1. Topic 04 Gastrointestinal infections II
Lecture for 3rd-year students
General medicine
Ondřej Zahradníček (with use of prof. Votava‘s last year sources)
23rd of October, 2015

2. Revision 1
Gastrointestinal tract is an area interesting from microbiology side of view
It contains two important ecosystems, where bacteria are normally found and use of antibiotic is often contraproductive:
Oral cavity with typical biofilm system (supragingival and subgingival plaque)
Intestine (especially large intestine) with about 1 kg of microbes, mostly anaerobic
On the other hand, other areas contain less or no microorganisms (oesophagus, stomach…)

3. Revision 2 Organ Normal flora Pathogens Oral cavity
Oral streptococci, neisserias, haemophili, anarobes (Prevotella, Porphyromonas), Actinomyces, Lactobacillus, Treponema denticola
Overmultiplied some species from left;
Candida, Herpes simplex virus, rarely something else
Oeso-phagus
None
Candida albicans, CMV (only immunocompromised)
Sto-mach
Helicobacter pylori
Bile bladder
Escherichia coli, Salmonella enterica

4. Revision 3 Organ Normal flora Pathogens Liver None
Hepatitis viruses (HAV to HEV); Granulomatous hepatitis: Coxiella (Q-fever), TB, Brucella; parasites: Entamoeba histolytica, Plasmodium sp., Leishmania sp., Schistosoma sp.
Perito-neal cavity
Those comming from colon (anaerobes + Enterobacteriaceae)
All*
Salmonella enterica ser. Typhi, Paratyphi A, B, C, Listeria (foetus) etc.
*but starting in the intestine

5. Revision 4: Intestinal syndromes
Bacterial overgrowth syndrome: after surgery, during depressed peristalsis or gastric achlorhydria bacteria may overgrow in the small intestine → steatorrhoea, deficiency of vitamin B12, diarrhoea, malabsorption of vitamins A and D
Diarrhoea: increase in daily amount of stool water – common intestinal response to many agents
Dysentery: acute inflammation of the colon → abdominal pain & small-volume stools with blood, pus and mucus

6. Revision 4: Intestinal pathogens
Bacteria: they may cause
Intestinal infection (e. g. salmonellosis)
Enterotoxicosis (staphylococcal, Bacillus cereus etc.)
Viruses: quite common, often not diagnosed
Fungi: present normally in the intestine, but sometimes overmultiplied
Parasites: not always causing diarrhoea, sometimes it can be even obstipation

7. Enterotoxicosis (food poisoning)
The term „food poisoning“ is commonly used for different situations, including infections. Nevertheless, it is necessary to understand that diarrhoea related with bacteria can be
1) Enterotoxicosis = intoxication due to a toxin produced by bacteria in the food; there is no incubation
Staphylococcus aureus enterotoxicosis (a superantigen)
Clostridium perfringens: heat-labile enterotoxin
Bacillus cereus: heat-stable enterotoxin and vomiting toxin (mostly in rice)
Clostridium botulinum: heat-labile neurotoxin
2) Infection caused by bacteria that produce toxins inside the intestine (e. g. enterotoxic strains of Escherichia coli); these are not considered an enterotoxicosis, incubation period exists
3) Infection caused by bacteria that do not produce toxins, but are invasive (e. g. enteroinvasive strains of Escherichia coli)

8. Enteric infection caused by E. coli
Escherichia coli strains causing diarrhoeal disease:
ETEC (enterotoxic E. coli): children in developing countries, traveller´s diarrhea; 2 enterotoxins (heat-labile and heat-stable)
EPEC (enteropathogenic E. coli): approx. 12 serotypes, for example O55, O111; small infants (< 3yo); disruption of microvillus structure; E. coli strains are tested for EPEC
EIEC (enteroinvasive E. coli): similar to Shigella invasion → dysentery-like disease ; invasion of colonic cells
STEC (shigatoxigenous E. coli): also known as VTEC (verotoxigenous E. coli); for more information see further
EAggEC (enteroaggregative E. coli) and other pathogenicity groups
Do not forget that majority of E. coli strains are „normal“ strains helping us by production of vitamins etc.!

