Bacteriophage amplification assay for detection of Listeria spp. using virucidal laser treatment

A protocol for the bacteriophage amplification technique was developed for quantitative detection of viable Listeria monocytogenes cells using the A511 listeriophage with plaque formation as the end-point assay. Laser and toluidine blue O (TBO) were employed as selective virucidal treatment for destruction of exogenous bacteriophage. Laser and TBO can bring a total reduction in titer phage (ca. 10(8) pfu/mL) without affecting the viability of L. monocytogenes cells. Artificially inoculated skimmed milk revealed mean populations of the bacteria as low as between 13 cfu/mL (1.11 log cfu/mL), after a 10-h assay duration. Virucidal laser treatment demonstrated better protection of Listeria cells than the other agents previously tested. The protocol was faster and easier to perform than standard procedures. This protocol constitutes an alternative for rapid, sensitive and quantitative detection of L. monocytogenes.

A cólera no Estado do Rio Grande do Norte--Brasil--perfil sorológico e de sensibilidade do Vibrio cholerae a diferentes antimicrobianos / Cholera in Rio Grande do Norte State--Brazil: sorology and sensitivity of Vibrio cholerae to different antimicrobials

PURPOSE: The emergence of multiple resistance to antimicrobials in Vibrio cholerae isolated in the state of Ceará, Brazil, alerted researchers in this area to the sensitivity to antimicrobials of strains isolated in Rio Grande do Norte (RN), Brazil. METHODS: One hundred and four strains of V. cholerae of human origin, isolated by Laboratório Central de Saúde Pública Dr. Almino Fernandes, were serologically typified and evaluated for in vitro sensitivity to eight antibiotics belonging to different groups (polymyxine, tetracycline, chloramphenicol, nitrofurantoin, sulphazotrin, pefloxacine, erythromycine, ampicillin). The strains were collected from patients suspected of contracting choleric diarrhea in the year 1999, in Natal/RN/Brazil. RESULTS: From the sample total, 100 were identified as V. cholerae, serogroup O:1, biotype El Tor, with 99 (95.3%) belonging to serovar Ogawa and only 1 (0.9%) to serovar Inaba. The 4 remaining were characterized as non O:1 V. cholerae, with 3 (2.9%) biochemically identified as Heiberg type I and 1 (0.9%) as type II. All the V. cholerae serogroup O:1 strains were sensitive to tetracycline, chloramphenicol, sulphazotrin, pefloxacine, erythromycine and resistant to polymyxine. In relation to nitrofurantoin, only 1 was sensitive. Only 1 was resistant to ampicillin. The non O:1 V. cholerae strains were resistant to polymyxine. CONCLUSIONS: The results showed sensitivity in 100% of the V. cholerae serogroup O:1 strains to tetracycline, an elective drug in the treatment of cholera, and an absence of multiple resistant strains in our environment. An interesting finding was the frequency of serovar Ogawa in 1999, considering the greater incidence of serovar Inaba in other years of cholera outbreaks in RN.

Bone marrow fibroblasts in patients with advanced lung cancer

In a previous study we demonstrated that the incidence of fibroblast colony-forming units (CFU-F) was very low in bone marrow primary cultures from the majority of untreated advanced non-small lung cancer patients (LCP) compared to normal controls (NC). For this reason, we studied the ability of bone marrow stromal cells to achieve confluence in primary cultures and their proliferative capacity following four continuous subcultures in consecutive untreated LCP and NC. We also evaluated the production of interleukin-1ß (IL-1ß) and prostaglandin E2 (PGE2) by pure fibroblasts. Bone marrow was obtained from 20 LCP and 20 NC. A CFU-F assay was used to investigate the proliferative and confluence capacity. Levels of IL-1ß and PGE2 in conditioned medium (CM) of pure fibroblast cultures were measured with an ELISA kit and RIA kit, respectively. Only fibroblasts from 6/13 (46 percent) LCP confluent primary cultures had the capacity to proliferate following four subcultures (NC = 100 percent). Levels of spontaneously released IL-1ß were below 10 pg/ml in the CM of LCP, while NC had a mean value of 1,217 + or - 74 pg/ml. In contrast, levels of PGE2 in these CM of LCP were higher (77.5 + or - 23.6 pg/ml) compared to NC (18.5 + or - 0.9 pg/ml). In conclusion, bone marrow fibroblasts from LCP presented a defective proliferative and confluence capacity, and this deficiency may be associated with the alteration of IL-1ß and PGE2 production

Vibrio cholerae resistant to 2,4-diamino-6,7-diisopropylpteridine (O/129) isolated from patients with enteritis in Ceará, Brazil.

This paper reports the characterization of clinical Vibrio cholerae resistant to vibriostatic agent O/129, using classical and plasmid analysis. In a study conducted during December 1991-September 1993, two of 7,058 V. cholerae strains, obtained from patients suspected to have cholera in the State of Ceará, northeast Brazil, were resistant to 150 micrograms of the vibriostatic agent O/129 (2,4-diamino-6,7-diisopropylpteridine). One strain was identified as V. cholerae O1 El Tor Inaba and the other one as serogroup O22. Only one O1 strain harboured a plasmid of 147 kb transferable to Escherichia coli K12, and five strains of V. cholerae O1 and non-O1 were sensitive to O/129 and plasmid-negative at a frequency between 8 x 10(-2) and 3.6 x 10(-5). Additionally, O/129-resistant strains of V. cholerae O1 and O22 were resistant to trimethoprim/sulphamethoxazole.

