RESUMO
Leaf number, dry weight, and nutrient composition of Lactuca sativa L. cv. Waldmann's Green leaves were compared following 9 days of treatment in a controlled environment room under various combinations of photosynthetic photon flux (PPF:350 vs 800 micromoles m-2 s-1), atmospheric CO2 level (ambient vs 1500 micromoles mol-1), and single-strength (1X:15 mM) vs double-strength (2X:30 mM) nitrogen (N) as NO3- alone or as NH4(+) + NO3- (1:5 molar ratio). CO2 enrichment greatly enhanced leaf number under all PPF and N conditions, but increased leaf dry weight only at high PPF. Conditions favoring high photosynthesis enhanced leaf starch content 3-fold, and protein content increased as much as 64% with 2X NH4(+)+NO3-. Free sugar content was 6 to 9% of leaf dry weight for all treatment combinations, while fat was 1.5 to 3.5%. Ash content varied from 15 to 20% of leaf dry weight. Modified controlled environments can be used to enhance the nutritional content as well as the yield of crops to be used for life support in space-deployed, self-sustaining human habitats. Leaf lettuce is a useful model crop for demonstrating the potential of nutritional value added by environmental manipulation.
Assuntos
Biomassa , Dióxido de Carbono/farmacologia , Ambiente Controlado , Lactuca/crescimento & desenvolvimento , Luz , Nitrogênio/farmacologia , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Lactuca/efeitos dos fármacos , Lactuca/efeitos da radiação , Nitratos/farmacologia , Valor Nutritivo , Fótons , Fotossíntese/efeitos dos fármacos , Fotossíntese/efeitos da radiação , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/efeitos da radiação , Compostos de Amônio Quaternário/farmacologiaRESUMO
Five greenhouse-grown cowpea [Vigna unguiculata (L.) Walp] cultivars were tested in a generalized random complete-block design to determine the effect of early leaf harvest on dry weight and protein concentration of plant parts at maturity. The most recent, fully expanded leaves on each branch from one group of plants were harvested at 5 and 7 weeks after planting. On the other groups of plants, no early leaf harvest was performed. Dry weight and protein concentration (dry weight basis) were determined for leaves, stems, and seeds at maturity and for leaves harvested early. Weight and protein concentration of seeds, leaves, and stems differed significantly between cultivars; protein concentration of leaves harvested at 5 or 7 weeks did not. Dry weight of leaves harvested at 5 vs. 7 weeks did not differ significantly, but leaf protein concentration was significantly higher at 5 weeks compared to 7 weeks. Across all cultivars, early leaf harvest had no significant effect on leaf or stem weight per plant at maturity. However, there was a significant decrease in seed weight when leaves were harvested early. Results suggest that even limited leaf harvest at 5 and 7 weeks has detrimental effects on yield, but not on protein concentration, of cowpea seeds harvested at maturity.
Assuntos
Sistemas Ecológicos Fechados , Fabaceae/crescimento & desenvolvimento , Sistemas de Manutenção da Vida , Folhas de Planta/crescimento & desenvolvimento , Proteínas de Plantas/metabolismo , Plantas Medicinais , Sementes/crescimento & desenvolvimento , Agricultura/métodos , Biomassa , Fabaceae/metabolismo , Valor Nutritivo , Tamanho do Órgão , Folhas de Planta/metabolismo , Caules de Planta/crescimento & desenvolvimento , Caules de Planta/metabolismo , Sementes/metabolismoAssuntos
Vírus da Dengue/imunologia , Dengue/prevenção & controle , Vírus da Encefalite Japonesa (Espécie)/imunologia , Encefalite Japonesa/prevenção & controle , Vacinas Virais , Animais , Variação Antigênica , Vírus da Dengue/genética , Vírus da Encefalite Japonesa (Espécie)/genética , Epitopos/imunologia , Variação Genética , Humanos , Imunidade Celular , Vacinas Atenuadas , Vacinas SintéticasRESUMO
Serum specimens collected during a prospective study of dengue infections among schoolchildren in Bangkok were tested for their ability to enhance dengue 2 (DEN-2) virus growth in human monocytes in vitro. Two groups of dengue-immune sera were compared: 32 dengue antibody positive serum specimens from children who subsequently developed asymptomatic secondary dengue infections; and 9 dengue antibody positive serum specimens from children who subsequently developed severe symptomatic secondary dengue infections, 8 of which were clinically diagnosed as dengue hemorrhagic fever. Antibody-dependent enhancement of virus growth was quantitated by measurement of virus yields in supernatant fluids of normal human monocyte cultures that were infected with DEN-2 virus in the presence of undiluted test serum. Only 4 of 32 (12%) preinfection sera from asymptomatic children, but 6 of 9 (67%) preinfection sera from symptomatic children, had significant enhancing activity (P less than 0.001). High serum DEN-2 antibody dependent enhancing activity is a significant (relative risk = 6.2) risk factor for severe illness among children in a dengue hemorrhagic fever endemic region. Dengue antibodies can be neutralizing and therefore protective, or they can be enhancing and increase the risk of dengue hemorrhagic fever.
