Spirochetes

Aashutosh Nama
M.Sc Microbiology Sem -1
Dr. B Lal Institute of Biotechnology
 • Background
• Speira = coil Chaite = hair
• Gram negative bacteria (differs from other
gram negative bacteria due to presence of
“endoflagella”).
• Thin, flexible, elongated spirally coiled helical
bacilli.
• Tightly coiled bacteria typically slender and
flexuous shpe.
 Endoflagella
• Endoflagella do not protrude outside, but
present in the periplasmic space between
peptidoglycan layer and outer membrane.
 Motility of spirochetes
• Endoflagella are responsible for various
motility of spirochetes such as:

1. Flexion – extension type

2. Corkscrew type rotatory

movement

3. Translatory type motility

 Pathogenes

• Most of the spirochetes are saprophytes.

Only there genus of them are major humans
pathogenes –
1. Borrelia
2. Treponema
3. Leptospira
Morphological differences between
Treponema, Borrelia and Leptospira
Spirochetes - Classification
TREPONEMA
 TREPONEMA PALLIDUM
• ™Morphology: extremely thin and delicate
with tapering ends
1. ™Size: 6–14 μm × 0.2 μm
2. ™Spirals: 6–12 spirals at intervals of 1 μm
3. ™Motility: flexion extension, translatory and
corkscrew motility
4. ™Endoflagella: About 3–4 flagella - motility
& highly antigenic
 PATHOGENESIS OF SYPHILIS
• Mode of transmission:
- Venereal
- Non-venereal - direct contact, blood
transfusion or transplacental
• ™Spread: T. pallidum penetrates through
mucosa or abraded skin  Enter lymphatics
and blood  systemic  primary lesion
• ™Incubation period: Variable (9–90 days)
Inversely proportional to the number of
organisms inoculated
 Microscopy & Culture
• Dark ground or phase contrast microscope
• ™Staining: Do not take up Gram stain
- Fluorescence staining
- Sliver impregnation methods
• ™Cultivation: Pathogenic treponemes cannot be
grown in artificial culture media. Maintained in
rabbit testes. E.g, „Nichols strain
• Non-pathogenic Treponemes – grow in Smith
Noguchi medium under strict anaerobic
conditions. E.g: Reiter’s strain & Noguchi strain
 LABORATORY DIAGNOSIS OF
SYPHILIS
• Direct Microscopy
• Dark Ground Microscopy (DGM)
• Direct Fluorescent Antibody Staining
• Silver Impregnation Staining
 Serology (Antibody Detection)
Cardiolipin antigens test
• VDRL test – Venereal Diseases Reference
Laboratory
• RPR test – Rapid Plasma Reagin Treponemal
antigen test
• TPHA test (Treponema pallidum
haemagglutination test)
 VDRL v/s RPR
RPR
Blood,
VDRL
plasma, serum, and Blood,
RPR
plasma and serum
CSF can be tested can be tested but not CSF
Rotation of slide is done for Rotation of card is done for
4 mins 8 mins

Sensitivity in primary Sensitivity in primary

syphilis is 78%
It is cheaper; one vial of
VDRL antigen can be used
for 250 tests. It is preferred
for field studies and for
antenatal screening
syphilis is 86%
RPR is expensive than
VDRL. It is preferred when
sample load is less.
• Borrelia
 Microscopy
• Larger than other spirochaetes
• 10-30 µm x 0.2-0.5 µm
• Irregular, wide coils (5-8)
• Motile, Gram negative, pointed ends
• Commensal in mouth or genitals
  Medically important borreliae
• B. recurrentis – relapsing fever
• B. vicentti – Vincent’s angina
• B. burgdoferi – Lyme disease
 Relapsing fever
• Relapsing fever is characterized by recurrent
episodes of fever and nonspecific symptoms
following exposure to insect vector carrying
Borrelia species.
B. Burgdoferi
B. vincentii
 LABORATORY DIAGNOSIS OF
SYPHILIS
• Serology :– Done for detection of antibodies
• ELISA and IFA( indirect fluorescence assay)
are available to detect serum antibodies.
• Molecular methods :- by RT- PCR (has been
developed targeting 16s rRNA and GlpQ genes
to identify the various species of borrelia.
 Lyme disease
• It is caused by Borrelia burgdoferi.
• it is transmitted by tick bite (ixodes tick)
• Lyme disease occur in 3 stages:-
1. Early localized infection:-
An annular maculopapular
lesion developed at the site of
the tick bite called “erythema
migrans”(after incubation
period 3-32 days).
2. Early disseminated infection :- B. burgdorferi
spreads hematogenously to many site resulting
in secondary annular skin lesions, arthralgia ,
malaise and neurological abnormalities.

