Search results for: resistant microbial strains

Authors: Rama Devi Kyatham, D. Ashok, K. S. K. Rao Patnaik, Raju Bathula

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We have shown the importance of pyrazoles as anti-microbial chemical entities. These compounds have generally been considered significant due to their wide range of pharmacological acivities and their discovery motivates new avenues of research.The proposed pyrazoles were synthesized and evaluated for their anti-microbial activities. The Synthesized compounds were analyzed by different spectroscopic methods.

Keywords: pyrazoles, validation, resistant microbial strains, anti-microbial activities

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Authors: R. Gounina-Allouane, I. Moussaoui, N. Boukahel

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Bacillus antimicrobial metabolites, especially those of Bacillus thuringiensis (Bt), are of great interest for research because of health risks generated by the excessive use of chemical additives as well as the propagation of resistant microbial strains, caused by the massive treatment with antibiotics. The objective of this study was the selection of Bt strains producing antimicrobial peptides (bacteriocins), and the partial purification of the most powerful bacteriocins, then the determination of their spectra of antimicrobial action. A collection of twenty one Bt strains isolated from soil at Boumerdès (northern Algeria) was used for screening strains having an antagonistic activity against phylogenetically closed bacteria. Spectra of antagonistic activity of two selected strains was determined against other Bt strains, Gram positive and Gram negative bacterial strains of clinical origin and others from ATCC collection as well as yeasts isolated in human dermatology. Bacteriocins of these two strains were partially purified and their effect on the kinetics of growth of the most sensitive microbial strains was studied. The bacteriocinogenic strains were biochemically characterized and their sensitivity to antibiotics was studied.

Keywords: antimicrobial peptides, Bacillus thuringiensis, bacteriocin, partial purification

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Authors: R. Gounina-Allouane, I. Moussaoui, N. Boukahel

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Bacillus antimicrobial metabolites, especially those of Bacillus thuringiensis (Bt), are of great interest for research because of health risks generated by the excessive use of chemical additives as well as the propagation of resistant microbial strains, caused by the massive treatment with antibiotics. The objective of this study was the selection of Bt strains producing antimicrobial peptides (bacteriocins), and the partial purification of the most powerful bacteriocins, then the determination of their spectra of antimicrobial action. A collection of twenty one Bt strains isolated from soil at Boumerdès (northern of Algeria) was used for screening strains having an antagonistic activity against phylogenetically closed bacteria. Spectra of antagonistic activity of two selected strains was determined against other Bt strains, Gram positive and Gram negative bacterial strains of clinical origin and others from ATCC collection as well as yeasts isolated in human dermatology. Bacteriocins of these two strains were partially purified and their effect on the kinetics of growth of the most sensitive microbial strains was studied. The bacteriocinogenic strains were biochemically characterized and their sensitivity to antibiotics was studied.

Keywords: antimicrobial peptides, Bacillus thuringiensis, bacteriocin, partial purification

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Authors: Ji-Chan Jang

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Tuberculosis is still major health problem because there is an increase of multidrug-resistant and extensively drug-resistant forms of the disease. Therefore, the most urgent clinical need is to discover potent agents and develop novel drug combination capable of reducing the duration of MDR and XDR tuberculosis therapy. Three reference strains H37Rv, CDC1551, W-Beijing GC1237 and six clinical isolates of MDRTB were tested to daptomycin in the range of 0.013 to 256 mg/L. Daptomycin is resistant to all tested M. tuberculosis strains not only laboratory strains but also clinical MDR strains that were isolated at different source. Daptomycin will not be an antibiotic of choice for treating infection of Gram positive atypical slowly growing M. tuberculosis.

Keywords: tuberculosis, daptomycin, resistance, Mycobacterium tuberculosis

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Authors: A. Qasmaoui, F. Ohmani, Z. Zaine, I. El Akrad, J. Hamamouchi, K. Halout, B. Belkadi, R. Charof

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The evolution of antibiotic resistance is a crucial aspect of the problem related to the intensive use of these substances in medicine for humans and animals. The production of ESBL extended spectrum β-lactamase enzymes is the main mechanism of resistance to β-lactam antibiotics in Escherichia coli. The objective of our work is to determine the resistance genes in E. coli strains.ESBL coli stored at the epidemic diseases laboratory of the National Institute of Hygiene. The strains were identified according to the classic bacteriological criteria. An antibiogram was performed on the strains isolated by the Mueller Hinton agar disc diffusion method. The production of ESBL in the strains was detected by the synergy assay technique and confirmed for the presence of the blaCTX-M1, blaCTX-M2, blaTEM, blaSHV, blaOXA-48 genes by gene amplification . Of the 27 observed strains of E.coli, 17 isolated strains present the phenotype of extended-spectrum Beta-lactamase with a percentage of 63%.. All 18 cefotaxime-resistant strains were analyzed for an ESBL phenotype. All strains were positive in the double-disc synergy assay. The fight against the emergence and spread of these multi-resistant antibiotic-resistant strains requires the reasonable use of antibiotics.

Keywords: E coli, BLSE, CTX, TEM, SHV, OXA, résistance aux antibiotique

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Authors: Recep Kesli, Merih Simsek, Cengiz Demir, Onur Turkyilmaz

