Multi-drug resistant ESKAPE pathogens and the uses of plants as their antimicrobial agents.

Infections by ESKAPE (Enterococcus sp., Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.) pathogens cause major concern due to their multi-drug resistance (MDR). The ESKAPE pathogens are frequently linked to greater mortality, diseases, and economic burden in healthcare worldwide. Therefore, the use of plants as a natural source of antimicrobial agents provide a solution as they are easily available and safe to use. These natural drugs can also be enhanced by incorporating silver nanoparticles and combining them with existing antibiotics. By focussing the attention on the ESKAPE organisms, the MDR issue can be addressed much better.

Genetic and Chemical Screening Reveals Targets and Compounds to Potentiate Gram-Positive Antibiotics against Gram-Negative Bacteria.

Gram-negative bacteria are intrinsically resistant to a plethora of antibiotics that effectively inhibit the growth of Gram-positive bacteria. The intrinsic resistance of Gram-negative bacteria to classes of antibiotics, including rifamycins, aminocoumarins, macrolides, glycopeptides, and oxazolidinones, has largely been attributed to their lack of accumulation within cells due to poor permeability across the outer membrane, susceptibility to efflux pumps, or a combination of these factors. Due to the difficulty in discovering antibiotics that can bypass these barriers, finding targets and compounds that increase the activity of these ineffective antibiotics against Gram-negative bacteria has the potential to expand the antibiotic spectrum. In this study, we investigated the genetic determinants for resistance to rifampicin, novobiocin, erythromycin, vancomycin, and linezolid to determine potential targets of antibiotic-potentiating compounds. We subsequently performed a high-throughput screen of ∼50,000 diverse, synthetic compounds to uncover molecules that potentiate the activity of at least one of the five Gram-positive-targeting antibiotics. This led to the discovery of two membrane active compounds capable of potentiating linezolid and an inhibitor of lipid A biosynthesis capable of potentiating rifampicin and vancomycin. Furthermore, we characterized the ability of known inhibitors of lipid A biosynthesis to potentiate the activity of rifampicin against Gram-negative pathogens.

Can Gram-Negative Bacteria Develop Resistance to Antimicrobial Blue Light Treatment?

Antimicrobial blue light (aBL) treatment is considered low risk for the development of bacterial resistance and tolerance due to its multitarget mode of action. The aim of the current study was to demonstrate whether tolerance development occurs in Gram-negative bacteria. We evaluated the potential of tolerance/resistance development in Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa and demonstrated that representative Gram-negative bacteria may develop tolerance to aBL. The observed adaption was a stable feature. Assays involving E. coli K-12 tolC-, tolA-, umuD-, and recA-deficient mutants revealed some possible mechanisms for aBL tolerance development.

An Efflux Pumps Inhibitor Significantly Improved the Antibacterial Activity of Botanicals from Plectranthus glandulosus towards MDR Phenotypes.

Bacterial multidrug resistance causes many therapeutic failures, making it more difficult to fight against bacterial diseases. This study aimed to investigate the antibacterial activity of extract, fractions, and phytochemicals from Plectranthus glandulosus (Lamiaceae) against multidrug-resistant (MDR) Gram-negative phenotypes expressing efflux pumps. The crude extract after extraction was subjected to column chromatography, and the structures of the isolated compounds were determined using spectrometric and spectroscopic techniques. Antibacterial assays of samples alone and in the presence of an efflux pump inhibitor (phenylalanine-arginine ß-naphthylamide, PAßN) were carried out using the broth microdilution method. The phytochemical study of P. glandulosus plant extract afforded seven major fractions (A-G) which lead to the isolation of seventeen known compounds. The ethanol extract of P. glandulosus was not active at up to 1024 µg/mL, whereas its fractions showed MICs varying from 32 to 512 µg/mL on the studied bacteria. Fraction C of P. glandulosus showed the lowest MIC (32 µg/mL) on E. coli ATCC8739 strain. Fraction D presented the highest activity spectrum by inhibiting the growth of 90% (9/10) of the studied bacteria. The presence of PAßN has improved the activity of extract and all fractions. Overall, the tested phytochemicals showed low activity against the studied bacteria. The overall results obtained in this study show that some fractions from P. glandulosus, mainly fractions C and D, should be investigated more for their possible use to fight against MDR bacteria.

