Genetic Characterization of Carbapenem Resistant Acinetobacter baumannii in Tertiary care settings of Lahore, Pakistan

Background: Acinetobacter baumannii is major cause of ventilator associated pneumoniae (VAP) as it is an opportunistic nosocomial organism. The current study was to find out the antibiotic resistance pattern of Acinetobacter baumannii, its phenotype and the genetic characterization of Metallo-β-Lactamase (MBL) genes that are responsible for carbapenem resistance.Methods: One hundred and fifty Carbapenem resistant Acinetobacter baumannii (CRAB) specimens were isolated and PCR amplification of organism specific bla-OXA-51gene was performed and antibiotic susceptibility was checked. Phenotypic susceptibility analysis was performed by Modified Hodge Test (MHT) and Imipenem-EDTA Double Disc Synergy Test (IMP-EDTA DDST). The carbapenemases and MBL producing genes were amplified by PCR.Results: CRAB showed high resistance against piperacillin/tazobactam (99.3%), cefepime and ceftazidime (99.3% each), amikacin (91.3%), ciprofloxacin (96.7%) and levofloxacin (96.7%). Only one isolate showed resistance to colistin. The isolates positive for both MHT and DDST (n=70) were further characterized to detect metallo-β-lactamase genes. Molecular characterization revealed the presence of bla-OXA-51 gene in all tested isolates (100%) followed by bla-VIM 89%, bla-OXA-23 64%, respectively and so on. Few genes coexisted with each other including bla VIM, bla OXA 23, bla OXA 51 and bla NDM-1.  None of the isolate was found positive for bla-IMP gene.Conclusion: It is concluded that CRAB isolates exhibited a high rate of resistance towards antimicrobials because of the presence of drug hydrolyzing enzymes, carbapenemases and MBLs. This is among the rare study reported recently indicating CRAB isolates co-harboring many resistant genes are very difficult to treat. There is a dire need to develop novel antibiotics against resistant A. baumannii to minimize its prevalence. Moreover, it is recommended that colistin treatment in the clinical settings should be continuously monitored in order to prevent the development of resistance

Microbial profiling and risk factors assessment for Otitis Media and Otitis Externa

Background: Otitis media and otitis externa are common otological manifestations in all generations especially in children. There is lack of accurate identification of the causative agent and thus poor diagnosis for such infections. Therefore, it leads to permanent anatomical disabilities including poor speech and defects in balancing and hearing. The study was conducted to isolate, characterize and identify the microbes causing otitis media and otitis externa. Methods: A total of 250 patients having otitis media and otitis externa were enrolled in the study from March 2011 to October 2011. All patients were examined through clinical examination and detailed history was collected. Pus samples from the discharging ears were plated on MacConkey’s, Chocolate and Blood agar for 24 to 48 hours. Isolates were identified on the basis of morphology, staining reactions and various biochemical tests. Results: In this study, only 6% cases yielded no growth, 14% yielded mixed cultures while 80% cases yielded pure cultures. The presumptive diagnosis for ear swabbing was otitis media (76%) and otitis externa (24%). The most common bacterial isolates obtained were Staphylococcus aureus (43.3%) followed byPseudomonas aeruginosa (25%) in the diagnosed cases of otitis media. While for the cases of otitis externa,Pseudomonas aeruginosa was the predominant organism with 52.2%. Infection of otitis media was most common among children and the persons having low socioeconomic conditions. Conclusion: Pseudomonas aeruginosa was identified as the principal pathogen followed by Staphylococcus aureus. To circumvent the painful effects of acute and chronic ear infections, an accurate microbial profiling may play pivotal role

Augmented cytotoxic, mutagenic and genotoxic response triggered by carvedilol and celecoxib combinations

It is understood that drugs regardless of their order of administration can exhibit drug interactions. Established on the fact that treatment of hypertension may last for decades and prolong usage of multiple drug regimen may induce substantial pathophysiological changes. Hence, This study was designed to evaluate the possible synergistic toxic effects of anti-hypertensive (carvedilol), and anti-inflammatory drug (celecoxib) alone and in combinations. Well-established MTT assay, Single Cell Gel Electrophoresis (SCGE) and Ames assay were employed to evaluate the toxicity at cellular level. Results from MTT assay on Vero cell line revealed that drug combinations have more pronounced anti-proliferative activity with combine IC50 value of 13.7:47.8 µg/mL. Likewise, exposure of peripheral blood mononuclear cells with drug combinations revealed significant (

Prevalence of Black Quarter in Association with Patho-Morphological Alterations in Affected Tissues at Different Zones of Punjab-Pakistan

