**4.6.3 Searching for** *Salmonella* **bacteria**

Same amount of water is added into tubes including selenite f medium which was prepared with one portion concentrate. After waiting at 37°C for 24 hours, planting is performed from every tube to SS agar. Furthermore, one cm3 each from tubes are taken and mixed with bismuth sulphite agar and poured into petri plates. When reproduction occurs, it is searched whether it is from *Salmonella* group. (551,602,611)

#### **4.7 Meat**

136 Salmonella – A Dangerous Foodborne Pathogen

Although "normal saline" (0.85% NaCl; Merck 1.06404) has been used as diluting solution for general purpose, "Maximum Recovery Diluent (Merck 1.12535) which has the formula of Normal Saline (0,85%) + Peptone (0,1%) in accordance with ISO 6887 is used recently. This solution is also referred as "peptone-saline". Furthermore, 0.1% peptone solution (Merck 1.07214) and "Buffered Peptone Water" (Merck 1.07228) which is used in *Salmonella* analysis

To drink and to use waters contaminated with stool or sewage leakages is very important because they may cause infections such as typhoid, dysentery, cholera. In contaminated water, typhoid germ may exist together with intestinal bacteria. However, number of these bacteria is very less in general and it is impossible to obtain. On the other hand, commensal bacteria such as coliform bacteria, Streptococcus faecalis and Clostridium perfringens (welchii) are always obtained in contaminated water easily. Such water containing these bacteria means that the water was contaminated with the stool an this may contain typhoid germ. Coliform bacteria shows the contamination with the stool with the safest manner. The most important of them is Escherichia coli which is main commensal bacteria of the intestine. Existence of spore forming anaerobe bacteria in the absence of other bacteria

Colifrom bacteria also exists in water contaminated with stool of various animals. But they are less. More bacteria are present in the water contaminated with sewage water. It is important to

Detection of alive bacteria count gives information about quantity and type of organic substances. The trial is performed both at 37°C and 22°C. Bacteria which mix from human and animal origin organic substances reproduce at 37°C in particular. Those which reproduce in lower temperature are saprophyte bacteria that mix from the soil and plants or

The water to be examined should be taken into colorless, preferably glass cap bottles with a volume of 250 cm3 . Orifice and cap of the bottle are wrapped with separate papers and sterilized in the autoclave. If water will be taken from running water, the orifice of the tap is burned with spirit flame and the water is put into the bottle after leaving the water run for five minutes. The cap of the bottle is closed by caring the sterility conditions. To take water from the creek or river, the bottle is hold from the bottom, it is immersed into the water upside down to 30 cm deep. The orifice of the bottle is turned to the flow direction and water is filled with water without touching. To take water from dead water, lakes and depots, the bottle that a ballast was hung to the bottom and bounded with ropes from the neck and the cap is immersed into the water with a desired depth. The rope is pulled and the cap is opened, after the bottle is filled with water, it is pulled to up and the cap is closed. If a period more than 3 hours will pass from taking the water sample until the examination,

If sample will be taken from chlorinated water, chlorine should be neutralized immediately. For this, one sodium thiosulphate crystal is put into the bottle or 0,2 cm3 from 1 g of crystallized sodium thiosulphate solution which was dissolved in 100 cm3 sterile water

To mix the water well before the assay, the bottle that water sample is taken is shaken.

detect the bacteria count in the water to determine the level of the contamination.

the bottle should be kept in ice. It may be sent to far places only in ice.

**4.5 Homogenization and dilution solutions** 

are used for dilution in accordance with ISO 6887.

shows an old contamination with the stool.

exist in the water normally.

before sterilization.

**4.6.1 Taking the water sample** 

**4.6 Water** 

Meat may include many germ types such as bacillus suptilis, Escherichia, proteus, *Salmonella* , staphylococcus species and fungus and especially anerobe spore forming bacteria show fundamental change. These bacteria cause formation of bad odor by making putrefaction in proteins. It is not very possible to decide on the status of the meat by bacteria on it.

