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Chapter 4 Microbiological Contaminants. Waterworks Operations WQT 111 Lecture 4. E. coli are bad?. True False. Objectives. Compare virus (small), bacteria (medium), and protozoa (large) Review Total Coliform Rule. Go Over Total Coliform Analytical Procedures.

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Chapter 4 microbiological contaminants

Chapter 4 Microbiological Contaminants

Waterworks Operations

WQT 111

Lecture 4


E coli are bad
E. coli are bad?

  • True

  • False


Objectives
Objectives

  • Compare virus (small), bacteria (medium), and protozoa (large)

  • Review Total Coliform Rule.

  • Go Over Total Coliform Analytical Procedures.

  • Perform Colilert Test of Unknown sample.


Viruses
Viruses

  • Smallest living entities 10 - 25 nm .  Viruses can be observed only with the aid of an electron microscope.

  • Viruses are obligate intracellular parasites - they cannot live or multiply outside of a host cell. 

  • More resistant to chlorine relative to bacteria

  • Viruses are not cellular organisms.  Many consist of a protein coat or capsid and internal nucleic acid, either DNA or RNA. 

  • Viruses do not produce enzymes for metabolism.

  • In the process of multiplying, viruses kill the infected host cells.   Important viral waterborne infections include hepatitis A and gastroenteritis (norwalk and rotavirus)





In the processes of viral metabolism enzymes are exuded which attack the host cell
In the processes of viral metabolism enzymes are exuded which attack the host cell?

  • True

  • False



Bacteria
Bacteria nucleus?

  • Smallest cellular organisms.  A typical bacterium is about 1 micrometer in size.  Bacteria have a unique cell construction that sets them apart from all other living organisms.

  • The vast majority of bacteria are free living soil and water microorganisms. 

  • Bacteria can exist in conditions that no other living organism can tolerate.  Conan the bacterium

  • Bacteria cause many diseases in animals and man.  Important waterborne infectious diseases include campylobacterosis, legionnaires disease, shigellosis, typhoid, enterovirulent E. coli 0157, and cholera.


E coli
E. coli nucleus?

http://pathport.vbi.vt.edu/pathinfo/pathogens/E.coli_O157H7.html

http://www.onr.navy.mil/media/view_image.asp?ID=63&SubID=64

  • E.coli are rod shaped, facultative anaerobic, gram-negative bacillus

  • Waste processing, Vitamin B and K production, and food absorption

  • They are indicators of fecal coliform

    • 10 billion-100 trillion per day per person


E coli are necessary for human survival
E. coli nucleus? are necessary for human survival?

  • True

  • False


E coli are rod shaped facultative anaerobic gram negative bacillus
E.coli nucleus? are rod shaped, facultative anaerobic, gram-negative bacillus?

  • True

  • False


Escherichia coli type 0157 h7 bacteria enteric pathogen
Escherichia coli nucleus?type 0157:H7 (bacteria, enteric pathogen)

  • The illness is characterized by severe cramping (abdominal pain) and diarrhea which is initially watery but becomes grossly bloody. Occasionally vomiting occurs.

  • Fever is either low-grade or absent.

  • The illness is usually self-limited and lasts for an average of 8 days. Some individuals exhibit watery diarrhea only.

  • Infectious dose is a few as 10 bacteria cells.

http://en.wikipedia.org/wiki/2006_United_States_E._coli_outbreak


"E Coli H0157" is the bacterium that is the best known serotype of "Enterohemorrhagic E. Coli"and known by its more formal scientific name of Escherichia coli 0157:H7. Transmitted from undercooked hamburger meat and raw milk, it releases verotoxin, which causes inflammation of the lower gastrointestinal tract, often resulting in bloody bowel movements, potentially several other illnesses and/or death.

  • True

  • False


Bacteria1
Bacteria serotype of "Enterohemorrhagic E. Coli"


Bacteria2
Bacteria serotype of "Enterohemorrhagic E. Coli"

rod

coccus

spirillum


Most total coliforms are rod shaped
Most total coliforms are rod shaped? serotype of "Enterohemorrhagic E. Coli"

  • True

  • False


Protozoa
Protozoa serotype of "Enterohemorrhagic E. Coli"

  • Protozoa are mostly single celled organisms that have a complex cell construction (5-100 mm).

