Hybrid POCT/Core Lab Approach Nam K. Tran, PhD, HCLD (ABB), FACB - - PowerPoint PPT Presentation

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Hybrid POCT/Core Lab Approach Nam K. Tran, PhD, HCLD (ABB), FACB - - PowerPoint PPT Presentation

Sep 13, 2022 144 likes •942 views

Detecting Sepsis via Molecular Testing Using a Hybrid POCT/Core Lab Approach Nam K. Tran, PhD, HCLD (ABB), FACB Associate Professor of Pathology and Laboratory Medicine Director of Clinical Chemistry, Special Chemistry/Toxicology, and POCT

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SLIDE 1

Detecting Sepsis via Molecular Testing Using a Hybrid POCT/Core Lab Approach

Nam K. Tran, PhD, HCLD (ABB), FACB

Associate Professor of Pathology and Laboratory Medicine Director of Clinical Chemistry, Special Chemistry/Toxicology, and POCT
  • Dept. of Pathology and Laboratory Medicine
University of California, Davis Health
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SLIDE 2

Learning Objectives

  • Describe current challenges in sepsis recognition, pathogen detection, and

management.

  • Identify the strengths and weakness of microbiological techniques.
  • Describe the types of rapid pathogen detection systems available.
  • Identify potential roles for point-of-care molecular pathogen detection and the

concept of the “hybrid” laboratory.

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SLIDE 3

Sepsis: The Clinical Problem

  • Sepsis is defined as life-threatening organ dysfunction caused by a dysregulated

host response to infection

Kost GJ, Tang Z, Tran NK, et al. Scand J Clin Lab Invest 2003;63:15

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SLIDE 4

Sepsis: The Clinical Problem

  • Sepsis is defined as life-threatening organ dysfunction caused by a dysregulated

host response to infection

  • Over 750,000 patients in the United States experience sepsis each year.
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SLIDE 5

Sepsis: The Clinical Problem

  • Sepsis is defined as life-threatening organ dysfunction caused by a dysregulated

host response to infection

  • Over 750,000 patients in the United States experience sepsis each year.
  • Mortality ranges from 28-50% and can be as high as 90% in cases of septic

shock. https://business.kaiserpermanente.org/

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SLIDE 6

Sepsis: The Definition Problem

  • Sepsis definitions evolving – highlights the complexity of the disease process.
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SLIDE 7

Sepsis: The Definition Problem

  • Sepsis definitions evolving – highlights the complexity of the disease process.
  • Children vs. adults are different, high risk patients vs. everyone else (?)
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SLIDE 8

Sepsis: The Definition Problem

  • Sepsis definitions evolving – highlights the complexity of the disease process.
  • Children vs. adults are different, high risk patients vs. everyone else (?)

https://www.acepnow.com/article/acep-endorses-latest-surviving-sepsis-campaign- recommendations/?singlepage=1&theme=print-friendly

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SLIDE 9

Sepsis: Antimicrobial Problem

Empiric antimicrobials necessary since time matters in sepsis. Odds of non-survival increases by 7.6% for every hour delay in treating ”severe sepsis”.

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SLIDE 10

Sepsis: Antimicrobial Problem

Empiric antimicrobials necessary since time matters in sepsis. Odds of non-survival increases by 7.6% for every hour delay in treating ”severe sepsis”.

Unnecessary use of antimicrobials leads to: ▪ Antimicrobial resistance,

  • C. difficile colitis, ESBL, CRE

▪ Toxicities and adverse drug events ▪ Increased morbidity and longer hospital stays ▪ Delays in starting appropriate antibiotic ▪ Reduced cost-effectiveness of health care delivery

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SLIDE 11

Sepsis: Antimicrobial Problem

Empiric antimicrobials necessary since time matters in sepsis. Odds of non-survival increases by 7.6% for every hour delay in treating ”severe sepsis”.

  • f antimicrobial use in acute

care hospitals is unnecessary

UP TO

50%

Unnecessary use of antimicrobials leads to: ▪ Antimicrobial resistance,

  • C. difficile colitis, ESBL, CRE

▪ Toxicities and adverse drug events ▪ Increased morbidity and longer hospital stays ▪ Delays in starting appropriate antibiotic ▪ Reduced cost-effectiveness of health care delivery

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SLIDE 12

How Antimicrobial Resistance Spreads

Overuse and non- judicious prescription exerts antimicrobial pressure to promote resistance!

  • Realization that Urgent

Care Centers lack any stewardship practices.

