SPIROCHETES & OTHER SPIRAL

MICROORGANISMS
MICROBIOLOGY LECTURE SERIES

LUZ GREGORIA LAZO-VELASCO, MD

SPIROCHETES
Spiral, motile bacteria
Family Spirochaetaceae Borrelia, Treponema
Family Leptospiraceae Leptospira, Leptonema,Turneriella

Many structural characteristics in common:

long, slender, helically coiled, spiral or corkscrew-shaped bacilli

Treponema pallidum has an outer sheath or glycosaminoglycan coating, outer

membrane (contains peptidoglycan; maintains structural integrity of the organism)
Endoflagella (axial filaments)- flagella-like organelles in the periplasmic space encased
by the outer membrane; begin at each end of the organism & wind around it,
extending to and overlapping at the midpoint

SPIROCHETES
Many structural characteristics in common:

inside the endoflagella- inner membrane (cytoplasmic membrane) that provides

osmotic stability & covers the protoplasmic cylinder
series of cytoplasmic tubules (body fibrils) near the inner membrane
Treponemes reproduce by transverse fission

TREPONEMA

T pallidum subspecies pallidum (syphilis)

T pallidum subspecies pertenue (yaws)
T pallidum subspecies endemicum (endemic syphilis, bejel)

T pallidum subspecies carateum (pinta)

TREPONEMA PALLIDUM
Slender spirals, 0.2 mm (W) x 5-15 mm (L)
Spiral coils regularly spaced at a distance of 1 mm from one another
Actively motile
Not readily seen unless immunofluorescent stain or dark-field illumination is used
Do not stain well with aniline dyes, can be seen in tissues when stained with silver

impregnation method
Not grown on bacteriologic media or in cell culture

Grows very slowly- antibiotics must be present at an effective level for several weeks

to kill the organism and cure the disease

TREPONEMA PALLIDUM
Microaerophilic, survives best at 3-5% oxygen

Nonpathogenic- Reiter strain (saprophytes, can be cultured anaerobically; grows on defined

medium of 11 AA, vitamins, salts, mierals, serum albumin)

May remain motile for 3-6 days at 25oC in proper suspending fluids & in the presence of

reducing substances

Remain viable for at least 24hrs in whole blood or plasma stored at 4oC
Killed rapidly by drying and elevation of temperature to 42oC
Rapidly immobilized & killed by trivalent arsenical, mercury and bismuth
Penicillin- treponemicidal in minute concentrations
Estimated division time: 30hours

TREPONEMA PALLIDUM
Antigenic structure:
Outer membrane surrounds the periplasmic space & the peptidoglycan-

cytoplasmic membrane complex

Membrane proteins, lipids, endoflagella
T pallidum sp pallidum- (+) hyaluronidase
Cardiolipin
Humans with syphilis develop antibodies capable of staining T pallidum by indirect

immunofluorescence
Reagin (distinct antibody-like substance which gives positive complement fixation

& flocculation test results with aqueous suspensions of cardiolipin extracted from
normal mammalian tissues)

TREPONEMA PALLIDUM
Pathogenesis, Pathology & Clinical Findings:

A. Acquired Syphilis
Transmitted by sexual contact; infectious lesion is on the skin or mucous

membranes of genitalia in 10-20% of cases, primary lesion- intrarectal, perianal

or oral
PRIMARY SYPHILIS: Spirochetes multiply locally at the site of entry, some

spread to nearby LN bloodstream

Papule (2-10 weeks after infection) hard chancre (ulcer with clean, hard
base; predominance of lymphocytes and plasma cells

Heals spontaneously

TREPONEMA PALLIDUM
Pathogenesis, Pathology & Clinical Findings:

A. Acquired Syphilis
TERTIARY SYPHILIS: development of granulomatous lesions (gummas) in

the skin, bones & liver, degenerative changes in the CNS (meningovascular
syphilis, paresis, tabes), cardiovascular lesions (aortitis, aortic aneurysm, aortic
valve insufficiency)

treponemes very rare

exaggerated immune response attributed to hypersensitivity to the organism

TREPONEMA PALLIDUM
Pathogenesis, Pathology & Clinical Findings:

