The drug of choice for the treatment of HSV encephalitis is high-dose intravenous acyclovir which should be administered as early as possible for 14 to 21 days. A clinical trial is currently assessing longer courses of therapy using oral valacyclovir. There are no clinical trials regarding the use of antivirals for VZV encephalitis but acyclovir for up to 3 weeks is recommended for severe infections like encephalitis. A longer course of therapy may be considered for immunocompromised patients. Foscarnet is the preferred agent against HHV-6 whereas combination therapy with foscarnet and ganciclovir is recommended as initial treatment of CMV encephalitis (Table 1).

Antivirals have not been proven effective for enterovirus encephalitis. The drug pleconaril is an inhibitor of viral replication and may be an option for patients with severe Enterovirus infections. Use of oseltamivir is appropriate for severe influenza. There is also no specific treatment for most causes of encephalitis although experimental therapies may be considered [13, 113].

There is a paucity of data regarding treatment and management of viral infection. Supportive care is the current mainstay of therapy for most viral infections, particularly for respiratory viruses. Though broad-spectrum antibiotic therapy may be prudent until a bacterial source for sepsis has been definitively ruled-out, sustained antibiotic treatment has no role in the management of viral sepsis except in the case of bacterial coinfections. Many viral infections can be prevented with the use of hand hygiene, environmental decontamination, use of personal protective equipment, elimination of second-hand smoke, and isolation of infected children (131). Additional protection can be conferred by administering vaccines for common communicable viruses. These preventive strategies are of particular importance in high-risk patients. As the scope of available vaccines and anti-viral therapies remains rather limited, development of novel vaccines and treatment is critical (131).

For RSV infection, management is currently limited to passive immunization for at-risk infants. Palivizumab, an RSV-specific monoclonal antibody, is Food and Drug Administration (FDA) approved for the prevention of infection in high-risk infants during RSV season. The American Academy of Pediatrics has issued more clear recommendations for palivizumab use, stating that it should be administered as a monthly injection during RSV season in children born less than 29 weeks, 0 days gestation and are less than 12 months of age or in children with congenital heart disease, chronic lung disease (132). Studies have shown variable efficacy of palivizumab, with reduction in RSV hospitalization rate by approximately 60% (133). Currently, aerosolized ribavirin is the only FDA-approved treatment available for the management of RSV infection, though its use remains controversial (134). To date, RSV vaccines and antiviral therapies remain an active area of investigation (135). A randomized, controlled trial performed in adult patients with RSV infection compared the RSV entry inhibitor GS-5806 to placebo and demonstrated a decrease in both viral load and the clinical severity of infection in patients treated with GS-5806 (136). Similar fusion inhibitors such as ALX-0171 (137), JNJ-2408068 (138), MDT-637 (139), and VP14637 (138) demonstrate efficacy in vitro, and ALX-0171 is undergoing a phase II clinical trial in infants hospitalized for RSV (clinicaltrials.gov registration no. NCT02979431). The use of ALS-008176, an RSV polymerase inhibitor, has similarly been shown to reduce viral load, rapidly clear RSV, and improve the severity of disease in adults with RSV infection (140). ALN-RSV01 is a lipid-based nanoparticulate system, containing small-interfering RNA (siRNA] that demonstrates promising antiviral effects against RSV in lung transplant patients (141) by targeting the mRNA of the RSV nucleocapsid protein, thereby limiting viral replication (142). However, until these novel treatments have undergone appropriate clinical trials, the pediatric medical community must continue to wait for effective RSV antiviral therapy.

