Gi Tract

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Colonization of the GI tract

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Normal Commensal Flora of the GI Tract
- Mouth: 10^10 bacteria (>500 species)

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- Streptococcus, Neisseria, Actinomyces, Veillonella &
Lactobacillus, some yeasts, transient viruses
- Eruption of 1st teeth: Porphyromonas, Prevotella &
Fusobacterium
- Growth of Teeth: S. sanguis*, S. mutans* & S. salivarius *Dental plaque

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Normal Commensal Flora of the GI Tract
Stomach: Sterile (0 bacteria/ml) or 103 bacteria/ml

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– Streptococcus, Staphylococcus, Lactobacillus &
Peptostreptococcus
- Helicobacter pylori: Gastritis; Peptic/duodenal ulcers
N.B. 105-107 bacteria/ml: - abnormality (achlorhydria or malabsorption syndrome)

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Normal Commensal Flora of the GI Tract
- Small Intestine: Sparse
- Duodenum: 0-104.5 bacteria/ml

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- (increases distance away from stomach)
Duodenum: Fluctuating transients: aerobic Streptococci, Staphylococci, Lactobacilli, yeasts
N.B Complete absence of coliforms & Bacteroides

Front 5

Normal Commensal Flora of the GI Tract
- Jejunum-Ileum: 0-105/ml - 107 bacteria/ml
- AT Ileocecal junction 106 - 108 organisms/ml
- Large Intestine: Colon 1010 - 1012 bacteria/ml
(richest & complex >400 species identified)

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– High counts Enterobacteriaceae – some Streptococcus, Staphylococcus, Lactobacillus, Bacteroides, Bifidobacterium, Clostridium
Large gut: 95-99% Anaerobic: Bacteroides, Bifidobacterium, Eubacterium, Peptostreptococcus & Clostridium Plus Enterobacteriacea

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Factors Affecting Microbial Composition
ALLOGENIC: originate outside ecosystem
- E.g., “Western” diet vs Native, vegetarian diet (Africa) Inc Bacteroides, Dec enterococci (other aerobes)

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- Diet: nature of meal (gastric emptying); milk
AGE, GEOGRAPHIC REGION
ANTIBIOTIC THERAPY: removal of normal flora (colonization by pathogenic org’s E.g., C. difficile)
Surgery: Alters bacterial pop. - Ileostomy effluents - unique ecological niche NOT corresponding ileum or colon

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Factors Affecting Microbial Composition
AUTOGENIC: arise within ecosystem
- Environment: opt growth 37oC, anaerobic

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H+ concentration (gastric acid) Peristalsis (directory flow rate), Shedding of epithelium, Mucus, Conjugated Bile salts? Immunological response (IgA)?

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ii. Activities of Microorganisms:

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Nutritional competition, Prodn bacterial inhibitors: bacteriocins, antibiotics Toxic metabolic end products
H2S prodn, Competition for attachment sites
Maintenance of low oxidation-reduction potentials

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I. Upper GI Infections: Oral Disease (Herpes & Mumps
II. Lower GI Tract Infections - (Food/Water)

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I. – Dental Caries – Periodontal disease – Abscess – Systemic Infection
II. – Changes in human demographics – Changes in food preferences – Changes in food production – Changes in food distribution – Microbial adaptation

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Food Hazards
- U.S figures ONLY include Botulism, E. coli O157:H7(STEC), Cholera, Salmonellosis, Typhoid fever, Shigellosis, Listeriosis (from 2002) & Vibriosis (from 2007)

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1. Microbial contamination
2. Naturally occurring toxicants
3. Environmental contaminants (e.g., metals)
4. Nutritional problems (i.e., malnutrition, undernutrition) 5. Pesticide residues
6. Food additives

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Major causes of "gastroenteritis"
- Norovirus (49%)
- Bacteria (40%)

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Front 12

Route of infection: Entery, disease, Exit
- Incubation period

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Front 13

FOOD-POISONING: TOXEMIA
FOOD-ASSOCIATED INFECTIONS (food-borne)

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consumption of food containing toxins (chemical or microbial) C. botulinum, S. aureus, B. cereus (1 form)
Fungal & Marine toxins
consumption of food containing organism (acts as vehicle for entry) A wide variety of pathogens

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Enteritis
“Gastro” enteritis
Colitis
Enterocolitis
Dysentery
Diarrhoea

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inflammation of intestinal mucosa
inflammation of stomach & intestinal linings
inflammation of large intestine
inflammation of small & large intestine
inflammation of GI tract with blood & pus in faeces
frequent and/or fluid stool (>3 loose stools) - Acute, Persistent, Chronic

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Etiologic Agents to Consider for Various Manifestations of Food Related Illnesses

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Front 16

Types of Diarrhoea

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Front 17

Food Toxemia: “Food Poisoning” “Non-inflammatory Gastroenteritis”

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Aetiological Agents
• Bacterial: S. aureus, B. cereus (1 type) & C. botulinum
• Fungal: Wild Mushrooms (Amanita, Clitocybe &
Psilocybes) Aflatoxin (Aspergillus sp.)
• Marine/Algae: Ciguatera, Scromboid & “Shellfish”

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Food poisoning:Toxemia Ingestion of preformed toxins (NOT INFECTION) NO microbial growth within human GI tract

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Symptomology: usually rapid (minutes-hours)
(C. botulinum: 6hrs-8days)
Lack of fever; no faecal leukocytes
Toxins Affect:
CNS (C. botulinum)
Both CNS & Intestines (S. aureus & B. cereus)

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Staphylococcus aureus: Gram +ve cocci (0.5-1.5mm)
Arranged in singles, pairs & clusters Aerobic or facultative Coagulase +ve, Catalase +ve, ST Enterotoxin production
Mode of action: UNKNOWN - act on gut receptors; stimulate vomiting (vagus & sympathetic nerves) – NO stimulation of adenylate cyclase

