THE ENTEROBACTERIACEAE FAMILY

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THE ENTEROBACTERIACEAE

FAMILY

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• DOMAİN: Bacteria

• PHYLUM: Proteobacteria

• CLASS: Gammaproteobacteria

• ORDER: Enterobacteriales

• FAMİLYA: ENTEROBACTERIACEAE

• GENUS: Escherichia

There are 41 genus

Salmonella

Yersinia

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GENERAL CHARACTERISTICS

• Gram negative

• Rod

• Non-spore

• Aerobic or facultative anaerob

• 1/3 of the veterinary pathogens are in this family

• Usually motile

(Except S. Pullorum and S. Gallinarum)

• Catalase positive

• All : oxidase negative, fermentative

(4)

Escherichia

• Echerichia coli: Most important agent

• Other factors : E. vulneris, E. blattae, E. fergusonnii, E. hermannii

• E. coli; Gram negative rod, has peritric flagella

• S-typed colony at 24 hours at 37 ° C, general medium

• Homogeneous turbidity at the broth

• Some strains are hemolytic, fermente Lactose

• With this characteristic they form pink colony on MacConkey Agar;

metallic red colour on EMB agar

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• INDOL TEST and METHYL RED TEST POSITIVE;

• UREA, HYDROGEN SULFUR, VOGES

PRESKAUER TEST, CITRATE TEST NEGATIVE

• IMVIC: ++--

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Antigenic Structure

• SOMATIC 'O' ANTIGEN (Incubation at 120 ° C for 2 hours)

• Heat resistant, lipopolysaccharide structure, 188 chromosomal

• FLAGELLAR 'H' ANTIGEN

• Sensitive to heat, in protein form, 53, chromosomal

• CAPSULES 'K' ANTIGEN (Masks O antigen)

• Polysaccharide (N-acetyl neurominic acid), 67 chromosomal

• FIBRIAL (PILUS) ANTIGENS (Masks O antigen)

• Protein form, was called the L fraction of the K antigen at first

• 4 sexes, 200 adhesion pills (EPECs defined)

TYPE- I: MANNOZ-SENSITIVE: hemagglutine, ex; F1 antigens

TYPE-II: MANNOZ-RESISTANT: hemagglutination does not inhibit with mannose, ex; F4 (K88), F5(K99), F6(987P), F41,

Other adhesives : CS31A, intimin

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Toxins

• Enterotoxin, neurotoxin, endotoxin, CNF

ENTEROTOXINS

• Synthesized by E. coli which is responsible of young animals’ (calf, lamb, piglet) intestinal infections

• Enterotoxin-producing E. coli are called ETEC (Enterotoxigenic E. coli)

• Those that do not produce are called NETEC

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• Enterotoxins are form in 2 fractions

– STABLE TOXIN (ST) heat resistant

• STa: activates quanil cyclase in intestinal epithelium, intracellular

guanosine monophosphate increase resulting the increase of fluid

and electrolyte secretion, and intestinal absorption is decreased

• STb: not determined

– LABEL TOXIN (LT) sensitive to heat

• Similar to cholera toxin, activates adenylate cyclase that catalyzes intracellular cAMP. Causes electrolyte (Na, Cl and water) release to lumen

• LT1 and LT2

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Determination of Enterotoxins

• INTESTINAL LOOP TEST

– The culture filtrate is injected into the rabbit intestines in 10 cm sections, followed by fluid and gas formation for 24 hours

• INFANT MOUSE TEST

– Cultural filtration is given to 3-4 years old mice with 1% Evans Blue and after 4 hours bowel weight is proportion to body weight

• TISSUE CULTURE TEST

– Culture supernatant, inoculation to Y1 adrenal cell cultures

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• Neurotoxines : Lipoprotein structure. It causes enterotoxemia in pigs, makes CNS impairment in mouse

• Endotoxines : They are lipoproteins in the cell wall (lipid A). Causes shock symptoms and severe fever in septicemia

• Cytotoxic Necrotizing Factor : Multi-nucleation in cell

culture, produces necrosis of rabbit skin, CNF1, CNF2

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• Aerobactin: In order to utilize iron, E. coli gains the iron from the iron-bound molecules such as

transferrin or lactoferrin in the organism by means of siderophors with the low-molecular compound in the structure.

– Phenolate siderophor from EPEC and ETEC

– Hydrosomate siderophor is released from EIECs

• Colisin: It is a bacteriocin, Up to 20 colisins have been

detected up to A-V

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Pathotypes of E. coli

• APEC (Avian Pathogenic)

• SEPEC (Septicaemiatic)

• UPEC (Uro Pathogenic)

• STEC (Shiga Toxigenic)

• EPEC (Entero Pathogenic)

• ETEC (Entero Toxigenic)

• EIEC (Entero Invaziv)

• EAEC (Entero Agregativ)

• MPEC (Mamo Pathogenic)

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Infections caused by E. coli

• Intestinal Infections (Colibacillosis):

