Onderzoek persister cells

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VerlorengezondheidM
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Lid geworden op: Za 19 Sep 2015, 23:59
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Re: Onderzoek persister cells

Berichtdoor VerlorengezondheidM » Ma 15 Jul 2019, 07:30

Henriëtte schreef:In een nieuwe publicatie van Rudenko en collega’s over persistentie wordt uitgebreid besproken hoe de Borrelia bacterie onder stressvolle omstandigheden verandert in persistente vormen. 14-06-2019

Persisters kunnen verschillende groottes en vormen aannemen, waarbij ze van de bekende beweeglijke spirocheet vorm veranderen naar ronde vormen, L-vormen of biofilmachtige microkolonies. Deze persistente vormen van Borrelia blijven ondanks agressieve antibioticabehandeling levensvatbaar en kunnen weer worden omgezet in de beweeglijke spirocheetvorm in een gunstige groeiomgeving


https://lymevereniging.nl/2019/06/14/me ... irocheten/

Henriëtte


Kunnen stressvolle omstandigheden ook opgelopen co-infecties zijn (met een synergetische werking op Borrelia) al dan niet veroorzaakt door tekenbeten; of is dat soms die ongemakkelijke waarheid?

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Chronic Lyme disease is due to the persistence of Borrelia within tissues but is also probably due to other micro-organisms responsible for co-infections.”

— Dr. Christian Perronne

Bron: Huffingtonpost, lees meer
Lyme Test Negative?
It Still May Be Lyme Disease!

Na 19 jaar onbehandeld rondlopen met een klinisch ziektebeeld: Chronische Lyme, Q-koorts, Bartonella, Babesia, Anaplasma, Chlamydia, Mycoplasma, Rickettsia, Yersinia, Coxsackie A16+ B, HHV-6.

Henriëtte
Berichten: 1665
Lid geworden op: Di 19 Mei 2015, 10:05

Re: Onderzoek persister cells

Berichtdoor Henriëtte » Vr 19 Jul 2019, 07:06

Lymevereniging

In muizen is in wetenschappelijk onderzoek weer uiterst zorgvuldig aangetoond dat Lyme-bacteriën 18 maanden antibiotica kunnen overleven. Wij mensen zijn helemaal klaar met de tunnelvisie dat Lyme na een maand antibiotica genezen zou zijn. Deze tunnelvisie veroorzaakt wereldwijd onnoemelijk veel leed. Het is de hoogste tijd voor een open mind!


The Generality of Post-Antimicrobial Treatment Persistence of Borrelia burgdorferi Strains N40 and B31 in Genetically Susceptible and Resistant Mouse Strains;

https://www.ncbi.nlm.nih.gov/m/pubmed/31308087/

Henriëtte

Sproetje
Berichten: 3143
Lid geworden op: Di 28 Okt 2014, 20:33

Re: Onderzoek persister cells

Berichtdoor Sproetje » Vr 02 Aug 2019, 21:26

The Functional and Molecular Effects of Doxycycline Treatment on Borrelia burgdorferi Phenotype

John R. Caskey1,2†, Nicole R. Hasenkampf1, Dale S. Martin1, Vladimir N. Chouljenko2, Ramesh Subramanian2, Mercedes A. Cheslock1 and Monica E. Embers1*
2019


https://www.frontiersin.org/articles/10 ... 00690/full

Recent studies have shown that Borrelia burgdorferi can form antibiotic-tolerant persisters in the presence of microbiostatic drugs such as doxycycline. Precisely how this occurs is yet unknown. Our goal was to examine gene transcription by B. burgdorferi following doxycycline treatment in an effort to identify both persister-associated genes and possible targets for antimicrobial intervention. To do so, we performed next-generation RNA sequencing on doxycycline-treated spirochetes and treated spirochetes following regrowth, comparing them to untreated B. burgdorferi. A number of genes were perturbed and most of those which were statistically significant were down-regulated in the treated versus the untreated or treated/re-grown. Genes upregulated in the treated B. burgdorferi included a number of Erp genes and rplU, a 50S ribosomal protein. Among those genes associated with post-treatment regrowth were bba74 (Oms28), bba03, several peptide ABC transporters, ospA, ospB, ospC, dbpA and bba62. Studies are underway to determine if these same genes are perturbed in B. burgdorferi treated with doxycycline in a host environment.


