Serratia marcescens forms a new type of cytolysin

Most Serratia marcescens strains produce a new type of cytolysin (hemolysin) which is also found in other Serratia species. The hemolytic polypeptide ShlA (M(r) 162 101) is secreted across the outer membrane through the help of the ShlB protein which also involves conversion of an inactive precursor in an hemolytically active form. Both proteins are synthesized with signal sequences which are released during export across the cytoplasmic membrane. Mutants expressing inactive ShlB derivatives are impaired in activation and secretion suggesting a tight coupling between both processes. The region of ShlA for activation and secretion is confined to the N-terminal 16% of the polypeptide which contains the sequence NPNG which is also found in the Proteus hemolysin, the Bordetella pertussis filamentous hemagglutinin and two highly expressed outer membrane proteins of Haemophilus influenzae. Substitution of the first asparagine (N) residue by isoleucine converts the Serratia hemolysin into an inactive secretion incompetent form. It is concluded that this region is recognized by ShlB for activation and secretion of ShlA. The Serratia hemolysin forms defined pores in erythrocyte membranes.     

Using selective media to assess aerosolization damage and ultraviolet germicidal irradiation susceptibility of Serratia marcescens

This study determined whether selective media, McConkey agar (MC) and minimal salt glucose agar (MA) are suitable for monitoring aerosolization damage of airborne Serratia marcescens in our laboratory aerosol exposure system and assessed the relationship between bacterial culturability in these media and ultraviolet germicidal irradiation (UVGI) susceptibility of the bacteria. Two types of bacterial cultures were prepared. The first culture was taken from bacteria growing on Tryptic soy agar (TSA) as complete medium (fresh culture), which provided nearly 100% of MC/MA tolerant bacteria, while the second one was prepared from freezing the fresh culture (frozen culture), which produced 55 and 81% of MC and MA tolerant bacteria, respectively. We monitored bacterial culturability in TSA, MC and MA from these cultures in the nebulizer reservoir and bioaerosls collected on a six-stage Andersen cascade bio-impactor. The results indicated that both concentration and percentage of MC/MA tolerant bacteria maintained at a similar level during nebulization. For the bioaerosols, although the concentration recovered in the media from the fresh culture was higher than that from the frozen culture, the percentage of MC/MA tolerant bacteria was similar to that before aerosolization. We concluded that MC and MA are not suitable for monitoring aerosolization damage of the bacteria. Moreover, culturability of the bacteria in MC and MA has no effect on their survival after aerosolization. With respect to the bacterial susceptibility to UVGI, MC/MA sensitive and tolerant population as well as the fresh and frozen cultures showed the same susceptibility.     

生防细菌CNY-04筛选、鉴定及其对灰霉病菌的防效

【目的】为研究土壤细菌对蔬菜灰霉病的生防价值, 从辽宁、山东等地区的蔬菜种植基地采集土壤样本56份, 分离、筛选出对灰霉病具有稳定拮抗作用的细菌9株。【方法】采用平板对峙培养法进行初筛、复筛, 用抑菌圈法测定其抑菌效果, 并进行离体果实试验验证其对蔬菜灰霉病的防治效果, 通过形态学特征、生理生化特征及16S rRNA基因序列分析研究其分类地位。【结果】细菌CNY-04对蔬菜灰霉病的拮抗能力最强且遗传稳定, 抑菌圈直径达到34 mm; 初步鉴定该菌株为格氏沙雷菌(Serratia grimesii), 尚未见该菌在生防上的报道; CNY-04液体菌剂对离体番茄果实灰霉病的防效为69.23%, 50%多菌灵防效为75.39%, 24 h时接种CNY-04处理的番茄发病率为40.0%, 而48 h时接种处理的发病率为51.1%。【结论】CNY-04是一株较为理想的拮抗菌, 丰富了生防资源。     

粘质沙雷菌裂解性噬菌体ФSM9-3Y的分离和鉴定

目的从医院污水中分离粘质沙雷菌噬菌体,并分析其生物学特性,为进一步研究针对耐药性粘质沙雷菌的噬菌体制剂提供依据。方法采用双层琼脂平板法分离纯化针对粘质沙雷菌的裂解性噬菌体,观察噬菌体对宿主菌的裂解特异性,通过负染法电镜观察噬菌体的形态结构,提取噬菌体核酸进行酶切电泳,测定噬菌体的最佳感染复数和一步生长曲线,SDS-PAGE电泳初步分析噬菌体的结构蛋白和非结构蛋白。结果从医院污水分离出7株可裂解粘质沙雷菌的噬菌体,对其中一株噬菌体(命名为ФSM9-3Y)的生物学特征进行了初步研究。电镜显示噬菌体呈蝌蚪状,头部为20面体立体对称、直径约70 nm;尾部长约50 nm。ФSM9-3Y的最佳感染复数为1。一步生长曲线表明;ФSM9-3Y的潜伏期约30 min,暴发时间70 min,暴发量为629 PFU/cell。凝胶电泳显示噬菌体基因组为双链DNA、大小约54 kb。SDS-PAGE呈现至少包括13种蛋白,相对分子质量范围在25～130 kD,其中主要蛋白的相对分子质量约为48 kD。结论此次分离的噬菌体ФSM9-3Y为裂解性噬菌体,根据形态和结构特征,粘质沙雷菌噬菌体ФSM9-3Y属于有尾病毒目,肌尾噬菌体科。     