Newborn intestinal infections of calves, lambs, piglets, foals and chicks

• Extra Intestinal Infections :

a) Urogenital system infections

Cystitis, pyelonephritis in cats and dogs; genital system diseases in dogs and horses

b) Breast gland infections

c) Lung inflammations

d) Wound infections

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Colibacillosis

• Calf septicemia :

O8, O9, O78, O45, O117, O35 serotypes

Diarrhea, septicemia, toxemia, sudden death ETECs cause; K99 ya da K99+F41

Oral way; feces and food and water

contaminated with feces are taken into body

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• Hygienic conditions are not good, prenatal stress, increased numbers of E. coli in the intestine, climate changes, nutritional

irregularities, taken pathogenic E. coli to the

body are the reasons for the disease

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Pathogenesis

• The newborn intestine is sterile

• The bacteria in the vaginal and perianal flora of the mother immediately colonize the intestinal tract through the mouth and rectum

• These bacteria are lactobacilli and compulsory anaerobic bacteria

• Lactobacilli lower the pH and suppress E. coli growth.

• Colostrum carries E. coli specific antibodies

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• Hygiene, stress, climate, predisposition to ETEC invasion

• ETECs are colonized by attaching K99 or K99 + F41 adhesins to microvilli of enterocytes

• Sta toxin is caused by acute diarrhea, dehydration and acidosis, coma and death (2-6 hours) (Enterotoxemic colibasillosis)

• Internal organs in the calf septicemia caused by EIEC and

EPEC, joints and nerves lesions, pathogenesis not clear (CNF

role?) Can not develop with the infected animal

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• Lambs; similar to calf colibacillosis

• In pigs; K88 antigen

• In poultry; causes colibacillosis, choligranuloma

(hygiene disease), enteritis, salpingitis, arthritis, air sac ileus and yellow incision ulcer and omphalitis

• In cats and dogs; extraintestinal, pyelonephritis and cystitis

• In cattle; it is also isolated from acute mastitis

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SALMONELLA

• Common in environment, in normal intestinal flora

• Daniel Elmer Salmon (1885)

• Carry the general characteristics of the

Enterobacteriaceae family (Gram(-), rod , without spore, without capsule, facultative)

• Facultative intracellular

• They are motile except for S. Pullorum and S. Gallinarum

• Can not fermente the Lactose. Form hydrogen sulphide

• Media were developed using these properties

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• Zoonotics, cause food poisoning

• Can survive for a long in cold food

• All species are pathogens

• Transmission is always through digestion

• In poultry S. Pullorum, S. Gallinarum are important

• According to the pathology they have formed

– Major Salmonellozis: S. Typhi, S. Paratyphi, only humans typhoid fever – Minor Salmonellozis: All other salmonella, non-typhoidal, toxic-infectious

food poisonings, gastrointestinal infections in different animal species

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• There are 2 species in the Salmonella genus according to the last classification. These:

S. enterica

S. bongori

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• S. enterica; there are 7 subgroups. There are about 2600 serotypes in total. These;

S. enterica subsp. enterica (I) (1454 serotypes) S. enterica subsp. salamae (II) (489)

S. enterica subsp. arizonae (IIIa) (94)

S. enterica subsp. diarizonae (IIIb) (324) S. enterica subsp. houtenae (IV) (79)

S. enterica subsp. bongori (V) (20)

S. enterica subsp. indica (VI) (12)

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• Many of the Salmonella agents isolated from

humans and animals are included in subgroup of S.

enterica subsp. enterica

• Nomenclature; according to the city or the host they were first isolated

– S. Dublin: city

– S. Typhi: typhoid fever

– S. Gallinarum: because it is isolated from chicken

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Salmonella General Characteristics

TEST CHARACTERISTIC

Gram Stain -

Catalase +

Oxidase -

Lactose -

Sucrose -

Glucose +

O/F Fermentative

H₂S +

Gas +

Indole -

Urea -

Lysine Decarboxylase +

Citrate +

Methyl Red +

Voges- Proskauer -

IMVIC -+-+

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MacConkey Agar

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Salmonella- E. coli MacConkey Agar

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Eosin Mehylene Blue (EMB) Agar

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• It is easy to produce in cases of septicemia and abort

• The chance of isolation is increased by using

enrichment methods when the isolation is

made from feces and feed

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ISO 6579

• Pre-Enrichment (the tetrotiyonatl, TPS)

– At 37 , 18-20 hours

– Increase the number of Salmonella

• Selective Enrichment

– Rappaport-Vassiliadis; including 20 mg/l novobiocin; Half of the floor; 41.5 hours 24 hours – Malasitic greenish suppresses other bactericides, Magnesium Chloride enhances Salmonella

• Culture on Differential Media (XLD)

– Xylose lysine deoxycholate (XLD) agar; at 37, 24 hours

– If xylose, lactose, sucrose are fermented, the pH is lowered, red columns due to phenol red – H2S black colony with ferric ammonium

– As lysine decarboxylase increases pH rises around pink colony colony – As a result, if you see black-centered red colonies, Salmonella

• Biochemical tests for Salmonella

•Urea, Indol, TSI agar, Beta galaktosidaz, Lizin dekarboksilaz, VP

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Antigenic Properties

• Antigenic structures of Salmonella agents are required, especially for their identification

• According to the Kauffman-White Scheme, about 2600 Salmonella serotypes have been identified

• First, Salmonella with polyvalent O antisera is

investigated then the other factors are examined

with group specific sera.

