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Time-3 hours. Maximum Marks: 100
Instruction-
1) All questions are compulsory.
2) Draw diagram wherever necessary.
3) Answer of questions and sub question must be written strictly according to serial order of question papers.
**Q.1 Very Short Answer Questions**
**1) Two important aldehydes used for disinfection and their uses:**
- **Formaldehyde**: Used for sterilizing surgical instruments and preserving biological specimens.
- **Glutaraldehyde**: Employed for cold sterilization of heat-sensitive medical equipment like endoscopes.
**2) Two important culture media:**
- **Nutrient Agar**: A general-purpose medium for growing non-fastidious bacteria.
- **MacConkey Agar**: Selective and differential medium for Gram-negative bacteria, particularly lactose fermenters.
**3) Four Gram-positive bacteria:**
1. *Staphylococcus aureus*
2. *Streptococcus pneumoniae*
3. *Bacillus subtilis*
4. *Clostridium tetani*
**4) Define antigen:**
An antigen is a molecule (e.g., protein, polysaccharide) that triggers an immune response, leading to the production of antibodies.
**5) Four DNA viruses and diseases caused:**
1. **Herpes Simplex Virus (HSV)**: Cold sores, genital herpes.
2. **Hepatitis B Virus (HBV)**: Hepatitis B.
3. **Varicella-Zoster Virus (VZV)**: Chickenpox and shingles.
4. **Human Papillomavirus (HPV)**: Warts and cervical cancer.
**6) Four important protozoan parasites:**
1. *Entamoeba histolytica* (amoebiasis)
2. *Plasmodium spp.* (malaria)
3. *Giardia lamblia* (giardiasis)
4. *Toxoplasma gondii* (toxoplasmosis)
**7) Diagram of cyst of *Entamoeba histolytica*:**
The cyst is spherical with 1-4 nuclei. Key features:
- **Chromatoid bodies** (rod-shaped structures).
- **Glycogen vacuole** (stains with iodine).
- Thick cyst wall for environmental survival.
**8) Constituents of MacConkey Agar:**
- **Bile salts** and **crystal violet** (inhibit Gram-positive bacteria).
- **Lactose** (carbohydrate source).
- **Neutral red** (pH indicator: pink/red for lactose fermenters).
- **Agar** (solidifying agent).
**9) Causative agent of malaria and vector:**
- **Agent**: *Plasmodium* spp. (e.g., *P. vivax*, *P. falciparum*).
- **Vector**: Female *Anopheles* mosquito.
**10) Three viruses transmitted by blood transfusion:**
1. **Human Immunodeficiency Virus (HIV)**
2. **Hepatitis B Virus (HBV)**
3. **Hepatitis C Virus (HCV)**
**Q.2 Short Answer Questions**
**1. Difference Between Cell Wall of Gram-Positive and Gram-Negative Bacteria**
Gram-positive and Gram-negative bacteria exhibit fundamental structural differences in their cell walls, which influence their staining properties, pathogenicity, and antibiotic susceptibility.
- **Gram-Positive Bacteria**:
- **Thick Peptidoglycan Layer (20–80 nm)**: Composed of alternating N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) linked by peptide chains. This dense layer retains the crystal violet-iodine complex during Gram staining, appearing purple under a microscope.
- **Teichoic Acids**: Embedded in the peptidoglycan, these polymers regulate cation movement, provide rigidity, and act as virulence factors by mediating host cell adhesion.
- **Absence of Outer Membrane**: Lacks an additional lipid bilayer, making them more permeable to hydrophobic molecules and susceptible to lysozyme and β-lactam antibiotics (e.g., penicillin).
- **Examples**: *Staphylococcus aureus*, *Streptococcus pneumoniae*.
- **Gram-Negative Bacteria**:
- **Thin Peptidoglycan Layer (5–10 nm)**: Surrounded by a hydrophobic **outer membrane** containing lipopolysaccharides (LPS). The LPS (endotoxin) triggers strong immune responses (e.g., septic shock).
- **Periplasmic Space**: A gel-like compartment between the plasma and outer membranes containing hydrolytic enzymes (e.g., β-lactamases) that degrade antibiotics and nutrients.