  • Patients often ask for

antimicrobials without medical background and physicians comply

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SLIDE 13

Sepsis: Pathogen Detection Problem

Rapid pathogen detection is the ”common denominator” for sepsis. Early pathogen recognition accelerates treatment appropriate decisions and improves outcomes. Unfortunately...

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SLIDE 14

Sepsis: Pathogen Detection Problem

Rapid pathogen detection is the ”common denominator” for sepsis. Early pathogen recognition accelerates treatment appropriate decisions and improves outcomes. Unfortunately... Microbiological culture remains the primary means for pathogen detection.

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SLIDE 15

Sepsis: Pathogen Detection Problem

Rapid pathogen detection is the ”common denominator” for sepsis. Early pathogen recognition accelerates treatment appropriate decisions and improves outcomes. Unfortunately... Microbiological culture remains the primary means for pathogen detection.

  • Blood culture detection limits range from 3.2 to 3,000

CFU/mL

  • In theory detects anything that grows in the specific

media.

  • Results may be affected by antimicrobial therapy.
  • Median analytical turnaround time (TAT) not

compatible with efforts for early recognition. ✓ Collection → Gram Stain: 10.4 hours ✓ Collection → Speciation: 26.4 hours ✓ Collection → MIC: 43.7 hours

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SLIDE 16

“Modern” Microbiology Laboratory

Microbiology hasn’t changed too much → concept remains the same up until recently. Grow the pathogen and determine the phenotype. Biochemical Testing

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SLIDE 17

“Modern” Microbiology Laboratory

Microbiology hasn’t changed too much → concept remains the same up until recently. Grow the pathogen and determine the phenotype. Antimicrobial Susceptibility Testing Biochemical Testing

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SLIDE 18

Patient is a 20 year old man status post motor vehicle accident with 90% TBSA 3rd and 4th degree burns and C1 pedicle and C4 foraminal fracture.

Day 1 2 3 4 8 23 30 31 32 33 37 38 39

BCA: Negative RC1: Collected RC1: H. influenzae WC1: Collected

15

WC1: MSSA, E. faecalis, Strep. viridans Added Linezolid

10

RC2: Collected Mold observed during dressing change. WC2: Collected Discontinued Linezolid, Meropenem; Added Vancomycin, WC2: A. fumigatus, Rhizopus sp. RC3: Collected Ampho B soaks

27

RC3: Collected BCB: Collected Added Voriconazole, Meropenem BCB: P. aeruginosa RC3: P. aeruginosa Septic Shock MAP: 40-50mmHg Started 4 Vasopressors Green exudate on wounds Platelet: 88,000 BCC: Negative +Tobramycin Patient expired Epinephrine Started Ceftazidime Added Posaconazole

Pathogen Detection in Burn Patients

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SLIDE 19

“Modern” Microbiology Laboratory

Microbiology hasn’t changed too much → concept remains the same up until recently. Grow the pathogen and determine the phenotype. Antimicrobial Susceptibility Testing Biochemical Testing

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SLIDE 20

Modern Microbiology Laboratory

Over the last 10 years, there’s been new innovations that have helped overcome the microbiology “TAT” problem. This includes automation, mass spec, and molecular diagnostics. Automated Culture Plating Antimicrobial Susceptibility Testing

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SLIDE 21

Modern Microbiology Laboratory

Automated Culture Plating Antimicrobial Susceptibility Testing MALDI-TOF-MS Over the last 10 years, there’s been new innovations that have helped overcome the microbiology “TAT” problem. This includes automation, mass spec, and molecular diagnostics.

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SLIDE 22

Modern Microbiology Laboratory

Automated Culture Plating Antimicrobial Susceptibility Testing MALDI-TOF-MS Molecular Dx Species ID Resistance (?) Over the last 10 years, there’s been new innovations that have helped overcome the microbiology “TAT” problem. This includes automation, mass spec, and molecular diagnostics.

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SLIDE 23

Modern Microbiology Laboratory

Automated Culture Plating Antimicrobial Susceptibility Testing MALDI-TOF-MS Molecular Dx Species ID Resistance (?) Over the last 10 years, there’s been new innovations that have helped overcome the microbiology “TAT” problem. This includes automation, mass spec, and molecular diagnostics.

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SLIDE 24

Rapid Pathogen Detection: The Promise of Molecular Diagnostics

  • Molecular approaches such as PCR offer highly sensitive and specific alternatives

to existing microbiological tests.

  • Provides potential to pick up certain resistance genes (e.g., mecA, kpc, NDM-1, etc)
  • Multiplex system scan detect up to 22 viruses and bacteria in 45 mins to 60 minutes

depending on the platform.

