The recently identified causative agent of White-Nose Syndrome (WNS), Geomyces destructans, has been responsible for the mortality of an estimated 5.7 million North American bats since its emergence in 2006. A primary focus of the National Response Plan, established by US Fish and Wildlife in 2011, was the identification of biological and chemical control options. In an effort to identify potential biological and chemical control options for WNS, six previously described bacterially produced volatile organic compounds (VOCs) and multiply induced Rhodococcus rhodochrous DAP96253 were screened for anti-Geomyces destructans activity. Geomyces destructans conidia and mycelial plugs were exposed to the VOCs and induced Rhodococcus in a closed air space at 15°C and 4°C and evaluated for inhibition of conidia germination and mycelial extension. Additionally, in situ application methods for induced Rhodococcus, such as fixed cell catalyst and fermentation cell paste in non-growth conditions, were screened with positive results. Rhodococcus was assayed for ex vivo activity via exposure to bat tissue ex-plants inoculated with G. destructans conidia. All VOCs inhibited radial growth of mycelial plugs and growth from conidia at both temperatures, with the greatest effect at low temperature (4°C). Induced Rhodococcus completely inhibited growth from conidia at 15°C and had a strong fungistatic effect at 4°C. Induced Rhodococcus inhibited Geomyces destructans growth from conidia when cultured in a shared air space with bat tissue explants inoculated with Geomyces destructans conidia. During the evaluation diffusible brown pigment was observed in G. destructans cultures exposed to induced Rhodococcus or select VOCs. The pigment was induced by light and oxidative challenge and hypothesized to be melanin. Traditional microbiological methods, as well as copper sulfide-silver staining and ultraviolet-visible spectroscopy, were utilized to confirm this hypothesis. This was a noteworthy result as melanin is a known virulence factor in other pathogenic fungi and may play a significant role in WNS. The identification of bacterially produced VOCs and inducible biological agents with anti-Geomyces destructans activity expands the pool of potential biological and chemical control options for WNS and provides wildlife management personnel with tools to combat this devastating disease.

Aflatoxin contamination accounts for millions of dollars worth of losses for corn and cotton in Texas. Two atoxigenic strains of Aspergillus flavus, AF36 and Afla-Guard, are labeled for its management. The purpose of this study was to measure differences in the ability of these strains to sporulate and to track movement of their conidia in corn and cotton fields. Sporulation was evaluated by incubating the two strains on their commercial formulations (inoculated on cereal grains) at six constant humidity levels ranging from 0-100%, using closed chambers with saturated salt solutions. Conidial production by Afla-Guard was 3-fold greater than that of AF36 at 100% humidity. Sporulation of the two strains was also evaluated on one substrate by inoculating their conidia on sterile, hulled barley. After 3 days, there was a 234-fold increase in conidia recovered from the barley inoculated with Afla-Guard, compared with a 21-fold increase in conidia recovered from the AF36-inoculated barley. These data suggest that the Afla-Guard strain sporulates better than the AF36 strain, which may be a factor in effectiveness for biological control. An in vitro de-Wit competition experiment showed that sporulation by the Afla-Guard strain was not affected by co-inoculation with either AF36 or the wildtype NRRL3357 toxin producing strain. To measure conidial movement, an Afla-Guard nitrate non-utilizing mutant colonizing autoclaved corn seed, was placed at one point in a field of cotton and corn. For detection, aliquots washed from leaf samples were plated onto a medium containing potassium chlorate. The mutant was recovered at a maximum distance of 6.4 m in corn fields along the same row and as far as 10.2 m across rows from the point source. In cotton fields, the mutant was recovered at 9.1 meters along the same row and 6.1 m across rows from the point source. There was no recovery at 24.3 m from the point source - the maximum distance evaluated. The experiment was repeated in a second year with similar results. These data suggest that plots in field trials may not need wide separation in order to avoid cross contamination. To assess the viability of a toxigenic and atoxigenic strain of A. flavus over time, polycarbonate packets containing conidia and sclerotia of both strains were buried in Ships Clay soil with the matric potential held constant at -24 kPa or -154 kPa. After 10 months, viable conidia were recovered in all treatments. After 14 months, viability of the atoxigenic strain incubated at -154 kPa ψm was lost, while other treatments remained viable. Ears of corn were inoculated via silk channel at different stages of silk senescence. Sclerotia were enumerated from the