Jib 221 assignment 2

DEBBRA MARCEL JP/8544/13 ASSIGNMENT 2

JIB 221 MICROBIOLOGY

1. The human gastrointestinal tract (GIT) contains a bacterial ecosystem that is

fundamental to our health.

a. Explain the environmental factors (temperature, oxygen, pH, pressure)

affecting this bacterial ecosystem. (Section 7.3)

ANSWER:

i. Temperature: There are three cardinal temperatures that affect survival

of the microorganisms, including the bacteria in human GIT. Minimum

temperature is the lowest temperature for the bacterial growth and

metabolism, while maximum temperature is the highest limit of

temperature for the bacterial growth and metabolism. However, the

optimum temperature is the best temperature for the bacteria, as it

promotes and allows fastest rate of growth and metabolism of the

bacteria. In human GIT, mesophiles bacteria commonly found as their

optimum temperature is 20 - 40 , as human body temperature is fall

within this range (+37 .

ii. Oxygen: The requirement for oxygen is depends on the ecological

needs of human GIT bacteria for their metabolism. Metabolized oxygen

will be transformed into several toxic products (chemicals), commonly

neutralized by most bacteria enzymes (example: catalese). Therefore,

only bacteria that can detoxify the by-product of oxygen such as

hydrogen peroxide can utilize the oxygen, and this type of bacteria is

called aerobic bacteria. Obligate (strict) aerobes are the bacteria that

cannot survive without oxygen, while microaerophilic bacteria can survive

in minimum amount of oxygen. Meanwhile, the bacteria that cannot deal

with toxic oxygen is forced to live in oxygen free habitat, and this type of

bacteria is called anaerobic bacteria. Obligate anaerobes can be

damaged or killed by oxygen as they lack of enzyme to detoxify oxygen,

therefore cannot survive with the presence of oxygen. However,

aerotolerant anaerobes do not use oxygen but still can survive with the

presence of oxygen. As it is very limited oxygen in human GIT, the

anaerobic bacteria predominate the populations.



iii. pH: The acidity and alkalinity of human GIT affect the activity and

integrity of enzymes and the structures of the bacteria cell that presence

in it. According to Evans et al. (1988), the pH for normal human GIT vary

from gastric (highly acidic; ranged pH 1.0-2.5), small intestine (slightly

acidic; pH + 6.6) and terminal ileum (alkaline; pH +7.5). The optimum pH

for most microbes ranged between pH 6 and pH 8 (neutrophiles), as

found in ileum. While in gastric (stomach) area, most bacteria are

acidophiles as they grow in acidic environment. Meanwhile,

alkalinophiles bacteria is very rare to be found as they only grow in

extreme alkaline environment.

iv. Pressure: As most microbes in human GIT exist under hypotonic and

isotonic conditions, some are also need to be osmotolerant due to the

constant changes of human GIT chemical composition caused by

metabolic process, dehydration and consumption. Recent studies also

found that halophiles (require high concentration of salt) archaea

presence in the colon of human who suffer inflammatory bowel disease,

even though this organ not considered as salty environment. The

survival of these halophilic microbes may be associated to the mucosal

microbiota (Oxley et. al, 2010)

b. List 6 [SIX] species of bacteria that can be found in the GIT.

ANSWER:

i. Bacteroides sp.

ii. Lactobacillus sp.

iii. Escherichia coli (E. coli)

iv. Salmonella sp.

v. Helicobacter pylori

vi. Campylobacter jejuni



c. What nutritional types are the microbes most likely found in the large

intestine? Briefly, explain your answer. (Section 7.1)

ANSWER:

Bacteria are the most dominant microbes found in large intestine. The

nutritional types are mainly parasitic. This is because they live within host

(human) to survive (grow and reproduce). These microbes are

endoparasites (parasite of internal organ) and utilize the tissue and fluids of

the large intestine. There are several degrees of microbes parasitism in

human GIT, including the large intestine. Pathogens are the microbes that

damage the host cells/tissues of host, which are also known as harmful

microbes as they can cause disease, inflammation and the worse, death.

