Cost modelling of Pseudomonas fluorescens and Pseudomonas chlororaphis as biocontrol for competitive exclusion of Salmonella enterica on tomatoes

Published research on process-based models for biocontrol of foodborne pathogens on produce is limited. The aim of this research was to develop cost model estimates for competitive exclusion (CE) process using Pseudomonas fluorescens and Pseudomonas chlororaphis (non-plant pathogenic and non-human pathogen) as biocontrol against Salmonella enterica on tomatoes. Cost estimates were based on material inputs, equipment, facilities, and projected processing conditions of post-harvest packaging of tomatoes. The microbiological data for inactivation of S. enterica was based on published papers. The small-scale processing facility was assumed to have a processing capacity of 2000 kg of tomatoes/hour for 16 h per day, operational 6 days a week, and for 3-months /year. The large-scale facility was assumed to have a processing capacity of 100,000 kg of tomatoes/hour. Estimated initial capital investment costs for small and large-scale models (production facility) were US$391,000 and US$2.1 million. Application of CE for biocontrol of S. enterica on tomatoes was estimated at US$0.0058–0.073/kg of tomatoes during commercial processing operations. This exceeds chlorine wash technology estimated at US$0.00046/kg and is competitive with gaseous chlorine dioxide at US$0.02–0.21/kg. For high-value produce, CE may complement existing technologies increase food safety, reduce storage loses, and extend shelf life of produce.     

Inactivation of Salmonella serovars by Pseudomonas chlororaphis and Pseudomonas fluorescens strains on tomatoes

Salmonella enterica and its serovars have been associated with pathogen contamination of tomatoes with numerous outbreaks of salmonellosis. To improve food safety, pathogen control is of immediate concern. The aim of this research was to assess the populations of natural microflora (aerobic mesophilic bacteria, lactic acid bacteria, yeasts and moulds and Pseudomonas species) on tomatoes, and evaluate the efficacy of Pseudomonas fluorescens (Pf) and Pseudomonas chlororaphis (Pc) for inactivation of Salmonella on tomatoes. Microflora were determined on sanitised and unsanitised produce and enumerated on Plate Count Agar, de Man, Rogosa and Sharpe medium, Potato Dextrose Agar and Pseudomonas Agar F media. The efficacy of Pc and Pf for inactivation of S. enterica serovars Montevideo, Typhimurium and Poona was determined on spot-inoculated tomato stem scars. The effects of storage time on bacterial populations were also investigated. On unsanitised tomatoes, lactic acid bacteria, Pseudomonas sp., aerobic mesophilic bacteria and yeasts and moulds ranged from 3.31–4.84, 3.93–4.77, 4.09–4.80 and 3.83–4.67 log CFU/g of produce, respectively. The microflora were similar at 0 and 24 storage hours on sanitised produce. The suppression of Salmonella Montevideo by P. chlororaphis and P. fluorescens on tomatoes ranged from 0.51 to 2.00 log CFU/g of produce. On Salmonella Montevideo and S. Typhimurium, the suppressive effects ranged from 0.51 to 0.95 and 0.46 to 2.00 log CFU/g of produce, respectively. The pathogen suppressive effects may be attributed to competition ability of Pseudomonas relative to Salmonella strains. Pseudomonas strains may be effective against Salmonella strains as a post-harvest application, but strain synergy is required to optimise pathogen reductions.     

