Respiratory Pathogens in Infants Diagnosed with Acute Lower Respiratory Tract Infection in a Tertiary Care Hospital of Western India Using Multiplex Real Time PCR.

OBJECTIVE: To determine the frequency of respiratory pathogens in infants diagnosed with acute lower respiratory tract infections. METHODS: A prospective cross-sectional observational study was conducted in infants hospitalized with a diagnosis of acute lower respiratory tract infection (ALRTI), in a tertiary care hospital in a metropolitan city of Western India. Nasopharyngeal swabs were analyzed by multiplex real time polymerase chain reaction, for 18 viruses and 3 bacteria (H. influenzae type b, C. pneumoniae and M. pneumoniae). The entire data was entered in Microsoft excel sheet and frequencies were determined. RESULTS: One hundred eligible infants were enrolled. Pathogens were detected in 82 samples, which included Respiratory syncytial viruses (RSV) A / B (35.4%), Human rhinovirus (25.6%), Adenovirus (22%), Human Parainfluenza viruses (11%), Human bocavirus (9.8), Human metapneumovirus A / B (8.5%), Influenza A (H1N1) pdm 09 (6.1%), Parechovirus (3.7%), Human coronaviruses (3.66%), Haemophilus influenzae type b (6.1%), Chlamydia pneumoniae (2.4%) and Mycoplasma pneumoniae (2.4%). Influenza A (other than H1N1), Influenza B, Human Coronavirus 229E and Enterovirus were not detected. The rate of coinfection was 34% and rhinovirus was the most common of the multiple pathogens. CONCLUSIONS: Spectrum of viral etiologies of ALRTI is highlighted. Etiological diagnosis of ALRTI would enable specific antiviral therapy, restrict antibiotic use and help in knowing burden of disease.

New clinical and seasonal evidence of infections by Human Parainfluenzavirus.

Human Parainfluenzaviruses (PIVs) account for a significant proportion of viral acute respiratory infections (ARIs) in children, and are also associated with morbidity and mortality in adults, including nosocomial infections. This work aims to describe PIV genotypes and their clinical and epidemiological distribution. Between December 2016 and December 2017, 6121 samples were collected, and submitted to viral culture and genomic quantification, specifically Parainfluenza 1-4 (PIV1-4), Influenza A and B, Respiratory Syncytial Virus (RSV) A and B, Adenovirus, Metapneumovirus, Coronavirus, Rhinovirus, and Enterovirus. Normalized viral load, as (log10) copies/103 cells, was calculated as virus Ct, determined by multiple qRT-PCR, as a function of the Ct of ß-globin. PIV was confirmed in 268 cases (4.37%), and linked to both upper and lower respiratory tract disease, being more frequent in children than in adults (5.23 and 2.43%, respectively). PIV1 and PIV3 were most common (31 and 32.5%, of total PIV positive samples, respectively), with distribution being similar in children and adults, as was viral load. PIV type was correlated with seasonality: PIV3 being more frequent in winter and spring, PIV1 in summer, and PIV 4 in fall. No correlation between vial load and clinical severity was found. Novel findings were that PIV viral load was higher in fall than in other seasons, and PIV4, classically linked to mild respiratory symptoms, was circulating, in children and adults, at all levels of symptoms throughout the year.

Dynamics of Sendai Virus Spread, Clearance, and Immunotherapeutic Efficacy after Hematopoietic Cell Transplant Imaged Noninvasively in Mice.

