Etiket Arşivleri: Agar

Ogye Agar Base

OGYE AGAR BASE (7655)

Intended Use

OGYE Agar Base is used with oxytetracycline in the detection and isolation of yeasts and molds from foods.

Product Summary and Explanation

Mossel, Visser, and Mengerink1,2 described Oxytetracycline-Glucose Yeast Extract (OGYE) Agar for the detection and enumeration of yeasts and molds in clinical specimens, foods, and other specimens of sanitary significance. Mossel et al.1,2 described improved recovery of yeasts and molds compared to acidified agar. This medium is supplemented with oxytetracycline as a selective agent, rather than relying on the low pH of acidified agar to suppress bacterial growth. Acidified agar is commonly used for detection of yeasts and
molds in foods and dairy products, and OGYE Agar Base provides an alternate medium.

Principles of the Procedure

Yeast Extract provides essential vitamins to stimulate the growth of yeasts and molds in OGYE Agar Base. Dextrose is the carbon energy source. Agar is the solidifying agent. The supplement, oxytetracycline, is a selective agent used to inhibit bacterial growth.


Source: https://dotscientific.com/pdf_library/ogye_agar_base_7655.pdf

Coliform Analysis in Water/ Membrane Filtration

General introduction:

Membrane  filtration  systems  are  especially  used  when  the  sample  like  drinking  water  or  fruit j uices   which  can  easily  pass through the filter  and  contain  really small  amount  of  microorganisms  less than   1 CFU/ ml.

About coliform bacteria and why do we do this test:  

Coliform   bacteria  are   present   in  the  environment  and  feces  of  all  warm-blooded  animals  and   humans.  Coliform  bacteria  are  unlikely  to  cause  illness.  However,  their  presence  in  drinking  water   indicates that  disease-causing  organisms  (pathogens)  could  be  in the  water  system.  Most  pathogens   that  can  contaminate  water  supplies  come  from  the  feces  of  humans  or  animals.  Testing  drinking   water   for   all   possible   pathogens   is   complex,   time-consuming,   and   expensive.   It   is   easy   and   inexpensive to test for coliform  bacteria.  If testing detects coliform  bacteria  in a water sample, water   services  search  for  the  source  of  contamination  and  restore  safe  drinking  water.  There  are  three   groups  of  coliform  bacteria.  Each  is  an  indicator  of  drinking  water  quality  and  each  has  a  different   level  of  risk. Total  coliform  is  a  large  collection  of  different  kinds  of  bacteria.  Fecal  coliform  are types   of  total  coliform  that  exist  in  feces.  E.  coli  is  a  subgroup  of  fecal  coliform.  Labs  test  drinking  water   samples for total  coliform.  If total  coliform  is  present, the  lab  also tests the sample for fecal  coliform   or E. coli, depending on the lab testing method.    Total  coliform  bacteria  are  common  in  the  environment  (soil  or  vegetation)  and  are  generally   harmless.  If  a  lab  detects  only  total  coliform  bacteria  in  drinking  water,  the  source  is  probably   environmental  and  fecal  contamination  is  unlikely.  However,  if  environmental  contamination  can   enter  the  system,  pathogens  could  get  in,  too.  It  is  important  to  find  and  resolve  the  source  of  the   contamination.     Fecal  coliform  bacteria  are  a  subgroup  of  total  coliform  bacteria.  They  exist  in  the  intestines  and   feces  of  people  and  animals.  The  presence  of  fecal  coliform  in  a  drinking  water  sample  often   indicates  recent fecal  contamination. That  means there  is  a  greater  risk that  pathogens  are  present.   E.  coli  is  a  subgroup  of  the  fecal  coliform  group.  Most  E.  coli  bacteria  are  harmless  and  exist  in  the   intestines  of   people  and  warm-blooded  animals.  However,  some  strains  can  cause   illness.  The   presence  of  E.  coli  in  a  drinking  water  sample  usually  indicates  recent  fecal  contamination.  That   means there is a greater risk that pathogens are present.    Note:  E.  coli  outbreaks  receive  a  lot  of  media   coverage.  A  specific  strain  of  E.  coli  bacteria   known as E. coli O157:H7 causes most of those   outbreaks.  When  a  drinking  water  sample  is   reported as  “E. coli  present,”  it does  not  mean   that   O157:H7   is   present.   However,   it   does   indicate  recent fecal  contamination.  Boiling  or   disinfecting       contaminated         drinking      water   destroys      all   forms      of   E.    coli,   including   O157:H7.

