185 research outputs found
Genotypic and Phenotypic Heterogeneity in Alicyclobacillus acidoterrestris: A Contribution to Species Characterization.
Alicyclobacillus acidoterrestris is the main cause of most spoilage problems in fruit juices
and acidic products. Since soil borne species often contaminate fruit juices and do not need
strict extreme requirements for survival, it is a great concern to investigate whether and how
soil species could evolve from their ecological niches in microbial community to new environments as fruit juices. In this study, 23 isolates of thermo-acidophilic, spore-forming bacteria from soil were characterized by cultural and molecular methods. In addition, 2 strains
isolated from a spoilage incident in pear juice were typed. Strains phenotyping showed that
they could be grouped into 3 different clusters, and some isolates showed identical or quite
similar patterns. Analyzing pH and temperature ranges for growth, the majority of strains
were able to grow at values described for many species of Alicyclobacillus. Qualitative utilization of lysine, arginine and indole production from tryptophan revealed, for the first time,
deamination of lysine and decarboxylation of arginine. Resistance to 5% NaCl as well as
the ability to hydrolyze starch and gelatin, nitrate reduction, catalase and oxidase activities
confirmed literature evidences. Examining of 16S rRNA, showed that isolates were divided
into three blocks represented by effectively soil species and strains that are moving from
soil to other possible growing source characterized by parameters that could strongly influence bacterial survival. RAPD PCR technique evidenced a great variability in banding patterns and, although it was not possible to obtain genotypically well-distinguished groups, it
was feasible to appreciate genetic similarity between some strains. In conclusion, the investigation of a microbial community entails a combination of metagenomic and classic culturedependent approaches to expand our knowledge about Alicyclobacillus and to look for new
subspecies
The DNA damage response promotes Polyomavirus JC infection by nucleus to cytoplasm NF-Kappa B activation.
Background: Infection of glial cells by human neurotropic polyomavirus JC (JCV), the causative agent of the CNS
demyelinating disease progressive multifocal leukoencephalopathy (PML), rapidly inflicts damage to cellular DNA.
This activates DNA damage response (DDR) signaling including induction of expression of DNA repair factor Rad51.
We previously reported that Rad51 co-operates with the transcription factor NF-ÎşB p65 to activate JCV early
transcription. Thus Rad51 induction by JCV infection may provide positive feedback for viral activation early in JCV
infection. DDR is also known to stimulate NF-κB activity, a phenomenon known as nucleus to cytoplasm or “insideout” NF-κB signaling, which is initiated by Ataxia telangiectasia mutated (ATM) protein, a serine/threonine kinase
recruited and activated by DNA double-strand breaks. Downstream of ATM, there occurs a series of posttranslational modifications of NF-ÎşB essential modulator (NEMO), the Îł regulatory subunit of inhibitor of NF-ÎşB (IÎşB)
kinase (IKK), resulting in NF-ÎşB activation.
Methods: We analyzed the effects of downstream pathways in the DDR by phosphospecific Western blots and
analysis of the subcellular distribution of NEMO by cell fractionation and immunocytochemistry. The role of DDR in
JCV infection was analyzed using a small molecule inhibitor of ATM (KU-55933). NEMO sumoylation was
investigated by Western and association of ATM and NEMO by immunoprecipitation/Western blots.
Results: We show that JCV infection caused phosphorylation and activation of ATM while KU-55933 inhibited JCV
replication. JCV infection caused a redistribution of NEMO from cytoplasm to nucleus. Co-expression of JCV large Tantigen and FLAG-tagged NEMO showed the occurrence of sumoylation of NEMO, while co-expression of ATM and
FLAG-NEMO demonstrated physical association between ATM and NEMO.
Conclusions: We propose a model where JCV infection induces both overexpression of Rad51 protein and activation
of the nucleus to cytoplasm NF-ÎşB signaling pathway, which then act together to enhance JCV gene expression
Results, questions, perspectives of a study on human polyomavirus BK and molecular actors in prostate cancer development
Background: Prostate cancer (PC) is a common tumor in Western countries. Several risk factors play significant roles. MYC, BIRC5/survivin, CDC25 and P53 may contribute to PC risk. As demonstrated, human Polyomavirus BK (BKV) could affect cellular homeostasis contributing to PC pathogenesis. Materials and Methods: Biological samples were collected from PC patients. Viral
RNA was searched using quantitative polymerase chain reaction (PCR), whereas a qualitative PCR was employed to find particular viral sequences. Proper size amplicons were analyzed. Single nucleotide polymorphisms (SNPs) were detected in p53 coding regions by means of a specific PCR.