Genetic diversity and risk factors for the transmission of antimicrobial resistance across human, animals and environmental compartments in East Africa: a review.

BACKGROUND: The emergence and spread of antimicrobial resistance (AMR) present a challenge to disease control in East Africa. Resistance to beta-lactams, which are by far the most used antibiotics worldwide and include the penicillins, cephalosporins, monobactams and carbapenems, is reducing options for effective control of both Gram-positive and Gram-negative bacteria. The World Health Organization, Food and Agricultural Organization and the World Organization for Animal Health have all advocated surveillance of AMR using an integrated One Health approach. Regional consortia also have strengthened collaboration to address the AMR problem through surveillance, training and research in a holistic and multisectoral approach. This review paper contains collective information on risk factors for transmission, clinical relevance and diversity of resistance genes relating to extended-spectrum beta-lactamase-producing (ESBL) and carbapenemase-producing Enterobacteriaceae, and Methicillin-resistant Staphylococcus aureus (MRSA) across the human, animal and environmental compartments in East Africa. MAIN BODY: The review of the AMR literature (years 2001 to 2019) was performed using search engines such as PubMed, Scopus, Science Direct, Google and Web of Science. The search terms included 'antimicrobial resistance and human-animal-environment', 'antimicrobial resistance, risk factors, genetic diversity, and human-animal-environment' combined with respective countries of East Africa. In general, the risk factors identified were associated with the transmission of AMR. The marked genetic diversity due to multiple sequence types among drug-resistant bacteria and their replicon plasmid types sourced from the animal, human and environment were reported. The main ESBL, MRSA and carbapenem related genes/plasmids were the blaCTX-Ms (45.7%), SCCmec type III (27.3%) and IMP types (23.8%), respectively. CONCLUSION: The high diversity of the AMR genes suggests there may be multiple sources of resistance bacteria, or the possible exchange of strains or a flow of genes amongst different strains due to transfer by mobile genetic elements. Therefore, there should be harmonized One Health guidelines for the use of antibiotics, as well as regulations governing their importation and sale. Moreover, the trend of ESBLs, MRSA and carbapenem resistant (CAR) carriage rates is dynamic and are on rise over time period, posing a public health concern in East Africa. Collaborative surveillance of AMR in partnership with regional and external institutions using an integrated One Health approach is required for expert knowledge and technology transfer to facilitate information sharing for informed decision-making.

Consequences Of Long-Term Bacteria's Exposure To Silver Nanoformulations With Different PhysicoChemical Properties.

PURPOSE: Resistance to antibiotics is a major problem of public health. One of the alternative therapies is silver - more and more popular because of nanotechnology development and new possibilities of usage. As a component of colloid, powder, cream, bandages, etc., nanosilver is often recommended to treat the multidrug-resistant pathogens and we can observe its overuse also outside of the clinic where different physicochemical forms of silver nanoformulations (e.g. size, shape, compounds, surface area) are introduced. In this research, we described the consequences of long-term bacteria exposure to silver nanoformulations with different physicochemical properties, including changes in genome and changes of bacterial sensitivity to silver nanoformulations and/or antibiotics. Moreover, the prevalence of exogenous resistance to silver among multidrug-resistant bacteria was determined. MATERIALS AND METHODS: Gram-negative and Gram-positive bacteria strains are described as sensitive and multidrug-resistant strains. The sensitivity of the tested bacterial strains to antibiotics was carried out with disc diffusion methods. The sensitivity of bacteria to silver nanoformulations and development of bacterial resistance to silver nanoformulations has been verified via determination of the minimal inhibitory concentrations. The presence of sil genes was verified via PCR reaction and DNA electrophoresis. The genomic and phenotypic changes have been verified via genome sequencing and bioinformatics analysis. RESULTS: Bacteria after long-term exposure to silver nanoformulations may change their sensitivity to silver forms and/or antibiotics, depending on the physicochemical properties of silver nanoformulations, resulting from phenotypic or genetic changes in the bacterial cell. Finally, adaptants and mutants may become more sensitive or resistant to some antibiotics than wild types. CONCLUSION: Application of silver nanoformulations in the case of multiple resistance or multidrug-resistant bacterial infection can enhance or decrease their resistance to antibiotics. The usage of nanosilver in a clinic and outside of the clinic should be determined and should be under strong control. Moreover, each silver nanomaterial should be considered as a separate agent with a potential different mode of antibacterial action.

