Redefining CT perfusion-based ischemic core estimates for the ghost core in early time window stroke.

BACKGROUND AND PURPOSE: In large vessel occlusion (LVO) stroke patients, relative cerebral blood flow (rCBF)<30% volume thresholds are commonly used in treatment decisions. In the early time window, nearly infarcted but salvageable tissue volumes may lead to pretreatment overestimates of infarct volume, and thus potentially exclude patients who may otherwise benefit from intervention. Our multisite analysis aims to explore the strength of relationships between widely used pretreatment CT parameters and clinical outcomes for early window stroke patients. METHODS: Patients from two sites in a prospective registry were analyzed. Patients with LVOs, presenting within 3 hours of last known well, and who were successfully reperfused were included. Primary short-term neurological outcome was percent National Institutes of Health Stroke Scale (NIHSS) change from admission to discharge. Secondary long-term outcome was 90-day modified Rankin score. Spearman's correlations were performed. Significance was attributed to p-value ≤.05. RESULTS: Among 73 patients, median age was 66 (interquartile range 54-76) years. Among all pretreatment imaging parameters, rCBF<30%, rCBF<34%, and rCBF<38% volumes were significantly, inversely correlated with percentage NIHSS change (p<.048). No other parameters significantly correlated with outcomes. CONCLUSIONS: Our multisite analysis shows that favorable short-term neurological recovery was significantly correlated with rCBF volumes in the early time window. However, modest strength of correlations provides supportive evidence that the applicability of general ischemic core estimate thresholds in this subpopulation is limited. Our results support future larger-scale efforts to liberalize or reevaluate current rCBF parameter thresholds guiding treatment decisions for early time window stroke patients.

Interferon-gamma ameliorates experimental autoimmune encephalomyelitis by inducing homeostatic adaptation of microglia.

Compelling evidence has shown that interferon (IFN)-γ has dual effects in multiple sclerosis and in its animal model of experimental autoimmune encephalomyelitis (EAE), with results supporting both a pathogenic and beneficial function. However, the mechanisms whereby IFN-γ may promote neuroprotection in EAE and its effects on central nervous system (CNS)-resident cells have remained an enigma for more than 30 years. In this study, the impact of IFN-γ at the peak of EAE, its effects on CNS infiltrating myeloid cells (MC) and microglia (MG), and the underlying cellular and molecular mechanisms were investigated. IFN-γ administration resulted in disease amelioration and attenuation of neuroinflammation associated with significantly lower frequencies of CNS CD11b+ myeloid cells and less infiltration of inflammatory cells and demyelination. A significant reduction in activated MG and enhanced resting MG was determined by flow cytometry and immunohistrochemistry. Primary MC/MG cultures obtained from the spinal cord of IFN-γ-treated EAE mice that were ex vivo re-stimulated with a low dose (1 ng/ml) of IFN-γ and neuroantigen, promoted a significantly higher induction of CD4+ regulatory T (Treg) cells associated with increased transforming growth factor (TGF)-ß secretion. Additionally, IFN-γ-treated primary MC/MG cultures produced significantly lower nitrite in response to LPS challenge than control MC/MG. IFN-γ-treated EAE mice had a significantly higher frequency of CX3CR1high MC/MG and expressed lower levels of program death ligand 1 (PD-L1) than PBS-treated mice. Most CX3CR1highPD-L1lowCD11b+Ly6G- cells expressed MG markers (Tmem119, Sall2, and P2ry12), indicating that they represented an enriched MG subset (CX3CR1highPD-L1low MG). Amelioration of clinical symptoms and induction of CX3CR1highPD-L1low MG by IFN-γ were dependent on STAT-1. RNA-seq analyses revealed that in vivo treatment with IFN-γ promoted the induction of homeostatic CX3CR1highPD-L1low MG, upregulating the expression of genes associated with tolerogenic and anti-inflammatory roles and down-regulating pro-inflammatory genes. These analyses highlight the master role that IFN-γ plays in regulating microglial activity and provide new insights into the cellular and molecular mechanisms involved in the therapeutic activity of IFN-γ in EAE.