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Objective: Goal of this study was to determine the antibiotic resistance of Staphylococcus aureus (S. aureus) and Methicillin resistant staphylococcus aureus (MRSA) strains isolated at Medical Microbiology Laboratory of ANS Application and Research Hospital, Afyon Kocatepe University, Turkey. Methods: S. aureus strains isolated between October 2012 and September 2016, from various clinical specimens were evaluated retrospectively. S. aureus strains were identified by both the conventional methods and automated identification system -VITEK 2 (bio-Mérieux, Marcy l’etoile, France), and Meticillin resistance was verified using oxacillin disk with disk-diffusion method. Antibiotic resistance testing was performed by Kirby-Bauer disc diffusion method according to CLSI criteria, and intermediate susceptible strains were considered as resistant. Results: Seven hundred S.aureus strains which were isolated from various clinical specimens were included in this study. These strains were mostly isolated from blood culture, tissue, wounds and bronchial aspiration. All of 306 (43,7%) were oxacillin resistant. While all the S.aureus strains were found to be susceptible to vancomycin, teicoplanin, daptomycin and linezolid, 38 (9.6 %), 77 (19.5 %), 116 (29.4 %), 152 (38.6 %) and 28 (7.1 %) were found to be resistant aganist to clindamycin, erythromycin, gentamicin, tetracycline and sulfamethoxazole/trimethoprim, retrospectively. Conclusions: Comparing to the Methicillin sensitive staphylococcus aureus (MSSA) strains, increased resistance rates of, trimethoprim-sulfamethoxazole, clindamycin, erythromycin, gentamicin, and tetracycline were observed among the MRSA strains. In this study, the most effective antibiotic on the total of strains was found to be trimethoprim-sulfamethoxazole, the least effective antibiotic on the total of strains was found to be tetracycline.

Keywords: antibiotic resistance, MRSA, Staphylococcus aureus, VITEK 2

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Authors: Maria Manzoor, Iram Gul, Muhammad Arshad

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Bacterial strains were isolated from contaminated soil collected from Rawalpindi and Islamabad. The strains were investigated for lead resistance and their effect on Pb solubility and PGPR activity. Incubation experiments were carried for inoculated and unoculated soil containing different levels of Pb. Results revealed that few stains (BTM-4, BTM-11, BTM-14) were able to tolerate Pb up to 600 mg L-1, whereas five strains (BTM-3, BTM-6, BTM-10, BTM-21 and BTM-24) showed significant increase in solubility of Pb when compared to all other strains and control. The CaCl2 extractable Pb was increased by 13.6, 6.8, 4.4 and 2.4 folds compared to un-inoculated control soil at increased soil Pb concentration (500, 1000, 1500 and 200 mg kg-1, respectively). The selected bacterial strains (11) were further investigated for plant growth promotion activity through PGPR assays including. Germination and root elongation assays were also conducted under elevated metal concentration in controlled conditions to elucidate the effects of microbial strains upon plant growth and development. The results showed that all the strains tested in this study, produced significantly varying concentrations of IAA, siderophores and gibberellic acid along with ability to phosphorus solubilization index (PSI). The results of germination and root elongation assay further confirmed the beneficial role of the microbial strains in elevating metal stress through PGPR activity. Among all tested strains, BTM-10 significantly improved plant growth. 1.3 and 2.7 folds increase in root and shoot length was observed when compared to control. Which may be attributed to presence of important plant growth promoting enzymes (IAA 74.6 μg/ml; GA 19.23 μg/ml; Sidrophore units 49% and PSI 1.3 cm). The outcome of this study indicates that these Pb tolerant and solubilizing strains may have the potential for plant growth promotion under metal stress and can be used as mediator when coupled with heavy metal hyperaccumulator plants for phytoremediation of Pb contaminated soil.

Keywords: Pb resistant bacteria, Pb mobilizing bacteria, Phytoextraction of Pb, PGPR activity of bacteria

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Authors: M. Raissy, M. Shahrani

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The present study was done to evaluate the presence of tetracycline resistance genes in Lactococcus garvieae isolated from cultured rainbow trout, West Iran. The isolates were examined for antimicrobial resistance using disc diffusion method. Of the 49 strains tested, 19 were resistant to tetracycline (38.7%), 32 to enrofloxacin (65.3%), 21 to erythromycin (42.8%), 20 to chloramphenicol and trimetoprim-sulfamethoxazole (40.8%). The strains were then characterized for their genotypic resistance profiles. The results revealed that all 49 isolates contained at least one of the tetracycline resistance genes. Tet (A) was found in 89.4% of tetracycline resistant isolates and the frequency of other gene were as follow: tet (E) 42.1%, tet (B) 47.3%, tet (D) 15.7%, tet (L) 26.3%, tet (K) 52.6%, tet (G) 36.8%, tet (34) 21%, tet (S) 63.1%, tet (C) 57.8%, tet (M) 73.6%, tet (O) 42.1%. The results revealed high levels of antibiotic resistance in L. garvieae strains which is a potential danger for trout culture as well as for public health.

Keywords: Lactococcus garvieae, tetracycline resistance genes, rainbow trout, antimicrobial resistance

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Authors: Zunnu Raen Akhtar, U. Irshad, M. Majid

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Due to the widespread cultivation of transgenic cotton, intense selection pressure resulted in resistant allele in pink bollworm, Pectinophora gossypiella (Gelechiidae: Lepidoptera). A resistant strain of pink bollworm against transgenic cotton has become a challenge to Integrated Resistance Management (IRM) in the World. Laboratory and field studies were conducted to determine the resistant strains of pink bollworm by performing bioassay, extracting the DNA, conducting PCR of both laboratory as well as field collected pink bollworms to observe the developed resistance. In all of the studies, two Bt varieties FH-142 and FH-118 expressing Cry1Ac compared to non-Bt (Control) were tested against pink bollworm. In the laboratory, bioassay results showed that there was no significant mortality difference between Bt and non-Bt varieties. Similar mortality percentage was observed in transgenic and non-transgenic (control) variety. Insects which were survived after bioassay, as well as those collected from the Bt cotton fields, were selected for further molecular studies. DNA extraction followed by PCR was conducted to check the resistant strains in pink bollworm. In field studies, we also observed the population dynamics of pink boll worms on Bt as compared to non-Bt varieties. Laboratory and field studies confirmed that resistant strains occurs in Pakistani Bt cotton fields. Different strategies should be adopted to combat that serious prevailing resistance issues.

