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Diagnosis of Infectious Disease


Maria T. Vazquez-Pertejo

, MD, FACP, Wellington Regional Medical Center

Reviewed/Revised Oct 2022
Topic Resources

Doctors suspect an infection based on the person's symptoms, physical examination results, and risk factors. First, doctors confirm that the person has an infection rather than another type of illness. For example, a person with a cough and difficulty breathing may have pneumonia (a lung infection). However, the person may instead have asthma or heart failure, which are not caused by infection. In such a person, a chest x-ray can help doctors distinguish pneumonia from the other possible disorders.

Once doctors confirm that the person has an infection, they usually need to know which specific microorganism is causing the infection. Many different microorganisms can cause a given infection. For example, pneumonia can be caused by viruses, bacteria, or, rarely, fungi. The treatment is different for each microorganism.

Many different types of laboratory tests can identify microorganisms. Laboratory tests use a sample of blood, urine, sputum, or other fluid or tissue from the body. This sample may be

No single test can identify every microorganism, and tests that work well for one microorganism often do not work well for another. Doctors must choose the test based on which microorganisms they think are most likely to cause a disorder.

Sometimes several different tests are done, typically in a specific order, based on the results of the previous test. Each test further narrows the possibilities. If the right test is not done, doctors may not identify the cause of infection.

Samples for Testing

A sample is taken from an area of the person's body likely to contain the microorganism suspected of causing the infection. Samples may include

  • Blood

  • Sputum

  • Urine

  • Stool

  • Tissue

  • Cerebrospinal fluid

  • Mucus from the nose, throat, or genital area

Some samples sent for testing, such as sputum, stool, and mucus from the nose or throat, normally contain many types of bacteria that do not cause disease. Doctors need to distinguish between these bacteria and those that could cause the person's illness.

Other samples come from areas that normally do not contain any microorganisms (that are sterile), such as urine, blood, or cerebrospinal fluid (the fluid that surrounds the brain and spinal cord). Finding any bacteria in such samples is abnormal as long as the area from which the sample was taken was first cleaned with an antiseptic to prevent contamination.

Staining and Examination Using a Microscope

Doctors sometimes can identify a microorganism simply by looking at it under a microscope.

Most samples are treated with stains. Stains are special dyes that color the microorganisms, causing them to stand out from the background. Some microorganisms have a distinctive size, shape, and stained color that enable doctors to recognize them.

However, many microorganisms look alike and cannot be distinguished using a microscope. Also, there must be enough of them, and they must be large enough to be seen with a microscope. For example, viruses cannot be identified using a microscope because they are too small.

For bacteria, doctors often first use Gram stain (a violet-colored stain). Bacteria are classified as follows:

  • Gram-positive (they look blue because they retain the violet Gram stain)

  • Gram-negative (they look red because they do not retain the stain)

Doctors can sometimes decide what antibiotics to use based on whether bacteria are gram-positive or gram-negative. Gram-positive and gram-negative bacteria tend to be susceptible to different groups of antibiotics.

In addition to Gram stain, other stains can be used depending on the microorganisms thought to be present.

Culture of Microorganisms

Many samples contain too few microorganisms to be seen using a microscope or to be identified using other tests. Thus, doctors usually try to grow the microorganism in a laboratory (called culture) until there are enough to identify.

The sample is placed on a sterile dish (plate) or in a test tube that contains specific nutrients to encourage growth of microorganisms. Different nutrients are used depending on which microorganism doctors suspect is causing the infection. Often, doctors add substances to the dish or test tube to stop the growth of microorganisms that do not cause the disease doctors suspect.

Many microorganisms, such as the bacteria that cause urinary tract infections or strep throat, can easily be grown in a culture. Some bacteria, such as the bacteria that cause syphilis, cannot be cultured at all. Other bacteria, such as those that cause tuberculosis, and fungi can be cultured but take weeks to grow. Some viruses can be cultured, but many cannot.

After the microorganisms are cultured, tests to identify them and to determine susceptibility and sensitivity to antimicrobial drugs are done.

Testing a Microorganism's Susceptibility and Sensitivity to Antimicrobial Drugs

Although doctors know in general which antimicrobial drugs are effective against different microorganisms, microorganisms are constantly developing resistance to drugs Antibiotic Resistance Antibiotics are drugs used to treat bacterial infections. They are not effective against viral infections and most other infections. Antibiotics either kill bacteria or stop them from reproducing... read more that were previously effective. Thus, susceptibility testing is done to determine how effective various antimicrobial drugs are against the specific microorganism infecting the person. This testing helps doctors determine which drug to use for a particular person's infection (see Selecting an Antibiotic Selecting an Antibiotic Antibiotics are drugs used to treat bacterial infections. They are not effective against viral infections and most other infections. Antibiotics either kill bacteria or stop them from reproducing... read more ).

Cultures are often used for susceptibility testing. Once a microorganism has been grown in a culture, doctors add different antimicrobial drugs to see which ones kill the microorganism. They also test how sensitive the microorganism is to a drug—that is, whether a small or a large amount of a drug is needed to kill the microorganism (sensitivity testing). If a large amount is needed to kill the microorganism in the laboratory, doctors usually do not use that drug.

