COVID-19 in donation and transplant.

SARS-CoV-2 infection has had a major impact on donation and transplantation. Since the cessation of activity two years ago, the international medical community has rapidly generated evidence capable of sustaining and increasing this neccesary activity. This paper analyses the epidemiology and burden of COVID-19 in donation and transplantation, the pathogenesis of the infection and its relationship with graft-mediated transmission, the impact of vaccination on donation and transplantation, the evolution of donation in Spain throughout the pandemic, some lessons learned in SARS-CoV-2 infected donor recipients with positive PCR and the applicability of the main therapeutic tools recently approved for treatment among transplant recipients.

Ceftolozane-tazobactam in nosocomial pneumonia.

Ceftolozane is a potent antimicrobial against Pseudomonas aeruginosa, including carbapenem-resistant and multidrug-resistant strains, and is also active against Enterobacteriaceae. It MIC (minimal inhibitory concentration) and MPC (mutant preventive concentration) are close together, allowing to avoid the mutant selection window specifically in the treatment of Pseudomonas aeruginosa infection. The molecule is time-dependent and stable when reconstituted at room temperature, facilitating safe and effective dosage optimization in frail and critically ill patients. It has been shown to be non-inferior to meropenem in the treatment of nosocomial infection in the ASPECT-NP study but superior in post-hoc studies in the subgroup of patients with ventilator-associated pneumonia, without the emergence of resistance during treatment. It is FDA approved at a dose of 3 g every 8 hours in the treatment of nosocomial pneumonia (HABP/VABP) in adults.

Ceftazidime-avibactam.

Ceftazidime is a 3rd generation cephalosporin active against Pseudomonas aeruginosa. Avibactam is an inhibitor of class A, C and some class D ß-lactamases. The antibacterial spectrum of ceftazidime-avibactam covers 95% of P. aeruginosa isolates and >99% of enterobacteria, including strains carrying extended-spectrum ß-lactamases (ESBLs). Selection of resistant mutants in Klebsiella pneumoniae and Enterobacter cloacae strains producing KPC-3 or KPC-2 after exposure to ceftazidime-avibactam has been described by the appearance of one or more amino acid changes in the Ω-loop of the ß-lactamase. These strains usually regain susceptibility to meropenem. There is evidence of a shorter multidrug-resistant organisms colonization period in patients treated with this antimicrobial, which could be beneficial in the treatment of infections caused by bacteria carrying ESBLs or carbapenemases.

Update in Infectious Diseases 2019.

The IX Course of Antimicrobials and Infectious Diseases update included a review of the main issues in clinical microbiology, epidemiology and clinical aspects for a current approach of infectious pathology. The present introduction summarizes about the most important meetings related to infectious diseases during 2018 (ECCMID, IAS, ASM and ID Week). In addition, the course provides a practical information to focus on nosocomial infection models, with immunosuppressed patients or complex multidrug-resistant pathogens. The closing lecture of this year reviewed the infection during donation process.

Response of macrophages and neural cells in contact with reduced graphene oxide microfibers.

Graphene-based materials are revealing a great promise for biomedical applications and demonstrating attractiveness for neural repair. In the context of neural tissue damage, the dialogue between neural and immune cells appears critical for driving regeneration, thus making the understanding of their relations pivotal. Herein, the acute response of RAW-264.7 macrophages on nanostructured reduced graphene oxide (rGO) microfibers has been evaluated through the analysis of cell parameters including proliferation, viability, intracellular content of reactive oxygen species, cell cycle, apoptosis, and cell size and complexity. The influence of the direct contact of rGO microfibers on their polarization towards M1 and M2 phenotypes has been studied by analyses of both M1 (CD80) and M2 (CD163) markers and the secretion of the inflammatory cytokines TNF-α and IL-6. Finally, the capability of these rGO microfibers to regulate neural stem cell differentiation has been also evaluated. Findings reveal that rGO microfibers inhibit the proliferation of RAW-264.7 macrophages without affecting their viability and cell cycle profiles. The presence of M1 and M2 macrophages on these microfibers was confirmed after 24 and 48 h, respectively, accompanied by a decrease in TNF-α and an increase in IL-6 cytokine secretion. These rGO microfibers were also able to support the formation of a highly interconnected neural culture composed of both neurons (map2+ cells) and glial cells (vimentin+ cells). These findings encourage further investigation of these microfibers as attractive biomaterials to interact with immune and neural cells, attempting to support wound healing and tissue repair after implantation.

