Sewage Treatment

 

Sewage and Industrial Effluent Treatment

Sewage treatment (municipal wastewater treatment) and industrial effluent treatment are processes designed to remove physical, chemical, and biological contaminants from wastewater before its discharge into natural water bodies or reuse. The primary objective of wastewater treatment is to protect public health, prevent environmental pollution, and comply with regulatory discharge standards. These treatment systems generally involve physical (primary), biological (secondary), and chemical or advanced (tertiary) treatment processes.

Overall Treatment Flow

Influent → Screening → Grit Removal → Skimming → Primary Sedimentation → Biological Treatment (Trickling Filter / Activated Sludge / Oxidation Ditch) → Secondary Sedimentation → Tertiary Treatment → Treated Effluent

1. Physical Treatment (Preliminary and Primary Treatment)

Physical treatment involves the removal of large, floating, settleable, and suspended solids using mechanical and gravitational methods. It mainly reduces the load on subsequent biological treatment processes.

1.1 Preliminary Treatment

Preliminary treatment is the first step in wastewater treatment and aims to remove gross solids, grit, and floating materials that may damage equipment or hinder treatment efficiency.

a. Screening

Screening is used to remove large floating and suspended materials such as rags, plastics, paper, cloth, sticks, cans, and dead animals. Wastewater is passed through bar screens made of vertical or inclined steel bars spaced about 5 cm apart. Coarse and fine screens are used depending on the treatment requirement that trap large objects. The collected screenings are disposed of by incineration, composting, or landfilling.

b. Grit Removal

Grit consists of heavy, non-biodegradable inorganic particles such as sand, gravel, silt.  Grit removal is carried out in grit chambers by controlling the flow velocity so that grit settles while organic matter remains in suspension. Common units include horizontal flow grit chambers, aerated grit chambers, and vortex grit chambers. Grit removal prevents abrasion of pumps and accumulation of sediments in tanks.

c. Skimming

Skimming is the process of removing floating fats, oils, grease, waxes, and soap scum from wastewater. In skimming tanks, air is bubbled from the bottom, causing oily materials to float to the surface, where they are mechanically skimmed off. Removal of grease prevents interference with oxygen transfer during biological treatment.

1.2 Primary Treatment

Primary treatment focuses on removing remaining suspended solids and reducing the organic load of sewage.

Sedimentation (Primary Clarification)

Sedimentation tanks allow wastewater to flow slowly (1–2 feet per minute), enabling suspended solids to settle at the bottom as primary sludge, while lighter materials float as scum. Sludge scrapers are used to remove settled solids periodically. Primary sedimentation removes approximately 50–70% of suspended solids and 25–40% of BOD.

Note- Sometimes there is the provision of Mechanical and chemical Flocculation

Mechanical Flocculation- Mechanical flocculation improves the removal of fine suspended and colloidal particles. Wastewater is gently stirred using rotating paddles at controlled speeds (~0.43 m/s), causing small particles to aggregate into larger flocs that settle more easily.

Chemical Coagulation and Flocculation- Chemical treatment involves the addition of coagulants such as alum, ferric chloride, or lime. Alum forms aluminum hydroxide [Al(OH)₃] precipitates that trap suspended and colloidal particles, forming larger flocs which settle at the bottom. This method is effective when wastewater contains high colloidal content.

2. Biological Treatment (Secondary Treatment)

Secondary treatment removes dissolved and colloidal organic matter using microorganisms. Microbial metabolism converts organic pollutants into stable inorganic compounds, significantly reducing biochemical oxygen demand (BOD).

MajorBiological Changes

• Organic carbon → CO₂ + H₂O
• Organic nitrogen → NH₃ → NO₃⁻ (nitrification)
• Colloidal matter → biologically coagulated and settled

2.1 Aerobic Treatment (With Oxygen)

Aerobic microorganisms degrade organic matter in the presence of oxygen.

a. Trickling Filter

A trickling filter is an attached-growth system consisting of a filter bed (gravel, stones, or plastic media), rotating spray arms, and a collection system. The sewage effluent or wastewater is sprayed over the bed by using an overhead sprayer, which is rotating at a constant speed; the spraying saturates the effluent with O2. There is a development of a gelatinous microbial layer called the zoogleal layer (bacteria, algae, protozoa, fungi). The fed surface becomes covered with aerobic microbial population comprising bacterial species, including Sphaerotilus natans, Beggiatoa, Flavobacterium, Achromobacter, Zooglea and Pseudomonas, microalgae, microfungi and protozoa. As wastewater trickles through or percolates over the media, organic matter is oxidized. The aerobic microflora decomposes the organic materials into small soluble molecules. Later, the treated effluent collected at the bottom of the tank is passed through the sedimentation tank. Natural airflow provides oxygen. Treated effluent is usually passed to a secondary clarifier.

