BACTE-LESSON-1 (1)

Microbiology

• study of living organisms of microscopic size

• Study of microbes

Pathogen

cause disease

• also known as infectious agents

• distinct shapes

• produce toxin

• spread easily

• live everywhere

Non pathogen

• do not cause disease

• harmless

• beneficial

• live on surface of the skim

Acellular microbes & Cellular microbes

two major categories of microbes

Acellular microbes

infectious particles

Cellular microbes

• microorganisms
  example: bacteria, protozoa and archaea

infectious disease

pathogen colonizes a person’s body
example: neisseria, mycobacteria, streptococcus

Microbial intoxications

pathogens produces toxins that once ingested by the host can cause intoxication

Toxin

poisonous molecules produced by microorganisms capable of causing harm to a host

Pathogenicity

ability of the organisms to cause disease to host or organisms

Virulence

refers to the degree of pathogenicity, a pathogen that can cause severe disease

Pathogenicity & Virulence

Under microbial intoxication

Communicable disease

• infectious disease

  • Capable of spreading from person to person

Pestilence

• earliest known infectious diseases

• occurred in egypt about 3180 BC

  • first recorded epidemic

Bubonic Plague (Yersinia)

• around 1900 BC

• near the end trojan war

• the greek army was decimated by an epidemic

Eber Papyrus

• epidemic fevers

• tomb in thebes, egypt

• around 1500 BC

Epidemics of plague

• rome in 790,710 and 640 BC

  • Greece around 430 BC

Christopher Columbus (1492-1892)

• Syphilis (1493)

• first appearance in europe in 1493

  • syphilis was carried to europe by native americans who were brought to portugal

Lucretius (98-55), Girolamo fracastorio (1478-1553)

suggested that disease were caused by “invisible creatures”

Girolamo Fracastoro (1478-1553)

• italian

• wrote poems about STDs (syphilis)

Antonie Van Leeuwenhoek (1632-1723)

• First True microbiologist

• first person to observe and accurately describe living microorganisms

• Father of bacteriology and protozoology

• animalcules or tiny living and moving cells seen under the microscopes

Abiogenesis

• life arise from non-living material if it contain “Pneuma” (vital heat)

• organisms do not descend from other organisms or from a parent

Francesco Redi

demonstrated that maggots were the offspring of flies, not products of spontaneous generation

John Needham

microbes arose spontaneously in broth from a “life force”

Lazzaro Spallanzani

Heated but sealed flasks remained clear, without any signs of spontaneous growth, unless the flasks were subsequently opened to the air.

Louis Pasteur

• Suggested that the exposure of a broth to air was not introducing a “LIFE FORCE” to the broth but rather airborne microorganisms.

• He boiled broth to sterilize it

• “Omne vivum ex vivo” (“Life only comes from life”)

Biogenesis (louis pasteur)

• Making new living things

• he demonstrated that microorganisms are present in the air

  • he proposed the use of heat in killing microorganisms (aseptic technique)

PASTEUR’S CONTRIBUTION TO MICROBIAL SCIENCE

 He disproved the theory of spontaneous generation

 He developed the vaccine against anthrax (1881) and rabies (1885)

 He improved the wine-making process (Fermentation and pasteurization)

Rudolf Virchow (1821-1902)

 Proposed the THEORY OF BIOGENESIS

 Which states that living cells can arise only from pre-existing living cells.

Theory of of antisepsis

• Ignaz Semmelweis (1816-1865)

• Joseph Lister (1827-1912)

• Robert Koch (1843-1910)

• Edward Jenner (1749-1823)

• Louis Pasteur (1882-1895)

• Piere Paul Emile Roux (1853-1933)

IGNAZ SEMMELWEIS (1816 - 1865)

 He demonstrated that routine handwashing can prevent the spread of diseases (PUERPERAL FEVER)

NOTE:

 How long should we Handwash?