Choice of therapeutic interventions and outcomes for the treatment of infections caused by multidrug-resistant gram-negative pathogens: a systematic review.

Background: Antimicrobial resistance is an increasingly serious threat to public health, and the increased occurrence of multidrug-resistant (MDR) bacteria is a concern in both high-income and low- and middle-income countries. The purpose of this systematic review was to identify and critically appraise current antimicrobial treatment options for infections with MDR Gram-negative bacteria. Methods: A literature search for treatment of MDR extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae, A. baumannii, and P. aeruginosa was conducted in MEDLINE in January 2019. Relevant studies published in English, German, and French that evaluated clinical success, microbiological success, and 30-day mortality outcomes were included. The population of interest was adult patients. Results: Of 672 studies, 43 met the inclusion criteria. Carbapenems are the most common antibiotics used for the treatment of ESBL-producing Enterobacteriaceae. The clinical and microbiological success was similar for group 1 carbapenems (imipenem, meropenem, or doripenem), group 2 carbapenems (ertapenem), and non-carbapenem antibiotics. Mortality data were contradictory for group 1 carbapenems compared to group 2 carbapenems. The most common treatment option for A. baumannii and P. aeruginosa infections was intravenous colistin, regardless of infection site. Clinical success and mortality were similar in A. baumannii infections treated with colistin combination therapy vs. colistin monotherapy, whereas heterogeneous results were found with respect to microbiological success. Monotherapy and colistin combination therapy were used against P. aeruginosa with clinical and microbiological success (70-100%) depending on the infection site and severity, and the antibiotic used. Ceftazidime-avibactam therapy for ESBL-producing Enterobacteriaceae and P. aeruginosa showed good clinical success in one study. Conclusion: We did not find robust evidence for antibiotic treatment of any infection with MDR Gram-negative bacteria, including ESBL-producing Enterobacteriaceae, A. baumannii, and P. aeruginosa, that would lead to a firm recommendation for one specific antibiotic over another or for monotherapy over combination therapy. The choice of antibiotic treatment should be based on susceptibility testing balancing the expected clinical success rate against the risk of development of antibiotic resistance and the risk of severe side effects.

High Prevalence of Antimicrobial Resistance in Gram-Negative Bacteria Isolated from Clinical Settings in Egypt: Recalling for Judicious Use of Conventional Antimicrobials in Developing Nations.

This study was designed to investigate, at the molecular level, the antimicrobial resistance mechanisms of different antimicrobial resistance genes, including, extended-spectrum ß-lactamases, AmpC ß-lactamases, class 1 and 2 integrons, and plasmid-mediated quinolone resistance genes of Gram-negative bacteria isolated from clinical settings in Egypt. A total of 126 nonduplicate Gram-negative isolates were recovered from different clinical samples taken from hospitalized patients in Egypt in 2014. Antimicrobial susceptibility testing showed that, 93.6% (118/126) of the isolates had a multidrug-resistant phenotype. Interestingly, we reported a high level of antimicrobial resistance nearly for all tested antibiotics; to our knowledge, this is the first report from Egypt indicating very high level of antibiotic resistance in Egypt. Polymerase chain reaction screening and DNA sequencing revealed that, 75.4% (95/126) of the isolates harbored at least one extended-spectrum ß-lactamase-encoding gene, with blaCTX-M being the most prevalent (65.9%), followed by blaSHV (46.8%). The AmpC ß-lactamase, blaCMY, was detected in 7.1% (9/126) of bacterial isolates, with blaCMY-42 being the most prevalent. Class 1 integrons were detected in 50.8% (64/126) of the isolates, and class 2 integrons were detected in 2.4% (3/126) of the isolates. The plasmid-mediated quinolone resistance gene, qnr, was detected in 58.7% (74/126) of the tested isolates, with qnrS being the most prevalent. Several antimicrobial resistance determinants were identified in Egypt for the first time, such as SHV-27, SHV-28, SHV-33, SHV-63, SHV-71, SHV-82, SHV-142, CMY-42, CMY-6, and the new CMY-72 like. This study highlights the importance of the conscious use of conventional antimicrobials to overcome the multidrug resistance problem.

