Lecture 15 - Gram negative rods (Synchronous)

Introduction

Aston University, located in Birmingham, UK, emphasizes the important study of Gram-negative facultative anaerobic rods, particularly focusing on the pathogenicity mechanisms of foodborne pathogens.

Aims and Objectives

  • Understanding Testing Importance: The curriculum emphasizes why testing foodborne pathogens is crucial for safety and health.

  • Pathogenic Mechanisms: Students will learn about the mechanisms through which pathogens like Salmonella and E. coli cause disease.

  • Characteristics Differentiation: The objectives include distinguishing characteristics and diagnostic tests specific to Salmonella typhimurium and E. coli O157:H7.

Enterobacteriaceae Family Overview

  • Species Count: The Enterobacteriaceae family consists of over 100 species.

  • Gram Negative Nature: These organisms are categorized as Gram-negative and are frequently implicated in cases of foodborne illness (Alnajar and Gupta, 2017).

Transmission Routes of Foodborne Illness

Foodborne illnesses can arise from multiple transmission routes:

  • Contamination During Preparation: Cross-contamination in food handling or preparation stages.

  • Cooking Contamination: Failure to adequately cook food can lead to illness.

  • Animal Contact: Interacting with infected pets and livestock can harbor pathogens.

  • Pre-existing Contamination: Foods that are already contaminated, such as eggs, pose a risk.

Prevalence of Foodborne Illness

  • UK Cases: The Food Standards Agency (FSA) estimates approximately 2.4 million cases annually in the UK.

  • Common Pathogens: The NHS indicates that Salmonella enterica Typhimurium is the most prevalent serotype, while E. coli O157:H7 is significant due to its potential fatality.

Serotypes and Classification

Understanding serotypes is essential for identification and diagnosis:

  • Kauffman–White Classification:

    • Flagellar (H) Antigens: Heat-labile proteins, with classifications of phase 1 and phase 2.

    • Somatic (O) Antigens: Heat-stable and alcohol resistant, involved in the lipopolysaccharide membrane structure.

Focus on Pathogenic Organisms

Salmonella enterica Typhimurium

  • Sources: Commonly found in both domestic and wild animals, particularly chickens.

  • Symptoms and Incubation: Exhibits symptoms such as diarrhea, fever, and abdominal cramps within 6 hours to 6 days of infection.

Escherichia coli Characteristics

  • Strain Context: Most E. coli strains are harmless, residing in the lower gastrointestinal tracts of warm-blooded animals.

  • Symptoms and Incubation: Symptoms include diarrhea (sometimes bloody), nausea, abdominal cramps, and vomiting, with an incubation period of 2-5 days.

Salmonella Pathogenesis Overview

Successful Salmonella pathogens exhibit a three-step process:

  1. Colonization: They penetrate epithelial cells.

  2. Evasion: They hijack phagocytes to avoid immune detection.

  3. Damage: They produce proteins leading to gastroenteritis by causing inflammation.

Salmonella Pathogenicity Islands (SPIs)

  • SPI Structure: There are five significant SPIs, notably SPI-1 and SPI-2, which encode proteins for the type III secretion system (T3SS). This system facilitates direct protein transfer from Salmonella to host cells.

  • SPI Function:

    • SPI1 facilitates adherence to intestinal cells.

    • SPI2 allows survival within phagocytes.

    • SPI3, SPI4, and SPI5 further regulate adherence and host cell dynamics.

Pathogenesis Processes

This pathway details how S. typhimurium progresses from adhesion to replication:

  1. Adhesion to epithelial cells occurs with the help of SPI4.

  2. Host cell signaling activates all 5 major SPI genes, prompting cytoskeletal rearrangements.

  3. S. typhimurium can be internalized into dendritic cells, facilitating its replication within specialized compartments (SCVs).

  4. This process includes mechanisms that prevent apoptosis of epithelial cells, ensuring pathogen survival.

Escherichia coli Pathogenesis

The Shiga toxin-producing E. coli (STEC), particularly E. coli O157:H7, poses significant clinical risks:

  • Colonization and Damage: E. coli O157:H7 binds to intestinal epithelial cells, activating pathways that suppress toxin removal and leading to severe conditions such as hemorrhagic colitis and hemolytic uremic syndrome.

Diagnosis in the Enteric Laboratory

The diagnostic procedures focus on:

  • Sample Types: Fecal samples processed in a Class II Biological Safety Cabinet.

  • Gram Stain Results: Typically reveal Gram-negative rod arrangements for Enterobacteriaceae.

  • Catalase and Oxidase Tests: Indicate catalase-positive and oxidase-negative results, essential for differentiation.

Media Utilization in Enteric Lab

Blood vs Chocolate Agar

  • Composition Differences:

    • Blood agar retains red blood cells for detecting hemolytic activity.

    • Chocolate agar lyses blood cells to release growth factors.

  • Hemolysis Types: Alpha (partial), beta (complete), and gamma (none) hemolysis classifications.

Species Identification

  • Salmonella vs E. coli on Media: Their appearance on blood agar and MacConkey agar exhibits differences in size and hemolytic properties, turquoise and pink coloration indicating lactose fermentation.

Conclusion

In summary, this session explored crucial aspects of Gram-negative foodborne pathogens. Further exploration into Gram-positive foodborne illnesses will follow in advanced studies.