Cloning the Metagenome to Access the Biodiversity of Unculturable Bacteria

See Table 1 and Flow Chart 1 for results of Bacteria # 1 and Table 2 and Flow Chart 2 for results of Bacteria # 2.

The purpose of this study project was to carefully isolate and identify two unknown bacteria from a mixed culture. The ability to properly evaluate biochemical test results is also necessary for the identification to be successful. The goal was to apply all of the methods and techniques that have been learned in the microbiology laboratory course for the proper identification of unknown bacteria. A certain amount of bacteria that were used throughout the course were possible bacteria that could be found in a mixed culture. The bacteria that were identified in the mixed culture were Staphylococcus Aureus and Kocuria Rhizophila.

Often scientists work with bacteria that do not come in a labeled test tube— for example, bacterial samples taken from infected human tissue or from the soil—and the scientist must then identify the unknown microorganism in order to understand what behavior to expect from the organism, for example, a certain type of infection or antibiotic resistance. However, because of the relatively few forms of bacteria compared to animals and because of the lack of bacterial fossil records due to their asexually reproductive nature, the taxonomy used to classify animals cannot be applied to bacteria (Brown 275). In order to classify unknown bacteria, a variety of physiological and metabolic tests are available to narrow a sample down from the fathomless number of possibilities into a more manageable range. Once these tests have been performed, the researcher can consult Bergey’s Manual of Determinative Bacteriology, a systematically arranged and continually updated collection of all known bacteria based on their structure, metabolism, and other attributes.

The purpose of this lab was to identify two unknown bacteria cultures using various differential tests. The identification of these unknown cultures was accomplished by separating and differentiating possible

Bacteria are ubiquitous; they can be found on the skin, in the soil, and inside the body. Because of the very nature of this ubiquity, it is important to be able to determine between different strains of bacteria. An example of this is determining the causative agent for a disease so that the patient will be treated with the appropriate antibiotics. It may be important to determine the bacteria in a certain region, because like with enteric bacteria, it is normal to find them in the digestive tract as they are in a symbiotic relationship with our bodies in this area; however, they also cause opportunistic infections in places outside of the digestive tract to our detriment, such as with a urinary tract infection. Some strains of bacteria are common to nosocomial infections, and identifying these bacteria as such helps create the guidelines for healthcare workers in antiseptic technique. All of the morphology and characteristics of each strain of bacteria help us to better understand the role of bacteria in the body as well as helps us understand how they can cause illness, and what treatment regimen to set in place. In lab this semester, a sample of unknown

The purpose of the following study is to determine where the two unknown bacteria acquired in Microbiology lab should be classified in regards to temperature, pH level, and osmoregularity. It is important to classify bacteria in order to identify them. Identification of bacteria is important because they are not only useful but potentially dangerous as well. The identification of bacteria can lead to breakthroughs in healthcare regarding treatment of old and new diseases alike. Identifying bacteria can also be used in many other areas from better crop production through microbial pesticides to biological warfare. Their uses are endless as are their abilities to evolve and adapt to changing environments. That is why it is so important

The purpose of this lab was to identify two unknown bacteria cultures using various differential tests. The identification of these unknown cultures was accomplished by separating and differentiating possible bacteria based on specific biochemical characteristics. Whether the tests performed identified specific enzymatic reactions or metabolic pathways, each was used in a way to help recognize those specifics and identify the unknown cultures. The differential tests used to identify the unknown cultures were oxidase, catalase, lactose and sucrose fermentation, Kugler/iron agar, nitrate reduction, gelatin hydrolysis, starch hydrolysis, manitol salt, MR-VP, citrate, bile esculin,

In this experiment, an unknown bacterium was given to each individual student. The main purpose of this lab was to identify the given unknown bacteria going through a series of biochemical tests as one of the gram negative bacteria among six different Gram negative bacteria Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa and Salmonella typhimurium. At the very beginning, streaking method; T-streak technique was used to isolate the pure colonies. For the morphological identification of unknown bacteria, Gram Stain Method was done. Biochemical tests that were conducted for the experiment

The purpose of this study was to identify the unknown bacterium using biochemical tests and various methods that had been learned from previous the microbiology laboratory class. Identifying the unknown bacterium was determined by separating and differentiating possible

PCR is the amplification of DNA by denaturing, annealing, and extension of a DNA template. Specific sequences can be amplified using single-stranded DNA that complements the target sequences known as primers. This process heats DNA until the strands separate, then primers bind to the target regions. DNA polymerase enzymes and single base nucleotides (dNTPs) are used to synthesize new strands of DNA to the target sequence. The end product will contain large quantities of the target sequence (Bean et al. 2015). The most notable in phylogenic studies is the 16S rRNA gene, because of it’s highly conserved primer-binding site and hyper variable regions that provide species-specific sequences within bacteria and archaea (Kolbert and Persin 1999). This gene is a component of the 30S small subunit of prokaryotic ribosome’s and serves as the primary site of protein synthesis. (Woese and Fox 1977). The 16S rRNA sequence can be amplified and matched to national databases provided by the National Center for Biotechnology Information (NCBI) using software termed Basic Local Alignment Search Tool (BLAST) to find regions of similarity between biological sequences for bacterial identification. Thus, providing a cost effective and timely method when compared to biochemical

Purpose of Bergy’s Manual: based on ribosomal RNA sequences, which presumably reflect phylogenetic (evolutionary) relationships. Used for the identification of prokaryotes. 2nd edition on classification of prokaryotes.

The ten most abundant bacterial families detected in our study were Koribacteraceae (Acidobacteria), Acidobacteriaceae (Acidobacteria), Sphingobacteriaceae (Bacteroidetes), Geobacteraceae (proteobacteria), Auto67-4W (Pedosphaerae), Acetobacteraceae (proteobacteria),

A highly conserved gene will be used to identify a prokaryotic species isolated from the body. Fundamental lab techniques will be also explored and utilized, such as amplifying using PCR, cloning, and transforming the gene into a host cell. DNA electrophoresis and specific substrate plating will serve as analysis check points. The final product will be sequenced and compared to similar species to observe phylogenetic relationships.

2. Introduction: Each student was given unknown bacteria and was instructed to perform a variety of experimental tests that would help to identify their bacteria. During the process of identification, the unknown bacteria was added to many different testing medias using aseptic technique. They are as follows: lactose fermentation on eosin methylene blue (EMB), TSI (Triple Sugar Iron agar), Phenol red sucrose, the SIM test, H2S by SIM, IMViC (indole, motility, voges-proskauer, and citrate), Urease (urea broth), PDase (Phenylalanine Deaminase), Lysine Decarboxylase, and Ornithine Decarboxylase. Colonial morphology on EMB was used to

Each mixed culture that was tested had one gram positive and one gram negative bacterial species. The possible species of bacteria that could have been isolated from the mixtures included the following: Bacillus subtilis, Corynebacterium glutamicum, Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, Enterobacter aerogenes, Salmonella enterica, and Pseudomonas aeruginosa. The identities of the unknown species were determined through comparing the experimental data against data acquired from earlier experimentation.