CHARACTERISTICS

OF BACTERIA
FST 221
MICROBIOLOGY
D A N I E L L E T H E O L . M A RT I N E Z
INTRODUCTION
What is Bacteria?
Bacteria are small single-celled organisms. Their control center, containing the genetic
information, is contained in a single loop of DNA. Some bacteria are harmful (pathogenic), but
most serve a useful purpose. They support many forms of life, both plant and animal, and they
are used in industrial and medicinal processes. Bacteria are thought to have been the first
organisms to appear on earth, about 4 billion years ago. The oldest known fossils are of bacteria-
like organisms.
TYPES OF BACTERIA
There are many different types of bacteria. One way of classifying them is by shape.
Spherical: Bacteria shaped like a ball are called cocci, and a single bacterium is a coccus. Example
is Staphylococcus aureus
TYPES OF BACTERIA
Rod-shaped: These are known as bacilli (singular bacillus). Some rod-shaped bacteria are curved.
These are known as vibrio. Example is Lactobacillus Casei.
TYPES OF BACTERIA
Spiral: These are known as spirilla (singular spirillus). If their coil is very tight they are known as
spirochetes. Example is Campylobacter.
TYPES OF BACTERIA
Filamentous: the Filamentous bacteria are long thin filament-like bacteria that can branch and
form networks called mycelium. An example is Candidatus savagella
GROWTH OF BACTERIAL
POPULATIONS
Bacterial colonies progress through four phases of growth: the lag phase, the log phase, the
stationary phase, and the death phase.

The lag phase is an adaptation period, where the bacteria are adjusting to their new conditions.
Once cells have accumulated all that they need for growth, they proceed into cell division. The
exponential or log phase of growth is marked by predictable doublings of the population, where
1 cell become 2 cells, becomes 4, becomes 8 etc. Conditions that are optimal for the cells will
result in very rapid growth (and a steeper slope on the growth curve), while less than ideal
conditions will result in slower growth. Cells in the exponential phase of growth are the
healthiest and most uniform, which explains why most experiments utilize cells from this phase.
GROWTH OF BACTERIAL
POPULATIONS
At some point the bacterial population runs out of an essential nutrient/chemical or its growth is
inhibited by its own waste products or lack of physical space, causing the cells to enter into the
stationary phase. At this point the number of new cells being produced is equal to the number of
cells dying off or growth has entirely ceased, resulting in a flattening out of growth on the growth
curve.

In the last phase of the growth curve, the death or decline phase, the number of viable cells
decreases in a predictable (or exponential) fashion. The steepness of the slope corresponds to
how fast cells are losing viability. It is thought that the culture conditions have deteriorated to a
point where the cells are irreparably harmed, since cells collected from this phase fail to show
growth when transferred to fresh medium.
CHARACTERISTICS OF THE
SPORES
Bacterial species have different coping mechanisms with selective harsh environmental
conditions. One of the most common coping mechanisms for bacteria is forming spores to
protect themselves against ecological degrading agents. Bacterial spores are the most dormant
form of bacteria since they exhibit minimal metabolism and respiration, as well as reduced
enzyme production.
OBJECTIVE
To familiarize the techniques used in the examination of the cultural, morphological,
physiological and biochemical tests for the characterization of bacteria.
MATERIALS
MATERIALS:
Bacteria Cultures
Petri Plates
Pipettes
Glass Slides and covers
Microscope
Culture Media
Test Tubes
CULTURAL CHARACTERISTICS
Prepare Nutrient Broth (NB) and Nutrient Agar (NA)
Mix 18g of agar for your slant and 8g for your stab, 3g of beef extract, 5g of peptone in 1L of
distilled water. Heat to dissolve, then transfer into test tubes and sterilize for 15 mins in an
autoclave at 121­°C. For Nutrient Broth medium, don’t add the agar to the mixture.
LIQUID MEDIUM GROWTH
Inoculate a bacteria culture onto the Nutrient Broth. Incubate for 24 hrs at 35­°C. After
incubation, observe the culture for its cultural characteristics; Surface Growth, Clouding, Odor
and Sediments.

