- Types of bacterial cultures
- Polymicrobial bacterial cultures
- Pure bacterial culture
- Mixed bacterial cultures
- Characteristics of bacterial cultures
- Examples of colonies according to their size
- Example of colonies according to their appearance
- Examples of colonies according to their texture
- Example of colonies according to their shape
- Example of colonies according to their borders
- Example of colonies according to their color
- Bacterial culture methods
- Facultative aerobic or anaerobic bacterial culture
- Anaerobic bacterial culture
- Bacterial culture in microaerophilia
- Materials and requirements
- materials
- Requirements to perform a bacteria culture
- Nutrients
- Hydrogen ion concentration (pH)
- Temperature
- Oxygen requirement
- Culture media
- Bacterial culture collection
- References
A bacterial culture is the result of the seeding of these microorganisms on nutritive media, in order for them to reproduce, resulting in the appearance of bacterial colonies in solid media and cloudiness in liquid media.
The sowing method is very important to be able to distribute the inoculum in such a way that the bacteria present are dispersed and can develop properly isolated colonies.
Various crops planted by the author in the Bacteriology Laboratory of the Biomedical and Technological Sciences Building, University of Carabobo. Source: composition of photos taken by the author MSc. Marielsa Gil.
The colonies obtained in the solid culture medium are the consequence of the proliferation of the microorganism that was seeded. Each colony starts from a single bacterium, which can multiply exponentially to form a macroscopically visible population.
The same occurs in liquid culture media but in this case bacterial growth is observed by turbidity.
Bacterial growth is possible when the chosen culture medium meets the nutritional and pH conditions necessary for the development of a particular bacterium. In addition, it is necessary to control other variables, such as temperature, incubation time, oxygen concentration, CO 2, among others.
Not all bacterial populations need the same requirements, although some may be common to almost all, such as that the medium contains peptones, it is also true that there are more demanding microorganisms that additionally require other specific substances, called growth factors.
For example, it can be mentioned that some Haemophilus need the presence of factor X (hemin) and factor V (NAD) to grow.
Types of bacterial cultures
Bacterial cultures can be polymicrobial, pure or mixed.
Polymicrobial bacterial cultures
Polymicrobial cultures are those that come from the sowing of a sample obtained from a place that has a normal microbiota, in which a certain pathogen can also be found.
Example: when developing a throat swab culture, a pathogen such as Streptococcus pyogenes may be found, but it will be accompanied by a common microbiota in the area.
In this case, a good striation is essential to allow the growth of the different types of bacteria present in isolation.
The colony that has characteristics compatible with a Streptococcus pyogenes strain will be carefully touched with the platinum loop and then planted in a virgin culture medium and thus obtain a pure culture of that microorganism.
From this pure culture, all the tests required to identify the bacteria can be carried out.
Pure bacterial culture
In order to efficiently identify a microorganism, one must work from a pure culture.
Pure cultures can be obtained as in the case of the previous example by reseeding an isolated colony of a polymicrobial culture, or when a sample that comes from a normally sterile place is planted and only one pathogen exists (growth of a single type of bacteria). Example: when a CSF is seeded from a patient with bacterial meningitis.
Another way is when reseeding from an old pure culture to obtain a renewed pure culture.
Mixed bacterial cultures
These cultures are composed of a mixture of microorganisms, as they occur in nature. In some industrial processes the use of mixed cultures is convenient.
It can be obtained through the union of several pure cultures, since these allow the study of bacterial interrelationships.
They are useful, for example, in the degradation of xenobiotics in industrial wastewater, since they act as biodegradable agents.
Characteristics of bacterial cultures
The morphological characteristics of the colonies obtained in bacterial cultures can be very varied.
Colonies can be very small, moderate, or large and may appear dry or mucoid, shiny or dull. Depending on the texture, it can vary between smooth and rough and, depending on the shape, they can be circular, flat, convex.
Depending on the color they can be: colorless, white, yellow, pink, fuchsia, red, orange, beige, grayish, greenish, brown, black or with a metallic luster, depending on the bacteria involved and the culture medium used.
The borders of the colonies can be regular or irregular. Others, on the other hand, may present a uniform film that is distributed in almost the entire medium called "swarming". This is characteristic of Proteus sp.
Some bacterial cultures emit odors that are quite characteristic of the species involved. For example, a culture of Pseudomonas aeruginosa has a characteristic fruit odor, while the genus Proteus has a characteristically putrid odor.
Examples of colonies according to their size
Extremely small: Mycoplasma pneumoniae, Francisella tularensis.
Small: Streptococcus sp, Enterococcus sp.
Moderate: Family Enterobacteriaceae
Large: Bacillus cereus, Pseudomonas aeruginosa.
Example of colonies according to their appearance
Dry: Lactobacillus confusus.
Mucoids: Klebsiella pneumoniae, Pseudomonas aeruginosa.
Brilliant: Klebsiella pneumoniae.
Opaque: Enterococcus faecalis, some strains of Neisseria gonorrhoeae.
Examples of colonies according to their texture
Smooth colonies: Micrococcus luteus
Rough colonies: Sarcina ventriculi on nutrient agar.
Example of colonies according to their shape
Circulars: Listeria murrayi and Micrococcus luteus on nutrient agar.
Flat: Staphylococcus equorum on nutrient agar.
Convex: Staphylococcus aureus, Klebsiella pneumoniae.
