Induction of the Phenylpropanoid Pathway by Acibenzolar-S-Methyl and Potassium Phosphite Increases Mango Resistance to Ceratocystis fimbriata Infection.

This study used both microscopic and biochemical analyses to investigate the possible defense responses induced by acibenzolar-S-methyl (ASM) and potassium phosphite (Phi) in mango plants inoculated with Ceratocystis fimbriata. Disease development was evaluated in the stems of inoculated mango plants and these were examined using fluorescence and light microscopy. High-performance liquid chromatography (HPLC) was used to quantify secondary metabolites in the stem sections. Spraying the plants with ASM and Phi reduced internal necrosis and disease development. The ASM and Phi induced many microscopic defense responses in the stem tissues against C. fimbriata infection. HPLC analysis revealed that the concentrations of two alkaloids (theobromine and 7-methylxanthine) and 10 phenolic compounds (catechin, epicatechin, epigallocatechin, gallic acid, myricetin, p-coumaric acid, p-hydroxybenzoic acid, phloridzin, sinapinic acid, and salicylhydroxamic acid) were higher in the stem tissues of plants sprayed with ASM or Phi than in inoculated control treatment. The concentrations of phenolic compounds were higher in the stem tissues of inoculated plants than in noninoculated plants, while the inverse was observed for alkaloids. Higher concentrations of secondary metabolites in the stem tissues were detected in the early stages of fungal infection, especially in plants treated with inducers. Taken together, the results from the present study clearly support the concept that the phenylpropanoid pathway in the stem tissues of mango plants infected by C. fimbriata can be induced by ASM and Phi.

The anti-oestrogen fulvestrant (ICI 182,780) reduces the androgen receptor expression, ERK1/2 phosphorylation and cell proliferation in the rat ventral prostate.

This study proposed to investigate further the role of oestrogens during pubertal growth of rat ventral prostate, by analysing the effect of anti-oestrogen fulvestrant (ICI 182,780) on the expression of androgen (AR) and oestrogen receptors (ESR1 and ESR2), mitogen-activated protein kinase (ERK1/2) phosphorylation, and expression of Ki-67, a biomarker for cell proliferation. Ventral prostates were obtained from 90-day-old rats treated once a week for 2 months with vehicle (control) or ICI 182,780 (10 mg/rat, s.c.). Transcripts for AR, ESR1 and ESR2 were evaluated by quantitative real-time polymerase chain reaction. Expression of AR, ESR1, ESR2, total and phospho-ERK1/2 was analysed by Western blot or immunofluorescence. Ki-67-positive cells and myosin heavy chain were detected by immunohistochemistry. Cylindrical epithelial cells slightly taller, epithelial dysplasia and an increase in smooth muscle layer were observed in the ventral prostate from ICI 182,780-treated rats. ICI 182,780 did not change the mRNA, but decreased the protein levels for AR in the ventral prostate. The expression of ESR1 (mRNA and protein) was upregulated by ICI 182,780, but no changes were observed on ESR2 expression (mRNA and protein). ICI 182,780 decreased the phosphorylation state of ERK1/2, with no changes in total ERK1/2 levels. Ki-67-positive cells in the ventral prostate were also decreased by ICI 182,780. In conclusion, ICI 182,780 induces downregulation of AR expression and may block the translocation of ESR1 and ESR2 from the nucleus to the plasma membrane, decreasing ERK1/2 phosphorylation and prostatic epithelial cell proliferation. These findings provide a basis for physiological roles of oestrogen in the ventral prostate. Further studies with fulvestrant are necessary in benign prostate hyperplasia and prostatic cancer models.

Genetic diversity, anti-microbial resistance, plasmid profile and frequency of the Vi antigen in Salmonella Dublin strains isolated in Brazil.

