الفهرس 
Chronic obstructive pulmonary diseaseOnly 14 pages are availabe for public view
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Abstract
Chronic Obstructive Pulmonary disease (COPD) is a common preventable and treatable disease, characterized by persistent airflow limitation that is usually progressive and associated with an enhanced chronic inflammatory response in the airways and the lung to noxious particles or gases. Exacerbations and comorbidities contribute to the overall severity in individual patients.
Acute exacerbation of COPD (AECOPD) is the most frequently triggered by bacterial infection as infection of patients with COPD with Gram-negative bacteria represents a problem of a great clinical relevance, because the antibiotic resistance mechanisms in these bacteria are increasingly being reported worldwide.
Dissemination of quinolone resistance among Gram-negative pathogens such as K. pneumoniae poses a considerable threat to public health and is a growing problem worldwide. Plasmid-mediated quinolone resistance (PMQR) can arise from the expression of proteins encoded by the qnrA, -B, and -S genes that are able to protect the DNA gyrase
The aim of this work was to determine the incidence of aerobic bacteria isolated from patients at Menoufia University Hospitals and to determine the resistant pattern of K. pneumoniae isolated from sputum of patients admitted to chest departments and clinic at Menoufia University Hospitals (MUHs) and to detect quinolone resistance of the isolated K.pneumoniae and to determine minimum inhibitory concentration. Also, to detect the ability of K.pneumoniae to form biofilm in vitro and its relation with antimicrobial resistance pattern.
This study was conducted over the period from November 2016 to October 2017 and its protocol was approved by the local ethics committee of the Menoufia University. This study included three groups; group (1), 25 patients with stable COPD were presented to Chest Out patient’s Clinic at Menoufia University Hospital [MUH} (without history of exacerbations in the last three months), group (2), 50 patients with AECOPD and group (3), 25 patients with different chest diseases as a control group admitted to Chest departments at MUH. All the selected patients were subjected to full history taking and thorough clinical examination. The Global Initiative for chronic Obstructive Lung Disease (GOLD) classification was used for evaluating disease severity.
One hundred sputum samples were evaluated based on Bartlett’s grading system. The specimens were processed according to standard microbiological methods. The grown K. pneumoniae isolates were identified by standard methods and Vitek-2 compact system. Antimicrobial susceptibility was determined by disk diffusion method and results were interpreted according to the Clinical and Laboratory Standard Institute (CLSI) guidelines. K. pneumoniae isolates were tested for MIC of ciprofloxacin and levofloxacin by agar dilution method. All clinical isolates of K. pneumoniae isolates were examined for the presence of PMQR genes by multiplex-PCR.
The results obtained in this study revealed that from 100 acceptable specimens, 99 isolates were obtained. 2 specimens showed mixed cultures (2 isolates for each) while 3 specimens showed no bacterial growth.
The most frequent isolated organism in AECOPD was Klebsiella spp. representing 27.3% of all isolates, followed by S. aureus 12.1% and P aeureginosa 9.1%. VITEK2 compact system showed that K. pneumoniae was the predominant Klebsiella species (94.3%), while only two isolates were identified as K. oxytoca (5.7%).
Studying the risk factors associated with AECOPD revealed that AECOPD was more common among old patients (P<0.001). Males (82.0%) were more affected than females (P<0.001).
Smoking was a high statistically significant risk factor for AECOPD (P<0.001). Higher rates of AECOPD were found in patients with duration of hospital stay more than 14 days (P=0.005). Higher rates of AECOPD were found in patients with a history of antibiotic administration (p<0.0001). AECOPD were found more in patients with history of associated co-morbidities (76%) compared to those with no history of associated co-morbidities (p= 0.006).
A highly significant relation was found between the severity of COPD (according to GOLD classifications) and K. pneumoniae isolation. In group I, K. pneumoniae was isolated only from patients with very severe stage (2/27, 7.4%).while, in group II, K. pneumoniae was isolated in all stages, but the highest rate was found in severe stage of AECOPD (70.4%).
Antimicrobial susceptibility tests using disk diffusion method revealed that all K. pneumoniae isolates were markedly resistant to amoxicillin (28/33, 84.9%), amoxicillin/clavulanic acid (25/33 75.8%) followed by ciprofloxacin (21/33, 63.6%) and levofloxacin (19/33. 57.6%).
Regarding biofilm detection, about 78.8% and 84.8% of K. pneumoniae isolates were biofilm producers by CRA and MCRA method repectively with no statistical difference.
The results obtained in this study revealed that, regarding the screening tests, the disk diffusion method revealed that, 63.6% of K.pneumoniae isolates were ciprofloxacin-resistant compared to 54.5% of K. pneumoniae isolates were ciprofloxacin -resistant isolates detected by MIC method.
For levofloxacin, 19/33 (57.6%) isolates were resistant to levofloxacin by disk diffusion method compared to 18/33 (54.5%) isolates by MIC method.
Risk factors for quinolone resistance, the highest rate of quinolones resistant K. pneumoniae by MIC was found in patients with age group more than 56 years (72.2%) than age group less than 56 years (27.8%) without significant difference (P>0.05). Also, quinolones resistant K. pneumoniae were isolated from males (66.7%) more than females (33.3%) with no statistical significant difference (P>0.05).
There was a statistically significant difference between quinolones resistant K. pneumoniae and a history of associated co-morbidities (77.8%, p=0.003). No significant difference regarding previous mechanical ventilation (6.7%, p=0.18)
Also, resistant K. pneumoniae were more isolated from smoking patients (72.2%) than others with no statistical significant difference (P=0.70).
Previous admission to hospital was a significant factor for isolation of quinolones resistant K. pneumoniae (P=0.009). The highest rate of resistant K. pneumoniae was found in patients with duration of hospital stay more than 14 days (61.5%), while the lowest rate with duration of hospital stay less than 7 days (7.7%) with statistical significant difference (P=0.02).
Quinolones resistant K. pneumoniae was more frequently isolated from patients with history of antibiotic administration (72.2%) more than those without history of antibiotic administration (27.8%) but without statistical significant difference (P=0.26).
A significant relationship was detected between biofilm production and multidrug resistance, where 89.3%, 75.0% and 67.9% biofilm forming K. pneumoniae strains were markedly resistant to amoxicillin/clavulanic acid, ciprofloxacin and levofloxacin respectively
Regarding PMQR genes in K. pneumoniae isolates, 36.4% of K. pneumoniae isolates were positive for qnrA1-6 gene, 57.6% were positive for qnrB1-3, 5, 6, 8 gene and 18.2% were positive for qnrS1-2 gene but no isolate had qnrB4. Out of (33) K. pneumoniae isolates, 15/33 K. pneumoniae isolates were susceptible to both ciprofloxacin and levofloxacin by MIC but only one isolate had qnrB1-3, 5, 6, 8 gene by multiplex PCR. In 18/33 resistant K. pneumoniae isolates to both ciprofloxacin and levofloxacin by MIC, qnrB1-3, 5, 6, 8 (as a single gene) was present only in 6 K. pneumoniae isolates by multiplex PCR. While combined genes qnrA1-6 & qnrB1-3, 5, 6, 8 and qnrA1-6 & qnrB1-3, 5, 6, 8 & qnrS1-2 were detected in 6 K. pneumoniae isolates by multiplex PCR.
Considering PCR as the gold standard, the sensitivity of MIC was 94.7%, specificity 100% and diagnostic accuracy 96.97% in relation to multiplex PCR for detection of quinolone resistance among K. pneumoniae isolates.