Characteristics of an ammonia-oxidizing bacterium with a plasmid isolated from alkaline soils and its phylogenetic relationship

An ammonia-oxidizing bacterium, strain TCH716, was isolated from alkaline soil at Harbin city, China. The cells of strain TCH716 are lobate (0.8-1.5 x 1.0-2.0 microm), gram-negative, obligately aerobic, and nonmotile. Colonies (1-2 mm in diameter) on gellan gum plate culture are reddish, circular, and smooth. The G + C content of DNA is 54.78 mol%. Its percentage of 16S rRNA gene sequence similarity (%) to Nitrosolobus multiformis ATCC 25196T (type strain) is 98.56%. This bacterium has an optimal growth temperature and pH at 30 degrees C and 8.0-8.5, respectively. The concentration of ammonium sulfate in the HEPES medium for optimum growth of this bacterium is 38 mM. Strain TCH716 was found to have a plasmid (approximately 6.5 kbp) that possessed a plasmid-linked gene for sulfonamide resistance. Phosphoglycerate kinase, RubisCO and PEPC were found to possess high specific activities compared to the activities of these enzymes in strain ATCC 25978T. In identification of strain TCH716, both morphological characteristics (compartmentalized cells) and the phylogenetic relationship based on 16S rRNA gene sequence are important. Based on results obtained, strain TCH716 belongs to the genus Nitrosolobus, and designated as Nitrosolobus sp. TCH716.     

Two kinds of ammonia-oxidizing bacteria isolated from biologically deodorizing plants in cold district

Two kinds of ammonia-oxidizing bacteria isolated from biologically deodorizing plants in cold districts in Japan were identified as Nitrosomonas sp. IWT202 and Nitrosomonas sp. IWT514. The optimum pHs for growth were 8.0 (IWT202) and 7.5 (IWT514). Although rockwool samples for isolation were collected from the same plants, the optimum temperature for growth of strain IWT202 (37 degrees C) differed from that of strain IWT514 (30 degrees C). The bacteria had a higher (IWT202, 37 degrees C) and lower (IWT514, 20 degrees C) growth temperature than is usually the case. Both strains were shown to differ completely in regard to the effect of the ammonium sulfate concentration in the medium for a 20 degrees C culture and 30 degrees C culture. The inoculation of these bacteria provides the possibility of recovering ammonia-oxidizing activity, when the ammonia-oxidizing activity is lowered in biological deodorizing plants in cold districts. It seems that these strains are suitable for application to deodorization.     

Nitrosomonas communis strain YNSRA, an ammonia-oxidizing bacterium, isolated from the reed rhizoplane in an aquaponics plant

An ammonia-oxidizing bacterium (strain YNSRA) was isolated from the rhizoplane of the reed (Phragmites communis) used in an aquaponics plant which is a wastewater treatment plant. Strain YNSRA was identified as Nitrosomonas communis by taxonomic studies. The hydroxylamine-cytochrome c reductase (HCR) of strain YNSRA was found to have a higher activity (25.60 u/mg) than that of Nitrosomonas europaea ATCC25978T (8.94 u/mg). Ribulose-1,5-bisphosphate carboxylase (RubisCO) activity was detected at very low levels in strain YNSRA, whereas strain ATCC25978T had definite activity.     

Characterization of a new keratin-degrading bacterium isolated from deer fur

A keratin-degrading bacterium was isolated from soil containing deer fur. An axenic culture of the keratin-degrading bacterium was obtained in liquid culture using a keratin enrichment technique. The isolated bacterium was gram negative and catalase- and oxidase-positive. Transmission electron microscopic observations showed that the bacterium was rod-shaped, 1.0-1.3 microm long and 0.7 microm in diameter. Phylogenetic analysis of 16S rDNA revealed that the new isolate has only 90.6% homology with Stenotrophomonas nitritireducens. Hence, this new bacterium was designated as Stenotrophomonas sp. D-1. The optimum temperature was determined to be 20 degrees C for maximum growth and keratinolytic enzyme production. Amino acid data, obtained after treating keratin powder with the supernatant culture, suggest that the major free amino acids resulting from keratin degradation are phenylalanine, tyrosine and valine. In addition, native chicken feather was degraded completely at 20 degrees C in 2.5 d by this bacterium.     

