The aim of this work is to research milk and kefir microbiological indicators according to microbiological risk factors in the milk processing company A. Work tasks: to analyze drinking milk, ultra high temperature pasteurized milk and kefir descriptions and factors of the risk in technological process schemes; microbiological factors of the risk control schemes; samples of these products microbiological indicators according to company’s self-control plan. Conclusion: The main microbiological risk factors in drinking milk is total bacterial count, coliform bacteria, Salmonella; in the kefir – coliform bacteria, yeasts and moulds. Principals control points in drinking milk, ultra high temperature pasteurized milk and kefir technological schemes are: reception of the raw cow’s milk; freezing of the raw cow’s milk, storage and keeping; pasteurization of milk and freezing, pasteurized milk compound storage and keeping; milk warming, deaeration, homogenization, sterilization and refreshing (only for ultra high temperature pasteurized milk); repeated thermal treatment, cooling up to inoculation temperature (only for kefir); inoculation and incubation (only for kefir); storage. In the company’s A self-control plan preview principal control points microbiological risk factors are controlled by keeping a check of heating and freezing stages temperature and duration, monitoring of microbiological parameters are made not less than 4 times per year. Coliform bacteria and total bacteria... [to full text]

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.

No description available.

The pervious pavement system has been identified as an effective source control device capable of removing urban stormwater pollution by trapping pollutants within the system and biodegradation. Recent studies have further demonstrated that the pervious pavement system could be used as a source of renewable energy capable of reducing household energy bill by about 80%. In view of ever increasing demand for water and the continued reduction in available fresh water resources in the world, stormwater has been recognized as a potential valuable source of water which could be harnessed. The overall aim of this multi disciplinary research was to evaluate the suitability of a modified pervious pavement system (PPS) for water harvesting and re-use, particularly focussing on potential third world applications and taking advantage of the latest developments in materials that are available for such applications. The aim was a holistic one in which water re-use was examined in terms of both the potential advantages from an irrigation point of view without ignoring the very important public health concerns that are often of concern when water is stored in circumstances which do not fit the normally used criteria for potable supplies. The results of this study confirmed the pollution control capability of the porous pavement system as earlier determined by previous studies. Also, a novel experimental rig was designed to reproducibly create very high and realistic rainfall events over model pavement structures. Furthermore, the performance of a new geotextile, Inbitex Composite® in the pervious pavement system was determined for the first time. Furthermore, this study also tested for the first time, the performance of a pervious pavement system modified by the incorporation of Inbitex Composite® geotextile with slits and made prescriptions as to how this new geotextile could be best installed in a modified pervious pavement system in order to achieve high infiltration without compromising pollution control. This study tested the practical use of the pervious pavement system for water harvesting and storage for reuse in irrigation. In order to achieve this, the author took what could be considered as a holistic approach to water quality issues and determined the chemical, electrochemical and microbiological quality of water stored in the system as well as investigated the public health concern of the potential of pathogenic organisms in waters stored in unconventional water storage system as the pervious pavement system. It also determined that the pervious pavement system have the capability to recycle water with physical, chemical and microbiological qualities that will meet international standards for irrigation and that the system does not offer a conducive environment for potential pathogenic organisms if contamination incident occurs from adjoining areas. This study also became the first to practically relate Sustainable Urban Drainage System (SUDS) to agricultural benefit by demonstrating how a SUDS device (pervious pavement system) could be used in addition to its urban drainage control role, as a source of supply of high quality irrigation water to cultivate crops fit for human and animal consumption despite high application of pollutants. This study determined contrary to the observation of earlier studies that the use of slow-release iv fertilizer could lead to eutrophication problems in cases where the water is channeled to natural water courses. Furthermore, active response of potential pathogenic bacteria to the presence of slow-release fertilizer was observed in this study. This raises a huge question on the need to add fertilizer to the pervious pavement system. Coupe, (2004) had demonstrated that oil degrading microbes would respond positively to food sources in the system and hence, there was no significant need for simulation by nutrient addition, the author concluded in the study presented here that fertilizer addition should only be conducted if the waters are to be used for irrigation where the nutrients would be beneficial to the plants and that even in this case, the microbiological water quality should be constantly monitored and the addition suspended if the risk of contamination from adjoining areas is high.

