HCMV infection in immunocompromised and congenital infection : epidemiology , diagnosis and prevention

1. Human cytomegalovirus (HCMV)
In congenital and in immunocompromised patients
Presented by :
Mohammed Al Toubi

2. Outline
 History of HCMV
 Classification
 Structure and replication
 Latency/persistency
 Transmission
 Epidemiology
 Clinical feature
 Diagnosis
 Prevention

3. History of CMV
 Observation : 1881 scientists observed large-inclusion bearing cells in the
kidneys of stillborn infant with syphilis that has a recognizable owl’s eye
cytopathology.
 Reporting of similar finding: scientists described similar inclusions in
human and rabbit following varicella infection and herpes simplex virus.
The condition was named “salivary gland virus disease”
 Laboratory investigation : carried out in vivo using guinea pigs and in
vitro using primary cell culture which provided evidence of a viral
etiology.
 Using microscopy, scientists could see virus-like particles of 199nm in
diameter in the nucleus and cytoplasm of pancreatic cells from an infant.

4. History of CMV
 In 1950s, use of human embryonic cell culture provided the breakthrough
that allowed three investigators namely, Smith, Weller and Rowe to
independently isolate human cytomegalovirus from different sources
 Wyatt et al suggested the name, “generalized cytomegalic inclusion
disease (CID)
 Weller et al (1960) proposed the term “cytomegalovirus” which means
large cell virus
 1970s, it was established that HCMV is a significant pathogen of human
fetus and later it was considered as an important opportunistic infection
of immunocompromised hosts.

5. Classification
 CMV belongs to the family Herpesviridae which contains three
subfamilies : α-, β- and γ
 Cytomegalovirus is a member of the Betaherpesvirinae subfamily
 β- subfamily characteristics :
o species specificity
o slow replication in cell culture
o cell-association
o cell-tropism
 Also β subfamily , greater level of genetic and evolutionary divergence
than the other two subfamilies α-, and Y

6. Structure

7. HCMV genome
S domainLarge domain
Among human herpesviruses, HCMV (HHV-5) has the largest genome of nearly 236
kb of dsDNA that encodes 173 genes

8. gH / gL / UL128-131
gH / gL/gO
Receptor
Receptor
HCMV
Epithelial/endothecilal
cell
Fibroblasts
HCMV has a broad – cell tropisms infecting different human cells

9. HCMV Replication
Main characteristic:
- Entry by
fusion/endocytosis
- Virus DNA replication in
the nucleus
- Slow replication cycle
48-72 hours
- Results in
nuclear/cytoplasmic
inclusion which looks like
owl’s eye

10. Latency , persistent and reactivation
Primary infection –
early life
• Clinically
asymptomatic
• Immunity
controlled
Persistency
• Shedding of virus in some
sites (saliva , urine ) for
months and years
Latency
• viral genome are maintained as
episomes in some cells
• low level or absence of detectable
virus replication
• Limited to myeloid cells
Reactivation

11. Transmission
 Horizontal
-occurs with direct contact with body fluids such as saliva, urine, tears, semen and cervical
secretions
- transmission rate increases in setting where more direct contact occurs in (example
:kindergarten, day care center, sex partners)
- also occurs in blood transfusion and transplantation, particularly organ transplantation.
 Vertical
mother to fetus by three routes:
- transplacental likely caused by recurrent infection
*medically more important
- intrapartum depends on virus shedding patterns
-postnatal - at vaginal
- during breast feeding

12. How many of you are infected
with HCMV??
Is infection high in men or in
women ??

13. Epidemiology
Prevalence
 Seroprevalence of CMV infection is high, with world-wide seroprevalence
between 45 and 100% in adult population.
 For example; in US studies showed CMV prevalence to be 59%
 In general , it is influenced by many geographical and demographical
factors such as location, age, sex, socioeconomic status (SES) and race.
 Risk of infection is highly correlated with the presence of children , and
the more of them the higher the risk.

14. Epidemiology :
immunocompromised patients
 In organs transplant patients
risk of HCMV infection depends on :
- the organ (e.g lung & heart transplant poses high risk =~40%)
-donor/recipients HCVM status
- high risk : (D+R-)
-intermediate risk : (D+R+)
-low risk : (D-R-)
 In congenitally infected patients
-HCMV is the leading cause of congenital infection
-high morbidity , mortality and disease burden
- prevalence reported to be 0.64-1%.