To detect bacteria, one gram is weighed, it is meshed in mortar with sand. 1000, 10.000, 100.000 dilutions are prepared. Colony count is performed with these in the petri plate. Diagnose of bacteria is performed if required.

#### **4.8 Egg**

Fresh egg is not always sterile. Gran (+) coccus, gram (-) bacillus and some fungus species may be present in the egg. Sometimes, egg may include *Salmonella* bacteria. (617, 618)

The shell of the egg has a porous structure. Gases and microscopic particles may pass through these pores. Bacteria always exist on the shell of the egg. Escherichia coli is present almost on all shells. Humidity causes bacteria to pass through pores in humid and dirty eggs. Therefore, eggs which sill be stored for a while must not be washed.

The egg which will be examined is cleaned well by washing with brush, water and soap. It is kept in 0,1% sublimated solution for 30 minutes. It is taken from here by a sterile spoon and put into 200 cm3 of sterile water. It is kept in the water for 10 minutes and put into 95% alcohol and left for 5 minutes. The egg is taken from the alcohol and left for drying and

Isolation and Identification of Salmonellas from Different Samples 139

For identification of various samples, methods which alternate and support each other.

Culture method in identification of *Salmonellas* is conducted with pre-enrichment and

Identification studies are same regardless from the source of the culture. Variety of mediums used in the culture may depend on characteristics of the sample examined. Especially when number of *Salmonellas* are less and other organisms are more, very careful study is required. If extra clinical samples are processed such as heating, drying and radiation or they are frozen or kept for a long time or pH level is low although clinical samples are examined as fresh, nonselective pre-enrichment culture is applied. Because these processes weakened *Salmonellas* and made them semi-selective. The purpose is to provide this kind of bacteria to their normal reproduction period before contacting inhibitor substances. Because selective substances may make a toxic effect for "weakened" *Salmonellas* . While enrichment bouillon culture facilitates reproduction of *Salmonellas* , it also provides inhibiting or decreasing effect for reproduction of other organisms. Accompaniment organisms mainly include coliforms, proteus species and pseudomonas. As the proportion of these organisms is more than *Salmonellas* in particular, selective enrichment process gains importance. However, there are differences between *Salmonella* types in terms of inhibitor substance sensitivity. Therefore, it is impossible to say which selective enrichment bouillon is the most suitable definitely for today. Selective agar mediums generally include inhibitor substances and an inhibitor system. Indicator system either changes the color of colonies or the color of agar area around the colony changes. Thus,

Brillant green agar Brillantgreen Sulphadiazine agar Brillant Green Mac Conkey agar Desoksicholate Citrate agar *Salmonella* -Shigella Agar,(SS) Bismuth Sulphite agar EMB

Samples are taken into non-selective enrichment medium (lactone bouillon) according to their clinical or extra clinical sample characteristics and incubates at 35-37 °C for 24 to 48 hours, then 1 ml from them is taken and taken into selective enrichment medium and

1. Clinical and other samples are taken into lactose bouillon with appropriate amounts. If the sample food is also solid, it is mixed in the blender. Ot is transferred into 500 ml of

(Selenite F, tetrathionate bouillon) A and incubated at 35-37 °C for 24 hours.

2. It is incubated at 35-37°C for 48 hours and passed to selective enrichment.

**5. Culture and identification methods of** *Salmonellas* 

Full automatic bacteria identification device

it helps to identify suspicious *Salmonella* colonies.

The following Agar Mediums are used in various countries.

These are:

Invic test

Culture methods

 Triple tube method Api method

 Serological tests Grubul Widel

**5.1 Culture methods** 

AGAR ENDO AGAR

**5.1.1 Non selective enrichment** 

erlenmayer flask or flasks.

selective medium planting.

broken from the wide edge by a sterile holder. The content is transferred into a sterile beaded jar. Egg white and yolk are mixed by shaking well. Microorganisms are searched by planting into various mediums.
