JOURNAL CLUB :

SYNDROME EVALUATION SYSTEM (SES) VERSUS

BLOOD CULTURE (BACTEC) IN THE DIAGNOSIS
AND MANAGEMENT OF NEONATAL SEPSIS -
A RANDOMIZED CONTROLLED TRIAL.

Bhat BV ,Prasad P ,Ravi Kumar VB ,Harish BN ,Krishnakumari K ,Rekha A , Manjunath

G ,Adhisivam B ,Shruthi B.

INDIAN JOURNAL OF PEDIATRICS, MAY 2016

 ABSTRACT :

OBJECTIVE:
To compare the clinical outcome of a multiplex polymerase
chain reaction (PCR) based molecular diagnostic method --
SYNDROME EVALUATION SYSTEM (SES) directed treatment
strategy vs. standard of care (BLOOD CULTURE) directed
treatment strategy for neonatal sepsis.
 INTRODUCTION

Sepsis is the leading cause of death in NICU

There is a need for rapid , accurate and effective diagnosis
Treatment may be started empirically but appropriate, of choice
treatment is also required
28 day mortality in adults was studied between appropriately treated
and inappropriately treated group by Harbarth et al , rates being 24%
and 39% respectively
Thus rapid detection of causative pathogens is of immense importance
GOLD STANDARD INVESTIGATIONS like BLOOD CULTURE has low
sensitivity and takes longer time ( 24-72 hrs )
 A number of rapid molecular diagnostic methods have
evolved capable of detection of causative pathogens of
neonatal sepsis including fungi , viruses and slow growing ,
fastidious bacteria with great accuracy and rapidity .
Two types of molecular techniques are generally used
1. Species identification and antibiotic resistance
detection in positive blood culture
2. Direct identification of pathogen from blood samples
( real time PCR )
 MOLECULAR DIAGNOSTIC SERVICES SYNDROME
EVALUATION SYSTEM (SES)

A patented PCR system technology that comprises of rapid

MULTIPLEX amplification and accurate identification of the virulence
associated genes of the causative agents or organisms
The accurate and rapid identification of the causative agents (7
hours of process time) empowers the clinician to initiate targeted
therapy
XCYTON offers SES as Molecular Diagnostic Services for various
critical illnesses in three verticals - Eye Infections, Brain Infections
and Sepsis or Blood stream infections.
 THE XCYTON ADVANTAGE
Rapid: Process time of 7 hours
Higher Accuracy: detects more number of cases than conventional
methods, 10-15% by conventional methods vs 75% by SES
Cost effectiveness: Avoids multiple testing and unnecessary
investigations and reduces hospital stay
Provides Direct evidence for the presence of infection - detects
DNA of pathogens
Comprehensive: Detects fungi, viruses, parasites and bacteria in a
SINGLE TEST. It also detects uni-microbial or poly-microbial infections
 THE TECHNOLOGY
Isolation of the genetic material of the causative
agent from the given specimen .
Simultaneous amplification of the "Syndrome
Specific Signature genes" of all the probable
causative agents .
Followed by "Syndrome Specific Hybridization".
 CNS INFECTIONS
SES-MENINGITIS
SES-MENINGO ENCEPHALITIS
SES- ACUTE ENCEPHALITIC SYNDROME
CYSTI CheX
BLOOD STREAM INFECTIONS
SES- SEPSIS( ADULTS, NEONATES & BURNS)
SES- PNEUMONIA
SES- ANTIBIOTIC RESISTANCE
SES ACUTE PYREXIA
SES- DENGUE-CHIKUNGUNYA
Detects 14 different bacterias and 3 species of fungi
Detects 5 different DNA viruses, 4 leading bacterias, 1
parasite and 1 fungal spp.
Detects 14 different RNA viruses, 5 DNA Viruses, 4
leading Bacterias, 1 parasite and 1 fungal spp.
Detects Neurocysticercal Antibodies
Detects 17 different bacterias and 2 species of fungi
Detects 17 different bacterias, 2 species of Fungi and 4
DNA viruses
Detects Gene markers in bacterias that confers resistance
for 3rd and 4th generation cephalosporins, carbapenems,
Vancomycin, Methicillin
Detects the commonest fever causing pathogens 3
bacterias, 2 parasites and 2 RNA viruses
Detects the 2 RNA viruses that cause seasonal fevers
 INFECTIONS IN
IMMUNOSUPPRESSED
Detects 17 different bacterias, 2 species of Fungi and 4
SES- FEBRILE NEUTROPENIA
DNA viruses
Detects 17 different bacterias, 2 species of Fungi and 4
SES-TRANSPLANT
DNA viruses
OPHTHALMIC INFECTIONS

