются также у архей и эукариот.

    В природе плазмиды обычно содержат гены, повышающие устойчивость бактерии к неблагоприятным внешним факторам (в т. ч. устойчивость к антибиотикам), нередко они могут передаваться от одной бактерии к другой (иногда даже к бактерии другого вида) и, таким образом, служат средством горизонтального переноса генов.

    Попадание плазмиды в клетку может осуществляться двумя путями: либо при непосредственном контакте клетки-хозяина с другой клеткой в процессе конъюгации, либо путём трансформации, то есть искусственного введения в клетку плазмиды, которому предшествует изменение экспрессии определённого гена клетки-хозяина (приобретение клеткой компетентности).

    Искусственные плазмиды используются как векторы в клонировании ДНК, причём благодаря их способности к репликации обеспечивается возможность репликации рекомбинантной ДНК в клетке-хозяине.

    ..Количество плазмид на одну клетку также варьирует. Если у некоторых плазмид число копий на клетку составляет одну или несколько, то у других, главным образом мелких, плазмид их число может доходить до десятков и даже сотен. Численность плазмид зависит от характера их репликации[3]. Плазмиды способны реплицироваться независимо от хромосомы, однако они могут находиться под строгим или ослабленным контролем от неё...

    Плазмида Borrelia burgdorferi - lp25
    геометрия плазмиды – линейная
    размер плазмиды (тыс. пар оснований) - 24,2
    число копий плазмиды на клетку – 1-2,
    (см.таблицу по ссылке)

    ...
    Репликация плазмид


    Как отмечалось выше, важным свойством плазмид является их способность к автономной репликации, в той или иной мере проходящей под контролем хромосомы. Этот контроль обусловлен тем, что плазмиды содержат не все необходимые для репликации гены, и поэтому в их репликации также принимают участие некоторые ферменты клетки[5]. Механизмы этого контроля подробнее освещены ниже...

    И очень-очень "интересная" информация..:

    Использование

    Плазмиды широко используются в генной инженерии для переноса генетической информации и генетических манипуляций. Для этого создаются искусственные плазмиды — векторы, состоящие из частей, взятых из разных генетических источников, а также из искусственно созданных фрагментов ДНК. :unsure: :nailbiting:

    Плазмиды — Википедия
     
    maya нравится это.
  4. Julia Göteborg
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    Julia Göteborg Ветеран форума

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    Ну и, естественно, у боррелии не одна плазмида..


    Plasmid analysis of Borrelia burgdorferi, the Lyme disease agent.

    A simple procedure for extraction of plasmid-enriched DNA from borreliae was used in a plasmid analysis of 13 strains of the Lyme disease agent, Borrelia burgdorferi. The extracted DNA was subjected to low-percentage agarose gel electrophoresis and examined either directly by ethidium bromide staining or after hybridization of the plasmids in situ with a DNA probe for the gene encoding the major outer membrane protein OspA. Each isolate had four to seven discernible plasmids of various sizes. Only 2 of the 13 strains had the same plasmid profile. The ospA gene probe hybridized to large plasmids to strains from both North America and Europe. A strain which had been passaged many times was found to have lost two of the six plasmids originally present. These findings indicate the potential usefulness of plasmid analysis as a strain-typing procedure and for identifying possible plasmid-conferred virulence factors.
    Plasmid analysis of Borrelia burgdorferi, the Lyme disease agent.

    Linear plasmids of the bacterium Borrelia burgdorferi have covalently closed ends

    The genetics of spirochetes, a division of eubacteria, has been little studied. Double-stranded linear plasmids were found in Borrelia burgdorferi, the agent of Lyme disease. A 49-kilobase linear plasmid contained the ospA and ospB genes, which encode the major outer membrane proteins of strain B31. Molecules of the 49-kilobase plasmid rapidly reannealed after alkaline denaturation; rapid renaturation was prevented if the 49-kilobase plasmids were first treated with S1 nuclease. When denatured plasmid molecules were examined directly, single-stranded circles of approximately 100-kilobase circumference were seen. These studies provide direct visual evidence that the linear plasmids have covalently closed ends. This form of DNA occurs in some animal viruses, but it has not heretofore been described in prokaryotic organisms.
    Linear plasmids of the bacterium Borrelia burgdorferi have covalently closed ends | Science

    Correlation between plasmid content and infectivity in Borrelia burgdorferi

    Infectivity-associated plasmids were identified in Borrelia burgdorferi B31 by using PCR to detect each of the plasmids in a panel of 19 clonal isolates. The clones exhibited high-, low-, and intermediate-infectivity phenotypes based on their frequency of isolation from needle-inoculated C3H/HeN mice. Presence or absence of 21 of the 22 plasmids was determined in each of the clones by using PCR primers specific for regions unique to each plasmid, as identified in the recently available genome sequence. Southern blot hybridization results were used to confirm the PCR results in some cases. Plasmid lp25 exhibited a direct correlation with infectivity in that it was consistently present in all clones of high or intermediate infectivity and was absent in all low-infectivity clones. lp28–1, containing the vmp-like sequence locus, also correlated with infectivity; all clones that lacked lp28–1 but contained lp25 had an intermediate infectivity phenotype, in which infection was primarily restricted to the joints. Plasmids cp9, cp32–3, lp21, lp28–2, lp28–4, and lp56 apparently are not required for infection in this model, because clones lacking these plasmids exhibited a high-infectivity phenotype. Plasmids cp26, cp32–1, cp32–2 and/or cp32–7, cp32–4, cp32–6, cp32–8, cp32–9, lp17, lp28–3, lp36, lp38, and lp54 were consistently present in all clones examined. On the basis of these results, lp25 and lp28–1 appear to encode virulence factors important in the pathogenesis of B. burgdorferi B31.
    Correlation between plasmid content and infectivity in Borrelia burgdorferi