Nanotechnology-Based Drug Delivery Strategies to Repair the Mitochondrial Function in Neuroinflammatory and Neurodegenerative Diseases.

Mitochondria are vital organelles in eukaryotic cells that control diverse physiological processes related to energy production, calcium homeostasis, the generation of reactive oxygen species, and cell death. Several studies have demonstrated that structural and functional mitochondrial disturbances are involved in the development of different neuroinflammatory (NI) and neurodegenerative (ND) diseases (NI&NDDs) such as multiple sclerosis, Alzheimer's disease, Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis. Remarkably, counteracting mitochondrial impairment by genetic or pharmacologic treatment ameliorates neurodegeneration and clinical disability in animal models of these diseases. Therefore, the development of nanosystems enabling the sustained and selective delivery of mitochondria-targeted drugs is a novel and effective strategy to tackle NI&NDDs. In this review, we outline the impact of mitochondrial dysfunction associated with unbalanced mitochondrial dynamics, altered mitophagy, oxidative stress, energy deficit, and proteinopathies in NI&NDDs. In addition, we review different strategies for selective mitochondria-specific ligand targeting and discuss novel nanomaterials, nanozymes, and drug-loaded nanosystems developed to repair mitochondrial function and their therapeutic benefits protecting against oxidative stress, restoring cell energy production, preventing cell death, inhibiting protein aggregates, and improving motor and cognitive disability in cellular and animal models of different NI&NDDs.

Solitary Cystic Psoas Muscle Metastasis From Rectosigmoid Adenocarcinoma.

The most common subtype of colon cancer is colorectal adenocarcinoma. Compared with other subtypes, such as signet-ring and mucinous, colorectal adenocarcinoma has been found to have lower rates of metastasis. Approximately 20% of colorectal cancer cases present with metastatic disease on initial evaluation. The most common locations for metastasis are the liver, lung, peritoneum, bone, and extra-regional lymph nodes. Metastatic disease to the skeletal muscle, however, is considerably rare. We present a clinical case of a 52-year-old female found to have a cystic iliopsoas muscle metastasis from rectosigmoid adenocarcinoma, initially classified as an infected fluid collection.

Cellular and Virtualization Technologies for UAVs: An Experimental Perspective.

The Unmanned Aircraft System (UAS) ecosystem is exponentially growing in both recreational and professional fields to provide novel services and applications to consumers from multiple engineering fields. However, this technology has only scraped the surface of its potential, especially in those cases that require fast reaction times. Accordingly, the UAS Traffic Management (UTM) project aims at efficiently managing the air traffic for Unmanned Aerial Vehicle (UAV) operations, including those cases where UAVs might be remotely managed from a completely different geographical location. With these considerations in mind, this article presents a cellular-assisted UAVs testbed used to complete a mission managed beyond the radio line-of-sight (BRLoS), as well as introducing a virtualization platform for deploying services using containerization technology. In addition, the article conducts a communication performance evaluation in order to determine if the testbed equipment meets the requirements to carry out this BRLoS management. Finally, indoor flight operations are carried out to demonstrate the feasibility and proper operation of the testbed.

Integration of 5G Experimentation Infrastructures into a Multi-Site NFV Ecosystem.