same plants following harvest of the crop. Sclerotial production by A. flavus was greatest from ears with silks inoculated at senescence, compared with inoculation when silks were green. The isolation frequency of Penicillium sp. from surface-sterilized kernels at harvest was the highest from ears that were inoculated with A. flavus when silks were fresh, as compared with A. flavus inoculation of ears with senescent silks. A Fusarium and Penicillium species was isolated from harvested kernels, and their sterile Czapek-Dox broth culture filtrates were tested for their effect on development of three strains of A. flavus on agar. The Penicillium broth filtrate greatly reduced sclerotial numbers relative to the control and the Fusarium filtrate (P<0.05). When A. flavus was grown in the presence of autoclaved Penicillium culture filtrate, there was no effect on sclerotial production. The Penicillium filtrate increased the rate of radial hyphal growth of the A. flavus isolates on agar compared to the control and the Fusarium culture filtrate.

Bdellovibrionaceae (BD) are unique, predatory, endoparasitic, Gram-negative bacteria. As the world's smallest living hunter they prey on other Gram-negative bacteria giving them potential as biological control agents. Prior to this study, however, there were no reports of BD in New Zealand. The overall aim of this research was to isolate BD from New Zealand sources, characterise them and investigate their potential role as a biological control agent. The history, characteristics, life cycle and mechanism of predation of this organism are reviewed and the possibility of the industrial applications of BD, are discussed. In this study, a halophilic species of BD was isolated from fourteen coastal sea water sites around New Zealand. Thirteen isolates were characterised using proven characterisation techniques including general, microscopic and molecular techniques. It was found that the isolates were taxonomically identical or very closely related to each other and belong to the genus Bacteriovorax. The predation pattern of BD isolates was examined against a group of Gram negative bacteria in solid and liquid media. The predation patterns and efficiencies of the different BD isolates were similar, which confirms that the BD isolates are closely related, are selective in their predation, and prey on some Gram-negative bacteria but not all. The rapid loss of culture viability of BD is well known, but no studies have been reported to date on the survival of pure cultures of BD at different temperatures. The survival rate of BD in dense suspensions at different temperatures without host bacteria was investigated and it was observed that pure BD cultures can be stored with minimal reduction in numbers at temperatures ranging from 4°C to 20°C. However, significant reductions in numbers were observed at -1 8"C, 30°C and 37°C after 13 to 16 days. The effects of the 13 New Zealand BD isolates on the growth of a population of Photobacterium phosphoreum were examined to select the best isolate for in vitro application. All of the isolates tested had considerable reduction effect against P. phosphoreum. Some isolates were more effective than others, despite their taxonomic similarity to each other. The isolate OT2 was selected for further studies based on these results. The in vitro efficacy of BD was assessed against late exponential cultures of a seafood spoilage bacterium, P. phosphoreum, originally isolated from Cod fillets from Denmark. Loglo reductions of P. phosphoreum and some other Gram-negative bacteria ranged from 4.5 to 4.8 after 9 h of incubation at 25OC. BD was effective in reducing the numbers of P. phosphoreum at pH 5.5 to 8.5 and salinity 0.9 to 4.5% (wlv). A significant interaction was observed between the prey and predator concentrations and nutrient concentration. Prey concentrations were observed to be the most vital factor in predation and the most favourable predation conditions were at a prey concentration of -8 loglo colony forming units (CFU)/mL, together with a predator concentration of 3 - 7 loglo plaque forming units (PFU)/mL and a prey : predator ratio of >5.0. The thresholds of the prey and predator concentrations for predation were observed to be 3.7 loglo CFUImL and 3.9 loglo PFUImL, respectively. The trials carried out in this study focused on the efficiency of BD on a pure culture of one organism, P. phosphoreum and not on mixed cultures of Gramnegative spoilage bacteria, the normal condition observed in saltwater fish. There has been very little research in this field and the results of these trials suggest further investigation into the effect of BD on mixed cultures of Gram-negative spoilage organisms is warranted. Since only one isolate of BD (OT2) was examined against only one spoilage bacterium (P. phosphoreum) in liquid medium, the evidence of these findings must be restricted to these particular conditions. Future studies, using a range of BD isolates against a mixture of spoilage and pathogenic organisms in solid medium are warranted. The biopreservation capability of BD in extending the shelf life of king salmon was evaluated. A significant effect was observed at 20°C but not at 10°C. At 20°C the shelf life was extended through extension of the lag phase of growth of the prey bacteria and a reduction in total numbers attained. Sensory evaluation of the salmon product being tested confirmed that the shelf life was extended. However, at 10°C there was no reduction in prey organisms, which suggested that the strain of BD used is ineffective at refrigeration temperatures.