西藏牦牛发酵乳中乳酸菌及酵母菌的特性与相互作用

研究了西藏牦牛发酵乳中的优势菌株:发酵乳杆菌166A、乳酒假丝酵母37、酿酒酵母100、郎比克假丝酵母58在牛乳中的生长特性。探讨了发酵乳杆菌与酵母菌间的相互作用,在发酵过程中3株酵母菌均对发酵乳杆菌166A的生长有促进作用,发酵乳杆菌166A能抑制乳酒假丝酵母37的生长。酵母菌与发酵乳杆菌166A共同接种有利于保持产品冷藏期间活菌数的稳定,菌株之间可能存在共生作用。     

Purification and comparison of triosephosphate isomerases from ammonia-oxidizing bacteria isolated from terrestrial and marine environments

Triosephosphate isomerases [TIMs, EC 5.3.1.1] were purified from two ammonia-oxidizing bacteria: Nitrosomonas sp. K1 (K1), Nitrosomonas sp. TNO632 (TNO). The molecular masses of the native enzymes were estimated to be about 53.6 (K1-TIM) and 51.9 kDa (TNO-TIM) by gel filtration, whereas SDS-PAGE produced one band for each enzyme with M(r) values of 27.1 (K1-TIM) and 26.4 kDa (TNO-TIM), respectively, suggesting that the enzymes consist of identical subunits. The apparent K(m) for d-glyceraldehyde-3-phosphate (GAP) and dihydroxyacetone phosphate (DHAP) were about 1.19 and 4.78 mM (K1-TIM), and 0.41 and 6.01 mM (TNO-TIM), respectively. The two TIMs had different pH-activity curves with an optimum pH range of 6.5 (K1-TIM) and 8.0 (TNO-TIM). The temperature optima of K1-TIM and TNO-TIM were 50-60 and 60-65 degrees C, respectively. Both enzymes were strongly inhibited by 5,5'-ditiobis at 1.0 mM. The N-terminal amino acid sequences of K1-TIM and TNO-TIM were MRAGFVAGNWKMHG (K1-TIM) and MVRTGLVAGNWKMNG (TNO-TIM). A homology of 74.1% was observed between K1-TIM and TNO-TIM.     

Inhibition of Listeria monocytogenes by a lactic acid bacterium isolated from Italian salami

Listeria monocytogenes is an opportunistic psychrotroph foodborne pathogen that has been used as a model organism to study the efficacy of many different preservation methods. This work aimed to test the antilisterial activity of lactic acid bacteria isolated from Italian salami and study the development of resistance. Isolates were obtained from naturally fermented Italian salami and cultures that retained activity in the supernatants after pH neutralization and catalase treatment were further characterized. The isolate showing highest inhibitory activity (PD 6.9) was tested for sensibility to proteases, heat and pH. To evaluate if resistance developed, sensitive strains were transferred with sub-lethal doses of the partially purified inhibitory substance and then inoculated into media containing higher doses of the extract. Isolate PD 6.9 inhibited several L. monocytogenes strains obtained from different origins and retained its activity over a wide range of pH and temperature. When increasing concentrations (10-100 AU ml(-1)) of the partially purified inhibitory substance were added to culture media, growth of L. monocytogenes did not occur even after 12 h of incubation. Cultures of Listeria that were transferred with sub-lethal doses (10 AU ml(-1)) of the partially purified inhibitory substance could resist higher doses of the extract (50 AU ml(-1)), but were inhibited when the concentration was further increased (100 AU ml(-1)). These results indicate that isolate PD 6.9 could potentially be used as a bioprotective culture for salami fermentation.     

A novel cyanobacteriolytic bacterium, Bacillus cereus, isolated from a Eutrophic Lake

Isolation and screening of cyanobacteriolytic bacteria were carried out. Fifteen strains of cyano-bacteriolytic bacteria were isolated by the double layer method using the cyanobacterium, Microcystis, as a sole nutrient. The isolate, N-14, showing the highest cyanobacteriolytic activity was identified as Bacillus cereus based on the 16S rRNA sequence. Components among the extracellular products in the culture supernatant of B. cereus were responsible for the cyanobacteriolytic activity. Lytic assay tests of culture supernatants indicated that the major substances for lytic activity could be non-proteinaceous, and hydrophilic, heat stable, and with a molecular weight of less than 2 kDa. The highest lytic activity was obtained under alkaline conditions, indicating an advantage for the practical application of water bloom control in eutrophic lakes where the pH is usually in the alkaline region. The lytic substance of B. cereus N-14 were compared with enterotoxins and an emetic toxin produced by a pathogenic strain of B. cereus, and also with a known algicide produced by Bacillus brevis, gramicidin. From these results, the lytic substance seemed to be a novel algicide.     

