Concentration Of Bacterial Suspension Research Articles

Microbiological Mechanisms of Collaborative Remediation of Cadmium-Contaminated Soil with Bacillus cereus and Lawn Plants.

Severe cadmium contamination poses a serious threat to food security and human health. Plant-microbial combined remediation represents a potential technique for reducing heavy metals in soil. The main objective of this study is to explore the remediation mechanism of cadmium-contaminated soil using a combined approach of lawn plants and microbes. The target bacterium Bacillus cereus was selected from cadmium-contaminated soil in mining areas, and two lawn plants (Festuca arundinacea A'rid III' and Poa pratensis M'idnight II') were chosen as the target plants. We investigated the remediation effect of different concentrations of bacterial solution on cadmium-contaminated soil using two lawn plants through pot experiments, as well as the impact on the soil microbial community structure. The results demonstrate that Bacillus cereus promotes plant growth, and the combined action of lawn plants and Bacillus cereus improves soil quality, enhancing the bioavailability of cadmium in the soil. At a bacterial suspension concentration of 105 CFU/mL, the optimal remediation treatment was observed. The removal efficiency of cadmium in the soil under Festuca arundinacea and Poa pratensis treatments reached 33.69% and 33.33%, respectively. Additionally, the content of bioavailable cadmium in the rhizosphere soil increased by up to 13.43% and 26.54%, respectively. Bacillus cereus increased the bacterial diversity in the non-rhizosphere soil of both lawn plants but reduced it in the rhizosphere soil. Additionally, the relative abundance of Actinobacteriota and Firmicutes, which have potential for heavy metal remediation, increased after the application of the bacterial solution. This study demonstrates that Bacillus cereus can enhance the potential of lawn plants to remediate cadmium-contaminated soil and reshape the microbial communities in both rhizosphere and non-rhizosphere soils.

The optimized microbial-induced calcium carbonate precipitation process to fabricate underwater superoleophobic mesh for efficient oil-water separation

Microbial-induced calcium carbonate precipitation (MICP) is an environmentally-friendly and easy operation technique for separating oil-water mixtures. However, the effect of environmental factors on the wettability and oil-water separation performance of MICP remains unclear. In this study, three primary factors including mineralization solution (MS) concentration, bacterial suspension (BS) concentration, and temperature were carefully investigated. Under these varying conditions, stainless steel meshes (SSM) were chosen as the substrates for the growth of CaCO3 precipitations. Experimental results demonstrate a strong dependence of the morphology and wettability of CaCO3 precipitations on these factors. Higher MS concentration and temperature contribute to enhanced hydrophilicity of CaCO3 coatings due to the increased surface roughness. The water flux of MICP-coated SSMs decreases with the increase of BS concentration, MS concentration, and temperature. The MICP-coated SSM obtained with MS concentration of 0.15 M, BS concentration of 0.1 × 108 cells/mL, and mineralization reaction at 25 ℃ for 24 hours demonstrated the optimal comprehensive performance, exhibiting underwater superoleophobicity with an underwater oil contact angle (UWOCA) exceeding 151°, an ultra-high oil intrusion pressure (> 3.0 kPa), ultrahigh flux (28,644 L m−2 h−1), and a preferable oil rejection rate (> 99.9%). Additionally, the as-prepared mesh exhibited superior anti-oil-fouling property and reusability, while concurrently demonstrating excellent chemical and thermal stability. This work contributes to advancing our knowledge of the influence of various environmental factors on the wetting characteristics of MICP and provides a feasible solution for the highly efficient and low-cost preparation of oil-water separation membranes based on the MICP process.

