Background Severe acute respiratory infections (SARI) are a leading cause of hospitalizations in children, especially due to viral pathogens. We studied the prevalence of respiratory viruses among children aged <5 years hospitalized with severe acute respiratory infections (SARI) in Kashmir, India. Methods We conducted a prospective observational study in two tertiary care hospitals from October 2013 to September 2014, systematically enrolling two children aged <5 years with SARI per day. We defined SARI as history of fever or measured fever (≥38°C) and cough with onset in the last 7 days requiring hospitalization for children aged 3-59 months and as physician-diagnosed acute lower respiratory infection for children aged <3 months. Trained study staff screened children within 24 hours of hospitalization for SARI and collected clinical data and nasopharyngeal swabs from enrolled participants. We tested for respiratory syncytial virus (RSV) A and B, influenza viruses, rhinoviruses (HRV)/enteroviruses, adenovirus (AdV), bocavirus (BoV), human metapneumovirus (hMPV) A and B, coronaviruses (OC43, NL65, C229E), and parainfluenza viruses (PIV) 1, 2, 3 and 4 using standardized duplex real-time polymerase chain reaction. Results Among 4548 respiratory illness admissions screened from October 2013 to September 2014, 1026 met the SARI case definition, and 412 were enrolled (ages = 5 days to 58 months; median = 12 months). Among enrolees, 256 (62%) were positive for any virus; RSV was the most commonly detected (n = 118, 29%) followed by HRV/enteroviruses (n = 88, 21%), PIVs (n = 31, 8%), influenza viruses (n = 18, 4%), BoV (n = 15, 4%), coronaviruses (n = 16, 4%), AdV (n = 14, 3%), and hMPV (n = 9, 2%). Fifty-four children had evidence of virus co-detection. Influenza-associated SARI was more common among children aged 1-5 years (14/18, 78%) while most RSV detections occurred in children <12 months (83/118, 70%). Of the RSV viruses typed (n = 116), the majority were type B (94, 80%). Phylogenetic analysis of G gene of RSV showed circulation of the BA9 genotype with 60bp nucleotide duplication. Conclusions Respiratory viruses, especially RSV, contributed to a substantial proportion of SARI hospitalizations among children <5 years in north India. These data can help guide clinicians on appropriate treatment and prevention strategies. © 2022. The Author(s)

Influenza virus non-structural protein 1 (NS1) counteracts host antiviral innate immune responses by inhibiting Retinoic acid inducible gene-I (RIG-I) activation. However, whether NS1 also specifically regulates RIG-I transcription is unknown. Here, we identify a CCAAT/Enhancer Binding Protein beta (C/EBPbeta) binding site in the RIG-I promoter as a repressor element, and show that NS1 promotes C/EBPbeta phosphorylation and its recruitment to the RIG-I promoter as a C/EBPbeta/NS1 complex. C/EBPbeta overexpression and siRNA knockdown in human lung epithelial cells resulted in suppression and activation of RIG-I expression respectively, implying a negative regulatory role of C/EBPbeta. Further, C/EBPbeta phosphorylation, its interaction with NS1 and occupancy at the RIG-I promoter was associated with RIG-I transcriptional inhibition. These findings provide an important insight into the molecular mechanism by which influenza NS1 commandeers RIG-I transcriptional regulation and suppresses host antiviral responses.

Historical specimens collected from hospitalized children were tested for the following 13 viruses: influenza A and B; respiratory syncytial virus (RSV); parainfluenza viruses 1-3; human metapneumovirus; rhinovirus; coronaviruses 229E, OC43, NL63 and HKU1 and Adenovirus using monoplex real-time reverse transcriptase polymerase chain reaction (rRT-PCR). They were retested using TaqMan Array Card (TAC), a micro-fluidic system, capable of simultaneous multi-pathogen testing, to evaluate its sensitivity and specificity against monoplex rRT-PCR. TAC showed high sensitivity (71%-100%) and specificity (98%-100%) for these viruses in comparison to monoplex rRT-PCR. Multi-specimen detection with high sensitivity and specificity makes TAC a potentially useful tool for both surveillance and outbreak investigations.

Background & objectives: Respiratory tract infections are common among Hajj and Umrah pilgrims which pose a public health risk of spread of respiratory infections. Influenza has been reported from Indian Hajj and Umrah returning pilgrims, but data on other respiratory pathogens are sparse in India. Here we report the presence of common respiratory viral pathogens in returning Hajj and Umrah pilgrims suffering from acute respiratory illness (ARI) in 2014-2015. Methods: Respiratory specimens (nasopharyngeal and throat swabs) were collected from 300 consenting pilgrims with ARI in the past one week and tested for influenza and Middle East Respiratory Syndrome coronavirus (MERS-CoV) and other respiratory viruses using in-house standardized quantitative real-time reverse-transcription polymerase chain reaction. Clinical features among the pathogen positive and negative patients were compared. The patients received symptomatic treatment and antivirals where appropriate and were followed telephonically to collect data on illness outcome. Results: Ninety seven (32.3%) of the 300 participants were tested positive for any virus, most common being influenza viruses (n=33, 11%). Other respiratory viruses that were detected included human coronaviruses [n=26, 8.7%; OC43 (n=19, 6.3%) and C229E (n=7, 2.3%)], rhinovirus (n=20, 6%), adenoviruses (n=8, 2.6%), parainfluenza viruses (n=7, 2.3%), respiratory syncytial virus (n=3, 1%) and bocaviruses (n=2, 0.6%). Clinical features observed in pathogen positive and pathogen negative patients did not differ significantly. Eighteen influenza positive patients were treated with oseltamivir. Interpretation & conclusions: Pilgrims returning from mass gatherings are often afflicted with respiratory pathogens with a potential to facilitate transmission of respiratory pathogens across international borders. The study reinforces the need for better infection prevention and control measures such as vaccination, health education on cough etiquette and hand hygiene.

