Background and ObjectivesHepatitis E virus (HEV) is a known transfusion‐transmissible agent. HEV ... more Background and ObjectivesHepatitis E virus (HEV) is a known transfusion‐transmissible agent. HEV infection has increased in prevalence in many developed nations with RNA detection in donors as high as 1 in 600. A high proportion of HEV infections are asymptomatic and therefore not interdicted by donor exclusion criteria. To manage the HEV transfusion‐transmission (TT) risk some developed nations have implemented HEV RNA screening. In Australia, HEV is rarely notified; although locally acquired infections have been reported, and the burden of disease is unknown. The purpose of this study was to determine the frequency of HEV infection in Australian donors and associated TT risk.Materials and MethodsPlasma samples (n = 74 131) were collected from whole blood donors during 2016 and screened for HEV RNA by transcription‐mediated amplification (TMA) in pools of six. Individual TMA reactive samples were confirmed by RT‐PCR and, if positive, viral load determined. Prevalence data from the ...
Aspects of Developmental and Comparative Immunology, 1981
ABSTRACT Previous studies on infectious bursal disease virus (IBDV) have demonstrated that it has... more ABSTRACT Previous studies on infectious bursal disease virus (IBDV) have demonstrated that it has a significant effect upon the avian humoral immune system. In the present study the effect of infectious bursal disease has been assessed sequentially by tests both for humoral and for cell mediated immunity (CMI). Day old chickens were infected with an Australian strain of IBDV of intermediate virulence by the ocular route. Reduced haemagglutination titres to sheep red blood cells and haemagglutination inhibition titres to lentogenic Newcastles disease virus indicates that this virus appears to affect mainly mercaptoethanol resistant antibodies. In addition the results of assays for CMI showed that this strain causes an impairment of this component of the immune system. It was found by immunofluorescent studies of bursa, spleen, thymus and peripheral blood that there was a significant reduction of light chain positive cells in the peripheral blood of infected chickens between two weeks and four weeks. Repopulation of the infected bursa by lymphocytes occurred by four weeks and the serum antibody levels to the above antigens returned to near normal levels by five weeks following infection but the CMI remained depressed.
Background An emerging infectious disease (EID) refers to a disease that has recently appeared in... more Background An emerging infectious disease (EID) refers to a disease that has recently appeared in a population or one that has rapidly increased in incidence or geographic range. Examples of EIDs include Ebola, HIV/AIDS, hepatitis E and the vector-borne diseases caused by Zika and dengue virus. EIDs pose a risk to transfusion safety, which can be direct, if the agent can be transmitted through blood transfusion, or indirect, where outbreaks reduce the pool of available donors. Although examples of both have been observed, the former will be the focus of this review. Many transfusion risks have become global due to globalization and increased international travel; however, unique region-specific concerns exist. The Asia-Pacific region varies in size depending on context, but for the purposes of this review will be defined as SouthEast Asia, East Asia and Oceania. This region is a hotspot for genetic diversity, as well as the emergence of a number of infectious diseases. The Asia-Pacific region is also geographically, culturally, socioeconomically and climatically diverse. Moreover, the level of sophistication in blood operators within this region varies, from a national supplier based on voluntary blood donors as is seen in Australia to hospital-based services relying on replacement donors, which is seen in many nations across the Pacific. Aims To review current EID risks to blood transfusion safety in the Asia-Pacific region. Methods Review of available public health reports and literature for the occurrence of EIDs in the Asia-Pacific region that pose a transfusion risk. Results EIDs that pose a transfusion risk occur across the Asia-Pacific region, with differing levels of endemicity between and within countries. Reports of hepatitis E virus (HEV) sero-positivity vary from 2Á2% in Fiji to 15Á2% in Papua New Guinea. As a result of high HEV prevalence in the Hokkaido region of Japan, HEV nucleic acid amplification testing (NAT) has been implemented for screening blood donations. Dengue infection is widespread across the Asia-Pacific, with major outbreaks occurring in 2014 in Fiji, Malaysia, the Philippines and Thailand. In Australia, dengue is episodic in the northeast , resulting in donation restrictions in affected areas. Zika virus emerged in the Asia-Pacific in early 2007 and outbreaks have spread across the Pacific, with epidemics in French Polynesia, the Cook Islands, Easter Island and New Caledonia. Due to the theoretical transfusion risk, several preventative procedures were implemented in French Polynesia during an outbreak in 2013, including NAT.
Duffy blood group phenotypes can be predicted by genotyping for single nucleotide polymorphisms (... more Duffy blood group phenotypes can be predicted by genotyping for single nucleotide polymorphisms (SNPs) responsible for the Fy(a) /Fy(b) polymorphism, for weak Fy(b) antigen, and for the red cell null Fy(a-b-) phenotype. This study correlates Duffy phenotype predictions with serotyping to assess the most reliable procedure for typing. Samples, n = 155 (135 donors and 20 patients), were genotyped by high-resolution melt PCR and by microarray. Samples were in three serology groups: 1) Duffy patterns expected n = 79, 2) weak and equivocal Fy(b) patterns n = 29 and 3) Fy(a-b-) n = 47 (one with anti-Fy3 antibody). Discrepancies were observed for five samples. For two, SNP genotyping predicted weak Fy(b) expression discrepant with Fy(b-) (Group 1 and 3). For three, SNP genotyping predicted Fy(a) , discrepant with Fy(a-b-) (Group 3). DNA sequencing identified silencing mutations in these FY*A alleles. One was a novel FY*A 719delG. One, the sample with the anti-Fy3, was homozygous for a 14-bp deletion (FY*01N.02); a true null. Both the high-resolution melting analysis and SNP microarray assays were concordant and showed genotyping, as well as phenotyping, is essential to ensure 100% accuracy for Duffy blood group assignments. Sequencing is important to resolve phenotype/genotype conflicts which here identified alleles, one novel, that carry silencing mutations. The risk of alloimmunisation may be dependent on this zygosity status.
