水产源乳酸菌的多样性及抑菌活性研究

为探究常见水产品中乳酸菌的分布情况,以及筛选出抑制水产常见致病菌的乳酸菌,从市场采集不同水产品的肠道,用乳酸菌分离筛选培养基分离乳酸菌;用16SrDNA鉴定对所分离的乳酸菌分类鉴定;应用打孔法抑菌试验测试乳酸菌对嗜水气单胞菌、副溶血弧菌、溶藻弧菌等15种水产致病菌的抑菌活性。从市场采集不同南亚热带鱼类、虾、贝、蛤等30个样品的肠道,分离出156株乳酸菌,并研究了这些乳酸菌的种属分布和抑菌活性。结果显示:菌株的革兰氏染色下的菌体形态主要有杆状、短杆状、球状、链状4种。16SrDNA鉴定主要包括乳酸乳球菌、戊糖片球菌、格氏乳球菌等11个种属。其中抑菌谱广、效果好的菌属有:假肠膜明串珠菌、肠膜明串珠菌、戊糖片球菌、乳酸乳球菌、乳明串珠菌、沙克乳杆菌和融合魏斯氏菌。本研究结果为乳酸菌在水产养殖上的应用提供理论依据。     

鲢嗜麦芽窄食单胞菌分离及药敏试验

为调查福建某养殖场鲢大面积死亡原因，对发病死亡的鲢肠道内容物、肛门出血分泌物及养殖塘池水中进行病原菌分离、鉴定，并对分离菌进行致病性及药敏试验。结果表明，分离鉴定X1为嗜麦芽窄食单胞菌（Stenotrophomonas maltophilia）,X2为嗜水气单胞菌（Aeromonas hydrophila），且这2株菌对鲢具有致病性。在药敏试验中，X1菌株对15种抗生素耐药；X2菌株对新霉素、氯霉素、青霉素G等11种抗生素敏感，对阿莫西林、复方新诺明表现耐药性。     

鲫肠道乳酸菌的分离及益生特性

为获得鱼源益生乳酸菌,本研究从健康鲫肠道内分离鉴定得到38株乳酸菌,并选择其中8株乳酸菌进行体外益生特性分析。结果显示,8株乳酸菌均能在p H=4.5和10%鲫胆汁环境中存活,对嗜水气单胞菌、豚鼠气单胞菌、无乳链球菌和副溶血弧菌均有较强的抑菌能力,但菌体表面疏水性和自凝集能力具有菌株特异性。选择5株疏水性较高(63%~89%)、自凝集能力强(37%~45%)的乳酸乳球菌进行体外黏附能力测定,结果显示,黏附能力在菌株间具有差异性(4.5%~7.9%),但均能显著降低嗜水气单胞菌NJ-35的体外黏附率,黏附抑制率达30%~35%;最后对筛选出的3株高黏附力的乳酸乳球菌进行安全性评价,发现3株菌对鲫均无致病力。鱼源乳酸菌的筛选,为鲫养殖生产中益生菌的实际应用提供了理论依据。     

金鱼竖鳞病病原菌的分离鉴定及药敏试验

为进一步了解引起金鱼竖鳞病的原因和病原,从患竖鳞病的濒死金鱼体内分离出3株细菌,经细菌形态、培养特征、生理生化特性鉴定,3株细菌均为嗜水气单胞菌。感染试验证实,分离菌具有较强的致病性。药敏试验结果显示,该菌对氟苯尼考、菌必治、左氧氟沙星、氯霉素、新霉素、恩诺沙星等药物高度敏感。     

耐喹诺酮类药物嗜水气单胞菌的分离鉴定及电镜观察

本研究对吉林省内12个水域采集和送检的66尾病鱼进行细菌分离鉴定,共检出46株嗜水气单胞菌疑似株,通过生化试验和PCR扩增气溶素基因aer鉴定法,确定其中22株为嗜水气单胞菌。进一步采用纸片扩散法和双倍稀释法测定嗜水气单胞菌分离株对多种抗生素的耐药性,其中部分菌株存在不同程度的耐药性,22株嗜水气单胞菌中有4株对喹诺酮类药物耐药,并且为交叉耐药。其对喹诺酮类药物的抑菌圈均小于19mm。嗜水气单胞菌耐喹诺酮类药物的检出率为18 2%。将病料中分离的8株嗜水气单胞菌在电镜下观察发现,2株耐药菌表面发现均匀分布的球形结构,而6株敏感菌表面未见此结构。     