Identification, Biocontrol Activity, and Field Application Effect of Bacillus velezensis Yb-1

Colletotrichum capsici is one of the most important pathogens on chili peppers. Unreasonable application of chemical fungicides will lead to threats to human and animal health, environmental damage, and increased fungicide resistance to pathogens. As an alternative strategy, biological control has been paid more and more attention by academics. In this study, the Yb-1 strain was isolated from healthy cucumber leaves, which could significantly restrict the mycelium growth of C. capsici and Botrytis cinerea, with inhibition rates of 93.49% and 74.03%, respectively. Strain Yb-1 was identified as Bacillus velezensis by morphological features and 16S rRNA gene, gyrA gene, and gyrB gene sequence analysis. When pepper seeds were treated with different concentrations of bacterial suspension, we found that a medium-concentration treatment (104 CFU/mL and 105 CFU/mL) could promote seed germination and growth, while high-concentration treatments (108 CFU/mL) limited seed germination and growth. In addition, the root-irrigation method, acupuncture-leaf method, and leaf-cutting method were used to evaluate the safety of strain Yb-1 on peppers. The results indicated that Yb-1 did not affect the normal growth of peppers. The results of further field trials showed that the control effect of strain Yb-1 on pepper anthracnose was 59.45%. Thus, the strain B. velezensis Yb-1 has great biocontrol potential for pepper anthracnose and promotes plant growth.

Agrobacterium rhizogenes mediated genetic transformation in Ficus religiosa L. and optimization of acetylcholinesterase inhibitory activity in hairy roots

The use of decoctions prepared from stem bark to treat neurodegenerative problems is amongst the widespread applications of Ficus religiosa L., a tree of great ethno-medicinal importance in the Indian sub-continent. The stem bark of the tree has been reported to have significant acetylcholinesterase inhibitory activity (AChEI). Latter is one of the most effective modalities used for the treatment of Alzheimer's disease (AD). The development of any commercial treatment modality for AD from this tree faces serious challenges for sample consistency in terms of quality (being strongly influenced by environmental and developmental stages) and quantity. Hairy root (HR) cultures are an excellent choice for such studies. Since no study has been reported for HR culture systems for F. religiosa , the present study was targeted to develop one. Three different HR lines were induced and cultured on different media. Root extracts, obtained from HRs on different media, were subjected to an AChEI assay to select the best conditions where HRs exerted higher AChEI. The wild-type strain of A. rhizogenes was used to induce hairy roots in different explants. The method of genetic transformation was optimised by changing concentrations of bacterial suspension, acetosyringone and calcium chloride in the infection and co-cultivation medium. The assay for measuring AChEI activity was modified from the assays described by Ellman et al. (1961) and Ingkaninan et al. (2000) . These methods recommend the use of IC 50 (half maximal inhibitory concentration) value for comparing AChEI activities in different samples. We developed a protocol for induction and long-term maintenance of HR cultures of F. religiosa L. In vitro conditions were optimized where hairy root extracts exerted higher AChEI activity (IC 50 value of 0.16 µg/ml) compared to that in the stem segments of the mother plants (IC 50 value of 0.49 µg/ml). The present work has been granted a patent by the Indian Patent Office, Government of India (Patent No360050, Date of grant 02/03/2021) (Siwach and Gill, 2021). The present study confirms the utility of F. religiosa in AD treatment and provides a standardized protocol for large-scale production of HR cultures (75ml medium in Erlenmeyer flask of 250 ml volume) that have higher AChEI activity; these cultures offer excellent platforms for future characterization and commercial preparation.

Identification of Two Bacillus Strains with Antimicrobial Activity and Preliminary Evaluation of Their Biocontrol Efficiency

Cabbage Fusarium Wilt (CFW) is a serious disease caused by Fusarium oxysporum f. sp. conglutinans in many parts of the world. The use of chemical fungicides has placed a heavy burden on the environment and is prone to drug resistance in plant pathogens. As a method with great potential, biological control has attracted the attention of many academics both at home and abroad. In this study, we have found that strains B5 and B6 had a strong inhibitory effect on various pathogens and significantly inhibited mycelium growth. They were both identified as Bacillus velezensis by morphological features, biochemical determinations, 16S rRNA gene and gyrA gene sequence analysis. When different concentrations of bacterial suspension were applied to cabbage seeds, hypocotyl and taproot length increased to varying degrees. The in vivo results showed that B5 and B6 decreased the incidence of cabbage seedling wilt disease, with B6 performing significantly better. Furthermore, B. velezensis B6 had the ability to colonize cabbage plants and rhizosphere soil. Thus, strain B6 has great potential for biocontrol development and this research could lead to the development of a promising biological agent for CFW.

