Taurai Mutanda and Faizal Bux

Institute for Water and Wastewater Technology Durban University of Technology Durban, South Africa

CONTENTS

4.1 Introduction……………………………………………………………………………………………….. 45

4.2 Sampling and Culturing Microalgae…………………………………………………………… 46

4.3 Microalgal Preservation……………………………………………………………………………… 46

4.4 Enumeration Methods……………………………………………………………………………….. 47

4.4.1 Spectrophotometric Analysis………………………………………………………….. 47

4.4.2 Gravimetric Analysis………………………………………………………………………. 47

4.4.3 Counting Chambers……………………………………………………………………….. 48

4.4.4 Flow Cytometry……………………………………………………………………………… 48

4.5 Conclusion………………………………………………………………………………………………… 48

Acknowledgments………………………………………………………………………………………………. 50

References………………………………………………………………………………………………………….. 50

4.1 INTRODUCTION

The estimation of a microalgal population size is no easy task due to the micro­scopic size of the cells. Consequently, it is impossible to physically count them with the naked eye. The size of microalgae falls within the size of other microbes (e. g., bacteria) and, as a result, most of the methods used for microbial cell counting are also applicable to microalgae. In general, conventional microbiological proto­cols are available and are sufficient for cell enumeration despite the proliferation of modern and advanced techniques. It is recommended that the researcher choose a method after assessing the costs involved because some of the latest methods require very sophisticated and expensive equipment.

According to Caron et al. (2003), “the identification and enumeration of micro — organismal species in natural aquatic assemblages is an essential prerequisite for ecological studies of these populations.” Effective ecological studies of populations of colonial freshwater phytoplankton species are hampered by a lack of methods for cell enumeration (Box, 1981). Closely related microalgal groups must be accurately distinguished, and this is very crucial when these species pose health and environ­mental risks (Caron et al., 2003).

Microalgal cell population size is important when studying growth kinetics. It is also important to know the amounts of biochemical constituents such as pigments (e. g., chlorophyll-a). Microalgal cells can be enumerated directly using techniques such as light absorption and/or indirectly via surrogate determination of dry or wet biomass and/or measurement of cell components such as organic nitrogen, phosphorus, etc. (Guillard and Sieracki, 2005).

However, despite their common usage, direct methods and gravimetric measure­ments of microalgal cell dry weights are tedious, time consuming, and prone to errors that may exceed ±10% (Elnabarawy and Welter, 1984). Techniques for the estimation of the abundance of microalgal cells in natural microalgal samples are improving at a fast pace with the advent of electronic counting methods. Despite the progress made in the development of these advanced and sophisticated techniques, they still have major drawbacks, such as cost and the requirement of highly skilled personnel to operate the equipment. This chapter describes some of the popular methods that are available for the enumeration of microalgal cultures. The methods that are widely used are spectrophotometry, dry weight determinations, light micros­copy, haemacytometry, and flow cytometry.