The role of killer cell immunoglobulin-like receptors (KIRs) in the transmission of HIV-1 has not been extensively studied. Here, we investigated the association of KIR gene content polymorphisms with perinatal HIV-1 transmission. The KIR gene family comprising 16 genes was genotyped in 313 HIV-1 positive Kenyan mothers paired with their infants. Gene content polymorphisms were presented as presence of individual KIR genes, haplotypes, genotypes and KIR gene concordance. The genetic data were analyzed for associations with perinatal transmission of HIV. There was no association of infant KIR genes with perinatal HIV-1 transmission. After adjustment for gravidity, viral load, and CD4 cell count, there was evidence of an association between reduction in perinatal HIV-1 transmission and the maternal individual KIR genes KIR2DL2 (adjusted OR = 0.50; 95% CI: 0.24-1.02, P = 0.06), KIR2DL5 (adjusted OR = 0.47; 95% CI: 0.23-0.95, P = 0.04) and KIR2DS5 (adjusted OR = 0.39; 95% CI: 0.18-0.80, P = 0.01). Furthermore, these maternal KIR genes were only significantly associated with reduction in perinatal HIV transmission in women with CD4 cell count >/= 350 cells/ mul and viral load <10000 copies/ml. Concordance analysis showed that when both mother and child had KIR2DS2, there was less likelihood of perinatal HIV-1 transmission (adjusted OR = 0.44; 95% CI: 0.20-0.96, P = 0.039). In conclusion, the maternal KIR genes KIR2DL2, KIR2DL5, KIR2DS5, and KIR2DS2 were associated with reduction of HIV-1 transmission from mother to child. Furthermore, maternal immune status is an important factor in the association of KIR with perinatal HIV transmission.

BACKGROUND: The increasing reports of Middle East Respiratory Syndrome (MERS) caused by MERS coronavirus (MERS-CoV) from many countries emphasize its importance for international travel. Muslim pilgrimages of Hajj and Umrah involve mass gatherings of international travellers. We set out to assess the presence of influenza and MERS-CoV in Hajj/Umrah returnees with acute respiratory infection. . METHODS: Disembarking passengers (n = 8753) from Saudi Arabia (October 2014 to April 2015) were interviewed for the presence of respiratory symptoms; 977 (11%) reported symptoms and 300 (age 26-90, median 60 years; 140 male) consented to participate in the study. After recording clinical and demographic data, twin swabs (nasopharyngeal and throat) were collected from each participant, pooled in viral transport media and tested by real-time RT PCR for MERS-CoV and influenza A and B viruses and their subtypes. RESULTS: The participants had symptoms of 1-15 days (median 5d); cough (90%) and nasal discharge (86%) being the commonest. None of the 300 participants tested positive for MERS-CoV; however, 33 (11%) tested positive for influenza viruses (A/H3N2 = 13, A/H1N1pdm09 = 9 and B/Yamagata = 11). Eighteen patients received oseltamivir. No hospitalizations were needed and all had uneventful recovery. CONCLUSION: Despite a high prevalence of acute respiratory symptoms, MERS coV was not seen in returning pilgrims from Hajj and Umrah. However detection of flu emphasises preventive strategies like vaccination.

METHODS: We analyzed influenza surveillance data from nine countries around southern and southeastern Asia spanning latitudinal gradient from equatorial to temperate zones to further characterize influenza type specific seasonality in the region. We calculated proportion of positives by month out of positives during that year and adjust for variation in samples tested and positivity in these countries. RESULTS: Influenza A epidemics were identified between November-March during winters in areas lying above 30o N latitude; during monsoon months of June-November in areas between 10o -30o N latitude, and no specific seasonality for influenza A virus circulation in areas lying closer to the equator. Influenza B circulation coincided with influenza A circulation in areas lying above 30o N latitude; however in areas south of 30o N Asia, influenza B circulated year round at 3-8% of annual influenza B positives during most months with less pronounced peaks during post-monsoon period. CONCLUSION: Even though influenza B circulates round the year in most areas of the tropical southern and southeastern Asia region, the most appropriate time for influenza vaccination would be prior to the monsoon season conferring protection against influenza A and B peaks using the most recent WHO recommended vaccine. This article is protected by copyright. All rights reserved.