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Habitat: Human & animal pathogens (skin), 50% humans carry Staphylococci
8 Exotoxins (A, B, C1, C2, C3, D, E, H) Water-soluble, low mw proteins, ST (chromosomal)
Infective dose 105 – 108orgs/g food (A & D common)
Neurologic (vomiting) & Enteric (diarrhoea) effect

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Staph Aureus: Clinical symptoms

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- Self-limiting illness: EMESIS within 6hr ingestion (mean 4.4hr) (BUT Not all vomit) - Recovery 24-48 hours
nausea, abdominal cramps, diarrhoea (watery), headaches, muscular cramping and/or prostration

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Staph Aureus - Incriminated Foods:
Poor Handling of Food
Outbreaks
Incidence

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cooked meat (fish, poultry), bakery foods (cream-filled),
dairy produce, fruit, vegetables & salads.
– Self-limiting disease (no incentive to report)
– US National Surveillance System (1-5% cases)
Highest: Summer, 2nd: November/December: Holiday

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Staph Aureus: Isolation & Identification

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Variety of Media available (MSA)
Baird-Parker(selective, diagnostic, recovery) - Lithium chloride & tellurite (selective agents) Egg yolk & pyruvate (recovery) Reduction of tellurite  shiny, jet-black colonies Surrounded by clearing zone
Confirm: Coagulase test

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Bacillus cereus: Habitat: air, soil, water & dust
Gram +ve rods (0.7mm x 3-10mm long) Arranged in chains, Aerobic or facultative, Spore former, Emetic toxin & Enterotoxin
Easily spread to food: Cross-contamination
• 2 types of Gastroenteritis

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Emetic (vomiting): Resembles Staph. aureus Short incubation: 2-3 hrs, Duration: 6-24 hr, ST Neurotoxin (peptide) prodn by cells in food
Incriminated Foods: rice & pulses, Incidence is under-reported: mild illness 27,000 cases/year (may include diarrhoeal type)

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Bacillus cereus: Isolation & Identification

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Implicated food contains >105 org/g
• Non-selective medium: Blood agar (sometimes + polymyxin (suppress Gram-ve)

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Clostridium botulinum: Variable size Gram +ve rods Anaerobic, Ferment range of CH2O’s  Gas Spore former, Produce exotoxins Susceptible to penicillin
Habitat: soil (fertilized animal excreta), lower GI tract humans & animals

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Botulism - Food poisoning (1800’s)
• Infant (1976): Most common botulism in U.S
• Food poisoning Association - Originally: contaminated meat (sausage) Now: Home-canning, vegetables, fish, fruits & condiments
• Major concern food processors & consumers

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Clostridium botulinum: pathogenesis

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NEUROTOXIN
• 8 types (A, B, C1, C2, D, E, F, G) Proteins
- Toxin A (potent) 10-8g KILL HUMAN
• Humans: A, B, E, rarely F (Animals: C & D)
• U.S frequent isolate type A, then B & E
• Europe frequent isolate type B (A rare)

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Food poisoning Botulism: Clinical Symptoms Vary
- Mild illness (disregarded or misdiagnosed)
- Serious disease (fatal within 24hr)
GI disturbances
1/3 patients (toxin A or B) & Almost all toxin E

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Incubation time: 18-36hr, Dependant: amount & antigenic toxin type Includes: nausea, vomiting & abdominal pain Diarrhoea often present: Constipation may occur
Toxemia symptoms then apparent, No fever in absence of complicating infections

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Clostridium botulinum: Diagnosis
• Fatal toxemia (rule out botulism)
• REPORTABLE DISEASE

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Presumptive diagnosis – Presence of rapidly descending paralysis – History of ingestion of home canned or fermented food?
Confirmative diagnosis – Demonstration of botulinum toxin in serum/faeces or incriminating food (mouse toxin-neutralization test)

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Differential Diagnosis
- Guillain-Barré syndrome: ascending paralysis
- Myasthenia gravis: descending paralysis
- Other microbial food poisonings & gastroenteritis
- Chemical (& non-microbial) food poisonings

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GB: Paresthesias or other sensory abnormalities, elevated cerebral spinal fluid protein
MG: Accentuation of muscle fatigability during exercise and positive response to endrophomium
No cranial nerve involvement, Symptoms occur within minutes

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Infant Botulism - NOT Food Poisoning
- Infants (2 weeks-6months age)
- Implicated Types A & B
- Symptoms: illness & constipation (overlooked)
- Later: Head control lost, Infant becomes flaccid
- Severely Infected: Respiratory Arrest

Back 30

Spore Germination (GI tract) - Vegetative cells - replicate & release toxin
Proceeds: lethargy, sleeps more than normal Suck & gag reflexes diminish, Dysphagia becomes evident as drooling
4-15% cases sudden death

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Clostridium botulinum: Diagnosis & Treatment
Diagnosis: is difficult
Requires: prompt action for survival
- Differential Diagnosis: neurological + GI Toxin
- Demonstration in faeces

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Treatment
Botulism Antitoxin Heptavalent (A, B, C, D, E, F, G)-(Equine) Supportive measures: maintain respiration
Baby Botulism Immune Globulin (BIG-IV) for A & B toxins

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Mushroom (Fungal) Toxin: Not common
- Short-acting: Wild mushrooms
- Long-acting: Mushrooms (uncultivated)
- CAN BE FATAL

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Short: Toxin: Museinol, Muscarine, Psilocybin, Coprius artemetaris, Ibotenic acid, Incubation <2hrs: Vomiting, diarrhoea
Toxin: Amantia, Incubation 4-8hrs: Diarrhoea, abdominal cramps

