Search

The present study was designed to identify the potential bioactive compound from endophytic actinomycetes (EA) Nocardiopsis sp. GRG 2 (KT 235641) against selected extended spectrum beta lactamase (ESBL) producing Pseudomonas aeruginosa (P. aeruginosa) and Klebsiella pneumoniae (K. pneumoniae). Initially, the multi drug resistance (MDR) effect of selected uropathogens was confirmed by respective UTI panel of Hexa antibiotics disc methods. The zone of inhibition ≤22 mm for ceftazidime, ≤ 27 mm for cefotaxime and ≤8 mm zone of MIC stripe against both the uropathogens of phenotypic methods confirmed, the selected strains were ESBL producer. Among the various EA extracts, GRG 2 extract showed excellent antibacterial activity against both ESBL producing P. aeruginosa and K. pneumonia by agar well diffution method. The molecular identification of selected GRG 2 strain was named as Nocardiopsis sp. GRG 2 (KT235641). The antibacterial metabolites present in the TLC elution was exhibited at 274 nm by UV visible spectrometer. The partial purification of preparative HPLC fraction 3 showed 14, 16 mm against P. aeruginosa and K. pneumoniae, respectively. Based on the antibacterial effect, the FT-IR, GC-MS and LC-MS analysis of fraction 3 was confirmed as 1, 4-diaza-2, 5-dioxo-3-isobutyl bicyclo[4.3.0]nonane (DDIBN). Further, the dose dependent inhibition of DDIBN against both ESBL producing pathogens was observed at 75 μg/mL by minimum inhibition concentration (MIC) and minimum bactericidal concentration (MBC). The increased cell death and disrupted cell membrane integrity were observed at MIC of DDIBN by confocal laser scanning electron microscope (CLSM) and scanning electron microscope (SEM). The results were proved that the DDIBN has potential antibacterial metabolites against ESBL producing pathogens and it can be applied for various other biomedical fields.

The aim of the current study is to identify bioactive compound from marine endophytic actinomycetes (MEA) isolated from Gulf of Mannar region, Southeast coast of India. Among the isolated actinomycetes, strain GRG 4 exhibited excellent ability to inhibit isolated colistin resistant (CR) Pseudomonas aeruginosa (P. aeruginosa) and Klebsiella pneumoniae (K. pneumoniae), which is a emerging threat to the world. The strain was identified as Streptomyces coeruleorubidus GRG 4 (KY457708), based on morphological, biochemical, phenotypic and genotypic characters. The bioactive metabolites present in the methanolic extract were partially purified by TLC and preparative HPLC. The active HPLC fraction 2 showed 15, 20 mm zone of inhibition against both CR P. aeruginosa and K. pneumoniae respectively. Analytical HPLC and FT-IR results of fraction 2 showed with carbonyl group. Both GC-MS and LC-MS results confirmed that the fraction 2 contained chemical constituents of Bis (2-Ethylhexyl) Phthalate (BEP). The compromised structure with loosely integrated and ruptured cell wall of BEP treated CR bacteria were observed by confocal laser scanning microscope (CLSM) and scanning electron microscope (SEM) at 75 μg/mL of minimum inhibitory concentration (MIC) dose. Further, cytotoxic effect of BEP against A549 human lung cancer cells revealed complete inhibition by cell proliferation and apoptosis was observed at 100 μg/mL in 24 h treatment. In addition, irreversible ROS dependent oxidative damage was clearly observed at the IC50 concentration of BEP. The toxicity of BEP was also studied against Vibrio fischeri (V. fischeri) and found to be highly toxic after 15 and 30 min of treatment. Based on the results it could be concluded that the identified compound BEP is a potent inhibitor for CR bacteria and A549 lung cancer cells.

The multidrug resistant Gram negative bacteria (MDRGNB) is an emerging burden and now represents a daily challenge for the management of antimicrobial therapy in healthcare settings. The present study was aimed to detect the prevalence of TEM and CTX-M type genes from GNB on urinary tract infection (UTIs). The ciprofloxacin resistant uropathogens were detected by HEXA UTI 5 disc diffusion method. The phenotypic detection of uropathogens producing extended spectrum beta lactamases (ESBLs) was confirmed by double disc combination test (DDCT) and phenotype confirmation test (PCT). The prevalence of TEM and CTX-M genes of uropathogens was identified by multiplex PCR analysis. The in vitro antimicrobial susceptibility of E. coli producing ESBL (26), 21 isolates of P. mirabilis, 17 P. aeruginosa, 14 K. pneumoniae and 6 Enterobacter sp. were detected. Based on the extension of the cephalosporin zone edge towards augmentin disc in the DDST method proved 84% of the isolates were ESBL positive. Similar results were obtained in phenotypic confirmatory test (PCT) by the increases of ≥5 mm zone of inhibition in the combination disc when compared with ceftazidime disc alone. The prevalence of TEM and CTX-M genes were determined from multidrug resistance uropathogens (MDU) respectively as 83%, 75%, 71%, 63%, 60%, 55%, 54%, 50%. The most prevalent (TEM + CTX-M) genes were also detected in ciprofloxacin resistant strains P. mirabilis BDUMS1 (KY617768) and E. coli BDUMS3 (KY617770). Due to the increase of ESBL genes in uropathogens, sustained supervision for using favorable antibiotics and decreasing the infection is essential.

