Prevalence and determinants of resistance mutations in HIV-1-infected patients exposed to integrase inhibitors in a large Italian cohort.

OBJECTIVES: The aim of the study was to analyse the prevalence of integrase resistance mutations in integrase strand transfer inhibitor (INSTI)-experienced HIV-1-infected patients and its predictors. METHODS: We selected HIV-1 integrase sequences from the Antiviral Response Cohort Analysis (ARCA) database, derived from INSTI-experienced patients between 2008 and 2017. Differences in the prevalence of resistance to raltegravir (RAL), elvitegravir (EVG) and dolutegravir (DTG) were assessed by χ2 test and predictors of resistance were analysed by logistic regression. RESULTS: We included 462 genotypes from INSTI-exposed individuals: 356 'INSTI-failing' patients and 106 'previously INSTI-exposed' patients (obtained a median of 42 weeks after INSTI discontinuation [interquartile range (IQR) 17-110 weeks]). Overall, at least low-level resistance (LLR) to any INSTI (Stanford 8.5 algorithm) was detected in 198 (42.9%) cases. The most frequent INSTI resistance mutation was N155H, followed by Q148H/K/R, G140A/C/S, E138A/K/T and Y143C/H/R. Y143R and E138A were more prevalent in viral subtype B versus non-B [5.2 versus 1.5%, respectively (P = 0.04), and 3.1 versus 0%, respectively (P = 0.02)]. Overall, the Q148H/K/R plus G140A/C/S and/or E138A/K/T pattern, defining an intermediate level of resistance to DTG, was detected in 70 (15%) cases. Independent predictors of at least LLR to any INSTI were current use versus past use of INSTIs, a lower genotypic sensitivity score (GSS) for contemporary antiretroviral drugs used, and having an integrase sequence obtained in calendar year 2016 as compared to 2008-2009. CONCLUSIONS: The results support integrase resistance testing in INSTI-experienced patients. Emergence of INSTI resistance is facilitated by the reduced genetic barrier of the regimen as a consequence of resistance to companion drugs. However, INSTI resistance may become undetectable by standard population sequencing upon INSTI discontinuation.

Further evidence about the crucial role of CSF biomarkers in diagnosis of posterior cortical atrophy.

Cerebrospinal fluid (CSF) biomarkers (protein tau, phosphorylated tau and amyloid Beta 1-42) are recognized as a supportive feature in diagnosis of Alzheimer's disease (AD) and their role in identifying atypical variants of AD is currently under investigation. We dosed these proteins in nine patients clinically and instrumentally affected by posterior cortical atrophy (PCA), a rare disorder characterized by a progressive neurodegenerative process that involves primarily the posterior brain regions. We compared the obtained values with a large group of AD patients (N = 117), recruited in our neurological department. Our data revealed no differences in the CSF profile between PCA and AD, showing abnormal values of protein tau, phosphorylated tau and amyloid Beta 1-42 in both groups of patients. This study underlines the diagnostic importance of CSF biomarkers in PCA patients, supporting the hypothesis that PCA is an atypical variant of AD with an onset before the age of 65.

Double heterozygosity for BRCA1 and hMLH1 gene mutations in a 46-year-old woman with five primary tumors.

Germline mutations in BRCA1 and BRCA2 genes predispose to hereditary breast cancer, whereas carriers of mutations in any of the mismatch repair genes (MMR; hMLH1, hMSH2, hMSH6, hPMS2) are highly susceptible to Lynch syndrome. In the present study, we describe a woman affected by unilateral breast cancer at the age of 35 years. After 4 years, during the follow-up she developed synchronous (and asymptomatic) endometrial cancer, ovarian carcinoma and renal clear cell carcinoma. After 7 years (at age 46), the patient developed an infiltrating carcinoma of the contralateral breast and died in a few months of metastatic disease. Initial investigations led to the detection of a constitutional mutation in the BRCA1 gene. The extended genealogical tree disclosed a suspected history of colorectal carcinoma in the maternal branch. Endometrial cancer of the proband was investigated for microsatellite instability (MSI) and immunohistochemical expression of MLH1, MSH2 and MSH6 proteins. An high MSI status and lack of expression of MLH1 protein were detected. hMLH1 gene sequencing revealed the presence of a constitutional mutation, which was also found in the mother of the proband. Loss of the wild-type hMLH1 allele was detected in both breast tumors, thus suggesting that the MMR defect contributed to the development of the breast cancer.