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Mycotoxigenic Fungi
Contamination of:
- Tree nuts, peanuts, oilseeds (corn & cotton)
Responsible for:
- Acute necrosis, cirrhosis & carcinoma (liver)

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Mycotoxins: 2 metabolites; Aspergillus, Fusarium & Penicillium
AFLATOXINS: Aspergillus flavus & A. parasiticus
- favourable conditions (temp & humidity)

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Marine Toxins: Large predatory reef fish: barracuda, grouper & amberjacks
Neurologic symptoms: circumoral & extremity paresthesia, severe pruritus, hot/cold temp reversal

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Ciguatera poisoning: Caribbean/Tropical Pacific
Dinoflagellates: Gambierdiscus toxicus: Ciguatoxin
Acute GI symptoms: 3-6hrs after ingestion
- Watery diarrhoea, nausea, abdominal pain (12 hr)

Front 35

Scromboid poisoning: Non-allergic histamine
Scrombridae Fish: tuna, mahi-mahi, marlin & bluefin
Other symptoms: dizziness, urticaria (rash), facial flushing, generalised pruritus, paresthesias

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Bacteria: Stenotrophomonas maltophilia, M. morganii
Histadine - Histamine (Scrombotoxin
- Burning sensation in mouth, a metallic taste
Acute GI symptoms: mins-3hrs after ingestion (<1hr) - - Watery diarrhoea, nausea, lasting 3-6hrs

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Neurologic & Paralytic Shellfish Poisoning
- Dinoflagellate algae: Karenia brevis

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Brevetoxins
• Incubation: <1-3 hours; Duration: 24-73 hours
• Paresthesia, mouth numbness, tingling sensation of
mouth & extremities, GI upset

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Paralytic shellfish poisoning
- Tingling & numbness of mouth spreading to extremities, GI symptoms less common, ataxia (muscular in-coordination) Severe cases: muscular paralysis, respiratory paralysis

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Dinoflagellate algae: Alexandrium spp., Gymnodinium catenatum, Pyrodinium bahamense, Gonyaulax spp.
• Saxitoxins: Incubation: <2hrs; Duration: 3 days

Front 38

Non-Inflammatory Diarrhoea (Aetiological Agents)
• Bacterial – E. coli (ETEC; EPEC), V. cholerae, Clostridium perfringens, Bacillus cereus (diarrhoeal)
• Viral- Rotavirus, Noroviruses, Adenoviruses, Others
- Toxins Affect: Enterotoxins (bacteria)

Back 38

Food associated: foodborne infection
Ingestion of organisms in food, Toxins produced (bacteria) (NO bacterial INVASION)
Symptomology: Acute watery diarrhoea
Longer incubation (than toxemia) due to colonisation With/without fever

Front 39

Escherichia coli: Member of normal (commensal) intestinal flora
- Family Enterobacteriacea, Gram -ve bacilli (Rods)
Facultative anaerobes
- Biochemical reactions, Complex antigenic structure
- Prodn variety of toxins (virulence factors)
- major opportunistic pathogen

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Front 40

ENTEROTOXIGENIC E. coli (ETEC)
Transmission: contaminated food & water
Infective dose: 100 million - 10 billion cells
1st cause of “Traveller’s Diarrhoea”
20-55,000 cases/year in U.S.

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Front 41

2 Plasmid-encoded ENTEROTOXINS
LT: mw80,000; Similar to Cholera; adenylate cyclase
ST: mw1,500-4,000; guanylate cyclase

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Front 42

ENTEROPATHOGENIC (EPEC)
“Infantile Diarrhoea”: Childhood Diarrhoea
Developing countries (50% mortality)
• Management: Rehydration therapy
• Antibiotic therapy: Trimethoprim/Fluoroquinolones

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Pathogenesis
- NOT fully understood (No ST or LT or CFA)
- Plasmid-borne (EAF) Bundle-forming Pilus (BFP)
- Effacement of microvilli

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Vibrio cholerae: Family Vibrionaceae
– Single curved Gram-ve rods, 2 - 4mm long
– (May be linked end to end) forming “S” shapes
– Motile (single polar flagellum)
– Non-spore forming
– Oxidase +ve

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– O & H antigens
• Serogroup: O1 & O139
– Ferment sucrose & mannose NOT arabinose
– Acid sensitive
– Halotolerant

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Vibrio cholerae: pathogeneis
- Infective dose: 108 - 1010 orgs
- Vibrio cells aligning close to microvilli of SI
- CHOLERA TOXIN - Bacteriophage encoded, AB toxin

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Receptor: Ganglioside GM1

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Vibrio cholerae: pathophysiology
- Result: Diarrhoea occurs (20-30 L/day) “RICE WATER
STOOL”

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Vibrio cholerae: treatment

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Management: REPLACE IONIC LOSS
- Oral and/or IV admin glucose
Antibiotic therapy: Tetracycline Reduce duration diarrhoea
Prevention: Parentally administered vaccine
(109 killed Vibrio cells)
- Lasts 3-6 months Only effective O1 serotype
- Not Recommended by FDA Sanitation & Hygiene

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Vibrio cholerae: Diagnosis
- Haiti (after earthquake)
- Sucrose +ve V. cholerae
- Sucrose -ve V. parahaemolyticus V. vulnificus
- Cholera antigens: Serotypes: Inaba, Ogawa &
Hikojima, Biotypes: “Classic” & El Tor

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• Clinical presentation (Cholera)
• Screening of stool samples Oxidase activity
• Thiosulphate-citrate-bile salts-sucrose (TCBS) agar
Sucrose (differentiating agent)