However, most microbes in large intestine are not fatal but also have

symbiotic relationship with the environment in the large intestine.

d. What ecological associations amongst the microbes can be found in the

GIT? (Section 7.4)

ANSWER:

According to the book entitled “Microbial Ecological Theory” (Oglive and

Hirsch, 2012), humans perform a wide range of relationship with resident

and transiently colonising microbes, from symbiotic, antagonistic to

mutualistic. The densest populations of microbes are found within the

human GIT, as this site produce rich nutrients for the habitat of bacteria,

fungi and few protozoa. The microbiomes association shows the key

example of host-microbe interactions in GIT. The microbial associations

within GIT are divided into symbiotic and non-symbiotic. Symbiotic

associations are the close nutritional relationships that happen between the

microbes within the human GIT. There are three types of symbiotic

relationship occur between microbes and human GIT namely mutualism,

commensalism and parasitism. Mutualism relationship shows the obligatory

dependant which both members gain the benefit, for example like

Lactobacillus sp. and Bacteroides sp. that assist in enzymatic production

and aid digestion as well as maintain pH in GIT. Commensalism



relationship shows that the bacteria get benefits but the GIT not harmed, just

like billions of bacteria found live on the waste in large intestine and co-

evolved with the host. When body of host change (get sick or antibody level

decreased), the commensal microbes can evolve and convert into parasite

in body, including GIT (example: E. coli). Meanwhile, non-symbiotic

association also presence in human GIT, which refers to free living microbes

that live within GIT but not require relationship with host for survival. This

association can be further divided into synergism and antagonism.

Synergism is an interrelationship between two or more microbes in human

GIT or the microbes’ relationship with the GIT itself, and this relationship

shows both side cooperate and share nutrients. Whereas, antagonism is a

type of competitive relationship, which occurs when the microbes inhibit and

destroy the others.

e. How does the microbial community communicate with each other in the

GIT? Why is this important? (Section 7.4)

ANSWER:

The microbial biofilms are among the most common and prolific associations

of microbes in human GIT. It is a result of the symbiotic and cooperative

relationship of microorganisms in the GIT microbiota. The biofilms may be

formed by a single type of microbes, but it is said that most biofilms are

observed as polymicrobial (association of more than one type of microbes)

which shows the participation of true bacteria, fungi and many other types of

microorganisms. The development and behavior of the microbial biofilms in

GIT can be explained by quorum sensing, a series of process which involve

formation, induction and expression. The process is commonly divided into

5 stages. Stage 1 shows the free-swimming of microbes settle on GIT

surface and remain there through weak, reversible adhesion via Van der

Waals forces. Stage 2 shows a sticky matrix synthesized by the microbes’

cells to hold them tightly to the substrate. Stage 3 shows the cells of

microbes release inducer (causing effect) molecules when biofilm reach

certain density (quorum, a minimum of group members) to coordinate

response. Stage 4 shows an occurrence of genetic induction when the cell



enlarged, where the inducer molecules stimulates specific genes on their

chromosomes to begin expression and synthesize a protein product such as

enzyme. Finally in stage 5, the unison form of enzymes secreted by the

cells performed when enzymatic reaction occurs. The formation of

microbiofilm in GIT allows the diverse microbes to react as a unit, affecting

the host either positively (promote health) or negatively (cause disease).

The study of microbiofilms in human GIT is important because it will lead to

a greater understanding of their involvement in infection and their

contributions to treatment such as disinfectant and drug. Each microbe that

involved in the formation of the biofilms plays their specific role as they rich

in different microhabitats and metabolic niches. For example, the mucosal

biofilms in human colon has been the interest of recent study, especially the

bacteria that cause inflammatory bowel disease because bacteria growing in

biofilm are resistant to antibiotic and hard to be eradicated (Macfarlane and

Dillon, 2007). Therefore, by understanding the characters and effects of the

biofilms especially in human GIT, we will find a better way to improve the

quality of human health.

f. In order to thrive in the human stomach, list 4 [FOUR] characteristics that a

microbe should have? Briefly explain your answer.

ANSWER:

i. Anaerobic: As human stomach has very low oxygen concentration,

the microbe must be able to perform anaerobic respiration and

fermentation to increase their survival.

ii. Acidophiles: Can survive and grow in harsh acidic environment of

human stomach that contains gastric juice (hydrochloric acid).

iii. Chemoautotroph: Can utilize the chemical substances in the human

stomach for the microbe nutritional sources.

iv. Can form microbial biofilm: The microbes that live within the stomach

must be able to form biofilm to promote their sustainability and

adhesion on the stomach surface.

v. Mesophilic: Must have the optimum temperature for their metabolism

and growth that ranged within human body temperature (+ ).



