FUN2: 9:00-10:00Scribe: Ryan O’Neill

Monday, November 10, 2008Proof: Brannon Heape

Dr. WaitesSpirochetes and RickettsiaePage1 of 4

RPR – rapid plasma reagent, VDRL – venereal disease research laboratory, TP – Treponema pallidum

  1. Introduction [S1]:
  2. Objectives[S2]
  3. Objectives are listed on slide.
  4. Spirochetes
  5. Treponema pallidum
  6. Borrelia spp.
  7. Leptospira spp.
  8. Rickettsiae
  9. Rickettsiae
  10. Ehrlichia spp.
  11. Treponema pallidum [S3]: agent of syphilis.
  12. Are helical organisms with a spiral appearance. 5-15 microns in length.
  13. These organisms are motile. We have spirochetes in many different parts of our body (e.g. mouth), but these are thecommensal, non-pathogenic spirochetes.
  14. The important thing about syphilis is that it has not been successfully cultured on artificial media (unlike the staphylococci and streptococci). You can grow them in vivo in rabbit epithelial cells. The fact that you can’t cultivate these organisms in vitro complicates diagnostic methods.
  15. Treponema pallidum are susceptible to environmental conditions. This is why to spread syphilis you have to have venereal transmission (mucosal membrane to mucosal membrane). They don’t exist free in nature.
  16. Treponema pallidum do not stain with aniline dyes like the gram stain. The cell wall of the organisms is too thin and delicate to take up staining. This also complicates diagnosis.
  17. Stages of Syphilis [S4]
  18. Syphilis is indeed a sexually transmitted disease.
  19. Primary Stage - The lesion that you see following sexual contact with an infected person is a hard chancre or ulcer that will appear on the genitalia or whatever body part contacted the Spirochetes.
  20. Not many bacteria can penetrate normal intact skin, but Spirochetes can.
  21. This will happen 1-3 weeks after encounter with the organism.
  22. It is usually painless and will heal on its own. This is considered the primary stage of syphilis.
  23. Secondary Stage – A rash forms that may on the palms of the hands, soles of the feet and possibly throughout the entire body. A number of other different lesions such as mucous patches and condyloma around the genitals may also form.
  24. At this stage you have a spirochetemia. The spirochetes have spread from the lesion and traveled through the blood stream and the body’s immune system causes these reactions.
  25. [S5]: Stages of Syphilis continued
  26. Latent stage - after the secondary stage your body begins to mount an immune response and then typically everything will go away. Then you will go into a latent period (divided into early latent and late latent depending on how long after the initial infection).
  27. Although the disease is normally treated, this is the path it would follow if not treated.
  28. Tertiary syphilis is the stage that follows the latent period. This is when you’ve had the disease for several years in the untreated form. Much of the manifestations are the response of the body and the immune system’s response to the microorganism and you can get lesions in the heart on the aortic valve, granuloma type lesions called “gummas,” central nervous system syphilis when the spirochetes invade the brain and the spinal fluid.
  29. Tertiary syphilis stage may also alter your cognitive function and eventually lead to paresis or lead to tabes dorsalis leads to a positional unawareness due to nerve reception problems (example: not knowing where your feet are as you walk up the steps).
  30. Oral & Genital Chancres [S6]
  31. Examples of the lesions you will see with syphilis.
  32. Chancre on the penis of someone who has contracted syphilis.
  33. Lesions always appear where the actual contact of the spirochetes takes place.
  34. Skin Lesions of Secondary Syphilis [S7]
  35. Example of the rash lesions due to the spread of the spirochetes throughout the blood stream.
  36. Palms of the hands and soles of the feet are typical of syphilis.
  37. Condyloma lesions that you see in secondary syphilis.
  38. [S8]: Tertiary Syphilis “Gumma”
  39. Example of a granuloma in the tissue that surrounds the mouth – a “gumma”
  1. Congenital Syphilis [S9]
  2. This organism can cross the placenta during pregnancy if the timing is right. The baby can actually be born with secondary syphilis and have the “snuffles” where mucous is expressed through the nose that contains motile spirochetes. It can affect the entire body.