  • Protozoa are classified into groups according to their method of motility:

    • flagellates, move with the aid of one or more whip-like flagella.

    • ciliates, move with the aid of a group of short hair-like cilia.

    • amoeba, move with the aid of temporary cell projections called pseudopods (false feet).

  • Most protozoa are free living water and soil microorganisms.

  • Protozoa produce important diseases in animals and man.  Important waterborne diseases include giardiasis, cryptosporidiosis and amoebic dysentery.


Protozoa like to eat bacteria
Protozoa like to eat bacteria? serotype of "Enterohemorrhagic E. Coli"

  • True

  • False


A cilia is a whip like tail structure in a protozoa
A cilia is a whip like tail structure in a protozoa? serotype of "Enterohemorrhagic E. Coli"

  • True

  • False


Protozoa intestinal parasites water borne diseases giardia lamblia cryptosporidium parvum
Protozoa Intestinal Parasites & Water Borne Diseases serotype of "Enterohemorrhagic E. Coli"Giardia lambliaCryptosporidium parvum


Giardiasis intestinal parasite
Giardiasis (intestinal parasite) serotype of "Enterohemorrhagic E. Coli"

  • Giardia lamblia (intestinalis) is a single celled animal, i.e., a protozoa, that moves with the aid of five flagella.

  • Giardia found in domestic animals (dogs and cats) and wild animals (beavers, ducks, and bears)

  • Most frequent cause of non-bacterial diarrhea in North America. ~ 25% of the cases of gastrointestinal disease

  • Giardiasis is most frequently associated with the consumption of contaminated water . 2% of US population. Oregon second most reported cases 1980s

  • Ingestion of one or more cysts may cause disease

  • Illness lasts for 1 to 2 weeks (chronic infections can last months to years). Treat with Flagyl=antibiotic

http://www.cfsan.fda.gov/~mow/chap22.html


Giardia lamblia is a bacteria that causes waterborne infection
Giardia lamblia serotype of "Enterohemorrhagic E. Coli" is a bacteria that causes waterborne infection?

  • True

  • False


Giardia is found in domestic animals dogs and cats and wild animals beavers ducks and bears
Giardia serotype of "Enterohemorrhagic E. Coli" is found in domestic animals (dogs and cats) and wild animals (beavers, ducks, and bears)?

  • True

  • False


Ingestion of one or more giardia cysts may cause disease
Ingestion of one or more giardia cysts may cause disease? serotype of "Enterohemorrhagic E. Coli"

  • True

  • False



Giardia lamblia protozoa
Giardia Lamblia/Protozoa in North America?


Cryptosporidiosis protozoa
Cryptosporidiosis (protozoa) in North America?

  • Cryptosporidium parvum, a single-celled animal, i.e., a protozoa, is an obligate intracellular parasite.

  • Infects many herd animals (cows, goats, sheep among domesticated animals, deer and elk among wild animals)

  • The infective stage (oocyst) is 3 µm in diameter or about half the size of a red blood cell.

  • Severe watery diarrhea lasting 2-4 days (no reliable treatment for cryptosporidiosis)

  • Serological surveys indicate that 80% of the population has had cryptosporidiosis.

http://www.cfsan.fda.gov/~mow/chap24.html


Cryptosporidiosis
Cryptosporidiosis in North America?

  • The new “superbug” Cryptosporidium parvum is first identified as an important waterborne infection in the 1990's.

  • Resistant to chlorination

  • Oregon reports waterborne outbreaks in 1992 Talent Oregon communities.

  • 1993 largest waterborne disease outbreak in US

  • ~400,000 Milwaukee Wisconsin

http://www.cfsan.fda.gov/~mow/chap24.html


Cryptosporidium protozoa
Cryptosporidium/Protozoa in North America?


Total coliform rule
Total Coliform Rule in North America?

  • Total Coliform Rule (TCR) on June 29, 1989

  • monitor for total coliforms at a frequency proportional to the number of people served

  • Further test that culture for the presence of either fecal coliforms or Escherichia coli;

  • Take samples at end of distribution systems

  • If positive occurs:

    • Re-collect and analyze at least 3-4 repeat samples within 24 hours :

    • 1 at the same tap as the positive

    • The others at sites located within 5 or fewer service connection adjacent (upstream and downstream) to the location of the routine positive sample; and

  • Take at least 5 routine samples the next month of operation.

