Human viral nucleic acids concentrations in wastewater solids from Central and Coastal California USA

Pathogen nucleic acids data in wastewater solids from 147 treatment plants in the United States: 2024–2025

We measured concentrations of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its variants, influenza A and B viruses, respiratory syncytial virus, human metapneumovirus, enterovirus D68, human parainfluenza types 1, 2, 3, 4a, and 4b in aggregate, norovirus genotype II, rotavirus, Candida auris, hepatitis A virus, human adenovirus, mpox virus, H5 influenza A virus, and pepper mild mottle virus nucleic acids in wastewater solids prospectively at 191 wastewater treatment plants in 40 states across the United States plus Washington DC. Measurements were made two to seven times per week from 1 January 2022 to 30 June 2024, depending on wastewater treatment plant staff availability. Measurements were made using droplet digital (reverse-transcription–) polymerase chain reaction (ddRT-PCR) following best practices for making environmental molecular biology measurements. These data can be used to better understand disease occurrence in communities contributing to the wastewater.

Enteric infections are important causes of morbidity and mortality, yet clinical surveillance is limited. Wastewater-based epidemiology (WBE) has been used to study community circulation of individual enteric viruses and panels of respiratory diseases, but there is limited work studying the concurrent circulation of a suite of important enteric viruses. A retrospective WBE study was carried out at two wastewater treatment plants located in California, United States. Using digital droplet polymerase chain reaction (PCR), we measured concentrations of human adenovirus group F, enteroviruses, norovirus genogroups I and II, and rotavirus nucleic acids in wastewater solids two times per week for 26 months (n = 459 samples) between February 2021 and mid-April 2023. A novel probe-based PCR assay was developed and validated for adenovirus. We compared viral nucleic acid concentrations to positivity rates for viral infections from clinical specimens submitted to a local clinical laboratory to assess concordance between the data sets. We detected all viral targets in wastewater solids. At both wastewater treatment plants, human adenovirus group F and norovirus GII nucleic acids were detected at the highest concentrations (median concentrations greater than 105 copies/g), while rotavirus RNA was detected at the lowest concentrations (median on the order of 103 copies/g). Rotavirus, adenovirus group F, and norovirus nucleic acid concentrations were positively associated with clinical specimen positivity rates. Concentrations of tested viral nucleic acids exhibited complex associations with SARS-CoV-2 and other respiratory viral nucleic acids in wastewater, suggesting divergent transmission patterns.IMPORTANCEThis study provides evidence for the use of wastewater solids for the sensitive detection of enteric virus targets in wastewater-based epidemiology programs aimed to better understand the spread of enteric disease at a localized, community level without limitations associated with testing many individuals. Wastewater data can inform clinical, public health, and individual decision-making aimed to reduce the transmission of enteric disease.

A comparative analysis of confirmed cases of human influenza virus (HIFV), human respiratory syncytial virus (HRSV), and human metapneumovirus(HMPV) was conducted to describe their clinical and epidemiological characteristics.During 2009-2021, active surveillance of acute respiratory infections (ARIs) was performed in nine provinces of China. Clinical and epidemiological information and laboratory testing results of HIFV, HRSV, and HMPV were analyzed.Among 11591 ARI patients, the single-infection rates of HIFV, HRSV, and HMPV were 15.00%, 9.59%, and 2.24%, respectively; the coinfection rate of these three viruses was 0.64%. HIFV infection was mainly in adults aged 15-59 years, accounting for 39.10%. HRSV and HMPV infections were mainly in children under 5 years old, accounting for 87.13% and 83.46%, respectively. Patients with HRSV infection were younger than HMPV. HRSV and HMPV had high similarities in clinical manifestations, presenting with lower respiratory symptoms. HIFV mainly presented with an upper respiratory infection. The epidemic peak of HRSV was earlier than that of HIFV, and that of HMPV was later than those of HRSV and HFIV. A total of 85.14% of coinfection cases were children under 5 years old. Coinfection might increase the risk of pneumonia in HIFV cases. During 2020-2021, the positive rates and seasonal patterns of these three viruses changed due to the impact of the COVID-19 pandemic.Certain clinical and epidemiological features were observed in HIFV, HRSV, and HMPV infections, which could be beneficial for guiding clinical diagnosis, treatment, and prevention of these three viruses in China.