- **Porins**: Channel proteins in the outer membrane selectively permit small molecules, contributing to antibiotic resistance (e.g., against vancomycin).
- **Examples**: *Escherichia coli*, *Pseudomonas aeruginosa*.
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**2. Bacterial Growth Curve**
The bacterial growth curve describes population dynamics in a closed culture system, divided into four phases:
1. **Lag Phase**:
- **Duration**: Minutes to hours.
- **Activity**: Cells synthesize enzymes, repair DNA, and adjust to environmental conditions (e.g., pH, temperature). No cell division occurs, but metabolic activity is high.
- **Factors Influencing Duration**: Nutrient availability, inoculum age, and stress adaptation.
2. **Exponential (Log) Phase**:
- **Characteristics**: Binary fission occurs at a constant rate, resulting in a logarithmic increase in cell number. Cells are metabolically active and highly susceptible to antibiotics targeting cell wall synthesis (e.g., penicillin).
- **Applications**: Used in industrial fermentation (e.g., insulin production) and research to study uniform microbial populations.
3. **Stationary Phase**:
- **Triggers**: Depletion of nutrients (e.g., carbon, nitrogen), accumulation of toxic waste (e.g., organic acids), and oxygen limitation.
- **Survival Strategies**: Cells produce secondary metabolites (e.g., antibiotics) and form endospores (in *Bacillus* and *Clostridium*).
4. **Death Phase**:
- **Decline**: Cell death exceeds replication due to prolonged nutrient scarcity and toxin buildup. Some persister cells may survive, leading to potential regrowth.
This model is critical for understanding microbial behavior in infections, bioremediation, and food preservation.
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**3. Hot Air Oven and Its Uses**
A hot air oven is a sterilization device that employs **dry heat** to eliminate all microbial life, including heat-resistant spores.
- **Principle**: Heat conduction at 160–180°C for 2–3 hours denatures proteins and oxidizes cellular components.
- **Sterilization Cycle**:
- **Preheating**: Ensures uniform temperature distribution.
- **Holding Time**: Maintains the target temperature to achieve sterility.
- **Cooling**: Prevents thermal shock to glassware.
**Applications**:
- **Laboratory Glassware**: Petri dishes, pipettes, and flasks.
- **Metals**: Surgical instruments (scalpels, scissors) and needles.
- **Heat-Stable Non-Aqueous Materials**: Paraffin wax, oils, and powders.
**Limitations**:
- Unsuitable for rubber, plastics, or liquids (deforms or evaporates).
- Less effective than autoclaves (moist heat penetrates faster).
**Validation**: Spore strips of *Bacillus subtilis* (non-pathogenic) are used to confirm sterilization efficacy.
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**4. Fixatives and Their Uses in Histology**
Fixatives preserve tissue architecture by inhibiting autolysis and bacterial decomposition. Key types include:
1. **Neutral Buffered Formalin (10%)**:
- **Mechanism**: Cross-links proteins via methylene bridges.
- **Use**: Routine histopathology due to excellent nuclear and cytoplasmic detail.
- **Drawback**: Over-fixation causes tissue hardening.
2. **Ethanol (70–100%)**:
- **Mechanism**: Dehydrates tissues and precipitates proteins.
- **Use**: Cytology smears (e.g., Pap tests) and glycogen preservation.
3. **Glutaraldehyde (2–4%)**:
- **Mechanism**: Stabilizes proteins and lipids via cross-linking.
- **Use**: Electron microscopy for ultrastructural clarity.
4. **Bouin’s Fluid**:
- **Composition**: Picric acid, formaldehyde, acetic acid.
- **Use**: Embryonic and delicate tissues (minimizes shrinkage).
5. **Zenker’s Fixative**:
- **Composition**: Mercuric chloride, potassium dichromate.
- **Use**: Enhances connective tissue (collagen) staining.
**Factors Affecting Fixation**: pH, temperature, tissue thickness, and duration. Optimal fixation ensures accurate diagnosis in pathology.
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**5. Life Cycle of *Plasmodium vivax* (Detailed Description)**
*Plasmodium vivax*, a malaria parasite, undergoes a complex life cycle in **two hosts**:
**A. Human Host (Asexual Cycle)**:
1. **Infection**: Infected female *Anopheles* mosquito injects **sporozoites** into the bloodstream during a blood meal.
