Predictors of mortality in critically ill patients with COVID-19 and diabetes

The COVID-19 pandemic has challenged the entire world, and patients with diabetes mellitus (DM) have been particularly affected. We aimed to evaluate predictors of mortality during the first 30 days of hospitalization in critically ill patients with COVID-19 and comorbid DM. This prospective study included 110 critically ill patients admitted with COVID-19 infection. Thirty-two (29%) patients had a previous diagnosis of DM. Clinical variables, laboratory tests, and vascular biomarkers, such as VCAM-1, syndecan-1, ICAM-1, angiopoietin-1, and angiopoeitin-2, were evaluated after intensive care unit (ICU) admission. A comparison was made between patients with and without DM. No difference in mortality was observed between the groups (48.7 vs 46.9%, P=0.861). In the multivariate Cox regression analysis, VCAM-1 levels at ICU admission (HR: 1 [1-1.001], P<0.006) were associated with death in patients with DM. Among patients with DM, advanced age (HR 1.063 [1.031-1.096], P<0.001), increased Ang-2/Ang-1 ratio (HR: 4.515 [1.803-11.308] P=0.001), and need for dialysis (HR: 3.489 [1.409-8.642], P=0.007) were independent predictors of death. Higher levels of VCAM-1 in patients with DM was better at predicting death of patients with severe COVID-19 and comorbid DM, and their cut-off values were useful for stratifying patients with a worse prognosis. Vascular biomarkers VCAM-1 and Ang-2/Ang-1 ratio were predictors of death in patients with severe COVID-19 and comorbid DM and those without DM. Additionally, kidney injury was associated with an increased risk of death.

Carbohydrate antigen 125 predicts pulmonary congestion in patients with ST-segment elevation myocardial infarction

Carbohydrate antigen 125 (CA125) has long been used as an ovarian cancer biomarker. However, because it is not specific for ovarian cells, CA125 could also be used to monitor congestion and inflammation in heart disease. Acute heart failure (HF) is used to identify patients with a worse prognosis in ST-segment elevation myocardial infarction (STEMI). We aimed to determine the association of CA125 with acute HF in STEMI and to compare CA125 with N-terminal pro brain natriuretic peptide (NTproBNP) with a cross-sectional study. At admission, patients were examined to define Killip class and then underwent coronary angioplasty. Blood samples, preferably taken in the hemodynamic ward, were centrifuged (1500 g for 15 min at ambient temperature) and stored at −80°C until biomarker assays were performed. Patients were divided into two groups according to the presence or absence of congestion. Patients in Killip class ≥II were in the congestion group and those with Killip <II in the absence of congestion group. We evaluated 231 patients. The mean age was 63.3 years. HF at admission was identified in 17.7% of patients. CA125 and NTproBNP levels were higher in patients with Killip class ≥II than those with Killip class <II (8.03 vs 9.17, P=0.016 and 772.45 vs 1925, P=0.007, respectively). The area under the receiver operator characteristic curve was 0.60 (95%CI 0.53−0.66, P=0.024) for CA125 and 0.63 (95%CI 0.56−0.69, P=0.001) for NTproBNP. There was no statistical difference between the curves (P=0.69). CA125 has similar use to NTproBNP in identifying acute HF in patients presenting with STEMI.

First record of the Lesser Snouted Treefrog Scinax nasicus (Cope, 1862) in Brazilian coast and new species records for the state of Rio Grande do Sul / Primeiro registro da perereca-de-banheiro Scinax nasicus (Cope, 1862) na costa do Brasil e novos registros para o estado do Rio Grande do Sul

Abstract Herein, we provide new occurrence records of Scinax nasicus (Cope, 1862) for the state of Rio Grande do Sul, Southern Brazil. All new records here provide are located on Southern half of the state. Besides this, we provide the first record for species in Brazilian coastal zone. Those records improve considerably our knowledge regarding species distribution in Southern Brazil.

Resumo Aqui, nós fornecemos novos registros de ocorrência de Scinax nasicus (Cope, 1862) para o estado do Rio Grande do Sul, Sul do Brasil. Todos os novos registros aqui fornecidos estão localizados na metade sul do estado. Além disso, nós fornecemos o primeiro registro para a espécie na zona costeira brasileira. Esses registros melhoram consideravelmente o nosso conhecimento sobre a distribuição da espécie no Sul do Brasil.

