Diagnosis of hantavirus infection in humans and rodents in Ribeirão Preto, State of São Paulo, Brazil

INTRODUCTION: Hantavirus pulmonary and cardiovascular syndrome (HPCS) is an emerging serious disease in the Americas. Hantaviruses (Bunyaviridae) are the causative agents of this syndrome and are mainly transmitted through inhalation of aerosols containing the excreta of wild rodents. In the Ribeirão Preto region (state of São Paulo, Brazil), HPCS has been reported since 1998, caused by the Araraquara virus (ARAV), for which Necromys lasiurus is the rodent reservoir. This study aimed to show diagnostic results relating to infection in humans and rodents, obtained at the Virology Research Center of the Ribeirão Preto School of Medicine, University of São Paulo, between 2005 and 2008. METHODS: HPCS was diagnosed by means of ELISA and/or RT-PCR in 11 (21.2%) out of 52 suspected cases, and 54.4% of these were fatal. Furthermore, 595 wild rodents (Necromys lasiurus, Akodon sp, Calomys tener and Oligoryzomys sp) were caught between 2005 and 2008. RESULTS: Fifteen (2.5%) of these rodents presented antibodies for hantavirus, as follows: Necromys lasiurus (4%), Calomys tener (1.9%) and Akodon sp (1.5%). Nucleotide sequences obtained through RT-PCR from one HPCS patient and one Calomys tener rodent were compared with hantavirus sequences from GenBank, which showed that both were homologous with ARAV. CONCLUSIONS: This work corroborates previous studies showing that ARAV is the hantavirus causing HPCS in the Ribeirão Preto region. It also shows that rodents infected with hantavirus represent a constant risk of transmission of this virus to man.

Serosurvey of hantavirus infection in humans in the border region between Brazil and Argentina

INTRODUCTION: According to reports by the Ministry of Health, in the far western region of the State of Santa Catarina, there have been no reports of hantavirus pulmonary syndrome, a zoonotic disease transmitted by feces of infected rodents. A seroepidemiological study of residents of this region, was conducted, with the aim of determining the presence of hantavirus infections. A total of 340 volunteers of both genus, from the towns of Belmonte and Paraíso, were studied. METHODS: The serum of these patients was collected and used to detect IgG antibodies against recombinant N protein of Araraquara hantavirus, by ELISA assay. The positive samples were then titrated and confirmed by immunofluorescence assay. RESULTS: This study demonstrated the presence of IgG antibodies against hantavirus N protein in 3.5% of the population. The most frequent occupation was farm worker, 81% had direct and indirect contact with rodents, 91.7% of positive cases were farm workers, indicating that the probable cause of infection occurred during barn cleaning. These antibodies are noteworthy, given that the levels of antibodies were verified in individuals whose contact with hantavirus may have occurred many years ago. CONCLUSIONS: This study shows the circulation of hantavirus in the region, a fact that until now, had not reported. All the serum reagents had contact with the pathogen, but did not develop pulmonary and cardiovascular syndrome. It is important to remain alert, because hantavirus is a serious and emerging disease of some relevance.

Study of hantavirus infection in captive breed colonies of wild rodents

Wild sigmondontine rodents are known to be the reservoir of several serotypes of New World hantaviruses. The mechanism of viral transmission is by aerosol inhalation of the excreta from infected rodents. Considering that the captive breed colonies of various wild mammals may present a potencial risk for hantaviral transmission, we examined 85 speciemens of Thrichomys spp. (Echimyidae) and 17 speciemens of Nectomys squamipes (Sigmodontinae) from our colony for the presence of hantavirus infections. Blood samples were assayed for the presence of antibodies to Andes nucleocapsid antigen using enzyme-linked immunosorbent assay (ELISA). Additionally, serum samples from workers previously exposed to wild rodents, in the laboratories where the study was conducted, were also tested by ELISA to investigate prevalence of anti-hantavirus IgG antibodies. All blood samples were negative for hantavirus antibodies. Although these results suggest that those rodent's colonies are hantavirus free, the work emphasizes the need for hantavirus serological monitoring in wild colonized rodents and secure handling potentially infected rodents as important biosafety measures.