Salmonella Dublin is strongly adapted to cattle causing enteritis and/or systemic disease with high rates of mortality. However, it can be sporadically isolated from humans, usually causing serious disease, especially in patients with underlying chronic diseases. The aim of this study was to molecularly type S. Dublin strains isolated from humans and animals in Brazil to verify the diversity of these strains as well as to ascertain possible differences between strains isolated from humans and animals. Moreover, the presence of the capsular antigen Vi and the plasmid profile was characterized in addition to the anti-microbial resistance against 15 drugs. For this reason, 113 S. Dublin strains isolated between 1983 and 2016 from humans (83) and animals (30) in Brazil were typed by PFGE and MLVA. The presence of the capsular antigen Vi was verified by PCR, and the phenotypic expression of the capsular antigen was determined serologically. Also, a plasmid analysis for each strain was carried out. The strains studied were divided into 35 different PFGE types and 89 MLVA-types with a similarity of ≥80% and ≥17.5%, respectively. The plasmid sizes found ranged from 2 to >150 kb and none of the strains studied presented the capsular antigen Vi. Resistance or intermediate resistance was found in 23 strains (20.3%) that were resistant to ampicillin, ciprofloxacin, chloramphenicol, imipenem, nalidixic acid, piperacillin, streptomycin and/or tetracycline. The majority of the S. Dublin strains studied and isolated over a 33-year period may descend from a common subtype that has been contaminating humans and animals in Brazil and able to cause invasive disease even in the absence of the capsular antigen. The higher diversity of resistance phenotypes in human isolates, as compared with animal strains, may be a reflection of the different anti-microbial treatments used to control S. Dublin infections in humans in Brazil.

Evaluation of motility enrichment on modified semi-solid Rappaport-Vassiladis medium (MSRV) for the detection of Salmonella in foods.

The detection and identification of Salmonella spp. is still troublesome and time consuming to the food industry. Employing the modified semi-solid Rappaport-Vassiliadis medium (MSRV), presumptive results for Salmonella can be obtained in 48 h, representing an interesting alternative to the standard methods. The specificity and sensitivity of the MSRV method were evaluated in this research. The efficiency of this method was also compared with the methodology recommended by the US Food and Drug Administration (FDA) using bismuth sulfite agar, XLT4 agar and Rambach agar. A total of 146 food samples comprised of 41 chicken thighs, 35 Brazilian fresh pork sausages, 35 samples of cocoa powder and/or granulated cocoa and 35 samples of grated fresh coconut, were examined. Overall, the rapid method (direct + indirect) and the standard culture detected 96.1% and 84.6% of the positive samples, respectively. No Salmonella was detected in the coconut or cocoa samples by any of the methods. Eighteen (43.9%) chicken thigh samples were contaminated with the microorganism. The rapid method (direct + indirect) and the standard culture detected 94.4% and 88.9% of these, respectively. Salmonella was detected in eight (22.8%) fresh pork sausage samples. The MSRV method detected Salmonella in all eight samples, while the standard gave positive results in six (75%). When compared with the standard method, the indirect method showed 86.4% sensitivity and 96.8% specificity, while the direct MSRV showed a sensitivity of 71.4% and specificity of 99.2%. Combined, both MSRV methods showed 95.5% sensitivity and 96.8% specificity. The MSRV medium also reduces the time necessary for the isolation of Salmonella from foods.

Prevalence and dissemination of Salmonella serotypes along the slaughtering process in Brazilian small poultry slaughterhouses.

Salmonella is the leading cause of human foodborne infections in Latin America, and poultry meat is one of the main vehicles. Small poultry slaughterhouses (fewer than 200 birds slaughtered per day) represent an important economic activity in certain regions. The slaughtering process in these abattoirs is manual and rudimentary, and frequently the hygienic conditions are poor. This study reports results of a detailed evaluation of the prevalence of Salmonella serotypes in carcasses, utensils, and environmental samples collected in 60 small Brazilian slaughterhouses. In the second step of the study, one of these slaughterhouses was selected to monitor the dissemination of Salmonella along the slaughtering process. For testing, conventional procedures were used: preenrichment in buffered peptone water (35 degrees C for 24 h), selective enrichment in Selenite-cystine (35 degrees C for 24 h), tetrathionate and Rappaport-Vassiliadis broths (42 degrees C for 24 h), plating on bismuth-sulfite and brilliant green agars (35 degrees C for 24 h), proper biochemical testing, and complete serotyping. Forty-one percent of samples harbored Salmonella spp., including 42% of carcasses, 23.1% of utensils, 71.4% of water, and 71.4% of freezers and refrigerators. Seventeen serotypes were detected. Salmonella Enteritidis predominated (30%), followed by Salmonella Albany (12%), Salmonella Hadar (12%), Salmonella Indiana (10%), and I 4,12:z:- (8%). All samples collected along the slaughtering process in the selected slaughterhouse were Salmonella positive. Five serotypes were detected, including Salmonella Albany, Salmonella Hadar, Salmonella Agona, Salmonella Emek, and Salmonella Indiana. More than 30% of the samples contained more than one serotype, and 12.5% presented three serotypes. The widespread occurrence of Salmonella in small slaughterhouses reinforces the need for implementation of effective control measures.

