In microbiology, streaking is a technique used to isolate a pure strain from a single species of microorganism, often bacteria. Samples can then be taken from the resulting colonies and a microbiological culture can be grown on a new plate so that the organism can be identified, studied, or tested.
The modern streak plate method has progressed from the efforts of Robert Koch and other microbiologists to obtain microbiological cultures of bacteria in order to study them. The dilution or isolation by streaking method was first developed in Koch's laboratory by his two assistants Friedrick Loeffler and Georg Theodor August Gaffky. This method involves the dilution of bacteria by systematically streaking them over the exterior of the agar in a Petri dish to obtain isolated colonies which will then grow into quantity of cells, or isolated colonies. If the agar surface grows microorganisms which are all genetically same, the culture is then considered as a microbiological culture.
Streaking is rapid and ideally a simple process of isolation dilution. The technique is done by diluting a comparatively large concentration of bacteria to a smaller concentration. The decrease of bacteria should show that colonies are sufficiently spread apart to affect the separation of the different types of microbes. Streaking is done using a sterile tool, such as a cotton swab or commonly an inoculation loop. Aseptic techniques are used to maintain microbiological cultures and to prevent contamination of the growth medium. There are many different types of methods used to streak a plate. Picking a technique is a matter of individual preference and can also depend on how large the number of microbes the sample contains.
The three-phase streaking pattern, known as the T-Streak, is recommended for beginners. The streaking is done using a sterile tool, such as a cotton swab or commonly an inoculation loop. The inoculation loop is first sterilized by passing it through a flame. When the loop is cool, it is dipped into an inoculum such as a broth or patient specimen containing many species of bacteria. The inoculation loop is then dragged across the surface of the agar back and forth in a zigzag motion until approximately 30% of the plate has been covered. The loop then is re-sterilized and the plate is turned 90 degrees. Starting in the previously streaked section, the loop is dragged through it two to three times continuing the zigzag pattern. The procedure is then repeated once more being cautious to not touch the previously streaked sectors. Each time the loop gathers fewer and fewer bacteria until it gathers just single bacterial cells that can grow into a colony. The plate should show the heaviest growth in the first section. The second section will have less growth and a few isolated colonies, while the final section will have the least amount of growth and many isolated colonies.
The sample is spread across one quadrant of a Petri dish containing a growth medium. Bacteria need different nutrients to grow. This includes water, a source of energy, sources of carbon, sulfur, nitrogen, phosphorus, certain minerals, and other vitamins and growth factors. A very common type of media used in microbiology labs is known as agar, a gelatinous substance derived from seaweed. The nutrient agar has a lot of ingredients with unknown amounts of nutrients in them. On one hand, this can be a very selective media to use because as mentioned bacteria are particular. If there is a certain nutrient in the media the bacteria could most certainly not grow and could die. On the other hand, this media is very complex. Complex media is important because it allows for a wide range of microbial growth. The bacteria growth can be supported by this media greatly due in part to the high amounts of nutrients. Choice of which growth medium is used depends on which microorganism is being cultured, or selected for.
Dependent on the strain, the plate may then be incubated, usually for 24 to 36 hours, to allow the bacteria to reproduce. At the end of incubation there should be enough bacteria to form visible colonies in the areas touched by the inoculation loop. From these mixed colonies, single bacterial or fungal species can be identified based on their morphological (size/shape/colour) differences, and then sub-cultured to a new media plate to yield a pure culture for further analysis.
Automated equipment is used at industrial level for streak plating the solid media in order to achieve better sterilization and consistency of streaking and for reliably faster work. While streaking manually it is important to avoid scratching the solid medium as subsequent streak lines will be damaged and non-uniform deposition of inoculum at damaged sites on the medium yield clustered growth of microbes which may extend into nearby streak lines.
Bacteria exist in water, soil and food, on skin, and intestinal tract normal flora. The assortment of microbes that exist in the environment and on human bodies is enormous. The human body has billions of bacteria which creates the normal flora fighting against the invading pathogens. Bacteria frequently occur in mixed populations. It is very rare to find a single occurring species of bacteria. To be able to study the cultural, morphological, and physiological characteristics of an individual species, it is vital that the bacteria be divided from the other species that generally originate in the environment. This is important in determining a bacterium in a clinical sample. When the bacteria is streaked and isolated, the causative agent of a bacterial disease can be identified.
Agar, or agar-agar, is a jelly-like substance consisting of polysaccharides obtained from the cell walls of some species of red algae, primarily from "ogonori" (Gracilaria) and "tengusa" (Gelidiaceae). As found in nature, agar is a mixture of two components, the linear polysaccharide agarose and a heterogeneous mixture of smaller molecules called agaropectin. It forms the supporting structure in the cell walls of certain species of algae and is released on boiling. These algae are known as agarophytes, belonging to the Rhodophyta phylum. The processing of food-grade agar removes the agaropectin, and the commercial product is essentially pure agarose.
An agar plate is a Petri dish that contains a growth medium solidified with agar, used to culture microorganisms. Sometimes selective compounds are added to influence growth, such as antibiotics.
Replica plating is a microbiological technique in which one or more secondary Petri plates containing different solid (agar-based) selective growth media are inoculated with the same colonies of microorganisms from a primary plate, reproducing the original spatial pattern of colonies. The technique involves pressing a velveteen-covered disk, and then imprinting secondary plates with cells in colonies removed from the original plate by the material. Generally, large numbers of colonies are replica plated due to the difficulty in streaking each out individually onto a separate plate.
A microbiological culture, or microbial culture, is a method of multiplying microbial organisms by letting them reproduce in predetermined culture medium under controlled laboratory conditions. Microbial cultures are foundational and basic diagnostic methods used as research tools in molecular biology.
