Enterobacteriaceae #2

Enterobacteriaceae Family Overview

  • Large Family of Bacteria
    • Currently recognized: 100 species.
    • Characteristics: Opportunistic and true pathogens.
    • Acquisition routes:
    • Environmental
    • Food and water
    • Fecal–oral route
    • Colonization
    • Healthcare-associated infections
    • Endogenous infections

Classification of Enterobacteriaceae

  • Classification based on:
    • Phenotypical characteristics
    • Biochemical reactions
    • Molecular & DNA sequencing
    • Notable change in naming conventions over the last 5-10 years due to improved understanding of genetic relationships.

Family Characteristics

  • Gram-negative bacilli & coccobacilli.
  • Fermentation Properties:
    • Ferment glucose.
    • Cytochrome oxidase negative (except Plesiomonas).
  • Most species reduce nitrates to nitrites.
  • Facultative anaerobes
  • Growth on media:
    • Grows well on blood agar (non-hemolytic) producing large, dull grey colonies.
    • Isolation techniques involve MacConkey, EMB, or HE media.
  • Motility:
    • Majority are motile (notable exceptions include Klebsiella, Shigella, and Yersinia).
  • Defined as true pathogens: always pathogenic in a host.

Serological Characteristics of Enterobacteriaceae

  • Somatic antigen (O):
    • Composed of LPS or endotoxin.
    • Heat stable; used to identify serogroups.
  • Envelope antigen (K):
    • Composed of a capsule.
    • Heat labile.
  • Flagellar antigen (H):
    • Located at the flagellum.
    • Defines a serotype when combined with O antigen; some are more immunogenic than others.

Isolation Techniques and Media Selection

  • Isolation Media:

    • Generally, blood agar with MacConkey or EMB for all specimens other than stools.
    • Initial observations of significant characteristics:
    • Swarming motility observed in Proteus.
    • Mucoid morphology in Klebsiella pneumoniae due to polysaccharide capsule.
    • Phenotypical Indicators:
    • Agar greening in presence of H2S production.

  • Specific media selections:

    • Differential and selective plates for specific Enterobacteriaceae species include:
    • Sorbitol MacConkey (SMAC) for verotoxic E. coli.
    • CIN agar for Yersinia enterocolitica.
    • XLD or HE agar for identifying Shigella and Salmonella.
    • Campy BAP for Campylobacter jejuni.
    • Blood agar and differential plates for urine cultures.

  • Colony count analysis is necessary, with specific focus on isolation success.

Identification and Testing

Gram Negative Identification Steps

  • For Gram-negative bacilli that grow on MAC or EMB and are oxidase negative, conduct the following tests to confirm Enterobacteriaceae:
    • Carbohydrate Fermentation Tests:
    • Test for lactose, sucrose, sorbitol, mannitol, xylose, and mannose.
    • Gas Production:
    • Assess for accumulation of hydrogen (H) + carbon dioxide (CO2).
    • Hydrogen Sulfide (H2S) Production:
    • IMViC Reactions:
    • Indole: Tests for tryptophanase enzyme; red color with Kovach’s reagent.
      • Methyl Red (MR): Tests for mixed acid fermentation; acidic pH indicates a positive MR reaction.
      • Voges-Proskauer (VP): Looks for acetoin; pink color with exposure to KOH and alpha-naphthol.
      • Citrate Test: Evaluates if citrate is utilized as the only carbon source; positive reaction involves growth and/or blue color in the presence of bromothymol blue.

Example of IMViC Testing

  • IMViC for Lactose-fermenting Enterobacteriaceae:
    • Lactose fermenters show distinct patterns across various tests, essential for phenotypic identification.

Further Biochemical Trials

Decarboxylase Testing

  • Focused on basic amino acids.
    • Lysine (LDC) → cadaverine.
    • Ornithine (ODC) → putrescine.
    • Arginine (ADH) → citrulline.

Urease Testing

  • Identifies enzymatic breakdown of urea; critical in differentiating Proteus and Providencia genera.

H2S Production Testing

  • Detected in TSI, LIA, and Hektoen Enteric media.
  • Identifies presence of sulfur sources; differentiative between Salmonella and Shigella based on H2S results.

Pathogens of Clinical Significance within Enterobacteriaceae

E. coli: Types and Infections

  • Uropathogenic E. coli (UPEC)
  • Neonatal Meningitis E. coli (NMEC)
  • Diarrheagenic E. coli: Further classified into 5 major syndromes based on symptoms, virulence factors, and serotypes:
    • ETEC: Enterotoxigenic
    • EPEC: Enteropathogenic
    • EIEC: Enteroinvasive
    • EHEC: Enterohemorrhagic (verocytotoxic E. coli)
    • EAEC: Enteroaggregative

Other Pathogenic Genera

  • Klebsiella: Known for hospital-acquired infections; requires extensive identification to isolate types.
  • Enterobacter: Related to urinary tract and respiratory infections, particularly in immunocompromised patients.
  • Serratia: Pathogenic in specific contexts, notably S. marcescens related to HAIs.

Salmonella: Characteristics and Clinical Significance

  • Salmonella spp. defined by unique serological interactions and biochemical testing.
  • Common human pathogens fall under:
    • S. Typhi: Typhoid fever.
    • S. Enteritidis: linked to foodborne illness.
  • Isolation and identification often achieved through selective media such as Hektoen Enteric and XLD agar.

Yersinia: Pathogenic Strain Significance

  • Recognized for causing diseases via specific species:
    • Y. pestis: Plague
    • Y. enterocolitica: Enterocolitis

Detection Techniques

  • PCR bases detection for antibiotic resistance genes, especially prevalent in clinical isolates.

Summary

  • The Enterobacteriaceae family covers a vast array of pathogens crucial in medical bacteriology. Ongoing advancements in genetic study have altered traditional classifications and identification strategies, informing both clinical and laboratory practices in pathogen detection and disease management.