There are no approved vaccines or virus-specific treatments for human parainfluenza viruses (HPIVs), which have recently been reclassified into the species Human respirovirus 1, Human respirovirus 3, Human rubulavirus 2, and Human rubulavirus 4 These viruses cause morbidity and mortality in immunocompromised patients, including those undergoing hematopoietic cell transplant (HCT). No small-animal models for noninvasive imaging of respiratory virus infection in the HCT host exist, despite the utility that such a system would offer to monitor prolonged infection, its clearance, and treatment options. We used a luciferase-expressing reporter virus to noninvasively image in mice the infection of murine respirovirus (strain Sendai virus [SeV]), the murine counterpart of HPIV1. Independent of disease severity, the clearance of infection began approximately 21 days after HCT, largely due to the recovery of CD8+ T cells. Immunotherapy with granulocyte colony-stimulating factor (G-CSF) and adoptive transfer of natural killer (NK) cells provided a limited therapeutic benefit. Treatment with a fusion (F) protein-specific monoclonal antibody arrested the spread of lung infection and reduced the disease severity even when treatment was delayed to up to 10 days postinfection but had little observable effect on upper respiratory tract infection. Adoptive transfer of virus-specific T cells at 10 days postinfection accelerated the clearance by 5 days, reduced the extent of infection throughout the respiratory tract, and reduced the disease severity. Overall, the results support investigation of the clinical treatment of respiratory virus infection in the HCT host with monoclonal antibodies and adoptive T-cell transfer; the imaging system should be extendable to other respiratory viruses, such as respiratory syncytial virus and influenza virus.IMPORTANCE Parainfluenza viruses are a major cause of disease and death due to respiratory virus infection in the immunocompromised host, including those undergoing bone marrow transplantation. There are currently no effective treatment measures. We noninvasively imaged mice that were undergoing a bone marrow transplant and infected with Sendai virus, a murine parainfluenza virus (respirovirus). For the first time, we show the therapeutic windows of adoptive T-cell therapy and treatment with a monoclonal antibody to the fusion (F) protein in clearing Sendai virus from the respiratory tract and reducing disease severity. Mice tolerated these treatments without any detectable toxicity. These findings pave the way for studies assessing the safety of T-cell therapy against parainfluenza virus in humans. Adoptive T-cell therapy against other blood-borne viruses in humans has been shown to be safe and effective. Our model of noninvasive imaging in mice that had undergone a bone marrow transplant may be well suited to track other respiratory virus infections and develop novel preventive and therapeutic strategies.

[Clinical analysis of children with pertussis and significance of respiratory virus detection in the combined diagnosis].

Objective: To analyze the clinical data of children with pertussis and explore the necessity of respiratory virus detection in the combined diagnosis so as to improve the clinician's understanding and standardize the diagnosis and treatment of pertussis in children. Method: Clinical data and laboratory examination of 195 suspected pertussis children between Jan. 2015 and Dec. 2016 in Children's Hospital Affiliated to Capital Institute of Pediatric were analyzed retrospectively. Result: The nasopharyngeal secretions were collected from 195 suspected pertussis children, PCR was employed to detect the nucleic acid of Bordetella pertussis. Meanwhile, 172 of 195 cases were screened for antigens of 7 common respiratory viruses by direct immunofluorescence (DIF) (respiratory syncytial virus(RSV), adenovirus(ADV), influenza virus A and B, parainfluenza viruses type Ⅰ-Ⅲ). (1) Eighty cases were positive in pertussis nucleic acid detection (positive rate was 41.0%), 47 males and 33 females, age ranged from one month to ten years, all of them had paroxysmal cough (100.0%), 50 cases with spasmodic cough (62.5%), 9 cases with vomiting after cough(11.2%), 22 cases with cyanosis after cough(27.5%), 13 cases with roaring after cough(16.2%), 4 cases with dyspnea(5.0%), 18 cases were diagnosed as pneumonia by chest radiography(22.5%) and 1 case was diagnosed as pertussis encephalopathy(1.2%); (2) 172 cases of respiratory virus DIF detection were completed and 69 of them were positive(positive rate was 40.1%), including 32 cases positive for RSV(18.6%), 29 cases for PIVⅢ(16.8%); (3) In 80 confirmed pertussis children, 66 cases of respiratory virus DIF detection were completed and 9 were positive(9/66, 13.6%), including 7 cases positive for PIVⅢ. The clinical manifestations were cyanosis after cough(6 cases), dyspnea(2 cases) and pneumonia were diagnosed by chest radiography in 3 cases, the clinical symptoms of these children were more prominent than children with general pertussis; (4) Patients were divided into three groups according to the pathogens: 57 cases in single pertussis group, 32 cases in RSV infection group, 22 cases in single PIVⅢ infection group.The cases of spasmodic cough in Pertussis group was 35 (61.4%), RSV infection group was 7(21.9%), single PIVⅢ infection group was 8(36.4%), compared with the other two groups, the incidence of spasmodic cough were higher in Pertussis group (χ(2) =12.850, 4.013, P<0.05). The cases of roaring in Pertussis group was 11 (19.3%), RSV infection group was 1(3.1%), single PIVⅢ infection group was 0, and the incidence were higher in Pertussis group (χ(2)=4.596, 4.932, P<0.05). The cases of dyspnea in Pertussis group was 2 (3.5%), RSV infection group was 11(34.4%), single PIVⅢ infection group was 0, and the incidence was higher in RSV infection group (χ(2)=15.654, 9.487, P<0.01). Conclusion: Pertussis is common in children, especially in unvaccinated or incompletely vaccinated infants. The typical clinical manifestation is paroxysmal spasmodic cough; complicated with PIVⅢ infection is a risk factor for sever pertussis. Early detecting of Bordetella by PCR is helpful for the diagnosis of pertussis, RSV and PIVⅢ are the main pathogen for Pertussis-like syndrome. The detection of respiratory virus is helpful for differential diagnosis and medication guidance.

