Biomedical subjects

M G Guzmán

Publications and source records attributed to M G Guzmán.

At least 19 recordsLinked to original sources

PubMed2006

Race: a risk factor for dengue hemorrhagic fever.

Cuban DHF/DSS outbreaks have provided evidence of a reduced risk of people of Negroid race for DHF/DSS compared to those of Caucasoid race. These observations from Cuban dengue outbreaks have significant epidemiological interest, as the differences in susceptibility to DHF/DSS among racial groups in Cuba coincide with that reported in African and Black Caribbean populations. In this article, we review the literature on race as a risk factor for DHF/DSS and discuss recent results from ongoing studies. Taking into consideration the origins of contemporary Cuban inhabitants, we propose that the Cuban, Caribbean Black and African populations share a common gene pool that could explain, at least partially, the low incidence of dengue hemorrhagic fever in Cuba and Caribbean and African countries. The central role played by immunological mechanisms in the pathogenesis of DHF/DSS has led us to consider that the polymorphic genes associated with the immune response must be carefully considered among those human genes regulating dengue disease severity that might be distributed unequally in Blacks and Whites.

Black People↗

PubMed2006

Kinetics of antibodies in sera, saliva, and urine samples from adult patients with primary or secondary dengue 3 virus infections.

OBJECTIVES: The kinetics of three serological markers (IgM, IgA, and IgG) in serum, saliva, and urine samples from adult patients with primary or secondary dengue infection were studied. DESIGN: Serum, saliva, and urine samples were collected from 22 patients with clinical and confirmed dengue 3 virus infection during the outbreak in Havana City in 2001. They were tested by capture IgM (MAC-ELISA), IgA (AAC-ELISA), and IgE (EAC-ELISA) and IgG ELISA inhibition method (EIM) to detect specific dengue antibodies. RESULTS: Similar kinetics were observed in IgM, IgA, and IgG antibodies in saliva and IgA and IgG in urine samples from secondary cases compared with kinetics in serum samples, although the values were lower. No IgG antibody was detected in saliva and urine samples in primary cases and IgM antibody was not detected in urine samples from either primary or secondary infection. All secondary cases were positive for IgG in saliva and urine samples at day 7. The kinetics of specific IgE antibodies in primary and secondary cases were different. CONCLUSIONS: The kinetics of three serological markers (IgM, IgA, and IgG) in serum, saliva, and urine samples from adult patients with primary or secondary dengue 3 virus infection were studied for the first time, showing its behavior and usefulness in dengue virus diagnosis. The specific IgE could play a role as a serological marker in secondary infections.

Adult↗

PubMed2006

Antibodies from patients with dengue viral infection mediate cellular cytotoxicity.

Acute and late convalescent sera (collected at day 5 of disease onset and 1 year later) from dengue fever (DF) and dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) laboratory confirmed cases, were tested for antibody-dependent cell-mediated cytotoxicity (ADCC) activity using dengue 1 (DENV-1) or dengue 2 (DENV-2) infected cells as target. All patients experienced their first dengue virus (DENV) infection 20 years before. ADCC activity was detected in acute sera from DHF/DSS but not in sera from DF patients. However, 1 year after illness, ADCC activity was observed in all cases. This preliminary report represents one of the few studies of ADCC in dengue patients and suggests that ADCC could be implicated in dengue pathogenesis.

Antibodies, Viral↗

PubMed2006

Detection and subtyping of dengue 1-4 and yellow fever viruses by means of a multiplex RT-nested-PCR using degenerated primers.

OBJECTIVE: Differential diagnosis of infections that cause similar diseases and may be active simultaneously in the same geographical areas is greatly needed. Dengue and yellow fever viruses (DENV and YFV) are transmitted by the same species of mosquito and both can cause haemorrhagic fever symptoms. These viruses are active mainly in regions where expensive and sophisticated technologies are not available. Our objective was to develop a simple, reliable and easy-to-perform method to detect and identify these viruses. METHODS: We slightly modified a generic RT-PCR able to detect the mentioned viruses and other members of this genus: specific primers for each one of these viruses were designed and included in the nested reaction instead of one of the generic ones. The reaction was optimized and viruses are amplified giving rise to bands of different sizes distinguishable in agarose gels. RESULTS: This test is able to detect and identify the four DENVs and YFV to a high level of sensitivity and specificity and can be used with clinical samples. This simple, reliable and easy-to-perform method able to detect and identify dengue 1-4 and YFV can be used in poor endemic countries.