基于根部年轮与导管特征的黄土高原苜蓿生长衰退分析

紫花苜蓿作为黄土高原退耕还林还草工程的重要人工草本物种,对控制水土流失、植被固碳和生态环境改善等方面都发挥着重要生态作用;但是苜蓿草地在黄土高原普遍出现生长不可持续的特征,在第6年左右植被出现明显生长衰退现象。本研究在黄土高原4个降水梯度(280、400、500 mm和550 mm)的地点采集紫花苜蓿和自然对照草本物种:委陵菜的根部年轮样品,并分析了紫花苜蓿与委陵菜的年轮宽度和导管大小随年龄的变化趋势特征及与气候要素的相关关系。研究结果表明:在气候暖干化环境下,紫花苜蓿和委陵菜的年轮宽度随年龄增加都有不断下降的趋势,且在降水量偏低样点下降更为显著,这是由于植株在降水量偏低地点易遭受严重水分胁迫,以致无法满足生长所需的水分而造成根部生长速率明显减缓。紫花苜蓿根部导管大小随年龄增加均有明显下降趋势,在降水量偏低的样点下降趋势更加显著;这表明水分胁迫随年龄增加逐渐加剧,紫花苜蓿导管大小逐年下降以致不能满足自身生长的水分需求,植被生长呈衰退趋势,且在降雨偏低地区,苜蓿生长衰退出现时间较早。自然草本委陵菜根部导管大小随年龄增加均有明显上升趋势且在降水量偏低样点上升趋势更显著,这表明委陵菜通过增大根部导管大小、增加输水能力来满足随年龄增加不断上升的水分需求,这也说明委陵菜导管结构特征较好的适应了黄土高原干旱半干旱环境,生长处于可持续发展状态。     

Effect of N concentration and N source on root colonization by Pseudomonas fluorescens 2-79RLI

Wheat (Triticum aestivum L.) was grown in nutrient solution with low or high N supply (NH₄NO₃ as N source). To further evaluate the influence of N form and its interaction with the nutrient solution pH, wheat plants were grown with NH ₄ ⁺ or NO ₃ ^- either in an conventional nutrient solution or in a nutrient solution in which the pH was maintained at pH 6.5 using a pH-stat system. The nutrient solution was inoculated with Pseudomonas fluorescens 2-79RLI, a genetically modified bacterium that contains lux genes activated by a ribosomal promoter. Cell numbers and physiological status of P. fluorescens 2-79RLI (length of the lag phase of bioluminescence) in the rhizosphere were determined at the root tip and in the lateral root zone. Nitrogen deficiency decreased both plant growth and root colonization by P. fluorescens 2-79RLI at the root tip while it had no effect on root colonization in the lateral root zone. The physiological status of P. fluorescens 2-79RLI was not affected by nitrogen deficiency. Ammonium nutrition increased root colonization by P. fluorescens 2-79RLI at the root tip and in the lateral root zone when the pH of the nutrient solution was allowed to change according to the N form provided. Under these conditions, the physiological status of P. fluorescens 2-79RLI was higher in the lateral root zone than at the root tip. In contrast, N source had no effect on root colonization or physiological status of P. fluorescens 2-79RLI in the nutrient solution maintained at pH 6.5. It is concluded that the stimulation of root colonization by NH ₄ ⁺ in the nutrient solution, not maintained at a constant pH, may be due to increased leakage of solutes into the rhizosphere as a result of impaired exudate retention by high H⁺ concentration in the rhizosphere or the apoplast. This revised version was published online in June 2006 with corrections to the Cover Date.     

Isolation, identification, and accumulation of 2-acetamidophenol in liquid cultures of the wheat take-all biocontrol agent Pseudomonas fluorescens 2-79

Pseudomonas fluorescens strain 2-79 (NRRL B-15132) is a classic biological control agent known to produce phenazine-1-carboxylic acid (PCA) as its primary means of suppressing take-all disease of wheat. In addition to PCA, an unknown metabolite was discovered in a liquid culture used to produce the biocontrol agent. The objective of the current study was to isolate, identify, and evaluate the accumulation of this compound in production cultures. Upon centrifugal fractionation of a production culture, thin-layer chromatography (TLC) analyses of extracts of the cells and cell-free supernatant indicated the compound to be primarily in the supernatant. Purified compound was obtained by extraction of culture supernatant, followed by flash chromatography of the extract and preparative TLC. The ¹H and ¹³C nuclear magnetic resonance and electron impact mass spectra indicated the compound to be 2-acetamidophenol (AAP). Measured by reversed-phase HPLC, the accumulations of AAP and PCA in cultures of strain 2-79 reached 0.05 g/l and 1 g/l, respectively. The accumulations of AAP and PCA in liquid cultures were linearly correlated (P < 0.001), as shown by studies of cultures stimulated to yield varying levels of PCA by controlling levels of oxygen transfer, pH, and growth medium composition. In this study, oxygen limitation, a defined amino-acid-free medium, and neutral pH stimulated maximal production of both AAP and PCA. Furthermore, a transposon mutant of 2-79 [2A40 2-79 (phz–)] unable to produce PCA did not accumulate AAP. These findings indicate that AAP and PCA are likely to share a common segment of biosynthetic pathway. This is the first report of AAP production by a strain of P. fluorescens. Possible routes of AAP production are discussed relative to current knowledge of the phenazine biosynthetic pathway of strain 2-79. The pertinence of AAP to the design of commercial seed inoculants of phenazine-producing bacteria for controlling wheat take-all is also considered. Received: 2 November 1999 / Received revision: 3 April 2000 / Accepted: 14 April 2000     

