Aim of the Study: Wastewater-based epidemiology is a powerful tool for monitoring the circu lation of pathogens in the population, including both asymptomatic and symptomatic infections. Human adenovirus (HAdV) is an enteric virus, responsible for various clinical manifestations, including upper respiratory tract infections, conjunctivitis, gastroenteritis, bronchitis and pneumonia, urinary tract infections, and skin infections. Occasionally, adenoviruses can cause more severe illnesses, including meningitis, encephalitis, and myocarditis. Adenovirus can be spread through water, making it a potential waterborne pathogen. Human HAdVs are classified into seven species (A-G) and more than 110 types. Species A and B are responsible for common respiratory infections, while species F includes serotypes 40 and 41, which are recognised as some of the most common causes of gastroenteritis. Methods: From 9 to 13 of January 2023, a total of 170 sewage samples were collected in 165 wastewater treatment plants (WTPs) in 18 regions and 2 autonomous provinces of Italy. HAdV was detected by amplification of a fragment of the HAdV hexon gene (714 to 855 bp long, depending on the HAdV type). Amplicons were sequenced by both Sanger (individual samples) and next-generation sequencing (in a pool of all PCR amplicons), using the Oxford Nanopore Technology (MinION platform). The long reads obtained by NGS were compared with 110 reference prototypes belonging to the Species HadV-A (n°=4), HadV-B (n°=17), HAdV-C (n°=7), HAdV-D (n°=78), HAdV-E (n°=1), HAdV-F (n°=2), and HAdV-G (n.=1). Results and Conclusions: The results showed a wide diversity of HAdVs in wastewater, indicating genetic diversity within the population at a national level. The nested PCR results showed the presence of HAdVs in 118 out of 170 samples (69.4%). Seventy-six samples were successfully sequenced and characterised by conventional Sanger sequencing as belonging to 5 HAdV Species and 7 types: Species A (HAdV-12, 2 samples), Species B (HAdV-3, 33 samples), Species C (HAdV-1, 2 and 89, 6 samples), Species D (HAdV-45, 1 sample), and Species F (HAdV 41, 34 samples). NGS analysis confirmed the results obtained by Sanger sequencing but was able to detect additional less represented types: HAdV-B21, HAdV-C5, HAdV-C89, HAdV-D23, HAdV-D46, HAdV-D49, HAdV-D83, and HAdV-F40. Other sequences could not be definitively assigned because they shared the same % nt identity with more than one HAdV prototype: HAdV-B7 and B66; HAdV- C1 and C104; HAdV- D45 and D73; HAdV- D48 and D81. Our findings highlight the diversity of HAdV species and types present in raw sewage, with the potential to impact surface waters and pose a significant threat to human health. By using next-generation sequencing, we were able to identify a broad range of “less prevalent” types in addition to those identified by Sanger sequencing. The results show that although the exon region is recognised as being able to discriminate among different types of human adenovirus, it is not actually able to identify certain types that have a high degree of nucleotide identity, particularly those belonging to the D Species.
