The purpose of the current study is to determine something of the nature of the heat resistance in Azotobacter, if in fact this is found to exist. An attempt is made to determine the specific physiological state associated with heat resistance as well as to resolve this resistance quantitatively.

The survival and germination of Clostridium perfringens spores in different heating media and at different heating rates was studied to determine the fate of C. perfringens spores during abusive cooking and cooling of pork products. The heat resistance (HR) of C. perfringens spores from three strains that were either previously heat shocked (HS) or non-heat shocked (NHS) was determined individually and as a cocktail in phosphate buffer solution (pH 7.4) (PBS), beef gravy (BG), ground pork (GP) and cured ground pork (CGP) at 75ºC. The effect of the heating rate on HR, germination and outgrowth of C. perfringens spores in CGP was determined by increasing the temperature from 20 to 75ºC at a rate of 4, 8, and 12ºC/h prior to heating and holding at 75ºC for 48 h. Heating rates at 4ºC/h in GP and CGP were repeated with additional cooling from 54.4 to 7.2ºC within 20 h (temperature abuse). Linear survival curves were observed on NHS spores in the four heating media, whereas HS spores showed linear curves when heated in PBS and BG, and biphasic curves when heated in GP and CGP. In general, HS spores were more heat sensitive than NHS spores. NHS spores heated in GP had greater HR than spores heated in CGP, BG or PBS. There were no significant differences (P>0.05) on the HR of C. perfringens spores in CGP heated from 20 to 75ºC at 4, 8, or 12ºC/h. Heating rates of 8 and 12ºC/h showed no difference in germination and outgrowth of inoculated spores, whereas at 4ºC/h, growth of C. perfringens occurred between 44 and 56ºC. Temperature abuse during cooling of GP resulted in 2.8 log CFU/g increase of C. perfringens counts. In CGP, C. perfringens counts decreased by 1.1 log CFU/g during cooling from 54.4 to 36.3ºC and then increased by 1 log CFU/g until the product reached 7.2ºC. However, with an initial inoculum in raw CGP of 5 log CFU C. perfringens spores/g, C. perfringens counts did not exceed 3.4 log CFU/g during a 20 h abusive cooling. These results suggest there is no risk associated with C. perfringens in cured pork products under the conditions tested. Results from the present study indicate that different behavior may be expected with different meat products.

Microbiological detection of foodborne pathogens is ineffective for monitoring critical control points (CCP) within a slaughter/processing Hazard Analysis and Critical Control Point (HACCP) system. Pathogens are usually absent from carcass surfaces and their uneven distribution makes it difficult to obtain a representative sample. However, microbiological testing can be applied within a HACCP plan to validate and verify the effectiveness of decontamination procedures designed to control hazards. With proper data collection, the reduction of an indicator group at a point in processing can indicate that a specific pathogen is being effectively controlled, especially when pathogen levels are too low to allow confirmation of process control, as they typically are in beef slaughter processing. Since E. coli O157:H7 has been shown to have some acid resistance, the ability of typical indicator organisms to accurately predict the reduction of this pathogen by carcass decontamination procedures has been a concern. Obtaining potential indicator bacteria from the same environmental reservoir as E. coli O157:H7 may provide non-pathogenic indicators with similar heat- and acid-resistance characteristics suitable for use in processing plant environments for validation and verification of carcass decontamination treatments within HACCP plans. Potential indicator bacteria were isolated from hides of cattle at slaughter facilities in Arizona, Georgia, and Texas and compared with isolates of E. coli O157:H7 from the same locations to determine similarity in acid- and heat-resistance characteristics. After evaluation at 2 heating temperatures (55 and 65??C) and 3 pH levels (3.0, 4.0, and 5.0), it was determined that several potential indicator bacteria were slightly more resistant than E. coli O157:H7 to heating and acid treatment. The greatest reduction in numbers for E. coli O157:H7 and indicator bacteria occurred at pH 3.0 and temperature of 65??C. Counts of bacteria grown at pH 4.0 and 5.0 were not significantly different. Testing indicated that several of the isolates from cattle hides would make good process control indicators since the indicator bacteria were reduced by heating or acid conditions at similar or greater rates when compared to E. coli O157:H7, providing an increased level of security that pathogens have been reduced in processing.

