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swiezy artykul citrosept

29.06.07, 13:08
Witam wszystkich uczestnikow Forum, dobrze, ze jestescie.
Znalazlam ciekawy artykul:

Infection. 2007 Jun;35(3):206-8. Related Articles, Links


Grapefruit Seed Extract is a Powerful in vitro Agent Against Motile and
Cystic Forms of Borrelia burgdorferi sensu lato.

Brorson O, Brorson SH.

Dept. of Microbiology, Sentralsykehus i Vestfold HF, Tønsberg, Norway.

PMID: 17565468 [PubMed - in process]

zdaje sie, ze jest to czasopismo niemieckie. czy moze ma ktos dostep do tego
artykulu, moze Artur?
Obserwuj wątek
    • franiolek1 Re: swiezy artykul citrosept 29.06.07, 13:44
      Oto caly artykul, nie moge dojsc do linku - znalazlam go chyba w pubmedzie

      Infection. 2007 Jun;35(3):206-8.
      Grapefruit Seed Extract is a Powerful in vitro Agent Against Motile and Cystic
      Forms of Borrelia burgdorferi sensu lato.
      Brorson O, Brorson SH.
      Dept. of Microbiology, Sentralsykehus i Vestfold HF, Tønsberg, Norway.

      PMID: 17565468 [PubMed - in process]



      Lyme borreliosis [1], caused by Borrelia burgdorferi sensu
      lato, may lead to long-term tissue infection, which may be
      difficult to cure.

      The outcome of Lyme borreliosis is highly
      dependent on the antibiotic treatment [2]. The observation
      of the ability of B. burgdorferi sensu lato to convert (and
      reconvert) to cystic forms [3–5] may explain why the infection
      sometimes is persistent and reactivating.

      Therefore, it
      might be important to eradicate all germative forms (not
      only the motile form) of the bacterium to obtain a proper
      treatment for Lyme borreliosis.

      Grapefruit-seed extract
      (GSE) contains bioactive flavenoids (e.g., hesperitin, resveratrol,
      and naringenin) and has been shown to possess
      anti-microbiological effect against bacteria and fungus [6,
      7].

      Many studies indicate that GSE is a substance whose
      therapeutic effect ranks equal to or better than other
      known anti-bacterial agents. Positive effects of GSE are decreased
      levels of TNF-£\, Nuclear factor Kb, NO, protection
      of the gastrointestinal tract against mechanical stress, and
      has anti-allergic and other antioxidative properties [8, 9].

      Naringenin, hesperidin and other citrus flavones have been
      found in plasma and tissue after ingestion [10]. Lactobacillus
      and bifidobacteria in the gut seems to be insignificantly
      affected by GSE [6], and no severe side effects have been
      observed. B. burgdorferi sensu lato has a gene for efflux
      mechanism which may be responsible for antibiotic resistance
      [11].

      GSE is an efflux inhibitor, which can be used
      to enhance the activity of antibacterial agents [12]. For
      the reasons mentioned above it is reasonable to test the
      hypothesis that motile and cystic forms of B. burgdorferi
      sensu lato will be susceptible to GSE, and this is the aim
      of our study.

      The bacterial strain used in our experiments was
      B. afzelii ACA-1. Production of mobile spirochetes and
      cystic forms was performed according to our previous procedure
      [13].

      Grape fruit seed extract 33% (Citrosept; Cintamani
      Europe AS, 2071 Råholt, Norway) was diluted in distilled
      water, sterile filtered by a 0.2 µm filter, and diluted geometrically
      in 5 ml Nalgene tubes from 0.33%–0.00064% in 2 ml
      of diluted BSK-H medium (dilution 1:100 in distilled water).

      The control was diluted BSK-H. Two ml suspension of
      cystic forms at an age of 1 h was added to each of the tubes
      giving a final GSE concentration of 0.165%–0.00032%.

      Susceptibility testing of mobile spirochetes to GSE
      was performed in a final dilution of GSE from 0.165% to
      0.00032% in BSK-H medium. Forty microliter of 107/ml
      bacteria in logarithmic growth was added, making the final
      volume 4 ml in each tube. One control with only BSK-H
      was used.

      To examine if GSE could prevent the conversion
      of mobile spirochetes to cystic forms, testing was also performed
      in distilled water for 1 h at 34 °C. One control with
      only distilled water was used. Motile bacteria in distilled
      water and BSK-H medium were incubated aerobically.