Microglia convert aggregated amyloid-ß into neurotoxic forms through the shedding of microvesicles.

Alzheimer's disease (AD) is characterized by extracellular amyloid-ß (Aß) deposition, which activates microglia, induces neuroinflammation and drives neurodegeneration. Recent evidence indicates that soluble pre-fibrillar Aß species, rather than insoluble fibrils, are the most toxic forms of Aß. Preventing soluble Aß formation represents, therefore, a major goal in AD. We investigated whether microvesicles (MVs) released extracellularly by reactive microglia may contribute to AD degeneration. We found that production of myeloid MVs, likely of microglial origin, is strikingly high in AD patients and in subjects with mild cognitive impairment and that AD MVs are toxic for cultured neurons. The mechanism responsible for MV neurotoxicity was defined in vitro using MVs produced by primary microglia. We demonstrated that neurotoxicity of MVs results from (i) the capability of MV lipids to promote formation of soluble Aß species from extracellular insoluble aggregates and (ii) from the presence of neurotoxic Aß forms trafficked to MVs after Aß internalization into microglia. MV neurotoxicity was neutralized by the Aß-interacting protein PrP and anti-Aß antibodies, which prevented binding to neurons of neurotoxic soluble Aß species. This study identifies microglia-derived MVs as a novel mechanism by which microglia participate in AD degeneration, and suggest new therapeutic strategies for the treatment of the disease.

Culture-independent analysis of endophytic bacterial communities associated with Brazilian sugarcane.

Sugarcane is an economically important culture in Brazil. Endophytic bacteria live inside plants, and can provide many benefits to the plant host. We analyzed the bacterial diversity of sugarcane cultivar RB-72454 by cultivation-independent techniques. Total DNA from sugarcane stems from a commercial plantation located in Paraná State was extracted. Partial 16S rRNA genes were amplified and sequenced for library construction. Of 152 sequences obtained, 52% were similar to 16S rRNA from Pseudomonas sp, and 35.5% to Enterobacter sp. The genera Pantoea, Serratia, Citrobacter, and Klebsiella were also represented. The endophytic communities in these sugarcane samples were dominated by the families Enterobacteriaceae and Pseudomonadaceae (class Gammaproteobacteria).

Differential effect of everolimus on progression of early and late cardiac allograft vasculopathy in current clinical practice.

Randomized trials showed that mTOR inhibitors prevent early development of cardiac allograft vasculopathy (CAV). However, the action of these drugs on CAV late after transplant is controversial, and their effectiveness for CAV prevention in clinical practice is poorly explored. In this observational study we included 143 consecutive heart transplant recipients who underwent serial intravascular ultrasound (IVUS), receiving either everolimus or mycophenolate as adjunctive therapy to cyclosporine. Ninety-one recipients comprised the early cohort, receiving IVUS at weeks 3-6 and year 1 after transplant, and 52 the late cohort, receiving IVUS at years 1 and 5 after transplant. Everolimus independently reduced the odds for early CAV (0.14 [0.01-0.77]; p = 0.02) but it did not appear to influence late CAV progression. High-dose statins were found to be associated with reduced CAV progression both early and late after transplant (p ≤ 0.05). Metabolic abnormalities, such as high triglycerides, were associated with late, but not with early CAV progression. By highlighting a differential effect of everolimus and metabolic abnormalities on early and late changes of graft coronary morphology, this observational study supports the hypothesis that everolimus may be effective for CAV prevention but not for CAV treatment, and that risk factors intervene in a time-dependent sequence during CAV development.

