Skip directly to site content Skip directly to page options Skip directly to A-Z link Skip directly to A-Z link Skip directly to A-Z link
Volume 21, Number 11—November 2015
Dispatch

Rotavirus P[8] Infections in Persons with Secretor and Nonsecretor Phenotypes, Tunisia

Tables
Article Metrics
25
citations of this article
EID Journal Metrics on Scopus
Siwar Ayouni, Khira Sdiri-Loulizi, Alexis de Rougemont, Marie Estienney, Katia Ambert-Balay, Serge Aho, Sabeur Hamami, Mahjoub Aouni, Mohamed Neji-Guediche, Pierre Pothier, and Gaël BelliotComments to Author 
Author affiliations: National Reference Center for Enteric Viruses, Dijon, France (S. Ayouni, A. de Rougemont, M. Estienney, K. Ambert-Balay, P. Pothier, G. Belliot); Monastir University, Monastir, Tunisia (S. Ayouni, K. Sdiri-Loulizi, M. Aouni); Public Hospital of Dijon, Dijon (S. Aho); Hospital Fattouma Bourguiba, Monastir (S. Hamami, M. Neji-Guediche)

Cite This Article

Abstract

To determine whether rotavirus infections are linked to secretor status, we studied samples from children in Tunisia with gastroenteritis. We phenotyped saliva for human blood group antigens and tested feces for rotavirus. Rotavirus was detected in 32/114 patients. Secretor genotyping showed that P[8] rotavirus infected secretors and nonsecretors, and infection correlated with presence of Lewis antigen.

Each year, millions of persons worldwide suffer from acute gastroenteritis. Group A rotavirus is the leading cause of acute gastroenteritis in children <5 years of age. The disease causes ≈453,000 deaths annually, mostly in developing countries (1); however, the number of cases has declined in industrialized countries where vaccines have been recommended (2).

Recent findings showed that human blood group antigens (HBGAs) might be involved in rotavirus attachment to intestinal cells (3,4,5,6). Expression of the HBGAs (A, B, H, and Lewis antigens) in saliva and on the surface of intestinal cells is driven by the FUT2 (A, B, and H antigens [secretor]) and FUT3 (Lewis antigens) genes, which express type 2 and type 3 fucosyltransferases, respectively. Approximately 20% of the white population is homozygous for a recessive point mutation of the FUT2 gene, which leads to the absence of A, B, and H antigen expression, also called the nonsecretor phenotype. There is also a Lewis-negative phenotype resulting from various mutations of the FUT3 gene (7).

The entry of rotavirus into cells involves several factors. Human and porcine rotaviruses could specifically interact with H antigen type 1, Lewis b antigen, or Lewis a antigen through their viral protein (VP) 8 and VP5 during the attachment phase (3,4,8). Of note, the HBGA binding profile is P genotype–dependent (4), and rotavirus infection correlates with the secretor and partial secretor phenotype (i.e., with active FUT2 gene status) (5,6,9). However, in some studies, no association has been observed between HBGAs from blood cells (10), including Lewis antigens (11), and rotavirus infection. A recent epidemiologic survey of children in the region of Monastir, Tunisia, gave us the opportunity to determine whether rotavirus infections in children could be linked to secretor status and HBGAs.

The Study

During November 2011–February 2012, feces and saliva samples were collected from 114 children <6 years of age who were seen for acute gastroenteritis at the Fattouma Bourguiba children’s hospital (Monastir). For 98 of these patients, blood samples were also collected at symptom onset for FUT2 genotyping by sequencing for the A385T and G428A nonsense mutations from total blood DNA (9). The study was approved by the Ethics Committee of the Fattouma Bourguiba University Hospital in Monastir, and informed consent was obtained from the parents of the 114 study participants.

The feces were first screened for the presence of group A rotavirus antigen by using the Premier Rotaclone detection kit (Meridian Bioscience, Inc., Paris, France). The remainder of the suspension was used for the extraction of nucleic acids by using a Nuclisens easyMAG system (bioMérieux, Marcy l’Étoile, France) according to the manufacturer’s instructions. RNA was eluted in a final volume of 110 μL and used for the molecular detection and typing of rotavirus. Norovirus PCR detection is described elsewhere (12).

Samples positive for rotavirus by ELISA were further confirmed and genotyped by PCR as described previously (Technical Appendix Table). Of the 114 patients, 32 had confirmed rotavirus infections by ELISA and PCR. Of the 32 confirmed cases, 24 (75%) occurred during the cold season, and 26 (80%) occurred in children <14 months of age; the mean age for infected persons was 8.1 months. We used ELISAs to screen the saliva of rotavirus-positive patients for A and B antigens (anti-A and anti-B mouse IgG from DIAGAST, Loos, France), H antigen (anti-H specific IgM from Thermo Fisher Scientific, Villebon sur Yvette, France), and Lewis antigens (anti-Lewis a [clone 7LE] and anti-Lewis b [clone 2–25LE] hybridoma supernatants; gift from Jacques Bara, INSERM U673). Among the secretor phenotype–positive rotavirus patients, no blood group antigen nor P or G genotypes (in feces specimens) were significantly overrepresented. For comparison, we assessed the distribution of ABO blood groups and Lewis antigens among patients with norovirus and rotavirus; no statistical difference was found (Table 1). Rotavirus infection was observed only in Lewis antigen–positive patients (p = 0.017, exact logistic regression); however, the prevalence of the Lewis antigen–negative phenotype in the population was low.