DNA Primers↗

PubMed2005

Generic RT-nested-PCR for detection of flaviviruses using degenerated primers and internal control followed by sequencing for specific identification.

Flaviviruses are a widespread and numerous group of arboviruses that can cause serious illness in humans. The continuous and slow spread of certain flaviviruses, such as Dengue viruses, and the recent entry and spread of West Nile virus to the American continent, point to the need to control these infections. This control requires the use of suitable techniques for diagnostic and surveillance programmes. A generic RT-nested-PCR that is, theoretically, able to detect each member of the group has been designed. The identification of the detected virus is carried out by sequencing. The introduction of an internal control would reduce the number of false negative results and could be used to quantify the viral load in clinical samples where the method works well.

False Negative Reactions↗

PubMed2004

Serological markers during dengue 3 primary and secondary infections.

BACKGROUND: The detection of the IgM antibody for the dengue virus in serum by ELISA has become one of the most important and useful methods for diagnosis of dengue using a single acute-phase serum sample. Currently, this system is an invaluable tool for the surveillance of dengue fever (DF) and dengue hemorrhagic fever (DHF). The usefulness of other serological markers such as IgA and IgE have been less studied. OBJECTIVE: To study the IgM, IgA and IgE specific antibody response in dengue 3 infected patients with different clinical picture and type of infection. STUDY DESIGN: One hundred and twenty-seven serum samples collected on days 5-7 at the onset of fever from clinically and serologically confirmed dengue cases were studied. Forty-two were classified as primary dengue fever cases, 48 as secondary dengue fever cases and 37 as secondary dengue hemorrhagic fever cases. All samples were tested by capture ELISA in order to detect dengue IgM, IgA and IgE antibodies. RESULTS AND CONCLUSIONS: In this study, significant differences were observed in the IgM, IgA and IgE response between the study groups. High IgA and IgE OD ratios in secondary dengue cases were found. The usefulness of serotype specific IgM antibody detection is also analyzed and discussed. A priority for future dengue research in terms of protection, recovery of infection and immunopathogenesis is to elucidate the role of these immunoglobulins. The cross reactivity response to IgM between dengue virus serotypes in primary and secondary cases should also be more studied.

Antibodies, Viral↗

PubMed2003

Carboxy-terminally truncated Dengue 4 virus envelope glycoprotein expressed in Pichia pastoris induced neutralizing antibodies and resistance to Dengue 4 virus challenge in mice.

We have expressed a recombinant Dengue 4 virus envelope glycoprotein (E4rec), truncated at its C-terminus by 53 amino acids, in Pichia pastoris. The presence of E4rec was confirmed by Western-blot using anti-DEN 4 hyper immune mouse ascitic fluid. E4rec migrated during SDS-PAGE as a 64 kDa protein. Treatment with endoglycosidases showed that the E protein was modified by the addition of short mannose chains and the absence of hyperglycosylation. When administered to BALB-C mice, E4rec elicited a DEN 4 neutralizing antibody response haemagglutination inhibition antibodies and specific memory T cell response. Mice immunized were also significantly protected against lethal DEN 4 virus challenge (86.6%, p < 0.001).

Animals↗

PubMed2001

Study of biologic attributes of Cuban dengue 2 virus after serial passage in primary dog kidney cells.

