Haemophilus, Bordetella, Brucella Flashcards
General characteristics of Haemophilus
◾ Gram-negative coccobacilli
◾ facultative anaerobes
◾ non-motile
◾ non-sporing
◾ oxidase positive
◾ catalase positive
Briefly discuss Haemophilus accessory growth factors.
(1) Factor X (Hemin): This is porphyin compound [similar to haem]. It is required for the synthesis of cytochrome and other enzymes e.g. catalase, peroxidase and oxidase. It is heat stable.
(2) Factor V (NAD or NADP): These are hydrogen acceptors in cellular metabolism. They are heat labile (destroyed at 120°C).
Briefly discuss Haemophilus virulence factors.
✔ Capsule: contains PRP (polyribosyl-ribitol phosphate), which prevents phagocytosis and opsonisation. This is the main virulence factor.
✔ Pili; adherence factor
✔ Outer membrane proteins e.g. porins, adhesins
✔ Lipooligosaccharide; may trigger strong inflammatory response
✔ IgA protease
How are Haemophilus influenzae serotypes classified?
H. influenzae strains are broadly classified as either:
(1) Encapsulated: These are further classified into six serotypes (a-f) based on the capsular polysaccharide. [These serotypes are not considered strains.]
(2) Non-encapsulated (NTHi): These lack a capsule and are not classified into serotypes.
✔ Strains: Both encapsulated (within a serotype) and non-encapsulated H. influenzae can be further classified into strains. Strains are defined primarily by genetic differences.
✔ Biotype: strains are futher subdivided into 8 biotypes (I-VIII) based on biochemical characteristics: urease activity, ornithine decarboxylase activity, and indole production.
Comment briefly on the epidemiology of H. influenzae.
◾ Mucosal organism spread through respiratory droplets.
◾ Present in 30-50% of healthy persons.
◾ Infections are seen frequently in children aged from six months to 4 years of age.
◾ In adults, it may produce secondary complications of severe primary illnesses.
◾ Immunocompromised adults are also at risk of developing infections.
Which Haemophilus influenzae serotype is responsible for majority of invasive diseases?
serotype b
Where is NTHi commonly found?
normal microbial flora of the upper respiratory tract
What types of infections are most commonly associated with NTHi?
It is usually involved in respiratory tract infections in children, otitis media, sinusitis, bronchitis and pneumonia.
Briefly describe the key steps in the pathogenesis of Haemophilus influenzae.
(1) H. influenzae attaches to the respiratory mucosa using adhesins and pili.
(2) The bacteria evade the host’s immune response via:
✔ IgA proteases (degrade IgA antibodies)
✔ Microcolony formation
✔ Phase/Antigenic variation (changes in surface antigens)
✔ Intracellular survival/invasion (in some cases)
(3) Encapsulated strains resist phagocytosis due to their capsule, allowing them to enter the bloodstream and cause invasive infections like, meningitis, epiglottitis, and septicaemia.
appropriate specimens for laboratory investigation of H. influenzae
Depends on the type of infection:
✔ Meningitis: CSF
✔ Epiglottitis: blood
✔ Pneumonia: sputum/tracheal aspirates
✔ Others: joint fluid, pleural fluid, middle ear fluid
State the most suitable agar for culturing of H. influenzae, and explain why it is preferred over other media.
Chocolate blood agar (CBA)
CBA is a nutrient agar enriched with heated blood, which releases Factor X and Factor V, making them available for the bacteria to grow.
Further notes:
Blood agar: While it contains blood, the red blood cells remain intact, and Factor X is not readily available. H. influenzae may grow on blood agar only as tiny satellite colonies around other bacteria that can lyse the red blood cells and release Factor X.
MacConkey agar: This is a selective medium for gram-negative bacteria, but it does not provide the necessary growth factors for H. influenzae.
What is the basis of the satellitism test for identifying Haemophilus influenzae?
◾ The satellitism test relies on the fact that H. influenzae requires specific growth factors, primarily NAD (nicotinamide adenine dinucleotide or Factor V), which it cannot produce on its own.
◾ Staphylococcus aureus produces NAD as a byproduct of its metabolism.
◾ Therefore, when the two bacteria are co-cultured, H. influenzae will grow as small “satellite” colonies only near the S. aureus streak.
