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  • Slide 1: Trends in diagnoses made in GUM clinics in the UK: 1998 – 2007 Trends over the 10 year period from 1998 to 2007 show a gradual rise in both the number of new STI diagnoses and other STI diagnoses (e.g. recurrent infections). Other diagnoses made in GUM clinics (e.g. candidosis & vaginosis) have remained relatively stable but rose sharply between the years 2006 and 2007.
  • Slide 3: Number of new diagnoses of selected STIs, GUM clinics, United Kingdom: 2007 Since 1998, diagnoses of uncomplicated chlamydia infections have increased by 150% from 48,726 in 1998 to 121,986 in 2007. This was followed by an increase in the viral infections genital warts (first attack) and genital herpes (first attack) between 2006 and 2007 of 7% and 20% respectively. Although there has been a 42% increase in gonorrhoea diagnoses since 1998, a continued decrease in the diagnoses has been observed since 2003. Between 2006 and 2007 there was no change in primary and secondary infectious syphilis.
  • Slide 13: Number of diagnoses of gonorrhoea by sex, GUM clinics, England & Wales*: 1925 – 2006 Diagnoses of gonorrhoea rose steadily during the 1960s and 70s and remained at high levels until 1985. Numbers decreased sharply during the early 1990s and diagnoses fell to their lowest levels since recording began. After 1997, diagnoses of gonorrhoea began to rise again in the United Kingdom; increasing by 46% between 1997 and 2006. However, since 2002 a decrease in diagnoses of 26% has been observed. The decline in the incidence of uncomplicated gonorrhoea between 1985 and 1988 may reflect changes in sexual behaviour brought about in response to the HIV epidemic. The subsequent rises observed from 1994 to 2002 suggest increases in unsafe sexual behaviours with people having increasing numbers of partners, and higher rates of partner change.
  • Slide 15: Rates of diagnoses of uncomplicated gonorrhoea by sex and age group, GUM clinics, United Kingdom: 1997 - 2006 Like other acute STIs, young people share a disproportionate burden of gonorrhoea. During 2006, in the United Kingdom, rates were highest in men aged 20-24 years (188/100,000 population) and women aged 16 to 19 years (128/100,000).
  • Slide 19: Number of new diagnoses of uncomplicated gonorrhoea by sex and men sexual orientation, GUM clinics, United Kingdom: 1996 - 2006 Since 1997, diagnoses of heterosexually and homosexually acquired gonorrhoea in men have risen by 30% (6,979 to 9,103) and 132% (1,954 to 4,524) respectively. During 2006, homosexually acquired gonorrhoea accounted for a third of diagnoses in men (4,524 of 13,627).
  • Slide 4: Rates of diagnoses of uncomplicated genital chlamydial infection by sex and country, GUM clinics, United Kingdom: 1997 - 2006 Rates of diagnoses of uncomplicated genital chlamydial infection have been increasing in all countries in the UK since 1997. In 2006, England had the highest rates of infection for both men (198/100,000 population) and women (196/100,000 population). Between 2005 and 2006, rates of chlamydia increased among men in all UK countries except Wales. Among women, an increase in rates was seen in Scotland and Northern Ireland.
  • Slide 5: Rates of diagnoses of uncomplicated genital chlamydial infection by sex and age group, GUM clinics, United Kingdom: 1997 – 2006 Rates of chlamydial diagnoses in GUM clinics have increased disproportionately between the age groups from 1996 to 2006. Among men, the rate of increase is especially high for those in the 20 to 24 year age group (1,144/100,000 in 2006), with lower increases seen among 16 to 19 and 25 to 34 year olds (544 and 444/100,000 in 2006). Higher rates (1,337 and 1,145/100,000) were observed among younger women (16-19 and 20-24 year olds), although the rates appear to be stabilising in this group since 2004.
