Rapid Diagnostic Tests for Infectious Diseases
Rapid Diagnostic Tests for Infectious Diseases
History and Scientific Foundations
Introduction
Before the era of molecular biology, determination of the identity of the cause of an infectious disease—the process of diagnosis—involved the observation of the symptoms of the disease, the culturing (growing and identifying) of the responsible bacteria, virus, or protozoa (which still is not always possible or may require a long time), and the results of a variety of biochemical tests. Diagnosis typically took days or weeks.
Beginning in the 1970s, the ability to detect target regions of bacterial and viral deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) has made it possible to both demonstrate the presence of microorganisms in blood, urine, and tissue samples from an infected person, and now to identify the microorganism even to the species level. Some of these molecular-based tests can be done in minutes.
Other rapid diagnostic tests are based on the presence of an antibody that is produced by the human immune system in response to the presence of a certain bacterial or viral component, usually a protein, that is generically termed an antigen. These immunotests are based on the detection of the binding of the sample antigen to the antibody.
History and Scientific Foundations
Rapid molecular tests rely on the detection of target regions of the microbial genetic material. Once the genetic sequence—the arrangement of the building blocks (nucleotides) of the DNA or RNA—of a variety of important microbial pathogens has been determined, target regions that are unique to a given organism or a gene that codes for the presence of an important diseasecausing contributor such as a toxin can be identified. The detection of these target regions can be proof of the presence of the microbe even in the absence of the actual isolation of the organism. Furthermore, comparison of the genetic sequence with sequences that have been saved in databases can identify the genus and sometimes even the species of the infecting microorganism.
The target genetic region may only be present in low quantities. A technique called polymerase chain reaction (PCR)—which was developed in the 1980s and which earned its discoverer, Kary Mullis, the 1993 Nobel Prize in Chemistry—enables the amplification of bits of DNA. Because each PCR cycle doubles the amount of the genetic material and because cycles can be done quickly (sometimes in minutes), literally billions of copies of the target DNA can be made in a few hours.
Immuno-based tests rely on the binding of a particular sample antigen to its corresponding antibody that is bound to a support such as a paper strip. Antigen-antibody binding is a specific reaction. Other antigens in a sample will not bind to the bound antibody unless they are almost identical both in the arrangement of amino acids that makes up the protein but in the three-dimensional shape adopted by the protein molecules in solution. As well, commercially available immunostrips contain controls that verify that the observed antigen-antibody binding, which is detected by the development of a color, is not a mistake.
Applications and Research
Rapid diagnostic tests have become popular in the diagnosis of infectious diseases. Since the 1980s, various antigen-antibody binding tests have been available for the examination of fluid specimens that include whole blood, the serum, and plasma components of blood, saliva, urine, and even fluid recovered from tissues.
The various tests, which can be capable of detecting as little as one nanogram of an antigen in the sample, include hepatitis B, human immunodeficiency virus (HIV), malaria (based on detection of an antigen of the malaria-causing microbe, Plasmodium falciparum), syphilis (based on detection of a Treponema pallidum antigen), Streptococcus (a common cause of a throat infection known as “strep throat”), urinary tract infections (based on the enhanced production of several enzymes during an infection), and influenza.
WORDS TO KNOW
ANTIBODY: Antibodies, or Y-shaped immunoglobulins, are proteins found in the blood that help to fight against foreign substances called antigens. Antigens, which are usually proteins or polysaccharides, stimulate the immune system to produce antibodies. The antibodies inactivate the antigen and help to remove it from the body. While antigens can be the source of infections from pathogenic bacteria and viruses, organic molecules detrimental to the body from internal or environmental sources also act as antigens. Genetic engineering and the use of various mutational mechanisms allow the construction of a vast array of antibodies (each with a unique genetic sequence).
ANTIBODY-ANTIGEN BINDING: Antibodies are produced by the immune system in response to antigens (material perceived as foreign). The antibody response to a particular antigen is highly specific and often involves a physical association between the two molecules. Biochemical and molecular forces govern this association.
ANTIGEN: Antigens, which are usually proteins or polysaccharides, stimulate the immune system to produce antibodies. The antibodies inactivate the antigen and help to remove it from the body. While antigens can be the source of infections from pathogenic bacteria and viruses, organic molecules detrimental to the body from internal or environmental sources also act as antigens. Genetic engineering and the use of various mutational mechanisms allow the construction of a vast array of antibodies (each with a unique genetic sequence).
CULTURE: A culture is a single species of microorganism that is isolated and grown under controlled conditions. The German bacteriologist Robert Koch first developed culturing techniques in the late 1870s. Following Koch's initial discovery, medical scientists quickly sought to identify other pathogens. Today bacteria cultures are used as basic tools in microbiology and medicine.
