Chapter 46 Spirochetes

Order Spirochaetales
Treponema
Borrelia
Brachyspira
Spirochaeta
Leptospira

General characteristics:
Long
Slender
Helically curved
Gram-negative bacilli

Unusual morphologic features of axial fibrils

-Flagella-like organelles that wrap around the bacterias cell walls
-enclosed within the outer sheath
-facilitate motility of the organism
-attached within the cell wall by insertion disks (a platelike structure) that is
located near the ends of the cells.
-protoplasmic cylinder rotating the fibrils for bacterial movement
-cork screw-like winding
-Differentiation of genera within the family is based on the number of axial
fibrils, the number of insertion disks present, and biochemical and metabolic
features.

Treponema slender with tight coils

Borrelia thicker with fewer and looser coils
Leptospira thicker with fewer and looser coils, hooked ends
Brachyspira comma-shaped/helical with tapered ends with four flagella at
each end

TREPONEMA
-Major pathogens:
T. pallidum subsp. Pallidum
T. pallidum subsp. Pertenue
T. pallidum subsp. Endemicum
T. pallidum subsp. Carateum

Infect humans and have not been cultivated for more than one
passage in vitro.
Most species stain poorly with gram staining/ giemsas methods
Best observed with dark-field or phase-contrast microscopy
Microaerophilic
T. vincentii
T. denticola
T. refringens
T. socranskii
T. pectinovorum

Normal inhabitants of the oral cavity or the human genital tract

Cultivable anaerobically on artificial media

Epidemiology and Pathogenesis

-Pathogenic treponems
Penetrate intact mucous membranes
Enter through breaks in the skin
T. pallidum is transmitted by sexual contact and vertically from mother
to the unborn fetus
Invades bloodstream and spreads to other body sites
Remarkable tropism to arterioles
Infection leads to endarteritis (inflammation of the lining of arteries)
and subsequent progressive tissue destruction.

-Spectrum of Disease
T. pallidum
Causes venereal syphilis (transmitted through sexual contact).
Stages:

Primary syphilis the appearance of chancre (painless ulcer) at the

site of inoculation, most commonly genitalia. Dissemination of the
organism occurs in this stage.

Secondary syphilis the patient is ill and in need of medical

attention. Systemic symptoms such as fever, weight loss, malaise, and
loss of appetite are present. The skin is the organ most commonly
affected (widespread rash with generalized lymphadenopathy). Aseptic
meningitis also occur.

Latent period the disease becomes inactive. Diagnosis can be

made using serologic methods. Relapses are common during this
stage.

Late Latent Syphilis asymptomatic and noninfectious

Tertiary syphilis tissue-destructive phase that appears 10 to 25

years after the initial infection in up to 35% of untreated patients.
Complications include central nervous disease (neurosyphilis),
cardiovascular abnormalities, eye disease and granuloma-like lesions,
 Gummas found in the skin, bones, or visceral organs. Congenital
syphilis is transmitted from mother to the unborn fetus during any
stage of infection. The unborn fetus may develop an asymptomatic
infection or symptomatic infection with damage to the bone and teeth,
deafness, neurosyphilis or neonatal death.

Laboratory diagnosis
Specimen collection
Samples from ulcers and lesions should not be contaminated with
blood, microorganisms, or tissue debris.

PCR samples
Collection: sterile Dacron or cotton swab
Container: cryotube containing nucleic acid transport medium or
universal transport medium.

Tissue or needle aspirates of lymph nodes place in 10% buffered

formalin at room temperature

Congenital syphilis small section of the umbilical cord is collected

and fixed in formalin or refrigerated until processed.

Direct Detection
Dark-field examination
Fluorescent antibody staining and microscopic examination
Gloves should always be worn.
Treponemes are long (8 to 10 microns) and consist of 8 to 14 tightly
coiled, even spirals.

Molecular Diagnostics
PCR for detection of T. pallidum
Useful for identification of organisms within exudate/lesions.

Serodiagnosis
For Treponematosis
2 types of antibodies
Treponemal antibodies produced against antigens of the
organisms themselves
Nontreponemal antibodies reagin antibodies produced in
infected patients against components of mammalian cells.

*Two most widely used nontreponemal serologic tests are:

Venereal Disease Research Laboratory (VDRL) Test quantitative
test; serum or CSD
 Rapid plasma region (RPR) Test

Other tests
EIA
Agglutination tests T. pallidum particle agglutination (TPPA)
Microhemagglutination assay (MHA-TP)
T. pallidum indirect hemaagluttination (TPHA)
Particle gel immunoassay (PaGIA)
Fluorescent treponemal antibody absorption (FTA-ABS)

Antimicrobial Susceptibility Testing and Therapy

Penicillin G is the drug of choice.
Ceftriaxone is also highly active in most cases of syphilis other than early
syphilis.
Tetracycline or doxycycline is the alternative for patients allergic to penicillin.
Treatment varies depending on the stage of disease and the host.

