MEDICAL MICROBIOLOGY II

Lesson 11

Rickettsia and Legionella

Rickettsiaceae

The family of microorganisms that cause typhus fever and related diseases mostly transmitted by arthropods

Closely related to viruses in their inability to grow on artificial culture medium and their being obligate intracellular parasites

True bacteria, have DNA and RNA, cell wall, are susceptible to antibiotics, and divide by binary fission. They are smaller than bacteria but not filterable

Rickettsiaceae

Contain 3 genera: Rickettsia, Coxiella and Rochalimaea

Rickettsia contains agents of typhus fever, spotted fever and scrub typhus

Rochalimaea contains the agent of trench fever (the microorganism differs from rickettsia agents: usually extracellular in the arthropod host and able to grow on blood agar

Rickettsiaceae

Coxiella contains agent of Q fever which differs from Rickettsia in being smaller, more resistant to physical and chemical agents and transmissible without an arthropod vector

Rickettsiaceae can also be subdivided according to the type of diseases they produce, the vectors that transmit them and their geographical distribution

Rickettsia

Small Gram negative pleomorphic rods, non-motile, non-capsulated (but R. prowazeki has an amorphous capsule)

Do not grow on cell-free media but R. quintana is able to grow on blood agar (32 -35 C)

Most are found in normally in the alimentary canal of insects - lice and mites

Rickettsia

R. prowazeki has man as its only natural maintenance host and the human body louse is the vector

The rickettsiae multiply in the gut of the louse and are found in the faeces of the louse

Infection occurs through contaminated louse faeces being rubbed in by scratching damaged skin

Rickettsia

Infection can also occur through inhalation of dried louse faeces, or through the conjunctiva or through blood transfusion

The vector for R. tsutsugamushi is the thrombiculid mite and its natural reservoirs are rats, mice, voles and quail

Infection occurs through the bite of an infected larval mite as the rickettsiae are contained in the saliva of the mite

Rickettsia

Rats are natural reservoirs of R. typhi (R. mooseri) and infection is transmitted by the rat flea, Xenopsylla cheopis

The rickettsiae multiply in the gut of the flea

Human infection is through inhalation of rickettsial particles in dried flea faeces

Rickettsia

The natural reservoirs for R. conori and R. siberica are dogs, rats, mice and probably rabbits

The rickettsiae are transmitted by hard ticks

Human infection is through the bite of infected tick

Pathogenicity

Pathogenesis begins with the invasion and multiplication of the rickettsiae in the endothelial cells of small blood vessels, smooth muscle cells and histiocytes

Tissues most involved are those of the skin, heart, brain and lungs

The invading organisms cause damage of tissues with a local cellular response in the infected cells become severely inflamed

Pathogenicity

Capillaries become blocked and there may be bleeding in severe cases

Most infections are characterised by high continuous fever, severe headache, general body pains and weakness

There is a characteristic macular or maculopapular rash appearing toward the end of the first week

Other signs may include oedema, nose bleeds and involvement of the eyes

Pathogenicity

Towards the second week the patient may become delirious, confused and eventually comatose

Fatality ranges from 15 - 70% and increases with age

Death usually occurs from general toxaemia, encephalitis, pneumonia or myocarditis

Typhus and spotted fever groups give the most severe symptoms

Pathogenicity

Following rickettsial infection, gangrene of the fingers, toes and other parts of the body may occur

In patients who recover, the organisms may remain latent in the lymphoid tissue and can be triggered off again in years to come

Laboratory Diagnosis

Most lab are restricted to performing only serological testing because of the difficulties in working with and the risk of infection with rickettsiae organism

Specimens: blood, tissues and sputum (should be labeled as biohazards)

Laboratory Diagnosis

1. Microscopy

Smears may be examined directly after staining by Giemsa, Castaneda or Macchiavellomethods

Also by IF

Organisms are pleomorphic, forming bacillary and coccal forms, small, measuring less than 0.5 m in diameter

