Commonly used methods for sickle cell disease screening

There are many different methods for screening for sickle cell disease. Then two most commonly used methods are isoelectric focusing and high performance liquid chromatography.

In isoelectric focusing, proteins are separated based on their specific isoelectric point. This point is determined by the pH in which the protein has no charge and does not migrate farther down the electric field. The specimen is added to a polyacrylamide gel attached to an electrical field and migrates from cathode to anode. The typical analytical run time for one test of ten samples to complete is 2.5 hours. The results are seen as sharp bands in their respective isoelectric points and are automatically calculated to show the percentages of each type of hemoglobin detected. Isoelectric focusing provides clearer banding patterns which are not affected by the analyte degrading in the blood spots. Because this method is not automated, the testing process is labor intensive and time consuming.

High performance liquid chromatography is ten times more expensive than isoelectric focusing. In this method, unknown hemoglobins are identified by comparing their retention times against a standard calibration curve. Test runs were considered normal with the presence of peaks in the zones representing HbF and HbA. Although variants are distinct, the presence of HbS in a specimen may interfere with the accurate HbA₂ measurement which identifies β-thalassemia. In a recent study, automated high performance liquid chromatography was calculated to have a coefficient of variation between 0.9-1.8% for the precision of retention times and a coefficient of variation of 4.4-17.5% for the precision of quantification. The retention times were shown to be within 0.01 minutes of the specified identification window. The results of this study showed this method to be an accurate and precise assay for the detection of abnormal hemoglobins.


Check out this journal article for more information

Confusing bacteria

Arcanobacterium haemolyticum is a gram positive rod with a matchbox or Chinese letter orientation. It was first discovered and described in 1946 in servicemen and people from the South Pacific. When plated out, the colony is small with a narrow zone of beta hemolysis and pits the agar. This organism has been a topic of controversy due to its resemblance to Corynebacterium pyogenes; because of this resemblance, A. haemolyticum was originally classified within the Corynebacterium genus with the subspecies hominis. Since this bacteria resembles Streptococcus pyogenes, the two are often confused and can be differentiated by the gram stain, inverse CAMP, and catalase reaction. This confusion was later resolved in 1982 when a new genus was created and this organism was renamed. The genus Arcanobacterium means secretive bacteria and is based on the characteristics of its fatty acid, peptidoglycan, and DNA.
Arcanobacterium haemolyticum typically infects teenagers and young adults aged 15 to 25 and is usually found in immunocompromised people. It is transmitted person to person through respiratory droplets and is found in the pharynx as well as the skin but there are no known risk factors. This bacteria causes pharyngitis, tonsillitis, and cellulitis. A rash believed to be caused by an exotoxin may be seen on the chest, abdomen, neck, and extremities in 20-25% of cases and can lead to diagnostic errors. The most common symptoms associated with A. haemolyticum are sore throat, pruritus, fever, pharyngeal erythema, and a nonproductive cough.

Arcanobacterium haemolyticum is usually treated with erythromycin, clindamycin, gentamicin, or cephalosporins. Serious infections may be treated with parenteral antimicrobial drugs.

Play this word search!

Introduction to Sickle Cell

Sickle cell disease is one of the most common monogenic diseases in the world with an approximated 7% of the world heterozygous carriers resulting in 300,000 newborns with severe hemoglobinopathies. It is an autosomal blood disease that causes the sickling of red blood cells. This sickle shape can lead to the development of cerebrovascular disease as well as cognitive impairment in children. If left untreated, it may also result in damage to organs such as the brain, kidneys, lungs, bones, and cardiovascular system. The sickling of red cells is caused at the sixth codon for the beta globin gene where glutamine is substituted for valine and has 900 different variants, most of which are not clinically significant. The sickling of these cells occurs due to a mutation in the hemoglobin gene and affects the ability of the hemoglobin to bind and release oxygen. This mutation results in crystallization that produces a polymer nucleus which grows and fills the cells, in turn disrupting the flexibility of the cell and causing dehydration. This can lead to a number of various complications, including a shortened life expectancy.

Sickle cell disease occurs in approximately 1 out of every 2500 newborns with black and Hispanic infants representing the majority. In infants, HbF is the major gene involved in determining the severity of the disease. Individuals who presented with a higher level of HbF generally had milder symptoms and fewer complications. Diagnosis of this disorder is based on the determination of variants of hemoglobin. Several newborn screening methods are currently being practiced; mainly isoelectric focusing and high performance liquid chromatography.

Check out this video for an animation of the sickle cell mutation in the blood.

Sick of the dorms

College can be a huge turning point in your life. New friends, new responsibilities, and new living accommodations. It can be very easy to get sick of the dorms, but it is even easier to get sick IN the dorms. One might assume that a terrible roommate is the biggest concern when moving into close quarters with other people, but airborne bacteria are actually far more serious. 

This week in lab, we isolated and identified bacteria from cerebrospinal fluid. My patient was a college student who presented with stiff neck and a petechial rash. He was said to be toxic, drowsy, and disoriented. 

These symptoms could be associated with almost any college student around exam time, but after several tests, he was found to have the bacteria Neisseria meningitides. Neisseria meningitides is the most common bacteria to cause bacterial meningitis in college aged students and the infection was most likely due to the close contact in the dormitories and could have been acquired from respiratory droplets from other asymptomatic carriers. The source of the infection is the bacteria in the nasopharynx and into the meninges. The bacteria then gain access into the CSF and cause inflammation in the subarachnoid space. Neisseria meningitides may lead to sepsis, organ damage, and even death; therefore treatment must be administered as quickly as possible after diagnosis. Penicillin is the preferred treatment option for confirmed cases of N. meningitides.  

Luckily, this disease is preventable with the proper vaccine. This vaccine is effective against 80% of the bacteria that cause meningitis and lasts about the same amount of time one spends in college. This is just one more way to prevent sickness in the dorms and ensure a happy, healthy college life.

Check out this brochure for information on the vaccine 

Welcome

Hello. My name is Eden Bray and this is my infectious diseases blog. Throughout the year I will be posting information regarding the current topics in class. For my first post,  I'll tell you a little bit about me. I'm from a small town in Kentucky and have been in Alabama a little less than a year. I received my Bachelor's degree in Chemistry from Kentucky Wesleyan College in Owensboro where I was also a member of the softball team. I'm currently enrolled in the Master's program for Clinical Laboratory Sciences at the University of Alabama at Birmingham. Some of my posts this year will about my graduate project, sickle cell disease. I hope you enjoy reading.