Microbiology Y1 factsheet:

 Yersina Pestis: Plague, blood-borne & transmitted by wild black rats & their fleas,
Gram -ve coccobacillus
 Average size of bacteria: 2-4 micrometers (um)

 Pseudomonas aeruginosa is an opportunistic pathogen

 Flagella is used for motility

 Fimbria/pilae- used to attach to surfaces
 Cell wall of fungi- chitin based (thus b-lactam antibiotics do not work)
 Cell membrane of fungi- ergosterol based (amphotericin B, nystatin and azoles target
ergosterol)

Gram staining is a fundamental technique used in bacteriology to classify bacteria into two
major groups: Gram-positive and Gram-negative. This distinction is based on the chemical and
physical properties of their cell walls. Here’s a step-by-step explanation of the Gram staining
process and the differences between Gram-positive and Gram-negative bacteria using
examples relevant for MBBS students.
Gram Staining Procedure:

 Fixation: A bacterial smear is prepared on a microscope slide and heat-fixed. This kills
the bacteria, making them adhere to the slide and enabling them to take up the stain.
 Crystal Violet Staining: The slide is covered with crystal violet dye for about one minute.
Both Gram-positive and Gram-negative bacteria absorb the crystal violet and appear
purple.
 Mordant (Iodine) Application: Iodine, which serves as a mordant, is added for one
minute. Iodine helps to fix the dye within the cell by forming a complex with the crystal
violet.
  Decolorization: Alcohol or acetone-alcohol is briefly applied to the slide. This is the
critical step that differentiates Gram-positive from Gram-negative bacteria. Gram-positive
bacteria retain the crystal violet-iodine complex due to their thick peptidoglycan layer,
while in Gram-negative bacteria, the outer membrane is disrupted, and the thinner
peptidoglycan layer does not retain the dye.

 Counterstain (Safranin or Fuchsine): After decolorization, the slide is washed and

counterstained with safranin or fuchsine for about 30 seconds to 1 minute. Gram-
negative bacteria, having lost the initial purple color, pick up this second dye and appear
red or pink, while Gram-positive bacteria retain the purple color.

Gram-Positive Bacteria
Gram-positive bacteria have a thick peptidoglycan layer (20-80 nm) in the cell wall that retains
the crystal violet-iodine complex even after the decolorization step. This results in them
appearing purple under the microscope.
Example: Staphylococcus aureus is a Gram-positive bacterium that appears as purple cocci
(spherical cells) in clusters. It is clinically significant as it can cause skin infections, pneumonia,
and food poisoning.

Gram-Negative Bacteria
Gram-negative bacteria have a more complex cell wall structure with a thin peptidoglycan layer
(7-8 nm) situated between an inner cytoplasmic membrane and an outer lipid membrane. They
lose the crystal violet stain during decolorization and take up the counterstain, appearing pink or
red.
Example: Escherichia coli is a Gram-negative bacterium that appears as red or pink rods under
the microscope. It is commonly found in the human intestine and is a frequent cause of urinary
tract infections and gastroenteritis.
Clinical Importance
Understanding whether a bacterium is Gram-positive or Gram-negative helps in determining its
pathogenic mechanisms and the appropriate antibiotic treatment. Gram-positive bacteria are
typically sensitive to antibiotics like penicillin, which disrupt the peptidoglycan layer. In contrast,
Gram-negative bacteria are often resistant to many antibiotics due to their outer membrane,
which acts as a barrier to many substances including antibiotics.
The Gram stain is a crucial diagnostic tool in microbiology, aiding in the initial identification and
characterization of bacteria in clinical specimens. It guides the choice of further tests and the
initial treatment approach before specific bacterial identifications are made.
 Microflora- ‘normal’ bacteria we have present in a perfect balance- beneficial to us

 Normal skin flora includes: staphylococcus epidermidis( dry exposed skin)

 Moist areas typically have: S. aureus, corynebacteria & Candida spp.

 Fatty areas (or with limited oxygen) the hair follicles, sebaceous glands and sweat
support anaerobic organisms e.g., Propionibacterium acnes and other diphteroids

 Pityriasis versicolor, also known as tinea versicolor, is a common superficial fungal

infection of the skin. It is caused primarily by yeast from the
genus Malassezia, which is normally found on the skin’s surface
part of the flora but its overgrowth due to various environmental
factors leads to the disease
  Some bacteria work together while in some cases only 1 takes over the region:
Streptococcus - signature bacterium in the oral cavity.
 Bacteroidetes – signature bacterium in the gut.
 Either you have Prevotella or you have Lactobacillus in the vagina.
 Either you have Propionibacterium or you have Staphylococcus on the skin.
 Either Streptococcus or Tannerella in the mouth