Development of SPR biosensor for simultaneous detection of multiplex respiratory viruses.

A surface plasmon resonance (SPR)-based biosensor was developed for specific detection of nine common respiratory virus, including influenza A and influenza B, H1N1, respiratory syncytial virus (RSV), parainfluenza virus 1-3 (PIV1, 2, 3), adenovirus, and severe acute respiratory syndrome coronavirus (SARS). The SPR biosensor was developed by immobilizing nine respiratory virus-specific oligonucleotides in an SPR chip. To increase the biosensor sensitivity, biotin was used to label the PCR primer and further amplify the signal by introducing streptavidin after hybridization. Throat swab specimens representing nine common respiratory viruses were tested by the innovative SPR-based biosensor to evaluate the sensitivity, specificity and reproducibility of this method. Results suggest that this biosensor has the potential to simultaneously identify common respiratory viruses.

Histochemical fluorescent staining of Sendai virus-infected cells with a novel sialidase substrate.

Sialidases, enzymes that remove terminal sialic acid residues, are pivotal in various biological processes such as malignancy and infection with pathogens. For histochemical staining of sialidase activity, we have developed a new synthetic sialidase substrate, sialic acid-conjugated fluorescent benzothiazolylphenol derivative (BTP3-Neu5Ac), for rapid, sensitive, and specific fluorescent staining of sialidase activity. Here, we showed the usefulness of BTP3-Neu5Ac for histochemical fluorescent staining of cells infected with Sendai virus (SV), which possesses sialidase activity. BTP3-Neu5Ac also visualised SV-infected regions of lung sections from SV-infected mice. We succeeded in histochemical fluorescent staining of SV both in vitro and in vivo. SV has been utilised in many virological and biotechnological studies such as developments of an oncolytic virus, a gene therapy vector, and a vaccine candidate. BTP3-Neu5Ac should contribute to rapid progress of such studies and researches on viral sialidase.

Multiplex real-time PCR for prompt diagnosis of an outbreak of human parainfluenza 3 virus in children with acute leukemia.

INTRODUCTION: Human parainfluenza virus type 3 (HPIV-3) causes significant morbimortality in immunocompromised patients. Outbreaks of severe pneumonitis have been previously described in this setting. MATERIALS AND METHODS: Retrospective observational study in children diagnosed with acute leukemia and a documented HPIV-3 infection in the context of a nosocomial outbreak occurred in a single center. RESULT: During summer 2012, an HPIV-3 infection was detected in six hospitalized children with acute leukemia. All the patients had respiratory symptoms and one of them suffered from parotitis. CONCLUSION: Early diagnoses using multiplex real-time polymerase chain reaction (PCR) let us control this outbreak. A phylogenetic analysis confirmed person-to-person transmission of a single HPIV-3 variant.

Use of data linkage to investigate the aetiology of acute lower respiratory infection hospitalisations in children.

