Production of mycotoxins byFusarium species isolated in Germany 2. Time course of deoxynivalenol and 3-acetyldeoxynivalenol formation byFusarium graminearum in different liquid media

Summary Several semisynthetic liquid media were examined for the large-scale production of deoxynivalenol (DON) und 3-acetyldeoxynivalenol (AcDON) byFusarium graminearum 183. Only in three of the eight media used could high toxin yields of DON and AcDON be detected. The maximum levels of DON in a medium according to Miller were 3 mg/l and of AcDON 32 mg/1. In glucose-yeast extract-peptone (GYEP) medium containing 1 % glucose, the AcDON concentrations reached 33 mg/1 and the DON yields were 19 mg/1. In a rice flour liquid medium, however, the mean levels of AcDON and DON increased to 170 mg/l and 9 mg/l, respectively. The maximum amounts observed were 480 mg/1 for AcDON and 65 mg/1 for DON. The addition of trifluoracetic acid sodium salt or malonic acid, which are suggested to cause an accumulation of acetyl-CoA by inhibiting enzymes of the tricarboxylic acid cycle, did not stimulate the toxin formation.

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Bildung von Mykotoxinen durch in Deutschland isolierteFusarium-Arten. 2. Zeitverlauf der Deoxynivalenol und 3-Acetyldeoxynivalenolbildung durchFusarium graminearum in verschiedenen Flüssigmedien

Zusammenfassung Zur Herstellung präparativer Mengen von Deoxynivalenol (DON) und 3-Acetyldeoxynivalenol (AcDON) durchFusarium graminearum 183 wurden verschiedene semisynthetische Flüssigme-dien untersucht. Nur in drei von acht eingesetzten Methen konnten hohe Toxingehalte an DON und AcDON nachgewiesen werden. Im Medium nach Miller betrugen die maximal erreichten Konzentrationen von DON 3 mg/1, von AcDON 32 mg/1. In GYEP-Medium mit einem Glucosegehalt von 1 % stieg die AcDON Konzentration bis 33 mg/1, die von DON bis 19 mg/1 an. Dagegen konnten in einem auf Reismehl basierenden Flüssigmedium durchschnittlich 170 mg/l AcDON und 9 mg/1 DON und maximal 480 mg/l AcDON und 65 mg/l DON erzielt werden. Der Zusatz von Natriumtrifluoracetat oder Malonsäure, die aufgrund einer Hemmung von Enzymen des Citratcyclus zu einer Anreicherung von Acetyl-CoA und damit zur vermehrten Bildung von Sekundärmetaboliten führen sollten, brachte keine Steigerung der Toxinbildung.

     

Production of mycotoxins by Fusarium species isolated in Germany. 2. Time course of deoxynivalenol and 3-acetyldeoxynivalenol formation by Fusarium graminearum in different liquid media

Several semisynthetic liquid media were examined for the large-scale production of deoxynivalenol (DON) und 3-acetyldeoxynivalenol (AcDON) by Fusarium graminearum 183. Only in three of the eight media used could high toxin yields of DON and AcDON be detected. The maximum levels of DON in a medium according to Miller were 3 mg/l and of AcDON 32 mg/l. In glucose-yeast extract-peptone (GYEP) medium containing 1% glucose, the AcDON concentrations reached 33 mg/l and the DON yields were 19 mg/l. In a rice flour liquid medium, however, the mean levels of AcDON and DON increased to 170 mg/l and 9 mg/l, respectively. The maximum amounts observed were 480 mg/l for AcDON and 65 mg/l for DON. The addition of trifluoracetic acid sodium salt or malonic acid, which are suggested to cause an accumulation of acetyl-CoA by inhibiting enzymes of the tricarboxylic acid cycle, did not stimulate the toxin formation.     

Production of trichothecene mycotoxins by Australian Fusarium species.