中华鳖腐皮病病原菌及防治药物研究

从具有典型腐皮病症状的中华鳖体中分离到Ｐ－１,Ｐ－２两株细菌,人工感染,经菌株形态特征和培养特性及生理生化反应的鉴定,证实均为中华鳖腐皮病的病原菌,且为嗜水气单有民菌。同时进行了该菌的药物敏感试验,筛选了１５种最佳防治药物。     

鲤鱼细菌性败血症病原菌的研究

从具有典型细菌性败血症症状的病鱼或濒死鱼中分离到2 株优势菌,经分离培养、人工感染试验、毒力因子、16S rRNA基因序列的测定及药敏试验,研究分离菌的生理生化特征、致病性及药物敏感性.根据2 株分离菌的菌落形态、生化特性和16S rRNA基因序列的测定结果(同源性为99%),确定为嗜水气单胞菌.人工感染试验的症状与自然发病相似,且再次感染后又分离到该菌株.2 株菌对阿米卡星、氨曲南、头孢噻肟等15 种药物均敏感,对氨苄西林均耐药,对复方磺胺表现出菌株差异.对其敏感的15 种药物可作为防治鲤鱼细菌性败血症的首选药物.     

团头鲂肠黏膜益生乳酸菌的分离及体外抗逆性研究

从团头鲂肠黏膜上分离筛选出2株对嗜水气单胞菌有较强的抑制作用的乳酸菌菌株JK-1和JK-2,通过耐受胆盐、耐受pH值、耐受蛋白酶和耐高温等试验对这2株乳酸菌进行体外抗逆性试验研究.其中在胆盐耐受性试验中,JK-1和JK-2在胆盐浓度0.4%、培养4 h时仍有35%以上的存活率;在pH值耐受性试验时,2株乳酸菌均可在一定的酸性和碱性条件下存活;胰蛋白酶对2株菌的存活率没有影响;在70 ℃的高温中,2株乳酸菌均有一定的耐受性.     

异育银鲫致病性嗜水气单胞菌的分离鉴定与药敏特性研究

对江苏盐城某养殖场患出血病的异育银鲫(Carassius auratus gibelio)进行细菌分离鉴定和药敏试验,为该病的防控提供科学依据.从病死及濒死异育银鲫的肝脏、脾、肾中分离病原菌,经过形态观察、生理生化试验、16S rRNA序列分析,最终进行细菌鉴定,采用纸片扩散法进行药敏实验,通过人工回归感染试验确定分离菌株的致病性.分离的2株细菌(编号为WJ11-3和WJ11-4)均为嗜水气单胞菌(Aeromonas hydrophila),均能导致异育银鲫死亡,并出现与自然条件下基本一致的症状.药物敏感性试验结果显示,嗜水气单胞菌分离株WJ11-3对苯唑西林、林克霉素、万古霉素显示耐药性,对恩诺沙星等28种药物高度敏感;WJ11-4对苯唑西林、林克霉素、新生霉素显示耐药性,对氟苯尼考等28种药物高度敏感.本次异育银鲫的出血病由嗜水气单胞菌感染引起,可选用恩诺沙星、阿洛西林、氧氟沙星、苯唑西林、林克霉素、新生霉素等多种药物进行防治.     

鳖三种气单胞菌的分离与药物敏感试验

在1997～2001年对广东、湖南等地70个鳖养殖场进行常见病的调查研究、分离纯化病原后进行生化试验，鉴定出11株嗜水气单胞菌、11株豚鼠气单胞菌、6株寡源气单胞菌等，经过毒力试验后．选出较强毒力的11株细菌，用15种药物进行敏感性试验筛选出敏感药物，为药物的临床应用提供依据。     

A Lactobacillus sp. from diseased female rainbow trout, Salmo gairdneri Richardson, in Newfoundland, Canada

Abstract. Post-spawning mortalities of 2- to 3-year old female rainbow trout, Salmo gairdneri Richardson, at a Newfoundland hatchery were studied. The fish were found to have a mixed bacterial infection involving primarily a Lactobacillus sp. (a Gram-positive, chain-forming coccobacillus) and to a lesser degree Enterobacteriaceae, Aeromonas hydrophila and Pseudomonas fluorescens. The bacteria were isolated from the visceral organs, kidney, and ascitic fluid present in the coelomic cavity. The fish had extensive degeneration and necrosis of the liver, spleen and kidney, and sloughing of intestinal epithelium, but whether these were caused directly by the bacterial infection was not established because the fish appeared to have received substantial stress from the mechanical stripping and from the retention of dead, unshed eggs in the coelomic cavity. Tissue sections suggested that substantial growth of the Lactobacillus sp. was occurring within these unshed eggs. The present study describes this unusual observation and compares the Lactobacillus sp. to two other Lactobacillus isolates from cultured trout.     

