Discussion Bacteria from the provided master stock plate were used for the gram stain test. Bacteria were then streak and grown from the master stock plate onto a working plate. After an incubation time of 24 hours at 37°C, the working plate bacteria were then used to perform the catalase and red blood cell hemolysis tests. After conducting the three tests, it was concluded that unknown #5 was Streptococcus agalactiae. Gram Stain Test The Gram Stain test was first performed to differentiate the bacteria based on the thickness of the peptidoglycan layer in its cell wall.
Thus, there may be low amount of amylase in saliva which cannot break down the starch completely which give dark-purple colour . In test tube C , the iodine solution change from brown to dark blue which is different from the expected . It is because the amylase is denatured at 75℃ that the the activity of amylase is low or even stop. Therefore, the starch is not broken down into maltose by amylase. In the test D, a dark-brown solution is seen in the test tube after adding the iodine as the pH of the 1ml 0.5M HCl is not an optimum pH for the activity of amylase that the starch is broken down into maltose .
Remove excess water from the surface of the gram stained glass slide and observed under 1000X oil immersion microscope. Gram-staining have performed for Staphylococcus aureus (control); Enterococcus faecalis (control); Nostril microflora in NA, MSA, and PYCa. Gram-staining provides results of the bacteria morphology, type of gram-stain. Catalase test was also done prior to gram-staining. MicrobactTM Biochemical Identification Kit was used for identifying gram-negative aerobic and facultatively anaerobic bacteria.
The tube was placed back in incubation for 96 more hours to observe any more positives. 2.10 Catalase Test A trypticase soy agar plate was used and after incubation, four drops of 3% Hydrogen Peroxide was added to the plate to flow over the bacterial growth. A presence of bubbling was observed. 2.11 Starch Hydrolysis A starch agar plate was inoculated with a streak of the unknown bacteria and then incubated. On the second day of incubation, the plate was removed from the incubator and placed over a hot plate heating Iodine solids.
al. 2014) The slide containing Micrococcus luteus produced bubbles when 3% hydrogen peroxide was dropped on it. (see Figure 9.3) This indicates that Micrococcus luteus produces catalase. Vigourous bubbling occurred when he slide containing the unknown microbe was dropped with 3% hydrogen peroxide. (See Figure 9.4) This indicates that the unknown microbe can produce catalase.
al.. Cyclodextrin glucosyltransferase (CGTase) producing alkalophilic bacteria were isolated from the water samples collected from Marneri pond of Tirunelveli, Tamil Nadu by serial dilution and plating method. Totally 22 bacteria were isolated from the collected water samples and screened for CGTase activity by Horikoshis medium II agar plate method. Among 22 bacterial isolates, 15 isolates showed CGTase activity and better zone producing strain was selected for further studies. The potential strain was identified as Bacillus circulans by 16S rDNA gene sequencing experiment. The best enzyme activity was observed at pH 10.5, 32°C, supplemented with cassava as carbon source an d beef extract as nitrogen source.
Gelatine hydrolysis test The tubes containing sterilized gelatine liquefaction medium was stab inoculated with fresh culture of bacterial isolates and incubated at 37±1o C for 24 h. After incubation the tubes were placed in refrigerator at 40 c for 30 min. Then these tubes were kept in ice cubes containing beaker and observed for Gelatine liquefaction by the bacterial isolates. The gelatine liquefied tubes were considered as positive reaction (Aneja, 2001). Composition of Gelatine liquefaction medium Peptone 5g Beef extract 3g Sodium chloride 5g Gelatine 120g Agar 20g Distilled water 1000ml PH 7 IV. Casein hydrolysis test The sterilized plates containing skim milk agar medium were point inoculated with fresh bacterial cultures and incubated
The procedure of staining will distinguishes organisms of the domain bacteria according to structure of cell wall. A thick peptidoglycan layer and stain blue to purple indicate as the Gram-positive cells while a thin peptidoglycan layer and stain red to pink indicate as Gram-negative cells. In bacteriology, the most frequently used staining procedure is the Gram stain, is a complex and differential staining procedure. Organisms in the Domain Bacteria are differentiated according to cell wall composition over and done with a series of staining and decolorization steps. Cell walls of Gram-positive bacteria contain thick layers of peptidoglycan which 90% of cell wall and this stain are purple.
The sterile medium was inoculated with known volume of inoculum and incubated at different temperatures (25, 30 and 350C) for different periods (48,72 and 96h) of fermentation. After fermentation the mouldy substrate was mixed with distilled water (1:4 w/v), agitated at through cheese cloth followed by centrifugation at 20,000 rpm for 20 minutes. The clear supernatant was used as crude enzyme. Assay of glucoamylase The assay of glucoamylase was carried out according to the method of Shazia Malik, Tehreema Iftikhar and Ikram Ul Haq13. One unit (U) of glucoamylase is defined as the amount that liberates 1 µmol of reducing sugar as glucose/ml/min under the assay condition.
The petri dish was incubated for 24Hrs at 35oC. The petri dish was observed to see if there was any bacterial growth and once the whole dish was full of bacterial colonies, it was observed under an UV lamp to see if there was fluorescence or not. I decided not to do the IMViC tests because the first two tests concluded on what the unknown bacteria