Wednesday, July 11, 2012

Psychrophiles in Commercially Produced Sauces, Glazes, and Marinades

 These are plates from the 2nd dilution of 13 different sauces on week 4 of our study.  We plated on July 3rd and did the 2nd dilution so that we would not have a potential of too many to count after being incubated for 48 hrs at 37C.  The plates were pulled out on July 5th and placed into the refrigerator at 4C.  I did not view the plates until July 9th and observed yellow, bacterial colonies growing.  These photos were taken on July 10th.

The colonies are small, circular, opaque yellow, entire, raised to convex, and glistening.  Perhaps tomorrow I can stop by the lab to streak for singles and the next day do a Gram stain.  From there we will decide what metabolic media to use in identifying this organism.

What is interesting about this growth is that it did not occur the first two weeks of tests.  Those plates had 1 or 2 yeast colonies growing on a few of the sauces.  For the most part the plates have been clean.  After seeing this growth, we looked at the plates from week 1 and week 2.  Week one showed a single bacterial colony growing on one streak for 2 different sauces.  Week 2 showed these yellow colonies on ALL of our plates.  It appears this organism is a psychrophile.

In preparing for this long-term shelf-life study, we aliquoted 25 ml of 13 different sauces into 50 ml conical tubes.  One set remains in the refrigerator for future testing if we see a need to do so.  The other sets were stored at room temp at 30C and 2 sets were placed in the incubator at 37C for week 1 and week 2.  So for week 1 and week 2, we ran a set of tests of the sauces stored at 30C and at 37C.  Based on previous results of these same sauces in a similar study, we did not plan for additional weeks at 37C.  Last week we completed week 4, which was only for the sauces stored at 30C.


Psychrophiles – Properties and Characteristics of Psychrophilic Bacteria
The best earliest review on psychrophiles is that of Ingraham and Stokes who describe these features of psychrophiles:
  • are typically Gram-negative bacteria and include members of the genera Pseudomonas, Flavobacterium, Psychrobacter, Colwellia and others.
  • often produce green, yellow, orange and purple-violet pigments at reduced temperatures
  • can tolerate elevated levels of salt
  • can grow well and sometimes optimally at temperatures up to about 20 degrees Centigrade
  • can grow at temperatures as low as -3 to -7 degrees Centigrade
  • produce extracellular enzymes that can degrade meats, milk, cheeses and other food products during cold conditions
I found a genus of bacteria that may fit but only testing will determine.  Chryseobacterium spp. (Flavobacterium) This genus is a Gram-negative, rod bacterium with yellow pigment.  It is non-motile.  C. indologenes tests positive for oxidase, negative for catalase, cannot ferment Mannitol or Urease, does not grow on MacConkey.  It weakly ferments glucose as seen on TSI slants.  They can grow at 4C but optimum growth is at 25C.  They are also heat stable and have a salt tolerance.  They are commonly found in soil, plants, foodstuffs, and water sources.

Like I said, we have to streak for singles and do a Gram stain first to determine next steps in identifying this organism. 


Tuesday, July 10, 2012

Multivariate Study on Antimicrobial Properties of Raw Honey, Stevia, and HFCS

Today we begin our next experiment to test the antimicrobial properties of raw honey, stevia, and high fructose corn syrup.  We will begin with extracting different groups of phytochemicals from the stevia plant and the stevia flower.  For the raw honey and the HFCS, we will make solutions of different concentrations.  We will be using disk diffusion to look for zone of inhibition against E. coli and S. aureus.

A couple of weeks ago we tested xylitol and HFCS.  We used the following concentrations:  0, 5, 20, and 25%.  There was no inhibition on the disk diffusion against E. coli and S. aureus.  But we did observe that the HFCS encouraged growth.

The other test we did with these two sweeteners was to see if E. coli and S. aureus could metabolize xylitol or HFCS.  We aliquoted 5 mL of PBS into 6 test tubes.  For 2 we added nothing else.  For another 2, we added 1.25 grams of xylitol and for the other 2, we added 1.25 mL of HFCS.  We inoculated 3 tubes with 100 microliters of E. coli and the other 3 with 100 microliters of S. aureus.  The results were interesting.  After incubating at 37 C for 48 hours, we observed growth of both organisms in the HFCS.  The xylitol also showed growth but not as much.  I plated 10 microliters onto a nutrient agar plate of each tube.  I did not do any dilutions.  After 24 hrs at 37 C, there was too many to count growth on the plate. Plus, either by my error or from contaminated PBS, there was also growth of yeast present on the plates.