Bdellovibrionaceae (BD) are unique, predatory, endoparasitic, Gram-negative bacteria. As the world's smallest living hunter they prey on other Gram-negative bacteria giving them potential as biological control agents. Prior to this study, however, there were no reports of BD in New Zealand. The overall aim of this research was to isolate BD from New Zealand sources, characterise them and investigate their potential role as a biological control agent. The history, characteristics, life cycle and mechanism of predation of this organism are reviewed and the possibility of the industrial applications of BD, are discussed. In this study, a halophilic species of BD was isolated from fourteen coastal sea water sites around New Zealand. Thirteen isolates were characterised using proven characterisation techniques including general, microscopic and molecular techniques. It was found that the isolates were taxonomically identical or very closely related to each other and belong to the genus Bacteriovorax. The predation pattern of BD isolates was examined against a group of Gram negative bacteria in solid and liquid media. The predation patterns and efficiencies of the different BD isolates were similar, which confirms that the BD isolates are closely related, are selective in their predation, and prey on some Gram-negative bacteria but not all. The rapid loss of culture viability of BD is well known, but no studies have been reported to date on the survival of pure cultures of BD at different temperatures. The survival rate of BD in dense suspensions at different temperatures without host bacteria was investigated and it was observed that pure BD cultures can be stored with minimal reduction in numbers at temperatures ranging from 4°C to 20°C. However, significant reductions in numbers were observed at -1 8"C, 30°C and 37°C after 13 to 16 days. The effects of the 13 New Zealand BD isolates on the growth of a population of Photobacterium phosphoreum were examined to select the best isolate for in vitro application. All of the isolates tested had considerable reduction effect against P. phosphoreum. Some isolates were more effective than others, despite their taxonomic similarity to each other. The isolate OT2 was selected for further studies based on these results. The in vitro efficacy of BD was assessed against late exponential cultures of a seafood spoilage bacterium, P. phosphoreum, originally isolated from Cod fillets from Denmark. Loglo reductions of P. phosphoreum and some other Gram-negative bacteria ranged from 4.5 to 4.8 after 9 h of incubation at 25OC. BD was effective in reducing the numbers of P. phosphoreum at pH 5.5 to 8.5 and salinity 0.9 to 4.5% (wlv). A significant interaction was observed between the prey and predator concentrations and nutrient concentration. Prey concentrations were observed to be the most vital factor in predation and the most favourable predation conditions were at a prey concentration of -8 loglo colony forming units (CFU)/mL, together with a predator concentration of 3 - 7 loglo plaque forming units (PFU)/mL and a prey : predator ratio of >5.0. The thresholds of the prey and predator concentrations for predation were observed to be 3.7 loglo CFUImL and 3.9 loglo PFUImL, respectively. The trials carried out in this study focused on the efficiency of BD on a pure culture of one organism, P. phosphoreum and not on mixed cultures of Gramnegative spoilage bacteria, the normal condition observed in saltwater fish. There has been very little research in this field and the results of these trials suggest further investigation into the effect of BD on mixed cultures of Gram-negative spoilage organisms is warranted. Since only one isolate of BD (OT2) was examined against only one spoilage bacterium (P. phosphoreum) in liquid medium, the evidence of these findings must be restricted to these particular conditions. Future studies, using a range of BD isolates against a mixture of spoilage and pathogenic organisms in solid medium are warranted. The biopreservation capability of BD in extending the shelf life of king salmon was evaluated. A significant effect was observed at 20°C but not at 10°C. At 20°C the shelf life was extended through extension of the lag phase of growth of the prey bacteria and a reduction in total numbers attained. Sensory evaluation of the salmon product being tested confirmed that the shelf life was extended. However, at 10°C there was no reduction in prey organisms, which suggested that the strain of BD used is ineffective at refrigeration temperatures.

