Luis F. de Salles-Gomes

Human vaccinia-like virus outbreaks in São Paulo and Goiás States, Brazil: virus detection, isolation and identification

Since October 2001, the Adolfo Lutz Institute has been receiving vesicular fluids and scab specimens of patients from Paraíba Valley region in the São Paulo and Minas Gerais States and from São Patricio Valley, in the Goiás State. Epidemiological data suggested that the outbreaks were caused by Cowpox virus or Vaccinia virus. Most of the patients are dairy milkers that had vesiculo-pustular lesions on the hands, arms, forearms, and some of them, on the face. Virus particles with orthopoxvirus morphology were detected by direct electron microscopy (DEM) in samples of 49 (66.21%) patients of a total of 74 analyzed. Viruses were isolated in Vero cell culture and on chorioallantoic membrane (CAM) of embryonated chicken eggs. Among 21 samples submitted to PCR using primers for hemagglutinin (HA) gene, 19 were positive. Restriction digestion with TaqI resulted in four characteristic Vaccinia virus fragments. HA nucleotide sequences showed 99.9% similarity with Cantagalo virus, described as a strain of Vaccinia virus. The only difference observed was the substitution of one nucleotide in the position 616 leading to change in one amino acid of the protein in the position 206. The phylogenetic analysis showed that the isolates clustered together with Cantagalo virus, other Vaccinia strains and Rabbitpox virus.

RC-IAL cell line: sensitivity of rubella virus grow

OBJECTIVE The rapid growth of the rubella virus in RC-IAL2 with development of cytopathic effect, in response to rubella virus infection, is described. For purposes of comparison, the rubella virus RA-27/3 strain was titered simultaneously in the RC-IAL, Vero, SIRC and RK13 cell lines. METHODS Rubella virus RA-27/3 strain are inoculated in the RC-IAL cell line (rabbit Kidney, Institute Adolfo Lutz). Plates containing 1.5x10(5) cells/ml of RC-IAL line were inoculated with 0.1ml s RA-27/3 strain virus containing 1x 10(4)TCID50/0.1ml. A 25% cytopathic effect was observed after 48 hours and 100% after 96 hours. The results obtained were compared to those observed with the SIRC, Vero and RK13 cell lines. Rubella virus was detected by immunohistochemistry. RESULTS With the results, it was possible to conclude that the RC-IAL cell line is a very good substrate for culturing rubella virus. The cells inoculated with rubella virus were examined by phase contrast microscopy and showed the characteristic rounded, bipolar and multipolar cells. The CPE in RC-IAL was observed in the first 48 hours and the curve of the increased infectivity was practically the same as observed in other cell lines. CONCLUSIONS These findings are important since this is one the few cell lines described in the literature with a cytopathic effect. So it can be used for antigen preparation and serological testing for the diagnosis of specific rubella antibodies.

RC-IAL: linhagem celular contínua de rim de coelho - características e substrato para replicação de vírus

A rabbit kidney cell line RC-IAL, isolated in 1976 and at present at 150a passage, has had its characteristics analysed. The cells presented morphology similar to fibroblasts throughout their culture. The cellular growth proportion remained unaltered from its isolation, with a cloning efficiency of around 9%. The line showed growth dependent on anchorage and chromosomic analysis presented the modal number of the species with small variations to about one chromosome, to a total of about 50%. The lines species of origin was confirmed through indirect immunofluorescence reaction and susceptibility to some viruses with cytopathic effect was verified with vaccinia, cowpox, herpes simplex types 1 and 2 and rubella viruses. This cellular substract is free from contaminating agents, thus satisfying the conditions for its use in scientific work, especially that relating to public health.Uma linhagem celular de rim de coelho (denominada RC-IAL), que foi isolada em 1976, e que atualmente esta na 150a passagem, teve suas caracteristicas analisadas. As celulas apresentaram morfologia semelhante aos fibroblastos desde o inicio de seu cultivo. A proporcao de crescimento celular nao se alterou desde seu isolamento, com uma eficiencia de clonagem ao redor de 9%. A linhagem mostrou crescimento dependente de ancoragem, e a analise cromossomica apresentou o numero modal da especie com pequenas variacoes para mais ou menos um cromossomo, resultando uma somatoria de 50%. Sua especie de origem foi comprovada atraves da reacao de imunofluorescencia indireta e a susceptibilidade da linhagem a alguns virus, com demonstracao do efeito citopatico, foi verificada com os virus da vacinia, cowpox, herpes simples tipo 1 e 2 e da rubeola. Esse substrato celular esta livre de contaminantes, satisfazendo assim, as condicoes para seu uso em trabalhos cientificos, principalmente os relacionadas a saude publica.