Keywords: transgenic cotton, resistance, pectinophora gossypiella, , integrated resistance management (IRM), polymerase chain reaction (PCR)

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Authors: Alouache Souhila, Messai Yamina, Torres Carmen, Bakour Rabah

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Objective: It has often been reported an association between antibiotic resistance and virulence. However, resistance to quinolones seems to be an exception, it tends instead to be associated with an attenuation of virulence, particularly in clinical strains. The purpose of this study was to evaluate the potential virulence of 28 quinolone-resistant E. coli strains recovered from water at the inflow (n=16) and outflow (n=12) of an urban wastewater treatment plant (WWTP). Methods: E. coli isolates were selected on Tergitol-7 agar supplemented with 2µg/ml of ciprofloxacin, they were screened by PCR for 11 virulence genes related to Extraintestinal pathogenic E. coli (ExPEC): papC, papG, afa/draBC, sfa/foc, kpsMTII, iutA, iroN, hlyF, ompT, iss and traT. The phylogenetic groups were determined by PCR and clonal relationship was evaluated by ERIC-PCR. Results: Genotyping by ERIC-PCR showed 7 and 12 DNA profiles among strains of wastewater (inflow) and treated water (outflow), respectively. Strains were assigned to the following phylogenetic groups: B2 (n = 1, 3.5%), D (n = 3, 10.7%), B1 (n = 10, 35.7%.) and A (n = 14, 50%). A total of 8 virulence-associated genes were detected, traT (n=19, 67.8%), iroN (n= 16, 57 .1%), hlyF (n=15, 53 .5%), ompT (n=15, 53 .5%), iss (n=14, 50%), iutA (n=9, 32.1%) , sfa/foc (n=7, 25%) and kpsMTII (n=2, 7.1%). Combination of virulence factors allowed to define 16 virulence profiles. The pathotype APEC was observed in 17.8% (D=1, B1=4) and human ExPEC in 7% (B2=1, D=1) of strains. Conclusion: The study showed that quinolone-resistant E. coli strains isolated from wastewater and treated water in WWTP harbored virulence genes with the presence of APEC and human ExPEC strains.

Keywords: E. coli, quinolone-resistance, virulence, WWTP

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Authors: Asho Ali

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Tuberculosis is a disease of grave concern which infects one-third of the global population. The high incidence of tuberculosis is further compounded by the increasing emergence of drug resistant strains including multi drug resistant (MDR). Global incidence MDR-TB is ~4%. Molecular epidemiological studies, based on the assumption that patients infected with clustered strains are epidemiologically linked, have helped understand the transmission dynamics of disease. It has also helped to investigate the basis of variation in Mycobacterium tuberculosis (MTB) strains, differences in transmission, and severity of disease or drug resistance mechanisms from across the globe. This has helped in developing strategies for the treatment and prevention of the disease including MDR.

Keywords: Mycobcaterium tuberculosis, molecular epidemiology, drug resistance, disease

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Authors: Seyed Davar Siadat, Samira Tarashi, Abolfazl Fateh, Arfa Moshiri

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Background: The global health issue of tuberculosis (TB) still affects patients in every country. TB control may not be as effective as it should be, especially when resistant strains are involved. In this regard, mycobacterial MTases play a major role in tuberculosis, but the mechanisms underlying their function have yet to be fully deciphered. Methods: Five resistant isolates of M.tb were accumulated. As a reference strain, M.tb H37Rv (ATCC 27249) was used. For this analysis, seven putative mycobacterial MTase genes (Rv0645c, Rv1694, Rv2966c, Rv3919c, Rv2756c, Rv1988, and Rv3263), as well as Rv1392 as SAM synthase, were selected. Comparing mutations and expression levels of MTases in different strains was accomplished by PCR-sequencing and qRT-PCR. The relative expression levels of these genes were calculated using the 2 -ΔΔCt method. Results: The Rv3919c gene (T to G in codon 341) and Rv1392 gene (G to A in codon 97) were the only mutations found in the INHR strain. In all sensitive and resistant isolates, Rv0645c, Rv3263, Rv2756c, and Rv2966c were overexpressed. However, the expression of Rv1988 and Rv3919c decreased in the sensitive strains, whereas the expression of Rv1694 increased. There was also a decreased expression of Rv1392 in the INHR isolate. Conclusion: The presence of mycobacterial MTases as well as resistance to antibiotics were found to be correlated in M.tb strains. Undoubtedly, there are some MTases that are associated with the virulence process. It is necessary to conduct additional studies to fully explore the impact of mycobacterial MTases within specific strains of M.tb to develop novel diagnostic and treatment strategies.

Keywords: mycobacterium tuberculosis, drug resistance, methyltransferases, s-adenosylmethionine

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Authors: Ashish Shukla, K. P. Mishra, Pushplata Tripathi

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This paper investigates the production and optimization of β-galactosidase enzyme using synthetic medium by isolated wild strains (S1, S2) mutated strains (M1, M2) through SSF and SmF. Among the different cell disintegration methods used, the highest specific activity was obtained when the cells were permeabilized using isoamyl alcohol. Wet lab experiments were performed to investigate the effects of carbon and nitrogen substrates present in Vogel’s medium on β-galactosidase enzyme activity using S1, S2, and M1, M2 strains through SSF. SmF experiments were performed for effects of carbon and nitrogen sources in YLK2Mg medium on β-galactosidase enzyme activity using S1, S2 and M1, M2 strains. Effect of pH on β-galactosidase enzyme production was also done using S1, S2, and M1, M2 strains. Results were found to be very appreciable in all the cases.

Keywords: β-galactosidase, cell disintegration, permeabilized, SSF, SmF

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Authors: Hani Belhadj, Daoud Harzallah, Seddik Khennouf, Saliha Dahamna, Mouloud Ghadbane

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In this study, Lactobacillus strains isolated from pollen were identified by means of phenotypic and genotypic methods, At pH 2, most strains proved to be acid resistants, with losses in cell viability ranging from 0.77 to 4.04 Log orders. In addition, at pH 3 all strains could grew and resist the acidic conditions, with losses in cell viability ranging from 0.40 to 3.61 Log orders. It seems that, 0.3% and 0.5% of bile salts does not affect greatly the survival of most strains, excluding Lactobacillus sp. BH1398. Survival ranged from 81.0±3.5 to 93.5±3.9%. In contrast, in the presence of 1.0% bile salts, survival of five strains was decreased by more than 50%. Lactobacillus fermentum BH1509 was considered the most tolerant strain (77.5% for 1% bile) followed by Lactobacillus plantarum BH1541 (59.9% for 1% bile). Furthermore, all strains were resistant to colistine, clindamycine, chloramphenicol, and ciprofloxacine, but most of the strains were susceptible to Peniciline, Oxacillin, Oxytetracyclin, and Amoxicillin. Functionally interesting Lactobacillus isolates may be used in the future as probiotic cultures for manufacturing fermented foods and as bioactive delivery systems.

