Transcription profiling of Density-dependent of Xanthomonas oryzae pv. oryzae PXO99A and PXO99∆ax21
ABSTRACT: Xanthomonas oryzae pv. oryzae strain PXO99A, so called Xoo, is disable to infect in rice cultivar carrying Xa21 gene. A set of experiments indicated that Ax21 is quorum sensing factor in PXO99 strain. To observe fine-tuned details how Ax21 controls expression of PXO99 in response to change of cell population density, transcriptional profiling analysis was perform by using published two channel oligo Xo microarray platform (Seo et al.,2008 BMC microbiology). Keywords: Comparative transcription profiling between low cell density and high cell density with same genetic background Three biological and dye-swap replicates, total six samples for each dataset. Two datasets contained; (i) PXO99 at 106 CFU/ml vs 108 CFU/ml, and (ii) PXO99∆ax21at 106 CFU/ml vs PXO99∆ax21 108 CFU/ml. 106 CFU/ml and 108 CFU/ml are cell density used in this study as representation of low and high cell density. All bacteria cultures were grown in PS (Peptone sucrose) broth media.
Project description:Xanthomonas oryzae pv. oryzae strain PXO99A, so called Xoo, is disable to infect in rice cultivar carrying Xa21 gene. A set of experiments indicated that Ax21 is quorum sensing factor in PXO99 strain. To observe fine-tuned details how Ax21 controls expression of PXO99 in response to change of cell population density, transcriptional profiling analysis was perform by using published two channel oligo Xo microarray platform (Seo et al.,2008 BMC microbiology). Keywords: Comparative transcription profiling between low cell density and high cell density with same genetic background Overall design: Three biological and dye-swap replicates, total six samples for each dataset. Two datasets contained; (i) PXO99 at 106 CFU/ml vs 108 CFU/ml, and (ii) PXO99∆ax21at 106 CFU/ml vs PXO99∆ax21 108 CFU/ml. 106 CFU/ml and 108 CFU/ml are cell density used in this study as representation of low and high cell density. All bacteria cultures were grown in PS (Peptone sucrose) broth media.
Project description:The objective of this study was to determine the comparative pharmacodynamics of four different carbapenems in combination with polymyxin B (PMB) against carbapenem-resistant Acinetobacter baumannii isolates using time-kill experiments at two different inocula. Two A. baumannii strains (03-149-1 and N16870) with carbapenem minimum inhibitory concentrations (MICs) ranging from 8 to 64?mg/L were investigated in 48-h time-kill experiments using starting inocula of 106?CFU/mL and 108?CFU/mL. Concentration arrays of ertapenem, doripenem, meropenem and imipenem at 0.25×, 0.5×, 1×, 1.5× and 2× published maximum serum concentration (Cmax) values (Cmax concentrations of 12, 21, 48 and 60?mg/L, respectively) were investigated in the presence of 1.5?mg/L PMB. Use of carbapenems without PMB resulted in drastic re-growth. All carbapenem combinations were able to achieve a ?3 log10 CFU/mL reduction by 4?h against both strains at 106?CFU/mL, whereas maximum reductions against strain 03-149-1 at 108?CFU/mL were 1.0, 3.2, 2.2 and 3.3 log10 CFU/mL for ertapenem, doripenem, meropenem and imipenem, respectively. None of the combinations were capable of reducing 108?CFU/mL of N16870 by ?2 log10 CFU/mL. Ertapenem combinations consistently displayed the least activity, whereas doripenem, meropenem and imipenem combinations had similar activities that were poorly predicted by carbapenem MICs. As doripenem, meropenem, or imipenem displayed similar pharmacodyanmics in combination, the decision of which carbapenem to use in combination with PMB may be based on toxicodynamic profiles if drastic discordance in MICs is not present.