  • Slide 24: Numbers of diagnoses of syphilis (primary, secondary and early latent) by sex, GUM clinics, England, Wales and Scotland*: 1931 - 2006 Diagnoses of infectious syphilis made at GUM clinics in England, Scotland and Wales peaked towards the end of World War II, and then fell sharply in the late 1940s. Men in England and Wales experienced increases in diagnoses throughout the 1960s and 70s, while female cases remained constant. During this period Scotland saw fluctuating figures which peaked in 1968 and 1978 in both men and women. The male to female ratio in diagnoses peaked at 8:1 in 1983 in England and Wales and was similarly high at this time in Scotland, reaching 10:1 in 1984. This suggests that sex between men was the most common route of acquisition. Diagnoses in men declined in the early to mid-1980s, coinciding with emerging awareness of HIV, adoption of safer sex practices, and a parallel fall in HIV transmission among homosexual men. Since the late 1990s there has been a 15 fold increase in syphilis diagnoses among men in England and Wales.
  • Slide 26: Number of diagnoses of infectious syphilis (primary and secondary) by sex and men sexual orientation, GUM clinics, United Kingdom: 1997 – 2006 Since 1997, the number of homosexually and heterosexually acquired infectious syphilis in men has increased sharply (75 and 11 times respectively). During 2006, homosexually acquired syphilis accounted for 58% (1,417 of 2,424) of diagnoses in men; in 1999 this figure was 32% (53 of 166). The large number of homosexually acquired cases are due to ongoing outbreaks in London, North West and South East regions.
  • Slide 27: Rates of diagnoses of infectious syphilis (primary & secondary) by sex and age group, GUM clinics, United Kingdom: 1997 - 2006 Unlike most other acute STIs, the burden of syphilis does not fall solely upon young people. Since 1997 rates have increased sharply in men aged 35 to 44 years (37 fold), 25 to 34 years (22 fold) and those aged 20 to 24 years (20 fold). Women experienced a markedly smaller increase for all age groups, but significant increases were made in the 16 to 19 and 20 to 24 age groups (7 and 6 fold respectively).
  • ppt

    1. 1. Bacterial Sexually Transmitted Infections Patrick Kimmitt
    2. 2. Today we are going to look at… <ul><li>Three distinct bacterial pathogens causing sexually transmitted infections </li></ul><ul><ul><li>Neisseria gonorrhoeae </li></ul></ul><ul><ul><li>Chlamydia trachomatis </li></ul></ul><ul><ul><li>Treponema pallidum </li></ul></ul>
    3. 3. We are going to consider… <ul><li>The organism, structure and physiology </li></ul><ul><li>The pathology of disease </li></ul><ul><li>Epidemiology </li></ul><ul><li>Laboratory diagnosis and treatment </li></ul><ul><li>There are many contrasts when looking at these uniquely adapted pathogens </li></ul><ul><li>You should be able to discuss each of these aspects </li></ul><ul><li>But first some background… </li></ul>
    4. 4. Trends in diagnoses made in GUM clinics in the UK: 1998 – 2007 Routine GUM clinic returns
    5. 5. Number of new diagnoses of selected STIs, GUM clinics, United Kingdom: 2007 Routine GUM clinic returns
    6. 6. Taking a sexual history and contact tracing is essential <ul><li>What did you do? </li></ul><ul><li>With whom? </li></ul><ul><li>When? </li></ul><ul><li>Where? </li></ul><ul><li>And what symptoms did it leave you with? </li></ul>
    7. 7. Gonorrhoea Neisseria gonorrhoeae
    8. 8. Clinical and epidemiological aspects <ul><li>2 nd commonest bacterial STI </li></ul><ul><li>2007: 18,710 cases reported to HPA </li></ul><ul><li>Most common age groups: males 20-24 females 16-19 </li></ul><ul><li>Males: usually symptomatic </li></ul><ul><li>Females: often asymptomatic </li></ul><ul><li>Complications: untreated females – PID, infertility, ectopic pregnancy </li></ul>