Unlike RSV, seasonal vaccines and several antiviral therapies are available to treat influenza viral infections. The seasonal influenza vaccine has demonstrated reasonable efficacy at attenuating influenza A and B viral disease (143). Currently, two forms of the influenza vaccine are available for use in children: a live attenuated vaccine in the form of a nasal spray and an inactivated vaccine in an injectable form. Antiviral agents used in the treatment and post-exposure prophylaxis of influenza infections include neuraminidase inhibitors (oseltamivir and zanamivir) and the adamantanes (amantadine and rimantadine). Oseltamivir is the most commonly used medication due to high prevalence of adamantane resistance. Oseltamivir has shown to be beneficial and tolerable in children with influenza if received within first 48 h of illness (144, 145). However, in cases of severe infection, initiation of oseltamivir beyond 48 h of symptom onset may still provide benefit (146). Nanotechnology-based vaccines are also being developed for influenza virus. InflexalR V and InfluvacR are two virosomal vaccines that have been shown to be efficacious against influenza infection (147, 148). STP702, another nanotherapeutic agent, is an siRNA under development designed to inhibit conserved regions in H1N1 and H5N1 strains of the influenza virus and prevent viral replication (116). Nanotraps such as sialylneolacto-N-tetraose c (LSTc)-bearing liposomal decoys bind to hemagglutinins on the influenza virus and prevent viral spread in vitro, demonstrating the potential have shown to be effective against influenza virus (117). The influenza polymerase inhibitor T-705 (favipiravir) has been demonstrated significant attenuation of influenza virus activity (149). Interestingly, at higher concentration, it has also shown to be effective against poliovirus, rhinovirus and RSV (149). Other agents under investigation include CS-8958, a long-acting neuraminidase inhibitor, and DAS181, an attachment inhibitor (150). Animal studies have also shown promising results with combination therapy (150). Various immunomodulatory agents have also been posited to temper the dysregulated host inflammatory response in severe influenza (151), including cyclooxygenase-2 inhibitors (152), doxycycline (153), glucocorticoids (154), macrolides (155, 156), peroxisome proliferator-activated receptor agonists such as gemfibrozil (157), sphingosine-1-phosphate (158), and the Tie2 receptor activator vasculotide (65). Further studies are needed to determine the efficacy of these treatments in human influenza infection.

Pleconaril, an orally administered viral capsid inhibitor, has shown to be effective against picornaviruses, especially enteroviruses and rhinoviruses (159). Abzug et al. reported greater survival in patients with neonatal enteroviral sepsis who received pleconaril (159). Similarly, patient with rhinovirus infection treated with pleconaril have shorter duration of symptoms, depending on susceptibility of the virus to the medication (160). No antiviral activity has been observed from pleconaril against HPeV (18). Intravenous immunoglobulin has shown potential benefit in management of enteroviral infections (161).

Prevention of neonatal HSV infection is more elusive as neonatal HSV disease often occurs after transmission from asymptomatic women with primary HSV infection (162). In cases of active maternal genital herpes, cesarean sections can decrease the incidence of neonatal HSV infection, especially when performed within 4 h of rupture of membranes (163). A subunit HSV vaccine has shown promising results in prevention of genital herpes and is currently under Phase III trial (164). Although not routinely recommended, antiviral prophylaxis with acyclovir in late pregnancy has been demonstrated to decrease viral shedding, leading to reduction in cesarean rates and recurrent herpes (165, 166). Patients with severe neonatal HSV infection (those with disseminated disease and CNS infection) should be treated with intravenous acyclovir for 21 days (167).

Viral sepsis may occur in HIV-infected children due to opportunistic or other secondary viral infections (19). Increasing use of highly active antiretroviral therapy (HAART) has significantly improved survival of HIV infected children by decreasing the progression to acquired immunodeficiency syndrome (AIDS), thereby maintaining host immunocompetence that protects against the development of viral sepsis (168–171). However, HAART is associated with potentially deleterious sequelae, making timing of the therapy very controversial in patients with active sepsis (19).

The need for informed consent was waived in view of the observational nature of the study with no interventions performed. The protocol and standardized clinical form, including the waiver of informed consent, were approved by the Asan Medical Center Institutional Review Board (IRB number: 2010-0079).

Of the 172 that tested positive for respiratory agents, there were 24 (14.0%) cases with mixed infections of two or three viruses. No patient was infected with more than three viruses. Twenty-one cases were infected with two viruses, with the most frequent mixture being adenovirus and influenza A virus (n = 5). Other frequent viral mixtures were adenovirus and rhinovirus A (n =3) and influenza A virus and coronavirus OC43 (n =3) (Table 3). Adenovirus was present in 62.5% (n = 15) of mixed infections which was almost a half (46.9%) of all detections for this virus. Respiratory syncytial virus B was the only aetiology that was not detected in a mixed infection.