A unique feature of influenza A virus (IAV) life cycle is replication of the viral genome in the host cell nucleus. The nuclear import of IAV genome is an indispensable step in establishing virus infection. IAV nucleoprotein (NP) is known to mediate the nuclear import of viral genome via its nuclear localization signals. Here, we demonstrate that cellular heat shock protein 40 (Hsp40/DnaJB1) facilitates the nuclear import of incoming IAV viral ribonucleoproteins (vRNPs) and is important for efficient IAV replication. Hsp40 was found to interact with NP component of IAV RNPs during early stages of infection. This interaction is mediated by the J domain of Hsp40 and N-terminal region of NP. Drug or RNAi mediated inhibition of Hsp40 resulted in reduced nuclear import of IAV RNPs, diminished viral polymerase function and attenuates overall viral replication. Hsp40 was also found to be required for efficient association between NP and importin alpha, which is crucial for IAV RNP nuclear translocation. These studies demonstrate an important role for cellular chaperone Hsp40/DnaJB1 in influenza A virus life cycle by assisting nuclear trafficking of viral ribonucleoproteins.

BACKGROUND: The burden estimation studies for respiratory syncytial virus (RSV) have been based on varied case definitions, including case-definitions designed for influenza surveillance systems. We used all medical admissions among children aged 0-59 months to study the effect of case definitions on estimation of RSV-associated hospitalizations rates. METHODS: The hospital-based daily surveillance enrolled children aged 0-59 months admitted with acute medical conditions from July 2009-December 2012, from a well-defined rural population in Ballabgarh in northern India. All study participants were examined and nasal and throat swabs taken for testing by real-time polymerase chain reaction (RT-PCR) for RSV and influenza virus. Clinical data were used to retrospectively evaluate World Health Organization (WHO) case definitions (2011) commonly used for surveillance of respiratory pathogens, ie, acute respiratory illness (WHO-ARI), severe ARI (SARI) and influenza-like illness (ILI), for determination of RSV-associated hospitalization. RSV-associated hospitalization rates adjusted for admissions at non-study hospitals were calculated. FINDINGS: Out of 505 children enrolled, 82 (16.2%) tested positive for RSV. Annual incidence rates of RSV-associated hospitalization per 1000 children were highest among infants aged 0-5 months (15.2; 95% confidence interval (CI) 8.3-26.8), followed by ages 6-23 months (5.3, 95% CI 3.2-8.7) and lowest among children 24-59 months (0.5, 95% CI 0.1-1.5). The RSV positive children were more likely to have signs of respiratory distress like wheeze, chest in-drawing, tachypnea, and crepitation compared to RSV-negative based on bivariate comparisons. Other less commonly seen signs of respiratory distress, ie, nasal flaring, grunting, accessory muscle usage were also significantly associated with being RSV positive. Compared to the estimated RSV hospitalization rate based on all medical hospitalizations, the WHO-ARI case definition captured 86% of the total incidence, while case definitions requiring fever like ILI and SARI underestimated the incidence by 50-80%. CONCLUSIONS: Our study suggests that RSV is a substantial cause of hospitalization among children aged <24months especially those aged <6 months. The WHO-ARI case definition appeared to be the most suitable screening definition for RSV surveillance because of its high sensitivity.

BACKGROUND: Influenza surveillance is an important tool to identify emerging/reemerging strains, and defining seasonality. We describe the distinct patterns of circulating strains of the virus in different areas in India from 2009 to 2013. METHODS: Patients in ten cities presenting with influenza like illness in out-patient departments of dispensaries/hospitals and hospitalized patients with severe acute respiratory infections were enrolled. Nasopharangeal swabs were tested for influenza viruses by real-time RT-PCR, and subtyping; antigenic and genetic analysis were carried out using standard assays. RESULTS: Of the 44,127 ILI/SARI cases, 6,193 (14.0%) were positive for influenza virus. Peaks of influenza were observed during July-September coinciding with monsoon in cities Delhi and Lucknow (north), Pune (west), Allaphuza (southwest), Nagpur (central), Kolkata (east) and Dibrugarh (northeast), whereas Chennai and Vellore (southeast) revealed peaks in October-November, coinciding with the monsoon months in these cities. In Srinagar (Northern most city at 34 degrees N latitude) influenza circulation peaked in January-March in winter months. The patterns of circulating strains varied over the years: whereas A/H1N1pdm09 and type B co-circulated in 2009 and 2010, H3N2 was the predominant circulating strain in 2011, followed by circulation of A/H1N1pdm09 and influenza B in 2012 and return of A/H3N2 in 2013. Antigenic analysis revealed that most circulating viruses were close to vaccine selected viral strains. CONCLUSIONS: Our data shows that India, though physically located in northern hemisphere, has distinct seasonality that might be related to latitude and environmental factors. While cities with temperate seasonality will benefit from vaccination in September-October, cities with peaks in the monsoon season in July-September will benefit from vaccination in April-May. Continued surveillance is critical to understand regional differences in influenza seasonality at regional and sub-regional level, especially in countries with large latitude span.

