Cosavirus Infection in Persons with and without Gastroenteritis, Brazil

Andreas Stöcker¹, Breno Frederico de Carvalho Dominguez Souza¹, Tereza Cristina Medrado Ribeiro¹, Eduardo Martins Netto, Luciana Oliveira Araujo, Jefferson Ivan Corrêa, Patrícia Silva Almeida, Angela Peixoto de Mattos, Hugo da Costa Ribeiro, Diana Brasil Pedral-Sampaio, Christian Drosten, and Jan Felix Drexler

Author affiliations: University of Bonn Medical Centre, Bonn, Germany (A. Stöcker, C. Drosten, J.F. Drexler); Federal University of Bahia, Salvador, Brazil (A. Stocker, B.F.C.D. Souza, T.C.M. Ribeiro, E.M. Netto, L.O. Araujo, J.I. Corrêa, P.S. Almeida, A.P. de Mattos, H.C. Ribeiro Jr, D.B. Pedral Sampaio)

Abstract

To determine possible cosavirus association with clinical disease, we used real-time reverse transcription PCR to test children and HIV-positive adults in Brazil with and without gastroenteritis. Thirteen (3.6%) of 359 children with gastroenteritis tested positive, as did 69 (33.8%) of 204 controls. Low prevalence, frequent viral co-infections, and low fecal cosavirus RNA concentrations argue against human pathogenicity.

The family Picornaviridae comprises 12 genera and includes several leading pathogens affecting human and animal health, e.g., foot-and-mouth-disease virus and polioviruses. With the advent of metagenomics, 3 novel human picornaviruses have been described since 2008: klassevirus, salivirus, and cosavirus (1–3).
Besides being detected in raw sewage (4), cosaviruses have been detected in human fecal specimens and were tentatively associated with gastroenteritis in 2 patients from Australia and Scotland (3,5). Cosavirus pathogenicity in humans has remained unknown, however, because detection rates in patients and healthy controls were similar in the only available cohort studies of patients with acute flaccid paralysis in Southeast Asia (3) and with gastroenteritis in China (6). We investigated cosavirus prevalence in 464 children and HIV-infected adults with gastroenteritis and 253 controls without gastroenteritis in Brazil.

The Study

Fecal specimens from 6 clinical cohorts in Salvador, northeastern Brazil, were analyzed (Table 1). Adult cohorts comprised HIV-infected patients with gastroenteritis (105 persons) and without (49 persons) gastroenteritis. Child cohorts included 359 children with gastroenteritis and 204 healthy children from child-care centers. Nasal swab specimens were obtained from controls attending child-care centers and from children with gastroenteritis and concomitant respiratory symptoms. All specimens were stored at −30° to −80°C until further processing.
Viral RNA was purified from ≈200 mg fecal specimen and 140 µL nasal swab specimen suspended in phosphate-buffered saline by using the Viral RNA Mini kit (QIAGEN, São Paulo, Brazil) as described (7). A nested reverse transcription PCR (3) detected cosavirus. After nucleotide sequencing of all PCR-positive specimens, we developed a specific real-time RT-PCR for quantifying viruses from Brazil (Table 2). Assay optimization and quantification relied on photometrically quantified cRNA in vitro transcripts, as described (8). After assay optimization, sensitivity was 6.8 copies per reaction.
In the adult cohorts, 1 of 105 HIV-infected persons with gastroenteritis had positive results for cosavirus. In the control group of 49 HIV-infected adults without gastroenteritis, none had positive cosavirus results (χ² 0.5; p = 0.49).
Considerably higher prevalence was detected among child cohorts. Of children with gastroenteritis, 13 (3.6%) of 359 patients were cosavirus positive. The proportion of cosavirus-positive controls without gastroenteritis, sampled in 2008, was significantly higher: 65 (49.2%) of 132 controls were cosavirus positive (χ² 149.1; p<0.0001). On resampling in 2011, 4 (6.5%) of 62 controls were cosavirus positive. Although the difference was not statistically significant (χ² 1.1; p = 0.3), this result was almost double that for patients. In another child-care center, none of 10 healthy children were cosavirus positive.