    Linear and Circular Plasmid Content in Borrelia burgdorferi Clinical Isolates

    The genome of Borrelia burgdorferi, the etiologic agent of Lyme disease, is composed of a linear chromosome and more than 20 linear and circular plasmids. Typically, plasmid content analysis has been carried out by pulsed-field gel electrophoresis and confirmed by Southern hybridization. However, multiple plasmids of virtually identical sizes (e.g., lp28 and cp32) complicate the interpretation of such data. The present study was undertaken to investigate the complete plasmid complements of B. burgdorferi clinical isolates cultivated from patients from a single region where early Lyme disease is endemic. A total of 21 isolates obtained from the skin biopsy or blood samples of Lyme disease patients were examined for their complete plasmid complements by Southern hybridization and plasmid-specific PCR analysis. All clinical isolates harbored at least six of the nine previously characterized cp32s. Fourteen isolates harbored all B31-like linear plasmids, and seven isolates simultaneously lacked lp56, lp38, and some segments of lp28-1. The distinctive plasmid profile observed in these seven isolates was specific to organisms that had ribosomal spacer type 2 and pulsed-field gel type A, which implies a clonal origin for this genotype. The presence of nearly identical complements of multiple linear and circular plasmids in all of the human isolates suggests that these plasmids may be particularly necessary for infection, adaptation, and/or maintenance in the infected host.
    Linear and Circular Plasmid Content in Borrelia burgdorferi Clinical Isolates


    Genome Stability of Lyme Disease Spirochetes: Comparative Genomics of Borrelia burgdorferi Plasmids


    Lyme disease is the most common tick-borne human illness in North America. In order to understand the molecular pathogenesis, natural diversity, population structure and epizootic spread of the North American Lyme agent, Borrelia burgdorferi sensu stricto, a much better understanding of the natural diversity of its genome will be required. Towards this end we present a comparative analysis of the nucleotide sequences of the numerous plasmids of B. burgdorferi isolates B31, N40, JD1 and 297. These strains were chosen because they include the three most commonly studied laboratory strains, and because they represent different major genetic lineages and so are informative regarding the genetic diversity and evolution of this organism. A unique feature of Borrelia genomes is that they carry a large number of linear and circular plasmids, and this work shows that strains N40, JD1, 297 and B31 carry related but non-identical sets of 16, 20, 19 and 21 plasmids, respectively, that comprise 33–40% of their genomes. We deduce that there are at least 28 plasmid compatibility types among the four strains. The B. burgdorferi ∼900 Kbp linear chromosomes are evolutionarily exceptionally stable, except for a short ≤20 Kbp plasmid-like section at the right end. A few of the plasmids, including the linear lp54 and circular cp26, are also very stable. We show here that the other plasmids, especially the linear ones, are considerably more variable. Nearly all of the linear plasmids have undergone one or more substantial inter-plasmid rearrangements since their last common ancestor. In spite of these rearrangements and differences in plasmid contents, the overall gene complement of the different isolates has remained relatively constant.
    Genome Stability of Lyme Disease Spirochetes: Comparative Genomics of Borrelia burgdorferi Plasmids


    Genomic sequence of a Lyme disease spirochaete, Borrelia burgdorferi


    The genome of the bacterium Borrelia burgdorferi B31, the aetiologic agent of Lyme disease, contains a linear chromosome of 910,725 base pairs and at least 17 linear and circular plasmids with a combined size of more than 533,000 base pairs. The chromosome contains 853 genes encoding a basic set of proteins for DNA replication, transcription, translation, solute transport and energy metabolism, but, like Mycoplasma genitalium, it contains no genes for cellular biosynthetic reactions. Because B. burgdorferi and M. genitalium are distantly related eubacteria, we suggest that their limited metabolic capacities reflect convergent evolution by gene loss from more metabolically competent progenitors. Of 430 genes on 11 plasmids, most have no known biological function; 39% of plasmid genes are paralogues that form 47 gene families. The biological significance of the multiple plasmid-encoded genes is not clear, although they may be involved in antigenic variation or immune evasion.
    Genomic sequence of a Lyme disease spirochaete, : Borrelia burgdorferi : Article : Nature
     
    maya нравится это.
  5. Melik
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    Melik Известный человек

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    здравствуйте; не могли бы вы и со мной поделиться своим опытом. спасибо
     
  6. Lana
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    Lana Ветеран форума

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    Давайте я поделюсь информацией, что говорила Клара: бруцеллез лечат либо риф плюс докси (бактериастатическая схема), либо стрептомицин плюс левомицитин (эта схема убивает инфекцию). Если начнете лечиться или раньше лечились одним из этих АБ - через 3 дня возникнет резистентность и это Аб станет бесполезен. Надо лечиться двумя АБ одновременно.


    Лечение длительное. Лечится хуже чем лайм по-моему еще.
     
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