Outer membrane protein A of Acinetobacter baumannii induces autophagy via Akt/mTOR/p70S6K signaling pathway in RAW264.7 cells

AIM:To analyze the effects of outer membrane protein A (OmpA) from Acinetobacter baumannii ATCC 19606 on the autophagy of RAW264.7 cells. METHODS:The RAW264.7 cell model stimulated by OmpA was established. The effects of OmpA on the autophagy of RAW264.7 cells were detected by immunofluorescence, Western blot and transmission electron microscopy. RESULTS:The OmpA increased the expression of LC3B-Ⅱ and reduced the phosphorylation levels of Akt, mTOR and p70S6K. Rapamycin further reduced the phosphorylation levels of mTOR and p-70S6K, and increased the expression of LC3B-Ⅱ induced by OmpA. CONCLUSION:The OmpA of Acinetobacter baumannii induces autophagy via Akt/mTOR/p70S6K signaling pathway in the RAW264.7 cells. This work provides a basis for further research on the molecular mechanism of autophagy induced by Acinetobacter baumannii to find a new method against the infection of Acinetobacter baumannii.     

Identification and pathogenicity study of emerging fish pathogens Acinetobacter junii and Acinetobacter pittii recovered from a disease outbreak in Labeo catla (Hamilton, 1822) and Hypophthalmichthys molitrix (Valenciennes, 1844) of freshwater wetland in West Bengal,India

The disease outbreaks in aquaculture system of wetlands are the major cause of fish mortality. Among various bacterial septicaemic diseases, fish mortality caused by Acinetobacter spp. is recently reported in different fish species. Fish disease outbreak was investigated in a wetland of West Bengal, India to identify the aetiological factors involved. The moribund fish were examined and subjected to bacterial isolation. Two bacterial causative agents were identified as Acinetobacter junii and Acinetobacter pittii by biochemical characterization and 16S rRNA gene amplification. Both the isolates were oxidase‐negative, nitrate‐negative, catalase‐positive and indole‐negative. The molecular identification using 16S rRNA gene sequencing and phylogenetic tree analysis further confirmed the two Acinetobacter spp. with 97%–99% similarity. The antibiotic resistance patterns of these two bacteria revealed that both of them were resistant to β‐lactam, cefalexin, cephalothin, amoxyclav, cefuroxime, cefadroxil, clindamycin, vancomycin and penicillin. In addition, A. pittii was also resistant to other antibiotics of cephams group such as ceftazidime and cefotaxime. In the challenge experiment, both A. junii and A. pittii were found to be pathogenic with LD₅₀ of 1.24 × 10⁵ and 1.88 × 10⁷ cfu/fish respectively. Histopathological examination of gill, liver and kidney revealed prominent changes supporting bacterial septicaemia. The investigation reports for the first time on concurrent infection by A. junii and multidrug‐resistant (MDR)‐A. pittii as emerging fish pathogens to cause severe mortality in Labeo catla and Hypophthalmichthys molitrix in a freshwater wetland.     

鲍曼不动杆菌雏鸡分离株CCGGD201101的分离、鉴定及致病性研究简

试验旨在鉴定吉林省某雏鸡孵育基地病死雏鸡组织中分离出的1株致病性菌CCGGD201101株并测定其致病性。对疑似致病菌进行生理生化试验、16S rDNA测序鉴定，并人工接种昆明鼠，测定其半数致死量，验证细菌毒力。经鉴定该菌为鲍曼不动杆菌（Acinetobacter baumannii）。以鲍曼不动杆菌CCGGD201101分离株为研究对象，并以鲍曼不动杆菌标准株（ATCC 19606）为对照，测得半数致死量，进一步证明鲍曼不动杆菌病死鸡分离株CCGGD201101具有较强致病性。     

