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4.1 Medical Microbiology > Bloodstream Infections > Flashcards

Bloodstream Infections Flashcards

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1
Q

Why should blood normally be sterile?

A

Lysozyme
Leucocytes
Immunoglobuline (sIgA)
Complement

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2
Q

What is a transient bloodstream infection?

A

This is wherby bacteria can enter the bloodsteam but only for a few minutes
They are quickly moped up by the leucocytes etc

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3
Q

What causes a transient blood infection to become a bloodstream infection?

A

If host defences are overwhelmed or evaded

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4
Q

What is the mortality associated with bloodstream infections?

A

Between 10 and 40%

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5
Q

Other than the high mortality rate associated with BSI, what is the main major concern with BSI?

A

Patients who survive sepsis have a x2 times risk of dying in the next 5 years

Patiens usually suffer physical and cognitive impairment after survival

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6
Q

What is the mortality of BSI based on?

A

It is based on what organism you are infected with vs host immune system

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7
Q

What are the four ways that a bloodstream infection can occur, give examples of each?

A

A focal point of infection within the body outside of the blood (most common) -> e.g. a UTI (urosepsis) or S. pneumonia pneumonia spread to blood

Normal flora of the skin/mucous membrane such as S. aureus enter through cuts or wounds/burns etc

Gut flora enter through GIT perforation, infection or diverticulitis e.g. E. Coli

Direct introduction e.g. IV drug users, intravenous catheters or contaminated medical equipment

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8
Q

What are the different names for BSIs, what do they mean, what one should you use?

A

Bacteraemia - just means bacteria present in the blood

Septicaemia - focuses on clinical symptoms

BSI should be used

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9
Q

What are the different names for BSIs, what do they mean, what one should you use?

A

Bacteraemia - just means bacteria present in the blood

Septicaemia - focuses on clinical symptoms

BSI should be used

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10
Q

What are the four steps to a bloodstream infection?

A

Systemic Inflammatory Response Syndrome (SIRS)

Sepsis

Sever Sepsis

Septic Shock

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11
Q

What is SIRS?

A

Systemic Inflammatory Response Syndrome

The initiation of clinical symptoms due to immune response to toxic bacteria or to their products

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12
Q

What is sepsis?

A

This is where there is clinical evidence of infection along with a systemic response

Two of the following:
- Fever
- Tachycardia
- Tachypnoea
- White cell count >12,000

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13
Q

What is sever sepsis?

A

Sepsis plus evidence of abnormal perfusion of any organ, indicating more sever organ dysfunction

i.e. sepsis + spread to one other organ

This is where we see te 5 year mortality

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14
Q

What is septic shock?

A

This is where patients with septic syndrome develop hypotension

This represents a critical stage that requires immediate intervention or the patient will die

This is one of the biggest hospital emergencies

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15
Q

What causes septic shock, what organisms, how does it happen?

A

It is most frequently seen with gram negative BSIs but less commonly can be seen in gram-positives

Gram negatives:
- Lipid A portion of endotoxin triggers a series of reactions including production of TNF, IL-1 and complement which contribute to shock response

Gram positives:
- Associated with pneumococal infections (GAS) or staphylococcal BSIs

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16
Q

What are the three types of BSIs?

A

Transient Bloodstream Infections

Intermittent Bloodstream infections

Continuous Bloodstream Infections

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17
Q

Define a transient bloodstream infection, when would you see it

A

The temporary presence of bacteria in the bloodstream lasting less than 20 minutes

Self limiting - may not cause symptoms

Often seen after dental work e.g. S. viridans, localised infections such as pneumococcal pneumonia, dental extraction or chewin with poor dental hygiene, or intravenous drug abuse

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18
Q

Define an intermittent BSI

A

Bacteria present in the bloodstream at irregular intervals

Lasts for hours or longer, occuring at specific times

Symptoms may recur with each episode of bacteraemia

E.g. undrained intra-abdominal abscesses, earlly in the course of a variety of systemic and localised infections such as pneumococcal pneumonia

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19
Q

Define continuous BSI, give some examples

A

The persistant presence of bacteria in the bloodstream, suggesting a severe infection that has overwhelmed host defences

Its ongoing and wont go away without treatment

Symptoms are persistant and can lead to sever illness

Infections related to catheters, S. aureus, HIV or cancer showing ongoing bacteraemia

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20
Q

What are the four most common infections that cause sepsis in adults?