Australian isolates of Fusarium species were grown on potato dextrose agar. Trichothecenes produced by these species were extracted by ethyl acetate followed by methanol and a silica gel column was used to clean-up the extract. The extracted samples were derivatized by acetylation with trifluoroacetic anhydride and the derivatives analysed by gas chromatography/mass spectrometry (GC/MS). Multiple ion detection was used to trace ions characteristic of the trichothecenes expected to be present. Quantitation of those found was based on a known mass of pentabromophenol that was added as an internal standard. Eight species of Fusarium (nineteen strains) were surveyed, of which three species, F. acuminatum, F. equiseti and F. sporotrichioides, produced the trichothecenes scirpentriol, diacetoxyscirpenol, neosolaniol, HT-2 toxin, T-2 toxin, T-2 tetraol and deoxynivalenol. Wheat samples were inoculated with four different species of Fusarium, F. acuminatum, F. equiseti, F. graminearum and F. sporotrichioides, and in these samples diacetoxyscirpenol, neosolaniol, HT-2 toxin and T-2 toxin were found.     

Production of alternaria mycotoxins by Alternaria alternata isolated from weather-damaged wheat

The production of Alternaria mycotoxins by Alternaria alternata isolated from Chinese weathered wheat kernels were first investigated on polished rice and durum wheat grains. These mycotoxins included alternariol (AOH) and its monomethyl ether (AME), altenuene (ALT), altertoxin I (ATX-I), and tenuazonic acid (TA). Of 25 isolates tested, all were AOH and AME producers, 21 (84%) coproduced ALT and ATX-I, and 8 (32%) produced TA in rice culture. TA was the most abundant toxin produced at a level ranging from 1,369 to 3,563 mg/kg. Much smaller amounts of AOH, AME, ALT, and ATX-I were present with average concentrations of 54, 40, 44, and 8 mg/kg, respectively. There were linear correlations between the level of AOH and AME (r = 0.846), alternariols (AOH plus AME) and ALT (r = 0.785), and ATX-I and TA (r = 0.553). Polished rice medium seems to support a bit more production of Alternaria metabolites than wheat but with an insignificant difference in concentrations (P > 0.05). A study of the time-course of toxin production by A. alternata isolates indicated that AOH production began faster than any other toxins monitored, and ALT production exhibited a progressive increase throughout the experiment. TA producers might reveal their considerably higher ability to produce toxin in the field despite their low frequency.     

Quantification of Fusarium poae DNA and associated mycotoxins in asymptomatically contaminated wheat

Recent surveys have identified increased predominance of Fusarium poae causing FHB (Fusarium Head Blight) of wheat in Europe. Several studies revealed a correlation between levels of F. poae DNA and nivalenol (NIV) and enniatins (ENNs) in highly contaminated cereal grain. In this study, F. poae specific TaqMan assays and mycotoxin analysis were performed on 48 asymptomatically contaminated wheat grain samples obtained from six different locations in northern Poland in 2006 and 2007. TaqMan assays revealed the presence of F. poae DNA in all samples analyzed, however the amounts of target DNA between the samples differed. Mycotoxin analysis revealed the presence of 13 toxins in the grain analyzed, however only ENN B and B1 were detected at quantifiable concentrations. A significant positive correlation was revealed between F. poae DNA (R = 0.75) and monthly mean rainfalls recorded in May (a month before wheat anthesis) in both years. Additionally, a significant positive correlation was found between levels of ENN B+B1 (R = 0.49) and rainfalls in May, however, no correlation was found between the quantity F. poae DNA and the level of ENN B+B1. This paper confirms the increasing importance of F. poae in the FHB complex of wheat in Poland.     

链格孢霉毒素的分析方法及其毒理机制研究进展

链格孢霉毒素是由链格孢霉属(Alternaria species)产生的次级代谢产物,因其大都具有细胞毒性、基因毒性、诱变性、致畸致癌性等毒性,食用被其污染的食物后会对人体健康造成严重危害。因此,筛选快速、简便的分析方法和研究其毒性作用机制都是当今的热点。本文主要概述了链格孢霉毒素的分类及理化性质,常见的分析方法及其毒理机制,旨在为链格孢霉毒素的风险评估和中毒防治等研究提供参考,同时对于人和动物的健康安全也具有重要意义。     

Short communication: Streptococcus species isolated from mastitis milk samples in Germany and their resistance to antimicrobial agents