Example of colonies according to their borders
Rounded edge: Staphylococcus aureus, Escherichia coli.
Irregular border: Klebsiella pneumoniae, Pseudomonas aeruginosa.
Lobed edges: Bacillus sp.
Example of colonies according to their color
Transparent or colorless: Shigella sp on XLD agar.
White: Staphylococcus hominis on blood agar.
Beige: Bacillus sphaericus and Lactobacillus confusus on nutrient agar.
Yellow: Staphylococcus aureus and Elizabethkingia meningoseptica on blood agar, Pediococcus sp, Staphylococcus lentus on nutrient agar, Pink: Escherichia coli on XLD agar, Micrococcus roseus on nutrient agar.
Fuchsia: Escherichia coli on Mac Conkey agar.
Violet: Chromobacterium violaceum.
Reddish: Serratia marcescens at room temperature on nutrient agar.
Orange: Kurthia zopfii on nutrient agar.
Greyish: Sporosarcina ureae in soil extract agar.
Greenish: Pseudomonas aeruginosa on BHI agar.
Brown: Enterococcus sp on bile esculin agar.
Black: Salmonella enteritidis on SS agar.
With metallic luster: Escherichia coli on Endo agar.
Bacterial culture methods
Facultative aerobic or anaerobic bacterial culture
It is the most widely used cultivation method. This is due to the fact that, in the first place, most of the bacteria pathogenic for man are aerobic or facultative anaerobic; and secondly, it is much cheaper and easier than anaerobic cultures. Example: Mycoplasma pneumoniae and Escherichia coli respectively.
Anaerobic bacterial culture
This type of culture requires complete oxygen suppression. Culture media for anaerobic bacteria should generally contain reducing substances, such as: ascorbic acid, thioglycollate, cysteine or sulfur to eliminate the toxic effects of oxygen contained in the air.
Some contain indicators such as resazurin which is blue in the presence of oxygen and colorless in anaerobiosis. Example: Clostridium tetani.
Bacterial culture in microaerophilia
The culture plates are placed in a microaerophilic hood with a lit candle, covered. The candle consumes the oxygen and goes out. In this condition, some bacteria grow, such as Streptococcus sp.
Materials and requirements
materials
The materials required to perform a bacterial culture are: Petri dishes or tubes with culture media or broth, platinum loop, sample, Bunsen burner or oven.
Requirements to perform a bacteria culture
The culture of bacteria requires many elements that must be taken into account, such as the nutrients in the medium, pH, temperature, oxygen concentration, CO2, humidity, among others.
Nutrients
Bacterial cultures require culture media that contain macroelements and microelements. Among the macroelements, organic substances such as peptones, amino acids, carbohydrates as sources of carbon and nitrogen can be mentioned.
Among the microelements are inorganic substances or trace elements manganese, zinc, nickel, boron, chlorine, selenium, silicon, cobalt, copper, among others.
Hydrogen ion concentration (pH)
The concentration of hydrogen ions (H +) and hydroxyl ions (OH -) in the medium where the culture will develop is of vital importance, as these determine the pH.
The most commonly used pH is neutral (pH = 7.0), however there may be means where an acidic or alkaline pH is convenient, for example if you want to isolate an acidophilic or alkaliphophilic bacteria respectively.
Temperature
Temperature is a very important factor that influences the growth of bacterial cultures. Bacteria can be psychrophilic (grow at temperatures <20 ° C, mesophilic (between 20 ° C and 42 ° C), thermophilic (40 ° C to 70 ° C), hyperthermophilic (70 ° C - 105 ° C).
Oxygen requirement
Aerobic: they grow in the presence of oxygen.
Microaerophilic: they grow in the presence of 5-10% CO 2.
Strict anaerobes: grows in the absence of oxygen.
Facultative anaerobes: grows in the presence of oxygen or without it.
Aerotolerant: they grow well without oxygen and tolerate little oxygen.
Culture media
Culture media are special nutritional preparations that are prepared in the laboratory to obtain microbial growth or culture. These media vary in consistency, composition, and function. Each laboratory will prepare the type of culture medium that suits their needs.
Bacterial culture collection
There are institutions or organizations that are dedicated to the collection, characterization, maintenance and distribution of bacterial strains of clinical, environmental or industrial importance.
These strains are used for research work and to carry out quality controls on the culture media.
Example: the strains American Type Culture Collection, Venezuelan Center for Collection of Microorganisms (CVCM) and strains of the National Institute of Hygiene, Epidemiology and Microbiology of Cuba (INHEM), among others.
References
- Benavides G, Hermida A. Isolation and identification of native bacterial flora from the soil of the Cruz Verde and Guasca páramos. (Cundinamarca). 2008. Bogotá. Available at: javeriana.edu.co/biblos
- Hans S. General Microbiology. Omega editions. 1991. Barcelona, Spain. Available in: biolprocariotas.files
- Weng Z, Junco R, Díaz R. Collection of microbial cultures: Notes on their development. Rev Cubana Hig Epidemiol, 2003; 41 (1). Available at: scielo.sld.cu/scielo.
- Koneman E, Allen S, Janda W, Schreckenberger P, Winn W. (2004). Microbiological Diagnosis. 5th ed. Editorial Panamericana SA Argentina.
- Forbes B, Sahm D, Weissfeld A. (2009). Bailey & Scott Microbiological Diagnosis. 12 ed. Editorial Panamericana SA Argentina.