الفهرس 
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Abstract
The emergence and rapid dissemination of carbapenemase‐producing Carbapenem ResistantEnterobacteriaceae (CRE) worldwide is a cause for concern.
Resistance to carbapenems has increased dramatically among Enterobacteriaceae.
Outbreaks of CRE, primarily Klebsiellapneumoniae, have been reported recently in several regions worldwide.
The aim of this study was directed to investigate the effect ofin-vitro efficacy of synergistic antibiotic combinations in treatmentof carbapenem resistant klebsiellaspps. in neonatal ICU in Beni-Suef university hospital and diagnosis of their main mechanisms of resistance by phenotypic and genotypic detection of (OXA-48, KPC, OMP-35 and OMP36)genes.
Totally, 228 clinical samples were cultured positive for Enterobacterieacea.Among theseisolates, theincidenceof Klebsiellaspps.was (120/228;52,6%) ,of which (100/120;83,3%)were CRK.
CRK isolates were collected from different specimens. The most common specimens were sputum (62/100;62%) followed by urine (22/100;22%) and blood(16/100;16%).
Hundred patients (100%) had history of recent antimicrobial treatment with; (15/100;15%)Quinolones, (58/100;58%)Carbapenems, (7/100;7%) B-lactams and (20/100;20%) other antibiotics.
Antibiotic susceptibility was performed for CRK isolates by disk diffusion method ,the resistance to Imipenem10 µg (IPM) was (100%) ,Meropenem10 µg (MEM) (100%)and to Ertapenem10 µg (ETP) (100%), while the resistance for beta –lactams was (100%), Aztreonam10 µg(100%),Amikacin30 µg (83/100; 83%),Ciprofloxacin 5 µg (95%) andGentamicin (86/100 ;86%).
Phynotypic screening of isolates for capabenemeasesusing :
1-Modified Hodge test for KPC detected 12(12%) isolates were positive for KPC , while 88(88%) were negative .
The MHT (Modified Hodge Test) detected 3 (3%) of blaKPC positive isolates, thus the sensitivity of the MHT was 18.3% and specificity was 89.3% in relation to PCR as gold standard.
2- IP/IPI E-Test for metallo B lactamases
Thirty isolates were taken randomlyand tested for Metallo B lactamases production by the E test and we found that (21/30;70%) isolates were positive,(4/30;13.3%) were negative and (5/30;16.7%) were undeterminable.
3-Boronic acid combination disc test (BACDT) for KPC and Amp-C
KPC and Amp-C were detected byBoronic acid combination disctest 62(62%) isolates were positive while 38(38%) were negative.
Conventional Polymerase chain reaction (PCR) for detection of OXA-48 gene, KPC gene, OMP 35 and OMP 36:
Frequency of OXA -48 gene was (14/100;14%) of CRK isolates positive, while (86/100;86%) were negative.KPC gene was positive in (16/100;16%)while (84/100;84%) were KPC negative.OMP-35 gene was positive in (18/100;18%) of CRKp isolates while (82/100;82%) were OMP-35 negative.OMP-36 gene was positive in (6/100;6%) ofCRKp isolates while (94/100;94%) were OMP-36 negative.
Synergy studies of antibiotics on CRK isolates was done by MIC and we found that 64% of klebsiella isolates were sensitive to GM+CIP combination therapy at different concentrations (4/4mg/ml, 2/2mg/ml, 8/8mg/ml and 16/16mg/ml),26% were sensitive to MEM+CIP at concentrations (0.5/2mg/ml ,1/4mg/ml, 2/8mg/ml and 4/16mg/ml),16% were sensitive to AK+1PM (8/0.5mg/ml and 128/8mg/ml) while only 4% were sensitive to other combinations (IPM+TZP ,IPM+MEM and IPM +OMP)and colistin alone.
Conclosions
Rapid detection of differenentgenes is of the most importance, since these MDR organisms have the potential to spread rapidly in hospital environments and cause nosocomial infections with high mortality rates.
Combined with Modified Hodge Test, which is not in agreement with PCR for detection of KPC, detection will be improved.
Molecular methods for detection of OXA-48, KPC, OMP-35 and OMP-36 producingKlebsiellaSpps.are the gold standard, however, they are expensive, time consuming, and require specialized equipment and expertise.
Also, OXA-48, KPC, OMP-35 and OMP-36 were circulating among K.pneumonie in our Beni-Suef university hospital (Neonatal ICU) in different numbers.
However, because of insufficient clinical information, we are currently unable to ascertain the exact mechanism of spread.
Combination therapy at gentamycin and ciprofloxacin may be effective in treatment of CRK at different concentrations.
Invitro data suggest that combination therapy can be effective even if the bacteria are resistant to the individual drugs, but the results vary greatly between studies. A better appreciation of the mechanisms of synergy would facilitate the understanding of results obtained and help to predict the effects of other antibiotic combinations.
Recommendations
We suggest that there is an urgent need for further studies to clearly map out the epidemiology and to conduct molecular characterization of carbapenemase producing isolates in healthcare facilities in the country to be aware of the emergence of these multidrug resistant isolates, as these are of significant public health concern both in the hospital and community setting.
Because resistance to carbapenems and other broad-spectrum beta-lactams is increasing, and because there is a lack of new antibiotics, it is urgent to explore the potential of combination therapy to enhance the antibacterial effects of available drugs.
Molecular detection of metallo b lactamases in our hospital is important due to high incidence by phynotypic detection.