Anomalous Clinical Pictures of Varicella.

THE clinical picture of varicella, like that of other infectious diseases, presents wide variation. Besides the known irregularities in rash distribution, anomalies of the prodromal period and of the exanthem have been described (Schamberg and Kolmer, 1928a; Ronaldson and Kelleher, 1938). In the present paper, the relevant features of five cases are presented and the apparent association of the anomaly with a previous skin condition is pointed out in three cases. All five cases were observed between 1954 and 1955 when no laboratory technique was available for identification of varicella virus or antibody. In three cases, fluid collected from vesicles was inoculated on the chick-embryo chorioallantois after treatment with a mixture of penicillin and streptomycin. Reinoculations of the original inocula and serial blind passages were made to eliminate the possibility of variola virus being the agent involved since this virus may sometimes fail to provoke typical lesions (MacCallum, I954). Besides, the failure to find chorioallantoic lesions supported the clinical diagnosis since varicella virus does not provoke lesions (Irons, Bohls, Cook and Murphy, I941; MacCallum, 1954). Biopsies of representative skin lesions were made in two cases and the histological findings were interpreted in the light of recent reviews of the pathology of viral pocks (Lever, 1949; McNair Scott, Blank, Coriell and Crouse, I950).

Observations on Vaccinial Eruptions and Differential Diagnosis of Generalized Vaccinia

IN spite of literature on generalized vaccinia, the differential diagnosis of this condition has received little attention while the clinical picture of eruptions following vaccination against smallpox continuously offers field for new findings. In the present paper the relevant characteristics of four cases of laboratory-confirmed vaccinial eruption are recorded and some reflections on the differential diagnosis of generalized vaccinia are made after observation of more than 50 cases.

CLINICAL AND SUBCLINICAL VARIOLA MINOR IN A WARD OUTBREAK.

INTRODUCTION Missing links in the chain of variola transmission are usually ascribed to unrecognized instances of clinically manifest infection or, less frequently, to instances of subclinical infection. Published virological studies of variola epidemics do not include a systematic search for subclinical infections, and in only two reports are isolated instances recorded. Verlinde & van Tongeren (1952) reported isolation of variola virus from the throat of a healthy contact of a patient with overt variola, a finding questioned by Dixon (1962) because of the long interval after contact. Rodrigues-da-Silva, Rabello & Angulo (1963) found significant complementfixation titres in six asymptomatic contacts of housemates or schoolmates with overt variola minor. The present paper describes an outbreak of variola minor occurring in a hospital ward and, particularly, a serological survey aimed at disclosing subclinical variola.

Generalized Vaccinia in Patients with Pemphigus Foliaceus

ALTHOUGH there are numerous reports of individual cases, only a few papers deal with general aspects of the clinical picture of generalized vaccinia (Tedder, I936; Clark, Seiler, Joe, Gammie, Tait and Jack, I944; Fries, Borne and Barnes, I948). During two outbreaks of generalized vaccinia occurring, after two mass vaccinations, in a hospital for patients with pemphigus foliaceus, some clinical observations were madefrom time to time while more clinical data could be collected in retrospect. Despite its limitations, the evidence so obtained is presented in this paper in view of the paucity of the pertinent literature.

Comparação de métodos para testar a citotoxicidade "in vitro" de materiais biocompatíveis

OBJECTIVE A comparison of the sensitivity of the agar diffusion method with that of extraction using cell-lines RC-IAL (fibroblastic of rabbit kidney) and HeLa (epithelial carcinoma cells from the cervix uteri of the human uterus), in the in vitro evaluation of materials of medical and hospital. MATERIALS AND METHODS Fifteen samples chosen at random, from among the already known positives and negatives in our stock, were tested and identified as cotton, form, latex, cellulose and acrylic. Besides the samples mentioned, many SDS (GIbco) concentrations were tested experimentally in RC-IAL and HeLa cell cultures. RESULTS Of the 50 samples tested, 44(88%) were positive by both methods. However, when the SDS were compared by using the two methods, positive results were noted in the concentrations of from 0.5 to 0.05 microgram/ml in the agar diffusion ans extraction methods. A cytotoxic effect was only noted in the concentrations of up to 0.25 microgram/ml. CONCLUSION When the SDS was used, differences favorable to the agar diffusion method were observed in the two cell lines, in two concentrations; that is, the sensitivity of this method was quantitatively greater on inspection than that of the extraction method, as well as being the simpler method to use.