2. Like all other organisms, microbes perform a multitude of biochemical reactions

to consume, utilize and store energy.

a. Give 2 [TWO] examples of catabolic and 2 [TWO] examples of anabolic

reactions that can be performed by these microbes. (Section 8.1)

ANSWER:

Catabolic reactions:

i. Respiration: Most heterotrophic microbes such as E. coli bacteria

perform anaerobic respiration as below and release high energy:

NO3- + NADH + H+ NO2

- + H2O + NAD+

ii. Fermentation: The fermentative microbes in ruminant stomach (rumen

part) work together in anaerobic manner by producing variety of

enzymes by fermentation to hydrolyze the complex chain of cellulose

into digestible glucose and release small amount of energy.

Anabolic reactions:

i. Photosynthesis: The photosynthetic microbes such as Euglena

(protozoa) make use of light as radiant energy source to oxidize water

releasing oxygen as by-product and make use of carbon dioxide to

synthesize carbohydrate as their own food:

6CO2 + 6H2O C6H12O6 + 6O2

ii. Chemosynthesis: Chemoautotrophic bacteria utilize the inorganic

substances and carbon dioxide to produce methane gas and water:

4H2 + CO2 CH4 + 2H2O



b. Enzymes are proteins that catalyze metabolic reactions by lowering the

activation energy.

List 6 [SIX] classes of enzymes based on the types of reactions they catalyze

and briefly state the reactions they perform.

ANSWER:

i. Oxidoreductases: Transfer electrons form one substrate to another. While

dehydrogenases transfer a hydrogen from one compound to another:

Example: Lactate dehydrogenase catalyzes pyruvic into lactic acid.

ii. Transferases: Transfer functional groups from one substrate to another.

Example: DNA polymerase synthesize a strand of DNA using the

complementary strand as a model, with DNA nucleotides as a substrate.

iii. Hydrolases: Performing cleave bonds on molecules with the addition of

water. Example: Lactase breaks down lactose into glucose and galactose.

iv. Lyases: Add groups to or remove groups from double-bonded substrates.

Example: Isocitrate lyase (adolase group) that involved in the glycoxylate

cycle, where it converts isocitrate to succinate.

v. Isomerases: Change substrate into its isomeric form. An isomer is a

compound that has the same molecular formula as another compound but

has different arrangement of atoms. Example: Glucose-6-phosphate

isomerase that catalyzes the conversion of glucose-6-phosphate in to

fructose-6-phosphate during the second step of glycolysis.

vi. Ligases: Catalyze the formation of bonds with the input of ATP and the

water removal. Example: DNA ligase catalyzes the ligation between breaks

in DNA by forming a phosphodiester bond.



c. Constitutive enzymes are always present and produced in equal amounts at

equal rates regardless of the amount of substrate. Whereas regulated enzymes

are only produced in response to cell requirements. What are the enzymes

involved in the Krebs cycle and how are they regulated by the microbial cell so

that there is no wastage through overproduction of metabolic energy? (Sections

8.1 and 8.3)

ANSWER:



Constitutive enzyme: Enzyme that involved in utilizing a chemical components

such as glucose which is very important in metabolism.

Regulated enzyme: Enzyme that induced (turned on) or repressed (turned off)

by degrees of genes that expressed (stimulated by substrate).

Regulation of 8 enzymes involved in Krebs cycle are as follow:

Enzyme name Substrate Reaction occurs by enzyme

Citrate synthase

Acetyl CoA + oxaloacetate

Aldol condensation happen during combination of 2C acetyl CoA + oxaloacetate + H2O forming 6C citrate and release CoA to allow it join with another group

Aconitase Citrate Dehydration of citrate forming isocitrate, a preparation for decarboxylation and redox in the next step.

Isocitrate dehydrogenase

Isocitrate Isocitrate is converted to 5C α-ketoglutarate + CO2 and generates NADH (equivalent of 2.5 ATP)

α-ketoglutarate dehydrogenase

α-ketoglutarate Oxidation of α-ketoglutarate and NADH+ with high energy compound 4C succinyl CoA regenerated

Succinyl coA synthase

Succinyl-CoA Succinyl-CoA converted to succinate + CoA + GTP (energy that captured in ATP).

Succinate dehydrogenase

Succinate Oxidative cause succinate to lose 2H+ and 2 e-, forming fumarate and generating FADH2 from FAD

Fumarase Fumarate Hydration cause fumarate to form malate

Malate dehydrogenase

Malate

Oxidation convert malate to final product called oxaloacetate, with addition of NADH. This final step of Krebs cycle may repeat when oxaloacetate react with acetyl CoA.

