Kristina Tag

Measurement of biodegradable substances using the salt-tolerant yeast Arxula adeninivorans for a microbial sensor immobilized with poly(carbamoyl) sulfonate (PCS) part I: construction and characterization of the microbial sensor

A microbial biosensor based on the yeast Arxula adeninivorans LS3 has been developed for measurement of biodegradable substances. Arxula is immobilized in the hydrogel poly(carbamoyl) sulfonate (PCS). The immobilized yeast membrane is placed in front of an oxygen electrode with -600 mV versus Ag/AgCl. Arxula is salt tolerant; it can give a stable signal up to 2.5 M NaCl in sample (120 mM in measuring cell). The sensors measurements are highly correlated to BOD5 measurements. It has a very high stability which can last for 40 day without any decrease in signal. The linear range of the sensor is up to a corresponding BOD value of 550 mg/l.

Arxula adeninivorans based sensor for the estimation of BOD

Abstract A microbial amperometric sensor for the determination of biochemical oxygen demand (BOD) using the yeast Arxula adeninivorans immobilized in polyvinylalcohol has been developed. The sensor with this microbial species has a wide substrate spectrum and allows BOD measurement with very short response times (70 sec), with an operation stability over 1 month, and a serial coefficient of ±5% when a standard solution containing 275 mg/L BOD was employed. A linear range was obtained up to 550 mg/L BOD using a glucose standard. The BOD-sensor was used to determine the BOD of various waste waters.

Measurement of biodegradable substances using the salt-tolerant yeast Arxula adeninivorans for a microbial sensor immobilized with poly(carbamoyl)sulfonate (PCS) Part II: Application of the novel biosensor to real samples from coastal and island regions

A microbial sensor for rapid measurement of the amount of biodegradable substances based on the salt-tolerant yeast Arxula adeninivorans LS3 has been developed especially for coastal and island regions. Our parameter, the so-called sensorBOD, that is available after only a few minutes, agrees with the 5-day value for the biochemical oxygen demand (BOD5) very well. We have employed the Arxula sensor in the short-time estimation and supervision of the BOD of both domestic and industrial wastewater with high salinity. The novel sensor makes it possible to monitor the different types of wastewater rapidly without pretreatment, and it can be used for an active process control of sewage treatment works. Compared to a commercially available sensor, the novel sensor achieves better agreement between sensorBOD and BOD5 measurements with salt containing samples.

Application of a wide-range yeast vector (CoMed™) system to recombinant protein production in dimorphic Arxula adeninivorans, methylotrophic Hansenula polymorpha and other yeasts

BackgroundYeasts provide attractive expression platforms in combining ease of genetic manipulation and fermentation of a microbial organism with the capability to secrete and to modify proteins according to a general eukaryotic scheme. However, early restriction to a single yeast platform can result in costly and time-consuming failures. It is therefore advisable to assess several selected systems in parallel for the capability to produce a particular protein in desired amounts and quality. A suitable vector must contain a targeting sequence, a promoter element and a selection marker that function in all selected organisms. These criteria are fulfilled by a wide-range integrative yeast expression vector (CoMed™) system based on A. adeninivorans- and H. polymorpha- derived elements that can be introduced in a modular way.ResultsThe vector system and a selection of modular elements for vector design are presented. Individual single vector constructs were used to transform a range of yeast species. Various successful examples are described. A vector with a combination of an rDNA sequence for genomic targeting, the E. coli- derived hph gene for selection and the A. adeninivorans-derived TEF1 promoter for expression control of a GFP (green fluorescent protein) gene was employed in a first example to transform eight different species including Hansenula polymorpha, Arxula adeninivorans and others. In a second example, a vector for the secretion of IL-6 was constructed, now using an A. adeninivorans-derived LEU2 gene for selection of recombinants in a range of auxotrophic hosts. In this example, differences in precursor processing were observed: only in A. adeninivorans processing of a MFα1/IL-6 fusion was performed in a faithful way.ConclusionrDNA targeting provides a tool to co-integrate up to 3 different expression plasmids by a single transformation step. Thus, a versatile system is at hand that allows a comparative assessment of newly introduced metabolic pathways in several organisms or a comparative co-expression of bottleneck genes in cases where production or secretion of a certain product is impaired.

Measurement of biodegradable substances with a mycelia-sensor based on the salt tolerant yeast Arxula adeninivorans LS3

Abstract The microbial sensor based on budding cells of the dimorphic yeast Arxula adeninivorans LS3 is one of the best suitable sensors for rapid measurement of biodegradable substances. However, Arxula is able to change its morphology (budding cells ⇔ mycelia) in response to the cultivation temperature. The suitability of such mycelia was tested as microbial sensor component. The mycelia-based sensor possess a similar linear response, stability, limit of determination and detection compared with the conventional budding cell-based sensor. However, differences exist in the storage capacity and substrate specificity: the mycelia-sensor has with six months a better storage stability as the conventional sensor, achieves a higher sensitivity for specific amino acids or sugars, shows a better correlation between sensorBOD and the BOD 5 -value with yeast extract as model wastewater and is more suitable for measurement of salt water.

Arxula adeninivorans LS3 as suitable biosensor for measurements of biodegradable substances in salt water

A microbial amperometric sensor based on the yeast Arxula adeninivorans was tested to determine its suitability for measuring biochemical oxygen demand (BOD) in salt water. The viability of cells immobilized onto the sensor membrane was hardly influenced up to 10% (w/v) NaCl in the sample, although the solubility of oxygen was affected. NaCl concentrations higher than 10% (w/v) caused a marked decrease in the oxygen solubility and deactivated the sensor. This outcome depended on the substrates used, e.g., alanine-, galactose- and acetic acid-sensor signals were influenced by any salt concentration whereas glucose-, glycerol-, maltose- and arginine-sensor signals were influenced only by higher salt concentrations. Sensor signals from yeast extract as well as glucose correlated with the quantity of these substances and with the salt concentration contained in the water. This correlation was linear up to 10% (w/v) NaCl and 0·125% (w/v) yeast extract or up to 10% (w/v) NaCl and 0·125% (w/v) glucose in the sample. The sensor signals are therefore influenced only by NaCl-determined solubility of oxygen and not by the physiological parameters of the immobilized cells. However, an increase of yeast extract- or glucose-concentrations in the presence of NaCl caused physiological effects on the sensor cells.