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Which bacteria won't be seen on a gram stain?

- Myobacteria


Why does myobacteria not show up on gram stains?

Although there is peptidoglycan in the cell wall, this is joined to a layer of mycolic acid and beyond this a layer of outer lipids. Waxy, hydrophobic components of their cell wall make it difficult for aqueous stains to penetrate.


How would you stain mycobacteria?

The Ziehl - Neelsen stain (ZN stain)􏰉


What is The Ziehl - Neelsen stain (ZN stain)􏰉?

This staining technique utilises strong carbol fuchsin at a concentration greater than that used in the Gram stain, to visualise mycobacteria.
After they are stained, they appear red as Bacteria containing mycolic acid resist decolorisation with an acid-alcohol mix and remain red


What are the important cellular properties of myobacteria?

Rod shaped bacteria with peptidoglycan wall.

Layer of myopic acid around cell wall

Slow growth


What is ABF and what is its relevance?

Acid-fast bacilli = myobacteria.

They are called AFBs as they resist the avid decolorisation step of the ZN stain.


How are myobacteria classified? (nomenclature)

1.􏰉 Mycobacterium tuberculosis (the cause of tuberculosis or TB)
2.􏰉 Mycobacterium leprae (the cause of leprosy)
3.􏰉 “Atypical mycobacteria”, also referred to by the following acronyms:
􏰚􏰉 NTM - non tuberculous mycobacteria
􏰚􏰉 MOTB - mycobacteria other than TB
􏰚􏰉 MOTT - mycobacteria other than tuberculosis


What bacteria are part of the Mycobacterium tuberculosis complex?

􏰚􏰉 M. tuberculosis
􏰚􏰉 M. bovis
􏰚􏰉 M. bovis BCG
􏰚􏰉 M. africanum
􏰚􏰉 others


What are the names given to drug-resistant TB?

􏰙􏰉 MDR-TB - multi drug resistant TB
􏰙􏰉 XDR-TB - extensively drug resistant TB


How is TB acquired?

AIRBORNE. Inhalation from another person with active pulmonary TB.

- The transmitter sheds tubercle bacilli from the lung in large amounts when coughing, sneezing, shouting or singing.
- Particles are of a size defined as “aerosol particles” – 0.5 to 5.0 μm


What are the potential disease manifestations following M. tuberculosis infection?

- 90% - asymptomatic
- 5% - develop symptoms in 2 years (primary)
- 5% - >2 years (secondary)

Gradual, nonspecific onset:
􏰙􏰉 fever
􏰙􏰉 night sweats
􏰙􏰉 tiredness
􏰙􏰉 weight loss
􏰙􏰉 persistent cough ± blood stained sputum

Slowly progressive involvement of the lungs may occur, resulting in cavity formation and scarring. The process usually involves the upper lobes.


What is the Mantoux test?

Tuberculin skin test.

This involves the intradermal injection of a bleb purified preparation of M. tuberculosis antigen (PPD – purified protein derivative). People who have been infected with MTB will develop a local skin reaction which is measured after 48 hours. A positive test does not differentiate between latent infection and active infection.


What drugs are used to treat TB?

The first line drugs used to treat susceptible TB are:
􏰙􏰉 isoniazid
􏰙􏰉 rifampicin
􏰙􏰉 pyrazinamide
􏰙􏰉 ethambutol
The standard course of treatment is 6 months.


How much of the worlds population is infected with M. tuberculosis?



What effects does HIV have on TB?

- higher progression rate
- risk factor


What is the main tissue infected by M leprae?

Nervous - schwann cells


What is the gram stain appearance of Neisseria meningitides?

gram negative cocci, but sometimes coccobacilli (in-between rod and cocci)


What is one of the important virulence factors of Neisseria meningitides and why?

Polysaccharide capsule.


What are the usual manifestations of Neisseria meningitides?

• meningitis characterised by fever, meningism (headache, neck stiffness, photophobia), and pus in the CSF
• bacteraemia characterised by fever and rash (petechiae evolving into purpura and ecchymoses)
• meninigitis and bacteraemia together


Where is Neisseria meningitides usually found?

nasopharynx of 5-15% of the healthy population
- bacteria can cross the mucosal barrier and cause bacteraemia


What do you do with people who have been exposed to Neisseria meningitides (household contacts - 'kissing cousins')

Ciprofloxacin/rifampicin prophylaxis


What are common morphologies of gram negative rods?

