The skin microflora comprises a wide array of inhabitants that play key roles in maintaining human health. The multitude of different habitats helps promote this diversity and combined with constant exposure to the outside environment, the skin microbiome is one of the more complex human microbiomes. On an individual basis, bacterial communities exhibit significant interpersonal variation with temporal stability at a variety of different skin locations ( ref). The constant shedding of these organisms, similar to that of skin cells, produces a microbial fingerprint that is left upon touching a surface. Furthermore, recent evidence suggests that each individual emits a unique microbial cloud that may prove useful for forensic identifications …show more content…
Briefly, cells obtained from a well isolated colony on an agar plate were resuspended in 50 l of cell lysis solution (.05 M Tris (pH 8), 1 mM 0.5M EDTA, 1% Triton X-100) contained in a 0.2ml PCR tube. The cell solution was lysed by incubation at 94℃ for 10 minutes in a T100 Thermal Cycler (Bio-Rad Laboratories, Irvine, CA). After centrifugation at 10000 × g for 5 minutes, 5 l of the supernatant was used as DNA template for amplification by the Polymerase Chain Reaction (PCR). Reaction mixtures consisted of DNA template, .25 µM each of 8F and 1492r primers (ref??), and 2X GoTaq Green Master Mix (Promega, Fitchburg, WI) in a final reaction volume of 25 µl. Amplification consisted of an initial denaturation step of 94oC for 5 min, followed by 30 cycles of 94oC for 1 min, 50oC for 1 min, and 72oC for 1 min 30 sec, with a final extension of 72oC for 5 min. Electrophoresis on 1% agarose gel under standard conditions was used to visualize an approximate 1600 base pair amplicon. DNA sequencing of the amplified 16S rRNA gene was performed by MCLabs (South San Francisco, CA) followed by analysis using the Basic Local Alignment Search Tool (BLAST) to make genus and species …show more content…
Twenty-nine total isolates of Staphylococcus epidermidis with approximate equal distribution among the three individuals (designated with either an F, M, or Y in the strain name) were randomly chosen. Total DNA was isolated from each isolate using a ZR Fugal/Bacterial DNA MicroPrep kit (Zymo Research, Irvine, CA) according to the manufacturer’s instructions. Absorbance at 260 and 280 nm using a P330 Nanophotometer (Implen, Westlake Village, CA) was used to measure the purity and concentration of isolated DNA. A MLST scheme examining seven different housekeeping genes previously implemented for clinical Staphylococcus epidermidis isolates was employed in this study (Thomas et al 2007). Briefly, DNA templates prepared from each of the 29 isolates and were used in the amplification of araC, aroE, gtr, mutS, pyrR, tpi, and yqiL using gene specific primers as described (Thomas 2007) and 2X GoTaq Green Master Mix in a final reaction volume of 25 µl. Electrophoresis on 1% agarose gel under standard conditions was used visualize gene specific amplicons followed by DNA sequencing at MCLabs. Phylogenetic analysis were conducted using MEGA version 7 (Kumar
After 48 hours of incubation the agar plates were viewed. Individual colonies were tested for successful isolation by gram staining and then viewing the stained bacteria under a microscope. Isolation was successful. One colony of each unknown bacteria was transferred to an agar slant for growth. The agar slants were stored at room temperature over the weekend so that they would not grow too much.
The purpose of this study is to identify an unknown bacterium from a mixed culture, by conducting different biochemical tests. Bacteria are an integral part of our ecosystem. They can be found anywhere and identifying them becomes crucial to understanding their characteristics and their effects on other living things, especially humans. Biochemical testing helps us identify the microorganism present with great accuracy. The tests used in this experiment are rudimentary but are fundamental starting points for tests used in medical labs and helps students attain a better understanding of how tests are conducted in a real lab setting. The first step in this process is to use gram-staining technique to narrow down the unknown bacteria into one of the two big domains; gram-negative and gram-positive. Once the gram type is identified, biochemical tests are conducted to narrow down the specific bacterial species. These biochemical tests are process of elimination that relies on the bacteria’s ability to breakdown certain kinds of food sources, their respiratory abilities and other biochemical conditions found in nature.