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Clostridium perfringens: 2 different diseases
1. Necrotic enteritis (Darmbrand & Pig-Bel)
- RARE (Papua New Guinea) C. perfringens strain
type C
2. Type A food-borne infection
- Major cause food-borne infection in U.S
C. perfringens strain type A

Back 48

Diagnosis
• Case history & symptoms
• Large # (>106/g) C. perfringens spores in faeces
• Large # vegetative cells (same serotype) in
incriminated food (>106/g)
• Presence of enterotoxin in faeces

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Bacillus cereus: Diarrhoeal (1948):
Resembles C. perfringens
Characterization: diarrhoea & abdominal pain
Incubation: 8-16hrs, Duration: 12-24 hr
LT Enterotoxin(s) prodn vegetative growth (late exponential phase) (in SI) adenyl acyclase-cAMP

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(Foods: meat & vegetable dishes, sauces, pasta, desserts & dairy products)

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Parasitic Causes of Diarrhoea

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Cryptosporidium spp.
Cyclospora cayetanensis
Entamoeba histolytica
Giardia intestinalis

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Viral Causes of Diarrhoea

Back 51

Rotavirus
Noroviruses (Norwalk & Norwalk-like Viruses) Adenovirus
Small round structured Astroviruses
Hepatitis A & E

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Rotavirus: Family Reoviridae: Respiratory Enteric Orphan
- 8 Genera Wheel-shape, 70nm dia, Non-enveloped
- 11 segments ds RNA (6 Structural, 5 Non-structural)
- 10 human rotavirus serotypes (G1-G4 important)

Back 52

Common genotypes: G1, 2, 3, 4 & 9 with [P8] & [P4]
Group 1: Worldwide distribution
Non-group 1 (2,3): Limited distribution (2 - China)

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Rotavirus Surveillance/year
Asia, Africa & LatinAmerica
- 140 million cases, >870,000 deaths (severe dehydration & electrolyte loss)

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• Factors for High Incidence & Mortality
– Unsafe water
– Inadequate sanitation
Age: <6months & >5 years ASYMPTOMATIC Protection against diarrhoeal infection

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Rotavirus: Family Reoviridae: Pathogenesis
- Seasonal? Temperate, Developed countries: “Winter Gastro” Tropical, Developing countries: Year long (Summer)

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Transmission: Faecal-oral route, Water & Air-borne
Incubation period: <48hrs (1-3days)
Replication: epithelial cells of SI
• Faeces: 108-1010 virus particles/ml, Shedding MAY persist for 10 days or more Peak within 8 days

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Rotavirus: Family Reoviridae: Pathogenesis

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Histopath studies: shortening & blunting of villi patchy, irregular intact mucosa mononuclear cell infiltration of lamina propria
Diarrhoea results from the loss of absorptive area & the flux of water/fluid across damaged surface

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Rotavirus: Family Reoviridae: - Clinical Manifestations
Detection- Virus in stool (peak at day 3/4 of diarrhoea)
• Latex agglutination, EIA (for characterisation),, Electron Microscopy (labour intensive, insensitive), Electrophoresis of RNA segments

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Illness: Sudden onset watery diarrhoea
- With/out vomiting
- Up to 6 days (longer: immunocompromised)
Complications
- Dehydration, severe & life-threatening

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Rotavirus: Family Reoviridae: Vaccines
- 1st Rotavirus Vaccine (Rotashield®)

Back 57

Rotashield: Live Oral Tetravalent Vaccine, 3 x 2.5ml doses @ 2, 4 & 6 months of age
- Prevent 50% rotavirus cases 70% severe cases; 100% associated dehydration
- Adverse Reactions: Intussusception
- CDC discontinuing use.

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Rotavirus Vaccine: RotaTeq®
Clinical study: 11 countries (3 continents); 72,324 infants
– 6 cases intussusception in RotaTeq® group
– 5 cases in placebo group

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Live, oral, pentavalent (G1, G2, G3, G4 & P8)
3 doses: 6-32 weeks of age
– 1st dose between 6-12 weeks of age,
- 3rd dose NOT given after 32 weeks of age
– US $63.96/dose

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Rotavirus Vaccine: Rotarix®
• Clinical study
– Latin America & Finland: 63,225 infants
– 6 cases intussusception Rotarix® group
– 7 cases in placebo group

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Live, attenuated oral, G1P8
• 2 doses
– 1st 2 months of age
– 2nd 4 months of age
– US $92.15/dose (2)

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Norwalk virus (Norovirus): Family Calciviridae (2002, Noroviridae)
4 Genera (Norovirus, Sapovirus, Lagovirus & Vesivirus)

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Diarrhoea stool specimens from gastroenteritis patients
Small (27nm dia), Non-enveloped, Amorphous surface: feathery, ragged outline Virion: ss +ve sense RNA (7.5kb), Single structural protein (60 kDa)

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Norwalk-like viruses: Sapoviruses
• Considerable genetic homology with Norwalk
• Shared virological characteristics

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Family Calciviridae
• Children & Adults (Elderly): 5 genogroups (GI-GV), Humans I II IV & V
- 66% gastroenteritis outbreaks in Long term Care facilities

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Sapoviruses: Epidemiology & Pathogenesis
- – Winter Vomiting Disease
- Transmission: 1 Faecal-oral route, Water-borne,
Food-borne (raw shellfish)

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Estimated 50% outbreaks of acute, nonbacterial gastroenteritis (US)
Older children & adults (Children <5yrs
• Virus multiplies in small intestine
• Produces transient lesions of intestinal mucosa
• Spares large intestine (NO faecal leukocytes)
• Shed in faeces

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Sapoviruses: Clinical & Diagnosis
- Mild & Brief (Fatalities are RARE): 24-48hr following ingestion, Lasts 24-60hrs