Thesis (MSc)--University of Stellenbosch, 2004. / ENGLISH ABSTRACT: The complexity of microbial systems has presented serious obstacles to the quantification of fermentation processes. Using computer modelling techniques progress has been made in monitoring, controlling and optimising microbial systems using material balancing techniques and empirical process models. The Monod equation is among the most commonly used models and is based on empirical findings with no mechanistic basis. Monod presents a simple model to describe the growth of a cell in a defined nutrient environment. The Monod equation is mathematically analogous to the formula that was proposed by Michaelis and Menten to describe enzyme kinetics. Both equations describe a hyperbolic function with a half-saturation constant (K_s in the monod equation and K_m in the Michaelis Menten equation) but the meaning of the two saturation constants K_s and K_m is different. In number of studies K_s and K_m are used as if they are equivalent. In contrast to Michaelis-Menten kinetics, which describes a process catalysed by a single enzyme, Monod kinetics describes an overall process involving thousands of enzymes. The Monod equation describes the specific growth rate of a microbial cell as the function of a limiting substrate concentration. The aim of this study was to test this principle, for Saccharomyces cerevisiae VIN13 under glucose limited aerobic chemostat conditions. The VIN13 was observed to follow the Monod description and when compared with other growth kinetic models gave one of the best fits to the data. A functional relationship between the half-saturation constant, K_s, and Michaelis Menten constant, K_m, was there after derived. This was achieved by using metabolic control analysis (MCA) to explain when K_m of the transporter becomes equal to the K_s. Using the deductions obtained from MCA a core kinetic model was then formulated to demonstrate that the K_s can either be smaller, equal or higher than the K_m of the transporter, depending on the flux control distribution in the model. / AFRIKAANSE OPSOMMING: Die kwantifisering van fermentasieprosesse word ernstig belemmer deur die kompleksiteit van mikrobiale sisteme. Deur gebruik te maak van rekenaar-ondersteunde modelleringstechnieke vir die opstelling van massa balans vergelykings en empiriese prosesmodelle is vordering gemaak in die waarneming, beheer en optimalisering van mikrobiale sisteme. Die Monod vergelyking is een van die mees gebruikte groeimodelle en is gebaseer op empiriese bevindings - die model het nie ‘n meganistiese grondslag nie. Die Monod vergelyking is wiskundig ekwivalent aan die vergelyking wat opgestel is deur Michaelis en Menten vir die beskrywing van ensiemkinetika. Beide vergelykings beskryf ‘n hyperboliese kurwe met ‘n konstante wat die halfversadigingswaarde aangee vir substraat (Ks in die Monod vergelyking en Km in die Michaelis-Menten vergelyking), maar die betekenis van die twee versadigingskonstantes is verskillend. In ‘n aantal studies word die Ks en Km waardes gebruik asof hulle gelyk is aan mekaar. In teenstelling met die Michaelis- Menten kinetika wat ‘n enkel ensiem-gekataliseerde reaksie beskryf, beskryf die Monod vergelyking ‘n proses wat duisende ensieme behels. Die Monod vergelyking beskryf die spesifieke groeitempo van ‘n bakteriële sel as ‘n funksie van die beperkende substraatkonsentrasie. Die doel van hierdie studie was om hierdie beginsel te toets vir Saccharomyces cerevisiae VIN13 wat onder glukose beperkte, aerobiese kondisies in ‘n chemostat gekweek word. Die VIN13 groei kon goed beskryf word met die Monod model, wat in vergelyking met ander groeimodelle een van die beste passings vir die meetpunte het gegee. Vervolgens is ‘n funksionele verwantskap afgelei tussen Ks en Km; deur gebruik te maak van metabole kontrole analise (MCA) kon verduidelik word wanneer die Ks gelyk is aan die Km van die transporter vir die beperkende substraat. Deur gebruik te maak van die MCA analise is ‘n eenvoudige kinetiese model opgestel om aan te toon dat die Ks kleiner, gelyk aan of groter kan wees as die Km van die transporter, afhanklik van die fluksie-kontrole verdeling in die model.

