Orientador: Meuris Gurgel Carlos da Silva / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia Quimica / Made available in DSpace on 2018-08-13T14:49:52Z (GMT). No. of bitstreams: 1 Serenotti_Fernando_D.pdf: 4169220 bytes, checksum: 8a060bd79b54b3181aca4449a7bf96f3 (MD5) Previous issue date: 2009 / Resumo: O lodo de esgoto, um dos principais componentes das águas residuárias geradas mundialmente, é um resíduo com alto conteúdo de umidade e grande carga de microrganismos. O uso da secagem apresenta-se como uma poderosa ferramenta na área ambiental devido a possibilidade de redução de volume e inativação microbiológica. O presente trabalho teve como objetivo avaliar o processo de secagem empregando um secador convectivo direto de fluxo ascendente, para redução de volume e inativação microbiológica de lodo de ETE. O lodo utilizado foi oriundo do tratamento primário da ETE - Tatu, da cidade de Limeira - SP. Foram desenvolvidos os seguintes estudos: determinação das características físico-químicas, ensaios de secagem a partir do planejamento experimental 3² em duplicata (duas variáveis - temperatura e vazão do ar de secagem, e três níveis) com análise estatística dos dados, a avaliação da redução de volume do lodo seco, ajuste de modelos matemáticos aos dados experimentais, verificação do efeito da sazonalidade do lodo de esgoto no processo de secagem, e avaliação da inativação microbiológica. Pelos resultados do trabalho verificou-se que as características físicoquímicas não sofreram alterações significativas antes e após o processo de secagem, e com isso, pode-se considerar que não ocorreu emissão atmosférica destes componentes. A cinética do processo de secagem se caracterizou inicialmente pelo período de aquecimento do material seguindo-se pelos períodos de secagem à taxa constante, 1.ª e 2.ª taxas decrescentes. O início do 2.º período de taxa decrescente foi marcado pela quebra ou fissura da torta, com aumento significativo da taxa de secagem, que é uma característica específica da secagem de materiais como o lodo. A análise estatística mostrou que a temperatura foi a variável mais significativa, indicando que o mecanismo de secagem do lodo de esgoto foi predominantemente difusivo. Contudo, devido o comportamento do 2.º período de secagem, os modelos difusivos não se ajustaram adequadamente, necessitando de modelos empíricos para descrever a 2.ª taxa decrescente. A redução de volume do material foi satisfatória, no caso cerca de 50%. A verificação da sazonalidade mostrou uma discreta diferença nos valores de pH e umidade inicial dos lodos estudados. As análises microbiológicas mostraram que após o tratamento térmico, houve a inativação microbiológica em praticamente todas as condições de processo estudadas, sendo o binômio tempo de exposição - temperatura o fator predominante para a esterilização e desinfecção do lodo de ETE. Com os resultados obtidos é possível considerar que o processo de secagem deste trabalho apresenta bom potencial de aplicação como tratamento de lodo de ETE, mas também de outros materiais que possuam características similares. Palavras-chave: secagem, lodo de esgoto, redução de volume, inativação microbiológica. / Abstract: Sludge, one of the main components of wastewater generated worldwide, is a waste with great quantity of moisture and microorganisms. The usage of drying has been a powerful tool in the environmental area due to the possibility of reduction of volume and microbiological inactivation. The present study aims to assess the drying process using an upflow direct convection dryer to reduce sludge volume and microbiological inactivation of Sludge Treatment Plants (STP). The used sludge has been taken from the primary treatment of STP - Tatu, city of Limeira, São Paulo. The following studies have been carried out: determination of physicochemical characteristics, drying experiments from the experimental design 3² in duplicate (two variables - temperature and drying air flow, and three levels) with statistical analysis of data, assessment of volume reduction of dried sludge, fitting of mathematical models to experimental data, verification of the effect of the seasonality of the sludge in the drying process, and assessment of microbiological inactivation. With the results of this study it could be verified that the physicochemical characteristics have not suffered significant changes nor before neither after the drying process, therefore, it can be said that atmospheric emission of these components have not occurred. The kinetics of the drying process was initially characterized by the heating period of the material followed by the drying periods of constant rate, first and second falling rates. The beginning of the second period of falling rate was marked by the