BACKGROUND: Despite the high mortality and morbidity resulting from acute respiratory infections (ARI) globally, there are few data from low-income countries on costs of ARI to inform public health policy decisions We conducted a prospective survey to assess costs of ARI episodes in selected primary, secondary, and tertiary healthcare facilities in north India where no respiratory pathogen vaccine is routinely recommended. METHODS: Face-to-face interviews were conducted among a purposive sample of patients with ARI from healthcare facilities. Data were collected on out-of-pocket costs of hospitalization, medical consultations, medications, diagnostics, transportation, lodging, and missed work days. Telephone surveys were conducted two weeks after medical encounters to ask about subsequent missed work and costs incurred. Costs of prescriptions and diagnostics in public facilities were supplemented with WHO-CHOICE estimates of hospital bed costs. Missed work days were assigned cost based on the national annual per capita income (US$1,104). Non-medically attended ARI cases were identified from an ongoing community-based ARI surveillance project in Faridabad. RESULTS: During September 2012-March 2013, 1766 patients with ARI were enrolled, including 451 hospitalized patients, 1056 outpatients, and 259 non-medically attended patients. The total direct cost of an ARI episode requiring outpatient care was US$4- $6 for public and $3-$10 for private institutions based on age groups. The total direct cost of an ARI episode requiring hospitalized care was $54-$120 in public and $135-$355 in private institutions. The cost of ARI among those hospitalized was highest among persons aged > = 65 years and lowest among children aged < 5 years. Indirect costs due to missed work days were 16-25% of total costs. The direct out-of-pocket cost of hospitalized ARI was 34% of annual per capita income. CONCLUSIONS: The cost of hospitalized ARI episodes in India is high relative to median per capita income. Data from this study can inform evaluations of the cost effectiveness of proven ARI prevention strategies such as vaccination.

BACKGROUND: Rapid point-of-care (POC) tests provide an economical alternative for rapid diagnosis and treatment of influenza, especially in public health emergency situations. OBJECTIVES: To test the performance of a rapid influenza diagnostic test, QuickVue (Quidel) as a POC test against a real-time polymerase chain reaction (RT-PCR) assay for detection of influenza A and B in a developing country setting. STUDY DESIGN: In a prospective observational design, 600 patients with influenza-like illness (ILI) or with severe acute respiratory illness (SARI) who were referred to the Influenza Clinic of a tertiary care hospital in Srinagar, India from September 2012 to April 2013, were enrolled for diagnostic testing for influenza using QuickVue or RT-PCR. All influenza A-positive patients by RT-PCR were further subtyped using primers and probes for A/H1pdm09 and A/H3. RESULTS: Of the 600 patients, 186 tested positive for influenza A or B by RT-PCR (90 A/H1N1pdm09, 7 A/H3 and 89 influenza B), whereas only 43 tested positive for influenza (influenza A = 22 and influenza B = 21) by QuickVue. Thus, the sensitivity of the QuickVue was only 23% (95% confidence interval, CI: 17.3-29.8) and specificity was 100% (95% CI: 99.1-100) with a positive predictive value (PPV) of 100% (95% CI 91.8-100) and a negative predictive value (NPV) of 74.3% (95% CI: 70.5-77.9) as compared to RT-PCR. CONCLUSIONS: The high specificity of QuickVue suggest that this POC test can be a useful tool for patient management or triaging during a public health crisis but a low sensitivity suggests that a negative test result need to be further tested using RT-PCR.

We estimate the contribution of influenza to hospitalizations for acute exacerbations of chronic obstructive pulmonary disease (AECOPD) in Kashmir, India. Prospective surveillance for influenza among patients hospitalized with AECOPD was conducted at a tertiary care hospital. Patients had clinical data collected and nasal/throat swabs tested for influenza viruses. Outcomes among patients with and without influenza were compared with logistic regression adjusting for age and underlying conditions. During October 2010-September 2012, 498 patients hospitalized with AECOPD were enrolled, of whom 40 (8%) had received influenza vaccine. Forty (8%) had influenza; influenza virus detection peaked in winter (January-March). Patients with influenza were more likely to die during hospitalization (adjusted OR 3.4, CI 1.0-11.4) than those without.

The seasonality of influenza in the tropics complicates vaccination timing. We investigated influenza seasonality in northern India and found influenza positivity peaked in Srinagar (34.09 degrees N) in January-March but peaked in New Delhi (28.66 degrees N) in July-September. Srinagar should consider influenza vaccination in October-November, but New Delhi should vaccinate in May-June.

BACKGROUND: The global burden of influenza is increasingly recognized, but data from India remain sparse. We conducted a multi-site population-based surveillance study to estimate and compare rates of influenza-associated hospitalization at two rural Indian health and demographic surveillance system (HDSS) sites at Ballabgarh and Vadu during 2010-2012. METHODS: Prospective facility-based surveillance for all hospitalizations (excluding those for trauma, elective surgery and obstetric, ophthalmic or psychiatric reasons) was conducted at 72 health facilities. After collection of clinical details, patients had nasopharyngeal swabs taken and tested by reverse transcription polymerase chain reaction for influenza viruses. Annual healthcare utilization surveys (HUS) were conducted in HDSS households to identify proportion of hospitalizations occurring at non-study facilities to adjust for hospitalizations missed through facility-based surveillance. RESULTS: HUS showed that 69% and 67% of hospitalizations occurred at study facilities at Ballabgarh and Vadu respectively. Overall, 6,004 patients hospitalized with acute medical illness at participating facilities were enrolled (1,717 from Ballabgarh; 4,287 from Vadu). The proportion of patients with influenza was higher at Vadu than Ballabgarh annually (2010: 21% vs. 5%, p<0.05; 2011: 18% vs. 5%, p<0.05; 2012: 23% vs. 5%, p<0.05). Annual adjusted influenza-associated hospitalization rates were 5-11 fold higher in Vadu (20.3-51.6 per 10,000) versus Ballabgarh (4.4-6.3 per 10,000). At both sites, influenza A/H1N1pdm09 and B predominated during 2010, A/H3N2 and B during 2011, and A/H1N1pdm09 and B during 2012. CONCLUSION: The markedly different influenza hospitalization rates by season and across communities in India highlight the need for sustained multi-site surveillance system for estimating national influenza disease burden. That would be the first step for initiating discussions around Influenza prevention and control strategies in the country.