Thesis (PhD)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: The wildlife industry in Namibia continues to grow as the production and consumption of game meat increases. However, the health risks posed by the trade in wildlife and related by-products to livestock and humans have not been fully assessed. The main objective of this study was to investigate the potential health risks related to the increased consumption of game meat and relevant by-products by assessing the quality of game meat, as well as determine the role of game meat species in the transmission of zoonoses. The microbiological quality and safety of export game meat was assessed. No differences in the aerobic plate count (APC) were observed between the years (2009 and 2010), but the mean Enterobacteriaceae count was 1.33±0.69log10 cfu/cm2 compared to 2.93±1.50log10 cfu/cm2 between the years. Insignificant heterotrophic plate count (HPC) levels were detected in 9/23 field water samples, while faecal bacteria (coliforms, Clostridium perfringens and enterococci) were not isolated in all samples. Seven serogroups, with the exception of O26, were detected in exotic species. A white tailed deer sample had a serotype belonging to O45 which confirmed positive for stx1 gene. In springbok, 5/15 pools of faecal samples tested positive for the intimin gene. No Salmonella spp were isolated, and all E. coli isolates from the meat samples were negative for STEC virulence genes (i.e. stx1, stx2, eae and hlyA). A linear regression analysis was conducted on selected variables to identify the main predictors and their interactions affecting pH of meat 4 hours post-slaughter. In an increasing order of magnitude during winter time, the pH reached at 16-36hr post slaughter in springbok heart, liver, spleen, kidney and lungs was significantly higher than pH 6.0, while no significant differences were observed from the regulatory reference (pH 6.0) in the heart. There was a positive association between the pH of game meat 4hr post-slaughter, and liver congestion. The pH of game meat 4hr post slaughter, increased by 0.11 units per mL increase in liver congestion, and decreased by 0.04 units per minute increase in the shooting to bleeding interval, irrespective of the species. Worm eggs of strongylids, Strongyloides papillosus, Toxocara spp, Trichuris spp and coccidia were found in variable numbers in both springbok and gemsbok faeces, indicating a potential risk of transmission to other species in the ecosystem. On examination of carcasses, a novel parasite, Skjabinodera kuelzii, was identified and noted to be associated with inguinal fascia and renal fat, but the public health significance remains unclear. Nevertheless, S. kuelzii should be considered as of potential significance during routine game meat inspection. A total of 12 310 springbok were harvested from 26 commercial farms over a period of two years. Tissue samples (i.e. 60 livers, 41 kidneys and 52 hindquarter muscles) were collected from randomly selected healthy animals. The mean values (i.e. above the detection limit) of cadmium (Cd) and lead (Pb) were 0.10±0.05mg/kg and 1.04±0.21mg/kg in the liver, respectively; and 0.33±0.22mg/kg and 0.905±0.51mg/kg in the kidney of springbok, respectively. The levels of cadmium and lead in the hindquarter muscles were below the detection limit. Serum samples (n=1692) collected from sheep, goats and cattle from four presumably at-risk farms, and 900 springbok (Antidorcas marsupialis) serum samples collected from 29 mixed farming units, were screened for Brucella antibodies by using the Rose-Bengal test (RBT). Positive cases were confirmed by complement fixation test (CFT). To assess the prevalence of human brucellosis, 137 abattoir employees were tested for Brucella antibodies using the standard tube agglutination test (STAT), and the enzyme linked immunosorbent assay (ELISA). Cattle and sheep from all four farms were negative by RBT and CFT, but two of the four farms carried 26/42 and 12/285 seropositive goats, respectively. Post mortem examination of seropositive goats revealed no gross pathological lesions. Culture for brucellae from organs of seropositive animals was negative. None of the wildlife sera tested positive by either RBT or CFT. Occurrence of confirmed brucellosis in humans was linked to the consumption of unpasteurized goat milk, home-made goat cheese and coffee with raw milk and prior contact with goats. All abattoir employees (n=137) tested negative by STAT, but 3 were positive by ELISA. The three abattoir workers were clinically normal, and lacked historical connections with clinical cases. This study illustrates the importance of microbiological, parasitic and residue monitoring as critical components of a hazard analysis and critical control point based system for game meat. The study also provides the basis for increased integrated health research, surveillance and meat safety risk analysis. / AFRIKAANSE OPSOMMING: Die Namibiese wildbedryf raak toenemend groter soos die produksie en verbruik van wildsvleis toeneem. Die verwante gesondheidsrisiko’s wat die gebruik van wildsvleis en verwante produkte vir mens en dier inhou, is nog