Evaluación de los efectos del LED en el tejido dérmico de ratones / The effect of application of red LED on the dermis of rats

Diodos Emisores de la Luz- Light Emitting Diodes (LED) son fuentes de luz que están siendo introducidos en los tratamientos del envejecimiento. El objetivo del estudio fue evaluar los efectos del LED rojo en la dermis de ratones wistar. Se trata de un estudio experimental controlado y randomizado, fueron utilizados 20 ratones wistar divididos en cinco grupos. Los grupos fueron: G1 en donde se aplicó 10 sesiones de diez minutos de fototerapia por LED. El G2 con 20 sesiones de 10minutos, el G3 con 10 sesiones de 20 minutos, el G4 con 20 sesiones de 20 minutos y el G5 fue el grupo control. Después de las aplicaciones se recolectó el material biológico de la zona, siendo realizado el análisis histológico de las áreas que recibieron las aplicaciones, sometiéndolas a las coloraciones de HE, tricómico de Masson y Verhoeef. Fue observada la presencia de micro vasos dérmicos con resultados significativos entre el G2 y el G5. Con relación a la presencia de mastocitos hubo resultados significativos entre el G1 y el G3. El análisis de las fibras elásticas presentó resultados significativos en los grupos G1, G2, G3, G4 en comparación al grupo control (G5). La evaluación de los efectos del LED en las fibras colágenos, los grupos tratados comparados al grupo control no presentaron ninguna diferencia significativa.

The Light-Emitting Diodes Light Emitting Diodes (LED) are a source of light wich are being introduced into the treatment of aging process. This study evaluate the effect of application of red LED on the dermis of Wistar rats. Twenty Wistar rats were used in this randomized controlled experimental study. Those rats were divided randomly into five groups. The G1 group underwent ten sessions of phototherapy by LED lasting ten minutes each. The G-2, 20 sessions of 10 minutes, the G-3, 20 minutes, the fourth group, 20 sessions of 20 minutes and the G-5 group was the control group. After the application, the biological material was collected and the histological analysis of the areas that suffered application were done, being subjected to HE staining, and Masson trichrome & Verhoeff. It was observed the presence of dermal microvascular, with significant results among G2 and G5 groups. Regarding the presence of mast cells, significant results were observed among G1 and G3 groups, however, that was not the case with the G5. By analyzing the elastic fibers, significant results were observed among G1, G2, G3, G4 groups, but the same results weren’t observed in the fifth group. Regarding the assessment of the effects of LED on the collagen fibers, the treated groups, when compared to the control group, showed no significant difference.

Estudo in vitro da atividade do extrato etanólico de sementes de bacupari (Rheedia gardneriana Planch. & Triana) e das frações no crescimento de Streptococcus mutans / In vitro study of the activity of ethanol extract from bacupari (Rheedia gardneriana Planch. & Triana) seeds and its fractions on Streptococcus mutans growth

Streptococcus mutans, principal microrganismo da cavidade oral, desempenha papel preponderante na formação de placas dentárias, sendo considerado o agente etiológico primário da cárie. Rheedia gardneriana, conhecida popularmente como bacupari, é uma planta utilizada com fins medicinais para o tratamento de diversas patologias, e por apresentar atividade antimicrobiana de compostos das folhas contra bactérias Gram-positivas e Gram-negativas. O objetivo do presente trabalho foi avaliar o efeito de extrato de semente de R. gardneriana sobre a cepa S. mutans UA159. Os testes foram conduzidos com o extrato etanólico bruto e as frações obtidas com os solventes diclorometano, etanol-água, metanol e hexano, em ensaios de inibição in vitro. O extrato bruto (100 por cento) apresentou halos de inibição com diâmetro similar ao obtido com solução de digluconato de clorexidina 0,12 por cento, usada como controle. Os ensaios com a fração diclorometano exibiram atividade inibitória 35 por cento menor comparado com o controle, enquanto nenhum efeito antimicrobiano foi observado com a fração etanol-água. Contrariamente, os resultados obtidos com as frações hexânica e metanólica demonstraram claramente a atividade antimicrobiana por inibição do crescimento bacteriano. Na fração metanólica a formação de halos de inibição foi similar ao do controle. Estes dados apresentam atividade antimicrobiana de R. gardneriana contra S. mutans.