AIM: To document the aetiology of acute lower respiratory infection (ALRI) hospitalisations in Western Australian children by linking population-based laboratory data with hospital morbidity data. METHODS: Data from all ALRI hospitalisations and laboratory records related to respiratory pathogens between 2000 and 2005 were extracted and linked through a population-based record linkage system. The proportion of specimens that were positive for each respiratory viral or bacterial pathogen was documented. RESULTS: Eight thousand nine hundred and eighty (45.2%) ALRI hospitalisations were linked to a laboratory record. Admissions to a private hospital and admissions from non-metropolitan areas were less likely to have a linked laboratory record. In 57.9% of linked hospitalisations, a respiratory virus and/or a bacterial pathogen was identified. Frequently identified viral pathogens included respiratory syncytial virus (RSV; n= 3226; 39.5% of those tested), influenza viruses (n= 664; 8.5%), parainfluenza virus type 3 (n= 348; 4.6%), picornaviruses (n= 292; 22.3%) and adenoviruses (n= 211; 2.7%). RSV was identified in 63.7% of bronchiolitis admissions in those aged under 6 months and 33.1% of pneumonia admissions in those aged under 12 months. Influenza viruses were identified in 81.6% of influenza-coded admissions. When a test was requested, Bordetella pertussis was identified in 21.2% of ALRI hospitalisations (n= 354), including 86.8% of whooping cough-coded admissions. CONCLUSIONS: This is the first report of population-based data linkage between statewide laboratory data and hospitalisation records and demonstrates proof of principle. RSV continues to be an important pathogen in ALRI. As pathogens were identified across all diagnoses, relying on hospital diagnosis coding alone may not accurately estimate the burden of different categories of ALRI.

Treatment of parainfluenza 3 infection with DAS181 in a patient after allogeneic stem cell transplantation.

Parainfluenza virus (PIV) can cause significant morbidity after allogeneic stem cell transplantation (SCT). We report the first use of inhaled DAS181 for PIV in an allogeneic SCT recipient. Symptoms, oxygenation, and pulmonary function tests improved. Nasopharyngeal samples showed a reduction in viral load. DAS181 should be systematically evaluated for severe PIV infection.

Human parainfluenza virus type 3 (HPIV 3) community-acquired pneumonia (CAP) mimicking pertussis in an adult: the diagnostic importance of hoarseness and monocytosis.

BACKGROUND: There are relatively few causes of acute community-acquired pneumonias (CAPs) in adults associated with prolonged cough. In adults the most common acute CAPs with a prominent and persistent nonproductive cough are due to Mycoplasma pneumoniae, Chlamydophilia (Chlamydia) pneumoniae, or Bordetella pertussis (pertussis). Pertussis is an underrecognized and underappreciated cause of CAP in adults. Different from classic pertussis in children, pertussis in adults presents with prolonged dry cough, that is, the "100-day cough." In pertussis, the characteristic nonspecific laboratory findings are leukocytosis and relative lymphocytosis. Dry cough accompanied by hoarseness with CAP in an adult should suggest C. pneumoniae or a respiratory virus (eg, influenza, parainfluenza, respiratory syncytial virus). METHODS: We present the case of a young woman who presented with a prominent and persistent pertussis-like cough with hoarseness. She had no leukocytosis or relative lymphopenia, which argued against the diagnosis of pertussis. Notably, she had persistent monocytosis. Her protracted pertussis-like cough that persisted during her hospitalization was so impressive that the diagnostic impression was pertussis. Direct fluorescent antibody (FA) and throat cultures were negative for pertussis. Furthermore, her hoarseness suggested the possibility of C. pneumoniae, but her C. pneumoniae immunoglobulin-M titer was negative. RESULTS: Because C. pneumoniae was ruled out, her hoarseness suggested a respiratory viral cause. A respiratory FA viral panel and viral throat cultures were obtained. The respiratory FA viral panel was negative for influenza A/B, respiratory syncytial virus, metapneumovirus, adenovirus, cytomegalovirus, and parainfluenza viruses. However, her viral throat cultures grew parainfluenza virus type 3 (HPIV 3), confirming the diagnosis. CONCLUSION: To the best of our knowledge, this is the first case of HPIV 3 CAP presenting with a prominent and persistent pertussoid cough in an adult mimicking pertussis with hoarseness and monocytosis.