nie volledig geassesseer nie. Die doelwit van die studie was om ondersoek in te stel na die potensiële gesondheidsrisiko's wat wildsvleis en verwante neweprodukte vir mens en dier inhou deur middel van die assessering van vleisgehalte en die bepaling van die rol van die wildsvleis spesies in die oordrag van soönoses. Die mikrobiologiese gehalte en veiligheid van uitvoer wildsvleis was geassesseer. Geen verskille in die aerobiese plaat telling (APC) vir monsters versamel tydens 2009 en 2010 is aangeteken nie. Die gemiddelde Enterobacteriaceae telling was 1.33± 0.69log10 cfu/cm2 in vergelyking met 2.93±1.50log10 cfu/cm2 tussen die jare. Onbeduidende heterotrofe plaattelling (HPC) vlakke is waargeneem in 9/23 water monsters, terwyl fekale bakterieë (d.i. kolivorme, Clostridium perfringens en enterokokke) nie in enige van die monsters geïsoleerd is nie. Sewe serogroepe, met die uitsondering van O26, is aangeteken vir die eksotiese spesies. Monsters verky van ʼn white tailed deer is as positief vir 'n serotipe van O45 getoets, en die teenwoordigheid van die stx1 geen is bevestig. In springbok het 5/15 poele van fekale monsters positief getoets vir die intimien geen. Geen Salmonella spp is geïsoleer nie en alle E. coli geïsoleer in die vleismonsters was negatief vir die Stec virulensie geen (d.i. stx1, stx2, EAE en hlyA). ʼn Liniêre regressie-analise is op geselekteerde veranderlikes wat as die belangrikste indikators kan dien, en enige moontlike interaksie wat die pH van wildsvleis 4 uur na-slag kan beïnvloed, uitgevoer. In 'n toenemende orde van grootte gedurende die winter tyd, die pH teen 16-36hr na slagting in springbok hart, lewer, milt, niere en longe was aansienlik hoër as die pH 6.0, terwyl geen beduidende verskille waargeneem is wanneer dit met die regulasie verwysingswaarde van die hart (pH 6.0) vergelyk is nie. Daar was 'n positiewe assosiasie tussen die pH van wildsvleis 4 uur na-slag en mate van aansameling in die lewer. Die pH van wildsvleis 4 uur na-slag, het toegeneem met 0.11 eenhede per mL toename in lewer aansameling en afgeneem met 0.04 eenhede per minuut toename in die skiet tot uitbloei interval, ongeag die spesie. Wurmeiers van rondewurms, Strongyloides papillosus, Toxocara spp, Trichuris spp en koksidia het in verskillende ladings in die mis van beide springbok en gemsbok ontlasting, voorgekom. Dit dui op 'n potensiële risiko van oordrag na ander spesies in die ekosisteem. Die voorkoms van ʼn nuwe parasiet, Skjabinodera kuelzii, in wildskarkasse is aangeteken en was geassosieer met inguinale fascia en renale vet, maar die openbare gesondheidsrisiko bly onduidelik. Daar word aanbeveel dat dié parasiet as ʼn potensiële risiko faktor tydens roetine vleisinspeksies beskou moet word. ʼn Totaal van 12 310 springbokke is oor 'n tydperk van twee jaar van 26 kommersiële plase geoes. Weefselmonsters (d.i. 60 lewers, 41 niere en 52 agterkwart spiere) is ewekansig versamel van gesonde diere. Die gemiddelde waardes (d.i. hoër as die opsporingslimiet) van kadmium (Cd) en lood (Pb) was 0.10 ± 0.05mg/kg en 1.04 ± 0.21mg/kg in die lewer onderskeidelik en 0.33 ± 0.22mg/kg en 0.905 ± 0.51mg/kg in die niere van springbok, onderskeidelik. Die vlakke van kadmium en lood in die agterkwart spiere was laer as die opsporingslimiet. Serum monsters (n=1692) is van skape, bokke en beeste van vier vermoedelik hoë risiko plase en springbok (Antidorcas marsupialis, n=900) van 29 gemengde boerdery sisteme versamel en getoets vir die teenwoordigheid van Brucella teenliggaampies deur middel van die Rose-Bengal-toets (RBT). Positiewe gevalle is bevestig deur die komplement binding toets (CFT). Die voorkoms van menslike brusellose is bepaal deur 137 abattoir werknemers te toets vir Brucella teenliggaampies deur gebruik te maak van die standaard buis agglutinasie toets (STAT) en die ensiembinding immunosorberende toets (ELISA). Beeste en skape van die vier hoë risiko plase het negatief getoets met die RBT en CFT metodes, maar bokke van twee van die vier plase het seropositief getoets (26/42 en 12/285 onderskeidelik). Nadoodse ondersoek van seropositief bokke het geen patologiese letsels aangedui nie. Die kultuur van orgaanmonsters van seropositief diere vir Brucellae was negatief. Die monsters versamel van wild het negatief getoets deur middel van die RBT en CFT toets metodes. Die voorkoms van brusellose in mense in die studie was geassosieer met die gebruik van ongepasteuriseerde melk, tuisgemaakte bokmelkkaas en koffie met ongepasteuriseerde melk, asook direkte kontak met bokke. Alle abattoir werknemers (n=137) het negatief getoets met die STAT metode, maar drie werknemers het positief getoets met die ELISA metode. Die drie abattoir werkers was klinies normaal en het nie vorige kontak met bevestigde kliniese gevalle gehad nie. Hierdie studie bevestig die belang van mikrobiologiese, parasitiese en residu monitering as kritieke komponente van 'n gevaar-analise en kritiese kontrolepunt gebaseerde stelsel vir die produksie en verbruik van wildsvleis. Die studie verskaf ʼn basis vir toekomstige navorsing gefokus op ʼn geïntegreerde benadering van mens- en diergesondheid, monitering en vleis veiligheid risiko-analises.