Two weeks later, the PBS shows no growth for either organism.  The xylitol shows small growth and the HFCS shows moderate growth. We plan to plate for growth but this time will probably take it to the 2nd dilution.

I read some interesting peer-reviewed research papers on the antimicrobial properties of stevia so I am excited about testing this.  I will also be learning how to do protein gel electrophoresis and will be doing chromatography.

Sunday, July 8, 2012

ASLT Continues

I am in my second semester of research in food microbiology.  The Spring semester of 2012 was dedicated to accelerated shelf-life testing of numerous sauces for a local company that manufactures these sauces for big-named customers.  Our first assay showed mixed results.  Our second assay of 5 sauces that were fresh off the line and sealed performed much better.  The attached photo shows growth of yeast in small numbers.  We did not have any bacterial growth.  All 5 sauces held up at the 37 C temp with no changes to color or odor.

Since we did not repeat results from the first assay to the second, we discovered that the sauces given to us in the first one were significantly old and were not hermetically sealed.  Additionally, I cultured one of the plates with bacterial growth and did a Gram stain.  We found Gram + and a Gram - organisms with a diplococci morphology.  We did some tests to try and identify the bacteria.  Motility, TSI, citrate and some others I am not recalling at the moment.  The results did not lead to any common bacteria that we, as budding microbiologists, were familiar with.  Dr. Bates, our supervising instructor, concluded it was environmental as in dirt.  We did take a pic of the Gram stain and I need to ask for a copy so I can post.

Right now we are repeating the first assay with the same 19 sauces.  This time the sauces are fresh off the line and hermetically sealed.  They are performing just as well as the sauces did in our second assay.  No bacteria and low yeast growth with some no growth at all.  No significant odor changes and a few of the sauces showing color change.  This is the start of a long-term study requested by the company.  Although we gave them our ASLT results, they tell us their customer wants real-time data so this study is set up to take place over the course of 12 months.

The next post will be about the scientific experiments we designed that have nothing to do with the shelf-life studies.  My research partner and I created a spreadsheet of all the ingredients in all of the sauces so we could analyze the results to see which ingredient we want to isolate and experiment with.  Right now we are getting ready to test the antimicrobial properties of sweeteners.  Raw honey, xylitol, high fructose corn syrup, and stevia.  More to come.

Monday, April 23, 2012

Questions for Next Experimental Design

In thinking of additional tests for our long-term sauce study, I think we need to consider what we've learned from Micro and apply that.  I'm thinking in terms of chemistry - functional groups, decomposition reactions/catabolism, and organic compounds - carbs, lipids, and proteins.  And also cell structure, enzymes & metabolism, and microbial growth.  If we have an understanding of how and why some of the sauces have a change in color, odor, and viscosity, then I think that would lead us to our next experimental design.  On Saturday, as I was driving to Little Rock, I had a lot of time (3 1/2 hours) to think about our research and I came up with a series of questions. 

  • What is the chemistry behind the change in color & odor that we’ve observed?  Are we seeing a significant change in viscosity or texture?  How come we are not seeing a significant change in pH?
  • What are the organic compounds in the sauces?  What type of chemical breakdown is happening?  How do we measure this?
  • Regarding proteins – what is the source of protein in the sauces?  Could it be cell components from ingredients like wheat or soy?  What enzymes are at work breaking proteins down?  Proteases or peptidases?  Are the proteins being denatured or deaminated?  As for enzymes, they are influenced by temp, pH, and substrate concentration so how does this tie into the data we’ve collected so far?
  • Catabolism requires energy so where is that energy coming from?  Our current sauces show that yeast is present and Dr. Bates said it is probably Saccharomyces.  Yeast ferments producing 2 ATP in glycolysis and then ethanol and CO2 as end products.   Could this be what I smelled in some of our previous sauces? 
  • What are the requirements for microbial growth?  Is aerobic or anaerobic respiration occurring?  What is being metabolized – carb, lipid or protein? 
  • Consider the ETC where O2 is the final electron acceptor and H2O is formed.  Does this water then become available for microbial growth or other chemical reactions?  Is this the water activity?

