-
VYSOKÉ UČENÍ TECHNICKÉ V BRNĚBRNO UNIVERSITY OF TECHNOLOGY
FAKULTA CHEMICKÁ
ÚSTAV CHEMIE POTRAVIN A BIOTECHNOLOGIÍ
FACULTY OF CHEMISTRY
INSTITUTE OF FOOD SCIENCE AND BIOTECHNOLOGY
SALMONELOVÉ INFEKCE, JEJICH ZDROJ ADIAGNOSTIKA
INFECTION OF SALMONELOSIS, THEIR SOURCE AND DIAGNOSIS.
BAKALÁŘSKÁ PRÁCEBACHELOR'S THESIS
AUTOR PRÁCE MARTIN VALKUSAUTHOR
VEDOUCÍ PRÁCE Mgr. DANA VRÁNOVÁ, Ph.D.SUPERVISOR
BRNO 2009
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-
Vysoké učení technické v BrněFakulta chemická
Purkyňova 464/118, 61200 Brno 12
Zadání bakalářské práce
Číslo bakalářské práce: FCH-BAK0203/2007 Akademický rok: 2008/2009
Ústav: Ústav chemie potravin a biotechnologií
Student(ka): Martin Valkus
Studijní program: Chemie a technologie potravin (B2901)
Studijní obor: Biotechnologie (2810R001)
Vedoucí bakalářské práce: Mgr. Dana Vránová, Ph.D.
Konzultanti bakalářské práce:
Název bakalářské práce:
Salmonelové infekce, jejich zdroj a diagnostika
Zadání bakalářské práce:
1. Vypracování literární rešerše na zadané téma.
2. Výběr metod pro experimentální část a jejich aplikace
3. Zpracování výsledků a jejich zhodnocení.
Termín odevzdání bakalářské práce: 29.5.2009
Bakalářská práce se odevzdává ve třech exemplářích na sekretariát ústavu a v elektronické formě
vedoucímu bakalářské práce. Toto zadání je přílohou bakalářské práce.
- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -Martin Valkus Mgr. Dana Vránová, Ph.D. doc. Ing. Jiřina Omelková, CSc.
Student(ka) Vedoucí práce Ředitel ústavu
- - - - - - - - - - - - - - - - - - - - - - -V Brně, dne 1.12.2008 doc. Ing. Jaromír Havlica, DrSc.
Děkan fakulty
-
3
ABSTRAKT
rodem Salmonella, jeho
i
V
.
ABSTRACT
thesis deals with infection of salmonelosis
diagnosis. Theoretical part includes general information about Salmonella species and its
morphology. It describes pathogenesis, source of infection, clinical symptoms, treatment and
diet recommendations. It analyzes statistics of number of infection occurrences in Czech
Republic. It also deals with prevention of infection origin on schools, population
foreknowledge and climatic effects.
Experimental part of this thesis contains description of optimization procedure of
identification Enterobacteriaceae by adjusting of pH of prepared medium MIU.
Salmonel
KEYWORDS
Infection of samonelosis, prevention, incidence, source of infection, cultivation, diagnosis
-
4
v
hlediska obsahu majetkem
...............................................
Podpis studenta
-
5
OBSAH
.................................................................................................................................. 7
..................................................................................................... 8
..................................................................................................... 8
2.1.1 Morfologie ................................................................................................................. 8
2.1.1.1 Rod Salmonella .................................................................................................. 9
.................................................................................................................... 9
.............................................................................................. 10
........................................................................................ 10
2.2 Patogeneze ...................................................................................................................... 10
................................................................................................. 11
2.2.2 Postup infekce ......................................................................................................... 11
...................................................................................................... 12
2.2.3.1 Tyfus a paratyfus .............................................................................................. 12
............................................................. 12
2.2.3.3 Bakteriemi .......................................................... 12
............................................................................... 12
2.2.4 Produkce endotoxinu ............................................................................................... 13
2.3 Zdroje infekce ................................................................................................................ 14
......................................................................................................... 15
..................................................................... 15
...................... 16
2.5.1. Prevence ................................................................................................................. 17
................................................................................ 17
2.5.1.2 Prevence v ........................................................................ 17
2.5.1.3 Prevence v ......................................................................... 17
........................................................................................................ 18
................................................................ 18
............................................................................................................... 19
........................................................................... 21
potravin ..................................................................................................... 21
........................................................................................ 21
2.6.1.2 Sterilizace ......................................................................................................... 22
.............................................................................. 22
..................................................................... 23
........................................... 23
........................................... 24
..................................................................................................... 28
................................................................................................ 28
.......................................................................................... 28
............................................................................................... 29
2.6.2.4 Kultivace .......................................................................................................... 29
....................................................... 29
ST ....................................................................................... 30 ..................................................................................... 30
-
6
.............................................................................................................. 30
................................................................................................. 30
.................................................................................................. 30
.................................................................................................................. 30
E .......................................................................................... 32 ......................................................................................................................... 32
4.1.1 Salmonella Enteritidis ............................................................................................. 32
4.1.2 Shigella sonnei ........................................................................................................ 33
4.1.3 Escherichia coli ....................................................................................................... 33
4.1.4 Klebsiela pneumonie ............................................................................................... 34
4.1.5 Proteus mirabilis ...................................................................................................... 34
............................................................................................................... 35
.............................................................................................................................. 36
....................................................................................................... 38
........................................... 41
-
7
kruhu po
aj.
supe
potrav .
v
projevy.
bi .