B. Congenital Syphilis
a pregnant woman with syphilis can transmit T pallidum to the fetus through the
placenta beginning in the 10th-15th weeks of gestation
fetal death, miscarriage, stillborn at term

congenital syphilis in childhood: interstitial keratitis

Hutchinsons teeth
saddle nose

periostitis
CNS anomalies

TREPONEMA PALLIDUM
Diagnostic Laboratory Tests:

Specimens: tissue fluid expressed from early surface lesions (demonstration of spirochetes

by either dark-field microscopy or immunofluorescence; nucleic acid amplification); blood

(serologic tests), CSF (Venereal Disease Research Laboratory testing-VDRL)
Dark-Field examination

Immunofluorescence fluorescein-labelled antitreponeme antibody

Nucleic acid amplification
Serologic Tests for Syphilis

TREPONEMA PALLIDUM
Diagnostic Laboratory Tests:
Serologic Tests for Syphilis

1. Nontreponemal tests - screening; not very sensitive to early syphilis; antigens used:
cardiolipin, cholesterol, purified lecithin to react with syphilitic reagin antibodies;
VDRL, unheated serum reagin (USR) detect flocculation by microscopic
examination; rapid plasma regain (RPR) test, toluidine red unheated serum
test (TRUST)
2. Treponemal antibody tests measures antibodies against T pallidum antigens

T pallidum particle agglutination (TP-PA) gelatin particles

T pallidum hemagglutination (TPHA) sheep erythrocytes
Microhemagglutination T pallidum (MHA-TP)

TREPONEMA PALLIDUM
Diagnostic Laboratory Tests:
Serologic Tests for Syphilis

2. Treponemal antibody tests measures antibodies against T pallidum antigens

Fluorescent treponemal antibody absorbed (FTA-ABS) uses indirect
immunofluorescence to detect reactive antibodies

Treatment
Penicillin drug of choice

Tetracycline, erythromycin

DISEASES RELATED TO SYPHILIS

All give (+) treponemal and nontreponemal serologic test results for syphilis
BEJEL: T pallidum subsp endemicum (Africa, Middle East, Southwest Asia)

highly infectious skin lesions

tx: Penicillin
YAWS: T pallidum subsp pertenue

endemic in many humid, hot tropical countries

1o lesion: ulcerating papule on the arms or legs

transmission by person-to-person contact in children <15yrs old
scar formation, bone destruction common

DISEASES RELATED TO SYPHILIS

PINTA: T pallidum subsp carateum; endemic in Mexico, Central & South America, Philippines,

some areas of the Pacific

restricted to dark-skinned individuals
1o lesion: non-ulcerating papule on exposed areas

later: flat, hyperpigmented lesions appear on skin, depigmentation & hyperkeratosis

years afterward
transmitted by nonsexual means- direct contact or through flies or gnats

BORRELIA RECURRENTIS
Epidemic Relapsing fever transmitted by the human body louse
Endemic Relapsing fever caused by borreliae transmitted by ticks of the genus Ornithodoros

Form irregular spirals 10-30 mm long, 0.3 mm wide; distance between turns vary from 2-4

mm

Highly flexible, move both by rotation and twisting

Stain readily with bacteriologic dyes as well as with blood stains (Giemsa, Wright stain)

Culture: fluid media containing blood, serum or tissue

Antigenic Structure: antibodies develop in high titer after infection

BORRELIA RECURRENTIS
Pathogenesis & Clinical Findings:
Incubation period: 3-10 days

Sudden onset of chills, fever (spirochetes abound in the blood)

Fever persists for 3-5 days, declines (4-10 days), followed by second attack of chills, fever,

intense headache, malaise

3-10 recurrences, diminishing in severity

Antibodies against spirochetes appear during the febrile stage

BORRELIA RECURRENTIS
Diagnostic Laboratory Tests:
Specimens: blood during the rise in fever for smears and animal innoculation
Smears: thin or thick blood smears stained with Wright or Giemsa stain reveal large,

loosely coiled spirochetes among the red cells

Animal inoculation: white mice or young rats inoculated intraperitoneally with blood.