    9. 9. Number of diagnoses of gonorrhoea by sex, GUM clinics, England and Wales*: 1925 – 2006 * Scotland & Northern Ireland data are excluded as they are incomplete from 1925 - 2003 Routine GUM clinic returns
    10. 10. Rates of diagnosis of uncomplicated gonorrhoea by sex and age group, GUM clinics, United Kingdom: 1997 - 2006 Routine GUM clinic returns Males Females
    11. 11. Number of diagnoses of uncomplicated gonorrhoea by sex and male sexual orientation, GUM clinics, United Kingdom: 1997 - 2006 Routine GUM clinic returns
    12. 12. Symptoms (if present) <ul><li>Males: urethral discharge, severe burning on urination </li></ul><ul><li>Females: vaginal discharge, yellow or blood-stained, pain on urination </li></ul><ul><li>Rectal infection gives rise to pain and discharge </li></ul><ul><li>Pharyngeal infection, sore throat </li></ul>
    13. 13. Symptoms 2 <ul><li>Both sexes: disseminated infection on rare occasions – usually as septic arthritis* </li></ul><ul><li>Infection during pregnancy may lead to ophthalmia neonatorum of baby (conjunctivitis)- blindness </li></ul><ul><li>May see dual genital infection with Chlamydia trachomatis – usual to treat for both at time of gonorrhoea diagnosis </li></ul>* For more info – Kimmitt et al Journal of Travel Medicine (2008); 15; 369-371
    14. 14. Neisseria gonorrhoeae <ul><li>The causative organism is Neisseria gonorrhoeae , a Gram-negative diplococcus i.e. often see cells as a pair. The genus Neisseria contains one other pathogenic species, N. meningitidis, which is the principle cause of bacterial meningitis. There are also many non-pathogenic species of Neisseria, often found in the pharynx </li></ul>
    15. 15. Neisseria gonorrhoeae <ul><li>N. gonorrhoeae is phagocytosed by polymorphonuclear neutrophils but resists intracellular destruction, remaining intact within the neutrophil. </li></ul><ul><li>It is fastidious , sensitive to desiccation and requires aerobic incubation with 5% carbon dioxide for growth. It grows as a small colony, often requiring 48 hours incubation. The colonies are grey, shiny, often with an irregular edge. The organism is catalase positive and rapidly oxidase positive . </li></ul><ul><li>No protective antibody response to gonorrhoea: recurrent infections are common in people who are at risk. </li></ul>
    16. 16. Laboratory methods <ul><li>Culture is required - for identification and antibiotic sensitivity tests </li></ul><ul><li>Urethral, cervical, rectal or pharyngeal swab </li></ul><ul><li>Use selective medium containing antibiotics and growth supplements (look this up) </li></ul><ul><li>e.g. Thayer Martin or New York City media </li></ul><ul><li>Molecular tests have been developed for the direct detection of N. gonorrhoeae infection and a single swab may be used in a double test to detect N. gonorrhoeae and Chlamydia trachomatis. </li></ul><ul><li>Commercial tests include the COBAS Amplicor and SDA tests </li></ul>
    17. 17. Identification tests <ul><li>Once you have cultured your samples you need to perform tests on single colonies to check/confirm identification </li></ul><ul><li>Oxidase test - result? </li></ul><ul><li>Gram stain – what are you looking for? </li></ul><ul><li>Phadebact GC* – uses a specific monoclonal antibody </li></ul><ul><li>API NH – utilizes carbohydrates plus enzymes activity, similar to API 20E </li></ul>* N. gonorrhoeae is often referred to as a gonococcus or GC
    18. 18. Treatment <ul><li>There is increasing resistance to penicillin and now ciprofloxacin </li></ul><ul><li>The recommended treatment of gonorrhoea is now either ceftriaxone (injectable) or cefixime (oral). As yet, no resistance has been reported to these third generation cephalosporins. In either case, a single dose is all that is necessary for the treatment of non-disseminated gonorrhoea </li></ul>
    19. 19. Chlamydia Chlamydia trachomatis