Streptococcus mutans, which is the main microorganism of the oral cavity, plays a preponderant role in dental plaque formation and is considered the primary etiologic agent regarding caries. Commonly known as "bacupari", Rheedia gardneriana is a plant used for medicinal purposes in the treatment of several pathologies; besides, its leaves have compounds that present antimicrobial activity against Gram-positive and Gram-negative bacteria. The aim of this work was to evaluate the effect of R. gardneriana seed extract on S. mutans strain UA159. The tests were carried out with crude ethanol extract and the fractions obtained with the solvents dichloromethane, ethanol-water, methanol, and hexane in in vitro inhibition assays. The crude extract (100 percent) presented inhibition halos with diameter similar to that obtained by using 0.12 percent chlorhexidine digluconate solution as control. Assays with the fraction dichloromethane showed an inhibitory activity 35 percent lower than that of the control, whereas no antimicrobial effect was observed with the ethanol-water fraction. Conversely, the results obtained with the fractions hexane and methanol clearly demonstrated antimicrobial activity by inhibiting the bacterial growth. In the methanol fraction, the formation of inhibition halos was similar to that in the control. These data present antimicrobial activity of R. gardneriana against S. mutans.

Investigación de metapneumovirus humano en pacientes hospitalizados: estudio multicéntrico / Research of human metapneumovirus in hospitalized patients: a multicenter study

Introducción: las infecciones respiratorias agudas bajas (IRAB) son la primer causa de hospitalización a lo largo del año. La etiología viral es la más frecuente. El Metapneumovirus humano (MNVh) ha sido vinculado a las IRAB con aspirado negativo para Virus respiratorio sincicial (VRS) y Adenovirus (AD). Objetivo: determinar la prevalencia, epidemiología, clínica y severidad de las infecciones por MNVh, e intentar detectar un patrón radiológico relacionado con el mismo. Métodos: se estudiaron niños entre 0 y 2 años de edad internados por IRAB en el Centro Hospitalario Pereira Rossell, Hospital Central de las Fuerzas Armadas, Hospital Policial, Hospital Británico y Asociación Española Primera de Socorros Mutuos en el período 1 de abril al 30 de noviembre de 2006. Los aspirados nasofaríngeos fueron analizados en la Sección Virología de la Facultad de Ciencias. Resultados: se estudiaron 185 pacientes, obteniéndose 17 resultados positivos para MNVh (9,2%), con 9 coinfecciones con VRS. La mayoría de los pacientes tenían 6 meses o menos de edad. Las manifestaciones clínicas principales fueron polipnea, tirajes y sibilancias. Los principales hallazgos radiológicos fueron infiltrado intersticial difuso e hiperinsuflación. Ningún paciente requirió internación en unidad de cuidados intensivos y no hubo casos fatales. Conclusiones: la prevalencia, características clínicas y evolutivas de las infecciones por MNVh no mostraron diferencias frente a las producidas por el VRS.

Brote epidémico de meningitis viral causado por echovirus tipo 4 en la provincia de Misiones / Outbreak of viral meningitis caused by echovirus type 4 in Misiones province

Se realizó un estudio retrospectivo a fin de describir un brote epidémico de meningitis causado por enterovirus, que comprometió a 143 niños de 1 mes a 14 años internados en el Hospital Pediátrico de Posadas (Misiones) con diagnóstico de meningitis aséptica, entre agosto y diciembre de 2005. Se observó un aumento de casos entre las semanas 33 a 50, con un pico máximo entre las semanas 47 y 48, lo que confirmó el brote. La mediana de edad de los niños afectados fue de 8 años y el 55,2% fueron varones. El 80% de los casos se observó entre escolares (5 a 14 años). El promedio del tiempo de internación fue de 4,5 ± 1,7 días, y no se registraron fallecidos. Los LCR se estudiaron mediante examen citoquímico y estudios bacteriológicos y virológicos (aislamiento viral, RT- PCR anidada e identificación molecular mediante secuenciación génica). Los recuentos de células en LCR variaron entre 6 y 5040 células /mm3, el 92% fueron inferiores a 500 células/mm3 y el 43,5% mostró predominio linfocitario. El 56% presentó concentraciones de glucosa normal, con proteínas ligeramente elevadas. El 28% de las muestras estudiadas por cultivo (17/60) mostró efecto citopático, compatible con enterovirus. La RT-PCR anidada permitió detectar enterovirus en un 73% de las muestras (43/59), con 6 casos que se tipificaron como echovirus tipo 4. El índice de positividad al combinar ambas técnicas alcanzó el 83%.