Thursday, April 5, 2012

First Assay in Short-term Study for Food Microbiology

We completed our first assay for the short-term study we are doing for a company who makes sauces.  Their customers asked them how long they can store the large totes of various sauces at room temperature in their warehouses before it spoils.  Our task, as undergrad research students, was to set up an experiment for accelerated shelf-life to determine which sauces may be stored at ambient temps.

This is the first time any of us conducted scientific research.  Although we tested samples of sauce over a two week period, it took us longer to complete the study as we had to learn new lab techniques.  We collected data for pH, viscosity, odor & color change, and CFU's for 18 different sauces at three different temperatures:  4, 30, 37.

We are now in the process of writing our first scientific paper, which we will be presenting at a conference in the Fall.  Today we started a second assay on 5 new sauces.  This time we will be collecting data over a four week period.  Eventually, we will be setting up a long-term shelf-life study.

I'll be posting our paper when it is finished.  But we've got a ways to go in turning our basic rough draft into an actual scientific paper.  It's a learning curve.

Monday, March 26, 2012

UTI's and Uropathogenic E coli - A Term Paper

The urinary system is comprised of two kidneys, two ureters, the bladder and urethra.  It has three primary functions:  (1) the removal of organic waste materials from body fluids through excretion, (2) the discharge of these waste materials through elimination; and (3) the homeostatic regulation of blood volume, blood pressure, and the concentration of solutes in plasma.  Other homeostatic mechanisms work to stabilize blood pH, conserve nutrients through reabsorption, and assist the liver in detoxification.  

A urinary tract infection (UTI) is caused when a pathogen invades the urothelial cells anywhere along the urinary tract.  The location of where the infection takes place has different names, such as cystitis for a bladder infection or pyelonephritis for a kidney infection.  This paper will focus on uncomplicated acute cystitis.  

Urinary tract infections are the second highest occurring infection in humans, following respiratory tract infections.  UTIs result in as many as seven million office visits and more than one million emergency room visits each year.  UTIs are more common in women than in men because of differences in anatomy, and changes in sexual maturation, pregnancy and childbirth.  50% of all women will experience an UTI within her lifetime, while 25% of those women will experience recurrence.  Others at risk are those with catheters, obstructions such as kidney stones, or those with diabetes mellitus.  For uncomplicated acute cystitis, 90% of those infections are caused by uropathogenic Escherichia coli (UPEC).

There are several symptoms that occur when the bladder is infected.  These include a strong, frequent urge to urinate even though the bladder is empty, pain or burning when urinating, pressure or cramping in the lower abdomen, and the presence of cloudy or bloody urine that may have a strong odor.  If left untreated, the infection can spread to the kidneys resulting in more severe symptoms including chronic renal failure.  Speaking from personal experience, UTIs are aggravating and they do hurt.  The frequent, strong urges to pee can really disrupt one’s day making it nearly impossible to get any work done.


The body has several defenses in place to combat invasion by pathogens in order to maintain homeostasis.  With the UTI, such defenses include the composition of urine, the micturition reflex, and the innate and adaptive immune responses.

In The Journal of Clinical Investigation, Donald Kaye, in his article Antibacterial Activity of Human Urine, wrote, “In discussing the kinetics of urinary tract infection O’Grady and Cattell emphasized as a host defense mechanism the importance of the mechanical effect of urine flow in diluting and removing bacteria from the urinary tract . . . in the present study, urine from normal individual is often inhibitory and sometimes bactericidal for growth of these organisms.”  From Kaye’s research study, it is noted that the pH and the concentration of urea provide an inhibitory effect on bacteria.  The data showed that at a pH of 5.0, the strongest inhibitory effect was found.  Additionally, the greater the concentration of urea, the greater the inhibitory effect on bacterial growth.