Usually seen in tissue smears near cells or free within the cytoplasm

Laboratory Diagnosis

2. Culture

Can be cultivated in animals, egg yolk or cell culture (only to be performed in well equipped lab)

3. Serology (ELISA of IF using specific rickettsial antigens

The immunological response to rickettsial infections is firstly the production of IgM antibodies

Followed in the later stages of the disease by IgG antibodies (may remain in circulation for years)

Laboratory Diagnosis

4. Weil-Felix reaction

A non-specific agglutination test in which rickettsial sera are tested for agglutinins to the O antigens of certain non-motile strains of Proteus vulgaris (OX19 and OX2) and Proteus mirabilis (OXK)

It is based on cross-reaction between rickettsial antibodies and the antigens of the Proteusstrains

Simple and useful test to diagnose acute rickettsial infections

Laboratory Diagnosis

Laboratory Diagnosis

5. Antibiotic sensitivity

Usually sensitive to erythromycin, tetracycline, chloramphenicol

Usually resistant to penicillin

Legionella

Gram negative, aerobic, non-sporing, non acid-fast and encapsulated bacteria, 0.3 - 0.7 m

Catalase and oxidase variable

Primary isolation may take up to 10 days incubation at 35 C

Growth is enhanced in an atmosphere containing 5% carbon dioxide

Most members are motile and produce beta-lactamase

Legionella

Type species: L. pneumophilia, L. micdadei, L. bozemanii, L. longbeachae, L. dumoffii, and L. gormanii

Pathogenicity

L. pneumophilia causes Legionnaires disease or Legionellosis - an atypical pneumonia with high fatality

It was first known as Legionnaires disease bacterium (LDB)

It derived its name from the epidemic at the American Legion convention in Philadelphia in 1976 (29 death out of 183 cases)

First named L. legionella, then later changed to L. pneumophila

Pathogenicity

L. pneumophila is an environmental microorganism

Human infection usually occurs from transmission through aerosols of organisms that have colonised water supplies and air conditioning systems

Person to person transmission is rare Legionellosis usually presents as non-

productive cough, chest pain and high fever Other symptoms include mental confusion,

diarrhoea, nausea and vomiting

Laboratory Diagnosis

Specimens: sputum, bronco-alveolar lavage, respiratory tissue and rarely pleural effusion and blood for culture. It may be necessary to culture samples from hospital water supplies and samples from cooling systems of air conditioners. Specimens must be sent ASAP but if not, specimen should be kept in refrigerator

Laboratory Diagnosis

1. Microscopy

Gram stained sputum - faintly stained Gram negative bacilli

Direct fluorescent antibody method are also valuable though they may not be totally specific as occasionally cross-reactions occur with non-legionella bacteria

Laboratory Diagnosis

2. Culture

Fastidious and slow growing, require growth supplements and cysteine for growth

Primary isolation medium is the buffered charcoal yeast extract (CYE) agar, medium is made selective by addition of vancomycin, polymixin B and anisomycin

Medium is inoculated with 1:10 dilution of sputum or lavage, and incubated at 35 C

Laboratory Diagnosis

Colonies of about 1 - 2 mm may appear after 3 days incubation

The colonies are round, greyish-pink and opalescent with entire edges

Laboratory Diagnosis

3. Identification test

Presumptive identification of Legionella is made when the suspected isolate is found to be cysteine dependent and faintly Gram negative bacillus

When sub-cultured on a cysteine deficient medium, the organisms fail to grow

4. Serology

Indirect immunoflourescent method is usually used to detect L. pneumophilia antibodies

Laboratory Diagnosis

A rapid microagglutination test is commercially available

5. Antibiotic sensitivity Sensitive to erythromycin, rifampicin,

cotrimoxazole and 4-fluoroquinolones

Most strains are resistant to penicillin due to their beta-lactamase production

Aminoglycosides are also effective therapy

THE END