Listeria monocytogenes est un pathogène d'origine alimentaire à multiples facettes caractérisé par sa capacité d'adaptation dans des conditions défavorables et par sa prolifération dans une vaste gamme d'environnements, du sol aux cellules hôtes des mammifères. L'hétérogénéité génétique de L. monocytogenes se reflète dans sa structure clonale diversifiée, ce qui corrèle, dans une certaine mesure, avec des traits phénotypiques tels que la virulence ou la résistance au stress. La thèse portait sur deux phénotypes les plus éminents, la formation d'un biofilm et le potentiel de virulence, sous différents angles et à l'aide des technologies les plus récentes. Tout au long des études, des grands panels d'isolats ont été utilisés pour représenter la diversité intraspécifique. Stimulants défavorables tels que le choc froid et la privation d'éléments nutritifs induits par l'étape d'adhésion bactérienne. L'ajout de NaCl aux cultures de croissance a stimulé la production de biofilm et, de manière surprenante, il a considérablement intensifié la maturation du biofilm de cellules privées de nutriments. Un degré élevé de variation de la productivité relative du biofilm a été observé parmi les sérotypes, les génotypes, de même que les isolats selon les conditions de culture. Cependant, un certain génotype (complexe clonal 26) a révélé de manière caractéristique une production de biofilm plus élevée à froid (10°C), suggérant une association du génotype avec le phénotype du biofilm. Pan-GWAS a identifié un certain nombre de gènes parmi lesquels ceux impliqués dans des fonctions telles que la ‘transformation/compétence’, les ‘gènes liés aux phages’ et le ‘métabolisme du phosphate’ devront faire l'objet d'études plus approfondies sur leur rôle dans la formation du biofilm. L'analyse du séquençage de l'ARN a révélé une grande hétérogénéité intraspécifique dans les profils de transcriptome basal qui mettaient en évidence le rôle du réseau de régulation, y compris certains facteurs transcriptionnels avec des rôles clés dans la virulence tels que σB, PrfA, et CodY. La plasticité transcriptomique entre les lignées I et II ainsi que les génotypes hyper et hypovirulents ont confirmé les caractéristiques évolutives et épidémiologiques de L. monocytogenes. De plus, la voie métabolique centrale a été impliquée dans l'infection dans le système modèle de Galleria mellonella. En conclusion, la thèse a exploré la diversité intraspécifique de L. monocytogenes et a donné lieu à de nombreux résultats phénotypiques, génomiques et transcriptomiques. Grâce à l'approche intégrative des omiques en listeriologie, le présent travail contribuera à dévoiler la physiologie et la pathogenèse de la bactérie. / Conditions and proliferation in a wide range of environments from soil to mammalian host cells. The genetic heterogeneity in L. monocytogenes is reflected on its diversified clonal structure which correlates, to some extent, with phenotypic traits such as virulence or stress resistance. The thesis investigated two most prominent phenotypes, biofilm formation and virulence potential, from various perspectives using state-of-the art technologies. Throughout the studies, large panels of isolates were used to represent the intraspecific diversity. Unfavourable stimuli such as cold shock and nutrient deprivation induced bacterial adhesion step. Addition of NaCl to growth cultures stimulated biofilm production and, surprisingly, it significantly intensified biofilm maturation of nutrient-deprived cells. High degree of variation in relative biofilm productivity was observed among serotypes, genotypes, as well as isolates across culture conditions, however, certain genotype (clonal complex 26) revealed distinctively higher biofilm production under cold temperature (10°C) suggesting an association of genotype with biofilm phenotype. Pan-GWAS identified a number of genes among which those implicated in functions such as ‘transformation/competence’, ‘phage-related genes’, and ‘metabolism of phosphate’ will need further investigations for their roles in biofilm formation. RNA sequencing analysis revealed high intraspecific heterogeneity in basal transcriptome profiles that featured the role of regulatory network including certain transcriptional factors with key roles in virulence such as σB, PrfA, and CodY. The transcriptomic plasticity between lineage I and II as well as hyper- and hypovirulent genotypes supported the evolutionary and epidemiological characteristics of L. monocytogenes. Moreover, the central metabolic pathway was implicated in the infection in Galleria mellonella model system. Conclusively, the thesis explored intraspecific diversity in L. monocytogenes and resulted in ample phenotypic, genomic, and transcriptomic findings. With the integrative omics approach in listeriology, the present work will contribute to unveiling the physiology and pathogenesis of the bacterium.