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Antigenic Characteristics

• Somatic 'O' Antigens

• It's in all Salmonella. It is heat resistant. 1, 2, 3,… are expressed in numbers.

• Those containing common antigenic factors are called A, B, C .... Z in the same group

• Because the letters are not enough, some groups are indicated by letters + numbers. There are 67 groups

• Ex: S. Typhi, S. Enteritidis, S. pullorum and S. gallinarum D1, which

contain factors 9 and 12,

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Antigenic Characteristics

• Flagellar 'H' Antigens:

• Motile Salmonella has thşs antigen. Heat sensitive (inactive at 60 ° C)

– Phase-1 Antigens : They are specific. It is found only in one or several Salmonella serovars which are close to each other

a, b, c ... z are represented by the letters, when not enough z1, z2, z3 ...

– Phase-2 Antigens: It is found in many Salmonella serovars. 1, 2, 3, ... they are called

– The ones that contain one are monophasic, the ones that carry

both are called diphasic

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Antigenic Characteristic

• Superficial Antigens : are antigens outside of the cell wall of the

bacterium. They take the name of Vi antigen (S. Typhi, S. Paratyphi A and C)

• Those carrying this antigen do not agglutinate with anti-O sera, masks should be heated to 60 ° C for this reason

• When a Salmonella effect is defined according to the antigenic

formula, first the somatic antigen, if there is superficial antigen, if it is motile and diphasic, then the Phase-1 and Phase-2 antigenic factors are written in order

• S. Typhi, S. Paratyphi A, S. Paratyphi C

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Antigenic Formulation

AGENTS NAME GROUP NAME ANTIGENIC

FORMULA MADE OF INFECTION

S. Typhi D1 9,12 (Vi): d:- Human Typhoid

S. Paratyphi A A 1,2,12:a: (1,5) Human Paratyphi

S. Paratyphi B B 1,4,(5), 12:b: 1, 2 Human Paratyphi

S. Paratyphi C C1 6,7,(Vi):c: 1, 5 Human Paratyphi

S. Abortus equi B 4, 12:-:e, n, x Abortus in Mares

S. Abortus ovis B 4,12:c:1,6 Abortus in Sheep

S. Typhimurium B 1,4,(5),12:i: 1, 2 Gastroenteritis in humans and

animals

S. Pullorum

S. Gallinarum D1 1,9,12:-:- White diarrhea and

winged typhoid in poultry

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• S. Pullorum and S. Gallinarum have the same antigenic form and are biovar.

• S. pullorum ornithine decarboxylase test

positive

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O and H symbols

• Edmund Weil and Arthur Felix

• Found that a flagellated Proteus sp. formed a thin film layer on the agar surface

• These motile strains, “Hauch” means film-producing in German

• For those who are non-motile “Ohne Hauch” means

non producing film

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• Endotoxins, Enterotoxins and Cytotoxins

• Endotoxins, intestinal mucosa destruction and severe acute toxemia

• Enterotoxins, increase the cAMP such as the LT of E. coli, resulting in loss of fluid and electrolyte

• Cytotoxins, are similar to Shigella cytotoxin

• O-1 phage lyses Salmonella by 97%

(41)

Typhoid

• Agent S. Typhi

• Contaminated water and contaminated food

• Dispersion with body secretions

• Eye, ear nerves, heart, brain, kidney, lungs and liver are affected

• High fever, headache, diarrhea

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Special Salmonella Infections in Animals

• S. Abortus equi: Sporadic Abort in mare

• S. Abortus ovis: Abort in Sheep and Goats

• S. Cholera suis: Swine paratyphoid

• S. Pullorum: Chick White Disease

• S. Gallinarum: Fowl Typhoid

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Infections Caused by Other Salmonella Factors

• S. Typhimurium S. Infantis

• S. Enteritidis S. Anatum

• S. Dublin

• S. Derby

• S. Newport

• These agents cause gastroenteritis in animals

• S. Typhi: human typhoid

• S. Enteritidis and S. Typhimurium food poisoning

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Yersinia Infections

• Gram negative, cocoid rod

• Without spore, without capsules

• Have peritric flagellae (Excluding Y. pestis)

• Motile at 22-25 degrees; They are non-motile at 37

• Y. pestis is non-motile

• Y. pestis, Y. enterocolitica, Y. pseudotuberculosis, Y. ruckeri

(45)

• Y. enterocolitica

Zoonotic infections that cause acute gastroenteritis in humans and animals and can be transmitted to humans by animals

• Y. pseudotuberculosis

It is mainly seen in rodents, horse, cattle, pigs, sheep, goats, cats and dogs. Canary and turkey is sensitive. Enteric

septicemia is seen.

Tuberculosis-like nodules in the chronic form of liver, spleen,

kidney, lung, and lymph nubs

(46)

THE ENTEROBACTERIACEAE FAMILY