-
8
2.1
Rod Salmonella Enterobacteriaceae
Salmonella enterica
poddr
[1]
Bakterii salmonely objevil roku 1885 doktor Theobald Smith z
mikrobiologie D. E. Salmona. [2]
2.1.1 Morfologie
Enterobacteriaceae ch,
, nesporu ch
rody: Citrobacter, Escherichia, Hafnia, Klebsiella, Pantoea,
Proteus, Morganella, Providentia, Salmonella, Shigella a Yersinia. [3]
Obr. 1: Bakterie Salmonelly
-
9
2.1.1.1 Rod Salmonella
Rod Salmonella
Kaufmann-
T fmann- [1, 25]
Skupina
A Salmonella Paratyphi A
B Salmonella Paratyphi B
Salmonella typhimurium
C Salmonella cholera-suis
Salmonella Paratyphi C
D
Salmonella Typhi
Salmonella Enteritidis
Salmonella gallinarum-pulorum
E Salmonella anatum
s je Salmonella
enterica subsp. enterica r
, aby se v - Salmonella
Enteritidis. [3]
Salmonel
:
p
ejce);
d - p [1]
V almonely hoj
st
da, kt
lze je izolovat z
ny
Salmonella Typhi a Salmonella
P je
nalezneme Salmonella Paratyphi B a z
nebo dobytka). [5]
-
10
Z
Salmonella cholerae-suis prase
Salmonella dublin dobytek
Salmonella gallinarum-pullorum
Salmonella Enteritidis d , dobytek
Salmonella abortus-equi
Salmonella abortus-ovis ovce
Salmonella typhimurium dobytek, holubi
tab.
velkou
produkci su
y salmonel
[5, 22]
2.1.2.2
.
s
nakazit z
p - v je krmivo vyrobeno z ;
s - ebo
( ).
je voda z
mi vodami. [8]
2.2 Patogeneze
Ke vzniku salmonelou erie
v
v
v
v 2-3
,
-
11
um a usmrcuje bakterie a
kvasinky.
[5]
2.2.1 infekce
K se salmonely
potv neinfikovali,
k tomu, aby se infekce rozvinula. Rozvoj infekce po expozici salmonelou j
I [5]
na vehikulu infekce. Bakterie v
ch kyselost v
je izolace
salmonel z vzor ,
em
2.2.2 Postup infekce
Jakmile se salmonel K
s
[5]
-
12
pii: [5]
tyfus a paratyfus;
gastroent ;
b ;
d .
. [23]
2.2.3.1 Tyfus a paratyfus
Jsou Salmonellou Typhi nebo Salmonellou Paratyphi A, B
7-
salmo do krve a ze do
20%. [5]
fekce
(symptomy jsou r d
, ch -
moc dokonce [5]
kytovat i u
osoby s mi mi , nebo osoby
Salmonelly Enteritidis. [5, 27]
Bakterie salmonely se za
paratyfu se
s , u osob nad 50 let je to
a u . [5]
-
13
2.2.4 Produkce endotoxinu
Endotoxiny jsou lipopolysacharidy (LPS)
specifickou tk
hydro
. U
bakt je jeho stru
metody.[10, 26]
monocyty- - monocyt cca 12-
Cirkuluje v -8%
krev . Monocyto-
( );
neutrofily
schopnost poh - jeho
jed
v
p ;
a
s
m
;
e
DIC diseminovan i
[10]
Obr. 2: St LPS Salmonelly Typhimurium [13]
-
14
2.3 Zdroje infekce
Salmonel
-41 -32 5- ). Pro
. Vzhledem k tomu, salmone , krom
, i
pokrmy z ;
c ;
n .
Salmonella ovoce (a to jak ze slupky, tak z ).
V roce 2001 propukla infekce
sestry. Bylo
z
V y
(racci a holubi)
-
15
Jednou z batole
organismu (dehydr
m
(gastroenteritidou) p ale naopak
p .
bakteriemie. [6]
v m
v [7]
Na 1
hodinu.
P d
1
ho
cukru.
B
B
.
N .
N .
