Molecular characterization of Hellenic variants of Apple scar skin viroid and Pear blister canker viroid in pome fruit trees

Apple scar skin viroid (ASSVd) and Pear blister canker viroid (PBCVd) are members of the genus Apscaviroid (family Pospiviroidae). In order to study the nucleotide sequence and secondary structure of Hellenic variants of these viroids, a large number of collected samples were initially screened by imprint hybridization; then ASSVd and PBCVd positive samples were assayed for the viroids by RT-PCR. Total RNA extracts were reverse-transcribed and amplified by polymerase chain reaction using two different specific primer pairs for each viroid. Purified RT-PCR products were directly sequenced or cloned into the pGEM-T and pCR® II vectors and then sequenced. Fourteen Hellenic full length ASSVd variants from 3 apple, 3 wild apple (Malus sylvestris), 1 wild pear (Pyrus amygdaliformis) and 3 pear trees are 330-335 nucleotides long. They differ from the reference sequences of ASSVd (ASSCS and Y00435) at 15-29 and 3-36 sites, respectively. Fifteen nucleotide changes (differences from ASSCS) are common among all Hellenic variants. Hellenic ASSVd variants share high identity (97-100%) with ASSVd isolates from Asian apples. Three Hellenic variants, deriving from different hosts and areas, are identical with each other (wild apple and apple from Pella [Macedonia] and pear from Achaia [Peloponnesus]) and with another group of 3 apple variants from China (Liaoning, AM1 and B-9). Sixteen full length Hellenic PBCVd variants from 12 trees (4 apples, 1 wild apple, 5 pears, 1 wild pear and 1 quince) are 314-316 nucleotides long. There are 6-50 nucleotide changes among all Hellenic variants and the prototype PBCVd isolate (NC001830). Twenty-two (22) changes are identical among the majority of the Hellenic variants, regardless of origin, and 28-35 changes occur in PBCVd sequences obtained from apple and wild apple samples. In addition, 2 Hellenic PBCVd variants are 97-98% homologous to some Australian and European (Bosnian) PBCVd pear isolates, whereas the remaining 14 share 86-94% identity with Australian PBCVd isolates from pear, quince and Japanese pear (Pyrus pyrifolia). This is the first detailed molecular study of ASSVd and PBCVd in Hellenic cultivated and wild pome fruit trees.Keywords: ASSVd, PBCVd, pome fruit, molecular characterizatio

Pospiviroidae viroids in naturally infected stone and pome fruits in Greece

Viroid research on pome and stone fruit trees in Greece is important, as it seems that such viroids are widespread in the country and may cause serious diseases. Our research dealt with three Pospiviroidae species infecting pome and stone fruit trees, namely Apple scar skin viroid (ASSVd), Pear blister canker viroid (PBCVd) and Hop stunt viroid (HSVd). Tissue-print hybridization, reverse transcription-polymerase chain reaction (RT-PCR), cloning and sequencing techniques were successfully used for the detection and identification of these viroids in a large number of pome and stone fruit tree samples from various areas of Greece (Peloponnesus, Macedonia, Thessaly, Attica and Crete). The 58 complete viroid sequences obtained (30 ASSVd, 16 PBCVd and 12 HSVd) were submitted to the GenBank. Our results showed the presence of ASSVd in apple, pear, wild apple (Malus sylvestris), wild pear (Pyrus amygdaliformis) and sweet cherry; HSVd in apricot, peach, plum, sweet cherry, bullace plum (Prunus insititia), apple and wild apple; and PBCVd in pear, wild pear, quince, apple and wild apple. This research confirmed previous findings of infection of Hellenic apple, pear and wild pear with ASSVd, pear, wild pear and quince with PBCVd and apricot with HSVd. Our findings also revealed for the first time the natural mixed infection of apple and wild apple with (ASSVd+PBCVd+HSVd), of apple and pear with (ASSVd+PBCVd), and of wild apple with (ASSVd+HSVd), as well as the natural infection of Hellenic sweet cherry, peach, bullace plum and plum with HSVd. To our knowledge, this is the first published report of detecting HSVd and PBCVd in infected apple and wild apple, and ASSVd in sweet cherry. Keywords: ASSVd, PBCVd, HSVd, stone fruit, pome fruit, Greec

