International Journal of Infectious Diseases 26 (2014) e96–e97
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Infection with Saint Louis encephalitis virus in the city of Ribeirao Preto, Brazil: report of one case Felipe Gonc¸alves Motta Maia, Juliana Helena Cha´vez, William Marciel de Souza, Marilia Farignoli Romeiro, Luiza Antunes de Castro-Jorge, Benedito Antoˆnio Lopes da Fonseca, Luiz Tadeu Moraes Figueiredo * Virology Research Center, School of Medicine of Ribeirao Preto, University of Sao Paulo, Av. Bandeirantes, 3900, 14049-900, Ribeirao Preto, Sa˜o Paulo, Brazil
A R T I C L E I N F O
Article history: Received 18 March 2014 Received in revised form 19 May 2014 Accepted 20 May 2014 Corresponding Editor: Eskild Petersen, Aarhus, Denmark Keywords: Saint Louis encephalitis virus Arbovirus Flavivirus Brazil
S U M M A R Y
Saint Louis encephalitis virus (SLEV) is a mosquito-borne ﬂavivirus from the Americas. In this report we describe aspects of the laboratory diagnosis of a patient with an acute febrile illness induced by SLEV that was initially diagnosed as dengue by positive IgM-ELISA. Infection with this virus is probably not rare in Brazil, but cases remain undiagnosed. It is necessary to improve the surveillance system, including laboratories, for the diagnosis of SLEV in Brazil. ß 2014 The Authors. Published by Elsevier Ltd on behalf of International Society for Infectious Diseases. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/bync-nd/3.0/).
1. Introduction Saint Louis encephalitis virus (SLEV) is an important mosquitoborne ﬂavivirus in the Americas. When infecting man, SLEV can produce disease ranging from a mild febrile illness to severe encephalitis. SLEV was ﬁrst identiﬁed in Saint Louis, Missouri, USA, in 1933.1 Several outbreaks have recently occurred in the Americas, and the fatality rate has ranged from 5% to 20%. There is no speciﬁc treatment or vaccine available for SLEV.2,3 The seroprevalence of SLEV in the Brazilian population ranges from 3% to 43%.4,5 SLEV has also been reported to infect wild birds, mosquitoes, and marsupials, and was recently isolated from the brain of a horse with encephalitis in Minas Gerais State.6,7 Human disease induced by SLEV has rarely been reported in Brazil. The ﬁrst case with a serological diagnosis occurred in Belem, Para State, in 1978.8 Another case, diagnosed by RT-PCR, was reported in Sao Paulo State (SP) in 2005.9 In 2006, an outbreak of acute febrile disease with meningoencephalitis and/or hemorrhagic manifestations involving at least 14 cases was reported in Sao Jose do Rio Preto, SP. Interestingly, this outbreak occurred
* Corresponding author. Tel.: +55 16 3602 4580; fax: +55 16 3602 3376. E-mail addresses: [email protected]
(W.M. de Souza), ltmﬁ[email protected]
concomitantly with a large dengue outbreak.10,11 In this outbreak, one case of co-infection with SLEV–dengue type 3 virus was observed.12 Herein, we report SLEV infection in one patient living in Ribeirao Preto, SP, Brazil. 2. Case report In April 2008, a retired 74-year-old man sought medical care after 24 h of fever, headache, diarrhea, muscle pain, arthralgia, and lipothymia. The patient lived in the urban area of Ribeirao Preto, had not been out of town in the last 15 days, and had been vaccinated for yellow fever in 2003. The patient was clinically diagnosed for dengue infection and a blood sample was collected for laboratory diagnosis. The patient probably recovered without sequelae. The patient’s serum was ﬁrst tested by ELISA (Panbio, Brisbane, Australia), showing IgM and IgG antibodies to dengue virus. The serum was then tested for the presence of NS1 dengue antigen by capture ELISA Platelia (Bio-Rad Laboratories, France), giving a negative result. The dengue virus genome was not ampliﬁed in the patient’s serum by conventional RT-PCR.13 Further investigations were done for other ﬂaviviruses by ELISA and RT-PCR. Anti-SLEV antibodies were detected by an in-house ELISA using recombinant domain III (rDIII) of the envelope protein of the virus as antigen, as reported previously.14 Corroborating these results, neutralizing
http://dx.doi.org/10.1016/j.ijid.2014.05.018 1201-9712/ß 2014 The Authors. Published by Elsevier Ltd on behalf of International Society for Infectious Diseases. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).