- Capsule shaped (i.e. Enterobacteriaceae)
- Curved,(comma9shaped(rods,(eg.( genus(Vibrio&
- Helical(rods,(eg:( genus&Campylobacter& genus&Helicobacter&


What are beta-lactamases?

Enzymes that some bacteria (i.e. gram negative rods) produce which destroy beta-lactam antibiotics (penicillins, cephalosporins, carbapenems).


What are beta-lactam antibiotics?

penicillins, cephalosporins, carbapenems


What are the 3 broad groups of beta-lactamases of most concern in Gram negative rods

1. ESBLs - Extended Spectrum Beta-Lactamases
2. AmpC beta lactamases
3. Carbapenemases


What are the broad groups of gram negative rods?

- Enterobacteriaceae
- non-fermantative gram negative rods
- oxidase positive fermenters
- Miscellaneous, fastidious gram negative rods
- Campylobacteria
- Helicobacter
- Legionella
- Haemophilus


What are the Enterobacteriaceae?

- Escherichia
- Enterobacter
- Klebsiella
- Proteus
- Serratia
- Salmonella
- Shigella
- Yersinia


What are the non-fermantative gram negative rods?

- Pseudomonas
- Acinetobacter
- Stenotrophomonas
- Burkholderia

Its members don’t utilise glucose by fermentative biochemical pathways


What are the oxidase positive fermenters?

- Vibrio
- Aeromonas


What are the Miscellaneous, fastidious gram negative rods?

- HACEK group
- Pasteurella
- Bordetella
- Brucella
- Bartonella


What test/result can distinguish Enterobacteriaceae from other gram negative rods?

Oxidase Test
It is negative


What common diseases can Escherichia coli cause?

- particular strains cause most UTI's
- These strains are called uropathogenic because they possess virulence factors which enable them to colonise the urethra and survive in the urinary tract.

- nosocomial infections
- diarrhoeal illness (diarrhoeagenic strains)


Which bacteria can cause UTIs?

Gram -ve rods:
- Escherichia coli
- Proteus mirabilis



What virulence factors do certain Escherichia coli have?

- diarrhoeagenic
- uropathogenic


What are the key differences between community acquired and hospital acquired gram -ve rods?


Which bacteria cause typoid (enteric) fever?

• Salmonella Typhi
• Salmonella Paratyphi

Typhoid occurs when Salmonella invades the bowel wall to produce a serious systemic disease


What are the typical infections of non-fermantative gram negative rods?

NOTE: they tend to be inherently antibiotic resistant

NOSOCOMAL (hospital)/Community infections:
- Pseudomonas aeruginosa (acquatic or moist environments) - can also be community acquired
- Stenotrophomonas maltophilia (if patient is debilitated)
- Acinetobacter (and community acquired pneumonia)

• lower respiratory tract infections
• urinary tract infections
• surgical infections
• septicaemia
- otitis externa (P)
- chronic wounds (P)
- pneumonia (P)

Burkholderia pseudomallei.


What is the gram stain appearance of Neisseria gonorrhoea?

Gram- negative diplococcus


What are the usual manifestations of Neisseria gonorrhoea?

• Urethritis (males and females)
• Cervicitis (females)
• Pharyngitis
• Proctitis (anorectal infection)

- ascending infection can cause PID
- disseminated infection - arthritis, skin lesions

- purulent conjunctivitus in perinatal transmission


What are the important chlamydias?

- Chlamydia trachomatis
- Chlamydophila pneumoniae
- Chlamydophila psittaci


What is the first line of treatment for Neisseria gonorrhoea?

Rural WA - still penicillin

Metropolotian - The recommended first-line treatment in metro areas is a single dose of intramuscular ceftriaxone.

Some places in world - resistant even to ceftriaxone


What are some of the features of Chlamydiaceae from a cellular perspective?

• No peptidoglycan in cell wall, therefore:
- not seen on Gram stain
- not responsive to β-lactam antibiotics

• Obligate intracellular parasites
- can only grow in living cells
- energy parasites
- will not grow on artificial media


What are the main disease caused by various strains of Chlamydia trachomatis?