Assuntos
Anticorpos Antivirais/fisiologia , Vírus da Dengue/crescimento & desenvolvimento , Dengue/etiologia , Monócitos/microbiologia , Células Cultivadas , Criança , Dengue/microbiologia , Humanos , Testes de Neutralização , Estudos Prospectivos , Fatores de Risco , Síndrome , Ensaio de Placa ViralRESUMO
The recently established virus family Flaviviridae contains at least 68 recognized members. Sixty-six of these viruses were tested by cross-neutralization in cell cultures. Flaviviruses were separated into eight complexes [tick-borne encephalitis (12 viruses), Rio Bravo (six), Japanese encephalitis (10), Tyuleniy (three), Ntaya (five), Uganda S (four), dengue (four) and Modoc (five)] containing 49 viruses; 17 other viruses were not sufficiently related to warrant inclusion in any of these complexes.
Assuntos
Antígenos Virais/análise , Flavivirus/classificação , Reações Cruzadas , Flavivirus/imunologia , Soros Imunes/imunologia , Testes de NeutralizaçãoAssuntos
Vírus da Dengue/imunologia , Dengue/prevenção & controle , Vacinas Virais , Animais , Vírus da Dengue/genética , Vírus da Dengue/patogenicidade , Epitopos/imunologia , Flavivirus/genética , Flavivirus/imunologia , Flavivirus/patogenicidade , Genes Virais , Vacinas Atenuadas , Vacinas Sintéticas , Proteínas Virais/imunologia , Organização Mundial da SaúdeRESUMO
Twenty-one dengue (DEN) viruses isolated from the Caribbean (Dominica and Jamaica) during the 1981-1982 epidemic year were distinct serological and genetic variants of DEN-4 virus. These isolates were clearly identified as DEN-4 viruses using type-specific monoclonal antibodies in indirect immunofluorescence assays. However, they either were not neutralized, or were neutralized poorly using hyperimmune mouse ascitic fluids (HMAF) or rhesus monkey serum directed against the H-241 prototype strain of DEN-4 virus isolated in the Philippines in 1956. HMAF prepared against a representative Caribbean isolate, however, neutralized with similar effectiveness the homologous virus, the H-241 prototype strain, and virus strains isolated from the Pacific and Southeast Asian areas from 1973 to 1984. The Caribbean isolate exhibited no more than 30% and 16% oligonucleotide spot homology with the H-241 and Bangkok viruses, respectively, by RNA fingerprint analysis, while demonstrating 82% and 89% homology with the Gilbert and Niue Island isolates, respectively. The isolation of dengue viruses which are serologically and genetically distinct from the prototype virus emphasizes the need for continued dengue virus surveillance. The recognition of unique dengue isolates should allow the selection of reference strains and vaccine candidate strains which will induce antibodies that are equally effective in neutralizing viruses from all geographic areas.