Keywords: probiotics, lactobacillus, pollen, bile, acid tolerance

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Authors: R. M. Noah, N. M. Nasir, M. R. Jais, M. S. S. Wahab, M. H. Abdullah, A. S. S. Raj

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Infectious diseases are getting more difficult to treat due to the resistant strains of bacteria. Current generations of antibiotics are most likely ineffective against multi-drug resistant strains bacteria. Thus, there is an urgent need in search of natural antibiotics in particular from medicinal plants. One of the common medicinal plants, Melaleuca cajuputi, has been reported to possess antibacterial properties. The study was conducted to evaluate and justify the presence of antibacterial activity of Melaleuca cajuputi essential oil (EO) against the multi-drug resistant bacteria. Clinical isolates obtained from the teaching hospital were re-assessed to confirm the exact identity of the bacteria to be tested, namely methicillin-resistant staphylococcus aureus (MRSA), carbapenem-resistant enterobacteriaceae (CRE), and extended-spectrum beta-lactamases producer (ESBLs). A well diffusion method was done to observe the inhibition zones of the essential oil against the bacteria. Minimum inhibitory concentration (MIC) was determined using the microdilution method in 96-well flat microplate. The absorbance was measured using a microplate reader. Minimum bactericidal concentration (MBC) was performed using the agar medium method. The zones of inhibition produced by the EO against MRSA, CRE, and ESBL were comparable to that of generic antibiotics used, gentamicin and augmentin. The MIC and MBC results highlighted the antimicrobial efficacy of the EO. The outcome of this study indicated that the EO of Melaleuca cajuputi had antibacterial activity on the multi-drug resistant bacteria. This finding was eventually substantiated by electron microscopy work.

Keywords: melaleuca cajuputi, antibacterial, resistant bacteria, essential oil

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Authors: Patrícia Branco, Catarina Prista, Helena Albergaria

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Industrial fuel-ethanol fermentations are carried out under non-sterile conditions, which favors the development of microbial contaminants, leading to huge economic losses. Wild yeasts such as Brettanomyces bruxellensis and lactic acid bacteria are the main contaminants of industrial bioethanol fermentation, affecting Saccharomyces cerevisiae performance and decreasing ethanol yields and productivity. In order to control microbial contaminations, the fuel-ethanol industry uses different treatments, including acid washing and antibiotics. However, these control measures carry environmental risks such as acid toxicity and the rise of antibiotic-resistant bacteria. Therefore, it is crucial to develop and apply less toxic and more environmentally friendly biocontrol methods. In the present study, an industrial fuel-ethanol starter, S. cerevisiae Ethanol-Red, was genetically modified to over-express AMPs with activity against fuel-ethanol microbial contaminants and evaluated regarding its biocontrol effect during mixed-culture alcoholic fermentations artificially contaminated with B. bruxellensis. To achieve this goal, S. cerevisiae Ethanol-Red strain was transformed with a plasmid containing the AMPs-codifying genes, i.e., partial sequences of TDH1 (925-963 bp) and TDH2/3 (925-963 bp) and a geneticin resistance marker. The biocontrol effect of those genetically modified strains was evaluated against B. bruxellensis and compared with the antagonistic effect exerted by the modified strain with an empty plasmid (without the AMPs-codifying genes) and the non-modified strain S. cerevisiae Ethanol-Red. For that purpose, mixed-culture alcoholic fermentations were performed in a synthetic must use the modified S. cerevisiae Ethanol-Red strains together with B. bruxellensis. Single-culture fermentations of B. bruxellensis strains were also performed as a negative control of the antagonistic effect exerted by S. cerevisiae strains. Results clearly showed an improved biocontrol effect of the genetically-modified strains against B. bruxellensis when compared with the modified Ethanol-Red strain with the empty plasmid (without the AMPs-codifying genes) and with the non-modified Ethanol-Red strain. In mixed-culture fermentation with the modified S. cerevisiae strain, B. bruxellensis culturability decreased from 5×104 CFU/mL on day-0 to less than 1 CFU/mL on day-10, while in single-culture B. bruxellensis increased its culturability from 6×104 to 1×106 CFU/mL in the first 6 days and kept this value until day-10. Besides, the modified Ethanol-Red strain exhibited an enhanced antagonistic effect against B. bruxellensis when compared with that induced by the non-modified Ethanol-Red strain. Indeed, culturability loss of B. bruxellensis after 10 days of fermentation with the modified Ethanol-Red strain was 98.7 and 100% higher than that occurred in fermentations performed with the non-modified Ethanol-Red and the empty-plasmid modified strain, respectively. Therefore, one can conclude that the S. cerevisiae genetically modified strain obtained in the present work may be a valuable solution for the mitigation of microbial contamination in fuel-ethanol fermentations, representing a much safer and environmentally friendly preservation strategy than the antimicrobial treatments (acid washing and antibiotics) currently applied in fuel-ethanol industry.

Keywords: antimicrobial peptides, fuel-ethanol microbial contaminations, fuel-ethanol fermentation, biocontrol agents, genetically-modified yeasts

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Authors: Bello Gonzalez T. D. J., Setten Van M., Essen Van A., Brouwer M., Veldman K. T.