Among the 32 rotavirus isolates, 30 were genotype P[8] and 2 were genotype P[4] (Table 2). G9, G3, and G1 were the most common genotypes and were detected in 13 (40%), 8 (25%), and 7 (21%) of the cases, respectively. The G genotype could not be determined for 1 P[8] genotype isolate. Genotypes G1, G3, G4, and G9 were all associated with the P[8] genotype, and genotype G2 was associated with the P[4] genotype. Rotavirus G9P[8] strains were predominant (n = 12), followed by G3P[8] (n = 8) and G1P[8] (n = 7) strains.

For 3 G9P[8] and 1 G3P[8] rotavirus-positive patients, saliva samples were negative for Lewis b antigen (mean absorbance at 450 nm was 0.25) and positive for the presence of Lewis a antigen (mean absorbance at 450 nm was 3.67), suggesting that the patients were Lewis-positive and nonsecretors. A total of 98 blood samples were genotyped by sequencing of the FUT2 gene. Homozygous secretor, heterozygous secretor, and nonsecretor genotypes represented 23.47%, 54.08%, and 22.45% of the cohort, respectively. All nonsecretors (n = 22) harbored the G428A mutation. The A385T mutation was absent. Blood samples were available for 28 of the 32 rotavirus-positive patients. Of these 28 patients, 24 were homozygous and heterozygous secretors and 4 were nonsecretors. Because rotaviruses were P- and G-typed by PCR using VP4- and VP7-specific primers, we further confirmed the presence of rotavirus in samples from nonsecretor patients. For 3 of the samples, we used a TaqMan-based quantitative reverse transcription PCR with VP2-specific primers to detect rotavirus (13), and for 2 rotavirus isolates (GenBank accession nos. KP862856 and KP862857) for which feces samples were still available, we confirmed the P[8] genotype by sequencing.

Conclusions

Our findings show that rotaviruses can infect secretor and nonsecretor Lewis antigen–positive persons, which suggests that rotavirus infection is not associated with the secretor phenotype or HBGA type. However, it should be noted that one limitation of our study was the small size of our cohort. A larger number of cases might provide new insights about the affinity of rotavirus toward certain types of HBGAs; a larger study with a more robust statistical analysis might confirm that rotavirus infections only occur in Lewis antigen–positive persons. In addition, we detected genotype P[8] rotavirus infection in both secretor and nonsecretor patients; this finding was not observed in previous studies (5,6,9). P[8] infection of nonsecretors might be associated with preexisting health conditions, and healthy nonsecretors might never be infected by P[8] rotavirus.

We and others (3,4,6) have characterized the secretor and Lewis phenotypes related to infection by rotaviruses. The interaction between rotavirus particles and HBGAs might constitute the first step in the attachment to the cell before internalization of the virus particle, after binding with integrins (4,5). However, other types of ligand, such as non-HBGA ligands and bacteria from intestinal flora, might also play a role during the infection process, as recently shown for noroviruses (14,15).

In conclusion, our data and that of others show that rotavirus infection might be correlated with genetic factors, such as HBGAs. Further studies will be required to determine the exact role of HBGA ligands and other ligands in rotavirus infection.

Ms. Ayouni is currently a PhD student at the National Reference Center for Enteric Viruses (Dijon, France). Her research interests focus on the relation between human blood group antigens and enteric viruses, mainly rotavirus and norovirus.

Top

Acknowledgments

We thank Jacques Bara for kindly providing us with Lewis-specific monoclonal antibodies; Philippe Daval, Céline Fremy, and the National Reference Center for Enteric Viruses for their technical support; and Philip Bastable for editorial assistance.

This study was supported by the National Reference Center for Enteric Viruses and the Public Hospital of Dijon, France. A fellowship from Campus France was awarded to S.A. (PHC-UTIQUE program).