Update in Infectious Diseases 2018.

VIII Updating Course of Antimicrobials and Infectious Diseases has reviewed useful microbiological, epidemiological and clinical aspects for a current approach of infectious pathology. Present manuscript summarizes a chronicle about the main infection related meetings during 2017 (ECCMID, IAS, ASM and ID Week). In addition, the course proposed a practical approach for understanding different type of pathogens and our selected topics this year were the epidemiology of bacterial nosocomial infection, a practical approach to Clostridium difficile infection patients, a two year selection of the top ten papers about fungal infection and an update in fungal biofilms. Finally, proffesors made a practical approach by main clinical syndromes like sepsis, infections in oncohematological patients, CNS infections in immunosuppressed patients and reviewed the top ten papers in transplant infectious diseases and infection control during the last two years.

Current aspects in sepsis approach. Turning things around.

The incidence and prevalence of sepsis depend on the definitions and records that we use and we may be underestimating their impact. Up to 60% of the cases come from the community and in 30-60% we obtain microbiological information. Sometimes its presentation is ambiguous and there may be a delay in its detection, especially in the fragile population. Procalcitonin is the most validated biomarker for bacterial sepsis and the one that best discriminates the non-infectious cause. Presepsin and pro-adrenomedullin are useful for early diagnosis, risk stratification and prognosis in septic patients. The combination of biomarkers is even more useful to clarify an infectious cause than any isolated biomarker. Resuscitation with artificial colloids has worse results than crystalloids, especially in patients with renal insufficiency. The combination of saline solution and balanced crystalloids is associated with a better prognosis. Albumin is only recommended in patients who require a large volume of fluids. The modern molecular methods on the direct sample or the identification by MALDI-TOF on positive blood culture have helped to shorten the response times in diagnosis, to optimize the antibiotic treatment and to facilitate stewardship programs. The hemodynamic response in neonates and children is different from that in adults. In neonatal sepsis, persistent pulmonary hypertension leads to an increase in right ventricular afterload and heart failure with hepatomegaly. Hypotension, poor cardiac output with elevated systemic vascular resistance (cold shock) is often a terminal sign in septic shock. Developing ultra-fast Point-of-Care tests (less than 30 minutes), implementing technologies based on omics, big data or massive sequencing or restoring "healthy" microbiomes in critical patients after treatment are the main focuses of research in sepsis. The main benefits of establishing a sepsis code are to decrease the time to achieve diagnosis and treatment, improve organization, unify criteria, promote teamwork to achieve common goals, increase participation, motivation and satisfaction among team members, and reduce costs.

Update on management of invasive candidiasis.

Given the growing incidence of invasive candidiasis in critically ill and haemato-oncological patients and its poor outcomes, an early diagnosis and treatment are need for get a better prognosis. This document reviews the current ap-proaches that help in diagnosis of invasive candidiasis based on culture-independent microbiological tests. The combination of clinical prediction scores with fungal serological markers could facilitate the approach in antifungal therapy, optimiz-ing it. This article also reviews the epidemiology and primary risk factors for invasive candidiasis in these patients, updating the therapeutic approach algorithms in both clinical contexts based on the main evidence and international guidelines.

Update in Infectious Diseases 2017.

Antimicrobial resistance in complex models of continuous infection is a current issue. The update 2017 course addresses about microbiological, epidemiological and clinical aspects useful for a current approach to infectious disease. During the last year, nosocomial pneumonia approach guides, recommendations for management of yeast and filamentous fungal infections, review papers on the empirical approach to peritonitis and extensive guidelines on stewardship have been published. HIV infection is being treated before and more intensively. The implementation of molecular biology, spectrometry and inmunology to traditional techniques of staining and culture achieve a better and faster microbiological diagnosis. Finally, the infection is increasingly integrated, assessing non-antibiotic aspects in the treatment.

MC3T3-E1 pre-osteoblast response and differentiation after graphene oxide nanosheet uptake.