OBJECTIVE: The serial passage of dengue viruses in primary dog kidney (PDK) cells has resulted in selection of attenuated viruses. However, the molecular changes responsible for loss of virulence are not well characterized. This article describes the isolation and biologic attributes of one dengue 2 virulent strain as a first step to allow the study of determinants of virulence at the molecular level. METHODS: A15 dengue 2 Cuban strain was isolated from the viremic plasma of a patient with uncomplicated dengue fever during the 1981 epidemic. This was then subjected to serial passage in PDK cells. Viruses resulting from several PDK passages were compared to the parent strain for plaque size and temperature sensitivity, neurovirulence in newborn mice, and cytopathogenic effects on LLC-MK(2) and C6/36-HT cell lines. RESULTS: A15 dengue 2 Cuban strain was successfully propagated in PDK cells. Primary dog kidney 52 to 53 viruses exhibited several biologic attributes, such as small plaques, temperature sensitivity, reduced mouse neurovirulence, and cytopathic effect in permissive cell lines. CONCLUSIONS: These results represent the first step to allow attenuation of this strain of dengue 2 virus.

Animals↗

PubMed2001

Mechanism of enterovirus involvement in epidemic neuropathy: hypothesis regarding pathophysiology.

During the epidemic of optic and peripheral neuropathy which occurred in Cuba in 1992-1993, viruses antigenically related to the Coxsackie viruses were isolated from cerebrospinal fluid of patients. Concurrently with the virologic studies, epidemiologic, toxicologic, nutritional, immunologic, and histopathologic investigations were also carried out. Although it was demonstrated that the illness was associated with toxic and nutritional risk factors, it has not been possible to identify a specific etiology for the symptoms observed. Taking into consideration the results obtained in all of the various investigations, we have formulated an integral, multifactorial hypothesis which attempts to explain a pathophysiologic mechanism by which the viruses isolated could participate in the pathogenesis of the illness. We propose that the viral agents produce a persistent infection, and the possibility that they may act as mediator of an autoimmune pathogenic process.

Coxsackievirus Infections↗

PubMed2000

Epidemiologic studies on Dengue in Santiago de Cuba, 1997.

A small, isolated outbreak of dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) due to dengue virus type 2 (DEN-2) was documented in Santiago de Cuba on the island of Cuba beginning in January 1997. There were 205 DHF/DSS cases, all in persons older than age 15 years. All but three had evidence of a prior dengue infection, with the only known opportunity being the islandwide dengue virus type 1 (DEN-1) epidemic of 1977-1979. Virtually complete clinical and laboratory surveillance of overt disease was achieved. From December 1997 to January 1998, a random, age-stratified serum sample was obtained from 1,151 persons in 40 residential clusters in Santiago. Sera were tested for DEN-1 and DEN-2 neutralizing antibodies. The prevalence of DEN-2 antibodies in children age 15 years and under, born after the 1981 DEN-2 epidemic, was taken as the 1997 DEN-2 infection rate. This was adjusted slightly to accommodate observed cases, resulting in an estimated infection rate of 4.3%. Dengue fever and DHF/DSS attack rates were calculated from estimated total primary and secondary DEN-2 infections. Only 3% of 13,116 primary infections were overt. The DHF/DSS attack rate for adults of all ages was 420 per 10,000 secondary DEN-2 infections.

Adolescent↗

PubMed2000

Do escape mutants explain rapid increases in dengue case-fatality rates within epidemics?

During the Cuban dengue epidemics of 1981 and 1997, significant monthly increases were observed in the proportion of total cases that presented as dengue haemorrhagic fever or dengue shock syndrome (DHF/DSS), and in case-fatality rates for both dengue fever and DHF/DSS. We believe that theses increases can be explained by the hypothesis that some of the population of antibodies against dengue 1 virus raised after natural primary infections react with "neutralisation" determinants found on dengue 2 viruses. These heterotypic antibodies do not prevent secondary dengue 2 infections, but serve to down-regulate the disease to mild illness or symptomless infections. A population of dengue 2 viruses that replicates in dengue-1-immune hosts escape heterotypic neutralisation. When inoculated into a new dengue-1-immune host, these viruses are free to interact with the more abundant infection-enhancing antibodies to produce severe disease.

Antibodies, Viral↗

PubMed2000

Human Dengue antibodies against structural and nonstructural proteins.