Reduction of Salmonella on alfalfa seeds using peroxyacetic acid and a commercial seed washer is as effective as treatment with 20 000 ppm of Ca(OCl)2

Aims: The efficacy of a commercial seed washer and 1 and 3% peroxyacetic acid or 20 000 ppm calcium hypochlorite for reducing Salmonella on alfalfa seeds was investigated. Methods and Results: Alfalfa seeds were inoculated with Salmonella Stanley to achieve c. 5 log CFU g^?1. Seeds were then treated with 1 or 3% peroxyacetic acid or 20 000 ppm calcium hypochlorite for 15 min in a commercial seed washer that uses air to enhance contact of the sanitizer with the seed. Experiments were also conducted using industry and laboratory methods. An c. 1‐log reduction in number of Salm. Stanley was demonstrated regardless of the chemical treatment or method of treatment. Although this 1‐log reduction was significant (P < 0·05), differences among the treatments were not significant. Treating the seed with 1 and 3% peroxyacetic acid resulted in similar Salm. Stanley reductions of 1·77 and 1·34 log, respectively, not being statistically significant (P > 0·05). Conclusions: These results suggest that under conditions tested, 1 or 3% peroxyacetic acid solutions are equally effective as 20 000 ppm of Ca(OCl)₂ in the reduction of Salm. Stanley on alfalfa seed when used in conjunction with a commercial seed washer. Significance and Impact of the Study: A 1% peroxyacetic acid solution could potentially be used in place of 20 000 ppm of Ca(OCl)₂ for treatment of seeds used for sprouting_. The commercial seed washer did not enhance removal of Salm. Stanley from alfalfa seeds, but did facilitate removal of excess soil from seeds.     

Growth and maintenance respiratory costs of cucumber fruits as affected by temperature, and ontogeny and size of the fruits

The rates of dry weight increase and respiration of fruits were measured throughout fruit ontogeny at 20, 25 and 30°C in cucumber ( Cucumis sativus L. cv. Corona). By maintaining one or five fruits per plant, which strongly affected fruit dry weight but not ontogeny, the effects of fruit size and ontogeny on respiration could be studied separately. The respiration rate per fruit followed a sigmoid curve during fruit ontogeny, while the specific respiration rate (respiration rate per unit dry weight) declined with time after anthesis. The specific respiration rate was almost linearly related to the relative growth rate. The specific respiratory costs for both growth and maintenance were highest in young fruits, but were not affected by fruit size. The average specific respiratory costs for growth and maintenance at 25°C were 3.3–3.9 mmol CO₂ g⁻¹ and 4.0 nmol CO₂ g⁻¹ s⁻¹, respectively. An increase in temperature had no effect on the specific respiratory costs for growth, while the costs for maintenance increased with a Q₁₀ of about 2. The costs for growth agreed reasonably well with theoretical estimates based on the chemical composition of the fruits but not with estimates based on only the carbon and ash content. The respiratory losses as a fraction of the total carbon requirement of a fruit changed during fruit ontogeny, but were independent of temperature and were similar for slow- and fast-growing fruits. The cumulative respiratory losses accounted for 13–15% of the total carbon requirement.     

