Transcript
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Endang S. Rahayu Fak. Teknologi Pertanian UGM

Mikrobiologi Dasar Mikrobiologi Dasar ((Fundamental Microbiology)Fundamental Microbiology)

Endang S. RahayuEndang S. Rahayu

Fakultas Teknologi PertanianFakultas Teknologi Pertanian

Universitas Gadjah MadaUniversitas Gadjah Mada

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Jadwal KuliahJadwal KuliahMikrobiologi DasarMikrobiologi Dasar

Semester II Th. 2005/2006Semester II Th. 2005/2006

Minggu Isi Kuliah Dosen

1 Pendahuluan : definisi dan aspek mikrobiologi; tujuan mempelajari mikrobiologi; sejarah mikrobiologi; mikrobiologi dan lingkungan; mikrobiologi sebagai ilmu dasar dan aplikasinya.

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2 Evolusi, sistematik, dan taksonomi mikrobia, penggolongan organisme dan taksonomi mikrobia : sistim 2-, 3-, 5- kingdoms; prokariot dan eukariot; taksonomi modern (pilogeni)

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3 Struktur sel : komponen-komponen sel dan fungsinya; perbedaan prokariot dan eukariot; membran sitoplasma, dinding sel, membran luar Gram negatif; Struktur internal (organel), flagella, pili, cilia, fimbriae, glycocalyx; bentuk dorman dari mikrobia (spora dan kista)

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4 Pertumbuhan mikrobia : pertumbuhan sel dan reproduksi; populasi mikrobia; cara pengukuran pertumbuhan; siklus pertumbuhan; factor lingkungan yang berpengaruh pada pertumbuhan (oksigen, pH, air/ aktivitas air, suhu

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5 Metabolisme : proses yang menghasilkan energi (energi kimiawi dan transfer energi, fosforilasi, jalur-jalur degradasi: nutrient komplek, glikolisa, regenerasi NAD, fermentasi dan respirasi); Proses yang membutuhkan energi (biosintesa komponen nitrogen, biosintesa karbohidrat, biosintesa lipida)

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6 Genetika mikrobia : Informasi genetik, replikasi, transkripsi, dan translasi ESR

Jadwal KuliahJadwal KuliahMikrobiologi DasarMikrobiologi Dasar

Semester II Th. 2005/2006Semester II Th. 2005/2006

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Lecture 1Lecture 1

• Introduction to microbiologyIntroduction to microbiology

• What is microbiologyWhat is microbiology

• Why study microbiologyWhy study microbiology

• The Historical Foundations of MicrobiologyThe Historical Foundations of Microbiology

• Abiogenesis – BiogenesisAbiogenesis – Biogenesis

• The golden age of microbiologyThe golden age of microbiology

• Anthrax & BioterrorismAnthrax & Bioterrorism

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What is Microbiology?What is Microbiology?

• The science of microorganisms (very The science of microorganisms (very small, unicellular organisms)small, unicellular organisms)

• The discipline is just over a century oldThe discipline is just over a century old

• Has given rise to molecular biology and Has given rise to molecular biology and biotechnologybiotechnology

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What are Microbes?What are Microbes?

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Sel ProkariotSel Prokariot

Sel EukariotSel Eukariot

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Sel prokariot dan eukariotSel prokariot dan eukariot

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Why study Microbiology (1)?Why study Microbiology (1)?

• Microorganisms were the first life on earthMicroorganisms were the first life on earth• Microorganisms created the biosphere that Microorganisms created the biosphere that

allowed multicellular organisms to evolveallowed multicellular organisms to evolve• Multicellular organisms evolved from Multicellular organisms evolved from

microorganismsmicroorganisms• >50% of the biomass on earth is >50% of the biomass on earth is

comprised of microorganismscomprised of microorganisms• Microorganisms will be on earth foreverMicroorganisms will be on earth forever

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4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0

BYA

Formation of O2 atmosphere

Cyanobacterial microfossils(oxygenic photosynthesis)

Earth formed

First microorganisms?

algae,marine invertebrates insects

mammals

No free O2

Reducing environment

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All cells of the present time organisms share a All cells of the present time organisms share a common ancestorcommon ancestor

An evolutionary phylogenetic treeAn evolutionary phylogenetic tree

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Why study Microbiology (2)?Why study Microbiology (2)?