cake break or fissure with the significant increase of the drying rate which is a specific characteristic of the drying of materials such as sludge. The statistic analysis has shown that the temperature was the most significant variable indicating that the sludge drying mechanism has predominantly been diffusive. However, due to the behavior of the second period of drying, the diffusive models did not adjust adequately, being necessary the use of empiric models to describe the second falling rate. The reduction of material volume was satisfactory, 50%, in this case. The verification of the seasonality has shown a discrete difference in the pH values and in the moisture content of the studied sludge. The microbiological analyses showed that after the thermal treatment there was microbiological inactivation practically in all the studied conditions of the process, being the binomial time of exposition - temperature the predominant factor for the sterilization and disinfection of sludge of STP. With the obtained results it is possible to consider that the drying process of this study shows a good potential of application for sludge treatment of STP, and also for other materials that have similar characteristics. / Doutorado / Engenharia de Processos / Doutor em Engenharia Química

Orientador: Expedito Tadeu Facco Silveira / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-03T22:51:31Z (GMT). No. of bitstreams: 1 Bedendi_RafaelaFerrari_M.pdf: 1078798 bytes, checksum: 4d00a8483869fd9a13387496464d8a85 (MD5) Previous issue date: 2003 / Resumo: Este trabalho teve como objetivo avaliar a eficiência de sistemas de embalagem com atmosfera modificada na preservação da qualidade de presunto cozido fatiado. Para tanto, acompanhou-se a estabilidade do produto a 4 :t 1°C, acondicionado em uma embalagem barreira a gases denominada "master pack" , com três tipos de atmosferas modificadas (100% CO21 60% CO2 I 40% N2 e 25% CO2 I 75% Nz). Comparativamente, foi avaliada a estabilidade do produto no sistema convencional sob vácuo. Bandejas de poliestireno não expandido contendo cerca de 200g de presunto cozido fatiado foram acondicionadas em embalagens de um filme poliolefínico selado a quente, sendo denominadas como embalagens primárias. Em seguida, seis embalagens primárias, de presunto cozido fatiado, foram colocadas dentro de um saco barreira a gases chamado de "master pack" (embalagem secundária), constituído de polietileno de baixa densidade e copolímeros de etileno e álcool vinílico. A taxa de permeabilidade ao oxigênio do material foi de 1,87 cm3 (CNTP)/m2/atm/dia a 23°C e a seco. No produto convencional, as bandejas foram envoltas em um filme termo-encolhível. Periodicamente, os produtos foram avaliados quanto à qualidade sensorial (aparência e odor), qualidade microbiológica (contagens de enterobactérias, psicrotróficas aeróbias, bactérias lácticas e Pseudomonas sp.) e características físicas e químicas (coloração e pH). As embalagens com atmosfera modificada foram periodicamente avaliadas quanto à composição gasosa do espaço-livre. Não foram verificadas alterações físicas e químicas no presunto cozido fatiado, nos diversos sistemas de acondicionamento, durante os períodos estudados. A estabilidade do produto para cada sistema de acondicionamento foi determinada com base em alterações sensoriais, tendo sido definidas como limitantes as classificações: cor característica do presunto fresco e odor característico de presunto fresco moderados. A vida útil do presunto no sistema convencional sob vácuo foi de 15 dias e no final deste período, as contagens de enterobactérias, psicrotróficas aeróbias, bactérias lácticas e Pseudomonas sp. foram de 4,0 109 UFC/g, 5,1 109 UFC/g, 4,5 109 UFC/g e < 1 109 UFC/g, respectivamente. No produto em atmosfera de 25% CO2/75% N2, o período de vida útil foi de 22 dias, ao final da qual, o presunto apresentava contagens de enterobactérias, psicrotróficas aeróbias, bactérias lácticas e Pseudomonas sp. de < 1, 5,1, 4,5 e < 1 log UFC/g, respectivamente. Contudo, constatou-se um prolongamento significativo da durabilidade do presunto cozido fatiado quando acondicionado sob atmosferas contendo altas concentrações de gás carbônico, em relação ao sistema convencional, uma vez que o período de estabilidade determinado foi de 29 dias (aumento de 93,3%), para os produtos em atmosfera 100% CO2 e 60% CO2/ 40% N2. Quanto à ação bacteriostática do gás carbônico, comprovou-se retardamento no desenvolvimento de bactérias psicrotróficas aeróbias e inibição do crescimento de enterobactérias e Pseudomonas sp. quando a atmosfera foi de 100% CO2. Quando esta atmosfera foi de 60% CO2 I 40% N2, houve um retardamento tanto no desenvolvimento de psicrotróficas aeróbias como no de Pseudomonas sp., sendo que