Besiyeri Hazırlama ( MEGEP )

1. BESİYERİ (ORTAM, VASAT)

1.1. Besiyeri Çeşitleri

1.1.1. Fiziksel Özelliklerine Göre

1.1.2. Kaynaklarına Göre

1.1.3. Kullanım Amacına Göre

1.2. Birçok Besiyerinin Bileşimine Giren Temel Maddeler

1.2.1. Agar

1.2.2. Pepton

1.2.3. Maya Ekstraktı (Yeast Extract)

1.2.4. Et Ekstraktı (Beef Extract)

1.2.5. Tuz (NaCl)

1.2.6. Jelâtin

1.2.7. Karbohidratlar

1.2.8. Kan

1.2.9. Tampon Maddeler (Buffer)

1.2.10. İnorganik maddeler

1.3. Besiyerinin Sahip Olması Gereken Özellikler

1.4. Besiyeri Formülasyonları

1.5. Etiket Bilgisini Kullanma

1.5.1. Etikette Yer Alan Bilgiler

1.5.2. Etiket Bilgilerine Göre Hesaplama Örnekleri

1.6. Besiyeri Hazırlık Aşamaları

1.6.1. Tartım

1.6.2. Kaba Aktarma

1.6.3. Çözündürme

1.6.4. Berraklaştırma

1.6.5. Besiyeri pH’sının Ayarlanması

1.6.6. Besiyeri pH’ının Ölçülmesi

1.7. Besiyeri Hazırlanan Kaplara Hazırlama Bilgilerini Yazma

1.8. Besiyerlerini Sterilizasyona Hazırlama Aşamaları

2. BESİYERLERİNİN STERİLİZASYONU VE MUHAFAZASI

2.1. Besiyeri Sterilizasyonu ve Aşamaları

2.1.1. Otoklavda Sterilizasyon

2.1.2. Su Banyosunda Sterilizasyon

2.1.3. Filtre ile Sterilizasyon:

2.1.4. Mikrodalga Fırında Sterilizasyon

2.2. Sterilizasyonla İlgili Dikkat Edilecek Noktalar

2.3. Sterilizasyon Sonrası İşlemler

2.3.1. Katkı Maddelerinin Eklenmesi

2.3.2. Sterilize Besiyerinin Petri Kutularına Dökülmesi

2.3.3. Agarlı Besinyerlerinin Yüzeylerinin Kurutulması

2.3.4. Tüplerdeki Besiyerlerine Yapılan İşlemler

2.3.5. Agarlı Besiyerlerinin Yeniden Eritilmesi

2.4. Sterilizasyon Kontrolü

2.5. Besiyerlerinin Muhafazası

2.6. Hazır Ticari Besiyerleri ve Muhafazası

Kaynak: http://www.megep.meb.gov.tr/?page=moduller

Besiyerleri ( Prof.Dr. A.Kadir Halkman )

