RESUMO
Introduction: There is not known if a viraemia post-oral polio vaccine (OPV) is detectable by modern molecular techniques. Such viraemia could affect the performance of the real time-polymerase chain reaction (PCR) for non polio enterovirus (EV) detection, technique of growing clinical use for the study of febrile infants. Objective: To determine viraemia post-first dose of OPV in healthy infants, by molecular techniques. Patients and Methods: 50 infants less than three months without previous VPO were randomized in 5 groups: a control group with pre-vaccination blood sample (BS), group 1 BS at day 2, group 2 BS at day 4, group 3, BS at day 6 and group 4, BS at day 8 post-vaccination. Conventional and specific PCR for poliovirus and real time PCR for non polio EV were performed in BS and in OPV samples. Results: No genetic material of poliovirus was detected in any infant, while in 9 of them (18%) non polio EV was identified. Real time PCR for EV did not amplify poliovirus from OPV samples. Discussion: Results suggest that no post VPO viraemia detectable by molecular methods exists. Considering that real time PCR for EV does not allow to identify polio virus, no false positives of the test are expected as a result of a recent VPO vaccination. We documented presence of non polio EV in blood of healthy asymptomatic infants.
Introducción: No existen estudios que indiquen si la vacuna polio oral (VPO) produce viremia detectable mediante métodos moleculares. Una eventual viremia podría afectar el rendimiento de la RPC tiempo real para detectar enterovirus (EV) no polio, examen de creciente uso clínico en lactantes pequeños con fiebre sin foco. Objetivo: Determinar viremia post VPO en lactantes sanos, por métodos moleculares. Métodos: 50 menores de 3 meses, al momento de recibir su primera VPO se distribuyeron en forma aleatoria en 5 grupos: control, muestra de sangre pre-vacunación; grupo 1, muestra al 2° día; grupo 2, al 4° día; grupo 3, al 6° día y grupo 4, al 8° día post-vacunación. Se realizó RPC convencional específica para virus polio y RPC tiempo real para EV no polio en las muestras de sangre y en muestras de VPO. Resultados: No se identificó presencia de material genético de virus polio en lactante alguno, mientras que en 9 (18%) se identificó presencia de EV no polio. La RPC tiempo real para EV no polio no amplificó material genético a partir de las muestras de VPO. Discusión: Los resultados sugieren que no existe viremia post-VPO detectable por métodos moleculares. Considerando que la RPC tiempo real de EV no polio de uso clínico no permite identificar la presencia de virus polio, estos hallazgos indican que no existirán falsos positivos de este examen como resultado de una vacunación VPO reciente. Adicionalmente se documentó presencia de EV no polio en sangre de lactantes asintomáticos.
Assuntos
Feminino , Humanos , Lactente , Masculino , Anticorpos Antivirais/sangue , Enterovirus/isolamento & purificação , Poliovirus , Poliomielite/prevenção & controle , Vacina Antipólio Oral/imunologia , Enterovirus Humano B/genética , Enterovirus Humano B/isolamento & purificação , Enterovirus/genética , Poliomielite/imunologia , Poliovirus/genética , Poliovirus/imunologia , Reação em Cadeia da Polimerase em Tempo RealRESUMO
INTRODUCTION: There is not known if a viraemia post-oral polio vaccine (OPV) is detectable by modern molecular techniques. Such viraemia could affect the performance of the real time-polymerase chain reaction (PCR) for non polio enterovirus (EV) detection, technique of growing clinical use for the study of febrile infants. OBJECTIVE: To determine viraemia post-first dose of OPV in healthy infants, by molecular techniques. PATIENTS AND METHODS: 50 infants less than three months without previous VPO were randomized in 5 groups: a control group with pre-vaccination blood sample (BS), group 1 BS at day 2, group 2 BS at day 4, group 3, BS at day 6 and group 4, BS at day 8 post-vaccination. Conventional and specific PCR for poliovirus and real time PCR for non polio EV were performed in BS and in OPV samples. RESULTS: No genetic material of poliovirus was detected in any infant, while in 9 of them (18%) non polio EV was identified. Real time PCR for EV did not amplify poliovirus from OPV samples. DISCUSSION: Results suggest that no post VPO viraemia detectable by molecular methods exists. Considering that real time PCR for EV does not allow to identify polio virus, no false positives of the test are expected as a result of a recent VPO vaccination. We documented presence of non polio EV in blood of healthy asymptomatic infants.