Anatomical brain connectivity can assess cognitive dysfunction in multiple sclerosis.

BACKGROUND: Brain disconnection plays a major role in determining cognitive disabilities in multiple sclerosis (MS). We recently developed a novel diffusion-weighted magnetic resonance imaging (DW-MRI) tractography approach, namely anatomical connectivitity mapping (ACM), that quantifies structural brain connectivity. OBJECTIVE: Use of ACM to assess structural connectivity modifications in MS brains and ascertain their relationship with the patients' Paced-Auditory-Serial-Addition-Test (PASAT) scores. METHODS: Relapsing-remitting MS (RRMS) patients (n = 25) and controls (n = 25) underwent MRI at 3T, including conventional images, T1-weighted volumes and DW-MRI. Volumetric scans were coregistered to fractional anisotropy (FA) images, to obtain parenchymal FA maps for both white and grey matter. We initiated probabilistic tractography from all parenchymal voxels, obtaining ACM maps by counting the number of streamlines passing through each voxel, then normalizing by the total number of streamlines initiated. The ACM maps were transformed into standard space, for statistical use. RESULTS: RRMS patients had reduced grey matter volume and FA, consistent with previous literature. Also, we showed reduced ACM in the thalamus and in the head of the caudate nucleus, bilaterally. In our RRMS patients, ACM was associated with PASAT scores in the corpus callosum, right hippocampus and cerebellum. CONCLUSIONS: ACM opens a new perspective, clarifying the contribution of anatomical brain disconnection to clinical disabilities in MS.

From primary progressive aphasia to corticobasal syndrome: two clinical and rCBF functional reports.

We describe two cases, both presenting with a 2-year history of isolated language disorders, one compatible with logopenic variant and the other with non-fluent variant of primary progressive aphasia (PPA). Afterwards, each developed a corticobasal syndrome (CBS) with alien limb phenomenon and a multi-domain cognitive impairment. Regional cerebral perfusion (rCBF) study using 99mTc-ECD single photon emission computed tomography (SPECT) revealed hypoperfusion patterns consistent with these aphasia types and with the presence of limb apraxia. We report two cases of PPA variants associated with CBS and we suggest that SPECT rCBF correlates can be useful in making a differential diagnosis within the PPA spectrum.

White matter damage in frontotemporal lobar degeneration spectrum.

White matter (WM) tract damage was assessed in patients with the behavioral variant frontotemporal dementia (bvFTD) and the 3 primary progressive aphasia (PPA) variants and compared with the corresponding brain atrophy patterns. Thirteen bvFTD and 20 PPA patients were studied. Tract-based spatial statistics and voxel-based morphometry were used. Patients with bvFTD showed widespread diffusion tensor magnetic resonance imaging (DT MRI) abnormalities affecting most of the WM bilaterally. In PPA patients, WM damage was more focal and varied across the 3 syndromes: left frontotemporoparietal in nonfluent, left frontotemporal in semantic, and left frontoparietal in logopenic patients. In each syndrome, DT MRI changes extended beyond the topography of gray matter loss. Left uncinate damage was the best predictor of frontotemporal lobar degeneration diagnosis versus controls. DT MRI measures of the anterior corpus callosum and left superior longitudinal fasciculus differentiated bvFTD from nonfluent cases. The best predictors of semantic PPA compared with both bvFTD and nonfluent cases were diffusivity abnormalities of the left uncinate and inferior longitudinal fasciculus. This study provides insights into the similarities and differences of WM damage in bvFTD and PPA variants. DT MRI metrics hold promise to serve as early markers of WM integrity loss that only at a later stage may be detectable by volumetric measures.