Die antimikrobielle Wirkung eines Pflanzenextraktes aus fermentiertem Rooibos (Aspalathus linearis) und eines Extraktes aus Hopfen (Humulus lupulus) auf Milchsäurebildner (Lactobacillus spp., Carnobacterium spp., Leuconostoc carnosum), Verderbniserreger (Bacillus spp., Brochothrix spp., Pseudomonas fluorescens) und pathogene Mikroorganismen (Listeria monocytogenes, Salmonella Enteritidis) wird unter In-vitro-Bedingungen über einen Lagerungszeitraum von 28 Tagen bei Temperaturen von 10 °C und 25 °C untersucht. In den sich anschließenden Challengeversuchen wird evaluiert, ob sich die antimikrobiellen Wirkungen beider Pflanzenextrakte gegen Listeria monocytogenes auf die Lebensmittelmatrix Rotschmierkäse übertragen lassen. Mögliche herstellungsbedingte und sensorische Probleme aufgrund der Anwendung der Pflanzenextrakte im Lebensmittel „Weichkäse“ werden erörtert. Die Ausgangskeimzahl beträgt je Keim 102–103 KbE/ml. Der Rooibosextrakt wird in einer Konzentration von 5 mg/ml zugesetzt und der Hopfenextrakt in einer Konzentration von 30 µg/ml. Unter In-vitro-Bedingungen ist eine statistisch abgesicherte bakteriostatische Wirkung des Rooibosextraktes auf Listeria monocytogenes bei einer Lagerungstemperatur von 10 °C in den ersten 14 Tagen nachweisbar. Die Reduzierung der Verderbniserreger in den ersten 24 Stunden nach Zugabe des Rooibosextraktes bei beiden Lagerungstemperaturen bzw. der Milchsäurebildner bei 25 °C ist statistisch nicht abzusichern und bei den weiteren Probenahmen nicht mehr darstellbar. Der Rooibosextrakt hat keine nachweisbare antibakterielle Wirkung auf Salmonella Enteritidis. Alle grampositiven Testkeime lassen sich durch die Zugabe des Hopfenextaktes statistisch signifikant in ihrem Wachstum beeinflussen. Eine antibakterielle Aktivität des Hopfens zeigt sich bei Listeria monocytogenes in einer verlängerten lag-Phase und anschließendem bakteriostatischen Effekt bei 10 °C. Der Hopfenextrakt hat keinerlei Wirkung auf die gramnegativen Teststämme Pseudomonas fluorescens und Salmonella Enteritidis. Der Extrakt aus Hopfen zeigt auch in der Lebensmittelmatrix Rotschmierekäse eine signifikante Wachstumshemmung auf Listeria monocytogenes. Der Extrakt aus fermentierten Rooibos zeigt keinen antilisteriellen Effekt. Beide Pflanzenextrakte beeinflussen leicht die sensorischen Eigenschaften der Käse, die Käsereifung nicht. Erstmals liegen antibakterielle In-vitro-Studien der getesteten Pflanzenextrakte gegen lebensmittelrelevante Mikroorganismen über 28 Tage vor. Eine Anwendung von Pflanzenextrakten als antimikrobieller Lebensmittelzusatz ist erst nach entsprechender Validierung im jeweiligen Einzelprodukt möglich.