1. Asif Noor, MD* 2. Theresa Fiorito, MD* 3. Leonard R. Krilov, MD*,† 1. *Department of Pediatrics, Children's Medical Center, NYU Winthrop Hospital, Mineola, NY 2. †Department of Pediatrics, State University of New York, Stony Brook School of Medicine, Stony Brook, NY * Abbreviations: AAP: : American Academy of Pediatrics Adv: : adenovirus CDC: : Centers for Disease Control and Prevention FDA: : Food and Drug Administration hMPV: : human metapneumovirus PCR: : polymerase chain reaction PIV: : parainfluenza virus RSV: : respiratory syncytial virus RV: : rhinovirus Clinicians must learn to identify viral infections in children during the winter months and must practice caution with the use of unnecessary medications in such cases. Recognition of the clinical pattern of viral infection (eg, bronchiolitis) in conjunction with judicious use of viral tests (either office-based immunoassays or newer molecular tests) may assist in epidemiological monitoring, cohorting patients in the hospital, withholding unnecessary therapies, and providing a definitive diagnosis. After completing this article, readers should be able to: 1. Review the epidemiological aspects and clinical signs and symptoms of common cold weather viruses. 2. Recognize situations in which viral testing is indicated. 3. Recognize situations in which treatment is indicated. In early November you are evaluating a 9-month-old boy born at 33 weeks of gestation. The infant presents with 2 days of fevers (101°F–102°F [38.3°C–38.8°C]), copious rhinorrhea, and 1 day of coughing with difficulty breathing. He is otherwise feeding well and has had adequate urination. His 4-year-older sister has an upper respiratory tract infection. On physical examination, the infant has a respiratory rate of 45 breaths/min without chest wall retractions. On auscultation there is good air entry with scattered rhonchi bilaterally. What is the most appropriate next step in management? 1. Obtain respiratory syncytial virus (RSV) and influenza antigen testing. 2. Obtain a chest radiograph to look for focal infiltrate. 3. Provide supportive care with nasal …