[Image 1] [Image 2]
main prevention method against H. influenzae
Vaccination. Currently the only available H. influenzae vaccine is the Hib vaccine.
What is Haemophilus influenzae biogroup aegyptius (Hae), and what is its primary clinical manifestation?
◾ Hae is a biogroup of non-typeable H. influenzae (NTHi) with a strong predilection for the conjunctiva.
◾ Its primary clinical manifestation is acute purulent conjunctivitis, a contagious eye infection characterized by pus discharge, often occurring in seasonal epidemics.
Besides conjunctivitis, what other, less common, disease is associated with Haemophilus influenzae biogroup aegyptius?
Brazilian Purpuric Fever: conjunctivitis, high fever, haemorrhagic skin lesions, abdominal pain, nausea, vomiting, septic shock, death [Image]
Briefly discuss clinical implications of Haemophilus ducreyi infection.
◾ Causative agent of chancroid: a sexually transmitted ulcer
◾ May cause lymphadenitis and bubo [large, tender lymph nodes in the groin] formation
◾ Can spread to other sites through autoinoculation
Progression and clinical presentation of chancroid
✔ Tender erythematous papule: The initial lesion usually appears as a small, red, and tender bump (papule) at the site of infection [4-7 days after exposure to H. ducreyi].
✔ Papule develops into a pustule
✔ Pustule ruptures within 2-3 days, forming a painful, shallow ulcer
✔ The ulcer has a soft, raised border, and a base covered with a grayish or yellowish, pus-like exudate
common sites for chancroid lesions in men and women
List the HACEK organisms.
◾ Haemophilus species [other than H. influenzae]
◾ Aggregatibacter species
◾ Cardiobacterium hominis
◾ Eikenella corrodens
◾ Kingella species
What do the HACEK organisms have in common?
◾ They are all part of the normal flora of the oral cavity.
◾ They are opportunistic microorganisms; require immunocompromised host.
◾ They can enter the bloodstream and cause infection, particularly in people with damaged heart valves or other underlying health conditions.
◾ The are responsible for 3% of cases of infective endocarditis, and are often a cause of culture-negative endocarditis.
3 clinically relevant Bordetella species
✔ Bordetella pertussis
✔ Bordetella parapertussis
✔ Bordetella bronchiseptica
general characteristics of Bordetella
◾ small, Gram-negative coccobacilli
◾ strict aerobes
◾ non-motile [B. bronchiseptica is motile]
◾ capsulated
◾ non-spore forming
Bordetella virulence factors
(1) Adhesins: filamentous haemagglutinin (FHA), pertactin, fimbriae
(2) Toxins: pertussis toxin, adenylate cyclase (CyaA) toxin, dermonecrotic toxin, tracheal cytotoxin
(3) Secretion systems: Type III secretion system, Type IV secretion system
Make a brief note on the important virulence secretion systems in Bordetella.
✔ Type III secretion system: Bordetella species possess a T3SS, a needle-like structure that allows them to inject effector proteins directly into the cytoplasm of host cells.
✔ Type IV secretion system: The T4SS forms a channel through which PT is transported across the bacterial outer membrane.
Briefly discuss the pathogenesis of Bordetella pertussis, focusing on its causation of Whooping Cough.
◾ Transmitted via the respiratory route.
◾ Primarily targets ciliated epithelial cells of the respiratory tract.
◾ Adhesins like FHA, pertactin and fimbriae facilitate attachment to these cells.
◾ Once achored, bacterium produces tracheal cytotoxin which stops the cilia from beating. This prevents the body from clearing debris from the lungs; the patient gets into a coughing fit. This is also contributed by excessive mucus production due to pertussis toxin.
◾ Toxins and other bacterial components trigger inflammation in the respiratory tract.
mechanism of action of pertussis toxin
🧨 comprises 2 subunits: A [enzymatically active] and B [mediates binding and entry]
🧨 internalized by endocytosis
🧨 travels through the cell’s internal transport network, making its way to the ER
🧨 in ER, A subunit is released from B subunit
🧨 A subunit mediates ADP-ribosylation of the inhibitory subunit of G protein complex called Gi/o.