Assuntos
Vírus da Dengue/genética , Aedes/microbiologia , Animais , Anticorpos Monoclonais , Anticorpos Antivirais/imunologia , Anticorpos Antivirais/isolamento & purificação , Antígenos Virais/imunologia , Antígenos Virais/isolamento & purificação , Células Cultivadas , Vírus da Dengue/imunologia , Vírus da Dengue/isolamento & purificação , República Dominicana , Imunofluorescência , Humanos , Jamaica , Camundongos , Testes de Neutralização , RNA Viral/isolamento & purificação , Ensaio de Placa ViralRESUMO
The relative binding sites of dengue serotype-specific, dengue subcomplex-specific, dengue complex-specific, flavivirus subgroup-reactive, and flavivirus group-reactive monoclonal antibody preparations were identified by using competitive antibody binding assays. A dengue complex-specific epitope, capable of mediating infection enhancement, was identified on a 20,000 dalton protein found on intracellular virions. The other epitopes were assigned relative positions on the E glycoprotein by competitive antibody binding. These could be grouped into 3 linkage groups based on the ability of some monoclonal antibodies to block contiguous binding sites. Some antibodies were able to increase or "promote" the binding of antibodies from other linkage groups. These results suggest that a continuum of antigenic reactivities exist on the E glycoprotein of the dengue viruses, and that the conformation of this glycoprotein may be altered after antibody binding.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos Virais/imunologia , Vírus da Dengue/imunologia , Epitopos/imunologia , Aedes/citologia , Animais , Anticorpos Antivirais/imunologia , Ligação Competitiva , Células Cultivadas , Receptores Fc/imunologia , Vírion/imunologiaRESUMO
Peripheral blood mononuclear cells (PBMC) from humans without antibodies to dengue 2 virus lysed dengue 2 virus-infected Raji cells to a significantly greater degree than uninfected Raji cells. The addition of mouse anti-dengue antibody increased the lysis of dengue-infected Raji cells by PBMC. Dengue 2 immune human sera also increased lysis of dengue-infected Raji cells by PBMC. These results indicate that both PBMC-mediated cytotoxicity and antibody-dependent cell-mediated cytotoxicity (ADCC) can cause significant lysis of dengue-infected Raji cells. The lysis of infected Raji cells in the ADCC assay correlated with the dilution of dengue-specific antibody which was added, indicating the dengue virus specificity of the lysis of dengue virus-infected Raji cells. Alpha interferon (IFN alpha) was detected in the culture supernatant of PBMC and dengue-infected Raji cells. However, enhanced lysis of dengue-infected Raji cells by PBMC may not be due to the IFN produced, because neutralization of all IFN activity with anti-IFN alpha antibody did not decrease the lysis of dengue-infected cells, and effector cells pretreated with exogenous IFN alpha also lysed dengue-infected cells to a greater degree than uninfected cells. The effector cells responsible for lysis of dengue virus-infected Raji cells in the natural killer and ADCC assays were analyzed. Nonadherent PBMC caused more lysis than did adherent cells. Characterization of nonadherent cells with monoclonal antibodies showed that the predominant responsible effector cells were contained in OKM1+ and OKT3- fraction in the natural killer and ADCC assays.
Assuntos
Citotoxicidade Celular Dependente de Anticorpos , Citotoxicidade Imunológica , Vírus da Dengue/imunologia , Células Matadoras Naturais/imunologia , Animais , Anticorpos Monoclonais , Linfoma de Burkitt/imunologia , Linhagem Celular , Culicidae , Dactinomicina/farmacologia , Dengue/imunologia , Imunofluorescência , Humanos , Interferons/imunologiaRESUMO
Virion RNAs of 12 geographically distinct dengue type 1 (DEN-1) virus isolates were clearly unique by RNA fingerprinting. Isolates from the same geographic area were very similar but differed from those of other areas, allowing us to establish three geographical groupings based upon percent shared oligonucleotides. Three Caribbean strains were virtually identical (85-91% homologous oligonucleotides) whereas Pacific/S.E. Asian strains exhibited considerably less homology to one another (44-49%). The Pacific/S.E. Asian strains exhibited little relationship (20-30%) to the Caribbean and African strains. A Sri Lankan isolate displayed a relatively high degree of homology to Nigerian isolates (60-66% homologous oligonucleotides), suggesting that the Sri Lanka DEN-1 infection originated from Africa. A 1978 Nigerian DEN-1 isolate and the 1969 Sri Lankan strain each exhibited greater than 50% homology with a 1977 Jamaican strain. The similarities observed between the African/Sri Lankan and Jamaican strains suggest that the DEN-1 virus which caused the 1977 Jamaican epidemic may have originated from Africa or Sri Lanka. The RNA fingerprint is a unique characteristic of DEN-1 strains from a particular geographic region, suggesting this technique as a useful tool for dengue epidemiological investigations.
