The construction of a whole-cell biosensor for phosphonoacetate, based on the LysR-like transcriptional regulator PhnR from Pseudomonas fluorescens 23F

The phnA gene that encodes the carbon-phosphorus bond cleavage enzyme phosphonoacetate hydrolase is widely distributed in the environment, suggesting that its phosphonate substrate may play a significant role in biogeochemical phosphorus cycling. Surprisingly, however, no biogenic origin for phosphonoacetate has yet been established. To facilitate the search for its natural source we have constructed a whole-cell phosphonoacetate biosensor. The gene encoding the LysR-type transcriptional activator PhnR, which controls expression of the phosphonoacetate degradative operon in Pseudomonas fluorescens 23F, was inserted in the broad-host-range promoter probe vector pPROBE-NT, together with the promoter region of the structural genes. Cells of Escherichia coli DH5a that contained the resultant construct, pPANT3, exhibited phosphonoacetate-dependent green fluorescent protein fluorescence in response to threshold concentrations of as little as 0.5 µM phosphonoacetate, some 100 times lower than the detection limit of currently available non-biological analytical methods; the pPANT3 biosensor construct in Pseudomonas putida KT2440 was less sensitive, although with shorter response times. From a range of other phosphonates and phosphonoacetate analogues tested, only phosphonoacetaldehyde and arsonoacetate induced green fluorescent protein fluorescence in the E. coli DH5a (pPANT3) biosensor, although at much-reduced sensitivities (50 µM phosphonoacetaldehyde and 500 µM arsonoacetate).

Intracellular accumulation of polyphosphate by the yeast Candida humicola G-1 in response to acid pH

Cells of a newly isolated environmental strain of Candida humicola accumulated 10-fold more polyphosphate (polyP), during active growth, when grown in complete glucose-mineral salts medium at pH 5.5 than when grown at pH 7.5. Neither phosphate starvation, nutrient limitation, nor anaerobiosis was required to induce polyP formation. An increase in intracellular polyP was accompanied by a 4.5-fold increase in phosphate uptake from the medium and sixfold-higher levels of cellular polyphosphate kinase activity. This novel accumulation of polyP by C. humicola G-1 in response to acid pH provides further evidence as to the importance of polyP in the physiological adaptation of microbial cells during growth and development and in their response to environmental stresses.

Antifungal photodynamic therapy

In photodynamic antimicrobial chemotherapy (PACT), a combination of a sensitising drug and visible light causes selective destruction of microbial cells. The ability of light-drug combinations to kilt microorganisms has been known for over 100 years. However, it is only recently with the beginning of the search for alternative treatments for antibiotic-resistant pathogens that the phenomenon has been investigated in detail. Numerous studies have shown PACT to be highly effective in the in vitro destruction of viruses and protozoa, as well as Gram-positive and Gram-negative bacteria and fungi. Results of experimental investigations have demonstrated conclusively that both dermatomycetes and yeasts can be effectively killed by photodynamic action employing phenothiazinium, porphyrin and phthatocyanine photosensitisers. Importantly, considerable setectivity for fungi over human cells has been demonstrated, no reports of fungal resistance exist and the treatment is not associated with genotoxic or mutagenic effects to fungi or human cells. In spite of the success of cell culture investigations, only a very small number of in vivo animal. and human trials have been published. The present paper reviews the studies published to date on antifungal applications of PACT and aims to raise awareness of this area of research, which has the potential to make a significant impact in future treatment of fungal infections. (c) 2007 Elsevier GmbH. All rights reserved.

Specific growth rate plays a critical role in hydrogen peroxide resistance of the marine oligotrophic ultramicrobacterium Sphingomonas alaskensis strain RB2256