Diversity of 16S rRNA genes from bacteria of sugarcane rhizosphere soil

Sugarcane is an important agricultural product of Brazil, with a total production of more than 500 million tons. Knowledge of the bacterial community associated with agricultural crops and the soil status is a decisive step towards understanding how microorganisms influence crop productivity. However, most studies aim to isolate endophytic or rhizosphere bacteria associated with the plant by culture-dependent approaches. Culture-independent approaches allow a more comprehensive view of entire bacterial communities in the environment. In the present study, we have used this approach to assess the bacterial community in the rhizosphere soil of sugarcane at different times and under different nitrogen fertilization conditions. At the high taxonomic level, few differences between samples were observed, with the phylum Proteobacteria (29.6 percent) predominating, followed by Acidobacteria (23.4 percent), Bacteroidetes (12.1 percent), Firmicutes (10.2 percent), and Actinobacteria (5.6 percent). The exception was the Verrucomicrobia phylum whose prevalence in N-fertilized soils was approximately 0.7 percent and increased to 5.2 percent in the non-fertilized soil, suggesting that this group may be an indicator of nitrogen availability in soils. However, at low taxonomic levels a higher diversity was found associated with plants receiving nitrogen fertilizer. Bacillus was the most predominant genus, accounting for 19.7 percent of all genera observed. Classically reported nitrogen-fixing and/or plant growth-promoting bacterial genera, such as Azospirillum, Rhizobium, Mesorhizobium, Bradyrhizobium, and Burkholderia were also found although at a lower prevalence.

Diversity of 16S rRNA genes from bacteria of sugarcane rhizosphere soil.

Sugarcane is an important agricultural product of Brazil, with a total production of more than 500 million tons. Knowledge of the bacterial community associated with agricultural crops and the soil status is a decisive step towards understanding how microorganisms influence crop productivity. However, most studies aim to isolate endophytic or rhizosphere bacteria associated with the plant by culture-dependent approaches. Culture-independent approaches allow a more comprehensive view of entire bacterial communities in the environment. In the present study, we have used this approach to assess the bacterial community in the rhizosphere soil of sugarcane at different times and under different nitrogen fertilization conditions. At the high taxonomic level, few differences between samples were observed, with the phylum Proteobacteria (29.6%) predominating, followed by Acidobacteria (23.4%), Bacteroidetes (12.1%), Firmicutes (10.2%), and Actinobacteria (5.6%). The exception was the Verrucomicrobia phylum whose prevalence in N-fertilized soils was approximately 0.7% and increased to 5.2% in the non-fertilized soil, suggesting that this group may be an indicator of nitrogen availability in soils. However, at low taxonomic levels a higher diversity was found associated with plants receiving nitrogen fertilizer. Bacillus was the most predominant genus, accounting for 19.7% of all genera observed. Classically reported nitrogen-fixing and/or plant growth-promoting bacterial genera, such as Azospirillum, Rhizobium, Mesorhizobium, Bradyrhizobium, and Burkholderia were also found although at a lower prevalence.

Zirconia toughened alumina (ZTA) powders: ultrastructural and histological analysis.

Ceramic materials, as Alumina and Zirconia, has made an improvement in the choice of new biomaterials for the load bearing application in dental and orthopaedic implants. These materials has shown mechanical resistance to high stress related to weight bearing and low debris in time. For this reason they are indicated on young patients implant, with high demanding activities and long life expectance. In literature however the risk of chronic inflammation due to chronic wear debris release and the possibility of carcinogenesis, is still to be definitively investigated. Another point to investigate is the acute reaction of the tissue in case of acute release of powders of these materials. The aim of this study was to investigate the possible local and systemic acute effects of ceramic precursors in form of powders of different size when released into articular joint. Powders of ZTA were implanted in the knee joint of twenty-four New Zealand white adult rabbits, that were sacrificed at 1,3,6, and 12 months. Radiographic, histological and immunoistochemestry analysis were conducted on periprosthetic tissue and peripheral organs, to verifying local host response and systemic toxic effects.

Changes in exercise capacity induced by heart transplantation: prognostic and therapeutic implications.