Assuntos
Vírus da Dengue/classificação , RNA Viral/análise , Vírus da Dengue/genética , Hibridização de Ácido Nucleico , Oligorribonucleotídeos/análiseRESUMO
Type-specific monoclonal antibodies prepared against the four dengue (DEN) virus serotypes were evaluated for their ability to identify low-passage human and mosquito isolates from Jamaica and West Africa by an indirect immunofluorescence assay. Serotyped human isolates from Jamaican dengue fever patients included 12 DEN-1, two DEN-2, and five DEN-4 viruses. Viruses from West Africa included 84 DEN-2 mosquito strains as well as two DEN-1 and one DEN-2 from humans. Results obtained using the immunofluorescence assay were consistent with virus identifications obtained using the more classical but costly and time-consuming plaque-reduction neutralization test. More viral isolates and higher virus yields were obtained using the C6/36 clone of Aedes albopictus cells rather than LLC-MK2 (monkey kidney) cells. Dengue type-specific monoclonal antibodies detected prototype viral antigens 24-48 hours postinfection in C6/36 cells. This is the first time that monoclonal antibodies have been used to serotype low-passage flavivirus isolates.
Assuntos
Anticorpos Monoclonais , Anticorpos Antivirais , Vírus da Dengue/classificação , Sorotipagem/métodos , Aedes , Animais , Antígenos Virais/análise , Linhagem Celular , Vírus da Dengue/imunologia , Imunofluorescência , HaplorrinosRESUMO
Colonized Aedes aegypti mosquitoes were fed on voluntary recipients of an experimental, live, attenuated, dengue type 2 (PR 159/S-1) vaccine to estimate the frequency of vector infection and the stability of the virus in mosquitoes. Two volunteers were viremic at the time of mosquito feeding, but only two of 114 mosquitoes that took a viremic blood meal became infected with the vaccine virus. Strains of virus recovered from the bodies of the mosquitoes and the volunteer's blood retained the temperature sensitivity and small plaque growth characteristics of the vaccine virus. Dengue viral antigen was not detectable in any of the mosquito heads by direct immunofluorescence and in vitro virus transmission by droplet feeding was not observed. This experiment showed that vector mosquitoes can be infected with vaccine virus by feeding on viremic vaccinees. Furthermore, the virus is sufficiently stable to retain the in vitro growth characteristics associated with the vaccine virus.
Assuntos
Aedes/parasitologia , Vírus da Dengue/imunologia , Insetos Vetores , Vacinas Virais/administração & dosagem , Animais , Dengue/imunologia , Dengue/microbiologia , Dengue/transmissão , Vírus da Dengue/crescimento & desenvolvimento , Feminino , Vacinas Atenuadas/administração & dosagem , Ensaio de Placa ViralRESUMO
Monoclonal antibodies produced against the four dengue virus serotypes identified four categories of reactions by immunofluorescence: flavivirus group reactive, dengue complex specific, dengue subcomplex specific (DEN-1, DEN-3), and dengue serotype specific. This is the first time that monospecific antibodies have been available for all of these unique antigenic determinants. Hybridoma cell lines producing dengue type-specific antibodies have been deposited in the Hybridoma Cell Bank of the American Type Culture Collection (Rockville, MD) for general distribution.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Vírus da Dengue/imunologia , Flavivirus/imunologia , Animais , Imunofluorescência , Humanos , Hibridomas/imunologia , Camundongos , Especificidade da EspécieRESUMO
Dengue type 2 virus replication was detected in the U-937 human monocyte cell line when the virus inoculum and the culture medium contained flavivirus antibodies diluted beyond their neutralizing titers. This was in marked contrast to yellow fever virus, which replicated very well in the absence of antibodies; however, 10-fold-higher yields of yellow fever virus could be obtained in the presence of flavivirus antibodies. These infection-enhancing antibodies were obtained from either a dengue type 2 human antiserum or reference hyperimmune obtained from either a dengue type 2 human antiserum or reference hyperimmune mouse ascitic fluid. The infection enhancement phenomenon, previously shown to be due to infection of Fc receptor-bearing cells with virus-antibody complexes, was completely blocked by preincubation of the cells with aggregated gamma globulin. The blocking results suggested an Fc receptor-mediated infection of the U-937 cells as well. A panel of monoclonal antibodies, previously characterized as either virus type specific or flavivirus cross-reactive and with mouse immunoglobulin subclasses G1 and G2a in both categories, were tested for their infection enhancement characteristics. A type-specific neutralizing monoclonal antibody preparation that was diluted beyond its neutralization titer did not cause infection enhancement, nor did low-level neutralizing monoclonal antibodies that were dengue serotype specific by the hemagglutination inhibition test. Only flavivirus cross-reactive monoclonal antibodies caused infection enhancement, irrespective of whether the immunoglobulins were G1 or G2a. These cross-reactive flavivirus determinants may reside at the tips of the glycoprotein projections on the virus particles, enabling the Fc ends of the cross-reactive antibodies attached to these determinants to interact with Fc receptors on susceptible cells.