Stained films of tail blood are examined for spirochetes 2-4 days later
Serology: spirochetes grown in culture can serve as antigens for CF tests

BORRELIA RECURRENTIS
Immunity: short duration
Treatment:

tetracyclines, erythromycin, penicillin

Prevention:

avoidance of exposure to ticks and lice

delousing (cleanliness, insecticide)

BORRELIA BORGDORFERI & LYME DISEASE

LYME DISEASE: Named after the town OF Lyme, Connecticut where clusters of cases

in children were identified

Transmitted to humans by the bite of a small Ixodes tick
Erythema migrans characteristic skin lesion with flulike symptoms (early)

Late manifestations arthralgia, arthritis

BORRELIA BORGDORFERI
Spiral organisms, 20-30mm long and 0.2-0.3 mm wide
Distance between turns varies from 2-4 mm
Variable numbers of endoflagella (7-11); highly motile

Stains readily with acid and aniline dyes and by silver impregnation techniques
Culture & Growth Characteristics:

Barbour-Stoenner-Kelly medium (BSK II) complex liquid medium

Rifampin, fosfomycin (Phosphonomycin), amphotericin B can be added to BSK II
to reduce

LEPTOSPIRA & LEPTOSPIROSIS

Leptospira interrogans pathogenic (>200 serovars)

Leptospira biflexa free-living, non-pathogenic (>60 serovars)

Tightly coiled, thin, flexible spirochetes 5-15 mm long, with very fine spirals 0.1-0.2

mm wide; one end often bent, forming a hook

Actively motile
Best seen using a dark-field microscope

EM show thin axial filament and delicate membrane

Does not stain readily, can be impregnated with silver

LEPTOSPIRA & LEPTOSPIROSIS

Culture: grow best under aerobic conditions at 28-30oC in semisolid medium

(Ellinghausen-McCullough-Johnson-Harris EMJH) in 10mL test tubes with 0.1%

agar and 5-fluorouracil
after 1-2 weeks, produce a diffuse zone of growth near the top of the tube and
later a ring of growth at a level in the tube corresponding to the level of the
optimal oxygen tension for the organisms
Growth Requirements: derive energy from oxidation of long-chain FA and

cannot use amino acids or carbohydrates as major energy sources

ammonium salts main source of nitrogen

can survive for weeks in water

LEPTOSPIRA & LEPTOSPIROSIS

Antigenic Structure:

outer envelope containing large amounts of lipopolysaccharide

Pathogenesis and Clinical Findings:

human infection comes from leptospires, often in bodies of water, entering the
body through breaks in the skin (cuts and abrasions) and mucous
membranes (mouth, nose, conjunctivae)

incubation period: 1-2 weeks

fever (spirochetes in the bloodstream) hemorrhage, necrosis of tissue
dysfunction of organs (liver, kidneys)- jaundice, haemorrhage, nitrogen
retention

LEPTOSPIRA & LEPTOSPIROSIS

Pathogenesis and Clinical Findings:

biphasic illness- after initial improvement, second phase develops when IgM
antibody titer rises
second phase aseptic meningitis (intense headache, stiff neck, pleocytosis of
the CSF); nephritis; hepatitis, skin, muscle, eye lesions
Diagnostic Laboratory Tests:

Specimens: blood, CSF, tissue, urine

Microscopic Examination: Dark-field examination, thick smears stained by
Giemsa technique, Fluorescein- conjugated antibodies or other immunohistochemical techniques

LEPTOSPIRA & LEPTOSPIROSIS

Diagnostic Laboratory Tests:

Culture: whole fresh blood or urine can be cultured on semisolid medium

Serology: the diagnosis of leptospirosis in most cases is confirmed
serologically
agglutinating antibodies appear 5-7 days after infection, develop slowly, peak at
5-8 weeks; very high titers may be attained (>1:10,000)
Treatment:

Oral doxycycline, ampicillin or amoxicillin (mild)

IV Penicillin, ampicillin