General Principles of Medical Bacteriology

Dongwoo Shin Laboratory of Molecular Bacteriology Department of Molecular Cell Biology Sungkyunkwan University School of Medicine

Features of Bacteria

-

Unicellular organism

-

Small size: 1~2 µm in length

-

Haploid: number of a gene (or allele) is one

-

Short generation time: in the case of E. coli, ~20 min

-

Asexual replication

Structure of a Bacterial Cell

Classification of Bacteria

Introduction

Never Underestimate the Power of Bacteria!



Why bacteria are once again in the public health spotlight

• Antibiotics -first introduced into widespread clinical use in the 1950s -miracle drugs -appeared to give humans the final victory over bacteria • However....

-during the late 1980s, an increase in bacterial infections -by 1995, bacterial infectious diseases became one of the top five causes of death in the US (e.g. pneumonia, sepsis)

Introduction



Why bacteria are once again in the public health spotlight

• Emerging infectious diseases -power of rapid evolution -acquire genes that confer new virulence traits or resistance to antibiotics • Unfortunately....

-human victory over bacteria had not occurred and is not going to occur anytime in the future

Introduction



Current public health issues caused by bacterial infections

• Emerging infectious diseases -new diseases caused by newly discovered bacteria:

Legionella pneumophila

-long-known pathogens acquiring traits that make them more dangerous: antibiotic-resistant bacteria (

Streptococcus pneumoniae, Staphylococcus aureus

) • Food-borne infections -Outbreaks of

Escherichia coli

O157:H7 infections: in Japanese schoolchildren, 1996

Introduction



Current public health issues caused by bacterial infections

• Modern medicine as a source of new diseases -transplantation, cancer chemotherapy -suppressed immune systems of patients -increase in the chance of opportunistic infection:

Pseudomonas aeruginosa

• Microbiota shift diseases -Outgrowth of pathogens in the large intestine by antibiotics treatment

Human Microbiota

Commensal and Pathogenic Microbial Flora in Humans

The adult human intestine is...

- Home to an inconceivable number of microorganisms : The size of population is up to 100 trillion, ~10 times greater than the total number of our somatic and germ cells “Thus, it is appropriate to view ourselves as a composite of many species and our genetic landscape as an amalgam of genes embedded in our Homo sapiens genome and in the genomes of our affiliated microbial partners (the microbiome)”

- Jeffrey I. Gordon, Science (2005)

Commensal and Pathogenic Microbial Flora in Humans



In a healthy human,

- The internal tissues (e.g. brain, blood, muscles): normally free of microorganisms - Conversely, the surface tissues (e.g. skin and mucous membrane): constantly in contact with environmental microorganisms and become colonized by certain microbial species - Normal microbiota (= microflora, normal flora): mixture of microorganisms regularly found at any anatomical site - Bacteria make up most of the normal microbiota over the fungi and protozoa

Commensal and Pathogenic Microbial Flora in Humans



Exposure of an individual to an organism

1. Transient (hours or days) colonization 2. Permanent colonization 3. Disease production (Infection) Note: colonization vs. infection

Commensal and Pathogenic Microbial Flora in Humans The Relationship between Normal Microbiota and the Host



Normal microbiota

- participates in the metabolism of food products → provides essential growth factors - protects against infections with virulent microorganisms - stimulates immune response “Without these organisms life would be impossible”

Commensal and Pathogenic Microbial Flora in Humans The Relationship between Normal Microbiota and the Host



Normal microbiota for causing disease

- opportunistic pathogens : members of the patient’s normal microbiota; no disease in normal setting but disease when introduced into unprotected sites (e.g. blood, tissues); immunocompromised patients are more susceptible

Staphylococcus aureus Escherichia coli

- cf. strict pathogens : a few infections

Mycobacterim tuberculosis

(tuberculosis)

Neisseria gonorrhoeae

(gonorrhea)