Prevention
No vaccines are available for the treponematoses.
Prevention is best accomplished by early and appropriate treatment, thereby
preventing person to person spread.

BORRELIA
Borreliosis
-relapsing fever
-transmitted by a human-specific body louse or a tick
-organisms are composed of 3 to 10 loose coils
-actively motile
-contain endoflagella beneath the outer membrane
-contain protoplasmic cylinder
-stain well with giemsas stain
-microaerophilic or anaerobic

Epidemiology and Pathogenesis

-causative agents of tickborne and louseborne relapsing fever and tickborne
Lyme disease in humans.

-Relapsing fever
Transmitted to humans by the bite of a louse or tick
B. recurrentis responsible for louseborne or epidemic relapsing
fever
- Pediculus humanus humanus
Humans are the only reservoir for B. recurrentis
Other species of ticks transmitting the organism: Ornithodoros (soft
tick)
 Depending on the orgnisms and the disease, their reservoir is either
humans or rodents in most cases.

Exhibit antigenic variability that may account for the cyclic fever
patterns associated with this disease.

-Lyme Disease
B. burgdorferi
B. garinii
B. afzelii
B. spielmanii
B. lusitaniae
B. valaisiana

Agents of lyme disease and are transmitted by the bite of Ixodes

(hard) ticks.
Ticks natural hosts are deer and rodents.
All stages of ticks larve, nymph, and adult can harbor the spirochete
and transmit disease.
Ticks require a period of attachment of at least 24 hours before they
transmit disease.
B. burgdorferi is able to avoid the human hose response.
Lyme disease are also believed to be due to the release of host
cytokines initiated by the presence of the organism.

Spectrum of Disease
-Relapsing Fever
2 to 15 days following infection, patients have an abrupt onset of fever,
headache, myalgia that lasts for 4 to 10 days.

Physical findings: Petechiae, Diffuse abdominal tenderness, and

conjunctival effusion.

Organisms hide from the antibody in different organs during the

afebrile period.

Subsequent relapses are usually milder and of shorter duration.

Louseborne relapsing fevers tend to be more severe.

Jarisch-Herxheimer reaction associated with the clearance of the

organisms from the bloodstream and release of cytokines within hours of
antibiotic treatment. Tachycardia, chills, rigors, hypotension, fever,
diaphoresis.

-Lyme Disease
 Characterized by three stages:
First Stage Erythema migrans (EM) red, ring-shaped skin lesion
with a central clearing that first appears at the site of the tick bite but may
develop at distant sites as well. May experience headache, fever, muscle and
joint pain, malaise.

Second stage arthritis, neurologic disorders, carditis. Result of the

hematogenous spread of spirochetes to organs and tissues. Neurologic
symptoms and infection may occur in the meninges, spinal cord, peripheral
nerves, and brain.

Third stage chronic arthritis, acrodermatitis chronica atrophicans

(ACA) diffuse skin rash, and may continue for years.

Laboratory Diagnosis
-Specimen Collection, Transport, and Processing
Relapsing fever peripheral blood for direct detection
Lyme disease serum

For stain/culture:
Blood
Biopsy specimen
Join and cerebrospinal fluids > transported without any preservatives
Tissue biopsy > placed in sterile saline to prevent drying

-Direct Detection Method

Relapsing Fever direct observation in the peripheral blood; dark- or
bright- field illumination; staining thick or thin films with Wrights or Giemsas
stains

Lyme Disease B. burgdorferi stained with Warthin-starry silver stain

- PCR dependent on the state of illness.

-Cultivation
Best specimens for culture: EM ring lesion/synovial tissue.
Kellys medium incubated at 30 to 34 degrees Celsius for up to 12
weeks under microaerophilic conditions.

-Seriodiagnosis
Relapsing Fever have not demonstrated reproducible or reliable
data for diagnosis because of the many antigenic shifts undergo during the
course of the disease.

Lyme Disease the standard for the diagnosis of Lyme disease.

Antibiotic Susceptibility Testing and Therapy
Relapsing fever tetracycline
Lyme Disease Doxycycline, amoxicillin, cefuroxime and parenteral
cephalosphorins are drugs of choice during the first stage.
- Ceftriaxone or cefotaxime for patient that failed initial
treatment or present in later stages
- Symptomatic treatment failures in patients with chronic
lyme disease have been reported.

Prevention
Recombinant outer surface protein A vaccine has been licensed for use
in humans against Lyme disease caused by infection with organisms
belonging to the B. burgdorferi complex.

Avoid tick-infested areas

Wear protective clothing
Check your clothing, body, and pets for ticks and removing them
promptly will also assist in the prevention of infection.

No vaccines against infections caused by other Borrelia species.

BRACHYSPIRA
General Characteristics

Brachyspira aalborgi; anaerobic; has not been isolated from animals

B. pilosicoli colonizes the intestine of variety of animal species;
resides in the brush border within the intestine.

Epidemiology and Pathogenesis

B. aalborgi transmitted via fecal-oral contamination
B. pilosicoli - ingestion of water contaminated with feces from
infected animals.
No pathogenic mechanisms have been identified
Clinical significance must be carefully correlated with patient signs and
symptoms
Laboratory Diagnosis
Specimen Collection and Direct Detection
Fresh stool
Rectal swabs
*Examined by dark-field microscopy
 Tissue biopsy specimens histologic examination using periodic
acid-Schiff (PAS) or hematoxylin and eosin staining.