Mastitis is one of the most frequent infectious diseases in dairy cattle and is a reason for antimicrobial drug usage in dairy cows. The bacteria involved in bovine mastitis are mainly Streptococcus spp., Staphylococcus spp., and coliforms. The aim of this study was to determine antimicrobial resistance among Streptococcus spp. isolated from bovine mastitis milk. Antimicrobial resistance in Strep. uberis (n = 227), Strep. dysgalactiae (n = 49), and Strep. agalactiae (n = 3) was determined for 9 antimicrobial agents using the broth microdilution method in accordance with Clinical and Laboratory Standards Institute recommendations. Of all Streptococcus spp., 13% were multidrug resistant. The rate of multidrug resistance was higher among Strep. uberis (15%) than among Strep. dysgalactiae (6%) and Strep. agalactiae (0%). Resistance to tetracycline was the most common, followed by resistance to erythromycin, pirlimycin, and gentamicin. Resistance rates were higher on farms with more than 80 cows compared with those with fewer than 20 cows. β-Lactams should remain the drugs of choice in the treatment of streptococcal mastitis. The slightly elevated minimum inhibitory concentrations determined for these antibiotics may indicate, however, the emergence of resistant streptococci. To identify such changes in susceptibility as early as possible, antimicrobial resistance in streptococci should be surveyed regularly.     

Production of trichothecenes and zearalenone by Fusarium species isolated from wheat

Fusaria isolates from wheat were tested for ability to produce trichothecenes and zearalenone. Four isolates of F. culmorum out of 13 produced vomitoxin (DON) and 3 Ac-DON, one produced diacetoxysirpenol and 12 zearalenone. Particularly high yield of zearalenone was observed in cultures of sever pathogenic isolates. Higher temperature (20 °C) during first week of incubation favoured yield of zearalenone. About 50% of zearalenone was produced by surface mycelium.     

基于Meta分析对全球小麦及小麦粉中多种真菌毒素污染特征的比较研究

目的 针对小麦及小麦粉中脱氧雪腐镰刀菌烯醇（DON）、黄曲霉毒素（AFs）、玉米赤霉烯酮（ZEN）、雪腐镰刀菌烯醇（NIV）、T-2毒素（T2）、HT-2毒素（HT2）、赭曲霉毒素A(OTA)、伏马菌素（FBs）、交链孢酚（AOH）、交链孢酚单甲醚（AME）、腾毒素（TEN）及交链孢菌酮酸（TeA）等真菌毒素的全球污染情况进行Meta分析。方法 数据主要来自PubMed、Web of Science、知网及万方数据库。结果 通过文献筛选,共纳入69篇文献22 308个样本。小麦及小麦粉中以上真菌毒素的全球总体污染率为58%(95%CI:51%～66%),其中,TeA为99%、TEN为88%、DON为85%、AFs为57%、ZEN为42%、T2为39%、AOH为30%、AME为29%、NIV为28%、HT2为25%、OTA为21%及FBs为16%;小麦及小麦粉中真菌毒素的全球总体污染水平为32.80μg/kg(95%CI:24.96～43.10μg/kg),DON在小麦及小麦粉中的含量最高,为317.53μg/kg,其次为TeA 117.37μg/kg及FBs 45.09μg/kg。DO...     

Production of -aminobutyric acid (GABA) by Lactobacillus paracasei isolated from traditional fermented foods

Lactobacillus strains that accumulated γ-aminobutyric acid (GABA) in culture medium were screened to determine strains with high GABA-producing ability. One strain, NFRI 7415, which was isolated from a Japanese traditional fermented fish (funa-sushi), showed the highest GABA-producing ability among the screened strains. Identification tests (i.e., 16S rDNA sequencing and sugar assimilation ability) indicated that NFRI 7415 belongs to Lb. paracasei. The GABA production was further improved by the addition of pyridoxal phosphate to the culture medium and pH regulation of culture medium at pH 5.0. Under optimal cultivation conditions, strain NFRI 7415 produced GABA at a concentration of 302 mm when the glutamate concentration in the culture medium was 500 mm.     