A

36% of patients had a lung infection
25% had a UTI
11% had a gut infection
11% had a skin infection

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21
Q

What sepsis is associated with oropharynx carriage?

A

N. meningitidis colonises the oropharynx
This can spread to blood = sepsis
This can then invade meninges to cause meningitis

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22
Q

What sepsis is associated with oral carriage?

A

Dentistry manipulation of teeth can lead to oral streptococci entering the blood

Occult bacteraemia -> can set up infection in damaged heart valves

results in endocarditis

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23
Q

What sepsis is associated with long-term central line catheters?

A

Coagulase-negative staphylococci
These can form biofilms and give rise to a continuous bacteraemia
-> line needs to be removed

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24
Q

What sepsis can be associated with bacterial pneumonia?

A

S. pnemonia caused pneumonia
Results in a transient bacteraemia

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25
Q

What sepsis is associated with cellulitis

A

Staphylococcus aureus

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26
Q

What sepsis is associated with liver abscesses

A

Liver abscesses tend to be polymicrobial -> usually sourced from appendix
Can cause a ‘dirty’ poymicrobial bacteraeia -> coliforms and such

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27
Q

What sepsis is associated with pyelonephritis

A

Urosepsis

E. Coli

Can cause septic shock (think Gram neg)

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28
Q

What kind of sepsis is associated with necrotisin fasciitis?

A

Group A streptococcus

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29
Q

What are the six most common causes of community acquired BSI in the immuno-cometent?

A

E. Coli (UTIs)
S. pneumoniae (pneumoniae)
S. aures (skin infections)
Enterobacteriaeceae
Neisseria (nasal colonisation)
B-haemalytic strep (S. pyogenes)

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30
Q

What are the nine most common causes of healtchare acquired BSIs in the immuno-competent

A

E. Coli (UTIs)
S. aureus (skin infections)
Enterobaceraceae -> Klebsiella
CNS -> biofilms/catheters etc
P. aeruginosa -> same as meningitis -> Beaumont etc
Enterococcus
Anaerobes -> Clostridiodes
S. pneumo -> pneumonia
Yeast -> candida/aspergillus

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31
Q

What are the some BSI pathogens associated with the immuno-compromised

A

Same organisms as in immuno-competent +
- L. monocytogenes (elderly)
- Corynebacterium sp
- Candida species + other fungi
- Mycobacterium

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32
Q

What are the some BSI pathogens associated with the immuno-compromised

A

Same organisms as in immuno-competent +
- L. monocytogenes (elderly)
- Corynebacterium sp
- Candida species + other fungi
- Mycobacterium

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33
Q

What is the most common ddrug/treatment associated with BSIs?

A

Chemo -> constant bacteramiae seen

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34
Q

What is a communinty associateddd BSI?

A

A BSI which occurs in an individual who was previously healthy

They are typically linked to focal infections:
- urinary tract infections - urosepsis
- pneumococal pneumonia - spread from lung
- skin and soft tissue infetion - staph

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35
Q

What is one of the reasons why S. aureus BSIs are so common?

A

S. aureus focal point might not be evident
S. aureus can cause osteomyelitis -> skin infection -> spread to bone, this infection might not be detectable i.e. might be treated and thought cleared -> metastatic spread from bone to blood etc etc

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36
Q

What are the relative frequencies of each causative organism of CA BSI in adults?