Lactic acid bacteria effective for regulating the growth of contaminant bacteria during the fermentation of <Emphasis Type="Italic">Undaria pinnatifida</Emphasis> (Phaeophyta)

ABSTRACT:   Lactic acid fermentation of seaweed is a recent topic and quite limited information is available on culture conditions. To know the suitable strains for use as a starter culture for seaweed fermentation, 14 lactic acid bacteria (LAB) strains, including 11 species, were tested in culture conditions prepared with or without salt. A commercial product of Undaria pinnatifida powder was used as a substrate for fermentation without sterilizing. Starter-suitability of the LAB strains was assessed from their predominance after culture. Among the tested strains, Lactobacillus brevis, Lactobacillus plantarum, Lactobacillus casei and Lactobacillus rhamnosus showed high (>90%) predominance in their cultures, while control cultures prepared without inoculation of LAB did not show any detectable growth of acid producing bacteria and spoiled. A total of 102 strains not showing acid producing activity were isolated and characterized from spoiled cultures, and all the isolates were observed as Bacillus strains, including 64 strains (62.7%) and 16 strains (15.7%) of a Bacillus cereu s-related and B. fusiformis -related species, respectively. The Undaria substrate before fermentation contained culturable microorganisms at 1.4–3.1 × 10² CFU/g, but the Bacillus cereu s-related strain was not a major composition, suggesting a concern of selective growth of the Bacillus cereu s-related strain during the spoiled fermentation.     

刺参池塘养殖系统中1株副干酪乳杆菌的分离及其生物学特性研究

本研究利用MRS培养基,从山东东营刺参(Apostichopusjaponicus)养殖池塘环境中分离到56株乳酸菌。以刺参腐皮综合征2种重要致病菌灿烂弧菌(Vibrio splendidus)和假交替单胞菌(Pseudoalteromonas nigrifaciens)为指示菌,采用牛津杯法测定所分离的乳酸菌对2株致病菌的抑制效果,筛选出1株具有较强抑菌活性的乳酸菌CSND-6,并对该细菌的形态、胞内产物和胞外产物的抑菌活性、生理生化实验、16S r DNA序列分析、菌株生长特性及对刺参安全性进行了研究。结果显示,菌株CSND-6对灿烂弧菌和假交替单胞菌的生长具有较好的抑制作用。其中,胞内产物的抑菌圈分别为17、22 mm,胞外产物的抑菌圈分别为25、31 mm。该菌株对刺参的高浓度浸浴胁迫实验结果表明,所分离的菌株对刺参是安全的。通过生理生化和16S rDNA序列分析表明,CSND-6属于乳杆菌科(Lactobacillaceae),与副干酪乳杆菌(LactobacillusparacaseiJCM1171)的相似性为99.83%。菌株CSND-6在30℃~44℃、pH为6~8范围内生长较快,24 h后进入对数生长期,28~32 h时达到生长高峰,最高值达2.10×10⁹CFU/ml。本研究筛选的副干酪乳杆菌属于刺参池塘养殖环境中土著益生菌,对刺参的主要病原菌抑制效果较好,并表现出了较好的生长特性,可为养殖池塘刺参疾病的生态防控和乳酸菌开发提供参考。     

Microflora of Arctic charr, Salvelinus alpinus (L.): gastrointestinal microflora of free-living fish and effect of diet and salinity on intestinal microflora

Abstract. The adherent aerobic bacterial flora present in the gastrointestinal tract and faeces of free-living Arctic charr, Salvelinus alpinus (L.), from Lake Takvatn, Northern Norway, were identified both qualitatively and quantitatively. Approximately 10⁵ bacteria g⁻¹ were found in both the small and large intestines. The predominant bacterial species were identified as Aeromonas , Enterobacteriaceae, Micrococcus and Lactobacillus. Other microorganisms isolated included Acinetobacter, Cytophaga, Flavobacterium, Moraxella, Pseudomonas, Vibrio , Coryneforms and Streptococcus. The intestinal microflora of free-living fish was dominated by Aeromonas and Lactobacillus , but the intestinal bacterial flora of wild fish transferred to hatchery was affected by feeding them either a capelin roe diet or a commercial feed in fresh and sea water. Approximately 55% of the bacterial flora in intestinal contents from fish fed the capelin roe diet was Enterobacteriaceae when the fish were held in fresh and sea water. However, when the wild-caught charr were fed a commercial diet in fresh water, Aeromonas and Pseudomonas dominated in faeces, while Vibrio and Pseudomonas were predominant in the diet group held in sea water.     