F.G.M. Maia et al. / International Journal of Infectious Diseases 26 (2014) e96–e97 Table 1 Diagnosis of Saint Louis encephalitis virus in a patient with suspected dengue fever Method
Dengue IgM capture ELISA Dengue IgG indirect ELISA Dengue NS1 antigen capture ELISA Dengue RT-PCR SLEV in-house IgG ELISA Classic neutralization test Flavivirus RT-PCR
Panbio Panbio Platelia-Bio-Rad
Reactive Reactive Non-reactive
Lanciotti et al., 2002 Chaves et al., 2013 Shope and Sather, 1979 Bronzoni et al., 2005
Negative Reactive Neutralizing Positive
remain undiagnosed, it is necessary to improve the surveillance system, including laboratories, for the diagnosis of SLEV in Brazil. SLEV circulates in the state of Sao Paulo, the most populated region of Brazil. This virus, probably brought in by birds, is transmitted to man in urban areas by Culex mosquitoes. However, the epidemiology of SLEV in urban areas is completely unknown. Further studies are necessary to better understand the epidemiology of SLEV in Brazil. Acknowledgements
SLEV, Saint Louis encephalitis virus.
anti-SLEV antibodies (logarithm neutralization index of 2.14) were obtained in the serum by a classic neutralization test in baby mice (Table 1).15 In addition, RNA was extracted from the patient’s serum using the QIAamp viral RNA mini kit (Qiagen, Hilden, Germany) and was subjected to an RT-PCR using non-structural protein 5 gene (NS5) ﬂavivirus generic primers. An amplicon of approximately 800 base pairs (bp) was obtained, as expected for a ﬂavivirus.16 The amplicon was puriﬁed with the QIAquick gel extraction kit (Qiagen, Hilden, Germany) and sequenced using the same ﬂavivirus generic primers and the Big Dye Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems, Foster City, CA, USA) in a sequencer Genetic Analyzer 3130 (Applied Biosystems, Foster City, CA, USA). An 804-bp sequence was obtained and its SLEV origin was conﬁrmed by BLAST (Basic Local Alignment Search Tool). This nucleotide sequence was deposited in GenBank (accession number KJ528400) and also compared to other NS5 SLEV sequences. The sequence showed a very high identity (100%) with that of the SLEV strain BeAn 246262 (GenBank accession number EU088424.1), identiﬁed in a Didelphis marsupialis black-eared opossum in Belem, Para State, in 1973. A high identity (100%) of the sequence with 800 nucleotides of the NS5 of SLEV identiﬁed in Sao Jose do Rio Preto (EF219166.1; DQ836337.1; DQ836336.1) and high homology with other Brazilian strains of SLEV was also observed.