Influenza A virus (IAV), similar to other viruses, exploits the machinery of human host cells for its survival and replication. We identified alpha-actinin-4, a host cytoskeletal protein, as an interacting partner of IAV nucleoprotein (NP). We confirmed this interaction using co-immunoprecipitation studies, first in a coupled in vitro transcription-translation assay and then in cells either transiently co-expressing the two proteins or infected with whole IAV. Importantly, the NP-actinin-4 interaction was observed in several IAV subtypes, including the 2009 H1N1 pandemic virus. Moreover, immunofluorescence studies revealed that both NP and actinin-4 co-localized largely around the nucleus and also in the cytoplasmic region of virus-infected A549 cells. Silencing of actinin-4 expression resulted in not only a significant decrease in NP, M2 and NS1 viral protein expression, but also a reduction of both NP mRNA and viral RNA levels, as well as viral titers, 24 h post-infection with IAV, suggesting that actinin-4 was critical for viral replication. Furthermore, actinin-4 depletion reduced the amount of NP localized in the nucleus. Treatment of infected cells with wortmannin, a known inhibitor of actinin-4, led to a decrease in NP mRNA levels and also caused the nuclear retention of NP, further strengthening our previous observations. Taken together, the results of the present study indicate that actinin-4, a novel interacting partner of IAV NP, plays a crucial role in viral replication and this interaction may participate in nuclear localization of NP and/or viral ribonucleoproteins.

OBJECTIVE: To characterize influenza seasonality and identify the best time of the year for vaccination against influenza in tropical and subtropical countries of southern and south-eastern Asia that lie north of the equator. METHODS: Weekly influenza surveillance data for 2006 to 2011 were obtained from Bangladesh, Cambodia, India, Indonesia, the Lao People's Democratic Republic, Malaysia, the Philippines, Singapore, Thailand and Viet Nam. Weekly rates of influenza activity were based on the percentage of all nasopharyngeal samples collected during the year that tested positive for influenza virus or viral nucleic acid on any given week. Monthly positivity rates were then calculated to define annual peaks of influenza activity in each country and across countries. FINDINGS: Influenza activity peaked between June/July and October in seven countries, three of which showed a second peak in December to February. Countries closer to the equator had year-round circulation without discrete peaks. Viral types and subtypes varied from year to year but not across countries in a given year. The cumulative proportion of specimens that tested positive from June to November was > 60% in Bangladesh, Cambodia, India, the Lao People's Democratic Republic, the Philippines, Thailand and Viet Nam. Thus, these tropical and subtropical countries exhibited earlier influenza activity peaks than temperate climate countries north of the equator. CONCLUSION: Most southern and south-eastern Asian countries lying north of the equator should consider vaccinating against influenza from April to June; countries near the equator without a distinct peak in influenza activity can base vaccination timing on local factors.

BACKGROUND & OBJECTIVES: Community outbreaks of disease amongst nomadic populations generally remain undocumented. Following a reported increase in acute respiratory tract infections (ARI) in May 2011 in a nomadic population of Sangerwini in Jammu & Kashmir, India, we examined the patients with ARI symptoms and their nasal swabs were tested for influenza virus. METHODS: Patients with ARI (n=526) were screened from May 14 to 23, 2011 and nasopharyngeal swabs collected from 84 with Influenza like illness (ILI) for bacterial cultures and influenza virus testing. Samples were tested for influenza A and influenza B by real time (RT)-PCR. RESULTS: Twelve (14.3%) of the 84 patients tested positive for influenza B, compared to only one (0.9%) of 108 patients with ILI in a parallel survey performed in Srinagar during the same period, suggesting a localized outbreak in the isolated nomadic community. All presented with respiratory symptoms of less than seven days. Familial clustering was seen in 40 per cent (25% of influenza B positives). Average daytime temperatures ranged from 15-16 degrees C compared to 22 degrees C in Srinagar. Four patients developed pneumonia whereas others ran a mild course with a total recovery with oseltamivir and symptomatic therapy. INTERPRETATION & CONCLUSION: Our report of confirmed influenza B in this underprivileged nomadic population argues for routine surveillance with efforts to improve vaccination and infection control practices.