Bdellovibrionaceae (BD) are unique, predatory, endoparasitic, Gram-negative bacteria. As the world's smallest living hunter they prey on other Gram-negative bacteria giving them potential as biological control agents. Prior to this study, however, there were no reports of BD in New Zealand. The overall aim of this research was to isolate BD from New Zealand sources, characterise them and investigate their potential role as a biological control agent. The history, characteristics, life cycle and mechanism of predation of this organism are reviewed and the possibility of the industrial applications of BD, are discussed. In this study, a halophilic species of BD was isolated from fourteen coastal sea water sites around New Zealand. Thirteen isolates were characterised using proven characterisation techniques including general, microscopic and molecular techniques. It was found that the isolates were taxonomically identical or very closely related to each other and belong to the genus Bacteriovorax. The predation pattern of BD isolates was examined against a group of Gram negative bacteria in solid and liquid media. The predation patterns and efficiencies of the different BD isolates were similar, which confirms that the BD isolates are closely related, are selective in their predation, and prey on some Gram-negative bacteria but not all. The rapid loss of culture viability of BD is well known, but no studies have been reported to date on the survival of pure cultures of BD at different temperatures. The survival rate of BD in dense suspensions at different temperatures without host bacteria was investigated and it was observed that pure BD cultures can be stored with minimal reduction in numbers at temperatures ranging from 4°C to 20°C. However, significant reductions in numbers were observed at -1 8"C, 30°C and 37°C after 13 to 16 days. The effects of the 13 New Zealand BD isolates on the growth of a population of Photobacterium phosphoreum were examined to select the best isolate for in vitro application. All of the isolates tested had considerable reduction effect against P. phosphoreum. Some isolates were more effective than others, despite their taxonomic similarity to each other. The isolate OT2 was selected for further studies based on these results. The in vitro efficacy of BD was assessed against late exponential cultures of a seafood spoilage bacterium, P. phosphoreum, originally isolated from Cod fillets from Denmark. Loglo reductions of P. phosphoreum and some other Gram-negative bacteria ranged from 4.5 to 4.8 after 9 h of incubation at 25OC. BD was effective in reducing the numbers of P. phosphoreum at pH 5.5 to 8.5 and salinity 0.9 to 4.5% (wlv). A significant interaction was observed between the prey and predator concentrations and nutrient concentration. Prey concentrations were observed to be the most vital factor in predation and the most favourable predation conditions were at a prey concentration of -8 loglo colony forming units (CFU)/mL, together with a predator concentration of 3 - 7 loglo plaque forming units (PFU)/mL and a prey : predator ratio of >5.0. The thresholds of the prey and predator concentrations for predation were observed to be 3.7 loglo CFUImL and 3.9 loglo PFUImL, respectively. The trials carried out in this study focused on the efficiency of BD on a pure culture of one organism, P. phosphoreum and not on mixed cultures of Gramnegative spoilage bacteria, the normal condition observed in saltwater fish. There has been very little research in this field and the results of these trials suggest further investigation into the effect of BD on mixed cultures of Gram-negative spoilage organisms is warranted. Since only one isolate of BD (OT2) was examined against only one spoilage bacterium (P. phosphoreum) in liquid medium, the evidence of these findings must be restricted to these particular conditions. Future studies, using a range of BD isolates against a mixture of spoilage and pathogenic organisms in solid medium are warranted. The biopreservation capability of BD in extending the shelf life of king salmon was evaluated. A significant effect was observed at 20°C but not at 10°C. At 20°C the shelf life was extended through extension of the lag phase of growth of the prey bacteria and a reduction in total numbers attained. Sensory evaluation of the salmon product being tested confirmed that the shelf life was extended. However, at 10°C there was no reduction in prey organisms, which suggested that the strain of BD used is ineffective at refrigeration temperatures.

