What is the benefit of using molecular biology techniques for identifying bacteria rather than the traditional way List 3 reasons?
Need a crash course in methods for identifying microbes? Well, look no further: here, you’ll find an overview of the methods available for the identification of bacteria, yeast, or filamentous fungi to the species level. Show
Identifying Microbes at the Species level: The WhySpecies-level identification allows you to discriminate between two species from the same genus, which is often essential in the treatment of infectious diseases. For example, the bacterial genus Yersinia contains approximately 15 species, some of which form part of the normal human microflora, but others of which are serious pathogens and require medical treatment (e.g. Yersinia pestis is the causative agent of bubonic plague). Before we get into identification, let’s outline some of the main applications of accurate microbial identification:
So, how do we go about identifying microbes? Traditional methods rely on phenotypic identification using staining, culturing, and simple biochemical tests. Nowadays, more powerful molecular, immunological, and biochemical analytical methods complement and sometimes replace traditional methods. We’ll look at both traditional and modern methods for identifying microbes below. Traditional Methods for Identifying Microbes1. Macroscopic FeaturesMacroscopic features encompass the overall appearance of a microorganism, including its shape, size, color, and smell (i.e. the features that you can see with the naked eye). You can often determine the type of microorganism by examining the gross morphological/macroscopic features on an agar culture. Examining Agar Cultures
2. Microscopic FeaturesThe following questions can take you a long way in identifying microbes using the microscope:
3. Staining and MicroscopyStains enable easier visualization under a microscope. Cytology microscopes have specific requirements to ensure clear differentiation between stained cells. Here’s an overview of the most popular microbiological stains. Gram StainingGram staining is often the go-to test in bacterial identification. This purple stain, based on the crystal violet dye, is named after the Danish bacteriologist Hans Christian Gram, who developed it. Typical Gram-positive bacteria include Bacillus, Staphylococcus, Streptococcus, and Clostridium spp., while Escherichia, Helicobacter, and Salmonella spp. are Gram-negative. Certain bacteria are Gram-variable and, therefore, aren’t amenable to Gram-staining. Endospore StainingThis involves applying a stain to a bacterial sample to check for the presence of spores. Because not all bacteria produce spores, this information can be useful in identification. Several spore stains are available, but malachite green is probably the most popular. Ziehl-Neelsen StainingThis is an important tool for the staining of Mycobacterium tuberculosis (TB), which can’t be Gram-stained. The red stain carbol fuchsin is used first, followed by a counterstain such as methylene blue. M. tuberculosis stain red while other bacteria stain blue. Stains for Fungi and YeastSeveral fungal stains exist, although they are generally non-specific. They help visualize fungal elements for identification rather than discriminate between fungal species. Examples of fungal stains include:
4. Simple Biochemical TestsCatalase Testing
Oxidase Testing
Substrate Utilization Tests
These tests overlap with the concept of selective growth media. A wide range of selective media exists for the isolation and identification of bacterial and fungal species. Physiological Requirements for GrowthAddressing the following questions can help you when identifying microbes:
5. Dichotomous Identification KeysDichotomous keys contain a series of steps, with each step presenting descriptions of two distinguishing features (e.g. Gram-positive or Gram-negative), with a direction to the next step in the key, until the identity is known. The idea is that you use dichotomous identification keys alongside the methods outlined above to help you identify your organism of interest. Modern Methods for Identifying MicrobesAlthough still widely used, traditional methods for identifying microbes suffer from two major drawbacks. First, they are applicable only to organisms that can be cultured in vitro. Second, some strains exhibit unique biochemical characteristics that don’t fit the pattern of any known genus and species. Fortunately, many modern methods for identifying microbes aren’t dependent on live cultures, and they can often reveal minute differences between organisms that escape detection by traditional means. 6. Identifying Microbes Using PCR
7. Microarray-Based Identification
8. Immunological Identification
9. Chemical/Analytical IdentificationFatty Acid Profiling
Metabolic Profiles/Chemo-Profiling
Do you use any other methods for identifying microbes? Just drop us a line in the comments section and let us know! Resources
Originally published July 27, 2017. Reviewed and updated February 2021. Why is using a molecular technique for identification advantageous?With regard to infectious disease, molecular diagnostics has major advantages over the sole use of serology. Molecular diagnostics is a more sensitive method allowing detection of lower amounts of infectious agents and therefore giving the ability to detect infections earlier than was previously possible.
What are the advantages of molecular diagnosis over the traditional method for detecting infectious diseases?In addition to direct detection from clinical specimens, molecular methods can confirm a positive culture within a day compared to approximately four weeks using phenotypic methods. This has shortened the time for laboratory confirmation of suspected tuberculosis even for smear-negative but culture-positive cases.
What are the advantages of molecular based over the conventional blood culture method?Molecular assays performed on positive blood culture bottles has improved sensitivity compared to conventional culturing methods, and has decreased turnaround times compared to routine culture [81].
Why bacteria is used in molecular biology?The advantages of using bacteria for these studies include their simple noncompartmented structure, the accessibility of their genetic material, and the possibility of correlating the expression of a gene in the intact cell with its expression in a system composed of highly purified components.
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