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Resistance to fluoroquinolones (FQ) has evolved increasingly over the years, posing a significant challenge for the treatment of human infections, particularly gastrointestinal tract infections caused by zoonotic bacteria transmitted through the food chain and environment. In broiler chickens, a relatively high proportion of FQ resistance has been observed in Escherichia coli indicator, Salmonella and Campylobacter isolates. We hypothesize that flumequine (Flu), used as a secondary choice for the treatment of poultry infections, could potentially be associated with a high proportion of FQ resistance. To evaluate this hypothesis, we used an in vitro fermentation chicken caeca model. Two continuous single-stage fermenters were used to simulate in real time the physiological conditions of the chicken caeca microbial content (temperature, pH, caecal content mixing, and anoxic environment). A pool of chicken caecal content containing FQ-resistant E. coli obtained from chickens at slaughter age was used as inoculum along with a spiked FQ-susceptible Campylobacter jejuni strain isolated from broilers. Flu was added to one of the fermenters (Flu-fermenter) every 24 hours for two days to evaluate the selection and maintenance of FQ resistance over time, while the other served as a control (C-Fermenter). The experiment duration was 5 days. Samples were collected at three different time points: before, during and after Flu administration. Serial dilutions were plated on Butzler culture media with and without Flu (8mg/L) and enrofloxacin (4mg/L) and on MacConkey culture media with and without Flu (4mg/L) and enrofloxacin (1mg/L) to determine the proportion of resistant strains over time. Positive cultures were identified by mass spectrometry and matrix-assisted laser desorption/ionization (MALDI). A subset of the obtained isolates were used for Whole Genome Sequencing analysis. Over time, E. coli exhibited positive growth in both fermenters, while C. jejuni growth was detected up to day 3. The proportion of Flu-resistant E. coli strains recovered remained consistent over time after antibiotic selective pressure, while in the C-fermenter, a decrease was observed at day 5; a similar pattern was observed in the enrofloxacin-resistant E. coli strains. This suggests that Flu might play a role in the selection and persistence of enrofloxacin resistance, compared to C-fermenter, where enrofloxacin-resistant E. coli strains appear at a later time. Furthermore, positive growth was detected from both fermenters only on Butzler plates without antibiotics. A subset of C. jejuni strains from the Flu-fermenter revealed that those strains were susceptible to ciprofloxacin (MIC < 0.12 μg/mL). A selection of E. coli strains from both fermenters revealed the presence of plasmid-mediated quinolone resistance (PMQR) (qnr-B19) in only one strain from the C-fermenter belonging to sequence type (ST) 48, and in all from Flu-fermenter belonged to ST189. Our results showed that Flu selective impact on PMQR-positive E. coli strains, while no effect was observed in C. jejuni. Maintenance of Flu-resistance was correlated with antibiotic selective pressure. Further studies into antibiotic resistance gene transfer among commensal and zoonotic bacteria in the chicken caeca content may help to elucidate the resistance spread mechanisms.

Keywords: fluoroquinolone-resistance, escherichia coli, campylobacter jejuni, in vitro model

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Authors: K. Ali, M. F. Qamar, M. A. Zaman, M. Younus, I. Khan, S. Ehtisham-ul-Haque, R. Tamkeen, M. I. Rashid, Q. Ali

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This study centers on molecular identification of Haemonchus contortus and isolation of Benz-imidazoles (BZ) resistant strains. Different abattoirs’ of two geographic regions of Punjab (Pakistan) were frequently visited for the collection of worms. Out of 1500 (n=1500) samples that were morphologically confirmed as H. contortus, 30 worms were subjected to molecular procedures for isolation of resistant strains. Resistant worms (n=8) were further subjected to DNA gene sequencing. Bio edit sequence alignment editor software was used to detect the possible mutation, deletion, replacement of nucleotides. Genetic diversity was noticed and genetic variation existing in β-tubulin isotype 1 of the H. contortus population of small ruminants of different regions considered in this study. H. contortus showed three different type of genetic sequences. 75%, 37.5%, 25% and 12.5% of the studied samples showed 100% query cover and identity with isolates and clones of China, UK, Australia and other countries, respectively. Interestingly the neighbor countries such as India and Iran haven’t many similarities with the Pakistani isolates. Thus, it suggests that population density of same genetic makeup H. contortus is scattered worldwide rather than clustering in a single region.

Keywords: Haemonchus contortus, Benzimidazole resistant, β-tubulin-1 gene, abattoirs

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Authors: C. A. Ologunde

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Escherichia coli have been a major microorganisms associated with, and isolated from feacal samples either in adult or children all over the world. Strains of these organisms are resistant to cephalosporins and fluoroquinolone (FQ) antimicrobial agents among hospitalized patients and FQs are the most frequently prescribed antimicrobial class in hospitals, and the level of resistant of E. coli to these antimicrobial agents is a risk factor that should be assessed. Hence, this study was conducted to determine the prevalence and risk factors for colonization with fluoroquinolone (FQ)-resistant E. coli in hospitalized patients in Ado-Ekiti. Rectal swabs were obtained from patients in hospitals in the study area and FQ-resistant E. coli were isolated and identified by means of Nalidixic acid multi-disk and a 1-step screening procedure. Species identification and FQ resistance were confirmed by automated testing (Vitek, bioMerieux, USA). Individual colonies were subjected to pulse-field gel electrophoresis (PAGE) to determine macro-restriction polymorphism after digestion of chromosomal DNA. FQ-resistant E. coli was detected in the stool sample of 37(62%) hospitalized patient. With multivariable analyses, the use of FQ before hospitalization was the only independent risk factor for FQ-resistant E. coli carriage and was consistent for FQ exposures for the 3-12 months of study. Pulsed-field gel electrophoresis of FQ-resistant E. coli identified conal spread of 1(one) strain among 18 patients. Loss (9 patients) or acquisition (10 residents) of FQ-resistant E. coli was documented and was associated with de novo colonization with genetically distinct strains. It was concluded that FQ-resistant E. coli carriage was associated with clonal spread. The differential effects of individual fluoroquinolone on antimicrobial drug resistance are an important area for future study, as hospitals manipulate their formularies with regard to use of individual fluoroquinolone, often for economic reasons.