Análise dos anticorpos vibriocidas e aglutinantes na população urbana do Município do Manacapura/AM (1992/1993) / Vibriocidal and agglutinating antibodies in the urban population in the municipality of Manacapuru/AM (1992-1993)

Foi realizado um estudo sorológico em 1.196 indivíduos residentes na área urbana do município de Manacapuru/AM, visando analisar o perfil dos anticorpos vibriocidas e aglutinantes. Empregou-se o procedimento de amostragem aleatória sistemática na obtenção da amostra populacional. Um ano após, obteve-se uma 2ª amostra de soro de 120 indivíduos (10% da amostra inicial), escolhidos aleatoriamente entre os participantes do inquérito, com o objetivo de avaliar o comportamento dos anticorpos nesse intervalo de tempo. Foram empregados os métodos de microtitulação de anticorpos vibriocidas e de soroaglutinação em tubos. A associação entre os anticorpos estudados foi determinada pelo coeficiente de correlação, calculado com base na distribuição de freqüência dos títulos detectados. A análise dos resultados revelou forte correlação positiva entre os anticorpos (r = 1,0) e queda nos títulos em grande proporção das amostras após um ano.

A serological study was carried out involving 1,196 individuals residents in the urban area of Manacapuru--Amazonas, to evaluate the behavior of vibriocidal and agglutinating antibodies. A systematic random sampling procedure was employed to obtain the sample. A year later a 2nd sample of serum was obtained from 120 individuals selected among the participants of the survey. Vibriocidal antibodies microtitulation and seroagglutination in tubes were employed. The correspondence between the studied antibodies was determined by the correlation coefficient calculated according to the frequency of the titles detected. The analysis of the results revealed positive correlation between the antibodies (r = 1.0) and a decrease in titles in a large proportion of the positive samples one year later.

Meningite por Listeria monocytogenes. Relato de casos em pacientes do Distrito Federal / Listeria monocytogenes meningitis. Case reports in patients from the Federal District

São descritos os primeiros casos de meningite por Listeria monocytogenes comprovados bacteriologicamente, em pacientes do Distrito Federal. Alguns comentários foram realizados com base nos achados laboratoriais, assim como, sobre certas peculiaridades clínico-epidemiológicas da listeriose humana.

It has been shown the role of Listeria monocytogenes as a etiological agent identified by bacteriological analysis among cases of human meningitis in Distrito Federal, Brazil. Laboratorial characteristics and some clinical and epidemiological aspects are reported.

Pesquisa de Vibrio cholerae O1 em amostras fecais da população urbana de Manacapuru, AM / Vibrio cholerae O1 in fecal samples from the urban population of Manacapuru, AM

O estudo foi realizado com o objetivo de identificar portadores assintomáticos do vibrião colérico em Manacapuru, AM, 1249 amostras fecais foram obtidas por swab retal e submetidas à análise bacteriológica. Vibrio cholerae O1 não foi detectado. Foram isolados e identificados: V. funissii em 12 (0,9%) amostras, V. fluvialis, em 4 (0,3%) e V. hollisae em 1 (0,1%).

The study was carried out to identify asymptomatic carriers of V. cholerae O1 in Manacapuru, AM. 1249 feces samples was obtained by rectal swab and cultivated. Had no growth of V. cholerae. On the other hand were isolated and identified: V. furnissii in 12 (0.9%) samples, V. fluvialis in 4 (0.3%) and V. hollisae in 1 (0.1%).

Evaluation of virulence factors in environmental isolates of Vibrio species

Strains of Vibrio parahaemolyticus, Vibrio fluvialis and Vibrio mimicus isolated from seafood and seawater were examined for characteristics related to infectivity, such as enzymatic activity and animal assays. All strains hydrolysed DNA, starch, gelatin and chitin. Variable results were obtained with the haemolysin, chondroitin, collagen, elastin and lecithin tests. Production of thermostable direct haemolysin by V. parahaemolyticus was detected in 7.1 per cent strains derived from seafood and 2 per cent from seawater. In the animal assays, strains of V. fluvialis showed positive results at skin PF (75 per cent ), mouse lethality (100 per cent ), but no fluid accumulation in the suckling mice model was noted. Concerning V. mimicus, results showed skin PF (100 per cent ), mouse lethality (100 per cent ) and fluid accumulation in suckling mice (66.6 per cent )

Manipuladores de alimentos em restaurantes de Belo Horizonte, portadores de Salmonella. / Food handlers in restaurants of Belo Horizonte, carriers of Salmonella

