What is the problem? CMV in pregnancy - how much testing is enough. Babies are such a nice way to. Bill Rawlinson

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1 in pregnancy - how much testing is enough Babies are such a nice way to start people. ~Don Herrold Bill Rawlinson w.rawlinson@unsw.edu.au What is the problem? Congenital (c) is the most common viral cause of congenital malformation in Australia Estimated annual incidence of congenital infection & disease in Aust & NSW Category Live births Primary maternal infection rate (1.5%) infected infants (40%) Symptomatic infection at birth (15%) Those with fatal infection (30%) Those with severe sequelae (70%) Asymptomatic infection at birth (85%) Those with later sequelae (15%) Total with sequelae Est figure in Australia ~246,000 3,690 1, , Est figure in NSW 86,5000 1,

2 What is the current status? WA 2 NT 3 SA 6 QLD 9 TAS 0 NSW 36 VIC 11 ACT 3 Transmission of c vertically In utero infection in % children born each year in Australia Significant numbers of the 250,000 women with concern regarding the risk serology is not part of routine antenatal screening What is the problem? 90% of babies with c will be clinically normal Pregnant women cannot be adequately retrospectively diagnosed using current tests c Aust Clinical symptoms Neurological abnormality Microcephaly Intracranial calcification Seizures Sensorineural hearing loss Chorioretinitis Developmental delay Total affected infants No. affected /42 % of affected children

3 c Aust Clinical symptoms Abnormality Neonatal death Maternal symptoms Maternal illness No. affected Delayed consequences of infection Later - sensorineural hearing loss (<5yr) - neurodevelopmental delay (<5yr) Eye disease Immunological effects Latency CLINICAL - Late complications Rejection Bacteraemia Fungal infection - aspergillosis Vascular effects - atherosclerosis [Nicols 2001] 3

4 in Neonates Transfusion - seropositive blood 10/74 infection 5/74 infection - seronegative blood 0/90 infection Higher risk - weight < 1250 g - BTF amount > 50 ml - mother seronegative [Yeager, 1974] - prematurity Premature seroconversion 1%-10% [Galea, 1993] Herpesvirus EM (Linda Stannard, University of Cape Town, S.A.) Virus Name Other Name Sub- Typical illness family HHV1 HSV 1 α Cold sores HHV2 HSV 2 α Genital ulcers HHV3 VZV α Chickenpox, Shingles HHV4 EBV β Glandular fever, Nasopharyngeal carcinoma HHV5 H β Mononucleosis, Pneumonia (immunocompromised) HHV6 HHV 6 β Exanthem subitum in children HHV7 HHV 7?β Unknown HHV8 HHV 8 γ 2 Kaposi's sarcoma 4

5 Vertical Transmission Location Trophoblasts infected with detected after in-situ PCR. Trophoblast cells Fetal capillary Endothelial cells Lacunae Stromal layer Branched villous x % (11/94) Cell types in placenta +++ trophoblasts (m) + stromal fibroblasts (m) +/- endothelial cells (f) in placenta Placental infection - reactivation - higher viral load - timing (T1 - T2 - T3) - transient maternal leucocytes - DC in placenta Transmission to placenta spermatozoum / ovum (undifferentiated, receptors) cervix maternal leucocytes Trophoblasts epithelial HLA II Vimentin neg Annexin II pos [Mathys, 1991; Witz, 1999; Trincado, 2002] Placenta - IS-PCR Placenta 5

6 Heart Lung Stillbirth - IS-PCR Intestine Liver Congenital Diagnosis Clinical suspicion IgM seropositive IgG seropositive IgG avidity Maternal urine culture Maternal blood NAT Affected child Amniocentesis NAT Q-NAT Culture Affected child Usually after 4 wks age Dried blood spot Antibody Level Weeks Antibody CF IgM- EIA Time IgG- EIA *Latex Reactivation Years Multiplex PCR % Positive IgG < > 69 Age Range (yrs) Red Cross Cohort SCIP Cohort Simultaneous detection of pathogens without the need for guidance by clinical manifestations High sensitivity and specificity Rapid Smaller sample size required 6