    20. 20. Clinical and epidemiological aspects <ul><li>The most common bacterial sexually transmitted infection, with 121,986 cases reported to the Health Protection Agency in 2007 </li></ul><ul><li>The causative organism is Chlamydia trachomatis </li></ul><ul><li>The number of cases has risen steadily since the mid 1990s </li></ul>
    21. 21. Rates of diagnoses of uncomplicated genital chlamydial infection by sex and country, GUM clinics, United Kingdom: 1997 - 2006 Routine GUM clinic returns Males Females
    22. 22. Rates of diagnoses of uncomplicated genital chlamydial infection by sex and age group, GUM clinics, United Kingdom: 1997 - 2006 Routine GUM clinic returns Males Females
    23. 23. Chlamydia STIs <ul><li>We have known about Chlamydia causing STIs for many years but it is only in the last 10-15 years where we have seen it emerge as a major pathogen </li></ul><ul><li>Most common age groups: males 20-24 </li></ul><ul><li>females 16-19 </li></ul><ul><li>Government screening for Chlamydia in under 25’s announced in 2003 </li></ul>
    24. 24. National Chlamydia Screening Programme <ul><li>NCSP aimed to screen at least 15% of sexually active 16-24 year olds. </li></ul><ul><li>£70m invested – aim to reduce the burden of disease due to Chlamydia </li></ul><ul><li>Hospital labs have seen a dramatic increase in their Chlamydia testing workload </li></ul><ul><li>Is it working? </li></ul>
    25. 25. Chlamydial disease <ul><li>Also known as NSU or NGU </li></ul><ul><li>The infection has a longer incubation period than gonorrhoea, of 1 to 3 weeks compared to 2-3 days (usually) </li></ul><ul><li>As symptoms for gonorrhoea appear first this is why treatment for both infection is usually offered </li></ul><ul><li>Asymptomatic Chlamydial infection is common in both sexes – at least 50% in males and 70% in females </li></ul>
    26. 26. Symptoms (when present) <ul><li>Females: </li></ul><ul><li>unusual vaginal discharge </li></ul><ul><li>bleeding (intramenstrual) </li></ul><ul><li>pain on urination </li></ul><ul><li>lower abdominal pain </li></ul><ul><li>Males: </li></ul><ul><li>urethral discharge </li></ul><ul><li>burning and itching in genital area </li></ul><ul><li>pain on urination </li></ul>
    27. 27. Symptoms <ul><li>In some cases the symptoms subside after a few days </li></ul><ul><li>In either sex, complications may ensue in the case of untreated infection </li></ul><ul><li>In males, untreated infection may lead to epididymitis and Reiter’s Syndrome (arthritis) </li></ul><ul><li>In females, the consequences of untreated infection are pelvic inflammatory disease (PID) in 10 to 40% of cases </li></ul>
    28. 28. Symptoms <ul><li>In up to 20% of patients with PID, infertility develops and the risk of ectopic pregnancy increases </li></ul><ul><li>The risk of infertility also increases if there has been more than one episode of PID </li></ul><ul><li>Infection in pregnancy can lead to infection of the baby - trachoma inclusion conjunctivitis or pneumonia </li></ul>
    29. 29. Chlamydia lifecycle <ul><li>Chlamydia is an unusual bacterial genus – it is an obligate intracellular pathogen </li></ul><ul><li>Over time it has lost the capacity to replicate independently </li></ul><ul><li>How would this affect laboratory diagnosis? </li></ul>
    30. 30. Lifecycle <ul><li>During their lifecycle Chlamydia may be found in two forms – elementary bodies and reticulate bodies </li></ul><ul><li>the infective form of Chlamydia is the Elementary Body (EB), a dense, circular body, about 0.3 μ m in diameter. EBs are fairly inert and can survive outside the cell </li></ul>
    31. 31. Life cycle <ul><li>EBs carry glycosaminoglycan molecules on their surfaces that bind to receptors on the surface of certain cells </li></ul><ul><li>after attachment, the EB is taken into the cell by endocytosis and remains inside the endocytotic vacuole for the next phase of the life cycle </li></ul>