Assuntos
Vírus da Dengue/imunologia , Flavivirus/imunologia , Monócitos/microbiologia , Anticorpos Monoclonais , Anticorpos Antivirais , Antígenos Virais , Linhagem Celular , Reações Cruzadas , Vírus da Dengue/crescimento & desenvolvimento , Humanos , Imunoglobulina G/imunologia , Receptores Virais/fisiologia , Replicação ViralRESUMO
Monoclonal antibodies directed against antigenic determinants of the New Guinea C strain of dengue-2 virus were obtained from lymphocyte hybridomas produced by fusing immune mouse lymphocytes with mouse myeloma cells. Hybridoma cell culture supernatants were screened by using a radioimmunoassay employing detergent-solubilized dengue-2 infected cell antigens. Monoclonal antibodies in ascitic fluids induced by 22 selected hybridomas were characterized by the hemagglutination-inhibition, plaque reduction neutralization, immunofluorescence, and complement-fixation tests. Both type-specific and broadly cross-reactive antibodies were observed, and immunoglobulin subclasses IgG1 and IgG2a were represented in both groups. At least three distinct antigenic determinants on the virion were defined using these antibodies. A single hybridoma produced antibody which recognized a dengue-2 virus type-specific determinant and exhibited high titered neutralization but had a low titer by hemagglutination inhibition. Four preparations reacted with a type-specific determinant and exhibited hemagglutination inhibition but did not neutralize. Seventeen hybridomas produced antibodies which were broadly cross reactive in all tests. Only two preparations reacted by complement fixation with dengue-2 antigens; both were cross reactive. Immunofluorescence specificity or cross reactivity correlated with neutralization and/or hemagglutination-inhibition. The dengue-2 virus type-specific antibody useful for identification of dengue-2 infected cells by immunofluorescence has been deposited in the Hybridoma Cell Bank of the American Type Culture Collection.
Assuntos
Antígenos Virais/imunologia , Vírus da Dengue/imunologia , Animais , Anticorpos Monoclonais , Anticorpos Antivirais , Reações Cruzadas , Vírus da Dengue/classificação , Epitopos , Hibridomas/imunologia , Imunoglobulina G/imunologia , Camundongos , SorotipagemRESUMO
Calves experimentally infected with Eimeria bovis and E zuernii were used in a controlled experiment to determine the anticoccidial activity of lasalocid sodium. Eleven-week-old Holstein calves were given an inoculum of 300,000 E bovis and 200,000 E zuernii oocysts; medication was initiated on the day of inoculation and continued for a 4-week period. The progress of parasitic infection was monitored with quantitative fecal oocyst examinations for the 6 weeks before calves were inoculated, and then in the 4-week treatment period and a 3-week observation period. The calves were given different doses of lasalocid sodium (0.5, 0.75, 1.0, and 3.0 mg/kg) and were compared with both nonmedicated inoculated calves and controls (nonmedicated, noninoculated calves). There were overall numerical reductions of oocysts produced in the medicated groups when compared with the nonmedicated inoculated controls.