7 DNA1 DNA2 RNA PRO HSV VZV T.gondii PVB, HSV1 VZV, toxoplasma HHV6, HHV7 Adenovirus, AAV Rubella, EV HCV, L GpB Streptococcus Ureaplasma, mycoplasma HHV-6 HHV-7 HHV-8 H Parvovirus B19 Mouse mammary tumour virus Influenza C. trachomatis L Rubella Hep C EV Improved Diagnostics NAD commercial Quantitation Genotyping Better reference standards Better interpretation Peripheral measures to replace amniocentesis ITALIAN MONITORING OF INFECTION IgG/IgM Uninfected Gestation -/- (Weeks) <10 +/- Latent Infection High -/+ +/+ -/+ +/- Repeat IgG/IgM +/+ -/ IgG avidity Seronegative Uninfected Low (Primary Infection) Recurrent Infection Amniocentesis (20ml) PCR Culture 7

8 SCIP serostatus -/- +/- +/+/+ +/+/- Serology 600 pregnant women IgG - IgM - IgG Avidity ND <20 wks gestation 169 >20 wks gestation 90 Total ND % 47% Low High Low avidity <35% 0.8% 6.8% 0.8% Antibody IgG Avidity IgM CF Congenital + +/- + + Primary infection +/ Secondary infection + - +/- + 8

9 Cytomegalovirus % primary infection in pregnancy 10-30% placental transmission to fetus 30% symptomatic - CID, retardation, blindness etc. 18% long-term sequelae - hearing loss, learning difficulties Quantitation of Benefits Surrogate measure of resistance Better correlation with disease Measure of viral load in blood vs other tissue Association with prognosis in some diseases Simplified sample Problems Cost Lack of correlation with some disease Lower sensitivity that qualitative Availability Sample size for testing Untreated Outcomes Antiviral Agents Asymptomatic 90% Symptomatic Issues - Infertility - Developmental delay up to 5 yrs Antiviral agent Aciclovir (ACV) Ganciclovir (GCV) Analogue of nucleoside nucleoside Mode of action Inhibits DNA pol DNA chain termination " Virus HSV VZV HSV VZV Foscarnet (PFA) PPi Inhibits DNA polymerase HSV VZV 9

10 HSV ACV GCV *tk *tk *UL97(pk) ACV-monoP GCV-monoP ACV-diPP ACV-triPPP GCV-diPP ACV-triPPP *DNA polymerase* direct DNA chain inhibition termination Principles Virus intimately related to the cell Development Combinatorial chemistry Fortuitous Specific agents for specific virus Acceptable agents Target essential process low toxicity * usual sites of resistance Issues Should all pregnant women be screened for? How best to monitor the affected child What is the best form of treatment for congenital Prophylaxis? Combination therapy? When will vaccines be available? 10

11 Questions? Some solutions tried D/R matching Blood transfusion screening Hyperimmune Ig ACV prophylaxis VCV prophylaxis GCV prophylaxis GCV pre-emptive therapy VGCV prophylaxis and therapy Combined therapy Prevention of post-transfusion - Pathogen Symptoms leucoreduction or screening Varicella-Zoster virus Herpes simplex virus Parvovirus B19 Toxoplasma gondii Hepatitis C Enterovirus Lymphocytic choriomeningitis virus Human herpesvirus type 6 Human herpesvirus type 7 Human herpesvirus type 8 Limb hypoplasia Neurodevelopmental disability Hydrops fetalis Similar to Chronic hepatitis Fetal death,?diabetes Similar to Australian high risk pts -BMT - stem cell tx - newly diagnosed leukaemics - pts likely to proceed to BMT - seronegative allograft recipients - neonates - HIV seropositive (unproven) [Vox Sanguinis 2002] Every day in Australia at least one child is born with congenital infection, every month, two children are born with perinatal HSV disease, it is likely there are >1000 more children born every year with undiagnosed congenital or perinatal viral infection 11