According to the textbook used for Anatomy and Physiology, the normal range for urine pH is 4.5 – 8.0, osmolarity ranges from 855-1335 mOsm/L, and the concentration of urea is typically 1.8g/dL.  The research study referenced above showed in their data that antimicrobial properties of urine is optimal at 5.0 but still demonstrated inhibitory activity up to 6.0.  At a pH greater than 6.0, the inhibitory effect against bacteria declined.  It also demonstrated that as the osmolarity increased, so did the concentration of urea.  Kaye concluded, “The results of the present study provide evidence for the role of urea in human urine as an antibacterial agent.  They also suggest that within the ranges of concentration commonly achieved in human urine, antibacterial activity is more a function of urea content than of osmolality, organic acid concentration, or ammonium concentration.”

As water is reabsorbed in the nephron, the concentration of urea increases where membranes in the ascending limb of the nephron loop, the DCT, and the collecting ducts are impermeable to urea.  The volume of urine is dependent on the movement of water within the nephron.  85% of this water is involved in obligatory water reabsorption.  The other 15%, about 27 liters per day, is under the influence of ADH, which is secreted by the hypothalamus.  This hormone inserts water channels (aquaporins) along the membranes.  This affects the osmotic concentration and as ADH increases, more water channels appear and more water is reabsorbed, thus concentrating the urine with solutes because it is less diluted by water.  In the case of a UTI, based on the study mentioned above where the concentration of urea had the greatest inhibitory effect, the body could respond by increasing ADH, which decreases the volume of urine making it more concentrated with urea.  This could inhibit bacterial growth in the bladder restoring homeostasis.  But on the other hand, increasing urine volume to cause a person to pee more will provide more bacterial cleansing of the bladder and urethra.

Aside from the composition of urine, the flow of urine through the micturition reflex is another important defense against invading pathogens.  Urine is the end product after the blood has gone through filtration in the nephrons, the functional units of the kidneys.  After the bodily fluid is filtered in the proximal convoluted tubule, it then goes through reabsorption in the Loop of Henle in which water, ions, metabolites and nutrients are reabsorbed into the vasa recta.  Finally, in the distal convoluted tubule, the active secretion of ions, acids, drugs and toxins takes place.  The fluid that is left is what makes up urine as it now flows into the collecting ducts of the nephron, then to the papillary duct and on to the minor calyx.  The minor calyces join to form a major calyx and two or three major calyces form the renal pelvis, which is connected to the ureter.  Urine then flows down the ureters, one from each kidney, to fill the urinary bladder.

The micturition reflex begins when the stretch receptors in the bladder are stimulated as the bladder become full.  This sends an impulse along the afferent fibers in the pelvic nerve to the sacral spinal cord.  Parasympathetic motor neurons are stimulated and send the signal along the interneurons that relay sensation to the thalamus, then through the projection fibers and on to the cerebral cortex.  This is when one feels the urge to urinate and the conscious decision is made to go to the bathroom.  While sitting on the toilet, one voluntarily relaxes the external urethral sphincter, which then causes the internal urethral sphincter to relax as well.  At the same time, parasympathetic preganglionic motor fibers in the pelvic nerve delivers an impulse to the postganglionic neurons that stimulate the detrusor muscle to contract.  (The detrusor muscle is the name for the muscle fiber of the bladder.) This elevates the hydrostatic pressure in the bladder and with the sphincters relaxed, urination takes place.  It is this act of urination that works to flush out any pathogens that may be present; thus maintaining homeostasis. 

With a UTI, a patient can experience dysuria, which is pain as a result of inflammation.  Moreover, urinalysis from a patient with a UTI will often show a higher than normal count of white blood cells (pyuria) as well as a higher than normal count of red blood cells (hematuria).  All of this is evidence of the body’s defenses working to restore homeostasis upon bacterial infection.

Attached to this report is an actual urinalysis from a family member who recently recovered from her first acute bout of cystitis.  She is a health-conscience, physically fit, 39 year old female with a very busy schedule who is under a lot of stress.  She called me to ask about some symptoms she was experiencing.  She described pain in her lower abdomen and the constant urge to pee with hardly anything coming out.  I asked her if she noticed her urine appearing cloudy or red.  She said yes to both and added that there were blood clots in her urine as well.  I advised her to go to her doctor as soon as possible because she had a bladder infection and she needed a course of antibiotics to help her immune system fight off the bacteria.  The battle waging inside her bladder was between UPEC and her body’s innate and adaptive immune responses working to restore homeostasis.

