Treponema

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Study OF BACTERIAL PATHOGENS

• SPIROCHETES

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TREPONEMA

• Trepos turn, nema thread.
• Treponemes are relatively short slender
  spirochetes with fine spirals and pointed or
  rounded ends.
• Few are pathogenic, while others occur as
  commensals in mouth, intestine and genetalia.
• Treponemes cause the following disease
• Veneral Syphillis T. pallidum
• Endemic Syphillis T. pallidum
• Yaws T.pertenue
• Pinta T.careteum

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TREPONEMA PALLIDUM

• It is the causative agent of syphilis, was
  discovered by Schaudinn and Hoffman (1905) in
  inguinal lymph nodes of Syphilitic patients.
• The name pallidum refers to its pale staining.

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T. pallidum
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Morphology

• Thin delicate spirochetes with tapering ends,
  about 10 µ long and 0.1 to 0.2 µ wide.
• It has 10 regular spirals, sharp and angular, at
  regular intervals.
• Actively motile, exhibiting rotation around long
  axis, backward and forward.
• Cannot be seen under light microscope in wet
  films but can be made out by negative staining
  with Indian ink.
• Its morphology and motility can be seen under
  dark ground and phase contrast microscopy.

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• It does not take ordinary bacterial strains but
  stains light rose red with prolonged giemsa
  stain.
• Can be stained by silver impregnation methods.
• Fontanas method is used for staining films and
  levadities for tissue section.
• Ultra structurally, cytoplasm of T. pallidum is
  surrounded by trilamminar cytoplasmic membrane,
  enclosed by cell wall containing peptidoglycan.
• It is morphologically indistinguishable from T.
  pertenue commensal spirochetes of mouth
  genitalia like T. microdentium T. mucosum

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T. pallidum under dark field microscopy
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T. pallidum stained by Giemsa stain
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CULTIVATION

• Pathogenic treponemes do not grow in artificial
  media, however can be maintained in motile
  virulent form for 10-12 days in complex media
  under anaerobic conditions.
• Virulent T. pallidum can be maintained by serial
  passage in rabbit testes.

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RESISTANCE

• Very delicate, being readily inactivated by
  drying or heat (41-42C for 1 hour).
• Fomites are hence of little importance.
• Susceptibility of T. pallidum to heat was basis
  of fever therapy in syphilis.
• Killed in 1-3 days at 0-4C
• Stored frozen at -70C in 10 glycerol,or in
  liquid nitrogen it remains viable for 10-15
  years.
• Inactivated by contact with oxygen, D/W, soap,
  arsenicals, mercurial, bismuth, common antiseptic
  agents antibiotics.

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Antigenic structure

• Treponemal infection induces at least 3 types of
  antibodies
• 1st is antibody that reacts in std. or non
  specific serological tests for syphilis, such as
  Wassermann, Kahn VDRL in which a lipid hapten
  extracted from beef heart is used as the antigen.
• The hapten is known as Cardiolipin is
  chemically a diphosphatidyl glycerol.
• 2nd antigen is a protein antigen present in T.
  pallidum as well as in non-pathogenic treponemes
  such as the Reiter treponeme.

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• This appears to be a group antigen.
• Antibodies to the group antigen appear in
  syphilis are demonstrable by serological tests
  in which the Reiter strain is used as an antigen.
• This antibody is not specific for syphilis
  false positive reaction are common.
• 3rd antigen, probably polysaccharide is species
  specific.
• Antibody to this antigen is demonstrated by T.
  pallidum immobilization test which is positive
  only in sera of patients infected with pathogenic
  treponemes.

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Pathogenicity

• Natural infection with T. pallidum occurs only in
  man
• Experimentally monkeys may be infected.
• A disease resembling syphilis can be produced
  experimentally in chimpanzees, with typical
  lesions.
• Rabbits can be infected by intradermal
  intratesticular inoculation.
• Hamsters are also susceptible.

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Syphilis

• Origin is not definitely known.
• Venereal syphilis is acquired by sexual contact.
• Spirochete enters body through minute abrassions
  on skin or mucosa. Multiplies at the site of
  entry.
• Clinical disease sets in after an incubation
  period of about a month.
• Clinical manifestation fall into 3 stages
• Primary lesion is chancre formation.

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• In all but a few chancre is genital, other sites
  include mouth nipples or in uterine cervix.
• chancre is painless, relatively avascular,
  circumscribed, indurated, superficially ulcerated
  lesion.
• Chancre is covered with a thick, glary exudate,
  very rich in spirochete.
• Regional lymph nodes are swollen, discrete,
  rubbery nontender.
• Even before chancre appears, spirochete spread
  from site of entry into lymph blood stream, so
  that the patient may be infectious during late
  incubation period.
• Chancre invariably heals in 10 to 40 days, even
  without treatment, leaving a scar.