3. Late persistent infection :- after the developed

arthritis of large joints(e.g. knees).
 Laboratory diganosis
• Culture :- B.burgdorferi can be grown on BSK
media (barbour- stoener-kelly ) through the
CSF, blood, skin lesions.
• Molecular methods :- it detects B.burgdorferi
flaB, ospA gene(outer surface of lipoprotein)
and 16s rRNA.
• Serology (by antibody detection) :- it is done by
ELISA method (if found positive, it has to be
western blot).
leptospirosis
• Lepto – fine or thin + spira – coil
• obligate aerobe spirochete
• Actively motile, delicate, closely wound coils,
Characteristic hooked ends
• Don’t stain readily, can be seen under
DGM(dark ground microscope)
• Zoonotic disease – leptospirosis
• Largely secreted in urine
• Survive many weeks in soil and water
 Classification
• Leptospria is antigenically complex; compries
two species: a.)L.interrogans
b.) L. biflexa
 Transmission
• Leptospirosis is zoonotic direct human to
human transmission does not occur.it is
transmitted by:
1. Direct or indirect contact with urine of
infected animal
2. Enter damaged skin which has immersed for
a long time in water or mud contaminated
with infected urine
 Laboratory diagnosis
• Microscopy
• Wet films: they may be observed under dark
ground microscope or phase contrast
microscope.
• Staining: they do not stain by ordinary stain,
but can be easily stained by sliver
impregnation stains.
• L. interrogans is 6-12 μm long; tightly and
regularly coiled, with hooked ends like
umbrella handle.
 Culture
• Culture condition : Leptospria is obligate aerobe
and slow growing. culture should be incubated at
30⁰C for 4-6 weeks.
• Culture media : requires enriched media such as –
1. EMJH liquid (ellinghausen, Mccullough,
johnson, harris).
2. Korthof’s media
3. Fletcher’s – semisolid medium.
 Test
1. Serology test – it detect the IgM (appears in 3
or 4 weeks) and IgG (appears later than IgM
antibody).
• Elisa: it detects IgM and IgG separately.
• Immunochromatographic test (ICT): : it
detects IgM and IgG antibody separately.
2. Antimicrobial sensitivity test- high doses of
penicilin will be effective against L.interrogans.
3. pcr method – it detects the lipL32 genes of
L.interrogans.
 Pathogenesis
• Two distinct phase
1. Septicemic phase – after entering thorough
mucosa or abraded skin L.interrognas spill
the bloodsteram and then attack on organ like
brain, liver, lung, heart and kidney.
2. Immune phase – as antibodies developed ,
antigen antibody complexes deposited in
various organ.
Real colonization – bacilli becomes adherent
to the proximal renal tubular brush border
and are excreted in urine.
 References
• Books
1. Apurba S Sastry, Sandhya Bhat, 2020, Essentials of Medical Microbiology ;
Jaypee Brothers Medical Publishers; Third edition.
2. Microbiology: An Introduction by Gerard J. Tortora, Berdell R. Funke , Christine
L. Case
3. Prescott L.M., Harley J.P. and Klein D.A., 2007, Microbiology 7th Edition, Mc
Grow Hill.

• Internet
1. Phylogenetic foundation of spirochetes B J Paster 1, F E Dewhirst
https://pubmed.ncbi.nlm.nih.gov/11075904/
2. Recent discoveries and advancements in research on the Lyme disease
spirochete Borrelia burgdorferi
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6545822/
3. Spirochete Flagella and Motility by Shuichi Nakamura
https://www.mdpi.com/2218-273X/10/4/550
4. https://microbeonline.com/spirochetes-morphology-classification-disease/
5. https://www.slideshare.net/AliaNajiha1/f-43-spirochaetes
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