a contagem de Pseudomonas sp. foi de 4,7 log UFC/g, no 50° dia de estocagem e inibição do crescimento de enterobactérias. Com base nos resultados obtidos confirmou-se o efeito positivo do acondicionamento sob atmosfera modificada com gás carbônico na extensão da vida útil de presunto cozido fatiado, sendo mais eficiente o seu efeito quanto maior for a concentração desse gás no espaço-livre / Abstract: The purpose of this study was to evaluate the efficiency of packaging systems with modified atmosphere to preserve the quality of sliced cooked ham. Thus the quality stability of the product at 4 +/_:t 1°C, packed in a gas barrier package with three kinds of modified atmosphere (100% CO2, 60% CO2/40% N2 and 25% CO2 I 75% N2), was studied. The shelf-life of each condition of atmosphere was analysed. Comparatively, the shelf-life of the same product packed in conventional under vacuum was also evaluated. No expanded polystyrene trays with about 200g of sliced cooked ham were placed in a hot sealed poliolefinic film packages, nominated as primary packages. After that, six primary packages of sliced cooked ham were placed inside a barrier bag called "master pack" (secondary packages), formed of low density polyethylene and copolymer of ethylene vinylindene alcohol. The material permeability rates to oxygen was 1,87 cm3 (CNTP)/m2/atmlday at 23°C and dry. For the conventional product, the trays were wrapped in a thermoform film. The products were periodically evaluated as to their organoleptie quality (appearance and odour), microbiological quality (counts of Enterobacteriaceae, aerobic psychrotrophic microorganisms, lactic acid bacteria and Pseudomonas sp.) and physical and chemical characteristics (colour and pH). The headspace gas composition of the modified atmosphere packages were periodically evaluated. The cooked ham did not present any physical or chemical alteration in any of the packaging systems during the analysed period. The shelf-life for each packaging system was established based on organoleptic alterations that defined as the limitin classification: characteristic collor of the fresh ham and characteristic odour of the fresh ham moderates. The shelf life of the ham in the conventional under vacuum was 15 days. At the end of this period, the counts of Enterobacteriaceae, aerobic psychrotrophie microorganisms, ladie acid bacteria and Pseudomonas sp. were 4,0 log CFU/g, 5,1 log CFU/g, 4,5 log CFU/g and < 1 CFU/g, respectively. The product placed in a 25% CO2/75% N2 atmosphere, the shelf-life of the was 22 days and during this period, the counts of Enterobacteriaceae, aerobic psychrotrophie microorganisms, lactic acid bactería and Pseudomonas sp. were < 1, 5,1, 4,5 and < 1 109 CFU/g, respectively. higher it's effect. However, it was verified a significant shelf-life increase of sliced cooked ham in atmospheres with high concentrations of carbon dioxide in relation to the conventional under vacuum, as fo 11 ows: 29 days (93,3% increase) for products in 100% CO2 and 60% CO21 40% N2 atmosphere. Due to the bacteriostatic action of the carbon dioxide, the development of aerobic psychrotrophics microorganisms was retarded and the growth of Enterobacteriaceae and Pseudomonas sp. was inhibited in the 100% CO2 atmosphere. When the atmosphere was 60% CO2 I 40% N2, the development of both aerobic psychrotrophic microorganisms and Pseudomonas was retarded, and the counts of Pseudomonas sp. was 4,7 log CFU/g, after 50 days of storage and the development of Enterobacteriaceae was inhibited. Based on the obtained results, it was confirmed the positive effect of the packaging in modified atmosphere with carbon dioxide on the shelf-life increase of sliced cooked ham. It has been proved that, the higher the concentration of carbon dioxide in the headspace / Mestrado / Mestre em Tecnologia de Alimentos

Contamination of meat with microorganisms during slaughter is inevitable. Hygiene management systems (HMSs) such as the Hygiene Assessment System (HAS) and Hazard Analysis Critical Control Point (HACCP) are used to prevent the contamination of beef with both spoilage and pathogenic microorganisms during slaughter. This study compared the effect of the HAS alone and a combination of HAS + HACCP on the microbiological quality of beef and investigated the survival of Escherichia coli O157:H7 co-cultured with different levels of Pseudomonas fluorescens and Lactobacillus plantarum on fresh beef. HAS alone and HAS combined with HACCP systems were each represented by two abattoirs. Sponge swab samples were collected from chilled beef carcasses for indicator organisms: Aerobic Plate Counts (APC), Enterobacteriaceae, Pseudomonas spp., and lactic acid bacteria. Swabs were also collected for pathogenic bacteria: E. coli