Nano-graphene oxide (GO) and its functionalized derivatives have aroused a great interest for drug delivery, tissue engineering and photothermal cancer therapy, but their biocompatibility has not yet been fully assessed. The aim of the present study was to evaluate the proliferation and differentiation of MC3T3-E1 pre-osteoblasts after the uptake of GO nanosheets (c.a. 400nm), functionalized with poly(ethylene glycol-amine) (PEG) and labelled with fluorescein isothiocyanate (FITC). Significant proliferation decrease and apoptosis increase were observed 3days after incorporation of FITC-PEG-GO by MC3T3-E1 cells. However, alterations on healthy pre-osteoblast differentiation into cells exhibiting osteoblast phenotype were not observed, as they showed normal alkaline phosphatase levels and matrix mineralization 12days after nanosheet uptake. The results suggest that 40µg/mL concentrations of these nanosheets would not affect the differentiation of healthy pre-osteoblasts, thus these PEG-GO nanosheets have potential to be used for biomedical applications after their internalization, as the induction of local hyperthermia on bone cancer.

Effects of nanocrystalline hydroxyapatites on macrophage polarization.

Silicon substituted and nanocrystalline hydroxyapatites have attracted the attention of many researchers due to their up-regulation in osteoblast cell metabolism and enhanced bioreactivity, respectively. On the other hand, the biomaterial success or failure depends ultimately on the immune response triggered after its implantation. Macrophages are the main components of the innate immune system with an important role in healing and tissue remodelling due to their remarkable functional plasticity, existing in a whole spectrum of functional populations with varying phenotypic features. The effects of nanocrystalline hydroxyapatite (nano-HA) and nanocrystalline silicon substituted hydroxyapatite (nano-SiHA) on the macrophage populations defined as pro-inflammatory (M1) and reparative (M2) phenotypes have been evaluated in the present study using RAW 264.7 cells and mouse peritoneal macrophages as in vitro models. M1 and M2 macrophage phenotypes were characterized by flow cytometry and confocal microscopy by the expression of CD80 and CD163, known as M1 and M2 markers, respectively. The polarization of primary macrophages towards the M1 or M2 phenotype was induced with the pro-inflammatory stimulus LPS or the anti-inflammatory stimulus IL-10, respectively, evaluating the biomaterial effects under these conditions. Our results show that both nano-HA and nano-SiHA favour the macrophage polarization towards an M2 reparative phenotype, decreasing M1 population and ensuring an appropriate response in the implantation site of these biomaterials designed for bone repair and bone tissue engineering.

Effects of bleaching on osteoclast activity and their modulation by osteostatin and fibroblast growth factor 2.

HYPOTHESIS: Dental bleaching with H2O2 is a common daily practice in dentistry to correct discoloration of anterior teeth. The aim of this study has been to determine whether this treatment of human teeth affects growth, differentiation and activity of osteoclast-like cells, as well as the putative modulatory action of osteostatin and fibroblast growth factor 2 (FGF-2). EXPERIMENTS: Previously to the in vitro assays, structural, physical-chemical and morphological features of teeth after bleaching were studied. Osteoclast-like cells were cultured on human dentin disks, pre-treated or not with 38% H2O2 bleaching gel, in the presence or absence of osteostatin (100 nM) or FGF-2 (1 ng/ml). Cell proliferation and viability, intracellular content of reactive oxygen species (ROS), pro-inflammatory cytokine (IL-6 and TNFα) secretion and resorption activity were evaluated. FINDINGS: Bleaching treatment failed to affect either the structural or the chemical features of both enamel and dentin, except for slight morphological changes, increased porosity in the most superficial parts (enamel), and a moderate increase in the wettability degree. In this scenario, bleaching produced an increased osteoclast-like cell proliferation but decreased cell viability and cytokine secretion, while it augmented resorption activity on dentin. The presence of either osteostatin or FGF-2 reduced the osteoclast-like cell proliferation induced by bleaching. FGF-2 enhanced ROS content, whereas osteostatin decreased ROS but increased TNFα secretion. The bleaching effect on resorption activity was increased by osteostatin, but this effect was less evident with FGF-2. CONCLUSIONS: These findings further confirm the deleterious effects of tooth bleaching by affecting osteoclast growth and function as well as different modulatory actions of osteostatin and FGF-2.

Is Streptococcus bovis a urinary pathogen?