一株墨鱼干源中度嗜盐菌的系统发育分析与生长特性

从市售墨鱼干表面分离纯化出一株中度嗜盐细菌NYJ01。16S rDNA序列测定和BLAST分析结果表明其16S rDNA与盐单胞菌属和栖盐田菌属中一些菌株的序列一致性高达99%。系统发育分析显示它与栖盐田菌属中4个种的模式菌株聚类在一个进化枝,表明菌株NYJ01是一种栖盐田菌。其耐盐范围是1%~20%NaCl,最适浓度为3%~10%NaCl;生长温度范围是10~45℃,最适温度为25~37℃;该菌在0.5%~15%的NaNO3培养液中生长良好,但不具有硝酸盐还原作用。可利用0.025%双乙酸钠、0.05%山梨酸钾、0.05%苯甲酸钠或0.05%脱氢乙酸钠等食品防腐剂抑制菌株NYJ01的生长。      

Salmonella serotypes isolated from chicken meat in Albania.

A total of 31 salmonellae, belonging to four different serogroups and nine serotypes, were isolated from 30 out of 461 (6.5%) chicken meat samples, taken during a 3-year period, from 1996 to 1998. There were no significant differences among years in Salmonella positive samples, ranging from 5.1% to 8%. The most frequently encountered serotype was Salmonella enteritidis, isolated in 16 out of 30 Salmonella positive samples (53.3%), but its predomination was clearly evident only in 1996. The other isolated serotypes were S. senftenberg (three isolates), S. newport (two isolates), and S. abony, S. agona, S. banana, S. brancaster, S. infantis and S. oslo with only one isolate each. Four other Salmonella strains were not fully serotyped.     

养猪场源致泻大肠埃希菌毒力基因和致病型研究

目的 了解养猪场源致泻大肠埃希菌(DEC)毒力基因及致病型分布情况,为从猪肉生产源头防控该菌引发的食源性疾病提供参考数据.方法 采用多重荧光实时定量聚合酶链式反应(PCR)检测分离 自12个养猪场的生猪、养殖环境和养殖工人的大肠埃希菌毒力基因,鉴定DEC的致病型.结果 985株分离 自养猪场的大肠埃希菌中,DEC占比2...     

Production of equol from daidzein by gram-positive rod-shaped bacterium isolated from rat intestine

Isoflavones (mainly daidzein and genistin) belong to the flavonoid group of compounds and are classified as phytoestrogens. In the intestine, daidzin is converted to daidzein by beta-glucosidase, and then daidzein is converted to O-desmethylangolensin (O-DMA) or equol via dihydrodaidzein by enzymes of intestinal bacteria. We isolated, for the first time, an anaerobic gram-positive rod-shaped strain capable of producing equol from daidzein. Its 16S rDNA gene sequence (1428 bp) showed 99% similarity with that of the human intestinal bacterium SNU-Julong 732 (AY310748) and 93% similarity with that of Eggerthella lenta ATCC 25559(T) (AF292375). This strain converted daidzein to equol via dihydrodaidzein in an equol-assay medium anaerobically. The addition of butyric acid and arginine increased the conversion ratio of daidzein to equol 4.7- and 4.5-fold, respectively.     

Survival of Campylobacter jejuni in biofilms isolated from chicken houses

Campylobacter jejuni is a thermophilic and microaerophilic enteric pathogen associated with poultry. Biofilms may be a source of C. jejuni in poultry house water systems since they can protect constituent microorganisms from environmental stress. In this study, the viability of C. jejuni in biofilms of gram-positive chicken house isolates (P1, Y1, and W1) and a Pseudomonas sp. was determined using a cultural method (modified brucella agar) and direct viable count (DVC). Two-day biofilms grown on polyvinyl chloride (PVC) coupons in R2A broth at 12 and 23 degrees C were incubated with C. jejuni for a 6-h attachment period. Media were then refreshed every 24 h for 7 days to allow biofilm growth. Two-day biofilms of P1, Y1, and Pseudomonas spp. enhanced attachment (P < 0.01) of C. jejuni (4.74, 4.62, and 4.78 log cells/cm2, respectively) compared to W1 and controls without preexisting biofilm (4.31 and 4.22 log cells/cm2, respectively). On day 7, isolates P1 and Y1 and Pseudomonas biofilms covered 5.4, 7.0, and 21.5% of the surface, respectively, compared to 4.9% by W1. Viable C. jejuni on the surface decreased (P < 0.05) with time, with the greatest reduction occurring on surfaces without a preexisting biofilm. The number of viable C. jejuni determined by DVC was greater than that determined by the cultural method, indicating that C. jejuni may form a viable but nonculturable state within the biofilm. Both DVC and the cultural method indicate that biofilms enhance (P < 0.01) the survival of C. jejuni during incubation at 12 and 23 degrees C over a 7-day period.     