IMMUNO-BASED TEST: An immuno-based test is a medical technology that tests for the presence of a disease by looking for reaction between disease organisms that may be present in a tissue or fluid sample and antibodies contained in the test kit.
PCR (POLYMERASE CHAIN REACTION): The Polymerase Chain Reaction, or PCR, refers to a widely used technique in molecular biology involving the amplification of specific sequences of genomic DNA.
The rapid detection of influenza is noteworthy since influenza viruses are characterized by their changing outer surface, and so their antigenic composition, from year to year. The rapid test targets a viral component that has proven to be more stable over time.
Research continues to refine the molecular and immuno-based rapid diagnostic tests, both in terms of their accuracy and the spectrum of microbial diseases that can be detected. For example, research published in 2006 reported on a PCR-based system that allows different types of hemorrhagic fevers to be distinguished. Since speed is vital is the treatment of people suffering from hemorrhagic diseases such as Ebola, this advance will help increase the survival rate of this traditionally lethal suite of diseases.
Impacts and Issues
Being able to rapidly diagnose infectious diseases can help initiate treatment faster, which can be key in combating a swiftly spreading infection. Furthermore, for diseases such as influenza that are caused by a virus, a rapid diagnosis curtails the misuse of antibiotics, which are useless against viral infections but which can stimulate the development of antibiotic resistance in resident bacteria. Such antibiotic misuse has been a key factor in the development of bacterial antibiotic resistance, which is increasingly making diseases such as tuberculosis more difficult and expensive to treat.
Molecular techniques require specialized (and expensive) equipment and trained personnel. This can be a limitation for smaller clinics in developed countries and can be completely impractical for rural clinics in developing and underdeveloped countries. Immuno-based tests are less expensive, the test strips are easier to transport and store as refrigeration is usually not required, and the results are clear and do not require interpretation.
The use of immuno-based strip tests has brought rapid diagnostic testing to rural clinics in underdeveloped and developed regions. Staff at these clinics can be easily trained to carry out and interpret the test results. The tests are also useful to field staff of agencies including the World Health Organization and the United States Centers for Disease Control and Prevention, which respond to illness outbreaks. The rapid detection of a disease and its geographical scope can be vital in combating an outbreak.
IN CONTEXT: TERRORISM AND BIOLOGICAL WARFARE
The Advanced Diagnostics Program is funded by the Defense Advanced Research Projects Agency of the United States government (DARPA). Its objective is to develop tools and medicines to detect and treat biological and chemical weapons in the field at concentrations low enough to prevent illness. Challenges to this task include minimizing the labor, equipment, and time for identifying biological agents. One area of interest includes development of field tools that can identify many different agents. To accomplish this goal, several groups funded under the advanced diagnostics program have developed field-based biosensors that can detect a variety of analytes including fragments of DNA, various hormones and proteins, bacteria, salts, and antibodies. These biosensors are portable, run on external power sources, and require very little time to complete analyses.
A second focus of the advanced diagnostics project is the identification of known and unknown or bioengineered pathogens and development of early responses to infections. A final goal is to develop the ability to continuously monitor the body for evidence of infection. Researchers are addressing this goal in two ways. The first involves engineering monitoring mechanisms that are internal to the body. In particular, groups funded under the initiative are developing bioengineered white blood cells to detect infection from within the body. Often genetic responses to infection occur within minutes of infection so analysis of blood cells provides a very quick indication of the presence of a biological threat. The second method involves the development of a wearable, non-invasive diagnostic device that detects a broad-spectrum of biological and chemical agents.
Rapid diagnostic tests are also being increasingly used to detect microbial contamination of food and water, especially since the deliberate release of anthraxladen letters in the autumn of 2001 in the United States. The realization that the nation's food and drinking water supplies are vulnerable to malicious contamination has spurred efforts to establish safeguards.
See AlsoFood-borne Disease and Food Safety.
BIBLIOGRAPHY
Books
Brunelle, Lynn, and Barbara Ravage. Bacteria. Milwaukee, WI: Gareth Stevens Publishing, 2003.
DeGregori, Thomas R. Bountiful Harvest: Technology, Food Safety, and the Environment. Washington, DC: Cato Institute, 2002.
Periodicals
Greenwald, Jeffrey L., Gale R. Burstein, Jonathan Pincus, and Richard Branson. “A Rapid Review of Rapid HIV Antibody Tests.” Current Infectious Disease Reports 8 (2006): 125–131.
Palacios, Gustavo, et al. “Masstag Polymerase Chain Reaction for Differential Diagnosis of Viral Hemorrhagic Fevers.” Emerging Infectious Diseases 12 (2006): 692–695.
Brian Hoyle