Respiratory virus infections have a major impact on health. Acute respiratory illnesses, mostly caused by viruses, are the most common illness experienced by otherwise healthy children and adults worldwide. Upper respiratory tract infections (URIs) such as common cold are exceedingly prevalent in infants and young children and continue to be common in older children and adults. Infants and young children may have 3–8 episodes of cold per year; those who attend daycare centers may have many more episodes per year [1–4]. URI can lead to complications such as acute otitis media, asthma exacerbation, and lower respiratory tract infections (LRIs). While LRIs such as pneumonia, bronchitis, and bronchiolitis occur much less frequently, they cause higher morbidity and some mortality, thus they are associated with high impact and greater healthcare costs. Approximately one third of children develop an LRI in the first year of life; LRI incidence decreases to 5%–10% during early school year, and 5% during preadolescent to healthy adult years [5, 6].

Common respiratory viruses include influenza A and B, respiratory syncytial virus A and B, parainfluenza virus types 1–3, adenovirus, rhinovirus, human metapneumovirus, and coronavirus types OC43 and 229E. Less common respiratory viruses include parainfluenza virus type 4, influenza virus C, and specific types of enteroviruses. The significance of more recently discovered viruses such as human bocavirus, coronavirus NL63, and HKU1 has yet to be elucidated [7, 8].

Clinical presentations of respiratory virus infections overlap among those caused by various viruses. In addition, clinical manifestations may mimic those of diseases caused by bacteria. Therefore, antibiotics are most often used in these infections, most of them unnecessarily. Furthermore, LRIs often require hospitalization for management such as intravenous antibiotics and symptomatic and supportive treatment. Specific antiviral treatment for respiratory virus infections is only available for influenza. Respiratory viral diagnosis is an integral part of patient management. Accurate diagnosis of specific respiratory virus infection not only improves the knowledge of disease the patient has but also can affect patient management and help prevent secondary spread of the infection. Rapid viral diagnosis may result in discontinuation of unnecessary antibiotics, initiation of antiviral drug for influenza, reduction of costs related to reduction of unnecessary investigations, and shortened hospital stay [9–11].

This paper describes up-to-date information on laboratory methods presently available in the diagnostic virology laboratories and those upcoming for detection of respiratory viruses.

The study was performed at a medical ICU of the Asan Medical Center, a tertiary referral hospital in Seoul, Republic of Korea. This university-affiliated teaching hospital has 2700 beds and eight ICUs. During the study period, most of the adult patients with severe HAP requiring ICU care were referred to the medical ICU. The medical ICU is a closed 28-bed unit managed by five board-certified intensivists. All intensivists attend structured twice daily bedside rounds. Fiberoptic bronchoscopy with bronchoalveolar lavage (BAL) was preferably performed on patients with bilateral interstitial pattern infiltration or non-resolving pneumonia, at the discretion of the physician’s judgment. The BAL protocol has been described in detail elsewhere.

This study reported a high prevalence of respiratory viruses in children ≤ 5 years using a custom assay and an FTD assay. Good concordance was observed for all the viruses between both assays except for RSVA/B. However larger numbers of positive samples need to be tested for thorough evaluation of less prevalent viruses. The custom primer and probe mix was much more economical than the commercial FTD kit. Our study suggests that this custom multiplex real-time RT-PCR can be used for simultaneous and rapid detection of multiple viruses in resource limited settings. This will help prevent unnecessary use of antibiotics and permit timely initiation of supportive therapy/antiviral drugs if available.

Treatment consists of supportive care as there are no licensed antivirals against HMPV. Two potential treatments that have been investigated are ribavirin and immunoglobulin. Ribavarin is a nucleoside with activity against RNA viruses and exhibits

in vitro activity against HMPV

119 and exhibited some efficacy in mice

120. Commercial intravenous immunoglobulin (IVIG) contains neutralizing activity against HMPV

119, and as noted above, antibodies alone exhibit efficacy both prophylactically and therapeutically in mice

93–

96. There are anecdotal reports of human use of ribavirin and IVIG

121 but no controlled trials and no guidelines to recommend the use of these measures.