The disease listeriosis is caused by Listeria monocytogenes. This common food-borne disease has been responsible for about 0.1 to 10 cases per million inhabitants per year. However, this disease is serious with its high fatality rates of 20% - 30%, and 40% of all cases reported have been in pregnant women suffered from a foetal abortion. Recently the organism has acquired resistance to antibiotic treatment and the development of an alternative treatment is necessary. Class IIa bacteriocins such as leucocin A have been shown to be active against L. monocytogenes. However, the leucocin A receptor molecule responsible for growth inhibition within L. monocytogenes remains unclear. Various studies have implicated the mannose PTS permease (EIIt Man) of L. monocytogenes as the putative receptor for class IIa bacteriocins. The results from studies reviewed indicate that the EIIt Man of L. monocytogenes could be the chiral receptor needed for bacteriocin interaction at the surface of targeted cells. Specifically, the membrane associated IIDMan and IICMan subunits were implicated in direct interaction with class IIa bacteriocins. Our study focused on cloning, expression and purification of the subunits of the mannose PTS permease of L. monocytogenes EGD. Primers were designed to amplify the subunit genes of the mptACD operon. The mptC, mptD and mptAB genes which were then successfully cloned into pET28a expression vector and transformed into E. coli JM109(DE3) host strain. Recombinant plasmids were screened using colony PCR. Subsequently recombinant pET28-C, pET28-D and pET28-AB was once again transformed and expressed in the E. coli BL21(DE3) pLysS expression host strain. After an induction at 30°C for 5 hours, IICMan and IIDMan were found to be expressed in the cell membrane, whilst IIABMan was expressed in the cytosol of the host expression strain. Membrane proteins His-IICMan, His- IIDMan, and cytosol associated His-IIABMan were purified using Ni2+-NTA affinity chromatography. Results for His-IICMan yielded a 28 kDa protein and a 55 kDa co-purified protein. Results for His-IIDMan yielded a 31 kDa protein and a 60 kDa co-purified protein. Results for His-IIABMan yielded a 35 kDa protein and a 68 kDa co-purified protein. A western blot analysis revealed that all proteins purified carried an attached His-tag as detected by an anti-mouse peroxidase conjugate anti-His-tag antibody. / Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2010.

Autorius: Aušra Lozoraitytė Tema: Listeria monocytogenes padermių, išskirtų iš mažmeninėje rinkoje parduodamų žuvų produktų, identifikavimas ir serotipavimas molekuliniais tyrimo metodais Darbo vadovas: prof. dr. Artūras Stimbirys Atlikimo vieta: Darbas 2012-2014 metais atliktas Lietuvos sveikatos mokslų universitete, Veterinarijos akademijoje, Maisto saugos ir kokybės katedroje. Darbo dydis: 52 puslapiai, 10 lentelių, 17 paveikslų. Darbo tikslas: palyginti Listeria monocytogenes identifikavimui naudotus standartinį ir molekulinį metodus. Atlikto darbo tyrimų objektas yra Listeria monocytogenes, kuri buvo išskiriama iš mažmeninėje rinkoje parduodamų šaltai rūkytų žuvų (lašišų) produktų. Tyrimai atlikti tarpusavyje derinant standartinius mikroorganizmų kultivavimo metodus ir molekulinius DNR analizės metodus. Tyrimų metu buvo ištirta 160 šaltai rūkytų žuvų produktų, t. y. lašišų papilvės, lašišų gabalėliai, lašišų file ir lašišų nugarėlės. Tyrimo metu nustatyta, kad šaltai rūkytų žuvų užkrėstumas listerijomis priklauso nuo produkto rūšies. Šaltai rūkytų lašišų papilvėse užkrėstumas listerijomis yra 7,5 karto didesnis nei lašišų nugarėlėse (p<0,05), 1,8 karto nei lašišų gabalėliuose (p<0,05) ir 30 kartų didesnis nei lašišų file (p<0,05). Mikrobiologinių tyrimų metu nustatyta, kad 32,5 % (p<0,05) žuvų produktų mėginių buvo užkrėsti L. monocytogenes, o atlikus dauginę PGR nustatyta, kad L. monocytogenes buvo užkrėsti 23,13 % mėginių (p<0,01). Tai įrodo, kad molekuliniai