15. Clinical feature:
In immunocompetent
usually asymptomatic.
 symptomatic infection occasionally results in a
febrile, mononucleosis-like illness such as fever (39
°C to 40°C), sore throat, malaise, myalgias, headache,
fatigue, splenomegaly, lymphocytosis and rash

16. Clinical feature:
in immucompromised
 HCMV cause severe to a life-threatening disease, particularly in AIDS and HCT
recipients patients
-due to reactivation or reinfection of HCMV
 In organ transplant patients
- Direct : end-organ disease (EOD) such as pneumonitis, gastrointestinal lesions,
hepatitis, retinitis, pancreatitis, myocarditis, and rarely encephalitis
-Indirect: graft rejection, accelerated coronary artery vasculopathy in heart
transplants and increased risk of bacterial and fungal infection, renal impairment
 In AIDS patients
-cause CMV retinitis (most common)
-pneumonitis and gastroenteritis

17. Clinical feature:
in congenitally infected
 HCMV is the most common nongenetic cause of childhood sensorial
hearing loss (SHL)
 outcome depends on : -Trimester (second and third )
-primary (40% transmission )/non-primary infection
-CMV viral load
 Congenital infection
Symptomatic
12-15 %
Asymptomatic
85-87 %
25 to 56 % develop SHL
4-10% death
30 to 35% CNS sequelae
10-15% develop
sequalae later in
life

18. HCMV diagnosis:
congenital infection
 Laboratory diagnosis is more reliable than clinical
 Tests used for pregnant women include :
- IgG avidity test
- IgM immunoblot
-neutralizing antibody testing
-CMV DNA real-time PCR (qPCR)
* anti-CMV specific IgM antibodies + low avidity anti-CMV IgG= gold
standard test for diagnosis of primary CMV infection in pregnant women
 The presence of CMV IgM may indicate one of the following:
• primary infection
• re-infection
• reactivation
• false-positive test result

19. HCMV diagnosis:
congenital infection
 Screening of pregnant women ???
-not recommended
Reasons - no effective vaccine
-no effective treatment
- infection occurs in seropositive women before pregnancy
 what about TORCH screening for congenital infection ?
- low utility ( very low positive incidence in many screening
program)
- high expense
- not cost-effective

20. HCMV diagnosis
congenital infection
 Prenatal diagnosis
- CMV DNA PCR of amniotic fluid
*sample should be collected 20 or more weeks of gestation & 6 to 8
weeks from the onset of maternal infection
- Ultrasound
* To diagnose systemic infection of fetus by observation of abnormal
extracerebral findings
 Postnatal diagnosis
- CMV DNA PCR using dried blood spots (DBS), Swab , Saliva,Urine
* testing should be done within the first 3 weeks after birth, if not
then infection is not congential

21. HCMV diagnosis:
immunocompromised patients
 Most reliable and valuable laboratory diagnostic tests are quantitative tests
Example : - CMV DNA quantitative qPCR
*Has to be standardized & results are in IU/ml
* use to diagnose active infection , screen for pre-emptive antiviral therapy and
monitor response to antiviral therapy
- direct antigen detection (DAD) or antigenemia (early antigens in blood
leukocytes).
* less reliable and less sensitive
 In Solid organ transplants recipients (SOP)
screen for CMV before transplant with
- CMV-specific immunoglobulin-G (IgG)
- CMV DNA in blood/Urine
-CMV –specific T cell in blood

22. Prevention : congenital infection
 HCMV infection and disease can be prevented, however there are challenges in
preventing horizontal transmission
- continuous shedding of the virus for months and years
- infection is silent in immunocompetent,
 Congenital infection Prevention can be done by
-Hand washing after contact with a child’s saliva or urine
-Avoidance of sharing drink and food with young children
-Avoidance of kissing young children on the mouth
 What abut CMV-Ig ??
should not be routinely administered to pregnant women with primary infection
 What abut antiviral ?
not recommended

23. Prevention
in immunocompromised
 In organ transplants patients
prophylactic: administer anti-CMV drugs to all who are at
increased risk of CMV infection
preemptive therapy: drugs are given only when viral
replication is detected by PCR
* monitor frequency and threshold level of viremia