Activation of otherwise unreactive substrates is a common strategy in chemistry and in nature. Adenylate-forming enzymes use adenosine monophosphate to activate the hydroxyl of their carboxylic substrate, creating a better leaving group. In a second step this reactive group is replaced in a nucleophilic elimination reaction to form esters, amides or thioesters. Recent studies have revealed that NRPS- independent siderophore (NIS) synthetases are also adenylate-forming enzymes, but are not included in the current superfamily description. NIS enzymes are involved in biosynthesis of high-affinity iron chelators which are used for iron acquisition by many pathogenic microorganisms. This is an important area of study, not only for potential therapeutic intervention, but also to illuminate new enzyme chemistries. Here the structural and biochemical studies of AcsD from Pectobacterium chrysanthemi are reported. AcsD is a NIS synthetase involved in achromobactin biosynthesis. The co-complex structures of ATP and citrate provide a mechanism for the stereospecific formation of an enzyme-bound citryl-adenylate. This intermediate reacts with L-serine to form a likely achromobactin precursor. A detailed characterization of AcsD nucleophile profile showed that it can not only catalyze ester formation, but also amide and possibly thioester formation, creating new stereospecific citric acid derivatives. The structure of a N-citryl-ethylenediamine product co-complex identifies the residues that are important for both recognition of L-serine and for catalyzing ester formation. The structural studies on the processive enzyme AlcC, which is involved in the final step of alcaligin biosynthesis of Bordetella pertussis, show that it has a similar topology to AcsD. It also shows that ATP is coordinated in a manner similar to that seen in AcsD. Biochemical studies of a substrate analogue establish that AlcC is not only capable of synthesizing substrate dimers and trimers, but also able to assemble the respective dimer and trimer macrocycles. A series of docked binding models have been developed to illustrate the likely substrate coordination and the steps along dimerization and macrocyclization formation. Structural and mechanistic comparison of NIS enzymes with other adenylate-forming enzymes highlights the diversity of the fold, active site architecture, and metal coordination that has evolved. Hence, a new classification scheme for adenylate forming enzymes is proposed.

O objetivo deste estudo foi avaliar o efeito inibitório de Lactobacillus rhamnosus sobre as contagens de Staphylococcus aureus e Listeria monocytogenes, aspergidos isoladamente ou em combinação sobre a superfície de queijo Minas Frescal, durante armazenamento por 21 dias a 7ºC. O delineamento consistiu em esquema fatorial 2x2x2, sendo 8 tratamentos com 4 repetições. As características físico-químicas (pH, atividade de água, umidade, teor de gordura, proteína e perfil de textura) foram determinadas nos queijos dos tratamentos sem adição de L. rhamnosus ou contendo este probiótico (T1 e T2, respectivamente). Verificaramse as contagens de L. rhamnosus, S. aureus e L. monocytogenes nos queijos de todos os tratamentos nos dias 1, 7, 14 e 21 de armazenamento. Foram também analisados os percentuais de sobrevivência dos microrganismos submetidos a condições de simulação do trato gastrointestinal (TGI) utilizando ensaios in vitro. Não houve efeito significativo (P>0,05) entre os parâmetros físico-químicos dos queijos dos tratamentos T1 e T2. As contagens de L. rhamnosus aumentaram (P<0,05) em todos os tratamentos a partir do dia 7 de armazenamento, estabilizando ao redor de 108 UFC/g, sendo que a presença concomitante de L. monocytogenes e/ou S. aureus nos queijos não influenciou a contagem de L. rhamnosus. L. rhamnosus diminuiu em cerca de 1 ciclo log as contagens de L. monocytogenes, e não exerceu efeito inibitório sobre S. aureus após 21 dias. S. aureus não sobreviveu ao teste de simulação ao TGI. No entanto, L. rhamnosus e L. monocytogenes apresentaram percentuais de sobrevivência entre 74,6% a 86,4%, e entre 75,8% a 94,1%, respectivamente. Os resultados demonstraram que a adição de L. rhamnosus não alterou as características físico-químicas dos queijos Minas frescal, porém exerceu efeito inibitório sobre L. monocytogenes, mas nenhum efeito sobre S. aureus. A utilização de L. rhamnosus como probiótico apresenta um potencial para inibição de L. monocytogenes na fabricação de queijos Minas frescal. São necessários estudos sobre os mecanismos envolvidos na competição entre as bactérias por substratos no alimento, bem como sua sobrevivência nas condições do TGI em ensaios in vivo. / The aim of this study was to evaluate the effects of Lactobacillus rhamnosus