V ,
y nebo
pomno
-
16
2.5 tu
Salmonella v
2008 pod v grafu 1:
Tab.3:
Rok salmonelami
1999 44845
2000 40233
2001 33594
2002 27964
2003 26899
2004 30724
2005 32927
2006 25102
2007 18204
2008 11009
Graf 1: salmonelami v letech
2008
0
10000
20000
30000
40000
50000
1999 2000 2001 2002 2003 2004 2005 2006 2007 2008
Roky
1999 - 2008
P klesl
-
17
2.5.1. Prevence
oblasti
prevenci v
kniha) a v V
se v [18]
2.5.1.1
formou
v
ovanost
2.5.1.2 Prevence v
V
2.5.1.3 Prevence v
podkapitoly:
-
-
-
18
v
2.5.1.4 Vliv m
ozhlas a
v
V
kontrolovalo zda:
n
a
muse
HACCP slo
, a mohou
stanice. [19, 29]
-
19
2.5.1.6
i
almonely v po
, Toto riziko
roste potravin z
22 meteorolo
z S
do tabulky (Tab.4 grafu (Graf 2).
Tab.4: [14]
Rok
1999 8,45
2000 9,22
2001 7,88
2002 8,79
2003 8,52
2004 8,09
2005 7,89
2006 8,45
2007 9,22
2008 8,96
Graf 2:
7,00
7,50
8,00
8,50
9,00
9,50
1999 2000 2001 2002 2003 2004 2005 2006 2007 2008
Rok
-
20
letech
letech
letech,
-
21
2.6 Diagnostika rodu Salmonella
Salmonella
Roli hraje i aktivita vody
pod [15]
potravin
[15]:
p ;
p
tetrathionanem a novobiocinem);
v
(
p
to, aby se nerozp
y ve
nabobtna
tuhost
o 2,5-
[15]
-
22
2.6.1.2 Sterilizace
(je uvedeno v popisu
na obal
po dobu 30-ti
minut ve varu
deionizovanou vodu a nav
v [15]
2.6.1.3 F
na 45
p
v ,
po rozli
zkumavek lze d
petriho misky p j ipraveny
k je
zkumavky
inou v [15]
-
23
o
.
v
Pokud
, a nelze ji po
m
k
.
.
.
[15]
vzorku
inkubujeme v
dobu
du Salmonella
-
redukuje
-
24
Tab.5.: Biochemi
Mikrob pohyb plyn z
Utilizace
Tvorba
H2S
Tvorba
indolu
Salmonella
Typhi + - - - - + -
Salmonella
spp. + + - - + + -
Shigella spp. - - - - - - V
Escherichia
coli + + + - - - +
Citrobacter
spp. + + + - + V V
Klebsiella spp. - + + + + - V
Proteus spp. + V - + V V V
-
Pohyb n h testovat na
pohyb,
na 24 hodin do termostatu, a oblasti
vpichu nebo v
Plyn z bakterie rodu Salmonella
nitol 2vrstvami MIU a agaru podle Haj
se realizuje v tzv. -
2:1.
Salmonelly Salmonelly
Indii, kde byly 2 % z
. [28]
Klebsielly nebo Protea a projevuje se
n Toto bylo
-
-
25
Tvorba H2S j
- (Kligler iron
agar
- Manitol
agar obr. 3 - obr. 4 Lysin
obr. 5)
[16]
-
26
nteritidis na XLD [16]
zkumavce s
Na obr. 6 Salmonella Enteritidis a
Escherichia coli.
Escherichia coli. Je
je
tvorba plynu.
Salmonellou En
Obr. 6: Kombinace agaru podle Hajna a MIU
-
27
Tvorba indolu
Na obr. 7 byl proveden test na tvorbu indolu u Escherichia coli a Salmonelly
Enteritidis. V ce Escherichie coli a v
Salmonelly Enteritidis.
Obr. 7
-
28
[5]:
s
dle Stuarta,
Blaira).
Tampon se do zkumavky z [5]
patogen, , 20]
Transport
Kultivace, izolace
-
29
2.6.2.3
, a to jak pro pacienta, tak
z [5]
2.6.2.4 Kultivace
v kapitole a.
je spojeno s , proto t ch
24 hodin. Pacienti s
a [5]
Na obr. 8 je
Obr.
-
30
ST
east extrakt (IMUNA
Pharm a.s., SR), Gluk ), K2HPO4 (M
- - 121 V
(Chirana, o),
pH metr
pipeta, Petriho misky
10 %) vu pH.
roztok
2HPO4
a 7,5 g agaru b 5
vody bylo
a
-
31
vypnuto a Roztok se 20 minut.
Tab. 6:
Erlenmayerova
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11.