24. Prevention
in immucompromised
 In SOT patients
prophylactic and preemptive treatment are beneficial to intermediate
risk (D+R- ) and high risk (D+R+) group
 In HCT patients , prophylactic drugs for all R+ and D+/R- HCT recipient
from the time of engraftment until 100 days post-transplantation
 HIV patients
Intravenous ganciclovir (IGCV) is recommended as prophylaxis for HIV-
infected adults, adolescents and children who are HCMV seropositive and
who have CD4+ T-lymphocyte counts less than 50 cells/μl

25. Prevention :
Vaccine
 No commercially licensed vaccines available against HCMV, however
there are ongoing clinical trials for some vaccines
 Vaccine development approaches:
-virus derived- vaccine (ex: attenuated virus )
-protein subunit
- gene-based approaches
 3 HCMV antigens that appear to be of greatest interest for a potential
vaccine:
gB glycoprotein
pentameric complex
pp65 tegument protein
Are the hot topics
in HCMV research

26. Thank you

27. References
 Public Health England , UK Standards for Microbiology Investigations:
Investigation of cytomegalovirus infection
 Ho, M., 2008. The history of cytomegalovirus and its diseases. Medical
Microbiology and . Immunology, 197(2), pp.65–73.
 Crough, T. and Khanna, R., 2009. Immunobiology of Human Cytomegalovirus:
from Bench to Bedside. Clinical Microbiology Reviews, 22(1), pp.76–98.
 Avery, Robin K. et al., 2018. Cytomegalovirus infections in lung and
hematopoietic cell transplant recipients in the Organ Transplant Infection
Prevention and Detection Study: A multi‐year, multicenter prospective cohort
study. Transplant Infectious Disease, 20(3), p.n/a–n/a.
 Rawlinson, W. D., Boppana, S. B., Fowler, K. B., Kimberlin, D. W., Lazzarotto, T.,
Alain, S., Daly, K., Doutré, S., Gibson, L., Giles, M. L., Greenlee, J., Hamilton, S. T.,
Harrison, G. J., Hui, L., Jones, C. A., Palasanthiran, P., Schleiss, M. R., Shand, A.
W., & van Zuylen, W. J., 2017. Congenital cytomegalovirus infection in
pregnancy and the neonate: consensus recommendationsfor prevention,
diagnosis, and therapy. The Lancet Infectious Diseases, 17(6), pp.e177–e188.

28. Oman statistic

29.  Why glycoproteins heavily studied ?
its Role :
- infectivity
- spread of the virus
- induction of the host immune response
 Due to its role , glycoproteins are potential targets for vaccine & antivirals

30. HCMV Envelope
 A lipid bilayer envelope that is composed of mostly
virus-encoded glycoproteins
 As many as 23 glycoproteins found associated with
purified virion
 Main function is attachment and entry, cell fusion,
cell-to-cell transmission and modulation of the host
cell response to the infection.
 Grouped in three glycoprotein complexes:
gC-I (gB)
gC-II ( gM & gN)
gC-III (gH & gH)

31. Glycoproteins Genetic Variability
 Amino Acid (AA) variability of gB was determined to be
resulting in 4 variants
 gN AA variability determined to be resulting in 8 identified
variants
 AA variability of gH approximately resulting in 2 variants (H1,
H2)
 gL AA variability is resulting in 4 variants
 gM is less studied glycoprotein

32. Glycoproteins Genetic Variability
HCMV gene Glycoprotein Amino Acid
variability %
Variants
UL55 gB 9.5% (86/ 906
codons
4 (gB1-4)
UL73 gN 50% (69/138 aa) 8 (gN-1, gN-2,
gN-3a, gN-3b,
gN-4a, gN-4b,
gN-4c, and gN-
4d)
UL100 gM No data
UL75 gH 4.6% (34 aa/743
total aa)
2 (gH1 & gH2)
UL115 gL < 2% (18/ 278 aa) 4 ( gL 1-4 )

33. gL/gH complex
 HCMV has a broad range tropism for human cells
gH/gL
gO
UL128 -UL131
Trimeric
complex
Pentameric
complex
Cell tropism to
fibrolasts & in
vivo infection
Cell tropism
to epithelial
endothelial
and myeloid
cells