Nat Sim 1

Hepatitis B (HBV)
Hepatitis B virus is a hepadnavirus -> hepa from hepatotropic (attracted to the liver) and
dna because it is a DNA virus

Can be transmitted through:
Needle prick
Sexual contact
Blood transfusion
Perinatal infection
IDUs (Intravenous drug users)

Hepa B Profile
HBsAg -> Usually used to screen for the presence of the virus
Anti HBs -> Reactive results means a post exposure to the virus, non reactive results
means a pre exposure to the virus
HBeAg -> The presence of HBeAg in a hosts serum is associated with much higher rates of
viral replication and enhanced infectivity. Reactive results mean the patient is infectious,
non-reactive results means the patient is not infectious
Anti HBe -> Rise in the serum means decline in viral replication
HBc Ag -> Not detected in the blood, can be obtained through liver biopsy
Anti HBc Total
IgG (chronic infection or past infection)
IgM (acute infection or recent infection)

Healthy carriers: infected person who is reactive to HBsAG, anti HBe, and anti HBc IgG, but
is non reactive to HBe Ag. Which means the person is not infectious

Serological test for hepatitis B
Principle: Ag + Ab = Ag Ab complex

Screening:
a) EIA (Enzyme immunoassay)
b) ICT (Immuno chromatographic test)
c) Dot blot
d) CLIA (Chemiluminescence assay)

Confirmatory: Neutralization

Monitoring: HBV DNA (PCR)






Screening
EIA (Enzyme immunoassay)
Sandwich technique
Target analyte: Ag
Carrier / support: micro wells
End point: color production
Process:
a) Separated serum is placed in the micro wells coated with HBV antibodies
b) Conjugate (enzyme is added) and is incubated for 90 min
c) Wash is done to remove unbound Ag /Ab
d) Substrate, a light sensitive reagent, which wound react to the conjugate is added
to form color
e) Stopping solution is added to stop the reaction
Result: Reactive (Blue or Purple), Non reactive (Pink or Clear)

ICT (Immuno chromatographic test)
Target analyte: Ag
Carrier / support: Nitrocellulose paper
End point: bond formation

Dot blot
Target analyte: Ag
Carrier / support: Nitrocellulose paper
End point: Dot formation

Confirmatory:
Neutralization
The HbsAg confirmatory assay uses the principle of specific antibody neutralization to
confirm the presence of HbsAg. The confirmatory reagent (human antibody to HbsAg)
is incubated with the specimen in solution. If HbsAg is present in the specimen the
confirmatory reagent will bind it. The neutralized HbsAg is subsequently blocked from
binding to the antibody-coated bead. This results in a reduction of signal when
compared to the non-neutralized specimen in which the negative control is used in
place of the confirmatory reagent. By definition, a specimen is confirmed as positive if
the reduction in signal of the neutralized specimen is at least 50% and the non-
neutralized control generates a signal greater than or equal to the assay cut-off.

Monitoring:
HBV DNA (PCR)
Specific
Measures amount of virus

Notes by Maam Phoebe

Nat Sim 2
Hepatitis C (HCV)

Can be transmitted through:
Post transfusion
Needle prick
Sexual contact
Blood transfusion
Perinatal infection
IDUs (Intravenous drug users)


Principle: Ag + Ab = Ag Ab complex

Screening:
a) EIA (Enzyme immunoassay)
b) ICT (Immuno chromatographic test)
c) Dot blot
d) CLIA (Chemiluminescence assay)

Confirmatory:
a) SIA (Strip immunoassay)
b) Western blot

Monitoring: HCV DNA (PCR)

Screening
EIA (Enzyme immunoassay)
Sandwich technique
Target analyte: Ab
Carrier / support: micro wells
End point: color production
Result: Reactive (Blue or Purple), Non reactive (Pink or Clear)

ICT (Immuno chromatographic test)
Target analyte: Ab
Carrier / support: Nitrocellulose paper
End point: bond formation

Dot blot
Target analyte: Ab
Carrier / support: Nitrocellulose paper
End point: Dot formation




Confirmatory:
Western blot
Target analyte: Ab
Carrier / support: Nitrocellulose paper
End point: bond formation






































Notes by Maam Phoebe
Nat Sim 3
HIV

HIV is a member of the genus Lentivirus part of the family Retroviridae

Species Virulence Infectivity Prevalence Inferred origin
HIV-1 High High Global Common chimpanzee
HIV-2 Lower Low West Africa Sooty Mangabey

Major enzymes of the virus to get in CD4 cell:
a) Reverse transcriptase for DNA transcription when the virus enters the cell
b) Integrase Enzyme used by the virus to go out the nucleus
c) Protease Enzyme used by the virus to go in the nucleus

Some proteins of the virus:
a) p24 (Capsid) First to rise in an infection even in the early stage of infection, most readily
detected
b) gp 120 (Docking glycoprotein)
c) gp41 (Transmembrane glycoprotein)
d) p17 (Matrix)

Serology
Performance of medical tests is often described in terms of:
Sensitivity: The percentage of the results that will be positive when HIV is present
Specificity: The percentage of the results that will be negative when HIV is not present.