- chlamydia
- trachoma
- LGV (lymphogranuloma venereum) - elephantitis may result


What are the symptoms of chlamydia?

- urethritis
- epididymitis
- prostatitis

- urethritis
- cervicitis
- upper genital tract infection (PID – pelvic inflammatory disease - the latter is a major cause of infertility


What is the source of human infection with C. psittaci and what does it cause?

Bird contact

• abrupt onset of fever
• malaise
• myalgia
• dry cough
• pronounced headache
• sweats & rigors


Which bacteria causes syphilis?

Treponema pallidum


What are some of the features of Treponema pallidum from a cellular perspective?

• spiral morphology
• obligate intracellular pathogen – require a host cell,
cannot be grown in vitro on artificial media
• cannot be seen on Gram stain


What are the causes of syphilis?

1. Congenital

2. Venereal
Acquired by direct skin or mucous membrane contact, usually anogenital, with an individual in the primary or secondary stage of the disease.


What are the stages of syphilis?

1. Primary
3-10 weeks

2. Secondary
3-6 weeks
a) early - year (NOT INFECTIOUS)

3. Tertiary
- years to decades. 25% of untreated cases.


What are the main clinical features of primary syphilis?

Chancre (painless ulcer with raised firm edges)


What are the main clinical features of secondary syphilis?

- typically begins with skin rash - often palms or soles

- condyloma lata - large broad-based gray or white warty lesions on moist regions
Highly infectious

Mucous patches
• ulcerated areas of mucosal breakdown
• tongue, palate, tonsils, inside lips and cheeks, vulva, vagina
• highly infectious

Systemic features:
• fever
• pharyngitis
• muscle aches, joint aches
• fatigue
• lymphadenopathy
• headache, aseptic meningitis with 7th & 8th cranial nerve palsies
• weight loss
• patchy hair loss


What are the three possible forms of tertiary syphilis?

1. Central nervous system syphilis – the form most commonly seen now:
- progressive dementia
- tabes dorsalis
2. Cardiovascular syphilis
3. Gummatous syphilis – least common form in the antibiotic era


What are the key cellular differences between mycoplasmas (and Ureaplasmas) and other bacteria?

- smallest form of free-living life
- They are too small to be visualised using light microscopy and they have no peptidogycan in their cell envelopes.
- They can be grown on agar media in laboratories


What are the genital mycoplasmas?

• Mycoplasma hominis
• Mycoplasma genitalium
• Ureaplasma urealyticum


What is the most important mycoplasma and what does it do?

Mycoplasma pneumoniae

It causes the following types of infections:
• tracheobronchitis
• bronchiolitis
• pneumonia
These may be accompanied by upper respiratory involvement:
• pharyngitis
• otitis media

"Walking pneumonia"


What is the broad term for fungal diseases?



Who is susceptible to opportunistic mycoses?

patients with impaired immunity – localised impairment or generalised impairment.


What is the difference between deep-seated and opportunistic mycoses?

The immune system of the host

NOT whether it is deep in the body


What are the causes of opportunistic mycoses?

1. Filamentous saprophytic fungi
• Aspergillosis
• Zygomycosis (= mucormycosis)
• Pheohyphomycosis
• Hyalohyphomycosis

2. Candida

3. Pneumocysts jiroveci


What are the manifestations of invasive pulmonary aspergillosis?

• in the severely immunocompromised • necrotising bronchopneumonia
• septic emboli to other organs (brain) • high mortality


What are the manifestations of invasive disseminated aspergillosis?

• in the severely immunocompromised
• to many other organs


What are the host factors that make patients susceptible to candidaemia?

- debilitation, recent surgery, immunosuppression
- recent or current antibiotics
- intravascular cannulas (portals of entry)
- concurrent skin or mucous membrane candidiasis
- patient on TPN (total parenteral nutrition)


What are the clinical manifestations of systemic candidiasis?

Once in the bloodstream, Candida can settle out into virtually any tissue or organ where it produces:
- nodules,
- abscesses and
- tissue destruction.


What type of funds is Cryptococcus neoformans􏰉?