Production of mycotoxins by Penicillium expansum inoculated into apples

We investigated the production of mycotoxins in apple fruits inoculated with spores of 40 strains of apple blue mold, Penicillium expansum. Patulin and citrinin contents in the extracts from apples stored at 25 degrees C for 12 days after inoculation were determined by high-performance liquid chromatography (HPLC) analysis with UV and fluorescence detection. Patulin and citrinin were produced by 90% (36) and 80% (32) of the 40 strains, indicating that P. expansum is a consistent producer of these mycotoxins. The patulin content in the extracts was substantially higher than the citrinin content. Other mycotoxins whose production in pure culture has been reported were simultaneously detected with high-resolution liquid chromatography-mass spectrometry (LC-MS) analysis with the positive ion mode of electrospray ionization. Along with patulin and citrinin, expansolides A and B were identified based on the HPLC and LC-MS spectral data and detected in 88% (35) of the extracts. The results indicate that P. expansum is a consistent producer of expansolides A and B in rotten areas of apple fruits. The findings raise the possibility that products from decayed apples might contain expansolides A and B in addition to patulin and citrinin.     

荧光光谱法研究链格孢霉毒素TeA与血清白蛋白的互作机理

链格孢霉毒素是由链格孢霉菌属(Alternaria species)产生的次级代谢产物,在水果、蔬菜以及粮食作物中污染率极高,可通过多种途径进入人类食物链,引发食品安全问题.为研究链格孢霉毒素细交链格孢菌酮酸(Tenuazonic Acid,TeA)在动物体内的吸收、分布、转运及代谢,本文采用稳态荧光光谱、同步荧光光谱...     

Comparison of antimicrobial resistance in Salmonella and Campylobacter isolated from turkeys in the Midwest USA

The current study was carried out to determine the antimicrobial resistance profiles and evaluate some molecular characteristics of a set of Salmonella and Campylobacter isolates recovered from production line turkeys in the Midwest region of the United States. A total of 94 birds identified as being positive for both Salmonella and Campylobacter spp. were selected for study. All Salmonella isolates were examined for antimicrobial resistance using the methods employed in the National Antimicrobial Resistance Monitoring System (NARMS). Campylobacter isolates were subjected to similar analysis using the Etest^®. In addition, polymerase chain reaction (PCR) was carried out to determine the presence of the antimicrobial resistance associated genes, integrase (int1), class 1 integrons (Salmonella and Campylobacter) and a multidrug efflux pump (Campylobacter spp.). Results from the study showed that the Salmonella and Campylobacter isolates examined displayed resistance to a number of antimicrobials, with Salmonella and Campylobacter isolates being resistant to at least three antimicrobials while some isolates showed resistances to 6 or 8 different antimicrobials. In addition, 68.1% of the Salmonella isolates tested were found to be positive for the class I integrase gene (int1), 28.7% possessed a 1000 bp gene cassette and 17% possessed an 800 bp gene cassette. All Campylobacter isolates were negative for int1, but 36.2% tested positive for the Campylobacter multidrug efflux pump (CmeB). A considerable number of Salmonella and Campylobacter isolates tested displayed varying degrees of antimicrobial resistance as well as the presence of some factors associated with the carriage and persistence of antimicrobial resistance. Similarities in the types of antimicrobial resistance observed in Campylobacter and Salmonella strains was evident. The results of this study suggest that prescribing practice at the farm level may be a factor in promoting antimicrobial resistance in more than one species of organism. Such practices may, therefore, contribute to the potential health risk for consumers should micro-organisms carrying multiple antimicrobial resistances enter the food chain. This study may be one of the first to report on the incidence of the multidrug efflux pump (CmeB) in Campylobacters recovered from processed turkeys. The antimicrobial resistance and molecular characteristics of Salmonella and Campylobacter is discussed.     

Characterization of Vibrio cholerae O139 isolated in Taiwan

Vibrio cholerae O139 Bengal is the second aetiologic agent of the cholera epidemic that emerged in late 1992 in India which has subsequently spread to other nations. A few sporadic cases of V. cholerae O139 infection have been reported in Taiwan since 1997. The V. cholerae O139 strains isolated from clinical and environmental sources in Taiwan were characterized based on subspecies molecular typing, toxin production, and susceptibility to antibiotics and environmental stresses, and were compared to several O1 and other non-O1/non-O139 strains. Typing by pulsed-field gel electrophoresis revealed that the domestic O139 strains were identical to three of the four Japanese strains. Most of these domestic O139 strains were alpha-haemolytic and produced highly variable amounts of cholera toxin. The presence of cholera toxin gene was confirmed in all of these domestic strains by polymerase chain reaction. Meanwhile, most of the domestic O139 strains were resistant to nitrofurantoin, streptomycin, furazolidone and trimethoprim–sulfamethoxazole, while the O1 strains were not. Susceptibility of selected O139 strains to temperature stresses at 55°C and −20°C, acid treatment at pH 3·0, and N-alkyl dimethyl benzyl ammonium treatment did not differ significantly from other strains examined. Accordingly, these Taiwanese O139 strains were genetically close to the Japanese strains and also shared some common biological traits.     