The bacterium Clostridium perfringens is a gram-positive anaerobe responsible for many diseases in man and other animals, the most common of which is acute food poisoning (AFP). It is estimated that nearly 240,000 cases of AFP occur each year in the U.S. The C. perfringens spore plays an important role in this infection. The heat resistance of the spore allows the organism to survive the cooking process, grow in the cooling food, and infect the victim. Despite the occurrence of this disease and the importance of the spore to this process, little work has been performed to determine how heat resistance is obtained and maintained by C. perfringens spores. In this work we study the spores and sporulation process of C. perfringens to determine what factors are most important in the formation of a heat resistant spore. We analyzed the spores produced by nine wild-type strains, including five heat-resistant food poisoning isolates and four less heat-resistant environmental isolates. We determined that threshold core density and a high ratio of cortex peptidoglycan relative to germ cell wall were necessary components of a highly heat-resistant spore. In order to test these observations, we constructed two mutant strains. The first could not achieve the necessary level of core dehydration and rapidly lysed in solution. The second mutant had a reduced amount of cortex relative to germ cell wall, and suffered a corresponding decrease in heat resistance as compared to our wild-type strains. The mutant strains supported the observations drawn from our wild-type strains. Dipicolinic acid is a major component of bacterial spores and is necessary for spore heat resistance. The Cluster I clostridia, including C. perfringens, lack the known DPA synthase operon, spoVF. We developed an in vitro assay for detecting DPA synthetase activity and purified the active enzyme from sporulating C. perfringens crude extract and identified the proteins with mass spectrometry. These results identified the electron transfer flavoprotein alpha chain (EtfA) as the DPA synthase of C. perfringens. Inactivating the etfA gene in C. perfringens resulted in a strain that could begin, but not complete, the sporulation process and produced dramatically lower amounts of DPA than the wild-type. The purified enzyme was shown to produce DPA in vitro and utilized FAD as a preferred cofactor. The results of this research may lead to future techniques to decrease the occurrence of the diseases caused by C. perfringens spores and treatments which may carry over to the diseases caused by similar organisms. / Ph. D.

Current test methodologies used to evaluate the performance of protective clothing do not adequately determine the provided level of protection. The heat fluxes imposed by current evaluation methods are not specifically related to fire environments typical to those the clothing is designed provide protection against. The U.S. Navy is in the process of developing an improved process for testing the fire resistance of daily wear uniforms and protective gear. The first phase of this project involves evaluating currently used evaluation methods and identifying the severity of fire environments that would be expected aboard Navy ships. The examination of the test protocols currently in use identifies major weaknesses, providing the justification for a new test protocol. The first step in developing an improved test protocol is to determine the types of fire scenarios that would be expected aboard Navy vessels. The nearly infinite number of possible fires are reduced to 6 typical cases involving spray fires, pool fires and furniture fires in both compartmented and unconfined cases. An analysis of the environments produced by these types of fires is presented. The effects of compartmentation parameters are also investigated to determine the critical factors that affect the expected fire environment. Expected heat fluxes for all scenarios are presented at a number of distances from the fire.