Network Function Virtualization (NFV) has been regarded as one of the key enablers for the 5th Generation of mobile networks, or 5G. This paradigm allows to reduce the dependence on specialized hardware to deploy telecommunications and vertical services. To this purpose, it relies on virtualization techniques to softwarize network functions, simplifying their development and reducing deployment time and costs. In this context, Universidad Carlos III de Madrid, Telefónica, and IMDEA Networks Institute have developed an NFV ecosystem inside 5TONIC, an open network innovation center focused on 5G technologies, enabling the creation of complex, close to reality experimentation scenarios across a distributed set of NFV infrastructures, which can be made available by stakeholders at different geographic locations. This article presents the protocol that has been defined to incorporate new remote NFV sites into the multi-site NFV ecosystem based on 5TONIC, describing the requirements for both the existing and the newly incorporated infrastructures, their connectivity through an overlay network architecture, and the steps necessary for the inclusion of new sites. The protocol is exemplified through the incorporation of an external site to the 5TONIC NFV ecosystem. Afterwards, the protocol details the verification steps required to validate a successful site integration. These include the deployment of a multi-site vertical service using a remote NFV infrastructure with Small Unmanned Aerial Vehicles (SUAVs). This serves to showcase the potential of the protocol to enable distributed experimentation scenarios.

Intranasal delivery of interferon-ß-loaded nanoparticles induces control of neuroinflammation in a preclinical model of multiple sclerosis: A promising simple, effective, non-invasive, and low-cost therapy.

Multiple sclerosis (MS) is an autoimmune disease affecting the central nervous system (CNS). Interferon (IFN)-ß constitutes one of the first-line therapies to treat MS, but has limited efficacy due to the injectable systemic administration, short half-life, and limited CNS access. To address these limitations, we developed IFN-ß-loaded chitosan/sulfobutylether-ß-cyclodextrin nanoparticles (IFN-ß-NPs) for delivery of IFN-ß into the CNS via the intranasal (i.n.) route. The nanoparticles (NPs) (≈200 nm, polydispersity ≈0.1, and zeta potential ≈20 mV) were prepared by mixing two aqueous solutions and associated human or murine IFN-ß with high efficiency (90%). Functional in vitro assays showed that IFN-ß-NPs were safe and that IFN-ß was steadily released while retaining biological activity. Biodistribution analysis showed an early and high fluorescence in the brain after nasal administration of fluorescent probe-loaded NPs. Remarkably, mice developing experimental autoimmune encephalomyelitis (EAE), an experimental model of MS, exhibited a significant improvement of clinical symptoms in response to intranasal IFN-ß-NPs (inIFN-ß-NPs), whereas a similar dose of intranasal or systemic free IFN-ß had no effect. Importantly, inIFN-ß-NPs treatment was equally effective despite a reduction of 78% in the total amount of weekly administered IFN-ß. Spinal cords obtained from inIFN-ß-NPs-treated EAE mice showed fewer inflammatory foci and demyelination, lower expression of antigen-presenting and costimulatory proteins on CD11b+ cells, and lower astrocyte and microglia activation than control mice. Therefore, IFN-ß treatment at tested doses was effective in promoting clinical recovery and control of neuroinflammation in EAE only when associated with NPs. Overall, inIFN-ß-NPs represent a potential, effective, non-invasive, and low-cost therapy for MS.

Las ciencias y la pandemia del Covid-19 / The sciences and the Covid-19 Pandemic

Durante la etapa de confinamiento social decretado para evitar el contagio de la pandemia del COVID-19, se ha redactado el presente ensayo partiendo de ejercitar la crítica a posiciones que desde algunas instituciones científicas -las academias de ciencias nacionales, en particular- se han tomado con relación a la situación actual que se vive en el país y a las decisiones tomadas por las autoridades gubernamentales para evitar la propagación de la epidemia en el territorio nacional. El cuerpo del ensayo busca mostrar, con la presentación de algunos de los campos científicos asociados a la atención científica de la pandemia ­ la virología, la epidemiología y la medicina-, la complejidad, las inconmensurabilidades y las posibles colisiones que existen entre ellas, situación que dificulta la obtención de soluciones apoyadas solo en las ciencias. Se concluye llamando la atención sobre algunas enseñanzas que se pueden desprender de la situación que ha tocado vivir en casi todos los países del mundo y en particular, el papel que Estado le ha tocado manejar en Venezuela(AU)