The Streptococcus bovis group (SBG) comprises several microorganisms associated with human infections. They have been associated with bacteremia, endocarditis, biliary tract infection, meningitis, and colorectal cancer, but their role as urinary pathogens is not well known. The objective of this investigation was to discover the incidence and clinical significance of the bacteriuria associated with this complex. A retrospective analysis of all adult patients with bacteriuria caused by SBG during the period 1995-2012 was carried out. During the study period, SBG was isolated in 153 adult patients, who had a mean age of 67 years, most of them being women (80%). Most of our patients (65%) had some underlying disease, with urologic disease being the most common (37%), followed by diabetes mellitus (27%) and neurologic disease (25%). Among the 88 patients in whom we were able to correctly assess symptoms, 45% had asymptomatic bacteriuria, 35% had lower urinary tract infection, and 20% had upper urinary tract infection. In 14 cases (9%), SBG was also isolated in blood cultures. Most of the isolates of SBG (72%) were S. gallolyticus subsp. pasteurianus. All isolates were susceptible to penicillin, 98% to nitrofurantoin, and 77% to fosfomycin. Although SBG bacteriuria is uncommon, it should not always be taken as a contaminant, mainly when S. pasteurianus is isolated, because it may cause urinary tract infections and, occasionally, sepsis, whereas when S. gallolyticus is isolated from urine, it may be a marker of underlying endocarditis and colorectal cancer.

In vitro evaluation of graphene oxide nanosheets on immune function.

HYPOTHESIS: Graphene oxide (GO) has attracted the scientific community attention due to its novel properties and wide range of potential applications including hyperthermia cancer therapy. However, little is known about the GO effects on the immune function which involves both innate and adaptive defence mechanisms through the activation of different cell populations and secretion of several cytokines. The effect of different GO nanosheets designed for hyperthermia cancer therapy on macrophage and lymphocyte function should be determined before using GO for this application. EXPERIMENTS: The effects of GO nanosheets with 1 (1-GOs) and 6 arms (6-GOs) of polyethylene glycol on RAW-264.7 macrophages and primary splenocytes (as approximation to the in vivo situation) were evaluated through the proinflammatory cytokine secretion and the modulation of cell proliferation in the presence of specific stimuli for either T-lymphocytes (concanavalin A, anti-CD3 antibody) or B-lymphocytes/macrophages (lipopolysaccharide). FINDINGS: 6-GOs significantly increased the secretion of TNF-α by RAW-264.7 macrophages without alteration of IL-6 and IL-1ß levels. The treatment of primary splenocytes with 1-GOs and 6-GOs in the presence of concanavalin A, anti-CD3 antibody and lipopolysaccharide, produced significant dose-dependent decreases of cell proliferation and IL-6 levels, revealing weak inflammatory properties of GOs which are favourable for hyperthermia cancer therapy.

Endocytic mechanisms of graphene oxide nanosheets in osteoblasts, hepatocytes and macrophages.

Nano-graphene oxide (GO) has attracted great interest in nanomedicine due to its own intrinsic properties and its possible biomedical applications such as drug delivery, tissue engineering and hyperthermia cancer therapy. However, the toxicity of GO nanosheets is not yet well-known and it is necessary to understand its entry mechanisms into mammalian cells in order to avoid cell damage and human toxicity. In the present study, the cellular uptake of pegylated GO nanosheets of ca. 100 nm labeled with fluorescein isothiocyanate (FITC-PEG-GOs) has been evaluated in the presence of eight inhibitors (colchicine, wortmannin, amiloride, cytochalasin B, cytochalasin D, genistein, phenylarsine oxide and chlorpromazine) that specifically affect different endocytosis mechanisms. Three cell types were chosen for this study: human Saos-2 osteoblasts, human HepG2 hepatocytes and murine RAW-264.7 macrophages. The results show that different mechanisms take part in FITC-PEG-GOs uptake, depending on the characteristics of each cell type. However, macropinocytosis seems to be a general internalization process in the three cell lines analyzed. Besides macropinocytosis, FITC-PEG-GOs can enter through pathways dependent on microtubules in Saos-2 osteoblasts, and through clathrin-dependent mechanisms in HepG2 hepatocytes and RAW-264.7 macrophages. HepG2 cells can also phagocytize FITC-PEG-GOs. These findings help to understand the interactions at the interface of GO nanosheets and mammalian cells and must be considered in further studies focused on their use for biomedical applications.

Triggering cell death by nanographene oxide mediated hyperthermia.

Graphene oxide (GO) has been proposed as an hyperthermia agent for anticancer therapies due to its near-infrared (NIR) optical absorption ability which, with its small two-dimensional size, could have a unique performance when compared to that of any other nanoparticle. Nevertheless, attention should be given to the hyperthermia route and the kind of GO-cell interactions induced in the process. The hyperthermia laser irradiation parameters, such as exposure time and laser power, were investigated to control the temperature rise and consequent damage in the GOs containing cell culture medium. The type of cell damage produced was evaluated as a function of these parameters. The results showed that cell culture temperature (after irradiating cells with internalized GO) increases preferentially with laser power rather than with exposure time. Moreover, when laser power is increased, necrosis is the preferential cell death leading to an increase of cytokine release to the medium.