Survival and exercise performance are key targets of heart transplantation (HT). We designed this study to help in identifying (1) patients with chronic heart failure (CHF) at risk of poor exercise capacity after HT and (2) HT recipients presenting risk factors modifiable with exercise showing a potential impact on outcome. We enrolled 49 HT recipients (age 52 ± 12 years, 84% males) who underwent a cardiopulmonary exercise test before (9 ± 6 months) and after (20 ± 14 months) HT. In the CHF phase, lower peak oxygen consumption (VO(2) ) (odds ratio 0.69, P=0.017) independently predicted peak VO(2) improvement after HT. In the post-HT phase, body mass index (BMI) [adjusted hazard ratio (HR) 1.16, P=0.034] and VE (ventilation)/VCO(2) (carbon dioxide production) slope (adjusted HR 1.07, P=0.031) independently predicted mortality. In conclusion, CHF patients with only a moderate impairment of peak VO(2) are at a risk of failing to achieve a significant improvement of exercise performance after HT. In the post-HT phase, a BMI≥28 and/or a VE/VCO(2) slope ≥47 represent risk factors for death, which are potentially modifiable with exercise. Prospective randomized studies are needed to analyze the effects of training on functional capacity and outcome in the different subsets of HT recipients.

In vivo characterization of Zirconia Toughened Alumina material: a comparative animal study.

The development of a new chromia-doped Zirconia Toughened Alumina (ZTA) material was previously reported as displaying mechanical properties suitable for implants with load bearing applications, such as orthopaedic and dental implants. This type of biomaterial is expected to be in contact with living tissues for a long period of time and its long-term toxicity must be carefully evaluated. In this study the suitability of this ZTA material as a candidate biomaterial for orthopaedic implants and dental devices was further investigated in vivo in comparison to alumina and zirconia, which are currently used in orthopaedic and dental surgery. Cylinders of the materials were implanted in vivo in white rabbits, and local and systemic tissue reactions were analyzed at different time intervals after surgery. Radiologic examinations displayed the absence of radiolucence around cylinders and no signs of implant loosening up to twelve months. No tumours developed in the animals either locally (at the site of implantation), or systemically in the peripheral organs. The results obtained suggest that this new ZTA material does not display any long term pathogenic effect in vivo. These findings extend our previous observations on the biocompatibility and the absence of any long-term carcinogenic effect in vitro of this material which displays interesting properties for biomedical applications. In conclusion, we report the in vivo characterization of a new chromia-doped ZTA material and confirm its suitability as a candidate biomaterial for orthopaedic implants and dental devices since it does not give any local nor systemic toxicity even after a long period of time after implantation.

Treatment of Wernicke's encephalopathy with high dose of thiamine in a patient with pyloric sub-stenosis: description of a case.

Wernicke's encephalopathy (WE) is an acute or subacute syndrome that results from a deficiency in vitamin B1 (thiamine). The syndrome is characterised by a classical triad of symptoms: nystagmus and ophthalmoplegia,mental-status changes, and unsteadiness of stance and gait. When patients with WE are inappropriately treated with low doses of thiamine, mortality rates average out at 20% and Korsakoff's Psychosis develops in about 85% of survivors(Sechi and Serra in Lancet Neurol 6(5):442­455,2007). We report the case of a patient with a pyloric substenosis that developed a WE, and was treated with high doses of thiamine showing after few days of treatment a great improvement of neurological and neuroradiological assessment, even though cognitive impairment was still severe at discharge and at 6 months follow-up.

Langerhans cell histiocytosis: bilateral temporal bone involvement in an adult with diabetes insipidus.

OBJECTIVE: To present a clinical case of an adult affected by Langerhans cell histiocytosis with bilateral, non-simultaneous, involvement of the temporal bone, associated with diabetes insipidus and to review the literature. METHODOLOGY: A rare case of bilateral temporal bone involvement of Langerhans cell histiocytosis in a 42-year-old woman affected by diabetes insipidus is reported. We present patient's clinical history supported by radiologic, histopathologic and audiologic findings. RESULTS: The patient was submitted to a series of otologic surgical procedures due to the progression of the disease. Ossicular chain was always preserved, so that conservative surgery (canal wall-up technique) was performed, permitting the achievement of good hearing results, bilaterally. CONCLUSIONS: Temporal bone involvement of Langerhans cell histiocytosis may lead to a progressive chronic disease. However, the ossicular chain can remain uninvolved, making a conservative surgical treatment possible. Careful follow-up is essential for detecting new lesions and serial CT scans are mandatory.