Características clínicas y epidemiológicas de la infección por virus parainfluenza en niños hospitalizados / Clinical and epidemiological manifestations of parainfluenza infection in hospitalized children

Los virus parainfluenza del ser humano (VPIh) son patógenos importantes de enfermedad respiratoria en niños; pese a ello, existe escasa información publicada en Sudamérica dirigida a caracterizar esta infección. Objetivo: Describir las manifestaciones clínicas y epidemiológicas específicas de los VPIh en niños hospitalizados. Pacientes y Métodos: Se revisaron todas las hospitalizaciones respiratorias (HR) efectuadas en el Hospital de la Pontificia Universidad Católica, Santiago, Chile, durante el período 2001-2004 y sus respectivos estudios virales obtenidos de secreciones nasofaríngeas en aquellos con sospecha de infección viral. Resultados: Se identificaron 3.043 HR siendo 64 (2,1 por ciento) VPUrh La edad promedio fue 13 meses (rango: 1 m-12 a) siendo 77 por ciento) de edad inferior a dos años. VPIh-2 fue el serotipo prevalente (47 por ciento), observándose una tendencia estacional para los serotipos 2 y 3. Las presentaciones más frecuentes fueron sibilancias asociadas a virus (40 por cientoo) y neumonía (30 por ciento). Todas las bronquiolitis se presentaron asociadas a VPIh serotipos 2 y 3. Sólo 17 por ciento de los hospitalizados por VPIh+ (44 por ciento VPIh-1) desarrollaron laringitis. Conclusión: Virus parainfluenza humano puede ser responsable de HR en niños, mostrando una tendencia estacional VPIh-2 y el serotipo 3. Aunque son poco frecuentes como causa de HR, confirmamos su participación como etiología específica de laringitis, bronquiolitis y neumonía, especialmente en niños pequeños.

Background: Human parainfluenza viruses (hPIV) are a common cause of respiratory illness of children but published data on clinical characteristics of hPIV infection in South America is scarce. Objective: To review the clinical presentation and epidemiological features of hPIV in a series of hospitalized children in Chile. Patients and Methods: Retrospective review of clinical charts from all pediatric admissions with a diagnosis of respiratory disease (between January 2001 to December 2004) at the Catholic University Hospital, Santiago, Chile. Nasopharyngeal secretions were tested for hPIV in children admitted with suspected respiratory viral infections. Results: A total of 3,043 respiratory admissions were recorded during the study period; 64 children (2.1 percent) were hPIV positive. Average age was 13 months (range: lm to 12y) and 77 percent> were younger than 2 years. HPIV-2 was the most common type identified (47 percent). A seasonal trend was noted for serotypes hPIV-2 and 3. Acute wheezing (40 percento) and pneumonia (30 percent) were the most common clinical diagnosis in hPIV positive children and 17 percent> hPIV positive children (44 percent> for hPIV-1) were associated with laryngitis. All hPIV positive bronchiolitis were due to serotypes hPIV-2 and 3. Conclusion: hPIV can cause respiratory disease requiring hospitalization; serotypes hPIV-2 and 3 displayed a seasonal trend. Although hPIV is an uncommon cause of severe respiratory infecion requiring hospitalization in children, it should be considered in the differential diagnosis of laryngitis, bronchiolitis and pneumonia, especially in younger children.

Respiratory virus infection in infants and children.

The recent availability of diagnostic polymerase chain reaction-based assays for the detection of viral pathogens has allowed us to identify the infectious agents present in a large percentage of culture-negative specimens. The information gained by these studies has led to a reassessment of the prevalence of individual viruses in lower respiratory tract infections in both normal and immunocompromised children. Pathologic examination of less severe cases identified by more sensitive methodologies in immunocompetent children is now possible and has prompted the realization that many of the classic features of particular viral infections in the lung are in fact hallmarks of fulminant disease. In this article we review the body of literature discussing these anatomic findings, the approaches taken by those investigators, and the types of reagents that are commercially available.

