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Preliminary investigations into ostrich mycoplasmas : identification of vaccine candidate genes and immunity elicited by poultry mycoplasma vaccinesVan der Merwe, Elizabeth Frances 12 1900 (has links)
Thesis (MSc)--University of Stellenbosch, 2006. / ENGLISH ABSTRACT: Ostrich farming is of significant economical importance in South Africa. Three ostrich mycoplasmas,
Ms01, Ms02 and Ms03 have been identified previously, and were provisionally named ‘Mycoplasma
struthiolus’ (Ms) after their host Struthio camelus. Ostrich mycoplasmas are the major causative
organisms of respiratory diseases, and they cause stock losses, reduced production and
hatchability, and downgrading of carcasses and therefore lead to large economic losses to the
industry. In order to be pathogenic to their host, they need to attach through an attachment
organelle, the so-called tip structure. This structure has been identified in the poultry mycoplasma,
M. gallisepticum, and is made up of the adhesin GapA and adhesin-related CrmA. Currently, no
ostrich mycoplasma vaccine is commercially available and for this reason the need to develop one
has arisen. Therefore the first part of this study was dedicated to the identification and isolation of
vaccine candidate genes in the three ostrich mycoplasmas. Four primer approaches for polymerase
chain reactions (PCR’s), cloning and sequencing, were used for the identification of adhesin or
adhesin-related genes from Ms01, Ms02 and Ms03. The primer approaches revealed that the target
genes could not be identified due to the high diversity of sequences that were generated. Therefore
sequences were also compared with those of other mycoplasma species in BLAST searches.
Results showed that the most significant hit was with the human pathogen M. hominis oppD, which
is located in the same operon as the membrane protein P100 involved in adhesion. Other hits were
with ABC transporters which may also play a role in cytadhesion.
The second part of this study was aimed at testing whether two poultry mycoplasma vaccines, M.
synoviae and M. gallisepticum, can be used in ostriches to elicit immune responses until an ostrich
mycoplasma vaccine has been developed. Ostriches on three farms of different age groups in the
Oudsthoorn district were therefore vaccinated with these vaccines in a vaccine trial. The enzymelinked
immunosorbent assay (ELISA) was used to test the level of antibody response. Results
showed that both vaccines elicited an immune response in all three age groups. A high percentage
of the ostriches reacted positively, which indicates that both vaccines elicit antibody responses and
may therefore give protection against ostrich mycoplasma infections. / AFRIKAANSE OPSOMMING: Volstruisboerdery is ‘n belangrike ekonomiese sektor in Suid-Afrika. Drie volstruismikoplasmas,
Ms01, Ms02 en Ms03, is voorheen geïdentifiseer en voorlopig ‘Mycoplasma struthiolus’ (Ms)
benaam na aanleiding van hul gasheer, Struthio camelus. Volstruismikoplasmas is die grootste
oorsaaklike organismes van respiratoriese siektes, kudde verliese en die afgradering van karkasse
wat lei tot groot ekonomiese verliese in die volstruisbedryf. Ten einde patogenies vir die gasheer te
wees, moet mikoplasmas deur middel van ‘n aanhegtingsmeganisme vasheg – die sogenaamde
puntvormige struktuur. Hierdie struktuur is in die pluimvee mikoplasma M. gallisepticum
geïdentifiseer, en bestaan uit aanhegting proteïen GapA en die aanhegting verwante proteïen
CrmA. Tans is geen volstruismikoplasma entstof kommersieel beskikbaar nie, en derhalwe het die
behoefte ontstaan om so ‘n entstof te ontwikkel. Die eerste gedeelte van hierdie studie is dus gewy
aan die identifisering en isolering van entstof kandidaat gene in al drie volstruismikoplasmas. Vier
inleier benaderings vir polimerase ketting reaksies (PKR), klonering asook geenopeenvolging
bepalings vir die identifisering van aanhegting of aanhegting verwante gene vanuit Ms01, Ms02 en
Ms03 is gebruik. Die inleier benaderings het getoon dat die teikengene nie geïdentifiseer kon word
nie as gevolg van hoë variasie in die gegenereerde geenopeenvolgings. Derhalwe is
geenopeenvolgings met ander mikoplasma spesies deur middel van BLAST soektogte vergelyk.
Resultate het getoon dat die betekenisvolste ooreenstemming dié met die menslike patogeen M.
hominis oppD was, wat deel vorm van die membraan proteïen P100 operon wat betrokke is by
aanhegting. Ander ooreenstemmings sluit ABC transporters in wat moontlik betrokke kan wees by
aanhegting.
Die tweede gedeelte van hierdie studie het ten doel gehad om te toets of twee pluimvee
mikoplasma entstowwe, M. synoviae en M. gallisepticum, gebruik kan word in volstruise om
immuunresponse te ontlok tot tyd en wyl ‘n volstruismikoplasma entstof ontwikkel is. Volstruise
vanaf drie plase in verskillende ouderdomsgroepe in die Oudtshoorn distrik was ingeënt met hierdie
entstowwe in ‘n entstof proefneming. Die ensiem-afhanklike immuno-absorpsie essaï (ELISA) was
gebruik om antiliggaam response te toets. Die resultate het getoon dat beide entstowwe
immuunresponse ontlok het in al drie ouderdomsgroepe. ‘n Groot persentasie van die volstruise het
positief gereageer wat ‘n aanduiding is dat beide entstowwe immuunresponse ontlok het en kan
dus beskerming bied teen volstruismikoplasma infeksies.