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abdominal cramps, myalgias, malaise, headache, nausea, low grade fever & 1-2 days diarrhoea
Diagnosis (not sensitive)
– 1) Mean (or median) illness duration of 12-60 hrs
– 2) Mean (or median) incubation period of 24-48 hrs – - 3) > 50% of people with vomiting
– 4) No bacterial agent previously found

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Sapoviruses: Detection
- Virus in stool (peak at day 2/5 after onset)
• Difficult to culture, RT-qPCR assays, High Sensitivity (10-100 virus copies/reaction), RT-PCR (for genotyping), EIA (not sensitive)

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Cruise Ship Outbreaks - sanitisation to cleaning & disinfection

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Norovirus Vaccine
2 doses: 3 weeks apart – IM
47% efficacy against illness – Illness less severe, shorter duration – No observed side effects

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LigoCyte® Pharmaceuticals Inc.
– norovirus virus like particle (VLP) vaccine (with
chitosan and monophosphoryl lipid A as adjuvants)

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Adenoviruses: Adenoviridae
- 2 Genera: Mastadenovirses & Aviadenoviruses
- Small 70-100nm dia, Icosahedral protein shell, 252
capsomeres, Protein core: ds DNA, 12 vertices
PENTONS each with fibre, 2 serotypes associated: 40
& 41 (Group F)

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Pathogenesis: Respiratory Tract
- Infect: Epithelial cells of Pharynx, Conjunctiva, Small
Intestine & occasionally other organ systems
Spread beyond regional lymph nodes NOT usual
• Replicate in intestine & present in stool
• Diarrhoea with or without vomiting
• 5-15% cases diarrhoea (2o Rotavirus)

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Astroviruses: Diarrhoea in Scotland
• Family Astroviridae
• Small 27-30nm dia
• Non-enveloped
• Smooth or slightly indented outer shell
• Inner 5 or 6 pointed star shaped core
• 6.8kb ss +ve sense RNA

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• 2-8% sporadic cases in infants (3o Rotavirus)
• Infection through year: Peak in winter
• 7 human serotypes
UK: Type 1 prevalent (65% cases)
Mexico: 6% cases, Type 2 prevalent (31%)
Japan: Type 6

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Small Round Structured Viruses

Toroviruses: Emerging GI Pathogen (similar to coronavirus)
• At risk?: Aged, immunosuppressed & hospitalised

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Status uncertain
• Some bona fide Calciviruses or Astroviruses
• Require more molecular data

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Hepatovirus: Hepatitis A virus (HAV)
• 27nm icoshedral particle (Picornaviridae structure)
• Non enveloped symmetrical ss (+ve) RNA
• 1983 Enterovirus 72: Biochemical & Biophysical
- Virus shedding in faeces (10-14d after exposure)

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Spread: Faecal-Oral route, person-to-person Poor sanitation & overcrowding (strawberrys, clams, etc)
- Frozen Organic Antioxidant Blend Products

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Hepatitis E Virus (HEV)
• 32-34nm icoshedral particle (similar Calciviruses)
• Non enveloped symmetrical (+ve) RNA
• Final taxonomic classification?

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Enterically-transmitted non-A, non-B hepatitis
• Incubation longer than HAV (mean 6 wks)

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Inflammatory Diarrhoea

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– Shigella spp.
– E. coli (EIEC)
– Salmonella Serotypes
– Campylobacter spp.
– Yersinia spp.
– V. parahaemolyticus & V. vulnificus
– EAEC & STEC (EHEC) (Non invasive organisms - cytotoxin)

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Inflammatory Diarrhoea
Food associated: foodborne infection
- Ingestion of organisms present in food
- Colonisation & Invasion of intestines (Except EAEC &
EHEC)

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Symptomology: Bloody diarrhoea (May begin as watery diarrhoea)
Longer incubation due to colonisation
Fever may be present
Toxins Affect: Enterotoxins &/or Cytotoxins

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Inflammatory Diarrhoea: Shigella sp.
Genus: 50 species, into 4 groups “O” antigens
Virulence factors:
• Endotoxin(Oantigen)
• Exotoxin: Enterotoxin acts as neurotoxin
Causes: meningismus & coma, ulceration
• NAD glycohydrolase Destroys all NAD in human cells
Shuts down metabolismm Cell death

Back 73

Closely related to E. coli (antigens & toxin-capabilities))
GROUP A: Shigella dysenteriae
GROUP B: Shigella flexneri
GROUP C: Shigella boydii
GROUP D: Shigella sonnei

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Back 74

Front 75

Bacillary Dysentery: Shigella dysenteriae type 1 (Shiga bacillus)

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Shiga toxin (cytotoxin) - inhibits protein synthesis
Enterotoxin - produces diarrhea
Exotoxin - inhibits sugar & Aa absorption in SI Neurotoxin - affects CNS

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Shigellosis by other Shigella sp.
= Readily transmitted faecal-oral route: Sanitation breaks down (4 F’s)
• Bloodstream invasion RARE

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• Shigella sonnei - children <5 years (day-care)
• Shigella flexneri – men who have sex with men
• Shigella boydii - rare
Management: Antibiotics: chloramphenicol, ampicillin, tetracycline most common

Front 77

Shigellosis by other Shigella sp.
- Diagnosis, Isolation & Identification
• Isolation from stools, water & food
- MacConkey agar pale/colourless colonies
- S-S agar (Salmonella-Shigella agar)

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Non-motile
Gram-ve rod
NO fermentation lactose
NO utilization citric acid
NO H2S production (except. S. flexneri) NO gas from glucose