medRxiv (Cold Spring Harbor Laboratory), Apr 22, 2021
ABSTRACTThere have been no comprehensive studies of a full range of blood group polymorphisms wit... more ABSTRACTThere have been no comprehensive studies of a full range of blood group polymorphisms within the Australian population. The problem is compounded by the absence of any databases carrying genomic information on chronically transfused patients and low frequency blood group antigens in Australia. Here, we use RBCeq, a web server-based blood group genotyping software, to identify unique blood group variants among Australians and compare the variation detected versus global data. Whole genome sequencing data was analysed from for 2796 healthy older Australians from the Medical Genome Reference Bank and compared with data from 1000G phase 3 (1KGP3) databases comprising 661 African, 347 American, 503 European, 504 East Asian, and 489 South Asian participants. There were 688 rare variants detected in this Australian sample population, including nine variants that had clinical associations. Notably, we identified 149 variants that were computationally predicted to be novel and deleterious. No clinically significant rare or novel variants were found associated with the genetically complex ABO blood group system. For the Rh blood group system, one novel and 16 rare variants were found. Our detailed blood group profiling results provide a starting point for the creation of an Australian blood group variant database.Key pointsWe identified unique blood group variants among the healthy older Australian population compared with global data using RBCeq software.Our detailed blood group profiling result may be a starting point for the creation of an Australian blood group variant database.
Background Australia is theoretically at risk of epidemic chikungunya virus (CHIKV) activity as t... more Background Australia is theoretically at risk of epidemic chikungunya virus (CHIKV) activity as the principal vectors are present on the mainland Aedes aegypti) and some islands of the Torres Strait (Ae. aegypti and Ae. albopictus). Both vectors are highly invasive and adapted to urban environments with a capacity to expand their distributions into south-east Queensland and other states in Australia. We sought to estimate the epidemic potential of CHIKV, which is not currently endemic in Australia, by considering exclusively transmission by the established vector in Australia, Ae. aegypti, due to the historical relevance and anthropophilic nature of the vector. Methodology/Principal findings We estimated the historical (1995–2019) epidemic potential of CHIKV in eleven Australian locations, including the Torres Strait, using a basic reproduction number equation. We found that the main urban centres of Northern Australia could sustain an epidemic of CHIKV. We then estimated future tre...
Background Since 2015, Zika virus (ZIKV) outbreaks have occurred in the Americas and the Pacific ... more Background Since 2015, Zika virus (ZIKV) outbreaks have occurred in the Americas and the Pacific involving mosquito-borne and sexual transmission. ZIKV has also emerged as a risk to global blood transfusion safety. Aedes aegypti, a mosquito well established in north and some parts of central and southern Queensland, Australia, transmits ZIKV. Aedes albopictus, another potential ZIKV vector, is a threat to mainland Australia. Since these conditions create the potential for local transmission in Australia and a possible uncertainty in the effectiveness of blood donor risk-mitigation programs, we investigated the possible impact of mosquito-borne and sexual transmission of ZIKV in Australia on local blood transfusion safety. Methodology/Principal findings We estimated 'best-' and 'worst-' case scenarios of monthly reproduction number (R 0) for both transmission pathways of ZIKV from 1996-2015 in 11 urban or regional population centres, by varying epidemiological and entomological estimates. We then estimated the attack rate and subsequent number of infectious people to quantify the ZIKV transfusiontransmission risk using the European Up-Front Risk Assessment Tool. For all scenarios and with both vector species R 0 was lower than one for ZIKV transmission. However, a higher risk of a sustained outbreak was estimated for Cairns, Rockhampton, Thursday Island, and theoretically in Darwin during the warmest months of the year. The yearly estimation of the risk of transmitting ZIKV infection by blood transfusion remained low through the study period for all locations, with the highest potential risk estimated in Darwin. Conclusions/Significance Given the increasing demand for plasma products in Australia, the current strategy of restricting donors returning from infectious disease outbreak regions to source plasma PLOS NEGLECTED TROPICAL DISEASES
Summary Background While blood transfusion is an essential cornerstone of hematological care, pat... more Summary Background While blood transfusion is an essential cornerstone of hematological care, patients requiring repetitive transfusion remain at persistent risk of alloimmunization due to the diversity of human blood group polymorphisms. Despite the promise, user friendly methods to accurately identify blood types from next-generation sequencing data are currently lacking. To address this unmet need, we have developed RBCeq, a novel genetic blood typing algorithm to accurately identify 36 blood group systems. Methods RBCeq can predict complex blood groups such as RH, and ABO that require identification of small indels and copy number variants. RBCeq also reports clinically significant, rare, and novel variants with potential clinical relevance that may lead to the identification of novel blood group alleles. Findings The RBCeq algorithm demonstrated 99·07% concordance when validated on 402 samples which included 29 antigens with serology and 9 antigens with SNP-array validation in 14 blood group systems and 59 antigens validation on manual predicted phenotype from variant call files. We have also developed a user-friendly web server that generates detailed blood typing reports with advanced visualization (https://www.rbceq.org/). Interpretation RBCeq will assist blood banks and immunohematology laboratories by overcoming existing methodological limitations like scalability, reproducibility, and accuracy when genotyping and phenotyping in multi-ethnic populations. This Amazon Web Services (AWS) cloud based platform has the potential to reduce pre-transfusion testing time and to increase sample processing throughput, ultimately improving quality of patient care. Funding This work was supported in part by Advance Queensland Research Fellowship, MRFF Genomics Health Futures Mission (76,757), and the Australian Red Cross LifeBlood. The Australian governments fund the Australian Red Cross Lifeblood for the provision of blood, blood products and services to the Australian community.