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SLIDE 25

Rapid Pathogen Detection: The Promise of Molecular Diagnostics

Challenges:

✓ Current systems test directly from culture positive specimens rather than whole blood. ✓ Throughput limited and high upfront cost for individual instruments limit use at the enterprise-wide level. ✓ High cost per multiplex test: ~$100/test and billable to the patient could be thousands of dollars! ✓ Majority of pathogens are not needed.

  • Molecular approaches such as PCR offer highly sensitive and specific alternatives

to existing microbiological tests.

  • Provides potential to pick up certain resistance genes (e.g., mecA, kpc, NDM-1, etc)
  • Multiplex system scan detect up to 22 viruses and bacteria in 45 mins to 60 minutes

depending on the platform.

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SLIDE 26

Rapid Pathogen Detection: The Promise of Molecular Diagnostics

Challenges:

✓ Current systems test directly from culture positive specimens rather than whole blood. ✓ Throughput limited and high upfront cost for individual instruments limit use at the enterprise-wide level. ✓ High cost per multiplex test: ~$100/test and billable to the patient could be thousands of dollars! ✓ Majority of pathogens are not needed.

  • Molecular approaches such as PCR offer highly sensitive and specific alternatives

to existing microbiological tests.

  • Provides potential to pick up certain resistance genes (e.g., mecA, kpc, NDM-1, etc)
  • Multiplex system scan detect up to 22 viruses and bacteria in 45 mins to 60 minutes

depending on the platform.

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SLIDE 27

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

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SLIDE 28

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

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SLIDE 29

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

Growth Time (10.4 hrs)

PCT Lactate IL-6 (?) CRP (?)

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SLIDE 30

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

Growth Time (10.4 hrs)

PCT Lactate IL-6 (?) CRP (?)

MALDI-TOF-MS (10 mins) Molecular Testing (1-2.5 hour) AST (24 hours)

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SLIDE 31

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

Growth Time (10.4 hrs)

PCT Lactate IL-6 (?) CRP (?)

MALDI-TOF-MS (10 mins) Molecular Testing (1-2.5 hour) AST (24 hours) Diagnostic Blind Zone Species ID Genotypic Resistance Phenotypic Resistance

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SLIDE 32

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

Growth Time (10.4 hrs)

PCT Lactate IL-6 (?) CRP (?)

MALDI-TOF-MS (10 mins) Molecular Testing (1-2.5 hour) AST (24 hours) Diagnostic Blind Zone Species ID Genotypic Resistance Phenotypic Resistance

We can conceivably get species ID and genetic resistance data for select targets within 24 hours now

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SLIDE 33

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

Growth Time (10.4 hrs)

PCT Lactate IL-6 (?) CRP (?)

MALDI-TOF-MS (10 mins) Molecular Testing (1-2.5 hour) AST (24 hours) Diagnostic Blind Zone Species ID Genotypic Resistance Phenotypic Resistance

However genotypic resistance may not always = phenotypic resistance.

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SLIDE 34

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

Growth Time (10.4 hrs)

PCT Lactate IL-6 (?) CRP (?)

MALDI-TOF-MS (10 mins) Molecular Testing (1-2.5 hour) AST (24 hours) Diagnostic Blind Zone Species ID Genotypic Resistance Phenotypic Resistance

A diagnostic blind spot still remains due to needing cultures to grow out as well

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SLIDE 35

Timeline for Testing Today:

Molecular Enhanced Pathogen Detection

Growth Time (10.4 hrs)

PCT Lactate IL-6 (?) CRP (?)

MALDI-TOF-MS (10 mins) Molecular Testing (1-2.5 hour) AST (24 hours) Diagnostic Blind Zone Species ID Genotypic Resistance Phenotypic Resistance

Is there clinical value getting faster than this?