Assuntos
Anticorpos Antivirais/sangue , Enterovirus/isolamento & purificação , Poliomielite/prevenção & controle , Vacina Antipólio Oral/imunologia , Poliovirus , Enterovirus/genética , Enterovirus Humano B/genética , Enterovirus Humano B/isolamento & purificação , Feminino , Humanos , Lactente , Masculino , Poliomielite/imunologia , Poliovirus/genética , Poliovirus/imunologia , Reação em Cadeia da Polimerase em Tempo RealRESUMO
The emergence and rapid spread of the 2009 H1N1 pandemic influenza virus showed that many diagnostic tests were unsuitable for detecting the novel virus isolates. In most countries the probe-based TaqMan assay developed by the U.S. Centers for Disease Control and Prevention was used for diagnostic purposes. The substantial sequence data that became available during the course of the pandemic created the opportunity to utilize bioinformatics tools to evaluate the unique sequence properties of this virus for the development of diagnostic tests. We used a comprehensive computational approach to examine conserved 2009 H1N1 sequence signatures that are at least 20 nucleotides long and contain at least two mismatches compared to any other known H1N1 genome. We found that the hemagglutinin (HA) and neuraminidase (NA) genes contained sequence signatures that are highly conserved among 2009 H1N1 isolates. Based on the NA gene signatures, we used Visual-OMP to design primers with optimal hybridization affinity and we used ThermoBLAST to minimize amplification artifacts. This procedure resulted in a highly sensitive and discriminatory 2009 H1N1 detection assay. Importantly, we found that the primer set can be used reliably in both a conventional TaqMan and a SYBR green reverse transcriptase (RT)-PCR assay with no loss of specificity or sensitivity. We validated the diagnostic accuracy of the NA SYBR green assay with 125 clinical specimens obtained between May and August 2009 in Chile, and we showed diagnostic efficacy comparable to the CDC assay. Our approach highlights the use of systematic computational approaches to develop robust diagnostic tests during a viral pandemic.
Assuntos
Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Influenza Humana/diagnóstico , Compostos Orgânicos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Virologia/métodos , Benzotiazóis , Chile , Primers do DNA/genética , Diaminas , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Neuraminidase/genética , Quinolinas , Sensibilidade e Especificidade , Coloração e Rotulagem/métodos , Estados Unidos , Proteínas Virais/genéticaRESUMO
Hantavirus cardiopulmonary syndrome (HCPS) is a highly pathogenic emerging disease (40% case fatality rate) caused by New World hantaviruses. Hantavirus infections are transmitted to humans mainly by inhalation of virus-contaminated aerosol particles of rodent excreta and secretions. At present, there are no antiviral drugs or immunotherapeutic agents available for the treatment of hantaviral infection, and the survival rates for infected patients hinge largely on early virus recognition and hospital admission and aggressive pulmonary and hemodynamic support. In this study, we show that Andes virus (ANDV) interacts with human apolipoprotein H (ApoH) and that ApoH-coated magnetic beads or ApoH-coated enzyme-linked immunosorbent assay plates can be used to capture and concentrate the virus from complex biological mixtures, such as serum and urine, allowing it to be detected by both immunological and molecular approaches. In addition, we report that ANDV-antigens and infectious virus are shed in urine of HCPS patients.
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Antígenos Virais/urina , Síndrome Pulmonar por Hantavirus/urina , Orthohantavírus/imunologia , beta 2-Glicoproteína I/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Antígenos Virais/sangue , Antígenos Virais/imunologia , Chlorocebus aethiops , Ensaio de Imunoadsorção Enzimática , Síndrome Pulmonar por Hantavirus/sangue , Síndrome Pulmonar por Hantavirus/imunologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Microesferas , RNA Viral/análise , Células VeroRESUMO
BACKGROUND: Andes virus (ANDV) infection, which has a case fatality rate of 37% in Chile, often occurs in household clusters and may be transmitted from person to person. METHODS: To determine the incidence and risk factors for additional household cases, we conducted a prospective study among recent household contacts of persons with hantavirus cardiopulmonary syndrome (HCPS) in Chile, including testing of serum for anti-hantavirus antibodies and blood cells for ANDV RNA by reverse-transcription polymerase chain reaction (RT-PCR). RESULTS: We enrolled 76 index case patients and 476 household contacts, of whom 16 (3.4%) developed HCPS; 32.6% of 92 cases occurred in household clusters. The risk of HCPS was 17.6% among sex partners of index case patients, versus 1.2% among other household contacts (P<.001). Person-to-person transmission was definite in 3, probable in 9, and possible in 2 of the 16 additional household case patients. We detected ANDV RNA by RT-PCR in peripheral blood cells 5-15 days before the onset of symptoms or the appearance of anti-hantavirus antibodies. CONCLUSIONS: In recent household contacts of persons with HCPS in Chile, the risk of HCPS was greatest among sex partners. Among the household contacts who developed HCPS, viremia preceded the onset of symptoms and the appearance of anti-hantavirus antibodies by up to 2 weeks.
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Busca de Comunicante , Características da Família , Síndrome Pulmonar por Hantavirus/transmissão , Orthohantavírus/isolamento & purificação , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/sangue , Criança , Pré-Escolar , Chile/epidemiologia , Feminino , Orthohantavírus/genética , Orthohantavírus/imunologia , Síndrome Pulmonar por Hantavirus/epidemiologia , Síndrome Pulmonar por Hantavirus/virologia , Humanos , Incidência , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , RNA Viral/sangue , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Risco , Viremia/epidemiologia , Viremia/transmissão , Viremia/virologiaRESUMO
We conducted a 16S rRNA nested PCR for the genus Ehrlichia and Ehrlichia spp. with blood samples from 30 ill dogs in Chile. Phylogenetic analysis was performed by using groESL gene amplification. We identified Anaplasma platys as 1 of the etiologic agents of canine ehrlichiosis.