This cycle occur in the cytoplasm of prokaryotic microorganism and mitochondria

matrix of eukaryotic microorganism, and the process is essential for generating

small organic molecules that microbes require for synthesis.

http://en.wikipedia.org/wiki/Aldol_condensation

http://en.wikipedia.org/wiki/Acetyl_CoA

http://en.wikipedia.org/wiki/Oxaloacetic_acid

http://en.wikipedia.org/wiki/Citric_acid

http://en.wikipedia.org/wiki/Citrate

http://en.wikipedia.org/wiki/Dehydration_reaction

http://en.wikipedia.org/wiki/Isocitric_acid

http://en.wikipedia.org/wiki/Alpha-Ketoglutaric_acid

http://en.wikipedia.org/wiki/Oxidation

http://en.wikipedia.org/wiki/Aldol_condensation



d. Describe the substrate used, metabolic reactions that occur, and products that

are formed by the fermentation of yeast in the making of bread? (Section 8.3)

ANSWER:

i. Substrates – 2 pyruvate (pyruvic acid) molecules that produced from the

glycolysis process of glucose.

ii. Metabolic reactions :

Phase 1 : Without the presence of oxygen, anaerobic fermentation force the

2 molecules of pyruvate to be decarboxylased into 2 molecules of

acetyldehyde catalyzed by the enzyme called pyruvate decarboxylase, and

released 2 molecules of carbon dioxide

Phase 2: The 2 molecules of acetyldehyde then reduced into 2 molecules of

ethanol catalyzed by the enzyme called alcohol dehydrogenase, with the

oxidation of 2 molecules of NADH (from glycolysis) into 2 molecules of NAD+.

This reaction is reversible.



iii. Products – 2 molecules of ethanol (ethyl alcohol) and 2 molecules of carbon

dioxide (bubbles that causing the bread to rise up).



e. Photosynthesis is said to be the reverse of glycolysis. But this is not exactly true,

what are some of the major differences between the two metabolic reactions?

(Section 8.5)

ANSWER:

PROCESS

CHARACTERISTICS

Photosynthesis Glycolysis

Function

Capture radiant energy in

the form of light and store it

into sugar (glucose)

Release energy that

was stored as sugar

(glucose)

Location

Only takes place at

2 sites:

i. Light reaction within

the thylakoid of

chloroplast

ii. Calvin cycle within the

stroma of chloroplast

Takes place in the

cytoplasm of a cell

(a part of cellular

respiration)

Reactants Water and carbon dioxide Oxygen and glucose

Type of metabolism Anabolism Catabolism

Main products Oxygen and glucose

Water and carbon

dioxide. Pyruvic acid

(pyruvate) is the final

product before entering

next step of cellular

respiration.

Basic equation 6CO2 + 6H2O + energy

(light) → C6H12O6 + 6O2

C6H12O6 + 6O2 → 6CO2

+ 6H2O + energy

Type of organism

Photosynthetic organism

such as algae and

cyanobacteria

Both photosynthetic and

non-photosynthetic

organism



REFERENCES:

1) D. F., Evans, G., Pye, R., Bramley, A. G., Clark, T. J., Dyson and J. D.,

Hardcastle. (1988). Measurement of gastrointestinal pH profiles in normal

ambulant human subjects. Gut. 1988 August; 29(8): 1035–1041.

2) Oxley, A. P. A., Lanfranconi, M. P., Würdemann, D., Ott, S., Schreiber,

S., McGenity, T. J., Timmis, K. N. and Nogales, B. (2010), Halophilic

archaea in the human intestinal mucosa. Environmental Microbiology,

12: 2398–2410. doi: 10.1111/j.1462-2920.2010.02212.x

3) S., Macfarlane and J.F., Dillon. (2007). Microbial biofilms in the human

gastrointestinal tract. The Society for Applied Microbiology, Journal of

Applied Microbiology 102 (2007) 1187–1196.

4) N.A., Campbell & J.B., Reece. (2008). Biology. 6th ed. San Francisco

(CA): Benjamin Cummings. p. 1247

5) Talaro, K. P and Chess, B. 2012. Foundations in Microbiology. 8th

Edition. McGraw-Hill. p. 828

6) L. A., Ogilvie and P. R., Hirsch. (2012). Microbial Ecological Theory:

Current Perspectives. Caister Academic Press (Norwich, UK). p. 120

Jib 221 assignment 2

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