• Our understanding of life has arisen Our understanding of life has arisen largely from studies of microorganisms largely from studies of microorganisms (biochemistry and genetics)(biochemistry and genetics)

• Studies of microorganisms continue to Studies of microorganisms continue to contribute to fundamental knowledge of contribute to fundamental knowledge of life processeslife processes

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Figure 1.4

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Why study Microbiology (3)?Why study Microbiology (3)?

• HealthHealth

• AgricultureAgriculture

• FoodFood

• EnvironmentEnvironment

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The Historical Foundations of The Historical Foundations of MicrobiologyMicrobiology

• The Development of the MicroscopeThe Development of the Microscope• The Debate over Spontaneous The Debate over Spontaneous

GenerationGeneration– RediRedi– NeedhamNeedham– SpallanzaniSpallanzani– VirchowVirchow– PasteurPasteur

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History of MicrobiologyHistory of Microbiology

• Robert Hooke- First person to peer into the microbial Robert Hooke- First person to peer into the microbial world, used compound lenses (1664)world, used compound lenses (1664)

• Anton van Leeuwenhoek- First true microbiologist, Anton van Leeuwenhoek- First true microbiologist, Dutch -Simple MicroscopeDutch -Simple Microscope

– single lens microscopes capable of 300 to 500X single lens microscopes capable of 300 to 500X (1632 - 1723)(1632 - 1723)

– drew and published (Proceeding of the Royal drew and published (Proceeding of the Royal Society in London) accurate pictures microscopic Society in London) accurate pictures microscopic algae, protozoa, bacteria, demonstrated “wee algae, protozoa, bacteria, demonstrated “wee beasties” (“wee animalcules”)beasties” (“wee animalcules”)

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Spontaneous GenerationSpontaneous Generation

• Spontaneous Generation- the idea that life Spontaneous Generation- the idea that life arose from non-living matter - Abiogenesisarose from non-living matter - Abiogenesis– Aristotle (384-322 BC) believed some simpler Aristotle (384-322 BC) believed some simpler

invertebrates could arise by spontaneous invertebrates could arise by spontaneous generationgeneration

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Spontaneous Generation Spontaneous Generation DebateDebate

• Francesco Redi (1626-1697)Francesco Redi (1626-1697)

– belief at the time- maggots spontaneously belief at the time- maggots spontaneously arose from rotting meatarose from rotting meat

– demonstrated that maggots were dependent demonstrated that maggots were dependent on the presence of fly eggs and of rotting on the presence of fly eggs and of rotting meatmeat

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Controversy ContinuesControversy Continues

• Leeuwenhoek’s discovery added fuel to the Leeuwenhoek’s discovery added fuel to the

controversycontroversy• Some proposed microorganisms could Some proposed microorganisms could

arise from spontaneous generation even if arise from spontaneous generation even if

higher organisms did nothigher organisms did not– They pointed out that boiled extracts of hay and They pointed out that boiled extracts of hay and

meat could give rise to microorganismsmeat could give rise to microorganisms

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Spontaneous Generation Spontaneous Generation (continued debate)(continued debate)

• In 1748, English Priest In 1748, English Priest John NeedhamJohn Needham reported the results of his experimentsreported the results of his experiments– boiled mutton broth and then tightly sealed the boiled mutton broth and then tightly sealed the

flaskflask– eventually many of the sealed flask became eventually many of the sealed flask became

cloudy with microorganismscloudy with microorganisms– he proposed that organic matter possessed a he proposed that organic matter possessed a

‘vital force’ that could give rise to life‘vital force’ that could give rise to life

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Spontaneous Generation Spontaneous Generation disproveddisproved

• Lazzaro Spallanzani - improved on Needham’s Lazzaro Spallanzani - improved on Needham’s experimental designexperimental design

• first- sealed seeds and water in glass flaskfirst- sealed seeds and water in glass flask• second- boiled themsecond- boiled them• no growth took place as long as the flasks no growth took place as long as the flasks

remained sealedremained sealed• proposed that air carried microbes to the cultureproposed that air carried microbes to the culture

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Controversy finally settledControversy finally settled• 1859, one of the fathers of modern microbiology, 1859, one of the fathers of modern microbiology, Louis Louis

PasteurPasteur offered strong evidence against Spontaneous offered strong evidence against Spontaneous GenerationGeneration

• Pasteur was a chemist by training but became interested Pasteur was a chemist by training but became interested in biology during his early studies on fermentations.in biology during his early studies on fermentations.

