Microbiology 2011 May 13, 2011 Yu Chun-Keung DVM, PhD

Chapter 42

Treponema, Borrelia, and Leptospira

Chapter 43

Mycoplasma and Ureaplasma

Chapter 42 Treponema, Borrelia, and Leptospira

疏螺旋體 密螺旋體 鉤端螺旋體

Spirochete

螺旋菌目

:

細長、彎曲、有運動力的細菌

Order Spirochaetales Human disease

Treponema Borrelia

密螺旋體 疏螺旋體

Leptospira

鈎端螺旋體 梅毒 Syphilis 貝傑 Bejel 斑病 Yaws 莓疹病 Pinta 回歸熱 Relapsing fever 萊姆氏病 Lyme disease Leptospirosis 鈎端螺旋體症 Etiologic agent T. pallidum T. endemicum T. pertenue T. carateum B. recurrentis B. burgdorferi L. interrogans

T. pallidum

Strict human pathogen, experimental syphilis only in rabbits, cannot grown in cell-free cultures

Syphilis (

梅毒

)

, sexually transmitted disease Syphilis : syn - together, philis - love

T. pallidum

Virulence factors:  Outer membrane protein for adherence,  Hyaluronidase for perivascular infiltration, )  Coated with fibronection for protection against phagocytosis Immune response to infection: tissue destruction

Clinical course

Primary phase: papule ( 丘疹 ), hard chancre ( 硬 性下疳 , painless ulcer) at site of inoculation, lymphadenopathy, endarteritis and periarteritis, heal spontaneously within 2 months, highly infectious, Ab(-).

Secondary phase: flu-like syndrome, generalized skin rash and condylomas ( 扁平濕疣 month, highly infectious, Ab(+).

), meningitis, hepatitis, etc; subside spontaneously week to Latent: 2 or 3-10 yr, no clinical sign, non infectious.

Clinical course

Late (tertiary) phase: all tissues may be involved, cardiovascular lesions (aneurysm 主動脈瘤 lesions (gummas, 80-85%), CNS degeneration (5-10%), granulomatous 梅毒腫 ) in skin, bone and liver. 30% of cases completely cure after 1 2 nd stage.

st or phase; another 30%, latent, serologic test (+); remainder, progress to tertiary

Congenital syphilis

Treponemas invade fetus at the fifth month of gestation Causes abortion, stillbirth, or death soon after delivery Congenital abnormality: interstitial keratitis, Hutchinson's teeth, saddle nose, and eighth nerve deafness

Epidemiology

Worldwide; in USA Neisseria gonorrhoeae > Chlamydia > Syphilis Extremely sensitive to drying or disinfectants, cannot be spread through contact with inanimate objects Not highly contagious, 30% change of infection after a single sexual contact Can be acquired congenitally or by transfusion; bacteremia can persist for > 8 years Incidence of late syphilis has markedly decreased, primary and secondary syphilis remain high

Diagnosis of syphilis

Direct detection of spirochetes : Darkfield microscopy (motile bugs + experience + prompt examination) Silver stain Culture : not used Serology: non-specific and specific tests

Non-treponemal tests

Antigen: cardiolipin (beef heart) + lecithin + cholesterol Detect nonspecific antibody (Reagin): a mixture of IgM & IgG direct against some normal tissue antigens VDRL (Venereal Disease Research Laboratory) test for serum and CSF samples

Advantage of VDRL:

cheap, easy to perform quantitative, screen test monitor disease course trace theraputic effect, become “ ” in 6-18 m after effective treatment.

Disadvantage of VDRL

Transient false positive: acute febrile disease, vaccination, pregnant women Long-term false positive: chronic autoimmune diseases, liver diseases False negative: progressive tertiary syphilis

Treponemal tests

Antigen:

T. pallidium

Detect specific antibody to T. pallidium (1) Fluorescent treponemal antibody (FTA) test Killed organism + patient's serum + labeled anti-human Ig = fluorescence “+” if Ab present (2) Micro-hemagglutination for (MHA-TP) test T. pallidum (RBC + treponemas) + patient's serum = “clumps” if Ab present

Treponemal tests

Confirmative tests for syphilis Influenced less by therapy False positive: autoimmune diseases Positive serologic test in infants: mean passive transfer of antibody OR congenital infection Re-test 6 months later

T/P/C

Penicillin: especially for neurosyphilis for pregnant women Tetracycline, doxycyline No vaccine, safe sex

Other Treponemes

–

cause

Nonvenereal treponemal diseases

Bejel (endemic syphilis) Yaws Pinta

Bejel (endemic syphilis)

T. pallidum subsp

endemicum

A highly infectious skin lesion, gummas of skin, bones, and nasopharynx Occur among children in Africa, Asia, and Australia.

