Isolation of lactic acid-related bacteria from the pig mucosal proximal gastrointestinal tract, including Olsenella umbonata sp. nov. and Veillonella magna sp. nov. [Elektronische Ressource] / vorgelegt von Mareike Kraatz

Mareike Kraatz

Informations

• Publié par: freie_universitat_berlin
• Publié le: 01 janvier 2011
• Nombre de lectures: 34
• Poids de l'ouvrage: 4 Mo

Extrait

Aus dem
Institut für Tierernährung
des Fachbereichs Veterinärmedizin
der Freien Universität Berlin

Isolation of lactic acidrelated bacteria
from the pig mucosal proximal gastrointestinal tract,
including Olsenella umbonata sp. nov.
and Veillonella magna sp. nov.

Inaugural-Dissertation
zur Erlangung des Grades eines
Doktors der Veterinärmedizin
an der
Freien Universität Berlin

vorgelegt von
Mareike Kraatz
Tierärztin aus Berlin

Berlin 2011

Journal-Nr.: 3431 Gedruckt mit Genehmigung
des Fachbereichs Veterinärmedizin
der Freien Universität Berlin

Dekan: Univ.-Prof. Dr. Leo Brunnberg
Erster Gutachter: Univ.-Prof. a. D. Dr. Ortwin Simon
Zweiter Gutachter: Univ.-Prof. Dr. Lothar H. Wieler
Dritter Gutachter: Univ.-Prof. em. Dr. Dr. h. c. Gerhard Reuter

Deskriptoren (nach CABThesaurus):
anaerobes; Bacteria; catalase; culture media; digestive tract; digestive tract mucosa; food
chains; hydrogen peroxide; intestinal microorganisms; isolation; isolation techniques;
jejunum; lactic acid; lactic acid bacteria; Lactobacillus; Lactobacillus plantarum subsp.
plantarum; microbial ecology; microbial flora; mucins; mucosa; mucus; new species;
Olsenella; Olsenella profusa; Olsenella uli; oxygen; pigs; propionic acid; propionic acid
bacteria; species composition; stomach; symbiosis; taxonomy; Veillonella; Veillonella ratti

Tag der Promotion: 21. Januar 2011
Diese Dissertation ist als Buch (ISBN 9783832527891) über den Buchhandel oder online
beim Logos Verlag Berlin (http://www.logos-verlag.de) erhältlich.
This thesis is available as a book (ISBN 9783832527891) via the book trade or online at
Logos Verlag Berlin (http://www.logos-verlag.de).








Allen gewidmet, von denen ich lernte
—Günter Grass, Beim Häuten der Zwiebel (2006)