Commensal and Pathogenic Microbial Flora in Humans



Microbiota IN and ON the human body

- A continual state of flux: determined by age, diet, hormones, health, and hygiene - Human fetus: in sterile environment - Newborn human: exposed to microbes from the mother and the environment; skin → oropharynx → gastrointestinal (GI) tract → other mucosal surfaces - Microbial population continues to change throughout the life

Commensal and Pathogenic Microbial Flora in Humans



Current knowledge of composition of human microbiota

- Based on culture methods: only a small proportion of the microbes can be cultivated - Human Microbiome Project (HMP) : initiated 2008; a large scale project for characterization of comprehensive human microbiota and analysis of its role in human health and disease; genomic techniques

Commensal and Pathogenic Microbial Flora in Humans



Respiratory Tract and Head



Upper Respiratory Tract: Mouth, Oropharynx, and Nasopharynx

The most common anaerobic bacteria:

Peptostreptococcus

,

Veillonella

,

Actinomyces

,

Fusobacterium

- The most common aerobic bacteria:

Streptococcus

,

Haemophilus

,

Neisseria

- Rarely associated with disease unless introduced into sterile sites (e.g. sinuses, middle ear, brain) - Potential pathogens:

Streptococcus pyogenes

,

Streptococcus pneumoniae, Staphylococcus aureus, Neisseria meningitidis, Heamophilus influenzae,

and Enterobacteriaceae

Commensal and Pathogenic Microbial Flora in Humans



Ear

The most common bacteria colonizing out ear: coagulase (-)

Staphylococcus

- Potential pathogens:

Streptococcus pneumoniae, Pseudomonas aeruginosa,

and Enterobacteriaceae

Commensal and Pathogenic Microbial Flora in Humans



Eye

The surface of the eye: colonized with coagulase (-)

Staphylococcus

-

S. pneumoniae, S. aureus, H. influenzae, N. gonorrhoeae, P. aeruginosa, Chlamydia trachomatis,

and

Bacillus cereus

: associated with disease 

Lower Respiratory Tract

Larynx, trachea, bronchioles, and lower airways: generally sterile - Acute disease of lower airways: virulent bacteria in mouth;

S. pneumoniae, S. aureus, Klebsiella

- Polymicrobial disease by chronic aspiration:

Peptostreptococcus

, anaerobic cocci, and anaerobic gram-negative rods

Commensal and Pathogenic Microbial Flora in Humans



Gastrointestinal Tract

 Colonized with microbes at birth  Home for diverse population of organisms throughout the life of the host  The populations remains relatively constant: But, antibiotic treatment disrupting the balanced flora 

Esophagus

Most organisms isolated from the esophagus: transient colonizers - Bacteria rarely cause disease of esophagus 

Stomach

Only small numbers of acid-tolerant bacteria: HCl, and pepsinogen -

Helicobacter pylori

: gastritis and ulcerative disease

Commensal and Pathogenic Microbial Flora in Humans



Small intestine

Colonized with many different anaerobic bacteria:

Peptostreptococcus

,

Porphyromonas

, and

Prevotella

- Gastroenteritis-causing bacteria:

Salmonella

and

Campylobacter

; present in small numbers as asymptomatic residents - Obstruction of small intestine (e.g. after surgery) → blind loop syndrome ( 막힌 고리 증후군 ): colonization and proliferation of the microbes in the large intestine → malabsorption syndrome ( 음식물 흡수 저해 )

Commensal and Pathogenic Microbial Flora in Humans



Large intestine

The biggest populations of microbes in the human body: ~10 11 bacteria / g of feces - The most common bacteria:

Bifidobacterium, Eubacterium, Bacteroides, Enterococcus

, and Enterobacteriaceae (e.g.