on growth of Staphylococcus aureus and Listeria monocytogenes in Minas frescal cheese during 21 days of storage at 7ºC. The experimental design was totally randomized, in a 2x2x2 factorial arrangement with 8 treatments and 4 replicates per treatment. Physical chemical parameters such as pH, moisture, water activity, fat, protein and texture profile analysis were carried out in cheeses where no microorganism were inoculated (T1) and in the cheeses inoculated with the probiotic bacteria, L. rhamnosus (T2). The counts of L. rhamnosus, S. aureus and L. monocytogenes were examined on days 1, 7, 14, 21 of storage. Survival percentage of the bacteria after exposure to simulated gastrointestinal conditions was studied in vitro. Statistical analysis indicated that there were no significant differences (P>0,05) among the means of the physical chemical parameters analyzed in treatments 1 and 2. From day 7 on, the counts of L. rhamnosus increased (P<0,05) in all treatments, stabilizing and reaching up to 108 CFU/g. It was noticed that the concurring presence of L. monocytogenes and/or S. aureus in the cheese samples did not show influence in the counts of the probiotic bacteria. The L. rhamnosus caused about 1 log cycle reduction in the counts of L. monocytogenes, but showed no inhibitory effect on S. aureus at the end of the period of storage. S. aureus did not survive the exposure to simulated gastrointestinal conditions. However, L. rhamnosus and L. monocytogenes showed survival percentages varying from 74,6% to 86,4%, and from 75,8% to 94,1%, respectively. The results showed that the addition of L. rhamnosus had no influence on the physical chemical characteristics of the Minas frescal cheese and no inhibitory effect on S. aureus, nevertheless demonstrated inhibitory effect on L. monocytogenes. The addition of probiotic strains of L. rhamnosus in Minas frescal cheese represents potential for L. monocytogenes inhibition. It is essential to carry out studies on the mechanisms involved in the competition for substrate by bacteria, as well as their survival to simulated gastrointestinal conditions in in vivo experiments.

A linfadenite caseosa é uma doença infectocontagiosas, de ocorrência mundial, que acomete caprinos e ovinos, caracterizada pela formação de abscessos em gânglios linfáticos superficiais, e alguns casos órgãos e linfonodos internos. É uma enfermidade causada pelo Corynebacterium pseudotuberculosis, responsável por grandes perdas econômicas na caprinocultura e ovinocultura. O presente trabalho teve por objetivo desenvolver um teste sorológico sensível e específico para detectar anticorpos anti-C. pseudotuberculosis em ovinos. Os animais foram classificados em dois grupos, com sintomatologia aparente para linfadenite caseosa (n=103) colhidos a campo, onde foram coletados amostras de soro e punção de linfonodos aumentados; e animais sem sintomatologia aparente (n=50) dos quais as amostras de soro e fragmentos de pulmão e linfonodo mediastínico foram colhidos em frigorífico. Em ambas as amostras a confirmação da presença e ausência da doença foi realizada através das provas de cultivo microbiológico e PCR, considerados nesse estudo como padrão para classificação dos animais. O resultado do cultivo microbiológico dos animais com sintomatologia aparente foi 53,5% (55/103) identificadas como C. pseudotuberculosis através de provas bioquímicas, e com a utilização da PCR, 46,5% (48/103) das amostras foram positivas para C. pseudotuberculosis. Com relação aos animais sem sintomatologia aparente, todas as amostras foram negativas no cultivo microbiológico e PCR (0/50). Na padronização do ELISA indireto foram utilizados 42 soros positivos e 43 soros negativos confirmados no cultivo microbiológico e PCR para linfadenite caseosa. A média de absorbância foi 1,88 com desvio padrão de 0,43, para as amostras positivas e, para as amostras negativas a média de 0,71 e desvio padrão 0,18. Dessa forma foram consideradas positivas, as amostras que apresentaram na reação de ELISA valor da DO> 1,1 e negativas de <1,1. A análise gráfica empregada no presente trabalho, curva ROC, permitiu encontrar o valor do ponto de corte associado à combinação dos parâmetros de sensibilidade e especificidade, assim, a acurácia do teste pôde descriminar indivíduos doentes de não doentes. A utilização de técnicas sorológicas no diagnóstico da linfadenite caseosa permitirá o controle epidemiológico da doença, porém não substitui o cultivo microbiológico, técnica considerada padrão ouro, e pode ser utilizada como teste de triagem ou mesmo na comercialização de animais, visto que a doença muitas vezes é de caráter inaparente, o que inviabiliza o diagnóstico clínico e microbiológico. / Caseous lymphadenitis is an infectious disease