We investigated the role that erythroviruses (parvovirus B19 and erythrovirus genotypes 2 and 3) play in the lives of immunosuppressed HIV-infected patients with chronic anemia. We screened the serum samples of 428 patients by specific ultrasensitive real-time polymerase chain reaction assay. Sixteen patients had circulating DNA, with no apparent clinical impact. Erythrovirus-associated anemia is an extremely rare event in HIV-infected patient

Bernasconi, Enos

Calmy, Alexandra

Dang, Thanh

Delhumeau, Cécile

Elzi, Luigia

Ferry, Tristan

Hirschel, Bernard

Meylan, Pascal

Rauch, Andri

Schmid, Patrick

Weber, Rainer

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HIV/AIDS • CID 2010:50 (1 January) • 115H I V / A I D SB R I E F R E P O R TInfrequent Replication of ParvovirusB19 and Erythrovirus Genotypes 2and 3 among HIV-Infected Patientswith Chronic AnemiaTristan Ferry,1,8 Bernard Hirschel,1 Thanh Dang,2 Pascal Meylan,2Ce´cile Delhumeau,1 Andri Rauch,3 Rainer Weber,4 Luigia Elzi,5Enos Bernasconi,6 Patrick Schmid,7 and Alexandra Calmy,1for the Swiss HIV Cohort Studya1Geneva University Hospital, Geneva, 2University Hospital Center and Universityof Lausanne, 3University Clinic of Infectious Diseases, University Hospital Bern,and University of Bern, Bern, 4Zu¨rich University Hospital, Zu¨rich, 5BaselUniversity Hospital, Basel, 6Lugano University Hospital, Lugano, and 7St GallenHospital, St Gallen, Switzerland; 8Infectious and Tropical Diseases Unit, HospiceCivils de Lyon, Claude Bernard University Lyon 1, Lyon, Francea Members of the study group are listed at the end of the text.We investigated the role that erythroviruses (parvovirus B19and erythrovirus genotypes 2 and 3) play in the lives ofimmunosuppressed HIV-infected patients with chronic ane-mia. We screened the serum samples of 428 patients by spe-cific ultrasensitive real-time polymerase chain reaction assay.Sixteen patients had circulating DNA, with no apparent clin-ical impact. Erythrovirus-associated anemia is an extremelyrare event in HIV-infected patients.Parvovirus B19 (PVB19), by targeting human erythroid pro-genitors, is responsible for acute anemia among patients withhemolytic disorders [1]. For severely immunocompromisedpatients, including HIV-infected patients with low CD4 cellcounts, the case reports and case series of chronic anemia dueto high levels of PVB19 replication associated with pure redcell aplasia have been reported [2–5]. Recently, 2 variants ofPVB19 belonging to erythroviruses in the Parvovividae family(erythrovirus genotypes 2 and 3, formerly called erythrovirusvariants A6 and V9, respectively) have been described. However,their pathogenic role has not been established, even amongimmunocompromised patients [6, 7].Received 18 May 2009; accepted 17 August 2009; electronically published 1 December2009.Reprints or correspondence: Dr. Tristan Ferry, Infectious Diseases Unit, Hospice Civils deLyon, Croix-Rousse Hospital, 93 Grande rue de la Croix-Rousse, 69004 Lyon, France (tristan.ferry@univ-lyon1.fr).Clinical Infectious Diseases 2010; 50:115–8 2009 by the Infectious Diseases Society of America. All rights reserved.1058-4838/2010/5001-0020$15.00DOI: 10.1086/649004Chronic anemia is common among HIV-infected patientsand is a predictor of shorter survival [8]. PVB19 is consideredto be a rare cause of acute or chronic anemia in the HIV-infected population, despite a high seroprevalence [9]. How-ever, in the study performed by Naides et al [10] in 1993, manypatients who were intolerant to azidothymidine (AZT) hadPVB19 infection. Recent data revealed that, by using a highlysensitive PVB19 polymerase chain reaction (PCR) detectionassay, low titers of PVB19 DNA were associated with mild tosevere chronic anemia for a selection of patients who receiveda renal transplant [11] and that the newly described