Keywords: E. coli, fluoroquinolone, risk factors, feacal carriage, hospitalized patients, Ado-Ekiti

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Authors: Naima Nur, Safa Islam, Saeema Islam, Faridul Alam

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Background: Drug-resistant pulmonary tuberculosis (DR-PTB), particularly multidrug-resistant tuberculosis (MDR-TB) and pre-extensive drug-resistant (pre-XDR), is a major challenge in effectively controlling TB, especially in developing. This study aimed to identify the strains of M. tuberculosis complex (MTC) and drug resistance patterns among the pulmonary tuberculosis patients. Methods: The study used a cross-sectional design, and 815 patients were recruited randomly in three study periods. In the first-period, 210 treated PTB patients, who were completed their treatment, received their diagnoses using light microscopy, fluorescence microscopy and cultured on Lowenstein-Jensen (L-J) slant, and then strains were identified as MTC by biochemical tests, and then sensitivity test in National Institute of Diseases of the Chest and Hospital. In the second-period, 220 re-treated PTB patients, who were completed their treatment, received their diagnoses using culture on L-J slant, line probe assay (LPA), and GeneXpert in the same hospital. In the last-period, during treatment, 385 MDR-PTB patients received their diagnoses using culture on L-J slant and LPA in the same hospital. Results: Among sixty-two (29.5%) PTB patients, 13% were sensitive to all first-line anti-TB drugs, 26% were MDR-TB patients, and 14.2% were pre-XDR-TB among 14 MDR-TB patients. After three years, 31% were MDR-TB among 220 re-treated PTB patients. After five years, 16.4% was pre-XDR-TB among 385 MDR-TB patients. Compared to females, male patients were significantly higher at all times. Conclusion: The current study demonstrated that in three study periods, the proportions of DR-TB, MDR-TB, and pre-XDR patients were an alarming issue and increasing daily.

Keywords: multi-drug resistant, drug-resistant, pre-extensive drug resistant, pulmonary tuberculosis

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Authors: Fatima Bouazza, Rachida Hassikou, Lamiae Amallah, Jihane Ennadir, Khadija Khedid

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This study evaluated the in vitro resistance and susceptibility of Enterobacteriaceae (Escherichia coli and Klebsiella oxytoca strains) and Staphylococci strains, isolated from sheep’s milk, against antibiotics and essential oils from Thymus satureioides and Mentha pulegium. Antibiotic resistance tests were done using disc diffusion while essential oils were extracted by steam distillation, and yields were calculated relative to plant dry matter. Gas chromatography-mass Spectrometry (GC-MS) was used to analyze each oil's chemical composition. The AMC, CTX, FOX, NA, CN, CIP, and OFX were very effective against the E. coli strains tested. Half of the strains were resistant to AMC, 60% to TIC, and 80% to TE. The K. oxytoca was resistant against AMC, FOX, and TIC (100%). Antibiotic-resistant testing on Staphylococci strains indicated Staphylococcus capitis and Staphylococcus chromogenes as the most sensitive. Staphylococcus aureus, Staphylococcus xylosus, and Staphylococcus cohnii ureal exhibited less resistance to OX, TE, PT, E, and P. The M. pulegium resulted in a higher yield of essential oil of 3.2% oil compared to T. satureioides with only 1.85% yield. Staphylococcus aureus, Staphylococcus xylosus, and Staphylococcus cohnii ureal had lower OX, TE, PT, E, and P resistance. M. pulegium yielded 3.2% essential oil compared to 1.85% for T. satureioides. The monoterpene oxygenated derivatives, monoterpene hydrocarbons, and phenols are found in essential oil extracts. T. satureioides essential oil had high antibacterial activity even at low concentrations (0.2; 0.55 g/mL). The Minimal Bactericidal Concentration (MBC) values indicate that the essential oils from the plants analyzed had bactericidal effects on all strains tested and are similar to the Minimal Inhibitory Concentration (MIC) values. The high antibacterial properties of these medicinal plants, against bacteria isolated from sheep’s milk, provide an opportunity to use these medicinal plants in the breeding sector as additives and preservatives in the dairy industry.

Keywords: antibiotic resistance, medicinal plants, essential oils, enterobacteriaceae, staphylococci, sheep milk

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Authors: Ranjith Kumar Kankala, Wei-Zhi Lin, Chia-Hung Lee

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Antibiotic resistance in bacteria has become an emergency situation clinically. To improve the efficacy of antibiotics in resistant strains, advancement of nanoparticles is inevitable than ever. Herewith, we demonstrate a design by immobilizing tetracycline (TET) in copper substituted mesoporous silica nanoparticles (Cu-MSNs) through a pH-sensitive coordination link, enabling its release in the acidic environment. Subsequently, MSNs are coated with silver nanoparticles stabilized polyethyleneimine (PEI-SNP) to act against drug-resistant (MDR) bacterial strains. Silver ions released from SNP are capable of sensitizing the resistant strains and facilitate the generation of free radicals capable of damaging the cell components. In addition, copper ions in the framework are also capable of generating free radicals through Fenton-like reaction. Furthermore, the nanoparticles are well-characterized physically, and various antibacterial efficacious tests against isolated multidrug resistant bacterial strain were highly commendable. However, this formulation has no significant toxic effect on normal mammalian fibroblast cells accounting its high biocompatibility. These MSN trio-hybrids, i.e., SNP, tetracycline, and copper ions result in synergistic effects, and their advancement could bypass resistance and allow synergism for effective treatment of antibiotic clinically.