Screening:
a) ICT (Immuno chromatographic test)
b) ELISA (Enzyme Linked Immuno Sorbent Assay)

Confirmatory: Western blot

Monitoring:
a) Immunophenotyping (CD4 count)
b) Viral load

Screening: (Sensitive)
ICT (Immuno chromatographic test)
Target analyte: Ab
Carrier / support: Nitrocellulose paper
End point: bond formation





ELISA (Enzyme Linked Immuno Sorbent Assay)
Target analyte: Ab
Genscreen (EVOLIS machine)

Result: Reactive or not reactive

Confirmatory: (Specific)
Western blot
Target analyte: Ab
Carrier / support: Nitrocellulose paper
End point: bond formation
Viral proteins are separated first and immobilized. In subsequent steps, the binding of
serum antibodies to specific HIV proteins is visualized

Result: Positive or negative

Monitoring:
Viral load
Measures amount of virus in a host. Viral load is one of the monitoring tests for HIV

Immunophenotyping (CD4 count)
Principle: Flow cytometry
Measures amount of CD4 left in a host. Immunophenotyping or CD4 count is one of the
monitoring tests for HIV
Specimen of choice: EDTA anticoagulated whole blood

CD4 cell
Helper T-cell or inducer cell
Co receptor of the major histocompatibility complex class II molecule
Coordinator of the immune system

Role in HIV
Assess the degradation of immune deterioration and speed of progression towards
AIDS
Monitor the efficacy of the treatment
Decision points to initiate ART
200 and below CD4 count -> AIDS





Notes by Sir Paul and Maam Zai

Nat Sim 4
Syphilis

Syphilis is a sexually transmitted infection caused by the spirochete bacterium Treponema
pallidum sub species pallidum.
Treponema pallidum sub species pallidum is a cork screw shape bacteria that cant be
cultured in vitro
The primary route of transmission is through sexual contact; it may also be transmitted
from mother to fetus during pregnancy or at birth, resulting to congenital syphilis.
Other human disease caused by related Treponema pallidum include:
a) Yaws (sub species pertenue)
b) Pinta (sub species carateum)
c) Bejel (sub species endemicum)
The signs and symptoms of syphilis vary depending in which of the four stages it presents
(primary, secondary, latent and tertiary).
The Primary stage classically presents with a single chancre (a firm, painless, non-
itchy skin alceration)
The Second stage syphilis classically presents with a diffuse rash, which frequently
involves the palms of the hands and soles of the feet
The Latent syphilis classically presents with little to no symptoms
The tertiary syphilis classically presents with gummas, neurological, or cardiac
symptom.
Specimen of choice: Serum in red or yellow top tube
Vortex is used to homogenize specimen prior to testing
Centrifuge is used to eliminate substances that can cause false negative results

Serology

Screening: RPR (Rapid Plasma Reagin)

Confirmatory:
TPHA (Treponema Pallidum Hemaglutination Assay)
TPPA (Treponema Pallidum Particle Agglutination Assay)

RPR (+), TPHA (+) -> Syphilis
RPR (-), TPHA (+) -> Not active, recent exposure (treatment needed)
RPR (+), TPHA (-) -> Biological false positive

Screening
RPR (Rapid Plasma Reagin)
18 mm circle
Principle: Flocculation
Positive in RPR does not mean positive for syphilis because of other biological factors
The term "reagin" means that this test does not look for antibodies against the actual
bacterium, but rather for antibodies against substances released by cells when they are
damaged by T. pallidum
In addition to screening for syphilis, an RPR level (also called a "titer") can be used to track
the progress of the disease over time and its response to therapy
Confirmatory
TPHA (Treponema Pallidum Hemaglutination Assay)
Avian erythrocyte

TPPA (Treponema Pallidum Particle Agglutination Assay)
Gelatin particles coated with Treponema pallidum
Synthetic particles
In the test, gelatin particles are sensitized with T. pallidum antigen. Patient serum is mixed
with the reagent containing the sensitized gelatin particles. The particles aggregate to
form clumps when the patient serum is positive for syphilis. In other words, the patient's
serum contains antibodies to T. pallidum. A negative test shows no clumping of gelatin
particles. This is a type of specific treponemal test for syphilis.






























Notes by Maam Ada
Nat Sim 5

Gonorrhea

Gonorrhea (also spelled gonorrhoea or gonorrha; colloquially known as the clap) is a common
human sexually transmitted infection caused by the bacterium Neisseria gonorrhoeae. The usual
symptoms in men are burning with urination and penile discharge. Women, on the other hand, are
asymptomatic half the time or have vaginal discharge and pelvic pain. In both men and women, if
gonorrhea is left untreated, it may spread locally, causing epididymitis or pelvic inflammatory
disease or throughout the body, affecting joints and heart valves.

Diagnosis: (GS/CS)
Gram stain
Culture and sensitivity

Neisseria gonorrhoeae

Neisseria gonorrhoeae, also known as gonococci (plural), or gonococcus (singular), is a species
of Gram-negativecoffee bean-shaped diplococci bacteria responsible for the sexually transmitted
infection gonorrhea

Neisseria gonorrhoeae grow on chocolate agar and is isolated on Thayer Martin Agar. Further
testing to differentiate the species include testing for oxidase (positive).

Neisseria gonorrhoeae are motile (twitching motility and possess type IV pili to adhere to surfaces.
The type IV pili operates mechanistically similar to grappling hook. Pili extend and attach to a
substrate that signals the pilus to retract, dragging the cell forward