  • Genel Bilgiler

  • Kısa Tarihçe

  • Besiyeri Hazırlama Şekilleri

  • Formülden Hazırlama

  • Dehidre Besiyerlerinden Hazırlama

  • Kullanıma Hazır Besiyerleri

  • Özel Uygulamalar

  • Besiyeri Bileşimine Giren Maddeler

  • Su

  • Peptonlar

  • Maya Ekstraktı

  • Et Ekstraktı

  • Malt Ekstraktı

  • Beyin ve Kalp Ekstraktı

  • Agar

  • Karbohidratlar

  • Tuz

  • Tampon Maddeler

  • pH İndikatörleri

  • Redoks İndikatörleri

  • Diğer İndikatörler

  • İnhibitörler

  • Diğer Maddeler

  • Besiyeri Çeşitleri

  • Genel Besiyerleri

  • Selektif Besiyerleri

  • Diferansiyel (Ayırt Edici; Fark Ettirici) Besiyerleri

  • Zenginleştirme Besiyerleri

  • İdentifikasyon Besiyerleri

  • Diğerleri

  • Besiyerlerinin Hazırlanması

  • Cam Malzeme

  • Tartım ve Eritme

  • pH Ayarlama

  • Katkıların İlavesi

  • Petri Kutularına Döküm

  • Kurutma

  • Yeniden Eritme

  • Depolama

  • Dehidre Besiyerlerinin Depolanması

  • Çözeltilerin Depolanması

  • Katkıların Depolanması

  • Hazırlanmış Besiyerlerinin Depolanması

  • Çözeltilerin Hazırlanması

  • Besiyeri Kalite Kontrolü

  • Besiyeri Hataları

  • Tehlikeli Maddeler

Besiyerleri Sunum

BESİYERLERİ

Peptonlar
Maya ekstraktı
Agar
Karbonhidratlar
Genel amaçlı besiyerleri
Seçici besiyerleri
Zenginleştirici besiyerleri
Ön zenginleştirme besiyerleri

Kaynak: http://www2.bayar.edu.tr/muhendislik/gida/docs/databank/Uygulama-besiyerleri.pps

Laboratory‎ > ‎Culture Media & Culture Methods ( Babitha ELIAS )