市售鸡肉中沙门菌分离株多重耐药谱测定

为了解我国市售鸡肉中沙门菌多重耐药状况及耐药程度，对国家食源性疾病监测网从市售鸡肉中分离到的51株沙门菌进行耐药检测，并对多重耐药谱进行分析。利用NccLs(National Committee of Clinical Labaratory Standard)推荐的纸片法进行耐药检测，51株分离菌株全部对至少3种以上的抗生素耐药，属多重耐药株，耐5种抗生素的9株(17.6％)，耐6～9种抗生素的12株(23．5％)，耐10种以上抗生素的9株(17．6％)。51株沙门菌对万古霉素、红霉素、苯唑西林都具有耐药性(100％)，对其它抗生素的耐药情况是：萘啶酮酸(52.9％)、磺胺(35．3％)、链霉素(33．3％)、四环素(29.4％)、氨苄青霉素(23．5％)、羧苄西林(21.6％)、阿莫西林(19．6％)、吡拉西林(19．6％)、美唑西林(17．6％)、强力霉素(17．6％)、氯霉素(13．7)和头孢噻吩(9．8％)。有3株菌对庆大霉素耐药，有2株菌对丁胺卡那霉素和甲氧苄胺嘧啶耐药，有1株菌对复方新诺明耐药。分离出1株耐15种抗生素的鼠伤寒沙门菌，其耐药谱与超级耐药鼠伤寒沙门菌DTl04的耐药谱类似。从8省市市售鸡肉中分离的51株沙门菌均为多重耐药株，提示我国畜牧养殖业在使用抗生素方面存在严重问题，应加强对饲料添加剂使用的安全性管理，合理使用抗生素。     

乌鸡酶解液的脱苦脱腥及其冻干粉制作

研究乌鸡酶解液的脱苦脱腥方法,并对脱腥后的酶解液进行冷冻干燥制得冻干粉.结果表明,与活性碳和β-环糊精相比,活性干酵母是乌鸡酶解液较好的脱腥剂.1.5％酵母在28℃下发酵1h,5000r/min离心20min后过滤,加0.2％的柠檬酸处理,处理后的酶解液腥味值为1.0,苦味值为1.5.冷冻干燥后酶解冻干粉呈淡黄色,略带酵母香味,基本无腥苦味,复水效果良好.干燥粉中水分含量4.18％,灰分为4.61％,多肽质量分数为66.67％,氮回收率为91.26％.     

Tailpipe greenhouse gas emissions from tank trucks transporting raw milk from farms to processing plants

An analysis of greenhouse gas emissions (carbon dioxide equivalents, CO₂e) was conducted from 2007 databases for 211,216 round trips of tank trucks that delivered raw milk from farms to processing plants in the United States of America. The total amount of milk was 4.81 × 10⁹ kg, or about 17.4% of the 2007 total USA production for use as fluid milk products. Average round trip distance was 850 km resulting in tailpipe emissions of 0.050 kg CO₂e kg⁻¹ milk delivered or 0.071 kg CO₂e kg⁻¹ milk consumed representing 3.5% of the total greenhouse gas emissions for fluid milk consumed. Based on this we estimate the total emissions for fluid milk delivery from farm to processor in the US at 1.3 × 10⁹ kg CO₂e y⁻¹. Some overall reduction in total delivery distance could be realized by realigning farm-to-processor relationships, especially in regions where farms are equally distant from multiple processors.     

乌鸡酶解液的脱苦脱腥及其冻干粉制作

研究乌鸡酶解液的脱苦脱腥方法,并对脱腥后的酶解液进行冷冻干燥制得冻干粉。结果表明,与活性碳和β-环糊精相比,活性干酵母是乌鸡酶解液较好的脱腥剂。1.5%酵母在28℃下发酵1h,5000r/min离心20min后过滤,加0.2%的柠檬酸处理,处理后的酶解液腥味值为1.0,苦味值为1.5。冷冻干燥后酶解冻干粉呈淡黄色,略带酵母香味,基本无腥苦味,复水效果良好。干燥粉中水分含量4.18%,灰分为4.61%,多肽质量分数为66.67%,氮回收率为91.26%。      

Virulence profiles of Klebsiella pneumoniae isolated from 2 large dairy farms in China

