Keywords Doppler indices - early-onset neonatal sepsis - internal carotid artery - middle cerebral
artery - transcranial Doppler - vertebral artery
Introduction
Annually, worldwide one million neonatal deaths are attributed to neonatal sepsis
alone.[1 ] Neonatal sepsis is an important cause of morbidity and mortality amongst newborns.[2 ], [3 ] Neonatal sepsis is defined classically as a clinical syndrome characterized by systemic
signs of infection commonly accompanied by bacteremia. Two types of neonatal sepsis
have been described: early-onset neonatal sepsis (EONS) and late-onset neonatal sepsis
(LONS).
When features of sepsis appear within the first 72 h of birth or when there is bacteremia
or bacterial meningitis occurring at ≤72 h in infants it is described as EONS. On
the contrary, when features of sepsis manifest beyond 72 h of birth it is termed as
late-onset sepsis.[2 ], [4 ], [5 ]
There is paucity of literature regarding early alteration of the cerebral blood flow
(CBF) in neonatal sepsis. The brain of the neonates is highly susceptible to blood
flow fluctuations such that moderately elevated CBF can increase the risk of cerebral
hemorrhage, whereas moderate hypoperfusion can expose the brain to ischemic damage.[4 ], [6 ], [7 ]
Changes in the CBF during early hours of life may play a pivotal role in perinatal
brain damage and cause acute and long-term morbidity which can be assessed by ultrasound
Doppler examination which is a real time, relatively safe, noninvasive, and sensitive
method to evaluate these vascular changes.[8 ] Our study aims to document these changes by ultrasound Doppler examination by studying
changes in values of peak systolic velocity (PSV) and end diastolic velocity (EDV),
resistivity index (RI), and pulsatility index (PI) in neonates with EONS compared
to age-matched neonates without any signs of sepsis and to evaluate the predictive
accuracy of CBFV by using ultrasound Doppler as a diagnostic marker of EONS.
Methods
This cross-sectional analytical study was conducted over a period of 2 years with
123 neonates enrolled in the study. The neonates were divided into two groups: Group
I had 54 neonates - neonates with EONS and group II –gestational age-matched neonates
without any signs of sepsis which comprised 69 neonates.
Ethical clearance was obtained from the institutional ethical clearance committee.
Informed consent was obtained from the parents/guardians of the enrolled subjects.
Inclusion criteria for neonates with early-onset neonatal sepsis
Neonates presenting within 72 h of birth with either clinical signs of sepsis or with
positive septic screen and/or culture positive sepsis were included in group I:[6 ], [7 ], [8 ]
Clinical signs of sepsis include features like hypothermia or fever, lethargy, poor
cry, refusal to suck, poor perfusion, prolonged capillary refill time, hypotonia,
absent neonatal reflexes, brady/tachycardia, respiratory distress, apnea and gasping
respiration, hypoglycemia, hyperglycemia, metabolic acidosis, bulging anterior fontanelle,
seizures, stupor/coma, vomiting, diarrhea, paralytic ileus, necrotizing enterocolitis,
direct hyperbilirubinemia, acute renal failure, petechiae, purpura, bleeding, multiple
pustules, umbilical redness and discharge, abscess.
Septic screen positive: When two or more of the following parameters are positive
[Table 1 ].
Table 1
Components of the septic screen
Components
Abnormal value
VLBW: Very Low Birth Weight, ESR: Erthrocyte Sedimentation Rate, AGA: Appropriate
for Gestational Age
Total leukocyte count
<5000/mm3
Absolute neutrophil count
Low counts as per Manroe chart for the term and Mouzinho’s chart for Very Low Birth
Weight infants
Immature/total neutrophil
>0.2
Micro-Erthrocyte
>15 mm in the first hour
Sedimentation Rate
C reactive protein
>1 mg/dl
Clinical workup
Detailed antenatal and intranatal history was taken. At birth, a thorough clinical
examination was done to exclude any systemic or congenital malformation. Laboratory
estimation of liver and kidney function tests, serum electrolytes, blood glucose,
coagulation profile, and arterial blood gas analysis was done to exclude inborn errors
of metabolism on basis of clinical suspicion.