erythro-virus genotype 3 has been detected in 3 HIV-infected patientswith chronic anemia [6]. We aimed to assess the prevalence ofcirculating erythrovirus DNA in immunocompromised HIV-infected patients with chronic anemia.Methods. HIV-infected patients included in the Swiss HIVCohort Study during the period from January 1998 throughApril 2008 were screened for erythrovirus DNA by use of ahighly sensitive real-time PCR method. Inclusion criteria were(1) Agence Nationale de Recherches sur le SIDA et les He´patitesVirales grade 1 (or higher) persistent anemia (with a hemo-globin level !10.5 g/dL [2 consecutive results with at least 3months between each sampling]), (2) immunosuppression (CD4cell count 500 cells/mL) during the episode of anemia, and (3)availability of at least 1 serum or plasma sample at the time ofthe determination of hemoglobin level or between the 2 deter-minations of hemoglobin level. Patients with ribavirin exposureat any time time between the 2 determinations of hemoglobinlevel were excluded. Patients exposed to myelosuppressive drugswere not excluded, because they might be associated with eryth-rovirus reactivation.DNA was extracted from frozen serum or plasma with theQiagen DNA mini kit (Qiagen). PVB19 and erythrovirus ge-notype 2 or 3 DNA were detected and quantified by an ultra-sensitive real-time PCR targeting the VP1 gene, a well-con-served region of the erythrovirus genomes, using an extractvolume corresponding to 10 mL of plasma in duplicate, withan extra reaction spiked with 1000 copies of PVB19 DNA asan inhibition control. Two forward primers (B19_2785_F1: 5′-CTTACACAAGCCTGGGCAAGTTAG-3′; and B19_2785_F2:5′-CTTACACAAGCCTGGGCGAGTTAG-3′; 50 nM each) and3 reverse primers (B19_2867_R1: 5′-GGGGCCCAGCTTGTA-GCTCATT-3′; B19_2867_R2: 5′-GAGGCCCAGCTTGTAGCT-CATT-3′; and B19_2867_R3: 5′-GAGGCCCAGCTTGTAACT-CATT-3′; 33 nM each) were used. The analytical sensitivity,using a 17-base minor-groove binder probe (B19_2817_P FAM-116 • CID 2010:50 (1 January) • HIV/AIDSTable 1. Baseline Characteristics of Patients With or Without Erythrovirus ReplicationCharacteristicPatients witherythrovirus replication( )np 16Patients withouterythrovirus replication( )np 412 PAge,a years 43.5 (40–46) 44 (40–50) .422Male sex 5 (31.3) 160 (38.8) .541White 12 (75.0) 260 (63.1) .332European ancestry 13 (81.3) 290 (70.4) .416Intravenous drug user 8 (50.0) 145 (35.2) .225Men who have sex with men 3 (18.8) 59 (14.3) .714CD4 cell count,a cells/mL 191.0 (156.5–322.0) 184.5 (77.0–290.0) .223Median log10 HIV-1 RNA level,a copies/mL 4.13 (3.66–11.04) 6.68 (3.66–11.06) .819Patients without HIV-1 replicationa,b 6 (40.0) 150 (36.7) .793Tested positive for HBsAg 2 (13.3) 28 (7.3) .313Tested positive for hepatitis C antibody 7 (43.8) 173 (42.6) .928Duration of AIDS, years 10.1 (4.1–15.7) 8.2 (3.1–13.6) .332Diagnosis of AIDSa,b 8 (57.1) 214 (67.1) .563Received ARTa 11 (68.8) 281 (68.4) .910Received ART with AZTa 6 (37.5) 133 (32.3) .651Hemoglobin level,a g/dL 9.6 (9.3–9.9) 9.4 (8.5–10.0) .574Hemoglobin level !9.5 g/dL 7 (43.8) 206 (50.1) .617NOTE. Data are no. (%) of patients or median value (interquartile range [IQR]). HBsAg, hepatitis B surface antigen;ART, antiretroviral therapy; AZT, azidothymidine; IQR, interquartile range.a During episode of anemia.b The calculation of the percentages was done after considering missing values.TACCCGGTACTAACTAT-MBG-DQ; 100 nM), an ABgeneQPCR Rox mix AB-1139b, and a 15-s, 95C, 1-min, 60C cycleon a taqMan 7900 machine, was close to 1 input copy of viralDNA (the World Health Organization International Standardfor parvovirus B19 DNA was used for calibration—NationalInstitute for Biological Standards code 99/800) [12]. In eachrun, the linearity of the Ct/input DNA relationship was verifiedfrom 10 to 100,000 copies/reaction using appropriate standards.From the entire patient