• At that time virtually nothing was known about At that time virtually nothing was known about microorganisms – except that they existed (this was microorganisms – except that they existed (this was discovered by Anton van Leeuwenhoek using home-discovered by Anton van Leeuwenhoek using home-made microscopes in 1684).made microscopes in 1684).

• It was known that microorganisms could be found in It was known that microorganisms could be found in decaying matter but it was not clear what role they were decaying matter but it was not clear what role they were playing in the decomposition process.playing in the decomposition process.

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Louis Pasteur (1822-1895)Louis Pasteur (1822-1895)

"Do not put forward "Do not put forward anything that you anything that you cannot prove by cannot prove by

experimentation"experimentation"

Drew out the necks of Drew out the necks of flasks so that they flasks so that they remained open to the airremained open to the airadded broth and boiled added broth and boiled it to destroy any it to destroy any microbes presentmicrobes presentflasks were incubated flasks were incubated for months and for months and remained sterileremained sterile

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Pasteur’s Swan Neck Flask Experiment

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The Golden Age of MicrobiologyThe Golden Age of Microbiology

• Fermentation and PasteurizationFermentation and Pasteurization

– PasteurPasteur

• The Germ Theory of DiseaseThe Germ Theory of Disease

– Agostino Bassi and PasteurAgostino Bassi and Pasteur

– Joseph LiterJoseph Liter

– Robert KochRobert Koch

• VaccinationVaccination

– Edward JennerEdward Jenner

– PasteurPasteur

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Rapid Advances 1857 -1914Rapid Advances 1857 -1914

• Rudolf Virchow introduced the concept of Rudolf Virchow introduced the concept of biogenesis: living cells can arise only from biogenesis: living cells can arise only from preexisting cells (1857).preexisting cells (1857).

• Louis Pasteur demonstrated that Louis Pasteur demonstrated that microorganisms are in the air everywhere and microorganisms are in the air everywhere and offered proof of biogenesis (1861).offered proof of biogenesis (1861).

• Agostino Bassi (1835) and Pasteur (1865) Agostino Bassi (1835) and Pasteur (1865) showed a causal relationship between showed a causal relationship between microorganisms and disease. microorganisms and disease.

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Pasteur’s Contributions to Pasteur’s Contributions to ScienceScience

• 1800- French Government (Napoleon III) 1800- French Government (Napoleon III) enlisted his help in their Wine Industryenlisted his help in their Wine Industry

– showed the decline in quality was due to a showed the decline in quality was due to a contaminating microbecontaminating microbe

– process of pasteurizationprocess of pasteurization

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Pasteur’s Contributions to Pasteur’s Contributions to ScienceScience

• Recognized that chickens became Recognized that chickens became immune to a bacterial pathogen if injected immune to a bacterial pathogen if injected with a weaken avirulent strain.with a weaken avirulent strain.

• Developed vaccines against rabies and Developed vaccines against rabies and anthraxanthrax

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• Pasteur’s perplexing problemPasteur’s perplexing problem

• while working on spontaneous generation he while working on spontaneous generation he experienced an anomalyexperienced an anomaly

• no matter how long some flasks were boiled, no matter how long some flasks were boiled, they always produced growththey always produced growth

• German botanist-German botanist-Ferdinand CohnFerdinand Cohn discovered discovered the existence of heat resistant bacterial sporesthe existence of heat resistant bacterial spores

• Pasteur reasoned that the flask that produced Pasteur reasoned that the flask that produced growth contained heat resistant bacterial growth contained heat resistant bacterial sporesspores

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• English physicist English physicist John TyndallJohn Tyndall dealt the dealt the absolute final blow to Spontaneous absolute final blow to Spontaneous Generation and resolved Pasteur’s problem, Generation and resolved Pasteur’s problem, he demonstrated the dust did indeed carry he demonstrated the dust did indeed carry microbes and if the dust were absent, the microbes and if the dust were absent, the broth remained sterile.broth remained sterile.

• He developed the process ofHe developed the process of tyndallization tyndallization which is fractional steam sterilizationwhich is fractional steam sterilization

• similar to the process we use today in the lab.similar to the process we use today in the lab.