Production and characterization of a cold-active and n-hexane activated lipase from a newly isolated Serratia grimesii RB06-22

Abstract

A lipase-producing bacterium isolated from raw milk was identified as Serratia grimesii based on 16S rRNA sequence analysis. The extracellular lipase was partially purified by ammonium sulfate precipitation and ultrafiltration. Maximal activity was observed at 10°C, the optimum pH was 8.0 and the enzyme was stable at 5–30°C for 1 h. The K_m and V_max values were 1.7 mM and 0.3 mM/min respectively. It was found that the lipase had the highest hydrolytic activity towards sunflower oil and soybean oil. CaCl₂ had a stimulatory effect on lipase activity, while EDTA and iodoacetic acid slightly inhibited the lipase activity and the enzyme was strongly inhibited by PMSF. The enzyme was compatible with various non-ionic surfactants as well as sodium cholate and saponin. In addition, the enzyme was relatively stable towards oxidizing agents. This lipase exhibited maximum activity in 35% n-hexane retaining about 2191% activity for 1 h.     

Y-linked variation for autosomal immune gene regulation has the potential to shape sexually dimorphic immunity

Sexually dimorphic phenotypes arise from the differential expression of male and female shared genes throughout the genome. Unfortunately, the underlying molecular mechanisms by which dimorphic regulation manifests and evolves are unclear. Recent work suggests that Y-chromosomes may play an important role, given that Drosophila melanogaster Ys were shown to influence the regulation of hundreds of X and autosomal genes. For Y-linked regulatory variation (YRV) to facilitate sexually dimorphic evolution, however, it must exist within populations (where selection operates) and influence male fitness. These criteria have seldom been investigated, leaving the potential for dimorphic evolution via YRV unclear. Interestingly, male and female D. melanogaster differ in immune gene regulation. Furthermore, immune gene regulation appears to be influenced by the Y-chromosome, suggesting it may contribute to dimorphic immune evolution. We address this possibility by introgressing Y-chromosomes from a single wild population into an isogenic background (to create Y-lines) and assessing immune gene regulation and bacterial defence. We found that Y-line males differed in their immune gene regulation and their ability to defend against Serratia marcescens. Moreover, gene expression and bacterial defence were positively genetically correlated. These data indicate that the Y-chromosome has the potential to shape the evolution of sexually dimorphic immunity in this system.     

基于易错PCR技术的黏质沙雷氏菌脂肪酶基因LipA的定向进化

利用易错PCR技术对黏质沙雷氏菌脂肪酶基因LipA进行定向进化,经过筛选最终获得一个比活力比野生酶提高了425 U/mg的突变体ep1,测序分析ep1有5个氨基酸发生了突变,与野生酶相比ep1的最适pH值由原来的8.5降低为7.5,Tm值提高3℃,Km值由原来的40 mg/mL降低为12.5 mg/mL.对其三维结构进行分析,推测酶学性质的改变可能与处在活性中心右前方双螺旋发卡结构上的158A、和处在下部β卷曲折叠拐角处的S375G的突变有关.     

A Stilbene Optical Brightener can Enhance Bacterial Pathogenicity to Gypsy Moth (Lepidoptera: Lymantriidae) and Colorado Potato Beetle (Coleoptera: Chrysomelidae)

Stilbene optical brighteners were first investigated to protect biological control agents such as viruses, fungi, and nematodes against ultraviolet light. Some are known to enhance the activity of insect viruses in Lepidoptera. In this work, one stilbene brightener, Tinopal LPW, also increased mortality of gypsy moth and Colorado potato beetle larvae when treated with bacteria/optical brightener combinations. This increase in mortality, however, did not occur for every bacteria/insect combination. In gypsy moth, a significant increase in larval mortality was observed only with Bacillus thuringiensis combined with Tinopal LPW. In Colorado potato beetle, however, the addition of Tinopal LPW increased larval mortality with all bacteria tested (B. thuringiensis, Serratia marcescens, Photorhabdus luminescens, and Chromobacterium sp.). The brightener also decreased the time to kill for these pathogens. This decrease in LT₅₀ was observed not only for bacteria+Tinopal LPW combinations, but also for combinations of Chromobacterium sp. toxin+Tinopal LPW. The mechanism for increase in bacterial toxicity by optical brighteners is compatible with mechanisms proposed for enhancement based on viral/lepidopteran/optical brightener systems that are not dependent on replication.