琼氏不动杆菌GXP04的苯酚羟化酶基因的克隆及序列分析

提取琼氏不动杆菌GXP 04的总DNA,采用限制性内切酶E coRⅠ酶切处理后,构建以pLAFR 3为载体的基因组文库。通过TA IL-PCR扩增出苯酚羟化酶的上游基因,对菌落原位杂交获得的11个转化子进一步鉴定,最终确定其中的8个转化子含有完整的苯酚羟化酶基因,选取其中的pLAFR 3-7对GXP 04的苯酚羟化酶基因的核苷酸序列进行分析,并对其蛋白质基本性质进行预测。     

银鲫不动杆菌病原菌鉴定及药敏试验

从患病的异育银鲫体内分离到5株可疑菌株,经API 20NE细菌生化鉴定,YCS07-04株鉴定率为99.5%,T值符合率0.9,菌株鉴定为鲍曼不动杆菌。采用3种不同浓度进行感染试验,5.0×106cfu/ml浓度组注射感染14 d死亡率达100%,浸泡感染14 d死亡3尾;对照组未出现病症和死亡。菌株在营养琼脂菌落生长良好,在营养肉汤培养基中混浊,有菌膜及沉淀生长;血琼脂平皿菌落呈灰白色,直径2~3 mm,不溶血;麦康凯琼脂菌落半透明圆形;在SS琼脂培养基中不生长。经负染电镜观察,菌株末端圆形短杆状,偶尔可见不规则丝状体,大小为(1.0~1.5)μm×(1.5~2.5)μm,无芽孢、荚膜和鞭毛,表面凹凸不平,静止期呈球形。药敏试验表明,菌株对丁胺卡那霉素、链霉素、妥布霉素、庆大霉素、左氟沙星、复方新诺明和洛美沙星高度敏感;对四环素中度敏感;对羧苄青霉素、氧哌嗪青霉素、苯唑青霉素、头孢他啶、头孢噻肟、头孢哌酮、头孢三嗪和氨曲南等耐药。     

1株奶牛源约翰逊不动杆菌的分离鉴定

旨在了解从奶牛垫料中分离得到1株革兰阴性菌的病原特征,本研究通过形态学观察、生化试验、染色镜检、16S rDNA同源性分析等方法对分离菌进行鉴定,并采用药敏试验测定药物敏感性,小鼠攻毒试验确定分离菌株毒力情况。结果表明,分离菌为革兰阴性菌;扩增的16S rDNA序列长度为1 283 bp,与约翰逊不动杆菌的核苷酸相似性大于99%;分离菌株对小鼠有较强的致病性,试验组小鼠死亡率达80%。分离菌对链霉素、复方新诺明等6种药物敏感,对头孢噻肟、头孢呋辛中度敏感,对青霉素、红霉素等4种药物耐药。本研究从奶牛垫料中分离到1株约翰逊不动杆菌,为奶牛源约翰逊不动杆菌的防治和研究提供了参考依据。     

牛源皮特不动杆菌对小鼠的致病性分析

为探究来源于牛呼吸道疾病综合征发病牛的皮特不动杆菌对小鼠的致病性,对其进行相关毒力基因的检测和小鼠致病性试验,试验内容主要包括临床症状观察、病理剖检、组织病理学观察,以及半数致死量(LD₅₀)和各器官载菌量的测定。结果显示,牛源皮特不动杆菌ZZCNC1807-6和ZZCSF1807-9菌株对小鼠均具有较强的致死性,对小鼠的LD₅₀分别为4.487×10⁷ CFU·mL^-1和3.177×10⁸ CFU·mL^-1。剖检死亡小鼠可见,牛源皮特不动杆菌主要造成小鼠皮下充血、肺淤血肿胀、肠道积气积液等症状,可引起心、肝、脾、肺、肾、脑等多种器官感染。组织病理学观察显示,小鼠各器官病变均较轻微,主要表现在肝、肺中有少量炎性细胞浸润,脾中白髓细胞坏死和纤维增生,其中ZZCNC1807-6攻菌组的病变较ZZCSF1807-9攻菌组相对严重。载菌量分析结果显示,2株菌对小鼠肝、心、脾、肺和肾的侵袭力差异不大,但ZZCNC1807-6对脑的侵袭力强于ZZCSF1807-9。综上,ZZCNC1807-6菌株的毒力强于ZZCSF1807-9。     