Sometimes genetic testing Tests That Detect Genetic Material in Microorganisms Infectious diseases are caused by microorganisms, such as bacteria, viruses, fungi, and parasites. Doctors suspect an infection based on the person's symptoms, physical examination results,... read more can be used to detect genes in the microorganism that cause resistance to certain antimicrobial drugs. For example, methicillin-resistant Staphylococcus aureus (MRSA) bacteria can be identified by testing for the mecA gene.

Because susceptibility testing occurs in the laboratory, the result does not always match what happens in the person's body when a drug is given. Factors related to the person receiving the drug can influence how effective a drug is (see also Overview of Response to Drugs Overview of Response to Drugs Everyone responds to drugs differently. The way a person responds to a drug is affected by many factors, including Genetic makeup Age Body size Use of other drugs and dietary supplements (such... read more ). They include the following:

  • How well the person's immune system is working

  • How old the person is

  • Whether the person has other disorders

  • How the person's body absorbs and processes the drug

Tests That Detect Antibodies to or Antigens of Microorganisms

Some microorganisms, such as the bacteria that cause syphilis, cannot be cultured. To diagnose such infections, doctors may use a variety of tests called immunologic tests. These tests detect one of the following:

Antibody tests

Antibody tests are usually done on a sample of the infected person’s blood because antibodies circulate in the blood. They also can be done on samples of cerebrospinal fluid or other body fluids.

Antibodies Antibodies One of the body's lines of defense ( immune system) involves white blood cells (leukocytes) that travel through the bloodstream and into tissues, searching for and attacking microorganisms and... read more Antibodies are substances produced by a person's immune system to help defend against infection. They are produced by certain types of white blood cell when these white blood cells encounter a foreign substance or cell. It typically takes several days to produce the antibody.

An antibody recognizes and targets the specific foreign substance (antigen Overview of the Immune System ) that triggered its production, so each antibody is unique, made for a specific type (species) of microorganism. If a person has antibodies to a particular microorganism, it means that the person has been exposed to that microorganism and has produced an immune response. However, because many antibodies remain in the bloodstream long after an infection has resolved, finding antibodies to a microorganism does not necessarily mean the person is still infected. The antibodies may remain from a previous infection.

Did You Know...

  • Finding antibodies to a microorganism in a person's blood does not necessarily mean that the person is still infected because the antibodies may remain from a previous infection.

Doctors may test for several antibodies, depending on which infections they think are likely. Sometimes doctors just test whether an antibody is present or not. But usually they try to determine how much antibody is present. They determine the amount of antibody by repeatedly diluting the sample in half until it no longer tests positive for the antibody. The more dilutions it takes until the test is negative, the more antibody there was in the infected person's sample.

Because it takes several days to weeks for the immune system to produce enough antibody to be detected, diagnosis of an infection may be delayed. Antibody tests done right after people become ill are often negative. Thus, doctors may take one sample immediately and then take another one several weeks later to see whether antibody levels have increased. If levels of an antibody are low on the first test after people become ill, finding an increase in the antibody levels several weeks later suggests an active, current or recent (rather than a previous) infection.

Antigen tests

Antigens are substances that can trigger an immune response in the body. Microorganisms (and all cells) have antigens on their surface and inside them. Each type of microorganism has different, unique antigens. Thus, finding one of those antigens means that specific microorganism is present. Antigen tests detect the presence of a microorganism directly, so that doctors can diagnose an infection quickly, without waiting for a person to produce antibodies in response to the microorganism. Also, these tests can be used in people whose immune system cannot produce many antibodies, such as people who have recently had bone marrow transplantation or who have AIDS.

To do antigen tests, doctors take a sample from a person and mix it with a test antibody to the suspected microorganism. If there are antigens from that microorganism in the person's sample, they attach to the test antibody. Different methods can be used to detect the antigen-antibody combination. But whatever method is used, the presence of the antigen means that the microorganism is present and probably is the cause of the infection.

Tests That Detect Genetic Material in Microorganisms

Tests that detect genetic material from microorganisms are called

  • Nucleic acid–based tests

If a microorganism is difficult to culture or identify by other methods, doctors can do tests to identify pieces of the microorganism’s genetic material. This genetic material consists of nucleic acids: deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). Some of the DNA and RNA in each organism are unique to that organism. Thus, finding some of that DNA or RNA means that specific microorganism is present.

The polymerase chain reaction (PCR) is an example of this type of test. The PCR technique is used to produce many copies of a gene from a microorganism, making the microorganism much easier to identify.

Each genetic test is specific to only one specific microorganism. That is, a genetic test for hepatitis C virus detects only that virus and not any other. Thus, these tests are done only when a doctor already suspects a particular disease.

Most nucleic acid–based tests are designed to identify the presence of a microorganism (called qualitative testing). However, for certain infections, such as HIV and hepatitis C, tests also can measure how much of the microorganism's genetic material is present (called quantitative testing) and thus determine how severe the infection is. Quantitative tests can also be used to monitor how well treatment is working.

Nucleic acid–based tests can sometimes be used to check the microorganisms for genes or gene mutations that make the microorganism resistant to a drug. However, these tests are not completely accurate because not all resistance mutations are known. Thus, tests cannot check for all the genes for resistance that may be present. Also, these tests are expensive, not widely available, and available for only a few microorganisms.

Other Tests Used to Identify Microorganisms

Tests that identify certain other unique characteristics of microorganisms are sometimes called

  • Non-nucleic acid–based identification tests

For example, tests can be done to identify the following:

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