Bibliography / Thesis (PhD (Pathology. Medical Microbiology))--University of Stellenbosch, 2010. / ENGLISH ABSTRACT: Overview: Mycoplasmas and ureaplasmas are not routinely diagnosed and
are under researched in South Africa. Prevalence, population shifts especially
concerning genital flora and implications in infection or other conditions are
unknown. Information pertaining to Mycoplasma pneumoniae in respiratory
disease is similarly lacking. There is little information on antimicrobial
susceptibilities and resistance development against Sexually Transmitted
Infections (STI) syndromic management approaches.
Aims: a) Elucidate mycoplasmal and ureaplasmal prevalence and
contributing factors concerning cervical colonisation or preterm delivery in
conjunction with HIV and Chlamydia trachomatis b) Investigate prevalence of
M. pneumoniae in respiratory infections in conjunction with HIV,
Mycobacterium tuberculosis and Pneumocystis jiroveci. c) Determine
antimicrobial susceptibilities of mycoplasmas and ureaplasmas and analyse
resistance genes. d) Assess the inter-generic transfer potential of resistance
gene (tetM) between Ureaplasma spp. and Neisseria gonorrhea.
Genital setting: The prevalence of genital mycoplasmas, ureaplasmas and
Chlamydia on women attending their first prenatal visit, in conjunction with
preterm labour or HIV status was investigated. For preterm labour (2003), 199
women were monitored for preterm delivery (<37 weeks); for colonisation and
HIV (2005), 219 women were screened. Microbial detection was performed on
DNA extracted from endocervical swabs employing PCR techniques.
Colonisation was seen to be highest in the 14-20 year group from 2003. In
women aged ±21 years, co-colonisation was 13% although there was a shift
from co-colonisation with Mycoplasma hominis and Ureaplasma spp. in 2003
to other dual/triple combinations in 2005. Overall major trends from both
collection periods were that the prevalence of Ureaplasma spp. tended to be
higher in women ±26 years, whilst prevalence of C. trachomatis and M.
hominis were lower. No association was evident between colonisation with M.
hominis, U. urealyticum, Ureaplasma parvum and labour outcome. HIV status had no effect on the prevalence/co-colonisation of M. hominis, Ureaplasma
spp. or C. trachomatis.
Respiratory setting: Studies were conducted to determine the prevalence of
community acquired atypical pneumonias in adults (M. pneumoniae and P.
jiroveci) and neonates (mycoplasmas, ureaplasmas and Chlamydia
trachomatis) in order to improve treatment management programmes in the
Port Elizabeth region. Sputum specimens from 102 adult patients presenting
with pneumonia/symptoms of pneumonia admitted to hospitals were assessed
by PCR. Details of patient’s gender, age, HIV and Mycobacterium
tuberculosis status were provided by the hospitals. Women were seen to be at
high risk for community-acquired P. jiroveci colonisation. Overall, prevalence
of P. jiroveci was 52.9% (54/102 patients). P. jiroveci was mainly associated
with HIV (25/74) (P. jiroveci and HIV positive patients in patient sample for
which clinical data and HIV status was available) and co-infection with M.
tuberculosis was observed in 12 HIV cases and one HIV negative patient. No
DHPS (20) or DHFR (17) resistance associated mutations were found in P.
jiroveci. M. pneumoniae was detected in one patient. For prevalence studies
(2007-2008) on atypical pneumonia in neonates, 69 endotracheal aspirates
were obtained. PCR detection of M. hominis, U. urealyticum and C.
trachomatis was performed and U. parvum detected in two specimens.