Project description:Kefir is a functional beverage that contains lactic and acetic acid bacteria (LAB, AAB) and yeasts. This work's aim was to study the chemical, microbial, and functional characteristics of kefir produced from cow's milk and soy milk. After fermentation, free amino acids were 20.92?mg 100?mL-1 and 36.20?mg 100?mL-1 for cow's milk and soy milk kefir, respectively. Glutamic acid was majority in both, suggesting that microbial proteolysis leads to an increase in free amino acids including glutamic acid. 108-109?CFU?mL-1 LAB, 106-107?CFU?mL-1 AAB, and 106-107?CFU?mL-1 yeasts were counted in cow's milk kefir, whereas soy milk kefir contained greatly lower yeasts and AAB. Lactococcus lactis, Kazachstania unispora, and Saccharomyces cerevisiae were isolated as major microorganisms in both kefirs. Acetobacter orientalis only existed in cow's milk kefir. Cow's milk and soy milk showed ACE inhibitory activity, which significantly increased after fermentation. Both kefirs also exhibited antioxidant activity and bactericidal activity against Escherichia coli, Salmonella Typhimurium, and Staphylococcus aureus.
Project description:Thirty-four strains of bacteria were isolated from Phellodendron amurense. Using Nectria haematococca as an indicator strain, the best strain, B18, was obtained by the growth rate method. The morphological, physiological and biochemical characteristics of strain B18 and its 16S DNA gene sequence were identified, and the biocontrol effect of strain B18 was assessed in pot and field tests, as well as in a field-control test. Drilling methods were used to determine the antibacterial activity of metabolites from strain B18 and their effects on the growth of pathogen mycelia and spores. The best bacteriostatic rate was 85.4%. B18 can hydrolyse starch and oxidize glucose but does not produce gas; a positive result was obtained in a gelatine liquefaction test. According to 16S DNA gene sequencing, strain B18 is Bacillus methylotrophicus (GenBank accession number: MG457759). The results of pot and field-control trials showed 98% disease control when inoculating 108 cfu/ml of the strain. The disease control effect of the B18 culture liquid (concentrations of 108, 2 × 106, 106, 5 × 105 and 2.5 × 105 cfu/ml) in the field-control test was higher than 80%, and the cure rate of the original delivery solution was 96%. Therefore, in the practical forestry production, a 2.5 × 105 cfu/ml culture liquidshould be applied in advance to achieve good control effects.
Project description:Infections due to carbapenem-resistant NDM-producing Escherichia coli represent a major therapeutic challenge, especially in situations of pre-existing colistin resistance. The aim of this study was to investigate combinatorial pharmacodynamics of colistin and tigecycline against E. coli harboring bla NDM- 5 and mcr-1, with possible mechanisms explored as well. Colistin disrupted the bacterial outer-membrane and facilitated tigecycline uptake largely independent of mcr-1 expression, which allowed a potentiation of the tigecycline-colistin combination. A concentration-dependent decrease in colistin MIC and EC50 was observed with increasing tigecycline levels. Clinically relevant concentrations of colistin and tigecycline combination significantly decreased bacterial density of colistin-resistant E. coli by 3.9 to 6.1-log10 cfu/mL over 48 h at both inoculums of 106 and 108 cfu/mL, and were more active than each drug alone (P < 0.01). Importantly, colistin and tigecycline combination therapy was efficacious in the murine thigh infection model at clinically relevant doses, resulting in >2.0-log10cfu/thigh reduction in bacterial density compared to each monotherapy. These data suggest that the use of colistin and tigecycline combination can provide a therapeutic alternative for infection caused by multidrug-resistant E. coli that harbored both bla NDM- 5 and mcr-1.