Serological evidence of hantavirus infection in rural and urban regions in the state of Amazonas, Brazil

Hantavirus disease is caused by the hantavirus, which is an RNA virus belonging to the family Bunyaviridae. Hantavirus disease is an anthropozoonotic infection transmitted through the inhalation of aerosols from the excreta of hantavirus-infected rodents. In the county of Itacoatiara in the state of Amazonas (AM), Brazil, the first human cases of hantavirus pulmonary and cardiovascular syndrome were described in July 2004. These first cases were followed by two fatal cases, one in the municipality of Maués in 2005 and another in Itacoatiara in 2007. In this study, we investigated the antibody levels to hantavirus in a population of 1,731 individuals from four different counties of AM. Sera were tested by IgG/IgM- enzyme-linked immune-sorbent assay using a recombinant nucleocapsid protein of the Araraquara hantavirus as an antigen. Ten sera were IgG positive to hantavirus (0.6%). Among the positive sera, 0.8% (1/122), 0.4% (1/256), 0.2% (1/556) and 0.9% (7/797) were from Atalaia do Norte, Careiro Castanho, Itacoatiara and Lábrea, respectively. None of the sera in this survey were IgM-positive. Because these counties are distributed in different areas of AM, we can assume that infected individuals are found throughout the entire state, which suggests that hantavirus disease could be a local emerging health problem.

Evidence of hantavirus infection in wild rodents captured in a rural area of the state of São Paulo, Brazil

Hantaviruses are the etiological agents of the Hantavirus Cardio-Pulmonary Syndrome, a serious rodent-borne disease in Brazil. In order to investigate the occurrence of hantavirus infection in wild rodents, a survey was conducted in three different suburban areas of the municipality of Pedreira, State of São Paulo, Brazil. Of the 145 wild animals captured belonging to 12 different species identified by morphology and karyological analysis, 107 were rodents of the following species: Akodon montensis, Bolomys lasiurus, Calomys tener, Oligoryzomys nigripes, Oligoryzomys flavescens, and Myocastor coypus. Blood samples from these rodents were assayed for the presence of antibodies against hantavirus by IgG ELISA using Andes recombinant nucleocapsid antigen. Antibody reactive to Andes virus was found in two different species, O. nigripes and O. flavescens. These results indicate a potential risk for hantavirus transmission to humans in this area, where reservoir rodents are present in peridomestic settings.

HANTAVIRUS PULMONARY SYNDROME (HPS) IN GUARIBA, SP, BRAZIL: REPORT OF 2 CASES

Human infections caused by a hantavirus were reported in different regions of the State of São Paulo (SP), Brazil during the first six months of 1998. Two cases of fatal pulmonary syndrome occurred in May of 1998 in the City of Guariba, located in the Northeastern Region of SP. Both patients worked in a corn storage barn infested by rodents. These patients, after 2 or 3 days of non-specific febrile illness, developed a severe interstitial pneumonia spreading widely in both lungs, causing respiratory failure and death. At autopsy both patients showed lung interstitial edema with immunoblast-like mononuclear cell infiltrates, consistent with a viral etiology. Hantavirus infection was diagnosed by ELISA in both cases and by RT-PCR in one of the patients. Aspects of the clinical presentation, physiopathology and differential diagnosis of Hantavirus Pulmonary Syndrome are discussed.