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SLIDE 36

ABA – MCTG COMBAT CASUALTY GRANT:

“Rapid, Quantitative, PCR-Based Detection of Staphylococcus aureus in Burn Sepsis Patients”

PI: Nam K. Tran, PhD

NIH Clinical Trials Registration Number: NCT01140269 UCD IRB Approval Number: 200918586 USAMRMC HRPO Log Number: A-15774.0 (Core Protocol)

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SLIDE 37

Study Model: Randomized Controlled Trial

RECRUITMENT BLOCK RANDOMIZATION (240 Patients) Inclusion Criteria Exclusion Criteria Age18 years Age<18 years 20% TBSA burns <20% TBSA burns Unable to consent IV Antibiotic allergies Non-survivable injuries CONTROL Observational group No PCR testing Routine laboratory testing Standard of care treatment EXPERIMENTAL Treatment group PCR testing for Staphylococcus aureus Routine laboratory testing Standard of care treatment Quantitation of positive PCR results (blinded) CONTROL (120) EXPERIMENTAL (120)

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SLIDE 38

PARTICIPATING SITES

UCDMC Sacramento, CA Torrance Memorial Torrance, CA

  • U. Cincinnati

Cincinnati, OH

  • U. Miami

Miami, FL Nathan Speare Philadelphia, PA

  • U. Washington

Seattle, WA

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SLIDE 39

GeneXpert PCR Near-Patient Testing

RESULTS Time (min) 2 60 - 70 Nasal Wound BC

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SLIDE 40

Staphylococcus aureus

▪ Gram positive cocci found in groups. ▪ Coagulase and catalase positive ▪ Produces capsules (types 5 and 8 are common human pathogens) ▪ Expresses beta-lactamase to confer penicillin resistance ▪ Colonizes 10 to 20% of adults ▪ Methicillin resistant strains (MRSA) associated with higher mortality.

Lowy FD. N Eng J Med 1998;339:520-532.

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SLIDE 41

Proof-of-Concept: Serial Quantitative PCR Testing

History: Patient is a 40 year old man with 20% total body surface area burns to the face, head, neck, left upper back, bilateral hands, and lower left extremity from a house fire. Blood cultures, respiratory cultures, and wound cultures were collected on day 5 for clinical suspicion of burn sepsis (American Burn Association Sepsis Trial) Day 5 Day 6 Day 7 Day 8 Day 9 0945: Vancomycin Started 750mg IV q8H 1900: Wound culture (WC) Collected PCR swab sample collected 2005: PCR detects 1,029 CFU of MSSA from wound swab 1,380 CFU of MSSA 2030: Mupirocin started on wounds 792 CFU of MSSA 661 CFU of S. aureus 0934: WC report 2+ Gram Positive Cocci 1015: WC report S. aureus (MIC pending) 1205: WC report MSSA Point-of-care PCR testing provided definitive results 4 days faster than

  • culture. Quantitative PCR correlated with

treatment efficacy

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SLIDE 42

There’s more than S. aureus

Gram Positive Gram Negative Fungi CoNS Acinetobacter baumannii Aspergillus fumigatus Enterococcus faecium Enterobacter aerogenes/ Candida albicans Enterococcus faecalis cloacae Candida glabrata

  • Staph. aureus
  • E. coli

Candida krusei

  • Strep. pneumoniae

Klebsiella pneumoniae/ Candida parapsilosis

  • Strep. sp.
  • xytoca

Candida tropicalis MRSA Proteus mirabilis Pseudomonas aeruginosa Serratia marcescens Stenotrophomonas maltophilia

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SLIDE 43

SeptiFast (not available in US)

Gram Positive Gram Negative Fungi CoNS1 Acinetobacter baumannii Aspergillus fumigatus Enterococcus faecium Enterobacter aerogenes/ Candida albicans Enterococcus faecalis cloacae4 Candida glabrata

  • Staph. aureus
  • E. coli

Candida krusei

  • Strep. pneumoniae

Klebsiella pneumoniae/ Candida parapsilosis

  • Strep. sp.2
  • xytoca4

Candida tropicalis MRSA3 Proteus mirabilis Pseudomonas aeruginosa Serratia marcescens Stenotrophomonas maltophilia

1-Staphylococcus hemolyticus, epidermidis = CoNS 2-Streptococcus agalaciae, pyogenes, viridans = Strep. Sp. 3-Separate test kit 4-No differentiation between these two subspecies

LAB BASED

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SLIDE 44

Patient is a 20 year old man status post motor vehicle accident with 90% TBSA 3rd and 4th degree burns and C1 pedicle and C4 foraminal fracture.

Day 1 2 3 4 8 23 30 31 32 33 37 38 39

BCA: Negative RC1: Collected RC1: H. influenzae WC1: Collected

15

WC1: MSSA, E. faecalis, Strep. viridans Added Linezolid

10

RC2: Collected Mold observed during dressing change. WC2: Collected Discontinued Linezolid, Meropenem; Added Vancomycin, WC2: A. fumigatus, Rhizopus sp. RC3: Collected Ampho B soaks

27

Septic Shock MAP: 40-50mmHg Started 4 Vasopressors Green exudate on wounds Platelet: 88,000 Patient expired Epinephrine Started Ceftazidime Added Posaconazole

Pathogen Detection in Burn Patients

RC3: Collected BCB: Collected Added Voriconazole, Meropenem BCB: P. aeruginosa RC3: P. aeruginosa BCC: Negative +Tobramycin

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SLIDE 45

Patient is a 20 year old man status post motor vehicle accident with 90% TBSA 3rd and 4th degree burns and C1 pedicle and C4 foraminal fracture.