Project description:Listeria monocytogenes is a common cause of bacterial meningitis. We developed an animal model of listerial meningitis.In survival studies, C57BL/6 mice received intracisternal injections with different L. monocytogenes sequence type 1 (ST1) colony forming units per milliliter (CFU; n?=?48, 105, 106, 107, 108, and 109 CFU/ml). Second, mice were inoculated with 108 CFU/ml ST1 and sacrificed at 6 h and 24 h (n?=?12/group). Outcome parameters were clinical score, CFUs, cyto- and chemokine levels, and brain histopathology. Third, 84 mice were inoculated (109 CFU/ml ST1) to determine optimal antibiotic treatment with different doses of amoxicillin and gentamicin. Fourth, mice were inoculated with 109 CFU/ml ST1, treated with amoxicillin, and sacrificed at 16 h and 24 h (n?=?12/group) for outcome assessment. Finally, time point experiments were repeated with ST6 (n?=?24/group).Median survival time for inoculation with 108 and 109 CFU/ml ST1 was 46 h and 40 h; lower doses of bacteria led to minimal clinical signs of disease. Brain levels of IL-6, IL-17A, and IFN-? were elevated at 24 h, and IL-1?, IL-6, IL-10, IFN-?, and TNF-? were elevated in blood at 6 h and 24 h. Histopathology showed increased meningeal infiltration, vascular inflammation of meningeal vessels, hemorrhages, and ventriculitis. In the treatment model, brain levels of IL-6 and IL-17A and blood levels of IL-6 and IFN-? were elevated. Compared to ST6, infection with ST1 led initially to higher levels of IL-1? and TNF-? in blood and more profound neuropathological damage. At 16 h post inoculation, IL-1?, IL-10, and TNF-? in blood and IL-6, IL17A, TNF-?, and IFN-? levels in brain were higher in ST1 compared to ST6 without differences in CFUs between STs. At 24 h, neuropathology score was higher in ST1 compared to ST6 (p?=?0.002) infected mice.We developed and validated a murine model of listerial meningitis. ST1-infected mice had a more severe inflammatory response and brain damage as compared to ST6-infected mice.
Project description:The influence of spore concentration on the ability of a Trichoderma consortium to colonize the Passiflora caerulea phyllosphere was evaluated by determining the effects of foliar treatments with two spore concentrations, in two repeated treatments, on the morphological, physiological, and ultrastructural characteristics, and on the yield and quality of P. caerulea. The studied crop quality features were related to its nutraceutical use: the accumulation of polyphenols and flavonoids, antioxidant activity, and effects on mouse fibroblast L929 cells. The Trichoderma consortium consisted of two strains, T. asperellum T36b and T. harzianum Td50b, and the concentrations used were 106 colony forming units (cfu)/mL and 108 cfu/mL. As a reference treatment, a commercial product that was based on herbs and algal extracts was used. As compared to the negative control, the treatment with the Trichoderma consortium at 108 cfu/mL concentration determines the accumulation of higher level of polyphenols and flavonoids and increased antioxidant activity. This enhancement of P. caerulea quality characteristics after treatment with the higher concentration of Trichoderma consortium was associated with larger leaves, increased number and size of chloroplasts, improved plant physiology characteristics, and an increased yield. The treatment with high concentration of Trichoderma consortium spores promotes phyllosphere colonization and benefits both crop yield and quality.
Project description:Galleria mellonella larvae were administered an inoculum of Candida albicans and the cellular and proteomic response to infection over the first 24 hours was monitored. The yeast cell density in infected larvae declined initially but replication commenced six hours post-infection. The hemocyte density decreased from 5.2 × 106/ml to 2.5 × 106/ml at 2 hours but increased to 4.2 × 106 at 6 hours but decreased subsequently. Administration of β - glucan to larvae also resulted in an increase in hemocyte density (5.1 ± 0.22 × 106/ml to 6.25 ± 0.25 × 106/ml, p < 0.05) and the population showed an increase in the density of small, granular type cells at 24 hours (p < 0.05). Hemocytes from larvae inoculated with β - glucan showed faster killing of C. albicans cells (53 ± 4.1% (p < 0.01), 64 ± 3.7%, (p < 0.01), respectively) than hemocytes from control larvae (24 ± 11%) at 60 min. Proteomic analysis indicated increased abundance of immune related proteins cecropin-A (5 fold) and prophenoloxidase-activating proteinase-1 (5 fold) 6 hours post infection but by 24 hours there was elevated abundance of muscle (tropomyosin 2 (141 fold), calponin (66 fold), troponin I (62 fold), troponin T (61 fold)) and proteins indicative of cellular stress (glutathione-S-transferase-like protein (114 fold)), fungal dissemination (muscle protein 20-like protein (174 fold)) and tissue breakdown (mitochondrial cytochrome c (10 fold)). Proteins decreased in abundance at 24 hour included β - 1,3 - glucan recognition protein precursor (29 fold), prophenoloxidase subunit 2 (25 fold) and lysozyme-like protein 1 (5 fold). By characterizing the early responses to infection, G. mellonella larvae may be used to model the initial stages of infection processes in mammals.