Antibiotic susceptibilities and resistance genes: The following
investigations on clinical isolates of U. parvum and U. urealyticum were
conducted (i) antibiotic susceptibility profiles, (ii) detection of drug target gene
mutations, or gene acquisitions and (iii) inter-generic resistance gene transfer
potential to Neisseria gonorrhoeae. Culture techniques applied to 132
endocervical specimens provided 66 Ureaplasma cultures (35 U. parvum, 9
U. urealyticum, 22 U. parvum + U. urealyticum). MIC determinations to
ofloxacin, erythromycin, tetracycline, doxycycline, azithromycin and josamycin
were performed. Thirty-seven ureaplasma cultures were fully susceptible to all
antibiotics tested; 21 showed intermediate resistance to erythromycin,
azithromycin and ofloxacin; while seven were resistant to tetracycline, three of
which were also resistant to doxycycline and one also resistant to azithromycin. Concerning ofloxacin resistance directed at quinolone
resistance determining regions, a substitution of Ser83Leu in ParC was
demonstrated in one intermediately-resistant Ureaplasma (MIC 4 µg/ml) while
a triple substitution of Asp112Glu in GyrA along with Ala125Thr and
Ala136Thr in ParC was found in six further intermediately-resistant strains. No
mutations were found in strains with MICs 1 µg/ml. No mutations were
detected in 23S rRNA operons, L4 or L22 proteins. TetM and int-Tn genes
were found in seven tetracycline-resistant strains. On screening 59
tetracycline-susceptible and -intermediate strains, eleven whilst possessing
an int-Tn gene lacked a large region of tetM and 48 only contained small
regions of tetM. The tetM genes of the seven tetracycline-resistant strains
were sequenced and comparisons performed against GenBank sequences of
Neisseria gonorrhoeae, Streptococcus pneumoniae and U. urealyticum. For
five strains tetM was seen to be highly mosaic in structure containing regions
that were similar to those of the GenBank strains and others that were unique.
In the tetM leader region, four hot spot recombination sites were identified that
could certainly influence the formation of the mosaic structures, upstream
insertion sequences/open reading frames and transposon regions that
regulate expression. On characterising the int-Tn genes of the seven
tetracycline-resistant strains, three types were present indicating transposons
from different origins had integrated into ureaplasma genomes. Reciprocal
tetracycline resistance gene transfer between ureaplasmas and N.
gonorrhoeae were unsuccessful. However, low-level tetracycline resistance
(MICs 4-8 µg/ml) was transferred to a U. parvum recipient from one U.
urealyticum and three U. parvum donors that carried tetM with MICs 16-64
µg/ml. On tetM PCR analysis, tetM was not detected in the transformants.
Conclusions: The importance of genital mycoplasmas, ureaplasmas and C.
trachomatis in long term aetiologies requires further investigations, certainly in
relation with syndromic management regimens that fail to reduce colonisation
rates. The high prevalence of P. jiroveci, the presence of M. pneumoniae in
cases of pneumonia and detection of U. parvum in two cases of neonatal
pneumonia investigated emphasises that in the absence of definitive
diagnoses, it is crucial to monitor treatment responses carefully, especially when first line antibiotic preferences are ß-lactams, in order to ensure
adequate and informed delivery of medical care. The finding of transposon
and/or tetM regions in all ureaplasmas investigated with or without full
expression of tetracycline resistance, in conjunction with tetM gene diversity,
certainly places ureaplasmas strongly in the picture for intra- and inter-generic
exchange of antibiotic resistance genes. / AFRIKAANSE OPSOMMING: Oorsig: Mikoplasma en ureaplasma word nie roetineweg gediagnoseer nie
en in Suid Afrika is nog min navorsing daaroor gedoen. Prevalensie,
populasie verskuiwings, veral in genital flora, en die impliksies van infeksie en
ander toestande is onbekend. Inligting rakende Mycoplasma pneumoniae in
respiratoriese siekte is ook gebrekkig. Daar is min inligting beskikbaar
rakende die antimikrobiale vatbaarheid en die ontwikkeling van
weerstandigheid gesien teen die benadering tot sindromiese hantering van
seksueel oordraagbare siektes.
Doelwitte: a) Om inligting te verskaf oor die prevalensie van mikoplasma en
ureaplasma en bydraende faktore betreffende voortydige kraam tesame met
MIV en Chlamydia trachomatis. b) Ondersoek van die prevalensie van M.
pneumoniae in respiratoriese infeksies tesame met MIV, Mycobacterium
tuberculosis en Pneumocystis jiroveci. c) Bepaling van die antimikrobiale
vatbaarheid van mikoplasma en ureaplasma en analisevan weerstandigheids
gene. d) Bereken die inter-genetiese oordrag potensiaal van
weerstandigheids gene (tetM) tussen Ureaplasma spp. en Naisseria
gonorrhoeae.