The marine oligotrophic ultramicrobacterium Sphingomonas alaskensis RB2256 has a physiology that is distinctly different from that of typical copiotrophic marine bacteria, such as Vibrio angustum S14. This includes a high level of inherent stress resistance and the absence of starvation-induced stress resistance to hydrogen peroxide. In addition to periods of starvation in the ocean, slow nutrient-limited growth is likely to be encountered by oligotrophic bacteria for substantial periods of time. In this study we examined the effects of growth rate on the resistance of S. alaskensis RB2256 to hydrogen peroxide under carbon or nitrogen limitation conditions in nutrient-limited chemostats. Glucose-limited cultures of S. alaskensis RB2256 at a specific growth rate of 0.02 to 0.13 h(-1) exhibited 10,000-fold-greater viability following 60 min of exposure to 25 mM hydrogen peroxide than tells growing at a rate of 0.14 h(-1) or higher. Growth rate control of stress resistance was found to be specific to carbon and energy limitation in this organism. In contrast, V. angustum S14 did not exhibit growth rate-dependent stress resistance. The dramatic switch in stress resistance that was observed under carbon and energy limitation conditions has not been described previously in bacteria and thus may be a characteristic of the oligotrophic ultramicrobacterium, Catalase activity varied marginally and did not correlate with the growth rate, indicating that hydrogen peroxide breakdown was not the primary mechanism of resistance. More than 1,000 spots were resolved on silver-stained protein gels for cultures growing at rates of 0.026, 0.076, and 0.18 h(-1). Twelve protein spots had intensities that varied by more than twofold between growth rates and hence are likely to be important for growth rate-dependent stress resistance. These studies demonstrated the crucial role that nutrient limitation plays in the physiology of S. alaskensis RB2256, especially under oxidative stress conditions.

Changes in antibiotic susceptibility in staphylococci habituated to sub-lethal concentrations of tea tree oil (Melaleuca alternifolia)

Aims: To investigate the effect of sub-lethal challenge with tea tree oil (TTO) on the antibiotic resistance profiles of staphylococci.

The application of colloidal gas aphrons in the recovery of fine cellulose fibres from paper mill wastewater

Colloidal gas aphrons (CGAs) are micron-sized gas bubbles of 25–30 µm in diameter produced by a high-speed stirrer in a vessel containing dilute surfactant solution. These bubbles, because of their small size, exhibit some colloidal properties. In this work, CGAs were used to separate fine fibres from a lean slurry of cellulosic pulp in a flotation column. The pulp fibres were recovered as foamate from the top. Sodium dodecyl sulphate at a concentration of 2.0 kg/m3 was used as a surfactant to generate the CGAs in a spinning disc apparatus. The results indicated that up to 70% flotation efficiency could be obtained within a short column height of 0.3–0.35 m. This technique can be applied to recover fine cellulosic pulp from paper-machine backwater.

Adsorption of Ni(SO4) on Malaysian rubber-wood ash

The possible use of wood ash as an adsorbent of nickel sulphate from dilute solutions and the effect of operating parameters were investigated in this study. The rate constants of adsorption were determined at different concentrations and temperatures. The applicability of the first-order reversible equation and an empirical kinetic model were tested to understand the kinetics of nickel sulphate removal at different concentrations. Pore diffusion was found as the rate-controlling step. The Langmuir and Freundlich isotherms were applied to find out the adsorption parameters. The activation energy of adsorption was -11.54 kJ mol-1. The value of the enthalpy change was ?H=-10.35 kcal mol-1.

Mass transfer correlation in flotation of palm oil by colloidal gas aphrons

Colloidal gas aphrons (CGAs) are micron-sized bubbles, which are produced by stirring a dilute surfactant solution at a high speed. In this work, CGAs have been used to clarify oily wastewater by flotation technique. The CGAs sparging rate was a critical factor that governed the efficiency of the process. A model for the determination of the mass transfer coefficient is also developed for the purpose of process design.

Respiratory syncytial virus (RSV) vaccines - Two steps back for one leap forward

Respiratory viruses are among the most important causes of morbidity and mortality worldwide. From a vaccine viewpoint, such viruses may be divided into two principle groups-those where infection results in long-term immunity and whose continued survival requires constant mutation, and those where infection induces incomplete immunity and repeated infections are common, even with little or no mutation. Influenza virus and respiratory syncytial virus (RSV) typify the former and latter groups, respectively. Importantly, successful vaccines have been developed against influenza virus. However, this is not the case for RSV, despite many decades of research and several vaccine approaches. Similar to natural infection, the principle limitation of candidate RSV vaccines in humans is limited immunogenicity, characterised in part by short-term RSV-specific adaptive immunity. The specific reasons why natural RSV infection is insufficiently immunogenic in humans are unknown but circumvention of innate and adaptive immune responses are likely causes. Fundamental questions concerning RSV/host interactions remain to be addressed at both the innate and adaptive immune levels in humans in order to elucidate mechanisms of immune response circumvention. Taking the necessary steps back to generate such knowledge will provide the means to leap forward in our quest for a successful RSV vaccine. Recent developments relating to some of these questions are discussed. (C) 2007 Elsevier B.V. All rights reserved.