Project description:Postharvest diseases of potato lead to significant food and economic losses worldwide. The exogenous application of eco-friendly methods plays an important role in the control of postharvest decay. In this work the effects of endophytic bacteria B. subtilis (10-4, 26D) were studied in the context of two application parameters: concentration, with a range between 103-108 CFU/mL tested, and synergistic effects of the signal molecule salicylic acid (SA) (0.05 mM) on potato tubers' resistance to Phytophthora infestans and Fusarium oxysporum during storage. The experiments were carried out on hydroponically grown potato (Solanum tuberosum L.) mini-tubers. This study demonstrates the suppressive effect of B. subtilis (10-4, 26D) on diseases of potato during storage and reveals that this effect happens in a dose-dependent manner, both individually and in combination with SA. The most effective concentrations of B. subtilis for suppression of both Ph. infestans and F. oxysporum are 108 CFU/mL (10-4 and 26D), 107 CFU/mL (10-4 + SA) and 106 CFU/mL (26D + SA). The ability of B. subtilis (10-4, 26D) to effectively penetrate and colonize the internal tubers' tissues when applied immediately prior to storage, and the ability of SA to accelerate these processes, have been proven. B. subtilis (10-4, 26D), individually and in compositions with SA, increased ascorbic acid content and decreased pathogen-induced proline accumulation and lipid peroxidation in tubers. This indicates a protective effect conferred to cells against reactive oxygen and an extension of aging processes, manifested by a prolonged shelf life and extended preservation of fresh appearance.
Project description:Xanthomonas oryzae pv. oryzae strain PXO99A, so called Xoo, is disable to infect in rice cultivar carrying Xa21 gene. Disrupted mutant of raxR gene, response regulator of two-component regulatory system (TCS), in Xoo was previously shown to partially retrieve back the bacterial capability to establish in Xa21 rice. RaxR was shown to mediate the expression of other rax gene operon members and also its expression is changed dependent on cell population density. In this study, we investigated the regulatory mechanisms mediated by RaxR using whole-genome transcriptional profiling analysis in comparison of (i) PXO99R (PXO99 strain lacking RaxR) vs. PXO99, (ii) PXO99Rox (PXO99 strain overexpressing RaxR) vs. PXO99, and (iii) PXO99Rox vs. PXO99R. As a result of array analysis, we revealed that RaxR is not only required for AvrXa21 activitiy, it also plays roles in regulatory functions, for example, pathogenicity, motility, and stress tolerance. Then, we generated knock out mutants of RaxR regulon members to validate regulatory functions of RaxR and to extend other biological impacts of RaxR beyond the Xoo AvrXa21 activity. The combined interpretation from array analysis and mutant functional validation presents the complexity of regulatory pathways between AvrXa21 activity and other biological activities in Xoo. Keywords: Comparative transcription profiling between modified genetic mutant and wild type Three biological and dye-swap replicates, total six samples for each dataset. Three datasets contained; (i) raxR knockout mutant vs. wildtype PXO99A in PSB, nutrient rich media (ii) RaxR constitutively express mutant vs. wildtype PXO99A in PSB, nutrient rich media, and (iii) raxR knockout mutant vs. RaxR constitutively express mutant in PSB, nutrient rich media