Prevalence of metallo-ß-lactamase acquired genes among carbapenems susceptible and resistant Gram-negative clinical isolates using multiplex PCR, Khartoum hospitals, Khartoum Sudan.

BACKGROUND: The increased prevalence of carbapenem-resistant Gram-negative isolates caused by Metallo-ß-lactamase (MBL) is worrisome in clinical settings worldwide. The mortality rate associated with infections caused by MBLs producing organisms ranging from 18 to 67%. This study aimed to determine the prevalence of Metallo-ß-lactamase genes among some Gram-negative clinical isolates (Carbapenems susceptible and resistant). METHODS: This paper describes a descriptive cross-sectional study carried out to detect MBL genes such as (blaVIM, blaIMP and blaNDM) by multiplex PCR mixture reaction among 200 Gram-negative clinical isolates (Citrobacter spp, Escherichia coli, Enterobacter spp, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis, Proteus valgaris). Khartoum hospitals during 2015 to 2016. LIMITATION: The study organisms were not evaluated for non-MBL carbapenemases, such as KPC and OXA-48. RESULTS: The prevalence of MBL genes by multiplex PCR assays among 200 Gram-negative clinical isolates was 72(36.1%). MBL positive genes among 100 carbapenems sensitive and 100 resistant isolates were 27(27%) and 45(45%) respectively. There was a statistically, significant association between the antimicrobial susceptibility and the presences of MBL genes (P.value = 0.008). E.coli was the predominant species possessing MBL genes 26(36.1%), with 22(30.7%) species having a combination of MBL genes. Verona integron Metallo beta-lactamase (VIM) was the most frequent genes 28(38.9%) out of 72 MBL detected genes, followed by imipenemase (IMP) was 19(26.4%), and consequently, New Delhi Metallo beta lactamase was 3(4.2%). CONCLUSION: This study revealed a high prevalence of MBL genes in some Gram-negative isolates from Khartoum State Hospitals which were not previously established in these hospitals.

Microbiota of Honeybees, Apis mellifera Adansonii (Hymenoptera: Apidae) from Selected Ecozones, South West Nigeria.

BACKGROUND AND OBJECTIVES: Honeybees explore their environment in search of nectar and pollens including water whose sources are contaminated by variety of microbes. This has both negative and positive economic implications on health status of honeybees, the consumers of the products including the quality of the products. Hence, the study was designed to investigate the effect of geographical locations on the occurrence and diversity of microbiota on the external integument and the digestive gut of adult worker honeybees, Apis mellifera adansonii. MATERIALS AND METHODS: The honeybee samples were sourced from Ondo, Ogun, Oyo and Osun of South West Nigeria. The samples were subjected to microbiological analyses using standard techniques. Two-ways ANOVA was used for statistical test of the data. RESULTS: The microbiota of the studied samples includes Corynebacterium kutsceri, Corynebacterium xerosis, Lactobacillus bulgaricus, Lactobacillus casei, Lactobacillus fermentum, Micrococcus luteus, Micrococcus varians and Staphylococcus aureus which were gram-positive bacteria, while gram-negative bacteria include Aeromonas veronii, Citrobacter diversus, Citrobacter freundii, Klebsiella oxytoca, Klebsiella pneumonia and Pseudomonas aeruginosa. Fungi isolates include Aspergillus parasiticus, Aspergilus flavus, Aspergilus fumigatus, Aspergilus niger and Rhizopus stolonifer. Most microbiota were not cosmopolitan across selected ecozones of south west, Nigeria except for Aspergillus fumigatus. The number of identified species of microbiota either in the digestive guts or on the external integument of worker honeybees from selected ecozones varies from one ecozone to another. There was significant species diversity of microbiota on external integument than in digestive guts of adult workers of A. mellifera adansonii from selected ecozones in South West, Nigeria. CONCLUSION: The study showed that both the external integument and the digestive guts of A. mellifera adansonii exhibited variety of microbial flora providing research exploration opportunity for probable microbes of economic and geographical importance.

