Using measurements of muscle cell nuclear RNA with flow cytometry to improve assessment of larval condition of Walleye Pollock (Gadus chalcogrammus)

Nuclear RNA and DNA in muscle cell nuclei of laboratory-reared larvae of Walleye Pollock (Gadus chalcogrammus) were simultaneously measured through the use of flow cytometry for cell-cycle analysis during 2009–11. The addition of nuclear RNA as a covariate increased by 4% the classification accuracy of a discriminant analysis model that used cell-cycle, temperature, and standard length to measure larval condition, compared with a model without it. The greatest improvement, a 7% increase in accuracy, was observed for small larvae (<6.00 mm). Nuclear RNA content varied with rearing temperature, increasing as temperature decreased. There was a loss of DNA when larvae were frozen and thawed because the percentage of cells in the DNA synthesis cell-cycle phase decreased, but DNA content was stable during storage of frozen tissue.

Cultural methods of detection for microorganisms: recent advances and successes

Most microbiological methods require culture to allow organisms to recover or to selectively increase, and target organisms are identified by growth on specific agar media. Many cultural methods take several days to complete and even then the results require confirmation. Alternative techniques include the use of chromogenic and fluorogenic substances to identify bacteria as they are growing, selective capture using antibodies after short periods of growth, molecular techniques, and direct staining with or without flow cytometry for enumeration and identification. Future microbiologists may not use culture but depend on the use of specific probes and sophisticated detection systems.

Recent progress in the identification and determination of freshwater phytoplankton in the natural environment

The biomass of the phytoplankton and its composition is one of the most important factors in water quality control. Determination of the phytoplankton assemblage is usually done by microscopic analysis (Utermöhl's method). Quantitative estimations of the biovolume, by cell counting and cell size measurements, are time-consuming and normally are not done in routine water quality control. Several alternatives have been tried: computer-based image analysis, spectral fluorescence signatures, flow cytometry and pigment fingerprinting aided by high performance liquid chromatography (HPLC). The latter method is based on the fact that each major algal group of taxa contains a specific carotenoid which can be used for identification and relative quantification of the taxa in the total assemblage. This article gives a brief comparative introduction to the different techniques available and presents some recent results obtained by HPLC-based pigment fingerprinting, applied to three lakes of different trophic status. The results show that this technique yields reliable results from different lake types and is a powerful tool for studying the distribution pattern of the phytoplankton community in relation to water depth. However, some restrictions should be taken into account for the interpretation of routine data.

Immunodetection of planktonic algae

Interest in the identification and characterisation of cyanobacteria and dinoflagellates in aquatic environments is increasing rapidly due to the perceived roles of these organisms in primary production and nuisance aspects in terms of water treatment and public health. Techniques for the identification and quantification of these organisms currently are limited, and the application of molecular approaches provides fundamental taxonomic information and techniques of practical value. Antigenic properties of algal cells may be useful taxonomic markers. Immunodetection techniques utilise the specificity of the antibody/antigen association as a probe for recognising and distinguishing between microorganisms according to their cell- surface chemistry. Immunofluorescent detection of unicellular cyanobacteria and dinoflagellates has been studied with success in marine and freshwater ecosystems and a range of techniques and results are presented and discussed. The most recent advances in the study of planktonic algae have come with the application of continuous flow cytometric methods (CFC). Flow cytometry makes use of the autofluorescence properties of the algal cells, which alone can be used to demonstrate their presence and permit their quantification in natural water samples. When used in conjunction with immunolabelling techniques, the potential of CFC analysis is broadened to study the serological/strain composition of plankters in natural populations. Changes in algal strains represented within and between waters over periods of time are reported and discussed, along with the ecological issues thus raised.