Dunging gutters filled with fresh water in finishing barns had no effect on the prevalence of Salmonella enterica on Brazilian swine farms.

Our aim was to assess the importance of dunging gutters filled with water in finishing barns for the prevalence of pigs shedding Salmonella enterica. Some finishing barns in Brazil are provided with a dunging-gutter system which consists of a continuous water flow at the back of solid-floored adjacent pens. Because there is transfer of faecal material between adjacent pens by water in this system and the faecal-oral route of transmission is so important for enteric pathogens, we tested the hypothesis that the presence of this kind of dunging-gutter system in finishing barns affects the prevalence of slaughter-age pigs shedding salmonella organisms in their faeces. The cross-sectional study was conducted on six farms each having barns with and barns without a dunging-gutter system. Breeding, management, nutritional and seasonal factors were similar in both barns on each farm. The two systems did not differ in prevalence of pigs shedding salmonella organisms. Five S. enterica serotypes were isolated: S. Agona, S. Javiana, S. Rissen, S. Sandiego and S. Senftenberg.

Progression of Salmonella enteritidis phage type 4 strains in São Paulo State, Brazil.

A total of 574 S. Enteritidis strains (383 from human sources and 191 from non-human sources) isolated between 1975-95, in São Paulo State, Brazil, were phagetyped. Among the strains isolated during the period of 1975-92, 80.9% of them belonged to phage type 8 (PT-8), but in 1993 strains of PT-4 accounted for 65.2% of all the S. Enteritidis isolates. In the following years, PT-4 strains accounted for 99.7% and 98.4% of phagetyped S. Enteritidis strains. The results obtained suggested that the current epidemic of S. Enteritidis in São Paulo State is clearly associated with the progression of PT-4 strains.

Changing patterns of Salmonella serovars: increase of Salmonella enteritidis in São Paulo, Brazil.

Serovars of a total of 5,490 Salmonella strains isolated during the period of 1991-95, from human infections (2,254 strains) and from non-human materials (3,236 strains) were evaluated. In the studied period, 81 different serovars were determined among human isolates. Salmonella Enteritidis corresponded to 1.2% in 1991, 2% in 1992, 10.1% in 1993, 43.3% in 1994, and 64.9% in 1995 of all isolates. A significant rise on the isolation of this serovar was seen since 1993 linked to food poisoning outbreaks. It is reported also an increase on the isolation of S. Enteritidis from blood cultures, associated mainly with patients with immunodeficiency syndrome. S. Enteritidis was prevalent among one hundred and thirty different serovars isolated from non-human sources. Increasing number of isolation of this serovar was seen from shell eggs, breeding flocks and from environmental samples. It is also reported a contamination of commercial feed stuffs by S. Enteritidis which represents a major concern for Brazilian poultry industry.

The role of public health laboratory in the problem of salmonellosis in São Paulo, Brazil.

From 1950 to 1990 a total of 45,862 strains (31,517 isolates from human sources, and 14,345 of non-human origin) were identified at Instituto Adolfo Lutz. No prevalence of any serovars was seen during the period 1950-66 among human sources isolates. Important changing pattern was seen in 1968, when S. Typhimurium surprisingly increased becoming the prevalent serovar in the following decades. During the period of 1970-76, S. Typhimurium represented 77.7% of all serovar of human origin. Significant rise in S. Agona isolation as well as in the number of different serovars among human sources strains were seen in the late 70' and the 80's. More than one hundred different serovars were identified among non-human origin strains. Among serovars isolated from human sources, 74.9%, 15.5%, and 3.7% were recovered from stool, blood, and cerebrospinal fluid cultures, respectively. The outbreak of meningitis by S. Grumpensis in the 60's, emphasizes the concept that any Salmonella serovars can be a cause of epidemics, mainly of the nosocomial origin. This evaluation covering a long period shows the important role of the Public Health Laboratory in the surveillance of salmonellosis, one of the most frequent zoonosis in the world.

[Epidemiologic markers of Salmonella typhimurium and Salmonella agona]. / Marcadores epidemiológicos de Salmonella typhimurium e Salmonella agona.