CULTURE MEDIA & CULTURE METHODS
Babitha Elias
Bacteria have to be grown (cultured) for them to be identified.
By appropriate procedures they have to be grown separately (isolated) on culture media and obtained as pure for study.
History
The original media used by Louis Pasteur – urine or meat broth
Liquid medium – diffuse growth
Solid medium – discrete colonies.
Colony – macroscopically visible collection of millions of bacteria originating from a single bacterial cell.
Cooked cut potato by Robert Koch – earliest solid medium
Gelatin – not satisfactory
– liquefy at 24oC
Agar
Frau Hesse
Used for preparing solid medium
Obtained from seaweeds.
No nutritive value
Not affected by the growth of the bacteria.
Melts at 98oC & sets at 42oC
2% agar is employed in solid medium
Types of culture media
I.Based on their consistency
a) solid medium
b) liquid medium
c) semi solid medium
II.Based on the constituents/ ingredients
a) simple medium
b) complex medium
c) synthetic or defined medium
d) Special media
Special media
Enriched media
Enrichment media
Selective media
Indicator media
Differential media
Sugar media
Transport media
Media for biochemical reactions
III.Based on Oxygen requirement
– Aerobic media
– Anaerobic media
Solid media – contains 2% agar
Colony morphology, pigmentation, hemolysis can be appreciated.
Eg: Nutrient agar, Blood agar
Liquid media – no agar.
For inoculum preparation, Blood culture, for the isolation of pathogens from a mixture.
Eg: Nutrient broth
Semi solid medium – 0.5% agar.
Eg: Motility medium
Simple media / basal media
– Eg: NB, NA
– NB consists of peptone, meat extract, NaCl,
– NB + 2% agar = Nutrient agar
Complex media
Media other than basal media.
They have added ingredients.
Provide special nutrients
Synthetic or defined media
Media prepared from pure chemical substances and its exact composition is known
Eg: peptone water – 1% peptone + 0.5% NaCl in water
Enriched media
Substances like blood, serum, egg are added to the basal medium.
Used to grow bacteria that are exacting in their nutritional needs.
Eg: Blood agar, Chocolate agar
Enrichment media
Liquid media used to isolate pathogens from a mixed culture.
Media is incorporated with inhibitory substances to suppress the unwanted organism.
Eg:
Selenite F Broth – for the isolation of Salmonella, Shigella
Alkaline Peptone Water – for Vibrio cholerae
Selective media
The inhibitory substance is added to a solid media.
Eg:
Mac Conkey’s medium for gram negative bacteria
TCBS – for V.cholerae
LJ medium – M.tuberculosis
Wilson and Blair medium – S.typhi
Potassium tellurite medium – Diphtheria bacilli
Indicator media
These media contain an indicator which changes its colour when a bacterium grows in them.
Eg:
Blood agar
Mac Conkey’s medium
Christensen’s urease medium
Differential media
A media which has substances incorporated in it enabling it to distinguish between bacteria.
Eg: Mac Conkey’s medium
Peptone
Lactose
Agar
Neutral red
Taurocholate
Distinguish between lactose fermenters & non lactose fermenters.
Lactose fermenters – Pink colonies
Non lactose fermenters – colourless colonies
Sugar media
Media containing any fermentable substance.
Eg: glucose, arabinose, lactose, starch etc.
Media consists of 1% of the sugar in peptone water.
Contain a small tube (Durham’s tube) for the detection of gas by the bacteria.
Transport media
Media used for transporting the samples.
Delicate organisms may not survive the time taken for transporting the specimen without a transport media.
Eg:
Stuart’s medium – non nutrient soft agar gel containing a reducing agent
Buffered glycerol saline – enteric bacilli
Anaerobic media
These media are used to grow anaerobic organisms.
Eg: Robertson’s cooked meat medium, Thioglycolate medium.
BIOCHEMICAL TEST & REACTIONS
They provide additional information for the identification of the bacterium.
The tests include:
Oxidase test
Triple sugar iron agar (TSI)
Indole test
Citrate utilization
Urease test
OXIDASE TEST
Detects the presence of an enzyme “oxidase” produced by certain bacteria which will reduce the dye – tetramethyl-p-phenylene diamine dihydrochloride.
Positive test is indicated by the development of a purple colour.
Oxidase positive – Pseudomonas, Vibrio, Neisseriae
Oxidase negative – Salmonella, Shigella
TRIPLE SUGAR IRON AGAR (TSI)
It is a composite media used to study different properties of a bacterium – sugar fermentation, gas production and H2S production.
In addition to peptone, yeast extract & agar, it contains 3 sugars – Glucose, Lactose, Sucrose.
The Iron salt – Ferric citrate indicates H2S production.
Phenol red is the indicator.
It is an orange red medium with a slant and a butt.
pH of the medium – 7.4
TSI REACTIONS:
Yellow – Acid
Pink – Alkaline
Yellow slant / Yellow butt (A/A) – Lactose fermenters.
Pink slant / Yellow butt (K/A) – Non lactose fermenters.
Pink slant / no colour change (K/K) – Non fermenters
Black colour – H2S production.
Gas bubbles or crack in the medium – gas production.