b. Activated Sludge Process

This is the most widely used aerobic biological treatment method. Wastewater is mixed with activated sludge (microbial biomass) in an aeration tank, where air or oxygen is supplied continuously. Microorganisms consume organic matter, forming flocs. The mixture is then sent to a secondary clarifier where sludge settles. A portion of sludge is recycled (return activated sludge), while excess sludge is removed. This process removes 85–95% of BOD and suspended solids.

Activated sludge system consists of an aeration tank, a settling tank and a sludge return system. At first, sewage from the primary treatment plant is mixed with sludge drawn from the previous batch, which is known as activated sludge or return sludge. The activated sludge contains a large number of microorganisms and serves as an inoculum of microorganisms. After mixing the activated sludge, sewage is placed in an aeration tank. In an aeration tank, Sewage is continuously aerated for 6-8 hours. During this period, microorganisms oxidizes the organic compounds to form CO₂, H₂0 and NO₃, etc.

So, activated sludge is a process for the treatment of domestic and industrial sewage using air and biological flocs composed of bacteria and protozoa that substantially reduce organic materials. This process starts when air is introduced into a sewage that is held in a large aeration tank, combined with the following aerobic microbial decomposers to develop biological floc that decomposes organic matter into simple soluble molecules, amino acids, ammonia, phosphorus, nitrates, CO₂, H₂O, etc.

·        Bacteria: Escherichia, Enterobacter, Achromobacter, Flavobacterium, Pseudomonas, Zooglea, Micrococcus, Sphaerotilus, Beggiatoa, Thiothrix , etc.

·        Protozoa: Amoaebe, Spriotrich and Verticillium. 

·        Filamentous fungi: Geotrichum, Cephalosporium, Penicillum and Cladodsporium.

After oxidation, sewage is passed to the settling tank and left undisturbed for 2-3 hours. Sludge settles to the bottom. This sludge is called activated sludge, which is fully oxidized and is very offensive. This activated sludge can be used as inoculum for the next batch of sewage. Most of the sludge is removed and some is returned to the aeration tank for the next round of treatment.

By the sludge digestion process, the BOD of sewage is reduced by 5-15%.

c. Oxidation Pond (Lagoon / Stabilization Pond)

Oxidation ponds, also known as lagoons, stabilization ponds, or reduced ponds, are large, shallow, open basins used for biological treatment of wastewater. This method is primarily aerobic in nature and is suitable for areas with warm climates and sufficient land availability. In this system, sewage from the primary treatment plant is retained in the pond for a long detention period, typically 10 to 40 days. During this time, microorganisms oxidize the organic matter present in the sewage.

A symbiotic relationship exists between algae and bacteria in oxidation ponds. Algae release oxygen during photosynthesis, which is utilized by aerobic bacteria to oxidize organic compounds. In turn, the carbon dioxide and nutrients released during bacterial oxidation are used by algae for photosynthesis. Additional oxygen is also supplied from the atmosphere since the pond is an open system. The oxidation pond generally remains aerobic during daytime and the early hours of night, allowing aerobic decomposition of organic matter. During late night hours, when photosynthesis ceases, oxygen levels may drop, leading to partial anaerobic conditions and anaerobic decomposition. Oxidation ponds are cost-effective, require minimal mechanical equipment, and provide satisfactory removal of BOD and pathogens.

 

2.2 Anaerobic Treatment (Without Oxygen)

Anaerobic treatment is used for high-strength wastewater and sludge stabilization.

a. Anaerobic Digestion

In sealed, oxygen-free digesters, anaerobic bacteria break down organic matter into biogas containing 60–70% methane (CH₄) and carbon dioxide. Anaerobic digestion reduces sludge volume, stabilizes organic matter, and produces renewable energy.

Anaerobic Treatment of Wastewater

Anaerobic treatment is a biological wastewater treatment process carried out in the absence of oxygen. It is mainly used for high-strength wastewater and for stabilization of sludge produced during primary and secondary treatment. This method is especially suitable for wastewater with high organic load, such as that from distilleries, food processing industries, and municipal sludge. Anaerobic treatment reduces organic matter, produces less sludge compared to aerobic processes, and generates useful energy in the form of biogas.