Top

References

  1. Tate  JE, Burton  AH, Boschi-Pinto  C, Steele  AD, Duque  J, Parashar  UD, 2008 estimate of worldwide rotavirus-associated mortality in children younger than 5 years before the introduction of universal rotavirus vaccination programmes: a systematic review and meta-analysis. Lancet Infect Dis. 2012;12:13641. DOIPubMed
  2. Lopman  BA, Payne  DC, Tate  JE, Patel  MM, Cortese  MM, Parashar  UD. Post-licensure experience with rotavirus vaccination in high and middle income countries; 2006 to 2011. Curr Opin Virol. 2012;2:434–42.
  3. Hu  L, Crawford  SE, Czako  R, Cortes-Penfield  NW, Smith  DF, Le Pendu  J, Cell attachment protein VP8* of a human rotavirus specifically interacts with A-type histo-blood group antigen. Nature. 2012;485:2569. DOIPubMed
  4. Huang  P, Xia  M, Tan  M, Zhong  W, Wei  C, Wang  L, Spike protein VP8* of human rotavirus recognizes histo-blood group antigens in a type-specific manner. J Virol. 2012;86:483343. DOIPubMed
  5. Nordgren  J, Sharma  S, Bucardo  F, Nasir  W, Gunaydin  G, Ouermi  D, Both Lewis and secretor status mediate susceptibility to rotavirus infections in a rotavirus genotype–dependent manner. Clin Infect Dis. 2014;59:156773. DOIPubMed
  6. Van Trang  N, Vu  HT, Le  NT, Huang  P, Jiang  X, Anh  DD. Association between norovirus and rotavirus infection and histo-blood group antigen types in Vietnamese children. J Clin Microbiol. 2014;52:136674. DOIPubMed
  7. Grahn  A, Elmgren  A, Aberg  L, Svensson  L, Jansson  PA, Lonnroth  P, Determination of Lewis FUT3 gene mutations by PCR using sequence-specific primers enables efficient genotyping of clinical samples. Hum Mutat. 2001;18:3589. DOIPubMed
  8. Blanchard  H, Yu  X, Coulson  BS, von Itzstein  M. Insight into host cell carbohydrate-recognition by human and porcine rotavirus from crystal structures of the virion spike associated carbohydrate-binding domain (VP8*). J Mol Biol. 2007;367:121526. DOIPubMed
  9. Imbert-Marcille  BM, Barbe  L, Dupe  M, Le Moullac-Vaidye  B, Besse  B, Peltier  C, A FUT2 gene common polymorphism determines resistance to rotavirus A of the P[8] genotype. J Infect Dis. 2014;209:122730. DOIPubMed
  10. Yazgan  H, Keles  E, Gebesce  A, Demirdoven  M, Yazgan  Z. Blood groups and rotavirus gastroenteritis. Pediatr Infect Dis J. 2013;32:7056. DOIPubMed
  11. Ahmed  T, Lundgren  A, Arifuzzaman  M, Qadri  F, Teneberg  S, Svennerholm  AM. Children with the Le(a+b-) blood group have increased susceptibility to diarrhea caused by enterotoxigenic Escherichia coli expressing colonization factor I group fimbriae. Infect Immun. 2009;77:205964. DOIPubMed
  12. Kamel  AH, Ali  MA, El-Nady  HG, de Rougemont  A, Pothier  P, Belliot  G. Predominance and circulation of enteric viruses in the region of greater Cairo, Egypt. J Clin Microbiol. 2009;47:103745. DOIPubMed
  13. Gutiérrez-Aguirre  I, Steyer  A, Boben  J, Gruden  K, Poljsak-Prijatelj  M, Ravnikar  M. Sensitive detection of multiple rotavirus genotypes with a single reverse transcription–real-time quantitative PCR assay. J Clin Microbiol. 2008;46:254754. DOIPubMed
  14. Jones  MK, Watanabe  M, Zhu  S, Graves  CL, Keyes  LR, Grau  KR, Enteric bacteria promote human and mouse norovirus infection of B cells. Science. 2014;346:7559. DOIPubMed
  15. Ruvoën-Clouet  N, Magalhaes  A, Marcos-Silva  L, Breiman  A, Figueiredo  C, David  L, Increase in genogroup II.4 norovirus host spectrum by CagA-positive Helicobacter pylori infection. J Infect Dis. 2014;210:18391. DOIPubMed

Top

Tables

Top

Cite This Article

DOI: 10.3201/eid2111.141901

Table of Contents – Volume 21, Number 11—November 2015

Comments

Please use the form below to submit correspondence to the authors or contact them at the following address:

Gaël Belliot, Centre National de Référence des Virus Entériques, Laboratoire de Virologie/Plateau Technique de Biologie/CHU Dijon, 2 rue Angélique Ducoudray, BP37013, 21070 Dijon CEDEX, France

Send To

10000 character(s) remaining.

Top

Page created: October 19, 2015
Page updated: October 19, 2015
Page reviewed: October 19, 2015
The conclusions, findings, and opinions expressed by authors contributing to this journal do not necessarily reflect the official position of the U.S. Department of Health and Human Services, the Public Health Service, the Centers for Disease Control and Prevention, or the authors' affiliated institutions. Use of trade names is for identification only and does not imply endorsement by any of the groups named above.
file_external