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The Recognition of the The Recognition of the Microbial Role in DiseaseMicrobial Role in Disease

• Agostino Bassi (1835)Agostino Bassi (1835)– linked a microorganism to diseaselinked a microorganism to disease

• Pasteur (1865)Pasteur (1865)– disease of silkwormsdisease of silkworms

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The Recognition of the The Recognition of the Microbial Role in DiseaseMicrobial Role in Disease

• Joseph ListerJoseph Lister– English surgeonEnglish surgeon– developed an antiseptic system of surgery developed an antiseptic system of surgery

and heat sterilized his instruments as well as and heat sterilized his instruments as well as using phenol with surgical dressingusing phenol with surgical dressing

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Robert Koch Robert Koch (1843 –1910)(1843 –1910)

Robert KochRobert Kochestablished a relationship between anthrax and the established a relationship between anthrax and the bacterium- bacterium- Bacillus anthracisBacillus anthracisKoch’s techniques are still used today Koch’s techniques are still used today Today we know these as Koch’s PostulatesToday we know these as Koch’s PostulatesReceived the Nobel prize 1905 for his work on Received the Nobel prize 1905 for his work on tuberculosistuberculosis

Direct evidence of the role of bacterial Direct evidence of the role of bacterial in disease transmissionin disease transmission

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Koch’s Postulates

On the basis of these studies Koch proposed a series of criteria On the basis of these studies Koch proposed a series of criteria that should be used to establish the link between disease and that should be used to establish the link between disease and microorganisms. These are now known as Koch’s postulatesmicroorganisms. These are now known as Koch’s postulates ..

1: Microorganisms must 1: Microorganisms must alwaysalways be present in the infected be present in the infected animal and be absent in healthy individuals.animal and be absent in healthy individuals.

2: Pure cultures of the microorganism must be obtained from 2: Pure cultures of the microorganism must be obtained from the diseased animal.the diseased animal.

3: Such a culture must be capable, 3: Such a culture must be capable, in itselfin itself, of causing disease , of causing disease in healthy animals.in healthy animals.

4: The microorganism should be re-isolated from the diseased 4: The microorganism should be re-isolated from the diseased animal and should retain the same properties in pure culture animal and should retain the same properties in pure culture as the original pure culture.as the original pure culture.

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Koch’s Tuberculosis studies

• In 1881 Koch turned his attention to the study of a human disease In 1881 Koch turned his attention to the study of a human disease – – Tuberculosis (TBC).Tuberculosis (TBC).

• At that time TB killed 1 in 7 people and even today it accounts for At that time TB killed 1 in 7 people and even today it accounts for more than 1 in 20 human deaths worldwide. TBC was known to be more than 1 in 20 human deaths worldwide. TBC was known to be contagious but no “contagion” had been identified.contagious but no “contagion” had been identified.

• TBC is a disease of the lungs caused by the bacterium TBC is a disease of the lungs caused by the bacterium Mycobacterium tuberculosisMycobacterium tuberculosis. It is acquired by inhalation of . It is acquired by inhalation of infected droplets or dust particles in the air. infected droplets or dust particles in the air.

• The bacterium can persist in the lungs for long periods. It survives The bacterium can persist in the lungs for long periods. It survives and grows within macrophages.and grows within macrophages.

• In individuals with a low resistance acute pulmonary infection can In individuals with a low resistance acute pulmonary infection can occur with extensive damage to lung tissue. Spread of the occur with extensive damage to lung tissue. Spread of the bacterium to other parts of the body can then lead to death.bacterium to other parts of the body can then lead to death.

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Robert Koch (1843 – 1910)Robert Koch (1843 – 1910)

• Developing pure culture techniqueDeveloping pure culture technique

• Use potato slices – first found individual Use potato slices – first found individual bacterial colonies growing with different bacterial colonies growing with different appearanceappearance

• Microscopic examination revealed cells Microscopic examination revealed cells within a single colony were similarwithin a single colony were similar

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Pure Culture Key to Studying Pure Culture Key to Studying MicrobesMicrobes

• Definition : Pure culture is a population of Definition : Pure culture is a population of organisms, all of which are the progeny of organisms, all of which are the progeny of a single organisma single organism