Background: Immunohematology reference laboratories provide red blood cell (RBC), platelet (PLT),... more Background: Immunohematology reference laboratories provide red blood cell (RBC), platelet (PLT), and neutrophil typing to resolve complex cases, using serology and commercial DNA tests that define clinically important antigens. Broad-range exome sequencing panels that include blood group targets provide accurate blood group antigen predictions beyond those defined by serology and commercial typing systems and identify rare and novel variants. The aim of this study was to design and assess a panel for targeted exome sequencing of RBC, PLT, and neutrophil antigenassociated genes to provide a comprehensive profile in a single test, excluding unrelated gene targets. Study Design and Methods: An overlapping probe panel was designed for the coding regions of 64 genes and loci involved in gene expression. Sequencing was performed on 34 RBC and 17 PLT/neutrophil reference samples. Variant call outputs were analyzed using software to predict star allele diplotypes. Results were compared with serology and previous sequence genotyping data. Results: Average coverage exceeded 250×, with more than 94% of targets at Q30 quality or greater. Increased coverage revealed a variant in the Scianna system that was previously undetected. The software correctly predicted allele diplotypes for 99.5% of RBC blood groups tested and 100% of PLT and HNA antigens excepting HNA-2. Optimal throughput was 12 to 14 samples per run. Conclusion: This single-test system demonstrates high coverage and quality, allowing for the detection of previously overlooked variants and increased sample throughput. This system has the potential to integrate genomic testing across laboratories within hematologic reference settings.
Background and Aims Although Ross River virus (RRV) and Barmah Forest virus (BFV) pose a risk to ... more Background and Aims Although Ross River virus (RRV) and Barmah Forest virus (BFV) pose a risk to transfusion safety, in Australia, blood donations are not screened for either. RRV and BFV pathogenesis is poorly understood; however, mannose binding lectin (MBL) levels have been associated with RRV disease severity. We investigated biological markers to identify early stages of infection in asymptomatic RRV or BFV infection. Methods Samples from 7001 blood donations from donors at risk for RRV and/or BFV were tested for anti-RRV (IgM) and/or anti-BFV (IgM). MBL level was assessed in 139 anti-RRV (IgM) positive samples, 143 anti-BFV (IgM) positive samples and clinical samples (10 RRV, 10 BFV). MCP-1, MIG, TNF-α, IFN-α, IFN-γ, IL-8, IL-10 and IP-10 were quantified in these samples and 50 seronegative samples. Results Anti-RRV (IgM) was detected in 2.3%, and anti-BFV (IgM) in 2.4%, of donations, consistent with asymptomatic infection. MBL deficiency was not associated with seropositivity, but higher MBL levels were evident in RRV patients. IP-10, MIG, MCP-1 and IL-8 were elevated in RRV patients and IFN-γ, MCP-1 and IL-8 higher in BFV patients. For both anti-RRV (IgM) and anti-BFV (IgM) positive donations, IL-8 was elevated compared to IgM negative donors. All statistical analysis unpaired T-test, 95% CI. Conclusions Asymptomatic RRV or BFV infection occurs in blood donors. The frequency of MBL deficiency was similar between samples from clinical and presumed asymptomatic RRV or BFV infection. Measurement of IL-8 may assist in the identification of early stage viral infection and function as a surrogate biomarker for early stage RRV or BFV infection.
Introduction: Dengue poses a problem for safe transfusion of blood components with confirmed repo... more Introduction: Dengue poses a problem for safe transfusion of blood components with confirmed reports of transfusion-transmission in Hong Kong and Singapore. The largest outbreak in 50 years occurred in North Queensland during 2008/2009 with more than 1,000 confirmed cases in Cairns and Townsville. During this outbreak, supplementary questioning for all donors was implemented, and fresh components were not manufactured from at risk donors. We aim to determine the seroprevalence of dengue exposure in this population during this epidemic. Methods: Samples were collected from blood donors during the 2008/2009 epidemic and 3 months after the last confirmed case. These samples were tested for anti-Dengue IgM, IgG and NS1 antigen with commercially available ELISA based assay kits from PanBio. Results: Initial analyses revealed 2.7% of samples from deferred donors were IgM repeat reactive. Of these, 16% were also positive for anti-dengue IgG, while none of these were positive for the NS1 viral antigen. However, two NS1 positives were found in samples collected from deferred donors. Conclusions: This initial analysis represents recent and cumulative past exposure in a presumed asymptomatic population, and will provide documentation of the rate of asymptomatic dengue infection during the epidemic. This data can also be used to assess the risk of dengue becoming endemic in North Queensland given that the mosquito vector is established in this region.
BACKGROUND: MNS hybrid GP(B-A-B) glycophorins are more commonly found in Southeast Asians and all... more BACKGROUND: MNS hybrid GP(B-A-B) glycophorins are more commonly found in Southeast Asians and alloantibodies to antigens they carry are clinically significant. Detection of hybrid glycophorins by serologic techniques is limited due to lack of commercial reagents. In this study, a genotyping method for GP(B-A-B) hybrid glycophorins based on high-resolution melting (HRM) analysis was applied for genotyping analysis in the Chinese Southern Han population. STUDY DESIGN AND METHODS: DNA samples from 3104 Chinese Southern Han blood donors were collected. GYP(B-A-B) genotypes were analyzed by HRM assay. Parts of samples (n 5 106) were also tested by multiplex ligation-dependent probe amplification (MLPA) assay. Direct sequencing was conducted in samples with variant melting curve profiles. RESULTS: A total of five GYP(B-A-B) genotypes (201/ 3104, 6.5%) were identified, which were GYP*Mur heterozygote (n 5 194), GYP*Mur homozygote (n 5 3), GYP*Bun heterozygote (n 5 2), GYP*HF heterozygote (n 5 1), and a novel GYP(B-A-B) hybrid allele (n 5 1). Genotyping results for GYP*Mur and wild-type GYPB samples obtained by HRM were consistent with MLPA, while GYP*Bun and GYP*HF heterozygote identified by HRM could only be identified to have one copy of 5 0 inactive splice site of GYPB Pseudoexon 3 by MLPA. In addition, 10 single-nucleotide polymorphisms (SNPs) including four known and six novel SNPs were identified in 31 samples. One sample was identified carrying both GYP*Mur and GYP*Sch alleles. CONCLUSION: The HRM assay could distinguish the GYP(B-A-B) hybrid alleles successfully. Polymorphisms identified within the GYPB gene should be taken into consideration when developing GYP(B-A-B) genotyping kits for the Chinese population.