Bacteriological water analysis is a method of analysing water to estimate the numbers of bacteria present and, if needed, to find out what sort of bacteria they are. It represents one aspect of water quality. It is a microbiological analytical procedure which uses samples of water and from these samples determines the concentration of bacteria. It is then possible to draw inferences about the suitability of the water for use from these concentrations. This process is used, for example, to routinely confirm that water is safe for human consumption or that bathing and recreational waters are safe to use.
A blood culture is a medical laboratory test used to detect bacteria or fungi in a person's blood. Under normal conditions, the blood does not contain microorganisms: their presence can indicate a bloodstream infection such as bacteremia or fungemia, which in severe cases may result in sepsis. By culturing the blood, microbes can be identified and tested for resistance to antimicrobial drugs, which allows clinicians to provide an effective treatment.
A growth medium or culture medium is a solid, liquid, or semi-solid designed to support the growth of a population of microorganisms or cells via the process of cell proliferation or small plants like the moss Physcomitrella patens. Different types of media are used for growing different types of cells.
Julius Richard Petri was a German microbiologist who is generally credited with inventing the device known as the Petri dish, which is named after him, while working as assistant to bacteriologist Robert Koch.
In microbiology, colony-forming unit is a unit which estimates the number of microbial cells in a sample that are viable, able to multiply via binary fission under the controlled conditions. Counting with colony-forming units requires culturing the microbes and counts only viable cells, in contrast with microscopic examination which counts all cells, living or dead. The visual appearance of a colony in a cell culture requires significant growth, and when counting colonies, it is uncertain if the colony arose from one cell or a group of cells. Expressing results as colony-forming units reflects this uncertainty.
Simmons' citrate agar is used for differentiating gram-negative bacteria on the basis of citrate utilization, especially for distinguishing Gammaproteobacteria of the family Enterobacteriaceae or even between species of the same genus. For example, Salmonella enteritidis would yield a positive (blue) result on Simmons’ agar and thus be distinguished from other Salmonella species like Salmonella typhi, Salmonella pullorum, and Salmonella gallinarum, which would yield a negative (green) result.
The Miles and Misra Method is a technique used in Microbiology to determine the number of colony forming units in a bacterial suspension or homogenate. The technique was first described in 1938 by Miles, Misra and Irwin who at the time were working at the LSHTM. The Miles and Misra method has been shown to be precise.
The disk diffusion test is a culture-based microbiology assay used in diagnostic and drug discovery laboratories. In diagnostic labs, the assay is used to determine the susceptibility of bacteria isolated from a patient's infection to clinically approved antibiotics. This allows physicians to prescribe the most appropriate antibiotic treatment. In drug discovery labs, especially bioprospecting labs, the assay is used to screen biological material and drug candidates for antibacterial activity. When bioprospecting, the assay can be performed with paired strains of bacteria to achieve dereplication and provisionally identify antibacterial mechanism of action.
In biology, axenic describes the state of a culture in which only a single species, variety, or strain of organism is present and entirely free of all other contaminating organisms. The earliest axenic cultures were of bacteria or unicellular eukaryotes, but axenic cultures of many multicellular organisms are also possible. Axenic culture is an important tool for the study of symbiotic and parasitic organisms in a controlled environment.
The Neogen Petrifilm plate is an all-in-one plating system made by the Food Safety Division of the Neogen Corporation. They are heavily used in many microbiology-related industries and fields to culture various micro-organisms and are meant to be a more efficient method for detection and enumeration compared to conventional plating techniques. A majority of its use is for the testing of foodstuffs.
Plate count agar (PCA), also called standard methods agar (SMA), is a microbiological growth medium commonly used to assess or to monitor "total" or viable bacterial growth of a sample. PCA is not a selective medium.
Fanny Hesse is best known for her work in microbiology alongside her husband, Walther Hesse. Following her initial suggestion of using agar as an alternative to gelatin, they were instrumental in pioneering agar's usage as a common gelling agent for producing media capable of culturing microorganisms at high temperatures.
In microbiology, the term isolation refers to the separation of a strain from a natural, mixed population of living microbes, as present in the environment, for example in water or soil, or from living beings with skin flora, oral flora or gut flora, in order to identify the microbe(s) of interest. Historically, the laboratory techniques of isolation first developed in the field of bacteriology and parasitology, before those in virology during the 20th century.
An inoculation needle is a laboratory equipment used in the field of microbiology to transfer and inoculate living microorganisms. It is one of the most commonly implicated biological laboratory tools and can be disposable or re-usable. A standard reusable inoculation needle is made from nichrome or platinum wire affixed to a metallic handle. A disposable inoculation needle is often made from plastic resin. The base of the needle is dulled, resulting in a blunted end.
Diagnostic microbiology is the study of microbial identification. Since the discovery of the germ theory of disease, scientists have been finding ways to harvest specific organisms. Using methods such as differential media or genome sequencing, physicians and scientists can observe novel functions in organisms for more effective and accurate diagnosis of organisms. Methods used in diagnostic microbiology are often used to take advantage of a particular difference in organisms and attain information about what species it can be identified as, which is often through a reference of previous studies. New studies provide information that others can reference so that scientists can attain a basic understanding of the organism they are examining.
In microbiology, colonial morphology refers to the visual appearance of bacterial or fungal colonies on an agar plate. Examining colonial morphology is the first step in the identification of an unknown microbe. The systematic assessment of the colonies' appearance, focusing on aspects like size, shape, colour, opacity, and consistency, provides clues to the identity of the organism, allowing microbiologists to select appropriate tests to provide a definitive identification.