Exclusion criteria
History of eclampsia or use of tocolytics or steroids in mothers
Neonates with medical or surgical illnesses: Perinatal asphyxia, congenital infections,
congenital heart disease, hemolytic anemia, metabolic disorders, surgical disorders
gross congenital malformations and meconium aspiration syndrome
Neonates with altered arterial blood gas analysis, with hemodynamic instability and
those requiring vasopressor support and mechanical ventilation, receiving anticonvulsants
or drugs like caffeine, theophylline at the time of Doppler examination.
Ultrasound Doppler examination of the neonate
Ultrasound Doppler examination was performed in neonates during the first 72 h of
birth. The examination was carried out in a thermoneutral environment. All the babies
were well fed before the examination. A total of 25% oral dextrose was used as a pacifier
with continuous monitoring of vitals.
The Doppler examinations were performed by a single radiologist on MyLab50 esaote
ultrasonography with color Doppler ultrasound machine with curvilinear (3.5-5 MHz
for ICA and MCA) and high frequency linear (7.5 MHz for VA) array transducer. PSV,
EDV, RI, and PI were measured in the ICA, MCA, and VA of either side. Angle corrected
velocities were taken. PI and RI were calculated as per formulae of the ultrasound
blood flow imaging technique. The examination was carried out through the anterior
fontanelle in the coronal plane [Figure 1 ]. The circle of Willis was located and the ICA and MCA were identified. The vertebral
artery was assessed through either side of the neck of the neonate [Figure 2 ].
Figure 1: Transcranial Doppler ultrasonography – examination of the neonate through the anterior
fontanelle
Figure 2: Doppler ultrasonography – assessment of vertebral artery through the side of the
neck
Statistical analysis
Data were analyzed using the statistical program SPSS version 23.0 (SPSS Inc., Chicago,
IL, USA). Frequencies and proportions of the different variable were expressed in
percentages. Comparison of numerical variables between the study groups was done using
Student t -test for independent samples in comparing two groups when normally distributed and
Mann–Whitney U test for independent samples when not normally distributed. Chi-square
test was used to compare proportions between the groups. Sensitivity, specificity,
PPV, NPV, and diagnostic accuracy were calculated at different selected cutoff values
for cerebral blood flow velocity (CBFV) parameters. A P value of < 0.05 was considered statistically significant.
Results
In our study out of total 123 neonates, 18% (22) were examined on day 1, i.e. within
24 h of life, 39% (48) were examined on day 2, i.e. within 24–48 h of life, and 43%
(53) were examined on day three, i.e. within 48–72 h of birth
Out of 69 neonates without sepsis, 29 were examined on day 2, 27 on day 3, and 13
on day one. Among neonates with sepsis 24 were examined on day 2, 21 on day 3, and
9 on day 1
In the present study out of 123 neonates, 68 (55.3%) were preterm and 55 (44.7%) were
term. Among neonates without sepsis there were 35 term and 34 preterm and in neonates
with sepsis, there were 34 preterm and 20 term neonates. Thus, the percentage of preterm
neonates (63%) was more as compared to term neonates (37%) in neonates with sepsis
Out of total 123 neonates, 46% (57) were females and 54% (66) were males. Among neonates
with sepsis there were 35 (64.8%) males and 19 (35.2) females. Among neonate without
sepsis there were 31 (44.9%) male and 38 (55.1%) female. In neonates with sepsis,
male patients were more as compared to females
In neonates without sepsis, 30 (43.5%) were Appropriate for Gestational Age and 39
(56.5%) had low birth weight, whereas in neonates with sepsis 17 (31.5%) were Appropriate
for Gestational Age and 37 (68.5%) had low birth weight. Thus, the percentage of low
birth weight babies was more in neonates with early onset neonatal sepsis
In the present study, we found there is statistically significant difference in Apgar
score at 5 min among neonates with early onset neonatal sepsis [median (interquartile
range)- 9 (8-9); min-max = 6-9] and neonates without early onset [median (interquartile
range)- 9 (9-9); min-max = 8-9] neonatal sepsis
Statistically significant difference is noted between the median values of PSV, EDV,
PI and RI of ICA, MCA and VA among neonates with sepsis and neonates without sepsis
as P value is less than 0.05The median and the range of PSV, EDV, RI, and PI of neonates
with sepsis and of neonates without sepsis of ICA, MCA, and the VA is as shown in
[Table 2 ] and [Figures 3 ], [4 ], [5 ], [6 ], [7 ], [8 ]
The predictive accuracy of measured parameters of ICA, MCA, and VA are as shown in
[Table 3 ]
Among the various parameters measured the diagnostic accuracy was highest for PI in
all the vessels
Sensitivity of the PI and the EDV of the ICA was 100%
The sensitivity of the PSV of the MCA was 100% and the diagnostic accuracy was 98.37%
for the PI
The sensitivity, specificity, PPV, NPV, and diagnostic accuracy of PI measured in
VA was 100%
The positive likelihood ratio was highest for PSV of VA, i.e., 71.42
The negative likelihood ratio was highest for PSV of ICA, i.e., 0.44.