Discussion

The results of the bioassay in mice demonstrate that after treatment with the MBC of doxycycline, the spirochetes were able to host adapt, and evade immune pressure to establish an infection. The number of spirochetes injected into each mouse was held constant, but viability was not assessed. Therefore, the number of actively growing B. burgdorferi was likely significantly reduced after doxycycline treatment. What is most interesting about the result is that no significant differences were observed in the spirochete infectivity when immunocompetent versus immunodeficient mice were used. This result suggests that exit from dormancy may occur very rapidly in vivo, allowing the immunoevasive phenotype to become established.


We have previously demonstrated that the persister cell phenotype appears to be generated stochastically and driven by slowed growth (Caskey and Embers, 2015). In this study, we aimed to identify gene expression patterns associated with the survival and re-growth of B. burgdorferi in the antibiotic environment. While a similar study in which RNASeq was applied to antibiotic treatment of B. burgdorferi was conducted (Feng et al., 2015), several important distinctions between that study and ours should be made. In that report by Feng et al, in vitro-cultured Bb were treated with either doxycycline or amoxicillin (50 μg/mL) for 6 days and then subjected to RNASeq, with comparison to the untreated control. Genes that were up-regulated by 2-fold or more were ascribed significant and the pathways affected were elucidated. In our study, we treated a slightly denser culture (5 × 107 versus 1 × 107) with doxycycline at the same dose (50 μg/ml) for 5 days. An aliquot of the treated cells was allowed to re-grow, such that we had 3 treatment groups. In addition, we performed RNASeq on duplicate samples and ascribed significance using both fold-change and p-values to account for variation between samples. Both studies shed light on the mechanisms that Bb may use to establish persistence and re-grow. The Feng study identified a large number of genes as upregulated following treatment, whereas we found that the vast majority of genes were down-regulated, likely owing to a global decline in transcription. In the Feng study, the ClpP protease was indicated as the most highly up-regulated gene in treated B. burgdorferi. It was up-regulated in our screen as well (0.43-fold), but not determined to be significant. The genes found to be significantly increased in treated versus control encoded several Erp proteins and a 50S ribosomal protein (bb0778). This was verified by standard RT-PCR, as shown in Figure 4. The results in Figure 4 are derived from amplifying transcript from an equal quantity of input RNA. We did not have a good constitutively transcribed gene that is consistently expressed in all groups equally to be used as a housekeeping gene control, based on the RNASeq data. Thus, these results are not fully quantitative but only meant for validation of the RNASeq results

We fully expected that genes categorized by involvement in the stress response (Bugrysheva et al., 2003; Drecktrah et al., 2015; Cabello et al., 2017) and DNA repair mechanisms (Fisher et al., 2017) would be significantly up-regulated with antibiotic treatment. However, this did not appear to be the case with either the Feng study or our own. The mechanisms governing the development of persister cells are not easily discerned by these RNASeq analyses, perhaps because the spirochetes stochastically enter dormancy (Caskey and Embers, 2015) and regrowth is determined by non-heritable traits. One supposition may be that instead, post-transcriptional, or even post-translational events such as lysine acetylation (Fisher et al., 2017; Bontemps-Gallo et al., 2018) govern the entry and exit from dormancy in the antibiotic environment. For example, in the toxin-mediated growth reduction within S. typhimurium, the toxins (TacT) add a post-translational modification (acetylation) to tRNA, which is reversed by a peptidyl tRNA hydrolase (Cheverton et al., 2016). A toxin-antitoxin system for Borrelia persister development has not been identified, but enzymatic modification of translational components could be involved in formation of persister cells and their re-growth. Nonetheless, we have identified a number of surface proteins which may be antigenic and serve as targets for novel immunotherapeutic strategies.
[…..]
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Henriëtte
Berichten: 1665
Lid geworden op: Di 19 Mei 2015, 10:05