Underestimation of Influenza Viral Infection in Childhood Asthma Exacerbations

Full-term and preterm infants admitted to neonatal intensive care units (NICUs) face a high risk of infections, due to the immaturity of their innate and adoptive immune systems, inadequate protection through maternal immunity and the need for repeated invasive procedures (1). In recent years, our knowledge of viral infections in neonates has increased, due to multiplex polymerase chain reaction (PCR)-based techniques. It is now possible to screen for as many as 17 viruses from a single mucus sample (2). However, the role of viral respiratory tract infections in the symptoms of infants admitted to NICUs at birth is still poorly understood. In a recent review, 32 respiratory viral outbreaks in NICUs were reported (3). These were caused by several different respiratory viruses, including respiratory syncytial virus (RSV) (89 patients in 11 outbreaks), enteroviruses (101 patients in 10 outbreaks) and adenovirus (79 patients in six outbreaks). In addition, outbreaks of coronavirus, rhinovirus, influenza A virus and parainfluenza virus infections have been reported (4–7). An epidemic may result in the temporary closure of a NICU. Recently, a NICU was closed for 28 days after adenovirus type 19 caused an outbreak of keratoconjunctivitis, which affected 12 NICU infants, two NICU staff members, two relatives of patients and two members of the ophthalmologic team (8). We report on an observational study on the use of a multiplex PCR in a NICU in Finland. The study was carried out at Turku University Hospital, the only tertiary level NICU in south-west Finland, which serves a population of about 750 000. An average of 600–700 infants are admitted to the NICU annually, resulting in approximately 6000 patient care days per year. About one-third of the infants are preterm, and 50–85 very low birth weight infants are treated annually. Two to four patients are treated in one room. Parents are encouraged to stay with their infant and provided with unlimited access. Visitors with respiratory tract infection symptoms are not allowed into the NICU, and siblings under school age cannot visit if there is an RSV outbreak in the community. Otherwise, healthy siblings are allowed access. During the study period from 1 January 2009 to 30 June 2011, 1589 infants were admitted to the unit and 76 (5%) were evaluated for respiratory viruses. A nasopharyngeal aspirate was taken from infants if they presented with symptoms of respiratory infection, such as rhinorrhea, sneezing, increased bronchial secretions, episodes of bradycardia and/or desaturations, or if they had been exposed to someone with a respiratory infection. We collected and analysed 139 samples from 76 infants for this study. Nasopharyngeal aspirates were collected in sterile tubes and analysed on the same or following day. Nucleic acids were extracted from the aspirates using Nuclisense easyMag extractor (Biomerieux, Boxtel, the Netherlands). Respiratory virus genomes were detected using Seeplex RV12 multiplex PCR assay for adenovirus, influenza A and B viruses, parainfluenza types 1–3 viruses, RSVa, RSVb, rhinovirus, human metapneumovirus, and coronaviruses 229E/NL63 and OC43/HKU1 (Seegene, Seoul, Korea). In addition, specimens were tested in a real-time PCR assay for enteroviruses, rhinovirus and RSV (9). During the two-and-a-half-year study period, no outbreaks of viral respiratory tract infection occurred in the NICU. Of the 139 samples taken from 76 infants, 28 samples (20%) from 15 infants (20%) were positive for one or more viruses. Six babies were positive for rhinovirus, five for parainfluenza type 3 virus, one for parainfluenza type 2

The burden of severe human metapneumovirus (HMPV) respiratory tract infections (RTIs) in European children has not been clarified. We assessed HMPV in Norwegian children and compared hospitalization rates for HMPV and respiratory syncytial virus (RSV). We prospectively enrolled children (<16 years old) hospitalized with RTI and asymptomatic controls (2006-2015). Nasopharyngeal aspirate samples were analyzed with polymerase chain reaction (PCR) tests for HMPV, RSV, and 17 other pathogens. We genotyped HMPV-positive samples and assessed shedding time in 32 HMPV-infected children. In children with RTI, HMPV was detected in 7.3% (267 of 3650) and RSV in 28.7% (1048 of 3650). Among controls, 2.1% (7 of 339) had low HMPV levels detected by PCR, but all were culture negative. HMPV primarily occurred from January to April and in regular epidemics. At least 2 HMPV subtypes occurred each season. The average annual hospitalization rates in children <5 years old with lower RTI were 1.9/1000 (HMPV) and 10.4/1000 (RSV). Among children with RTI, the median HMPV shedding time by PCR was 13 days (range, 6-28 days), but all were culture negative (noninfectious) after 13 days. HMPV appears in epidemics in Norwegian children, with a hospitalization rate 5 times lower than RSV. Low levels of HMPV are rarely detected in healthy children.