During stage social confine-ment decreed to avoid CO-VID-19 pandemic spread, this essay has been written basis on exercising criticism positions that from some scientific insti-tutions -the national science academies in particular- they have taken about country sit-uation current and decisions made by government authorities to prevent epidemic spread in the national territory. Essay body seeks to show, with the presentation of some scientific fields associated with the scientific care of the pandemic -virology, epidemiology, and medicine-, the complexity, the incommensurabilities and the possible collisions that exist between them, situation that makes it difficult to obtain solutions supported only in the sciences. It concludes by calling attention to some lessons that can be derived from situation that has had to be lived in almost all world countries and, in particular, the role that the State has had to deal with in Venezuela(AU)

Automated Deployment of an Internet Protocol Telephony Service on Unmanned Aerial Vehicles Using Network Functions Virtualization.

The Network Function Virtualization (NFV) paradigm is one of the key enabling technologies in the development of the 5th generation of mobile networks. This technology aims to lessen the dependence on hardware in the provision of network functions and services by using virtualization techniques that allow the softwarization of those functionalities over an abstraction layer. In this context, there is increasing interest in exploring the potential of unmanned aerial vehicles (UAVs) to offer a flexible platform capable of enabling cost-effective NFV operations over delimited geographic areas. To demonstrate the practical feasibility of utilizing NFV technologies in UAV platforms, a protocol is presented to set up a functional NFV environment based on open source technologies, in which a set of small UAVs supply the computational resources that support the deployment of moderately complex network services. Then, the protocol details the different steps needed to support the automated deployment of an internet protocol (IP) telephony service over a network of interconnected UAVs, leveraging the capacities of the configured NFV environment. Experimentation results demonstrate the proper operation of the service after its deployment. Although the protocol focuses on a specific type of network service (i.e., IP telephony), the described steps may serve as a general guide to deploy other type of network services. On the other hand, the protocol description considers concrete equipment and software to set up the NFV environment (e.g., specific single board computers and open source software). The utilization of other hardware and software platforms may be feasible, although the specific configuration aspect of the NFV environment and the service deployment may present variations with respect to those described in the protocol.

Transport-Layer Limitations for NFV Orchestration in Resource-Constrained Aerial Networks.

In this paper, we identify the main challenges and problems related with the management and orchestration of Virtualized Network Functions (VNFs) over aerial networks built with Small Unmanned Aerial Vehicles (SUAVs). Our analysis starts from a reference scenario, where several SUAVs are deployed over a delimited geographic area, and provide a mobile cloud environment that supports the deployment of functions and services using Network Functions Virtualization (NFV) technologies. After analyzing the main challenges to NFV orchestration in this reference scenario from a theoretical perspective, we undertake the study of one specific but relevant aspect following a practical perspective, i.e., the limitations of existing transport-layer solutions to support the dissemination of NFV management and orchestration information in the considered scenario. While in traditional cloud computing environments this traffic is delivered using TCP, our simulation results suggest that using this protocol over an aerial network of SUAVs presents certain limitations. Finally, based on the lessons learned from our practical analysis, the paper outlines different alternatives that could be followed to address these challenges.

Neurochemical and behavioral characterization of neuronal glutamate transporter EAAT3 heterozygous mice.