3. Discussion More than 10 pathogenic ﬂaviviruses have been described in Brazil, with dengue viruses being those most commonly diagnosed.6 The acute benign febrile illness induced by SLEV, as observed in our patient, could easily be confused with dengue in a country where all four dengue virus serotypes circulate endemically. Public health authorities should be concerned about crossreactions of IgG and IgM antibodies to SLEV and other Brazilian ﬂaviviruses in dengue serologic tests. This clinical presentation of SLEV infection has also been reported by other authors.10,12,17 Our patient did not develop meningoencephalitis, which is often reported in persons over 60 years of age.10,12,17 It is possible that many SLEV cases are not recognized mainly due to a cross-reaction among ﬂaviviruses in serologic tests, including the dengue IgM ELISA, as observed in the present patient. Thus, considering that SLEV infection is probably not rare in Brazil and that the cases
This study was supported by a grant from FAPESP – Fundac¸a˜o de Amparo a` Pesquisa do Estado de Sa˜o Paulo (No. 08/50617-6), Fellowship No. 06/01179-0, 12/02836-6, and 12/24150-9, and CNPq – Conselho Nacional de Pesquisa No. 301677/2013-1. Conﬂict of interest: The author declares no competing interest in publishing this case report. References 1. Reisen WK. Epidemiology of St. Louis encephalitis virus. Adv Virus Res 2003;61:139–83. 2. Tsai TF, Mitchell CJ. St Loius encephalitis. In: Monath TP, editor. The Arboviruses: Epidemiology and Ecology. Boca Raton, FL: CRC Press; 1988. p. 431–58. 3. Kopp A, Gillespie TR, Hobelsberger D, Estrada A, Harper JM, Miller RA, et al. Provenance and geographic spread of St. Louis encephalitis virus. MBio 2013;4:e00322–413. 4. Pauvolid-Correa A, Tavares FN, Costa EV, Burlandy FM, Murta M, Pellegrin AO, et al. Serologic evidence of the recent circulation of Saint Louis encephalitis virus and high prevalence of equine encephalitis viruses in horses in the Nhecolandia sub-region in South Pantanal, Central-West Brazil. Mem Inst Oswaldo Cruz 2010;105:829–33. 5. Pauvolid-Correa A, Campos Z, Juliano R, Velez J, Nogueira RM, Komar N. Serological evidence of widespread circulation of West Nile virus and other ﬂaviviruses in equines of the Pantanal, Brazil. PLoS Negl Trop Dis 2014;8:e2706. 6. Figueiredo LT. The Brazilian ﬂaviviruses. Microbes Infect 2000;2:1643–9. 7. Rosa R, Costa EA, Marques RE, Oliveira TS, Furtini R, Bomﬁm MR, et al. Isolation of Saint Louis encephalitis virus from a horse with neurological disease in Brazil. PLoS Negl Trop Dis 2013;7:e2537. 8. Pinheiro FP, LeDuc JW, Travassos da Rosa AP, Leite OF. Isolation of St. Louis encephalitis virus from a patient in Belem, Brazil. Am J Trop Med Hyg 1981;30:145–8. 9. Rocco IM, Santos CL, Bisordi I, Petrella SM, Pereira LE, Souza RP, et al. St. Louis encephalitis virus: ﬁrst isolation from a human in Sao Paulo State, Brazil. Rev Inst Med Trop Sao Paulo 2005;47:281–5. 10. Mondini A, Cardeal IL, Lazaro E, Nunes SH, Moreira CC, Rahal P, et al. Saint Louis encephalitis virus, Brazil. Emerg Infect Dis 2007;13:176–8. 11. Terzian AC, Mondini A, Bronzoni RV, Drumond BP, Ferro BP, Cabrera EM, et al. Detection of Saint Louis encephalitis virus in dengue-suspected cases during a dengue 3 outbreak. Vector Borne Zoonotic Dis 2011;11:291–300. 12. Mondini A, Bronzoni RV, Cardeal IL, dos Santos TM, Lazaro E, Nunes SH, et al. Simultaneous infection by DENV-3 and SLEV in Brazil. J Clin Virol 2007;40:84–6. 13. Lanciotti RS, Calisher CH, Gubler DJ, Chang GJ, Vorndam AV. Rapid detection and typing of dengue viruses from clinical samples by using reverse transcriptasepolymerase chain reaction. J Clin Microbiol 1992;30:545–51. 14. Chavez JH, Reis VP, Silva JR, Laure HJ, Rosa JC, Fonseca BA, et al. Production and diagnostic application of recombinant domain III of West Nile envelope protein in Brazil. Rev Soc Bras Med Trop 2013;46:97–9. 15. Shope RE, Sather GE. Arboviruses. In: Lennette E. H, Schmidt N.J. editors. Diagnostic procedures for viral, rickettsial and chlamydial infections. Washington, DC: Public Health Association; 1979. 16. de Morais Bronzoni RV, Baleotti FG, Ribeiro Nogueira RM, Nunes M, Moraes Figueiredo LT. Duplex reverse transcription-PCR followed by nested PCR assays for detection and identiﬁcation of Brazilian alphaviruses and ﬂaviviruses. J Clin Microbiol 2005;43:696–702. 17. Spinsanti L, Basquiera AL, Bulacio S, Somale V, Kim SC, Re V, et al. St. Louis encephalitis in Argentina: the ﬁrst case reported in the last seventeen years. Emerg Infect Dis 2003;9:271–3.