Serological survey of hantavirus in rodents in Uberlândia, Minas Gerais, Brazil

We conducted a serological survey to determine the presence of hantavirus infection in rodents in Uberlândia, Minas Gerais as well as to identify and characterize associated factors. Rodents were captured using Sherman live-capture traps set in rural and peri-urban environments. A total of 611 rodents were captured. There was a higher trap success in peri-urban areas (26.3%) and a higher prevalence of antibodies among rodents captured in rural areas (2.9%). Necromys lasiurus was the most common species (42.2%) and the more frequently infected (4.6%). One Calomys tener (1/141; 0.7%) and one Calomys sp. (1/14; 7.1%) were also positive for the hantavirus infection. In N. lasiurus, antibody prevalence correlated with population density (p < 0.01), age class (p = 0.003) and presence of scars (p = 0.02). The data confirm that horizontal transmission is the main mechanism that maintains the virus in nature. The higher seropositivity in N. lasiurus is consistent with genetic studies that associate this species with an Araraquara virus reservoir; the seropositivity of C. tener and Calomys sp. may indicate the occurrence of spillover infection or the presence of other circulating hantaviruses.

Prevalence of serum antibodies to hantavirus in a rural population from the southern state of Santa Catarina, Brazil

INTRODUCTION: Rodent-borne hantaviruses cause severe human diseases. We completed a serological survey of hantavirus infection in rural inhabitants of Turvo County, in the southern State of Santa Catarina, Brazil, in which seropositivity for hantavirus was correlated to previous disease in the participants. METHODS: The levels of IgG antibodies to hantavirus Araraquara in the sera of 257 individuals were determined using an immunoenzymatic assay. RESULTS: IgG antibodies to hantavirus were found in 2.3% of the participants. All seropositive participants reported previous disease with symptoms suggestive of hantavirus cardiopulmonary syndrome. CONCLUSIONS: Human infections causing unreported cardiopulmonary syndrome probably occur in the southern State of Santa Catarina.

Expression of a hantavirus N protein and its efficacy as antigen in immune assays

Hantavirus cardiopulmonary syndrome (HCPS) has been recognized as an important public heath problem. Five hantaviruses associated with HCPS are currently known in Brazil: Juquitiba, Araraquara, Laguna Negra-like, Castelo dos Sonhos, and Anajatuba viruses. The laboratory diagnosis of HCPS is routinely carried out by the detection of anti-hantavirus IgM and/or IgG antibodies. The present study describes the expression of the N protein of a hantavirus detected in the blood sample of an HCPS patient. The entire S segment of the virus was amplified and found to be 1858 nucleotides long, with an open reading frame of 1287 nucleotides that encodes a protein of 429 amino acids. The nucleotide sequence described here showed a high identity with the N protein gene of Araraquara virus. The entire N protein was expressed using the vector pET200D and the Escherichia coli BL21 strain. The expression of the recombinant protein was confirmed by the detection of a 52-kDa protein by Western blot using a pool of human sera obtained from HCPS patients, and by specific IgG detection in five serum samples of HCPS patients tested by ELISA. These results suggest that the recombinant N protein could be used as an antigen for the serological screening of hantavirus infection.

Prevalence of antibody to hantaviruses in humans and rodents in the Caribbean region of Colombia determined using Araraquara and Maciel virus antigens

We tested sera from 286 agricultural workers and 322 rodents in the department of Córdoba, northeastern Colombia, for antibodies against two hantaviruses. The sera were analysed by indirect ELISA using the lysate of Vero E6 cells infected with Maciel virus (MACV) or the N protein of Araraquara virus (ARAV) as antigens for the detection of antibodies against hantaviruses. Twenty-four human sera were IgG positive using one or both antigens. We detected anti-MACV IgG antibodies in 10 sera (3.5%) and anti-ARAV antibodies in 21 sera (7.34%). Of the 10 samples that were positive for MACV, seven (70%) were cross-reactive with ARAV; seven of the 21 ARAV-positive samples were cross-reactive with MACV. Using an ARAV IgM ELISA, two of the 24 human sera (8.4%) were positive. We captured 322 rodents, including 210 Cricetidae (181 Zygodontomys brevicauda, 28 Oligoryzomys fulvescens and 1 Oecomys trinitatis), six Heteromys anomalus (Heteromyidae), one Proechimys sp. (Echimyidae) and 105 Muridae (34 Rattus rattus and 71 Mus musculus). All rodent sera were negative for both antigens. The 8.4% detection rate of hantavirus antibodies in humans is much higher than previously found in serosurveys in North America, suggesting that rural agricultural workers in northeastern Colombia are frequently exposed to hantaviruses. Our results also indicate that tests conducted with South American hantavirus antigens could have predictive value and could represent a useful alternative for the diagnosis of hantavirus infection in Colombia.