Clostridioides difficile (C. difficile) is a bacterium that causes severe diarrhea and colitis, an
inflammation of the colon, most commonly affecting older adults in hospitals or in long-term care
facilities. The infections often occur after the use of antibiotic medications, which disrupt the
normal balance of bacteria in the gut. Dysbiosis refers to a disruption in the balance and
composition of the microbiota in the body, particularly in the gut. This imbalance can lead to a
range of health issues, including vulnerability to infections. Antibiotics, especially broad-
spectrum types, kill not only harmful bacteria but also beneficial bacteria that contribute to a
healthy gut microbiome. This loss of microbial diversity creates an environment where C. difficile
spores can thrive and proliferate. Result: Without the competition from other gut bacteria, C.
difficile spores germinate, producing toxins that cause symptoms ranging from mild diarrhea to
life-threatening inflammation of the colon.

Small Intestine Bacterial Overgrowth (SIBO)

Definition
Excessive bacterial growth in the small intestine, often involving non-native bacterial types.
Pathophysiology
Caused by motility issues, anatomical abnormalities, acid suppression, and immune
deficiencies.
Clinical Features
Symptoms: Bloating, abdominal pain, diarrhea, weight loss, nutrient deficiencies.
Malabsorption: Steatorrhea and vitamin deficiencies.
Diagnosis
Breath Tests: Measures hydrogen and methane production.
Small Intestinal Aspirate and Fluid Culture: Gold standard for detecting bacterial
overgrowth.
Additional Tests: Blood tests for anemia, imaging for structural issues.
Treatment
Antibiotics: Main treatment, typically rifaximin and metronidazole.
Dietary Changes: Low-FODMAP diet to reduce symptoms.
Prokinetics: Improve gut motility.
Correcting Underlying Causes: Surgical corrections, medication adjustments.
Prognosis and Management
High recurrence rate; may require repeated treatments.
Long-term management involves dietary adjustments and lifestyle changes.

Respiratory:

 Upper respiratory tract infections: Sinusitis, Nasopharyngitis, Otitis (media and externa),
Tonsillitis, Pharyngitis, Epiglottis, Laryngitis
 Lower respiratory tract infections: Tracheitis, Bronchiolitis, Bronchitis, Pulmonary
tuberculosis, Pneumonia, Empyema
 Invaders- are pathogens that can infect a 100% healthy body
 Opportunistic pathogens infect when your body’s immune system is not at its best they
are also known as secondary invaders

common opportunistic pathogens:

1. Candida spp.
2. Pseudomonas aeruginosa
3. Aspergillus spp.
4. Staphylococcus epidermidis
5. Cryptococcus neoformans
6. Cytomegalovirus (CMV)
7. Mycobacterium avium complex (MAC)

Common Cold

Main Agents: Rhinoviruses (50% of cases), coronaviruses, RSV, influenza virus, adenovirus.
Transmission
Methods: Direct contact with infected secretions, airborne droplets.

Process: Virus enters through mucous membranes, leads to inflammation and symptoms.
Symptoms: Runny nose, congestion, sneezing, sore throat, mild headache, cough; fever rare
in adults, more common in children.
Duration: Typically 7-10 days.

Diagnosis based On: Symptoms; lab tests usually unnecessary.

Treatment-Supportive care (rest, fluids, OTC pain relievers, decongestants).
Antibiotics: Not recommended (ineffective against viruses).

Epstein-Barr Virus (EBV) : A member of the Herpesviridae family; also known as human
herpesvirus 4 (HHV-4).
Primary Infection: Infectious mononucleosis (commonly known as mono or the "kissing
disease").
Transmission
Modes: Primarily through salivary contact; can also be transmitted via blood and genital
secretions.
Incubation Period: Typically 4 to 6 weeks.

Infectious Mononucleosis Symptoms: Fever, sore throat, swollen lymph nodes, fatigue,
splenomegaly.
Treatment : Mostly supportive (hydration, analgesics, rest).

Antivirals: Not routinely used; may be considered in severe or complicated cases.

Avoidance of Amoxicillin: Can cause rash in individuals with infectious mononucleosis.