Contents
Chapter I Scope and intention 10
Outline 12
Coauthors 14
Chapter II Lactobacilli and other lactic acidrelated bacteria in the mucosal
proximal gastrointestinal tract of pigs: a review of ecology for two
derivative approaches for isolation of novel species 19
Chapter III LHP: a new medium for the specific direct isolation of lactobacilli
and screening for strains of novel species with haemindependent
manganese catalase activity on hydrogen peroxide 51
Chapter IV PGMplus and PGM: new mucinbased media in the specific
isolation of lactic acidrelated bacteria from the mucosal stomach
and jejunum of a pig, including Olsenella umbonata sp. nov. and
Veillonella magna sp. nov. 78
Chapter V Olsenella umbonata sp. nov., a microaerotolerant anaerobic lactic
acid bacterium from the sheep rumen and pig jejunum, and
emended descriptions of Olsenella, Olsenella uli and Olsenella
profusa 98
Chapter VI Veillonella magna sp. nov., isolated from the jejunal mucosa of a
healthy pig, and emended description of Veillonella ratti 124
Chapter VII Lactic acidrelated bacterial community from the mucosal stomach
and jejunum of a pig, including Olsenella umbonata sp. nov. and
Veillonella magna sp. nov. 140
Chapter VIII General discussion 149
Chapter IX Summary / Zusammenfassung 189
References 191
Acknowledgements / Danksagung 192
Statement of originality / Selbständigkeitserklärung 193
v
Abbreviations
16S 16Svedberg
adj. adjective
Ala alanine
ANI Anaerobe Identification
AP alkaline phosphatase
approx. approximately
ATCC American Type Culture Collection
ATP adenosine triphosphate
BHG brainheartglucose
BLAST Basic Local Alignment Search Tool
bp base pair
°C degree Celsius
C4/C8/C14 4/8/14 carbon atoms
ca. circa
Ca calcium
CaCl calcium chloride 2
CCUG Culture Collection University of Gothenburg
CDC Centers for Disease Control
CFU colonyforming units
CIP Collection de l’Institut Pasteur
CO carbon dioxide 2
CoA coenzyme A
CuSO copper sulphate 4
d day (time)
D$ Dconfigurated
DDBJ DNA Data Bank of Japan
DIG digoxigenin
DNA deoxyribonucleic acid
DSM/DSMZ Deutsche Sammlung von Mikroorganismen und Zellkulturen
edn. edition
EDTA ethylenediaminetetraacetic acid
e.g. exempli gratia (for example)
EMBL European Molecular Biology Laboratory
et al. et alii, et aliae (and others)
FAA fastidious anaerobe agar
FCM$FISH flow cytometryfluorescent in situ hybridization
vi
fem. feminine (gender)
FEMS Federation of European Microbiological Societies
FeSO iron sulphate 4
Fig. figure
g gram
GC gas chromatography
G+C$content guanine and cytosine content
GI gastrointestinal
JCM Japan Collection of Microorganisms
Jr. junior
h hour (time)
HAP hyperammoniaproducing
HCl hydrochloric acid
H O water 2
H O hydrogen peroxide 2 2
HPLC highpressure liquid chromatography
H S hydrogen sulphide 2
i.e. id est (that is)
IJSEM International Journal of Systematic and Evolutionary Microbiology
INRA Institut National de la Recherche Agronomique
kDa kilodalton
K HPO potassium monohydrogen phosphate 2 4
KH PO potassium dihydrogen phosphate 2 4
l litre
L. Latin
L$ Lconfigurated
LAB lactic acidproducing bacteria
LAMVAB Lactobacillus Anaerobic MRS with Vancomycin and Bromocresol green
LARB lactic acidrelated bacteria
LDH lactate dehydrogenase
LHP Lactobacillushydrogen peroxide
LMG Laboratorium voor Microbiologie en Microbiële Genetica
6g microgram
6l microlitre
6m micrometer
m. masculine (gender)
M molar
max maximal
ME minimumevolution
vii
MEGA Molecular Evolutionary Genetics Analysis
meq milliequivalent
mg milligram
MgSO magnesium sulphate 4
min minute (time) or minimal (OD)
ml millilitre
mm millimetre
mM millimolar
mmol millimol
MnSO manganese sulphate 4
MP maximumparsimony
MRS de ManRogosaSharpe
MRS$Van MRSVancomycin
n number (quantity)
n. noun
N dinitrogen (nitrogen gas) 2
N$ nitrogen
NaCl sodium chloride
+
NAD /NADH nicotinamide adenine dinucleotide
Na HPO sodium monohydrogen phosphate 2
NaH PO sodium dihydrogen phosphate 2 4
NCBI National Center for Biotechnology Information
NCIMB National Collection of Industrial, food and Marine Bacteria
ng nanogram
NJ neighbourjoining
N.L. NeoLatin
nm nanometre
nmol nanomol
no. number
nov. nova (novel, new)
O dioxygen (oxygen gas) 2
O$ oxygenlinked
OD/OD optical density /maximal optical density max
p$ para
p./pp. page/pages
PBS phosphatebuffered saline
PCR polymerase chain reaction
PGM porcine gastric mucin
PGMM porcine gastric mucusmucosa
viii
pH pondus Hydrogenii; negative decadic logarithm of the molar
concentration of dissolved hydronium ions
PHYLIP PHYLogeny Inference Package
PJMM porcine jejunal mucusmucosa
pK negative decadic logarithm of the acid dissociation constant K a a
PY peptoneyeast extract
PYG peptoneyeast extractglucose
rDNA ribosomal deoxyribonucleic acid
RDP Ribosomal Database Project
Rogosa’s SL Rogosa’s selective Lactobacillus
rRNA ribosomal ribonucleic acid
s second (time)
SB Slanetz and Bartley
SCFA shortchain fatty acids
SD standard deviation
SEM scanning electron microscopy
sp./spp. species/species pluralis
subsp. subspecies
T type (type strain of species)
TES TrisEDTAsucrose
TMB tetramethylbenzidine
Tris tris(hydroxymethyl)aminomethane
UBM unsupportive basal medium
UK United Kingdom
UPGMA unweighted pairgroup method with arithmetic mean
URAS unreduced aerobically sterilised
UV ultraviolet
vol. volume
v/v volume/volume
wk week (time)
w/v weight/volume
ix
Chapter I
Scope and intention
The present thesis is about lactobacilli, other lactic acidproducing and lactic acidfermenting
bacteria [together termed herein ‘lactic acidrelated bacteria’ (LARB)] from the pig mucosal
proximal gastrointestinal (GI) tract. It describes the theoretical background for the design and
application of two approaches for isolation of novel species of these groups of bacteria from
the mucosae of the stomachs and jejuna of healthy domestic pigs. These approaches were
developed based on what is invariably present in the otherwise often empty (or jejune) pig
proximal gut: oxygen and mucins. As one approach was rewarding, the thesis also contains
the descriptions of two novel LARB species: Olsenella umbonata and Veillonella magna.

I decided on this scope, because I had learned from my veterinary studies, and especially from
the studies of animal nutrition, microbiology and pig diseases during my time of student
research (Kraatz et al., 2004; Taras et al., 2005; Kraatz et al., 2006; Taras et al., 2006), that
the intimate coexistence (symbiosis) of mucosaassociated lactic acidproducing bacteria
(LAB) in the proximal GI tract is utterly relevant for both pig nutrition and health.

Lactobacilli and other acidtolerant LAB are dominant members of the