Septic Tank

A septic tank is a simple anaerobic treatment system commonly used for domestic sewage in areas without centralized sewer systems. It is a closed underground tank where sewage is retained for several hours to days. During this detention period, heavier solids settle at the bottom forming sludge, while lighter materials such as oils and grease float on the surface forming scum. Anaerobic bacteria present in the tank partially digest the organic matter, converting it into gases and stabilized sludge. The clarified effluent flows out of the tank for further treatment or disposal through soak pits or drainage fields. Septic tanks are low-cost systems but provide only partial treatment.

Anaerobic Sludge Digestion

Anaerobic sludge digestion is used for stabilizing sludge generated from primary and secondary treatment units. The process takes place in sealed digesters under controlled temperature conditions (mesophilic or thermophilic). Anaerobic microorganisms decompose complex organic matter through stages such as hydrolysis, acidogenesis, acetogenesis, and methanogenesis. This process significantly reduces sludge volume, destroys pathogens, and produces biogas. Digested sludge is more stable, less odorous, and easier to dewater and dispose of safely.

Biogas Production

Biogas is a valuable by-product of anaerobic treatment and sludge digestion. It mainly consists of methane (60–70%), carbon dioxide (30–40%), and trace amounts of hydrogen sulfide and other gases. Methane-rich biogas can be used as a renewable energy source for cooking, heating, or electricity generation in wastewater treatment plants. Biogas production not only recovers energy but also improves the overall sustainability of wastewater treatment systems.

Advantages of Anaerobic Treatment

·        Suitable for high-strength wastewater

·        Low energy requirement (no aeration needed)

·        Produces renewable energy (biogas)

·        Low sludge production

 

3. Chemical and Advanced Treatment (Tertiary Treatment)

Tertiary treatment is employed when high-quality effluent is required, particularly for reuse or discharge into sensitive environments.

Major Processes

• Coagulation and Flocculation: Removal of fine particles and color
• Filtration: Sand filters, membrane filtration
• Disinfection: Chlorination, UV radiation, ozonation
• Nutrient Removal:
• Nitrogen: Nitrification–denitrification
• Phosphorus: Chemical precipitation using alum or lime

Sludge Treatment and Disposal

Sludge generated during primary and secondary treatment is thickened, digested (aerobically or anaerobically), dewatered, and safely disposed of or reused as fertilizer if free from pathogens and toxic substances.

 

Microscopy

 

Microscopy is the science of using microscopes to observe objects that are too small to be seen with the naked eye. In microbiology, microscopy is essential for studying microorganisms such as bacteria, fungi, protozoa, and viruses.

Importance of Microscopy in Microbiology

–       Study of morphology (shape, size, arrangement)

–       Identification and classification of microorganisms

–       Observation of stained and unstained specimens

–       Understanding cell structure and function

Microscope-

"Micro" refers to tiny, and "scope" refers to view or look at. Microscopes are tools used to enlarge small objects so that they can be observed and studied. They range from a simple magnifying glass to an expensive electron microscope.

In the field of microbiology, the invention of the microscope is mainly credited to Antonie van Leeuwenhoek (1632–1723). He was the first person to observe and describe microorganisms (which he called “animalcules”). He used a simple microscope with very high magnification (about 200–300×). In 1674–1683, he observed bacteria, protozoa, yeast, and sperm cells.

Note- Zacharias Janssen (c. 1590) → Invented the compound microscope

Antonie van Leeuwenhoek → First to use the microscope to study microorganisms

Classification of microscopes.

Microscopes can be classified based on various properties.  Based on the lens system, microscopes are of two types.

Simple microscope	Compound microscope
1. It has a single lens system.	1. It has two or more lens system.
2. Example:-Magnifying glass.	2. Example:-Compound microscope basically used in the laboratory.
3. Reading small letters, simple observation	3. Studying microorganisms, cells
4.Magnification Low (up to ~10×)	4. High (40×–1000×)

Based on the source of illumination, there are two types of microscope:

Light Microscope	Electron Microscope
1.   It uses light as a source of illumination.	1.   It uses an electron or electron gun as a source of illumination.
2.   Lens used in a glass lens	2.   The lens used is an electromagnetic lens.
3.   Here, a vacuum is not needed.	3.   A vacuum must be created since electrons can travel only in a vacuum.
4.   Resolution is low compared to the electron microscope.	4.   Resolution is very high, capable of viewing particles of size 0.1 to 0.2 nanometers (nm).
5.   Image can be directly seen by the human eye.	5.   Since the human eye can’t see electrons. Electrons are converted into an amazing image by striking a fluorescent screen.