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ENTEROINVASIVE E. coli (EIEC)
SE Asia/S America
Similar to Shigellosis less severe (Often mistaken)
NO Shiga toxin
Infective dose as few as 10 organisms

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• Pathogenesis: Invasion of enterocytes in LARGE INTESTINE, Inhibits protein synthesis, killing host cells
Causes dead WBC’s (pus), RBC’s and mucosal cells in stool
• Management Rehydration therapy

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Salmonella Genus: • Ubiquitous pathogens
2200 different types based on Vi antigens (Capsular) Salmonella enterica subspecies enterica serotype XXX

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• Salmonellosis: Clinical Manifestations (3 Types)
• Gastroenteritis
S. Typhimurium, S. Enteritidis, S. Newport
• Septicemia/Bacteremia(focalinfection)Rare
S. Cholerasuis
• Enteric (Typhoid) Fever
S. Typhi

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Enterocolitis (Gastroenteritis)
• Salmonella “food-borne infection”
Localized infection, Excessive fluid secretion from ileum & jejunum

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Salmonella Heidelburg: October 2013 - March 2014
Foster Farms Chicken

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Reptile-Associated Salmonellosis
8 Outbreaks: 473 individuals, 29% hospitalisations, 71% <10yo

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• Infants/children: direct or indirect contact
• Lizards, snakes, turtles & frogs
Turtles <4inches banned in US (1975)
77% redn in turtle-associated Salmonellosis

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Enteric Fever: S. Typhi
Important morbidity/mortality worldwide
US: ONLY seen in traveller’s to Asia, Mexico, India TYPHOID FEVER
Enteric fever: S. Paratyphi A, B or C

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Enteric Fever: ASYMPTOMATIC CARRIAGE
2-5% typhoid patients - excrete 1-1,000 mill. S. Typhi/g faeces
Carrier state important in transmission

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• Management: Chloramphenicol/Ciprofloxacin
• Prevention: 3 Developed vaccines (See CDC website)

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Outbreak: Salmonella Typhi
• Frozen Mamey Fruit Pulp
Motile
Gram-ve rod
NO fermentation lactose H2S production
Gas from glucose Serotyping

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Diagnosis, Isolation & Identification
Isolation from stools, water & food
MacConkey agar pale/colourless colonies
S-S agar (Salmonella-Shigella agar)

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Diagnosis of S. Typhi

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1) History of travel to endemic areas
2) Rose coloured spots on abdomen (2-4days)
3) Examination of blood (anaemia, leukopenia,
absence of eosinophils)
4) Isolation of S. Typhi on S-S agar
5) Positive Widal reaction (agglutination of O & H
antigens)

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Campylobacter species: exclusively veterinary disease, but common cause of diarrhea in humans
– C. jejuni (subsp. jejuni, doylei), C. coli
– C.lari,C.hyointestinalis

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Small, curved-spiral rods
1.5 - 3.5 mm long by 0.2 - 0.4mm wide
Gram -ve
Non-sporing
Motile (single polar flagellum)
Microaerophilic - req. 5% O2, 10% CO2 for growth DO NOT ferment CH2O
Catalase +ve
No growth at 25oC, but at 37oC, readily 42 - 43oC

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Campylobacter species: EPIDEMIOLOGY
Intestinal tract of wide variety of wild & domestic animals (Zoonotic)
• Faecal contaminated water

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Commercially raised poultry
Normal commensal of cows
Long-term commensal of sheep
Pigs (carriers of C. coli)
C. jejuni intestinal commensal
Cats & dogs

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60% all cases: from ingested contaminated liquid or solid food,
i.e., unpasteurized milk, raw/partially cooked poultry & contaminated water

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Campylobacter species:PATHOGENESIS
• Dose - 104 org (as few as 500 cells)
INVASION: Inflammation & Bacteremia
TOXIN:
– Endotoxin
– Enterotoxin: watery diarrhoea
– Cytotoxin: Verotoxin similar to Shiga toxin: (Haem
colitis) Significance not understood

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CLINICAL• Symptoms: 3-5 days after ingestion
Vomiting - slight
Diarrhea - often profuse (green?)
Abdominal pain - often severe
Prostration - often severe
Pyrexia - often present
Other symptoms - bloodstained faeces

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Campylobacter species: Management
Usually self-limiting
Erythromycin - eradicate C. jejuni from faeces
Severe abdominal pain - aminoglycoside, chloramphenicol, doxycycline

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Association/Complications
– Reactive arthritis (1% cases): Knee joint (6-12 mnths) – Acute Inflammatory Demyelinating Polyneuropathy:
Guillain-Barré syndrome (GBS) (30% cases) AKA:
Acute Motor Axonal Neuropathy

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Detection of C. jejuni

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Yersinia enterocolytica (Yersiniosis)
= Lesser cause Y. pseudotuberculosis
• Common in children <7 yrs (1-4 y); adults
• Rivals Salmonella - acute gastroenteritis (cooler
climates)
• -1 - +40oC (Psychrotroph – Facultative psychrophile)

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Pathogenesis (Poorly Understood)
• Invasive induces inflammatory response
Distal ileum (gut-associated lymphoid tissue)
Adjacent tissues & mesenteric lymph nodes also
infected (mimic appendicitis)
• (Chromosomal) ST Enterotoxin
• inc cGMP

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Yersinia enterocolytica (Yersiniosis)
- CLINICAL FEATURES
• Management: oxytetracycline or doxycycline

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Self-limiting enterocolitis
Incubation period 3-7 days
Lasts 14-21 days (Longer)
Symptoms: abdominal pain & diarrhoea Mild fever, vomiting rare

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Post-infective Reactive Arthritis (Autoimmunity Arthritis)
• Small proportion patients (poorly understood)
– Induced polyclonal T-cell stimulation (toxin)
– Non-specific immune stimulation of invasin binding
to b1 integrins on T lymphocyte
– Other bacterial antigens