In-Vitro and In-Vivo Tolerance and Therapeutic Investigations of Phyto-Fabricated Iron Oxide Nanoparticles against Selected Pathogens.

The Paeonia emodi (P. emodi)-mediated iron oxide nanoparticles (Fe2O3 NPs) were screened for in-vitro and in-vivo antibacterial activity against the Staphylococcus aureus (S. aureus) (ATCC #: 6538) and Escherichia coli (E. coli) (ATCC #:15224). The synthesized Fe2O3 NPs were characterized via nitrogen adsorption-desorption process, X-ray diffractometer (XRD), transmission and scanning electron microscopies (TEM and SEM), energy dispersive X-ray (EDX) and Fourier transform infrared (FTIR) spectroscopies. The SBET was found to be 94.65 m2/g with pore size of 2.99 nm, whereas the average crystallite and particles size are 23 and 27.64 nm, respectively. The 4 μg/mL is the MIC that inhibits the growth of E. coli, whereas those for S. aureus are below the detection limit (<1.76 μg/mL). The tolerance limit of the mice model was inspected by injecting different concentration of Fe2O3 NPs and bacteria suspensions. The 14 ppm suspension was the tolerated dose and the concentration above were proved lethal. The most severe infection was induced in mice with injection of 3 × 107 CFUs of both bacteria, while the inoculation of higher concentrations of bacterial suspensions resulted in the mice’s death. The histopathological and hematological studies reveals that the no/negligible infection was found in the mice exposed to the simultaneous inoculation of Fe2O3 NPs (14 ppm) and bacterial suspensions (3 × 107 CFUs).

Optimization of operational conditions using the experimental design method to remove Escherichia coli from contaminated groundwater by an adsorbent material prepared from Moroccan oil shales

The aim of the present study is to investigate the removal of Escherichia coli (E. coli) strain from contaminated groundwater using an adsorbent material prepared from Moroccan oil shales extracted in the Timahdit region (layer X). Several factors were studied and optimized using the experimental design method based on the Plackett-Burman Experimental plan (PBE); namely, the bacterial suspension concentration (105–108 CFU/mL), the adsorbent mass (1–30 mg), the pH (3–9), the temperature (20–37 °C), the contact time (5–360 min) and the stirring speed (45–150 rpm). Data analysis, by the PBE model, showed that the bacterial suspension concentration, the adsorbent mass and the stirring speed are the most significant factors that influence the removal of E. coli. The optimum process conditions were found with a suspension concentration below 5 × 107 CFU/mL, about 15.5 mg of the adsorbent material and a stirring speed over 97.5 rpm at 28.5 °C and pH 6 during 182 min. Under these conditions, the adsorption efficiency of E. coli reached 70.57%.

Effect of Particle Size on Mechanical Property of Bio-Treated Sand Foundation

In the field of geotechnical engineering, microbially induced calcium precipitation technology is feasible and sustainable alternative to improve the engineering characteristics of sand foundation under different geological conditions for a long time. However, it is unclear how the effects of different sand particle sizes on the engineering characteristics of bio-treated sand column. The method of intermittent injection in batches was used to develop a series of bio-treated sand columns. The results showed that the mechanical properties of the bio-treated column improved by increasing the particle size. Low concentration of bacterial suspension and cementation reagent leads to the increase of calcium carbonate and unconfined compressive strength. Additionally, the total injection times increased, thus risking time cost. Furthermore, the increase of sand particle size was beneficial to the uniformity of the spatial distribution of calcium carbonate in the bio-treated column. The coefficient of variation was reduced by up to 52.0%. Scanning electron microscopy results confirmed that the size and uniformity of calcite crystals on the surface of sand particles were related to the concentration of cementation solution.

AI-Blue-Carba: A Rapid and Improved Carbapenemase Producer Detection Assay Using Blue-Carba With Deep Learning.