A

E. Coli = 25% -> NB, most common in Mater
S. pneumo = 22%
S. viridans = 10%
S. aureus = 10%
Others (Haemophilus, Neisseria, GNBs etc) = 17%

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37
Q

What are the four most common CA BSIs in children

A

S. pneumo (pneumonia)
N. meningitidis (carriage?)
S. aureus ( skin infections)
E. Coli

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38
Q

What has been the most significant change in CA BSI in children over the last few years?

A

Since the introduction of the Hib vaccine, H. influenza associated BSIs have decreased significantly

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39
Q

What are some reasons for the increase in HA BSIs, give an example of a scenario explaining this?

A

Increase in invasive procedures

Prolonged hospital stays

Foreign bodies such as catheters/central lines etc

e.g. a healthy pregnant woman comes into hospital for a C section (invasive procedure), she will now stay 5 days recovery in hosp (prolonged stay), a catheter will be put in (foregin body) -> patient now at very high risk of BSI

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40
Q

What are five invasive procedures with the highest risk of BSI?

A

Endoscopy: 0-20%
- CoNS, streptococci, diptheroids
- ?perforates bowel

Colonoscopy: 0-20%
- E. coli, Bacteroides species
- ? perforates bowel

Barium enema: 0-20%
= Enterococci, aerobes + anaerobic GNBs
- ? perforates bowel

Dental extraction: 40-100%
- transient BSI
- GBS = S. viridans

Prostate Transurethral resection: 0-40%
- Coliforms, enterococci, s. aureus
-

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41
Q

Comment on the rates of HCA BSI in ireland

A

2012 = 13%
2017 = 10%
2023 = 10.4%

*HCA BSI increasing slightly over last 5 years

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42
Q

What is the breakdown of the causes of BSIs in Ireland, origins of BSIs?

A

31/64 = Primary infections due to catheters

33/64 = Secondary infections due to focal infections:
- 33% = UTIs
- 24% = DIG
- 12% = Skin/Soft TI
- 12% = Surgical site infections
- 9% = pulmonary infection
- 9% = oher infections

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43
Q

Why are CNS BSIs often difficult to diagnose?

A

CNS are commensals of the skin
=> hard to determine if a CNS is a true infecion or just a commensal contaminant => hence why we take 3 sets of BCBs

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44
Q

How are E. Coli, S. aureus, S. pneumo, Enterococci, GNBs HCA BSIs trending?

A

E. Coli trending upwards
S. aureus flatlining
S. pneumo was high prior to covid, drop during covid, rising again

Both Ent faecium and Faecalis were rising, faecium much higher than faecalis, faecium seems to have peaked

Klebsiella increasing, P. aeruginosa flatlines and acinetobacter steadily low numbers

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45
Q

Comment on E. Coli trends and relative resistant strains
(3)

A

Invasive E.Coli decreased by 7.7% from 2019 to 2023

Proportion of ESBL+ decreased from 11.3% to 8.7%
=> proportion of ESBL decreasing

Very few CPEs -> stable at 3-4 cases a year

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46
Q

Comment on S. aureus trends and relative resistance
(3)

A

S. aureus has increased by 9.2% from 2019-2023

MRSA has decreased to 9.7% -> lowest to date

MSSA BSIs are increasing though

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47
Q

Comment on K. pneumoniae trends and relative resistance

A

K. pneumonia decreased by 10% between 2019 and 2023

Slight decrease in ESBLs, quinolone resistance and MDRKP

Number and proportion of Carbapenem-resistant is stable -> overall low in Ireland but needs to be monitored

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48
Q

Comment on E. faecium trends and resistance

A

Since 2012 the proportion of VREfm BSI has been over 40% -> we used to be 3rd worst in Ireland

VRE BSI have decreased from a peak of 46% in 2015 down to 21.4% in 2023 -> massive decrease -> still above European average but way down