2. **Liver Stage (Exo-erythrocytic Schizogony)**:
- Sporozoites invade hepatocytes → Multiply into thousands of **merozoites** (7–10 days).
- Some remain dormant as **hypnozoites**, causing relapses.
3. **Erythrocytic Stage**:
- Merozoites infect red blood cells (RBCs) → Develop into **trophozoites** (ring stage) → Mature into **schizonts** → Rupture RBCs, releasing new merozoites (causing cyclical fever).
- Some merozoites differentiate into **gametocytes** (sexual forms).
**B. Mosquito Host (Sexual Cycle)**:
1. **Gametogenesis**: Gametocytes ingested by a mosquito during feeding → Gametes form in the midgut.
- Male gametocyte → Exflagellates into microgametes.
- Female gametocyte → Macrogamete.
2. **Fertilization**: Microgamete fertilizes macrogamete → Forms **zygote**.
3. **Ookinete Formation**: Zygote transforms into motile **ookinete** → Penetrates the midgut wall.
4. **Oocyst Development**: Ookinete becomes an **oocyst** → Releases **sporozoites** via sporogony (10–24 days).
5. **Transmission**: Sporozoites migrate to salivary glands, ready to infect a new human host.
**Clinical Correlation**: Fever coincides with RBC rupture; hypnozoites cause recurrent malaria.
**Q.3 Long type Questions**
**1) Various Classes of Antibodies and Structure/Properties of IgG**
Antibodies, or immunoglobulins (Ig), are classified into five primary isotypes based on their heavy chains: **IgG, IgA, IgM, IgE, and IgD**.
**Structure of IgG**:
IgG is a monomeric Y-shaped glycoprotein composed of **two identical heavy (γ) chains** and **two identical light chains** (κ or λ), interconnected by disulfide bonds. Each chain contains a **variable region** (antigen-binding site) and a **constant region** (determines effector functions). The molecule has two antigen-binding fragments (Fab) and a crystallizable fragment (Fc) that mediates interactions with immune cells. IgG is further divided into four subclasses (IgG1–IgG4) in humans, differing in hinge region flexibility and biological activity.
**Properties of IgG**:
- **Most abundant antibody**: Constitutes ~75% of serum antibodies.
- **Long half-life**: ~21–28 days, ensuring sustained immunity.
- **Crosses the placenta**: Provides passive immunity to neonates.
- **Effector functions**: Opsonizes pathogens for phagocytosis, activates the classical complement pathway, and neutralizes toxins/viruses.
- **Secondary immune response**: Predominantly produced during memory immune reactions.
**2) Important Nematodes and Life Cycle/Pathogenesis/Diagnosis of Ascaris lumbricoides**
Three medically significant nematodes are:
1. **Ascaris lumbricoides** (roundworm)
2. **Ancylostoma duodenale** (hookworm)
3. **Enterobius vermicularis** (pinworm)
**Life Cycle of *Ascaris lumbricoides***:
1. **Egg ingestion**: Fertilized eggs in contaminated soil are ingested.
2. **Larval hatching**: Larvae emerge in the small intestine, penetrate the intestinal wall, and migrate via blood to the lungs.
3. **Pulmonary phase**: Larvae ascend the bronchial tree, are coughed up, swallowed, and return to the intestine.
4. **Maturation**: Develop into adult worms (15–35 cm long) within the intestine, producing ~200,000 eggs/day.
**Pathogenesis**:
- **Larval migration**: Causes Löffler’s syndrome (eosinophilic pneumonia, cough, fever).
- **Intestinal obstruction**: Heavy worm burdens lead to blockage, malnutrition, or appendicitis.
- **Hypersensitivity**: Allergic reactions to worm antigens.
**Laboratory Diagnosis**:
- **Stool microscopy**: Detection of characteristic round, bile-stained eggs with a bumpy outer layer.
- **Adult worms**: Occasionally expelled in stool or vomit.
- **Imaging**: X-rays or ultrasounds for intestinal or biliary complications.
- **Serology**: ELISA to detect anti-Ascaris antibodies (less common).
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