A rapid and accurate detection of carbapenemase-producing Gram-negative bacteria (CPGNB) has an immediate demand in the clinic. Here, we developed and validated a method for rapid detection of CPGNB using Blue-Carba combined with deep learning (designated as AI-Blue-Carba). The optimum bacterial suspension concentration and detection wavelength were determined using a Multimode Plate Reader and integrated with deep learning modeling. We examined 160 carbapenemase-producing and non-carbapenemase-producing bacteria using the Blue-Carba test and a series of time and optical density values were obtained to build and validate the machine models. Subsequently, a simplified model was re-evaluated by descending the dataset from 13 time points to 2 time points. The best suitable bacterial concentration was determined to be 1.5 optical density (OD) and the optimum detection wavelength for AI-Blue-Carba was set as 615 nm. Among the 2 models (LRM and LSTM), the LSTM model generated the higher ROC-AUC value. Moreover, the simplified LSTM model trained by short time points (0–15 min) did not impair the accuracy of LSTM model. Compared with the traditional Blue-Carba, the AI-Blue-Carba method has a sensitivity of 95.3% and a specificity of 95.7% at 15 min, which is a rapid and accurate method to detect CPGNB.

Self-cleaning and flexible filters based on aminopyridine conjugated microporous polymers nanotubes for bacteria sterilization and efficient PM2.5 capture

The capture and elimination of harmful particulate matter (PM) both in air and water is of great importance for human health and environmental sustainability. Here, we demonstrate a novel strategy for the exploitation of conjugated microporous polymer bearing aminopyridine moiety (A-CMPs) as an advanced filter for bacteria sterilization and efficient PM capture. The A-CMPs network shows a hierarchically porous structure with mechanical robustness and flexibility, which facilitates to filtration especially for PM with different particle sizes. The capture efficiency of A-CMPs aerogels for PM2.5 and PM10 were respectively up to PM2.5 ≥ 99.57 ± 0.19% and PM10 ≥ 99.98 ± 0.01% in a long-term durability test and easy to be regenerated. Moreover, the A-CMPAs features excellent superhydrophobicity, which is difficult to saturate with water aerosols in humid air (RH: 89 ± 3%) and in turn shows superior stability and high-performance in terms to capture efficiency. More importantly, the A-CMP monolith exhibits excellent antimicrobial activity and high concentrations of bacterial suspension (e.g., using E. coli as probe bacterial) could be effectively captured and quickly killed during filtration, which endows the A-CMPs additional sterilization performance and thus is of great technological significance with remarkable potentials as a new kind of advanced filter for multifunctional filtration in both air and water.

Promising Bacillus subtilis strains BZR 336g and BZR 517 for biocontrol of blackcurrant against Septoria leaf spot under unfavorable climate conditions

Septoria leaf spot is the most serious blackcurrant disease in Western Siberia. Two Bacillus subtilis strains BZR 336g and BZR 517 isolated in geographically distant locality (south of Russia) were evaluated for the blackcurrant disease suppression in field experiments (Western Siberia). The field trials were conducted on two cultivars differed by their susceptibility to Septoria ribis in two consecutive years differed by environmental conditions. As a rule, plant treatment with bacterial suspensions (105 and 106 CFU/ml) significantly (P < 0.05) reduced the disease severity compared with the untreated blackcurrant plants. In some cases, the effect of bacterial strains was comparable to that of chemical fungicide Topaz, CE 0.1%. Biological control of blackcurrant disease caused by S. ribis depended on B. subtilis strain, concentration of bacterial suspension, plant cultivars, and weather conditions. For the first time, we showed that B. subtilis strains isolated in a region with warm climate effectively suppressed the main blackcurrant disease in geographically distant locality with cold climate.

Septicaemia of subterranean termites Coptotermes curvignathus caused by disturbance of bacteria isolated from termite gut and its foraging pathways.