Uropathogenic E. coli (UPEC) must face the innate immune defenses of the body upon invasion.  But one virulent strain has multiple ways to evade the host’s defenses.  The first step in the pathogenesis of UPEC is the portal of entry.  UPEC lives in the human gut’s flora and if microscopic amounts of feces come into contact with the urethra, the UPEC can then enter, traveling up the urethra to the bladder.  As mentioned earlier, UTIs affect more women than men.  Women can become infected if they wipe from back to front or as a result of sexual intercourse.

E. coli are a Gram-negative bacillus that grows well under various environmental conditions.  These rod-shaped bacteria have fimbriae, or pili, which function to bind to the receptor sites on the host cell membrane.  This report will focus on UPEC strain NU14.  The filamentous type 1 pili have a FimH adhesion which binds to mannose-containing glycoprotein receptors.  Research of Martinez et al, published in the European Molecular Biology Organization Journal states, “Interactions between FimH and receptors expressed on the luminal surface of the bladder epithelium appear to be critical to the ability of many uropathogenic E. coli (UPEC) strains to colonize the bladder and cause disease.”

As the photo illustrates, UPEC is able to penetrate into the cell through membrane ruffling and is able to evade host defenses by leaving the lumen of the bladder where flow of urine would flush them out of the body.  Once in the urothelial cell, the bacteria have a source for nutrients and the opportunity to proliferate.  

However, UPEC still has to overcome the obstacles of the body’s first line of defense, which are the pH and the concentration of urea present in urine as well as the flow of urine to flush out the bladder and urethra.  As will be discussed, UPEC RU14 has virulence factors that enable it to evade the host defenses but note that research has revealed “in recent years important advances in our understanding of the molecular mechanisms governing host resistance and in maintaining tissue homeostasis in the urinary tract to ultimately prevent constant microbial invasion have been made.”    

The body’s innate immune response to this strain of UPEC infection begins with the physical barriers of the urothelial cells.  In Billips’, et al. research on the Molecular Basis of Uropathogenic E. coli Evasion of the Innate Immune Response in the Bladder, it is reported that the body’s robust innate immune response produces the inflammatory cytokines and chemokines in the urothelium.  The production of these chemical messengers “results in the rapid recruitment of neutrophils into the bladder lumen and in bacterial clearance.”  The activation of this response in the urinary tract depends upon recognition by the pattern recognition receptors (PRR) of the pathogen-associated molecular patterns (PAMPs) such as lipopolysaccharide (LPS) or type 1 pili.

In addition to this recognition mechanism, toll-like receptors (TLRs) function in a similar way through a signaling cascade to produce inflammatory cytokines and chemokines.  However, some UPEC can suppress the activation of these components.  Specifically, UPEC strain NU14 can suppress the secretion of interleukin-6 (IL-6) and IL-8 and the proinflammatory transcription factor NF-KB.  When UPEC suppresses NF-KB, there is enhanced type 1 pilus-mediated apoptosis, which decreases the levels of inflammatory cytokines and thus neutrophil recruitment.  

The proinflammatory transcription factor NF-KB is an example of a localized inflammatory tissue response resulting in the release of histamine.  The cardinal signs of inflammation are redness, pain, heat, and swelling.  (The reason for dysuria with an UTI is because the inflammation causes the detrusor muscle of the bladder to spasm and that creates even more irritation.  Also the inflammation of the lining of the bladder and urethra is what causes the burning sensation upon urination.)  The adherence of UPEC to the bladder lining initiates this inflammatory response.  Mast cells respond by:
  •  Increasing blood flow resulting in swelling
  •  Activating phagocytes which engulf and destroy pathogens
  •  Increasing capillary permeability
  •  Activating complement for pore formation, enhanced phagocytosis, and histamine release
  •  Initiating the clotting reaction to wall off region (if there is hematuria)
  •  Increasing local temperature to disrupt pathogen
  •  Activating adaptive defenses with specificity, versatility and memory
Depending on the amount of inflammation, the vasodilation that results will affect the afterload of the heart, peripheral blood pressure, and capillary blood flow.  Vasodilation lowers blood pressure which decreases afterload, the amount of pressure needed by the ventricle to open the semilunar valve to eject blood.  A decrease in afterload provides less time for isovolumetric contraction and leads to a decrease in end-systolic volume (ESV).  This in turn equals to an increase in EDV, stroke volume and cardiac output.  This is another way a connected body system works to maintain homeostasis.