BACKGROUND & OBJECTIVES: Most studies on the clinical presentation with influenza viruses have been conducted in outpatient or inpatient medical facilities with only a few studies in community settings. Clinical differences between influenza A (H1N1) pdm 09 and influenza B virus infections have importance for community-based public health surveillance. An active community surveillance at the time of emergence of pandemic influenza provided us with an opportunity to compare the clinical features among patients infected with influenza A (H1N1) pdm09 virus and those with influenza B virus co-circulating in an active community-based weekly surveillance in three villages in Faridabad, Haryana, north India. METHODS: Active surveillance for febrile acute respiratory infection (FARI) was carried out in a rural community (n=16,182) in the context of an inactivated trivalent influenza vaccine trial (among children <11 yr). Individuals with FARI were assessed clinically by nurses and respiratory samples collected and tested for influenza viruses by real time RT-PCR from November 2009 to August 2010. Clinical symptoms of patients with influenza A (H1N1) pdm 09 and influenza B infection were compared. RESULTS: Of the 4796 samples tested, 822 (17%) were positive for influenza virus. Of these, 443 (54%) were influenza A (H1N1) pdm09, 373 (45%) were influenza B and six were other subtypes/mixed infections. The mean age was lower for patients with influenza B (16.4 yr) than influenza A (H1N1) pdm09 infection (18.7 yr; P=0.04). Among children aged 5-18 yr, chills/rigours (OR 4.0; CI 2.2, 7.4), sore throat (OR 6.8; CI 2.3, 27.3) and headache (OR2.0; CI 1.3, 3.3) were more common in influenza A (H1N1) pdm09 infection than in influenza B cases. Chills/rigours (OR 2.4; CI 1.4, 4.0) and headache (OR 1.7; CI 1.0, 2.7) were associated with influenza A (H1N1) pdm09 infection in those >18 yr. No significant differences were seen in children <5 yr. CONCLUSION: Our findings show that the differences in the clinical presentation of influenza A(H1N1)pdm09 and influenza B infections are not likely to be of clinical or public health significance.

OBJECTIVES: Though respiratory viruses are thought to cause substantial morbidity globally in children aged <5 years, the incidence of severe respiratory virus infections in children is unknown in India where 20% of the world's children live. METHODS: During August 2009-July 2011, prospective population-based surveillance was conducted for hospitalizations of children aged <5 years in a rural community in Haryana State. Clinical data and respiratory specimens were collected. Swabs were tested by RT-PCR for influenza and parainfluenza viruses, respiratory syncytial virus (RSV), human metapneumovirus, coronaviruses, and adenovirus. Average annual hospitalization incidence was calculated using census data and adjusted for hospitalizations reported to occur at non-study hospitals according to a comunity healthcare utilization survey. RESULTS: Of 245 hospitalized children, respiratory viruses were detected among 98 (40%), of whom 92 (94%) had fever or respiratory symptoms. RSV accounted for the highest virus-associated hospitalization incidence (34.6/10,000, 95% CI 26.3-44.7) and 20% of hospitalizations. There were 11.8/10,000 (95% CI 7.9-18.4) influenza-associated hospitalizations (7% of hospitalizations). RSV and influenza virus detection peaked in winter (November-February) and rainy seasons (July), respectively. CONCLUSION: Respiratory viruses were associated with a substantial proportion of hospitalizations among young children in a rural Indian community. Public health research and prevention in India should consider targeting RSV and influenza in young children.

BACKGROUND: Data on influenza illness rates with population denominators are needed to quantify overall morbidity and to prioritize public health intervention strategies. METHODS: The rates of influenza A(H1N1)pdm09 infection during pandemic phases were determined in a longitudinal community cohort study as part of an influenza vaccine study in a rural community of North India. RESULTS: During the 711 731 person-weeks of surveillance, a total of 1410/7571 (19%) febrile acute respiratory illness cases were positive for influenza. Of these, 749 (53%) were influenza A(H1N1)pdm09, 643 (46%) influenza B, and 18 (1%) influenza A (H3N2). The overall incidence rate of influenza-associated febrile acute respiratory illness was 128/1000 person-years. The incidence rates of influenza A(H1N1)pdm09 were high during both the pandemic phase (179/1000 person-years; November 2009 to January 2010) and post-pandemic phase (156/1000 person-years; August to October 2010), with children <18 years of age being at the greatest risk of influenza infection in the community. CONCLUSIONS: These findings provide important information for planning clinical and public health intervention strategies to mitigate the impact of influenza epidemics.

BACKGROUND: Influenza is vaccine-preventable; however, the burden of severe influenza in India remains unknown. We conducted a population-based study to estimate the incidence of laboratory confirmed influenza-associated hospitalizations in a rural community in western India. METHODS: We conducted active surveillance for hospitalized patients with acute medical illnesses or acute chronic disease exacerbations in Pune during pandemic and post pandemic periods (May 2009-April 2011). Nasal and throat swabs were tested for influenza viruses. A community health utilization survey estimated the proportion of residents hospitalized with respiratory illness at non-study facilities and was used to adjust incidence estimates from facility-based surveillance. RESULTS: Among 9,426 hospitalizations, 3,391 (36%) patients were enrolled; 665 of 3,179 (20.9%) tested positive for influenza. Of 665 influenza positives, 340 (51%) were pandemic A(H1N1)pdm09 and 327 (49%) were seasonal, including A/H3 (16%), A/H1 (3%) and influenza B (30%). The proportion of patients with influenza peaked during August 2009 (39%) and 2010 (42%). The adjusted annual incidence of influenza hospitalizations was 46.8/10,000 during pandemic and 40.5/10,000 during post-pandemic period with comparable incidence of A(H1N1)pdm09 during both periods (18.8 and 20.3, respectively). The incidence of both pH1N1 and seasonal hospitalized influenza disease was highest in the 5-29 year olds. CONCLUSIONS: We document the previously unrecognized burden of influenza hospitalization in a rural community following the emergence of influenza A(H1N1)pdm09 viruses in India. During peak periods of influenza activity circulation i.e during the monsoon period, 20% of all hospital admissions in the community had influenza positivity. These findings can inform development of influenza prevention and control strategies in India.