Re: Onderzoek persister cells

Berichtdoor Henriëtte » Zo 11 Aug 2019, 23:24

Onderzoek effect Doxycycline op Borrelia;

In het onderzoek bleek dat na de doxycycline behandeling de Borrelia’s in staat zijn zich aan te passen aan een nieuwe gastheer (in dit geval muizen), het afweersysteem te ontwijken en een infectie te veroorzaken. Sommige Borrelia genen werden geactiveerd, andere die te maken hadden met hergroei weer geremd. Er was geen verschil in infectiviteit tussen immuuncompetente en immuundeficiënte muizen, hetgeen zou kunnen betekenen dat het ontwaken uit slaaptoestand en herinfecteren erg snel kan gaan.

https://www.tekenbeetziekten.nl/onderzo ... -borrelia/

The Functional and Molecular Effects of Doxycycline Treatment on Borrelia burgdorferi Phenotype 18-04-2019

Recent studies have shown that Borrelia burgdorferi can form antibiotic-tolerant persisters in the presence of microbiostatic drugs such as doxycycline.


https://www.frontiersin.org/articles/10 ... 00690/full

Henriëtte

Sproetje
Berichten: 3143
Lid geworden op: Di 28 Okt 2014, 20:33

Re: Onderzoek persister cells

Berichtdoor Sproetje » Ma 19 Aug 2019, 17:48

Identification of new drug candidates against Borrelia burgdorferi using high-throughput screening

Venkata Raveendra Pothineni,1 Dhananjay Wagh,1 Mustafeez Mujtaba Babar,1 Mohammed Inayathullah,1 David Solow-Cordero,2 Kwang-Min Kim,1 Aneesh V Samineni,1 Mansi B Parekh,1 Lobat Tayebi,3 Jayakumar Rajadas1
2016

https://www.dovepress.com/identificatio ... ticle-DDDT

Discussion

doxycyclin, was unable to kill B. burgdorferi persisters in vitro.7,21 In addition to this, patients with Lyme who are diagnosed later and treated may develop arthritis. These patients treated later do not respond fully to a first course of antibiotic therapy.21,39
……….

Among the identified drug molecules, erythromycin and kitasamycin appeared to be very active. The drug erythromycin was already used clinically for the treatment of Lyme disease. In some laboratories, strains and clinical isolates of B. burgdorferi show resistance to erythromycin.40
……

One of the other drug molecules, linezolid, was found to possess ~98% inhibition of the stationary-phase Borrelia in the initial screen. Moreover, the drug was found to possess the MIC and MBC values of 0.4 μg/mL and 8.4 μg/mL, respectively.
…..,
Linezolid possesses commendable pharmacokinetic profile with up to 100% bioavailability after oral administration and has a high volume of distribution.48,49 It is efficiently distributed in most of the body tissues, including osteoarticular and central nervous system.50,51
…..
An interesting drug candidate identified in the study was the alcohol dehydrogenase inhibitor, disulfiram. Though the drug is used as a treatment of alcohol abuse, recently its anticancer potential has also been discovered. It has been known to make complexes with metal ions and cause the disruption of the proteasome, leading to death of the cancer cells. Similarly, the metabolites of the drug molecule have been known to significantly inhibit the growth of a number of bacterial species, including the biofilm-forming Pseudomonas aeruginosa.62 The drug molecule has a good bioavailability, and it passes the blood–brain barrier to show its effect in the central nervous system. Though most of the studies are concerned to the use of the drug molecule in alcoholism yet, being a safe, FDA-approved molecule, the drug can be repurposed for its antibacterial potential.

Henriëtte
Berichten: 1665
Lid geworden op: Di 19 Mei 2015, 10:05

Re: Onderzoek persister cells

Berichtdoor Henriëtte » Za 14 Sep 2019, 10:06

Persister cells still a problem for Lyme disease patients;

The presence of persisters may facilitate emergence of antimicrobial-resistant bacteria. “It should be noted that although persistence is phenotypic, the presence of persisters can also facilitate emergence of genetically antimicrobial-resistant bacteria, e.g., by mutation,” according to Cabello


http://danielcameronmd.com/persister-ce ... -patients/

Henriëtte


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