Cultivation
Brain heart infusion (BHI)
Tryptic soy agar containing 10% F bovine blood, 400 migrocrams
per mL of spectinoymycin, and 5 micrograms per ml of polymyxin in
anaerobic conditions at 37 degrees Celsius.
B. aalborgi is weakly beta-hemolytic on BHI medium.

Approach to Identification
B. aalborgi strong positive hippurate hydrolysis reaction and weak
indole reaction
B. pilosicoli indole negative, weak hippurate hydrolysis reaction

Antibiotic Susceptibility and therapy

B. pilosicoli susceptibility to Augmentin (amoxicillin-clavulanic
acid), ceftriaxone, chloramphenicol, meropenem, tetracycline,
metronidazole

LEPTOSPIRA
General Characteristics
Can be free-living or parasitic
Spiral-shaped
Right handed helices with hooked ends
Contain two axial filaments
Exhibit spinning motility (rapid back and forth movement)
Classified in two major groups:
Leptospira interrogans main species associated with human
leptospirosis
Leptospira biflexa saprophytic environmental strains

Epidemiology and Pathogenesis

Leptospirosis a zoonosis
Common in developing countries, warm climates where contact with
infected animals or water contaminated with urine is likely to occur.

Leptospira interrogans
Can infect most mammals, as well as reptiles, amphibians, fish, birds
and invertebrates.

Humans become infected through direct or indirect contact with the

urine or blood of infected animals.
 Enters the human host through breaks in the skin, mucous membranes
or conjunctivae.

Infection can be acquired in home and recreational settings or in

people who work in certain occupational settings (farmers, ranchers,
abattoir workers, trappers, veterinarians).

Pathogenic leptospires rapidly invade the bloodstream after entry and

spread throughout all sites in the body such as central nervous system
and kidneys.
Virulent strains show chemotaxis towards hemoglobin and the ability to
migrate through host tissues.

Spectrum of Disease
Symptoms begin abruptly 2 to 20 days after infection
Fever, headache and myalgia
Most common clinical syndrome is anicteric leptospirosis.
Self-limiting illness consisting of a septicemic stage with high fever and
severe headache that lasts 3 to 7 days followed by the immune stage.

Immune stage milder than septicemic stage

- Aseptic meningitis
- Weils disease (icteric leptospirosis) most severe
illness; symptoms caused by liver, kidney, or vascular
dysfunction with lethal pulmonary hemorrhage; death
can occur.

Laboratory Diagnosis
Specimen Collection, Transport, and Processing
First 10 days leptospires are present in the blood, CSF, and peritoneal
dialysate.
Urine specimen can be obtained beginning in the second week of
illness up to 30 days after the onset of symptoms.
Collected in citrate, heparin, or oxalate anticoagulants.
EDTA is the favored anticoagulant for molecular testing.
Urine should not be placed in preservatives and should be processed
within 1 hour for optimal results.
Direct Detection
Dark-field microscopy
Fluorescent antibody staining
PCR assay

Molecular Diagnostics
 Pulsed field electrophoresis (PFGE)
Restriction fragment length polymorphism (RFLP)
For identification of serovars.

Cultivation
Culture organisms from blood, CSF, or urine.
Sodium oxalate anticoagulated blood are inoculated into tubes of
semisolid media enriched with rabbit serum or bovine serum albumin.
Urine should be inoculated soon after collection. The acidity may harm
the spirochetes.
200 microgram/mL of 5-fluorouracil may prevent contamination by other
bacteria without harming the leptospires.
Ellinghausen-McCullough-Johnson-Harris (EMJHS) or Fletchers Medium
Incubate at room temperature of 30 degrees Celsius in the dark for 6 to 8
weeks.

Approach to identification
Number of coils and hooked ends
Environmental leptospires can grow in 10 degrees Celsius or lower than
pathogenic leptospires.
May be visualized using dark-field or immunofluorescence.

Serodiagnosis
Fourfold or greater rise in titer of agglutinating antibodies.
Microscopic agglutination (MA) test
Indirect hemagglutination
IgM-detection assays

Molecular Testing
PCR
Real-time PCR

Antibiotic Susceptibility and Therapy

Supportive management
Use of appropriate antibiotics
Ceftriaxone, penicillin, amoxicillin, doxycycline, tetracycline recommended
for treatment of leptospirosis
Standardized procedures for antibiotic susceptibility are limited by the
slow growth of the organisms and the need for serum during cultivation.

Prevention
Vaccination of domestic livestock and pet dogs.
 Protective clothing, rodent control measures, and preventing recreational
exposures, such as avoiding freshwater ponds, are indicated in preventing
leptospirosis.