SES- ENDOPHTHALMITIS Detects 10 different bacterias and 2 species of Fungi

SES- RETINITIS Detects 3 DNA viruses

SES UVEITIS Detects the Mycobacterium spp and 1 parasite

Detects the 3 RNA, 3 DNA viruses, 3 Mycobacterium spp.

SES- PAN UVEITIS
And 1 parasite

SES FUCHS Detects 3 DNA viruses, M.tuberculosis and 1 parasite

 MATERIAL AND METHODS:
STUDY DESIGN : Randomized controlled trial (RCT)
STUDY PLACE : NICU JIPMER , Pondicherry
STUDY PERIOD : January 2013 August 2013
SAMPLE SIZE : 385 neonates
Study approved by INSTITUTE SCIENTIFIC ADVISORY AND ETHICS
COMMITTEE
INCLUSION CRITERIA :
1. Institutional deliveries with early neonatal sepsis and antibiotic
nave
2. Babies with atleast two positive laboratory sepsis screening test
with clinical sepsis as per EMA guidelines
 At least 2 of the following signs symptoms had to be
present
1. Hypothermia or fever
2. Lethargy , poor cry , refusal to suck
3. Poor perfusion , prolonged CRT
4. Hypotonia , absence of neonatal reflexes
5. Respiratory distress , apnea and gasping respiration
6. Hypo/Hyperglycemia
 At least 2 of the following laboratory screening test to be
positive
1. Micro ESR > 15 mm/ 1st hr
2. TLC < 5000/cmm or > 20000/cmm
3. CRP > 4 mg/dl
4. I : T ratio > 0.2
5. Procalcitonin > 2 ng/ml
 EXCLUSION CRITERIA :
1. Birth weight < 1 kg 2. Gestational age < 28 weeks
3. APGAR score at 5 min < 6 4. Major congenital anomalies
385 neonates with sepsis were randomized by computer generated
random numbers into two groups -- SES and control (BACTEC).
Proportion outcome in control arm was set as 70% survival while SES
arm considered significant at 85% survival
TYPE I error set at two sided 5% and power at 90% , Dropout rate
assumed 25%
 Written informed consent was obtained
Two ml of blood was drawn from each baby
, 1ml of which was sent for blood culture,
and the remaining was sent for SES in
opaque sealed envelopes
Empirical antibiotics started after sending
blood samples and further escalations done
as per NICU protocols
Both tests were performed for all the
neonates.
In the SES group, the results of SES test
were revealed to the treating clinicians
immediately when available, while in the
control group, SES results were revealed
only after completing the entire study.
BACTEC results were not revealed to
XCYTON conducting SES test
 Further antibiotic changes, if required were done in SES and
control groups based on their respective reports.
Antibiotics stopped after 10 days if neonates were clinically normal
and sepsis screen negative
Necessary additional investigations and interventions done as per
NICU protocols when required
All babies followed up until 1 month after discharge
Comparison was made between the 2 groups on
Microbiological profile
Immediate outcome
Duration of hospital stay
Number of antibiotics used
Readmission within a month
SES PROCEDURE
Primer and probe design
biotin labeled primers and
probes used in SES designed
using full length genome
sequences from GenBank NCBI
Assay procedure 1. NUCLEIC
ACID EXTRACTION
2. NUCLEIC ACID
AMPLIFICATION
3. HYBRIDIZATION
OUTCOMES OF THE STUDY
PRIMARY :
Neonatal survival due to early detection of causative
agents
SECONDARY :
Mortality
Number of days in NICU
Duration of hospital stay
Antibiotic changes required
Duration of antibiotics and support measures
RESULTS
Significant reduction of mortality in SES compared to control ( p<
0.001) ( 33 - control , 6 - SES ) . No death in SES negative babies in
both groups .
SES resulted in significantly lesser number of days of NICU and
hospital stay
Readmission within 1 month was also lower in SES arm ( p< 0.001).
Babies in SES arm needed lesser antibiotics , lesser change of
antibiotics , lesser hours of ventilation , lesser dosage and duration of