Glyphosate application increased catabolic activity of gram-negative bacteria but impaired soil fungal community.

Glyphosate is a non-selective organophosphate herbicide that is widely used in agriculture, but its effects on soil microbial communities are highly variable and often contradictory, especially for high dose applications. We applied glyphosate at two rates: the recommended rate of 50 mg active ingredient kg-1 soil and 10-fold this rate to simulate multiple glyphosate applications during a growing season. After 6 months, we investigated the effects on the composition of soil microbial community, the catabolic activity and the genetic diversity of the bacterial community using phospholipid fatty acids (PLFAs), community level catabolic profiles (CLCPs), and 16S rRNA denaturing gradient gel electrophoresis (DGGE). Microbial biomass carbon (Cmic) was reduced by 45%, and the numbers of the cultivable bacteria and fungi were decreased by 84 and 63%, respectively, under the higher glyphosate application rate. According to the PLFA analysis, the fungal biomass was reduced by 29% under both application rates. However, the CLCPs showed that the catabolic activity of the gram-negative (G-) bacterial community was significantly increased under the high glyphosate application rate. Furthermore, the DGGE analysis indicated that the bacterial community in the soil that had received the high glyphosate application rate was dominated by G- bacteria. Real-time PCR results suggested that copies of the glyphosate tolerance gene (EPSPS) increased significantly in the treatment with the high glyphosate application rate. Our results indicated that fungi were impaired through glyphosate while G- bacteria played an important role in the tolerance of microbiota to glyphosate applications.

Treatment of infections caused by multidrug-resistant Gram-negative bacteria: report of the British Society for Antimicrobial Chemotherapy/Healthcare Infection Society/British Infection Association Joint Working Party.

The Working Party makes more than 100 tabulated recommendations in antimicrobial prescribing for the treatment of infections caused by multidrug-resistant (MDR) Gram-negative bacteria (GNB) and suggest further research, and algorithms for hospital and community antimicrobial usage in urinary infection. The international definition of MDR is complex, unsatisfactory and hinders the setting and monitoring of improvement programmes. We give a new definition of multiresistance. The background information on the mechanisms, global spread and UK prevalence of antibiotic prescribing and resistance has been systematically reviewed. The treatment options available in hospitals using intravenous antibiotics and in primary care using oral agents have been reviewed, ending with a consideration of antibiotic stewardship and recommendations. The guidance has been derived from current peer-reviewed publications and expert opinion with open consultation. Methods for systematic review were NICE compliant and in accordance with the SIGN 50 Handbook; critical appraisal was applied using AGREE II. Published guidelines were used as part of the evidence base and to support expert consensus. The guidance includes recommendations for stakeholders (including prescribers) and antibiotic-specific recommendations. The clinical efficacy of different agents is critically reviewed. We found there are very few good-quality comparative randomized clinical trials to support treatment regimens, particularly for licensed older agents. Susceptibility testing of MDR GNB causing infection to guide treatment needs critical enhancements. Meropenem- or imipenem-resistant Enterobacteriaceae should have their carbapenem MICs tested urgently, and any carbapenemase class should be identified: mandatory reporting of these isolates from all anatomical sites and specimens would improve risk assessments. Broth microdilution methods should be adopted for colistin susceptibility testing. Antimicrobial stewardship programmes should be instituted in all care settings, based on resistance rates and audit of compliance with guidelines, but should be augmented by improved surveillance of outcome in Gram-negative bacteraemia, and feedback to prescribers. Local and national surveillance of antibiotic use, resistance and outcomes should be supported and antibiotic prescribing guidelines should be informed by these data. The diagnosis and treatment of both presumptive and confirmed cases of infection by GNB should be improved. This guidance, with infection control to arrest increases in MDR, should be used to improve the outcome of infections with such strains. Anticipated users include medical, scientific, nursing, antimicrobial pharmacy and paramedical staff where they can be adapted for local use.