THE USE OF NANOPORE SEQUENCING TECHNOLOGY PROVIDES NEW INSIGHTS INTO THE WIDE RANGE OF HUMAN ADENOVIRUSES PRESENT IN ASTEWATER / Bonanno Ferraro, G.; Mancini, P.; Veneri, C.; Del Giudice, C.; Iaconelli, M.; Brandtner, D.; Bucciarelli, Giuseppe; Torlontano, Paolo; Aprea, Giuseppe; Scattolini, Silvia; Scattolini, Silvia; D’Angelantonio, Daniela; Migliorati, Giacomo; La Bianca, Michele; Anna Cifarelli, Rosa; Palma, Achille; e Giuseppe Lauria, Giovanna La Vecchia; Brienza, Rosanna; Montenegro, Patrizia; Malacaria, Eduardo; Folino, Giuseppe; Iannone, Michelangelo; Casaburi, Filomena; Bulotta, Giorgia; Barillari, Emanuela; Dragone, Melania; Sacco, Iolanda; Tomaino, Carmine; Felicetta, Cristina; Calabria, Adelaide; Varcasia, Ottavia; Stefanizzi, Francesca; Vizza, Concetta; D’Argenzio, Angelo; Cossentino, Luigi; Olivares, Renato; Pizzolante, Antonio; Fusco, Giovanna; Tosco, Alessandra; Porta, Amalia; Pennino, Francesca; Maria, Triassi; Angelini, Paola; Gentili, Lisa; De Lellis, Laura; Nasci, Daniele; Alborali, Giovanni; Formenti, Nicoletta; Guarneri, Flavia; Fontani, Nadia; Nani, Giulia; Palumbo, Franca; Borlone, Gianluca; Guercio, Marco; Mariuz, Marika; Trani, Gabriella; Pariani, Anna; Ancona, Carla; Barca, Alessandra; Serio, Flavia; Antonella Giorgi, Doriana; Ferrante, Irene; Monfrinotti, Monica; Riosa, Silvia; Capparuccini, Valeria; Teresa Scicluna, Maria; Cersini, Antonella; Arizzi, Mariaconcetta; Cecchini, Giancarlo; Ottaviano, Claudio; Nicosia, Elena; Fontani, Nadia; Nani, Giulia; Palumbo, Franca; Borlone, Gianluca; Guercio, Marco; Grasselli, Elena; Allaria, Giorgia; Dondero, Lorenzo; Rispo, Francesca; Izzotti, Alberto; Maria Bertolotto, Rosa; Nicosia, Elena; Rosatto, Stefano; Bellisomi, Marta; Tomesani, Irene; Tiso, Micaela; Ammoni, Emanuela; Cereda, Danilo; Nadia Losio, Marina; Bertasi, Barbara; Oliva, Desdemona; Giovanna Guiso, Maria; Ferrari, Fabio; Mundo, Maria; Martines, Antonino; Castiglioni, Sara; Schiarea, Silvia; Salmoiraghi, Giulia; Antonelli, Manuela; Azzellino, Arianna; Malpei, Francesca; Turolla, Andrea; Binda, Sandro; Laura, Pellegrinelli; Primache, Valeria; Cocuzza, Clementina; Franzetti, Andrea; Musumeci, Rosario; Martinelli, Marianna; Bertanza, Giorgio; Luisa Callegari, Maria; Bolognini, Luigi; Filippetti, Fabio; Paniccia', Marta; Ciuti, Francesca; Briscolini, Sara; Magi, Silvia; Colitti, Michele; Montanaro, Carmen; Aprea, Giuseppe; Scattolini, Silvia; D’Angelantonio, Daniela; Migliorati, Giacomo; Grazia Cerroni, Maria; Griglio, Bartolomeo; Berruti, Renza; Cravero, Mauro; Costa, Angela; Bianchi, Manila; Decastelli, Lucia; Romano, Angelo; Tramuta, Clara; Carraro, Elisabetta; Pignata, Cristina; Bonetta, Silvia; Richiardi, Lisa; Di Vittorio, Giuseppe; Mongelli, Onofrio; De Giglio, Osvalda; Apollonio, Francesca; Triggiano, Francesco; Teresa Montagna, Maria; Ungaro, Nicola; Palermo, Mario; Massimo Maida, Carmelo; Mazzucco, Walter; De Grazia, Simona; Giammanco, Giovanni; Purpari, Giuseppa; Ferrante, Margherita; Agodi, Antonella; Barchitta, Martina; Cala', Piergiuseppe; Carducci, Annalaura; Verani, Marco; Federigi, Ileana; Lauretani, Giulia; Muzio, Sara; Ramazzotti, Matteo; Antonelli, Alberto; Santoro, Giovanni; Federici, Ermanno; Petricciuolo, Maya; Barigelli, Sofia; Ruffier, Mauro; Borney, Francesca; Grange, Eric; Damasco, Florida; Russo, Francesca; Pitter, Gisella; Groppi, Vanessa; Rigoli, Franco; Zampini, Marco; Baldovin, Tatjana; Amoruso, Irene; Zago, Lorella; Stenico, Alberta; Demetz, Lea; Pizzo, Francesco; Schiavuzzi, Alessandra; Mengon, Elena; Cadonna, Maria; Postinghel, Mattia; Cutrupi, Francesca; Foladori, Paola; Manara, Serena; Suffredini, E.; La Rosa, G.. - (2023), pp. 228-228. (Intervento presentato al convegno 7th National Congress of the Italian Society for Virology - One virology one health tenutosi a Brescia nel 25-27 giugno 2024).