inotropes and lesser transfusion of blood products ( p< 0.001).
 SES was better than BACTEC in identifying the causative organism in both
the groups (68% vs. 18% in SES group and 72% vs. 18% in control
group).
SES had 100% concordance with blood culture by BACTEC.
Detection of bacteria and fungi were four and ten-fold higher respectively
with SES when compared to BACTEC culture.
Microbiological diagnosis was rapid with SES compared to BACTEC (7h vs.
72h).
Treatment based on SES resulted in significantly less mortality (3% vs.
18%).
 Causative agents could be identified in only 18% babies by culture method
whereas SES was positive in 68% and 72 % in SES and control arms
respectively
SES detected more microbes (p<0.001) , more fungi ( 18 & 30 in SES and
control arm ) ( 48 candida , 4 aspergilus vs BACTEC 5 candida , none
aspergilus )
2 cases of Serratia sp. Detected by BACTEC which is not a part of SES panel
All culture positive cases (66) were monomicrobial on BACTEC but out of
these 66 cases 43 were found to be polymicrobial by SES .
134 cases out of 258 SES positive cases were monomicrobial .
 Occurrence of dual and triple organisms.
 There were 7 cases where SES detected 4 pathogens
All babies in the study had early neonatal sepsis ( meningitis : 7
SES , 6 culture ) ( UTI : 7 - SES , 9 - culture )
Out of 33 deaths in CONTROL arm , 30 were positive on SES of
which 11 had candidemia , while only 11 were positive on culture
with 1 candidemia
Out of 6 deaths in SES arm all were positive on SES with 3
candidemia , while 4 were only positive on culture with no
candidemia
 All cases from both arms were tested on SES antibiotic
resistance .
CTXM1 was detected in 50 cases (out of 185) in SES arm , 46
cases ( out of 183 ) in control arm [Klebsiella pneumoniae with
ESBL phenotype]
OXA-23 detected in 5 cases in SES arm and 6 cases in control
arm [Acinetobacter baumannii] , 1 was Meropenem resistant
2 cases showed van A gene on SES arm , one of which was
culture positive for Enterococcus faecium Vancomycin resistant
phenotype
Methicillin resistance was not detected in the study
 CONCLUSIONS :
Mortality rate in control arm was 17.8% compared to 3.2 % in SES arm
signifies the importance of early switch to targeted therapy .
Blood culture positivity in the study was only 8 %. Frequent isolation of
skin commensal was another problem which is not present with SES .
SES had 100 % concordance with culture except for Serratia sp.
SES was negative in 110 cases ( 60 SES arm , 50 control arm ) and
there was no deaths among them signifying SES had not missed even
a single case of significant sepsis ( 100% NPV )
Average antibiotic usage was 2.6 in SES arm compared to 5.8 in
control , ultimately reducing antibiotic resistance
 However less frequent but serious pathogen like Serratia is not
part of SES panel
Authors believe that SES is an add on diagnostic tool along with
blood culture
Impact of SES on extramural and previously treated babies has
not been studied
Detailed analysis of SES negative cases should be done .
 DRAWBACKS OF THE STUDY
Only Inborn babies admitted in NICU considered
Early neonatal sepsis considered as < 7 days
Antibiotic nave babies studied
ELBW babies excluded
No place for hepatosplenomegaly and abdominal distension in clinical sepsis criteria
used
ANC not considered as a part of sepsis screen
Randomisation though claimed to be efficient , divided 385 babies into unequal groups
( 190 SES and 195 control )
Methicillin resistance not tested , overall antibiotic resistance testing results were non
conclusive
Antibiotic stoppage in SES / culture negative babies controversial
Mortality statistics not properly elaborated or tabulated .
THANK YOU.