These innate immune defenses are critical to homeostasis.  Additional immune responses by the body to remove pathogens in order to maintain health involve the cell-mediated and the antibody-mediated response of adaptive immunity.  When the uropathogenic E. coli enter the host cell, the host releases IL-6 and IL-8 which operate to recruit macrophages to the affected area.  Additionally, the infected cell will present a major histocompatibility complex (MHC) on its surface.  This is a signal for a lymphocyte to bind and activate.

T cell lymphocytes are cell-mediated so upon encountering an infected cell presenting a Class I MHC, the T cell will bind at its CD8 marker to the MHC glycoprotein and will activate.  Upon activation, the T cells divide forming active cytotoxic T cells (Tc) and memory Tc cells.  The cytotoxic T cell destroys the infected cell in one of three ways.  First the Tc can release perforin, which essentially punches holes in the cell membrane causing water to flow into the cell until it bursts.  Second, the release of cytokines may stimulate apoptosis.  Finally, lymphotoxins may be released, which interfere with cellular metabolism.

Other T cells with CD4 markers will bind to infected cells that present a Class II MHC on its surface.  When activated, they divide into helper T cells (TH) and memory helper T cells.  The TH cell secretes cytokines which stimulates T cell division, attracts macrophages, attract and stimulate the action of Tc cells, and promotes the activation of B cells.  B cells are responsible for antibody-mediated immunity.  They are sensitized when exposed to antigens in the interstitial fluid.  The antibodies of the B cell bind to the antigens and then present them on their cell membrane.  When the activated TH cell encounters this antigen-antibody binding, it releases cytokines that costimulate the B cell resulting in B cell activation.  The activated B cell then divides producing memory B cells and the plasma cells that secrete the antibodies.

There are 5 classes of antibodies:   IgM, IgD, IgA, IgG, and IgE.  With the antigen-antibody complexes, there are 7 ways in which these antibodies will eliminate the antigen.  They are:
  • Prevention of bacterial and viral adhesion
  • Neutralization
  • Agglutination
  • Attraction of phagocytes
  • Stimulation of inflammation
  • Opsonization
  • Activation of Complement
Upon invasion by the uropathogenic E. coli that causes the urinary tract infection in greater than 80% of all cases, the adaptive immune response recognizes the signals from the infected cell and activates to destroy that cell, which in turn will destroy the bacteria.  The cytotoxic T cells work directly to destroy the infected cell.  The helper T cell releases cytokines which sensitize and activate the B cells.  This is when the immunoglobulins go to work.  IgM is the first responder whose concentration will decline as IgG production increases.  Since the bladder is lined by a layer of mucosa, IgA will also be present as it will attack pathogens before they gain access to the internal tissue of the mucosa.

The graphic above provides a scenario of some of the innate and adaptive immune responses involved with a urinary tract infection.  The graphic depicts the blocking action of secretory IgA and a protein called a Tamm-Horsfall Protein.  These adhere to the fimbriae of the bacterium to prevent attachment to the bladder lumen leaving the bacteria suspended in the urine, which will be flushed out of the body resulting in bacterial clearance

For UPEC that make it past this blocking action to bind to and invade the host, the infected cell releases cathelicidin, defensins, and nitric oxide to directly combat the bacteria.  At the same time, the cell releases IL-6 and IL-8 to enlist the phagocytic action of the neutrophils.  Cathelicidin is a microbial peptide that is important in maintaining the integrity of the urinary tract by its action to destroy uropathogenic bacteria.  

In summary of homeostasis, the body has several lines of defense against the colonization of E. coli in the urinary bladder to restore balance back to a state of health.  With the UTI, such defenses include the composition of urine, the micturition reflex, and the innate and adaptive immune responses.


