The Lab Chronicles: My Experience in the Microbiology Laboratory – Week 1

For the past three years, I have attended the University of Ontario Institute of Technology (UOIT) pursuing a Bachelor of Health Science in Medical Laboratory Science. This degree is specific to the Medical Laboratory Technologist (MLT) profession. For my final year, I will be performing my clinical practicum at a hospital rotating between the five disciplines on the clinical lab: Microbiology, Biochemistry, Histopathology, Hematology, and Transfusion Science. I have already begun my first rotation in the Microbiology lab.

When first entering into the Microbiology lab, I noticed that it is separated into days of testing: Day 1, 2 and 3. Since bacterial growth takes at least 24 hours, there are usually several days of testing involved before a bacteria is identified and susceptibilities to antibiotics are reported.

During my first week in lab, I was scheduled to work on the Day 1 Bench. Alongside the lab technician, I received a variety of specimens into the lab, ranging from urines, stools, wound swabs, the list is endless. Once these specimens came into the lab, they were received through the Laboratory Information System (LIS), which is a software system that inputs lab specimens, test results and sends final reports to the doctor. While receiving, a barcode scanner scans the sample, and applicable information (such as sample collector identifier, date and time of collection) is inputted.

While most samples received in the laboratory are also tested there, other samples have to be sent out. One reason why specimens are sent out is that the hospital lab does not perform the testing for that microorganism. Another common reason is that there is an increased risk associated with working with certain microorganisms. Some examples of tests that are sent out are virus testing, parasitology, and chlamydia. These specimens are still received through the LIS at the hospital, then put in specific transport containers if applicable, and shipped out.

After specimens to be tested at the hospital are received, they need to be plated on appropriate media. For those who are not familiar with Microbiology, media refers to the agar plates or broth liquid that organisms are put on to help their growth. Many types of media exist and they are categorized as either enriched, differential, and/or selective. Enriched media contains nutrients to help bacteria thrive. Differential media uses colored indicators to differentiate between certain types of bacteria. Selective media have added substances that inhibit the growth of other bacteria, such as normal flora, in order for the pathogen to be isolated.

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Different media used in the Clinical Microbiology Laboratory

All of the specimens received are plated onto media under the Biological Safety Cabinet. This Biological Safety Cabinet is used for the protection against biohazardous materials. It has a High Efficiency Particulate Air (HEPA) filter to make sure that the air within the cabinet is being circulated. While plating specimens, I noticed that many of the blood plates required a ‘Staph Streak’ after they had been streaked for isolation. This Staph Streak is done using an American Type Culture Collection (ATCC) strain of Staphylococcus aureus bacteria. The S. aureus is put on the plate in the form of an X so that it touches all quadrants of the plate. This streak is done in order to grow the fastidious bacteria, Haemophilus influenzae. This bacteria requires both hemin and NAD to grow. The blood plate contains hemin, and the S. aureus strain excretes both NAD and hemin. Therefore, H. influenzae would be able to grow closely around the Staph Streak.

Staph Streak

Staph Streak

Another responsibility of the Day 1 Bench is reading clinical Gram stains. Clinical Gram stains are made directly from specimens. There are some specimens in which clinical Gram stains need to be performed and then read under the microscope. One example of this kind of specimen is sputum. Sputum is the mucus and saliva that accumulates as a result of an infectious process in the lower respiratory system. Since these specimens are not usually collected properly, the technologist checks under the microscope for its suitability. If there are many skin cells present, and no or few white blood cells present, the sputum has not been collected properly and must be rejected. However, if there are many white blood cells and few to no epithelial cells, the specimen can be cultured.

Sputum Gram stain with many epithelial cells and few white blood cells

Unacceptable sputum Gram stain with many skin cells and few white blood cells present

Acceptable sputum Gram stain with many white blood cells and no skin cells

Acceptable sputum Gram stain with many white blood cells and no skin cells present

Another common specimen where a clinical Gram stain is made is with vaginal swabs, for the diagnosis of Bacterial Vaginosis (BV). This is done by observing the levels of three different kinds of bacteria: Lactobacilli, Gardnerella, and Mobiluncus. Lactobacilli are Gram positive bacilli that are normal flora in the vagina. Whenever a woman is diagnosed with BV, the levels of these bacteria are low or absent. This decrease in Lactobacilli results in an increase of pH in the vagina, causing the overgrowth of pathogenic bacteria. A bacteria that is seen in high numbers in BV is Gardnerella, which is a small Gram variable bacteria. Finally, another bacteria present in large amounts is a curved Gram variable bacteria, named Mobiluncus. While the technologist is observing for these bacteria in the slide, they are also observing for the presence of any yeast that may be causing the BV.

Bacterial vaginosis Gram stain

Bacterial vaginosis Gram stain

Another stain that I performed in this week was the Auramine Rhodamine stain for acid fast bacilli (AFB), such as tuberculosis (TB). This directly stains the mycolic acid on the AFB. The first step is to stain with Auramine O for 15 minutes. Afterwards, the slide is decolorized with acid to get rid of any excess stain. Finally, the potassium permanganate, also known as a quencher, is added to remove any background staining. Positive and negative controls are also stained along with patient specimens. Once done staining, the slide is observed under a fluorescent microscope in a dark room. The positive and negative controls are read first to see if the stain worked properly. If it was done properly, the positive control would have AFB present, and the negative control would not. All slides stained for AFB must be examined under the microscope very carefully for the presence of any AFB. Even the presence of a single AFB is a positive diagnosis of TB. Any AFB slides must be reviewed by another technologist before the result is reported.

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Auramine rhodamine stain

At the end of the week, I performed my first venipuncture in six months. Since I have a requirement before I am done placement to perform fifty successful draws, it was suggested that I draw four tubes of blood from one person. I was very nervous drawing the blood, however I performed every step correctly, with the technician watching me. While I was drawing the fourth tube of blood, the person whom I was taking blood from said that they didn’t feel well, and then fainted. While everyone was rushing to help them I was distraught and thought that their fainting was my fault. The main lesson that I learned from this experience is that fainting is never the fault of the healthcare professional drawing the blood. Many people have different reasons as to why they faint, and you should never feel guilty to have caused a patient pain or stress. The outcomes to having blood drawn correctly (by being analyzed properly, and getting the correct diagnosis) far outweigh the brief discomfort or pain felt by the patient. Overall, if you ever have a negative experience with performing a task, you should always be willing to try again. As the saying goes, practice makes perfect! Even though this particular venipuncture might have been a negative experience for me, I will definitely try again soon to draw blood.

Overall, this week has been a real eye opener with how the Clinical Microbiology lab functions. While I face new challenges every day, I am slowly getting used to the new equipment and procedures of the laboratory. Everyone that I have encountered has been a great help in teaching me the fundamental concepts of the lab.