Table 2
Comparison of cerebral blood flow velocities and Doppler indices measured in the internal
carotid artery, middle cerebral artery, vertebral artery, on Doppler ultrasound among
neonate without sepsis and neonate with sepsis
Parameter
Neonate without sepsis [Median (range)]
Neonate with sepsis [Median (range)]
Statistical significance (P )
PSV (cm/s) ICA
35.40 (27.5-45.0)
54.00 (38.00-70.00)
0.00
EDV (cm/s) ICA
7.60 (2.40-12.30)
18.36 (8.64-31.4)
0.00
RI ICA
0.80 (0.70-0.92)
0.66 (0.50-0.84)
0.00
PI ICA
1.4 (1.20-1.80)
1.00 (0.60-1.24)
0.00
PSV (cm/s) MCA
25.00 (17.00-33.3)
45.00 (31.8-55.8)
0.00
EDV (cm/s) MCA
5.42 (3.15-24.00)
16.50 (7.60-24.00)
0.00
RI MCA
0.79 (0.70-0.86)
0.70 (0.40-0.82)
0.00
PI MCA
1.35 (1.00-1.67)
1.00 (0.55-1.20)
0.00
PSV (cm/s) VA
21.00 (12.00-30.00)
40.00 (30.00-55.00)
0.00
EDV (cm/s) VA
3.60 (1.40-7.00)
9.78 (4.10-19.20)
0.00
RI VA
0.82 (0.70-0.92)
0.73 (0.54-0.90)
0.00
PI VA
1.51 (1.24-2.10)
1.00 (0.89-1.20)
0.00
Table 3
Predictive accuracy of different parameters of cerebral blood flow velocity in the
internal carotid artery, middle cerebral artery, and vertebral artery
Parameter
Area under the curve (AUC)
Cut-off
Sensitivity
Specificity
Positive predictive value
Negative predictive value
Diagnostic accuracy (%)
Likelihood ratio (LR)+
LR_
PSV ICA
0.967
≥39.5
98.1
68.1
70.7
97.9
81.30
3.07
0.44
EDV ICA
0.988
≥8.52
100
65.2
69.2
100
88.48
2.873
-0.533
RI ICA
0.945
≤0.73
94.4
87
85
95.2
90.4
7.261
-0.085
PI ICA
0.998
≤1.25
100
95.7
94.7
100
97.5
23.25
-0.04
PSV MCA
0.994
≥31.4
100
91.3
90
100
95.12
11.49
-0.095
EDV MCA
0.982
≥7.64
98.1
88.4
86.9
98.4
92.6
8.456
-0.109
RI MCA
0.936
≤0.76
92.6
65.2
67.6
91.8
77.23
2.66
-0.420
PI MCA
0.995
≤1.15
98.1
98.6
98.1
98.6
98.37
70.071
0.005
PSV VA
0.999
≥29
100
98.6
98.2
100
99.18
71.42
0
EDV VA
0.969
≥4.36
96.3
72.5
73.2
96.2
82.92
3.501
-0.328
RI VA
0.802
≤0.80
81.5
66.7
65.7
82.1
73.17
2.44
-0.221
PI VA
1.000
≤1.22
100
100
100
100
100
0
Figure 3: Transcranial Doppler ultrasound examination of internal carotid artery in a neonate
with early-onset neonatal sepsis shows PSV-64.9 cm/s, EDV- 23.7 cm/s, RI- 0.63, PI-
0.93
Figure 4: Transcranial Doppler ultrasound examination of middle cerebral artery in a neonate
with early onset neonatal sepsis shows PSV- 38.3 cm/s, EDV-13.3 cm/s, RI- 0.65, PI-
1.07
Figure 5: Doppler ultrasound examination of vertebral artery in a neonate with early-onset
neonatal sepsis shows PSV- 46.4 cm/s, EDV- 15.1 cm/s, RI- 0.67, PI- 0.92
Figure 6: Transcranial Doppler ultrasound examination of internal carotid artery in a neonate
without sepsis shows PSV-35 cm/s, EDV- 10 cm/s, RI-0.71, PI-1.44
Figure 7: Transcranial Doppler ultrasound examination of middle cerebral artery in a neonate
without early-onset neonatal sepsis shows PSV-32.5 cm/s, EDV- 8.7 cm/s, RI- 0.73,
PI- 1.42
Figure 8: Doppler ultrasound examination of vertebral artery in a neonate without early onset
neonatal sepsis shows PSV-22.4 cm/s, EDV-2.6 cm/s, RI- 0.89, PI- 1.71
Discussion
In the present study, significantly lower resistance (PI and RI) and higher peak systolic