O157:H7, Staphylococcus aureus and Salmonella spp. There was no significant difference between the microbiological quality of beef carcasses processed in the abattoirs with the HAS and that of beef carcasses processed in abattoirs with combined HAS + HACCP. E. coli O157:H7 was isolated from carcasses processed in an abattoir with the combined HAS + HACCP system. Moreover, although overall S. aureus counts at all abattoirs were comparable, a higher incidence (47% of carcasses) was obtained from an abattoir with combined HAS + HACCP. Salmonella spp. was not detected during the study. The microbiological quality of beef at HAS abattoirs is not significantly different to that of beef processed at HAS + HACCP abattoirs. The combined HAS + HACCP did not prevent contamination of beef carcasses with E. coli O157:H7 and S. aureus. Effective implementation of HAS can reduce contamination of beef with spoilage and pathogenic microorganisms. The effect of different levels of P. fluorescens (102 and 106 log10 cfu/ml) and L. plantarum (102 and 104 log10 cfu/ml) on the survival of E. coli O157:H7 on beef loins was investigated. Sterile beef loins inoculated with E. coli O157:H7 and P. fluorescens were aerobically stored for 7 days at 4°C, while those inoculated with E. coli O157:H7 and L. plantarum were vacuum-packaged and stored for 8 weeks at 4°C. APC, E. coli O157:H7 and either P. fluorescens or L. plantarum counts were determined at different storage intervals. For the aerobically packaged beef loins, E. coli O157:H7 was detected throughout the 7-day storage period regardless of the P. fluorescens level in the inoculum. For the vacuum packaged beef loins, similar inoculum levels of E. coli O157:H7 and L. plantarum allowed E. coli O157:H7 to survive until week 5 of storage, while a higher inoculum level of L. plantarum inhibited E. coli O157:H7 from week 3. Once fresh beef has been contaminated with E. coli O157:H7 the level of P. fluorescens in the background flora does not inhibit its survival and growth. However, under vacuum storage, the application of L. plantarum as a biopreservative inhibits the survival of E. coli O157:H7 on beef. Comprehensive strengthening of preventive strategies is required to eliminate contamination of beef carcasses with E. coli O157:H7. Bacterial contamination of carcasses during slaughter is inevitable. Effective implementation of HAS at abattoirs produces beef carcasses of microbiological quality comparable to that produced through the use of combined HAS and HACCP. While the level of P. fluorescens on beef does not inhibit the survival of E. coli O157:H7 on aerobically stored beef, the combination of L. plantarum, and low storage temperature inhibits the survival of this pathogen on beef under vacuum storage. / Thesis (PhD)--University of Pretoria, 2011. / Food Science / unrestricted

Please read the abstract in the section 00front of this document / Dissertation (MSc (Microbiology))--University of Pretoria, 2006. / Microbiology and Plant Pathology / unrestricted

It is generally known that media containing simple sugars (sucrose, glucose) and organic nitrogen sources (ammonium) when buffered to acidic pH stimulates aflatoxin production in Aspergillus flavus & A. parasiticus while lactose, nitrate and an alkaline pH inhibit aflatoxin biosynthesis. It has been shown that pH of the growth medium is the most important regulatory factor for aflatoxin biosynthesis since media containing stimulatory carbon and/or nitrogen sources (sucrose and ammonia) do not enhance aflatoxin (or sterigmatocystin) production at alkaline pH. RNA interference (in A. flavus) of the pH regulatory transcription factor, PacC, resulted in aflatoxin production under acidic and alkaline pH conditions whilst wildtype Aspergillus flavus produced aflatoxins only under acidic conditions. This conclusively proved that PacC negatively regulates aflatoxin production at alkaline pH in A. flavus. However the exact mechanism involved in PacC repression of aflatoxin biosynthesis at alkaline pH still remains unknown. The AflR protein is essential for expression of several genes in the aflatoxin biosynthetic cluster. In the current study, sequence analysis of the aflR promoter indicated the presence of two putative PacC binding sites within the aflR promoter of A. flavus 3357WT located at positions -162 and -487 bp from the start codon. The presence of the PacC binding sites in the aflR promoter indicated a possible link between aflR expression and PacC regulation under alkaline conditions. Thus, in this study, it was hypothesized that at alkaline pH, PacC inhibits aflR expression by binding to one or both of the PacC binding sites within the aflR promoter. This in turn, would result in inhibition of aflatoxin biosynthesis