    32. 32. Life cycle <ul><li>the EB develops into a Reticulate Body (RB) which is larger (0.5 to 1.0 μ m) and metabolically active, although it uses host cell ATP-generating systems </li></ul><ul><li>inside the vacuole, the RB grows and replicates its DNA </li></ul><ul><li>during this phase, the contents of the vacuole are termed an “Inclusion Body” </li></ul>
    33. 33. Life cycle <ul><li>Staining of the Inclusion Body with iodine to demonstrate infection of cell cultures </li></ul>
    34. 34. Life cycle <ul><li>EB Formation and Release </li></ul><ul><li>after 18 to 24 hours, the RB reorganises into many EBs which are released on cell rupture (24 to 48 hours after infection) </li></ul>
    35. 35. Chlamydia trachomatis <ul><li>There are many different serotypes* and these can be grouped according to the type of disease that they cause – not all infections are STIs! </li></ul><ul><li>Serotypes A, B and C cause a serious eye infection that begins with conjunctivitis and may progress (particularly with repeated infection) to conjunctival scarring and blindness – trachoma </li></ul><ul><li>Serotypes D to K cause a less severe form of conjunctivitis that does not usually result in trachoma </li></ul>* Do you remember what a serotype is? – see PIP!
    36. 36. Trachoma <ul><li>Not an STI </li></ul><ul><li>very common in tropical countries and when sufferers don’t get treated for the initial infection </li></ul><ul><li>transmitted via hands etc. and via flies </li></ul>
    37. 37. C. trachomatis STIs <ul><li>The more common type of infection associated with serotypes D to K is sexually transmitted: </li></ul><ul><li>NGU (non-gonococcal urethritis) in males (also called NSU: non-specific urethritis) </li></ul><ul><li>urethritis, cervicitis, salpingitis in females </li></ul><ul><li>can lead to PID (pelvic inflammatory disease) and resulting infertility due to scarring of Fallopian tubes </li></ul><ul><li>also increased risk of ectopic pregnancy </li></ul>
    38. 38. Treatment <ul><li>Azithromycin (clamelle) is usually first choice – single dose is enough </li></ul><ul><li>Alternatively can use doxycycline (adults) or erythromycin (babies) - Treat for extended periods (1-3 weeks due to prolonged replication cycle) </li></ul>
    39. 39. Lymphogranuloma venereum <ul><li>C. trachomatis serotypes L1, L2 and L3 only cause LGV (lymhogranuloma venereum) </li></ul><ul><li>begins with a genital ulcer, infection spreads to inguinal lymph nodes which enlarge and break down, discharging pus </li></ul><ul><li>if untreated, can lead to enlargement & granulomatous hypertrophy of glands </li></ul>
    40. 40. LGV <ul><li>Was rare in developed nations before 2003 </li></ul><ul><li>386 cases in UK in 2007 (500% increase in 10 years) </li></ul><ul><li>Most often seen in men who are HIV positive </li></ul>
    41. 41. Diagnosis of Chlamydia infection <ul><li>You have two options – 1) Use highly trained professionals or 2) Have a go yourself at home </li></ul><ul><li>Which do you think is the most sensible? </li></ul>
    42. 42. Home Chlamydia testing <ul><li>While better than no testing at all there are concerns that some will not follow the procedure correctly – these tests need to be idiot proof! </li></ul><ul><li>Based upon an immunochromatography test on urine – positive = colour change </li></ul><ul><li>Such methods are not very sensitive so some positives will be missed! </li></ul>
    43. 43. Laboratory diagnosis <ul><li>Sample type may be a swab from the affected area (e.g. urethra) or urine is acceptable for some tests </li></ul><ul><li>Traditional laboratory methods include tissue culture assay, ELISA and immunofluorescence </li></ul><ul><li>These are now being replaced by molecular assays </li></ul>