O presente trabalho objetivou descrever o comportamento da cepa Salmonella Enteritidis (S. Enteritidis) PT4, frente ao tratamento térmico a 60,0º C por três minutos e meio, em ovo integral pasteurizado desidratado reconstituído, interrelacionando a termorresistência com temperatura prévia de incubação e concentração de microrganismos. Foram realizadas 150 análises, das quais 75 foram trabalhadas com cepa previamente incubada à 35ºC e 75 com cepa incubada à 43ºC. Cada um destes grupos foi subdividido em três subgrupos, inoculado-se as concentrações de 10³, 105 e 108 unidades formadoras de colônia (UFC)/mL de caldo BHI. Após inoculação, foi simulada pasteurização do ovo, a 60,0ºC por 3,5 minutos, em banho-maria, e, posteriormente, realizadas contagem em placa (UFC) e Número Mais Provável (NMP), para verificação da concentração de S. Enteritidis sobrevivente. O tempo de redução decimal (valor D) foi de 2,54 e 2,57 minutos para S. Enteritidis previamente incubada à 35ºC e inoculada nas concentrações de 105 e 108 UFC/mL de caldo BHI, respectivamente. Para S. Enteritidis previamente incubada à 43ºC e inoculada nas concentrações de 105 e 108 UFC/mL de caldo BHI, o valor D foi de 2,58 e 2,40 minutos, respectivamente. Não foi possível o cálculo do valor D para a concentração 10³ UFC/mL de caldo BHI, visto que não houve população sobrevivente. Não houve diferença significativa entre os resultados de contagem e NMP, comparando-se as duas temperaturas de incubação, entretanto, os resultados de contagem e NMP das três concentrações inoculadas foram diferentes entre si. Concluiu-se que a cepa pré incubada a 43ºC não apresentou maior resistência ao calor, em relação à cepa incubada a 35ºC; e não houve maior resistência ao calor, comparando-se as diferentes concentrações. / The purpose of this study was to describe the behavior of Salmonella Enteritidis (S. Enteritidis) PT4 undergoing thermal treatment at 60ºC for 3 ½ minutes in reconstituted dried-whole egg, correlating the heat resistance with a previous temperature of incubation and microrganism concentration. 150 analyses were conducted, 75 with S. Enteritidis PT4 previously incubated at 35ºC and 75 with S. Enteritidis PT4 previously incubated at 43ºC. Each of these two groups was subdivided in 3 sub-groups, adding the inoculum at concentrations of 10³, 105, 108 CFU/mL of BHI broth. After the inoculation, pasteurization of the egg was simulated at 60ºC for 3,5 minutes in a water bath. Then colony count and Most Probable Number (MPN) were carried out to check the concentration of surviving S. Enteritidis. The decimal reduction time (D-value) was 2.54 and 2.57 minutes for S. Enteritidis previously incubated at 35ºC and inoculated in the concentration of 105 and 108 CFU/mL of BHI broth, respectively. Concerning the S. Enteritidis previously incubated at 43ºC and inoculated in concentrations of 105 and 108 CFU/mL of BHI broth, the D-value was 2.58 and 2.40 minutes, respectively. It was not possible to calculate the D-Value for the 10³ CFU/mL concentration of BHI broth, because there was no surviving population. There was no significant difference between the results of colony count and MPN, comparing the two incubation temperatures, however the results of colony count and MPN of the three inoculated concentrations were different from each other. It was possible to conclude that the S. Enteritidis PT4 previously incubated at 43ºC hadn´t shown more heat resistance if compared with the S. Enteritidis PT4 incubated at 35ºC; and there wasn’t more heat resistance, comparing the different concentrations.