A CBC and/or a blood culture may be ordered by the doctor and certainly a urine sample is collected to run a urinalysis.   First the sample is visually examined for color and clarity.  A cloudy, red urine sample, for example, indicates abnormalities.  A urinalysis testing positive for a UTI may show abnormally high levels for white blood cell count and nitrites.  Nitrites indicate the presence of bacteria since bacteria can convert nitrates to nitrites.  Also the red blood cell count may be high due to inflammation causing blood vessels to leak RBCs into the urine.  If bacteria are present, a urine culture may be done to identify the type of bacteria, which knowing this is important in prescribing the correct antibiotic for treatment.

The results of the urinalysis completed upon my family member’s visit to the ER are listed below.  From her clean catch urine sample, the pathologist observed the color was red and the clarity was turbid.  This indicates the presence of both red blood cells (hematuria) and white blood cells (pyuria), which is typical when there is infection.  Some of the specific components that are indicators of infection with abnormally high levels in her urine were for nitrites, white blood cells, red blood cells and bacteria.  Upon the presence of bacteria, a urine culture was also done.  This showed the bacteria responsible for her infection to be E. coli with a too many CFUs (colony forming units) to count.

Also of interest with this urinalysis is the specific gravity.  A result of 1.026 is at the high end of normal.  This means there were more solutes in her urine making it more concentrated.  This also means she was not taking in enough fluids to be properly hydrated.  One risk of limited intake of fluids is the formation of kidney stones.  There needs to be enough water available to the urinary system in order to properly flush out the solutes as well as invading bacteria.

How was her body working to restore homeostasis?  First, the pH was within the normal range at 6.0.  According to the research cited above, this measure of pH was still at a level of activity where her body was working to inhibit the growth of the bacteria.  Second, was the frequent urge to urinate as this micturition reflex worked to flush out bacteria in the bladder.  Finally, her body’s innate and adaptive immune responses kicked into high gear as evident by the pain, cramping, bleeding and blood clots resulting from inflammation.  The inflammation was necessary to bring in the white blood cells to destroy the bacteria; to activate T cells, B cells and subsequent antibodies; and to wall off the infected area to prevent the bacteria from spreading.

If it is a case of complicated UTI, as seen with higher risk patients such as pregnant women, those on catheters, or those in nursing homes, additional tests may be ordered.  These tests include a CT scan of the abdomen, kidney ultrasound, intravenous pyelogram, or a voiding cystourethrogram.  These tests will look for other problems such as kidney infection (pyelonephritis), kidney stones, structural abnormalities, or, in men, chronic prostatitis.


Mild cases of acute cystitis may disappear on their own without treatment, but since there is a risk for kidney infection, treatment with antibiotics is usually recommended.  In the case of my family member, her urine culture noted that the best antibiotic to take to fight E. coli is a piperacillin such as tazobactam, which is a beta-lactam effective against Gram-negative bacteria.  An alternative to antibiotics for mild cases is to drink cranberry juice.  Studies have shown that the active compound of cranberries works to prevent bacterial adhesion.  This is also useful as a preventative measure.
The following antibiotics are used to treat UTIs:  (Source:
  • Beta-lactams, including penicillins and cephalosporins (for example, Amoxicillin, Augmentin, Keflex, Duricef, Ceftin, Lorabid, Rocephin, Cephalexin, Suprax, and others); many organisms have resistance to some of these drugs.
  • Trimethoprim-sulfamethoxazole combination antibiotic (for example, Bactrim DS and Septra); many organisms may show resistance.
  • Fluoroquinolones (for example, Cipro, Levaquin, and Floxacin) resistance is developing; also these should not be used in pregnancy or in the pediatric population.
  • Tetracyclines (for example, tetracycline, doxycycline, or minocycline) used most often for Mycoplasma or Chlamydia infections; like fluoroquinolones, they should not be used in pregnancy or by the pediatric population.
  • Aminoglycosides (for example, gentamycin, amikacin, and tobramycin) used usually in combination with other antibiotics to combat severe UTIs.
  • Macrolides (for example, clarithromycin, azithromycin, and erythromycin), used more often with some STD-caused urinary problems.
Treatment with antibiotics is usually successful and symptoms of the infection usually disappear within 24 -48 hours.  

Recurrence is a possibility for about 20-30% of women who’ve had one episode of UTI.  It is considered a recurrence if infection returns for three or more episodes with a 12 month period.  Recurrence is caused by different bacteria than the one that caused the first infection.  If a repeat episode happens because of the same organism, it is considered a relapse as opposed to a recurrence and occurs less often.

