Spirochetes are thin, spiral-shaped, Gram-negative bacteria characterized by endoflagella within their periplasmic space. The endoflagella allow for various motility types and are responsible for pathogenesis. The major pathogenic genera are Borrelia, Treponema, and Leptospira. Treponema pallidum causes syphilis while Borrelia burgdorferi causes Lyme disease. Leptospira interrogans causes leptospirosis, a zoonotic disease transmitted via contact with infected animal urine. Laboratory diagnosis involves microscopy, culture, and serological detection of antibodies.

1. Spirochetes
Aashutosh Nama
M.Sc Microbiology Sem -1
Dr. B Lal Institute of Biotechnology

2. • 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.

3. Endoflagella
• Endoflagella do not protrude outside, but
present in the periplasmic space between
peptidoglycan layer and outer membrane.

4. 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

5. Pathogenes
• Most of the spirochetes are saprophytes.
Only there genus of them are major humans
pathogenes –
1. Borrelia
2. Treponema
3. Leptospira

7. Morphological differences between
Treponema, Borrelia and Leptospira

8. Spirochetes - Classification

9. TREPONEMA

10. 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

11. 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

12. 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

13. LABORATORY DIAGNOSIS OF
SYPHILIS
• Direct Microscopy
• Dark Ground Microscopy (DGM)
• Direct Fluorescent Antibody Staining
• Silver Impregnation Staining

14. 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)

15. VDRL v/s RPR
VDRL RPR
RPR
Blood, plasma, serum, and
CSF can be tested
Blood, plasma and serum
can be tested but not CSF
Rotation of slide is done for
4 mins
Rotation of card is done for
8 mins
Sensitivity in primary
syphilis is 78%
Sensitivity in primary
syphilis is 86%
It is cheaper; one vial of
VDRL antigen can be used
for 250 tests. It is preferred
for field studies and for
antenatal screening
RPR is expensive than
VDRL. It is preferred when
sample load is less.

16. •Borrelia

17. 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

18. Medically important borreliae
• B. recurrentis – relapsing fever
• B. vicentti – Vincent’s angina
• B. burgdoferi – Lyme disease

19. Relapsing fever
• Relapsing fever is characterized by recurrent
episodes of fever and nonspecific symptoms
following exposure to insect vector carrying
Borrelia species.

20. B. Burgdoferi
B. vincentii

21. 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.

22. 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).

23. 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).

24. 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).

25. leptospirosis

26. • 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

27. Classification
• Leptospria is antigenically complex; compries
two species: a.)L.interrogans
b.) L. biflexa

28. 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

29. 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.

30. 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.

31. 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.

32. 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.

33. 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