Effects of Pseudomonas fluorescens strain UTPF5 on the mobility,mortality and hatching of root-knot nematode Meloidogyne javanica

The root-knot nematode Meloidogyne spp. includes important plant pathogens worldwide. This study has considered nematode Meloidogyne javanica second stage larvae activity in the extracts of Pseudomonas fluorescens strains UTPF5 and cytotoxic effect of the strain on the nematode. The movement of second stage larvae of nematodes in water agar medium at four concentrations of bacterial extracts and second stage larvae mortality rate of hatching nematode and bacterial strains in vitro were affected. Different concentrates of the strain UTPF5 effect nematode larvae movement and disposal of the same. Bacterial extraction kills almost 100% of the larvae hatching after 24?h and a complete ban on egg hatch of biocontrol nematodes and nematode indicated that root-knot nematode larvae movement on the right attract the bacteria P. fluorescens to extract in the first place.     

Inactivation of Salmonella Typhimurium,Escherichia coli O157:H7 and Listeria monocytogenes on alfalfa seeds by the combination treatment of vacuumed hydrogen peroxide vapour and vacuumed dry heat

We evaluated the combined effects of vacuumed hydrogen peroxide vapour (VHPV) and vacuum-sealed dry heat (vacuum heat, VH) to inactivate food-borne pathogens (Salmonella Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes) on alfalfa seeds. Alfalfa seeds inoculated with food-borne pathogens were sequentially treated initially with 1·0 ml of 0 or 30% VHPV for 1 min and later with dry heat (DH) or VH for 2 h, and the rate of seed germination was evaluated. The combination treatment decreased the populations of three food-borne pathogens below the limit of detection (1·0 log CFU per gram) on alfalfa seeds without decreasing germinability. The sequential treatment using VHPV and VH greatly reduced the total treatment time needed to inactivate pathogens on alfalfa seeds by more than 5 log CFU per gram. These results demonstrate that a combination of VHPV and VH has potentially employed as a new method for pasteurization of alfalfa seeds without affecting their germinability.     

不同来源和粒径的胶体对光合细菌生长的效应

天然胶体是近岸海域细菌和浮游植物可利用氮的一个重要来源 ,其含量的增加可促进藻类的繁殖和生长 ,有时甚至引发赤潮。研究显示 ,细菌在高分子量胶体存在下的生长和代谢速率比在低分子量中的提高 3～ 6倍 ,暴露在阳光下的胶体有机物可释放生物可利用的富氮组分 ,进而提高细菌对胶体的分解。利用错流超滤技术从河流、河口、海洋水体和微藻培养液中提取胶体 ,研究了胶体来源和粒径对光合细菌 (PSB,沼泽红假单胞菌 Rhodopseudomonaspalustris)生长的影响。结果表明 ,在一定的胶体有机碳浓度范围 ,河流胶体 (0～ 2 81.0μmol/ L )、河口胶体 (0～ 12 1.2μmol/ L )、海洋胶体 (0～ 88.8μmol/ L )和生源胶体 (7.7～5 4 8.6μmol/ L )都能促进 PSB的生长 ,分别使其相对增长率平均提高了 4 7.3%～ 196 .2 % ,3.6 %～ 9.3% ,8.1%～ 10 .4 %和2 .4 %～ 6 .9%。其中 ,PSB在河流胶体中的相对增长率的平均值 (Y)与有机碳浓度 (CDOC)呈对数相关 (Y (% ) =- 193.7 70 .7ln CDOC) ,表明高浓度的河流胶体 ,对 PSB生长的促进效果更显著。两种生源胶体中 ,海水小球藻胶体 (0～ 5 4 8.6μmol/ L )对PSB生长的促进作用大于球等鞭金藻胶体 (7.7～ 384 .4 μmol/ L) ;PSB在粒径为 10 k D～ 0 .2 2 μm的海洋胶体中的生长大于     

Interaction between Pseudomonas fluorescens and Meloidogyne incognita on tomato plant as influenced by the different levels of phosphorus