  3. Hutchinson’s incisors (notched) and Mulberry molars are dental lesions that result in transmission of the disease during the time of specific tooth development and the corresponding enamel.
  4. Some other manifestations of congenital syphilisare 8th nerveauditory deafness and the abnormality of the tibia of the lower extremity called saber shins that is bold like a sword because of the effect of spirochetes on the developing bone in the fetus.
  5. Diagnosis of Syphilis: Darkfield [S10]
  6. How do you diagnose syphilis?
  7. If you have a lesion due to syphilis, you can collect fluid and place it on a darkfield microscope and then look for the motile spirochetes. Only valuable if it is a genital lesion, not an oral lesion. This is because we have spirochetes in our mouth already.
  8. The value of a darkfield test for syphilis is you can look at it and know the answer right away.
  9. Darkfields are used if a patient has an active lesion of primary syphilis.
  10. Non Treponemal Tests (RPR, VDRL) [S11]
  11. You want to screen people for syphilis is they have had sexual relations with someone who shows signs of syphilis such as a rash or a chancre (even if it has come and gone).
  12. We need to look for an antibody response (serological response) because we cannot cultivate it in a laboratory.
  13. Two types of serological tests for syphilis:
  14. Screening tests – Non Treponemal Tests (RPR, VDRL)
  15. Confirmatory tests
  16. Screening Tests:
  17. Advantages: inexpensive, easy & quick
  18. Measure cardiolipin, lecithin and cholesterol antibodies. There are antigens based upon these compounds that are used to detect antibodies in the serum because when you develop an antibody response to the Treponema pallidumit will cross-react with these substances. Also, carbon particles with antigens attached mixed with the blood serum will cause an agglutination reaction if there is antibody present.A rising titer of antibody confirms the disease. If you get a positive screening test (such as cardiolipin), before you tell the patient they have syphilis, you need to confirm the test because there can be other things that cause a false positive (autoimmune diseases, pregnancy, etc.).
  19. You need to confirm it with a more specific Treponemal test (i.e. confirmatory test).
  20. Usually becomes reactive late in the primary stage (while the chancre is still there). If you have a normally functioning immune system, it will always be positive by the time you reach the secondary stage.
  21. If you have syphilis and you are not treated until the later part of the second stage, you may develop persisting IgG antibodies and have a positive serological test for the rest of your life.
  22. [SQ]: Are the two different tests you mentioned earlier the RPR and VDRL?
  23. [A]: No. RPR and VDRL are types of screening tests (Non-Treponemal). The other type of test is the confirmatory test (Treponemal). So screening and confirmatory tests are the two types that were mentioned.
  24. [S12]: RPR Nontreponemal Test
  25. Take a card and the patient serum and add the carbon particles that contain the cardiolipin antigens attached to them and mix together. If you see an agglutination reaction that means an antibody and antigen are reacting, which is a positive test.
  26. [S13]: Treponemal Tests
  27. Use antigens from the treponemal organism, so that you are making sure that the antibody that you are measuring is actually specific for the TP. These tests are more complex and expensive. These tests are only done after positive screening tests.
  28. Treponema pallidum Particle Agglutination (TPPA) is involved with gelatin particles sensitized with TP antigen. Note on the slide the agglutination reaction.
  29. This is what you do to confirm if you have a positive test with the RPR.
  30. [S14]: Fluorescent Treponemal Antigen (FTA) Test
  31. Take a microscope slide that contains the TP antigen, and then react this with the patient’s serum. If the patient has antibody against TP antigen, the antibody will bind to the antigen on the slide. Then you come back and add a fluorescent-labeled antibody against human IgG. The spirochetes will appear with fluorescence if positive.
  32. Disadvantage: expensive and require a lot of experience.
  33. However, the FTA is a good test and you only get a few false positives in autoimmune disease.
  34. You will also have a positive antibody for life. This may lead to a misdiagnosis later in life if a patient gets syphilis again and gets tested for it.