Improvements in blood group genotyping methods have allowed large scale population-based blood gr... more Improvements in blood group genotyping methods have allowed large scale population-based blood group genetics studies, facilitating the discovery of rare blood group antigens. Norfolk Island, an external and isolated territory of Australia, is one example of an underrepresented segment of the broader Australian population. Our study utilized whole genome sequencing data to characterize 43 blood group systems in 108 Norfolk Island residents. Blood group genotypes and phenotypes across the 43 systems were predicted using RBCeq. Predicted frequencies were compared to data available from the 1000G project. Additional copy number variation analysis was performed, investigating deletions outside of RHCE, RHD, and MNS systems. Examination of the ABO blood group system predicted a higher distribution of group A1 (45.37%) compared to group O (35.19%) in residents of the Norfolk Island group, similar to the distribution within European populations (42.94% and 38.97%, respectively). Examinatio...
Aim: The emergence of Zika virus (ZIKV) in the Americas has resulted in a public health emergency... more Aim: The emergence of Zika virus (ZIKV) in the Americas has resulted in a public health emergency. Three documented cases of ZIKV transfusion-transmission highlights that this virus is a potential threat to blood transfusion safety. An approach to manage this risk is pathogen inactivation, such as the THERAFLEX MB-PLASMA system. We examined the effectiveness of this system to inactivate ZIKV in plasma at different visible-light doses. Methods: ZIKV was spiked into pooled plasma (n = 3), then treated with the THERAFLEX MB-Plasma system. Pre-and post-treatment samples were taken at each illumination dose (0, 20, 40, 60, 120 J/cm 2) and viral infectivity determined by plaque assay. The reduction in viral infectivity was calculated. Results: Treatment of plasma with the THERFALEX MB-Plasma system resulted in 5.68 log 10 reduction in ZIKV infectivity at 120 J/cm 2 , with residual viral infectivity reaching the limit of detection of the assay with treatment at 40 J/cm 2. Discussion: Our study has shown the THERAFLEX MB-PLASMA system can reduce the infectivity of ZIKV to the limit of detection of the assay used at one third of the standard illumination dose. Our data suggest this system may be an effective option for managing ZIKV transfusion-transmission risk in plasma.
Objective: Maternal allo-antibody production is stimulated when fetal red blood cells are positiv... more Objective: Maternal allo-antibody production is stimulated when fetal red blood cells are positive for an antigen absent on the mother's red cells. The maternal IgG antibodies produced will pass through the placenta and attack fetal red cells carrying the corresponding antigen. Allo-immune hemolytic disease of the fetus and newborn caused by anti-E rarely occurs. Case summary: We report two cases of anti-E hemolytic diseases in neonates. One of the neonates had severe hemolysis presenting with severe anemia, thrombocytopenia, and conjugated hyperbilirubinemia, while the other had moderate anemia and unconjugated hyperbilrubinemia. Although both the neonates were treated by phototherapy and intravenous immunoglobulin, one of them received double volume exchange transfusion. Conclusion: There appeared to be an increase in the occurrence of hemolytic disease of the fetus and newborn caused by Rh antibodies other than anti-D. In this case report, both patients presented with anemia and hyperbilirubinemia but were successfully treated, with a favorable outcome.
aware, there are no previous published reports of a variant t(9;22;11) in a patient with CML and ... more aware, there are no previous published reports of a variant t(9;22;11) in a patient with CML and a history of CLL. Methods: A bone marrow aspirate was analysed by (1) G-band karyotype, (2) fluorescence in situ hybridisation (FISH) using dual fusion BCR-ABL1 and locus specific ASS probes (VYSIS), and (3) 8x60K CGH+SNP microarray using a Cancer Cytogenomic Microarray Consortium (CCMC)-based design (Agilent Technologies). Results: Karyotyping showed an unusual banding pattern of chromosome 4 short arm and a three-way balanced translocation between chromosomes 9, 22 and 11, and the BCR-ABL1 gene fusion was confirmed by FISH: 46,XY,add(4)(p15.2), t(9;22;11)(q34;q11.2;p15).ish (ASS+,ABL1+;BCR+,ABL1+; BCR+). CGH-array showed a 21.5 Mb deletion of chromosome segment 4p16.2->p15.2 but no other copy number abnormalities or loss of heterozygosity. Conclusion: The finding of BCR-ABL1 gene fusion as a result of a variant three-way t(9;22;1) was critical in the change of diagnosis to CML and resultant imatinib base therapy. The 21.4 Mb chromosome 4p deletion is of uncertain significance. Transformation from CLL to CML is not a recognised occurrence. As the patient had never been treated for CLL the development of CML does not appear to be therapeutically induced. Interestingly, since the implementation of imatinib the patient has had a major molecular response and his previous lymphocytosis has resolved.