Yeast with a very large capsule.


How is Cryptococcus acquired?

Infection is acquired by inhalation and is usually asymptomatic.

- can occur in any immune classification


How does Cryptococcus manifest?

- Pulmonary cryptococcosis
􏰺􏰉 a spectrum of features
􏰺􏰉 propensity to metastasize
to brain

- CNS cryptococcosis
􏰺􏰉 meningoencephalitis
􏰺􏰉 acute or insidious onset
􏰺􏰉 infection may not be suspected

- Cutaneous cryptococcosis
􏰺􏰉 nodular skin lesions
􏰺􏰉 few or many


What would you use India Ink for and how does it work?


This fungus is a yeast and is characterised by having a very large capsule. In clinical samples, the capsule can be microscopically visualised if the sample is put on a glass slide with India ink. The ink blocks out the light except where it is transmitted through the capsules.


What is the main cause of travellers' diarrhoea?



What are the main causes of febrile illnesses in returned travellers?

- malaria
- typhoid
- dengue


Name the genus and 5 species that cause malaria

Genus: Plasmodium

- P. falciparum
- P. vivax
- P. ovale
- P. malariae
- P. knowlesi


Name the vector genus of malaria

Anopheles mosquito (females only)


Outline the life cycles of Plasmodium in the human and the mosquito

In mosquito:
- macrogametocyte
- ookinete
- oocyst
- cyst ruptures which releases sporozoites
- injects into person

In person:
- goes to liver and infects liver cells
- schizont
- schizont ruptures and immature trophozoite is in blood
- gametocytes
- taken up by mosquito


Name the stages of Plasmodium in the human life cycle

- human liver stages
- human blood stages


Which Plasmodium species produce hypnozoites

P. vivax and P. ovale

Some of the sporozoites infecting the liver, instead of undergoing schizogony, develop into dormant forms called hypnozoites.

Hypnozoites can remain dormant for months or years, producing relapses of clinical disease when they “awake” to undergo schizogony and produce merozoites. Hypnozoites are not killed by the drugs routinely used to treat malaria and people infected with P. vivax or P. ovale require a special additional drug for this purpose.


What could cause traveller's diarrhoea?

1. Bacteria
i.e. E.coli, salmonella, shidella etc

2. Viruses
- rotavirus
- enteric adenovirus

3. Parasite
- protozoa
- round worms - nematodes
- tape worms - cestodes
- flukes - trematods


What is dysentery and what are the main causes?

infection of the intestines resulting in severe diarrhoea with the presence of blood and mucus in the faeces.

Main Causes:
- Shigella
- Entamoeba histolytica
- Campylobacter


What are the 3 main groups of helminths that infect humans?

􏰻􏰉 Cestodes (tapeworms from animals)

􏰻􏰉 Nematodes (roundworms)
- Hook worm = Necator, Ancyclostoma
- Strongyloids
- Enterobius (pinworm/threadworm)
- Trichuris (whipworm)
- Ascaris (giant round worm)

􏰻􏰉 Trematodes (flukes / flatworms) - lifecycle in water
- Schistosoma


What are Enterobius, Trichuris and Ascaris?

Modes of transmission?

Clinical features of infections?

Enterobius - pinworm/threadworm
Trichuris - whipworm
Ascaris - giant round worm

Transmission: faecal-oral: ingestion of ova. Adult worms develop in intestine, shed ova.

gastrointestinal problems


What is the mode of transmutation of a hookworm and strongyloides?

larvae penetrate intact skin. Adults develop in intestine of humans, dogs, cats. Ova shed in faeces.


Describe the consequences of heavy hookworm infestation in a malnourished child

A person with a heavy infection may experience abdominal pain, diarrhea, loss of appetite, weight loss, fatigue and anemia. The physical and cognitive growth of children can be affected. Even death.


Outline the life cycle of Schistosoma mansion and the disease it produces


- Larvae penetrate intact skin.
- Adult worms develop in venous plexuses of bladder or bowel.
- Ova shed in urine or faeces into water.
- Eggs hatch in water,
- larvae penetrate freshwater snails (intermediate host).

Disease: Schistosomiasis. The urinary tract or the intestines may be infected.