  35. The TPPA test is preferred over FTA due to these reasons.
  36. Borrelia Characteristics [S15]
  37. Note the Borrelia spirochete on the slide.
  38. They are larger than the syphilis organisms.
  39. In some cases you can see Borrelia in a blood smear because of the differences in the cell walls. They may even stain with the Giemsa stain. Generally not the gram stain though.
  40. Occasionally, you can see these organisms in a blood smear. This is usually not done because they are hard to see since they are so few in numbers.
  41. Unlike TP, you can sometimes cultivate in artificial media with some difficulty.
  42. Relapsing Fever: B. recurrentis & others [S16]
  43. Relapsing fever is a disease that Borreliarecurrentis
  44. Many of the Borrelia diseases are carried by arthropod vectors (e.g. ticks and lice) and can be transmitted by infected rats. This results in having periodic fever associated with the spirochetes in the bloodstream. Chills and headaches will occur.
  45. These organisms are mainly in the Western U.S.
  46. You can make the diagnosis by measuring antibody against the Borrelia.
  47. Diagnosis problem - These organisms can shift their surface antigens. This results in a problem of diagnosing these infections is that you can get a false negative because it doesn’t account for this antigen shifting.
  48. Lyme Disease Borrelia burgdorferi [S17]
  49. Another type besides the relapsing fever is Lyme Disease.
  50. Caused by Borrelia burgdorferi. The life cycle of this organism wasn’t understood until 1984 and it has been reported over 40 states. Not common in Alabama because we don’t have the right kind of ticks (Ixodes ticks). These ticks carry the Lyme spirochete and if they bite an animal the Borrelia from its saliva can enter into the animal and soon create a reservoir of the spirochete.
  51. Even though it is not common in Alabama, people travel to other places where these reservoirs are common.
  52. The spirochetes that are in the saliva of the tick are injected into the body and travel through the blood stream. They get into the nerves, heart and the joints.
  53. It is not a severe disease, but it is a slow onset problem where you have joint pains, not feeling well, neural pains. This causes a problem with the diagnosis. You need to measure antibody production against the spirochete. The problem that you run into is that people don’t often go to the doctor right away and that the antibody production varies in different people. If the antibody test comes back negative, but you still think this person has Lyme Disease.
  54. If you see something like this [S18] Erythema Chronicum means that it is very likely that you have the Lyme Disease and there is no need for further testing.
  55. Ultimately, you need to measure antibody production and their rise and fall. The problem is that people don’t often go to the doctor right away and that the antibody levels vary in different people as well as varieties in laboratories. You will need to test again within a month to compare the antibody levels.
  56. [S18]: Erythema Chronicum Migrans Rash
  57. [S19]: Ixodes Tick Vector for Lyme Disease
  58. Normal tick and blood-filled tick.Are arthropods, but not insects.
  59. Treatment is tetracycline or doxycycline that will eradicate the infection.
  60. There was a vaccine made for Lyme disease for the Western U.S. population, but side effects caused vaccine to fail.
  61. Leptospira [20]
  62. Third and final group of Spirochetes.
  63. Tightly spiraled, obligate anaerobes. It takes 4 weeks to grow them anaerobically.
  64. Leptospira interrogans is the prototype organism.
  65. Like the other previous examples, antibody detection is the primary means of detection of Leptospirosis.
  66. [S21]: Leptospirosis
  67. Catch from contact with animals (e.g. dogs, cats, pigs, etc.) and they carry the Leptospira in their kidneys.
  68. A typical way that a human might catch this disease is to walk barefoot through animal urine. These bugs like the spirochetes, can enter through mucous membranes or the skin and gain access to the bloodstream, kidney or liver.
  69. They cause fever of unknown origin (FUO), meningitis or jaundice. Overall hepatitis type symptoms.
  70. Unexplained hepatitis or meningitis that is not easily diagnosed might require ordering an antibody test.
  71. If you catch it early enough in the disease before Leptospiraspread, you can see the organisms in the spinal fluid. But usually the diagnosis rests of serology.