  • 16 different species of Total Coliforms!


Epa approved methods total coliforms
EPA APPROVED METHODS in North America?Total Coliforms

  • Presence–absence (P-A) The P-A test is a qualitative procedure that was developed as a sensitive, economical, and efficient means of analyzing drinking water samples

  • Membrane filter (MF) The MF procedure was introduced to bacteriological water analysis in 1951, after its capacity to produce results equivalent to those obtained by the MTF procedure was demonstrated

  • Multiple tube fermentation (MTF) procedures - The MTF procedure, in comparison with the MF procedure, lacks precision, is more difficult to perform, and takes longer to produce results; because of this, the latter has largely replaced it for routine examinations of drinking water.


Presence absence
Presence/Absence in North America?

  • Incubate tubes or bottles

  • YES OR NO?

  • Fluorescent end product from E.Coli fermentation

Positive

E.Coli


What is the most common method used in labs to test for total coliform and e coli
What is the most common method used in labs to test for total coliform and E. coli?

  • DMA

  • Green

  • Colilert

  • Lamp


Total coliforms
Total Coliforms total coliform and


Total coliforms m endo broth

Membrane Filtration total coliform and

Total Coliforms (m-Endo broth)

Total Coliform

Total Coliform

Total Coliforms (http://dl.clackamas.cc.or.us/wqt111/unit-8-coliformtest.htm)

  • Red colony with a metallic sheen within 24±2 hours at 35±0.2oC


Fecal coliforms m fc broth
Fecal Coliforms (m-FC broth) total coliform and

  • Blue colonies for fecal coliforms. Gray to cream colored are non fecal coliforms

Fecal Coliform

Fecal Coliforms (http://dl.clackamas.cc.or.us/wqt111/unit-8-coliformtest.htm)

  • Blue colony within 24±2 hours at 44.5±0.5oC

Fecal Coliform

MI-Media: Pure Culture of Escherichia coli ATCC 25922 with UV Light (http://www.whatman.com/products/?pageID=7.61.409.297)


In the membrane filter method the number of coliforms is estimated by the
In the membrane filter method, the number of coliforms is estimated by the:

  • Number of colonies grown

  • Number of negative tubes

  • Number of positive tubes

  • Sum of positive and negative tubes


Fecal coliforms are incubated at 44 5 o c for 24 hours
Fecal Coliforms are incubated at 44.5 estimated by the: oC for 24 hours

  • True

  • False


When conducting the fecal coliform membrane test count all the colonies that are in color
When conducting the fecal coliform membrane test, count all the colonies that are _______ in color.

  • Red

  • Yellow

  • Blue

  • Orange


Fecal coliforms are incubated in a water bath at this temperature for 24 hours
Fecal coliforms are incubated in a water bath at this temperature for 24 hours.

  • 44.5 oC

  • 35.0 oC

  • 103 oC

  • 37.5 oC


The recommended holding time for a fecal coliform sample prior to analysis is 24 hours or less at 4 deg. C.?

  • True

  • False


Multiple tube fermentation method
Multiple Tube Fermentation Method prior to analysis is 24 hours or less at 4 deg. C.?

Inoculate lauryl tryptose broth and incubate for 24h at 35oC

Gas or acidity

Transfer to BGBB

incubate 48 h at 35oC

No gas or acidity

Incubate

24 h at 35oC

No gas produced

- test. Coliform group absent

Gas produced

Confirm as in (1)

Acidic

growth produced

Confirm as in (1)

  • No gas or acidic

  • growth produced

  • test.

  • Coliform

  • group absent

Gas produced

Coliform group confirmed

Re-inoculate in fresh BGBB

Positive completed phase


Multiple tube fermentation method1
Multiple Tube Fermentation Method prior to analysis is 24 hours or less at 4 deg. C.?

  • Step 1 presumptive phase

  • Use lauryl tryptose broth

  • Grow “stressed” organisms

  • Confirm positives

  • Calculate MPN

Gas

Growth

http://www.bact.wisc.edu/Microtextbook/index.php?module=Book&func=displaychapter&chap_id=55&theme=printer


Habitats of fecal and nonfecal coliforms
Habitats of Fecal and NonFecal Coliforms prior to analysis is 24 hours or less at 4 deg. C.?