tyrimo... [toliau žr. visą tekstą] / Author: Ausra Lozoraityte Subject: The molecular research methods in the identification and serotyping of Listeria monocytogenes strains in fish products in retail Supervisor of the work: prof. dr. Arturas Stimbirys Location: The study was conducted in the Food Safety and Quality Department of Veterinary Academy of the Lithuanian University of Health Sciences, in 2012-2014. Scope of the work: 52 pages, 10 tables, 17 figures. The aim of the work: Identification of the Listeria monocytogenes by comparing molecular and standard methods. The study object is Listeria monocytogenes, which is found in the cold smoked fish (salmon) products, sold in the retail market. The study has been carried out combining standard micro-culture cultivation and molecular DNA analysis methods. 160 cold-smoked fish products, i. e. underbellies of the salmon, salmon pieces, salmon fillet and backs of the salmon was investigated during in the study. Also it was found that contamination of the cold smoked fish products with listeria depends on the type of the product. Contamination with listeria in the underbellies of the salmon is 7.5 times higher, than in the backs of the salmon (p<0.05), 1.8 times higher, than in the pieces of salmon (p<0.05) and 30 times higher, than in the salmon fillet (p<0.05). Microbiological researches showed that 32.5 % (p<0.05) of the fish products samples were infected with L. monocytogenes, and after the multiplex PCR was found that 23.13 % of samples (p<0.01) were... [to full text]

Tyrimo tikslas – nustatyti įvairių cheminių medžiagų įtaką Listeria monocytogenes bakterijoms mėsos (jautienos) žaliavoje. Mėsos žaliava (jautiena) buvo perkama įvairiose mėsos ir jos produktų prekybos vietose. Tyrimų laboratorijoje buvo ištirtas Listeria monocytogenes bakterijų atsparumas įvairioms cheminėms medžiagoms: pieno rūgščiai, acto rūgščiai, cinamaldehidui, chloro dioksidui. Buvo naudotos įvairios šių medžiagų koncentracijos: pieno rūgšties – 2 %, 3 %, 4 %, 5 %, acto rūgšties – 2 %, 3 %, 5 %, cinamaldehido - 0,5 %, 1,5 %, 3 %, chloro dioksido – 5 ppm ir 15 ppm. Atsparumas buvo vertintas užkrečiant žalią jautieną. Taip pat buvo nustatytas šių cheminių medžiagų poveikis bendram mikroorganizmų skaičiui. Atlikus tyrimą nustatyta, kad iš visų tyrime naudotų cheminių medžiagų didžiausią neigiamą poveikį Listeria monocytogenes ir BMS augimui daro pieno rūgštis, nes per 48 valandas L. monocytogenes skaičius sumažėjo 129 kartus, o bendras mikroorganizmų skaičius – 22,6 karto (p<0,05). Lyginant naudotų cheminių medžiagų poveikį nustatyta, kad pieno rūgštis ir chloro dioksidas stipresnį neigiamą poveikį darė L. monocytogenes bakterijoms, o cinamaldehidas ir acto rūgštis – bendram mikroorganizmų skaičiui. / The goal of the research is to determine the influence of various chemicals on Listeria monocytogenes bacteria in raw meat (beef). Raw meat was purchased in various markets of meat and meat products. The resistance of Listeria monocytogenes to various chemicals like lactic acid, acetic acid, cinnamaldehyde, chlorine dioxide was tested in the laboratory. These chemicals were used in various concentrations: lactic acid – 2 %, 3 %, 4 %, 5 %, acetic acid – 2 %, 3 %, 5 %, cinnamaldehyde – 0,5 %, 1,5 %, 3 %, chlorine dioxide – 5 ppm and 15 ppm. Resistance was evaluated by infecting raw beef. The effects of these chemicals to the total number of microorganisms was also determined. The research revealed that the most influencing chemical from the ones tested was lactic acid. Within 48 hours the amount of L. monocytogenes decreased 129 times and the total number of microorganisms decreased 22,6 times (p<0,05). While comparing the effects of the chemicals used in the research it was determined, that lactic acid and chlorine dioxide has a stronger negative impact on the growth of L. monocytogenes bacteria and cinnamaldehyde and acetic acid has a bigger influence on the total number of microorganisms.