population, we compared the char-acteristics of patients with erythrovirus replication with thosewithout erythrovirus replication, and from that patient pop-ulation, we subsequently compared the number of patients withAZT exposure. We used the x2 test to compare qualitative var-iables, and we used the Fisher exact test (as appropriate) andthe nonparametric Mann-Whitney test to compare continuousvariables. All statistical tests were 2 sided, and a threshold of5% was chosen as a cutoff for declaring statistical significance.We finally went back to the medical files of patients with eryth-rovirus replication to determine the characteristics of anemiaby collecting additional biological data (eg, the mean corpus-cular volume, the reticulocyte cell count, the iron and ferritinserum levels).Results. There were 428 patients who fulfilled the inclusioncriteria. At study entry, 165 (39%) were male, the median agewas 44 years (interquartile range [IQR], 40–50 years), and 153(36%) reported intravenous drug use. The median CD4 cellcount was 185 cells/mL (range, 77–290 cells/mL), with 150 pa-tients having a CD4 cell count of !50 cells/mL. The medianhemoglobin level was 9.4 g/dL (range, 8.5–10.0 g/dL), and 138patients (32%) were exposed to AZT at the time blood sampleswere obtained.Erythrovirus DNA was detected in 16 patients (4%). Theerythrovirus viral load ranged from 18 to 6820 copies/mL andwas detectable at a very low level (!500 copies/mL) in 13 (81%)of the 16 patients. No statistically significant differences in char-acteristics at the time of sampling were noticed after compar-ison of patients with erythrovirus replication with those withouterythrovirus replication (Table 1). Erythrovirus replication wasnot more frequently detected in patients with a CD4 cell countof !200 cells/mL (9 [56%] vs 7 [44%] of 16 patients witherythrovirus replication). Of note, the prevalence of intravenousdrug users seemed to be higher among patients with erythro-virus replication than among patients without erythrovirus rep-lication (8 [50%] of 16 patients vs 145 [35%] of 412 patients;). However, both groups of patients had similar me-Pp .225dian hemoglobin levels (hemoglobin level of 9.6 g/dL [IQR,9.3–9.9 g/dL] for patients with erythrovirus replication vs 9.4g/dL [IQR, 8.5–10.0 g/dL] for patients without erythrovirusreplication; ) and a similar prevalence of severe anemiaPp .574(7 [44%] of 16 patients with erythrovirus replication vs 206[50%] of 412 patients with erythrovirus replication; ).Pp .617Circulating erythrovirus DNA was detected in only 6 (4%) ofthe 139 patients on AZT, and no statistically significant differ-ence was noticed between patients with circulating erythrovirusHIV/AIDS • CID 2010:50 (1 January) • 117DNA and patients without circulating erythrovirus DNA (datanot shown).Finally, the origin of anemia for the 16 patients with erythro-virus replication was mainly considered to be carential (6 pa-tients experienced iron deficiency) or multifactorial (HIV in-fection with or without AZT therapy, cirrhosis, drug abuse,chronic renal insufficiency, and/or acute non–Hodgkin lym-phoma). None of our patients presented with pure red cellaplasia.Discussion. We found that PVB19 and erythrovirus ge-notype 2 or 3 DNA detection was infrequent in the bloodsamples of a large cohort of immunocompromised HIV-in-fected anemic patients, despite using highly sensitive real-timePCR methods. The typical clinical presentation of PVB19 in-fection for HIV-infected patients includes transfusion-depen-dent severe anemia with reticulocytopenia [2–5]. This clinicalform of anemia, which requires multiple infusions of intra-venous immunoglobulins and the introduction of highly activeantiretroviral therapy to recover, is rarely observed, despite ahigh PVB19 seroprevalence in HIV-infected individuals (eg,81% of HIV-infected