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49
Q

Comment on P. aeruginosa trends and resistance

A

P. aer BSI have been stable between 2019 and 2023

MDR have decreased from 6.4% in 2019 to 4.8% in 2023

Pseudo high before covid, droped, high again but lower than pre-covid

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50
Q

Comment on S. pneumo trends and resistance

A

Pre-pandemic high of 360 cases, drop to 179 in covid years, increase again

Number of penicillin resistant non wild type strains has decreased

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51
Q

Comment on Acinetobacter trends and resistance

A

There was no MDR Acinetobacter resistance detected in 2023 in Ireland

Major problem in Southern and Eastern Europe
- some countried reporting relative resistance of 75% or more
- what happens in southern Europe tends to happen to us

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52
Q

Comment on GAS trends

A

Between October 2022 and August 2023 there was an unusual and unreasonal upsurge in GAS disease, the majority of which in children (<18)

An established relationship between Varicella infection and subsequent GAS i.e. children who got chickenpox subsequently got GAS

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53
Q

What are the most common BSIs in newborns?

A

Group B streptococci (from mothers flora)
E. Coli
CNS
Candida

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54
Q

What are some issues with getting BSI samples for neonatal BSIs?

A

Specimen collection is so difficult
Small blood volume only
Contamination - samples nearly always contaminated

55
Q

Why has the spectrum of organisms we detect in immuno-compromised BSI changed over the years?

A

Lengthening periods of neutropenia and duration of hospital stay

Increased use of CVC

Increased use of broad-sepctrum antibiots -> chronically ill immuncompromised e.g chemo patients on long term broad spectrum

Were seeing a high incidence of infection with organisms that are gnerally non-virulent in the normal host and form part of the normal flora e.g. CNS, enterococci, candida etc etc

56
Q

What are five requirements of a good blood culture?

A

Utilise culture medium suitable for the recovery of small numbers of all potential pathogens including fastidious

Neutralise or remove antimicrobial substances e.g. resin/charcoal

Minimise risk of contamination

Facilitate earliest possible detection

Faciliate high specimen throughput

57
Q

What kind of systems are used for bood cultures?

A

Fully automated continuous monitoring systems e.g. BacT/Alert 3

58
Q

What is the principle of the BacT?

A

Colorimetric detection of CO2 production

59
Q

What is the maximum blood volume for the BacT?

A

10mmls

60
Q

What are some factors that affect pathogen recovery on automated BC systems?

A

Method of collection
Number and timing of samples

Volume of sample - underfilled/overfilled
Media used
Neutralisation of antimicrobial agents through resin
Incubation time and temperature
Agitation of media
Headspace atmosphere

61
Q

Why is the collecting of BCBs so important?

A

Need to collect three sets taken at three separate time points

Both aerobic and anaerobic bottles should be taken

Should be taken when temperature is peaking/on the rise

62
Q

How should a BSB sample be taken?

A

Collect blood sample via venepuncture site

Disinfect skin and septum of blood culture bottle with alcohol and air dry

Take blood sample and divide between aerobic/anaerobic )and fungal bottle if necessary)

63
Q

At what time should BCB be taken

A

Before antimicrobial therapy where possible
As soon as possible after a spike of fever
- except for query endocarditis

64
Q

What is different about taking a babies BCB

A

Blood can be taken from baby’s toe -> same applies to intravenous drug users with collapsed veins

65
Q

Why do we take 3 blood culture bottle sets?

A

These are taken to allow for recognition of contaminants

They are taken at 3 separate time points over several hours or days - this increases yield

66
Q

What are the most common BSI contaminants?

A

Coagulase-neg staphs
S. viridans
Diptheroids

67
Q

What is the most critical factor in blood culture collection?

A

Volume -> underfilled vs overfilled

68
Q

What is the most critical factor in blood culture collection?

A

Volume -> underfilled vs overfilled

69
Q

Why is volume the most critical factor in blood culture filling?