Fungos filamentosos com capacidade de produzir micotoxinas podem estar presentes em alimentos, desde o cultivo até o produto após industrialização. Devido a isso, estratégias para controlar o crescimento fúngico devem ser investigadas, a fim de evitar o desenvolvimento desses microrganismos, bem como a produção de suas toxinas nos alimentos. Neste trabalho, duas abordagens para o controle de fungos toxigênicos foram avaliadas. A primeira estratégia foi a utilização de bactérias provenientes de diferentes ambientes aquáticos, sendo que 10 linhagens de Bacillus spp. e a linhagem Pseudomonas sp. 4B foram testadas quanto à influência sobre os parâmetros de crescimento (taxas de crescimento micelial, esporulação e germinação de esporos) de fungos toxigênicos (Aspergillus e Penicillium) e formação de micotoxinas. Todas as bactérias foram capazes de inibir o crescimento dos fungos em meio de cultura, apresentando halos de inibição variando de 1,0 até 15,7 mm. Bacillus sp. P11 apresentou resultados mais expressivos em relação às demais linhagens do gênero Bacillus com maiores valores de redução na maioria dos parâmetros de crescimento. Além disso, Bacillus sp. P11 e Pseudomonas sp. 4B apresentaram efeito sobre as taxas de crescimento micelial, esporulação e germinação de esporos, com níveis de redução acima de 43,3, 32,1 e 84,1% respectivamente. Mesmo assim, as demais linhagens também apresentaram resultados satisfatórios sobre esses parâmetros. Estas bactérias também reduziram a síntese de aflatoxina B1 e ocratoxina A em mais de 94 e 63%, respectivamente, quando cultivadas simultaneamente com os fungos produtores de cada micotoxina. Adicionalmente, a capacidade de Bacillus sp. P11 em produzir os lipopeptídeos iturina A (167,9 mg/mL de extrato butanólico) e surfactina (361,9 mg/mL de extrato butanólico) foi confirmada. Estes compostos podem ter contribuído para a atividade antifúngica desta bactéria. A segunda estratégia investigada neste estudo para controlar o desenvolvimento de fungos toxigênicos foi o emprego de nanofibras de poli-ɛ-caprolactona (PCL) incorporadas com cetoconazol e natamicina como material antimicrobiano. Nesta abordagem, as nanofibras foram produzidas pela técnica de eletrofiação e posteriormente caracterizadas e avaliadas quanto ao seu potencial antifúngico. Nanofibras funcionalizadas com cetoconazol ou natamicina apresentaram atividade antifúngica contra os isolados toxigênicos uma vez que zonas de inibição variando de 6 a 44 mm foram observadas. Além disso, as análises de microscopia eletrônica e espectroscopia demonstraram que a incorporação dos antifúngicos não altera de forma expressiva as principais características das nanofibras. Também foi possível verificar a capacidade de liberação controlada dos antifúngicos durante 72 h de contato das nanofibras com diferentes soluções simulantes. Valores próximos a 80 e 45 μg/mL de cetoconazol e natamicina, respectivamente, foram observados em solução de Tween 20 (5%). Portanto, o processo de eletrofiação foi capaz de agregar propriedades antifúngicas às nanofibras de PCL. Os resultados demonstraram que