Microbial pathogens continue to attract a great deal of attention to manage the termite population. Every bacterium has its own mode of action and in fact, the mechanisms used by bacteria to attack termites remain elusive at the moment. Hence, the objective of this study was to evaluate the susceptibility of subterranean termites Coptotermes curvignathus to opportunistic pathogens using culturable aerobic bacteria isolated from the termite gut and its foraging pathways. Bacterial suspensions were prepared in concentrations of 103, 106 and 109 colony-forming units (CFU) ml−1 and introduced to the termites via oral-contact and physical contact treatment. The data show that contact method acted slower and gave lower mortality, compared to the oral-contact method. Coptotermes curvignathus were highly susceptible to Serratia marcescens and Pseudomonas aeruginosa. Serratia marcescens showed the highest mortality percentage of 68% and 54% at bacterial concentration of 109 CFU ml−1 via oral-contact and contact method, respectively. Serratia marcescens was also defined as the bacteria with the highest ability to induce the high mortality of C. curvignathus with the lowest concentration of bacterial suspension at a given time under laboratory condition. The results of this study indicate that P. aeruginosa and S. marcescens in particular may be attractive candidates worth further examination as a possible biocontrol agent against C. curvignathus in the field and to evaluate environmental and ecological risks of the biocontrol.

Development of a new protocol for freeze-drying preservation of Pseudoalteromonas nigrifaciens and its protective effect on other marine bacteria

Abstract Background Freeze-drying is known as one of the best methods to preserve bacterial strains. Protectant is the key factor affecting the survival rate of freeze-dried strains. In addition, salinity, bacterial suspension concentration, drying time, and other factors can also affect the survival rate of strains to varying degrees. At present, there are relatively few studies on freeze-drying preservation of marine bacteria. In the present study, we performed the freeze-drying protectant screening and optimized the preservation conditions for Pseudoalteromonas nigrifaciens, which is widely distributed in marine environment. The protective effects of the screened protectants were verified by 18 other marine bacterial strains. Results The results indicated that the combination of 5.0% (w/v) lactose, 5.0% (w/v) mannitol, 5.0% (w/v) trehalose, 10.0% (w/v) skim milk powder, 0.5% (w/v) ascorbic acid and 0.5% (w/v) gelatin was the best choice for the preservation of P. nigrifaciens. The suggested salinity and concentration of initial cell suspension were 10 g/L NaCl and 1.0 × 109 CFU/mL, respectively. Furthermore, stationary-phase cells were the best choice for the freeze-drying process. The highest survival rate of P. nigrifaciens reached 52.8% when using 5–10% (w/v) skim milk as rehydration medium. Moreover, the other 18 marine strains belonging to Pseudoalteromonas, Vibrio, Photobacterium, Planomicrobium, Edwardsiella, Enterococcus, Bacillus, and Saccharomyces were freeze-dried under the abovementioned conditions. Their survival rates were 2.3–95.1%. Conclusion Collectively, our results supported that the protectant mixture and parameters were beneficial for lyophilization of marine bacteria. How to cite: Zhang Z, Yu Y, Wang Y, et al. Development of new protocol for freeze-drying preservation of Pseudoalteromonas nigrifaciens and its protective effect on other marine bacteria. Electron J Biotechnol 2020;44. https://doi.org/10.1016/j.ejbt.2019.12.006 .

First isolation of Photobacterium damselae subsp. damselae from half-smooth tongue sole suffering from skin-ulceration disease

Climate change will likely promote the emergence of the mesophilic bacteria Photobacterium damselae subsp. damselae as a pathogen affecting global economic marine fish farming. In this study, we identified links between P. damselae subsp. damselae and skin ulcerations in commercially farmed half-smooth tongue sole (Cynoglossus semilaevis) in the Bohai Bay area. Nearly circular ulcerations appeared on the pigmented side of the diseased fish and were caused by damage to the epidermis, dermis, and muscular layers, with significant congestion found in the brain, liver, and intestine accompanied by significant pathological changes. Physiological and biochemical characterization and 16S rRNA gene sequencing identified six kinds of bacteria isolated from diseased fish, with artificial-infection testing and histopathologic analysis revealing P. damselae subsp. damselae as the cause of death and the variable clinical symptoms (abdominal swelling and skin ulcers) observed following injection with different concentrations of bacterial suspension (~104–106 CFU mL−1). Additionally, histological analysis showed that the pathological changes induced by the bacterial suspensions were similar to those found in naturally diseased fish; however, the degree of damage differed according to time after infection. Moreover, antimicrobial-susceptibility testing showed that P. damselae subsp. damselae exhibited different degrees of resistance to 11 of 13 types of antibacterial drugs but strong susceptibility to chloramphenicols, suggesting their efficacy for first-line treatment of diseased fish.