O presente trabalho objetivou descrever o comportamento da cepa Salmonella Enteritidis (S. Enteritidis) PT4, frente ao tratamento térmico a 60,0º C por três minutos e meio, em ovo integral pasteurizado desidratado reconstituído, interrelacionando a termorresistência com temperatura prévia de incubação e concentração de microrganismos. Foram realizadas 150 análises, das quais 75 foram trabalhadas com cepa previamente incubada à 35ºC e 75 com cepa incubada à 43ºC. Cada um destes grupos foi subdividido em três subgrupos, inoculado-se as concentrações de 10³, 105 e 108 unidades formadoras de colônia (UFC)/mL de caldo BHI. Após inoculação, foi simulada pasteurização do ovo, a 60,0ºC por 3,5 minutos, em banho-maria, e, posteriormente, realizadas contagem em placa (UFC) e Número Mais Provável (NMP), para verificação da concentração de S. Enteritidis sobrevivente. O tempo de redução decimal (valor D) foi de 2,54 e 2,57 minutos para S. Enteritidis previamente incubada à 35ºC e inoculada nas concentrações de 105 e 108 UFC/mL de caldo BHI, respectivamente. Para S. Enteritidis previamente incubada à 43ºC e inoculada nas concentrações de 105 e 108 UFC/mL de caldo BHI, o valor D foi de 2,58 e 2,40 minutos, respectivamente. Não foi possível o cálculo do valor D para a concentração 10³ UFC/mL de caldo BHI, visto que não houve população sobrevivente. Não houve diferença significativa entre os resultados de contagem e NMP, comparando-se as duas temperaturas de incubação, entretanto, os resultados de contagem e NMP das três concentrações inoculadas foram diferentes entre si. Concluiu-se que a cepa pré incubada a 43ºC não apresentou maior resistência ao calor, em relação à cepa incubada a 35ºC; e não houve maior resistência ao calor, comparando-se as diferentes concentrações. / The purpose of this study was to describe the behavior of Salmonella Enteritidis (S. Enteritidis) PT4 undergoing thermal treatment at 60ºC for 3 ½ minutes in reconstituted dried-whole egg, correlating the heat resistance with a previous temperature of incubation and microrganism concentration. 150 analyses were conducted, 75 with S. Enteritidis PT4 previously incubated at 35ºC and 75 with S. Enteritidis PT4 previously incubated at 43ºC. Each of these two groups was subdivided in 3 sub-groups, adding the inoculum at concentrations of 10³, 105, 108 CFU/mL of BHI broth. After the inoculation, pasteurization of the egg was simulated at 60ºC for 3,5 minutes in a water bath. Then colony count and Most Probable Number (MPN) were carried out to check the concentration of surviving S. Enteritidis. The decimal reduction time (D-value) was 2.54 and 2.57 minutes for S. Enteritidis previously incubated at 35ºC and inoculated in the concentration of 105 and 108 CFU/mL of BHI broth, respectively. Concerning the S. Enteritidis previously incubated at 43ºC and inoculated in concentrations of 105 and 108 CFU/mL of BHI broth, the D-value was 2.58 and 2.40 minutes, respectively. It was not possible to calculate the D-Value for the 10³ CFU/mL concentration of BHI broth, because there was no surviving population. There was no significant difference between the results of colony count and MPN, comparing the two incubation temperatures, however the results of colony count and MPN of the three inoculated concentrations were different from each other. It was possible to conclude that the S. Enteritidis PT4 previously incubated at 43ºC hadn´t shown more heat resistance if compared with the S. Enteritidis PT4 incubated at 35ºC; and there wasn’t more heat resistance, comparing the different concentrations.

No description available.

This diploma thesis is focused on determination of thermal properties of gloves by means of thermal manikin. The background research with relation to this topic together with derivation of the computational equations used for the following measurement are presented in the first part. The most significant part of this work is suggestion of procedure for measuring the heat resistance of gloves with thermal manikin based on procedure from ČSN EN 511. This methodics was verificated with 5 pairs of gloves together with analysing the uncertainties and testing the repeatability of this measurement. This work also contains the outline for all of the measured pairs with posibilities of their usage in specific situations and surrounding air temperatures. At the end of this work is mentioned brief summary of achieved results.

Bacterial spore peptidoglycan (PG) is very loosely cross-linked relative to vegetative PG. Theories suggest that loosely cross-linked spore PG may have a flexibility which contributes to the attainment of spore core dehydration. The structure of the PG found in fully dormant spores has previously been examined in wild type and many mutant strains. These analyses showed little correlation between the degree of spore PG cross-linking and core dehydration. However, these studies only examined the structure of PG from dormant spores and did not allow for the structural analysis of spore PG during sporulation when actual spore PG synthesis and core dehydration occur. Structural analyses of developing spore PG from wild type Bacillus subtilis and eight mutant strains are included in this study. Structural analyses of developing spore PG suggest the following: a) the germ cell wall PG is synthesized first next to the inner forespore membrane; b) cross-linking is relatively high in the first 10% of spore PG synthesized; b) a rapid decrease in cross-linking is observed during synthesis of the next 20% of the spore PG; and c) this decrease is followed by an eightfold rise in the degree of cross-linking during synthesis of the final 70% of the spore PG. This increasing gradient of cross-linking was previously predicted to contribute to the attainment of spore core dehydration. However, analyses of mutant strains indicate this cross-linking gradient is not required for the attainment of spore dehydration. / Master of Science