6,00 6,20 6,40 6,60 6,80 7,00 7,20 7,40 7,60 7,80 8,00
6,02 6,21 6,42 6,61 6,81 7,01 7,20 7,42 7,61 7,82 8,01
pH po sterilizaci 5,96 6,15 6,38 6,57 6,76 6,95 7,16 7,37 7,58 7,79 7,98
6,01 6,19 6,41 6,59 6,81 7,01 7,20 7,39 7,61 7,80 8,01
V takto
za a
Tab. 6.
jeho
etriho misku a
a n
22 zkumavek tak, aby sloupec
5 cm.
Salmonellou Enteritidis, Shigelou sonnei, Eschericia coli, Klepsielou pneumoniae
a Proteem mirabilis
-
32
E
Salmonella Enterobacteriaceae a to
bioch . :
XLD
MAL
DCA
Agar podle Hajna
Bizmut
TSI
detekci salmonel :
-
ONPG test
Jednou z
je komb . P
ohyb na hodnotu pH
Enteritidis v
Enterobacteriaceae.
4.1.1 Salmonella Enteritidis
Tab. 7: u Salmonelly Enteritidis
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11.
Pohyb - +/- + +/- + + + +/- +/- +/- +/-
Indol - - - - - - - - - - -
Urea - - - - - - - - - - -
+
-
+/-
-
33
v V
roz 7,2. Po
s
4.1.2 Shigella sonnei
Tab. 8: u Shigelly sonnei
Ozn
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11.
Pohyb +/- +/- +/- - +/- - - - - +/- +/-
Indol - - - - - - - - - - -
Urea - - - - - - - - - - -
+
-
+/-
se
Shigelly roz
7,0 7,6. P s
4.1.3 Escherichia coli
Tab. 9 Escherichia coli
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11.
Pohyb + + + + + + + + + + +
Indol + + + + + + + + + + +
Urea - - - - - - - - - - -
+
-
+/-
U Escherichie coli
-
34
V
bublinek tvorba plynu z
4.1.4 Klebsiela pneumonie
Tab. 10 Klebsiely pneumonie
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11.
Pohyb +/- +/- - - - - - +/- +/- - +/-
Indol - - - - - - - - - - -
Urea V V V V V V V V V V V
+
-
+/-
V
U Klebsiely pneumonie
v termos
Klebsiela pneumonie n 7,2.
Tvorba indolu byla negativ
4.1.5 Proteus mirabilis
Protea mirabilis
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11.
Pohyb + + + + + + + + + + +
Indol - - - - - - - - - - -
Urea + + + + + + + + + + +
Poz +
-
+/-
U Protea mirabilis
-
35
Tab. 11: Roz Enterobacteriaceae
Bakterie Pohyb Indol Urea
Salmonella Enteritidis + - -
Shigella sonnei - - -
Escherichia coli + + -
Klepsiela pneumonie - - +
Proteus mirabilis + - +
+
-
Tab. 12: ,
Bakterie
Pohyb Indol Urea
Salmonella Enteritidis 6,8-7,2 6,0-8,0 6,0-8,0
Shigella sonnei 7,0-7,6 6,0-8,0 6,0-8,0
Escherichia coli 6,0-8,0 6,0-8,0 6,0-8,0
Klepsiela pneumonie 6,4-7,2 6,0-8,0 6,0-8,0
Proteus mirabilis 6,0-8,0 6,0-8,0 6,0-8,0
Klepsielu
pneumonie (viz. kap. 4.1.4 Klebsiela pneumonie) .
Salmonelly Enteritidis, Shigelly sonnei a Klebsiely
pneumonie. v 7,2 (viz. Tab. 12)
Enterobacteriaceae Tab. 11.
-
36
V byla zhodno teristika Salmonella, jeho
klin
Byly
kupovat v dispozici.
letech 1999
V
dle a 561/2004 Sb. v
k
prodejen.
letech 1999-2008 z
Salmonella z potravin a
Jsou uvedeny
Salmonella.
V byl proveden pokus o hodnoty
7,6
-
37
a pH byla stanovena na 7,0 7,2
o na
minimum.
-
38
1. -270.
2.
s. 53-62. ISBN 80-902896-6-5
3.
metod v -165.
ISBN 80-210-2272-8.
4.
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s. 80-82.
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[cit. 22.4.2009].
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ISBN 80-247-0584-2.
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ISBN 80-7345-037-2.
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[cit. 19.5.2009] http://www.cqs.cz/haccp.php>.
-
41
ATB Antibiotika
BGA Brilliant Green Agar
DCA -
DIC
H2S
HACCP Hazard Analysis Critical Control Points (
)
HCl
HPCP Fenol-chloroform-benzol extrakce
ISO International Organization for Standardization
K2HPO4
KIA Kligler iron agar
LPS Lipopolysacharid
MAL Manitol- -
MIU Pohyb-urea-
NaOH
PS
Sb.
SBP
TSI Triple sugar iron agar
XLD r