Assuntos
Doenças dos Bovinos/tratamento farmacológico , Coccidiose/veterinária , Lasalocida/uso terapêutico , Animais , Bovinos , Coccidiose/tratamento farmacológico , Lasalocida/administração & dosagem , MasculinoRESUMO
Trypsin treatment of adherent human monocytes greatly reduced or eliminated the ability of these cells to support dengue virus replication. However, addition of dilute (nonneutralizing) antibody to the inoculum and the culture medium resulted in viral yields similar to those from monocytes not treated with trypsin. These results suggested that viral entry was facilitated by phagocytosis of immune complexes via Fc receptors on the monocytes. This concept was tested by (i) pretreating monocytes with aggregated gamma globulin, which resulted in a 40-fold reduction of viral yields after infection with dilute antibody-virus complexes and (ii) forming an immune complex with virus, antivirus F(ab')2 fragments, and rabbit anti-human Fab. Whereas F(ab')2 fragments alone would not enhance virus replication in trypsin-treated monocytes, the immune complex containing a rabbit Fc piece did increase the yield of dengue virus. These results suggest that dengue virus can infect a cultured monocyte in two ways: (i) through a viral receptor that is trypsin sensitive or (ii) through an Fc receptor that is not trypsin sensitive.
Assuntos
Complexo Antígeno-Anticorpo , Vírus da Dengue/crescimento & desenvolvimento , Monócitos/microbiologia , Receptores Fc/fisiologia , Receptores Virais/fisiologia , Anticorpos Antivirais , Células Cultivadas , Vírus da Dengue/imunologia , Humanos , Fragmentos Fab das Imunoglobulinas , Tripsina/farmacologia , Replicação Viral/efeitos dos fármacosRESUMO
Three passage levels of dengue-2 virus strain PR-159, obtained during the course of deriving the attenuated S-1 vaccine, were tested for their ability to replicate in subpopulations of human peripheral blood leukocytes: (i) 6th primary African green monkey kidney (PGMK) cell passage (parent virus); (ii) 19th PGMK cell passage of a small-plaque-forming clone derived from the parent virus (S-1 PGMK virus); and (iii) virus derived by four additional passages of the S-1 PGMK virus in diploid fetal rhesus lung cells (S-1 vaccine virus). Replication of these PR-159 viruses and another strain of dengue-2 virus adapted to Raji cells (16681-Raji virus) was measured in adherent and nonadherent mononuclear cells. All viruses except the S-1 PGMK virus replicated in monocytes. Occasional replication of the S-1 PGMK virus was associated with reversion to parent virus. The addition to the monocyte cultures of low concentrations of homologous dengue-2 antibody or non-neutralizing heterologous antibody increased the yield of the parent virus as much as 400-fold. This phenomenon of immune enhancement usually enabled the S-1 PGMK virus to replicate slowly in monocytes, but the progeny virus produced large plaques similar to the parent virus. Replication of the S-1 vaccine virus in cultured monocytes did not result in the appearance of large plaques. We could not recover S-1 vaccine virus from monocytes harvested from infected volunteers in the same manner that monocytes from natural human infections yield wild virus. The three passage levels of PR-159 virus were tested for replication in lymphocytes in comparison with the 16681-Raji virus. Only the 16681-Raji virus replicated in human lymphocytes cultured with or without enhancing antibody.
Assuntos
Vírus da Dengue/crescimento & desenvolvimento , Monócitos/microbiologia , Cultura de Vírus , Replicação Viral , Animais , Anticorpos Antivirais , Linfoma de Burkitt , Linhagem Celular , Chlorocebus aethiops , Meios de Cultura , Vírus da Dengue/imunologia , Haplorrinos , Humanos , Técnicas In Vitro , Rim , Linfócitos/microbiologia , Vacinas Virais , Vírus do Nilo Ocidental/imunologiaRESUMO
Dimethyl sulfoxide (DMSO)incorporated into an agar overlay containing DEAE-dextran enhanced plaque formation in Vero cells by Naples sandfly fever virus passaged in mouse brain or Vero cell cultures. No plaques were visible when DMSO was used without the DEAE-dextran, some plaques were rarely visible (less than 0.5mm) when DEAE-dextran was used without the DMSO, and up to 10-fold more plaques were clearly visible (0.5-1.5 mm) when both chemicals were used. The combined enhancing effect of DMSO and DEAE-dextran was also observed with mouse brain passaged, but not Vero passaged Sicilian sandfly fever virus. Other Phlebotomus group viruses produced a bit plaques (3-5 mm) and did not require DMSO for plaque formation, although an increase in plaque clarity was obtained with DMSO for some of them. Plaque reduction neutralization tests were assayed successfully under agar containing DMSO. The alphavirus Sindbis produced slightly larger plaques under agar containing DMSO, but there was no effect on clarity or size of plaques produced by the flavivirus dengue-2.