🧨 this modification causes disregulation of production of cAMP, resulting in increased cAMP levels
🧨 disruption of cellular processes: increased mucus production, lymphocytosis, histamine sensitization
state 3 stages of Bordetella pertussis infection
(1) Catarrhal stage
(2) Paroxysmal stage
(3) Convalescent stage
Briefly discus catarrhal stage of Bordetella pertussis infection.
runny nose, sneezing, low-grade fever, lacrimation, mild cough [that worsens in frequency and degree]
Briefly discus paroxysmal stage of Bordetella pertussis infection.
🧨 intense, uncontrollable coughing spells that can occur many times a day
🧨 at the end of a paroxysm, there is a massive inspiratory effort during which the classic “whoop” occurs
🧨 coughing fits may be accompanied by vomiting, cyanosis
Briefly discuss convalescent stage of Bordetella pertussis infection.
recovery phase; coughing gradually becomes less frequent and less severe
3 complications of bordetella pertussis infection
encephalitis, seizures, bronchopneumonia, respiratory failure, atelectasis [incomplete expansion or collapse of a lung (or part of a lung)]
Bordetella pertussis clinical specimen
nasopharyngeal swabs and aspirates/respiratory droplets on a cough plate
Bordetella pertussis growth media
(1) Bordet-Gengou (BG) agar
(2) Regan-Lowe agar [charcoal agar with cephalexin]
Bordetella pertussis growth conditions
Temperature: 35-37°C
Atmosphere: aerobic with increased humidity
Incubation time: 3-7 days [slow-growing]
Bordetella pertussis colonial morphology
small, pearl-grey, shiny (like mercury droplets), mucoid colonies [Image]
Bordetella pertussis biochemical tests
urease negative, negative nitrate reduction test, oxidase positive
Bordetella pertussis prevention
(1) Vaccination by DTaP vaccine
(2) Early recognition and isolation of patients to limit spread
(3) Prompt treatment of infected individuals
general characteristics of Brucella
✔ pleomorphic gram-negative bacilli/coccobacilli
✔ non-spore forming
✔ non-motile
✔ non-capsulated
✔ obligate aerobes
✔ oxidase, catalase, and urease positive
mode of transmission of Brucella
It is a zoonosis. May enter via:
(1) mucous membrane: droplets
(2) broken skin: direct or indirect contact with infected animals
(3) ingestion [e.g. unpasteurized dairy products]
Brucella species primarily considered to be pathogenic to humans
(1) B. melitensis
(2) B. abortus
(3) B. suis (biovars 1, 3 and 4)
(4) B. canis
Brucella virulence factors
✔ intracellular survival
✔ lipopolysaccharide (LPS)
✔ type IV secretion system
✔ outer membrane proteins
Briefly explain the pathogenesis of Brucella.
◾ After entry, the bacterium is taken up by macrophages.
◾ It avoids destruction by manipulating the host cell’s machinery to create a protective niche called teh “Brucella-containing vacuole”.
◾ This vacuole avoids fusion with lysosomes.
◾ The Brucella-containing vacuole becomes associated with the endoplasmic reticulum. Once inside this ER-associated compartment, the bacteria can establish chronic infection.
Brucella clinical manifestations
◾ Acute illness may be characterized by: undulant fever [fluctuating], generalized malaise, headache, lymphadenopathy, hepatosplenomegaly, muscle and joint pain.
◾ Chronic illness may have similar symtpoms as the acute illness.
Brucellosis complications
Diverse: osteomyelitis, endocarditis, neurobrucellosis [meningitis, encephalitis], epididymo-orchitis, miscarriage or premature birth, etc …
specimen for laboratory diagnosis of Brucellosis
blood (preferred), bone marrow
Brucella culturing
✔ Tryptic soy broth
✔ Usual culturing temperatures
✔ Facultative anaerobe, though it grows best in an atmosphere enriched with 5-10% CO2
✔ Cultures should be incubated for 4 weeks
✔ Colourless or grey colonies
✔ Highly infectious, so extra precautions must be observed
List serological methods that can be employed in the laboratory diagnosis of Brucellosis.
◾ ELISA
◾ Standard tube agglutination test
◾ Modified tube agglutination test
◾ Brucellin skin test
◾ PCR
Brucella prevention and control
🧨 Control of disease in animal hosts
🧨 Effective heat treatment of dairy produce
🧨 Hygienic precautions to prevent occupational exposure
antimicrobial agents effective against Brucella
doxycycline, streptomycin, rifampin