In-vitro antibacterial and anti-encrustation performance of silver-polytetrafluoroethylene nanocomposite coated urinary catheters

Catheter-associated urinary tract infections (CAUTIs) are among the most common hospital-acquired infections, leading to increased morbidity and mortality. A major reason for this is that urinary catheters are not yet capable of preventing CAUTIs. To develop an anti-infective urinary catheter. An efficient silver-polytetrafluoroethylene (Ag-PTFE) nanocomposite coating was deposited on whole silicone catheters, and two in-vitro bladder models were designed to test antibacterial (against Escherichia coli) and anti-encrustation (against Proteus mirabilis) performances. Each model was challenged with two different concentrations of bacterial suspension. Compared with uncoated catheters, coated catheters significantly inhibited bacterial migration and biofilm formation on the external catheter surfaces. The time to develop bacteriuria was an average of 1.8 days vs 4 days and 6 days vs 41 days when the urethral meatus was infected with 106 and 102 cells/mL, respectively. For anti-encrustation tests, the coated catheter significantly resisted encrustation, although it did not strongly inhibit the increases in bacterial density and urinary pH. The time to blockage, which was found to be independent of the initial bacterial concentration in the bladder, was extended from 36.2±1.1 h (uncoated) to 89.5±3.54 h (coated) following bacterial contamination with 103 cells/mL in the bladder. Moreover, the coated catheter exhibited excellent biocompatibility with L929 fibroblast cells. Ag-PTFE coated Foley catheters should undergo further clinical trials to determine their ability to prevent CAUTIs during catheterization.

Root promoting effect and its regulation mechanisms of endophytic Bacillus amyloliquefaciens YTB1407 in sweet potato

We isolated the endophytic Bacillus amyloliquefaciens YTB1407 from the root of Panax quinquefolium, which has both biological control and growth promoting effects. To investigate its potential applications, a pot experiment of sweet potato was tested to assess the capacity of endophytic colonization of YTB1407 and the selection of its optimum concentration by investigating the performance of root characteristics on three time points in the whole early growth phase after irrigating with different concentrations of bacterial suspensions with treatment of sterile water as control. The activities of endogenous hormone IAA, ZR, t-ZR and IAA oxidase (IAAO, PPO, POD) were analyzed. The results showed that YTB1407 promoted the specific colonization of root system, the elongation of adventitious root and branch roots, and root activity in the early growth stage of sweet potato. At later growth stage, it formed greater fresh mass of absorption root and lower aboveground/root system mass ratio. YTB1407 suspensions with OD600 of 0.50 (T0.50) had more significant effect, which induced the highest fresh tuber mass and the largest effective tuber numbers of per plant at top cover stage. YTB1407 promoted the differentiation of adventitious roots into tubers at initial point of tuberization by increasing IAA content and the ratio of (t-ZR+ZR)/IAA, decreasing IAAO activity and enhancing PPO activity. Moreover, it promoted the differentiated roots into tubers at tuberization stage by keeping the higher content of IAA, lower ratio of (t-ZR+ZR)/IAA, and decreasing IAAO and PPO activities.

Ecological applications of Pseudomonas as a biopesticide to control two-spotted mite Tetranychus urticae: chitinase and HCN production