velocity and end diastolic velocity in all the three major vessels ICA, MCA, and VA
have been documented within 72 h of birth in neonates with EONS. This indicates a
generalized increase in the CBF as an early response to sepsis with a decrease in
the blood flow resistance. As the earliest signs of sepsis are often subtle and nonspecific,
a high index of suspicion is needed for early diagnosis.[9 ]
In our study, we have examined the neonates within 72 h of birth as and when they
presented with features of EONS as well as neonates without sepsis taken as controls.
Due to inflammatory challenges of chorioamnionitis, there is activation of the fetal
immune system which is known as fetal inflammatory response syndrome (FIRS). FIRS
is initially subclinical. Amplification of the inflammatory response in neonates after
birth known as systemic inflammatory response syndrome (SIRS) due to chorioamnionitis
is clinically diagnosed as EONS.[10 ]
The pathophysiology of sepsis at the cellular and molecular level is unknown. Studies
have stated that there occurs impairment of cerebral autoregulation due to cytokine
response and lead to cerebral ischemia or overperfusion which is responsible for perinatal
white matter injury and cerebral hemorrhage. Damage to blood–brain barrier (BBB) results
in its increased permeability with the transfer of cytokines across the BBB with the
direct inflammatory effects of free radicals, oxidative stress, and cytokines on glial
cells.[11 ], [12 ], [13 ], [14 ]
It has also been shown that astrocytes and microglia are capable of producing proinflammatory
cytokines during inflammation which can modify the re-uptake of glutamate and stimulate
the release of free radicals or induce local production of nitric oxide which is a
potent vasodilator.[15 ], [16 ], [17 ], [18 ], [19 ]
As there has been a substantial improvement in the antenatal, perinatal care there
has been an increase in the survival of the premature and the low-birth weight babies.
Changes in CBFV may have a key role in perinatal brain damage, and both acute and
long-term morbidity may be closely related to rapid vascular changes during the early
hours of life.[6 ]
We measured the parameters of the vessels on either side as there are studies which
have shown that there is no significant difference in the values on both sides.[6 ], [7 ], [20 ]
Our results agreed with the study conducted by Sriparna Basu et al . that detected significantly lower resistance, vasodilatation, and higher peak systolic
velocity in the three major cerebral vessels (ICA, MCA, and VA) indicating generalized
increase in CBF as an early response to sepsis.[6 ]
Koch et al . also proved that their chorioamnionitis full terms had decreased resistance in most
of the major cerebral vessels compared with controls.[21 ]
Lower resistance and increased peak systolic velocity in neonates with EONS was also
stated by Rania Hashem et al . by evaluating the ACA and MCA at day 3 of life by transcranial Doppler[4 ] and by EL Shimmy[3 ]
et al . who concluded that CBF and cord blood neuron-specific enolase were elevated in the
neonates with EONS.