since expression of several aflatoxin biosynthetic pathway genes is dependent on activation by AflR. The aim and objective of this study was to test the validity of this hypothesis i.e. that at alkaline pH PacC binds to one or both of its recognition sites within the aflR promoter thereby inhibiting aflR expression which subsequently would result in inhibition of aflatoxin biosynthesis. This was done by first mutating each individual and then both PacC binding sites in the A. flavus 3357 aflR promoter via Single-Joint PCR (SJ-PCR) and fusing the wildtype and each mutated aflR promoter to the Green Fluorescent Protein (gfp) gene and the trpC terminator to yield a functional expression vector. These constructs were then transformed into A. flavus 3357.5. Positive transformants were confirmed to express GFP by fluorescence microscopy and spectrofluorometry. Quantification of GFP protein levels of the various transformants in this study indicated that PacC negatively regulated aflR promoter activity at alkaline pH. RT-qPCR was performed on positive transformants after growth on SLS medium at acidic and alkaline pH to determine if PacC negatively regulated aflR promoter activity at alkaline pH and to determine whether PacC binds preferentially to one or both recognition sites within the aflR promoter. RT-qPCR analysis suggest that PacC binds non-preferentially to both recognition sites within the aflR promoter on sucrose and lactose media at alkaline pH, although mutation of PacC binding site 2 results in a slightly higher expression compared to mutation of PacC binding site 1. Increasing the concentration of an aflatoxin conducive nitrogen source stimulated aflR promoter activity but this was not sufficient to overcome negative regulation by PacC. It is generally known that repression of aflR expression results in repression of aflatoxin biosynthesis irrespective of pH. The results of this study strongly suggest that PacC negatively regulates aflR promoter activity at alkaline pH by binding to one or both PacC recognition sites within the aflR promoter. Since aflR promoter activity is repressed by PacC at alkaline pH, this substantiates the hypothesis that PacC represses aflatoxin biosynthesis by inhibiting expression of aflR. Furthermore, the results of this study indicated that there may be some PacC protein present in the active form at acidic pH irrespective of the carbon source and nitrogen source used in the growth medium. RT-qPCR analysis indicated that any active PacC present at acidic pH may cause repression of the aflR promoter based on the position of the PacC binding site relative to the aflR start codon, although it appears that PacC may have a higher affinity for PacC binding site 2 (which is closer to the aflR start codon).

Um problema na indústria coureira é a deterioração de peles e couros devido ao desenvolvimento de microrganismos no processamento do couro. As peles e os couros contêm nutrientes adequados para o crescimento de microrganismos, como carboidratos, gorduras e proteínas, além das condições ambientais, alta umidade, temperatura de armazenagem e pH favoráveis. Alguns gêneros de bactérias e fungos sintetizam importantes substâncias deste substrato, causando modificações prejudiciais na superfície do couro e nas propriedades físico-mecânicas, deixando manchas pigmentadas de difícil remoção, afetando a qualidade do produto final e causando perda de valor comercial. Desta forma, surge a necessidade de desenvolver estratégias de controle dos microrganismos de modo a reduzir ou eliminar este problema. Para tanto, recorre-se comumente à utilização de microbicidas. No passado, a ação esperada dos agentes antimicrobianos era principalmente de fornecer uma proteção eficaz, mas em anos mais recentes, a preocupação com a sua toxicidade e com potenciais riscos ecológicos tornou-se também importante. Nos dias atuais uma grande preocupação mundial é o cuidado com a preservação do meio ambiente. Devido a isto, várias pesquisas estão voltadas para o desenvolvimento de novas tecnologias limpas e renováveis como também a otimização de processos. Tendo em vista a melhoria de processos no que diz respeito ao uso de microbicidas adicionados em peles e couros, para prevenir a contaminação dos mesmos por microrganismos, esta dissertação centrou-se na avaliação do desempenho de microbicidas comerciais convencionalmente utilizados na indústria do couro sendo eles 2-(tiocianometiltio) benzotiazole (TCMTB), isotiazolina, dispersão oleosa de 2-n-octil-4-isotiazolin-3-ona + carbendazim (OIT+BMC/óleo), dispersão aquosa de 2-n-octil-4-isotiazolin-3-ona + carbendazim (OIT+BMC/água), 2-n-octil-4-isotiazolin-3-ona (OIT) e para-cloro-meta-cresol (PCMC) contra as espécies de bactérias Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa e Streptomyces sp. e as espécies de fungos Aspergillus niger, Aspergillus flavus, Penicillium herguei e Penicillium chrysogenum. Os microbicidas foram aplicados nas etapas de remolho, píquel, curtimento de couro com cromo e curtimento/engraxe com tanino vegetal. Os efeitos antimicrobianos dos microbicidas foram avaliados através de ensaios microbiológicos acelerados de plaqueamento e de acondicionamento em câmara tropical e testes de biodeterioração no solo, seguidos de análises visual, Microscopia eletrônica de varredura e ensaio de tração. Também foi testada a sorção e wash-out dos microbicidas em couros wet-blue. Outro teste feito nos próprios microbicidas foi o de concentração inibitória mínima (MIC). Os resultados demonstraram baixa capacidade antibacteriana e antifúngica dos microbicidas selecionados quando aplicados no processo de remolho contra o ataque das bactérias Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa, Streptomyces sp. e no processo de engraxe para o couro curtido com tanino vegetal contra o ataque dos fungos Aspergillus niger, Aspergillus flavus. Dois dos microbicidas estudados, TCMTB e OIT+BMC/água aplicados no couro wet-blue, revelaram elevada capacidade antifúngica contra os quatro diferentes fungos testados. Dos microbicidas submetidos ao teste de absortividade e wash-out, o microbicida à base de TCMTB apresentou alta e rápida absortividade pelo couro wet-blue, além de possuir resistência à lavagem. / A problem in the leather industry is the deterioration of leather skins due to the development of microorganisms, in the processing of leather. The skin and leather containing nutrients suitable for the growth of microorganisms such as carbohydrates, fats and proteins , as well as environmental conditions, high humidity, storage temperature and pH favorable. Some genera of bacteria and fungi synthesize important ingredients of this substrate, causing harmful changes in the surface of the leather and the physical and mechanical properties, leaving pigmented spots are difficult to remove, affecting the quality of the final product and loss of commercial value. Thus, there arises the need to develop strategies for control of microorganisms in order to reduce or eliminate this problem, therefore, appeal commonly the use of microbicides. In the past, the expected action of antimicrobial agents was mainly to provide effective protection, but in more recent years, concerns about the toxicity and potential ecological risks has also become important. Nowadays a major global concern is the careful preservation of the environment, due to this many researches are focused on the development of new clean and renewable technologies as well as process optimization. In view of the improvement of processes in respect to the use of microbicides added to hides and skins to prevent contamination thereof by microorganisms , this work has focused on the evaluation of the performance of commercial microbicides conventionally used in the leather industry, 2-metiltiocianato benzothiazole (TCMTB) isothiazoline, oily dispersion of 2-n-octil-4-isotiazolin-3-ona + carbendazim (OIT+BMC/oil), water dispersion of 2-n-octil-4-isotiazolin-3-ona + carbendazim (OIT+BMC/water), 2-n-octil-4-isotiazolin-3-ona (OIT) and para-chloro-meta-cresol (PCMC), against the bacterial species Bacillus subtilis, Escherichia coli and Pseudomonas aeruginosas and Streptomyces sp. e species of fungi Aspergillus niger, Aspergillus flavus, and Penicillium herguei Penicillium chrysogenum, compared with the control. Microbicides were applied in steps of soaking, pickling, chrome tanning and grease/tanning with vegetable tannin. The antimicrobial effects of microbicides made for these applications were evaluated by accelerated plating microbiological testing and tropical chamber rain and biodegradation tests on the ground, followed by analysis (visual , SEM and tensile test) . Also was tested the absorptivity and wash-out of microbicides in wet-blue leather. Another test done on their own microbicides was the minimal inhibitory concentration (MIC). The results showed low capacity antibacterial and antifungal activities of selected microbicides when applied in the process of soaking the attack of the bacteria Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa, Streptomyces. sp. e in the process of grease for leather vegetable tannin against fungal attack Aspergillus niger, Aspergillus flavus. Two of microbicides studied TCMTB and OIT + BMC/water applied in wet-blue leather, high capacity antifungal against revealed four different fungi tested. For microbicides tested for absorbency and wash-out the microbicide based TCMTB showed high and rapid absorbency by wet-blue leather also has resistance to washing.