Among S. typhimurium and S. agona strains isolated during the period from 1971 to 1987, the biotypes, colicine types and resistance patterns were determined for 734 S. typhimurium and 631 S. agona strains. Among 734 S. typhimurium strains 65 biotypes were disclosed with prevalence of biotypes 1a (28.34%), 1b (29.84%) and 9 bi (18.28). Concerning S. agona, the biotype 1a represented by 87.16%, was the commonest clone among our strains. Although colicine typing added little information to characterize these serotypes, it should be usefull when applied in epidemiological study of outbreaks. It was observed multiply antimicrobial resistance mainly among human strains, particularly from nosocomial origins.

Etiology of acute diarrhea among children in Ribeirão Preto-SP, Brazil.

To study the main enteropathogens causing diarrhea in the region of Ribeirão Preto regarding serogroups and serotypes, the feces of 1836 children under 10 years old, from both sexes, attack of acute gastroenteritis, were analysed during a period of 4 years in Adolfo Lutz Institute - Ribeirão Preto, SP. The pathogens identified by standard methods were the following: Escherichia coli, Salmonella spp., Shigella spp., Campylobacter spp., Yersinia spp., and Cryptosporidium spp. Positive samples were 22.8% (419) with 1.7% association of pathogens. Larger isolates were mainly from children 0 to 11 months old. Enteropathogenic E. coli (EPEC) was most frequent (8.7%) with predominance of serogroup O119 (40.2%), followed by Shigella (6.2%), 63.6% of which S. sonnei.

[Food poisoning outbreak caused by Salmonella Enteritidis in the northwest of São Paulo State, Brazil]. / Surto alimentar por Salmonella Enteritidis no Noroeste do Estado de São Paulo, Brasil.

A foodborne outbreak which affected 211 persons occurred, in a School, in 1993. The epidemiological data obtained by interviewing the affected and non affected persons sampled showed as predominant symptoms: diarrhoea, fever (77.7%), abdominal cramps (67.7%), vomiting (65.8%), hot-and-cold sensations (54.5%) and headache (44.5%). The median incubation period was of 17 hours, the limits being 3 and 29 hours. The disease period was of from 3 to 4 days. The food concerned was a kind of paté, a mayonnaise mixture prepared with fresh eggs with boiled potatoes that was consumed with bread. The biological material analysis-3 coprocultures, and leftovers of the food revealed the presence of one and the same organism: Salmonella Enteritidis. In the food, the numbers of this bacterium per gram were sufficient to account for the manifestation of the disease (10(4) and 10(5)g). The antibiogram of all th isolates showed the same sensibility pattern. The preparation related to this outbreak suggests the endogenous contamination of the eggs; the cross contamination-the outbreak affected three school periods, as the food was prepared separately for each school period; and the conditions under which the food was kept during the time from preparation to consumption. The observation of the 3 food handlers, by successive coprocultures, for one week, indicates that they were not asymptomatic carriers nor were they affected as a result of this outbreak by the causal bacteria.

[Food borne disease outbreaks caused by Salmonella enteritidis]. / Surtos de enfermidades transmitidas por alimentos causados por Salmonella enteritidis.

OBJECTIVE: It is to describe outbreaks of salmonellosis reported from July 1993 through June 1997 in the Northwest region of S. Paulo State, Brazil, one of the areas where several foodborne outbreaks of salmonellosis have been recently detected. METHOD: Data of 19 epidemiological investigations were analysed; 87 stool specimens and 38 food samples (including 12 of shell eggs) were processed for microbiological analysis. Salmonella strains were identified by serotyping, phagetyping and antimicrobial susceptibility testing. RESULTS: There were 906 ill persons including 295 hospitalized patients. Phage type 4 (PT 4) Salmonella Enteritidis strains were isolated from 80.5% of stool samples, from all food samples and from 41.7% of eggs. Of the outbreaks, 95.7% were associated with the consumption of food containing raw or undercooked eggs. All strains were susceptible to the 13 antimicrobials, except the strains from the nosocomial outbreak. CONCLUSIONS: The results obtained show the need for the implementation of control measures regarding egg and storage, as well as for guidance to the public as to the risks involved in the consumption of inadequately prepared eggs.

Effects of the oestrogen receptor antagonist Fulvestrant on expression of genes that affect organization of the epididymal epithelium.

