الفهرس 
Microbial infectionOnly 14 pages are availabe for public view
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Abstract
Mixed infection is infection with more than one kind of organism at the
same time as in some abscesses, pneumonia, and infections of wounds.
Interspecies interaction between organisms is any process by which an organism
has an effect on an organism of a different species. Historically, interspecies
interactions have focused on growth-inhibitory interactions, yet a variety of
phenotypic outcomes other than antibiosis are possible, including changes in
metabolism (growth inhibition or stimulation, or production of new small
molecules) or morphological and developmental changes (alterations in cell
shape or morphology; production of biofilms or specialized processes such as
sporulation and germination).
The interactions between different species may be due to new roles for known
molecules such as peptidoglycan, antibiotics at subinhibitory concentrations or
signals used in signaling system crosstalk e.g. autoinducer-2 (a wellcharacterized quorum-sensing molecule) and some fatty alcohols and acids
where intraspecies small molecule signals may modulate the microbial
development of different species.
That is why the present study focused basically on studying the interactions
between certain microbial species isolated from mixed infection and how these
interactions affected their coexistence.
Therefore, clinical specimens showed mixed infections were collected (35 out of
142 collected clinical specimens with a percentage of about 25%). The recovered
isolates from mixed infection (72 out of 179 of total collected isolates with a
percentage of about 25 %) were collected from different clinical specimens in the
period of about one year of this study. Pus was the clinical specimen of the highest
prevalence (74%) of mixed infections in comparison to other clinical specimens.
Thirty three out of 35 clinical specimens with mixed infections (94.3%) harboured two microbial species while the other two specimens harboured three bacterial
species. Both Staphylococcus and Pseudomonas isolates coexisted in mixed
infection at the highest prevalence (42.8 %) compared to other coexisted bacterial
isolates.
Different Pseudomonas isolates that coexisted in mixed infection with
Staphylococcus isolates were assayed for their protease and lipase productivities
however, they showed no significant differences. Moreover, acylhomoserine
lactone (AHL) production as well as antibiogram analysis of the respective
isolates showed significant variation.
from the previous findings, two models of coexisting and Staphylococcus isolates
(SP12 and SP14 models) were selected for studying their interactions using
different physiological parameters. This selection was based on the results obtained
where the two Staphylococcus isolates, S12 and S14 were methicillin-resistant
(MRSA) and the two Pseudomonas isolates, P12 and P14 were identified as P.
aeruginosa however, they showed significant difference in their antibiogram
analysis as well as presence or absence of endogenous plasmids.
Although both Staphylococcus (S.) aureus isolates S12 and S14 were methicillin
resistant (exhibited cefoxitin resistance and shared the same resistance profiles to
two additional antimicrobial agents clindamycin and ceftazidime), they showed
different resistance profiles where S. aureus S12 showed resistance profile to
amikacin, cefoperazone, ciprofloxacin, piperacillin- tazobactam for which S. aureus
S14 was sensitive. Similarly, S. aureus S14 showed resistance to co-amoxiclav,
erythromycin and co-trimoxazole, the antimicrobial agents to which S. aureus S12
was sensitive.
In case of P. aeruginosa, although both isolates were resistant to clindamycin,
erythromycin, vancomycin, cefoxitin and co-trimoxazole, isolate P12 was resistant
to co-amoxiclav, azithromycin, cefipime, doxycycline, levofloxacin, cefoperazone,piperacillin-tazobactam, ceftazidime, cefpodoxime, sulbactam-ampicillin and
cefotaxime, the eleven antimicrobial agents to which isolate P14 was sensitive.
This antimicrobial profile of the coexisting microbial species in the two selected
models could represent good examples for studying their interaction. Results
showed that there was significant reductions in the viable count of both S. aureus
isolate S14 and P. aeruginosa isolate P14 when grown in co-culture as compared to
their growth in monocultures. While, there was no significant difference in the
growth of P. aeruginosa isolate P12 in monoculture and in co-culture with S.
aureus isolate S12. Moreover, there was no significant effect of different
physiological factors (incubation temperature and pH) on growth profile however,
optimum reduction effect of P14 on its coexisted S. aureus S14 was observed at
37ºC and initial pH 7.2 .
Furthermore, the culture supernatant of P. aeruginosa P14 (harboring no
plasmids) exerted a significant reduction effect on the biofilm formation (about
57% reduction) of the co-existing MRSA isolate (S14) however this effect was
not observed upon using the culture supernatant of P. aeruginosa P12 (harboring
plasmids) on the biofilm formation of the co-existing MRSA isolate (S12)
Quorum sensing controlling genes including acylhomoserine synthase (ahl), 2-heptyl-3-hydroxy-4(1H)-quinolone synthase (pqsH) and AraC family
transcription regulator (araC) of P. aeruginosa, particulary those involved in the
regulation and synthesis of major molecules of quorum sensing when coexisting
with Gram positive pathogens were detected using PCR and chromosomal DNA
as templates.
DNA sequencing and genetic analysis of the PCR products of the respective
genes were carried out. The quorum sensing controlling genes araC, PqsH and
ahl obtained in this study were submitted into the GenBank database under the
accession codes, KT693035, KT693034, KT693033, respectively. AraC was a
model of AraC transcription regulator with a conserved N-terminal arabinose binding domain and C-terminal H-T-H motive. PqsH was a model of putative 2-heptyl-3-hydroxy-4(1H)-quinolone synthase with a conserved domain of a NAD
(P)-binding Rossmann-like domain. LasI showed a conserved domain with the
acyl-homoserine-lactone synthase (LasI) of the protein family COG3916. The
open reading frames (ORFs) of the respective genes showed no mutation or
deviation in the predicted tertiary structures.
Domains and phylogenetic analysis as well as analysis of the predicated tertiary
structures respective gene products showed significant conservation (more than
80%) in the nucleotide/amino acid sequences with the respective homologous in
the GenBank database. Moreover, the the open reading frames of the respective
genes showed no mutation or any deviation on the entire genes. Therefore, to
confirm the difference in the inhibitory effects among the the two selected
models, plasmid transformation experiments were carried out followed by
testing the inhibitory effect of the culture supernatant of the obtained
transforamant P14 isolate.
Accordingly, plasmids extracted from P. aeruginosa clinical isolate P12 were used
to transform competent cells prepared from P. aeruginosa clinical isolate P14.
Results showed that only three plasmid bands out of six had been successfully
transformed into P. aeruginosa clinical isolate P14 host strain. Results showed the
culture supernatant of P. aeruginosa harbouring no endogenous plasmids (P14)
exerted a significant reduction effect on the biofilm formation of the co-existed
MRSA isolate (about 57% reduction) however, the culture supernatant of P.
aeruginosa harbouring endogenous plasmids (P12) showed an increase of the
biofilm formation of the co-existed MRSA isolate (about 5% increase). However,
plasmid acquisition significantly decreased the inhibitory effect of P. aeruginosa
isolate P14 on the biofilm formation of the co-existing MRSA isolate S14.