We recently reported on the diversity of Klebsiella pneumoniae isolated from dairy herds in China. In our previous work, isolates from subclinical mastitis (SCM) had lower indices of diversity when compared with bacteria from other sources, possibly due to a contagious-like spread of udder adapted strains. Here we explored the virulence profile and capsular types of K. pneumoniae isolated from different sources on 2 dairy farms in China. Our overarching goal was to gain insights on the role of virulence genes toward the severity of mastitis caused by K. pneumoniae. A total of 1,484 samples were collected from clinical mastitis (CM; n = 355), SCM (n = 561), bulk tank milk (BTM; n = 130), and environmental and extramammary (EE) sites (n = 438). From those, 431 K. pneumoniae isolates were obtained, including 129, 77, 66, and 159 isolates from CM, SCM, BTM, and EE samples, respectively. Polymerase chain reactions were used to determine the capsular types and to detect potential virulence genes in all isolates. No significant farm effects were observed when comparing the distribution of most virulence genes in K. pneumoniae isolated from each source. K57 was the most prevalent capsular type in K. pneumoniae from all sources, but with increased detection rate in isolates from CM. entB, kfu, fimH1, mrkD, and β-d-lacZ were frequently detected in K. pneumoniae from all sources. β-d-lacZ, entB, and ituA were more prevalent in isolates from CM, whereas kfu, allS, and nif were more frequently detected in isolates from SCM. ybtS, aerobactin, and rpmA had increased prevalence in K. pneumoniae from BTM when compared with bacteria from other sources. No association was detected between virulence genes and the severity of CM. K57 and the nif gene had the highest discriminatory power to classify isolates from CM and SCM, respectively. Based on our findings, it is likely that K57 is the dominant capsular type in K. pneumoniae causing CM in large Chinese dairy herds. Likewise, we demonstrated that β-d-lacZ is disseminated in K. pneumoniae isolated from large Chinese dairy farms, irrespectively of the source of bacteria. Our results also suggest a low contribution of the virulence profile of K. pneumoniae toward CM severity. Finally, the role of nif in increasing the adaptability to the udder and promoting a contagious-like spread of K. pneumoniae warrants further investigation.     

Lactobacilli isolated from chicken intestines: potential use as probiotics

Lactobacillus strains were tested for their in vitro probiotic properties. Cell surface hydrophobicity was found to be very high for Lactobacillus fermentum subsp. cellobiosus and Salmonella Gallinarum; high values could indicate a greater ability to adhere to epithelial cells. Studies on Lactobacillus animalis indicated relative cell surface hydrophobicities smaller than those of L. fermentum subsp. cellobiosus and L. fermentum. L. animalis and Enterococcus faecalis were able to coaggregate with L. fermentum subsp. cellobiosus and L. fermentum, respectively, but not with Salmonella Gallinarum. After mixed-culture studies for determining suitable growth behavior, the pair of strains L. animalis plus L. fermentum subsp. cellobiosus was selected for an attempted challenge against Salmonella Gallinarum. Double and triple mixed-culture studies indicated that selected lactobacillus strains were able to retain their beneficial characteristics in the presence of Salmonella Gallinarum such as presence of lectins, production of antimicrobial compounds, and ability to grow and compete. The selected microorganisms can be considered as potential ingredients for a chicken probiotic feed formulation intended to control salmonellosis and also improve poultry sanitation.     

Ribulose-1,5-bisphosphate carboxylase/oxygenase from an ammonia-oxidizing bacterium, Nitrosomonas sp. K1: purification and properties

The ammonia-oxidizing chemoautotrophic Nitrosomonas sp. strain K1 exhibited marked ribulose-1,5-bisphosphate carboxylase (RubisCO) activity. The RubisCO [EC 4.1.1.39] was purified as an electrophoretically homogeneous protein. The molecular mass of the enzyme was estimated to be about 460 kDa by gel filtration, and it consists of two subunits [large (L): 52.2 kDa; small (S): 13.3 kDa] as demonstrated by SDS-PAGE. This confirmed that the enzyme has an L(8)S(8) structure. The K(m) values of the enzyme for RuBP, NaHCO3, and Mg2+ were estimated to be 0.112, 0.415, and 1.063 mM, respectively. The optimum pH and temperature for its activity were approximately 7.0 and 45 degrees C. The enzyme was stable up to 45 degrees C and in a pH range from 7.0-9.0 (4 degrees C, 48 h). The enzyme activity was inhibited by Cu2+, Hg2+, N-ethylmaleimide, p-chloromercuribenzoate, and SDS (0.1 mM). The activity was also inhibited by ammonium sulfate at high concentrations (38-303 mM) but the stability of the enzyme showed no inhibition at the same ammonium sulfate concentrations. The N-terminal amino acid sequences of the large and small subunits are AIKTYQAGVKEYRQTYW QPDYVPL and AIQAYHLTKKYETFSYLPQM, respectively.