Abstract The two-spotted spider mite (Tetranychus urticae Koch) is an important pest of many horticultural crops. A study was conducted to evaluate the effect of three fluorescent Pseudomonas isolates obtained from rhizospheric soil of tomato (Solanum lycopersicon) in Agadir, Morocco: Q110B, Q036B and Q172B, as potential biological control agents for T. urticae. Both acaricide and repellent activities were assessed on homogenous adult mites. The acaricidal activity test evaluated five concentrations of bacterial suspensions: 0 (control), 102, 104, 106, 108, and 1010 cfu · ml−1, while only the 1010 cfu ∙ ml−1 concentration of each bacterium was used for the repellent bioassay. The mortality rate and repellentindex were recorded 24, 48 and 72 h after application. Results indicated that the survival rate of T. urticae was reduced (p ≤ 0.01) by all three bacterial isolates compared to control. Within the 24–72 h time period the mortality rates ranged from 8 to 87%, 16 to 99%, and 13 to 89%, for Q110B, Q036B and Q172B isolates, respectively. The isolate Q036B (LC50 = 0.598 cfu · ml−1) provided higher mortality rates than Q172B and Q110B with LC50 values of 90,846 and 169,585 cfu · ml−1, respectively. Repellent activity was also the highest with Pseudomonas Q036B having a 71% repellence index at 48 h after application. Regarding the mechanism of action, all three isolates produced hydrogen cyanide, and exhibited protease and cellulose activities, although only Q036B and Q172B had potential chitinase action. Identification analysis showed that the isolates were either Pseudomonas putida (Q172B) or P. fluorescens (Q110B and Q036B). Our results indicate that the P. fluorescens isolate Q036B is a promising candidate for biological control of T. urticae, and has potential to contribute to an integrated pest management program to control this important pest. Then the fruits produced will be qualified as safe for consumers and the environment. The present work was customized to give support for policy decision makers as an agroecological potential meeting needs of industries and ecological balance.

Establishment and evaluation of mouse models of Staphylococcus aureus and Klebsiella pneumoniae bloodstream infections

Objective To establish the reliable models for early diagnosis of bloodstream infections by common clinical bacteria and to provide reliable experimental models for screening and validation of biomarkers in the early stage of bacterial bloodstream infections. Methods Strains of Staphylococcus aureus (S.aureus) ATCC25923 and Klebsiella pneumoniae (K.pneumoniae) ATCC700603 were used as experimental strains. Different concentrations of bacterial suspensions were injected into mice through tail vein. LD50 (median lethal dose) was calculated by Karber method and the concentration of 1/2LD50 was used for injection. The established mouse models were evaluated based on the changes in physical signs, body weights, white blood cells (WBC), platelets and IL-6 as well as the results of blood culture, PCR and HE staining. Results LD50 of S. aureus and K. pneumoniae to ICR mice were 8.1×108 CFU/ml and 1.11×109 CFU/ml, respectively. Body weights of mice decreased significantly at the time point of 24 h after infection. WBC counts in normal mice were about (1.42±0.66)×109/L, but increased in model groups at 3 h after bacteria challenge. Compared with the control group, WBC counts in model groups increased significantly at 6 h (P<0.05), followed by decreases. Two days after infection, WBC counts in model groups increased again and remained at high levels till the 7th day. However, PLT counts in the model groups dropped dramatically within 2 days as compared with that in control group (P<0.05). Increase of IL-6 was detected in the early K. pneumoniae infection and the extent of that was larger than that in S. aureus infection. Results of the whole blood culture were all positive within 2 days after infection. PCR analysis confirmed that the bloodstream infection pathogens were S. aureus and K. pneumoniae. Alveolar exudation, edema and leukocyte infiltration were observed in lung and liver tissues with HE staining. Conclusion The mouse models of S. aureus and K. pneumoniae infections for early detection of bloodstream infections are successfully established. This study provides reliable animal models for further researches on early diagnosis of different pathogen-induced bloodstream infections. Key words: Staphylococcus aureus; Klebsiella pneumonia; Virulence factor; Bloodstream infection model

AgNO3 dependant modulation of glucose mediated respiration kinetics in Escherichia coli at different pH and temperature.

The inhibitory role of AgNO3 on glucose-mediated respiration in Escherichia coli has been investigated as a function of pH and temperature using Clark-type electrode, environmental scanning electron microscopy, and computational tools. In the given concentration of bacterial suspension (1 × 108 CFU/ml), E. coli showed an increasing nonlinear trend of tetra-phasic respiration between 1-133 μM glucose concentration within 20 min. The glucose concentrations above 133 μM did not result any linear increment in respiration but rather showed a partial inhibition at higher glucose concentrations (266-1066 μM). In the presence of glucose, AgNO3 caused a concentration-dependent (47-1960 μM) inhibition of the respiration rate within 4 min of its addition. The respiration rate was the highest at pH 7-8 and then was decreased on either side of this pH range. The inhibitory action of AgNO3 upon bacterial respiration was the highest at 37 °C. The observations of the respiration data were well supported by the altered bacterial morphology as observed in electron microscopic study. Docking study indicated the AgNO3 binding to different amino acids of all respiratory complex enzymes in E. coli and thereby explaining its interference with the respiratory chain. Copyright © 2016 John Wiley & Sons, Ltd.