For patient descriptions, we compared RSV-positive with RSV-negative patients within two groups of patients: all ARI patients and ARI patients with pneumonia. For the assessment of predictive factors for severity, patients with severe outcomes were compared with patients with no severe outcome within 6 groups of patients: all ARI patients, RSV-positive ARI patients, RSV-negative ARI patients, and those same three groups in patients less than 2-year old. Death, stay in ICU, oxygen use and severe pneumoniae were considered as indicators for severity. However, the mortality rate was too low to be analyzed and the analyses of ICU stay and oxygen use did not provide any added-value, so ultimately severe pneumonia alone was analyzed as an indicator of severe outcome. Data were double entered into an Access database (Microsoft Corporation). Statistical analysis was performed using Statistical Package for the Social Sciences version 23.0 for Windows (SPSS Inc., Chicago, IL, USA). For comparison of categorical data, the Pearson Chi-square (χ2) test and Fisher’s exact test were used as appropriate. The ANOVA test was applied to compare continuous variables. We fitted a binary logistic regression model, including a stepwise selection procedure, to assess predictive factors for severity. The level of significance was set at p < 0.05.

The tracheal aspirate specimens were first treated with an equal volume of Sputasol solution (Oxoid, Basingstok, UK) in a 37 °C water bath for liquefaction. The total RNA/DNA was then extracted from the liquefaction sample using the QIAamp Viral RNA Mini Kit (Qiagen, Hilden, Germany) per the manufacturer's instructions. The nucleic acids was stored in aliquots at −80 °C until use.

This study was approved by Nepal Health Research Council (reg. no. 180/2015).

All ethical issues including plagiarism, Informed Consent, misconduct, data fabrication and/or falsification, double publication and/or submission, redundancy, etc have been completely observed by the authors.

Perhaps one of the most significant advances in the detection of respiratory viruses is the recent introduction of novel swab types such as flocked swabs. Flocked swabs consist of nylon fibers perpendicular to the swab shaft. Unlike standard woven rayon swabs which trap sample material, the fibers of flocked swabs act like a brush to collect respiratory epithelial cells and efficiently release them in transport medium. Studies comparing nasal flocked swabs to nasal aspirates from children showed that the sensitivity of flocked swabs was at least equivalent for the detection of a variety of respiratory viruses [13, 14]. Similar studies comparing flocked swabs to woven rayon swabs in adult patients are needed. Another type of novel swab material is polyurethane foam. A recent study demonstrated that polyurethane foam swabs of the anterior nares performed better than flocked swabs for detection of influenza viruses in children by a rapid antigen test.

Acute respiratory tract infections are major causes of morbidity and mortality. In 2000, lower respiratory tract infections were globally the number one infectious cause of disability adjusted life-years. The commonest respiratory viruses that cause acute upper and lower respiratory tract infections and which are routinely tested for in most virus diagnostic laboratories are: influenza A virus (FLA); influenza B virus (FLB); respiratory syncytial virus (RSV); parainfluenza virus type 1 (PF1); parainfluenza virus type 2 (PF2); parainfluenza virus type 3 (PF3) and adenovirus (ADV). Additionally, human rhinoviruses (HRV) and coronavirus 229E (CoV-229E) are also linked to acute respiratory infection but less commonly included in laboratory reports; human metapneumovirus (hMPV) is not yet part of most United Kingdom virus laboratory test repertoires (personal feed-back from the United Kingdom Clinical Virology Network).

As part of service development it was necessary to provide an alternative to virus culture for testing immunofluorescence negative respiratory specimens. Historically and indeed currently immunofluorescence and virus culture are the main methods used to diagnose acute respiratory virus infections. Culture is accepted as more sensitive than immunofluorescence but slower and therefore less useful for direct patient management decisions. Using a standard culture technique for the culture of respiratory viruses our median reporting times for culture positive and culture negative specimens were 6 days (based on 407 specimens) and 7 days (based on 2159 specimens) respectively; virus identification by this technique required the use of monoclonal antibody staining of the cell monolayer in addition to observation for viral cytopathic effect. We therefore wished to develop a test capable of reporting on immunofluorescence negative specimens within a 24 hour period.