Genitale omgewing: Die prevalensie van genitale mikoplasma, ureaplasma
en Chlamydia in vroue tydens hul eerste prenatale besoek, tesame met
vroegtydige kraam en MIV status is ondersoek. In voortydige kraam (2003), is
199 vroue gemonitor vir voortydige kraam (<37 weke); vir kolonisasie en MIV
(2005), is 219 vroue getoets. Mikrobiale toetsing is gedoen deur DNS te win
vanaf endoservikale deppers met PKR tegnieke. Kolonisasie was die hoogste
in die ouderdomsgroep 14.20 jaar, in 2003. In vroue van ±21 jaar was medekolonisasie
13% alhoewel daar en verskuiwing was van mede-kolonisasie met
Mycoplasma hominis en Ureaplasma spp. in 2003 tot ander dubbel/trippel
kombinasies in 2005. Die oorkoepelende tendens in altwee die tydperke van
waarneming was dat die prevalensie van Ureoplasma spp. geneig was om
hoër te wees in vroue ±26 jaar, terwyl prevalensie van C. trachomatis en M.
hominis laer was. Geen assosiasie kon getoon word tussen koloniesasie met M. hominis, U. urealyticum, Ureaplasma parvum en uitkoms van kraam nie.
MIV status het geen effek gehad op die prevalensie/mede-kolonisasie van M.
hominis, Ureaplasma spp. of C. Trachomatis nie.
Respiratories: Studies is gedoen om die prevalensie van gemeenskaps
verworwe atipiese pneumonie in volwassenes (M. pneumoniae en P. jiroveci)
en neonate (mikoplasma, ureaplasma en Chlamydia trachomatis) te bepaal
om behandeling en hantering programme in die Port Elizabeth area te
verbeter. Sputum monsters van 102 volwasse pasiënte wat presenteer het
met pneumonie of simptome van pneumonie en wat tot hospitale toegelaat
was, is ontleed. Besonderhede van die pasiënte se geslag, ouderdom, MIV en
Mycobacterium tuberculosis status is deur die hospitale verskaf. PKR is
gedoen met inleiers gerig teen die volgende gene: P. jiroveci vir die aantoning
van mitokondriale groot subeenheid RNS en vir die analise van mutasies vir
ko-trimoksasool weerstandigheid dihydropteroaat sintetase (DHPS) en
dihydrofolaat reduktase (DHFR); M. pneumoniae vir die aantoning van P1
adhesien en 16S rRNS. Vroue het ‘n hoë risiko vir gemeenskapsverworwe P.
jiroveci kolonisasie gehad. In die algemeen was die prevalensie van P.
jiroveci 52.9% (54/102 pasiënte). P. jiroveci was hoofsaaklik geassosieerd
met MIV (25/74) (P. jiroveci en MIV positiewe pasiënte in die pasiënt monster
waarvoor daar kliniese data en MIV status bekend was) en mede-infeksie met
M. tuberculosis is gesien in 12 MIV gevalle en een MIV negatiewe pasiënt.
Geen DHPS (20) of DHFR (17) weerstandigheids geassosieerde mutasies is
gevind in P. Jiroveci nie. M. pneumoniae was aangetoon in een pasiënt. Vir
prevalensie studies (2007-2008) op atipiese pneumonie in neonate is 69
endotrageale aspirate verkry. PKR toetsing vir M. hominis, U. urealyticum en
C. trachomatis is gedoen met ‘primers’ soos voorheen gepubliseer.
Ureaplasma parvum is aangetoon in twee neonate met PKR met negatiewe
kultuur resultate.
Antibiotika sensitiwiteite en weerstandigheids gene: Die volgende toetse
is gedoen op kliniese isolate van U. parvum en U. urealyticum (i) antibiotika
sensitiwiteits profiele, (ii) aantoning van teiken geen mutasies, of geen
aanwinste en (iii) potensiaal vir inter-generiese weerstandigheids geen oordrag na Neisseria gonorrhoeae. Kultuur tegnieke toegepas op 132
endoservikale monsters het 66 Ureaplasma kulture gelewer (35 U. parvum, 9
U. urealyticum, 22 U. parvum + U. urealyticum). MIK bepaling vir ofloksasien,
eritromisien, tetrasiklien, doksisiklien, azitromisien en josamisien is gedoen.
