PATHOGENIC OR POTENTIALLY PATHOGENIC BACTERIA AS CONTAMINANTS OF FRESH-WATER FROM DIFFERENT SOURCES IN ARARAQUARA, BRAZIL

The microbiological quality of various fresh waters in the Araraquara region, state of São Paulo, Brazil was investigated. Ninety-nine water samples were taken from rivers, reservoirs, artesian and non-artesian wells, springs and tap waters, and these waters were analysed using: plate counts of heterotrophic microorganisms (per 1 ml); Most Probable Number (MPN) of fecal coliforms and E. coli (per 100 ml); tests for presence of Salmonella, Shigella, Yersinia, the E. coli pathogens of classes EPEC, ETEC and EIEC and Mycobacterium, Shigella, Yersinia and enteroinvasive E. coli (EIEC) were not isolated. The other types of microorganisms were isolated in varying proportions. We conclude that the waters investigated represent a potential microbiological health risk.

Use of Pyrosequencing and Denaturing Gradient Gel Electrophoresis to Examine the Effects of Probiotics and Essential Oil Blends on Digestive Microflora in Broilers Under Mixed Eimeria Infection

A protective digestive microflora helps prevent and reduce broiler infection and colonization by enteropathogens. In the current experiment, broilers fed diets supplemented with probiotics and essential oil (EO) blends were infected with a standard mixed Eimeria spp. to determine effects of performance enhancers on ileal and cecal microbial communities (MCs). Eight treatment groups included four controls (uninfected-unmedicated [UU], unmedicated-infected, the antibiotic BMD plus the ionophore Coban as positive control, and the ionophore as negative control), and four treatments (probiotics BC-30 and Calsporin; and EO, Crina Poultry Plus, and Crina PoultryAF). Day-old broilers were raised to 14 days in floor pens on used litter and then were moved to Petersime batteries and inoculated at 15 days with mixed Eimeria spp. Ileal and cecal samples were collected at 14 days and 7 days postinfection. Digesta DNA was subjected to pyrosequencing for sequencing of individual cecal bacteria and denaturing gradient gel electrophoresis (DGGE) for determination of changes in ileal and cecal MC according to percentage similarity coefficient (%SC). Pyrosequencing is very sensitive detecting shifts in individual bacterial sequences, whereas DGGE is able to detect gross shifts in entire MC. These combined techniques offer versatility toward identifying feed additive and mild Eimeria infection modulation of broiler MC. Pyrosequencing detected 147 bacterial species sequences. Additionally, pyrosequencing revealed the presence of relatively low levels of the potential human enteropathogens Campylobacter sp. and four Shigella spp. as well as the potential poultry pathogen Clostridiun perfringens. Pre- and postinfection changes in ileal (56%SC) and cecal (78.5%SC) DGGE profiles resulted from the coccidia infection and with increased broiler age. Probiotics and EO changed MC from those seen in UU ilea and ceca. Results potentially reflect the performance enhancement above expectations in comparison to broilers not given the probiotics or the specific EO blends as feed supplements.

CONDICOES BACTERIOLOGICAS DE LEITE CRU E DE LEITE PASTEURIZADO NA REGIAO DE ARARAQUARA, SP, BRASIL

Milk can be an important spreading vehicle of pathogenic agents mainly for young children who are an important group of milk consumers. 135 milk samples (77 of raw milk and 58 of pasteurized milk) were analysed in order to verify the number of heterotrophic bacteria, of Staphylococcus aureus and of total coliforms, as well as to determine presence of Salmonella, Shigella, enteropathogenic Escherichia coli (EPEC) and enteroinvasive Escherichia coli (EIEC). The results were negative for Salmonella, Shigella and EIEC. EPEC serotypes 0:28, 0:111 and 0:55 were isolated in 4 of raw milk samples. The heterotrophic bacteria was found in counts over 30.000 UFC/mL in 91%, 25% and 68,75% respectively of raw milk, milk grade B and grade C. Counts over 30.000 coliforms/mL were found in 70,13%, 6,25% and 6,24% of the raw milk, B and C, respectively. 32,40% of the raw milk had counts of S. aureus over 3.000 UFC/mL.

