The microbe that I highly suspected are listed below:
Bacteria


















Virus
















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Plates used
· Blood plate agar (BAP): Blood Plate agar contains blood from a mammal (usually sheep), and respires to typical transperent nature, typically at a concentration of 5–10%. BAP are an enriched, differential media used to isolate fastidious organisms and detect hemolytic activity. β-hemolytic activity will show complete lysis of red blood cells surrounding colony, while α-hemolysis will only partially lyse hemoglobin and will appear green. γ-hemolysis is the term referring to a lack of hemolytic activity.

· Salmonella-Shigella Agar modified (SS): Beef Extract, Enzymatic Digest of Casein, and Enzymatic Digest of Animal Tissue found in SS plate provide sources of nitrogen, carbon, and vitamins required for organism growth. Lactose is the carbohydrate present in Salmonella Shigella Agar. Bile Salts, Sodium Citrate and Brilliant Green inhibit Gram-positive bacteria, most coliform bacteria, and inhibit swarming Proteus spp., while allowing Salmonella spp. to grow. Sodium Thiosulfate and Ferric Citrate permit detection of hydrogen sulfide by the production of colonies with black centers. Neutral Red is the pH indicator.

· Campylobacter Selective medium: A blood free medium, which will support the growth of enteric Campylobacter species. The selective supplements cefaperazone and amphotericin make the medium selective for Campylobacter jejuni and Campylobacter laridis when incubated at 37°C. Incubation at 42°C is no longer necessary and higher recovery rates have been reported at 37°C. Blood is replaced in the medium with charcoal, ferrous sulphate and sodium pyruvate, which enhance the growth and aerotolerance of Campylobacter species.

· MacConkey Agar (MAC): MacConkey agar is a differential plating medium recommended for use in the isolation and differentiation of lactose-fermenting organisms from nonfermenting Gram-negative enteric bacteria. It is selective by the presence of specific inhibitors.

· Thiosulphate-citrate bile sucrose agar (TCBS): A selective isolation medium for pathogenic Vibrio species. Most Enterobacteriaceae other than Vibrio species are suppressed for at least 24h. Bile salts inhibit Gram-positive organisms. Sodium thiosulphate serves as a source of sulphur, which, in combination with ferric citrate, detects hydrogen sulphide production. When sucrose is fermented it produces acid changing the pH. This is indicated by bromothymol blue and thymol blue. The medium is also alkaline which enhances the recovery of Vibrio cholerae.

· Selenite broth: Selenite Broth contains enzymatic digest of casein and enzymatic digest of animal tissue that provides nitrogen and vitamin sources. The main carbohydrate that is present in the broth is lactose. Lactose is the fermentable carbohydrate. Sodium Phosphate in the broth acts as a buffer. A rise in pH decreases selective activity of Selenite. The acid produced by lactose fermentation helps to maintain a neutral pH. Sodium Selenite inhibits the growth of Gram-positive bacteria and many Gram-negative bacteria.

· Alkaline peptone water (APW): Alkaline Peptone Water is an enrichment medium used for the cultivation of Vibrio species from feces and other infected materials. Clinical materials containing small numbers of Vibrio should be inoculated into an enrichment medium prior to plating onto a selective medium, such as TCBS Agar. Alkaline Peptone Water is a suitable enrichment broth for this purpose. The relatively high pH of the medium (approximately 8.4) provides a favorable environment for the growth of vibrios.

· Cefsulodin-Irgasan-Novobiocin (CIN) agar for Yersinia enterocolitica: Cefsulodin-Irgasan-Novobiocin (CIN) agar is a differential and selective medium for the isolation of Yersinia enterocolitica. Fermentation of mannitol in the presence of neutral red produces characteristic "bull's-eye" colonies. These are colourless with a red centre. A zone of precipitated bile may also be present. Crystal violet, sodium desoxycholate, cefsulodin, Irgasan (triclosan) and novobiocin are inhibitory agents.

· Sorbitol MAC for enterohaemorrhagic E.coli: Sorbitol MacConkey Agar medium contains sorbitol instead of lactose and it is recommended for the detection of enteropathogenic strains of E. coli, which ferments lactose, but does not ferment sorbitol and hence produce colorless colonies. Sorbitol fermenting strains of E. coli 0157:H7 produce pink-red colonies. The red colour is due to production of acid from sorbitol, absorption of neutral red and a subsequent colour change of the dye when pH of the medium falls below 6.8.

· Loeffler’s methylene blue: This solution is a solution that can be used alone as a simple stain, positive stain or as the counterstain in the acid fast stain procedure (Ziehl-Nielsen). It can also be used in the staining procedure that detects metachromatic granules (volutin).

· Enterococcosel Agar with 6 microgram / ml of vancomycin (VRE): Enterococcosel Agar Enterococcosel Agar incorporates Bile Esculin Azide Agar to yield rapid, selective detection and enumeration of enterococci. The surveillance for Vancomycin-Resistant Enterococci (VRE) can be accomplished by plating stool cultures onto Enterococcosel Agar with Vancomycin (6 µg/mL).


Microscopy
A wet preparation is examined for the presence of leucocytes and erythrocytes. Their presence may indicate invasive disease. This test is only done on request as its usefulness is limited.

1. Place a drop of liquid faeces or saline suspension of the faecal specimen on a microscope slide. Any mucous or flecks of pus or blood that may be present should be included in the suspension as these are likely to harbour disease causing organisms
2. Mix 1 drop of Loeffler’s methylene blue stain with the faeces specimen. Note that there must be an equal volume of faeces to stain.
3. Place a cover slip over it.
4. Wait 2 – 3 minutes for the nuclei to stain and then read the preparation under high power on (40x)
5. Observe for predominating numbers of white blood cells (WBCs), which indicate an invasive pathogen.

Culture
Inoculated places are O2 incubated unless otherwise indicated in the table.

All faeces are inoculated onto the following media: BAP, MAC, SS, Selenite broth and Campylobacter selective agar for the isolation of salmonella, Shigella and Campylobacter spp.

In addition to the above:
· Bloody faeces are also inoculated onto Sorbitol MAC plate.
· Watery faeces are plated on TCBS and inoculated onto APW to culture for Vibrio spp.


After culturing, gram staining is performed to differentiate gram positive (purple) and negative (pink). Gram negative bacteria includes gram negative cocci, bacilli and coccibacilli while positive includes gram positive cocci and bacilli.


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