Sewe-en-dertig kulture was ten volle sensitief vir alle antibiotika wat getoets
is; een-en twintig het intermediere weerstandigheid teenoor eritromisien,
azitromisien en ofloksasien getoon, terwyl sewe weerstandig was vir
tetrasiklien, drie daarvan was ook weerstandig vir doksisiklien. Wat betref
ofloksasien weerstandigheid gemik teen kwinoloon weerstandigheids
bepalende gebiede, is vervanging van Ser83Leu in ParC gedemonstreer in
een intermedier weerstandige Ureaplasma (MIK 4 µml) terwyl en trippel
vervanging van Asp112Glu in GyrA saam met Ala125Thr en Ala136Thr in
ParC gevind is in ses ander intermedier weerstandige stamme. Geen
mutasies is gevind in stamme met MIKs van MICs 1 µg/ml nie. Geeneen van
die ureaplasma was weerstandig vir eritromisien/azitromisien nie en geen
mutasies is gevind in 23S rRNA operons , L4 of L22 proteine nie. TetM en int-
Tn gene is gevind in sewe tetrasiklien weerstandige stamme. 58 Tetrasiklien
sensitiewe en .intermediere stamme is getoets, waarvan elf en int-Tn geen
gekort het sowel as en groot deel van tetM, terwyl 48 slegs klein dele van
TetM bevat het. Die tetM gene van die sewe tetrasiklein-werstandige stamme
se geenvolgorde is bepaal en vergelykings is getref teenoor die GenBank
volgordes van Neisseria gonorrhoeae, Streptococcus pneumoniae en U.
urealyticum. In vyf stamme is gevind dat die tetM geen hoogs mosaiek in
struktuur was met areas wat ooreenstem met die in GenBank stamme, en
ander areas wat uniek is. In die tetM leier area, is vier ehot spot f
herkombinasie areas geidentifiseer wat sekerlik die vorming van die mosaiiek
strukture kon beinvloed, asook transposon areas wat geenuitdrukking bepaal.
Met karakterisering van die int-Tn gene van die sewe tetrasikleinweerstandlige
stamme, was drie tipes teenwoordig waarin transposons vanaf
verskillende oorsprong aangedui was, geintegreerd met die ureaplama
genome. Resiprokale tetrasiklien weerstandigheids geen oordrag tussen
ureaplasma en n. gonorrhoea was nie suksesvol nie. Lae-vlak tetrasiklien
weerstandigheid (MIK fs van 4 . 8 µg/ml) is wel suksesvol oorgedra na en U.
parvum ontvanger vanaf een U. urealyticum en drie U. parvum ontvangers wat tetM gedra het met MIKs van 16-64 µg/ml. Met die analise van tetM met
PKR, kon tetM nie aangetoon word in die transformante nie.
Gevolgtrekkings: Die belang van genitale mykoplasma, ureaplasma en C.
trachomatis in langtermyn etologie benodig verdere ondersoek, veral in die lig
van die sindromiese behandeling regimes wat nie kolonisasie verminder nie.
Die hoe prevalensie van P. jiroveci, die teenwoordigheid van M. pneumoniae
in gevalle van pneumonie en die aantoning van U. parvum in twee gevalle van
neonatale pneumonie benadruk dat, in die afwesigheid van en definitiewe
diagnose, dit noodsaaklik is om respons tot behandeling sorgvuldig te
moniteer, veral indien die eerste lyn antibiotika keuse ß-laktam antimikrobiale
middels of kefalosporiene is, sodat behoorlike en ingeligde gesondheidsorg
gelewer kan word. Die bevinding van transposon en/of tetM gebiede in alle
ureaplasma wat ondersoek is met of sonder volle uitdrukking van tetrasiklien
weerstandigheid, in samehang met tetM diversiteit, plaas verseker
ureaplasma sterk in die prentjie vir intra- en inter-generiese uitruiling van
antibiotika weerstandigheids gene. / Nelson Mandela Metropolitan University / National Research Foundation (NRF Thuthuka) / Medical Research Council
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/5430 |
| Date | 12 1900 |
| Creators | Govender, Sharlene |
| Contributors | Chalkley, Lynda J., Wasserman, Elizabeth, University of Stellenbosch. Faculty of Health Sciences. Dept. of Pathology. Medical Microbiology. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | xx, 148 p. : ill. |
| Rights | University of Stellenbosch |
Page generated in 0.0042 seconds