      The tubes with the mobile borrelia in BSK-H medium
      and the cysts in diluted BSK-H were examined by Dark
      Field Microscopy (DFM) (400°—wink after 1 h and 7 days to
      detect presence of eventual mobile spirochetes and intact
      cysts.

      Bacteria exposed to GSE in water were examined by
      DFM at 400°— to examine the ratio of cyst/bacteria. Vital
      staining was performed on bacteria exposed to GSE for
      1 week by mixing 10 µl of Live/dead BacLight™ bacterial
      viability kit (Molecular Probes L-13152 Eugene, OR, USA)
      with 10 µl of the culture. This mixture was placed on a glass
      slide protected with a coverslip. The BacLight-stained bacteria
      were examined by UV-microscopy (800°—wink.
      Infection 2007; 35: 206–208

      Infection 35 · 2007 · No. 3 © URBAN & VOGEL 207
      The following cultures of spirochetes and GSE were
      examined by transmission electron microscopy (TEM)
      as earlier described [13]:

      – motile spirochetes incubated for 1 week with GSE at
      a dilution of 0.0052%, 0.0026%, 0.0013% and a control
      without GSE in BSK-H medium,

      – motile spirochetes incubated for 1 h with GSE at a
      dilution of 0.165%, 0.0825%, 0.0413%, 0.01% and a
      control without GSE in BSK-H medium,

      – motile spirochetes incubated for 1 h with GSE at a dilution
      at 0.0413%, 0.0052%, 0.0013%, 0.00064% and a
      control without GSE in distilled water, and

      – 1 h old cysts incubated for 1 h with GSE at a dilution
      of 0.021%, 0.01%, 0.0052%,
      0.0013%, 0.00064% and a control without GSE.
      GSE-exposed cultures were recultivated in BSK-H
      medium as earlier described
      [13] to confirm or invalidate
      the existence of viable bacteria.

      MBC of the mobile
      spirochetes was determined by
      recultivation of GSE-ex posed
      spirochetes, and the lowest
      GSE concentration where no
      growth occurred was set as the
      MBC value.

      The MIC value for
      mobile spirochetes was determined
      according to the lowest
      GSE concentration, which
      gave reduced multiplication
      when examined in DFM.


      When the susceptibility testing for
      mobile spirochetes was performed in
      distilled water, the rate of conversion
      was strongly dependent on the GSE
      concentration.

      After incubation for
      1 h at 34 °C the number of spirochetes
      converted to cysts ranged from none at
      GSE concentration of 0.165%–0.0052%,
      10% at 0.0028%, 20% at 0.0013%, 95%
      at 0.00064%, and > 95% in the control
      when examined in DFM. By TEM, the
      dilution of 0.0013% showed a very few
      cysts; the dilution of 0.00064% showed
      many normal cysts but not as many as in
      the control.

      Susceptibility testing of normal mobile
      borrelia exposed to GSE at 34 °C
      for 1 h revealed motile bacteria at concentrations
      ? 0.01%. After 5 weeks of
      incubation in fresh BSK-H medium,
      motile spirochetes were observed only
      at the dilutions ? 0.021%. By TEM some
      bacteria with normal appearance (compared
      to the control) were observed in the concentration
      of 0.041%, which is set to be the MBC (Figure 1).

      When
      the mobile spirochetes were exposed to GSE for 1 week
      at 34 °C in fresh BSK-H medium the estimated MBC was
      0.0052% and MIC was 0.00032%. Four weeks of cultivation
      revealed 107 bacteria/ml in the 0.0026% dilution.

      However, BacLight™ showed green structures (green color
      indicates living organisms) only from the 0.0013% dilution.
      This corresponded well with results obtained by TEM.
      Rupturing was observed by TEM and DFM for
      100% of the 1 h old cysts which had been incubated in
      GSE from 0.165%–0.021%; for GSE-dilutions from
      0.01%–0.00064% rupturing was observed for 90%–5%


      Figure 1. (a) Spirochetes incubated for 1 h at 34 °C with 0.165% GSE diluted in
      BSK-H medium.
      Only a very few pycnotic bacteria were present. Most bacteria were completely
      dissolved.
      (b) Spirochetes exposed to 0.041% GSE. The bacteria have a normal
      ultrastructure.
      TEM. Bar = 500 nm.