LF – E.coli, Klebsiella
NLF – Salmonella, Shigella
H2S – Proteus
CITRATE UTILIZATION
Done in Simmon’s Citrate medium.
To detect the ability of certain bacteria to utilize citrate as the sole source of carbon.
Contains Sodium citrate and bromothymol blue as the indicator.
If citrate is utilized, alkali is produced which turns the medium to blue.
Citrate positive – blue colour
Citrate negative – green colour
Positive – Klebsiella
Negative – E.coli
UREASE TEST
Done in Christensen’s urease medium.
This test is used to detect organisms that produce urease.
Urease produced by the organisms split urea into ammonia and CO2.
Urease positive – pink colour
Urease negative – yellow colour
Positive – Proteus, Klebsiella
Negative – E.coli, Salmonella
CULTURE METHODS
Culture methods employed depend on the purpose for which they are intended.
The indications for culture are:
To isolate bacteria in pure cultures.
To demonstrate their properties.
To obtain sufficient growth for the preparation of antigens and for other tests.
For bacteriophage & bacteriocin susceptibility.
To determine sensitivity to antibiotics.
To estimate viable counts.
Maintain stock cultures.
Culture methods include:
Streak culture
Lawn culture
Stroke culture
Stab culture
Pour plate method
Liquid culture
Anaerobic culture methods
STREAK CULTURE
Used for the isolation of bacteria in pure culture from clinical specimens.
Platinum wire or Nichrome wire is used.
One loopful of the specimen is transferred onto the surface of a well dried plate.
Spread over a small area at the periphery.
The inoculum is then distributed thinly over the plate by streaking it with a loop in a series of parallel lines in different segments of the plate.
On incubation, separated colonies are obtained over the last series of streaks.
LAWN CULTURE
Provides a uniform surface growth of the bacterium.
Uses
For bacteriophage typing.
Antibiotic sensitivity testing.
In the preparation of bacterial antigens and vaccines.
Lawn cultures are prepared by flooding the surface of the plate with a liquid suspension of the bacterium.
STROKE CULTURE
Stroke culture is made in tubes containing agar slope / slant.
Uses
Provide a pure growth of bacterium for slide agglutination and other diagnostic tests.
STAB CULTURE
Prepared by puncturing a suitable medium – gelatin or glucose agar with a long, straight, charged wire.
Uses
Demonstration of gelatin liquefaction.
Oxygen requirements of the bacterium under study.
Maintenance of stoke cultures.
POUR PLATE CULTURE
Agar medium is melted (15 ml) and cooled to 45oC.
1 ml of the inoculum is added to the molten agar.
Mix well and pour to a sterile petri dish.
Allow it to set.
Incubate at 37oC, colonies will be distributed throughout the depth of the medium.
Uses
Gives an estimate of the viable bacterial count in a suspension.
For the quantitative urine cultures.
LIQUID CULTURES
Liquid cultures are inoculated by touching with a charged loop or by adding the inoculum with pipettes or syringes.
Uses
Blood culture
Sterility tests
Continuous culture methods
Disadvantage
It does not provide a pure culture from mixed inocula.
ANAEROBIC CULTURE METHODS
Anaerobic bacteria differ in their requirement and sensitivity to oxygen.
Cl.tetani is a strict anaerobe – grows at an oxygen tension < 2 mm Hg.
Methods:
Production of vacuum
Displacement of oxygen with other gases
Chemical method
Biological method
Reduction of medium
Production of vacuum:
Incubate the cultures in a vacuum desiccator.
Displacement of oxygen with other gases
Displacement of oxygen with hydrogen, nitrogen, helium or CO2.
Eg: Candle jar
Chemical method
Alkaline pyrogallol absorbs oxygen.
McIntosh – Fildes’ anaerobic jar
Consists of a metal jar or glass jar with a metal lid which can be clamped air tight.
The lid has 2 tubes – gas inlet and gas outlet
The lid has two terminals – connected to electrical supply.
Under the lid – small grooved porcelain spool, wrapped with a layer of palladinised asbestos.

Working:
Inoculated plates are placed inside the jar and the lid clamped air tight.
The outlet tube is connected to a vacuum pump and the air inside is evacuated.
The outlet tap is then closed and the inlet tube is connected to a hydrogen supply.
After the jar is filled with hydrogen, the electric terminals are connected to a current supply, so that the palladinised asbestos is heated.
Act as a catalyst for the combination of hydrogen with residual oxygen.
Gaspak
Commercially available disposable envelope.
Contains chemicals which generate H2 and CO2 on addition of water.
Cold catalyst – in the envelope
Indicator is used – reduced methylene blue.
Colourless – anaerobically
Blue colour – on exposure to oxygen
Biological method
Absorption of oxygen by incubation with aerobic bacteria, germinating seeds or chopped vegetables.
Reduction of oxygen
By using reducing agents – 1% glucose, 0.1% Thioglycolate
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