of worldwide occurrence that affects sheep and goats, characterized by the formation of abscesses in superficial lymph nodes, and sometimes internal organs and lymph nodes. It is caused by Corynebacterium pseudotuberculosis, responsible for great economic losses in goat and sheepproduction. This study aimed to develop a sensitive and specific serological test to detect anti- C. pseudotuberculosis in sheep. The animals were divided into two groups, the first one encompassing the ones with apparent caseous lymphadenitis symptoms (n = 103), where serum samples and puncture of enlarged lymph nodes were collected, and the second one with animals with no apparent symptoms (n = 50) of which samples (lung fragments, serum and and mediastinal lymph nodes) were inspected and harvested in the slaughterhouse. In both groups the presence and absence of the disease was carried through the evidence of microbiological culture and PCR, considered as the standard for the groups classification. Microbiological culture results of animals with apparent symptoms was 53.5% (55/103) identified as C. pseudotuberculosis by biochemical tests, and allied to PCR, 46.5% (48/103) of the samples were positive for C. pseudotuberculosis. Regarding animals without apparent symptoms, all samples were negative in microbiological culture and PCR (0/50). For the standardization of indirect ELISA we used 42 positive sera and 43 negative sera confirmed by microbiological culture and PCR for caseous lymphadenitis. The mean absorbance was 1.88 with standard deviation of 0.43 for the positive samples, and for negative samples the average was 0.71 and standard deviation 0.18. Thus we considered as positive, the samples with DO value > 1.1 and negative <1.1. The graphical analysis employed in this study, ROC curve, allowed us to find the cutoff value allied to the association of sensitivity and specificity parameters, thus the test accuracy was able to discriminate sick from not sick individuals. The use of serological techniques for the diagnosis of caseous lymphadenitis will allow the epidemiological control of the disease, but does not replace the microbiological culture, considered as the gold standard technique, and can be used as a screening test or animals trade, since the disease condition often is unapparent, which derails the clinical diagnosis and microbiological.

Os objetivos deste estudo foram desenvolver o protocolo do Método do Índice de Qualidade (MIQ) para avaliação da qualidade sensorial da pescada amarela parcialmente eviscerada e armazenada em gelo; avaliar as alterações sensoriais, químicas e microbiológicas da pescada amarela durante o armazenamento e indicar os parâmetros mais adequados para avaliação da qualidade propondo limites de aceitação para a espécie. Quatro lotes com peixes recém capturados em Praia Grande-SP foram transportados à Faculdade de Zootecnia e Engenharia de Alimentos (FZEA) e armazenados entre camadas de gelo (0-1ºC) em câmara fria. Periodicamente 5 peixes foram amostrados e submetidos às análises sensorial (para desenvolvimento do protocolo MIQ e posterior avaliação), microbiológicas (contagens de psicrotróficos, coliformes totais e fecais, Salmonella sp, estafilococos coagulase positiva) e químicas (bases nitrogenadas voláteis-BNV, trimetilamina-TMA, triptofano livre e uréia). A qualidade da pescada amarela comercializada em 14 pontos de venda nas cidades de Santos, São Vicente e Praia Grande foi avaliada utilizando-se o protocolo MIQ desenvolvido, análises microbiológicas e químicas. O protocolo MIQ desenvolvido apresentou aumento linear do índice de qualidade (IQ) ao longo do armazenamento. Nos peixes do lote 1, contagens de psicrotróficos atingiram 107ufc/g no 14º dia. Nos lotes 3 e 4 esse limite foi alcançado entre o 6º e 8º dia. Contagens de E.coli superiores a 102ufc/g foram observadas em 9,5% dos peixes do lote 1 após 21 dias de armazenamento e em 15,25% dos peixes do lote 3 no 6º e 9º dia. Staphylococcus aureus estava ausente em todos os peixes e Salmonella sp foi detectada foi detectada em 7,4% das pescadas amarelas analisadas. Os valores finais de BNV variaram de 23.8 mg/100g a 159 mg/100g em pescadas amarelas e foi dependente da carga microbiana inicial. Todos os lotes apresentaram aumento no teor de TMA, que variou entre 0,18 mg/100g e 15,44mg/100g. As amostras de pescada amarela comercializadas apresentaram IQ variando de 2 a 23, indicando um tempo provável pós captura de 3 até 14 dias. Todos os peixes analisados estavam livres de Salmonella sp e E.coli. Staphylococcus aureus foi determinado em níveis abaixo do limite estipulado pela legislação. Contagens de psicrotróficos acima de 107ufc/g foram observadas em 50% dos peixes. Coliformes totais variaram de 102ufc/g até 