A descriptive retrospective study was carried out to describe an epidemic outbreak of enteroviral meningitis in Misiones. We reviewed records of 143 children from 1 month to 14 years of age who were hospitalized with aseptic meningitis in the Pediatric Hospital of Posadas from August to December 2005. Increased number of cases was observed between weeks 33 to 50 which reached a maximum peak in weeks 47 and 48, confirming an outbreak. The median of age was 8 years old, 55.2% were males. Eighty percent of cases were in 5 to 14 years old children. The average length of time spent in the hospital was 4.5±1.7 days, no deaths were reported. We performed cell counts, chemical and bacterial studies of CSF, and culture or RT-Nested/PCR for enteroviruses. Isolates were serotyped by RT-PCR amplification and genetic sequencing. Cell counts were from 6 to 5040 cells/mm3. Ninety two percent had less than 500 cells/mm3 and 43.5% had lymphocyte predominance. Glucose levels were normal with slightly elevated protein counts in 56% of cases. Of the cultured samples, 28% (17/60) showed cytopathic effect compatible with enterovirus. RT-n-PCR detected enterovirus in 73% (43/59) of the analyzed CSF. Echovirus type 4 was identified in 6 of them. The positive indicator obtained by combining both techniques was 83% (58/70).

Análise da confiabilidade de um método de mensuração do ângulo de pico de torque ativo dos isquiossurais / Analysis of the reliability of a method for measuring hamstring active peak torque angle

INTRODUÇÃO: O comprimento muscular pode ser inferido através da determinação da relação comprimento-tensão do músculo. Essa relação é tradicionalmente investigada por meio da medida do torque máximo produzido pelo músculo e do ângulo em que esse torque é gerado. OBJETIVO: O presente estudo verificou a confiabilidade teste-reteste de um método de mensuração do ângulo de pico de torque ativo dos isquiossurais em jovens saudáveis. MÉTODO: Vinte e cinco indivíduos saudáveis (22,88 ± 1,67 anos) foram avaliados duas vezes em um intervalo de três semanas. Um dinamômetro isocinético foi utilizado no modo passivo para avaliar o torque passivo dos isquiossurais. A atividade muscular foi monitorada para garantir silêncio eletromio-gráfico. O dinamômetro foi utilizado no modo concêntrico para determinar o torque total dos isquiossurais. O torque ativo foi obtido subtraindo-se o torque passivo do torque total. O ângulo do pico de torque ativo foi utilizado para a análise. RESULTADO: Não houve diferença estatisticamente significativa entre as duas medidas realizadas (t= 1,009; p= 0,323). O Coeficiente de Correlação Intraclasse para os valores obtidos do ângulo de pico de torque ativo foi de 0,948 (p= 0,0001; IC 95 por cento 0,881 - 0,977). CONCLUSÃO: O presente estudo demonstrou que o método descrito é confiável para a quantificação deste ângulo, sugerindo que este método pode ser utilizado para avaliar mudanças da curva torque-ângulo promovidas por alterações de comprimento muscular.

INTRODUCTION: Muscle length can be inferred from the length-tension relationship of the muscle. This relationship is traditionally investigated by measuring the peak torque produced by the muscle and the angle at which it is generated. OBJECTIVE: The present study investigated the test-retest reliability of a method for measuring hamstring active peak torque angle in healthy young adults. METHOD: Twenty-five healthy individuals (22.88 ± 1.67 years) were assessed twice with an interval of three weeks. An isokinetic dynamometer was used in passive mode to assess hamstring passive torque. Muscle activity was monitored to ensure electromyographic silence. The dynamometer was also used in concentric mode to determine hamstring total torque. The active torque was obtained as the difference between total torque and passive torque. The active peak torque angle was used for the analysis. RESULTS: There was no significant difference between the two measurements (t= 1.009; p= 0.323). The intraclass correlation coefficient for the active peak torque angle values obtained was 0.948 (p= 0.0001; 95 percent CI: 0.883 - 0.977). CONCLUSION: This study has shown that the method described is reliable for the quantification of active peak torque angle, thus suggesting that this method can be used to evaluate shifts in the torque-angle curve produced by muscle length changes.