Antibodies against dengue virus type 2 and 4 proteins in acute-phase sera of 10 primary and 10 secondary dengue fever and dengue hemorrhagic fever patients were studied by Western blotting. In the first group the immune response was barely detectable, while in the second group more proteins were detected, with a very strong reaction. Anti-NS1 and -NS3 antibodies were detected mainly in secondary cases. Anti-E, -NS3, and -NS5 antibodies were detected in a high number of cases. The possibility of implementing early diagnostic assays for antigen detection is suggested.

Animals↗

PubMed2000

Comparison of rapid centrifugation assay with conventional tissue culture method for isolation of dengue 2 virus in C6/36-HT cells.

A rapid centrifugation assay was compared with conventional tube cell culture for dengue virus isolation in both sera and autopsy samples from dengue and dengue hemorrhagic fever/dengue shock syndrome fatal cases. The rapid centrifugation assay allowed isolation of virus from 16.6% more samples than the conventional method, and it shortened the time for dengue virus detection. Finally, it allowed the isolation of dengue 2 virus in 42.8% of tissue samples from five fatal cases. Our results suggest that the rapid centrifugation assay may be useful for detection of dengue virus in clinical specimens.

Aedes↗

PubMed1999

Fatal dengue hemorrhagic fever in Cuba, 1997.

OBJECTIVES: After more than 15 years without dengue activity, a dengue II epidemic was reported in Cuba in 1997. Three thousand and twelve serologically confirmed cases were reported, with 205 dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) cases and 12 fatalities. This report presents the clinical, serologic, and virologic findings in the 12 fatal DHF/DSS cases. METHODS: Serum and necropsy samples were studied by viral isolation in C636 cell line and polymerase chain reaction. Serum samples were tested by IgM capture enzyme-linked immunoassay (ELISA) and ELISA inhibition method (EIM). RESULTS: All 12 cases were classified as DHF/DSS according to the Pan American Health Organization Guidelines for Control and Prevention of Dengue and Dengue Hemorrhagic Fever in the Americas. All patients were older than 15 years. Women were more frequently affected. The symptoms and signs presented by these patients were similar to those previously described in DHF/DSS cases. Clinical deterioration occurred on average at day 3.75. Abdominal pain and persistent vomiting were the earliest and most frequent warning signs. Dengue infection was confirmed in all cases. IgM antibodies were detected in 11 of 12 cases, all of them with a secondary infection. Dengue II virus was detected by viral isolation in 12 samples and by polymerase chain reaction in 17. Virus or RNA was detected in various tissues, including kidney, heart, lung, and brain. CONCLUSION: The clinical, pathologic, and laboratory features of 12 cases of fatal dengue hemorrhagic fever were reviewed. The results obtained demonstrate that adults with a primary dengue infection are at risk of developing the severe disease (DHF) if they are infected with a different serotype.

Adolescent↗

PubMed1999

[Epidemiology of dengue and hemorrhagic dengue in Santiago, Cuba 1997].

A dengue epidemic that Cuba reported in 1997 registered more than 500,000 cases of dengue fever produced by viral serotype 1. In 1981, there was an epidemic of dengue hemorrhagic fever produced by serotype 2 of the virus. This time 344,203 clinical cases were reported, 10,312 of which were severe cases of hemorrhagic fever that led to 158 fatalities (101 of them among children). The reintroduction of dengue, and specifically of dengue viral serotype 2 (Jamaica genotype), was quickly detected in January 1997 through an active surveillance system with laboratory confirmation of cases in the municipality of Santiago de Cuba, in the province of the same name. The main epidemiological features of this outbreak are reported in this paper. A total of 3,012 cases were reported and serologically confirmed. These included 205 cases classified as dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS), 12 of which were case fatalities (all among adults). Secondary infection with dengue virus was one of the most important risk factors for DHF/DSS. Ninety-eight percent of the DHF/DSS cases and 92% of the fatal cases had contracted a secondary infection. It was the first time dengue hemorrhagic fever was documented as a secondary infection 16 to 20 years after initial infection. Belonging to the white racial group was another important risk factor for DHF/DSS, as had been observed during the 1981 epidemic. During the most recent epidemic it was demonstrated that the so called "fever alert" is not useful for early detection of an epidemic. Measures taken by the country's public health officials prevented spread of the epidemic to other municipalities plagued by Aedes aegypti.