Diversity of endophytic bacteria in Brazilian sugarcane.

Endophytic bacteria live inside plant tissues without causing disease. Studies of endophytes in sugarcane have focused on the isolation of diazotrophic bacteria. We examined the diversity of endophytic bacteria in the internal tissues of sugarcane stems and leaves, using molecular and biochemical methods. Potato-agar medium was used to cultivate the endophytes; 32 isolates were selected for analysis. DNA was extracted and the 16S rRNA gene was partially sequenced and used for molecular identification. Gram staining, catalase and oxidase tests, and the API-20E system were used to characterize the isolates. The strains were divided into five groups, based on the 16S rRNA sequences. Group I comprised 14 representatives of the Enterobacteriaceae; group II was composed of Bacilli; group III contained one representative, Curtobacterium sp; group IV contained representatives of the Pseudomonadaceae family, and group V had one isolate with an uncultured bacterium. Four isolates were able to reduce acetylene to ethylene. Most of the bacteria isolated from the sugarcane stem and leaf tissues belonged to Enterobacteriaceae and Pseudomonaceae, respectively, demonstrating niche specificity. Overall, we found the endophytic bacteria in sugarcane to be more diverse than previously reported.

Development of a new zirconia-toughened alumina: promising mechanical properties and absence of in vitro carcinogenicity.

High purity alumina as well as zirconia ceramics have been widely used as orthopaedic implant biomaterials and dental devices displaying optimal, but sometimes exclusive, mechanical properties. In order to combine the advantages of alumina and zirconia ceramic materials different types of composites have been developed in which either zirconia is dispersed in an alumina matrix or vice versa. Orthopaedic and dental implant biomaterials are expected to be in contact with living tissues for a long period of time and their long term toxicity must be carefully evaluated. In this study we report the development of a high performance chromia-doped zirconia toughened alumina (ZTA) material which displays promising mechanical properties in terms of hardness, strength and fracture toughness that make it suitable for prosthesis even for small joints. The long-term biocompatibility of this material was also evaluated, mainly in terms of DNA damage, mutagenicity and cancerogenetic potential in mammalian cells. The results obtained suggest that this new ZTA material does not display any longterm carcinogenic effect and it is suitable for biomedical applications from a cancerogenetic point of view. In conclusion, we report the development of a new chromia-doped ZTA material with interesting properties, both from a mechanical and a biocompatibility point of view which warrant further studies on its suitability as a candidate biomaterial for orthopaedic implants and dental devices.

Inclusion body myopathy, Paget's disease of the bone and frontotemporal dementia: recurrence of the VCP R155H mutation in an Italian family and implications for genetic counselling.

The acronym IBMPFD denotes a syndrome including inclusion body myopathy, Paget's disease of the bone (PDB) and frontotemporal dementia (FTD) as cardinal features, which is caused by missense mutations in the VCP gene. We studied the clinical characteristics and the histopathological features in two siblings and their mother who presented with adult-onset myopathy and presenile, rapidly progressive FTD. One sibling also showed PDB. Light and electron microscopy performed on muscle biopsies demonstrated degenerative changes with inclusion bodies and abnormal aggregates. Mutation analysis of the VCP gene on affected siblings revealed a heterozygous missense mutation (R155H) in a hot spot. This is the first Italian family with multiple individuals diagnosed as having IBMPFD and carrying the recurrent R155H mutation. The implications for genetic counselling were also discussed, with regard to the procedures that may be offered to families suffering from a multisystem disorder with high risk of cognitive decline.