Transcriptional Response of Multidrug-Resistant Klebsiella pneumoniae Clinical Isolates to Ciprofloxacin Stress.

BACKGROUND: The term "persisters" refers to a small bacterial population that persists during treatment with high antibiotic concentration or dose in the absence of genetic resistance. The present study was designed to investigate the transcriptional response in indigenous Klebsiella pneumoniae under the ciprofloxacin stress. METHODS: Isolation and identification of K. pneumoniae were carried out through standard microbiological protocols. The characterization of quinolone resistance was performed by estimating the quinolone susceptibility testing, MIC estimation, and detecting the QRDR and PMQR. Transcriptional response of the isolates to ciprofloxacin was determined using qPCR. RESULTS: Among 34 isolates, 23 (67%) were resistant to ciprofloxacin. Both QRDR (gyrA and gyrB) and PMQR (qnrA, qnrB, and qnrS) were detected in the isolates, and all were found resistant to ciprofloxacin. The mRNA levels of both mutS and euTu under the influence of ciprofloxacin were significantly increased. On ciprofloxacin exposure, the mRNA levels of the DNA damage response element (mutS) were raised in a time-dependent fashion. K. pneumoniae showed high-level resistance to ciprofloxacin in the presence of mutations in QRDR and PMQR genes. CONCLUSION: The transcriptional response revealed the upregulation of DNA repair and protein folding elements (mutS and euTu) in ciprofloxacin stress and delayed cell division. The ciprofloxacin was found to trigger various stress responses in a time- and concentration-dependent manner.

Molecular basis of quinolone resistance in clinical isolates of Klebsiella pneumoniae from Pakistan.

Antibiotic resistant Klebsiella pneumoniae, is associated with various nosocomial infections that are difficult to treat. This study is designed to find out the patterns of resistance against commonly used antibiotics in K. pneumoniae clinical isolates with special attention to fluoroquinolones. A total number of 200 K. pneumoniae clinical isolates collected from various tertiary care hospitals of Punjab, Pakistan for a span of 1 year were investigated. Isolates were identified biochemically and genetically using VITEK® system and species-specific PCR, respectively. Antibiogram of isolates was studied by using disc diffusion and broth micro-dilution assays. Highest infection of K. pneumoniae detected in urinary tract (43%) followed by respiratory tract (25.5%). Most of the isolates displayed strong resistance against ampicillin, cefotetan, tazobactam, cefuroxime, cefixime, ceftriaxone, ampicillin-sulbactam imipenem, meropenem, ciprofloxacin and moxifloxacin, while sensetive to cefotaxime. Chromosoaml mutation was deteted in gyrA gene, gyrA harbors a strong mutation which provides resistance against ciprofloxacin by substituting Ser83→Ile. However, no mutation was detected in gyrB gene. Moreover, qnrB1 plasmid born resistant gene was only detected among qnrA, qnrB and qnrS. The story depicts an alarming situation of antibiotic resistance among K. pneumoniae associated with various nosocomial infections.

The First blaKPC Harboring Klebsiella pneumoniae ST258 Strain Isolated in Pakistan.

Klebsiella pneumoniae is an important pathogen that causes pneumonia and bloodstream infections, especially in neonates and intensive care patients. The carbapenems remain an important therapeutic option for clinicians, particularly against cephalosporin-resistant Gram-negative pathogens. This increased use of carbapenems at clinics has resulted in the evolution and spread of carbapenem-resistant K. pneumoniae. In this study, we isolated six bla K. pneumoniae carbapenemase (KPC)-producing strains belonging to sequence type 258 (ST258) from clinical, environmental, and veterinary sources. Antibiotic susceptibility was performed on these isolates and the genes responsible for extended-spectrum beta-lactamases and metallo-beta-lactamase production were screened. The molecular typing was done using multilocus sequence typing. Isolated strains were resistant to various antibiotic classes, including carbapenems, and carried the carbapenem-resistant gene, blaKPC. All strains were susceptible to tigecycline and colistin. This is the first report detecting K. pneumoniae ST258 strains in Pakistan.

Emergence of bla NDM-1 Harboring Klebsiella pneumoniae ST29 and ST11 in Veterinary Settings and Waste of Pakistan.

INTRODUCTION: Intense livestock farming practices enforcing the farmers to use antibiotics as food supplements on a routine basis. Aberrant use of antibiotics is associated with the emergence of antibiotics resistance and resistant superbugs. Keeping in view the current scenario, the present study was designed for the first time from Pakistan with a specific aim to estimate the prevalence of the carbapenem-resistant Klebsiella pneumoniae in veterinary settings and the waste in Pakistan. METHODS: A total of 138 samples from various veterinary sources were collected by employing a nonprobability sampling technique. Isolation and phenotypic identification of carbapenem-resistant K. pneumoniae were performed according to the CLSI standard. Molecular detection of various antibiotic resistance genes (ARGs) was done through PCR by using specific primers against each ARG. According to the pasture scheme, the multilocus sequence typing (MLST) was performed to characterize the K. pneumoniae sequence types (STs). RESULTS: According to the results of the study, overall 9.4% (13/138) isolates were confirmed carbapenem-resistant K. pneumoniae. Among various carbapenem ARGs particularly, the bla NDM-1 was found in 92.3% (12/13) isolates followed by bla OXA-48 84.6% (11/13). MLST results revealed that overall 3 STs were found in the study which includes ST29, ST11, and ST258. Taking together, this is the first study to our best knowledge which demonstrated the prevalence of carbapenem-resistant K. pneumoniae and its various STs prevalent in veterinary settings and the waste of Pakistan. CONCLUSION: Based on the above-mentioned facts, we suggested that veterinary settings and waste are the potential source and reservoir of carbapenem-resistant K. pneumoniae, which may be disseminated to the environment and ultimately can affect the public and companion livestock health.

CRISPR-Cas system: a potential alternative tool to cope antibiotic resistance.

Antibiotic exposure leads to massive selective pressures that initiate the emergence and spread of antibiotic resistance in commensal and pathogenic bacteria. The slow process of developing new antibiotics makes this approach counterintuitive for combatting the rapid emergence of new antibiotic resistant pathogens. Therefore, alternative approaches such as, the development of nucleic acid-based anti-bacterial treatments, anti-bacterial peptides, bacteriocins, anti-virulence compounds and bacteriophage therapies should be exploited to cope infections caused by resistant superbugs. In this editorial, we discuss how the newly popular CRISPR-Cas system has been applied to combat antibiotic resistance.