 15 - 20 Seconds or 2 Happy birthday song

JOSEPH LISTER (1827 - 1912)

 He introduced the SYSTEM OF ANTISEPTIC SURGERY
 He pioneered promoting handwashing before and after operation

ROBERT KOCH (1843 - 1910)

• first to show irrefutable proof that bacteria cause diseases

 HE DISCOVERED:

 Bacillus anthracis (1876)

 Mycobacterium tuberculosis (1882)

• first to cultivate bacteria on boiled potatoes,

• He developed a culture media for observing bacterial growth isolated from the human body

Four (4) criteria to assess if a microorganism can cause a disease (Segre, 2013):

1. The microorganism must be found in diseased but not healthy individuals

2. The microorganism must be cultured from the diseased individual

3. Inoculation of a healthy individual with the cultured microorganism

4. The microorganism must be re-isolated from the inoculated, diseased individual and matched to the original microorganism

EDWARD JENNER (1749 - 1823)

introduced the smallpox vaccination through cowpox inoculation.

LOUIS PASTEUR (1882 - 1895)
PIERE PAUL EMILE ROUX (1853 - 1933)

Pasteur used the term “VACCINE” for an attenuated culture (REDUCED VIRULENCE) they made a series of experiments to produce an attenuated strains of bacteria

Modern Therapy: Magic Bullet

• Selman waksman (1888-1973)

• Alexander Fleming (1881-1955)

• Paul Ehrlich (1854- 1915)

• Ellie Metchnikoff (1845-1916)

• Ferdinand Cohn (1828-1898)

• Fanny Hesse (1850-1934)

• Julius Richard Petri (1852-1921)

• Robert Hooke, English-Man (1665)

SELMAN WAKSMAN (1888 - 1973)

He discovered STREPTOMYCIN and NEOMYCIN antibiotics

ALEXANDER FLEMING (1881 - 1955)

 He accidentally discovered the antibiotics PENICILLIN (Penicillium notatum) (fungi)
 He discovered LYSOZYME

PAUL EHRLICH (1854 - 1915)

He discovered SALVARSAN (Arsphenamine) for the treatment of syphilis.

ELLIE METCHNIKOFF (1845 - 1916)

First to describe the IMMUNE SYSTEM CELLS and the PROCESS OF PHAGOCYTOSIS.

JOHN TYNDALL (1820 - 1893)

Showed that dusts carry agents that could contaminate a sterile broth.

FERDINAND COHN (1828 - 1898)

He discovered that there are bacteria that could withstand a series of heating and boiling because of heat-resistant structures known as ENDOSPORES

FANNY HESSE (1850 - 1934)

She suggested the use of AGAR

 A solidifying agent, in the preparation of culture media

JULIUS RICHARD PETRI (1852 - 1921)

He developed the PETRI DISH

ROBERT HOOKE, ENGLISH-MAN (1665)

life’s smallest structural units were “LITTLE BOXES”, or “CELLS”, as he called them. Using his improved version of COMPOUND MICROSCOPE,
 Cell Theory
- The theory that all living things are composed of cells.

ANTON VAN LEEUWENHOEK, DUTCH MERCHANT AND AMATEUR SCIENTIST

• probably the first to actually observe live microorganisms

• He made his own microscope magnified to about 270 times and described the different shapes of bacteria.

• made detailed drawings of “ANIMALCULES” in rainwater, in his own feces, and in some material scraped from his teeth.

TAXONOMY

Provides a consistent means to classify, name, and identify organisms. This consistency allows biologist worldwide to use a common label for every organism studied within the multitude of biologic disciplines

three distinct but highly interrelated disciplines: (taxonomy)

 Classification

 Nomenclature (naming)

 Identification

CLASSIFICATION

It is the organization of microorganisms that have similar morphologic, physiologic, and genetic traits into specific groups or taxa.