Diss. (sammanfattning) Stockholm : Karol. inst., 2004. / Härtill 4 uppsatser.

Thesis (Ph. D.)--University of Washington, 1997. / Vita. Includes bibliographical references (leaves [93]-113).

Thesis (Ph. D.)--West Virginia University, 2006. / Title from document title page. Document formatted into pages; contains xvi, 250 p. : ill. (some col.). Vita. Includes abstract. Includes bibliographical references.

Este projeto foi desenvolvido no Laboratório de Patologia Experimental do Hospital de Clínicas de Porto Alegre (HCPA), com a aprovação da Comitê de Ética em Pesquisa do HCPA e com apoio financeiro parcial do Fundo de Incentivo à Pesquisa e Eventos do HCPA (FIPE). O experimento 1, chamado de projeto piloto, teve como objetivo implementar a técnica de IHQ para identificar a Listeria monocytogenes (L.m.), utilizando anticorpo policlonal antilisteria monocytogenes (Biodesig ). Vários testes foram realizados para acertar a diluição (1:1000) que foi diferente da preconizada pelo fabricante. Os blocos de parafina, de dez placentas provenientes de parto prematuro ou aborto foram utilizados para os cortes histológicos e a preparação das lâminas para a coloração Hematoxilina e Eosina (HE) e imunoistoquímica (IHQ). As lâminas foram identificadas por números para resguardar a identidade das pacientes. O resultado do HE mostrou alterações inflamatórias em oito placentas e L. m. foi identificada pelo IHQ em cinco dessas placentas. O objetivo do 2º experimento foi identificar a L. m. em tecido nervoso cerebral de ruminantes, utilizando a técnica implementada no projeto piloto. O material utilizado neste trabalho foi cedido pelo Setor de Patologia Veterinária do Departamento de Patologia da Universidade Federal de Santa Maria. Os casos estudados (2 ovinos, 1 caprino e 2 bovinos) tinham suspeitas clínicas diversas e as necropsias dos animais evidenciaram aspectos sugestivos da doença. Os cinco casos foram confirmados pelo IHQ, comprovando a importância da utilização desta técnica para o diagnóstico da listeriose no SNC de ruminantes. O 3º experimento objetivou identificar a L. m. em placentas encaminhadas ao Serviço de Patologia do HCPA no ano 2000. Da mesma forma que no experimento 1, as lâminas foram identificadas por números. Após o levantamento realizado nos registros dos exames anatomopatológicos (AP) deste setor, observou-se que 714 AP eram de placentas provenientes de aborto, parto prematuro e nascimento a termo examinados naquele período. Foram sorteados 254 AP para análise através de HE, revelando que 148 desses AP apresentavam alterações inflamatórias (corioamnionite, vilite e deciduite). Os blocos destas placentas foram utilizados para fazer as lâminas e realizar IHQ. A consulta aos prontuários dos casos com alterações inflamatórias permitiu observar que um deles tinha a confirmação bacteriológica de L. m. na placenta, tornando-se este o controle positivo. O controle negativo foi selecionado entre aqueles sorteados que não apresentavam alterações inflamatórias. A presença de L. m. foi identificada em 33,78% das placentas analisadas pela técnica IHQ. Corioamnionite e vilite foram as alterações inflamatórias que mostraram diferença estatística significativa nas placentas positivas. L. m. estava presente nas placentas de 1º, 2º e 3º trimestres gestacionais. A idade das gestantes, casos de aborto e/ou parto prematuro não mostraram diferença estatística significativa com a presença ou ausência de L. m. nas placentas. Abortos habituais