Day 1 2 3 4 8 23 30 31 32 33 37 38 39

PCRA: Negative BCA: Negative RC1: Collected RC1: H. influenzae WC1: Collected

15

WC1: MSSA, E. faecalis, Strep. viridans +Linezolid

10

RC2: Collected Mold observed during dressing change. WC2: Collected Discontinued Linezolid, Meropenem; Added Vancomycin WC2: A. fumigatus, Rhizopus sp. RC3: Collected Ampho B soaks

27

RC3: Collected PCRB: P. aeruginosa BCB: Collected Added Voriconazole, Meropenem Septic Shock MAP: 40-50mmHg Started 4 Vasopressors Green exudate on wounds Platelet: 88,000 Patient expired Epinephrine Added Ceftazidime

Whole-Blood PCR-Based Pathogen Detection

Added Posaconazole BCB: P. aeruginosa RC3: P. aeruginosa BCC: Negative +Tobramycin

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SLIDE 46

Where are the whole blood PCR tests?

Blood Culture Sample FDA-approved As noted, the majority of pathogen detection systems today for septicemia relies on blood culture as the specimen type. Reason:

  • Integrates

into microbiology workflow.

  • Don’t

have to worry about amplifying the signal.

  • Less questions about what you’re

detecting is “real” or not.

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SLIDE 47

Where are the whole blood PCR tests?

Blood Culture Sample FDA-approved As noted, the majority of pathogen detection systems today for septicemia relies on blood culture as the specimen type. Reason:

  • Integrates

into microbiology workflow.

  • Don’t

have to worry about amplifying the signal.

  • Less questions about what you’re

detecting is “real” or not.

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SLIDE 48

What to do with PCR+/BC– Cases: Is ”DNAemia” Real?

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SLIDE 49

Patient is a 20 year old man status post motor vehicle accident with 90% TBSA 3rd and 4th degree burns and C1 pedicle and C4 foraminal fracture.

Day 1 2 3 4 8 23 30 31 32 33 37 38 39

PCRA: Negative BCA: Negative RC1: Collected RC1: H. influenzae WC1: Collected

15

WC1: MSSA, E. faecalis, Strep. viridans +Linezolid

10

RC2: Collected Mold observed during dressing change. WC2: Collected Discontinued Linezolid, Meropenem; Added Vancomycin WC2: A. fumigatus, Rhizopus sp. RC3: Collected Ampho B soaks

27

RC3: P. aeruginosa PCRB: P. aeruginosa BCB: Collected Added Voriconazole, Meropenem BCB: P. aeruginosa Septic Shock MAP: 40-50mmHg Started 4 Vasopressors Green exudate on wounds Platelet: 88,000 BCC: Negative Added Tobramycin Patient expired Epinephrine Added Ceftazidime

Whole-Blood PCR-Based Pathogen Detection

Added Posaconazole

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SLIDE 50

Patient is a 20 year old man status post motor vehicle accident with 90% TBSA 3rd and 4th degree burns and C1 pedicle and C4 foraminal fracture.

Day 1 2 3 4 8 23 30 31 32 33 37 38 39

PCRA: Negative BCA: Negative RC1: Collected RC1: H. influenzae WC1: Collected

15

WC1: MSSA, E. faecalis, Strep. viridans +Linezolid

10

RC2: Collected Mold observed during dressing change. WC2: Collected Discontinued Linezolid, Meropenem; Added Vancomycin WC2: A. fumigatus, Rhizopus sp. RC3: Collected Ampho B soaks

27

RC3: Collected PCRB: P. aeruginosa BCB: Collected Added Voriconazole, Meropenem Septic Shock MAP: 40-50mmHg Started 4 Vasopressors Green exudate on wounds Platelet: 88,000 Patient expired Epinephrine Added Ceftazidime

Whole-Blood PCR-Based Pathogen Detection

Added Posaconazole BCB: P. aeruginosa RC3: P. aeruginosa PCRC: P. aeruginosa BCC: Negative Added Tobramycin

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SLIDE 51

Whole-Blood PCR-Based Pathogen Detection

Patient is a 42 year-old woman with a perforated jejunum status post exploratory laparotomy and small bowel resection, who developed septic shock 24-hours later.