唐菖蒲伯克霍尔德菌椰毒致病变种食物中毒分离株的遗传特性研究

目的 对1株引起聚集性米酵菌酸中毒的生吊面浆食品样品分离株唐菖蒲伯克霍尔德菌椰毒致病变种(Burkholderia gladioli pv. cocovenenans)DBJ进行全基因组测序并分析,解析菌株产毒及致病的基因特征。方法 提取唐菖蒲伯克霍尔德菌椰毒致病变种DBJ基因组DNA,对菌株进行测序获得全基因组完成图,利用生物信息学分析手段对序列进行挖掘分析。结果 菌株DBJ基因组由两个染色体(G1和G2)和一个质粒(P)组成,其中两个染色体大小均为4 Mb左右,质粒大小约为300 kb。两个染色体的GC含量也相仿,约为68.0%,质粒则稍低为63.0%。进一步分析发现,G2染色体上存在产米酵菌酸的bon基因簇。遗传进化分析后发现,菌株DBJ在亲缘关系上与加拿大玉米分离株UCD-UG_CHAPALOTE最为接近,两株菌处在同一进化分支。且255株菌种中有31株携带bon基因簇的菌株,较不携带该基因的唐菖蒲伯克霍尔德菌有明显遗传进化倾向。结论 唐菖蒲伯克霍尔德菌椰毒致病变种DBJ分离株基因组中携带产米酵菌酸的bon基因簇,是引起食物中毒的主要原因。     

解淀粉芽孢杆菌psd基因过表达对胞苷发酵的影响

以解淀粉芽孢杆菌BG-09(Bacillus amyloliquefaciens BG-09)为出发菌株,分析代谢途径中与细胞膜渗透性有关的基因psd,研究过表达基因psd对胞苷发酵的影响。以解淀粉芽孢杆菌BG-09基因组为模板克隆psd,构建了含psd的重组质粒pHT43-psd,通过化学转化法转入菌株大肠杆菌DH5α,验证成功后将其转入B.amyloliquefaciens BG-09,获得重组菌株B.amyloliquefaciens BG-09-psd。将B.amyloliquefaciens BG-09-psd菌株通过摇瓶发酵,研究过表达基因psd对菌体的生长、胞苷和尿苷产量积累的影响。结果显示,重组菌株B.amyloliquefaciens BG-09-psd发酵液中胞苷浓度为1.199 g/L,与对照菌株B.amyloliquefaciens BG-09相比,提高了15.51%,尿苷浓度为0.552 g/L,增加了6.56%,表明过表达基因psd可促进胞苷的积累。     

谈谈德国印刷行业中的安全生产管理

~~谈谈德国印刷行业中的安全生产管理@胡维友$合肥中德印刷培训中心~~     

基于钝齿棒杆菌argGH启动子改造的L-瓜氨酸高效合成

L-瓜氨酸是尿素循环的重要中间体,在argGH编码酶的催化下易分解为L-精氨酸,在微生物体内难以大量积累。作者通过启动子替换手段,以弱启动子P-dapAB6替换调控argGH表达的启动子P-argG,构建了弱化L-瓜氨酸分解代谢途径的重组菌株Corynebacterium crenatum H-7-PdapAB6:argGH。重组菌株的转录水平和酶活力结果显示,重组菌株分解途径中argG和argH的基因表达水平下调,精胺琥珀酸合成酶ASS(argG编码)和精胺琥珀酸裂解酶ASL(argH编码)酶活分别降低91.80%和55.35%。摇瓶发酵结果表明,L-瓜氨酸的产量、糖酸转化率和生产强度分别为33.85 g/L、0.25 g/g和0.34 g/(L·h),较原始菌株分别提高了4.91、5.00和4.86倍。通过启动子替换策略,构建了L-瓜氨酸分解代谢途径弱化的工程菌株,初步实现了L-瓜氨酸的高效合成。