The role of oestrogens in epididymal function is still unclear. Knockout of the oestrogen receptor ESR1 (Esr1(-/-) ) or treatment with the anti-oestrogen Fulvestrant affect epididymal milieu and sperm motility. We investigated the effect of in vivo treatment of rats with Fulvestrant on: (i) expression of genes that may be important for the architecture and function of the epididymal epithelium: prominins 1 and 2, metalloproteinase 7, claudin 7, beta-catenin and cadherin 13, and (ii) levels of oestradiol and testosterone, and expression of oestrogen and androgen receptors, in the initial segment (IS), caput, corpus and cauda epididymis. Fulvestrant (i) reduced gene expression of prominin 1 (variant 1) in the caput, reduced prominin 1 protein content in the caput epididymis and in the efferent ductules, and increased the localization of prominin 1 in microvilli of the caput and corpus; (ii) reduced gene expression of prominin 2 in the corpus and cauda epididymis; (iii) increased the metalloproteinase 7 content in the apical region of principal cells from IS/caput; (iv) reduced in the corpus epididymis, but increased in the efferent ductules, the cadherin 13 mRNA level; (v) reduced testosterone but increased oestradiol levels in the corpus and cauda; (vi) increased the androgen receptor protein content in all regions of the epididymis, and the oestrogen receptor GPER in the corpus and cauda epididymis. In conclusion, treatment with Fulvestrant induced regional-specific changes in hormonal and steroid receptor content, and affected expression of proteins important for epithelial organization and absorption/secretion. The mechanisms of oestrogen action may differ among epididymal regions, which may contribute to determine region-specific sperm functions.

Experimental varicocoele in rats affects mechanisms that control expression and function of the androgen receptor.

Varicocoele is an important cause of male infertility. Normal male reproductive function and fertility depends on a delicate balance between androgen receptor (AR) and the classic oestrogen receptors ESR1 (ERα) and ESR2 (ERß). Using a model of surgically induced varicocoele in rats, this study aimed to investigate the effects of varicocoele on the expression of AR, ESR1, ESR2 and G-protein coupled oestrogen receptor (GPER). Varicocoele did not affect the mRNA and protein expression of ESR1 and ESR2 in both testes. Varicocoele did not affect the mRNA and protein expression of GPER in the right testis, but slightly reduced the mRNA and increased the protein levels in the left testis. Varicocoele did not affect the mRNA for AR, but reduced the protein levels in both testes. A proteomic approach was used in an attempt to find differentially expressed targets with possible correlation with AR downregulation. Varicocoele caused the differential expression of 29 proteins. Six proteins were upregulated, including the receptor for activated C kinase 1 (RACK1), and 23 were downregulated, including dihydrolipoamide dehydrogenase, alpha-enolase and pyrophosphatase 1. Western blot analysis confirmed that varicocoele upregulated the expression of RACK1, a protein involved with tyrosine phosphorylation and regulation of AR transcriptional activity, AR metabolism and dynamics of the blood-testis barrier. In conclusion, this study suggests that varicocoele affects mechanisms that control AR expression and function. This regulation of AR may play an important role in the varicocoele-induced testicular dysfunction. Furthermore, varicocoele downregulates several other proteins in the testis that may be useful markers of spermatozoa function and male infertility.

Characterization and immunogenicity of meningococcal group C conjugate vaccine prepared using hydrazide-activated tetanus toxoid.

The steps to produce, purify and control an immunogenic Brazilian conjugate vaccine against group C meningococcus (MenCPS-TT) using hydrazide-activated tetanus toxoid were developed. The conjugation methodology reduced the reaction time easily allowing scale-up. One freeze-dried pilot vaccine lot purified by tangential filtration, showed satisfactory quality control results including safety and stability. The pilot vaccine was immunogenic in mice in a dose-dependent fashion generating a 10-20-fold rise in IgG response in mice. The vaccine also induced high bactericidal titers. Vaccine concentrations of 1 and 0.1 microg showed higher avidity indices, suggesting induction of immunologic memory. These results support initiation of Phase I clinical studies with the MenCPS-TT conjugate vaccine.