[Virology: the contribution of molecular biology in the microbiologic diagnosis of pediatric respiratory diseases]. / Virologie : l'apport de la biologie moléculaire dans le diagnostic microbiologique en pneumopédiatrie.

The conventionnal tools used for virological diagnosis include direct antigen detection by immunofluorescence (IFA) or an immunoenzymatic test (EIA), and viral isolation technique (VIT). In most cases, IFA and EIA have a slightly lower sensitivity than VIT but are also able to detect some VIT-negative samples. Results of several teams using RT-PCR technologies show that the molecular methods detect more positive cases than the conventional tools. Work is under way to expand the number of viruses detected by multiplex RT-PCR and to determine wether newly discovered viruses, such as human metapneumovirus, contribute to burden of paediatric lower respiratory infections. In conclusion, according to requirements of speed, low cost of the methods, and to achieve the highest rate of detection of respiratory viruses, the combined use of IFA and multiplex RT-PCR is today likely to be the best way to improve diagnosis of respiratory illnesses in children.

[Use of polymeric suspensions as a viral sorbent to detect cattle serum antibodies].

The paper shows it possible to use stained polymeric microspheres, 1.7 microm in diameter, that contain viruses onto the surface, in the latex agglutination test to detect antibodies to the bovine serum viruses of infective rhinotracheitis, parainfluenza-3, viral diarrhea, respiratory syncytial infection, and adenoviral infection.

Respiratory disease due to parainfluenza virus in adult leukemia patients.

Reports of human parainfluenza viruses (HPIV) in patients with leukemia have been limited to a few cases or as a portion of general surveys. In order to expand the knowledge of these infections in this patient group, the frequency and clinical course of HPIV infections was determined among 1,787 patients with leukemia treated at The University of Texas M.D. Anderson Cancer Center between July 1994 and December 1997. HPIV was isolated from 47 (6.2%) of the 770 patients who were cultured for respiratory viruses. HPIV type 3 accounted for 39 of the 47 HPIV infections. Twenty-six patients developed pneumonia, and the associated mortality was 27%. Multivariate analysis revealed that a low absolute lymphocyte count and pneumonia were associated with increased mortality. Concurrent respiratory and other infections were associated with an increased frequency of pneumonia. Only five patients with pneumonia received antiviral therapy and four of them survived the infection. HPIV infection in leukemic patients is frequently associated with pneumonia and the mortality rate from pneumonia is substantial among lymphopenic patients.

Human parainfluenza virus giant cell pneumonia following cord blood transplant associated with pulmonary alveolar proteinosis.

Giant cell pneumonia secondary to human parainfluenza virus 3 has been reported only rarely in immunocompromised hosts. The few cases documented after bone marrow transplant have resulted in significant morbidity and mortality. To our knowledge, this entity has not been described following umbilical cord blood transplant. Pulmonary alveolar proteinosis, a rare condition that has been reported with increasing frequency in association with immunocompromise and infections, has not been documented in the setting of either umbilical cord blood transplant or human parainfluenza viral infection. We report what we believe is the first documented case of giant cell pneumonia caused by human parainfluenza virus 3 in an umbilical cord blood transplant recipient. To our knowledge, a unique associated feature of this case, a pulmonary alveolar proteinosis-like reaction, has not been reported previously in association with human parainfluenza virus pneumonia.

Guillain-Barré syndrome after allogeneic hematopoietic stem cell transplantation.

Guillain-Barré syndrome is a rare complication in the setting of hematopoietic stem cell transplantation. We report three children with T cell lymphoma/leukemia in whom this syndrome developed soon after they received unrelated donor transplants. The rapid onset of symptoms raises the concern that the bone marrow transplant conditioning regimen (ie, total body irradiation, cyclophosphamide and cytosine arabinoside) might have precipitated the clinical syndrome of ascending polyneuropathy. Although central nervous system toxicity has been well described with high-dose cytosine arabinoside therapy, peripheral neuropathy of the Guillain-Barré type has been reported only infrequently. We review possible factors contributing to the development of this syndrome in these three patients.