  72. Rickettsiae [S22]
  73. Are similar to the Chlamydia in that they are obligate intracellular organisms
  74. Considered gram-negative bacteria, just like the Chlamydia.
  75. Most of these diseases are transmitted by arthropods. Exception: Coxiella
  76. Are true bacteria because they contain both types of nucleic acids (DNA and RNA) as opposed to viruses that only contain one or the other. Are also stainable with Giemsa stain & reproduce by binary fission.
  77. The Rickettsiae are very dangerous organisms and should never be grown in diagnostic labs because they can become aerosolized and infect the workers.
  78. Are susceptible to heat, drying and bactericidal chemicals, so you can eliminate them.
  79. Primary means of diagnosis is serology. Immunohistochemistry (antibody tagged to a dye) & PCR tests are becoming more popular in diagnosis as well.
  80. [S23]: Rickettsiae in Cytoplasm of Host Cell
  81. Arrows are pointing to the Rickettsiae in the cytoplasm. This whole cell is the host cell.
  82. The prototype disease caused by Rickettsiae is the Rocky Mountain Spotted Fever
  83. [S24]: Rickettsia: Pathology
  84. Upon biting from a tick, the Rickettsiae migrate to the endothelial cells where they invade and are taken up by the capillary endothelium where they multiple and they produce a vasculitis, which is why you get the spots in Rocky Mountain Spotted Fever.
  85. The cell begins to swell because the bacteria is growing inside of the cells and they take over the metabolism of the cell and the cells die and can clot the vessels which leads to more vasculitis. As the cells die, the bacteria are released into more parts of the body causing more lesions in more capillaries in other parts of the body.
  86. Ultimately, you can get disseminated intravascular coagulation.
  87. The organism can even replicate within phagocytic cells because it is mechanisms by which it cannot be killed by the cells. 20% of the people who get this may die.
  88. Rocky Mountain Spotted Fever treatment is usually tetracycline because you need to use an antibiotic that can get into cells. Whenever you have an intracellular infection you have to use a drug like this.
  89. [S25]: Selected Rickettsial Diseases of USA
  90. Rocky Mountain Spotted Fever is actually more common in the southeast. The tick is the vector but there are usually reservoirs in other types of animals.
  91. Typhus is caused by Rickettes and is a disease that breaks out in crowded situations where rats are living in the house with the family. Very rare in U.S.
  92. Ehrlichia are carried by ticks and infect white blood cells. Have monocytic and anaplasma (granulocytic) types.
  93. [S26]: Macular Rash of RMSF
  94. Rash of Rocky Mountain Spotted Fever.
  95. This is Ehrlichia chafeensis in a monocyte.
  96. [S27]: Ehrlichia chafeensis in a monocyte
  97. Note red cells and white cell’s monocyte nuclei.
  98. This is the Ehrlichia inclusion here (arrow).
  99. Note the mulberry lesion that is seen as an inclusion in white blood cells.
  100. See this in white blood cells and can usually be detected measuring antibodies.
  101. Unless you actually look at the slides, you may not be able to see these bacteria.
  102. Dr. Waites gave an example of a patient who had Ehrlichia and was given the wrong antibiotics and died because of it. Only tetracycline can combat these intracellular organisms.
  103. An antibody test may not be positive right away because it takes a few days for antibodies to develop.
  104. When doing antibody tests, you normally need to test two blood samples (early and late) so you can compare the rise or fall of antibody count…this is a disadvantage of an antibody test.
  105. A PCR assay has the advantage of showing results sooner than an antibody test, but these are readily available for uncommon conditions like these.
  106. Remember, you have to suspect it first before you run the test!
  107. [SQ]: Is there a particular part of the body that these organisms like to attack?
  108. [A]: Yes. This organism grows in the white blood cells. If you get enough of them you will see them.

[end 50 min]