1株罗曼粉鸡源申氏不动杆菌的分离鉴定及耐药性分析

为了解贵州省某养殖场罗曼粉鸡发病病原的特征及其耐药性,本试验从病死鸡中分离出一株革兰氏阴性菌,命名为GZ2019,并对其进行纯化培养、生化试验、16S rRNA序列分析、药敏试验及动物回归试验。分离菌在普通琼脂培养基上呈圆形凸起、表面光滑、半透明的灰白色小菌落,鲜血营养琼脂培养基上菌落形态与普通琼脂培养基上的一致,但不溶血;革兰氏染色结果镜检显示,分离菌为成双排列的阴性球杆菌,偶见单个存在或链状排列;生化试验结果显示,分离菌枸橼酸盐利用、过氧化氢酶反应为阳性,硝酸还原、氧化酶、葡萄糖发酵等反应为阴性,无运动性;16S rRNA序列分析结果显示,分离株与不动杆菌的核苷酸同源性在72.6%~99.9%之间,与申氏不动杆菌LUH 4760株（GenBank登录号：AJ275041）、SNSK 752株（GenBank登录号：MG584984）、MCDA01株（GenBank登录号：KY385627）及RP1株（GenBank登录号：MG461636）的核苷酸同源性高达99.9%;药敏试验结果显示,分离菌对头孢曲松和痢特灵敏感,对头孢哌酮、头孢呋辛、头孢他啶等5种药物中度敏感,对新霉素、环丙沙星、羧苄西林等17种药物耐药;动物回归试验结果显示,试验组小鼠隔离饲养1周仅死亡1只,死亡率为20%,表明该分离菌致病性较弱。本试验成功分离出1株低致病性申氏不动杆菌GZ2019,可为鸡源申氏不动杆菌病的预防和治疗提供一定的参考依据。     

Role of Some Plant Growth Promotory Bacteria on Enhanced Fe Uptake of Wheat

The quest for plant growth promotory bacteria that can help in sustainable agriculture has increased in the recent years owing to increasing cost of fertilizers. In the present study, the potential of three isolated bacteria having plant growth promotory effects in mobilizing the indigenous soil iron (Fe) in two different varieties of wheat was evaluated. Three bacterial isolates belonging to the genera Burkholderia and Acinetobacter were evaluated singly and in combinations for Fe uptake of wheat in a net house study. Results showed that bacterial isolates when used in combination performed better in comparison to their individual use. These isolates significantly increased the mean Fe concentration in grains and straw by 34.0% and 52.4% over the uninoculated controls and also increased the total Fe uptake per pot in comparison to uninoculated controls. The bacterial inoculation also showed a positive impact on the bioavailability of diethylenetriaminepentaacetic acid (DTPA) extractable Fe in soil.     

Bacterial Pleuritis with Thickened Mesothelial Hyperplasia in a Young Beagle Dog

A five-month-old male beagle dog suddenly became moribund. Bloody fluid accumulated in the thoracic and abdominal cavities, and soft yellow flecks were floating in the thoracic fluid. The mediastinum and pericardium became dark reddish with villous thickening. Other parietal and pulmonary pleurae were rough, and the organs adhered to each other. Histologically, most mediastinal pleura formed papillary projections covered by a single layer of mesothelial cells. Many macrophages and neutrophils infiltrated the submesothelial connective tissue. At the mediastinum adjacent to the pericardium, cuboidal mesothelial cells proliferated solidly and formed a thick surface stratum. The flecks consisted of gram-negative filamentous or small bacillary (coccoid) bacteria. In the right posterior lobe of the lung, neutrophilic infiltration and a large encapsulated abscess including a bacterial colony were present. We diagnosed this case as “bacterial pleuritis with thickened mesothelial hyperplasia”. The cause of the pleuritis might be a chronic pleural infection spread via the lung abscess.