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SLIDE 52

Whole-Blood PCR-Based Pathogen Detection

Patient is a 42 year-old woman with a perforated jejunum status post exploratory laparotomy and small bowel resection, who developed septic shock 24-hours later.

2 3 Day 1 4 5

Septic Shock (3 hours) T = 38.6, WBC = 3.1 BP = 81/61, HR = 129

6 7 8 9

PCRA → Klebsiella pneumoniae/oxytoca and Enterobacter aerogenes/cloacae

T = 38.6, WBC = 18.2 BCA → Negative UC1 → Negative Antibiotic dose increased in view of edema T = 39.0, WBC = 20.4 BCB → Negative OR Exploratory laparotomy Bowel resection Open abdomen Started Ceftriaxone + Flagyl Discontinued Ceftriaxone + Flagyl Started Piperacillin/Tazobactam T = 38.7, WBC = 16.2 Added ciprofloxacin and vancomycin

10 11 12 17

Transferred to Floor T = 37.0, WBC = 14.1 OR: Abdominal Closure T = 37.2, WBC = 13.1 CT: negative for abscesses

PCRB = K. pneumoniae/oxytoca (Catheter only) and

  • E. aerogenes/cloacae (Catheter + Peripheral)
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SLIDE 53

Where are the whole blood PCR tests?

Blood Culture Sample Whole Blood Sample Not FDA-approved

7 Gram pos 8 Gram neg 6 Fungi In Development

FDA-approved Quite a few whole blood-based systems exist, but are not FDA approved (SeptiFast CE Marked)

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SLIDE 54

Where are the whole blood PCR tests?

Blood Culture Sample Whole Blood Sample FDA-approved Not FDA-approved

Candida sp. (5 species)

  • E. faecium
  • E. coli
  • K. pneumoniae
  • P. aeruginosa
  • S. aureus

7 Gram pos 8 Gram neg 6 Fungi In Development

FDA-approved Small, but growing number of molecular tests available that can assay from whole blood. However, test menu remains limited.

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SLIDE 55

T2-Time Magnetic Resonance Pathogen Detection

Innovative technology that mitigates the challenges of whole blood matrix by using T2-time magnetic resonance.

  • Providers faster results compared to contemporary blood culture enhanced molecular /

mass spec testing.

  • Limits of detection are reasonable and down to 1 CFU/mL.
  • However limited panel.

https://www.t2biosystems.com/

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SLIDE 56

Where are the whole blood PCR tests?

Blood Culture Sample Whole Blood Sample FDA-approved Not FDA-approved

Candida sp. (5 species)

  • E. faecium
  • E. coli
  • K. pneumoniae
  • P. aeruginosa
  • S. aureus

7 Gram pos bacteria 8 Gram neg bacteria 6 Fungi In Development

FDA-approved Not FDA-approved Also some novel technologies are

  • n the horizon!
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SLIDE 57

Smart Particle Technology

Another innovative technology with the potential benefit to accelerate in vitro susceptibility results and speciation

  • Utilizes “smarticles” bioparticles to specifically bind to target bacteria.

https://diagnostics.roche.com/global/en/article-listing/smarticles-technology.html

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SLIDE 58

Smart Particle Technology

Another innovative technology with the potential benefit to accelerate in vitro susceptibility results and speciation

  • Utilizes “smarticles” bioparticles to specifically bind to target bacteria.
  • Effectively a “live-cell” molecular test.

https://diagnostics.roche.com/global/en/article-listing/smarticles-technology.html

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SLIDE 59

Smart Particle Technology

Another innovative technology with the potential benefit to accelerate in vitro susceptibility results and speciation

  • Utilizes “smarticles” bioparticles to specifically bind to target bacteria.
  • Effectively a “live-cell” molecular test.
  • Can also detect phenotypic drug resistance in vitro.

https://diagnostics.roche.com/global/en/article-listing/smarticles-technology.html

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SLIDE 60

Current Lab-Centric Solutions

So most molecular pathogen detection remains in the central laboratory space. Laboratory-based testing

  • Lab keeps revenue
  • Demand on lab staff
  • Ensures consistency
  • Minimizes regulatory
  • versight of waived users
  • However impacts ED/

ICU workflow

  • Not all detectable

pathogens are needed

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SLIDE 61

Current Lab-Centric Solutions

Constrained to the laboratory due to their complexity (CLIA) or reliance on blood culture samples. Good for the laboratory! Laboratory-based testing