      Figure 2. (a) One hour old cysts incubated for 1 h at 34 °C in BSK-H medium
      with 0.0013% GSE. A few
      normal and some dissolved cysts were present. (b) The same cysts as in A, but
      exposed to 0.00064%
      GSE. The number of normal cysts present was approximately the same as in the
      control. (c) The cysts
      in B were transferred into fresh BSK-H medium and incubated for 5 weeks in 34 °
      C. Many normal spirochetes
      were present. TEM. Bar = 1,000 nm.


      (most rupturing for the less diluted GSE). For the negative
      control > 98% cysts occurred intact. When transferred
      to
      • franiolek1 ciag dalszy 29.06.07, 13:45



        (most rupturing for the less diluted GSE). For the negative
        control > 98% cysts occurred intact. When transferred
        to BSK-H medium, motile bacteria were observed
        after following incubation time: 14 days for the control
        and GSE dilution 0.00032%; 5 weeks for the 0.00064%
        dilution; no re-growth for higher concentrations (Figure
        2). Therefore, the MBC was calculated to 0.0013%.

        The highest GSE concentrations made the bacteria
        and cysts disappear completely, leaving only small uncharacteristic
        fragments; at lower GSE-levels the membranes
        showed herniation and disruption, and the contents had
        leaked out.

        The MBC was strongly dependent on the length
        of the incubation. GSE was very active even for very short
        incubation times, in agreement with previous results [7].

        The MBC obtained by DFM for the motile bacteria agreed
        well with the TEM results. Presence of GSE reduced the
        conversion from spirochetes to cysts when the susceptibility
        testing was performed in distilled water. This study was
        performed in vitro and further studies are needed to demonstrate
        eventual effects in vivo. From our results it will be
        rational to test the hypothesis that a combination of GSE
        and antibiotics will be efficient in the treatment of resistant
        Lyme borreliosis.

        Ø. Brorson, S.-H. Brorson

        References
        1. Hengge UR, Tannapfel A, Tyring SK, Erbel R, Arendt G, Ruzicka T:
        Lyme borreliosis. Lancet Infect Dis 2003; 3: 489–500.

        2. Petrovic M, Vogelaers D, Van Renterghem L, Carton D,
        de Reuck J, Afschrift M: Lyme borreliosis — a review of the
        late stage and treatment of four cases. Acta Clin Belg 1998;
        53: 178–183.

        3. Gruntar I, Malovrh T, Murgia R, Cinco M: Conversion of Borrelia
        garinii cystic forms to motile spirochetes in vivo. APMIS 2001;
        109: 383–388.

        4. Hulínská D, Barták P, Hercogová J, Hancil J, Basta J, Schramlová J:
        Electron microscopy of Langerhans cells and Borrelia burgdorferi
        in Lyme disease patients. Zbl Bakt 1994; 280: 348–359.

        5. Preac Mursic V, Wanner G, Reinhardt S, Wilske B, Busch U,
        Marget W: Formation and cultivation of Borrelia burgdorferi
        spheroplast L-form variants. Infection 1996; 24: 218–225.

        6. Ionescu G, Kiehl R, Wichmann-Kunz F, Williams CH, Bauml L,
        Levine S: Oral citrus seed extract in atopic eczema: in vitro and
        in vivo studies on intestinal microflora. J Orthomolec Med 1990;
        5: 155–157.

        7. Heggers JP, Cottingham J, Gusman J et al: The effectiveness of
        processed grapefruit-seed extract as an antibacterial agent: II.
        Mechanism of action and in vitro toxicity. J Altern Complement
        Med 2002; 8: 333–340.

        8. Zayachkivska OS, Konturek SJ, Drozdowich D, Konturek PC, Brzozowski
        T, Ghegotsky MR: Gastroprotective effects of flavenoids
        in plant extracts. J Physiol Pharmacol 2005; 56(Suppl 1): 219–231.

        9. Tsai SH, Lin-Shiau SY, Lin JK: Suppression of nitric oxide synthase
        and the down-regulation of the activation of NF B in macrophages

        • franiolek1 Re: ciag dalszy 29.06.07, 13:49
          To dzieki badaniom Brorsonow wiemy, ze cysty borrelii mozna rozbic
          metronidazolem i plaquenilem.
          Tu jest link, ale do artykulow po francusku
          www.lymeinfo.net/francais/traitement_kystes.pdf

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