as bactérias e os nanomateriais investigados neste estudo são promissores para o controle de fungos toxigênicos e produção de micotoxinas. / Filamentous fungi that have the potential to produce mycotoxins may be present in food, from cultivation to after industrialization. Therefore, several strategies to control fungal growth must be investigated in order to avoid the development of these microorganisms and the production of their toxins in food. In this work, two approaches to toxigenic fungi control were evaluated. The first one was the use of bacteria from different aquatic environments as biocontrol agents in which 10 Bacillus spp. strains and the Pseudomonas sp. 4B strain were tested in relation to the effect on growth parameters (mycelial growth, sporulation and spore germination rates) of toxigenic fungi (Aspergillus and Penicillium) and mycotoxin formation. All bacteria were able to inhibit the fungal growth in culture medium with inhibition zones ranging from 1.0 to 15.7 mm. It was also observed that Bacillus sp. P11 had better results compared to other Bacillus strains with larger reduction values in most of growth parameters. Furthermore, Bacillus sp. P11 and Pseudomonas sp. 4B exhibited effect on mycelial growth, sporulation and spore germination rates with reduction values above of 43.3, 32.1 and 84.1%, respectively. Even so, the other strains also showed satisfactory results on these parameters. Finally, these bacteria reduced the synthesis of aflatoxin B1 and ochratoxin A at levels above 94 and 63%, respectively, when co-cultivated with each mycotoxin producing fungi. Additionally, the ability of Bacillus sp. P11 to produce lipopeptides such as iturin A (167.9 mg/ml of butanolic extract) and surfactin (361.9 mg/ml of butanolic extract) was confirmed. These compounds may have contributed to antifungal activity of this bacterium. The second investigation of this work in order to control the growth of toxigenic fungi was the use of poly-ε-caprolactone nanofibers incorporated with ketoconazole and natamycin as antimicrobial material. In this approach, nanofibers were produced by the electrospinning technique and subsequently characterized and evaluated for their antifungal potential. Both nanofibers functionalized with ketoconazole and natamycin showed antifungal activity against toxigenic isolates since inhibitory zones ranging from 6 to 44 mm were observed. In addition, scanning electron microscopy and infrared spectroscopy analysis showed that the antifungals incorporation does not change the characteristics of nanofibers. It was also possible to verify the ability of controlled drug release during 72 h of nanofibers contact with different simulants solutions. Values near 80 and 45 μg/ml of ketoconazole and natamycin, respectively, were observed in the solution containing 5% Tween 20. Therefore, the electrospinning process was able to provide antifungal properties to the nanofibers. The results showed that bacteria and nanomaterials investigated in this study are promising for developing control strategies of toxigenic fungi and mycotoxin production.