9. STEC/VTEC – more information
those strains produce 2 cytotoxic shigatoxins
mechanism of pathogenesis: destruction of microvilli
hemorrhagic colitis & hemolytic-uremic syndrome (HUS)
HUS – three typical signs:
so called mikroangiopatic anemia
trombocytopenia (lack of platelets)
acute renal failure
Letality in HUS is 5 %, almost in children < 5 yo; often lasting ill effects
in 2011 there was a strain O104:H4 in Gerrmany, more common is strain O157:H7
subgroup of STEC is EHEC (enterohaemorrhagic E. coli)

10. Escherichia coli – good and bad
www2.mf.uni-lj.si/~mil/bakt2/bakt2.htm

11. Salmonella infections
Two types of salmonella infections:
1) Systemic infections caused by anthropopathogenic serovars S. Typhi, S. Paratyphi A–C; the intestine is just gate of infection
2) Intestinal infections caused by zoopathogenic serovars S. Enteritidis, S. Typhimurium, S. Infantis etc.; there exist more than 4000 serotypes; in some cases even these may cause systemic infections, but in majority of them it is just intestinal infection
Pathogenesis of both starts with the invasion of intestinal epithelia
In 1) invasion continues and infection becomes generalized → little or no diarrhoea, but pronounced fever & other general symptoms; common carriership even for many years!
In 2) infection is localized to ileocaecal region → diarrhoea, nausea & vomiting, abdominal pain, temperature may be elevated

12. Salmonela on MAL agar
Foto O. Z.

13. Typhoid fever – diagnostics and treatment
Diagnostics and treatment of diarrhoeic infections will be described later, so here only for Typhoid fever:
Diagnostics:
Direct diagnostics – specimens of blood, urine (!) and stool tested for Salmonella, serotyping for S. Typhi
Indirect diagnostics – in later phases – Widal reaction (agglutination in test tubes)
Treatment: antibiotics (chloramphenicol, fluorochinolones, ampicillin, cotrimoxazol)

14. Campylobacter and Shigella
Campylobacter jejuni intestinal infections are recently even more frequent than salmonella infections; the microbe invades jejunal epithelium. Poultry is the reservoir
Shigella is a bacterium that does not exist according to recent genetic findings (it is just a „special case of E. coli). Nevertheless it is still considered existing for practical reasons. It contains four species: Shigella sonnei, S. flexneri, S. boydii, S. dysenteriae
Very low infectious dose → epidemic outbreaks
Transmitted only among humans
Invasion of cells of colon and rectum; in some strains also shiga toxin (the same as in STEC strains of E. coli)
The disease is called bacterial dysentery

15. Campylobacter jejuni

16. Yersinia and Vibrio
Yersinia enterocolitica causes gastroenteritis, in children also mesenterial lymphadenitis (mimicking acute appendicitis). Contaminated food is a vehiculum of infection; the microbe multiplies in refrigerator even at 4 °C
Vibrio cholerae acts through cholera toxin. This toxin activates adenylate cyclase → hypersecretion of water & electrolytes → death by dehydration and electrolyte abnormalities. V. cholerae flourishes in water & causes epidemics. Most important serovars are O1 (with biotypes Classis and El Tor) and O139
Vibrio parahaemolyticus: from raw fish & shell-fish

17. Vibrio cholerae

18. Post-antibiotic diarrhoea
Use of antibiotics makes considerable changes in the intestinal ecosystem; many parts of normal flora are destroyed and some other multiply much more than normally
Clostridium difficile (CLDI) infection: after any antibiotic, but most frequent after lincosamides (lincomycin or clindamycin)
dangerous pseudomembranous colitis
patients contaminate the hospital environment with resistant spores
diagnostics and treatment: see next slide
Infections due to other bacteria: common after tetracyclines; from excessively multiplied Staphylococcus aureus, Pseudomonas aeruginosa or also Candida albicans (an example of diarrhoea of mycotic origin)

19. CLDI infections: diagnostics
Diagnostics: stool is sent to the laboratory (stool, not rectal swab) and examined for CLDI toxin and CLDI antigen using an immunochromatography test. Possible results:
Toxin +, antigen + means clear infection
Toxin –, antigen + means that infection is still possible (the toxin detection might be false negative)
Toxin –, antigen – means that the infection is very unlikely

20. CLDI infections: treatment
metronidazol (milder cases)
vancomycin taken orally (!) (severe cases) and recently also fidaxomycin (new and very expensive drug)
it is also possible to use so called faecal bacteriotherapy – a stool donor (usually a relative) is tested for many infections; the stool is then mixed with saline and the „cocktail“ is usually administered using a nasogastric sond

21. Viral agents of diarrhoea
Generally: the most typical agents are small, acid- and bile-resistant non-enveloped viruses
Rotaviruses (Reoviridae family) cause serious diarrhoea of young children, epidemics in winter
Noroviruses and sapoviruses (formerly agents Norwalk and Sapporo, Caliciviridae family) epidemics in children and adults, too; norovirus infections are quite common, especially in maritime countries; seefood is common source and people travelling on oversea ships are often infected
Astroviruses (star-shaped virions)
Adenoviruses type 40 and 41
Small, round gastroenteritis viruses