Types of Microscope

Bright Field Microscopy

Bright Field Microscopy is the most basic, commonly used light microscopy technique where light passes directly through a sample, producing a dark image on a bright, illuminated background. It is essential for observing stained, fixed, or high-contrast,, colored samples in biology and materials science.

· Imaging Principle: Transmitted white light passes through the specimen, and the image is formed by the absorption of light in denser, stained areas.

· Applications: Commonly used in microbiology for examining stained bacteria and in cell biology for tissue sections.

Magnification

The microscope produces an enlarged image of the object that is examined through it. This enlargement is known as its magnification and is measured in diameter. Eg: A magnifying lens which gives an image 36 times as large as that of the object is said to have a magnification of 36 diameter or 36X(x=times).

Magnification is the process of enlarging the apparent size of an object. It tells how many times the image of the object is larger than the actual object.

Total Magnification=Magnification of Objective Lens×Magnification of Eyepiece (Ocular Lens)

Example:

• Objective lens = 40×
• Eyepiece = 10×
• Objective lens = 40×

Total Magnification=40×10=400×

Note: High magnification without good resolution gives a blurry image.

Resolution

Resolution (or resolving power) is the ability of a microscope to distinguish two points that are very close together as separate entities. In other words, it measures the clarity or detail of the image.

The ability of a microscope to distinguish two closely spaced objects as separate and distinct entities is called resolution. Eg. The human eye has the resolving power of 0.25mm, which means that two dots placed 0.25mm apart can be distinguished as two dots. If the distance between the two dots is less than 0.25, only one dot will be seen.

•        Resolving Power (R.P.) = λ/2 NA where λ= wavelength of light , N.A=Numerical Aperture

or

Formula for Resolution (d):  d=λ/2 NA

Where:

• d= minimum distance between two distinguishable points (in meters or micrometers)
• λ (lambda) = wavelength of light used (usually 400–700 nm for visible light)
• NA = Numerical Aperture of the objective lens

Interpretation:

The formula RP = λ / (2 x NA) indicates that the resolving power is influenced by both the wavelength of light and the numerical aperture.

The resolving power is inversely proportional to the wavelength of light (λ). As the wavelength decreases, the resolving power increases, allowing for better separation of closely spaced objects.

The resolving power is directly proportional to the numerical aperture (NA). Higher numerical apertures lead to greater resolving power. A larger NA allows the lens to capture more diffracted light, contributing to improved resolution.

• Smaller d → better resolution → can see finer details
• Higher NA or shorter wavelength → better resolving power

 

Malaria

                                                                         Malaria

Malaria is a blood-borne disease caused by the protozoan Plasmodium and is typically transmitted through the bite of an infected Anopheles mosquito. Infected mosquitoes carry the Plasmodium parasites. 4 species of malaria parasites can infect humans: Plasmodium vivax, P. ovale, P. malariae, and P. falciparum.

P. falciparum causes a more severe form of the disease and those who contract this form of malaria have a higher risk of death. Plasmodium falciparum is the most virulent species of Plasmodium in humans.

Habitat of Plasmodium Parasites
In Mosquitoes:
Plasmodium parasites develop in the gut and salivary glands of Anopheles mosquitoes. These mosquitoes require a warm and humid environment for their development, making tropical and subtropical regions ideal.

 
In Humans:
Inside humans, Plasmodium parasites inhabit the liver and red blood cells. The liver stage occurs in hepatocytes (liver cells), while the blood stage takes place within red blood cells.

Geographical Distribution
Tropical and Subtropical Regions: Malaria is predominantly found in tropical and subtropical regions where the climate supports the Anopheles mosquito population. This includes parts of Africa, South Asia, Southeast Asia, and Central and South America.

