Ebac 1

• Family Enterobacteriaceae
• General Characteristics

  • Often referred as ENTERICS
  • Consists of a large number of diverse organisms (Frequently encountered in the lab)
  • Normal flora of the GI tract of animals and humans
  • Gram negative bacilli or coccobacilli
  • Associated in infections almost throughout the body
  • Ubiquitous
• All Enterobacteriaceae

  • Facultative anaerobe, non-spore-former
  • Glucose fermenters
  • Cytochrome oxidase negative except: Plesiomonas
  • Reduce nitrates to nitrites except: Erwinia, Pantoea agglomerans, Photorhabdus, and Xenorhabdus
  • Motile except: Klebsiella, Shigella, Yersinia
  • Motile members have peritrichous flagella except: Tatumella ptyseos which has polar flagella
  • All grow luxuriantly on CHOC and BAP
  • Klebsiella and sometimes Enterobacter produce mucoid colonies
  • Beta-hemolysis is seen in some strains
  • All are catalase positive except for one group of Shigella species and Xenorhabdus
• Serotyping (Serogrouping) of Enterobacteriaceae


  Antigenic Structure Used For Identification Through Serotyping

  
• Serogroups of Escherichia coli

  Based On Its O And H Antigens

  
• 2 Broad Categories of Enterobacteriaceae

  • Opportunistic Pathogens
• Selective and Differential Media

  • Isolation of enteric bacteria from clinical specimen is primarily done with the use of selective and differential media
  • Selective: have inhibitors to suppress other bacteria particularly gram positive
  • Differential: provide (?) for organism to exhibit their unique biochem characteristics that can be use in differentiation

  
• Primary culture media used in the presumptive identification of enteric organisms in the laboratory

  • EMB, MAC, XLD, HEA, SSA

  
• Selective and Differential media is just a presumptive identification of enteric bacteria

  
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  Eosin–methylene blue (EMB) agar is a selective differential medium useful in the isolation and identification of gram-negative enteric bacteria

  
• MacConkey Agar (MAC)


  MacConkey agar (MAC) is a selective, differential, primary plating medium. It selects for Enterobacteriaceae and other gram-negative rods in the presence of mixed microbiota and differentiates them into lactose fermenters and non–lactose fermenters
• Xylose Lysine Deoxychoalte Agar (XLD)


  Xylose-lysine-desoxycholate (XLD) agar is a selective, differential, primary plating medium used to isolate Salmonella and Shigella spp. from stool and other specimens containing mixed biota

  
• Salmonelle-Shigella Agar (SS)


  Salmonella-Shigella (SS) agar is used to select for Salmonella and some strains of Shigella from stool specimens
• Identification of Salmonella and Shigella becomes more difficult
• Shigella dysenteriae and Shigella flexneri can occasionally be inhibited on XLD agar
• Some strains of Salmonella may fail to produce H2S and therefore resemble Shigella colonies
• On Salmonelle-Shigella Agar, blackening is more likely to occur when alkaline conditions exist
• Salmonella-Shigella (SS) agar

  Used to select for Salmonella and some strains of Shigella from stool specimens
• Salmonella-Shigella (SS) agar

  • Contains bile salts, sodium citrate, and brilliant green, which inhibit the growth of gram-positive and many lactose-fermenting, gram-negative rods normally found in stool
  • Lactose is the sole carbohydrate source in the medium, and neutral red is the pH indicator
  • Sodium thiosulfate is added as a source of sulfur for the production of H2S
• If an organism grows on SS agar and ferments lactose, it will produce acid and change the indicator to pink-red
• If H2S is produced on SS agar, it reacts with the ferric ammonium citrate present in the medium, forming a black precipitate in the center of the colony
• Shigella colonies on SS agar

  Appear colorless because these organisms do not ferment lactose or produce hydrogen sulfide
• Salmonella colonies on SS agar

  Are colorless, with a black center, because these organisms usually make H2S but do not ferment lactose
• Pink to red colonies on SS agar

  Indicate that the organism ferments lactose
• If Proteus grows on SS agar, swarming is inhibited
• Hektoen enteric (HE) agar


  A selective differential medium used for direct isolation of enteric pathogens from feces and for indirect isolation from selective enrichment broth
• Hektoen enteric (HE) agar

  • Contains bile salts at concentrations that not only inhibit the growth of gram-positive bacteria, but also inhibit the growth of many gram-negative organisms that are part of the normal intestinal microbiota
  • Contains lactose, salicin, and sucrose to determine fermentation patterns, detected by the pH indicator bromothymol blue
  • Contains ferric salts (sodium thiosulfate, ferric ammonium citrate) to detect the production of hydrogen sulfide gas
• HE agar should not be autoclaved, and overheating should be avoided
• Most nonpathogens ferment one or both of the sugars on HE agar, and colonies appear bright orange to salmon pink because of the low pH interaction with incorporated dyes
• Nonfermenters, such as Salmonella and Shigella spp., on HE agar

  Typically produce green to blue-green colonies
• Hydrogen sulfide gas production on HE agar is seen as a black precipitate that accumulates within colonies
• Serotyping


  A serologic test that determines species/subtypes of bacteria based on the antigens present on its surface

  
• Antigens


  Substances (proteins) that react with specific antibodies
• Antibodies (anti-sera)


  Proteins that react to antigens
• Antigenic variations


  Refers to the variations in the antigenic structures of antigens
• Salmonella/Escherichia/Shigella species may look the same under the microscope, but they can be separated into different many serotypes according to the variations of the antigens found on their surface

  
• Serotyping

  Determined by the agglutination of bacteria with specific antisera to identify variants of O and H antigens

  
• H antigen

  Flagellar antigen, a protein in nature that is heat labile
• O antigen


  Somatic antigen, heat stable, located in the cell wall, part of the lipopolysaccharide

  
• K antigen


  Capsular antigen/envelope antigen, a heat labile polysaccharide found in encapsulated strains that covers the O antigen