Microbiological quality of ice used to refrigerate foods

Ice used for human consumption or to refrigerate foods can be contaminated with pathogenic microorganisms and may become a vehicle for human infection. To evaluate the microbiological content of commercial ice and ice used to refrigerate fish and seafood, 60 ice samples collected at six different retail points in the city of Araraquara, SP, Brazil, were studied. The following parameters were determined: total plate counts (37° C and 4° C), most probable number (MPN) for total coliforms, fecal coliforms and Escherichia coli, presence of Salmonella spp., Shigella spp., Yersinia spp., E. coli, Vibrio cholerae and Aeromonas spp.. Results suggested poor hygienic conditions of ice production due to the presence of indicator micro-organisms. Fifty strains of E. coli of different serotypes, as well as one Y. enterocolitica biotype 1, serogroup 0:5, 27 and phage type Xz (Ye 1/05,27/Xz) and one Salmonella Enteritidis phage type 1 (PT1) were isolated. Aeromonas spp., Shigella spp. and V. cholerae were not detected. The presence of high numbers of coliforms, heterotrophic indicator micro-organisms and pathogenic strains suggested that commercial ice and ice used to refrigerate fish and seafood may rep resent a potential hazard to the consumer in our community. © 2002 Elsevier Science Ltd. All rights reserved.

Espectro de agentes etiológicos associados a surtos de doenças transmitidas por alimentos em núcleos receptores turísticos de três regiões geográficas do Estado de São Paulo

Foodborne and waterborne diseases are spread by the consumption of food or water contaminated with bacteria and/or their toxins, viruses, parasites or chemicals. The aim of the research reported here was to establish the spectrum of etiologic agents of foodborne outbreaks at 15 tourist resorts in three geographic regions of the State of Sao Paulo (Brazil). The study was based on the cases reported to the Epidemic Surveillance Center (CVE) of the Public Health Authority of the State of São Paulo (SES), from 2002 to 2005. The tourist centers were chosen at random in three regions of the state (Capital, Interior and Coast) and offered the following attractions: events, agribusiness, cultural history, shopping, town center, gastronomy, health and leisure, sun and sea. Among the bacteria, the results showed that Salmonella spp. were most frequently associated with outbreaks, followed by Shigella spp., enterotoxigenic Escherichia coli, Staphylococcus aureus, Clostridium perfringens, Bacillus cereus and Campylobacter spp. Viruses (Rotavirus and Hepatitis A) played a part in many of the cases, while the frequency of parasites and worm infestations was low in the foodborne disease outbreaks at these resorts. The mixed foods (rice, beans, liver, potatoes, barbecue, juice), fish and poultry were the three commonest vehicles implicated in the outbreaks.

Diarréia em bezerros da raça Nelore criados extensivamente: estudo clínico e etiológico

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Estudo da atividade antimicrobiana de algumas árvores medicinais nativas com potencial de conservação/recuperação de florestas tropicais

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Mastite bovina em rebanhos leiteiros no município de Parauapebas, mesorregião sudeste do estado do Pará

A mastite em sua forma subclinica é a responsável pelas maiores perdas de produção leiteira representando elevados prejuízos econômicos. Com o objetivo de estudar a etiologia da mastite subclinica bovina no município de Parauapebas-Pa, foram submetidas ao California Mastitis test. - CMT 174 (8,4%) vacas em sua maioria mestiças, aparentemente saudáveis de 15 propriedades leiteiras localizadas no referido município, situado na Mesorregião Sudeste do estado do Pará. Observou-se que 84 (48,33%) animais apresentaram resultados de +,++,+++ ao CMT. O leite de cada teto que reagiram ao CMT num total de 178 amostras foi analisado bacteriologicamente visando o isolamento e a identificação dos microorganismos, foram analisadas as características macroscópicas, microscópicas e bioquímicas das culturas isoladas. Do total de amostras foram isoladas 208 cepas de agentes microbianos em culturas puras ou em associações sendo todos provenientes de leite mamitico, das quais 141(67,79%) cepas eram cocos Gram positivos- S.aureus (29%) e S.epidermidis (19,14%) e 67 (32,21%) eram enterobactérias. Entre as enterobacterias destacaram-se Pseudomonas sp.com 12 (17,91%) cepas esisoladas, Citrobacter sp. com 12( 17,95%) cepas e Shigella sp. com 10(14,92%), Outras 15 cepas de enterobacterias que não foram identificadas. O isolamento dos agentes apresentou variação significativa, pois se consideraram as observações quanto ao manejo de ordenha dos animais estudados, as condições higiênicas sanitárias da obtenção do leite através da aplicação de um questionário visando a observação das seguintes variáveis: sistema de criação, manejo adotado na propriedade, higiene e nível de exposição, infecciosidade dos agentes isolados o que reafirma a complexidade da infecção na área de estudo e seu aspecto multifatorial, necessitando o investimento por parte dos órgãos fiscalizadores de melhores praticas de higiene e obtenção na atividade de exploração de leite.