EUKARYOTES

• “True nucleus” of eukaryotes (From Gr karyon “nucleus”) is only one of their distinguishing features

• The membrane-bound organelles, the microtubules, and the microfilaments form a complex intracellular structure

Flagella and Cilia

Complex multistranded structures

PROKARYOTES

 Organisms in which DNA is not physically separated from cytoplasm (no membrane bound organelles)

Distinguishing Characteristics:

 Relatively small in size (1um in diameter)

 Absence of nuclear membrane

 The DNA of almost all bacteria is a circle with a length of 1mm

 Most prokaryotes have only single chromosome

 The specialized region of cell containing DNA is termed as NUCLEOID and can be visualized by EM (Electron Microscope)  Have the capacity to exchange small packets of genetic information (This information is carried on PLASMIDS)

Peptidoglycan layer

or also called as the MUREIN or MUCOPEPTIDE LAYER

 NAG (N-acetylglucosamine)

 NAM (N-acetylmuramic acid)

MUSA

 Mycoplasma

 Urea plasma

 Spiro plasma

 Anaeroplasma

FAMILY

• contain multiple genera and consists of organisms with a common attribute

• The name of a family is formed by adding the suffix -aceae to the root name of one of the group’s genera, called the type genus

   Ex. Family - Streptococcaceae o Genus - Streptococcus

GENUS

Contains different species that have several important features in common.

 GENUS

o Streptococcus

o Escherichia

SPECIES

 (Abbreviated as SP., singular, or SPP., plural)

 Is the most basic of the taxonomic groups and can be defined as a collection of bacterial strains

SUB-SPECIES

• Serotype

• Biotype

SEROTYPE

Based on serologic differences.

o Ex. E. coli (>700 serotypes

BIOTYPE

Based on biochemical differences

o Ex. Treponema pallidum subsp. Pallidum

NOMENCLATURE

Naming of microorganisms according to established rules and guidelines set forth in the International Code of Nomenclature of Bacteria (ICNB) or the Bacteriological Code (BC)

(ICNB)

international Code of Nomenclature of Bacteria

(BC)

Bacteriological Code

Genus and Species Rules:

1. Genus designation

a. First letter is always capitalized

2. Species Designation

a. First letter is always lower case

3. Printed in italics

a. Ex. Staphylococcus aureus

b. Staphylococcus agalactiae

4. Underlined in script

a. Ex. Staphylococcus aureus

b. Staphylococcus agalactiae

5. The name may be abbreviated by using the upper case

form of the first letter of the genus designation followed by

a period (.) and the full species name, which is never abbreviated

a. Ex. S. aureus, S. agalactiae

6. Informal designation

a. May be used to label particular group of organisms. These designations are not capitalized or italicized.

i. Ex. Staphylococci, enterococci, streptococci

IDENTIFICATION

Process by which a microorganism’s key features are delineated.

GENOTYPIC CHARACTERISTICS

a. Organism’s genetic make-up, including the nature organisms gene and constituent nucleid acids.

i. DNA base composition ratio

PHENOTYPIC CHARACTERISTICS

Based on features beyond the genetic level and include both readily observable characteristics and those that may require extensive analytic procedures to be detected.

i. Macroscopic and microscopic morphology (inoculated)

ii. Staining characteristics (gram staining, any staining technique)

iii. Nutritional requirements (an/aerobic organism, needs oxygen, doesn’t need additional vitamins, etc. for growth)

iv. Biochemical testing (results)

Macroscopic morphology

the microbial growth patterns on artificial media as observed when inspected with the unaided eye.Micro

Microscopic Morphology

The size, shape, intracellular inclusions, cellular appendages, and arrangement of cells when observed with the aid of microscopic magnfication

Staining characteristics

The ability of an organisms to reproducibly stain a particular color with the application of specific dyes and reagents. Staining is used in conjunctions with microscopic morphology for bacterial identification

Environmental requirements

the ability of an organism to grow at various temperatures, in the presence of oxygen and other gases, at various pH levels, or in the presence of other ions and salts, such as NaCl

Nutritional requirements

the ability of an organisms to use various carbon and nitrogen sources as nutritional substrates when grown under specific environmental conditions