Morphology

The following are the diagnostic forms of parasites found in humans

Ring form (Early Trophozoite)
This is the young trophozoite found inside RBCs.
The name ring is derived from the morphological appearance of the stage, resembling a ring-like structure.
A small cytoplasmic rim and a chromatin dot (nucleus) are seen

Trophozoite (Mature)
RBC starts enlarging (especially in P. vivax and P. ovale).
Trophozoites are larger and more ameboid in shape.
They feed on hemoglobin, and their morphology includes a central nucleus and pigment granules.
Cytoplasm becomes more prominent, chromatin more condensed.
Pigment (hemozoin) may appear as brown-black granules.

Schizont
As the trophozoites mature, they form schizonts.
These structures contain multiple nuclei and are larger than trophozoites.
Contains multiple merozoites (number varies by species: e.g., P. falciparum: 16–32, P. malariae: 6–12).
They eventually rupture the red blood cell, releasing more merozoites into the bloodstream, which can infect new red blood cells.

Gametocytes (Sexual Forms)
Gametocytes are the sexual stage of the parasite and are infectious to mosquitoes.
P. falciparum: crescent or banana-shaped.
P. vivax, P. ovale, P. malariae: round or oval.
There are two types of gametocytes.
Microgamete: male form
Macrogamete: female form
Gametocytes are infective to mosquitoes.
• Male (microgametocytes) and female (macrogametocytes) gametocytes are taken up by mosquitoes during a blood meal.

Sporozoites
• The sporozoites are the infective form and are infectious to humans
• They are found in infected mosquitoes in the salivary glands of female Anopheles mosquitoes.
• Sporozoites are single-nucleated, sickle-shaped structures with equally pointed ends.
• The peripheral fibres serve as an organ of locomotion.
• These infectious forms are injected into the human host's bloodstream by an infected mosquito. They travel to the liver and invade hepatocytes, initiating the exoerythrocytic cycle.

Ookinete
The male and female gametocytes fuse to form the zygote, which then matures into a motile form called an ookinete.
They are elongated, spindle-like (sausage-shaped).
Ookinete invades the midgut wall of the mosquito to develop into an oocyst.

Oocyst
Once the ookinete successfully penetrates the midgut epithelium, it transforms into a rounded structure known as an oocyst.
They are spherical or oval in shape, can undergo sporogony to produce thousands of sporozoites inside.
When mature, the oocyst ruptures, releasing sporozoites into the mosquito hemocoel, which migrate to the salivary glands for the next transmission.

Life Cycle of Malarial Parasites

The life cycle of Plasmodium begins when an infected female Anopheles mosquito bites a human and injects sporozoites, the infective stage, into the bloodstream along with its saliva. These sporozoites circulate in the blood for about 20–30 minutes, after which they quickly leave the circulation and enter the liver cells (hepatocytes). This marks the beginning of the liver or exo-erythrocytic stage. Inside the liver cells, each sporozoite grows and undergoes repeated asexual division to form a large structure called a schizont, which contains thousands of daughter cells known as merozoites. After several days, the infected liver cells burst open, releasing the merozoites into the bloodstream.

Note-  In infections by P. vivax and P. ovale, some sporozoites do not immediately divide but instead become hypnozoites, dormant forms capable of reactivating after months or years and causing relapse.

Once released into the bloodstream, the merozoites initiate the erythrocytic (blood) stage by invading red blood cells (RBCs). Inside each RBC, the parasite first appears as a delicate ring-shaped trophozoite. The trophozoite feeds on hemoglobin and enlarges, eventually developing into a mature trophozoite, and then undergoes nuclear division to form another schizont filled with merozoites. When the schizont becomes mature, the RBC ruptures, releasing numerous merozoites into circulation, which then infect new RBCs. This cyclic rupture of RBCs, typically every 48–72 hours depending on the Plasmodium species, is responsible for the characteristic bouts of fever, chills, and rigors seen in malaria patients. This blood-stage multiplication continues repeatedly and is responsible for the clinical symptoms of the disease.

During these repeated asexual cycles, some merozoites differentiate into sexual forms called gametocytes. These gametocytes—male (microgametocytes) and female (macrogametocytes)—circulate in the bloodstream but do not cause symptoms.

When another female Anopheles mosquito bites the infected human, it ingests these gametocytes along with the blood meal, beginning the mosquito stage of the life cycle. Inside the mosquito’s gut, the gametocytes quickly mature: the microgametocyte produces several flagellated microgametes, while the macrogametocyte develops into a single macrogamete. Fertilization occurs when a microgamete fuses with a macrogamete to form a zygote, which then elongates into a motile form called an ookinete.