A

There is a direct relationship between blood volume and yield

20-30mls of blood are recommended for culture (total volume)

10mls of sample should be added to bottles

70
Q

Explain the relationship between blood volume and bacterial yield

A

Direct relationship

Per 1ml of blood there is a 3% increase in yield -> overfiled can result in too many leucocytes which can cause false positives??

71
Q

How many mls of neonatal blood needed for culture

A

Only 0.5-2.0mls needed

The magnitude of childhood BSI is usually higher than in adults

*Never take no more than 1% of a childs blood

72
Q

What is in a basic BC set?

A

Standard aerobic bottle + anaerobic bottle

Each contains 40mls of Tryptic Soy Broth Agae (TSB) Media +/- resin or activated charcoal

73
Q

What is in a basic BC set?

A

Standard aerobic bottle + anaerobic bottle

Each contains 40mls of Tryptic Soy Broth Agae (TSB) Media +/- resin or activated charcoal

74
Q

What is in a paediatric BC set?

A

20/30mls of peptrone enriched Tryptic Soy Broth agar
-> enriched TSB agar

Supplemented with brain/heart infusion solids (BHI) and activated charcoal

75
Q

Why do we enrich paed BC bottles with BHI solids and charcoal?

A

These improve microbial recovery by absorbing antimicrobials

76
Q

Why do we enrich paed BC bottles with BHI solids and charcoal?

A

These improve microbial recovery by absorbing antimicrobials

77
Q

What media is used for Mycobacteria?

A

Middlebrook 7H9 broth

78
Q

What is the ideal ratio of components of BC bottles?

A

Blood to broth ratio of 1:15
- needed to counteract antimicrobial effects

Blood to broth ratio of 1:5 with the addition of other specific agents that inhibit antibacterial properties

79
Q

What is the resin used in blood culture bottles?

A

Resin in the forms of free-floating beads

These mop up and eliminate any antibiotics that ma be present to increase sensitivity

They also provide a larger surface area for bacterial adhesion and replication

This facilitates lysis of host cells and the release of intracellular organisms

80
Q

What is the activated charcoal used in BCBs?

A

This enhances microbial recovery by absorbing antimicrobials creating a more favourable environment for bacterial growth

81
Q

What incubation is optimal for blood cultures?

A

35-37 degrees celsius for 5-7 days for routine cultures - optimal for most organisms

extended incubation if query endocarditis or Brucellosis/other fastidious organisms or fungi

82
Q

Why do a lot of places no longer extend blood cultures for query endocarditis anymore?

A

There has been a lot of research that proves this extenstion may be pointless

83
Q

Why do a lot of places no longer extend blood cultures for query endocarditis anymore?

A

There has been a lot of research that proves this extenstion may be pointless

84
Q

If you query a Brucella BSI what should you do?

A

10 days + extended incbation
Terminal subculture even if still negative

85
Q

What safety considerations should you take when processing blood cultures?

A

Subculture must be performed in a safety cabinet
-> if query brucella then in cat 3

sub-vent units for subculturing is preferred

Venting needles are now plastic and not metal as there was a lot of needle stick injuries with lab staff

86
Q

What care shouhld be taken between taking sample and loading it onto system?

A

Blood cultures should be transported to lab as soon as possible

Specimens should be loaded onto system as soon as possible

Sample can only be at room temperature for a max of 4 hours

Ambient temperature is preferable ro refrigeration

  • BCB not to be incubated at 37 degreses -> this can cause you to miss a positive
87
Q

What care shouhld be taken between taking sample and loading it onto system?

A

Blood cultures should be transported to lab as soon as possible

Specimens should be loaded onto system as soon as possible

Sample can only be at room temperature for a max of 4 hours

Ambient temperature is preferable ro refrigeration

  • BCB not to be incubated at 37 degreses -> this can cause you to miss a positive
88
Q

What did the BacT virtuo do with negative bottles?

A

If a bottle was negative after 5 days incubation the virtuo would simply throw the bottle off

Virtuo would let the LIS system know that the bottle was negative

Scientist will just have to report it

89
Q

What is the main reason for having to reject blood culture bottles?