C-MYC, BIRC5/survivin and CDC25 gene expression was investigated using a Retro Transcriptional Quantitative PCR. Results: Viral DNA copy number was higher in cancer tissues taken from Gleason score 9 patients with Gleason score 7. Different p53 mutated compared to patients exons
were found according to tumor advanced stage and a statistical significant correlation was found between Gleason score and p53 mutational rate. C-MYC, BIRC5/survivin and CDC25 expression was de-regulated according to the literature. Conclusion: The presence of BKV and its variants in transformed cells does not exclude viral pressure in cell immortalization. Expression of other target genes evidenced a significant change in their regulation, useful for cancer drug discovery and therapies
High frequency of JCV DNA detection in prostate cancer tissues
BACKGROUND: Prostate cancer (PC) represents the most frequently diagnosed cancer in men. Exposure to infectious agents has been considered to induce prostatic inflammation and cancerous transformation. Controversial data exist concerning the role of the human polyomaviruses BK (BKV) and JC (JCV) in PC etiology. Therefore, a possible association between these polyomaviruses and PC was investigated.
MATERIALS AND METHODS: Urine, blood and fresh prostatic tissue specimens were collected from 26 patients with PC. The presence of BKV and JCV, the possible non-coding control region (NCCR) variations and the genotyping analysis of viral protein 1 (VP1) of both viruses were assessed.
RESULTS: Data showed a preferential viral re-activation in the urinary compartment and a statistically significant prevalence of JC viruria and of BKV in PC tissues. A BKV DDP-like NCCR sequence was isolated in two patients, whereas JCV NCCR was consistently of an archetypal structural organization. A prevalence of the European genotypes was observed for both viruses.
CONCLUSION: Our data demonstrated the presence of JCV DNA in 14/24 (58.3%) cancerous prostatic tissue specimens, confirming the results obtained in a previous study, in which JCV has been defined as common inhabitant of the prostate, and opening the discussion about its potential role in PC
EFFICIENT PROPAGATION OF ARCHETYPE JC POLYOMAVIRUS IN COS-7 CELLS: EVALUATION OF REARRANGEMENTS WITHIN NCCR STRUCTURAL ORGANIZATION DURING TRANSFECTION.
John Cunningham virus (JCPyV) is an ubiqui-tous human pathogen that causes disease in immunocom-promised patients. The JCPyV genome is composed of an early region and a late region, which are physically sepa-rated by the non-coding control region (NCCR). The DNA sequence of the NCCR distinguishes two forms of JCPyV, the designated archetype and the prototype, which resulted from a rearrangement of the archetype sequence. To date, the cell culture systems for propagating JCPyV archetype have been very limited in their availability and robust-ness. Prior to this study, it was demonstrated that JCPyV archetype DNA replicates in COS-7 simian kidney cells expressing SV40 TAg and COS-7 cells expressing HIV-1 Tat. Based on these observations, the present study was conducted to reproduce an in vitro model in COS-7 cells transfected with the JCPyV archetype strain in order to study JCPyV DNA replication and analyze NCCR rear-rangements during the viral life cycle. The efficiency of JCPyV replication was evaluated by quantitative PCR (Q-PCR) and by hemagglutination (HA) assay after trans-fection. In parallel, sequence analysis of JCPyV NCCR was performed. JCPyV efficiently replicated in kidney-derived COS-7 cells, as demonstrated by a progressive increase in viral load and virion particle production after transfection. The archetypal structure of NCCR was maintained during the viral cycle, but two characteristic point mutations were detected 28 days after transfection. This model is a useful tool for analyzing NCCR rearrangements during in vitroreplication in cells that are sites of viral persistence, such as tubular epithelial cells of the kidne
Increased prevalence of Human Polyomavirus JC viruria in Chronic Inflammatory Rheumatic Diseases patients in treatment with anti-TNF α: a 18 month follow-up study.
Chronic inflammatory rheumatic diseases (CIRDs) are immune-mediated pathologies
involving joints. To date, TNFα-blocking agents administration is the most promising
therapy, although these treatments are associated with an increased Polyomavirus
JC (JCPyV) reactivation, the etiological agent of the Progressive Multifocal
Leukoencephalopathy (PML). The aim of this study was the recruitment and the
analysis of a CIRDs cohort in order to investigate a possible correlation between
JCPyV presence and the influence of anti-TNF-α agents on viral loads. Blood and urine
samples were collected from 34 CIRDs subjects prior the first anti-TNF-α infusion (T0)
and after 3 (T3), 6 (T6), 12 (T12), and 18 (T18) months. Results showed persistent JC
viruria significantly higher than JC viremia throughout the 18 month follow-up study
(p=0.002). In JCPyV positive samples, the non-coding control region (NCCR) was
analyzed. Results evidenced archetypal structures (type II-S) in all isolates with the
exception of a sequence isolated from a plasma sample, that corresponds to the type
II-R found in PML subjects. Finally, the viral protein 1 (VP1) genotyping was performed
and results showed the prevalence of the European genotypes 1A, 1B, and 4. Since
only few studies have been carried out to understand whether there is a PML risk in
CIRDs population infected by JCPyV, this study contributes to enrich literature insight on
JCPyV biology in this cluster. Further investigations are necessary in order to recognize
the real impact of biologics on JCPyV life cycle and to identify possible and specific viral
variants related to increased virulence in CIRDs patient
Riattivazione del Poliomavirus umano JC in soggetti affetti da Malattie Infiammatorie Croniche Intestinali trattati con farmaci biologici
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