ocorreram em pacientes com ou sem L. m. no tecido placentário. Conclusão: a técnica de imunohistoquímica pode ser utilizada para confirmar o diagnóstico histopatológico de listeriose em placentas e tecido nervoso central de ruminantes. / This project was developed at the Pathology Experimental Laboratory of Hospital de Clínicas de Porto Alegre (HCPA), after approval by the local Research Ethics Committee, and received partial financial support from the Research Incentive Fund (FIPE) of HCPA. The first experiment (or pilot scheme) aimed at implementing the immunohistochemistry (IHC) technique for the identification of Listeria monocytogenes (L.m.), by way of anti-listeria polyclonal antibody (Biodesign ). Several tests were performed in order to find the correct dilution (1:1000), which was different from that which had been recommended by the manufacturer. Paraffin-embedded blocks of placentas obtained from premature birth or abortion were used for the histological sections, and the slides were prepared for hematoxylin-eosin (HE) staining and immunohistochemistry. The slides were labeled with numbers in order to preserve patient anonymity. The result of HE staining revealed inflammatory disorders in eight placentas and L.m. was identified by IHC in five of them. The aim of the second experiment was to identify L.m. in the brain tissue of ruminants by employing the technique implemented in the pilot scheme. The material used in this study was provided by the Division of Veterinary Pathology of the Department of Pathology of Universidade Federal de Santa Maria. The animals studied (2 sheep, 1 goat and 2 cows) exhibited several clinical suspicions and the necropsy results were suggestive of the disease. The five cases were confirmed by IHC, highlighting the importance of this technique to the diagnosis of listeriosis in the CNS of ruminants. The aim of the third experiment was to identify L.m. in placentas and abortion specimens that had been referred to the Division of Pathology of HCPA in year 2000. As with the first experiment, the slides were numbered. After surveying the registries of pathoanatomical exams (PAs), we noted that 714 PAs corresponded to placentas, obtained from abortion, premature delivery and full-term birth, examined during that period. Two hundred fifty- four PAs were randomly selected (drawn out) for HE staining, and 148 of these PAs showed evidence of inflammatory disorders (chorioamnionitis, villitis and deciduitis). The blocks of these placentas were used for preparing the slides and carrying out IHC. The analysis of medical records in the cases with inflammatory disorders allowed us to observe that one of the records contained bacteriological confirmation of L.m. in the placenta (positive control). The negative control was chosen among the drawn-out PAs that did not present inflammatory disorders. L. m. was detected in 33.78% of the placentas analyzed by IHC. Chorioamnionitis and villitis showed significant statistical difference in the positive placentas. L. m. occurred in the 1st, 2nd and 3rd trimesters of pregnancy. The age of pregnant women, the cases of abortion and/or premature births were not statistically different as to the presence or absence of L. m. in the placentas. Habitual abortions occurred in patients with or without L. m in the placental tissue. Conclusion: Immunohistochemistry may be used to confirm the histopathological diagnosis of listeriosis in placentas and brain tissue of ruminants.