Background and ObjectivesHepatitis E virus (HEV) is a known transfusion‐transmissible agent. HEV ... more Background and ObjectivesHepatitis E virus (HEV) is a known transfusion‐transmissible agent. HEV infection has increased in prevalence in many developed nations with RNA detection in donors as high as 1 in 600. A high proportion of HEV infections are asymptomatic and therefore not interdicted by donor exclusion criteria. To manage the HEV transfusion‐transmission (TT) risk some developed nations have implemented HEV RNA screening. In Australia, HEV is rarely notified; although locally acquired infections have been reported, and the burden of disease is unknown. The purpose of this study was to determine the frequency of HEV infection in Australian donors and associated TT risk.Materials and MethodsPlasma samples (n = 74 131) were collected from whole blood donors during 2016 and screened for HEV RNA by transcription‐mediated amplification (TMA) in pools of six. Individual TMA reactive samples were confirmed by RT‐PCR and, if positive, viral load determined. Prevalence data from the ...
Aspects of Developmental and Comparative Immunology, 1981
ABSTRACT Previous studies on infectious bursal disease virus (IBDV) have demonstrated that it has... more ABSTRACT Previous studies on infectious bursal disease virus (IBDV) have demonstrated that it has a significant effect upon the avian humoral immune system. In the present study the effect of infectious bursal disease has been assessed sequentially by tests both for humoral and for cell mediated immunity (CMI). Day old chickens were infected with an Australian strain of IBDV of intermediate virulence by the ocular route. Reduced haemagglutination titres to sheep red blood cells and haemagglutination inhibition titres to lentogenic Newcastles disease virus indicates that this virus appears to affect mainly mercaptoethanol resistant antibodies. In addition the results of assays for CMI showed that this strain causes an impairment of this component of the immune system. It was found by immunofluorescent studies of bursa, spleen, thymus and peripheral blood that there was a significant reduction of light chain positive cells in the peripheral blood of infected chickens between two weeks and four weeks. Repopulation of the infected bursa by lymphocytes occurred by four weeks and the serum antibody levels to the above antigens returned to near normal levels by five weeks following infection but the CMI remained depressed.
Background An emerging infectious disease (EID) refers to a disease that has recently appeared in... more Background An emerging infectious disease (EID) refers to a disease that has recently appeared in a population or one that has rapidly increased in incidence or geographic range. Examples of EIDs include Ebola, HIV/AIDS, hepatitis E and the vector-borne diseases caused by Zika and dengue virus. EIDs pose a risk to transfusion safety, which can be direct, if the agent can be transmitted through blood transfusion, or indirect, where outbreaks reduce the pool of available donors. Although examples of both have been observed, the former will be the focus of this review. Many transfusion risks have become global due to globalization and increased international travel; however, unique region-specific concerns exist. The Asia-Pacific region varies in size depending on context, but for the purposes of this review will be defined as SouthEast Asia, East Asia and Oceania. This region is a hotspot for genetic diversity, as well as the emergence of a number of infectious diseases. The Asia-Pacific region is also geographically, culturally, socioeconomically and climatically diverse. Moreover, the level of sophistication in blood operators within this region varies, from a national supplier based on voluntary blood donors as is seen in Australia to hospital-based services relying on replacement donors, which is seen in many nations across the Pacific. Aims To review current EID risks to blood transfusion safety in the Asia-Pacific region. Methods Review of available public health reports and literature for the occurrence of EIDs in the Asia-Pacific region that pose a transfusion risk. Results EIDs that pose a transfusion risk occur across the Asia-Pacific region, with differing levels of endemicity between and within countries. Reports of hepatitis E virus (HEV) sero-positivity vary from 2Á2% in Fiji to 15Á2% in Papua New Guinea. As a result of high HEV prevalence in the Hokkaido region of Japan, HEV nucleic acid amplification testing (NAT) has been implemented for screening blood donations. Dengue infection is widespread across the Asia-Pacific, with major outbreaks occurring in 2014 in Fiji, Malaysia, the Philippines and Thailand. In Australia, dengue is episodic in the northeast , resulting in donation restrictions in affected areas. Zika virus emerged in the Asia-Pacific in early 2007 and outbreaks have spread across the Pacific, with epidemics in French Polynesia, the Cook Islands, Easter Island and New Caledonia. Due to the theoretical transfusion risk, several preventative procedures were implemented in French Polynesia during an outbreak in 2013, including NAT.
Duffy blood group phenotypes can be predicted by genotyping for single nucleotide polymorphisms (... more Duffy blood group phenotypes can be predicted by genotyping for single nucleotide polymorphisms (SNPs) responsible for the Fy(a) /Fy(b) polymorphism, for weak Fy(b) antigen, and for the red cell null Fy(a-b-) phenotype. This study correlates Duffy phenotype predictions with serotyping to assess the most reliable procedure for typing. Samples, n = 155 (135 donors and 20 patients), were genotyped by high-resolution melt PCR and by microarray. Samples were in three serology groups: 1) Duffy patterns expected n = 79, 2) weak and equivocal Fy(b) patterns n = 29 and 3) Fy(a-b-) n = 47 (one with anti-Fy3 antibody). Discrepancies were observed for five samples. For two, SNP genotyping predicted weak Fy(b) expression discrepant with Fy(b-) (Group 1 and 3). For three, SNP genotyping predicted Fy(a) , discrepant with Fy(a-b-) (Group 3). DNA sequencing identified silencing mutations in these FY*A alleles. One was a novel FY*A 719delG. One, the sample with the anti-Fy3, was homozygous for a 14-bp deletion (FY*01N.02); a true null. Both the high-resolution melting analysis and SNP microarray assays were concordant and showed genotyping, as well as phenotyping, is essential to ensure 100% accuracy for Duffy blood group assignments. Sequencing is important to resolve phenotype/genotype conflicts which here identified alleles, one novel, that carry silencing mutations. The risk of alloimmunisation may be dependent on this zygosity status.