Wild dengue virus types 1, 2 and 3 viremia in rhesus monkeys

Among the flaviviruses, dengue, with its four serotypes, has spread throughout the tropics. The most advanced vaccines developed so far include live attenuated viruses, which have been tested in humans but none has been licensed. Preclinical testing of dengue vaccine candidates is performed initially in mice and in nonhuman primates. In the latter the main criteria used to assay protection are neutralizing antibodies elicited by the vaccine candidate and the magnitude and duration of peripheral viremia upon challenge of previously immunized animals. Towards the identification of wild-type viruses that could be used in challenge experiments a total of 31 rhesus monkeys were inoculated subcutaneously of wild dengue types 1, 2, and 3 viruses. The viremia caused by the different viruses was variable but it was possible to identify dengue viruses useful as challenge strains.

Attenuation and immunogenicity of recombinant yellow fever 17D-dengue type 2 virus for rhesus monkeys

A chimeric yellow fever (YF)-dengue serotype 2 (dengue 2) virus was constructed by replacing the premembrane and envelope genes of the YF 17D virus with those from dengue 2 virus strains of Southeast Asian genotype. The virus grew to high titers in Vero cells and, after passage 2, was used for immunogenicity and attenuation studies in rhesus monkeys. Subcutaneous immunization of naive rhesus monkeys with the 17D-D2 chimeric virus induced a neutralizing antibody response associated with the protection of 6 of 7 monkeys against viremia by wild-type dengue 2 virus. Neutralizing antibody titers to dengue 2 were significantly lower in YF-immune animals than in YF-naive monkeys and protection against challenge with wild-type dengue 2 virus was observed in only 2 of 11 YF-immune monkeys. An anamnestic response to dengue 2, indicated by a sharp increase of neutralizing antibody titers, was observed in the majority of the monkeys after challenge with wild-type virus. Virus attenuation was demonstrated using the standard monkey neurovirulence test. The 17D-D2 chimera caused significantly fewer histological lesions than the YF 17DD virus. The attenuated phenotype could also be inferred from the limited viremias compared to the YF 17DD vaccine. Overall, these results provide further support for the use of chimeric viruses for the development of a new live tetravalent dengue vaccine.

Infección por virus BK en paciente pediátrico trasplantado renal / [BK virus infection in a pediatric renal transplant recipient]

BK Human Polyomavirus causes an asymptomatic primary infection in children, then establishing latency mainly in the urinary tratt. Viral reactivation can lead to renal pathology in individuals with impaired cellular immune response. This is particularly important in pediatric transplant recipients, who can suffer a primary infection when immunosupressed. We followed up the case of a 5 years old patient who received a renal transplant in October 2003, and presented damaged graft 45 days after the intervention. The patient suffered 3 episodes of renal function failure between October 2003 and June 2004. Blood, urine, renal biopsy and lymphocele liquid samples were analyzed. A differential diagnosis between acute rejection and infectious causes was established by testing for BK, CMV and ADV viruses, and the cytological study of renal tissue. Laboratory findings together with clinical signs suggest the patient was infected by BK virus. As a final consideration, the great importance of differentiating between acute rejection and BK infection is emphasized, since immunosuppressant management is opposite in each case.

Pico de flujo espiatorio en niños uruguayos sin enfermedad de 3 a 13 años / Peak espiatory flow in Uruguayan children without disease, from 3 to 13 years old

El pico de flujo espiratorio (PFE) ha sido incluido en las recomendaciones de consenso sobre control y tratamiento del asma desde la década de los 90. Dado que presenta variaciones según diferentes poblaciones, se determinaron los valores de PFE de niños uruguayos sin enfermedad. Se estudiaron 362 varones y 437 niñas procedentes de nueve escuelas, con edades de 3 a 13 años cumplidos, sin antecedentes respiratorios ni utilización de medicación antiasmática en su historia previa, y que al momento del estudio no evidenciaran anormalidades del examen clínico del aparato respiratorio. Los PFE fueron determinados mediante flujómetros Vitalograph con los niños en posición de pie y se registró el mayor valor logrado en tres o más pruebas sucesivas, sin oclusión nasal. Se estudiaron los niños agrupados por sexo y en intervalos de 5 kg, talla de 10 cm y edad en intervalos de un año. Se determinó el valor percentilar 10, 50 y 90 para cada intervalo de clase y se correlacionó mediante una regresión de cuadrados mínimos con un polinomio de 2º grado. El coeficiente de correlación (r²) para los valores de p.10, p.50 y p.90 fue de 0,99, 0,98 y de 1,00 para talla, siendo levemente inferiores para edad y peso. Los valores de PFE son mayores en los varones con respecto a las niñas a partir de un peso de 40 kg, una edad de 9 años y una talla de 140 cm. Se comparan los valores obtenidos en nuestro estudio con los publicados en la literatura.