Billips, Benjamin K., Anthony J. Schaeffer, and David J. Klumpp:  Molecular Basis of Uropathogenic Escherichia coli Evasion of the Innate Immune Response in the Bladder.  Infection and Immunity, Sept. 2008; Vol. 76, No. 9: 3891-3900.
Kaye, Donald:  Antibacterial Activity of Human Urine.  The Journal of Clinical Investigation, October, 1968; 47(10): 2374-2390.
Martinez, Juan J., Matthew A. Mulvey, Joel D. Schilling, Jerome S. Pinker and Scott J. Hultgren:  Type 1 Pilus-mediated Bacterial Invasion of Bladder Epithelial Cells.  The European Molecular Biology Organization Journal, 2000; Vol. 19 No. 12: 2803-2812.
Martini, Nath, and Bartholomew:  Fundamentals of Anatomy & Physiology, 9th edition.
Puri, Randhir and Jaideep Malhotra:  Recurrent Urinary Tract Infection (UTI) in Women.  South Asian Federation of Obstetrics and Gynecology, January – April 2009; 1(1): 10-13.
Saemann, Marcus D., Walter H. Horl and Thomas Weichhart:  Uncovering Host Defenses in the Urinary Tract:  Cathelicidin and Beyond.  Nephrology Dialysis Transplantation, October, 2008; 22:347-349.
Thumbikat, Praveen, Carl Waltenbaugh, Anthony J. Schaeffer, and David J. Klumpp:  Antigen-Specific Responses Accelerate Bacterial Clearance in the Bladder.  The Journal of Immunology, 2006; 176: 3080-3086.

Thursday, March 22, 2012

Just Starting Out

I am going to write this blog from the perspective of a student new to the world of microbiology.  I will share what I've explored and learned about microbes beginning from the undergraduate level.

I hold a BS degree in Special Education.  I taught at the high school level for 8 years in the area of learning disabilities and behavior disorders.  I left teaching for the corporate world where I worked for a very large company as a Webmaster and Knowledge Advisor.  When the economic downturn peaked in 2008, I was laid off from a job I loved.  I became one of the long-term unemployed as I was unable to find a job in the Chicago area for 2 years.  I relocated to Northwest Arkansas where there were more opportunities.  This is when I decided I was going back to school for a new degree and a new career.  I selected nursing.

Microbiology is a prerequisite for nursing school and little did I know that when I took this class last semester a whole new world would open up to me.  Not only was I very interested in Micro, but I also did exceedingly well in the class!  Currently, I have the privilege of being one of 6 students selected to conduct undergrad research in food microbiology at my school.  We just completed our short-term study and now we are in the process of writing up our research paper.  We will be presenting a poster at a science conference in Fayetteville this Fall.  I now have a passion for Micro and have created a new long-term goal for myself:  attend graduate school at the University of Arkansas to obtain a Ph.D. in Cell & Molecular Biology.

But my path to a Ph.D. is not a straight path as I am faced with some hurdles.  The financial aid I am receiving right now is for nursing school.  I have not figured out a way where I could forgo nursing to start immediately on taking the science classes I need for grad school.  You see, my BS degree is not in Biology so my transcript is deficient in Chemistry, Physics, and Life Science and I need most of these completed before I apply to grad school.  I only work part-time and do not earn enough to pay for these classes on my own.  I suppose I could change my degree program to Biology, but the problem there is the need for me to re-enter the workforce asap in order to help support myself, and in the event my boyfriend loses his job, to support both of us.  Nursing is the fastest route for me to get to a place where I am able to earn a salary again.

My plan right now is to proceed with nursing school.  I have applied to the nursing school at NWACC and expect to be accepted in May 2012 to start the next Fall.  I expect to be pinned as a nursing graduate in Spring 2014 and then I have to pass the NCLEX to become an R.N.  I would like my first nursing job to be involved with infection control so I can stay tied to my interest of microbes.  With a decent salary, I will be able to take the undergrad science classes I am deficient in so I can "beef up" my transcript.  Then I will apply to grad school, win some scholarships and maybe even a fellowship to help pay for grad school, and finally be on a straighter path towards a Ph.D.