• In nature, microbes almost never occur as In nature, microbes almost never occur as pure culturespure cultures

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Development of solidifying Development of solidifying agents and complex mediaagents and complex media

• Koch wanted to culture pathogens so he Koch wanted to culture pathogens so he used something similar to the body tissueused something similar to the body tissue

• Meat extractsMeat extracts

• Initially, gelatin (jell-o) was used to solidify Initially, gelatin (jell-o) was used to solidify beef extract broth but : (1) many beef extract broth but : (1) many organisms can digest gelatin and (2) it organisms can digest gelatin and (2) it melts at 37 melts at 37 ooC, the favored incubation C, the favored incubation temperature for most pathogens temperature for most pathogens

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Development of solidifying agentsDevelopment of solidifying agents

• Agar was suggested by Fannie Hesse wife of Agar was suggested by Fannie Hesse wife of Walther Hesse working in Koch LabWalther Hesse working in Koch Lab

• Story : Walther was working with gelatin plates Story : Walther was working with gelatin plates in summer and was having troublesin summer and was having troubles

• Asked wife : Why do your jellies and pudding Asked wife : Why do your jellies and pudding stay solid in warm weather?stay solid in warm weather?

• Fannie learned to use Agar-agar from a dutch Fannie learned to use Agar-agar from a dutch neighbour in New York who spent time in Asianeighbour in New York who spent time in Asia

• Agar-agar had been used as a gelling agent in Agar-agar had been used as a gelling agent in Asia for centuriesAsia for centuries

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AgarAgar

• Agar is complex polysaccharides derived Agar is complex polysaccharides derived from seaweedfrom seaweed

• Its only melts at 100 Its only melts at 100 ooC, and solidifies at C, and solidifies at 45 45 ooC C

• Non-toxic to most microorganismNon-toxic to most microorganism

• Stable at sterilization temperatureStable at sterilization temperature

• Only a very few bacteria have enzymes Only a very few bacteria have enzymes that digest itthat digest it

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Petri Dish

Petri dish was invented in 1877Petri dish was invented in 1877By an assistant of Koch –By an assistant of Koch –R.J. PetriR.J. Petri

Shallow glass plates Shallow glass plates

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Alexander FlemingAlexander Fleming

• Observed that the mold Observed that the mold Penicillium Penicillium inhibited the growth of a bacterial culture.inhibited the growth of a bacterial culture.

• Penicillin has been used clinically as anPenicillin has been used clinically as an

antibiotic since the 1940santibiotic since the 1940s

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Anthrax & BioterrorismAnthrax & Bioterrorism

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Bioterrorism 1Bioterrorism 1Use of biological material to inflict suffering Use of biological material to inflict suffering

and/or death on enemiesand/or death on enemies (board definition)(board definition)

• Biological warfare is not a new idea; In 184BC Hannibal Biological warfare is not a new idea; In 184BC Hannibal threw baskets of snakes onto enemy ships!threw baskets of snakes onto enemy ships!

• In 1346 Tartar soldiers catapulted plague victims into In 1346 Tartar soldiers catapulted plague victims into besieged Kaffka and successfully started an epidemic.besieged Kaffka and successfully started an epidemic.

• In 1763 the British army gave smallpox-contaminated In 1763 the British army gave smallpox-contaminated blankets to Indians who were loyal to the French.blankets to Indians who were loyal to the French.

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• In 1925 the Geneva convention outlawed the use of In 1925 the Geneva convention outlawed the use of biological weapons.biological weapons.

• In 1969 the US “officially” ended its program of developing In 1969 the US “officially” ended its program of developing biological weapons.biological weapons.

• In 1972 the Biological and Toxin weapons convention In 1972 the Biological and Toxin weapons convention approved a treaty to outlaw biological weapons approved a treaty to outlaw biological weapons (140 countries signed this treaty)(140 countries signed this treaty)

• It is likely that many countries still have It is likely that many countries still have the capacity for biological.the capacity for biological.

Bioterrorism 2Bioterrorism 2

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What is Anthrax?What is Anthrax?

• Anthrax is a disease that is caused by the bacterium Anthrax is a disease that is caused by the bacterium Bacillus anthracisBacillus anthracis (the link was first proved by (the link was first proved by Koch).Koch).

• B. anthracisB. anthracis is a Gram-positive, spore-forming is a Gram-positive, spore-forming common soil microorganism.common soil microorganism.