In short, the book 'Deadly Companions: How Microbes Shaped Our History' explains the dynamism, the changing interactions between humans and microbes, and the way and extent to which these interactions have influenced the human cultural history. This book is very useful because it tell us that we must strive to understand what makes microbes successful as we are faced with unrelenting microbial drug resistance. Microbes mutate to fatal human pandemics and it’s for this reason that we need to adopt a microbe centric world views.
Amplification reaction was done in a 25.0 µL reaction mixture containing 0.4 µL DNA (from DNA extraction), 5.0 µL of 10X PCR reaction buffer, 14.2 µL of sterelized dH2O, 2.0 µL of magnesium chloride (MgCl2, 25 mM), 1.0 µL nucleotide/dNTP mix (10 Mm), and 0.4 µL of 5 u/µL Taq DNA polymerase for each primer namely respectively. The components and the volume used for the amplification reactions are listed in Table 3.2. For the reaction, PCR reaction was performed in a programmable gradient-enabled thermocycler (Bio-Rad MyCycler™ Thermal Cycler).
Staphylococcus epidermidis is classified as a bacterium. This particular bacterium is identified in laboratory testing by its forms of clusters and a violet stain. The violet stain indicates that the staphylococcus epidermidis is gram- positive meaning that it is sphere shaped and does not contain an outer cell membran). This type of Staphylococcus does not typically cause disease in healthy people. However, S. epidermidis is quite the opportunist.
For the original analysis, the corrected pairwise distance will be calculated using the Jukes–Cantor and the Maximum Composite Likelihood Model. The Jukes–Cantor model assumes that the rate of nucleotide substitution or all nucleotides (C, A, T and G) are equal, that nucleotide frequencies are equal, that there is an equal rate of substitution among sites, and does not correct for the lower rate of transversion substitutes in comparison to transitional substitutions (Jukes and Cantor, 1969). The Maximum Composite Likelihood takes into account the phylogenic relationship between sequences, using the sum of the log likelihoods of the bases as the composite likelihood. Both pair wise distances and substitution parameters are estimated using the Maximum Composite Likelihood (Tamura et al. 2004). Both models should yield different maximum sequence divergence and average divergence that can then be compared to the original paper. With sequence divergence data, the temporal origin of the genus can be identified. The two alternate models to the Kimura-2 parameter will be analyzed to discuss which methods yield results closest to the expected time origin of the genus
Two unknown bacteria were provided that were labeled “12A” and “12B”. Both bacteria were then used individually following aseptic technique to create heat-fixed smears. Both bacteria were strategically placed on the slide in order to prevent cross contamination and inaccurate results. A total of three slides were made in order to ensure accuracy. The heat-fixed slides were then Gram stained using the procedure listed in the manual; followed by observation under a Leica Dm500 microscope. The slides were observed under 4X,10X,40X and oil immersion and both unknown reactions were double checked by Dr. McLaughlin.
“David M. Lodge // Department of Biological Sciences // University of Notre Dame.” University of Notre Dame. 2009. Web.1 Mar. 2010.
Most parents remembered hearing about the importance of protecting their children from the sun, yet children are still playing in the sun without sunscreen or protective clothing. Many people these days are not taking skin cancer seriously after knowing its significance. Skin cancer has been a growing problem in the United States and millions of people have suffered from it every year. The three most common skin cancers are Melanoma, Basal cell, and Squamous cell, which can cause bumps, sores, growths, etc. Skin cancer is a deadly disease with many causes, but the advantage is that it can be prevented.
Normal microbiota are bacteria that naturally habitat the human body without causing harm. There are two forms, residential and transient. Residential microbiota are the organisms that have been with you since the day you were born and shall remain with you for the rest of your life; meanwhile, transient microbiota are the harmless bacteria that you come in contact with on a daily basis. These can easily be removed from the body meaning that transient microbes only stay with you for a short period of time. Both forms of normal microbiota are usually found on the surface of the skin, mucous membranes, digestive tract, upper respiratory system, intestines,
The human gut microbiota composition is individual specific and serves as finger prints. The gut microbiota plays an indispensable role in the maintenance of host's homeostasis. The microbiota acts as a barrier against pathogens and their invasion with a highly dynam...