    44. 44. Tissue culture <ul><li>Tissue Culture in cycloheximide-treated McCoy cells – detection of inclusion bodies by iodine staining or IF </li></ul><ul><li>Cumbersome method </li></ul>
    45. 45. Other traditional tests <ul><li>Direct immunofluorscence using a labelled monoclonal antibody specific to the major outer membrane protein (MOMP) </li></ul><ul><li>ELISA tests to detect Chlamydia antigen e.g. IDEIA are useful and can be automated </li></ul><ul><li>However, molecular tests are rapid, specific and sensitive… </li></ul>
    46. 46. Molecular methods <ul><li>A number of molecular methods based on amplification of Chlamydia nucleic acids have been introduced. </li></ul><ul><li>These include assays based on PCR, NASBA, TMA, Strand displacement amplification, LCR etc </li></ul><ul><li>Most common method in UK is BD ProbeTec SDA assay </li></ul><ul><li>www.chlamydiae.com/diagnostics_index.asp </li></ul>
    47. 47. Syphilis Treponema pallidum
    48. 48. Clinical and epidemiological aspects <ul><li>We have seen an 870% increase in cases of syphilis since 1996 </li></ul><ul><li>In 2007, 2680 cases were reported to HPA </li></ul><ul><li>Age groups: Males 25-44 years, Females 20-24 </li></ul><ul><li>There are “hotspots” of cases in the UK e.g. London, Manchester </li></ul><ul><li>Most often seen in males, especially men who have sex with men (MSM) </li></ul>
    49. 49. Numbers of diagnoses of syphilis (primary, secondary and early latent) by sex, GUM clinics, England and Wales, and Scotland*: 1931 - 2006 * Equivalent Scottish data are not available prior to 1945. N. Ireland data from 1931 to 2000 are incomplete and have been excluded. Routine GUM clinic returns
    50. 50. Number of diagnoses of infectious syphilis (primary and secondary) by sex and male sexual orientation, GUM clinics, United Kingdom: 1997 – 2006 Routine GUM clinic returns
    51. 51. Rates of diagnoses of infectious syphilis (primary & secondary) by sex and age group, GUM clinics, United Kingdom: 1997 - 2006 Routine GUM clinic returns Males Females
    52. 52. Stages of disease <ul><li>There are four main stages of disease – progressively more destructive. </li></ul><ul><li>Treatment can prevent development of the next stage - </li></ul><ul><li>1. Primary </li></ul><ul><li>2. Secondary </li></ul><ul><li>3. Latent </li></ul><ul><li>4. Tertiary </li></ul>
    53. 53. Clinical aspects <ul><li>Caused by the spirochaete bacterium, Treponema pallidum ssp pallidum </li></ul><ul><li>Highly infectious </li></ul><ul><li>Starts with the development of one or more ulcers at the point of entry of the organism – CHANCRE </li></ul><ul><li>A chancre is the lesion of primary syphilis </li></ul><ul><li>Typically painless and will disappear even without treatment </li></ul>
    54. 54. Primary syphilis <ul><li>30% who come into contact with syphilis during sex will be infected </li></ul><ul><li>Only 40% show symptoms of classical appearance </li></ul><ul><li>90 day incubation period </li></ul><ul><li>Lesion will disappear within three weeks even without treatment. </li></ul><ul><li>Can be missed/dismissed by patient </li></ul>
    55. 55. Secondary syphilis <ul><li>Usually appears around 6 weeks after chancre disappears. </li></ul><ul><li>Can be up to 2 years before signs show </li></ul><ul><li>Multiple system involvement </li></ul><ul><li>Mucosal and skin involvement most common </li></ul><ul><li>Symptoms will resolve in most cases. </li></ul><ul><li>The most infectious stage of syphilis </li></ul>
    56. 56. Syphilis “the great mimic” <ul><li>The symptoms seen in patients with syphilis are highly variable and often similar to those seen in other diseases – “the great mimic” </li></ul><ul><li>Makes diagnosis without laboratory testing very difficult </li></ul><ul><li>Sir William Osler “the physician who knows syphilis knows medicine…&quot; </li></ul>