To study the epidemiological characteristics of respiratory virus infection and its relations to climatic factors in Suzhou. From 2006 to 2009, viral etiology surveillance was conducted among 6655 children hospitalized with acute respiratory tract infections (ARIs). Direct immunofluorescence method was used to test respiratory secretion samples for respiratory syncytial virus (RSV), influenza viruses A and B (Inf-A, Inf-B), parainfluenza virus types I, II, and III (Pinf-I, Pinf-II, Pinf-III) and adenovirus. Samples were tested for human metapneumovirus (hMPV) with reverse transcription polymerase chain reaction (RT-PCR). Samples from Jan 2006 to Dec 2009 were also tested for human bocavirus (HBoV). Climatic factors, including mean temperature, relative humidity, rainfall amount, sum of sunshine and mean wind velocity were collected monthly. The relationship between activity of each virus and climatic factors were analyzed by linear regression and stepwise regression analysis. From 2006 to 2009, in the total virus detection rate was 32.2% (2142/6655) in Suzhou. RSV was the most common virus and the average detection rate was 15.7% (1048/6655), followed by hMPV 8.9% (596/6655), HBoV 7.8% (148/1883), Pinf-III 2.7% (183/6655), Inf-A 2.4% (161/6655), ADV 1.3% (89/6655), Pinf-I 0.4% (29/6655), Inf-B 0.37% (25/6655) and Pinf-II 0.16% (11/6655). The positive rates of RSV, hMPV and ADV were significantly different in four years (χ(2) = 17.71, 33.23, 8.42, all P values < 0.05). Different virus has different epidemiological characteristics and distinct seasonality. The detection rate of RSV, hMPV, Inf-A were higher in Winter as 37.2%, 13.2%, 4.4%, respectively. ADV and Pinf-III were higher in summer as 2.3% and 4.6% respectively. The peak of HBoV existed in Autumn as 3.3%. The total virus detection rate showed significant inverse correlation with month average temperature (r = -0.732, P < 0.001) and a weak inverse correlation with average wind velocity was also found (r = -0.36, 0.01 < P < 0.05). The highest month total virus detection rate was from 47.6% to 84.4% when average temperature was from 3.2°C to 9.4°C and mean wind velocity was from 1.2 - 1.9 m/s. The associations of average temperature, sum of sunshine and wind velocity with RSV activity were statistical significant (r = -0.88, P < 0.001; r = -0.43, P < 0.01; r = -0.47, P < 0.01). The highest rate was from 24.3% to 58.2%, when mean temperature was from 5.3°C to 19.9°C, mean wind velocity was from 1.3 - 2.4 m/s and sum of sunshine was 61.0 to 153.4 hours. hMPV detection rate was inversely correlated with mean temperature and rain account (r = -0.43, P < 0.01; r = -0.29, P < 0.05). The rate was highest from 11.7% to 31.6% when mean temperature was from 5.3°C to 21.9°C and rain account was from 27.5 millimeter to 150.9 millimeter. Only mean temperature was positively correlated with Pinf-III (r = 0.53, P < 0.001). The rate was from 2.8% to 7.2% when mean temperature was between 11.9°C and 30.4°C. ADV detection rate was positively correlated with mean temperature and sum of sunshine, but negatively correlated with wind velocity (r = 0.35, P < 0.05; r = 0.30, P < 0.05; r = -0.32, P < 0.05). The rate was from 2.2% to 6.6% when mean temperature was between 15.9°C and 30.4°C, and sum of sunshine between 93 hours to 240.7 hours and mean wind velocity was from 1.1 - 2.8 m/s. Average temperature and relative humidity showed interactions on the detection rate of ADV (r = 0.36, P = 0.0093; r = -0.34, P = 0.016), but temperature showed higher effect on ADV detection rate. ADV detection rate was high at higher temperature (15.9 - 30.4°C) and low humidity (56% - 71%). RSV was one of the most common viruses among hospitalized children in Suzhou, and hMPV and HBoV also played an important role in respiratory tract infection of children. Different virus has different cycle and seasonality. Climatic factors, especially mean temperature, was the main factor affecting the virus prevalence.