Resistance profiles

the exhibition of a characteristic inherent resistance to specific antibiotics, heavy metals or toxins

Antigenic Properties

the profiles of microorganisms established by various serologic and immunologic methods to determine relatedness among various microbial groups

Subcellular properties

molecular constituents of the cell that are typical of particular taxon, or organism group, as established by various analytic methods

Chemotaxonomic properties

the chemical constituents of the cells, such as the structure of teichoic acids, fatty acid analysis and protein profiles, as determined by analytical methods

DNA base composition ratio

DNA comprises four bases (guanine, cytosine and thymine) The extent to which the DNA from two organisms is made up of cytosine and guanine

Nucleic acid ( DNA and RNA) base sequence characteristics, including those determined by hybridization assays

The order of bases along a strand of DNA or RNA is known as the base sequence. The extent to which sequences are homoglous (similar) between two microorganisms can be determined directly or indirectly by various molecular methos

Average nucleotide Identity (ANI)

this methods analyses multiple coding sequences in a microorganisms genome to determine the average nucleotide identity using genome sequencing and computer algorithms. The relatedness of microorganisms is accurate at 95%-96% threshold for organisms identification

Genome to Genome Distance

this is a computerized calculation that uses inference by in-silico genome comparison eliminating the liitation and errors associated with wet-lab techniques. Organisms are related with a GGD threshold score of 70% or greater

LABORATORY BIOSAFETY

Containment principles, technologies, and practices implemented to prevent unintentional exposure to pathogens and toxins, or their unintentional release.

“PROTECTING PEOPLE FROM DANGEROUS PATHOGENS”

Protect the users

 Protect those outside the labs

 Protect the environment

LABORATORY BIOSECURITY

Instituitional and personal security measures designed to prevent the loss, theft, misuse, diversion, or intentional release of pathogens and toxins.

“PROTECTING PATHOGENS FROM DANGEROUS PEOPLE”

PRACTICE AND PROCEDURES

a. Standard Practices

 Most important concept/strict adherence

 Aware of potential hazard

 Trained and proficient in techniques

 Supervisors responsible for: appropriate laboratory facilities, personnel and training

b. Special practices and considerations

SAFETY EQUIPMENT

 Primary containment barrier

 Minimize exposure to hazard

o Prevent contact/contain aerosols

 Engineering controls/equipment

 PPE

o Gown

o Gloves

o Respirator

o Face shield booties

 BSC

 Covered or ventilated animal cage systems

FACILITY DESIGN AND CONSTRUCTION

 Secondary barrier/engineering controls

 Contributes to worker protection

 Protects outside the laboratory

o Ex. Building and lab design, ventilation,

autoclaves, cage, wash facilities 4

INCREASING LEVELS OF PROTECTION

BIOSAFETY LEVELS 1-4.

Principle of Biosafety

1. Standard Practices

2. Safety Equipment

3. Facility design and construction

4. increasing levels of protection

Biohazard Symbol

1. Agent

2. Host

3. Source

4. Transmission

Agent

The type of microorganism, that causes infection or hazardous condition

Host

The organisms in which the microorganisms infect. The new host must be susceptible

Source

The host from which the microorganism originates. The carrier host might not show symptoms

TRANMISSION

The means of transmission, mostly direct or indirect. Some routes of transmission includes air, insect, direct contact and contaminated surfaces

Germs (agent)

• bacteria

• viruses

• parasites

Where germs live (reservoir)

• people

• animals/pets (dogs, cats, reptiles)

• wild animals

• food

• soil

• water

How Germs get out (portal of exit)

• mouth (vomit, saliva)

• cuts in the skin (blood)

• during diapering and toileting stool)

Germs get around (mode of transmission)

• contacts (hands, toys, sand)

• droplets (when you speak sneeze or cough)

How germs get in (portal of entry)

• mouth

• cuts in the skin

• eyes

Next sick person (susceptible host)

• babies

• chicken

• elderly

• people with a weakened immune system

• Unimmunized people

• anyone