Effect of estradiol on biofilm formation of staphylococcus epidermidis after breast implant surgery

?To investigate the effect of the estradiol hormones on biofilm formati on and structure of Staphylococcus epidermidis after breast implant surgery. ?The concentration of Staphylococcus epidermidis strains ATCC35984 was adjusted to 1×107 CFU/mL or 1×108 CFU/mL, and the type strains were incubated on the surface of silica gel in 125 pmol/L estradiol suspensions to prepare bacterial biofilms model in vitro. After cultured in vitro for 4, 6, 12, 24, 48, and 72 hours, bacteria growth and biofilm formation ability were assessed by means of the XTT and crystal violet staining respectively. According to the above results, the bacterial suspension concentration was selected for experiments. The experimental concentration of Staphylococcus epidermidis ATCC35984 suspension and the concentrations of 50, 125, 250, 500 pmol/L estradiol suspensions were mixed with silica gel respectively to prepare biofilm model in vitro, no estradiol suspension served as control group. The experimental concentration of Staphylococcus epidermidis ATCC12228 suspension was used to prepare the same model in the negative control. After cultured in vitro for 4, 6, 12, 24, 48, and 72 hours, the same methods were used to assess the bacteria growth dynamics and biofilm forming ability, and the scanning electron microscope (SEM) was used to observe bacterial biofilm structure cultured on the surface of silica gel; the laser scanning confocal microscope (CLSM) was used to measure bacterial biofilm thickness on the surface of silica gel after 6, 12, and 24 hours. ?According to the results of semi quantitative detection of crystal violet stain and XTT methods, the bacterial suspension of 1×107 CFU/mL was selected for the experiment. XTT results indicated that the growth rates of ATCC12228 strain (at 4, 6, 12, 24, and 72 hours ) and ATCC35984 strain (at 4, 6, 24, and 72 hours) in 125, 250, and 500 pmol/L estradiol were significantly faster than those in 0 and 50 pmol/L (P<0.05). The growth rate of 500 pmol/L group was significantly faster than 125 and 250 pmol/L groups at 4, 6, and 72 hours (P<0.05), and the growth rate of 250 pmol/L group was significantly faster than that of 125 pmol/L group at 72 hours (P<0.05), but there was no significant difference between 0 and 50 pmol/L groups (P>0.05). At the same time point and same estradiol concentration, the growth rates showed no significant difference between 2 strains (P>0.05). Semi quantitative detection of crystal violet staining showed no biofilm formed in ATCC12228 strain in all estradiol concentration groups at different time points. In ATCC35984 strain, the biofilm was found at 4 hours and gradually thickened with time, reached the peak at 24 hours. After cultured for 4 and 6 hours, the biofilm of 0 pmol/L groups were significantly thicker than that of 125, 250, and 500 pmol/L groups (P<0.05). At 12 hours, the 125 pmol/L group had the thickest biofilm, showing significant difference when compared with other groups (P<0.05). The CLSM showed ATCC35984 biofilm thickness of 125, 250, and 500 pmol/L was significantly less than that of 0 and 50 pmol/L groups at 6 hours (P<0.05), but difference was not significant between other groups (P>0.05). Then the thickness of the biofilm increased gradually, and the thickness of 125 pmol/L group was significantly larger than that of other concentration groups at 12 and 24 hours (P<0.05). The SEM observation showed that the biofilm of 125 pmol/L group was denser and thicker than that of the other concentration groups at each time point. ?High level estradiol can promote bacteria growth, biofilm formation, and biofilm maturity of Staphylococcus epidermidis.