Increased detection of G3P[9] and G6P[9] rotavirus strains in hospitalized children with acute diarrhea in Bulgaria Zornitsa Mladenova a,b,⇑,1 , Sameena Nawaz b,1 , Balasubramanian Ganesh c , Miren Iturriza-Gomara d a National Center for Infectious and Parasitic Diseases, Sofia, Bulgaria b Public Health of England, London, United Kingdom c National Institute of Cholera and Enteric Diseases, Kolkata, India d Institute of Infection and Global Health, University of Liverpool, Liverpool, United Kingdom article info Article history: Received 11 June 2014 Received in revised form 23 September 2014 Accepted 12 November 2014 Available online 20 November 2014 Keywords: Rotavirus Human strain Reassortant Zoonotic transmission Rotavirus vaccine abstract Rotavirus severe disease in children is now vaccine-preventable and the roll-out of vaccination programs globally is expected to make a significant impact in the reduction of morbidity and mortality in children <5 years of age. Rotavirus is also a pathogen of other mammals and birds, and its segmented RNA genome can lead to the emergence of new or unusual strains in human population via interspecies transmission and reassortment events. Despite the efficacy and impact of rotavirus vaccine in preventing severe diar- rhea, the correlates of protection remain largely unknown. Therefore, rotavirus strain surveillance before, during and after the introduction of immunization programs remains a crucial for monitoring rotavirus vaccine efficacy and impact. In this context, molecular characterization of 1323 Bulgarian rotavirus strains collected between June 2010 and May 2013 was performed. A total of 17 strains of interest were analyzed by partial sequence analysis. Twelve strains were characterized as G3P[9] and G6P[9] of poten- tial animal origin. Phylogenetic analysis and comparisons with the same specificity strains detected spo- radically between 2006 and 2010 revealed the constant circulation of these unusual human strains in Bulgaria, although in low prevalence, and their increased potential for person-to-person spread. Ó 2014 Elsevier B.V. All rights reserved. 1. Introduction Rotaviruses (RVs) are the leading cause of acute viral gastroen- teritis in children under 5 years of age. The total number of RV diarrhea cases has considerably declined since 2001, but RVs still account for the death of 420–494 thousand children 65 years old, as half of the cases occur in 5 developing countries (Tate et al., 2012; Parashar et al., 2009). In 2006 two RV vaccines were approved, and since then they have been included in many national immunization calendars around the globe. The implemen- tation of RV vaccination program in many countries has revealed the dramatic decrease of the RV hospitalizations and economic benefits connected with this (Rha et al., 2014; Linhares and Justino, 2014; Nakagomi et al., 2013a; De Oliveira et al., 2013; Raes et al., 2011). However, in countries without RV vaccine included into the national immunization programs, RVs are still responsible for high morbidity and mortality (Yen et al., 2011; Mladenova et al., 2011). RVs are classified as a genus of the family Reoviridae. The gen- ome of RVs, enclosed in a triple-layered protein capsid, consists of 11 double-stranded RNA segments which code for a total of 12 viral proteins, six structural (VP1–VP4, VP6, VP7) and six nonstruc- tural (NSP1–NSP6) (Estes and Kapikian, 2007). RVs are classified into groups A to G based on epitopes of the VP6 intermediate cap- side protein, as RVs group A (RVA) have the major human and vet- erinary health impact. The two outer capsid proteins of RVA, VP7 (glycosylated, or G-type) and VP4 (protease-sensitive, or P-type), elicit neutralizing antibody responses, and may have a role in homotypic protection. For this reason G and P-types are the basis for the traditional binary nomenclature of RVA (Hoshino et al., 1985). To date, at least 27 G and 37 P genotypes have been defined in human, mammals and birds according to molecular-genetic diversity of the VP7 and VP4, respectively (Matthijnssens et al., 2011a). At least, 12 G types (G1–G6, G8–G12 and G20) and 15 P types (P[1]–P[11], P[14], P[19], P[25], and P[28]) have been detected in RVA-infected humans. Recently, a novel genotyping classification scheme based on all 11 genome segments has been http://dx.doi.org/10.1016/j.meegid.2014.11.011 1567-1348/Ó 2014 Elsevier B.V. All rights reserved. ⇑ Corresponding author at (formerly): National Reference Laboratory of Entero- viruses, Department of Virology, National Center of Infectious and Parasitic Diseases, 44A, Stoletov Blvd., Sofia 1233, Bulgaria. Tel.: +359 2 931 23 22x247; fax: +359 2 943 30 75. E-mail address: zornitsavmbg@yahoo.com (Z. Mladenova). 1 The ZM and SN were equally contributed to the investigation reported in the present study and to the preparation of the draft. Infection, Genetics and Evolution 29 (2015) 118–126 Contents lists available at ScienceDirect Infection, Genetics and Evolution journal homepage: www.elsevier.com/locate/meegid