medRxiv (Cold Spring Harbor Laboratory), Apr 22, 2021
ABSTRACTThere have been no comprehensive studies of a full range of blood group polymorphisms wit... more ABSTRACTThere have been no comprehensive studies of a full range of blood group polymorphisms within the Australian population. The problem is compounded by the absence of any databases carrying genomic information on chronically transfused patients and low frequency blood group antigens in Australia. Here, we use RBCeq, a web server-based blood group genotyping software, to identify unique blood group variants among Australians and compare the variation detected versus global data. Whole genome sequencing data was analysed from for 2796 healthy older Australians from the Medical Genome Reference Bank and compared with data from 1000G phase 3 (1KGP3) databases comprising 661 African, 347 American, 503 European, 504 East Asian, and 489 South Asian participants. There were 688 rare variants detected in this Australian sample population, including nine variants that had clinical associations. Notably, we identified 149 variants that were computationally predicted to be novel and deleterious. No clinically significant rare or novel variants were found associated with the genetically complex ABO blood group system. For the Rh blood group system, one novel and 16 rare variants were found. Our detailed blood group profiling results provide a starting point for the creation of an Australian blood group variant database.Key pointsWe identified unique blood group variants among the healthy older Australian population compared with global data using RBCeq software.Our detailed blood group profiling result may be a starting point for the creation of an Australian blood group variant database.
Background Australia is theoretically at risk of epidemic chikungunya virus (CHIKV) activity as t... more Background Australia is theoretically at risk of epidemic chikungunya virus (CHIKV) activity as the principal vectors are present on the mainland Aedes aegypti) and some islands of the Torres Strait (Ae. aegypti and Ae. albopictus). Both vectors are highly invasive and adapted to urban environments with a capacity to expand their distributions into south-east Queensland and other states in Australia. We sought to estimate the epidemic potential of CHIKV, which is not currently endemic in Australia, by considering exclusively transmission by the established vector in Australia, Ae. aegypti, due to the historical relevance and anthropophilic nature of the vector. Methodology/Principal findings We estimated the historical (1995–2019) epidemic potential of CHIKV in eleven Australian locations, including the Torres Strait, using a basic reproduction number equation. We found that the main urban centres of Northern Australia could sustain an epidemic of CHIKV. We then estimated future tre...
Background Since 2015, Zika virus (ZIKV) outbreaks have occurred in the Americas and the Pacific ... more Background Since 2015, Zika virus (ZIKV) outbreaks have occurred in the Americas and the Pacific involving mosquito-borne and sexual transmission. ZIKV has also emerged as a risk to global blood transfusion safety. Aedes aegypti, a mosquito well established in north and some parts of central and southern Queensland, Australia, transmits ZIKV. Aedes albopictus, another potential ZIKV vector, is a threat to mainland Australia. Since these conditions create the potential for local transmission in Australia and a possible uncertainty in the effectiveness of blood donor risk-mitigation programs, we investigated the possible impact of mosquito-borne and sexual transmission of ZIKV in Australia on local blood transfusion safety. Methodology/Principal findings We estimated 'best-' and 'worst-' case scenarios of monthly reproduction number (R 0) for both transmission pathways of ZIKV from 1996-2015 in 11 urban or regional population centres, by varying epidemiological and entomological estimates. We then estimated the attack rate and subsequent number of infectious people to quantify the ZIKV transfusiontransmission risk using the European Up-Front Risk Assessment Tool. For all scenarios and with both vector species R 0 was lower than one for ZIKV transmission. However, a higher risk of a sustained outbreak was estimated for Cairns, Rockhampton, Thursday Island, and theoretically in Darwin during the warmest months of the year. The yearly estimation of the risk of transmitting ZIKV infection by blood transfusion remained low through the study period for all locations, with the highest potential risk estimated in Darwin. Conclusions/Significance Given the increasing demand for plasma products in Australia, the current strategy of restricting donors returning from infectious disease outbreak regions to source plasma PLOS NEGLECTED TROPICAL DISEASES
Summary Background While blood transfusion is an essential cornerstone of hematological care, pat... more Summary Background While blood transfusion is an essential cornerstone of hematological care, patients requiring repetitive transfusion remain at persistent risk of alloimmunization due to the diversity of human blood group polymorphisms. Despite the promise, user friendly methods to accurately identify blood types from next-generation sequencing data are currently lacking. To address this unmet need, we have developed RBCeq, a novel genetic blood typing algorithm to accurately identify 36 blood group systems. Methods RBCeq can predict complex blood groups such as RH, and ABO that require identification of small indels and copy number variants. RBCeq also reports clinically significant, rare, and novel variants with potential clinical relevance that may lead to the identification of novel blood group alleles. Findings The RBCeq algorithm demonstrated 99·07% concordance when validated on 402 samples which included 29 antigens with serology and 9 antigens with SNP-array validation in 14 blood group systems and 59 antigens validation on manual predicted phenotype from variant call files. We have also developed a user-friendly web server that generates detailed blood typing reports with advanced visualization (https://www.rbceq.org/). Interpretation RBCeq will assist blood banks and immunohematology laboratories by overcoming existing methodological limitations like scalability, reproducibility, and accuracy when genotyping and phenotyping in multi-ethnic populations. This Amazon Web Services (AWS) cloud based platform has the potential to reduce pre-transfusion testing time and to increase sample processing throughput, ultimately improving quality of patient care. Funding This work was supported in part by Advance Queensland Research Fellowship, MRFF Genomics Health Futures Mission (76,757), and the Australian Red Cross LifeBlood. The Australian governments fund the Australian Red Cross Lifeblood for the provision of blood, blood products and services to the Australian community.
