Upload
others
View
3
Download
0
Embed Size (px)
Citation preview
ENHANCING BLOOD SAFETY THROUGH NAT: THE FIRST BLOOD BANK EXPERIENCE FROM EASTERN INDIA
Dr. Ritam Chakrabarty, MD
Department of Transfusion Medicine
Apollo Gleneagles Hospitals, Kolkata
TRANSMEDCON 2016
Introduction
• Transfusion-transmitted infections are a major problem with blood
transfusion
• The benefit of Nucleic acid amplification test (NAT) over serology
has already been discussed elaborately by previous workers
• NAT has been able to detect infectious donors who were
otherwise designated normal serologically
• Here we share our experience of using NAT to screen blood
donors in Eastern India
Material and Methods • All blood donations between 23rd November 2013 and 31st August 2016
were included.
• Donors non-reactive for HBsAg, anti-HIV 1 & 2 and anti-HCV by CLIA
(VITROS, ECiQ, OCD) and negative for Syphilis and Malaria, were
subjected to NAT.
• NAT for HBV-DNA, HCV-RNA and HIV-RNA in the minipool of 6 samples
was performed using the Roche Cobas TaqMan MPX assay
• Individual sample of each positive pool was tested subsequently in the
same platform.
Material and Methods:
• Each positive sample was then investigated for viral
discrimination and viral quantitation from reference
laboratory.
• Sample positive for hepatitis- B virus (HBV) DNA was
further screened for anti-HBc antibody & antibody to
HBsAg (anti-HBs) by CLIA (VITROS, ECiQ, OCD).
• All results were documented & recorded as per the SOP.
Results • Of the total 24799 blood donations, HBsAg, anti-HCV, and anti HIV
by CLIA were detected in 168 (0.677%), 148 (0.596%) and 58 (0.233%)
donors respectively (Table 1)
• 172 donors (0.694%) tested positive for VDRL
• Out of 24253 samples tested for NAT, 9 (0.037%) were positive for
HBV DNA and 1 (0.004%) for HIV RNA
• The NAT yield was observed to be 1 in 2425 donations
• 7 NAT positive donors were sero-reactive for anti-HBc and 6
reactive for anti-HBs antibodies
• Viral load in each sample was < 6IU/ml (Figures)
Donor screening by NAT
Total blood donation
N = 24799
Serology / CLIA non reactive: 24253 Serology / CLIA reactive: 374
HIV 1& 2
N = 58
(0.233%)
HbsAg
N = 168
(0.677%)
HCV
N = 148
(0.596%)
NAT positive: 10 (yield = 1 / 2425)
HIV 1& 2: 1 HBV: 9 HCV: 0
Anti-HBc reactive: 7 Anti-HBs reactive: 6Viral load (N = 9): < 6IU/ml
VDRL +ve: 172
Viral discrimination & quantitation
Viral discrimination & quantitation
Viral discrimination & quantitation
DISCUSSION
HIV
HBVHCV
1996199419921990198819861984
1:100
1:1000
1:10 000
1:100 000
1:1 000 000
1998 2005
Ris
k p
er u
nit
Modified from Busch et al. JAMA 2013; 289: 959-62
2012
Evolution of Transfusion Risks near “eradication” of major viruses
Blood Safety in IndiaOut of 8 million annual donations, majority are first time donors
Heterogeneous structure of blood banking: Government / Private
/ NGO / Stand Alone
Marker General Population Blood Donor
HIV 0.36 % 0.3%
HBV 2.5 – 4% 1.4%
HCV 0.9% 0.7%
Marker
HIV
HBV
HCV
Antibody
Genome
Time of
infection
Time
Months/years
Antigen
Generalized course of TTI
30-70 days 3rd Gen ELISA
8-14 days
NAT
25-60 days 4th Gen ELISA/CLIA
Mandatory
Rs 100 - Rs 200
Additional Rs 200 - 300
Additional Rs 700 - 1000
Who will bear the economic burden?
What is NAT?• Nucleic Acid Technology (Nucleic Acid
Amplification Testing)
• All involve extraction or capture of nucleic acid,
amplification and detection
Commonly used systems are
PCR-based assays – NA amplification by
denaturation, annealing & primer extension
Transcription mediated amplification (TMA) assay
- Isothermal NA amplification in the same tube
Benefits of Implementing NAT
Decreases WP infection unlike serological assays
Increased sensitivity and specificity
Detects viral mutants and immunosilent carriers
Earlier donor counseling and patient care
Characterization of genotype is possible.
Future application may include additional viruses e.g.
CMV, Dengue etc
Genotype coverage in NAT
HIV-1 Group M, subtypes A, B, C, D, F, G, H
HIV-1 Group O (constitute 1-5% of all HIV)
HIV-2 (Increasing prevalence in India)
HBV, genotypes A, B, C, D, E, F, G
HCV, genotypes 1a, 1b, 2 a, 2b, 3a, 4a, 5a, 6a
NAT : Indian ScenarioStudy Sample
s testedType
of NATYield Remark
Makroo et al, 2008
12,224 ID 8 (1/1108)
1 HIV, 1 HIV-HCV, 6 HBV
Singh et al 2009
20,256 MP 11 (1/2000)
All HBV
Jain et al, 2012
23,779 MP 8 (1/2972)
8 HBV
Agarwal et al, 2013
73,898 ID 121 (1/610)
1 HIV, 37 HCV, 73 HBV, 10
co-infKoshy et al 2013
52,083 ID 47 (1/1108)
43 HBV, 3 HCV, 1 HIV
Cobas® TaqScreen MPX Test, v2.0 Simultaneous detection of multiple targets
FAMHIV-1-M, O, 2485/520 nm
HEXHBV
540/575 nm
JA270HCV
610/640 nm
CY5.5IC680/715 nm
Ch. 1
Ch. 3 Ch. 2
Ch. 4
Dyes and Filters
Real time multi-channel detection
Analytical SensitivityComparative analysis
Markers ProcleixUltrioElite
CobasTaqScreenMPX v2.0
Comments
HIV M Group 18 IU/mL 46.2 IU/mL Doubling Time of HIV (17 hrs) & HCV (10.8 hrs) virus is short& also viral load in window period is high for these RNA viruses, so lower sensitivity for HIV & HCV shall not miss any window period infections.
HIV O Group No claim 18.3 copies/mLHIV-2 10.4 IU/mL 7.9 IU/mLHCV 3.0 IU/mL 6.8 IU/mL
HBV 4.3 IU/mL 2.3 IU/mL Doubling time of HBV is long (2.6 days) & viral load in initial window period infection is not very high so probability of missing out HBV with a less sensitive assay is high
Viral load (copies/ul)HIV: upto 107
HCV:upto 107
HBV: 10 – 104
Conclusion • Introduction of NAT has successfully identified infectious donors
• Despite its cost effectiveness issues NAT will be a standard of
blood donor screening in the future
• With the implementation of NAT in our blood centre we could save
30 patients in 33 months who would have otherwise received the
infected blood components
• NAT could detect 9 occult hepatitis in otherwise healthy donors
who were then referred to gastroenterology for management
• 1 HIV donor was detected in window period
• Restraining these donors from further donations saved more lives
• Blood bank without NAT facility may start routine anti-HBc testing
Thank [email protected]