Background: Blackleg or Black Quarter (BQ) is a serious bacterial disease caused by Clostridium chauvoei. It causes edematous and gaseous changes in skeletal muscles of animals. The study was designed to find prevalence of BQ in 6 districts of Punjab, Pakistan from June 2018 to June 2019.Methods: Animals were randomly monitored, and selected in each union council regardless of age, sex and species, but lameness and hyperthermia were the selection parameters. A proforma was used to record the experimental readings of each animal. A total of 1500 swab samples from bovines (cattle and buffaloes) were processed through conventional PCR and culture sensitivity tests to examine the comparative diagnostic efficacy and suitability of the test.Results: Sensitivity and specificity of culture tests were 87.6% and 34.19% respectively, whereas PCR demonstrated 81% and 47.3% results for the said aspects, respectively. Alpha toxin gene (Ccta) was amplified at 52.2oC yielding an amplicon size of 1400 base pairs. Area wise and season wise prevalence of each animal was also determined. The prevalence of BQ in cattle was significantly higher (P 0.05) in Lahore (0%) and Nankana Sahib (8%) which belonged to the canal irrigated zones.Conclusion: There was higher prevalence of BQ in different areas of Punjab, Pakistan that show the higher risk of disease transmission. A systematic surveillance system is essential to regularly monitor the disease incidence and prevalence in these areas of Pakistan.Keywords: Black Quarter; Prevalence; Pathological; Ruminants;  Zone

Molecular characterization of ochratoxin a producing indigenous Aspergillus strains from poultry feed in Pakistan

Ochratoxin A (OTA) is nephrocarcinogenic and immunosuppressive toxin and OTA producing molds contaminate the food crops. Isolation and identification of ochratoxin producing fungi was carried out from poultry feed samples (n=120) followed by preliminary confirmation through macroscopic and microscopic characteristics. Purified fungal isolates identified as Aspergillus 1842(91.68%) followed by Penicillium 91 (4.53%), Mucor 52 (2.58), Alternaria 7 (0.35%), Cladosporium 6 (0.29%), Fusarium 4 (0.199%) and unidentified (07). OTA production was confirmed through thin layer chromatography (TLC) followed by high performance liquid chromatography (HPLC). Only 41 isolates (2.22%) out of 1842 Aspergillus isolates were able to produce toxin. At genetic level, characterization was performed through polymerase chain reaction (PCR) using species specific gene primers. From 41 isolates 27, 9 and 5 were characterized as Aspergillus terreus, Aspergillus parasiticus, and Aspergillus ochraceus, respectively. Physical and chemical factors were optimized for OTA production. Under the effect of 37 °C temperature and 7.5 pH of Sabouraud dextrose broth (SDB) medium, higher toxin (969.45±.03 μg/mL) production was observed from ASPO-6 isolate. ASPO-4 isolate produce higher toxin amount in SDB medium with supplementation of maize 5%, wheat 1% and rice 3%. OTA stability was determined by adjusting standard concentration of 100 μg/mL in organic solvents (chloroform, acetonitrile and methanol) and organic solids. Least percentage log reduction in OTA concentration and stability of OTA was observed in opaque vials with chloroform and sucrose and transparent vials with sucrose after 6 months. OTA can be used as indigenous standard for identification of OTA from field samples

Activity of plant essential oils against antibiotic resistant Enterococcus faecalis isolated from diarrheic children

Activity of plant essential oils and their fractions was evaluated against characterized isolates of antibiotic resistant Enterococcus faecalis recovered from diarrheic children. The isolates were confirmed by polymerase chain reaction (PCR) targeting 16S rRNA gene amplification followed by nucleotide sequencing and accession numbers retrieved were MW349990.1, MW349859.1, MW332122.1, MW356805.1, MW349975.1, MW349988.1, MW356790.1, MW356244.1, MW341593.1 and MW332549.1. These isolates were screened for antibiotic susceptibility to a wide range of antibiotic groups and mean zone of inhibition (ZOI) of all antibiotics were recorded. Antibacterial activity of plant essential oils (n=05) was checked against three antibiotic resistant isolates of E. faecalis. Three plant essential oils having higher ZOI including Cinnamomum verum, Syzygium aromaticum and Nigella sativa were used against resistant E. faecalis isolates to determine minimum inhibitory concentration (MIC). The lowest MIC observed was of S. aromaticum (11.39±3.94 mg mL-1). The S. aromaticum n-hexane plus chloroform fraction displayed higher mean ZOI (16.67±2.51 mm), while the lowest MIC was of n-hexane oil fraction. Based upon gas chromatography-mass spectrometry (GC/MS) analysis, the most effective fatty acid was eugenic acid which is present in higher proportion in both fractions. These fractions of essential oils proved safe for the treatment of antibiotic resistant diarrheic cases of children caused by E. faecalis

CMS physics technical design report : Addendum on high density QCD with heavy ions

Peer reviewe