Serological cross-reactivity of members of the Metapneumovirus genus

Objective To explore the viral pathogens in hospitalized children with acute respiratory tract infection in Weifang. Methods Nasopharyngeal secretion (NPS) samples were collected from hospitalized patients with acute respiratory tract infection from July 2016 to June 2017. The NPS samples were detected for 16 respiratory virus types/subtypes including influenza A virus (FluA), influenza B virus (FluB), seasonal influenza A HlNl virus (sH1N1), parainfluenza virus types 1, 2 and 3 (PIV-1, 2, 3), respiratory syncytial virus A (RSVA), respiratory syncytial virus B (RSVB), human rhinovirus (HRV), adenovirus (ADV), human metapneumovirus (HMPV), four coronavirus sybtypes (Cov-NL63, 229E, OC43, HKUl) and human bocavirus (BoV) by multiplex reverse transcription polymerase chain reaction (RT-PCR) assays based on automatic capillary electrophoresis. Results A total of 769 children with respiratory tract infection were enrolled, and the overall positive rate for the 16 common respiratory tract viruses was 33.68% (259/769). The positive rates were as follows: RSV (9.23%), PIV (7.93%), Flu (6.89%), HRV (4.68%), ADV (3.38%), HMPV (1.69%), CoV (0.91%), BoV (0.65%). The positive rate of viral detection showed significant differences among different age groups (χ2=8.724, P=0.033), and the highest positive rate was noted in the age group of 6 months to 1 year. The overall positive rate of viral detection showed a significant difference in terms of seasonal distribution, with a peak prevalence in winter. Conclusions RSV was the main respiratory tract virus among children in Weifang, especially in winter and spring. The distribution of viruses in children with respiratory tract infection was associated with age and season. Key words: Acute respiratory tract infection; Respiratory virus; Children

Objective To explore the viral pathogens of acute respiratory tract infection in Shanghai, and to provide reference data for diagnosis and treatment. Methods Between January 2012 and December 2013, 643 nasopharyngeal aspiration (NPA) samples were collected from children who had been hospitalized for acute respiratory tract infection at Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine, the Ninth People′s Hospital, Affiliated to Medical College of Shanghai Jiaotong University School and Putuo District Center Hospital Shanghai, China.Human rhinoviruses, Respiratory syncytial virus, Adenovirus, Influenza viruses A and B, Parainfluenza viruses 1-4, Human metapneumovirus, Human bocavirus, Human coronavirus HKU1 and Human coronavirus NL-63 were detected by reverse transcirption-polymerase chain reaction (RT-PCR) and standard polymerase chain reaction (PCR) methods.Epidemiology and clinical manifestations of each virus were analyzed. Results Six hundred and forty-three NPAs samples were collected from children (402 boys, 241 girls) with ARTI, The mean age was 12 months(range from 11 days to 12 years). Viral pathogens were identified in 369 (57.4%) samples.RSV was the highest detection rate in 2012 and ADV was the highest detection rate in 2013, HCoV-NL63 was not detected.6 months to 1 year old age group virus detection rate was highest, along with the age growth, virus detection rate was falling, virus detection throughout the year, diagnosis includes upper and lower respiratory tract infection. Conclusions Viral pathogens are the main etiology of ARTI in children in Shanghai area.RSV and ADV are the most frequent viral pathogens. Key words: Children; Acute respiratory tract infections; Viral etiology; Reverse transcription polymerase chain reaction; Polymerase chain reaction

BACKGROUND AND OBJECTIVES:Human metapneumovirus (hMPV) and the Netherlands human coronavirus (HCoV-NL63) have been isolated from children with respiratory tract infection. The prevalence of these viruses has not been reported from Saudi Arabia. We sought to determine whether hMPV and HCoV-NL63 are responsible for acute respiratory illness and also to determine clinical features and severity of illness in the hospitalized pediatric patient population.DESIGN AND SETTING:Prospective hospital-based study from July 2007 to November 2008.PATIENTS AND METHODS:Nasopharyngeal specimens from children less than 16 years old who were suffering from acute respiratory diseases were tested for hMPV and HCoV-NL63 by reverse transcriptase–polymerase chain reaction. Samples were collected from July 2007 to November 2008.RESULTS:Both viruses were found among Saudi children with upper and lower respiratory tract diseases during the autumn and winter of 2007 and 2008, contributing to 11.1% of all viral diagnoses, with individual incidences of 8.3% (hMPV) and 2.8% (HCoV-NL63) among 489 specimens. Initial symptoms included fever, cough, and nasal congestion. Lower respiratory tract disease occurs in immunocompromised individuals and those with underlying conditions. Clinical findings of respiratory failure and culture-negative shock were established in 7 children infected with hMPV and having hematologic malignancies, myelofibrosis, Gaucher disease, and congenital immunodeficiency; 2 of the 7 patients died with acute respiratory failure. All children infected with HCoV-NL63 had underlying conditions; 1 of the 4 patients developed respiratory failure.CONCLUSION:hMPV and HCoV-NL63 are important causes of acute respiratory illness among hospitalized Saudi children. hMPV infection in the lower respiratory tract is associated with morbidity and mortality in immunocompromised children. HCoV-NL63 may cause severe lower respiratory disease with underlying conditions.