Este estudo objetivou analisar o potencial de deterioração da microbiota psicrotrófica presente em presunto cozido fatiado comercializado entre Maio e Junho de 2015 e Fevereiro e Março de 2016 no mercado público da cidade de Porto Alegre/RS e avaliar a influência de fatores ambientais na qualidade microbiológica do mesmo. Os presuntos foram coletados em 4 bancas desse local e foram realizadas contagens de bactérias psicrotróficas de 8 amostras e pesquisa de Listeria monocytogenes. Selecionaram-se 134 colônias de psicrotróficos isolados de presunto fatiado, 71 deles apresentaram atividade proteolítica, 58 atividade lipolítica e 12 apresentaram produção de exopolissacarídeo. Selecionaram-se 2 bactérias com a presença dessas atividades para identificação molecular, as quais foram identificadas como Kluyvera sp. e Carnobacterium sp. Além delas, mais 2 Listeria monocytogenes isoladas nesse trabalho foram submetidas ao teste de produção de biofilme, resultando como fracas formadoras e também ao teste de aderência em aço inoxidável, todas apresentando capacidade de adesão. A pesquisa de Listeria monocytogenes nos presuntos fatiados mostrou 100% de presença, sendo que 50% foram identificadas como L. monocytogenes, as quais pertenceram aos sorotipos 1/2a (1), 1/2b (2), 1/2c (2). Realizou-se análise de presunto cozido inteiro, em sua embalagem original, sendo que não foram encontrados micro-organismos Tratou-se o presunto fatiado com extrato de hibisco a 40% e pediocina a 0,5% e 1,0% e realizou-se contagem de mesófilos, psicrotróficos, Listeria spp., S. aureus e E.coli. O extrato de hibisco reduziu a carga desses micro-organismos. Pediocina 0,5% e 1% apresentaram pouca ação frente ao controle de mesófilos, psicrotróficos e E. coli, mas mantiveram a carga de S. aureus controlada e foram eficazes contra Listeria spp. Foram realizadas também contagens para Listeria monocytogenes, E. coli, S. aureus, mesófilos e psicrotróficos em suabes oriundos de fatiador de alimentos, superfície de contato e utensílio utilizados nas bancas do mercado público. Em conclusão grande parte dos psicrotróficos apresentou atividade proteolítica e lipolítica, as quais alteram organolepticamente o alimento. Alguns apresentaram produção de biofilme e capacidade de aderência, fato indesejado, pois sua remoção é mais difícil no ambiente industrial, com isso nota-se que a legislação brasileira apresenta carência na contagem de psicrotróficos em produtos cárneos. / This study aimed to evaluate the microbiota present in sliced cooked ham sold in the public market in Porto Alegre/RS and evaluate the ability of compounds with antimicrobial activity of hibiscus extract and pediocin to control the microbiota found. Ham collected was stored refrigerated until to arrive the laboratory for analysis. Psychrotrophic bacteria counts were performed. Were selected 134 colonies of psychrotrophic microorganisms isolated for sliced ham and 71 of them showed proteolytic activity, 58 lipase activity, 12 showed production of exopolysaccharide. Two of these bacteria were selected for molecular identification which were identified as Kluyvera sp. and Carnobacterium sp. These two bacteria plus two Listeria monocytogenes isolated for sliced ham were subjected to testing of biofilm production (resulting as weak forming of biofilm) and were tested for adhesion in stainless steel and all showed this property. The research of Listeria spp. in sliced cooked ham showed 100% of presence, which 50% were identified as L. monocytogenes to serotypes 1/2a (1) 1/2b (2), 1/2c (2). Analysis was carried out of a whole piece of cooked ham in its original packaging and none microorganisms were found. The sliced ham was treated with hibiscus extract of 40% and pediocin of 0.5% and 1.0% and has been mesophilic, psychrotrophic, Listeria spp., S. aureus and E. coli counts The hibiscus extract reduced the quantity of these microorganisms. Pediocin 0.5% and 1,0% had little action against the control of mesophilic, psychrotrophic and E. coli, but in S. aureus counts were controlled bacteria charge and were effective against Listeria spp.. Also counts of Listeria monocytogenes, E. coli, S. aureus, mesophilic and psychrotrophic bacteria were performed from swabs of slicer food, contact surface and food tool (knife or spatula) used in public market stalls. In conclusion, most of the psychrotrophs presented proteolytic and lipolytic activity, which alter organoleptically the food. Some of them have presented biofilm production and adhesion capacity, undesirable fact because when the biofilm is formed is more difficult to remove it in the industrial environment. With this it is showed that the brazilian legislation presents a lack in the research of psychrotrophs in meat products kept refrigerated.