A

Doctors tend to send down blood cultures in batches

This means a bottle could have been left at room temp for more than 4 hours by the time we receive it in the lab

90
Q

How long should you leave a positive blood culture on the bacT, why is this significant?

A

Never leave a blood culture for more than 4 hours on the BacT

Certain organisms such as S. pneumo will undergo autolysis - this will result in a negative gram from these positive cultures

91
Q

What is done with positive blood cultures?

A

Gram and culture

92
Q

How do you gram and culture directly from a blood culture bottle?

A

Invert the positive bottle gently to mix contents

Wipe the septum of the bottle with 70% ethanol and allow to air dry

Using the safety, venting unit inoculate the agar plates: Blood, Chocolate, MH and MacConkey

Add your ring of blood cultture antimicrobials to the muller hinton plate

Inoculate a clean microscopy slide and allow to dry for gram

93
Q

What culture plates are put up or blood cultures

A

Always blood anaerobically with MTZ and chocolate in CO2 both at 37 degrees and ready daily

Additional plates include Muller Hinton, MacConkey

Candida chrom agar or malt agar for query yeast

Chromagar for query GNB

Bile aesculin for query enterococci

etc etc

94
Q

What do we do for a query staph in blood culture?

A

MH + cefoxitin

MRSA = cefoxtin resitant

95
Q

What do we do with a query S. pneumo in blood culture?

A

Slide latex pneumo test directly from blood culture

Can put up blood + optochin

96
Q

What antimicrobials are put up on Muller Hinton agar, why these?

A

Cefoxitin
Vancomycin
Ciprofloxacin
CPD (cefpodoxime)
Meropenem
TZP (piercillin/tazobactam)

These are not for a treatment plan they are just for notifying resistance

97
Q

How do we carry out AST for blood cultures?
(3)

A

We do AST straight from blood culture bottles, no subbing - direct AST

We cannot standardise the inoculum when using drops of blood so AST is only for guidance - prelim report

All suceptibility is repeated the following day with bacterial colonies from culture either by disc diffusion or E test or on Vitek

98
Q

Why do we do AST directly from blood culture bottles, why is this significant?

A

it is done to save time due to the serious nature of a blood stream infection

We can have a preliminary guide report out a day earlier than full AST

99
Q

What should a scientist doo with a positive blood culture vs negative

A

Positive culture:
- if gram positive phone results ASAP
- issue preliminary culture report
- when culture complete issue a final authorised report (after maldi ID)

Negatives:
- negative report is issues at 5 days and 7 days
- negatve final report issues for suspected IE culures still negative at 21 days

100
Q

What are the two main reasons for false positive blood cultures?

A

Metabolic activity of WBCs

Lysis of bacteria e.g. autolysis of S. pneumo

101
Q

What happens with S. pneumo o blood cultures

A

They grow but then they start to autolyse

By the time you take an S. pneumo off and look at gram the gram might already be negative

You will still incubate the plates but these also might still be negativ

102
Q

How do you interpret a BacT growth chart?

A

A sigmoid curve = positive growth
A constantly increasing curve which doesnt flatline = high white cells
Flatline = negative

103
Q

How do you deal with false positives due to high white cells?

A

These bottles will prematurely flag positive

The bacT will unload these thinking they are positive

Scientist will have to read growth chart and determine high whites and reload bottle

Bottle will have to be repeatedly reloaded until 5 days incubation is complete - this can be a pain

104
Q

How often is contamination seen in blood culture bottles

A

Before Leen Six Sigma and scientists with LEEN belts there was a lot more contamination:
- Between 15 and 30% of bottles were contaminated

With leen processing we not have special blood culture kits with sterile gloves and more training etc which has greatly reduced contamination
- Now only between 2 and 5% of bottles

105
Q

What are the most common contaminating organisms?

A

Skin flora:
- CNS (70% of contamination)
- Diptheroids
- Bacillus species

106
Q

How can we determine if an organism is a contaminant or not?