Person-to-person, contaminated eating utensil

Yaws

T. pallidum subsp

pertenue

A chronic skin disease among children; destructive lesions of skin, lymph nodes, bones Tropical areas of South America, Central Africa, Southeast Asia Person-to-person, direct contact Nodules on the elbow resulting from a pertenue Treponema bacterial infection. The Free Dictionary

Pinta

T. pallidum

subsp

carateum

A skin disease causing unsightly pigment changes Central and South America Person-to-person, direct contact

Chapter 42 Treponema, Borrelia , and Leptospira Borrelia recurrentis

Relapsing fever ( 回歸熱 )

Borrelia burgdorferi

Lyme disease (萊姆病)

Relapsing fever (

回歸熱

)

A bloodstream infection producing 3 to 10 recurrence of febrile and afebrile cycles. Incubation period about 1 week Bacteremic phase (release endotoxin)  3-7 days - fever, chills, muscle aches, headache, splenomegaly, hepatomegaly  The bacteria are rapidly removed by specific antibody (agglutination and complement mediated lysis)

Afebrile period

 1 week  Antigenic variation - alter serotype-specific outer envelope proteins through gene rearrangement

Relapse

Borrelia recurrentis : epidemic relapsing fever  Human body louse (no transovarian transmission), short lifespan  Single relapse, more severe, mortality 4%-40% Borrelia spp : endemic relapsing fever  Soft ticks (transovarian transmission), long lifespan  Repeated relapse, mortality <5% 體蝨 軟蜱 硬蜱

Lab diagnosis

Human blood → blood smear → Giemsas or Wright stain → Borellia are large enough to be detected by microscope Human blood → inject into to the abdomen of laboratory mouse → mouse blood → smear → stain → Detect Borellia

Lyme disease

(萊姆病)

It is caused by spirochete Borrelia burgdorferi It is transmitted to humans by the bite of a hard-shelled tick ( Ixodes ). Reservoir – rodents, deer, ticks (transovarian transmission)

Kenneth Todar University of Wisconsin-Madison Department of Bacteriology

Disease stages of Lyme disease

Initial stage

Incubation period 3-30 days

Erythema chronicum migrans

bite (游走性紅斑 ) develop at site of tick Flu-like illness – malaise, severe fastigue, headache, fever, chill Lymphadenopathy, >4 weeks

Late stage

80% patients develop late manifestation (if untreated)

First phase

– neurologic symptoms (meningitis, encephalitis … ), cardiac dysfunction

Second phase - arthralgia and

arthritis Low numbers of organisms present in skin lesion, immunopathology (?)

Sensitivity of Diagnostic Tests for Borrelia Infections

Test Test Sensitivity

Relapsing fever Lyme disease

Microscopy

Good

Poor Culture Poor Serology Not available * IFA, ELISA Poor

confirmatory tests

*

Treatment

Relapsing fever: Tetracycline, Erythromycin Lyme disease: doxycycline

Chapter 42 Treponema, Borrelia, and Leptospira

Leptospira

Leptospira interrogans (

問號型鉤端螺旋體

)

218 serovars; shaped like a question mark; pathogenic for wild and domestic animals and humans

Leptospira biflexa

63 serovars; twice bent, a free-living saprophyte, not associated with disease

Grown in medium with rabbit serum

or bovine serum albumin (neither treponema nor borrelia can) Natural reservoir: rodents (rats), farm animals

Colonize the renal tubules and

shed in urine

Streams, standing water, moist soil – source of infection, survive > 6 weeks

An zoonotic disease Recreational exposure, occupational exposure, flood; no person-to-person spread Penetrate intact mucous membranes or skin through cuts or abrasions, through blood stream, spread to all tissues Multiply rapidly and damage

endothelium of small blood vessels

Leptospirosis

Mild leptospirosis (90% of cases) Severe leptospirosis (Weil ’ s disease) Acute stage (septicemic phase) : the first week after organisms enter the bloodstream, leptospira can be cultured from the blood and cerebrospinal fluid. A short asymptomatic period Chronic stage (immune phase) : leptospirae are found only in the urine.

Jaundice (organisms invade liver); nephritis (organisms invade kidney). Death results from kidney failure.

Diagnostic tests for Leptospirosis

Test Method Microscopy Gram stain Darkfield Sensitivity Insensitivity Insensitivity

Culture

FA Blood CSF Urine Insensitivity “+” during 1st wk “+” during 1st or 2nd wk “+” after 1st wk

Serology

Microagglu.* Sensitive, specific *patient ’ s serum to agglutinate live leptospira (reference laboratory test)

T/P/C

Penicillin or ampicillin i.v. for severe cases Doxycycline, ampicillin p.o. for less severe cases Doxycycline for prophylaxatic Vaccination of livestocks and pets Rodent control

Wash Those Soda Cans This incident happened recently in North Texas.

On Sunday Monday , a woman went boating taking with her some cans of coke which she put into the refrigerator of the boat. On she became ill and was taken to the hospital and placed in the Intensive Care Unit. She died on straight to the shops without being cleaned.

mouth to avoid any kind of fatal accident.