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Diagnosis, Isolation & Identification
Special investigation
• Diagnosis from stool is possible
Often considered late: rising antibody titres in
paired serum
• MacConkey (pinpointcolonies/48hrs)
• Specialized Yersiniamedia

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NON - CHOLERA Vibrio’s
- common along the gulf coast

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Not agglutinated by anti O1 sera
Halophilic organisms (Common coastal waters)
V. parahaemolyticus
V. alginolyticus
V. vulnificus
V. cholerae (NOT toxigenic V. cholerae O1 or O139)

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Vibrio parahaemolyticus
• Ingestion of raw/poorly cooked seafood
Acute abdominal pain, vomiting & watery diarrhea
Japan - raw fish (# 1 Food-borne)
US - shellfish
• Treatment: tetracycline

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• May-October 2013
Clams & Oysters, Harvest Area Closures

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Vibrio vulnificus - Diarrhoea & infection of cuts (Salt water abrasions) -- Vibrio Illnesses after Hurricane Katrina --- Multiple States
Virulent strain: Management: tetracycline
Intense skin lesions (gastroenteritis & even severe bacteremia)

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Clinical presentation (Not Cholera)
Screening of stool samples Oxidase activity
Thiosulphate-citrate-bile salts-sucrose (TCBS) agar Sucrose (differentiating agent)
Sucrose -ve V. parahaemolyticus, V. vulnificus

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ENTEROAGGREGATIVE (EAEC)
• PATHOGENESIS: Not fully understood
NO EAF (Enteric Adherence factor)
Possess AAF (Aggregative Adherence factor)

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1) Initial adherence to intestinal mucosa and/or mucus layer (fimbriae)
2) Enhanced mucus prodnthick mucous biofilm
3) Cytotoxin prodn?damage to intestinal cells

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ENTEROHAEMORRHAGIC E. coli VEROTOXIN PRODUCING (VTEC)
SHIGA TOXIN PRODUCING (STEC)

causes: LIFE THREATENING CONDITIONS

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HAEMORRHAGIC COLITIS
HAEMOLYTIC UREMIC SYNDROME (8 -11% cases)
Acute renal failure, Thrombocytopenia, Microangiopathic haemolytic anaemia
THROMBOTIC THROMBOCYTOPENIA PURPURA

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Pathogenesis: Attachment (similar to EPEC)
Phage encoded: CYTOTOXIN - VEROTOXIN
- 2 types (VT1 & VT2) both AB toxins
- Shiga-like toxin (rRNA) blocks protein synthesis

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Diagnosis of E. coli’s: Routine stool culture

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1) MacConkey’s agar (Red-Pink colonies)
2) Sorbitol MacConkey’s agar (no fermentation STEC)
3) (ETEC) Inoculate mouse adrenal cells: stimulation of
adenylate cyclase by LT/ST
4) ELISA on toxin bound to antibody
5) DNA probe to detect toxin genes

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Other GI Tract Infections

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Antibiotic-Associated Diarrhoea - C. difficile
Gastritis/Duodenal/Gastric Ulcers - H. pylori
Gastrointestinal Abscess (Peritonitis, Appendicitis & Diverticulitis) - E. coli & Bacteroides spp. + others

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Clostridium difficile
• NOT Food-borne
• Normal flora 3% adults
• MajorNOSOCOMIALpathogen
• Spectrum of intestinal diseases

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• Antibiotic-Associated Diarrhoea
Usually cause only minor concern
Can evolve into a life-threatening enterocolitis
• Antibiotics
Ampicillin, cephalosporins, clindamycin & amoxicillin
• Induced Pseudomembranous Colitis
Antineoplastic agents: methotrexate

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Clostridium difficile

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2 toxins
Toxin A enterotoxin (fluid accumulation in bowel)
(weak cytotoxin most mammalian cells)
Toxin B potent cytotoxin
Decreases cellular protein synthesis & disrupts
microfilament system of cells (similar to diphtheria toxin)

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Clostridium difficile: Clinical Symptoms
• Vary: mild diarrhoea → severe abdominal pain
accompanied fever (>101oF) & severe weakness
• Diarrhoea: watery usually non-bloody (5-10% bloody)
excess mucus & pus (or blood)
Hypoalbumineia & Leukocytosis common

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Diagnosis: Difficult, Not distinguished from Ulcerative colitis & Crohn’s
Colonic examination (presence of pseudomembrane)
AND isolation C. difficile, associated antibiotic therapy Toxin presence

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Pseudomembranous Colitis: Management
Discontinue antibiotic - symptoms resolve 1-14 days

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If severe or no response; treat oral antibiotics:
• Vancomycin (“gold” standard) or Metronidazole (milder infections) - Relapses in 15-20% patients
• Dificid (fidaxomicin) bid 10 days (US $2,800 for 10d course)
• Faecal microbiota transplantation

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Helicobacter pylori: 1982 (Australia) from gastric biopsies Named Campylobacter pyloridis (C. pylori)
1989 Helicobacter pylori

Biological carcinogen

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MORPHOLOGY
Gram -ve
Non spore-forming
Curved to spiral (1-3 turns)
Motile - polar (5-6) flagella Microaerophilic 2-5%O2, 5-10%CO2 Catalase +ve; Urease +ve Coccoidal forms under culture

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H Pylori: ASSOCIATED CAUSE
• Gastritis (stomach atrum)
• Duodenal ulcers (& gastric ulcers)
• Gastric cancer

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Factors contributing to H. Pylori related gastric pathology

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H Pylori: Pathogenesis
• Gastric colonisation is common
• Route of infection UNCLEAR
• Role of cytotoxin, urease, mucinase, flagella
mechanisms UNDER INVESTIGATION