22. Rotavirus

23. Parasitic agents of diarrhoea
In previously healthy individuals:
Entamoeba histolytica: amoebic dysentery
Giardia lamblia: giardiasis
Cryptosporidium parvum: cryptosporidiosis
Cyclospora cayetanensis
In AIDS patients also:
Isospora belli (coccidium)
Strongyloides stercoralis hyperinfection (helminth)

24. Giardia intestinalis
CD-ROM „Parasite-Tutor“ – Department of Laboratory Medicine, University of Washington, Seatle, WA

25. Other intestinal parasites not causing diarrhoea (helminths)
Small intestine:
Ascaris lumbricoides (human roundworm)
Ancylostoma duodenale (Old World hookworm)
Necator americanus (New World hookworm)
Strongyloides stercoralis (threadworm)
Fasciolopsis buski (giant intestinal fluke)
Taenia saginata (beef tapeworm)
Taenia solium (pork tapeworm)
Hymenolepis nana (dwarf tapeworm)
Diphyllobothrium latum (fish tapeworm)
Large intestine:
Enterobius vermicularis (pinworm)
Trichuris trichiura (whipworm)

26. Ascaris lumbricoides (roundworm) egg
CD-ROM „Parasite-Tutor“ – Department of Laboratory Medicine, University of Washington, Seatle, WA

27. Fungi and diarrhoea
Candida albicans and other members of Candida genus are normally present in the intestine, but overmultiplying (after antibiotic treatment or radiotherapy) causes infections. Treatment is, nevertheless, also indicated in case of extraintestinal (commonly vaginal) infections with intestinal reservoir
Enterocytozoon bieneusi (microsporidium) is a causative agent in diarrhoea in AIDS patients. (Microsporidia is a strange group of fungi, formerly considered to be parasites)

28. Candida albicans

29. Intestinal infections × intestinal transmission
Not all faecal-oral route transmitted infections are intestinal infections. Example: poliomyelitis attacks neural system, but is faecal-orally transmitted
On the other hand not all intestinal infections are strictly faecal-orally transmitted
Faecal oral transmission has several subtyptes
alimentary (food contamination; infections with big infectious dose, e. g. salmonellosis)
dirty hands and fomites (shigellosis)
passive vectors (flies, cocroaches)

30. Diarrhoea treatment
Diarrhoea treatment is not related to the causative agent (with exception of parazital diarrhoea, where antiparazital drugs are used)
Main goal is water supply and care about organism
Antibiotic treatment is not used even in bacterial diarrhoea, the effect to the infection is weak, but they paradoxically prolonge the positivity for an infection (e. g. for Salmonella)
Exceptions: travellers (it is necessary to solve it), very serious infections (E. coli O104:H4); quinolones or some other drugs are used
Other treatment: charcoal, eventually drugs with only topic antimicrobial activity

31. Microflora care
In convalescence after diarrhoea, but also after systemic antibicrobial therapy we try to renew normal microflora
We use yogurt (non-sweet, non-fat), fermented cabbage, drugs (Hylac)
Some of them contain substrates for „good“ bacteria, these are prebiotics.
Some containg directly the bacteria, those are probiotics
Some contain both, those are symbiotics

32. Prevention of intestinal infections
Care about water sources
Good food hygiene (farms, stores, but also our kitchens!)
No cross contamination (do not store already prepeared meals with fresh eggs or meat)
Personal hygiene (children education)
Fighting passive vectors (flies, other insects)
Public health regulations in persons positive for some infections (no work with food, no attendance of school etc.)

33. How to také stool for individual examinations
Bacteria – Amies (or similar) transport medium
Yeasts – Amies transport medium, eventually a special fungal transport medium (FungiQuick)
Viruses – a hazel-nut sized specimen
Parasites – a hazel-nut sized specimen, traveller anamnesis, usually three specimens
CLDI toxin – a hazel-nut sized specimen
Threadworms – Graham method (see practical sessions)

34. Diagnostics of bacterial agents
Microscopy of specimen is usually not used
Cultivation uses specific media, almost for enterobacteria(see next slide)
Direct examination of toxins A a B (Clostridium difficile) and its antigen
Diagnostics of viral agents: usually antigen detection, eventually DNA detection
Diagnostika parasital and fungal agents: special parasitonogy and mycology methoda

35. Stool processing
Day 0. (sampling)
24 h
selenit
XLD
Endo
MCs KA
28°C
42°C
48 h
MAL
Endo
NaCl
CIN
CCDA
Selenit,e XLD, MAL – for Salmonella diagnostics
CIN – Yersinia
CCDA – Campylobacter
NaCl – staphylococci
MCS – some STECs
Endo – Enterobacteria
Blood agar – some more +
72hod
identifikace
Negative result – after 48 h
Positive result – 72 h and more
*If not mentioned specially, culture is performed at 37°C

36. Thanks for listening