First report and molecular analysis of Apple scar skin viroid in sweet cherry

Apple scar skin viroid (ASSVd) is a serious pathogen of pome fruits. Recently, it has been reported in Chinese apricot and Chinese peach. In the context of our research on fruit tree viroids in Greece, ASSVd was initially detected in a sweet cherry tree cv ‘Tragana Edessis’ from Florina (Macedonia) by RT-PCR and this finding was confirmed by direct sequencing. This tree is located at the edge of a newly established apple orchard, along with other sweet cherry and wild cherry (Prunus avium) trees. In order to verify this interesting finding, we examined for ASSVd four sweet cherry trees, two wild cherry trees and their neighboring apple trees in the same orchard.The examination was done by imprint hybridization using an ASSVd-specific DIG-labelled probe at stringent hybridization conditions and by RT-PCR using two different ASSVd specific primer pairs. We obtained ASSVdpositive results for all 6 cherry trees. No ASSVd was detected in the apple trees of the orchard. Purified ASSVdpositive RT-PCR products from the cherries were directly sequenced or cloned into the pGEM-T vector and then sequenced. ASSVd sequences were obtained from 5 trees. These sequences are 327-340 nucleotides long and share 96-99% identity with ASSVd isolates from Asian (Indian) apples. These results are similar to our data for other ASSVd variants from cultivated and wild pome fruit trees in Greece. The cherry ASSVd variants differ from the ASSCS prototype isolate of ASSVd at 18-29 sites. There are 15 nucleotide changes (differences from ASSCS) common to all Hellenic ASSVd variants, including cherry and pome fruit tree variants. There are no cherry-specific nucleotide changes in the ASSVd sequences obtained. To our knowledge, this is the first published report of natural infection of cherry by ASSVd. Keywords: ASSVd, cherry, molecular analysis, Hellenic sequence

Chronic inflammatory demyelinating polyneuropathy: A 6-year retrospective clinical study of a hospital-based population

We reviewed the clinical, electrophysiological, laboratory and neuroimaging features of 25 patients with chronic inflammatory demyelinating polyneuropathy (CIDP) admitted to Aeginition Hospital from 1996 to 2001. We also investigated the response to several treatment modalities. The aim was to reveal the clinical spectrum of the disease; the diagnostic criteria developed by the Ad Hoc Subcommittee of the American Academy of Neurology (AAN) in 1991 were used. The subjects consisted of 17 men (68%) and eight women (32%) aged 18-81 years (mean age: 48.5 years) with CIDP. Eighteen patients (72%) had a symmetric neuropathy, whereas seven (28%) had an asymmetric neuropathy. Two patients (8%) had a pure sensory neuropathy. Nine (36%) presented with cranial nerve involvement and only one (4%) had central nervous system demyelination. Most patients had a satisfactory response after treatment with corticosteroids, intravenous immunoglobulins, plasma exchange and azathioprine. In conclusion, CIDP is a clinically heterogeneous disorder. It is one of the few serious chronic neuropathies that has a good (although not permanent) treatment response. (C) 2006 Elsevier Ltd. All rights reserved

Low penetrance of antibiotics in the epithelial lining fluid. The role of inhaled antibiotics in patients with bronchiectasis.

Plasma drug concentrations, spectrum of antibacterial activity and minimum inhibitory concentration (MIC) had been widely considered as markers of the efficacy of antibiotics. Nonetheless, in several cases, antibiotics characterized by all these features were ineffective for the treatment of respiratory tract infections. A typical paradigm represented the case of patients with bronchiectasis who do not always benefit from antibiotics and thus experiencing increased sputum production, worse quality of life, more rapid forced expiratory volume in the first second (FEV1) decline, more frequent exacerbations and increased mortality rates, especially those with Pseudomonas aeruginosa (P. aeruginosa) chronic infection. Subsequently, penetrance of antibiotics in the epithelial lining fluid has gradually emerged as another key factor for the outcome of antibiotic treatment. Given that a plethora of antibiotics presented with poor or intermediate penetrance in the epithelial lining fluid, inhaled antibiotics targeting directly the site of infection emerged as a new option for patients with respiratory disorders including patients with bronchiectasis. This review article intends to summarize the current state of knowledge for the penetrance of antibiotics in the epithelial lining fluid and present results from clinical trials of inhaled antibiotics in patients with bronchiectasis of etiology other than cystic fibrosis