Copper oxide nanoparticles (CuO NPs) were synthesized biologically using leaf extract of Camilla japonica. The typical UV–visible spectral peak of CuO NPs was observed at a wavelength of ∼290 nm, which confirmed their successful synthesis. From scanning electron microscope (SEM) and transmission electron microscope (TEM) analyses, the synthesized CuO NPs were found to possess spherical shape. Energy dispersive X-ray analyzer (EDX) results revealed that the CuO NPs are almost pure with atomic percentages of 50.92 for Cu and 49.08 for O. Fourier transform infrared (FTIR) confirmed the presence of an absorption peak located at a wavenumber position of ∼480 cm−1 typical for highly pure CuO NPs. TEM images displayed that the particles are relatively uniform in size ∼15–25 nm. The P. aeruginosa and K. pneumonia showed complete resistance against Hexa 077 antibiotic discs. The result of ≤22 ceftazidime and ≤27 cefotaxime confirmed that both the uropathogens were ESBL producers. The ≥8 mm of the MIC stripe further confirmed that both the uropathogens were ESBL producers. Furthermore, the antibacterial activity of CuO NPs against selected ESBL producing P. aeruginosa and K. pneumoniae at minimum inhibition concentration (MIC) of 100 μg/mL. The decreased cell viability and damaged membrane construction of both the uropathogens were observed by confocal laser scanning microscope (CLSM) using AO/EB stains at desired MIC dose. The morphological damage of the bacterial cells was demonstrated by SEM analysis. Hence, based on the above in vitro findings, the results suggested that the CuO NPs are efficient antibacterial compounds against ESBL producing bacteria, and that the plant leaf mediated CuO NPs can be considered as novel and promising material to act against various infectious bacteria.

Urinary tract infections (UTIs) are diverse public health complication and caused by range of pathogens, however mostly Gram negative bacteria cause significant life threatening risks to different populations. The prevalence rate and antimicrobial resistance among the Gram negative uropathogens alarmed significantly heighten the economic burden of these infections. In this study, we investigated the antibiofilm efficiency of Pyrrolo [1,2-a] pyrazine-1,4-dione,hexahydro-3-(2-methylpropyl) extracted from endophytic actinomycetes Nocardiopsis sp. GRG 1 (KT235640) against P. mirabilis and E. coli. The extracted compound was characterized through TLC, HPLC, GC-MS, LC-MS and confocal laser scanning microscopy (CLSM), scanning electron microscopy (SEM). The compound, Pyrrolo [1,2-a] pyrazine-1, 4-dione, hexahydro-3-(2-methylpropyl) inhibits both bacterial biofilm formation as well as reduces the viability of preformed biofilms. Furthermore, CLSM image shows cell shrinkage, disorganized cell membrane and loss of viability. The SEM result also confirms the cell wall degradation in treated cells of the bacteria. Hence, the Pyrrolo [1,2-a]pyrazine-1,4-dione, hexahydro-3-(2-methylpropyl) is active against P. mirabilis and E. coli.

The accelerative outgrowth of extended spectrum β-lactamases (ESBLs) producing Escherichia coli (E. coli) and Proteus mirabilis (P. mirabilis) was mainly due to incessant relentless influence of antibiotics thereby increasing incidence and death rate which was obvious from the survey of ESBLs producing bacteria related health problem. In the present paper, we synthesized and characterized zinc oxide nanoparticles (ZnO NPs) employing using Camellia japonica leaf extract, bactericidal action of these NPs against extended spectrum β lactamases (ESBLs) positive E. coli and P. mirabilis clinical strains owing the minimal inhibitory concentration (MIC) percentage 83, 81% at 100 μg/mL concentration and minimum bactericidal concentration (MBC) final inhibiting concentration at 150 μg/mL. Moreover, confocal laser scanning microscopy (CLSM) and scanning electron microscope (SEM) results evident for loss of viability, cell shrinkage, disarrangement of cell membrane, and cell wall lysis activity of ZnO NPs against ESBLs positive E. coli BDUMS3 (KY617770) and P. mirabilis BDUMS1 (KY617768) strains. From the results, it was observed that the biologically synthesized ZnO NPs has stronger antibacterial effect against ESBLs producing bacterial strains. Nevertheless, current date there is no reports of antibacterial activity of metal oxide (ZnO) NPs against ESBL producing gram negative bacteria. Consequently, this finding is the first report in this respect and it shows band gap energy and ROS accumulation to damage the cell wall and inhibit the growth of ESBL producing gram negative strains.