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• UREASE allow H. pylori survival at pH 2.0
Able to split ammonia from urea = alkaline environment
• Virulence Factors: allowing for adhesion & damage to mucosa
- cagPAI: VacA cytotoxin; BabA; OipA

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H. Pylori Management
• H. pylori sensitive in vitro DO NOT WORK Monotherapy in vivo
• EffectiveTherapies
– Triple, Quadruple or Sequential regimens

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• Triple Therapy (clarithromycin resistance <15%)
– PPI (lansoprazole, omeprazole, pantoprozaole or esomeprazole) + amoxicillin (bid), and clarithromycin (bid) – 7-14 days
• Quadruple Therapy (clarithromycin resistance ≥ 15%)
– PPI, combined with bismuth (qid) + two antibiotics (e.g., metronidazole (qid) and tetracycline qid) – 10-14 days

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Helicobacter resistance to clarithromycin in different countries
Vaccination? - Helivax Inactivated, whole cell vaccine

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Initial Vaccine Studies:
Mice vaccinated - Oral vaccine H. felis sonicate + mucosal adjuvant cholera toxin (LT E. coli) = complete protection

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H. Pylori: Gastrointestinal Abscess
• Peritonitis
• Appendicitis & Diverticulitis
• Intra-abdominalabscess
• Liver abscess
• Pancreas abscess

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healthy GI tract >300 bacterial species
Pathogenesis
• Reduced O2 tension & Oxidation-reduction potential
• Impaired blood supply; necrosis of tissue, growth of facultative anaerobes

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H. Pylori
• Association: vascular disease, trauma, surgery, presence of foreign bodies, malignancy, radiation therapy, injection of vasoconstrictive agents, shock, cold or oedema
• Provides: anaerobic environment, impaired host defenses
• Enable opportunists to grow

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Clinical Symptoms & Causes
- E. coli. Also by tuberculosis, N. gonorrhoeae & C. trachomatis infections
Peritonitis: pain, abdominal distention, diffuse muscle
spasm, tenderness & rebound tenderness, decreased/
absent peristalsis, rigidity of abdominal wall, tenderness on rectal or vaginal exam, fever & leukocytosis
1st peritonitis (spontaneous bacterial peritonitis)
2nd peritonitis from spillage of bacteria

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H. Pylori: Appendicitis:
(See pseudoappendicitis: Y. enterocolytica)
• Pelvic abscess: pain, deep tenderness in 1 or both lower quadrants, fever, urinary frequency, dysuria & diarrhoea with passage of mucus in the first stools. Rectal or vaginal examination may reveal tenderness

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H. Pylori • Diverticulitis: Abdominal pain, peritoneal irritation, fever & leukocytosis
– Acute diverticulitus (peridiverticulitis)
– Microperforations of diverticula cause contamination of peritoneal cavity by aerobic and anaerobic flora resident to normal colon
– Most Common Causes: Bacteroides spp., E. coli & enterococci

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H. Pylori
Pylephlebitis & Liver abscess: Chills, fever, epigastric or right upper quadrant pain, nausea, vomiting, enlargement & tenderness of liver
• Pancreatic abscess: (acute) severe epigastric pain. Recent history of excessive ingestion of food & alcohol. Nausea & vomiting are common

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– Cholecystitis: obstruction of cystic duct with subsequent bacterial invasion of the gall bladder
– Most Common Causes: E. coli, Klebsiella sp., Enterobacter sp., Proteus sp., P. aeruginosa, enterococci, streptococci, staphylococci, Bacteroides sp., Clostridium sp., Fusobacterium sp., peptostreptococci

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H. Pylori Diagnosis:
– Type and location of pain
– WBC count
– Biochemistries
– Imaging (X-ray, computerised tomography, position emission tomography, ultrasound & radioisotope imaging - gallium or indium)
– Pathognomonic signs

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Treatment Principles:
Improve vascular perfusion (correct fluid & electrolytes)
Combat effects of bacteria & toxic metabolites
Reduce paralytic ileus
Eliminate primary source of infection
Aspirate infected exudate (drain primary lesion)
Treat local or distant complications

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Antimicrobial Therapy: Anaerobes resistant to penicillins, cephalosporins & most amino glycosides
Possibilities:
Chloramphenicol (succinate): Bacteroides fragilis Metranidazole: All Bacteroides spp.
Gentamycin, Tobramycin & Amikacin: useful Clindamycin: 60% Bacteroides spp sensitive

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Management of Diarrhoeal Disease
Oral rehydration (ORT): till normal rehydration restored (determined by: clinical condition/body weight) Sodium: 150-155mmol/l Glucose: 200-220mmol/l Potassium: 4-5mmol/l

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Intravenous rehydration: shock, exhaustion precluding oral feeding and oral rehydration failure
Antiemetic drugs: reduce fluid loss  oral rehydration effective
Antidiarrhoeal drugs: RARELY SUCCESSFUL (reduce gut motility - allow accumulation of fluid)

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Management of Food-borne Disease in the Community
• Developing countries: Poor water quality GI infections: waterborne
• Developed countries: Food important source Ideally: free from pathogenic bacteria
In practice: frequently contaminated

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Prevention of Infections
• Safe food production: healthy flocks & herds, avoid
sewage to fertilize crops
• Food manufacture processes: hygienic slaughter & meat packing, rodent-free storage of crops, store at chill or refrigeration temps, hygienic packaging, cold chain during distribution

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• Domestic & commercial food hygiene: adequate refrigeration, avoidance of cross-contamination, usage within spoilage dates, adequate decontamination of food by washing and/or cooking, personal & kitchen hygiene

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