• Primarily a disease of livestock; catlle, sheep etc.Primarily a disease of livestock; catlle, sheep etc.

• Pasteur devised the first vaccine against anthrax. Pasteur devised the first vaccine against anthrax.

• Today vacccination is still the primary means of Today vacccination is still the primary means of controlcontrol.

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Anthrax in HumansAnthrax in Humans

• Contrary to reports in the popular press – Contrary to reports in the popular press – B. anthracisB. anthracis is is not a good human pathogennot a good human pathogen

• Humans are fairly resistant to infection. Even textile Humans are fairly resistant to infection. Even textile workers exposed to infected animal products have a low workers exposed to infected animal products have a low infection rate.infection rate.

• Large numbers of spores (8,000-10,000) are required to Large numbers of spores (8,000-10,000) are required to infect humans. This exposure is very rare in the “normal” infect humans. This exposure is very rare in the “normal” environment.environment.

• Breathing air around infected animal carcasses would Breathing air around infected animal carcasses would lead to the inhalation of around 2-10 spores every 10 lead to the inhalation of around 2-10 spores every 10 minutes! minutes!

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Infection by Infection by B. anthracisB. anthracisThere are 3 routes of infection by this pathogen:There are 3 routes of infection by this pathogen:

1)1) Entry by ingestionEntry by ingestion

• Gastrointestinal anthrax can arise byGastrointestinal anthrax can arise by the ingestion of contaminated meat, usually from an the ingestion of contaminated meat, usually from an animal that died of anthrax.animal that died of anthrax.

• Infection reaches intestinal lymph nodes, then causes Infection reaches intestinal lymph nodes, then causes septacemia. Exact pathogenesis is not understood.septacemia. Exact pathogenesis is not understood.

• Very rare – but in cases that present clinically the Very rare – but in cases that present clinically the mortality rate is about 50%.mortality rate is about 50%.

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2) ) Entry through skin lesionsEntry through skin lesions.

• Spores enter damaged areas of skin. Spores enter damaged areas of skin.

• Erythematous papule after 2-5 days (red bump).Erythematous papule after 2-5 days (red bump).

• Centre of lesion darkens after 7-10 days.Centre of lesion darkens after 7-10 days.

• Usually heals without intervention (spread to blood occurs Usually heals without intervention (spread to blood occurs in less than 5% of cases).in less than 5% of cases).

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3) Entry by inhalationEntry by inhalation

• Inhalation of spores can lead to pulmonary anthrax – also Inhalation of spores can lead to pulmonary anthrax – also called “woolsorters disease”.called “woolsorters disease”.

• Malaise within 1-5 days, including cough, fever and chest Malaise within 1-5 days, including cough, fever and chest pain.pain.

• Difficulty with breathing, blueish skin colour, swelling of Difficulty with breathing, blueish skin colour, swelling of the chest.the chest.

• Once swelling is evident the disease beyond treatment Once swelling is evident the disease beyond treatment and death usually occurs.and death usually occurs.

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Anthrax and bioterrorismAnthrax and bioterrorism

Advantages:Advantages:

• Very easy to obtain and to grow.Very easy to obtain and to grow.• Spores very stable.Spores very stable.• Spores easily inhaled.Spores easily inhaled.• High fatality rate in cases of pulmonary anthrax. High fatality rate in cases of pulmonary anthrax.

Disadvantages:Disadvantages:

• Dispersing spores over a wide area is difficult.Dispersing spores over a wide area is difficult.• Not contagious – no person-to-person spread.Not contagious – no person-to-person spread.• If diagnosed early it is easily treated with antibiotics.If diagnosed early it is easily treated with antibiotics.

Page 60: ESR - FM 1 Introduction

Endang S. Rahayu Fak. Teknologi Pertanian UGM

Concept CheckConcept Check

Microorganisms can be both beneficial Microorganisms can be both beneficial and harmful to humans. Although we tend and harmful to humans. Although we tend to emphasize the harmful microorganisms to emphasize the harmful microorganisms (infectious and disease agents), many (infectious and disease agents), many more microorganisms are beneficial than more microorganisms are beneficial than harmful.harmful.

In what ways are microorganisms In what ways are microorganisms important in the food industries ?important in the food industries ?


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