New therapy from old drugs: synergistic bactericidal activity of sulfadiazine with colistin against colistin-resistant bacteria, including plasmid-mediated colistin-resistant mcr-1 isolates.

The recent emergence of colistin (COL) resistance, particularly mcr-1 plasmid-mediated COL resistance in Gram-negative bacteria, has led to renewed interest in antibiotic combinations to overcome clinical therapeutic impasses. The aim of this study was to evaluate the potential of the synergistic and bactericidal activity of COL in combination with sulphonamide compounds, including sulfadiazine (SDI), sulfamethoxazole (SMX) and trimethoprim/sulfamethoxazole (SXT), as well as trimethoprim (TMP) against clinical COL-resistant bacterial strains, including strains with the plasmid-encoded mcr-1 gene. A collection of 55 COL-resistant and -susceptible strains from different origins (Laos, Thailand and France) was used in this study. Several in vitro methods were used to determine the potential of the synergistic activity of these combinations, including Etest on agar pre-treated plates, the Etest cross method and the chequerboard assay. A time-kill assay was performed to evaluate the potential bactericidal activity of combinations in addition to synergistic activity. Significant synergistic activity was observed with all combinations tested. The combination of COL + SDI presented the highest synergistic effect against the various species of COL-resistant strains (92.7%). For the other combinations, a synergistic effect was also observed but with lower frequency for COL + SMX (33.3%), COL + TMP (47.3%) and COL + SXT (31.5%). Synergy was observed independently of the COL resistance mechanism. These in vitro results suggest that the combination of COL + SDI would appear to be justifiable in patients with multidrug-resistant bacterial infections that cannot be treated with COL monotherapy.

Antibacterial activity of Ilex paraguariensis (Yerba Mate) against Gram-positive and Gram-negative bacteria.

INTRODUCTION: The stems and leaves of Ilex paraguariensis are popularly used for tea infusions in South America and the Middle East. The health benefits have been previously studied, revealing anti-mutagenic, anti-oxidant, hepatoprotective, hypocholesteremic and glycemic improvement. Limited research was performed on the antibacterial activity of the aqueous extract of Yerba Mate on standard and clinical isolates of Gram-positive and Gram-negative bacteria. METHODOLOGY: Commercial Ilex paraguariensis stems and leaves were ground and extracted with sterile deionized water at 70°C. Four ATCC bacterial strains and twenty-five bacterial clinical strains were used for testing. To obtain the minimal inhibitory concentration (MIC), the Yerba Mate aqueous solution was serially diluted according to the microdilution method. For the minimal bactericidal concentration (MBC), the tubes with clear broth were sub-cultured. To identify the types of ESBLs present in the clinical isolates, a multiplex PCR was performed. RESULTS: An antibacterial activity was observed against most of tested strains, with a greater activity against Gram-positive bacteria. MIC and MBC values ranged between 0.468 mg/mL and 15 mg/mL of aqueous extract of Yerba Mate. CONCLUSION: The aqueous extract of the stems and leaves of Ilex paraguariensis extracted at 70°C showed a significant antibacterial activity. There was no correlation found between the different molecular resistance profiles and the antibacterial activity range.

Characterization of the Serralysin-Like Gene of 'Candidatus Liberibacter solanacearum' Associated with Potato Zebra Chip Disease.