Avaliação isocinetica da função muscular do quadril e do tornozelo em idosos que sofrem quedas / Isokinetic evaluation of hip and ankle muscle function among elderly people who suffer falls

Analisar o impacto da funcao muscular dos membros inferiores sobre as quedas em uma populacao de idosos. Metodos: os participantes foram 30 idosos, 14 que nao haviam sofrido quedas e 16 que ja haviam sofrido quedas no ultimo 6 meses, selecionados aleatoriamente no ambulatorio de geriatria de um hospital universitario. Todos foram submetidos a avaliacao demografica e clinica e ao teste de funcao muscular no Dinamometro Isocientico Biodex. Os avaliadores nao foram informados sobre o grupo a que pertencia cada idoso ate o final do estudo. Foram feitas analises estatisticas descritivas para todas as variaveis e para a comparacao entre os grupos foram utilizados o test t-Student, Mann-whitney, qui-quadrado ou teste exato de fisher, no nivel de significancia < 0,05. Resultados Em relacao a funcao muscular do tornozelo, os idosos que ja cairam apresentaram menor potencia media (p variado entre 0,005 e 0,001), menor trabalho propocional ao peso corporal (p variado entre 0,046 e 0,028) e o menor pico de torque propocional ao peso corporal( p=0.23). Para a articulacao do quadril nao houve diferencas estatisticamente significativas entre os grupos em nenhuma das variaveis testadas.Conclusoes: idosos que ja cairam apresentaram menores valores de pico de torque, trabalho proporcional ao peso corporal e potencia media para a articulacao de tornozelo em relacao aos que nao cairam. Nao houve diferencas estatisticamente significatyivas para a funcao muscular do quadril. Nossos achados mostram que na abordagem fisioterapeutica do idoso e necessario incluir rxercicios de fortalecimento para o tornozelo, contribuindo para a prevencao de quedas

Enfermedad neurologica por adenovirus / Neurologic disease due to adenovirus infection

El objetivo de este trabajo fue determinar la prevalencia de adenovirus (ADV) en las infecciones del sistema nervioso central (SNC). Se analizaron 108 muestras de líquido cefalorraquídeo (LCR) provenientes de 79 casos de encefalitis, 7 meningitis y 22 de otras patologías neurológicas, recibidas en el período 2000-2002. Cuarenta y nueve (47.35%) se obtuvieron de pacientes inmunocomprometidos. La presencia de ADV se investigó mediante reacción en cadena de la polimerasa en formato anidado (Nested-PCR). La identificación del genogrupo se realizó mediante análisis filogenético de la secuencia nucleotídica parcial de la región que codifica para la proteína del hexón. Se detectó la presencia de ADV en 6 de 108 (5.5%) muestras de LCR analizadas. Todos los casos positivos pertenecieron a pacientes con encefalitis que fueron 79, (6/79, 7.6%). No se observó diferencia estadísticamente significativa entre los casos de infección por ADV en pacientes inmunocomprometidos e inmunocompetentes (p>0.05). Las cepas de ADV detectadas se agruparon en los genogrupos B1 y C. En conclusión, nuestros resultados describen el rol de los ADV en las infecciones neurológicas en Argentina. La información presentada contribuye al conocimiento de su epidemiología, en particular en casos de encefalitis.

Estudio de sensibilidad antiviral de virus herpes simplex en pacientes trasplantados / [Antiviral sensitivity of herpes simplex virus in immunocompromised patients]