A

The type of organism (is it a skin flora organism)

No of bottles positive -> across all bottles or just one set

How long did it take to go positive - 5 days usually for a contaminant but only 48hrs for a tru BSI

Clinical information - could this be infectious etc

107
Q

What is the gold standard for detection of BSI pathogens

A

Blood culture

108
Q

How sensitive are blood cultures?

A

Low positivity rate: only between 30-40%, 60% at best of BSI are blood culture positive

109
Q

Why is the positivity rate for blood cultures so low?

A

Mostly due to patients being on antimicrobial treatment

Slow growing fastidious organism

110
Q

Other then poor positivity rate, what is the main pitfall of blood cultures?

A

The slow TAT
It takes atleast 24 hours for a preliminary ID -> if culture positive within 24hours - but usually its 48
Takes an additional 24hours to have AST guide and another day to have full AST
- very slow TAT

111
Q

Why are so many blood culture samples contaminated with antimicrobials?

A

For every hour that the treatment of a BSI is delayed there is a 7% increase in patient mortality

Because of this clinicians are absolutely not waiting around for blood cultures, they are treating the patient ASAP

Even if culture negative the clinicians will still treat the patient if there is high enough suspicion

112
Q

What are the three novel approaches to BSIs being looked at?

A

FISH
MALDI-TOF
Multiplex PCR both direct and indirect

113
Q

What does FISH stand for?

A

Fluorescence In Situ Hybridisation

114
Q

What is FISH?

A

A molecular diagnostic technique used to detect bloodstream infections by identifying the presence of pathogens directly in blood culture samples

It works using fluorescntly labelled probes that specifically bind to unique sequences of the taget organisms RNA or DNA

115
Q

Talk about the use of FISH for blood cultures

A

Can detect 20-25 pathogen species responsible for over 90% of BSI

Uses fluorescently labelled oligonucleotide probes specific for 16S rRNA designed for most of these pathogens

It can improve the speed of pathogen identification, it is often used alongside other diagnostic methods to overcome these limitations - but still need to culture for AST

116
Q

What are the main pros of FISH for BSI detection

A

Rapid identification within hours after a positive BCB

Direct detection striaght from BCB

Highly specific probes

Very effective for fastidious/slow-growing organisms

Can detect bacteria and fungi

Complementary to other techniques such as MALDI-TOF and PCR

117
Q

What are the cons of FISH for BSI detection

A

Limited target range - cant detect 10% of organisms causative of BSIs - only has certain targets

Cannot detect antibiotic resistance - AST still needed

Requires a positive blood culture - 24-48 hours wasted

Higher cost - probes are expensive compared to agar

Skilled personnel needed to interpret results

False negatives due to genetic mutations in target regions - probes wont bind

Limited quantification - can only detect if present or now etc

118
Q

How can MALDI be used for Blood cultures

A

Maldi can be used in two ways:
- by IDing from actual isolates/colonies done on a quickly grown first inoclum
- Directly from positive blood culture

119
Q

How do we use the MALDI to ID organisms straight from blood culture?

A

Microboial cells are harvested from blood culture

Cells are processed using the MALDI Sepsityper Kit

Cells are then identified by the MALDI Biotyper 3.0

120
Q

What are the pros and cons for using MALDI-TOF for BC

A

Rapid TAT following flags positive 20-70mins

Still have to wait for growth in blood culture bottles (but not sub plate)

Have to culture for sensitivities

Sensitivity of 76% but specificity of 96%

121
Q

What organisms does the MALDI-TOF struggle to ID from blood cultures

A

CNS: 24-97% sens
Enterococcus: 50-70%
Strep: 32-50%

It also struggles with polymicrobial infections

122
Q

What are the main benefits of the MALDI-TOF for BC ID

A

reduced TAT
Relatively cheap, less than 25 euro a test

123
Q

In reality very few labs use the sepsityper kit with MALDI, what do we do instead

A

A lot of labs do short-term incubation on solid media for blood cultures

Blood cutures positive in morning - streaked early on - growth till later in the day

GPCs and GNBs only take about 6hours to start growing

First inoculum starts to grow after a fw hours - hazy growth - can be IDd from this

124
Q

What PCR do we use for blood cultures?