THE USE OF NANOPORE SEQUENCING TECHNOLOGY PROVIDES NEW INSIGHTS INTO THE WIDE RANGE OF HUMAN ADENOVIRUSES PRESENT IN ASTEWATER
Francesca Cutrupi;Paola Foladori;Serena Manara;
2023-01-01
Abstract
Aim of the Study: Wastewater-based epidemiology is a powerful tool for monitoring the circu lation of pathogens in the population, including both asymptomatic and symptomatic infections. Human adenovirus (HAdV) is an enteric virus, responsible for various clinical manifestations, including upper respiratory tract infections, conjunctivitis, gastroenteritis, bronchitis and pneumonia, urinary tract infections, and skin infections. Occasionally, adenoviruses can cause more severe illnesses, including meningitis, encephalitis, and myocarditis. Adenovirus can be spread through water, making it a potential waterborne pathogen. Human HAdVs are classified into seven species (A-G) and more than 110 types. Species A and B are responsible for common respiratory infections, while species F includes serotypes 40 and 41, which are recognised as some of the most common causes of gastroenteritis. Methods: From 9 to 13 of January 2023, a total of 170 sewage samples were collected in 165 wastewater treatment plants (WTPs) in 18 regions and 2 autonomous provinces of Italy. HAdV was detected by amplification of a fragment of the HAdV hexon gene (714 to 855 bp long, depending on the HAdV type). Amplicons were sequenced by both Sanger (individual samples) and next-generation sequencing (in a pool of all PCR amplicons), using the Oxford Nanopore Technology (MinION platform). The long reads obtained by NGS were compared with 110 reference prototypes belonging to the Species HadV-A (n°=4), HadV-B (n°=17), HAdV-C (n°=7), HAdV-D (n°=78), HAdV-E (n°=1), HAdV-F (n°=2), and HAdV-G (n.=1). Results and Conclusions: The results showed a wide diversity of HAdVs in wastewater, indicating genetic diversity within the population at a national level. The nested PCR results showed the presence of HAdVs in 118 out of 170 samples (69.4%). Seventy-six samples were successfully sequenced and characterised by conventional Sanger sequencing as belonging to 5 HAdV Species and 7 types: Species A (HAdV-12, 2 samples), Species B (HAdV-3, 33 samples), Species C (HAdV-1, 2 and 89, 6 samples), Species D (HAdV-45, 1 sample), and Species F (HAdV 41, 34 samples). NGS analysis confirmed the results obtained by Sanger sequencing but was able to detect additional less represented types: HAdV-B21, HAdV-C5, HAdV-C89, HAdV-D23, HAdV-D46, HAdV-D49, HAdV-D83, and HAdV-F40. Other sequences could not be definitively assigned because they shared the same % nt identity with more than one HAdV prototype: HAdV-B7 and B66; HAdV- C1 and C104; HAdV- D45 and D73; HAdV- D48 and D81. Our findings highlight the diversity of HAdV species and types present in raw sewage, with the potential to impact surface waters and pose a significant threat to human health. By using next-generation sequencing, we were able to identify a broad range of “less prevalent” types in addition to those identified by Sanger sequencing. The results show that although the exon region is recognised as being able to discriminate among different types of human adenovirus, it is not actually able to identify certain types that have a high degree of nucleotide identity, particularly those belonging to the D Species.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione
