Goodbye for Now

Well, this week marks the end of our summer semester, and also the end of the assignment which brought about this blog.  I have enjoyed writing these entries more than I had originally expected and hope that those of you who have read them or followed me have enjoyed them and found them to be informative.  The main objective for this series of entries was to deliver information to the public in a manner that people, both familiar with the topics as well as  the average person who may not be, would have the opportunity to read and become informed with some of the various topics, activities, and procedures that go on in the medical laboratory setting.  This will probably be the last article posted for a while; I have not ruled out posting every now and then.  Until the next time, keep learning.

Alex Stroup

 

 

 

Ebola: Background and General Information

Certainly by now, many of you are probably aware of the ongoing Ebola virus outbreak occurring in western Africa.  Due to the severity and highly infectious nature of this particular strain of the virus in combination with other factors including the culture of the native peoples and poor economics, the diagnosis, treatment, and overall control of this current outbreak are proving to be extremely difficult.  I thought for this post I would share some information about Ebola including what type of methods are utilized in the diagnosis and treatment of this disease.

Ebola is a virus that, along with other closely related pathogens, causes viral hemorrhagic fever; which can be described as uncontrolled bleeding that when combined with vomiting and diarrhea, leads to extreme dehydration and anemia.¹  Symptoms of this disease range from less obvious, nonspecific signs including nausea, fever, and rash to more severe manifestations such as bleeding both internally and externally.¹  Ultimately, the extreme dehydration and low blood pressure brought on by these more severe symptoms are what typically result in the high mortality rate of this disease.¹

Diagnosis of the Ebola virus can be problematic, which aids in the spread of the disease.  Usually, other pathogens that cause similar symptoms are suspected which impedes and compromises the proper isolation and treatment of infected individuals, and also allows for others to become infected due to exposure to body fluids containing the pathogen, thus spreading the disease exponentially.¹  Typically, only in situations where the virus is suspected are the proper precautions and confirmation tests conducted in an adequate amount of time.¹  These include tests which detect antibodies developed by the patient in response to the virus, tests which look for antigens associated with the virus, molecular testing which identifies known nucleic acid sequences that are present in the pathogen, and viral isolation.¹

Treatment of the Ebola virus is currently limited to treating the side effects of the virus through fluids and blood transfusions in order to restore the patient’s electrolyte levels and blood pressure until the virus has ran its course.¹

Hopefully in the near future a more efficient treatment method or vaccine may be developed to aid in these ongoing outbreaks, which seem to pop up every few years and devastate many people.

For further information about this disease as well as up to date news about the current outbreak, check out the CDC website.  I have provided a link below in the works cited section for those of you who are interested.

Works Cited

1.       "Ebola Hemorrhagic Fever." Centers for Disease Control and Prevention. CDC, July 29, 2014. Web. August 1, 2014.

Main Article Link: http://www.cdc.gov/vhf/ebola/index.html

 

 

 

 

"Danger! Watch yourself!" Lab Safety Practices

With all the news concerning poor laboratory practices, such as the CDC anthrax incident and finding cultures of various diseases in storage closets, emphasis on proper safety in the laboratory environment is essential.

In any medical laboratory environment, there are inherent dangers which can be increased unnecessarily by poor safety protocols and practices.  As with any job or daily routine, a level of familiarity can lower one’s guard and open the door for potentially serious outcomes.

Here is a list, that may seem common sense and second nature to a laboratorian, which if followed could prevent many of these opportunities for danger.

1.      Treat every sample as a potentially dangerous pathogen.

Any sample that comes through a medical laboratory could contain any virus, bacteria, fungus, or parasite and should be processed with this kept in mind.

 

2.      Sterilize all equipment and properly discard contaminated materials

This serves the purpose of preventing cross contamination between samples or accidental exposure and keeps the work area uncluttered.

 

3.      Disinfect work spaces before and after use

This follows along with the previous rule, keeping the work space clean.

 

4.      WASH YOUR HANDS!

This is probably the easiest yet most important rule on this list.  The significance of proper hand washing technique and frequency is fundamental to lab safety.

 

5.      Never mouth pipette

This sounds very strange, but only 20 to 30 years ago, this was a common practice.  Now it has been phased out for the most part, but still worthy of mention for obvious reasons.

 

6.      No food or drinks in the laboratory

This is another rule that could become abused in the everyday workplace due to the daily routine.

 

7.      Clearly label everything

This practice could prevent accidental exposures do to treatment of a dangerous substance with insufficient safety protocols, and keeps everyone informed and on the same level as far as samples and reagents being used are concerned.

 

8.      Autoclave any contaminated materials

As with suggestions 2 and 3, the proper discarding of used materials which have been exposed to hazardous substances or dangerous pathogens is crucial in keeping what is in the lab from harming others.

 

9.      Follow proper procedures for spills and/or exposures

Any laboratory has strict protocols and procedures already set in place for any potential spills or exposures which could occur within the laboratory environment.  These preplanned actions are meant to minimize the risk of these hazardous events if they do occur.

 

Although a certain level of familiarity, comfort, and routine is expected in any workplace environment, it is the responsibility of everyone involved to be both vigilant and actively participate in the prevention and implementation of safety in the laboratory environment.

 

(The main bullet points where derived from a list from an article by Daniel E. James, “Nine Safe Practices for the Microbiology Laboratory”, located on the Carolina® Science and Math Support Website obtained 7/25/2014)

http://www.carolina.com/teacher-resources/Interactive/nine-safe-practices-for-the-microbiology-lab/tr11085.tr

 

"It Came From Africa"-A Case Study

As our semester comes to a close, we have been covering parasites of all shapes, sizes, and types.  I have really enjoyed reading the case studies on the CDC website, like the one I shared last week, and thought that I would continue by sharing another which involves someone who contracted a parasite during a trip to Africa.

As with any case study, the narrative and patient history are the most important parts, so that is where this begins.

Case #362 - December, 2013

“A 29-year-old female post-graduate student in Zoology went on an expedition to see the lowland gorillas in the Democratic Republic of Congo. She reported numerous insect bites while traveling but did take anti-malarial prophylaxis. Approximately one week after returning home she developed fever. About a month later, she started experiencing headaches, itchy skin, and swollen lymph nodes and sought medical attention. A blood specimen was collected; smears made and stained with Wright-Giemsa.”¹

 

 

 

"Case #362 – December, 2013." DPDx - Laboratory Identification of Parasitic Diseases of Public Health Concern (Figure D)

 

Based off the findings of the peripheral blood smear and the organisms which were observed as well as the case history, the diagnosis was Trypanosoma brucei; most probably the Trypanosoma brucei gambiense subspecies due to the geographic location of where the individual traveled.¹

These organisms are transferred to humans by the bite of the Tsetse fly, which allows an immature stage of the organism’s life cycle to be introduced to the human host.  Symptoms can range from headaches, like the individual in the case study, to coma/death once more central nervous system involvement occurs.

Treatment of this disease depends on what disease stage that the organism has been able to progress; such as whether it has crossed the blood brain barrier or is just in the blood stream.²  The drugs used to treat this type of infection are typically only available from the Centers for Disease Control.²  After treatment, an infected individual will need to have repetitive examinations of their cerebral spinal fluid in order to rule out a relapse of the disease.²

 

Works Cited

1.       "Case #362 – December, 2013." DPDx - Laboratory Identification of Parasitic Diseases of Public Health Concern. CDC, November 29, 2013. Web. 18 July 2014.

http://www.cdc.gov/dpdx/monthlyCaseStudies/2013/case362.html

 

2.       "Parasites - African Trypanosomiasis (also known as Sleeping Sickness)." Centers for Disease Control and Prevention. CDC, August 29, 2013. Web. 18 July 2014.

http://www.cdc.gov/parasites/sleepingsickness/treatment.html

 

 

Parasites, Laboratory Practices…. and a Case Study!

 Over the past week or so we have been covering material on protozoan parasites which included the standard procedures used in diagnosis as well as some specialized techniques.  As stated in my post from last week, these life forms can present difficulty in the processing, detecting, and diagnosis of disease compared to other organisms such as bacteria.

The standard techniques for the detection of intestinal parasites include a direct wet prep, a concentrated method, and a permanent stain.  The wet prep allows a technologist to look for motility of living, mobile forms of protozoan parasites known as trophozoites, while the concentrated prep increases the likelihood of observing a parasite, thus increasing the overall sensitivity. The permanent stain, usually a trichrome stain, aids in the ability of viewing cysts, eggs, etc by adding color to morphology which would normally not be easily observed. 

Some organisms may need special staining in order to be detected.  These include Cryptosporidium, Cyclospora, and Isospora which require an Acid-Fast staining technique to be utilized for their oocysts to be observed.

Here is a case study I came across on the CDC website which gives an example of laboratory methods in the detection of protozoan parasites.

“A 34-year-old missionary worker sought medical attention for abdominal pain, nausea, and watery diarrhea after returning from visiting friends in Central America. A stool specimen was collected for laboratory testing and a modified Kinyoun’s acid-fast stained smear was prepared.  The objects of interest measured 8-9 micrometers in diameter on average.”¹

 



Centers for Disease Control and Prevention-

DPDx Case #374-June,2014 (Figure A)¹

 

Based off the information provided in the case study along with the picture, it can be concluded that the causative pathogen was Cyclospora cayetanensis.  One of the clues which could aid in determining this is the fact that an acid-fast technique had to be used to view the oocyst.  Also, from our lectures I have learned that this organism causes traveler’s diarrhea and is endemic to Central America.

Here is the link to the case study I referenced in this post if you would like to see additional pictures.  There are also many other informative case studies available on their website which you might enjoy. 

http://www.cdc.gov/dpdx/monthlyCaseStudies/2014/case374.html

Works Cited

1. "Case #374 - June, 2014." DPDx - Laboratory Identification of Parasitic Diseases of Public Health Concern. CDC, June 2014. Web. 10 July 2014.

Cryptosporidium and Giardia Source Water Monitoring

This week in lecture we began covering material on protozoan parasites, which can cause a variety of diseases and can be harder to detect in the laboratory than other organisms such as bacteria.  In an attempt to take a slightly different angle on the subject, I thought that going over how some species can be monitored/detected in the environment would be beneficial. 

For the past seven years prior to entering the program, I worked as a biologist for an environmental testing laboratory, which participated in the EPA source water monitoring study of systems for Giardia and Cryptosporidium. During my time there I analyzed over 3,000 samples, so you could say that I somewhat familiar with the process.

The method is long, labor intensive, time consuming and could take up to a week to complete.  In general, a large sample of water is concentrated to just a few drops on a slide which is then stained and examined microscopically.

First, a roughly 10 liter sample of source water (which is the lake, river, etc that a water processing plant pulls from to make drinking water) is collected and filtered through a specialized filter that has pores small enough to catch the desired organisms.  Next, a soapy solution is added to the filter, which is then shaken to remove and resuspend any collected material.  This solution is then centrifuged to concentrate the dirt and any organisms into a pack pellet.  Then the supernatant is removed, the pellet is resuspended, and transferred to a small tube.  Next, a solution containing small iron beads coated in anti-Crypto and anti-Giardia antibodies as well as pH buffers are added to the tube with the sample.  The beads will grab onto the organisms, and in the presence of a magnet, allow the potential organisms to be separated from the remainder of the contents of the sample.   This process can then be reversed by the addition of acid, which elutes the organisms from the beads into the eluate and onto a microscope slide.  This can then be stained with fluorescent dyes and examined for the presence and enumeration of the cysts and oocysts which may be present.

For the EPA study, water systems were required to send these samples in once a month for two years.  After the required monitoring was completed, the data could be compiled to calculate the number of cysts/oocysts per liter which could be correlated to a risk for these organisms to contaminate the drinking water produced by the individual water system. 

Hopefully this was not too boring, but I thought that it would be useful for others to know how these organisms are detected from environmental samples in addition to their detection in human samples.  

Transfusion Reactions

This week in lab we combined elements of two of the classes we are taking this semester: Blood Bank and Infectious Disease.  I felt that the exercise also brought together several things we have learned since beginning the program.  We processed simulated samples from patients that were experiencing a transfusion reaction.  These reactions can occur for several reasons including factors with the recipient’s immune system and contamination of the blood product itself. 

The process of determining what could be responsible for a transfusion reaction began with reviewing the patient’s vital signs and symptoms, looking for clerical errors, processing several laboratory tests on the individual’s blood and urine, and finally setting up microbiological testing  from the donor material to see if a microbial organism was growing in the product itself.



The blood product that was being transfused in our case was a unit of platelets, which have the highest rate of contamination since they are incubated at room temperature up to five days.

The first step to analyzing the microbiological aspect of the laboratory was utilizing a Gram Stain to get an idea whether any organisms were present and a preliminary identification of those that were observed. The appropriate media for growth of the specimen included Sheep Blood Agar, MacConkey, Chocolate, and a time saving special plate that differentiates organisms by causing colonies of various species to appear specific colors thus saving time in the determination of the causative microbe.

When observing the Gram Stain of my specimen I noticed many Gram positive organisms with elongated protrusions growing out of their sides, which are known as germ tubes.  This led me to believe that the specimen had become contaminated by a Candida species yeast organism.  After 24 hours, I was able to observe the different media plates that were set up and determine that it was the Candida albicans species, based off its color on the specialty media that differentiated organisms of the Candida genus by the color of the colony growth.

As I stated before, I thought this laboratory exercise brought together several aspects of the program we have learned about so far, and was a good simulation of a real world circumstance.

Acanthamoeba, and you can too!

Although we have not covered material on parasites yet in our class, I wanted to talk a little this week about the significance of Acanthamoeba species, a protozoan genus that can commonly cause several different complications affecting different parts of the body.

Acanthamoeba species can be found in the soil and freshwater environments in many places around the world, and typically feed on bacteria which also reside in these environments.  However, some opportunistic species can become a health risk to people.  Depending on the site or method of entry into the body, this can result in keratitis and encephalitis/meningitis.  Amoeboid Keratitis occurs when the amoeba comes into contact with the eye, and can lead to blindness in severe cases; in many cases this results from improper use of tap water to clean contact lenses.  Acanthamoeba granulomatous encephalitis occurs when the amoeba enters the nasal mucus membrane and migrates to the brain to feed.  Although very rare, this type of infection usually results in death.¹

Since these organisms can encyst, they are extremely difficult for the host’s immune system to rid itself, and many drugs are ineffective as well.

Acanthamoeba species are also carriers for many pathogenically significant bacteria and virus species, and therefore may operate as a vector for infection for these pathogens which include Legionella species, Staphylococcus aureus, and Campylobacter species.¹ 

So remember, never use tap water to clean your contacts or flush out your sinuses!

Reference

1.      Wikipedia contributors. "Acanthamoeba." Wikipedia, The Free Encyclopedia. Wikipedia, The Free Encyclopedia, 11 Jun. 2014. Web. 20 Jun. 2014.

Monitoring Sepsis through the Detection of latent Viruses

The topic of my entry this week will combine the subject matter of two previous posts: septic infections and viruses.

 It may sound strange, but some study has been done that suggests that it may be possible to monitor progression/severity of septic infections through the detection of previously latent viruses, indicating a clinically significant depressed immune system.  In order to accomplish this, several serially collected blood and/or plasma samples were obtained from hospitalized patients over a period of time and monitored for an increase in viral DNA from species that are capable of becoming latent, or hiding within the host’s DNA.  Examples of these which were used in the study included cytomegalovirus (CMV), Epstein-Barr (EBV), and herpes-simplex (HSV).¹

In theory, when the immune system is suppressed to a certain extent, these latent viruses would be reactivated and enter the patient’s blood.  For the purpose of this study, an increase in the viral DNA present in the patient’s blood was cross correlated to secondary infections such as bacteria and fungi to see if a relationship could be established, which could be used to indicate the underlying status of the corresponding person’s immune system.¹

It was determined that this method of serially monitoring the amount of virus present in a septic patient’s blood could be useful in determining the severity of the depression of the immune system.  The authors suggest that this could be best accomplished through monitoring a panel of common viruses such as CMV, EBV, HSV as well as others, and observed for a marked increase across the board which could indicate that an individual has entered the immunosuppressive stage of sepsis meaning that they would be almost incapable of fighting off infections from various opportunistic pathogens such as bacteria and fungi.¹

If you would like to read the full article and see the figures, here is the link:

http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0098819

Reference:

1.      Walton AH, Muenzer JT, Rasche D, Boomer JS, Sato B, et al. (2014) Reactivation of Multiple Viruses in Patients with Sepsis. PLoS ONE 9(6): e98819. doi:10.1371/journal.pone.0098819

"So what is a virus?"

Over the past week we have been learning about viruses in our lecture portion of the course.  We discussed their overall morphology and biochemical components, as well as laboratory diagnosis methods for a wide variety of clinically significant viruses.  With this week’s post, I wanted to briefly cover some of this information.

So you may ask:

“So what is a virus?”

“How do they infect us?”

“What kind of treatment can I receive for an infection?”

These are just some of the commonly asked questions about viruses.  Primarily, they are composed of a little protein and one type of genetic material; unlike other organisms which have both RNA and DNA.  This genetic material can be either single strand DNA, double stranded DNA, single strand RNA, or double stranded RNA and is one method of categorization.  Viruses, believe it or not, are considered by some scientist to not even really be alive, since they are incapable of reproducing/replicating without high jacking the cellular components of a host. 

Since there is such a wide array of virus types, there likewise are widely variable methods of infection that are utilized by these pathogens.  This can range from respiratory, skin to skin, body fluids, and even in some cases vectors such as mosquitoes.  Once inside the body, viruses look for specific chemical markers on cells that they have specialized to utilize for their replication.

Especially with viruses, the old proverb “An ounce of prevention is better than a pound of cure” is applicable.  The most effective method to prevent the majority of clinically significant viruses is vaccination, because some treatment methods are not effective against them.  Antibiotics work only on cellular pathogens, and are therefore useless.  For some viruses, there are anti-viral medications that can be helpful, but these are usually only developed for more severe diseases such as HIV, Hepatitis C, and influenza; not for every virus.  In many cases, symptoms are treated, and the individual’s immune system will eventually rid itself of the virus.

Diagnosis can be made through several ways, which are best classified as either direct or indirect.  Direct methods look for the virus itself by detecting proteins or genetic material, while indirect look for antibodies developed by the immune system which are specific to antigens of the virus in question.

 

Legionnaire’s Disease

As many of you may have heard, recently there have been several cases of Legionnaire’s Disease diagnosed at UAB Hospital in Downtown Birmingham, Alabama.  I thought that for this week’s topic, I would give some background information about this condition and the causative pathogen for this disease.

Legionnaire’s Disease is a type of pneumonia brought about by an infection of a bacterium from the Legionella genus.  These organisms can be found in virtually any source of water, and are likewise transferred through inhalation of water droplets that contain the pathogen, not person to person.  The disease has an incubation time of two to ten days and is characterized by symptoms that are similar to other types of bacterial pneumonia which may include fever, headache, and nonproductive cough.  The individuals at the highest risk of infection for this disease include the elderly, smokers, and those who are immunocompromised, while those with a healthy immune system either fight off the bacteria and never get Legionnaire’s Disease or only get a milder case.¹

When you combine the facts, it is clear how an outbreak can easily happen in a hospital environment.  Many of the individuals have a weakened immune system, and if exposed to the Legionella bacterium in the water supply, they can easily contract this disease. 

Diagnosis of this disease is accomplished through a variety of means such as special fluorescent stains, DNA detection, and even a urine antigen test.  Since this organism is difficult to grow and isolate on standard media, special media must be utilized that can aid in its culture.¹

Hopefully this information helped in the understanding of this disease.

References:

1.      Tillie, Patricia M (2014). Legionella. Bailey and Scott’s Diagnostic Microbiology 13^th Edition (pp. 424-430). St. Louis, Missouri: Elsevier Mosby.

Bacteremia and Blood Culture Part 2

The presence an infection in the blood is detected through the use of blood cultures.  For this procedure, blood is collected into sterile bottles containing a media which when metabolized by any present organism will result in the release of carbon dioxide.  This alters the pH of the substrate on the bottom of the container, causing a change in color which is generally measured by a sensor in an automated incubator, usually monitored up to a five day span.  Once this color change is detected, the medical technologist will receive an alarm indicating which bottle has tested positive for an increase of carbon dioxide so that further culture and identification of the pathogen can occur.

In order to achieve this, blood from the culture bottle is inoculated onto media for isolation/identification of the organism as well as Gram stained so that a preliminary identification can occur and general treatment can begin.  After 24 hours the resulting growth on the media can be observed and processed using rapid testing such as oxidase or catalase to obtain a final identification.  If a final identification cannot be determined at this step, identification systems can be utilized to identify the pathogen.  Upon final identification, antibiotic susceptibility testing can be initiated in order to provide a more precise antibiotic treatment that can be used to rid the patient of the infection.

This is a brief description of the lecture information and laboratory exercise we performed during the first week of our course.  I processed a blood culture bottle by using this method of isolation culture, rapid testing, and identification systems, and was able to determine that the patient in question had bacteremia resulting from Escherichia coli.  I found this exercise to be quite informative, and hopefully my description can aid in the understanding of how this process works in a medical laboratory setting.

Bacteremia and Blood Culture Part 1

For my first official entries, I thought it would be good to review over some of the information and activities we have covered so far in the course during the first couple of weeks, particularly bacteremia and blood cultures.  This will be a two part post, with the second half being posted next week.

Under normal circumstances and in healthy individuals, the blood system within your body should not contain organisms of any kind, and is considered to be a sterile body fluid.  Inevitably, due to incidences such as cuts, IV catheters, and infections of other body sites, pathogens such as bacteria or viruses can gain access into the blood stream.  The immune system is able to normally rid itself of these invasions in most healthy adults.  Unfortunately for individuals with a weak or compromised immune system, this can lead to a more serious complication.  Since the blood system is basically a highway throughout the body, if undiagnosed or untreated this can lead to systemic infections that can affect various organs and may lead to death. 

The terminology used to describe these conditions is based on the responsible organism and the severity of the infection.  Some of these include bacteremia, viremia, and fungemia which indicate the presence of bacteria, viruses, and fungi in the blood, respectively.  Septicemia is another term that is used to indicate that pathogens are present and reproducing within the blood stream. 

Welcome

Welcome to Infection of Knowledge!  This blog was created for the purpose of giving an inside view to the world of the Microbiology/Infectious Disease scientific discipline from the perspective of a graduate student throughout the course of an entire semester.  The subject matter posted here will cover various issues ranging from current events related to the field to interesting topics or information discussed in some of the coursework and laboratory exercises that I am currently undertaking. Hopefully, this information can help others to better understand the biological medical laboratory, and shed light on some of the lesser known facts related to this area of expertise.