Predation of oral pathogens by Bdellovibrio bacteriovorus 109J.

Periodontal diseases are multifactorial infections elicited by a complex of primarily gram-negative bacteria that interact with host tissues and lead to the destruction of the periodontal structures. Bdellovibrio bacteriovorus is a gram-negative bacterium that preys upon other gram-negative bacteria. It was previously shown that B. bacteriovorus has an ability to attack and remove surface-attached bacteria or biofilms. In this study, we examined the host specificity of B. bacteriovorus strain 109J and its ability to prey on oral pathogens associated with periodontitis, including; Aggregatibacter actinomycetemcomitans, Eikenella corrodens, Fusobacterium nucleatum, Prevotella intermedia, Porphyromonas gingivalis and Tannerella forsythia. We further demonstrated that B. bacteriovorus 109J has an ability to remove biofilms of Ei. corrodens as well as biofilms composed of A. actinomycetemcomitans. Bdellovibrio bacteriovorus was able to remove A. actinomycetemcomitans biofilms developed on hydroxyapatite surfaces and in the presence of saliva, as well as to detach metabolically inactive biofilms. Experiments aimed at enhancing the biofilm removal aptitude of B. bacteriovorus with the aid of extracellular-polymeric-substance-degrading enzymes demonstrated that proteinase-K inhibits predation. However, treating A. actinomycetemcomitans biofilms with DspB, a poly-N-acetylglucosamine (PGA) -hydrolysing enzyme, increased biofilm removal. Increased biofilm removal was also recorded when A. actinomycetemcomitans PGA-defective mutants were used as host cells, suggesting that PGA degradation could enhance the removal of A. actinomycetemcomitans biofilm by B. bacteriovorus.

Predation of human pathogens by the predatory bacteria Micavibrio aeruginosavorus and Bdellovibrio bacteriovorus.

AIMS: The focus of this study was to evaluate the potential use of the predatory bacteria Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus to control the pathogens associated with human infection. METHODS AND RESULTS: By coculturing B. bacteriovorus 109J and M. aeruginosavorus ARL-13 with selected pathogens, we have demonstrated that predatory bacteria are able to attack bacteria from the genus Acinetobacter, Aeromonas, Bordetella, Burkholderia, Citrobacter, Enterobacter, Escherichia, Klebsiella, Listonella, Morganella, Proteus, Pseudomonas, Salmonella, Serratia, Shigella, Vibrio and Yersinia. Predation was measured in single and multispecies microbial cultures as well as on monolayer and multilayer preformed biofilms. Additional experiments aimed at assessing the optimal predation characteristics of M. aeruginosavorus demonstrated that the predator is able to prey at temperatures of 25-37°C but is unable to prey under oxygen-limiting conditions. In addition, an increase in M. aeruginosavorus ARL-13 prey range was also observed. CONCLUSIONS: Bdellovibrio bacteriovorus and M. aeruginosavorus have an ability to prey and reduce many of the multidrug-resistant pathogens associated with human infection. SIGNIFICANCE AND IMPACT OF THE STUDY: Infectious complications caused by micro-organisms that have become resistant to drug therapy are an increasing problem in medicine, with more infections becoming difficult to treat using traditional antimicrobial agents. The work presented here highlights the potential use of predatory bacteria as a biological-based agent for eradicating multidrug-resistant bacteria, with the hope of paving the way for future studies in animal models.

Affinity purification of HC-Pro of potyviruses with Ni2+-NTA resin.

The HC-Pro of zucchini yellow mosiac virus (ZYMV) was found to bind to Ni2+-NTA resin with or without His-tagging. The binding stringency was similar to that observed in proteins with a zinc finger motif like the HC-Pro. Using this characteristic we developed an efficient and rapid method (2-3 h) for purification of the HC-Pro of several potyviruses. A dominant protein of about 150 kDa was extracted and identified as the HC-Pro of ZYMV by means of immunoblotting. About 150 microg of HC-Pro were partially purified from the soluble fraction of 1 g of leaves. High titers of HC-Pro protein were obtained from plants infected with four potyviruses [ZYMV, watermelon mosaic virus II (WMVII), papaya ringspot virus (PRSV) and turnip mosaic virus (TuMV)]. The HC-Pros of potato virus Y (PVY) and tobacco vein mottling virus (TVMV) did not bind to the Ni2+-NTA resin. The ZYMV-HC-Pro purified by the Ni2+-NTA resin could bind in vitro to ZYMV virions blotted onto a membrane. All the HC-Pros which had been successfully purified by the Ni2+-NTA resin were bound in vitro to membrane-blotted ZYMV coat protein. However, only the HC-Pros of ZYMV and WMVII were able to mediate aphid transmission of purified ZYMV virions. The purification procedure described herein is efficient and convenient, and enables HC-Pro for a number of potyviruses to be obtained in larger amounts and at higher purity than possible by means of most existing methods, based on ultracentrifugation.

An excitatory and a depressant insect toxin from scorpion venom both affect sodium conductance and possess a common binding site.

Two insect selective toxins were purified by gel-permeation and ion-exchange chromatographies from the venom of the scorpion, Leiurus quinquestriatus quinquestriatus, and their chemical and pharmacological properties were studied. The first toxin (LqqIT1) induces a fast excitatory contraction paralysis of fly larvae and is about 40 times more toxic than the crude venom. It is a polypeptide composed of 71 amino acids, including 8 half-cystines and devoid of methionine and tryptophan, with an estimated molecular weight of 8189 and a pI value of 8.5. The second toxin (LqqIT2) induces a slow depressant, flaccid paralysis of fly larvae. It is composed of 72 amino acids, including 8 half-cystines, is devoid of proline methionine and histidine, and has an estimated molecular weight of 7990 and a pI value of 8.3. The contrasting symptomatology of these toxins is interpreted in terms of their effects on an isolated axonal preparation of the cockroach in current and voltage clamp conditions. LqqIT1 (0.5-4 microM) induced repetitive firing of the axon which was attributable to two changes in the sodium conductance, a small increase in the peak conductance and a slowing of its turning off. LqqIT2 (1-8 microM) caused a blockage of the evoked action potentials, attributable to both a strong depolarization of the axonal membrane and a progressive suppression of the sodium current. Neither toxin affected potassium conductance. The two toxins differ mainly in their opposite effects on the activatable sodium permeability. In binding assays to a preparation of insect synaptosomal membrane vesicles, the two toxins were shown to competitively displace the radioiodinated excitatory insect toxin derived from the venom of the scorpion, Androctonus australis [( 125I]AaIT), which strongly resembles, in its chemistry and action, the LqqIT1 toxin. The present two toxins have demonstrated a strong affinity closely resembling the AaIT, with KD values of 0.4, 1.9, and 1.0 nM for LqqIT1, LqqIT2, and AaIT, respectively. These data suggest the possibility that the excitatory and depressant insect toxins share a common binding site associated with sodium channels in insect neuronal membranes.