Among all the parameters studied in ICA, the sensitivity of pulsatility index was
highest in the present study with sensitivity of 100% and with a diagnostic accuracy
of 97.5%, whereas for MCA the diagnostic accuracy was highest for pulsatility index
(98.37%) and the sensitivity was highest for peak systolic velocity (100%). The pulsatility
index measured in vertebral artery had 100% sensitivity and diagnostic accuracy.
Blood culture is considered to be the gold standard for the diagnosis of neonatal
sepsis.[9 ] In our study, eight (14.8%) neonates had a positive blood culture.
Certain differences can be observed in the measurements of various parameters of the
ICA, MCA, and VA among neonates with and without EONS in the different studies. These
differences can be attributed to the time of performance of the Doppler measurements
and the number of culture-positive cases which were different in all the studies.
It has been documented that there occurs increase in the CBF before the clinical appearance
of the frank features of sepsis meaning that the inflammatory process first affected
the cerebral circulation before it could affect any other body system, thereby further
making the role of transcranial Doppler in assessment of EONS more important such
that CBF can be used as an early marker of SIRS and to identify the neonates who are
going to develop EONS.
There are contradictory reports regarding evidence of effect of sepsis on the CBF.
Some state that there occurs vasodilatation and increased CBF,[22 ], [23 ], [24 ], [25 ] whereas others have demonstrated decreased CBFV along with higher PI in patients
with sepsis.[26 ], [27 ], [28 ] Reduced CBF is ascribed to the vasoconstriction of the resistance arterioles.[28 ] Increased PI, a parameter for compliance of the vascular bed suggesting cerebral
edema or venous congestion, has been found to correlate with poor neurological outcome.[29 ]
Among the various demographic parameters and the clinical data, we found that the
preterm and low-birth weight neonates were affected more as compared to term which
are also the risk factors for EONS.[30 ], [31 ] Our study reveals male neonates to be more vulnerable to develop EONS as compared
to females which was similar with the study by Basu et al .[6 ] Studies have revealed that term male infants have higher incidence of sepsis than
term female infants.[31 ]
There are conflicting reports regarding the significance of low Apgar score at 5 min
as risk factor in development of neonatal sepsis. In our study, we found statistically
significant difference in Apgar score at 5 min among neonates with EONS and neonates
without EONS. However in the study conducted by Basu et al . there was no statistically significant difference in Apgar score at 5 min among
neonates with EONS and neonates without EONS.[6 ] However according to Kari Simonsen, low Apgar scores (score of 6 at 5 min) is a
risk factor for EONS.[2 ]
Our study was cross-sectional with single-time assessment of CBFV and therefore could
not assess the fluctuation of CBFV over a period of time and correlate it with the
adverse effect as studies have shown that children who developed neonatal sepsis were
three times more likely to have neuromotor and cognitive development alterations at
12 months of corrected age.[21 ] Therefore, the single-time assessment of the CBF velocity was one of the limitations
of our study.
Conclusion
Our study shows that the cerebral hemodynamics in neonates with EONS is altered with
an increase in the CBF and a decrease in the resistance.
Assessment of CBF at early hours of birth by Doppler ultrasound examination can be
adopted as a bedside, noninvasive investigation with immediate diagnostic, and late
prognostic significance. It can be considered as one of the complementary investigation
to the various laboratory test that requires sufficient amount of blood sample requiring
invasive procedure, which can be avoided by noninvasive Doppler examination. Apart
from being noninvasive it is portable, uses nonionizing radiation, accurate, cost-effective,
and allows simultaneous examination of cranial structures and measurement of absolute
CBFV with the added benefit of serial and repetitive examination with early institution
of treatment.
Declaration of patient consent
The authors certify that they have obtained all appropriate patient consent forms.
In the form the patient(s) parents/guardian has/have given his/her/their consent for
his/her/their images and other clinical information to be reported in the journal.
The patients understand that their names and initials will not be published and due
efforts will be made to conceal their identity, but anonymity cannot be guaranteed.