Objective To study the characteristics of etiology of lobar pneumonia in hospitalized children. Methods Medical history and sputum specimens were collected from 1 179 hospitalized children with lobar pneumonia from January 2006 to December 2015. Multiple pathogenic joint detection combined with the history data were used for analysis. Seven kinds of common respiratory virus were detected by direct immunofluorescence. Mycoplasma pneumoniae (MP), Chlamydia pneumoniae (CP) and human Bocavirus (hBoV) were detected by fluorescence quantitative polymerase chain reaction (PCR). Human Rhinovirus (HRV) and human Metapneumovirus (hMPV) were detected by reverse transcription PCR. Aspirates were cultured for bacteria. MP specific antibody IgG and IgM were tested by enzyme-linked immunosorbent assay (ELISA). Positive rates of each group were compared by χ2 test or Fisher exact test. Results Total etiology detection rate of lobar pneumonia in hospitalized children was 83.9% (989/1 179). The etiology detection rate of MP, virus, bacteria and streptococcus pneumoniae (SP) were 74.0%, 14.2%, 18.3% and 12.2%, respectively. The virus detection rate in 1—3 years old group was the highest, and that in ≥6 years old group was lower than other group (χ2=70.095, P 0.05). The MP detection rate was above 70% in every season. The detection rates of SP and hBoV were basically the same in every season. The detection rate of HI was higher in spring, Pinf 3 and SA were higher in summer, HRV was higher in autumn, and respiratory syncytial virus (RSV) and moraxella catarrhalis (MC) were higher in winter. Conclusions Lobar pneumonia occurs more common in elder children. MP is the major pathogen of lobar pneumonia, and SP is the second. The MP detection rate increases with age. The pathogen detection rate varies with age, but the effect of seasonal factor is not obvious on pathogen detection in lobar pneumonia. Key words: Pneumonia, lobar; Etiology; Age; Season; Children

Objective The study was to investigation the contribution of 9 respiratory viruses and clinical features of refractory pneumonia in children in Lanzhou area. Methods In total, 100 bronchoalveolar lavage fluid were collected from children younger than 14 years with refractory pneumonia between January 2013 and December 2013. The polymerase chain reaction (PCR) was used to screen human respiratory syncytial virus (HRSV), human metapneumovirus (HMPV), influenza viruses A and B (IFVA, IFVB), parainfluenza virus types 1 to 3 (PIV1-3), human rhinoviruses (HRVs), and human coronaviruses (HCoV-NL63 and HKU1), enterovirus(EV) using a standard reverse-transcription PCR technique, adenovirus(AdVs) and human Bocavirus (Bcov) using traditional PCR methods. All PCR-positive products were sequenced. Results Viral agents were identified in 53% (53/100) of specimens, including AdV in 40%(40/100), followed by RSV in 18%(18/100), IFVB in 5% (5/100), HRV in 4%(4 /100), EV-68 was found in one case. Among AdV positive cases, AdV7 (33/40) was detected most frequently, followed by AdV2 in 15% (6/40), AdV1 in 2.5%(1/40), The coinfection rate of positive sample was 24.53%(13/53). Conclusions Viruses play an important role in refractory pneumonia in children in Lanzhou area. AdV is an important viral agent in children with refractory pneumonia in Lanzhou City, especially AdV7. The frequency of fever, respiratory failure, cyanosis, and pulmonary consolidation were significant between AdV positive sample and negative sample. Key words: Refractory pneumonia; PCR; Viral pathogens; Children