 Group A streptococci/GAS/GABHS= Streptococcus pyogenes

Pneumonia: An inflammatory condition of the lung primarily affecting the alveoli, which fill with
fluid or pus.
Typical Bacterial Pathogens
1. Streptococcus pneumoniae: Most common cause of bacterial pneumonia.
2. Haemophilus influenzae: Second most common, especially in COPD patients and
children.
3. Staphylococcus aureus: Associated with post-viral infections and hospital-acquired
pneumonia.
4. Klebsiella pneumoniae: Common in alcoholics and immunocompromised individuals.
5. Pseudomonas aeruginosa: Typically seen in patients with chronic lung diseases and
cystic fibrosis.
Atypical Bacterial Pathogens
1. Mycoplasma pneumoniae: Causes "walking pneumonia," common in younger
populations.
2. Chlamydophila pneumoniae: Often mild symptoms; affects all ages.
3. Legionella spp.: Associated with contaminated water supplies (e.g., air conditioning
systems), causing Legionnaires' disease.
Viral Pathogens
1. Influenza virus: Seasonal outbreaks; can lead to secondary bacterial pneumonia.
2. Respiratory Syncytial Virus (RSV): Major cause in infants and elderly.
3. Adenoviruses: Typically cause mild symptoms but can be severe in
immunocompromised patients.
4. Coronaviruses (including SARS-CoV-2): Can range from mild respiratory symptoms to
severe pneumonia.
Fungal Pathogens
1. Pneumocystis jirovecii: Particularly affects individuals with HIV/AIDS.
2. Cryptococcus neoformans: Typically affects immunocompromised patients, such as
those with HIV/AIDS.
3. Histoplasma capsulatum: Acquired from environments contaminated with bat or bird
droppings.
Clinical Features
Symptoms: Cough, fever, chills, dyspnea, chest pain, and fatigue.
Signs: Crackles or wheezing on auscultation, tachypnea.
Diagnosis
Chest X-ray: Identification of infiltrates.

Microbiological Tests: Sputum culture, blood cultures, PCR, and antigen tests for specific
pathogens.
Serology: Useful for atypical pathogens like Mycoplasma and Legionella.
Treatment
Bacterial: Appropriate antibiotics (e.g., macrolides for atypical pathogens, beta-lactams for
typical pathogens).
Viral: Supportive care and antivirals where applicable.

Fungal: Specific antifungal treatment depending on the organism.

Complications
Pleural effusion, respiratory failure, and septicemia.

 Hospital-acquired pneumonia (HAP) occurs in patients who have been in a hospital for
48 hours or more, while community-acquired pneumonia (CAP) is developed outside of
hospital or healthcare environments.

Cardiology:

 Fever of unknown origin (FUO) refers to a prolonged fever of at least three weeks' duration, with
a temperature above 38.3°C (101°F) on several occasions, and a diagnosis that remains uncertain
after investigations for at least 3days

 FUO continuous- swings less than 1degree

 FUO Intermittent/recurrent swings by at least 1 degree
 Nosocomial FUO-hospital-acquired infection; patient w/o a fever upon admission gets a fever 72
hours after
 Neutropenic FUO- low neutrophil count
 HIV-associated FUO: lasting >4wks (as outpatient) or >3 days as hospital inpatient with HIV
 If NO FEVER or CELLULAR MATERIALS in laboratory samples – it is more likely a toxin-
based infection
 Infective endocarditis:

Inflammation of the inner lining of heart chambers and valves

Mainly affecting people with risk factors:
1. damaged or artificial heart valves
2. congenital/structural heart defects
3. implanted medical devices (heart or blood vessels)
4. IVD abusers
5. bloodstream infections

Increasing in number probably due to: 1. an ageing population with relatively greater prevalence of:
degenerative valve disease, prosthetic valves, intracardiac devices 2. more patients are receiving
hemodialysis 3. periodontal disease remains common.

Categories: Native valve, Prosthetic valve and IVD user

Symptoms : • commonly FUO (>38 °C) • chills/night sweats • heart murmurs • headaches • shortness of
breath • tiredness • muscle & joint pain • persistent cough • swelling of feet, legs or abdomen • paleness •
Splinter haemorrhages (vasculitis) • Osler’s nodes (vasculitis) • Janeway lesions( micro abscesses/septic
emboli) • Roth’s spots ( micro abscesses/septic emboli)

if untreated vegetations can lead to: o Rheumatic fever o Heart failure o Stroke

Key Pathogens:
Bacteria:
1. Streptococcus viridans: A group of oral streptococci, responsible for about 40-50% of cases;
commonly associated with subacute endocarditis.
2. Staphylococcus aureus: Causes acute endocarditis; most common cause in intravenous drug
users and one of the most virulent.
3. Enterococci (E. faecalis, E. faecium): Important in infections acquired in healthcare settings or
from gastrointestinal or genitourinary tracts.
4. Coagulase-negative staphylococci (e.g., S. epidermidis): Commonly associated with prosthetic
valve endocarditis.
5. HACEK group (Haemophilus spp., Aggregatibacter spp., Cardiobacterium hominis, Eikenella
corrodens, Kingella kingae): Typically cause endocarditis with a subacute course; rare but
important for their association with negative routine blood cultures.
Fungi:Candida spp. and Aspergillus spp.: More rare, associated with immunocompromised patients
and those with prosthetic valves.
To diagnose: blood culture, Echocardiography
Treatment- Long duration Antibiotics (gentamycin for gram -ve and vancomycin for gram +ve)