Atraves de coprocultura de 328 servidores de sete restaurantes de Belo Horizonte,foram detectados 65 portadores de Salmonella (19,82%). Cinquenta por cento deles eliminavam mais de um sorotipo por ocasiao da pesquisa. Houve relacao direta entre o estado de portador com o tempo de servico prestado em restaurante. Os meios de cultura utilizados para o isolamento foram: caldo tetrationato de Kauffmann;Rappaport, Rappaport-Hofer, agar EMB, agar Hektoen, meio de Costa-Vernin e os meios convencionais para os testes bioquimicos. As temperaturas de incubacao para os meios de enriquecimento e seletivo-indicadores foram de 37 graus C e 41 graus C. A frequencia dos sorotipos isolados foi a seguinte: S.newlands, 26,6%; S. derby, 25%; S. anatum, 25%; S. agona, 18,7%; S. typhimurium,14,1% S. newport, 12,5%; S. oranienburg, 10,9%; S. senftenberg, 7,8%; S. saint-paul, 4,7%; S. michigan, 4,7%; S. tshiongwe, 1,6%; S.morotai, 1,6%. Dos resultados obtidos verifica-se a necessidade de submeter os manipuladores de alimentos a um controle periodico, atraves da coprocultura, para que haja menor risco de contaminacao dos alimentos e consequentemente dos usuarios de restaurantes

Pesquisa de Listeria monocytogenes em solos. / Research of Listeria monocytogenes in soils

Foi investigada a presenca de L. monocytogenes em 20 amostras de solos da superficie, oriundas de diferentes areas ecologicas do Estado do Rio de Janeiro (perimetro urbano, pasto, mata e horta). A analise bacteriologica evidenciou 32 amostras do microrganismo, revelando maior incidencia no solo nao cultivado (mata) que representou 56,25% dos isolamentos, ficando em segundo plano a area urbana com 25%. No levantamento, destacam-se os sorotipos L1/2b (46,87%), L4b (25%) e L1/2a (18,75%) como os mais frequentes, embora incidindo de mameira heterogenea em relacao as areas analisadas. Entre os varios aspectos estudados, salienta-se que 50% das amostras se comportaram como patogenicas, em decorrencia da producao de ceratoconjuntivite em cobaia, detalhe este intimamente relacionado com a atividade hemolitica "in vitro". Confirma-se que tanto as formas virulenta e avirulenta de L. monocytogenes sobrevivem as condicoes impostas pelo solo, sendo provavelmente a causa primaria desta contaminacao os dejetos eliminados por diferentes especies de animais portadores, corroborada pela presenca de coliformes fecais nas areas analisadas

Salmonella typhi: lisotipia VI e biotipificaçäo em amostras oriundas de algumas regiöes do Brasil / Salmonella typhi: lysotype VI and biotyping in sample from some regions of Brazil

Fez-se uma análise da distribuicäo da frequência dos lisotipos VI e dos tipos fermentativos segundo o esquema de Kristensen, em 1.150 amostras de Salmonella typhi, isoladas de diferentes regiöes do Brasil (Pará, Pernambuco, Bahia, Minas Gerais, Rio de Janeiro, Säo Paulo e Rio Grande do Sul). No computo geral, observou-se a prevalência dos lisotipos A (38,1%); Ela (18,9%); amostras VI negativas (16,6%); D6 (8,7%) I + IV (4,6%); T (2,3%) e C1 (2,1%) e a ocorrência de alguns tipos fágicos característicos para determinadas áreas (B3, C4 e 40 na Bahia; E1b, F2, G1 e L1 em Säo Paulo; E4 e 28 no Rio de Janeiro). Quanto a classificacäo bioquímica, 55,2% das amostras caracterizaram-se no biotipo II (xilose e arabinose negativas), 44,2% no tipo fermentativo I (xilose positiva e arabinose negativas) e 0,52% no tipo III (xilose e arabinose positivas), respectivamente

Primeiro isolamento e identificacao de Vibrio parahaemolyticus no Brasil de infeccao gastrointestinal humana. / The first isolation and identification of Vibrio parahaemolyticus from human gastrointestinal infection in Brazil

Amostra de Vibrio parahaemolyticus sorotipo 05:k17 e Kanagawa positiva, foi isolada de fezes diarreicas de uma crianca de Cascavel, Ceara. Esta e a primeira referencia de isolamento de V. parahaemolyticus de origem humana no Brasil e, provavelmente da America do Sul

Drug resistance in salmonella strains isolated from raw sewage in Rio de Janeiro.

Amostras de Salmonella isoladas de esgotos do Rio de Janeiro foram examinadas para se verificar a presenca de especimes resistentes a drogas. Em contraste com um levantamento realizado em 1970, quando nao foram encontradas amostras resistentes, o presente trabalho revelou a ocorrencia de 31% (39/126) de amostras resistentes a pelo menos uma droga. Todas as linhagens multiresistentes foram capazes de transferir os marcadores respectivos para receptoras adequadas, Os isolados apresentaram o serotipo S. tryphimurium, con excecao de duas amostras, caracterizadas como S. agona. Duas dentre as 39 amostras resistentes e duas dentre 87 amostras sensiveis as drogas usadas eram colicinogenicas. Noventa por cento (29/32) dos determinantes transferiveis eram do tipo Fi