Adolescent↗

PubMed1998

[Polymerase chain reaction for rapid detection and serotyping of dengue virus in clinical samples].

This study describes the benefits of using reverse transcriptase polymerase chain reaction (RT-PCR) for the rapid detection and typing of dengue virus in clinical samples. Twenty-seven serum specimens from patients with dengue fever and dengue hemorrhagic fever in Colombia, Nicaragua, and Panama were directly subjected to RT-PCR for the detection of dengue virus. The resulting double-stranded DNA product was typed by a second round of PCR amplification (nested PCR) with type-specific primers, viral culture/indirect immunofluorescence (IIF), and enzyme-linked electroimmunoassay for IgM anti-dengue antibodies. The amplified virus genome was detected and typed within 8 hours. Nested RT-PCR, using viral culture and IIF as the gold standard, showed 100% sensitivity; 78% specificity; 69% positive predictive value, and 100% negative predictive value. It is noteworthy that two of the specimens whose results were positive with nested RT-PCR and negative with viral culture showed specific IgM antibodies. The results of the RT-PCR were in close agreement with those obtained through viral culture. This suggests PCR can greatly facilitate the rapid and early diagnosis of dengue infection.

Colombia↗

PubMed1998

[Advances in the development of a vaccine against dengue].

There are multiple factors responsible for the increase in the epidemics of dengue/dengue hemorrhagic fever in the tropical and subtropical regions. The characteristics of this disease, plus the absence of effective antiviral drugs; it is imperative to develop a vaccine against the dengue virus. To obtain such a vaccine, it is necessary an antigen that could bring protective immunity through life against the four dengue serotypes. Different strategies have been developed to obtain the vaccine, from the conventional one (attenuated virus), to the second and third generation of vaccines (immunisation with proteins and DNA respectively). The Envelope and the Membrane structural proteins, as well as the non structural NS1 and NS3, had been considered of major interest in the develop of the vaccine. In spite of the advances in this field, it is still much more research to be done, so it is considered that a vaccine against the dengue virus will not be available until the first decade of the next century.

Dengue↗

PubMed1997

Viral isolation from cases of epidemic neuropathy in Cuba.

OBJECTIVE: To investigate the possibility of a viral agent in the central nervous system of patients with epidemic neuropathy. DESIGN: Virus isolation attempts, in cell cultures and suckling mice, from cerebrospinal fluid (CSF) of neuropathy patients and controls undergoing lumbar puncture for unrelated reasons. Serologic studies in patients, contacts, and controls. SETTING: An epidemic of optic and peripheral neuropathy affected more than 50,000 people in Cuba in 1991 through 1993. Illness was associated with dietary limitations and increased physical demands accompanying the shortages of food and fuel experienced in Cuba since 1989. Most patients responded to parenteral vitamin therapy, and the epidemic began to subside when oral vitamin supplementation was begun for the entire Cuban population. RESULTS: Coxsackievirus A9 (five isolates) and a similar, less cytopathic virus (100 isolates) were recovered from 105 (84%) of 125 CSF specimens from neuropathy patients. The strains with light cytopathic effect were antigenically related to Coxsackieviruses A9 and B4 by cross-neutralization and immunoblotting assays. Virus persisted in CSF of some patients for 1 to 12 months. Cerebrospinal fluid from patients and both types of virus from cell culture produced illness, including complete posterior flaccid paralysis, in newborn mice, and virus was reisolated from the mice. Mouse tissues and sural nerve biopsy specimens from patients were stained by immunoperoxidase and colloidal gold techniques using hyperimmune rabbit antisera against the virus with light cytopathic effect. CONCLUSIONS: Coxsackievirus A9 or an antigenically related agent with a light cytopathic effect was present in CSF of 84% of 125 patients with epidemic neuropathy. The role of these agents, probably in combination with nutritional factors, in the pathophysiology of the disease requires further investigation.

Adult↗