Bacterial Growth and Measurement

Bacterial Growth

Appropriate environmental conditions are needed for bacterial growth including moisture, pH, temperature, osmotic pressure, atmosphere and nutrients. The time required for the production of daughter cells (generation time) is dependant on genetic and nutritional factors. Generation time can range from 20 mins to 20 hours.

Bacterial Nutrition

Bacterial growth requires nutrients to be available in the immediate environment. Bacteria are mainly chemoheterotrophs (using organic chemicals as energy). Specialised forms of agar plate containing different types of media have been developed to facilitate bacterial growth in controlled specific conditions. Therefore it is possible to determine the species of bacteria based on, amongst other things, the type of media they are able to grow on.
Most pathogenic bacteria can be grown on a nutrient medium at 37C, although they can grow between 20-45C. Most bacteria also grow optimally at a neutral pH and it is standard practice to buffer culture media so maintain it around pH 7. Another key determinant in bacterial growth is their preference for different types of atmosphere. Bacterial preference for oxygen means it is possible to assign all bacteria into four main groups; aerobes, anaerobes, facultative anaerobes and microaerophiles. Therefore anaerobic bacteria are unable to grow in an atmosphere containing oxygen.

Methods for Counting bacteria

Method	Technique	Comments
Microscopic Counting
Direct Smear	Smear on microscope slide from defined volume and dilution. Count performed using 50 microscope fields.	Slow and unreliable method and cannot differentiate viable and non-viable bacteria
Chamber Counting	Count fixed volume of bacterial suspension using a calibrated slide	No differentiation between viable and non-viable bacteria
Colony Counting
Spread Plate	Known vol of bacterial suspension spread onto agar plate and incubated for 24-48 hours	Number of colonies counted and expressed as colony-forming units (CFU)/ml of suspension
Pour Plate	A small vol of a known bactrial dilution is added to a petri dish with 20ml of molton agar at 45-48C and mixed	Colony counting carried out as above.
Miles-Misra	Diluted bacterial solution placed on plate in 5 seperate positions	Number of colonies counted and expressed as colony-forming units (CFU)/ml of suspension
Membrane Filtration	Following filtration of a known vol of bacterial dilution through a 0.22um pore size, filter is placed on an agar plate and incubated for 24-48 hours	The number of viable bacteria are expressed as CFU/ml of fluid
Other Counting Methods
Opacity Tubes	A bacterial suspension is matched visually with Mcfarland's opacity standard tubes	This test indicates the total bacterial cell numbers per ml
Electronic Counting	Counting machines such as the Coulter Counter can give rapid and accurate results	Reliability of results is dependant on quality control and test only gives a total cell count.

Quinn et al., P.J.. (2002) Veterinary Microbiology and Microbial Disease. Oxford: Blackwell Science Limited. pp.Bacterial Counting Techniques, P13.