Fungos filamentosos com capacidade de produzir micotoxinas podem estar presentes em alimentos, desde o cultivo até o produto após industrialização. Devido a isso, estratégias para controlar o crescimento fúngico devem ser investigadas, a fim de evitar o desenvolvimento desses microrganismos, bem como a produção de suas toxinas nos alimentos. Neste trabalho, duas abordagens para o controle de fungos toxigênicos foram avaliadas. A primeira estratégia foi a utilização de bactérias provenientes de diferentes ambientes aquáticos, sendo que 10 linhagens de Bacillus spp. e a linhagem Pseudomonas sp. 4B foram testadas quanto à influência sobre os parâmetros de crescimento (taxas de crescimento micelial, esporulação e germinação de esporos) de fungos toxigênicos (Aspergillus e Penicillium) e formação de micotoxinas. Todas as bactérias foram capazes de inibir o crescimento dos fungos em meio de cultura, apresentando halos de inibição variando de 1,0 até 15,7 mm. Bacillus sp. P11 apresentou resultados mais expressivos em relação às demais linhagens do gênero Bacillus com maiores valores de redução na maioria dos parâmetros de crescimento. Além disso, Bacillus sp. P11 e Pseudomonas sp. 4B apresentaram efeito sobre as taxas de crescimento micelial, esporulação e germinação de esporos, com níveis de redução acima de 43,3, 32,1 e 84,1% respectivamente. Mesmo assim, as demais linhagens também apresentaram resultados satisfatórios sobre esses parâmetros. Estas bactérias também reduziram a síntese de aflatoxina B1 e ocratoxina A em mais de 94 e 63%, respectivamente, quando cultivadas simultaneamente com os fungos produtores de cada micotoxina. Adicionalmente, a capacidade de Bacillus sp. P11 em produzir os lipopeptídeos iturina A (167,9 mg/mL de extrato butanólico) e surfactina (361,9 mg/mL de extrato butanólico) foi confirmada. Estes compostos podem ter contribuído para a atividade antifúngica desta bactéria. A segunda estratégia investigada neste estudo para controlar o desenvolvimento de fungos toxigênicos foi o emprego de nanofibras de poli-ɛ-caprolactona (PCL) incorporadas com cetoconazol e natamicina como material antimicrobiano. Nesta abordagem, as nanofibras foram produzidas pela técnica de eletrofiação e posteriormente caracterizadas e avaliadas quanto ao seu potencial antifúngico. Nanofibras funcionalizadas com cetoconazol ou natamicina apresentaram atividade antifúngica contra os isolados toxigênicos uma vez que zonas de inibição variando de 6 a 44 mm foram observadas. Além disso, as análises de microscopia eletrônica e espectroscopia demonstraram que a incorporação dos antifúngicos não altera de forma expressiva as principais características das nanofibras. Também foi possível verificar a capacidade de liberação controlada dos antifúngicos durante 72 h de contato das nanofibras com diferentes soluções simulantes. Valores próximos a 80 e 45 μg/mL de cetoconazol e natamicina, respectivamente, foram observados em solução de Tween 20 (5%). Portanto, o processo de eletrofiação foi capaz de agregar propriedades antifúngicas às nanofibras de PCL. Os resultados demonstraram que as bactérias e os nanomateriais investigados neste estudo são promissores para o controle de fungos toxigênicos e produção de micotoxinas. / Filamentous fungi that have the potential to produce mycotoxins may be present in food, from cultivation to after industrialization. Therefore, several strategies to control fungal growth must be investigated in order to avoid the development of these microorganisms and the production of their toxins in food. In this work, two approaches to toxigenic fungi control were evaluated. The first one was the use of bacteria from different aquatic environments as biocontrol agents in which 10 Bacillus spp. strains and the Pseudomonas sp. 4B strain were tested in relation to the effect on growth parameters (mycelial growth, sporulation and spore germination rates) of toxigenic fungi (Aspergillus and Penicillium) and mycotoxin formation. All bacteria were able to inhibit the fungal growth in culture medium with inhibition zones ranging from 1.0 to 15.7 mm. It was also observed that Bacillus sp. P11 had better results compared to other Bacillus strains with larger reduction values in most of growth parameters. Furthermore, Bacillus sp. P11 and Pseudomonas sp. 4B exhibited effect on mycelial growth, sporulation and spore germination rates with reduction values above of 43.3, 32.1 and 84.1%, respectively. Even so, the other strains also showed satisfactory results on these parameters. Finally, these bacteria reduced the synthesis of aflatoxin B1 and ochratoxin A at levels above 94 and 63%, respectively, when co-cultivated with each mycotoxin producing fungi. Additionally, the ability of