The ookinete penetrates the mosquito’s midgut wall and settles beneath its outer lining, where it develops into an oocyst. The oocyst gradually enlarges and undergoes repeated divisions to produce thousands of sporozoites. When the oocyst matures, it bursts, releasing sporozoites into the mosquito’s body cavity. These sporozoites then migrate to the mosquito’s salivary glands, where they are stored. When the mosquito next bites a human, the sporozoites are injected into the bloodstream, thus completing the cycle and initiating a new infection in another host.

 

BNS NAMS Microbiology

 

NATIONAL ACADEMY OF MEDICAL SCIENCES

BNS FIRST YEAR FINAL EXAM 2081

LEVEL:- Bachelor in Nursing Sciences (BNS)

SUBJECT:- Microbiology / Parasitology / Virology 104-2081

1.     Discuss the modes of transmission of the dengue virus and describe the clinical features of dengue virus infection. What preventive measures can be taken to reduce the risk of dengue virus transmission?

The dengue virus is primarily transmitted to humans through the bite of infected female mosquitoes, predominantly of the species Aedes aegypti and Aedes albopictus. The transmission occurs in the following ways:

• Mosquito-Borne Transmission: Aedes mosquitoes acquire the virus by biting an infected person during their viremic phase (when the virus is present in the bloodstream). The infected mosquito becomes a carrier and can transmit the virus to another person during subsequent bites.
• Maternal Transmission: The virus can be transmitted from a pregnant mother to her fetus, particularly during childbirth.
• Blood Transfusion: Infected blood products can potentially transmit the virus.
• Laboratory Transmission: Accidental exposure to the virus in laboratory settings.

Clinical Features of Dengue Virus Infection

The clinical presentation of dengue virus infection can range from mild to severe and is classified into three main forms:

1. Dengue Fever (Mild Form):
• Symptoms:
• Sudden high fever (104°F or higher).
• Severe headache, retro-orbital (behind the eyes) pain.
• Muscle and joint pain ("breakbone fever").
• Rash: Initially maculopapular (flat and raised areas) followed by petechiae (small red spots).
• Nausea, vomiting, and fatigue.
• The fever typically lasts 2–7 days.
2. Dengue Hemorrhagic Fever (DHF):
• Symptoms:
• High fever with increased capillary permeability.
• Abdominal pain and persistent vomiting.
• Hemorrhagic manifestations: Nosebleeds, gum bleeding, or blood in stool/urine.
• Thrombocytopenia: Low platelet count leading to bleeding tendencies.
• Complications: Fluid leakage can lead to shock.
3. Dengue Shock Syndrome (DSS):
• The most severe form, characterized by:
• Profound shock due to severe plasma leakage.
• Organ failure and potentially fatal outcomes if untreated.

Preventive Measures to Reduce the Risk of Dengue Virus Transmission

1. Mosquito Control:
• Eliminate Breeding Sites: Remove standing water from containers, tires, pots, and gutters where mosquitoes breed.
• Insecticide Use: Apply larvicides to stagnant water and spray insecticides in high-risk areas.
2. Personal Protection:
• Wear Protective Clothing: Long-sleeved shirts and pants reduce exposure to mosquito bites.
• Use Insect Repellents: Apply repellents containing DEET, picaridin, or IR3535 on exposed skin.
• Use Bed Nets: Particularly effective in areas with high mosquito populations.
3. Community Engagement:
• Conduct public awareness campaigns on the importance of mosquito control and protection.
• Encourage community cleanup drives to eliminate mosquito habitats.
4. Structural Measures:
• Install window and door screens to prevent mosquitoes from entering homes.
• Use air conditioning to reduce mosquito activity indoors.
5. Health Surveillance:
• Monitor and report dengue cases to identify and respond to outbreaks promptly.
• Implement vector control measures in affected areas.

2.     Define antimicrobial agents. Explain the major mechanisms by which bacteria become resistant to antibiotics. (2+8=10)

Antimicrobial agents are substances that kill or inhibit the growth of microorganisms, including bacteria, viruses, fungi, and parasites. These agents can be naturally derived (e.g., antibiotics from microorganisms), synthetic, or semisynthetic. Examples include penicillin, tetracycline, and sulfonamides.