Inoculating Liquid Bacterial Culture – YouTube 3:51 mins

SOLID MEDIUM GROWTH
Aerobic organisms should be observed on the agar slant. Inoculate the agar slant with the
bacteria culture using an inoculating needle and streak the bottom of the slant to the top.
Incubate for 24 hrs at 35°C. After incubation, observe the cultural characteristics of the growth.
Characteristics will differ based on the medium used.
Anaerobic organisms should be observed on the agar stab. Inoculate the medium with an
inoculating needle. Stab the center of the medium. Incubate for 24 hrs at 35°C. After incubation,
observe the cultural characteristics of the growth. Characteristics will differ based on the medium
used.
MORPHOLOGICAL
CHARACTERISTICS
Morphological Characteristics of the culture are observed by size and shape of the cell and
spore, stain reactions, spore formation, presence of capsules.
MORPHOLOGICAL
CHARACTERISTICS
Preparation of smears
Preparation of a bacterial smear for staining – YouTube 0:26 mins

Staining
Gram staining is a common technique used to differentiate two large groups of bacteria
based on their different cell wall constituents. The Gram stain procedure distinguishes between
Gram positive and Gram negative groups by coloring these cells red or violet. Gram positive
bacteria stain violet due to the presence of a thick layer of peptidoglycan in their cell walls, which
retains the crystal violet these cells are stained with. Alternatively, Gram negative bacteria stain
red, which is attributed to a thinner peptidoglycan wall, which does not retain the crystal violet
during the decoloring process.
STAINING
Methylene Blue Staining
1. Use the smear prepared in the previous procedure. Staining is done at the sink.
2. Add several drops of Methylene blue, enough to cover the smear, and wait 1 min.
STAINING
3. Rinse the slide with water from the squirt bottle
STAINING
4. Blot the slide with bibulous paper. Redraw the focus line on the top of the slide if necessary.

5. Observe.
STAINING
Gram Staining:
Gram Staining – YouTube 0:00 mins

Spore Staining
Spore Stain Procedure (Schaeffer-Fulton Method) – YouTube 0:00 mins

Capsule Staining
The capsule stain – YouTube 0:23 mins
STAINING
Flagella Staining
Flagella Stain – YouTube 0:00 mins
Physiological and Biochemical Test
Lithmus Milk Test
Litmus Milk Test - Amrita University – YouTube 0:56 mins

Hydrogen Sulfide Test

Hydrogen sulfide test (Biochemical assay) – YouTube 0:00 mins

Production of Ammonia
How to test ammonia production by a bacteria – YouTube 0:16 mins
Physiological and Biochemical Test
Production of Indole
Indole test - Microbiology (Microbial Biochemical test) – YouTube 0:42

Starch Hydrolysis
Starch Hydrolysis – YouTube 0:00

Gelatine Liquefication
How to perform Gelatinase (Gelatin Hydrolysis)Test – YouTube 0:31 mins
 Physiological and Biochemical Test
Catalase Test
Microbiology: the Catalase test – YouTube 0:00 mins

Nitrate Reduction Test

Microbiology: Anaerobic Nitrate Reduction Test – YouTube 3:04 mins in

Motility Test
Hanging Drop Method – YouTube 0:00 mins

Fermentation of Sugars
Sugar Fermentation by Bacteria - Microbiology – YouTube 0:00 mins
LACTIC ACID BACTERIA
Lactic acid bacteria (LAB) are heterogenous group of bacteria which plays a significant role in a
variety of fermentation processes. They ferment food carbohydrates and produce lactic acid as
the main product of fermentation. In addition, degradation of proteins and lipids and production
of various alcohols, aldehydes, acids, esters and sulphur compounds contribute to the specific
flavour development in different fermented food products.
LACTIC ACID BACTERIA
Are an order of gram-positive, low-GC, acid-tolerant, generally nonsporulating, non respiring,
either rod-shaped (bacilli) or spherical (cocci) bacteria that share common metabolic and
physiological characteristics. These bacteria, usually found in decomposing plants and milk
products, produce lactic acid as the major metabolic end product of carbohydrate fermentation,
giving them the common name lactic acid bacteria (LAB).
LACTIC ACID BACTERIA
During fermentation process Lactic Acid Bacteria (LAB) produces anti-microbial metabolites,
include organic acids like propionic, acetic acid and lactic acids as final products. They create
unfavorable environment to spoilage and pathogenic microorganism’s growth.