The Herpes simplex Virus (HSV) resistance to acyclovir (ACV) occurs in a 5

of the inmunocompromised patients, approximately. The treatment with analogs of nucleosides, causes the appearance of resistent HSV-ACV stocks (ACVr) which can be produced by alteration in genes coding for the TK or the DNA-polymerase. A previous large-scale clinical study on ACVr HSV strains isolated from patients infected with human immunodeficiency virus indicated that 96

of ACVr HSV mutants were low producers of, or deficient in, TK activity (TK-), with 4

being TK mutants with an altered substrate specificity. No DNA Pol mutants were isolated. The pirophosphate analogs generate resistance in the gene of DNA-polymerase by mutation. In this paper we show the methodology used for the determination of sensibilite profiles to ACV and Phoscarnet (PFA) in a population of inmunocompromised patients. We analized 46 HSV strain from vesicular injuries of transplanted patients. All samples, were inoculated in human fibroblasts and the HSV isolates were identified by inmunofluorescence whith monoclonal antibodies. These strains were amplified and the profile of susceptibility determinated in Vero cells, using 100 tissue culture inhibition dosis 50 (TCID50) of each Viral stock and the specific antiviral drugs in different concentrations. The cytopathic effect (CPE) was evaluated after 72hs. post infection. The 50

inhibitory concentration (CI50) was calculated from the percentage of inhibition of the ECP based on the concentration of the drug. From 46 isolations, 26 were HSV-1 and 20 were HSV-2. Two of them, one HSV-1 and one HSV-2, were resistant to ACV and none of the isolates were resistant to PFA.

Comparative sequence analysis of the P-, M- and L-coding region of the measles virus CAM-70 live attenuated vaccine strain

Measles virus is a highly contagious agent which causes a major health problem in developing countries. The viral genomic RNA is single-stranded, nonsegmented and of negative polarity. Many live attenuated vaccines for measles virus have been developed using either the prototype Edmonston strain or other locally isolated measles strains. Despite the diverse geographic origins of the vaccine viruses and the different attenuation methods used, there was remarkable sequence similarity of H, F and N genes among all vaccine strains. CAM-70 is a Japanese measles attenuated vaccine strain widely used in Brazilian children and produced by Bio-Manguinhos since 1982. Previous studies have characterized this vaccine biologically and genomically. Nevertheless, only the F, H and N genes have been sequenced. In the present study we have sequenced the remaining P, M and L genes (approximately 1.6, 1.4 and 6.5 kb, respectively) to complete the genomic characterization of CAM-70 and to assess the extent of genetic relationship between CAM-70 and other current vaccines. These genes were amplified using long-range or standard RT-PCR techniques, and the cDNA was cloned and automatically sequenced using the dideoxy chain-termination method. The sequence analysis comparing previously sequenced genotype A strains with the CAM-70 Bio-Manguinhos strain showed a low divergence among them. However, the CAM-70 strains (CAM-70 Bio-Manguinhos and a recently sequenced CAM-70 submaster seed strain) were assigned to a specific group by phylogenetic analysis using the neighbor-joining method. Information about our product at the genomic level is important for monitoring vaccination campaigns and for future studies of measles virus attenuation.

RT-nested PCR for the detection of enterovirus in biological samples from patients with suspected enteroviral infections

In this study, we have tested a reverse transcription (RT) nested polymerase chain reaction (nPCR) for detection of enterovirus (EV) RNA in cerebrospinal fluid (CSF), serum samples, and conjunctival swabs (CS) from patients with suspected enterovirus infections. A specific 113-bp fragment was amplified using primers designed based on 5' non coding region of the enterovirus genome. The enterovirus RT-nPCR was able to detect 0.001 plaque forming unit (pfu)/ml. Since no PCR product was detected in each of the CSF, CS and serum samples from patients with proven-non-enterovirus viral infections, this method was found to be specific. EV RNA was detected in all 30 culture-confirmed CSF samples and yielded positive results in 5 out of 7 additional cases of culture-negative CSF samples with other evidences of enterovirus infection. Overall, EV RNA was detected in 95 of the patients with clinical diagnosis of viral central nervous system (CNS) disease and confirmed enterovirus infection. Furthermore, we were able to detect EV RNA in 24 (47) out of 51 CSF samples from patients with clinical diagnosis of viral CNS disease and negative laboratory evidence of viral infection. The percentage of positive EV RNA detection in paired CSF and serum samples from 11 patients with an enterovirus isolate in CSF was 100 (11 of 11) and 73 (8 of 11), respectively. In addition, EV-specific IgM was detected in 64 (7 of 11) of the sera tested. The method was also tested against 136 samples of CS from patients with clinical diagnosis of acute hemorrhagic conjunctivitis. Ninety nine of them resulted positive (73), while only 27 (20) had been positive for viral culture. In summary, our study shows the importance of enterovirus RT-nPCR for the diagnosis of enterovirus associated disease in different kind of biological samples and different types of diseases.