Undergraduate honors paper--Mount Holyoke College, 2006. Program in Biochemistry. / Includes bibliographical references (leaves 105-111).

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.

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.

Universiẗat, Diss., 2004--Leipzig.

The purpose of this study was to focus attention on the physiology of the bdellovibrio-host interaction and to determine the metabolic requirements for this reaction. Since bdellovibrio is an aerobic organism, direct measurements of respiration, turbidity, and viable cell counts are reliable indications of the metabolic activity of the cells. It was determined that the metabolic requirements for the parasitic interaction are constituents from either metabolically active host cells or cells which are capable of at least some metabolic activity. The nutritional requirements of host-independent bdellovibrios suspended in buffer are not met by the presence or absence of viable or nonviable Enterobacter aegnes. Unlike the HD bdellovibrios, the HI bdellovibrios lack the ability to make economical use of their self-digesting processes.

Durch den exzessiven Gebrauch von Antibiotika haben sich in den letzten Jahren zunehmend Resistenzen herausgebildet. Eine potentielle Alternative zu konventionellen Antibiotika sind prädatorische Bakterien. Das Bakterium Bdellovibrio bacteriovorus hat einen zweiphasigen Lebenszyklus bestehend aus einer Angriffsphase, in der es andere gram-negative Bakterien jagt, und einer Wachstumsphase, in der es das Zytoplasma eines Wirtes für die eigene Reproduktion nutzt. Für einen künftigen Einsatz von B. bacteriovorus als Antibiotikum müssen die Prozesse des Lebenszyklus verstanden werden. Das Proteom von B. bacteriovorus wurde bisher jedoch nur sehr wenig untersucht. Daher wurden in dieser Arbeit mithilfe der Massenspektrometrie Proteine von verschiedenen Zeitpunkten des Lebenszyklus von B. bacteriovorus relativ quantifiziert. Es konnten zahlreiche Proteine identifiziert werden, die zu spezifischen Zeitpunkten des Lebenszyklus hoch- oder herabreguliert werden. Die größten Unterschiede im Proteinmuster konnten zwischen der Angriffs- und der Wachstumsphase beobachtet werden. In der Angriffsphase sind einige Proteine herabreguliert, die mit der Proteinexpression im Zusammenhang stehen. Weiterhin wurde bestätigt, dass sich junge und gealterte Zellen der Angriffsphase deutlich voneinander unterscheiden, womit die Angriffsphase eigentlich aus zwei Phasen besteht. Auf Grundlage der Ergebnisse und eines Vergleiches mit Transkriptionsdaten wurde die Vermutung aufgestellt, dass B. bacteriovorus Proteine, welche spezifisch für die Angriffsphase sind, bereits während der Wachstumsphase synthetisiert. Im Zusammenhang mit der Forschung an B. bacteriovorus konnten auch neue Impulse bezüglich der MeCAT-basierten massenspektrometrischen Proteinquantifizierung angestoßen werden. In dieser Arbeit wurde unter anderem ein MeCAT-Reagenz mit Acrylamidfunktionalität entwickelt, welches erfolgreich als interner Standard für die Laserablation-ICP-MS von Polyacrylamidgelen verwendet werden kann. / Due to the excessive use of antibiotics, antibiotic resistance has increased over the last years. A potential alternative to conventional antibiotics are predatory bacteria. The predatory bacterium Bdellovibrio bacteriovorus has a biphasic life cycle consisting of an attack phase in which it hunts other gram-negative bacteria, and a growth phase in which it uses the cytoplasm of a prey cell as a substrate for its own reproduction. For future application of B. bacteriovorus as an antibiotic, it is necessary to understand the processes that occur during the life cycle. However, almost no information has been obtained regarding the proteome of B. bacteriovorus yet. Using mass spectrometry and an isotopic labelling strategy, proteins from different time points in the life cycle of B. bacteriovorus were quantified relatively to each other in this work. Numerous proteins were identified that are up- or down-regulated at specific time points in the life cycle. The largest differences in protein pattern existed between the attack phase and the growth phase, whereas only minor differences occurred within the growth phase. For instance, several proteins that appear to be down-regulated during the attack phase are related to protein expression. Furthermore, it was confirmed that there is a significant difference between young and aged cells of the attack phase. Therefore, the attack phase actually consists of two phases. Based on the results and on a comparison with transcription data, it was suggested that attack phase specific proteins of B. bacteriovorus are already synthesized during the growth phase. In connection with the research on B. bacteriovorus, new impulses regarding the MeCAT based protein quantification with mass spectrometry could be initiated. In this work, a MeCAT reagent with acrylamide functionality was developed, which can be used successfully as an internal standard for laser ablation ICP-MS of polyacrylamide gels.