Prevalence of infection with hantavirus in rodent populations of central Argentina

We studied hantavirus seroprevalence and virus variability in rodent populations in Diego Gaynor, northwest of Buenos Aires province, Argentina. Rodent samplings were conducted in railroads and cropfield borders in March and July 1999, September and December 2000, and March 2001. Antibody detection was performed by an enzyme link immunosorbent assay (ELISA), using the recombinant nucleoprotein of Andes (AND) virus as antigen. Tissue samples were taken from positive antibody individuals in order to confirm the presence of hantavirus genomic material and to identify virus genotypes. Akodon azarae was the most abundant species, followed by Oligoryzomys flavescens, while Calomys laucha and C. musculinus were rarely caught. We found a rate of seroprevalence of 9.3% for a total sample of 291 A. azarae and 13.5% for 37 O. flavescens. After molecular analyses of hantavirus, we confirmed the presence of hantavirus genomic material in 16 individuals with ELISA (+) results and two individuals with ELISA (-). Four amplimers for each species were sequenced and compared to the corresponding sequences of representative hantaviruses. We identified the AND Cent Lec from three O. flavescens, and the Pergamino virus from four A. azarae and from one O. flavescens. A. azarae males had higher seroprevalence than females, and heavier individuals showed higher seroprevalence than lighter ones. We did not find seroprevalence differences according to sex in O. flavescens, although this result may have been produced by the low sample size. The lowest seroprevalence was found in a period of high rodent density, when juveniles prevailed in the population. We found higher seroprevalences than those detected in previous studies for other localities of central Argentina where cases of hantavirus pulmonary syndrome (HPS) have been reported. The presence of AND Cent Lec virus in rodent populations of the study area, which is responsible of HPS cases in central Argentina, suggests that human populations are at risk of HPS disease, although there were not reported cases of this disease until today.

Experimental infection of Rio Mamore hantavirus in Sigmodontinae rodents

This study shows an experimental spillover infection of Sigmodontinae rodents with Rio Mamore hantavirus (RIOMV). Necromys lasiurus and Akodon sp were infected with 103 RNA copies of RIOMV by intraperitoneal administration. The viral genome was detected in heart, lung, and kidney tissues 18 days after infection (ai), and viral excretion in urine and faeces began at four and six ai, respectively. These results reveal that urine and faeces of infected rodents contain the virus for at least 18 days. It is possible that inhaled aerosols of these excreta could transmit hantavirus to humans and other animals.

HANTAVIRUS PULMONARY SYNDROME IN THE PROVINCE OF RIO NEGRO, ARGENTINA, 1993-1996

Early in 1995 the first case of Hantavirus Pulmonary Syndrome was serologically confirmed in El Bolsón (Province of Río Negro, Argentina), corresponding to the third outbreak reported in Argentina. A total of 26 cases of HPS related to the Andean region of Rio Negro Province, were reported from 1993 to 1996, 17 in El Bolsón, 4 in San Carlos de Bariloche, and 5 in Buenos Aires. The incidence rate was 5.03 x 100000 with a mortality rate of 51.85 x 100. The occurrence of cases was mainly seasonal, with a significantly greater number in the spring, and the persons affected mainly lived in urban or periurban areas. In four cases, the affected individuals were members of a couple, spouses or live-in contacts. Seven cases were Health workers (physicians, nurses or administrative staff). Twelve cases were related among them, due to an outbreak of 80 days. Two of them did not visit the Andean region. A total of 139 rodents were captured and seven of them, Olygoryzomys longicaudatus, were found to be serologically positive. The possibility of infection by contact with rodents or fecal matter is being analyzed and also hypothesis related with interhuman transmission