    57. 57. Latent syphilis <ul><li>Two stages : </li></ul><ul><li>– Early latent- up to 2 years </li></ul><ul><li>Patient still infectious </li></ul><ul><li>– Late latent- after 2 years </li></ul><ul><li>Patient no longer sexually infectious although can still pass infection vertically </li></ul>
    58. 58. Tertiary syphilis <ul><li>3 - 20 years after primary infection </li></ul><ul><li>Benign gummatous phase - Characterized by slow growing granulomatous lesions </li></ul><ul><li>Infiltrative or destructive </li></ul><ul><li>Can affect any organ </li></ul>
    59. 59. Tertiary syphilis <ul><li>May also see cardiovascular complications e.g. aortic aneurysm </li></ul><ul><li>Tertiary syphilis is often associated with dementia – CNS involvement may also present as general paralysis of the insane, demyelination of the spinal cord resulting in pains, loss of feeling and difficulty walking. Changes in the joint - so-called Charcot's joints may develop owing to loss of nerve supply </li></ul>
    60. 60. Congenital syphilis <ul><li>If infection is acquired in pregnancy, usually miscarriage or still-birth ensues. However, if the foetus survives, it may show signs of congenital syphilis: the Hutchinson’s Triad – Hutchinson’s teeth (pointed), deafness & keratitis </li></ul><ul><li>There is a statutory requirement to screen all pregnant women for evidence of syphilis – antibody test (see later) </li></ul>
    61. 61. Treatment <ul><li>Syphilis is a potentially devastating disease that is easy to treat, but it is essential that it is caught in the early stages. </li></ul><ul><li>Benzathine penicillin is usually used. A single dose is sufficient to cure primary syphilis, although longer treatments are required for later stages, including the treatment of late latent syphilis. No penicillin resistance has been observed </li></ul>
    62. 62. Treponema pallidum ssp pallidum <ul><li>Treponema pallidum ssp pallidum is a very long, slender bacterium, which is about 0.1μm in diameter and 22μm in length </li></ul><ul><li>Since the maximum resolution of a bright-field microscope is 0.2μm, the organism cannot be seen by conventional microscopy </li></ul><ul><li>Cannot Gram stain this organism </li></ul>
    63. 63. Treponema pallidum ssp pallidum <ul><li>Can we culture this organism using artificial media? </li></ul><ul><li>NO! </li></ul><ul><li>The organism has undergone reductive evolution so it has lost many of the metabolic processes required for independent growth </li></ul><ul><li>This rules out using culture and identification as a diagnostic tool </li></ul>
    64. 64. Other subspecies <ul><li>There are three other subspecies of T. pallidum – these cause the non-venereal infections yaws, pinta and bejel </li></ul><ul><li>These are found in the Caribbean and W. Africa – they are now very rare </li></ul><ul><li>However, we need to bear these in mind as the antibody response to syphilis is identical to these 3 infections </li></ul><ul><li>Potential for misdiagnosis when interpreting serology results! </li></ul>
    65. 65. Laboratory diagnosis <ul><li>Diagnosis is usually confirmed using both clinical evidence and laboratory test results </li></ul><ul><li>Can we see syphilis down the microscope? </li></ul><ul><li>YES – using dark ground microscopy </li></ul><ul><li>What are the disadvantages of this test? </li></ul>
    66. 66. Dark ground microscopy <ul><li>Usually done in Genitourinary medicine clinics </li></ul><ul><li>Take fluid from an abraded ulcer – view sample against a dark background </li></ul><ul><li>Treponema is apparent by virtue of refractivity </li></ul><ul><li>Also often see characteristic corkscrew motility </li></ul>
    67. 67. Serology <ul><li>Can detect an antibody response to infection using serology </li></ul><ul><li>A major disadvantage of serology is the immune system takes a while to produce antibodies – so early infection will be missed </li></ul><ul><li>There are a number of serological tests for syphilis – BUT no one method is 100% reliable </li></ul><ul><li>This makes the interpretation of serological tests a bit tricky (but I will explain…) </li></ul>