  • Lab keeps revenue
  • Demand on lab staff
  • Ensures consistency
  • Minimizes regulatory
  • versight of waived users
  • However impacts ED/

ICU workflow

  • Not all detectable

pathogens are needed

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SLIDE 62

Current Lab-Centric Solutions

Constrained to the laboratory due to their complexity (CLIA) or reliance on blood culture samples. Good for the laboratory! How about point-of-care testing? Laboratory-based testing

  • Lab keeps revenue
  • Demand on lab staff
  • Ensures consistency
  • Minimizes regulatory
  • versight of waived users
  • However impacts ED/

ICU workflow

  • Not all detectable

pathogens are needed

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SLIDE 63

ED and ICU

Hybrid Laboratory: Molecular

Could we exist as a “hybrid lab” incorporating both centralized diagnostics and point-

  • f-care testing?

Laboratory-based testing

  • Lab keeps revenue
  • Demand on lab staff
  • Ensures consistency
  • Minimizes regulatory
  • versight of waived users
  • However impacts ED/

ICU workflow

  • Not all detectable

pathogens are needed

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SLIDE 64

ED and ICU

Hybrid Laboratory: Molecular

Could we exist as a “hybrid lab” incorporating both centralized diagnostics and point-

  • f-care testing?

Bedside Testing

  • Options limited to mainly

respiratory panels.

  • ”True” POCT (waived)

solutions typically tests for Flu A/B and RSV.

  • RN workflow?
  • $$$$

Laboratory-based testing

  • Lab keeps revenue
  • Demand on lab staff
  • Ensures consistency
  • Minimizes regulatory
  • versight of waived users
  • However impacts ED/

ICU workflow

  • Not all detectable

pathogens are needed

slide-65
SLIDE 65

Hybrid Laboratory: Molecular

Could we exist as a “hybrid lab” incorporating both centralized diagnostics and point-

  • f-care testing?

Bedside Testing

  • Options limited to mainly

respiratory panels.

  • ”True” POCT (waived)

solutions typically tests for Flu A/B and RSV.

  • RN workflow?
  • $$$$

ED and ICU Laboratory-based testing

  • Lab keeps revenue
  • Demand on lab staff
  • Ensures consistency
  • Minimizes regulatory
  • versight of waived users
  • However impacts ED/

ICU workflow

  • Not all detectable

pathogens are needed

slide-66
SLIDE 66

Laboratory-based testing

  • Lab keeps revenue
  • Demand on lab staff
  • Ensures consistency
  • Minimizes regulatory
  • versight of waived users
  • However impacts ED/

ICU workflow

  • Not all detectable

pathogens are needed Near-patient testing?

  • Lab keeps revenue
  • Demand on lab staff
  • Intermediate turnaround

time – some ED/ICU workflow issues

  • New space / new
  • perators?

Hybrid Laboratory: Molecular

Could we exist as a “hybrid lab” incorporating both centralized diagnostics and point-

  • f-care testing? Can we leverage multiple platforms to optimize clinical impact?

Bedside Testing

  • Options limited to mainly

respiratory panels.

  • ”True” POCT (waived)

solutions typically tests for Flu A/B and RSV.

  • RN workflow?
  • $$$$

ED and ICU

slide-67
SLIDE 67

Challenges Remain for POCT

Significant technological and regulatory barriers in the way of POCT molecular pathogen detection for sepsis. Bedside Testing

  • Options limited to mainly

respiratory panels.

  • ”True” POCT (waived)

solutions typically tests for Flu A/B and RSV.

  • RN workflow?
  • $$$$

ED and ICU

  • Whole blood remains a challenging

matrix.

  • Bloodstream pathogen concentrations

may be low (0.5 – 1.0 CFU/mL) in early sepsis.

  • Molecular panels remain limited (can
  • nly detect what you assay is designed

to detect).

  • Flu A/B, RSV, Strep A is easier to

diagnose versus sepsis.

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SLIDE 68

Challenges Remain for POCT

Significant technological and regulatory barriers in the way of POCT molecular pathogen detection for sepsis. Bedside Testing

  • Options limited to mainly

respiratory panels.

  • ”True” POCT (waived)

solutions typically tests for Flu A/B and RSV.

  • RN workflow?
  • $$$$

ED and ICU

  • Costs

for molecular tests remain relatively high.

  • Costs associated with POCT operators

poorly defined.

  • Over utilization of molecular assays is

a known problem → need mechanisms to optimize use.