individuals vs 57% of individuals in thegeneral population of Switzerland) [2–5, 8]. Previous studiesthat tried to detect circulating PVB19 DNA in the blood sam-ples of anemic HIV-infected patients were limited, because thestudies (1) included only a small number of patients; (2) wereperformed before the year 2000, mainly using qualitative dotblot PCR (which may have a low sensitivity) or nested PVB19PCR; and (3) did not detect the newly described erythroviruses,such as erythrovirus genotypes 2 and 3, that may also be in-volved in the process of chronic anemia in immunocompro-mised patients [13–15].Our study, which targeted both PVB19 and erythrovirus ge-notypes 2 and 3 DNA with a real-time PCR, demonstrates thatthe prevalence of erythrovirus replication was low for a largeselection of HIV-infected anemic patients, even for patientswith a low CD4 cell count. Moreover, the viral load of positivesamples was low, probably corresponding to circulating DNAwithout crude active infection and without any clinical signif-icance. The prevalence of 4% observed in our selected popu-lation of HIV-infected patients with chronic anemia was closeto the prevalence of 1% observed in US blood donors in 2000–2003 but seemed to be higher than the prevalence of 0.2%observed in German and Austrian blood donors in 2004–2006[16, 17]. It was noteworthy that, in the latter study, most ofthe patients with circulating PVB19 DNA exhibited a low viralload, !105 copies/mL [17]. This range of viral load may cor-respond to “viral dust,” probably following a symptomatic ornot primary PVB19 old infection. Our observation supportsthe hypothesis that (1) PVB19-induced anemia is an extremelyrare event for HIV-infected patients and (2) erythrovirus ge-notypes 2 and 3 may not have any pathogenic role to play inthis population. Of note, a low CD4 cell count per se does notseem to be sufficient to allow reactivation of PVB19 [18]. Pa-tients who were on AZT at the time of the sampling were notmore likely to experience erythrovirus replication, comparedwith patients who were on other antiretroviral drugs.Our study has some limitations, including the cross-sectionaldesign and the absence of indicators of the circumstances andcharacteristics of anemia (data on the reticulocyte count, themean corpuscular volume of red cells, and the level of serumferritin were not routinely collected in the Swiss HIV CohortStudy). In conclusion, PVB19-associated anemia is a rare eventamong HIV-infected patients, and erythrovirus genotypes 2 and3 may not have any pathogenic role to play in the process ofchronic anemia in such patients. This study does not supportthe extensive use of erythrovirus PCR for HIV-infected patientswith chronic anemia, except in the previously described specificcontext of severe anemia with pure red cell aplasia (which wasnot observed in our study).Swiss HIV Cohort Study. The members of the Swiss HIVCohort Study are as follows: M. Battegay, E. Bernasconi, J. Bo¨ni,H.C. Bucher, P. Bu¨rgisser, A. Calmy, S. Cattacin, M. Cavassini,R. Dubs, M. Egger, L. Elzi, M. Fischer, M. Flepp, A. Fontana,P. Francioli P (President of the Swiss HIV Cohort Study), H.Furrer (Chairman of the Clinical and Laboratory Committee),C.A. Fux, M. Gorgievski, H.F. Gu¨nthard (Chairman of theScientific Board), H.H. Hirsch, B. Hirschel, I. Ho¨sli, C. Kahlert,L. Kaiser, U. Karrer, C. Kind, T. Klimkait, B. Ledergerber, G.Martinetti, N. Mu¨ller, D. Nadal, F. Paccaud, G. Pantaleo, A.Rauch, S. Regenass, M. Rickenbach (Head of Data Center), C.Rudin (Chairman of the Mother and Child Substudy), P.Schmid, D. Schultze, J. Schu¨pbach, R. Speck, B.M. de Tejada,P. Taffe´, A. Telenti, A. Trkola, P. Vernazza, R. Weber R, and S.Yerly.AcknowledgmentsWe would like to acknowledge Nathalie Vallier and Cyril Andre for theirhelp.Financial support. This study has been financed in