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Immunological and epidemiological investigations in South African ostriches and penguinsBotes, Annelise 04 1900 (has links)
Dissertation (PhD)--University of Stellenbosch, 2004 / ENGLISH ABSTRACT: Newcastle disease (NO) and mycoplasma infections in ostriches have considerable
economic implications for the South African ostrich industry in that NO is a limiting
factor in the export of ostrich products to the European Union and mycoplasma
infections cause stock losses, reduced production, reduced hatchability and
downgrading of carcasses. In the first section of this dissertation, the role of passively
acquired and mucosal immunity in protection of ostrich chicks against Newcastle
disease virus (NOV) was investigated. Ostrich hen serum IgG and yolk IgY were
isolated and characterized, and the transfer of maternal anti-NOV antibodies to the
egg yolk was determined using an enzyme-linked immunosorbent assay (ELISA).
Results indicated that anti-NOV antibodies were successfully transferred from the
ostrich hen to the egg yolk. In addition, ostrich IgA was isolated, characterized and
rabbit anti-ostrich IgA antibodies produced and used for measuring mucosal anti-
NOV IgA antibodies produced in response to mucosal vaccination. Results indicated
that the live La Sota vaccine stimulates IgA production and thus mucosal immunity in
ostrich chicks. In the second section of this dissertation, ostrich mycoplasmas were
isolated and identified using 16S rRNA gene sequencing. These sequences indicated
that ostriches carry three unique mycoplasmas, which are phylogenetically quite
divergent. The 16S rRNA gene sequences of the ostrich mycoplasmas were
subsequently used for the development of specific primers for the detection and
diagnosis of mycoplasma infections in ostriches by PCR.
The last section of this dissertation focuses on avian malaria in African penguins and
the management of this disease during rehabilitation. The Foundation for the
Conservation of Coastal Birds (SANCCOB) is a seabird rescue and rehabilitation
centre, which is largely dedicated to the rehabilitation of diseased, injured and oiled
penguins. Significant mortalities due to avian malaria occur at this facility. The aim of
this study was the development of an ELISA for the purpose of assessing the natural
levels of anti-Plasmodium antibodies in African penguins on entry into the SANCCOB
facility and during rehabilitation. Results indicated significant increases in anti-
Plasmodium antibody levels after entry, which was not influenced by oiling. Infection
with malaria and not parasite recrudescence was viewed to be the cause of this
increase, indicating a possible role of the SANCCOB facility in exposing penguins to
avian malaria. / AFRIKAANSE OPSOMMING: Newcastlesiekte (NS) en mikoplasmainfeksies in voltruise het geweldige ekonomiese
implikasies vir die Suid-Afrikaanse volstruisbedryf. Die rede hiervoor is dat NS 'n
beperkende faktor in die uitvoer van volstruisprodukte na die Europese Unie is, en
mikoplasmainfeksies tot kudde verliese, verlaagde produksie en uitbroei asook lae
gradering van karkasse lei. In die eerste gedeelte van hierdie proefskrif is die rol van
passiewe- en mukosale-immuniteit in die beskerming van volstruiskuikens teen NS
virus (NSV) ondersoek. Volstruishenserum IgG en eier IgY is geïsoleer en
gekarakteriseer en die oordrag van maternale anti-NSV antiliggame na die eier
ondersoek met behulp van 'n 'enzyme-linked immunosorbent assay' (ELISA).
Resultate het getoon dat anti-NSV antiliggame suksesvol van die hen na die eier
oorgedra is. Volstruis IgA is ook geïsoleer, gekarateriseer en konyn anti-volstruis IgA
antiliggame geproduseer wat gebruik is vir die bepaling van mukosale anti-NSV IgA
antiliggame in reaksie op mukosale immunisering. Resultate het getoon dat
lewendige La Sota entstof IgA produksie stimuleer en dus tot mukosale-immuniteit in
volstruiskuikens lei. In die tweede gedeelte van hierdie proefskrif is
volstruismikoplasmas geïsoleer en geïdentifiseer met behulp van 16S rRNA
geenopeenvolgingsbepalings. Hierdie volgordes het getoon dat drie unieke
mikoplasmas in volstruise voorkom wat filogeneties verskillend blyk te wees. Die 16S
rRNA geenopeenvolgings van die volstruismikoplasmas is gebruik vir die
ontwikkeling van spesifieke inleiers vir die PKR identifisering en diagnose van
mikoplasmainfeksies in volstruise.
Die laaste gedeelte van hierdie proefskrif fokus op voëlmalaria in die Afrika pikkewyn
en die bestuur van hierdie siekte gedurende rehabilitasie. Die 'South African
Foundation for the Conservation of Coastal Birds' (SANCCOB) is 'n seevoëlreddingsen
rehabilitasie-sentrum vir siek, beseerde en ge-oliede pikkewyne. Hierdie sentrum
het egter aansienlike vrektes as gevolg van voëlmalaria. In hierdie studie is 'n ELISA
ontwikkel vir die bepaling van natuurlike anti-Plasmodium antiliggaamvlakke van
pikkewyne by aankoms en tydens rehabilitasie by SANCCOB. Resultate het 'n
toename in anti-Plasmodium antiliggaamvlakke getoon na toelating wat nie beïnvloed
is deur olie nie. Hierdie toename kan toegeskryf word aan nuwe malariainfeksies en
nie 'n heruitbraak van bestaande infeksies nie wat daarop dui dat pikkewyne aan
voëlmalaria blootgestel word by die SANCCOB-sentrum.
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