There are 16 different species of coliforms in nature
There are 16 different species of coliforms in nature? prior to analysis is 24 hours or less at 4 deg. C.?

  • True

  • False


E coli results must always be higher or lower than fecal coliform results
E. coli prior to analysis is 24 hours or less at 4 deg. C.? results must always be higher or lower than fecal coliform results?

  • Higher

  • Lower


What is an ideal indicator
What is an IDEAL INDICATOR? prior to analysis is 24 hours or less at 4 deg. C.?

  • Be applicable in all water types.

  • ALWAYS be present in water when pathogenic bacteria of fecal contamination are present.

    • Density or numbers should relate to amount/degree of pollution

    • Greater survival time in water

    • Disappears rapidly following the disappearance of pathogens


Coliform bacteria- prior to analysis is 24 hours or less at 4 deg. C.? Microorganisms predominantly inhabiting the intestines of humans and other animals, but also occasionally found elsewhere. They include all aerobic and facultative anaerobic, Gram-negative, nonspore-forming, rod-shaped bacteria that ferment lactose with the production of gas. Also included are all bacteria that produce a dark, purplish-green metallic sheen by the membrane filter technique used for coliform identification.

  • True

  • False


What is an ideal indicator1
What is an IDEAL INDICATOR? prior to analysis is 24 hours or less at 4 deg. C.?

  • Absent from a bacteriologically safe water

  • Easily analyzed for without any anomalies or false positives.

  • Greater survival time in water

  • Disappears rapidly following the disappearance of pathogens

  • Be harmless to man or other animals


Total and fecal coliform are all pathogenic
Total and fecal coliform are all pathogenic? prior to analysis is 24 hours or less at 4 deg. C.?

  • True

  • False


Coliform bacteria are
Coliform bacteria are: prior to analysis is 24 hours or less at 4 deg. C.?

  • More resistant to chlorination than pathogenic bacteria

  • Less resistant to chlorination than pathogenic bacteria

  • A group of pathogens causing hepatitis

  • A group of pathogens causing cholera


Which of the following are true about indicator organisms
Which of the following are true about indicator organisms? prior to analysis is 24 hours or less at 4 deg. C.?

  • Be present when pathogen is present

  • Be at lower numbers then the pathogen

  • Have a shorter lifespan then a pathogen

  • Be able to kill a pathogen and a lab technician in the same day


Fecal coliforms are a subset of total coliform bacteria
Fecal coliforms are a subset of total coliform bacteria? prior to analysis is 24 hours or less at 4 deg. C.?

  • True

  • False


Coliforms are used as indicator bacteria to verify the effectiveness of disinfection techniques because ______.

  • They are the most numerous of all wastewater pathogens

  • They cause the worst diseases of any of the wastewater pathogens

  • They are easier to detect and are harder to destroy than most pathogenic organisms

  • They have a symbiotic relationship with pathogens; one cannot survive without the other.


Coliform bacteria are1
Coliform bacteria are: effectiveness of disinfection techniques because ______.?

  • Commonly found throughout distribution systems

  • Desirable in storage tanks for iron digestion

  • Highly resistant to chlorine

  • Indicator organisms

  • Deadly


. effectiveness of disinfection techniques because ______.

# of organisms

  • A comparison of pathogens to indicator organism

  • Indicator organisms should outlive a pathogen

  • An indicator organism can coexist with a pathogen

  • All of the above

Time


This is the special sugar that fecal coliform bacteria can metabolize
This is the special sugar that fecal coliform bacteria can metabolize.

  • Glucose

  • Fructose

  • Lactose

  • Rosalic acid


Coliform bacteria and the procedures used to test for them are divided into the two categories of
Coliform bacteria and the procedures used to test for them are divided into the two categories of _______.

  • MF and MTF

  • LTB and BGB

  • Total and fecal coliforms

  • M-Endo and M-FC


Objective #4 on the syllabus (Understand the fundamentals of water microbiology, especially as it relates to waterborne infectious disease) has been met?

  • Strongly Agree

  • Agree

  • Disagree

  • Strongly Disagree


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