Background
Methods
Study groups and clinical specimens
Sample collection, clinical assessment, screening and serodiagnosis
Genomic DNA isolation and quantification
Estimation of viral load using real time PCR
Statistical analysis
Primer designing for conventional CMV PCR
Virus/genotypes Target genes | Primers and probes [5′ → 3′] |
---|---|
CMV-gB gene | External Primer: |
F: TTT GGA GAA AAC GCC GAC R: CGC GCG GCA ATC GGT TTG TTG TA | |
gB genotypes: gB1 | Internal Primer: |
F: ATG ACC GCC ACT TTC TTA TC R: GTT GAT CCA CRC ACC AGG C | |
gB2 | F: TTC CGA CTT TGG AAG ACC CAA CG R: GTT GAT CCA CRC ACC AGG C |
gB3 | F: TAG CTC CGG TGT GAA CTC C R: GTT GAT CCA CRC ACC AGG C |
gB4 | F: ACC ATT CGT TCC GAA GCC GAG GAG TCA R: GTT GAT CCA CRC ACC AGG C |
gB5 | F: TAC CCT ATC GCT GGA GAA C R: GTT GAT CCA CRC ACC AGG C |
Molecular diagnosis of gB genotypes by multiplex nested PCR [M-nPCR]
DNA sequencing
Bioinformatics and phylogenetic analysis
Results
Clinical manifestations | No. of NB affected [%] |
---|---|
Hepatosplenomegaly | 8 [47.0] |
Neonatal jaundice | 5 [29.4] |
Neonatal cholestasis | 7 [41.1] |
Bronchopneumonia | 3 [17.6] |
Septicemia | 2 [11.7] |
Thrombocytopenia | 6 [35.2] |
Intrauterine growth restriction [IUGR] | 4 [23.5] |
Microcephaly | 3 [17.6] |
Congenital cataract | 1 [5.8] |
Hearing impairment | 1 [5.8] |
Cerebral calcification | 1 [5.8] |
Sample No. | CMV load [log10/ml] | gB genotypes | Clinical significance |
---|---|---|---|
1. | 3.2 × 103 | gB1, gB2 | Hepatosplenomegaly, neonatal cholestasis, microcephaly thrombocytopenia, intrauterine growth restriction, |
2. | 2.7 × 102 | gB1 | Hepatosplenomegaly, neonatal jaundice, neonatal cholestasis |
3. | 1.8 × 104 | gB2 | Thrombocytopenia, intrauterine growth restriction, microcephaly |
4. | 3.6 × 103 | gB1 | Hepatosplenomegaly, neonatal jaundice, neonatal cholestasis |
5. | 5.2 × 103 | gB1, gB2 | Hepatosplenomegaly, neonatal cholestasis, microcephaly thrombocytopenia, intrauterine growth restriction, |
6. | 6.8 × 106 | gB2 | Thrombocytopenia, intrauterine growth restriction, microcephaly |
7. | 3.8 × 103 | gB1, gB3 | Hepatosplenomegaly, neonatal jaundice, neonatal cholestasis, bronchopneumonia, septicemia, thrombocytopenia |
8. | 2.8 × 104 | gB5 | Septicemia, bronchopneumonia, thrombocytopenia |
9. | 4.5 × 102 | gB3 | Bronchopneumonia, septicemia, thrombocytopenia |
10. | 3.8 × 103 | gB5 | Septicemia, bronchopneumonia, thrombocytopenia |
11. | 1.6 × 105 | gB3 | Bronchopneumonia, septicemia, thrombocytopenia |
12. | 6.1 × 103 | gB1 | Hepatosplenomegaly, neonatal jaundice, neonatal cholestasis |
13. | 5.8 × 103 | gB2, gB5 | Thrombocytopenia, microcephaly, septicemia, bronchopneumonia, hepatosplenomegaly, intrauterine growth restriction, |
14. | 3.4 × 103 | gB4 | Congenital cataract, hearing impairment, microcephaly, hepatosplenomegaly |
15. | 2.4 × 105 | gB2 | Thrombocytopenia, intrauterine growth restriction, microcephaly |
16. | 1.8 × 104 | gB3 | Bronchopneumonia, septicemia, thrombocytopenia |
17. | 2.6 × 103 | gB1 | Hepatosplenomegaly, neonatal jaundice, neonatal cholestasis |
FJ527563_AD169 | |||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
KY436004 | 1.9 | ||||||||||||||||
KY436005 | 1.9 | 0.0 | |||||||||||||||
KY436006 | 1.3 | 3.0 | 3.0 | ||||||||||||||
KY436007 | 1.1 | 3.0 | 3.0 | 0.7 | |||||||||||||
KY436008 | 1.1 | 3.0 | 3.0 | 0.7 | 0.0 | ||||||||||||
KY436009 | 1.3 | 3.7 | 3.7 | 1.0 | 1.8 | 1.8 | |||||||||||
KY436010 | 1.3 | 3.7 | 3.7 | 1.0 | 1.8 | 1.8 | 0.0 | ||||||||||
KY436011 | 1.1 | 2.6 | 2.6 | 0.8 | 0.1 | 0.1 | 1.9 | 1.9 | |||||||||
KY436012 | 1.3 | 3.7 | 3.7 | 1.0 | 1.8 | 1.8 | 0.0 | 0.0 | 1.9 | ||||||||
KY436013 | 1.1 | 3.0 | 3.0 | 0.7 | 0.0 | 0.0 | 1.8 | 1.8 | 0.1 | 1.8 | |||||||
KY436014 | 1.3 | 3.7 | 3.7 | 1.0 | 1.8 | 1.8 | 0.0 | 0.0 | 1.9 | 0.0 | 1.8 | ||||||
KY436015 | 1.3 | 3.7 | 3.7 | 1.0 | 1.8 | 1.8 | 0.0 | 0.0 | 1.9 | 0.0 | 1.8 | 0.0 | |||||
KY436016 | 1.9 | 0.0 | 0.0 | 3.0 | 2.6 | 2.6 | 3.7 | 3.7 | 3.0 | 3.7 | 2.6 | 3.7 | 3.7 | ||||
KY436017 | 1.3 | 3.7 | 3.7 | 1.0 | 1.8 | 1.8 | 0.0 | 0.0 | 1.8 | 0.0 | 1.8 | 0.0 | 0.0 | 3.7 | |||
KY436018 | 1.3 | 1.1 | 1.1 | 1.7 | 1.1 | 1.0 | 1.3 | 1.3 | 0.8 | 1.3 | 1.1 | 1.3 | 1.3 | 1.2 | 1.2 | ||
KY436019 | 1.7 | 3.7 | 3.7 | 1.0 | 1.3 | 1.3 | 1.0 | 1.0 | 1.2 | 1.0 | 1.3 | 1.0 | 1.0 |
n/c
| 1.0 | 1.1 | |
KY436020 | 1.1 | 3.0 | 3.0 | 0.7 | 0.0 | 0.0 | 1.8 | 1.8 | 0.1 | 1.8 | 0.0 | 1.8 | 1.8 | 2.6 | 1.8 | 1.1 | 1.3 |