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Anaplasma/isolamento & purificação , Anaplasmose/microbiologia , Doenças do Cão/microbiologia , Anaplasma/classificação , Anaplasma/genética , Anaplasmose/epidemiologia , Animais , Chile/epidemiologia , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , Doenças do Cão/epidemiologia , Cães , FilogeniaRESUMO
Amantadine has been used for prevention and treatment of influenza A infection. It blocks the proton through the M2 ion channel. Drug-resistant viruses appear quickly when this therapy is used. Single amino acids changes in the H2 protein can confer resistance, being the most frequent one in position 31. Different methods to detect resistant strains have been described. The objectives were to determine the existence of amantadine resistance of influenza A strains isolated in a virologic laboratory in Santiago, Chile, between 2001-2002, and to validate a new molecular method to detect resistant strains. A PCR restriction fragment length polymorphism analysis was employed for the detection of resistant viruses. In 31 processed strains no mutation in the position 31 was found. This result supports that amantadine resistance is very low or absent in Chile. This could be explained by a limited use of this drug in the study population. This method could be used as a monitoring system to survey resistant viruses.
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Amantadina/farmacologia , Antivirais/farmacologia , Farmacorresistência Viral , Vírus da Influenza A/efeitos dos fármacos , Proteínas da Matriz Viral/genética , Adolescente , Adulto , Idoso , Animais , Linhagem Celular , Criança , Pré-Escolar , Chile , Cães , Farmacorresistência Viral/genética , Feminino , Humanos , Lactente , Vírus da Influenza A/genética , Masculino , Pessoa de Meia-Idade , Mutação , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , RNA Viral/genética , RNA Viral/isolamento & purificaçãoRESUMO
BACKGROUND: The availability of a serologic test for cat scratch disease in humans has allowed the diagnosis of an increasing number of cases of this disease in Chile. AIM: To perform a serological survey for Bartonella henselae among cats in Chile. MATERIAL AND METHODS: Blood samples from 187 cats living in three Chilean cities were obtained. IgG antibodies against Bartonella henselae were measured using indirect immunofluorescence. Blood cultures were done in 60 samples. The presence of Bartonella henselae in positive cultures was confirmed by restriction fragment length polymorphism polymerase chain reaction (RFLP-PCR). RESULTS: The general prevalence of IgG antibodies against Bartonella henselae was 85.6%. No differences in this prevalence were found among cats younger or older than 1 year, or those infested or not infested with fleas. However domestic cats had a lower prevalence when compared with stray cats (73 and 90% respectively, p <0.01). Bartonella henselae was isolated in 41% of blood cultures. All the isolated were confirmed as Bartonella henselae by RFLP-PCR. CONCLUSIONS: This study found an important reservoir of Bartonella henselae in Chilean cats and therefore a high risk of exposure in humans who have contact with them.
Assuntos
Bartonella henselae/isolamento & purificação , Gatos/microbiologia , Reservatórios de Doenças/microbiologia , Animais , Anticorpos Antibacterianos/sangue , Bartonella henselae/imunologia , Distribuição de Qui-Quadrado , Chile , Técnica Indireta de Fluorescência para Anticorpo , Imunoglobulina G/sangue , Estudos SoroepidemiológicosRESUMO
An epidemiologic and seroprevalence survey was conducted (n=830) to assess the proportion of persons exposed to hantavirus in IX Region Chile, which accounts for 25% of reported cases of hantavirus cardiopulmonary syndrome. This region has three geographic areas with different disease incidences and a high proportion of aboriginals. Serum samples were tested for immunoglobulin (Ig) G antibodies by enzyme-linked immunosorbent assay against Sin Nombre virus N antigen by strip immunoblot assay against Sin Nombre, Puumala, Río Mamoré, and Seoul N antigens. Samples from six patients were positive for IgG antibodies reactive with Andes virus; all patients lived in the Andes Mountains. Foresting was also associated with seropositivity; but not sex, age, race, rodent exposure, or farming activities. Exposure to hantavirus varies in different communities of IX Region. Absence of history of pneumonia or hospital admission in persons with specific IgG antibodies suggests that infection is clinically inapparent.
Assuntos
Infecções por Hantavirus/epidemiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/sangue , Chile/epidemiologia , Feminino , Orthohantavírus/imunologia , Orthohantavírus/isolamento & purificação , Infecções por Hantavirus/imunologia , Humanos , Imunoglobulina G/sangue , Masculino , Pessoa de Meia-Idade , PrevalênciaRESUMO
Dengue virus was detected for the first time in Chile, in an outbreak of dengue fever on Easter Island. The virus was isolated in tissue culture and characterized by reverse transcription-polymerase chain reaction as being dengue type 1.