Escherichia coli

) -

E. coli

: present from birth to death, ~1% of the intestinal population but the most common aerobe responsible for intraabdominal disease -

Bacteroides flagilis

: a minor member of the intestinal flora but the most common anaerobe responsible for intraabdominal disease -

Bifidobacterium

and

Eubacterium

: the most common bacteria in large intestine but harmless

Commensal and Pathogenic Microbial Flora in Humans



Large intestine

Antibiotic treatment:  Rapidly alters the population  Causes proliferation of antibiotic-resistant bacteria (e.g. Enterococcus, Pseudomonas)  Rapid growth of

Clostridium difficile

colitis ( 거짓막염 ) → psedomembranous - Infection by enteric pathogens:  Shigella, and enterohemorrhagic E. coli  Disruption of the colonic flora → intestinal disease

Commensal and Pathogenic Microbial Flora in Humans



Genitourinary Tract

 Only anterior urethra and vagina: permanent colonization by microbes  Urinary bladder: transient colonization cleared by bactericidal activity of the uroepithelial cells and flushing action of voided urine 

Anterior urethra

-

Lactobacillus, Streptococcus

, coagulase (-)

Staphylococcus

- Transient colonization with fecal organisms:

Enterococcus

and Enterobacteiaceae urinary tract infection (UTI) -

N. gonorrhoeae

and

C. trachomatis

: common causes of urethritis

Commensal and Pathogenic Microbial Flora in Humans



Vagina

Diverse populations influenced by hormones - Newborn girls: Lactobacillus at birth → declined maternal estrogen →

Staphylococcus, Streptococcus

, and Enterobacteriaceae → estrogen production at puberty →

Lactobacillus

(predominant),

Staphylococcus, Streptococcus, Enterococcus

, Enterobacteiaceae -

N. gonorrhoeae

: common cause of vaginitis

Commensal and Pathogenic Microbial Flora in Humans



Skin

-Coagulase (-)

Staphylococcus

-

Clostridium perfringens

-

Acinetobacter

Power of Rapid Evolution of Bacteria

Bacterial Gene Transfer



Inter-strain transfer of DNA provides ‘genetic diversity’ of bacteria: Horizontal Gene Transfer

- e.g. Evolutionary process of pathogenic bacteria from nonpathogenic bacteria - e.g. Spread antibiotics resistance among bacteria



Three ways of bacterial gene transfer Conjugation : by physical interaction Transformation : transfer of a naked DNA Transduction : by bacteriophages

Bacterial Plasmids



Plasmid

-

Small and circular dsDNA molecules that can exist independently of host chromosome

-

Contains own replication origin

-

e.g. F plasmid

Bacterial Plasmids



Major types of plasmids Type Fertility Factor R plasmids Names Size (kb) Copy No.

F Factor 95-100 1-3 Hosts Phenotypes

E. coli, Salmonella

Conjugation R1 80 1-3 G(-) bacteria Ap R , Km R , Cm R 9 10-30

E. coli

Colicin production Col plasmids ColE1 Virulence plasmids Ent pZA10 83 56

E. coli S. aureus

Enterotoxin Enterotoxin B Metabolic plasmids TOL 75

Pseudomonas

Toluene degradation

Bacterial Conjugation



Conjugation - Transfer of genetic information by direct cell to cell contact



An electron micrograph of two

E. coli

in an early stage of conjugation cells

Bacterial Conjugation

e.g. F +

×

F mating - Donor strain (F + ) contains F factor carrying the genes for pilus formation and plasmid transfer

DNA Transformation



Transformation - Uptake of naked DNA by a cell



‘Competent’ cell - One able to take up DNA and be transformed

DNA Transformation

1. Natural transformation -

Streptococcus

,

Bacillus

,

Thermoactinomyces

,

Haemophilus

,

Neisseria

,

Moraxella

,

Acinetobacter

,

Helicobacter

,

Pseudomonas

- Mechanism in

S. pneumoniae

- DNA uptake machinery 2. Artificial transformation - Treatment of

E. coli

with CaCl 2 - Electric shock (electroporation)

Transduction

e.g. Generalized transduction

-

Occurs during the lytic cycle of phages and can transfer any part of the bacterial genome

-

Formation of transducing particle

-

E. coli

P1 and

Salmonella

phages P22



Genetic mechanism of evolution of pathogenic E. coli

Biological Mechanisms of Drug Resistance



Genetic mechanism of evolution of methicillin and vancomycin resistant Staphylococcus aureus