Keywords: antibiotic resistance, copper, mesoporous silica nanoparticles, Ph-sensitive release, polyethyleneimine, silver, tetracycline

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Authors: Zohra Benfodda, Valentin Duvauchelle, Chaimae Majdi, David Bénimélis, Catherine Dunyach-Remy, Patrick Meffre

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New antibiotics are necessary to treat microbial pathogens, especially ESKAPE pathogens that are becoming increasingly resistant to available treatment. Despite the medical need, the number of newly approved drugs continues to decline. The majority of antibiotics under clinical development are natural products or derivatives thereof. 43 juglone/naphthazarin derivatives were synthesized using Minisci-type direct C–H alkylation and evaluated for their antibacterial properties against various clinical and reference Gram-positive MSSA, clinical Gram-positive MRSA. Different compounds of the synthesized series showed promising activity against clinical and reference MSSA (MIC: 1–8 μg/ml) and good efficacy against clinical MRSA (MIC: 2–8 μg/ml) strains. The synergistic effects of active compounds were evaluated with reference antibiotics (vancomycin and cloxacillin), and it was found that the antibiotic combination with those active compounds efficiently enhanced the antimicrobial activity and consequently the MIC values of reference antibiotics were lowered up to 1/16th of the original MIC. These synthesized compounds did not present hemolytic activity on sheep red blood cells. In addition to the in silico prediction of ADME profile parameter which is promising and encouraging for further development.

Keywords: juglone, naphthazarin, antibacterial, clinical MRSA, synergistic studies, MIC determination

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Authors: Xing Ping Hu, Yuexun Tian, Jerome Hogsette

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Housefly, Musca domestica L., is a serious urban nuisance and public health/food safety concern. This study evaluated the topical toxicity of 17 essential oil components and 3 plant essential oils against permethrin-resistant adult females and insecticide-susceptible house fly strains. Results show that thymol had the lowest LD₅₀ values against permethrin-resistant strain (43.77 and 41.10 ug per fly) and permethrin-susceptible strain (35.19 and 29.16 ug per fly) at both 24- and 48-hours post treatments; (+)-Pulegone had the lowest LD₉₅ values against the permethrin-resistant strain (0.15 and 0.10 mg per fly) at 24- and 48-hours post treatments, whereas plant thyme oil had the lowest LD₉₅ value of 0.17 mg per fly at post-24h and post-48h against the permethrin-susceptible strain. Additionally, the LD₅₀s was slightly but not significantly negatively correlated with the boiling points of the compounds tested; but showed no correlation with the density and LogP. These results indicate that specific essential oils and compounds have topical insecticidal properties against house flies with low dose. They may have the potential for development as botanical insecticides.

Keywords: urban pest, public health, pest management, botanical chemical

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Authors: Elsa Makue Nguuffo, Esther Del Florence Moni Ndedi, Jacky Njiki Bikoï, Jean Paul Assam Assam, Maximilienne Ascension Nyegue

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Aims: The present study aims to evaluate the kinetic parameters of essential oils (EOs) and combinations fromDrypetes gossweileri Stem Bark, Ocimum gratissimum leaves, Cymbopogon citratusleaves after evaluation of their antibacterial activityonmultidrug-resistant strains ofShigella. Material and Methods:fiveclinical strains of Shigellaisolated from patients with diarrhoeaincluding Shigella flexneri, and 4 otherstrains of Shigella sppwere selected. Their antibiotic profile was established using agar test diffusion with seven antibiotics belonging to seven classes.EOs were extracted from each plant using hydrodistillation process. The activity of Ciprofloxacin®, OEs, and their combination formulatedinthe followingratios(w/w/w): C1: 1/1/1; C2: 2/1/1; C3: 1/2/1, C4:1/1/2 was evaluated microdilution assay. The various interactions of OEs in the different combinations were determined then the OE and the most active combination were retained to determine their kinetic parameters on S. flexneri. Results: Antibiotic susceptibility tests revealed that most Shigella isolates (n = 4) were resistant to six antibiotics tested. Ciprofloxacin (40%), Nalidixic acid (60%), Tetracycline (80%), Amoxicillin (100%), Cefotaxime (80%), Erythromycin (100%), and Cotrimoxazole (80%) were the profiles found in the different strains of Shigella. About the antibacterial activity of OEs, Drypetes gossweileriOE and C2 combination had shown a higher Shigellicide property with a Minimal Inhibitory Concentration(MIC) respectivelyranging from 0.078 mg/mL to 0.312 mg/mL and 0.012 to 1.562 mg/mL. Combinations of OEs showed various interactions whose synergistic effects were mostly encountered. The best deactivation was obtained by the combination C2 at 16 MIC withb= 1.962. Conclusion: the susceptibility of Shigella to OEs and their combinations justifies their use in traditional medicine in the treatment of shigellosis.

Keywords: shigella, multidrug-resistant, EOs, kinetic

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Authors: Goksen Arik, Mihriban Korukluoglu

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Lactic acid bacteria (LAB) has been used commonly in the food industry. They can be used as natural preservatives because acidifying carried out in the medium can protect the last product against microbial spoilage. Besides, other metabolites produced by LAB during fermentation period have also an antimicrobial effect on pathogen and spoilage microorganisms in the food industry. LAB are responsible for the desirable and distinctive aroma and flavour which are observed in fermented food products such as pickle, kefir, yogurt, and cheese. Various LAB strains are able to produce B-group vitamins such as folate (B11), riboflavin (B2) and cobalamin (B12). Especially wild-type strains of LAB can produce B-group vitamins in high concentrations. These cultures may be used in food industry as a starter culture and also the microbial strains can be used in encapsulation technology for new and functional food product development. This review is based on the current applications of B-group vitamin producing LAB. Furthermore, the new technologies and innovative researches about B vitamin production in LAB have been demonstrated and discussed for determining their usage availability in various area in the food industry.