BACKGROUND: Obsessive-compulsive disorder (OCD) is a severe neuropsychiatric condition affecting 1-3% of the worldwide population. OCD has a strong genetic component, and the SLC1A1 gene that encodes neuronal glutamate transporter EAAT3 is a strong candidate for this disorder. To evaluate the impact of reduced EAAT3 expression in vivo, we studied male EAAT3 heterozygous and wild-type littermate mice using a battery of behavioral paradigms relevant to anxiety (open field test, elevated plus maze) and compulsivity (marble burying), as well as locomotor activity induced by amphetamine. Using high-performance liquid chromatography, we also determined tissue neurotransmitter levels in cortex, striatum and thalamus-brain areas that are relevant to OCD. RESULTS: Compared to wild-type littermates, EAAT3 heterozygous male mice have unaltered baseline anxiety-like, compulsive-like behavior and locomotor activity. Administration of acute amphetamine (5 mg/kg intraperitoneally) increased locomotion with no differences across genotypes. Tissue levels of glutamate, GABA, dopamine and serotonin did not vary between EAAT3 heterozygous and wild-type mice. CONCLUSIONS: Our results indicate that reduced EAAT3 expression does not impact neurotransmitter content in the corticostriatal circuit nor alter anxiety or compulsive-like behaviors.

Respiratory viruses detected in Mexican children younger than 5 years old with community-acquired pneumonia: a national multicenter study.

BACKGROUND: Acute respiratory infections are the leading cause of mortality in children worldwide, especially in developing countries. Pneumonia accounts for 16% of all deaths of children under 5 years of age and was the cause of death of 935000 children in 2015. Despite its frequency and severity, information regarding its etiology is limited. The aim of this study was to identify respiratory viruses associated with community-acquired pneumonia (CAP) in children younger than 5 years old. METHODS: One thousand four hundred and four children younger than 5 years of age with a clinical and/or radiological diagnosis of CAP in 11 hospitals in Mexico were included. Nasal washes were collected, placed in viral medium, and frozen at -70°C until processing. The first 832 samples were processed using the multiplex Bio-Plex/Luminex system and the remaining 572 samples using the Anyplex multiplex RT-PCR. Clinical data regarding diagnosis, clinical signs and symptoms, radiographic pattern, and risk factors were obtained and recorded. RESULTS: Of the samples tested, 81.6% were positive for viruses. Respiratory syncytial virus (types A and B) was found in 23.7%, human enterovirus/rhinovirus in 16.6%, metapneumovirus in 5.7%, parainfluenza virus (types 1-4) in 5.5%, influenza virus (types A and B) in 3.6%, adenovirus in 2.2%, coronavirus (NL63, OC43, 229E, and HKU1) in 2.2%, and bocavirus in 0.4%. Co-infection with two or more viruses was present in 22.1%; 18.4% of the samples were negative. Using biomass for cooking, daycare attendance, absence of breastfeeding, and co-infections were found to be statistically significant risk factors for the presence of severe pneumonia. CONCLUSIONS: Respiratory syncytial virus (types A and B), human enterovirus/rhinovirus, and metapneumovirus were the respiratory viruses identified most frequently in children younger than 5 years old with CAP. Co-infection was present in an important proportion of the children.

Association of vitamin D deficiency, season of the year, and latent tuberculosis infection among household contacts.

OBJECTIVES: Vitamin D (VD) enhances the immune response against Mycobacterium tuberculosis in vitro, and VD deficiency has been described in patients with active tuberculosis (TB). However, the role of hypovitaminosis D in the pathogenesis of early TB infection acquisition is unclear. We aimed to evaluate the association of VD deficiency, season of the year, and latent TB infection in household contacts (HHC), given that this is a potentially modifiable condition often related to nutritional deficiencies and lack of sun exposure. METHODS: We prospectively enrolled new pulmonary TB cases (n = 107) and their HHC (n = 144) over a 2-year period in Santiago, Chile. We compared plasma 25-hydroxycholecalciferol (25OHD) levels and examined the influence of season, ethnic background, living conditions, and country of origin. RESULTS: Over 77% of TB cases and 62.6% of HHC had VD deficiency (<20 ng/ml). Median 25OHD concentration was significantly lower in TB cases than in HHC (11.7 vs. 18.2 ng/ml, p<0.0001). Migrants HHC had lower 25OHD levels than non-migrants (14.6 vs. 19.0 ng/ml, p = 0.026), and a trend towards a higher burden of latent TB infection (52.9% vs. 35.2%, p = 0.066). Multivariate analysis found VD deficiency in HHC was strongly associated with being sampled in winter/spring (adOR 25.68, 95%CI 7.35-89.7), corresponding to the seasons with lowest solar radiation exposure. Spring enrollment-compared with other seasons-was the chief risk factor for latent TB infection in HHC (adOR 3.14, 95%CI 1.28-7.69). CONCLUSIONS: Hypovitaminosis D was highly prevalent in TB cases and also in HHC. A marked seasonality was found for both VD levels and latent TB in HHC, with winter being the season with lowest VD levels and spring the season with the highest risk of latent TB infection.