The development of an automated instrument employing reflectance colorimetry was described. Several models were designed, assembled, and programmed to perform microbial and enzymic tests automatically. Samples were prepared manually or automatically by a Zymate™ II robot. These samples were incubated during the tests to maintain an optimum temperature for reactions and microbial growth. During incubation, color changes of appropriate indicator dyes in the sample/reagent mixtures were measured intermittently, recorded, and compared to previously defined end points. The computer-controlled instrument received data that related time of color changes with the initial numbers of microorganisms or the enzyme activity of the samples. Traditional pH and oxidation/reduction dyes were used. Suitable dyes and media were selected for fast estimation in the different assays studied . Applications of the instrument to evaluate raw milk for the total viable microbial count, abnormality, broad spectrum antibiotics, and coliforms were emphasized. The automated colorimeter system successfully quantitated total and coliform microflora in raw milk. Correlations between reflectance colorimetry and the spiral plate count method were .932 (using .12% TIC as indicator), .922 (using BCP as indicator), and .681 (using .04% TTC as indicator). A coefficient of correlation of .874 was obtained when reflectance colorimetry was compared with coliform numbers on violet red bile agar. The reflectance colorimetry system provided better precision than current reference methods. Preliminary incubation or larger sample volumes were required to estimate low numbers of microflora . Antibiotic residue detection was also evaluated using Lactococcus lactis ssp. cremoris UC 310+ with the automated colorimeter system. The following concentrations (ppb) could be detected: penicillin G:::;; 5, ampicillin ≤ 5, tetracycline ≤ 250 , sulfamethazine ≤ 30, streptomycin ≤ 1000, kanamycin ≤ 500, and chloramphenicol ≤ 500. Abnormal milk could be screened out by measuring the NAGase activity and chloride ion content in milk samples. Both methods had been integrated into the automated colorimeter system. The coefficient of correlation between somatic cell count and the NAGase activity as measured with the colorimeter was .802; a correlation of .792 could be obtained when chloride ion content was measured.

<p dir="ltr"><i>Pseudomonas aeruginosa</i> (<i>P. aeruginosa</i>) is a Gram-negative bacterium responsible for many infections in immunocompromised humans. This multi-drug resistance human pathogen can form biofilms, which help protect it from not only clinical treatment but also from main homeostasis and metabolism. Understanding biofilm structures is critical to help combat biofilm formation and develop better ways to treat <i>P. aeruginosa</i> infections. A molecule that helps biofilm formation and virulence infections for <i>P. aeruginosa</i> is pyocyanin, which is believed to be correlated with the invasiveness of the bacteria and the stabilization of biofilms. To better understand the role of pyocyanin in assisting <i>P. aeruginosa</i> with survival, we applied optical imaging to study pyocyanin in biofilms and under blue light treatment. Using nonlinear optical imaging methods, we could successfully detect the aggregation of pyocyanin in biofilms. Furthermore, we discovered that pyocyanin protects <i>P. aeruginosa</i> from blue light inactivation. In addition, we found that blue light treatment alters the structure of pyocyanin, leading to irreversible changes that produce distinct spectra in UV-Vis and fluorescence signals. <i>These results provide new insights into how pyocyanin protects </i><i>P. aeruginosa</i> in blue light treatment. Further investigation would lead to better treatment strategies for more effective treatment of <i>P. aeruginosa</i> and biofilms for various infections.</p>

Whole cell biosensors have been extensively used for monitoring toxicity and contamination of compounds in environmental biology and microbial ecology. However, their application in the pharmaceutical and cosmetics industries for preservative efficacy testing (PET) has been limited. According to several pharmacopoeias, preservatives should be tested for microbial activity using traditional viable count techniques; the use of whole cell microbial biosensors potentially provides an alternative, fast, and efficient method. The aim of the study was to construct and develop whole cell microbial biosensors with Pseudomonas aeruginosa ATCC 9027. Constitutive promoters: PlysS, Pspc, Ptat, Plpp and PldcC and the lux-cassette were inserted into plasmid pME4510 and transformed into P. aeruginosa ATCC 9027 cells to produce bioluminescent strains. Plasmids were found to be maintained stably (~50 copies per cell) throughout the growth and death cycle. The novel bioluminescent strains were validated in accordance with the pharmacopoeia using bioluminescence detection and quantification followed by comparison with the traditional plate counting method. The bioluminescent method was found to be accurate, precise and equivalent at a range of 103 – 107 CFU/mL, as compared with plate counting. Recovery of bacterial cells was quantified using bioluminescence; this method proved to be accurate with percentage recoveries between 70-130% for all bioluminescent strains. The method was also more precise (relative standard deviation less than 15%) than the traditional plate counting method or the ATP bioluminescent method. Therefore, the bioluminescent constructs passed/exceeded pharmacopoeial specified criteria for range, limit of detection, accuracy, precision and equivalence. Physiology of the validated bioluminescent strains was studied by assessing the growth and death patterns using