Determinação qualitativa de enterobactérias presentes em tartarugas da Amazônia (podocnemis expansa) de vida livre e cativeiro

A tartaruga da amazônia (Podocnemis expansa) corresponde a um recurso faunístico muito importante para as populações ribeirinhas da região amazônica, além de ser uma das principais espécies indicadas para produção em cativeiro. O consumo dessa espécie como alimento na região, gerou uma demanda de estudos quanto à questão sanitária e seu impacto na saúde pública. O principal objetivo deste trabalho foi avaliar a microbiota intestinal de tartarugas da amazônia de vida livre e cativeiro, verificando a ocorrência de bactérias da Família Enterobacteriaceae no trato intestinal desses animais. Para isso, foram utilizadas 116 tartarugas adultas, de ambos os sexos, sendo que, 51 foram capturadas na Ilha de São Miguel, município de Santarém (PA), 50 animais pertenciam a um cativeiro comercial e 15 eram provenientes de um criadouro conservacionista, localizados na região metropolitana de Belém, Pará. De cada animal, foi colhida amostra de material biológico cloacal, utilizando-se swabs estéreis para em seguida serem acondicionados em tubos com meios de transporte e enviados ao laboratório para análises bacteriológicas. Todas as amostras foram imersas em caldos Selenito e BHI durante 24 horas e posteriormente semeadas em Agar Shigella-Salmonella e Agar Mac Conkey na temperatura de 37ºC por 24 horas. As UFCs (Unidades formadoras de colônia) foram semeadas em Agar Muller Hilton por mais 24 horas em estufa a 37ºC e identificadas pelo sistema Vitek® totalmente automatizado. Do total de 116 amostras foram obtidos 245 crescimentos bacterianos nos quais 83 (33,87%) eram provenientes dos animais de vida livre, com a identificação de 20 espécies bacterianas. Nos animais mantidos em cativeiro, foram obtidos 162 (65,72%) isolamentos, identificando-se 10 espécies de bactérias. Oito espécies foram encontradas em ambos os ambientes e 14 espécies em apenas um deles. A espécie Klebsiella pneumoniae foi a mais frequente, com 52 isolamentos, totalizando 21,22% dos crescimentos bacterianos, seguida de Enterobacter cloacae (35/14,29%), Serratia marcescens (29/11,84%) e Salmonella species (24/9,80%). Nos quelônios de vida livre, os microrganismos mais isolados constituiram-se dos genêros Enterobacter, Klebsiella, Citrobacter e Aeromonas. Klebsiella pneumoniae, Serratia marcescens, Enterobacter cloacae e Salmonella spp. apresentaram frequências elevadas naqueles animais cativos. Este resultado evidencia uma maior diversidade de microrganismos entre os animais de vida livre e uma contaminação elevada por amostra nos animais de cativeiro. As espécies Salmonella sp., E. coli e Acinetobacter ssp., tiveram sua frequência aumentada provavelmente devido a influência do cativeiro, sendo portanto, sugeridas como indicativas da qualidade sanitária de populações da tartaruga da Amazônia.

Aspectos microbiológicos e epidemiológicos da doença diarréica aguda no município de Juruti, Pará

O presente estudo descreveu os aspectos epidemiológicos e etiológicos da diarréia aguda no município de Juruti, Pará, Brasil. Foram avaliadas 261 amostras de fezes (170 diarréicas e 91 controles) de pacientes atendidos em Unidades de Saúde Pública, no período de fevereiro a julho de 2009. Para o isolamento de bactérias enteropatogênicas, utilizou-se meios seletivos indicadores e de enriquecimento. A caracterização bioquímica foi realizada utilizando os Sistemas API-20E e a sorologia, através de antisoros polivalentes e monovalentes. Para a detecção das categorias de E. coli diarreiogênicas foram executados dois ensaios de PCR multiplex. A pesquisa de Campylobacter jejuni e Campylobacter coli e Rotavírus foi executada através da técnica de ELISA, nas amostras de fezes. No exame parasitológico foram utilizados os métodos diretos (salina/Lugol) e sedimentação espontânea. Das 154 amostras positivas (118 diarréicas e 36 controles), 75,4% eram de infecções únicas e 24,6% de infecções mistas. A maioria dos casos incluiu crianças menores de 10 anos de idade (55,9%), sem diferença significante entre os sexos feminino e masculino. Os enteropatógenos mais frequentes no grupo diarréico foram E. histolytica/E. dispar (26%), Shigella spp (15,7%), G. lamblia (13,3%) e E. coli diarreiogênicas (12,8%), com Shigella spp associada à diarréia aguda (p = 0,0028). Quanto às categorias patogênicas de E. coli, a ETEC (7,2%) foi o tipo mais frequente nos casos de diarréia aguda, seguido de EAEC (5,9%). Os agentes menos frequentes nos casos diarréicos foram representados por C. jejuni/C. coli (4,7%), Rotavírus (2,8%), Salmonella Panama, A. hydrophila e A. sóbria (0,5%). Os resultados encontrados fornecem subsídios importantes para a vigilância epidemiológica e ambiental da doença diarréica aguda, no município de Juruti.

