Lab 7 – Sampling The Human Body Ninoska Garcia-ortiz 063

Lab 7 – Sampling the Human Body Ninoska Garcia-Ortiz 063 053 2 Objective: • •

Learn how to aseptically sample micro-environments on the human body Compare the density and diversity of microbial populations in various environments

Theory: -

All parts of the body are colonized by microorganisms that they come into contact with external environment, either directly or indirectly Microbes are found: skin, oral cavity, upper respiratory tract, intestinal tract and urogenital tract

Skin - Not a favorable place for growth of many microorganisms, it’s a relatively dry ecosystem, (exception are areas that contain numerous hair follicles and sebaceous and sweat glands - Limited nutrients - Salt concentration may slow growth - Acidic (pH 4 – 6) secretions are also somewhat inhibitory - Sweat and sebaceous gland secretions promote growth of normal flora - Secretions provide water and liberate nutrients: amino acids, lipids - Many disease causing microorganisms are inhibited by the utilization of lipids and lipolytic flora that result in the release of fatty acids - Indigenous microflora is primarily Gram + - Includes coccoid forms such as micrococci and staphylococci and diphtheroids such as the corynebacteria and propionibacteria, as well as some yeasts, such as Pityrosporum sp., that require oily substances for growth Throat - Normal flora contains a variety of microorganisms - Most common indigenous population include members of: staphylococci, streptococci, corynebacteria, spirochetes, actinomycetes and mycoplasmas, yeasts and viruses - Infection diagnosis often achieved by plating throat swab material unto blood agar, which readily differentiates throat flora by hemolytic type Nasal - Nasal passage flora similar to external skin flora - Gram + - Salt-tolerant organisms: Staphylococcus aureus, S. epidermis, aerobic diphtheroids predominate - 10 – 30% individuals carry S. aureus, coagulase positive, permanently or sporadically

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Easily tested with TGA agar

GI Tract - Microorganisms outnumber us 10 to 1 on our body, most of which inhabit GI Tract, - Predominantly mouth and large intestine - Feces contain about 1012 bacteria per gram (about 40% by wt of total fecal mass - Bacteria predominantly obligate anaerobes such as Bacteroides Urinary Tract - Normal flora of UT restricted to about first cm of urethra - Microorganisms not normally found in upper urethra, bladder, ureters or kidneys - ♀ reproductive tract possesses a large vaginal microflora throughout - Vaginal flora of ♀ of reproductive age is a mix of: lactobacilli, anaerobic Bacteroides, Clostridium, aerobic diphtheroids, staphylococci and yeasts - Lactobacilli lower vaginal pH 3.5 – 4.5 - Before puberty, after menopause, lactobacilli are less numerous in response to different hormonal levels.

Questions: 1– Mannitol Salts Agar – selective media. • Specifically selects for micrococci and staphylococci by way of the nutrients that it contains. Macconkey Agar - both selective and differential. • Selective due to the fact that it allows Gram negative enteric bacilli to grow and inhibits the growth of Gram positive bacteria. • Differential because it allows for differentiation of lactose fermenting and non lactose fermenting Gram negative enteric bacilli such as E.coli and P.vulgaris Tellurite Glycine Agar - selective media. • specifically tests for coagulase-positive staphylococci (which will grow after 24hrs.) Coagulase-negative staphylococci and other organisms are inhibited. Blood Agar – differential media. • Enriched medium which will allow for the growth of most micro-organisms. • Serves to differentiate between alpha-hemolytic organisms that produce a green olive zone, beta-hemolytic organisms that will produce a clear zone and gamma haemolytic organisms that will exhibit no detectable change. 2–

Alpha hemolysis - caused by oxidative effect of peroxide waste on heme Beta-hemolysis – caused by lysis of the red blood cells by the bacterial exoenzymes known as hemolysis. 3– Macconkey Agar, which is both a selective and a differential media, could be used to determine the presence of E. coli. This medium allows for the differentiation of lactose fermenting and non lactose fermenting Gram negative enteric bacilli such as E.coli and P.vulgaris. 4– The results for all the plates were as expected, except the growth on the Tellurite Glycerine Agar plates. The plate with the S. eareus gave the expected results, however; the plate with the S. epidermis had similar results. There was not supposed to be any growth in this plate. The growth can perhaps be explained by the fact that the plate was not inspected in the recommended 24 hour time frame, but instead 1 week later. This theory is further enforced by the evident “slower” growth on the plate that should not have had any growth. Conclusion: The objective of the lab, • •

To learn the difference between selective and differential media, and To learn how to use both types of media,

was achieved. The use of four different types of Agar were used to attain the objective. In the case of the Tellurite Glycerine Agar, we saw the importance of analyzing results of time sensitive experiments. The assumption that the growth was due to prolonged exposure is based on the fact that we were already told that coagulase-positive staphylococci would grow black colonies within 24 hours. The fact that there was growth in both plates, is an indication that the experiment was not done correctly. In this case, we are able to identify the infraction, analysis one week later, instead of 24 hours later.