Increasingly however, the sensitivity of nucleic acid amplification techniques for diagnosis has become recognised. However widespread concerns about contamination issues and perceived cost have slowed their widespread adoption. An added problem for acute respiratory tract infections is the relatively large number of viruses that need to be accounted for, a problem which presents specific technical challenges.

One such challenge is the different optimal annealing temperatures of the primer sets for each prospective virus target. The ABI PRISM 7000 real-time facility from Applied Biosystems addresses this by using bundled software to design primer/probe combinations that use a common amplification protocol. However this approach is compromised by the inability of software to allow for target heterogeneity. In addition it does not allow users to adopt clinically validated primer sets from the literature.

To address these problems we adopted an alternative approach through the development of a generic touchdown amplification protocol. Touchdown protocols involve a pre-designed stepped reduction in the annealing temperature used for primer-to-template binding, which introduces a competitive advantage for specific base-pair priming over non-specific priming. A detailed knowledge of the optimum annealing temperature is therefore not required. The study protocol was empirically constructed and proved robust when applied to a large range of respiratory viral and bacterial targets, without compromising individual test sensitivity. It was designed for use with in-house primer master-mixes that recognise 12 common respiratory viruses.

Before deciding on the layout of the molecular strip, as described in the methods, we undertook a wide range of preliminary validation steps for each primer set. The complexity of the strip makes it impossible to fully evaluate using the classical approach of applying an individual gold standard to each virus type. Classically this approach works well where a single target is under investigation. However although the strip is putatively designed to identify 12 viruses, the actual number of individual types targeted is over one hundred and sixty because of the inclusion of generic primer sets for HRV and ADV respectively. The classical approach is further compounded for viruses (a) that cannot be grown or grown easily; (b) for which commercial IF sera are not available; (c) for which specimen panels are not available. We therefore adopted a phased validation, culminating in the present study. Sensitivity was ascribed by undertaking copy number determination on cloned targets and these ranged form 6 × 103 copies per ml for human rhinovirus type 1b to 4.2 × 103 copies/ml for RSV-A. Specificity was ascribed through reproducibility, i.e. specimens which were repeatedly positive, following our standard clinical reporting algorithm, were regarded as true positives; a similar approach was recently described for hMPV. In addition amplicon sequencing was used as an initial specificity check. The primers sets were tested on clinical respiratory specimens arising from a number of ethically approved studies. These included respiratory specimens from patients: (a) with chronic obstructive pulmonary disease; (b) with acute asthma; (c) on assisted ventilation in intensive care. They were also tested on respiratory specimens collected as part of an influenza spotter program as well as on laboratory specimens of known virus reactivity.

To test the feasibility of its routine use we needed to clinically validate its performance in a routine setting on specimens tested in parallel with our standard immunofluorescence protocol for the diagnosis of acute virus respiratory infections. Although the routine immunofluoresence panel lacked capacity for the detection of rhinoviruses, human metapneumovirus and CoV-229E, these were included on the strip for clinical reasons during the period of the study. These findings and their implications are reported.

A total of 356 nasopharyngeal aspirate and throat swab samples were collected from patients with SARI by a trained technician using a sterile nylon flocked swab and placed in viral transport medium (VTM), labelled and transported on ice at the earliest to Advanced research lab (ICMR Grade-1 Virology Lab) of SMS Medical college Jaipur for further processing and storage of the samples. The study was approved by the institutional ethics committee.

We found that viral infections were often neglected during this population-based “real life” study of suspected septic patients. The study was performed during the winter period, referred to as “the flu season”, when respiratory viral infections are most prevalent. This should have resulted in increased clinical suspicion. Yet, during the first four weeks of the influenza epidemic, very few clinical samples are requested. In this material, a viral respiratory infection was initially suspected by clinicians in only 30% of patients with viral findings by multiplex PCR. This was especially true when CRP was over 100 mg/L, or if there was a new infiltrate on the chest X-ray indicating pneumonia. This underestimation may lead to nosocomial spread or outbreaks of viral respiratory infections, as we have previously experienced in our own hospital. It may also lead to overuse of antibiotics, as well as underuse of antivirals, especially in risk groups that might benefit from such treatment.