constitutive gene expression linked with bacterial replication. Promoter strengths were evaluated at various stages of the growth and death pattern and related to promoter sequences. PlysS, Ptat and Plpp were relatively strong promoters whilst PldcC and Pspc were relatively weak promoters. Relative promoter strength decreased in the order of Plpp>Ptat>PlysS>PldcC>Pspc during the exponential phase whilst Ptat was stronger than Plpp during the stationary phase of growth. Plpp had its highest level of expression during the exponential phase, while Ptat had relatively stable lux expression during the stationary phase. Correlations between relative bioluminescence and CFU at 24 hours were greater than 0.9 indicating a strong relationship for all bioluminescent strains. Reduction in correlation coefficients to approximately 0.6 between relative bioluminescence and CFU and between relative fluorescence and CFU beyond 24 hours indicated that a certain proportion of cells were viable but non-culturable. Tat-pME-lux showed steady bioluminescence compared to CFU count (R>0.9) throughout 28 days of growth. Equivalence analysis showed no significant difference between the bioluminescence and plate count method throughout 28 days of growth for all five bioluminescent strains. Applicability of these novel bioluminescent strains was evaluated for preservative efficacy tests (PET) using bacterial replication and bioluminescence as a measure of constitutive gene expression. PET using benzalkonium chloride and benzyl alcohol showed no significant difference between the bioluminescent method and the plate count method. Good correlations between bioluminescence, CFU count and fluorescence were obtained for benzalkonium chloride (BKC) concentrations (R>0.9) between 0.0003% and 0.0025% against strains lysR25, lppR4 and tatH5. Similarly, good correlations (R>0.9) between the three parameters were obtained for benzyl alcohol (BA) concentrations between 0.125% and 2% against strains lysR25, lppR4 and tatH5. The bioluminescent method and traditional plate counting method were equivalent for concentrations of BKC (0.0003 - 0.02%) and BA (0.25 - 2%) during preservative efficacy tests. These bioluminescent constructs therefore are good candidates for selection for preservative efficacy testing. The bioluminescent method and traditional plate counting method were also found to be equivalent for construct tatH5 at a concentration of 0.125% BA. PET testing with BKC and BA showed that tatH5-pMElux (R>0.9) had consistently high correlation coefficients between CFU and relative bioluminescence. Together with the results from growth and death kinetics, where tatH5 showed the greatest constitutive expression, it can be concluded that P. aeruginosa ATCC 9027 tatH5-pMElux is the best construct for testing various antimicrobial agents. This study has shown that according to the pharmacopoeial requirements, the bioluminescent method is more accurate, precise and equivalent to the traditional plate counting method and therefore can be utilised instead of the traditional plate counting method for the purpose of preservative efficacy testing.

Surtos de listeriose têm sido causa de preocupação para indústrias de alimentos e profissionais da saúde. Os testes de rotina para detecção de Listeria sp em alimentos são demorados e envolvem o uso de meios de cultura seletivos para enriquecimento e semeadura. Neste estudo, a presença de Listeria sp, em amostras de produtos cárneos, foi investigada por testes imunológicos rápidos (Tecra® Listeria VIA, Tecra® Listeria UNIQUE e BioControl VIP® para Listeria) e pelo método clássico (Health Protection Branch, Canadá). A concordância entre o método clássico e os testes rápidos foi estabelecida com limite de 95% de confiança. Verificou-se que os testes VIA e VIP® são de fácil execução e rápidos, além de apresentarem desempenho comparável ao do método clássico, independentemente do alimento avaliado. O teste UNIQUE apresentou desempenho variável com a amostra e gerou um número elevado de resultados positivos falsos, o que dificulta seu emprego. / Outbreaks of human listeriosis have been a cause of concern to the food industry and health professionals. The routine methods for detecting Listeria sp in foods are time-consuming and involve the use of selective enrichments and culturing on selective agars. In this study, the presence of Listeria sp in 120 meat and meat products samples was investigated by three rapid immunoassays (Tecra® Listeria VIA, Listeria UNIQUE and BioControl VIP®.+ for Listeria) and a cultural procedure. The detection of Listeria sp by the cultural method was done according to Canada\'s Health Protection Branch Method and the detection using rapid tests was done following the manufacturers\' instructions. The agreement between cultural and rapid tests was established at a confidence limit of 95%. Eighty-one samples (67.5%) were Listeria sp positive by at least one of the four methods. L. monocytogenes was present in 49.2% of the samples. Sixty-four samples (53.3%) were positive by the cultural procedure. For the rapid tests, 62 (51.7%) were VIA positive, 65 (54.2%) were VIP® positive and 41 (50.2%) were UNIQUE positive. Fifty-five samples (45,8%) were positive by the cultural method and VIA simultaneously, 55 (45.8%) by the cultural method and VIP® and 35 (43.2%) by the cultural and UNIQUE. VIA detected 4 positive samples that were not detected by any of the other methods, while VIP detected 7 positive samples and UNIQUE only 2. VIA and VIP® tests are fast and easy to execute. They also performed similarly to the cultural method despite the food matrix. UNIQUE, on the other hand, was strongly influenced by the sample type and generated a high number of false positive results what makes its use unpractical.