The nonculturable bacterium 'Candidatus Liberibacter solanacearum' is the causative agent of zebra chip disease in potato. Computational analysis of the 'Ca. L. solanacearum' genome revealed a serralysin-like gene based on conserved domains characteristic of genes encoding metalloprotease enzymes similar to serralysin. Serralysin and other serralysin family metalloprotease are typically characterized as virulence factors and are secreted by the type I secretion system (T1SS). The 'Ca. L. solanacearum' serralysin-like gene is located next to and divergently transcribed from genes encoding a T1SS. Based on its relationship to the T1SS and the role of other serralysin family proteases in circumventing host antimicrobial defenses, it was speculated that a functional 'Ca. L. solanacearum' serralysin-like protease could be a potent virulence factor. Gene expression analysis showed that, from weeks 2 to 6, the expression of the 'Ca. L. solanacearum' serralysin-like gene was at least twofold higher than week 1, indicating that gene expression stays high as the disease progresses. A previously constructed serralysin-deficient mutant of Serratia liquefaciens FK01, an endophyte associated with insects, as well as an Escherichia coli lacking serralysin production were used as surrogates for expression analysis of the 'Ca. L. solanacearum' serralysin-like gene. The LsoA and LsoB proteins were expressed as both intact proteins and chimeric S. liquefaciens-'Ca. L. solanacearum' serralysin-like proteins to facilitate secretion in the S. liquefaciens surrogate and as intact proteins or as a truncated LsoB protein containing just the putative catalytic domains in the E. coli surrogate. None of the 'Ca. L. solanacearum' protein constructs expressed in either surrogate demonstrated proteolytic activity in skim milk or zymogram assays, or in colorimetric assays using purified protein, suggesting that the 'Ca. L. solanacearum' serralysin-like gene does not encode a functional protease, or at least not in our surrogate systems.

Antibiofilm efficacy of green synthesized graphene oxide-silver nanocomposite using Lagerstroemia speciosa floral extract: A comparative study on inhibition of gram-positive and gram-negative biofilms.

Biofilm architecture provides bacteria with enhanced antibiotic resistance, thus raising the need to search for alternative therapies that can inhibit the bacterial colonization. In the present study, we synthesized graphene oxide-silver nanocomposite (GO-Ag) by non-toxic and eco-friendly route using a floral extract of Legistromia speciosa (L.) Pers. The gas chromatography-mass spectrometry (GC-MS) analysis of plant extract revealed the presence of compounds which can simultaneously act as reducing and capping agents. The sub-inhibitory concentrations of synthesized GO-Ag reduced the biofilm formation in both gram-negative (E. cloacae) and gram-positive (S. mutans) bacterial models. Growth curve assay, membrane integrity assay, scanning electron microscopy (SEM) and confocal scanning laser microscopy (CSLM) revealed different mechanisms of biofilm inhibition in E. cloacae and S. mutans. Moreover, quantitative RT-PCR (qRT-PCR) results suggested GO-Ag is acting on S. mutans biofilm formation cascade. Biofilm inhibitory concentrations GO-Ag were also found to be non-toxic against HEK-293 (human embryonic kidney cell line). The whole study highlights the therapeutic potential of GO-Ag to restrain the onset of biofilm formation in bacteria.

Antimicrobial Activities of Ceftazidime-Avibactam and Comparator Agents against Gram-Negative Organisms Isolated from Patients with Urinary Tract Infections in U.S. Medical Centers, 2012 to 2014.

A total of 7,272 unique patient clinical isolates were collected from 71 U.S. medical centers from patients with urinary tract infections in 2012 to 2014 and tested for susceptibility to ceftazidime-avibactam and comparators by broth microdilution methods. Ceftazidime-avibactam inhibited >99.9% of all Enterobacteriaceae at the susceptible breakpoint of ≤8 µg/ml (there were only three nonsusceptible strains). Ceftazidime-avibactam was also active against Pseudomonas aeruginosa isolates (MIC50, 2 µg/ml; MIC90, 4 µg/ml; 97.7% susceptible), including many isolates not susceptible to meropenem, ceftazidime, and/or piperacillin-tazobactam.