Changing patterns of Salmonella serovars: increase of Salmonella enteritidis in Sao Paulo, Brazil

Foram avaliados os sorotipos de 5.490 cepas de Salmonella isolados no periodo, 1991-95, de infeccoes humanas (2.254 cepas) e de materiais de origem nao humana (3.236 cepas) bem como o perfil de sensibilidade aos agentes antimicrobianos de 131 cepas de S. enteritidis (92 de origem humana e 39 de origem nao humana). No periodo estudado, foram determinados 81 diferentes sorotipos. S. Enteritidis correspondeu a 1,2 por cento em 1991, 2 por cento em 1992, 10,1 por cento em 1993, 43,3 por cento em 1994 e 64,9 por cento em 1995. Um aumento significativo no isolamento de S. enteritidis foi verificado em 1993 associado a ocorrencia de surtos de enfermidades transmitidas por alimentos. E relatado tambem o aumento deste sorotipo a partir de hemoculturas, principalmente daquelas oriundas de pacientes com sindrome de imunodeficiencia. S. enteritidis foi tambem o sorotipo prevalente em materiais de origem nao humana, particularmente em ovos, aves (matrizes) e em amostras do meio ambiente. Ressalta-se a importancia da contaminacao, das materias primas, componentes de racao de aves, pela S. enteritidis, o que representa um preocupante problema para a avicultura brasileira

Progression of Salmonella enteritidis phage type 4 strains in São Paulo State, Brazil

A total of 574 S. Enteritidis strains (383 from human sources and 191 from non-human sources) isolated between 1975-95, in São Paulo State, Brazil, were phagetyped. Among the strains isolated during the period of 1975-92, 80.9 of them belonged to phage type 8 (PT-8), but in 1993 strains of PT-4 accounted for 65.2 of all the S. Enteritidis isolates. In the following years, PT-4 strains accounted for 99.7 and 98.4 of phagetyped S. Enteritidis strains. The results obtained suggested that the current epidemic of S. Enteritidis in São Paulo State is clearly associated with the progression of PT-4 strains.

The role of Public Health Laboratory in the problem of salmonellosis in Sao Paulo, Brazil

No periodo de 1950-90 foram identificadas 45.862 cepas de Salmonella, sendo 31.517 provenientes de infeccoes humanas e 14.345 de materiais de origem nao humana. O objetivo deste trabalho foi analisar as alteracoes ocorridas quanto a frequencia dos sorotipos isolados neste periodo. No periodo 1950-66, nao houve predominio evidente de nenhum sorotipo; entretanto, no perido 1970-76, com inicio em 1968, a S. Typhimurium passou a ser o sorotipo predominante, representando 77,7 por cento dos sorotipos isolados. Observou-se um aumento significativo da S. Agona, bem como de uma grande variedade de sorotipos. Quanto as salmonelas de origem nao humana, chama a atencao o grande numero (mais de 100) de sorotipos...

Surto alimentar por Salmonella Enteritidis no Noroeste do Estado de Säo Paulo, Brasil / Outbreak of Salmonella Enteritidis in northwest of S. Paulo State, Brazil

Em 1993 ocorreu um surto alimentar em escola, com 211 afetados. Os dados epidemiológicos levantados por entrevista de amostragem de afetados e näo afetados mostraram que os sintomas predominantes foram diarréia, febre (77,7 porcento), dor abdominal (67,7 porcento) e vômito (65,8 porcento), calafrios (54,5 porcento) e cefaléia (44,5 porcento). A mediana de incubaçäo foi de 17 horas, com limites entre 3 e 29 horas. A duraçäo da doença foi de 3 a 4 dias. O alimento consumido foi um tipo de patê, mistura de molho de maionese preparada com ovos crus com batata cozida, passado em päo. A análise de material biológico (3 coproculturas) e de restos de alimentos, revelou a presença do mesmo microorganismo, a Salmonella Enteritidis. No caso dos alimentos, o núnero encontrado desta bactéria por gramo de produto era compatível com a quantidade de células necessária para desencadear a doença (10*4 e 10*5/g). O antibiograma de todas as cepas isoladas revelou o mesmo padräo de sensibilidade. As falhas no preparo de alimento relacionadas com o levantamento indicam a possibilidade de contaminaçäo endógena dos ovos; contaminaçäo cruzada - o surto afetou três períodos escolares, sendo que para cada um o alimento foi preparado em separado - e as condiçöes de manutençäo do alimento após preparo e até o consumo. A observaçäo por uma semana seguida das 3 merendeiras envolvidas, através de coprocultura, näo indicaram que as mesmas fossem portadoras assintomáticas desta bactéria ou que tivessem sido envolvidas no surto em questäo