Keywords: B vitamin, food industry, lactic acid bacteria, starter culture, technology

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Authors: Ramesh Subramani, Mathivanan Narayanasamy, William Aalbersberg

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It is well accepted by last 35 years of research and on-going programmes that marine environment harbours abundant and unique biodiversity, which is currently playing as an important source in bioprospecting. It has become apparent that marine microorganisms are lead in the biodiscovery. Among marine organisms, actinobacteria are a target phylum for discovering novel antibiotics against increasing the multi-drug resistant human pathogens because of these taxa representing for novel genera and species. Marine actinomycetes are a proven source of new antibiotic leads and novel enzymes with important industrial applications. A total of 183 streptomycete and 25 non-streptomycete strains were isolated from different marine samples collected from north-eastern part of the Indian Ocean. Among them, 111 isolates displayed antibacterial activity against human pathogens and 151 exhibited antifungal activity against phytopathogens. Importantly, most of them produced various extracellular enzymes and 58 of them produced exopolysaccharides. Totally eight small bioactive compounds and a thermostable alkaline protease have been purified from a selected strain, Streptomyces fungicidicus. Besides, our on-going studies on non-streptomycete strains (rare actinomycetes) are most likely promising resource for new and unique compounds against current emerging drug-resistant pathogens. We have just recognised the chemical diversity in marine microorganisms. Therefore it is worthwhile to continue the exploration of marine microorganisms for new drug leads, novel enzymes and other bioprospecting research.

Keywords: bioactive compounds, industrial enzymes, marine actinobacteria, microbial metabolites, marine natural products

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Authors: Darnetty Baharuddin Salleh

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Fusarium ear rot disease is one of the most important diseases of maize and not only causes significant losses but also produced harmful mycotoxins to animals and humans. A total of 141 strains of Fusarium species were isolated from maize plants showing typical ear rot symptoms in Indonesia, and Malaysia by using the semi-selective medium (peptone pentachloronitrobenzene agar, PPA). These strains were identified morphologically. For strains in Gibberella fujikuroi species complex (Gfsc), the identification was continued by using biological identification. Three species of Fusarium were morphologically identified as Fusarium in Gibberella species complex (105 strains, 74.5%), F. verticillioides (78 strains), F. proliferatum (24 strains) and F. subglutinans (3 strains) and five species from other section (36 strains, 25.5%), F. graminearum (14 strains), F. oxysporum (8 strains), F. solani ( 1 strain), and F. semitectum (13 strains). Out of 105 Fusarium species in Gfsc, 63 strains were identified as MAT-1, 25 strains as MAT-2 and 17 strains could not be identified and in crosses with nine standard testers, three mating populations of Fusarium were identified as MP-A, G. moniliformis (68 strains, 64.76%), MP-D, G. intermedia (21 strains, 20%) and MP-E, G. subglutinans (3 strains, 2.9%), and 13 strains (12.38%) could not be identified. All trains biologically identified as MP-A, MP-D, and MP-E, were identified morphologically as F. verticillioides, F. proliferatum, and F. subglutinans, respectively. Thus, the results of this study indicated that identification based on biological identification were consistent with those of morphological identification. This is the first report on the presence of MP-A, MP-D, and MP-E on ear rot-infected maize in Indonesia; MP-A and MP-E in Malaysia.

Keywords: Fusarium, MAT-1, MAT-2, MP-A, MP-D, MP-E

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Authors: Sadaf Ilyas, Saba Riaz

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The extensive use of antibiotics has led to increases emergence of antibiotic-resistant organisms. Pseudomonas is a notorious opportunistic pathogen involoved in nosocomial infections and exhibit innate resistance to many antibiotics. The present study was conducted to assess the prevalence, levels of antimicrobial susceptibility and resistance mechanisms of Pseudomonas. A total of thirty clinical strains of Pseudomonas were isolated from different clinical sites of infection. All clinical specimens were collected from Chughtais Lahore Lab. Jail road, during 8-07-2010 to 11-01-2011. Biochemical characterization was done using routine biochemical tests. Antimicrobial susceptibility was determined by Kirby-Baeur method. The plasmids were isolated from all the strains and digested with restriction enzyme PstI and EcoRI. Transfer of Multi-resistance plasmid was checked via transformation and conjugation to confirm the plasmid mediated resistance to antibiotics. The prevalence of Pseudomonas in clinical specimens was found out to be 14% of all bacterial infections. IPM has shown to be the most effective drug against Pseudomonas followed by CES, PTB and meropenem, wheareas most of the Pseudomonas strains have developed significant resistance against Penicillins and some Cephalasporins. Antibiotic resistance determinants were carried by plasmids, as they conferred resistance to transformed K1 strains. The isolates readily undergo conjugation, transferring the resistant genes to other strains, illustrating the high rates of cross infection and nosocomial infection in the immunocompromised patients.

Keywords: pseudomonas, antibiotics, drug resistance, horizontal gene transfer

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Authors: Julie Connelly, Tanvi Toprani, Xin Xie, Dhinoth Kumar Bangarusamy, Kris C. Gunsalus

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The overuse of antibiotics worldwide has contributed to the development of multi-drug resistant (MDR) pathogenic bacterial strains. There is an increasing urgency to discover antibiotics to combat MDR pathogens. The microbiome of the Arabian Gulf is a largely unexplored and potentially rich source of novel bioactive compounds. Microbes that inhabit the Abu Dhabi coastal regions adapt to extreme environments with high salinity, hot temperatures, large temperature fluctuations, and acute exposure to solar energy. The microbes native to this region may produce unique metabolites with therapeutic potential as antibiotics and antifungals. We have isolated 200 pure bacterial strains from mangrove sediments, cyanobacterial mats, and coral reefs of the Abu Dhabi region. In this project, we aim to screen the marine bacterial strains to identify antibiotics, in particular undocumented compounds that show activity against existing antibiotic-resistant strains. We have acquired the ESKAPE pathogen panel, which consists of six antibiotic-resistant gram-positive and gram-negative bacterial pathogens that collectively cause most clinical infections. Our initial efforts of the primary screen using colony-picking co-culture assay have identified several candidate marine strains producing potential antibiotic compounds. We will next apply different assays, including disk-diffusion and broth turbidity growth assay, to confirm the results. This will be followed by bioactivity-guided purification and characterization of target compounds from the scaled-up volume of candidate strains, including SPE fraction, HPLC fraction, LC-MS, and NMR. For antimicrobial compounds with unknown structures, our final goal is to investigate their mode of action by identifying the molecular target.

Keywords: marine bacteria, natural products, drug discovery, ESKAPE panel

Procedia PDF Downloads 75