    68. 68. Serological tests <ul><li>Serological tests for syphilis can be divided into two general types </li></ul><ul><li>Non-specific tests – these rely on the fact that syphilis antibodies also bind (cross-react) to cardiolipin (found in ox heart) e.g. VDRL and RPR tests </li></ul><ul><li>Specific tests – e.g. TPPA (TPHA), ELISA and FTA (abs) </li></ul><ul><li>If positive with one method must confirm with a second method </li></ul>
    69. 69. Venereal Disease Reference Laboratory test <ul><li>Mix patient sera with antigen (cardiolipin) on a slide for 8 mins </li></ul><ul><li>Examine for agglutination (positive test) </li></ul><ul><li>Quantitative test – if positive test a dilution series of sera to obtain the highest dilution which is positive – antibody titre </li></ul>Positive Negative
    70. 70. VDRL <ul><li>VDRL becomes positive 1-2 weeks after chancre appearance (73%) and reaches high titres in secondary syphilis (100%) </li></ul><ul><li>BUT becomes negative in latent syphilis and also following treatment </li></ul><ul><li>Therefore this test is very important in monitoring the effect of treatment and stage/activity of disease </li></ul><ul><li>False positives are a problem (e.g. recent vaccination, connective tissue disease) </li></ul>
    71. 71. Treponema pallidum Particle Agglutination test (TPPA) <ul><li>Specific test for syphilis antibodies </li></ul><ul><li>Patient sera diluted in a microtitre plate </li></ul><ul><li>Gelatin particles – control particles </li></ul><ul><li>Gelatin particles coated with Treponema antigen – test particles </li></ul><ul><li>Added to different wells </li></ul><ul><li>Incubate </li></ul><ul><li>Observe for agglutination – indicates serum antibodies reacting with antigen on particles </li></ul><ul><li>If negative the particles will sink to the bottom of the well </li></ul>
    72. 72. TPPA
    73. 73. TPPA <ul><li>Becomes positive in primary syphilis (71%) and 100% positive in secondary </li></ul><ul><li>Remains positive for life – even if treated </li></ul><ul><li>Test used to be performed using sheep erythrocytes not gelatin particles – TPHA </li></ul><ul><li>TPPA is a cumbersome test to perform so used as a confirmatory test </li></ul><ul><li>For screening patients (e.g. in pregnancy) we use an ELISA test - automated </li></ul>
    74. 74. ELISA <ul><li>Some ELISA kits detect IgG only (OK), others detect IgM & IgG (best – as helps determine stage of disease) </li></ul><ul><li>Positive in 82% of cases of primary syphilis and 100% of secondary </li></ul><ul><li>Remains positive despite treatment (IgG) </li></ul><ul><li>If positive confirm usually with TPPA test </li></ul>
    75. 75. FTA (abs) <ul><li>Indirect immunofluorescence test </li></ul><ul><li>“ Gold standard” test – positive in 86% of primary syphilis and 100% of secondary </li></ul><ul><li>However it is a cumbersome and difficult test to do – so it is only performed in reference laboratories </li></ul>
    76. 76. Laboratory diagnosis of syphilis <ul><li>If patient presents with an ulcer – perform dark ground microscopy – if positive begin treatment and monitor by serology </li></ul><ul><li>If no ulcer or microscopy is negative we must rely on serology… </li></ul><ul><li>ELISA is used as a screening test as it is cheap, automated and rapid </li></ul><ul><li>If positive perform a TPPA to confirm a true positive. </li></ul>
    77. 77. Laboratory diagnosis of syphilis <ul><li>If positive by ELISA and TPPA begin treatment and perform VDRL test to ensure patient is clear from infection </li></ul><ul><li>What laboratory results would you see in a case of secondary syphilis? </li></ul><ul><li>Or latent syphilis? </li></ul>