Neurochemical and behavioral characterization of neuronal glutamate transporter EAAT3 heterozygous mice

BACKGROUND: Obsessive-compulsive disorder (OCD) is a severe neuropsychiatric condition affecting 1-3% of the worldwide population. OCD has a strong genetic component, and the SLC1A1 gene that encodes neuronal glutamate transporter EAAT3 is a strong candidate for this disorder. To evaluate the impact of reduced EAAT3 expression in vivo, we studied male EAAT3 heterozygous and wild-type littermate mice using a battery of behavioral paradigms relevant to anxiety (open field test, elevated plus maze) and compulsivity (marble burying), as well as locomotor activity induced by amphetamine. Using high-performance liquid chromatography, we also determined tissue neurotransmitter levels in cortex, striatum and thalamus-brain areas that are relevant to OCD. RESULTS: Compared to wild-type littermates, EAAT3 heterozygous male mice have unaltered baseline anxiety-like, compulsive-like behavior and locomotor activity. Administration of acute amphetamine (5 mg/kg intraperitoneally) increased locomotion with no differences across genotypes. Tissue levels of glutamate, GABA, dopamine and serotonin did not vary between EAAT3 heterozygous and wild-type mice. CONCLUSIONS: Our results indicate that reduced EAAT3 expression does not impact neurotransmitter content in the corticostriatal circuit nor alter anxiety or compulsive-like behaviors.

Programming of Dopaminergic Neurons by Neonatal Sex Hormone Exposure: Effects on Dopamine Content and Tyrosine Hydroxylase Expression in Adult Male Rats.

We sought to determine the long-term changes produced by neonatal sex hormone administration on the functioning of midbrain dopaminergic neurons in adult male rats. Sprague-Dawley rats were injected subcutaneously at postnatal day 1 and were assigned to the following experimental groups: TP (testosterone propionate of 1.0 mg/50 µL); DHT (dihydrotestosterone of 1.0 mg/50 µL); EV (estradiol valerate of 0.1 mg/50 µL); and control (sesame oil of 50 µL). At postnatal day 60, neurochemical studies were performed to determine dopamine content in substantia nigra-ventral tegmental area and dopamine release in nucleus accumbens. Molecular (mRNA expression of tyrosine hydroxylase) and cellular (tyrosine hydroxylase immunoreactivity) studies were also performed. We found increased dopamine content in substantia nigra-ventral tegmental area of TP and EV rats, in addition to increased dopamine release in nucleus accumbens. However, neonatal exposure to DHT, a nonaromatizable androgen, did not affect midbrain dopaminergic neurons. Correspondingly, compared to control rats, levels of tyrosine hydroxylase mRNA and protein were significantly increased in TP and EV rats but not in DHT rats, as determined by qPCR and immunohistochemistry, respectively. Our results suggest an estrogenic mechanism involving increased tyrosine hydroxylase expression, either by direct estrogenic action or by aromatization of testosterone to estradiol in substantia nigra-ventral tegmental area.

Unmanned Aerial Vehicles (UAVs) and Artificial Intelligence Revolutionizing Wildlife Monitoring and Conservation.