Bacillus sp. P11 to produce lipopeptides such as iturin A (167.9 mg/ml of butanolic extract) and surfactin (361.9 mg/ml of butanolic extract) was confirmed. These compounds may have contributed to antifungal activity of this bacterium. The second investigation of this work in order to control the growth of toxigenic fungi was the use of poly-ε-caprolactone nanofibers incorporated with ketoconazole and natamycin as antimicrobial material. In this approach, nanofibers were produced by the electrospinning technique and subsequently characterized and evaluated for their antifungal potential. Both nanofibers functionalized with ketoconazole and natamycin showed antifungal activity against toxigenic isolates since inhibitory zones ranging from 6 to 44 mm were observed. In addition, scanning electron microscopy and infrared spectroscopy analysis showed that the antifungals incorporation does not change the characteristics of nanofibers. It was also possible to verify the ability of controlled drug release during 72 h of nanofibers contact with different simulants solutions. Values near 80 and 45 μg/ml of ketoconazole and natamycin, respectively, were observed in the solution containing 5% Tween 20. Therefore, the electrospinning process was able to provide antifungal properties to the nanofibers. The results showed that bacteria and nanomaterials investigated in this study are promising for developing control strategies of toxigenic fungi and mycotoxin production.

An array of Bacillus licheniformis strains were isolated from a commercial powdered milk process. Bacteriophages exhibiting activity against B. licheniformis were isolated from cattle manure and effluent samples destined for a lagoon at a dairy farm. After sequencing, 8 of the 10 phages were found to be novel and genetically differentiated. Transmission electron scanning microscopy (TSEM) was performed. All bacteriophages were of the family Herelleviridae with contractile tail sheaths ranging from 80µm to 150µm and, surprisingly, survived a common fluid milk processing treatment used to inactivate vegetative cells. The survival of the phage after high temperature short time pasteurization of 73℃ for 20 s shows that the use of bacteriophages in milk to control B. licheniformis could be applied as a potential quality control, retarding the germination of spores and reduction of final spore counts in products with long run times such as dairy powders.

Four closely related species of leaf beetles (Diorhabda spp.; Coleoptera:Chrysomelidae) have been introduced into the western United States as biocontrol agents for the invasive Eurasian shrub tamarisk (Tamarix spp.; Violales:Tamaricaceae). These beetles have since continued to spread and establish throughout the western United States. Another exotic insect, the tamarisk leafhopper (Opsius stactogalus, Fieber;Hemiptera: Cicadellidae), had previously become established in these areas and now shares tamarisk as a host plant with the beetles. To assess more carefully the potential for interactions between leafhoppers and beetles, field censuses and cage studies were conducted to determine the phenologies and potential interactions of O. stactogolus and D. carinulata when attacking Tamarix ramosissima (Ledebour) in western Colorado. The leafhopper underwent development through at least three generations per season, whereas the beetle was shown to develop through two generations per season. Variation in leafhopper abundance was associated with the extent and type of foliar damage to tamarisk trees. Individual trees with greatest D. carinulata abundances and subsequent defoliation had significantly reduced O. stactogalus abundances thereafter. Abundance of O. stactogalus was also shown to vary significantly among tamarisk plants in cage settings where leafhoppers were given the choice of potted tamarisk plants with ~50% damage to foliage from D. carinulata versus undamaged plants. In contrast, D. carinulata abundance was not shown to differ strongly in response to O. stactogalus damage in the field or in cage experiments. Field results across sites, however, showed similar trends of reduced beetle abundance on plants more heavily attacked by leafhoppers, and larval growth tests suggested slight reductions in larval pupation and adult emergence of D. carinulata when grown on O. stactogalus-damaged tamarisk. It is not clear if slight tendencies in D. carinulata abundance along with much stronger responses in O. stactogalus abundance were the result of limited plant material, rather than an induced plant defense. It is clear, however, that these specialist herbivores are interacting in an asymmetric competitive fashion while feeding on the same host plant.

No description available.