A

Film array BC Id panel (we didnt have this in the mater)

This was used in the maternity hospitals not so much clinical hospitals (only put up really at request of docs)

125
Q

What targets are there on the film array for BCs

A

24 targets total:
- 8 gram positive
- 11 gram negativ
- 5 yeast

2 antimicorbial resistance genes (mecA, vanA/B and bla kp)

126
Q

Compare the film array vs maldi-tof for bc id

A

Film array is more accurate (91% vs 81%)
Film array is quicker (+2.4hrs vs 2.9hrs after positive virtuo)
Film array can detect some resistance genes, maldi cannot
Film array is way more expensive (120 euro vs 25 euro)

127
Q

What PCR methods can be used to indirectly test on blood
(3)

A

Lightcycler SeptiFast Test

SepsiTest

IRIDICA BAC BSI assay

128
Q

Talk about the Lightcycler SeptiFast Test for Blood Cultures, what kind of PCR is it, pros and cons.

A

Designed to detect and identify the 25 most important bacterial and fungal species causing BSI within 6 hours

It is a 3x multiplex amplificatin reaction: gram pos master mix, gram neg master mix and fungal master mix

Only needs 1.5mls of blood to extract DNA

TAT of only 6 hours total!!

Expensive at about 200 euro a test

Can only detect expected pathogens

129
Q

Talk about the SepsiTest for direct BSI detection. what kind of sequencing, sample type, pros and cons, compare to light cycler

A

1ml of EDTA-treated whole blood
Universal PCR and Sanger sequencing
Id species >200 genera bacteria and 65 genera of fungi
Costs about 190 euro
TAT of 8 hours

(more sensitive than LightCycler, slightly less expensive, a little slower)

130
Q

Talk about the IRIDICA BAC BSI assay for direct BSI detection, sample type, PCR type, targets, pros/cons, compare to other methods

A

5mls of EDTA-treated whole blood
Combines broad-range PCR + electrospray ionisation MS
Also detects mecA, vanA, vamB and blKPC determinants
Detects over 780 bacterial and candidal species
Costs about 235 euros for test
TAT of 6 hours

Most amount of targets, but most expensive, short TAT though and can detect resistance

131
Q

Why do we not use Septifast or SepsiTest in Ireland

A

When compared to blood culture the Septifast had a sens of 50% while the sepsitest had a 48% sense
- very low sensitivity which didnt meet hospital criteria

132
Q

Why do we not use the IRIDICA in ireland

A

When compared to BC the IRIDICA had a sens of 81% -> good sens

IRIDICA has the potential to greatly decrease TAT but it is really expensive at the moment

There also hasnt been enough evidence to prove that this reduced TAT actually impacts the treatment of the patient -> patients treated with broad spectrum antibiotics usually anyways as soon as BC positive is phoned (if not before) -> still have to wait for AST to switch to different antibiotics

133
Q

In general what are the limitations of PCR for BSIs

A

Low diagnostic sensitivity
High cost
Interpretation problems
- CNS and streptococci (contaminants vs infectious)
- DNAemia but not a BSI
No proven benefit on patient treatment, mortality, hospital stay etc

134
Q

What are the potential benefits of direct PCR for BSIs

A

Its better than conventiona BC -> BC has low sensitivity - PCR isnt great sensitivtiy but it is better than this

PCR is much better for anti-microbial pre-treated patints
PCR is preferred for immuno-compromised patients
PCR is preferred for neonatal sepsis - quick TAT

PCR is best used when combined with positive blood cultures

4.1 Medical Microbiology (30 decks)

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