A pot experiment was conducted on tomato (Solanum lycopersicum cv. Pusa Ruby) to assess the effect of different phosphorus (P) levels (0, 125, 250 and 500 mg/pot) and the plant growth promoting rhizobacterium, Pseudomonas fluorescens, on the growth of tomato and on the reproduction of Meloidogyne incognita. Maximum growth of tomato occurred at P rates of 125 mg/kg soil, irrespective of whether plants were uninoculated or inoculated with P. fluorescens or M. incognita or inoculated with both the agents. Nematodes per gram of roots, egg masses per root, eggs per egg mass and galls per root significantly increased by increasing levels of P. P. fluorescens performed better than other treatments and different P levels in improving tomato growth and reducing galling and multiplication of M. incognita.     

陇中黄土高原半干旱区苜蓿地土壤干燥化特征及适宜种植年限

研究了陇中黄土高原半干旱区不同种植年限紫花苜蓿地土壤水分特征及适宜种植年限.结果表明： 3、8、12和14年生苜蓿地0～300 cm土层土壤平均含水量均明显低于当地土壤稳定湿度值.12和14年生苜蓿地0～300 cm土层土壤含水量仅为9.2%和7.1%,甚至低于作物有效水分下限.1、3、8、12和14年生紫花苜蓿地0～300 cm土层干燥化指数分别为125.4%、30.5%、18.4%、-34.2%和-83.3%,除1年生苜蓿地土壤无干燥化现象之外,其余种植年限苜蓿地土壤均呈不同程度的干燥化.随苜蓿种植年限的延长,土壤干燥化程度加剧,但干燥化速率呈减缓趋势.综合苜蓿生产力动态和土壤水分状况,该区紫花苜蓿适宜的种植年限为8～10 年.     

Survival and colonization of Escherichia coli O157:H7 on spinach leaves as affected by inoculum level and carrier, temperature and relative humidity

Aims: To determine survival and colonization of Escherichia coli O157:H7 on spinach leaves as affected by inoculum level and carrier, temperature and relative humidity (r.h.). Methods and Results: Spinach leaves were inoculated with suspensions of E. coli O157:H7 in distilled water (DW) and 0·1% peptone water (PW) and incubated at 4, 12 and 25°C and 43, 85 and 100% r.h. The number of E. coli O157:H7 on leaves (5·6 or 1·9 log CFU per leaf) inoculated using DW as a carrier medium increased significantly at 25°C and 100% r.h. within 120 h but remained constant or decreased significantly under other test conditions. E. coli O157:H7 on leaves (5·4 log CFU per leaf) inoculated using PW as a carrier increased significantly within 72 and 24 h, respectively, at 12 or 25°C and 100% r.h.; counts using a low inoculum (2·2 log CFU per leaf) increased significantly within 24 h at 25°C. Conclusions: Escherichia coli O157:H7 can colonize on spinach leaves at 12 or 25°C in a 100% r.h. environment. Organic matter in the inoculum carrier may provide protection and nutrients which enhance survival and colonization. Significance and Impact of the Study: Colonization of E. coli O157:H7 on spinach leaves as affected by organic matter in the inoculum, temperature and r.h. was determined. These observations will be useful when developing strategies to prevent growth of E. coli O157:H7 on pre‐ and postharvest spinach.     

Performance,gut morphology and carcass characteristics of fattening rabbits as affected by particle size of pelleted diets

A review of past literature revealed inconsistencies in recommended feed particle size for optimal growth and productive performance of rabbits. Changing diet formulation and subsequent processing conditions may improve pellet texture and potentially affect rabbit performance. In the current study, two isoenergetic and isonitrogenous pelleted diets were formulated, which varied in the particle size of the concentrates (2 and 8 mm, respectively). The objective was to evaluate the effect of different particle sizes of compound diets on performance, nutrient utilisation, gut morphology, and carcass characteristics of fattening Italian White breed rabbits. The finely ground diet led to a significant improvement in feed efficiency and apparent digestibility of crude protein, ether extract, crude fibre and NDF, without any negative effect on gut morphology. Furthermore, a smaller particle size of concentrates in pelleted diets improved carcass traits. Meat colour parameters showed significant differences in longissimus lumborum and biceps femoris due to dietary treatments, but in both muscles pH values 1 h and 24 h after slaughter remained unchanged. It is concluded that a finely ground pelleted diet can be used to improve growth performance of rabbits without affecting carcass parameters.     