As in a comprehensive study on bacterial–viral respiratory tract illness over three winter seasons by Falsey et al. in 2013, influenza A virus was the most common viral finding, appearing in study samples almost two weeks earlier than in clinical samples. In only 35 of 96 cases of influenza A virus infection (36%) was influenza virus initially suspected as sole cause or contributing factor to the acute illness.

Respiratory syncytial virus and human metapneumovirus may cause critical respiratory illness and pneumonia, not only in children, but also in elderly. For example, human metapneumovirus was found to be the causative agent in an outbreak of pneumonia among elderly at an institution in the Netherlands. In this study, human metapneumovirus was a slightly more common finding than respiratory syncytial virus, especially in patients with long history of fever and respiratory tract congestion, combined with radiological signs of pneumonia.

Nasopharyngeal culture is generally discouraged or not recommended for etiological diagnosis of pneumonia. However, in a Swedish study by Strålin et al. in 2006, there was a good correlation between nasopharyngeal findings of these bacteria and the etiology of pneumonia, as has been seen in previous Swedish studies. In our study of patients in the emergency department suspected to be septic, there was a strong correlation between nasopharyngeal findings of S. pneumoniae or H. influenzae and X-ray findings of a new infiltrate, indicative of pneumonia. More so, these bacteria were not found in the nasopharynx of any of the 210 patients with a non-respiratory infection or no infection, and they were rarely found in patients with a respiratory tract infection but not pneumonia. The study results imply that nasopharyngeal findings of S. pneumoniae and H. influenzae in sepsis patients should be considered carefully for patient treatment. A recently published paper by Bjarnason et al. in 2017 demonstrates a good correlation between real-time PCR findings of S. pneumoniae or H. influenzae to pneumonia diagnosis in adults, which also builds support for a clinical relevance of these upper respiratory bacterial findings.

Co-infections of bacteria and respiratory viruses, mainly S. pneumoniae and influenza A or respiratory syncytial virus, are found in 3–40% of patients with CAP, depending on diagnostic methods used, with the higher end reflecting studies in which nasopharyngeal culture is included for etiological diagnosis [1, 2]. Using nasopharyngeal sampling only, we found indications of viral-bacterial co-infections in 28 of 137 (20%), a proportion we believe to be an underestimation. As in the study by Falsey et al. in 2013, S. pneumoniae was the bacteria most often associated with pneumonia and a viral co-infection. As many as 75% of patients with pneumonia and S. pneumoniae in the nasopharynx were positive for a respiratory virus, mainly influenza A virus, but also human metapneumovirus. The two youngest patients, with pneumonia and severe sepsis, aged 37 and 42 years respectively, were both positive for S. pneumoniae and human metapneumovirus in the nasopharynx. No other pathogens could be demonstrated by routine cultures.

In the clinical setting it is often difficult to determine whether a patient with respiratory symptoms has a viral infection, a bacterial infection, or a mixed viral-bacterial infection. No constellation of clinical symptoms, vital signs, biomarkers (such as white blood cell count, C-reactive protein, or procalcitonin) have adequate sensitivity and specificity. New tools to improve predictions of patient benefit from antibiotic treatment are urgently needed. Recently, whole blood analysis for the identification of host gene activation profiles has been able to discriminate viral infections from bacterial infections with high accuracy in severely ill infants, as described by Herberg et al. in 2016. In adults with lower respiratory tract infections, a similar technique seems able to discriminate viral from bacterial infections much better than procalcitonin, as shown by Suarez et al. in 2015. In the same study mixed viral-bacterial infections also elicited a characteristic gene activation profile.

Our study supports increased testing for respiratory viruses in patients believed to be septic, especially those presenting with respiratory tract symptoms. With current technology, results can be obtained within a few hours and have an impact on clinical decisions and patient logistics in the emergency department. Cost effectiveness should be investigated. A viral diagnosis may not only lead to fewer admissions and less antibiotic treatment if bacterial pneumonia is suspected or demonstrated, but may also decrease viral exposures for admitted patients. Patients with influenza A or B may benefit from antiviral treatment alone or in conjunction with antibacterial treatment, if bacterial pneumonia is suspected or demonstrated, perhaps even reducing viral contagiousness. Even in neutropenic patients, a viral finding and a favorable outcome in the first few days may safely allow for discontinuation of antibiotic treatment.