The lung microbiome dynamics between stability and exacerbation in chronic obstructive pulmonary disease (COPD): Current perspectives

Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory disorder with a course that is not uniform for all COPD patients. Although smoking is considered as the major cause of the disease, persistent or recurrent infections seem to play a particular role in the disease establishment and progression. COPD is characterized by dysregulated immunity that has been associated with the bacterial colonization and infections. The establishment of culture-independent techniques has shed new light on the relationships between bacterial ecology and health status and expanded our knowledge on the lung microbiome. Interactions between the host and lung microbiome result in inflammation and activation of resident cells. The lung microbiome contains populations of symbionts and pathobionts in balance which lose their equilibrium and disturb the balance of T-helper and regulatory T-cells (Treg) upon infection, or lung disease. In COPD factors such as disease severity, exacerbations, degree of inflammation, and type of treatment used (e.g inhaled or systemic steroids and antibiotics) affect the composition of lung microbiota. Recent data indicate that the presence of specific bacterial taxa in the airways has the potential to influence the host immune response and possibly to interfere with disease phenotype. Although, there is a growing body of evidence for the role of microbiome in COPD several unanswered questions still exist for its clinical relevance. © 201

Real-World Assessment of Quality of Life in Patients with Relapsing Remitting Multiple Sclerosis Treated with Teriflunomide for Two Years: Patient-Reported Outcomes from the AURELIO Study in Greece

Introduction: Multiple sclerosis (MS) is a highly heterogeneous inflammatory disease of the central nervous system. Patient-reported outcomes (PROs) in a real-world clinical setting can provide detailed information about MS from the patient's perspective. PROs were used here to assess quality of life (QoL), treatment satisfaction, clinical efficacy, and safety outcomes in a Greek cohort of relapsing remitting MS (RRMS) patients treated with oral teriflunomide (14 mg/day). Methods: AURELIO was a 2-year, prospective, observational study whose QoL primary endpoint was assessed with the Multiple Sclerosis Impact Scale (MSIS-29). Secondary endpoints included analyses of Patient Determined Disease Steps (PDDS), Treatment Satisfaction Questionnaire for Medication (TSQM), Expanded Disability Status Scale (EDSS), annualized relapse rate (ARR), adherence, and safety outcomes. Results: AURELIO enrolled 282 patients (62.8% female; mean age 44.8 [SD ± 11] years; EDSS 2.0 [SD ± 1.6]; 44.6% treatment-naïve), with 212 patients (75%) remaining on treatment at study end. MSIS-29 total scores remained stable, while the MSIS-29 psychological scale showed significant improvement (p = 0.0015) at 2 years vs. baseline. TSQM scores at 2 years showed significant improvements in effectiveness (+ 6.6, p = 0.0001), convenience (+ 1.9, p = 0.0256), and global satisfaction (+ 8.1, p = 0.0001) vs. baseline. Disease progression was stable as indicated by non-significant changes in PDDS and EDSS vs. baseline. The ARR was low at 0.065, with a slightly higher ARR in previously treated (0.070) vs. naïve patients (0.058). Adherence was high at > 90%. Overall, 91 patients (32.3%) in the study reported a total of 215 safety events (32 serious, of which 21 were classified as mild–moderate). No new safety signals were observed. Conclusions: These data highlight the importance of PROs to facilitate personalized treatment strategies in MS. In line with other teriflunomide studies, AURELIO showed stable QoL, efficacy and safety outcomes, and good treatment satisfaction both in treatment-naïve and previously treated patients in this Greek cohort of patients with RRMS. © 2022, The Author(s)