Estudos metabolômicos entre interações da rizosfera de Senna spectabilis e rizobacterias associadas

Pós-graduação em Química - IQ

Pesquisa de salmonella em mutuns (mitu mitu) mantidos em cativeiro

We collected 50 samples of curasow feces from the Scientific and Cultural Breeding Center in the city of Poços de Caldas. The samples were enriched in broth medium Tetrationato and Cystine-selenite and plated on Salmonella Shigella (SS), Mac Conkey (MC), endo-C (EC), Brilliant Green (VB) and Eosyn Methilen Blue EMB, remaining at 37 °C for 24h. Colonies suspected of Salmonella were inoculated in tubes containing Agar Triple Sugar Iron (TSI) and again incubated at 37 ºC for 24h. Tubes with characteristic growth were submitted to slide agglutination test with polyvalent somatic and flagellar serum. The medium SS, MC and VB were the most efficient for growing, and 36% of samples were positive for Salmonella spp. As birds are important reservoirs of Salmonella spp and it may represent a high risk to human health, there is a need to implement a cleaning routine in enclosures avoiding contamination between the enclosures and the consequent entrainment of these micro-organisms, by the keepers until their homes or other venues. The presence of Salmonella in breeding centers may be responsible for lower hatchability of eggs, undermining the purpose of the entity, which is to study and maintain the various birds species.

The prophenoloxidase system and in vitro interaction of Trypanosoma rangeli with Rhodnius prolixus and Triatoma infestans haemolymph

The presence of the prophenoloxidase (proPO) system in the haemolymph of Rhodnius prolixus and Triatoma infestans and the role played by Trypanosoma rangeli in the in vitro activation of proPO were tested. Both R. prolixus and T. infestans whole blood preparations showed a very active ProPO system. The proPO cascade of the two insect species were differentially activated by microbial-derived extracts: laminarin was a better activator of T. infestans haemolymph than of R. prolixus blood, and lipopolysaccharides from Shigella flexneri or Pseudomonas aeroginosa caused significant proPO activation of T. infestans haemolymph but not of R. prolixus preparations. For the two insect species, neither T. rangeli from culture nor parasite lysates were able to trigger proPO activation. The presence of the parasite in R. prolixus haemolymph/laminarin assays, however, significantly reduced the level of proPO activation to that of spontaneous activating controls. The immobilization of T. rangeli in vitro in haemolymph preparations occurred in both insect species and was dependent on the proPO activation intensity. Our results suggest that the susceptibility of R. prolixus to T. rangeli haemocoel infection may be explained, at least in part, by the suppression of the insect immune defence system i.e., inhibition of proPO in the presence of this protozoan parasite.

Laribacter hongkongensis: a potential cause of infectious diarrhea

In this study, we describe the isolation of Laribacter hongkongensis, a recently described genus and species of bacterium, in pure culture on charcoal cefoperazone deoxycholate agar from the stool of six patients with diarrhea. Three patients were residents of Hong Kong, and three of Switzerland. In none of the stool samples obtained from these six patients was Salmonella, Shigella, enterohemorrhagic Escherichia coli, Vibrio, Aeromonas, Plesiomonas, or Campylobacter recovered. Rotavirus antigen detection, electron microscopic examination for viruses, and microscopic examinations for ova and cysts were all negative for the stool samples obtained from the three patients in Hong Kong. Enterotoxigenic E. coli was recovered from one of the patients in Hong Kong. Unlike L. hongkongensis type strain HKU1, all the six strains were motile with bipolar flagellae. Sequencing of the 16S ribosomal RNA genes of the six strains showed that they all had sequences with only 0-2 base differences to that of the type strain. Pulsed field gel electrophoresis of the SpeI digested genomic DNA of the six isolates and that of the type strain revealed that the seven isolates were genotypically unrelated strains. More extensive epidemiologic studies should be carried out to ascertain the causative association between L. hongkongensis and diarrhea and to define the reservoir and modes of transmission of L. hongkongensis.