Lutibacter profundi sp. nov., isolated from a deep-sea hydrothermal system on the Arctic Mid-Ocean Ridge and emended description of the genus Lutibacter.

A bacterial strain designated LP1T was isolated from a microbial mat growing on the surface of a black smoker chimney at the Loki's Castle hydrothermal system, which is located on the Arctic Mid-Ocean Ridge. Phylogenetic analyses based on 16S rRNA gene sequences positioned strain LP1T within the family Flavobacteriaceae with Lutibacterholmesii as the closest relative (97.5 % 16S rRNA gene sequence similarity). Strain LP1T was rod-shaped, Gram-reaction-negative and non-motile. It grew in a modified artificial seawater medium supplemented with tryptone and vitamins at pH 5.5-7.5 (optimum pH 6.0-6.5), within a temperature range of 13-34 °C (optimum 23 °C), and under microaerobic conditions. The most abundant fatty acids (>10 %) were iso-C15 : 0 (25.2 %) and iso-C15 : 0 3-OH (14.5 %). The genome of strain LP1T has a DNA G+C content of 29.8 mol%. Based on the results of the polyphasic characterization presented here, strain LP1T is considered to represent a novel species of the genus Lutibacter, for which the name Lutibacter profundi sp. nov. is proposed. The type strain is LP1T (=DSM 100437T =JCM 30585T). An emended description of the genus Lutibacter is also provided to fit the description of strain LP1T.

Evaluation of microbial population and functional genes during the bioremediation of petroleum-contaminated soil as an effective monitoring approach.

This study investigated the abundance and diversity of soil n-alkane and polycyclic aromatic hydrocarbon (PAH)-degrading bacterial communities. It also investigated the quantity of the functional genes, the occurrence of horizontal gene transfer (HGT) in the identified bacterial communities and the effect that such HGT can have on biostimulation process. Illumina sequencing was used to detect the microbial diversity of petroleum-polluted soil prior to the biostimulation process, and quantitative real-time PCR was used to determine changes in the bacterial community and functional genes (alkB, phnAc and nah) expressions throughout the biostimulation of petroleum-contaminated soil. The illumine results revealed that γ-proteobacteria, Chloroflexi, Firmicutes, and δ-proteobacteria were the most dominant bacterial phyla in the contaminated site, and that most of the strains were Gram-negative. The results of the gene expression results revealed that gram-negative bacteria and alkB are critical to successful bioremediation. Failure to maintain the stability of hydrocarbon-degrading bacteria and functional gene will reduce the extend to which alkanes and PAHs are degraded. According to the results of the study, the application of a C:N:P ratio of was 100:15:1 in the biodegradation experiment resulted in the highest rate at which petroleum hydrocarbons were biodegraded. The diversity of pollutant-degrading bacteria and the effective transfer of degrading genes among resident microorganisms are essential factors for the successful biostimulation of petroleum hydrocarbons. As such, screening these factors throughout the biostimulation process represents an effective monitoring approach by which the success of the biostimulation can be assessed.

Antibiotic treatment algorithm development based on a microarray nucleic acid assay for rapid bacterial identification and resistance determination from positive blood cultures.

Rapid diagnosis of bloodstream infections remains a challenge for the early targeting of an antibiotic therapy in sepsis patients. In recent studies, the reliability of the Nanosphere Verigene Gram-positive and Gram-negative blood culture (BC-GP and BC-GN) assays for the rapid identification of bacteria and resistance genes directly from positive BCs has been demonstrated. In this work, we have developed a model to define treatment recommendations by combining Verigene test results with knowledge on local antibiotic resistance patterns of bacterial pathogens. The data of 275 positive BCs were analyzed. Two hundred sixty-three isolates (95.6%) were included in the Verigene assay panels, and 257 isolates (93.5%) were correctly identified. The agreement of the detection of resistance genes with subsequent phenotypic susceptibility testing was 100%. The hospital antibiogram was used to develop a treatment algorithm on the basis of Verigene results that may contribute to a faster patient management.