107ufc/g. Os valores de TMA e BNV foram maiores que os estabelecidos pela legislação brasileira em 50% e 65% dos peixes, respectivamente. No entanto, em relação à TMA, concentrações superiores a 4mg/100g foram encontradas em peixes que apresentaram contagens de psicrotróficos, concentração de BNV e IQ aceitáveis, sugerindo valores em torno de 12-14mg TMA/100g de peixe como limite adequado para consumo de pescada amarela O valor 30mg de BNV/100g recomendado pela legislação brasileira parece ser adequado como limite de comercialização de pescada amarela, porém a concentração de BNV não está diretamente relacionada ao tempo de armazenamento desde a captura. Triptofano livre e uréia não foram parâmetros químicos adequados para avaliação do frescor da pescada amarela. O protocolo MIQ foi eficiente para a avaliação da qualidade de pescada amarela, apresentando boa correlação entre a perda da qualidade sensorial e o tempo de estocagem. A análise de correlação entre os resultados das diversas análises sugere que a pescada amarela, quando eviscerada e resfriada em gelo, mantém características adequadas para consumo até o 8º dia de armazenamento. Diversos estabelecimentos comerciais de Santos, São Vicente e Praia Grande disponibilizam para os consumidores pescada amarela resfriada em condições inadequadas para consumo de acordo com a legislação brasileira. / The objectives of this study were to develop a Quality Index Method (QIM) scheme for sensory evaluation of partially gutted Acoupa weakfish; to evaluate the sensory, chemical and microbial changes during storage in ice and to suggest the best parameters for quality evaluation and limits of acceptability of this fish species. Four groups of Acoupa weakfish caught in Praia Grande, São Paulo state, were transported to the Faculdade de Zootecnia e Engenharia de Alimentos (FZEA) within 24h of their capture and stored between layers of ice (0-1ºC) in chilling room. Periodically, 5 fishes were sampled and evaluated by sensory (QIM), microbial (counts of psychrotrophic, total and faecal coliforms, Salmonella sp and coagulase-positive Staphylococcus) and chemical (volatile base nitrogen-VBN, trimethylamine-TMA, free tryptophan and urea) analysis. The quality of samples of Acoupa weakfish commercialized in fourteen markets in Santos, São Vicente and Praia Grande cities was also evaluated. The QIM scheme presented linear increase of QI during the storage. In fishes of group 1 the counts of psychrotrophic bacteria reached 107cfu/g at the 14th day of storage. In groups 3 and 4 this limit was reached between days 6 and 8. Counts of E. coli higher than 102cfu/g were detected in 9.5% of the fishes belonging to the group 1 after day 21. This level was also detected in 15.25% of the fishes of the group 3 after days 6 and 9. Staphylococcus aureus was not detected and Salmonella sp was present in 7.4% of the Acoupa weakfish in groups 1 and 2. VBN contents ranged from 23.8 mg/100g to 159 mg/100g and was affected by the microbial load at the beginning of the storage. All the groups of Acoupa weakfish in this study, presented increases in TMA contents during storage and the values ranged between 0.18 and 15.44mg/100g. The samples of Acoupa weakfish obtained from markets showed QI ranging from 2 to 23, suggesting a storage time between 3 to 14 days respectively. All the fishes were free from Salmonella sp and E. coli. Counts of Staphylococcus aureus were lower than the amount determined by the Brazilian legislation. Counts of psychrotrophic bacteria higher than 107cfu/g were observed in 50% of the fishes. Counts of total coliforms ranged from 102cfu/g till 107cfu/g. TMA and VBN contents were higher than the amount allowed by the Brazilian legislation in 50% and 65% of the fishes, respectively. Nevertheless, TMA contents higher than 4mg/100g, were detected in fishes presenting low counts of psychrotrophics, low VBN levels and low QI. Contents ranging from 12-14mg TMA/100g of fish are the suggested limits of consumption for Acoupa weakfish. The limit of 30mg of VBN/100g of fish, allowed by the Brazilian legislation is the appropriate amount for commercialization of Acoupa weakfish, although the VBN contents are not directly related to the storage time. Free tryptophan and urea were not useful as quality indices for Acoupa weakfish. The QIM scheme was effective for quality evaluation of Acoupa weakfish showing significant correlation with quality loss and storage time. Results of correlation analysis allow suggesting that Acoupa weakfish, when gutted and maintained in ice, is suitable for consuming until day 8. Several markets in Santos, São Vicente and Praia Grande commercialize chilled Acoupa weakfish that are unacceptable for consumption according to the Brazilian legislation.