This study has several limitations. It is a single-center study performed during one winter period only. The study was primarily not an etiological study of respiratory tract infections. We did not take ongoing antibiotic treatment into account. Only routine sampling from the nasopharynx was performed, albeit using a flocked swab for better yield. If sputum or nasopharyngeal aspirates had been analyzed with molecular techniques, we would have expected a higher yield for both bacteria and viruses [2, 30]. A further weakness is the open inclusion based on clinical suspicion of sepsis only, without specific criteria. Yet another was the subjectivity involved in deciding the relevance of the findings. What role do the viral findings play, both alone or in conjunction with bacterial findings? We can only show correlations between findings and clinical entities, yet the majority of findings do seem to correlate well to respiratory tract infections. Therefore, we believe that our conclusion, that significant viral disease in severely ill patients is underdiagnosed by clinicians, is warranted. Diagnosing these infections early may be of help for the clinical decision making process and thereby for the patients.

This was a retrospective, Institutional Review Board-approved study of patients who developed laboratory-confirmed respiratory viral infections and whose symptoms began during hospitalization at Rhode Island Hospital and Hasbro Children’s Hospital (HCH) between April 1, 2015 and April 1, 2016. Rhode Island Hospital is licensed for 719 beds, 87 of which are licensed for HCH. The two hospitals are located in adjoining buildings on the same campus as part of the Lifespan Hospital System.

A nosocomial respiratory viral infection was defined as a hospitalized patient who had a positive respiratory viral panel ([RVP] Luminex, Austin, TX), rapid influenza test (Xpert; Cepheid, Sunnyvale, CA), or rapid respiratory syncytial virus (RSV) test (Xpert; Cepheid) of a nasopharyngeal or bronchoscopic lavage specimen. The RVP assay included testing for influenza A and B, RSV A and B, coronavirus, parainfluenza, human metapneumovirus, adenovirus, as well as rhinovirus and enterovirus; however, the assay did not distinguish the latter 2 viruses, which heretofore will be noted as rhino/enterovirus. Hospitalized patients with a positive result in one of these tests during the study period were identified using our institutional infection control software system (TheraDoc; Premier, Charlotte, NC). A definite nosocomial respiratory viral infection was defined as a patient whose number of days from hospital admission to symptom onset exceeded the upper range for the incubation period of the identified virus (Table 1). A possible nosocomial respiratory viral infection case was defined as a patient in whom the number of days from hospital admission to symptom onset was within the range of the incubation period for the identified virus who were admitted without clinical signs or symptoms of a respiratory infection. A patient could be included more than once during the hospitalization if they had complete resolution of symptoms ascribed to the respiratory viral infection, recurrence of symptoms compatible with such an infection, and the time interval was greater than the above-noted incubation period. If the second such episode was due to a different virus, it was considered a definite case, and if it was due to the same virus, it was considered a possible case. Cases were assigned to a season based on the date when the symptoms of a respiratory viral infection were first documented in the medical record. Estimates of the incidence of nosocomial viral infections yearly in US acute care adult and pediatric hospitals were based on the number of such infections identified in the current study and the number of admissions to these hospitals during the 2014 fiscal reporting year based on data from the American Hospital Association.

A literature review was performed using the following PubMed search terms: nosocomial; hospital-acquired; and respiratory viral infections. Only studies written in the English language were included. Studies were excluded in our comparative analysis of the incidence of respiratory viral infections if the study was an outbreak investigation or if the reported incidence included nonrespiratory viral infections.

Data and swabs result from a surveillance system that received regulatory approvals, including the CNIL (National Commission for Information Technology and Civil Liberties Number 1592205) approval in July 2012. All the patients have received oral information and gave their consent for swab and data collection. Data were collected for surveillance purpose and are totally anonymous.