Background: Immunohematology reference laboratories provide red blood cell (RBC), platelet (PLT),... more Background: Immunohematology reference laboratories provide red blood cell (RBC), platelet (PLT), and neutrophil typing to resolve complex cases, using serology and commercial DNA tests that define clinically important antigens. Broad-range exome sequencing panels that include blood group targets provide accurate blood group antigen predictions beyond those defined by serology and commercial typing systems and identify rare and novel variants. The aim of this study was to design and assess a panel for targeted exome sequencing of RBC, PLT, and neutrophil antigenassociated genes to provide a comprehensive profile in a single test, excluding unrelated gene targets. Study Design and Methods: An overlapping probe panel was designed for the coding regions of 64 genes and loci involved in gene expression. Sequencing was performed on 34 RBC and 17 PLT/neutrophil reference samples. Variant call outputs were analyzed using software to predict star allele diplotypes. Results were compared with serology and previous sequence genotyping data. Results: Average coverage exceeded 250×, with more than 94% of targets at Q30 quality or greater. Increased coverage revealed a variant in the Scianna system that was previously undetected. The software correctly predicted allele diplotypes for 99.5% of RBC blood groups tested and 100% of PLT and HNA antigens excepting HNA-2. Optimal throughput was 12 to 14 samples per run. Conclusion: This single-test system demonstrates high coverage and quality, allowing for the detection of previously overlooked variants and increased sample throughput. This system has the potential to integrate genomic testing across laboratories within hematologic reference settings.
Background and Aims Although Ross River virus (RRV) and Barmah Forest virus (BFV) pose a risk to ... more Background and Aims Although Ross River virus (RRV) and Barmah Forest virus (BFV) pose a risk to transfusion safety, in Australia, blood donations are not screened for either. RRV and BFV pathogenesis is poorly understood; however, mannose binding lectin (MBL) levels have been associated with RRV disease severity. We investigated biological markers to identify early stages of infection in asymptomatic RRV or BFV infection. Methods Samples from 7001 blood donations from donors at risk for RRV and/or BFV were tested for anti-RRV (IgM) and/or anti-BFV (IgM). MBL level was assessed in 139 anti-RRV (IgM) positive samples, 143 anti-BFV (IgM) positive samples and clinical samples (10 RRV, 10 BFV). MCP-1, MIG, TNF-α, IFN-α, IFN-γ, IL-8, IL-10 and IP-10 were quantified in these samples and 50 seronegative samples. Results Anti-RRV (IgM) was detected in 2.3%, and anti-BFV (IgM) in 2.4%, of donations, consistent with asymptomatic infection. MBL deficiency was not associated with seropositivity, but higher MBL levels were evident in RRV patients. IP-10, MIG, MCP-1 and IL-8 were elevated in RRV patients and IFN-γ, MCP-1 and IL-8 higher in BFV patients. For both anti-RRV (IgM) and anti-BFV (IgM) positive donations, IL-8 was elevated compared to IgM negative donors. All statistical analysis unpaired T-test, 95% CI. Conclusions Asymptomatic RRV or BFV infection occurs in blood donors. The frequency of MBL deficiency was similar between samples from clinical and presumed asymptomatic RRV or BFV infection. Measurement of IL-8 may assist in the identification of early stage viral infection and function as a surrogate biomarker for early stage RRV or BFV infection.
Introduction: Dengue poses a problem for safe transfusion of blood components with confirmed repo... more Introduction: Dengue poses a problem for safe transfusion of blood components with confirmed reports of transfusion-transmission in Hong Kong and Singapore. The largest outbreak in 50 years occurred in North Queensland during 2008/2009 with more than 1,000 confirmed cases in Cairns and Townsville. During this outbreak, supplementary questioning for all donors was implemented, and fresh components were not manufactured from at risk donors. We aim to determine the seroprevalence of dengue exposure in this population during this epidemic. Methods: Samples were collected from blood donors during the 2008/2009 epidemic and 3 months after the last confirmed case. These samples were tested for anti-Dengue IgM, IgG and NS1 antigen with commercially available ELISA based assay kits from PanBio. Results: Initial analyses revealed 2.7% of samples from deferred donors were IgM repeat reactive. Of these, 16% were also positive for anti-dengue IgG, while none of these were positive for the NS1 viral antigen. However, two NS1 positives were found in samples collected from deferred donors. Conclusions: This initial analysis represents recent and cumulative past exposure in a presumed asymptomatic population, and will provide documentation of the rate of asymptomatic dengue infection during the epidemic. This data can also be used to assess the risk of dengue becoming endemic in North Queensland given that the mosquito vector is established in this region.
BACKGROUND: MNS hybrid GP(B-A-B) glycophorins are more commonly found in Southeast Asians and all... more BACKGROUND: MNS hybrid GP(B-A-B) glycophorins are more commonly found in Southeast Asians and alloantibodies to antigens they carry are clinically significant. Detection of hybrid glycophorins by serologic techniques is limited due to lack of commercial reagents. In this study, a genotyping method for GP(B-A-B) hybrid glycophorins based on high-resolution melting (HRM) analysis was applied for genotyping analysis in the Chinese Southern Han population. STUDY DESIGN AND METHODS: DNA samples from 3104 Chinese Southern Han blood donors were collected. GYP(B-A-B) genotypes were analyzed by HRM assay. Parts of samples (n 5 106) were also tested by multiplex ligation-dependent probe amplification (MLPA) assay. Direct sequencing was conducted in samples with variant melting curve profiles. RESULTS: A total of five GYP(B-A-B) genotypes (201/ 3104, 6.5%) were identified, which were GYP*Mur heterozygote (n 5 194), GYP*Mur homozygote (n 5 3), GYP*Bun heterozygote (n 5 2), GYP*HF heterozygote (n 5 1), and a novel GYP(B-A-B) hybrid allele (n 5 1). Genotyping results for GYP*Mur and wild-type GYPB samples obtained by HRM were consistent with MLPA, while GYP*Bun and GYP*HF heterozygote identified by HRM could only be identified to have one copy of 5 0 inactive splice site of GYPB Pseudoexon 3 by MLPA. In addition, 10 single-nucleotide polymorphisms (SNPs) including four known and six novel SNPs were identified in 31 samples. One sample was identified carrying both GYP*Mur and GYP*Sch alleles. CONCLUSION: The HRM assay could distinguish the GYP(B-A-B) hybrid alleles successfully. Polymorphisms identified within the GYPB gene should be taken into consideration when developing GYP(B-A-B) genotyping kits for the Chinese population.