the framework ofthe Swiss HIV Cohort Study, supported by the Swiss National ScienceFoundation.Potential conflicts of interest. E.B. has received honoraria or travelgrants from Abbott, Boehringer Ingeldheim, Gilead, GlaxoSmithKline,Merck Sharp & Dohme, Pfizer, and Tibotec. All other authors: no conflicts.References1. Heegaard ED, Brown KE. Human parvovirus B19. Clin Microbiol Rev2002; 15:485–505.2. Koduri PR, Kumapley R, Valladares J, Teter C. Chronic pure red cellaplasia caused by parvovirus B19 in AIDS: use of intravenous immu-noglobulin—a report of eight patients. Am J Hematol 1999; 61:16–20.3. Morelli P, Bestetti G, Longhi E, Parravicini C, Corbellino M, MeroniL. Persistent parvovirus B19-induced anemia in an HIV-infected pa-tient under HAART. Case report and review of literature. Eur J ClinMicrobiol Infect Dis 2007; 26:833–7.4. Plentz A, Hahn J, Holler E, Jilg W, Modrow S. Long-term parvovirus118 • CID 2010:50 (1 January) • HIV/AIDSB19 viraemia associated with pure red cell aplasia after allogeneic bonemarrow transplantation. J Clin Virol 2004; 31:16–9.5. Koduri PR. Parvovirus B19-related anemia in HIV-infected patients.AIDS Patient Care STDS 2000; 14:7–11.6. Nguyen QT, Wong S, Heegaard ED, Brown KE. Identification and char-acterization of a second novel human erythrovirus variant, A6. Virology2002; 301:374–80.7. Servant A, Laperche S, Lallemand F, et al. Genetic diversity within humanerythroviruses: identification of three genotypes. J Virol 2002; 76:9124–34.8. Belperio PS, Rhew DC. Prevalence and outcomes of anemia in individualswith human immunodeficiency virus: a systematic review of the literature.Am J Med 2004; 116(Suppl 7A):27S–43S.9. Vernazza PL, Pfister LA, Siegl G, Cassinotti P. High seroprevalence ofparvovirus B19 among patients infected with human immunodeficiencyvirus. Clin Infect Dis 1996; 22:198–9.10. Naides SJ, Howard EJ, Swack NS, True CA, Stapleton JT. ParvovirusB19 infection in human immunodeficiency virus type 1-infected per-sons failing or intolerant to zidovudine therapy. J Infect Dis 1993; 168:101–5.11. Bergallo M, Merlino C, Daniele R, Costa C, Ponzi AN, Cavallo R. Quan-titative competitive-PCR assay to measure human parvovirus B19-DNAload in serum samples. Mol Biotechnol 2006; 32:23–9.12. Saldanha J, Lelie N, Yu MW, Heath A. Establishment of the first WorldHealth Organization International Standard for human parvovirus B19DNA nucleic acid amplification techniques. Vox Sang 2002; 82:24–31.13. Abkowitz JL, Brown KE, Wood RW, et al. Clinical relevance of par-vovirus B19 as a cause of anemia in patients with human immuno-deficiency virus infection. J Infect Dis 1997; 176:269–73.14. Gyllensten K, So¨nnerborg A, Jorup-Ro¨nstro¨m C, Halvarsson M, YunZ. Parvovirus B19 infection in HIV-1 infected patients with anemia.Infection 1994; 22:356–8.15. Raguin G, Leruez-Ville M, Gregoire V, et al. Low prevalence of activeparvovirus B19 infection in HIV-infected patients. Eur J Clin MicrobiolInfect Dis 1997; 16:760–2.16. Kleinman SH, Glynn SA, Lee TH, et al. Prevalence and quantitationof parvovirus B19 DNA levels in blood donors with a sensitive poly-merase chain reaction screening assay. Transfusion 2007; 47:1756–64.17. Schmidt M, Themann A, Drexler C, et al. Blood donor screening forparvovirus B19 in Germany and Austria. Transfusion 2007; 47:1775–82.18. Manning A, Willey SJ, Bell JE, Simmonds P. Comparison of tissuedistribution, persistence, and moleculare epidemiology of parvovirusB19 and novel human parvovirus PARV4 and human bocavirus. J In-fect Dis 2007; 195:1345–52.

Infrequent Replication of Parvovirus B19 and Erythrovirus Genotypes 2 and 3 among HIV-Infected Patients with Chronic Anemia

Infrequent Replication of Parvovirus B19 and Erythrovirus Genotypes 2 and 3 among HIV-Infected Patients with Chronic Anemia

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