From October 2013 to June 2016 pilgrims and people returning from the Middle East presented with respiratory symptoms and having an epidemiological link were enrolled through active screening at the point of entry. After risk assessment by the medical team, suspected patients were admitted to the Kurmitola General Hospital isolation unit. In addition, travelers arriving with no clinical presentation received a health card mentioning the sign and symptoms of MERS-CoV infection. Self reported cases who developed symptoms within 14 days of arrival were included in sample collection along with admitted patients. The health care providers were instructed to use WHO standard Personal Protective Equipment (PPE) including N95 masks during handling of the patients.

In this study, we have focused on inward patients only. Therefore, we may have missed a significant number of outpatients, who would have had milder form of hMPV associated ARTI.

Viruses account for most of the respiratory tract infections in childhood [1, 12, 14]. Viral infections of the respiratory tract are often treated with antibiotics due to the absence of viral diagnostics to identify the viral aetiology. Thus, a proper diagnosis is crucial prior to initiating antibiotic treatment for bacterial ARTI or pneumonia [9, 15]. In developing countries, a lack of availability of diagnostic facilities contributes to the use of antibiotics and thus to development of antimicrobial resistance [9, 10]. The imaging studies and blood cell differential count may give a clue on the type of infective agent. However, in atypical pneumonias, getting an educated guess about the bacterial and viral causes are difficult. Hence, routine viral laboratory diagnosis is crucial and implementation of such facilities is highly warranted.

RSV is the most common respiratory viral pathogen causing hospitalization of thousands of children each year [2, 15]. Many of the affected children do not require hospitalization and some with severe respiratory disease are hospitalized or even managed in the intensive care unit (ICU). The children requiring ICU admission are typically young infants and those with co-morbidities. These children can be severely ill and require intubation and mechanical ventilation but most of the children recover and a very few succumb to the disease. Currently we are seeing the emergence of respiratory pathogens either due to change in antigenicity in influenza viruses or emergence and introduction of newly emerging viral pathogens like hMPV.

In this case series, hMPV infection showed a disease spectrum similar to that seen during RSV infection, common cold to life threatening pneumonia. Children delivered through LSCS appear to have less resistance to infection and in our study also, children delivered through LSCS had a high risk of developing hMPV/RSV co-infection. A child with a birth order > 3 had a high risk of getting hMPV/RSV co-infection and this might be due to lack of care to subsequent children in bigger families. In a few cases, even without co- morbidities, children experienced severe hMPV infection needing ICU care. In many cases, RSV/hMPV co-infection resulted in similar disease spectrum to that of RSV infection.

Specific aetiological diagnosis of childhood ARTI is not performed routinely in Sri Lanka. But if it is done routinely it will invariably guide the clinicians on the use of antibiotics including antivirals. This case series indicates the importance of establishing laboratory diagnosis for viral ARTI. Furthermore, hMPV is a potential pathogen that needs to be tested in children with ARTI. A detailed epidemiological study is in progress to elucidate the prevalence and seasonality of childhood ARTI caused by a wider range of respiratory viruses including hMPV in Sri Lanka.

This was a prospective and observational study conducted across multiple consecutive RSV seasons to determine the incidence rate of medically attended acute respiratory illness (MARI) or events leading to worsening cardiorespiratory status in adults with severe COPD and/or advanced CHF associated with RSV and other viral infections. Fifty‐seven sites in nine countries (Bulgaria, Canada, Czech Republic, France, Germany, Italy, Russia, Sweden, and the United States) in the Northern hemisphere participated in the study from fall 2011 through spring 2014. The protocol was approved by independent institutional review boards, and all subjects signed written informed consent at enrollment.

The study was approved by the National Ethics Committee for Health Research, Ministry of Health, Lao PDR, and the Oxford Tropical Research Ethics Committee. Methods were carried out in accordance with the relevant guidelines and regulations. Informed signed consent was obtained from their legal guardians for all children included in this study.

Detection of respiratory viruses, including HPIV 1–4, HRSV, HMPV, HCoV-OC43, HCoV-229E, HCoVNL63, HCoV-HKU1, HRV, HAdV, and HBoV was performed using a RT-PCR Kit «AmpliSens ARVI-screen-FL» (Interlabservice, Russia), and IFVA and IFVB virus detection was performed using a RT-PCR Kit « AmpliSens Influenza virus A/B-FL» (Interlabservice, Russia) according to the manufacturer's instructions. Positive and negative controls were included in each run.