Apoptosis induction is an antiviral host response, however, influenza A virus (IAV) infection promotes host cell death. The nucleoprotein (NP) of IAV is known to contribute to viral pathogenesis, but its role in virus-induced host cell death was hitherto unknown. We observed that NP contributes to IAV infection induced cell death and heterologous expression of NP alone can induce apoptosis in human airway epithelial cells. The apoptotic effect of IAV NP was significant when compared with other known proapoptotic proteins of IAV. The cell death induced by IAV NP was executed through the intrinsic apoptosis pathway. We screened host cellular factors for those that may be targeted by NP for inducing apoptosis and identified human antiapoptotic protein Clusterin (CLU) as a novel interacting partner. The interaction between IAV NP and CLU was highly conserved and mediated through beta-chain of the CLU protein. Also CLU was found to interact specifically with IAV NP and not with any other known apoptosis modulatory protein of IAV. CLU prevents induction of the intrinsic apoptosis pathway by binding to Bax and inhibiting its movement into the mitochondria. We found that the expression of IAV NP reduced the association between CLU and Bax in mammalian cells. Further, we observed that CLU overexpression attenuated NP-induced cell death and had a negative effect on IAV replication. Collectively, these findings indicate a new function for IAV NP in inducing host cell death and suggest a role for the host antiapoptotic protein CLU in this process.

Immunoglobulin (Ig) GM and KM allotypes, genetic markers of gamma and kappa chains, are associated with humoral immune responsiveness. Previous studies have shown the relationships between GM6-carrying haplotypes and susceptibility to malaria infection in children and adults; however, the role of the genetic markers in placental malaria (PM) infection and PM with HIV co-infection during pregnancy has not been investigated. We examined the relationship between the gene polymorphisms of Ig GM6 and KM allotypes and the risk of PM infection in pregnant women with known HIV status. DNA samples from 728 pregnant women were genotyped for GM6 and KM alleles using polymerase chain reaction-restriction fragment length polymorphism method. Individual GM6 and KM genotypes and the combined GM6 and KM genotypes were assessed in relation to PM in HIV-1 negative and positive women, respectively. There was no significant effect of individual GM6 and KM genotypes on the risk of PM infection in HIV-1 negative and positive women. However, the combination of homozygosity for GM6(+) and KM3 was associated with decreased risk of PM (adjusted OR, 0.25; 95% CI, 0.08-0.8; P = 0.019) in HIV-1 negative women while in HIV-1 positive women the combination of GM6(+/-) with either KM1-3 or KM1 was associated with increased risk of PM infection (adjusted OR, 2.10; 95% CI, 1.18-3.73; P = 0.011). Hardy-Weinberg Equilibrium (HWE) tests further showed an overall significant positive F(is) (indication of deficit in heterozygotes) for GM6 while there was no deviation for KM genotype frequency from HWE in the same population. These findings suggest that the combination of homozygous GM6(+) and KM3 may protect against PM in HIV-1 negative women while the HIV-1 positive women with heterozygous GM6(+/-) combined with KM1-3 or KM1 may be more susceptible to PM infection. The deficit in heterozygotes for GM6 further suggests that GM6 could be under selection likely by malaria infection.

Population-based active surveillance in India showed higher incidence rates for influenza A(H1N1)pdm09 among children during pandemic versus postpandemic periods (345 vs. 199/1,000 person-years), whereas adults had higher rates during postpandemic versus pandemic periods (131 vs. 69/1,000 person-years). Demographic shifts as pandemics evolve should be considered in public health response planning.

The burden of disease due to influenza is not well characterized for children in developing countries and the effectiveness of available influenza vaccines in lower resource settings has not been established. We initiated a prospective, longitudinal, phase IV, household-randomized, controlled, observer-blinded three year study (2009-2011) in a rural community of India to measure the total and indirect household protective effects of immunizing children ages 6 months through 10 years with seasonal inactivated trivalent influenza vaccine (TIV) or a control vaccine (n=3697). Active weekly surveillance was conducted year round with home visits for identification of febrile acute respiratory illness (FARI) conducted for all vaccine recipients and household members (n=18,220). Nasal and throat swabs were collected from each FARI episode for influenza detection by real-time reverse transcription polymerase chain reaction. The primary outcome was reduction in laboratory confirmed influenza infections in the influenza vaccine versus control vaccine group, with secondary outcome assessing indirect effects among the entire study population. This report describes the study site, cluster study design, choice of study and control vaccines, and the initial enrollment in the study.