Surveying threatened and invasive species to obtain accurate population estimates is an important but challenging task that requires a considerable investment in time and resources. Estimates using existing ground-based monitoring techniques, such as camera traps and surveys performed on foot, are known to be resource intensive, potentially inaccurate and imprecise, and difficult to validate. Recent developments in unmanned aerial vehicles (UAV), artificial intelligence and miniaturized thermal imaging systems represent a new opportunity for wildlife experts to inexpensively survey relatively large areas. The system presented in this paper includes thermal image acquisition as well as a video processing pipeline to perform object detection, classification and tracking of wildlife in forest or open areas. The system is tested on thermal video data from ground based and test flight footage, and is found to be able to detect all the target wildlife located in the surveyed area. The system is flexible in that the user can readily define the types of objects to classify and the object characteristics that should be considered during classification.

Rhinovirus is an important pathogen in upper and lower respiratory tract infections in Mexican children.

BACKGROUND: Most of the studies characterizing the incidence of rhinovirus (RV) have been carried out in hospitalized children and in developed countries. In those studies, RV-C has been associated with more severe respiratory tract infections than RV species A and B. In this study we determined the frequency and diversity of RV strains associated with upper and lower respiratory tract infections (URTI, LRTI) in Mexico, and describe the clinical characteristics of the illness associated with different RV species. METHODS: A prospective surveillance of 526 and 250 children with URTI and LRTI was carried out. Respiratory samples were analyzed by RT-PCR for viruses. The 5' untranslated region of the RV genome was amplified and sequenced. RESULTS: In the case of URTI, 17.5% were positive for RV, while this virus was found in 24.8% of LRTI. The RV species was determined in 73 children with URTI: 61.6% were RV-A, 37% RV-C and, 1.4% RV-B; and in 43 children with LRTI: 51.2% were RV-A, 41.8% RV-C, and 7% RV-B. No significant differences in clinical characteristics were found in patients with RV-A or RV-C infections. A high genetic diversity of RV strains was found in both URTI and LRTI. CONCLUSIONS: Both RV-A and RV-C species were frequently found in hospitalized as well as in outpatient children. This study underlines the high prevalence and genetic diversity of RV strains in Mexico and the potential severity of disease associated with RV-A and RV-C infections.

Is there still room for novel viral pathogens in pediatric respiratory tract infections?

Viruses are the most frequent cause of respiratory disease in children. However, despite the advanced diagnostic methods currently in use, in 20 to 50% of respiratory samples a specific pathogen cannot be detected. In this work, we used a metagenomic approach and deep sequencing to examine respiratory samples from children with lower and upper respiratory tract infections that had been previously found negative for 6 bacteria and 15 respiratory viruses by PCR. Nasal washings from 25 children (out of 250) hospitalized with a diagnosis of pneumonia and nasopharyngeal swabs from 46 outpatient children (out of 526) were studied. DNA reads for at least one virus commonly associated to respiratory infections was found in 20 of 25 hospitalized patients, while reads for pathogenic respiratory bacteria were detected in the remaining 5 children. For outpatients, all the samples were pooled into 25 DNA libraries for sequencing. In this case, in 22 of the 25 sequenced libraries at least one respiratory virus was identified, while in all other, but one, pathogenic bacteria were detected. In both patient groups reads for respiratory syncytial virus, coronavirus-OC43, and rhinovirus were identified. In addition, viruses less frequently associated to respiratory infections were also found. Saffold virus was detected in outpatient but not in hospitalized children. Anellovirus, rotavirus, and astrovirus, as well as several animal and plant viruses were detected in both groups. No novel viruses were identified. Adding up the deep sequencing results to the PCR data, 79.2% of 250 hospitalized and 76.6% of 526 ambulatory patients were positive for viruses, and all other children, but one, had pathogenic respiratory bacteria identified. These results suggest that at least in the type of populations studied and with the sampling methods used the odds of finding novel, clinically relevant viruses, in pediatric respiratory infections are low.