植生克雷伯氏菌（Klebsiella planticola 19-1）nifA-like基因的克隆

植生克雷伯氏菌(Klebsiella planticola 19-1)是从新疆鄯善地区玉米根际分离得到的一株联合固氮菌。在40℃高温下有较强的乙炔还原活性。 本工作利用Southern Blot分子杂交技术, 以Klebsiella pneumoniae的nifA为探针，证明了在K．planticola 19-1中存在nifA-like基因，由nifH-lacZ实验推论其nifA-like基因产物对高温相对稳定。经过大质粒电泳和Southern Blot分子杂交，发现nifA-like基因定位于染色体外的大质粒上。本工作进一步克隆了含有K．plonticola 19-1的nifA-like基因的DNA片段，做了它的限制性酶切图谱，并将nifA-like基因初步定位。

联合固氮菌Klebsiella oxytoca SA2和水稻幼苗的相互作用

从内蒙流沙地先锋植物沙竹（Psammochloa mongokca）内分离到一株内生细菌，经鉴定定名为Klebsiella oxytoca SA-2 K.oxytoca SA-2兼性厌氧固氮，NH4+抑制其固氮酶合成。部分抑制固氮酶活性;N03 -抑制其固氮酶的合成和活性。 60℃灭活K.oxytoca SA-2整体菌免疫兔子得到抗血清。免疫印迹表明此抗血清具K.oxytoca SA-2种特异性。石蜡切片免疫金银染色结合显微观察发现K.oxytoca SA-2定殖于沙竹叶鞘薄壁细胞和叶片的薄壁细胞内。 K.oxytoca SA-2在半固体培养基中接种水稻幼苗，限菌培养21天，根内重新分离的数量达l06 cfu/g．FWroot，但K.oxytoca SA-2在富养的土壤中生长良好，表现为兼性内生菌。 限菌培养水稻（Oryza sativa）幼苗，石蜡切片免疫金银染色结合显微观察研究了的K.oxytoca SA-2的侵染特性．K.oxytoca SA-2可以通过侧根发生处和表皮细胞胞间层进入根内，在皮层薄壁细胞间隙大量定殖，在解体和看似完整的薄壁细胞内也有定殖，在根和茎中柱内K.oxytoca SA-2进入了木质部导管。在根基，K.oxytoca SA-2大量侵入了已解体的内皮层和中柱鞘细胞，植物细胞在K.oxytoca SA-2侵入后解体，可能表现为严格的局部超敏反应。 接种K.oxytoca SA-2 21天，水稻地上苗部分没有发现肉眼和显微可见的病症。与对照相比，接种K.oxytoca SA-2显著促进限氮培养水稻幼苗的生长。由于K.oxytoca SA-2在限碳限氮培养基和水稻幼苗共培养时能分泌NH4+和植物激素，它可能通过向水稻幼苗提供氮素和分泌植物激素促进植物生长。而且用固氮酶铁蛋白抗血清进行免疫金银染色发现定殖在根基皮层薄壁细胞胞间层和细胞间隙，木质部导管和茎基木质部导管的K.oxytoca SA-2可以表达固氮酶，固氮参与了K.oxytoca SA-2在水稻幼苗中的内生。 培养基内碳源（苹果酸）和培养温度对K.oxytoca SA-2和水稻幼苗相互作用的影响也进行了研究。 研究表明，K.oxytoca SA-2作为兼性内生固氮菌，能够和植物紧密联合，并在植物体内开拓一个有利的生态位固氮，而且K.oxytoca SA-2可以分泌NH4+和植物激素，在和植物相互作用中使植物受益。

Diverse tetracycline resistant bacteria and resistance genes from coastal waters of Jiaozhou Bay

Environmental microbiology investigation was carried out in Jiaozhou Bay to determine the source and distribution of tetracycline-resistant bacteria and their resistance mechanisms. At least 25 species or the equivalent molecular phylogenetic taxa in 16 genera of resistant bacteria could be identified based on 16S ribosomal deoxyribonucleic acid sequence analysis. Enterobacteriaceae, Pseudomonadaceae, and Vibrionaceae constituted the majority of the typical resistant isolates. Indigenous estuarine and marine Halomonadaceae, Pseudoalteromonadaceae, Rhodobacteraceae, and Shewanellaceae bacteria also harbored tetracycline resistance. All the six resistance determinants screened, tet(A)-(E) and tet(G), could be detected, and the predominant genes were tet(A), tet(B), and tet(G). Both anthropogenic activity-related and indigenous estuarine or coastal bacteria might contribute to the tet gene reservoir, and resistant bacteria and their molecular determinants may serve as bioindicators of coastal environmental quality. Our work probably is the first identification of tet(E) in Proteus, tet(G) in Acinetobacter, tet(C) and tet(D) in Halomonas, tet(D) and tet(G) in Shewanella, and tet(B), tet(C), tet(E), and tet(G) in Roseobacter.

Dominant chloramphenicol-resistant bacteria and resistance genes in coastal marine waters of Jiaozhou Bay, China

Studies of abundance, diversity and distribution of antibiotic-resistant bacteria and their resistance determinants are necessary for effective prevention and control of antibiotic resistance and its dissemination, critically important for public health and environment management. In order to gain an understanding of the persistence of resistance in the absence of a specific antibiotic selective pressure, microbiological surveys were carried out to investigate chloramphenicol-resistant bacteria and the chloramphenicol acetyltransferase resistance genes in Jiaozhou Bay after chloramphenicol was banned since 1999 in China. About 0.15-6.70% cultivable bacteria were chloramphenicol resistant, and the highest abundances occurred mainly in the areas near river mouths or sewage processing plants. For the dominant resistant isolates, 14 genera and 25 species were identified, mostly being indigenous estuarine or marine bacteria. Antibiotic-resistant potential human or marine animal pathogens, such as Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis and Shewanella algae, were also identified. For the molecular resistance determinants, the cat I and cat III genes could be detected in some of the resistant strains, and they might have the same origins as those from clinical strains as determined via gene sequence analysis. Further investigation about the biological, environmental and anthropogenic mechanisms and their interactions that may contribute to the persistence of antibiotic-resistance in coastal marine waters in the absence of specific antibiotic selective pressure is necessary for tackling this complicated environmental issue.