BACKGROUND: Healthcare workers (HCWs) universally have a poor uptake of influenza vaccination. However, no data are available from India. OBJECTIVE: To explore knowledge, attitudes, and practices associated with influenza vaccination in HCWs in a temperate climate area in northern India. PATIENTS AND METHODS: A self-administered questionnaire was offered to all HCWs in three major hospitals of Srinagar and information sought on motivations, perceptions, preferences and practices regarding influenza vaccination. RESULTS: Of the 1750 questionnaires received, 1421 (81%) were returned. Only 62 (4.4%) HCWs had ever received influenza vaccination even as 1348 (95%) believed that influenza poses adverse potential consequences for themselves or their contacts; 1144 (81%) were aware of a vaccine against influenza and 830 (58%) of its local availability. Reasons cited by 1359 participants for not being vaccinated included ignorance about vaccine availability (435; 32%), skepticism about efficacy (248; 18%), busy schedule (166; 12%), fear of side effects (70; 4%), and a perception of not being-at-risk (82; 6%). Sixty-one percent (865) believed that vaccine programs are motivated by profit. Eighty-eight percent opined for mandatory vaccination for HCWs caring for the high-risk patients, as a part of 'employee health program'. Most of the participants intended to get vaccinated in the current year even as 684 (48%) held that vaccines could cause unknown illness and 444 (31%) believed their adverse effects to be underreported. CONCLUSION: Influenza vaccination coverage among HCWs is dismally low in Srinagar; poor knowledge of vaccine availability and misperceptions about vaccine effectiveness, fear of adverse effects and obliviousness to being-at-risk being important barriers. Multifaceted, adaptable measures need to be invoked urgently to increase the coverage.

OBJECTIVE: Clinical case definitions used for influenza surveillance among hospitalized patients vary and need systematic evaluation. DESIGN, SETTING AND SAMPLE: During July 2009-August 2011, we collected clinical data and specimens (nasal and throat swabs) from rural patients hospitalized for acute medical illnesses. Specimens were tested by rRT-PCR for influenza viruses. MAIN OUTCOME MEASURES: Case definitions evaluated the following: influenza-like illness (ILI: measured fever plus cough or sore throat); severe acute respiratory illness (SARI: ILI with difficulty breathing in ≥5 years, Integrated Management of Childhood Illness-defined pneumonia or severe pneumonia, or physician diagnosed lower respiratory infection in <5 years); acute respiratory infection (ARI: ≥1 of cough, nasal discharge, difficulty breathing or sore throat); febrile acute respiratory illness (FARI: fever plus either cough, sore throat, runny nose, difficulty breathing, or earache). Variants that included "reported fever" and additional sign-symptom combinations were also evaluated. RESULTS: We enrolled 1043 hospitalized patients, including 257 children <5 years of age (range 1 day-86 years). Seventy-four patients tested influenza virus positive (including 28 A(H1N1)pdm09). Sensitivity(95% CI) and specificity (95% CI) for influenza infection were 78% (67-87) and 60% (57-63) for ILI (measured/reported fever); 37% (26-49) and 78% (75-80) for SARI (measured/reported fever); 82% (72-90) and 57% (54-60) for FARI (measured/reported fever); 88% (78-94) and 45% (42-49) for ARI; and 74% (63-84) and 61% (58-64) for measured/reported fever plus cough. Case definitions including only measured fever had lower sensitivity. CONCLUSION: ILI and FARI with measured/reported fever provided good balance between sensitivity and specificity among hospitalized patients. The simpler case definition of measured/reported fever plus cough is suited for field surveillance.

Pregnant women have abundant natural killer (NK) cells in their placenta, and NK cell function is regulated by polymorphisms of killer cell immunoglobulin-like receptors (KIRs). Previous studies report different roles of NK cells in the immune responses to placental malaria (PM) and human immunodeficiency virus (HIV-1) infections. Given these references, the aim of this study was to determine the association between KIR gene content polymorphism and PM infection in pregnant women of known HIV-1 status. Sixteen genes in the KIR family were analyzed in 688 pregnant Kenyan women. Gene content polymorphisms were assessed in relation to PM in HIV-1 negative and HIV-1 positive women, respectively. Results showed that in HIV-1 negative women, the presence of the individual genes KIR2DL1 and KIR2DL3 increased the odds of having PM, and the KIR2DL2/KIR2DL2 homozygotes were associated with protection from PM. However, the reverse relationship was observed in HIV-1 positive women, where the presence of individual KIR2DL3 was associated with protection from PM, and KIR2DL2/KIR2DL2 homozygotes increased the odds for susceptibility to PM. Further analysis of the HIV-1 positive women stratified by CD4 counts showed that this reverse association between KIR genes and PM remained only in the individuals with high CD4 cell counts but not in those with low CD4 cell counts. Collectively, these results suggest that inhibitory KIR2DL2 and KIR2DL3, which are alleles of the same locus, play a role in the inverse effects on PM and PM/HIV co-infection and the effect of KIR genes on PM in HIV positive women is dependent on high CD4 cell counts. In addition, analysis of linkage disequilibrium (LD) of the PM relevant KIR genes showed strong LD in women without PM regardless of their HIV status while LD was broken in those with PM, indicating possible selection pressure by malaria infection on the KIR genes.

The influenza virus neuraminidase (NA) protein primarily aids in the release of progeny virions from infected cells. Here, we demonstrate a novel role for NA in enhancing host cell survival by activating the Src/Akt signaling axis via an interaction with carcinoembryonic antigen-related cell adhesion molecule 6/cluster of differentiation 66c (C6). NA/C6 interaction leads to increased tyrosyl phosphorylation of Src, FAK, Akt, GSK3beta, and Bcl-2, which affects cell survival, proliferation, migration, differentiation, and apoptosis. siRNA-mediated suppression of C6 resulted in a down-regulation of activated Src, FAK, and Akt, increased apoptosis, and reduced expression of viral proteins and viral titers in influenza virus-infected human lung adenocarcinoma epithelial and normal human bronchial epithelial cells. These findings indicate that influenza NA not only aids in the release of progeny virions, but also cell survival during viral replication.