Molecular Microbiology Laboratory

There are many reasons for identifying an unknown bacterium. The reasons range from medical purposes, such as determining if the unknown could cause ailments in living things or knowing what microorganisms are needed to make antibiotics. The experiment was done by applying methods in order to identify an unknown bacterium.

The purpose of this lab was to identify an unknown bacteria culture using differential tests. The identification of the unknown culture was accomplished by identifying the bacteria based on its specific metabolic characteristics and morphology. It is suggested that culture 11 is a sample of Enterobacter aerogenes.

And the unknown bacterial sample for Group 11 is… UNKNOWN. We were unable to identify our bacteria due to cross-contamination at some point during our research. This was first brought to our attention while running growth tests (Table 1). At that time, we discovered that most of the tests (EMB-lactose, PEA, Forms String in KOH) were showing that our sample was Gram-Positive, one test (Vancomycin) was showing that there was a Gram-Negative Bacteria growing.

The general principals for environmental cleaning are to ensure the hospital environment is as clean as possible to reduce the risk of infection, and that all precautions are taken in accordance to legislation and Healthcare policy’s and guidelines. “To prevent the transfer of micro-organisms which may cause infection, and to prevent the transfer of foreign protein which may cause adverse reaction and pose the risk of spreading diseases e.g. vCJD. “

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

This experiment illustrates the importance of handwashing and proves that hand washing is worth it. Since our hands are constantly coming into contact with ourselves and others, touching surfaces, grabbing objects, being sneezed into, etc., keeping our hands clean is one of the most effective, yet simple way we can take to avoid getting sick and spreading germs to others. Many diseases and conditions are spread by not washing hands with soap and clean, running warm water. “The human skin is a host to anywhere between 10,000-10,000,000 bacteria per square centimeter and since health care providers come into contact with pathogenic bacteria by being engaged in patient care, hand washing can reduce the risk of spreading diseases (page 3).” The objective of the experiment is to test the effectiveness of hand washing and demonstrate normal flora. This report presents the procedures and materials for the experiment, the experiment's results, and an analysis of those results.

My unknown organism #6 is Morganella morganii, which is a gram-negative bacillus rods commonly found in the environment and also in the intestinal tracts of humans, mammals, and reptiles as a normal flora. (3, 5) This bacterium Morganella morganii, was first discovered in the 1906 by a British bacteriologist named H. de R. Morgan. (2) Despite its wide distribution, it is an uncommon cause of community-acquired infection and is most often encountered inpostoperative and other nosocomial settings. (2, 3) Morganella morganii infections respond well to appropriate antibiotic therapy; however, its

Citrobacter Freundii is a species of bacteria that can be potentially harmful to humans. It is known to cause meningitis by protruding into the brain and replicating itself (1). The Citrobacter species has also been found as a cause of some urinary tract infections, diarrhea, and even gastrointestinal diseases and symptoms (3). C. Freundii can be located in a wide variety of soils and water (3). Lastly, it is also the cause of many nosocomial infections due to its presence in water (1).

The main idea of this experiment was to correctly identify the unknown bacteria, #3. Identification of unknown bacteria yields multiple benefits in many different areas in the research of microorganisms. In this experiment I performed many different test dealing with things such as the presence of enzymes, fermentation abilities and different chemical reactions. Observations made from the tests were then compared to a gram negative unknown chart in order to identify the bacteria. Based off of my results and the chart, I concluded the bacteria #3 was the bacteria Escherichia coli. E. coli is most commonly found in the intestines of warm blooded organisms. Most E. coli strands are non pathogenic however, there are strands

Two smears of the unknown bacterium #5 were inoculated while the second smear was used for a back up. The unknown bacterium dried for at least forty minutes. After the smears dried, the slides were heat fixed two times to ensure the stability of the organism. The slide was placed on top of the staining rack then over the small sink.

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

The main objective was to identify an unknown organism by utilizing skills we learned in our labs this semester. The purpose was to attain the possible identity of the unknown organism by actually performing biochemical tests and staining techniques we learned in lab. After performing and analyzing the results, we were able to use Bergey’s Manual of Systemic Bacteriology as a guideline to narrow down the genus of our organism test by test.

The bases of this experiment was to discover the identify of the unknown from three possible specimens: Klebsiella pneumonia, Escherichia coli, and Enterobacter aerogenes. Utilizaing the T streak technique, the bacteria was isolated into pure colonies for further study. The Gram Stain method was used to identity the morhphology of the bacteria such as the shape and whether the bacteria was Gram positive or Gram negative. Biochemical test were also used to help identify the unknown bacteria. The biochemical test used was the Triple Sugar Iron Agar, Sulfur Indole Motility test, Methyl Red test, Voges-Proskauer test, Citrate test, Urease test, and the Gelatin test. After observing the morphology of the bacteria using the Gram Stain method and utilizing all the possible biochemical test, the bacteria was identified to be Enterobacter aerogenes.

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.

The bacteria that was contained within Unknown tube #12 is believed to be Pseudomonas aeruginosa, Figure 1. The bacteria tested to be Gram Stain negative, producing a pink, red color retained from the staining process. When the species of bacteria was plated on nutrient media, the cells produced an irregular and spreading configuration as shown in Figure 2. This same plating test provided the margins and elevation, lobate and hilly, respectively. The specimen was stabbed in a Fluid Thioglycollate Medium (FTM) tube using an inoculated loop of the bacteria. The results of this experimentation indicate the type of oxygen requirement of the bacteria. The test found the bacteria to be aerobic as colonies of the bacteria began to form along the top of the FTM tube (Manual 2017).