de Man, Rogosa and Sharpe (MRS) agar is developed primarily for the cultivation of lactobacilli
from various sources with the intention of producing a defined medium as a substitute for
tomato juice agar.
LACTIC ACID BACTERIA
Although many genera of bacteria produce lactic acid as primary or secondary fermentation
products, typical lactic acid bacteria are those of the Lactobacillales order, including the following
genera: Lactobacillus, Carnobacterium, Lactococcus, Streptococcus, Enterococcus, Vagococcus,
Leuconostoc, Oenococcus, Pediococcus, Tetragonococcus, Aerococcus and Weissella
LACTIC ACID BACTERIA AS
SOURCE OF PROBIOTICS
Microorganisms considered as commercial probiotics are mainly of the Lactobacillus genus with
over one hundred species recognized, for example: L. acidophilus, L. rhamnosus, L. reuteri, L.
casei, L. plantarum, L. bulgaricus, L. delbrueckii, L. helveticus. Lactobacilli are Generally
Recognized As Safe (GRAS) organisms.
LAB ARE USED AS STARTER CULTURES
EITHER IN DAIRY OR NON-DAIRY PRODUCTS
LACTIC ACID BACTERIA IN WINE
MAKING
The density of lactic acid bacteria in the initial phases of winemaking (the must phase and onset
of alcoholic fermentation) ranges from approximately 10^3 to 10^4 CFU/mL. The bacteria are
from a variety of mostly homofermentative species.
The most abundant are L. plantarum, Lactobacillus casei, Lactobacillus hilgardii, Leuconostoc
mesenteroides, and Pediococcus damnosus.
Less common species include O. oeni and Lactobacillus brevis.
LACTIC ACID BACTERIA IN WINE
MAKING
The main role of Lactic Acid Bacteria (LAB) in wine is to conduct the malolactic fermentation
(MLF). This process can increase wine aroma and mouthfeel, improve microbial stability and
reduce the acidity of wine. The main role of lactic acid bacteria (LAB) in wine has traditionally
been to perform the conversion of malic acid to lactic acid.
Saccharomyces cerevisiae yeast and Oenococcus oeni LAB are the most commonly used option.
WHERE IS THE LACTIC ACID
BACTERIA USED IN WINE
MAKING?
WHERE IS THE LACTIC ACID
BACTERIA USED IN WINE
MAKING?
WHERE IS THE LACTIC ACID
BACTERIA USED IN WINE
MAKING?
WHERE IS THE LACTIC ACID
BACTERIA USED IN WINE
MAKING?
THANK YOU
REFERENCES
Bacteria | What is microbiology? | Microbiology Society
Bacteria (genome.gov)
Jill Seladi-Schulman, Ph.D. — Written by Yvette Brazier on February 12, 2019
BioExplorer.net. (2022, May 29). Explore 13 Different Shapes of Bacteria. Bio Explorer. https://www.bioexplorer.net/bacteria-shapes.html/.
Gram Staining (carleton.edu)
Experiment 2A Direct Stains | Lab02 | Virtual Edge| General Microbiology/Molecular Biology | Molb
2021 | College of Agriculture and Natural Sciences (uwyo.edu)
Marina Basta; Pavan Annamaraju, Bacterial Spores, Bacterial Spores - StatPearls - NCBI Bookshelf (nih.gov)
Linda Bruslind, Microbial Growth, January 4, 2021, 9: Microbial Growth - Biology LibreTexts
bacteria - Growth of bacterial populations | Britannica
Saez-Lara MJ, Gomez-Llorente C, Gil A (2015). "The role of probiotic lactic acid bacteria and bifidobacteria
in the prevention and treatment of inflammatory bowel disease and other related diseases: a systematic review of randomized human clinical tr
ials“
Rosario Muñoz, ... Blanca de las Rivas, in Molecular Wine Microbiology, 2011
A mini review fermentation and preservation: role of Lactic Acid Bacteria, Aizhan Rakhmanova, Zaid Ashiq Khan, Kamran Shah