Bacteria develop resistance to antibiotics through various mechanisms. These can be categorized into intrinsic resistance (naturally occurring) and acquired resistance (developed through mutations or gene acquisition). The major mechanisms include:

1. Enzymatic Degradation or Modification of Antibiotics- Bacteria produce enzymes that inactivate antibiotics by breaking them down or modifying their structure. Examples: Beta-lactamases: Enzymes that hydrolyze the beta-lactam ring in penicillins and cephalosporins.

2. Alteration of Target Sites-  Bacteria modify the target molecule or structure that the antibiotic binds to, reducing the drug's efficacy.Examples: Methicillin-resistant Staphylococcus aureus (MRSA): Alters penicillin-binding proteins (PBPs) to resist beta-lactams.

3. Efflux Pumps- Bacteria use efflux pumps to actively expel antibiotics from the cell, reducing intracellular drug concentration.Examples: Tetracycline resistance: Efflux pumps encoded by genes like tet(A) actively transport tetracycline out of the cell.

4. Reduced Permeability- Bacteria decrease antibiotic entry by modifying or reducing the number of porin channels in the cell membrane. Examples:Gram-negative bacteria: Reduced porin expression prevents entry of beta-lactams and fluoroquinolones.

5. Bypassing Metabolic Pathways- Bacteria develop alternative pathways to bypass the antibiotic's action. Examples: Resistance to sulfonamides and trimethoprim: Bacteria acquire alternate enzymes (e.g., dihydropteroate synthase) to continue folic acid synthesis.

 

3.     List common intestinal protozoa. Describe the life cycle, pathogenesis, and laboratory diagnosis of Entamoeba histolytica.

Some common intestinal protozoa are

·  Entamoeba histolytica: Causes amoebiasis.

·  Giardia lamblia: Causes giardiasis.

·  Cryptosporidium spp.: Causes cryptosporidiosis.

 

The life cycle of E histolytica is relatively simple and consists of infective cysts and the invasive trophozoite stage. The life cycle completes in a single host, i.e, human.

Humans become infected with E. histolytica cysts from contaminated food and water. The mature Cyst is resistant to the low pH of the stomach and remains unaffected by gastric juices. The cyst wall is then lysed by intestinal trypsin, and when the cyst reaches the caecum or lower part of the ileum, excystation occurs. The neutral or alkaline environment and bile components favor excystation.

Excystation of a cyst gives 4 trophozoites.

Trophozoites are active and carried to the large intestine by the peristalsis of the small intestine.

Trophozoites then gain maturity and divide by binary fission.

The trophozoites adhere to the mucus lining of the intestine by lectin and secrete proteolytic enzymes, which cause tissue destruction and necrosis.

Parasite, when it gains access to the blood, migrates and causes extra-intestinal diseases.

When the load of trophozoites increases, some of the trophozoites stop multiplying and revert to cyst form by the process of encystation.

These cysts are released in feces, completing the life cycle.

 

Pathogenesis

a.      Invasive Amoebiasis: Trophozoites invade the intestinal mucosa, causing tissue destruction and ulceration. Leads to amoebic colitis, characterized by dysentery (bloody diarrhea) and abdominal pain.

b.    Extraintestinal Amoebiasis: Trophozoites may enter the bloodstream and disseminate to other organs, particularly the liver, causing amoebic liver abscess.

c. Virulence Factors: Adhesion molecules: Mediate attachment to the intestinal epithelium. Cytotoxins: Induce cell death and tissue damage. Proteolytic enzymes: Degrade host tissues.

 The Laboratory Diagnosis includes

• Microscopic Examination:
• Direct Wet Mount: Detect motile trophozoites or cysts in fresh stool.
• Concentrated Stool Smear: Increases the sensitivity for cyst detection.
• Culture: Stool or tissue samples can be cultured to isolate the parasite.
• Serological Tests: Detect antibodies in cases of extraintestinal amoebiasis (e.g., liver abscess).
• Molecular Methods: PCR is highly sensitive and specific, distinguishing E. histolytica from non-pathogenic species like E. dispar.
• Imaging for Extraintestinal Disease: CT or ultrasound can identify liver abscesses.

4.     Define antigen and antibody. How are neonates protected from infections before their immune system has reached maturity? (2+2+6=10)

Antigen- An antigen is a molecule, usually a protein or polysaccharide, that is recognized by the immune system as foreign. It can trigger an immune response, including the production of antibodies. Examples include toxins, components of pathogens (bacteria, viruses, fungi), and allergens.