Wednesday .

The autopsy concluded she died of Leptospirosis. This was traced to the can of coke she drank from, not using a glass. Tests showed that the can was infected by dried rat urine and hence the disease Leptospirosis. Rat urine contains toxic and deathly substances. It is highly recommended to thoroughly wash the upper part of soda cans before drinking out of them.

The cans are typically stocked in warehouses and transported A study at NYCU showed that the tops of soda cans are more contaminated than public toilets (i.e.).. full of germs and bacteria. So wash them with water before putting them to the

Chapter 43

Mycoplasma and Ureaplasma

200 species; 16 colonize humans and 5 associated with diseases

Mycoplasma (

黴漿菌

)

M. pneumoniae M. hominis M. genitalium

Ureaplasma (

尿漿菌

)

U. urealyticum

Smallest

(0.1-0.3  m) and

simplest

free-living bacteria (about twice the genome size of certain large viruses) Small,

fried-egg-like

colonies (except

M. pneumoniae

)

Lack a cell wall

  Highly pleomorphic shapes

Resistant to penicillin

, cephalosporins, vancomycin, but sensitive to tetracycline, erythromycin.

Cell membrane contains sterols - rigid Anaerobic (except

M. pneumoniae

) Grow slowly in cell-free media, need sterols, use glucose as a source of energy (

ureaplasmas

require

urea

)

Epidemiology

M. pneumoniae

Strict human pathogen Worldwide disease with no seasonal incidence Most common in school-age

children and young adults (5-15y)

, but all age groups are susceptible Spread by respiratory droplets during coughing episodes in close contact among classmate or family members

U. urealyticum, M. hominis, and M. genitalium

Infants (females) are colonized with the agents Carriage does not persist. Only a small proportion of prepubertal children remains colonized

The incidence of genital mycoplasmas is associated with sexual activity

 Sexually active men and women 15% with

M. hominis

and

45-75% with Ureaplasma

Pathogenesis -

M. pneumoniae

Extracellular pathogen;

infect and colonize mucous membrane

(nose, throat, trachea, LRT).

Adheres to sialated glycoprotein receptor (1) at the base of cilia, (2) on surface of RBC by means of

P1 antigen

.

Pathogenesis -

M. pneumoniae

Causes

ciliostasis

, destroy cilia and ciliated epithelial cells; breakdown clearance activity, lead to LRT infection and

persistent cough

.

M. pneumoniae

contains

superantigen

, can attract inflammatory cells and induce cytokine secretion (TNF, IL-1, IL-6).

Clinical disease -

M. pneumoniae

Mostly asymptomatic carriage Cause mild URT disease (acute pharyngitis), low-grade fever, malaise, headache, dry and

nonproductive cough

, persist for > 2 weeks Tracheobronchitis with lymphocyte and plasma cell infiltration, and

atypical (walking) pneumonia

Secondary complication: hemolytic anemia, arthritis, myocarditis, pericarditis, neurologic abnormalities (e.g., meningoencephalitis)

Typical pneumonia

-

bacterial pneumonia

Abrupt, rigorous onset Productive cough, purulent sputum High fever, chest pain, stiffness in the neck Chest consolidation and rales.

Murray, et.al: Textbook of Respiratory Medicine

Atypical (walking) pneumonia

Chronic in both onset and recovery Flulike symptomes generalized aches, discomfort, headache, chill, dry cough, low grade fever Chest radiographs: patchy broncho pneumonia, interstitial pattern, not pneumonia Murray, et.al: Textbook of Respiratory Medicine

Diseases caused by U. urealyticum and M. genitalium and M. hominis

M. genitalium

: nongonococcal urethritis (NGU), pelvic inflammatory disease

U. urealyticum

: NGU, pyelonephritis, abortion, premature birth

M. hominis

: pyelonephritis, postpartum fever, systemic infection in immunocompromised patients

Lab diagnosis

Culture of mycoplasmas is not routinely attempted, and relatively insensitive 

M. pneumoniae

can grow in special medium with animal serum (sterols), yeast extract (nucleic acid), glucose, pH indicator, and penicillin. Colonies have a “mulberry-shaped”.



M. hominis

requires arginine for growth. Colonies have a fried-egg appearance.



Ureaplasma

requires urea for growth Microscope: no cell wall, stain poorly, no value

Serology –

for M. pneumoniae only

Complement fixation test : high false positive rate ELISA for detection of IgM and IgG Abs, more sensitive; need dual serum samples Cold agglutinins:  Non-specific IgM Abs that bind the I antigen on human RBC at 4 ° C, develop in 65% of the patients – insensitive and nonspecific.

Treatment / Prevention / Control

M. pneumoniae

: erythromycin, tetracycline (also good for chlamydia)

Ureaplasma

: use erythromycin, resistant to tetracycline

M. hominis

: resistant to erythromycin and tetracycline, use clindamycin Avoidance or safe sex for genital mycoplasma No vaccine available