Competition among native and invasive Phragmites australis populations: An experimental test of the effects of invasion status,genome size,and ploidy level

Among the traits whose relevance for plant invasions has recently been suggested are genome size (the amount of nuclear DNA) and ploidy level. So far, research on the role of genome size in invasiveness has been mostly based on indirect evidence by comparing species with different genome sizes, but how karyological traits influence competition at the intraspecific level remains unknown. We addressed these questions in a common‐garden experiment evaluating the outcome of direct intraspecific competition among 20 populations of Phragmites australis, represented by clones collected in North America and Europe, and differing in their status (native and invasive), genome size (small and large), and ploidy levels (tetraploid, hexaploid, or octoploid). Each clone was planted in competition with one of the others in all possible combinations with three replicates in 45‐L pots. Upon harvest, the identity of 21 shoots sampled per pot was revealed by flow cytometry and DNA analysis. Differences in performance were examined using relative proportions of shoots of each clone, ratios of their aboveground biomass, and relative yield total (RYT). The performance of the clones in competition primarily depended on the clone status (native vs. invasive). Measured in terms of shoot number or aboveground biomass, the strongest signal observed was that North American native clones always lost in competition to the other two groups. In addition, North American native clones were suppressed by European natives to a similar degree as by North American invasives. North American invasive clones had the largest average shoot biomass, but only by a limited, nonsignificant difference due to genome size. There was no effect of ploidy on competition. Since the North American invaders of European origin are able to outcompete the native North American clones, we suggest that their high competitiveness acts as an important driver in the early stages of their invasion.     

Effects of sulfate on lactate and C2-, C3- volatile fatty acid anaerobic degradation by a mixed microbial culture

The effects of sulfate on the anaerobic degradation of lactate, propionate, and acetate by a mixed bacterial culture from an anaerobic fermenter fed with wine distillery waste water were investigated. Without sulfate and with both sulfate and molybdate, lactate was rapidly consumed, and propionate and acetate were produced; whereas with sulfate alone, only acetate accumulated. Propionate oxidation was strongly accelerated by the presence of sulfate, but sulfate had no effect on acetate consumption even when methanogenesis was inhibited by chloroform. The methane production was not affected by the presence of sulfate. Counts of lactate- and propionate-oxidizing sulfate-reducing bacteria in the mixed culture gave 4.5×10⁸ and 1.5×10⁶ viable cells per ml, respectively. The number of lactate-oxidizing fermentative bacteria was 2.2×10⁷ viable cells per ml, showing that sulfate-reducing bacteria outcompete fermentative bacteria for lactate in the ecosystem studied. The number of acetoclastic methanogens was 3.5×10⁸ viable cells per ml, but only 2.5×10⁴ sulfate reducers were counted on acetate, showing that acetotrophic methanogens completely predominated over acetate-oxidizing sulfate-reducing bacteria. The contribution of acetate as electron donor for sulfate reduction in the ecosystem studied was found to be minor.     

食物浓度对角突臂尾轮虫种群增长、个体大小和卵大小的影响

以蛋白核小球藻为食物 ,应用群体累积培养法研究了食物浓度对角突臂尾轮虫种群增长、个体大小和卵大小等的影响 .结果表明 ,食物浓度对角突臂尾轮虫种群增长率、个体大小和卵大小均有极显著影响 .其中 ,轮虫种群增长率与食物浓度间呈曲线相关 ;当食物浓度为 8.2 4 5 3× 10 6cells·ml-1时 ,种群增长率达最大值 0 .6 0 85d-1;在所研究的食物浓度范围内 ,轮虫个体具有随食物浓度的增大而增大的趋势 ,轮虫卵体积在中等食物浓度范围 (6 .0× 10 6～ 9.0× 10 6cells·ml-1)时较大