The purpose of the study is to identify an unknown microorganism using multiple microbiology lab techniques. Through this process I will gain knowledge on how to perform these techniques as well as the importance of these tests on identifying unknown microorganisms. This is significant as the goal of this course is to familiarize ourselves with the common microbiology tests as well as the microorganisms we encounter in our daily activities.
Bacterial cells, like plant cells, are surrounded by a cell wall. However, bacterial cell walls are made up of polysaccharide chains linked to amino acids, while plant cell walls are made up of cellulose, which contains no amino acids. Many bacteria secrete a slimy capsule around the outside of the cell wall. The capsule provides additional protection for the cell. Many of the bacteria that cause diseases in animals are surrounded by a capsule. The capsule prevents the white blood cells and antibodies from destroying the invading bacterium. Inside the capsule and the cell wall is the cell membrane. In aerobic bacteria, the reactions of cellular respiration take place on fingerlike infoldings of the cell membrane. Ribosomes are scattered throughout the cytoplasm, and the DNA is generally found in the center of the cell. Many bacilli and spirilla have flagella, which are used for locomotion in water. A few types of bacteria that lack flagella move by gliding on a surface. However, the mechanism of this gliding motion is unknown. Most bacteria are aerobic, they require free oxygen to carry on cellular respiration. Some bacteria, called facultatibe anaerobes can live in either the presence or absence of free oxygen. They obtain energy either by aerobic respiration when oxygen is present or by fermentation when oxygen is absent. Still other bacteria cannot live in the presence of oxygen. These are called obligate anaerobes. Such bacteria obtain energy only fermentation. Through fermentation, different groups of bacteria produce a wide variety of organic compounds. Besides ethyl alcohol and lactic acid, bacterial fermentation can produce acetic acid, acetone, butyl alcohol, glycol, butyric acid, propionic acid, and methane, the main component of natural gas. Most bacteria are heterotrophic bacteria are either saprophytes or parasites. Saprophytes feed on the remains of dead plants and animals, and ordinarily do not cause disease. They release digestive enzymes onto the organic matter. The enzymes breakdown the large food molecules into smaller molecules, which are absorbed by the bacterial cells. Parasites live on or in living organisms, and may cause disease. A few types of bacteria are Autotrophic, they can synthesize the organic nutrients they require from inorganic substances. Autotrophic bacteria are either photosynthetic or Chemosynthetic. The photosynthetic bacteria contain chlorophyll that are different from the plant chlorophyll. In bacterial photosynthesis, hydrogen is obtained by the splitting of compounds other than water.
Tamura, K., Peterson, D., Peterson, N., Stetcher, G., Nei, M., and Kumar S. (2011) MEGA5: Molecular Evolutionary Genetics Analysis using Maximum Likelihood, Evolutionary Distance, and Maximum Parsimony Methods. Molecular Biology and Evolution. Retrieved February 01, 2014 from http://mbe.oxfordjournals.org/
Although many microbes become harmful, some remain harmless, and others become beneficial to eukaryotes. Microbes exist universally, so they live everywhere. Microbes that exist in soil may affect gardeners, farmers, or children. Children tend to chew on their nails and put their fingers in their mouth, this presents the microbe to the point of entry. Microbes in foods and water effects everyone. Bacteria replicates everywhere, including in food and water, however, viruses may only replicate inside a host cell. Necessities of life include food and water, so humans constantly digest bacteria. Once inside the body, microbes proliferate to cause disease or infection. When cooking a meal, the heat’s constantly destroying microbes, but it takes up to two hours of cooking to destroy all microbes. Heat destroys most microbes after 30 minutes at boiling point. Eating leftover foods will increase the risk of infectious disease. While frigerated, microbes continue to proliferate, but at a much slower rate. Beds and furniture contain many microbes. Think about it, when a person comes home from a long day of work or school, the first thing on their mind is to relax. So, they sit on their sofa, or lie on their bed, while still wearing the clothes they had on all day. The microbes in the air constantly attach to the external surface. Microbes become inhaled while breathing and crawl off macro-organisms to adhere onto everything. Like humans, microbes include parts of the life cycle. Many humans may not have much knowledge about microbes, since some don’t wash their hands, and some allow children to play in dirt or mud. Some people don’t wash their sheets often, and some don’t shower or brush their teeth as required. This shows the lack of knowledge that most individuals have about microbes. Microbes exist everywhere, yet many individuals still require hygiene