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• Secondary Syphilis sets in 2 - 6 months after
  primary lesion heals, during which period,
  patient is asymptomatic.
• Secondary Syphilis are due to widespread
  multiplication of Spirochetes and their
  dissemination through blood.
• Roseolar or papular skin rashes, mucous patches
  in oropharynx and condylomata at mucocutanious
  junction are characteristic lesions.
• Spirochetes are abundant in lesions and
  consequently patient is most highly infectious
  during secondary stage.
• Secondary lesions are very variable in
  distribution, intensity and duration, but they
  usually undergo spontaneous healing in some
  instances taking as long as 4 5 years.

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• After secondary lesions disappear, there is a
  period of quiescence known as Latent Syphilis.
• Diagnosis during this period is possible only by
  Serological tests.
• In many cases, this is followed by natural cure,
  but in others, after several years,
  manifestations of tertiary Syphilis appear.
• These consist of cardiovascular lesions including
  aneurysms chronic granulomata and meningovascular
  manifestations.
• Tertiary regions contains few Spirochetes and may
  represent manifestations of delayed
  hypersensitivity.
• In some symptoms developed, decades after the
  initial infection, known as quaternary Syphilis

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• In Syphilis acquired non venereally, natural
  evolution is as in venereal Syphilis, except that
  the primary chancre is extra genital usually on
  fingures.
• In congenital Syphilis, infection is transferred
  from Mother to offspring transplacentally, where
  the manifestations and the course are different.

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Chancre developed during veneral syphilis
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Laboratory Diagnosis

• It is important in assessing the cure after
  treatment.
• Laboratory diagnosis consists of demonstration of
  Spirochetes, under the microscope and of
  antibiotics in serum or CSF.
• Specimens should be collected with care as the
  lesions are highly infectious.
• The specimen is examined under dark field
  microscope.
• T.pallidum is identified by its slender spiral
  structure and slow movement.
• If examination cannot be done immediately, the
  exudate should be collected in capillary tubes,
  the end sealed and transported to the laboratory.
• A fluorescent antibody test on smears of exudate
  gives a higher positive rate.

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Fluorescent antibody test
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Serological Tests

• These are mainstay of laboratory diagnosis.
• They may be classified based on the antigens
  used
• Standard test for Syphilis (STS) Lipoidal or
  cardiolipid antigens used.
• Treponemel tests
• Using cultivable Treponemes as antigens.
• Pathogenic T.pallidum (Nichols Strain) as
  antigen.

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Standard Tests for Syphilis

• It was found that an alcoholic extract of Ox
  heart tissue was used as an antigen to which,
  Lecithin and cholesterol was added.
• This crude antigen was standardized by Pangborn
  (1942-45), who introduced cardiolipid, a purified
  extract of Beef heart.
• The standard tests usually employed are
  Wassermann, kahn venereal disease research
  laboratory (VDRL) tests.
• Wassermann reaction is a complement fixation
  test.
• It is as follows

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• Wassermann test is replaced due to it complexity
  by simpler tests as, Kahn VDRL.
• Kahn test is a tube flocculation test.
• It is as follows

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• VDRL has several advantages over Wassermann
  Kahn.
• Hence, it is the most widely used serological
  test for syphilis.

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Treponemal Tests

• A)
• The only test, in this group that is employed at
  present is Reiter protein complement fixation
  test (RPCF).
• Principle is the same, as the Wassermann Test,
  but the antigen is an extract of Reiter
  Treponeme.
• RPCF is less sensitive than the cardiolipid tests
  in early Syphilis, but is more sensitive in late
  Syphilis
• B)
• Tests using T.pallidum (Nichols Strain) may be
  classified in which the antigen used is live,
  killed or an extract of Spyrochetes.

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• i Tests using live T.pallidum
• T.pallidum immobilization (TPI) test employs live
  organism maintained anaerobically in a complex
  medium.
• The test serum is incubated aerobically with a
  suspension of Treponemes and complement.
• If antibodies are present, the Treponemes will be
  immobilized, when examined under dark field
  elimination.
• The test is considered reactive if more than 50
  percent of Treponemes are immobilized, doubtful
  if between 50 20 and non reactive if less than
  20.
• It is the most specific test available for
  Syphilis but is positive even for Yaws

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• ii Test using killed T.pallidum

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• iii Tests using T. pallidum extract
• T. pallidum haemagglutination (TPHA) employs an
  antigen tanned erythrocyte sensetized with an
  extract of T. pallidum obtained by freeze
  thawing.
• Cross reacting antibodies are absorbed from the
  sera before testing.
• In some cases of neurosyphilis, reagin antibodies
  may be absent in serum, but may be demonstrated
  in CSF.
• This is due to local production of antibodies in
  nervous system.

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Epidemiology

• Veneral syphilis is world wide in distribution.
• As originally described, it was very virulent
  disease with florid cutaneous manifestations.
• It was expected to be eradicated, on discovery of
  therapeutic response to penicillin, as the
  organism has no extra human reservoir.
• But there has occurred an increase in its
  incidence, due to changing customs, habits
  values in society.