四联活菌制剂对养殖水体中氨氮及亚硝酸盐的降解

利用四联活菌制剂,在室内进行了对养殖池塘水体中氨氮及亚硝酸盐的降解试验.结果表明,光合细菌、纳豆芽孢杆菌、乳酸菌、硝化细菌具有较好的氨氮、亚硝酸盐降解性能,随着添加质量浓度的增加,氨氮、亚硝酸盐的去除率增加;各菌株氨氮降解能力依次为:乳酸菌＞光合细菌＞硝化细菌＞纳豆芽孢杆菌;亚硝酸盐降解能力依次为:硝化细菌＞纳豆芽孢杆菌＞光合细菌＞乳酸菌.四联活菌制剂对养殖水体中氨氮及亚硝酸盐降解试验结果表明,乳酸菌、光合细菌、硝化细菌、纳豆芽孢杆菌的协同作用对氨氮、亚硝酸盐的降解效果更显著、快速.当制剂添加量分别为1.5、3.0、4.5 kg/hm~2时,5 d氨氮的去除率分别为52%、80%、74%,亚硝酸盐的去除率接近100%,结果均显著高于添加同剂量单一菌株时的氨氮、亚硝酸盐的去除率.     

Selection of lactic acid bacteria as candidate probiotics and in vivo test on Artemia nauplii

Lactic acid bacteria (LAB) were isolated from rotifer cultures in a marine hatchery to search for potential probiotics for marine animals. Fifteen strains were first selected among a total of 55, according to antibacterial activity against Vibrio sp. Among eight strains identified as Lactobacillus casei, four were highly adhesive, suggesting some ability for surface colonization. The other strains were identified as Lactobacillus plantarum, Lactobacillus dextrinicus, and Leuconostoc sp. To validate probiotic potential, Artemia were challenged against pathogenic Vibrio alginolyticus, with or without one of six selected LAB strains. The six strains protected Artemia against the pathogen, to some extent on condition that nutrient enrichment was provided. La. casei BR51 and X2 were preferred, as they were efficient even in the absence of nutrient supply. La. casei X2 was finally selected as candidate probiotic, due to the best growth performances of Artemia, with or without the pathogen.     

Isolation,Screening, and Identification of Proteolytic Lactic Acid Bacteria from Indigenous Chao Product

The aim of the study was to isolate, select, and identify proteolytic lactic acid bacteria (LAB) from chao, a traditional fermented fish from Pangkajene and Kepulauan Regency, South Sulawesi, Indonesia. LAB was isolated by poured plate method. Proteolytic LAB were selected using agar skim milk media. Protease activity of LAB was determined based on the amount of tyrosine released in unit/mL. Proteolytic LAB were identified using API 50 CHL kit and 16S rRNA gene sequence analysis. The result showed that a total of 60 isolates were obtained from chao, and 57% of them were cocci-shape. Fifteen isolates were halotolerant proteolytic LAB. Their R values and protease activity were 2.11–3.39 and 0.267–0.304 U/mL, respectively. Identification by API 50 CHL kit showed that four rod-shape isolates were Lactobacillus plantarum, and two others were Lactobacillus curvatus. Cocci-shape isolates could not be identified as cocci bacterium. Rep-PCR results showed that there were two kinds of bands, namely thick and thin. Two isolates were selected from two types of bands that had the highest R for the analysis of 16S rRNA gene sequences, namely Ags1-3 and Ags7-3. The results showed that Ags1-3 isolate was identified as Lactobacillus plantarum and Ags7-3 as Pediococcus acidilactici.     

微生物发酵对罗非鱼下脚料蛋白酶解液脱腥去苦效果比较

以嗜热链球菌(Streptococus thermophilus)、保加利亚乳杆菌(Lactobacillus bulgaricus)和瑞士乳杆菌(L.helveticus)及2种酿酒酵母菌(Saccharomyces cerevisiae)为菌种,对罗非鱼(Oreochromis niloticus)下脚料蛋白酶解液进行发酵,利用感官评定法,比较不同微生物发酵对酶解液脱腥去苦效果.试验结果表明,乳酸菌脱腥效果优于酵母菌,其中以嗜热链球菌脱腥效果最好,其适宜的发酵条件为接种量10%,发酵温度42℃,发酵时间9 h.微生物发酵去苦效果不明显.     

唾液乳杆菌NF19的液体发酵条件研究

从舟山海域海水中筛选到一株海洋细菌NF19,经16S rDNA分子鉴定和API鉴定结果显示菌株NF19为唾液乳杆菌属(L.salivarius)的一个种,将序列上传至NCBI获得GenBank登录号为FJ405036。通过对菌株NF19进行发酵条件优化及正交试验,得到菌株NF19的最佳发酵条件为:初始pH7.2,接种量为8%,在120 r/min下20℃培养48 h。此条件下发酵液有最佳抑菌效果。