Antibody- An antibody is a glycoprotein (also known as an immunoglobulin) produced by B cells in response to an antigen. It specifically binds to the antigen to neutralize it or mark it for destruction by immune cells.

Neonatal Protection from Infections

Neonates have an immature immune system at birth, making them vulnerable to infections. However, they are protected by passive immunity and other mechanisms until their immune system matures.

1. Maternal Antibodies (Passive Immunity)

• During pregnancy, maternal IgG antibodies are transferred across the placenta to the fetus via the neonatal Fc receptor (FcRn).
• These antibodies provide protection against infections the mother has encountered.
• Levels are highest at birth but decline over the first few months of life.

2. Breastfeeding

• Colostrum (the first milk) and breast milk provide antibodies, mainly IgA.
• IgA protects the mucosal surfaces of the respiratory and gastrointestinal tracts.
• Breast milk also contains immune cells, cytokines, and antimicrobial proteins like lactoferrin and lysozyme.

3. Innate Immunity

• Neonates rely on innate immune mechanisms, including:
• Phagocytic cells (e.g., neutrophils, macrophages).
• Complement proteins for pathogen lysis.
• Physical barriers like the skin and mucous membranes.

4. Vaccination-

• Early immunization helps stimulate the neonate’s adaptive immune system against specific pathogens (e.g., BCG for tuberculosis, Hepatitis B vaccine).

5. Write short notes on: (any two) (5 x 2 = 10)

a. Normal flora and their functions in the human body

b. Superficial mycoses

c. Safety Precaution in Microbiology lab

a. Normal flora, also known as microbiota, refers to the group of microorganisms (bacteria, fungi, and viruses) that reside on or in the human body without causing harm under normal conditions. Examples- Skin: Staphylococcus epidermidis, Gastrointestinal Tract: Escherichia coli, Lactobacillus.

Functions:

·       The normal flora synthesize and excrete vitamins in excess of their own needs, which can be absorbed as nutrients by their host. For example, in humans, enteric bacteria secrete Vitamin K and Vitamin B12, and lactic acid bacteria produce certain B-vitamins.

 

·       The normal flora prevent colonization by pathogens by competing for attachment sites or for essential nutrients. 

·      
The normal flora may antagonize other bacteria through the production of substances which inhibit or kill nonindigenous species. The intestinal bacteria produce a variety of substances like fatty acids and peroxides to highly specific bacteriocins, which inhibit or kill other bacteria. 

·       The normal flora stimulate the development of certain tissues, i.e., the caecum and certain lymphatic tissues (Peyer's patches) in the GI tract.

 

·       The normal flora stimulate the production of natural antibodies. Since the normal flora behave as antigens in an animal, low levels of antibodies produced against components of the normal flora are known to cross react with certain related pathogens, and thereby prevent infection or invasion. 

 b. Superficial Mycoses

Superficial mycoses are fungal infections that affect the outermost layers of the skin, hair, and nails without invading deeper tissues.

Common Types:

1. Tinea Versicolor: Caused by Malassezia species, leading to hypo- or hyperpigmented patches on the skin.
2. Tinea Nigra: Caused by Hortaea werneckii, resulting in dark patches on the palms or soles.
3. Black Piedra: Affects the hair shaft, caused by Piedraia hortae.
4. White Piedra: Affects the hair, caused by Trichosporon species.

Treatment:

• Topical antifungal agents (e.g., ketoconazole, terbinafine).
• Good hygiene practices to prevent recurrence.

c. Safety Precautions in a Microbiology Lab

Safety precautions are required to prevent contamination, infection, and accidental exposure to potentially harmful microorganisms.

Major Precautions:

1.     Personal Protective Equipment (PPE):

• Wear lab coats, gloves, and masks.
• Use eye protection when handling hazardous materials.

2.     Sterilization and Disinfection:

• Use autoclaves to sterilize equipment and culture media.
• Disinfect work surfaces before and after use.

3.     Proper Handling of Specimens:

• Avoid direct contact with samples.
• Use aseptic techniques to transfer cultures.

4.     Waste Disposal:

• Dispose of biological waste in designated biohazard bins.
• Sharps (e.g., needles) must be discarded in puncture-proof containers.

5.     Behavioral Practices:

• No eating, drinking, or smoking in the lab.
• Tie back long hair and avoid touching the face.

6.     Emergency Procedures:

• Know the location of eyewash stations and fire extinguishers.
• Report all accidents and spills immediately.