Kirsti Erkomaa

Detection of microcystins with protein phosphatase inhibition assay, high-performance liquid chromatography–UV detection and enzyme-linked immunosorbent assay: Comparison of methods

Abstract A colorimetric protein phosphatase inhibition assay (PPI assay), a commercial enzyme-linked immunosorbent assay (ELISA) test and different HPLC methods using UV detection were compared for the detection of cyanobacterial hepatotoxins, microcystins (MCYST) and nodularin. The suitability of the methods to detect different toxin variants was evaluated by using pure toxins and laboratory cultures as well as water and bloom samples of toxic cyanobacteria. The emphasis of the study was on the analysis of polar demethyl microcystin variants that are common in nature but for which there exist no commercial standards. The IC 50 values of MCYST-LR for the PPI assay and the ELISA test were 2.2–2.5 and 0.26–0.38xa0μgxa0l −1 , respectively. The most important factors that decreased toxin recovery in sample treatment were the use of C 18 cartridges and polypropylene containers. Good recoveries of toxins were obtained by using hydrophilic–lipophilic balanced (Oasis HLB, Waters) cartridges for concentrating the samples. The results obtained with the PPI assay, the ELISA test and HPLC correlated quantitatively well with the exception of [ d -Asp 3 ] microcystins. Concentrations of [ d -Asp 3 ]MCYST-RR measured with the PPI assay were only 5% of those obtained by the ELISA test and HPLC. Concentrations of hydrophobic microcystin variants were lower when analysed with ELISA than with the other methods. The World Health Organisation (WHO) has set a guideline value of 1xa0μgxa0l −1 for the world-wide most common microcystin variant, MCYST-LR in drinking water. Since the quantitative ranges of the PPI assay and the ELISA test are within microcystin concentrations in natural waters, and both tests are easy to perform, they show potential for routine use in the screening and monitoring of microcystins from drinking water supplies and from recreational waters.

Effects of phytosterols on zebrafish reproduction in multigeneration test

Zebrafish from mixed sex populations were exposed continuously across three generations to two phytosterol preparations both containing -sitosterol. The phytosterols were isolated from wood and soy beans. Blood vitellogenin levels and sex ratio changes were used as intermediate indicators of the reproduction failures. Both sterol preparations caused vitellogenin induction in the exposed fish. The wood sterol changed the sex ratio of the exposed fish. In generation F1, the predominant sex was male, and in generation F2 it was female. The soy sterol in the used test concentration was lethal to the exposed fish in generation F1. This multigeneration test evidenced that phytosterols containing -sitosterol disrupt the reproduction system of zebrafish by changing the sex ratios and by inducing the vitellogenin production in the exposed fish.

The impact of crop plant cultivation and peat amendment on soil microbial activity and structure

AbstractA rapid means for restoring soil fertility could be addition of peat to the plough layer. The impact of cultivation of eight different crops (the joint impact of plant and the management tailored for each plant), with and without soil amendment by peat treatment on soil microbiological, physical and chemical properties was assessed for two consecutive growing seasons. As a measure of the functional diversity of soil microbial community we estimated the activity of several different extracellular soil enzymes using the ZymProfiler® test kit. ATP content was measured to yield information on the amount of the active microbial biomass, and phospholipid fatty acid (PLFA) profiles were analysed to reveal the microbial community structure. The enzyme activity patterns of the soil samples indicated several differences due to the different crops and years but ATP content and PLFA profiles were rather stable. However, microbial biomass as total amount of PLFAs depended on the plant and peat treatment and ATP content varied between the years. The effects of the peat treatments were less clearly indicated by the biological parameters one or two years after the amendment, as only arylsulphatase and β-xylosidase activities were affected in both the years. Soil moisture, affecting enzyme activities, depended on the year and crop plant and peat addition increased it.n Abbreviations: AMC – 7-amino-4-methylcoumarin; AP – aminopeptidase; ATP – adenosine triphosphate; Cmic– microbial biomass carbon; DNA – deoxyribonucleic acid; EC – electrical conductivity; FAME – fatty acid methyl ester; fw – fresh weight; MUF – 4-methylumbelliferyl; na – not added; Nmic– microbial biomass nitrogen; PDE – phosphodiesterase; PLFA – phospholipid fatty acid; PME – phosphomonoesterase; SOM – soil organic matter

Virulence genes of Aeromonas isolates, bacterial endotoxins and cyanobacterial toxins from recreational water samples associated with human health symptoms

Exposure to cyanobacterial water blooms has been associated with various kinds of adverse health effects. In addition to cyanobacteria and their toxins, the bacteria associated with cyanobacteria could also be the etiological agents. We isolated Aeromonas strains (n = 176) from water samples (n = 38) taken from sites where cyanobacteria were suspected to have caused human health symptoms, of which fever and gastrointestinal symptoms were the most common. The isolates were screened by PCR for six virulence gene types (12 genes). The majority (90%) of the strains contained at least one of the virulence genes. Most common amplification products were those of genes (act/aerA/hlyA) that encode cytotoxic enterotoxin and haemolytic products. The genes encoding cytotonic enterotoxins (ast and alt), phospholipase (lip/pla/lipH3/alp-1), elastase (ahyB) and flagellin subunits (flaA/flaB) were also present in 5-37% of the Aeromonas strains. Analysed toxins (cyanobacterial hepatotoxins and neurotoxins, and bacterial endotoxins) were not detectable or were present in only low concentrations in the majority of the samples. The results indicated that the toxins were unlikely to be the main cause of the reported adverse health effects, whereas more attention should be paid to bacteria associated with cyanobacteria as a source of health effects.

Finnish experiences of QUASIMEME I

The authors outline the institutional benefits and improvements in analytical methodology arising as a result of their participation in the QUASIMEME project.