Streptococcus bovis group and biliary tract infections: an analysis of 51 cases.

Streptococcus bovis is a well-known cause of endocarditis, but its role in other infections has not been well described. We analysed prospectively all patients with biliary tract infections caused by S. bovis group during the period 1988-2011. We selected those cases associated with cholangitis and cholecystitis, defined according to Tokyo guidelines. Identification of the strains was performed using the API 20 Strep and the GP card of the Vitek 2 system, and was confirmed by molecular methods. Our series included 51 cases (30 cholangitis and 21 cholecystitis). The associated microorganisms were: Streptococcus infantarius (biotype II/1) 29 cases (57%), Streptococcus gallolyticus subsp. pasteurianus (biotype II/2) 20 cases (39%) and Streptococcus gallolyticus subsp. gallolyticus (biotype I) two cases (4%). The only difference found between S. infantarius and S. gallolyticus subsp. pasteurianus was a greater association of the first with malignant strictures of the bile ducts: 48% (14/29) versus 5% (1/20), p <0.001. Thirty-seven of the cases also had bacteraemia, causing 20% (37/185) of all S. bovis bacteraemia, with differences between S. gallolyticus subsp. gallolyticus (2/112; 2%) and the other two microorganisms: S. infantarius and S. gallolyticus subsp. pasteurianus (35/73; 48%; p <0.001). The vast majority of biliary tract infections due to S. bovis group are caused by S. infantarius and S. gallolyticus subsp. pasteurianus (S. bovis biotype II), and nearly half of the bacteraemia due to these two species has a biliary source (43% of the S. infantarius and 56% of S. gallolyticus subsp. pasteurianus).

In vitro biocompatibility and antimicrobial activity of poly(ε-caprolactone)/montmorillonite nanocomposites.

A triblock copolymer based on poly(ε-caprolactone) (PCL) and 2-(N,N-diethylamino)ethyl methacrylate (DEAEMA)/2-(methyl-7-nitrobenzofurazan)amino ethyl acrylate (NBD-NAcri), was synthesized via atom transfer radical polymerization (ATRP). The corresponding chlorohydrated copolymer, named as PCL-b-DEAEMA, was prepared and anchored via cationic exchange on montmorillonite (MMT) surface. (PCL)/layered silicate nanocomposites were prepared through melt intercalation, and XRD and TEM analysis showed an exfoliated/intercalated morphology for organomodified clay. The surface characterization of the nanocomposites was undertaken by using contact angle and AFM. An increase in the contact angle was observed in the PCL/MMT(PCL-b-DEAEMA) nanocomposites with respect to PCL. The AFM analysis showed that the surface of the nanocomposites became rougher with respect to the PCL when MMTk10 or MMT(PCL-b-DEAEMA) was incorporated, and the value increased with the clay content. The antimicrobial activity of the nanocomposites against B. subtilis and P. putida was tested. It is remarkable that the biodegradation of PCL/MMT(PCL-b-DEAEMA) nanocomposites, monitored by the production of carbon dioxide and by chemiluminescence emission, was inhibited or retarded with respect to the PCL and PCL/1-MMTk10. It would indicate that nature of organomodifier in the clay play an important role in B. subtilis and P. putida adhesion processes. Biocompatibility studies demonstrate that both PCL and PCL/MMT materials allow the culture of murine L929 fibroblasts on its surface with high viability, very low apoptosis, and without plasma membrane damage, making these materials very adequate for tissue engineering.

Cell uptake survey of pegylated nanographene oxide.

Graphene and more specifically, nanographene oxide (GO) has been proposed as a highly efficient antitumoral therapy agent. Nevertheless, its cell uptake kinetics, its influence in different types of cells and the possibility of controlling cellular internalization timing, is still a field that remains unexplored. Herein, different cell types have been cultured in vitro for several incubation periods in the presence of 0.075 mg ml(-1) pegylated GO solutions. GO uptake kinetics revealed differences in the agent's uptake amount and speed as a function of the type of cell involved. Osteoblast-like cells GO uptake is higher and faster without resulting in greater cell membrane damage. Moreover, the dependence on the commonly used PEG nature (number of branches) also influences the viability and cell uptake speed. These facts play an important role in the future definition of timing parameters and selective cell uptake control in order to achieve an effective therapy.