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Immunity

• Immune response is not well understood.
• A patient is said to be refractory to reinfection
  so long as the original infection persists.
• Susceptiblity to reinfection is seen after 1st
  infection has been cured during the early stages.
• This is called as pingpong syphilis.
• Patients develop humoral antibodies, IgM, IgG,
  IgA, following infection,when the primary chancre
  appears.
• Antibodies increase in titre during the next
  several months.

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• However, even high titre of humoral antibodies do
  not influence natural progression of infection.
• Hence cell mediated immunity is considered as
  more relevant.
• During the early stages of syphilis there occurs
  inhibition of cell mediated immune response
  which is activated after secondary stage.

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Prophylaxis

• Transmission is by direct contact, which can be
  prevented by avoiding sexual contact with an
  infected person.
• Use of prophylactic penicillin carries the danger
  that it may supress primary lesion without
  eliminating the infection so that recognition
  treatment of disease may become more difficult.
• No vaccine is available.

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Treatment

• Chemotherapy has been of great interest.
• Mercurial, iodides, bismuth arsenicals have
  been replaced by penicillin.
• Penicillin is effective, but it is necessary to
  give an adequate dose maintain drug level
  sufficient long to establish cure.
• In patients, allergic to penicillin, erythromycin
  or tetracycline may be used.
• Chloramphenicol is not recommended.
• Penicillin treatment sometimes induces reaction,
  Jarisch-Herxheimer reaction, consisting of fever,
  malaise exacerbation of symptoms.
• It is frequent, but harmless during primary or
  secondary syphilis.

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• But is a little dangerous, in late syphilis.
• It is believed to be due to liberation of toxic
  products from massive destruction of treponemes
  or due to hypersensitivity.

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Non venereal treponematoses

• These diseases occur in endemic foci in several
  parts of the world, in communities with poor
  standard of hygiene.
• Infection is usually transmitted by direct body
  to body contact.
• It has been suggested that these represent the
  ancient patterns between man and treponemes
• When civilization and the general use of clothing
  limited the chance of bodily contact among
  people, the treponemes may have become adapted
  for transmission through sexual intercourse
  resulting in venereal Syphilis

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Endemic syphilis

• Syphilis, transmitted non venereally, was
  endemic.
• With recognition and with mass treatment with
  Penicillin under WHO auspices, endemic Syphilis
  has become very rare.
• It has also been reported from India.
• Diseases common in young children.

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ulcer developed during endemic syphilis
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• Primary chancre is not usually seen except some
  times on nipples of mothers infected by their
  children.
• Disease is usually seen with manifestations of
  secondary Syphilis, such as mucous patches and
  skin eruptions.
• Disease progresses to tertiary lesions
  particularly gummatous lesions.
• Laboratory diagnosis and treatment are as for
  venereal Syphilis.

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Yaws

• Also known as framboesia, pian, parangi, is
  endemic in tropical areas of Africa, Asia and
  America.
• The causative agent is T.pertenue, which is
  morphologically and antigenically
  indistinguishable from T.pallidum.
• The primary lesion (Mother Yaw) is an extra
  genital papule which enlarges and breaks down to
  form an ulcerating granuloma.
• Destructive gummatous lesion of the bones are
  common.

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• Infection is by direct contact.
• Flies may act as mechanical vectors. For e.g.
  Hippolates pallippes has been found feeding on
  open source but epidemiological importance being
  unknown.
• Laboratory diagnosis and treatment are as for
  Syphilis.
• There appears to be some cross immunity between
  Yaws and Syphilis, so the venereal Syphilis is
  rare in communities where Yaws is endemic.

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Pinta

• Pinta is endemic in central and South America and
  in the neighbouring islands.
• Primary lesion is an extra genital papule which
  does not ulcerate, but develops into a lichenoid
  or psoriaform patch.
• Secondary skin lesions are characterized by hyper
  or hypo pigmentation.
• The causative agent is T.carateum.
• It is very closely related to T.pallidum, but is
  not antigenically identical so that cross
  immunity between Pinta and Syphillis is only
  partial.

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Non pathogenic treponemes

• Several commensal treponemes occur on the genital
  mucosa and may cause confusion in the diagnosis
  of syphilis by dark field examination.
• These include T.microdentium and T.macrodentium
  found in the mouth.
• T.refringens was observed by Schautinn and
  Hoffman, when they 1st discovered T.pallidum.
• It was found in syphilitic as well as
  non-syphilitic genital lesions.
• T.calligyrum which also may be found in
  genitilia, a natural venereal infection in
  rabbits is caused by T.cuniculi morphologically
  identical with T.pallidum.

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THANKYOU

• Surabhi R. Meppadath
• B.Sc Microbiology