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SLIDE 69

Case Example: Diagnosis of Respiratory Tract Infections (RTI) in the ED during Flu Season

SIRS RTI POCT Leveraging chemistry tests to enhance molecular performance and cost-effectiveness Linkage of molecular diagnostics with chemistry / immunoassays offers other options and may optimize utilization of expensive molecular tests.

Hybrid Laboratory: Immunoassays with Molecular Creates Value

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SLIDE 70

Case Example: Diagnosis of Respiratory Tract Infections (RTI) in the ED during Flu Season

SIRS RTI Procalcitonin Lactate CRP(?) IL-6(?) POCT Leveraging chemistry tests to enhance molecular performance and cost-effectiveness

Hybrid Laboratory: Immunoassays with Molecular Creates Value

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SLIDE 71

Case Example: Diagnosis of Respiratory Tract Infections (RTI) in the ED during Flu Season

SIRS RTI Procalcitonin Lactate CRP(?) IL-6(?) Bacterial vs. Non- Bacterial Sources Antibiotics vs. Antivirals

  • vs. Non-Antimicrobials

POCT Leveraging chemistry tests to enhance molecular performance and cost-effectiveness

Hybrid Laboratory: Immunoassays with Molecular Creates Value

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SLIDE 72

Case Example: Diagnosis of Respiratory Tract Infections (RTI) in the ED during Flu Season

SIRS RTI Procalcitonin Lactate CRP(?) IL-6(?) Bacterial vs. Non- Bacterial Sources Antibiotics vs. Antivirals

  • vs. Non-Antimicrobials

POCT LAB Leveraging chemistry tests to enhance molecular performance and cost-effectiveness Improves BOTH antimicrobial and diagnostic stewardship!

Hybrid Laboratory: Immunoassays with Molecular Creates Value

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SLIDE 73

Case Example: Diagnosis of Respiratory Tract Infections (RTI) in the ED during Flu Season

SIRS RTI Procalcitonin Lactate CRP(?) IL-6(?) Bacterial vs. Non- Bacterial Sources Antibiotics vs. Antivirals

  • vs. Non-Antimicrobials

POCT LAB Leveraging chemistry tests to enhance molecular performance and cost-effectiveness Improves BOTH antimicrobial and diagnostic stewardship!

Hybrid Laboratory: Immunoassays with Molecular Creates Value

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SLIDE 74

Hybrid Laboratory: PCT and IL-6

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SLIDE 75

Hybrid Laboratory: PCT and IL-6

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SLIDE 76 Linscheid et al. Endocrinology 144(12):5578–5584

Procalcitonin (PCT) Basics

▪ Pro-hormone to calcitonin ▪ Normally produced in C-cells (normal serum levels <0.05 ng/mL) ▪ Bacterial infections: PCT released into bloodstream uncleaved ▪ Viral infections: PCT suppressed by IFNγ ▪ Low in non-specific inflammation, neutropenia, viral/fungal infections

  • FIG. 5. Schematic diagram of CALC I expression in adipocytes and thyroidal C cells. In the classical neuroendocrine paradigm, the expression of CT mRNA is
restricted to neuroendocrine cells, mainly C cells of the thyroid. Initially, the 116-amino acid prohormone ProCT is synthesized and subsequently processed to the considerably smaller mature CT. In sepsis and inflammation, proinflammatory mediators induce CT mRNA. In contrast to thyroidal cells, adipocytes and other parenchymal cells lack secretory granules, and hence, unprocessed ProCT is released in a nonregulated, constitutive manner.
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SLIDE 77

Future of the Hybrid Laboratory for Sepsis Prediction, Detection, and Management

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SLIDE 78

Integrating Molecular Testing, Microbiology, Chemistry/Immunoassay, POCT and Data Sciences

Automated Chemistry / Immunoassay High-Throughput Automated Molecular Diagnostics Point-of-Care Molecular

Future Directions

Artificial Intelligence Proteomics

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SLIDE 79

Conclusions

Clinical Performance Value Diagnostic Stewardship Workflow Optimization Molecular provides superior performance versus microbiology for detectable pathogens. Early appropriate and targeted anti-infective therapy improves

  • utcomes.

Integration of molecular POCT and laboratory methods with chemistry/immunoassay techniques. Diagnostic stewardship is needed to optimize molecular testing due to the relatively high cost. Speed Future Future will involve integrating multiple test modalities (hybrid lab) with electronic decision support to optimize value and care. Molecular diagnostics and potentially POC pathogen detection reduces “diagnostic blind spot”.