Improvements in blood group genotyping methods have allowed large scale population-based blood gr... more Improvements in blood group genotyping methods have allowed large scale population-based blood group genetics studies, facilitating the discovery of rare blood group antigens. Norfolk Island, an external and isolated territory of Australia, is one example of an underrepresented segment of the broader Australian population. Our study utilized whole genome sequencing data to characterize 43 blood group systems in 108 Norfolk Island residents. Blood group genotypes and phenotypes across the 43 systems were predicted using RBCeq. Predicted frequencies were compared to data available from the 1000G project. Additional copy number variation analysis was performed, investigating deletions outside of RHCE, RHD, and MNS systems. Examination of the ABO blood group system predicted a higher distribution of group A1 (45.37%) compared to group O (35.19%) in residents of the Norfolk Island group, similar to the distribution within European populations (42.94% and 38.97%, respectively). Examinatio...
Aim: The emergence of Zika virus (ZIKV) in the Americas has resulted in a public health emergency... more Aim: The emergence of Zika virus (ZIKV) in the Americas has resulted in a public health emergency. Three documented cases of ZIKV transfusion-transmission highlights that this virus is a potential threat to blood transfusion safety. An approach to manage this risk is pathogen inactivation, such as the THERAFLEX MB-PLASMA system. We examined the effectiveness of this system to inactivate ZIKV in plasma at different visible-light doses. Methods: ZIKV was spiked into pooled plasma (n = 3), then treated with the THERAFLEX MB-Plasma system. Pre-and post-treatment samples were taken at each illumination dose (0, 20, 40, 60, 120 J/cm 2) and viral infectivity determined by plaque assay. The reduction in viral infectivity was calculated. Results: Treatment of plasma with the THERFALEX MB-Plasma system resulted in 5.68 log 10 reduction in ZIKV infectivity at 120 J/cm 2 , with residual viral infectivity reaching the limit of detection of the assay with treatment at 40 J/cm 2. Discussion: Our study has shown the THERAFLEX MB-PLASMA system can reduce the infectivity of ZIKV to the limit of detection of the assay used at one third of the standard illumination dose. Our data suggest this system may be an effective option for managing ZIKV transfusion-transmission risk in plasma.
Objective: Maternal allo-antibody production is stimulated when fetal red blood cells are positiv... more Objective: Maternal allo-antibody production is stimulated when fetal red blood cells are positive for an antigen absent on the mother's red cells. The maternal IgG antibodies produced will pass through the placenta and attack fetal red cells carrying the corresponding antigen. Allo-immune hemolytic disease of the fetus and newborn caused by anti-E rarely occurs. Case summary: We report two cases of anti-E hemolytic diseases in neonates. One of the neonates had severe hemolysis presenting with severe anemia, thrombocytopenia, and conjugated hyperbilirubinemia, while the other had moderate anemia and unconjugated hyperbilrubinemia. Although both the neonates were treated by phototherapy and intravenous immunoglobulin, one of them received double volume exchange transfusion. Conclusion: There appeared to be an increase in the occurrence of hemolytic disease of the fetus and newborn caused by Rh antibodies other than anti-D. In this case report, both patients presented with anemia and hyperbilirubinemia but were successfully treated, with a favorable outcome.
aware, there are no previous published reports of a variant t(9;22;11) in a patient with CML and ... more aware, there are no previous published reports of a variant t(9;22;11) in a patient with CML and a history of CLL. Methods: A bone marrow aspirate was analysed by (1) G-band karyotype, (2) fluorescence in situ hybridisation (FISH) using dual fusion BCR-ABL1 and locus specific ASS probes (VYSIS), and (3) 8x60K CGH+SNP microarray using a Cancer Cytogenomic Microarray Consortium (CCMC)-based design (Agilent Technologies). Results: Karyotyping showed an unusual banding pattern of chromosome 4 short arm and a three-way balanced translocation between chromosomes 9, 22 and 11, and the BCR-ABL1 gene fusion was confirmed by FISH: 46,XY,add(4)(p15.2), t(9;22;11)(q34;q11.2;p15).ish (ASS+,ABL1+;BCR+,ABL1+; BCR+). CGH-array showed a 21.5 Mb deletion of chromosome segment 4p16.2->p15.2 but no other copy number abnormalities or loss of heterozygosity. Conclusion: The finding of BCR-ABL1 gene fusion as a result of a variant three-way t(9;22;1) was critical in the change of diagnosis to CML and resultant imatinib base therapy. The 21.4 Mb chromosome 4p deletion is of uncertain significance. Transformation from CLL to CML is not a recognised occurrence. As the patient had never been treated for CLL the development of CML does not appear to be therapeutically induced. Interestingly, since the implementation of imatinib the patient has had a major molecular response and his previous lymphocytosis has resolved.
Uploads
Papers by Robert Flower