罗非鱼特定腐败菌生长动力学模型和货架期预测

以有氧冷藏养殖罗非鱼为研究对象,建立和验证了用于预测冷藏罗非鱼微生物学质量和剩余货架期的特定腐败菌生长动力学模型。感官、总挥发性盐基氮(TVBN)评价和微生物生长动态分析表明,罗非鱼在0～15℃贮藏中的特定腐败菌为假单胞菌,在0、5、10、15℃,感官货架期终点的假单胞菌数平均值为7.70±0.11log10cfu·g-1。假单胞菌在0、5、10、15℃的生长实验值用于建立生长动力学模型表明,Gompertz方程能很好地描述假单胞菌在0～15℃温度区域的生长动态。Nmax(最大菌数)受贮藏温度的影响不大,在4种温度下平均值在8.85±0.18log10cfu·g-1。温度对μmax(最大比生长速率)和Lag(延滞时间)的影响,采用Belehradek方程描述呈现良好线性关系。用贮藏在3、8℃假单胞菌的生长实验值,验证建立的模型,偏差度为0.97～1.01,准确度为1.02～1.04,货架期预测值和实测值的相对误差分别为3.47%和-7.91%,显示建立的模型可以快速可靠地实时预测0～15℃贮藏罗非鱼的微生物学质量和剩余货架期。     

大菱鲆腐败菌生长动力学研究和货架期预测

采集0、3、7、10℃贮藏大菱鲆中希瓦氏菌生长数据,建立了大菱鲆0~10℃贮藏中特定腐败菌生长动力学模型和剩余货架期预测模型。感官、化学和微生物生长动态分析表明,希瓦氏菌为大菱鲆0~10℃贮藏中的特定腐败菌。Gompertz方程能较好地描述希瓦氏菌的生长。采用Belehradek方程描述的温度对希瓦氏菌最大比生长速率和延滞时间的影响均呈较好线性关系。以5℃贮藏大菱鲆中希瓦氏菌生长数据,对所建立的生长动力学模型和剩余货架期预测模型的准确性进行验证,得到最大比生长速率、延滞时间、货架期的预测值相比实测值的相对误差分别为-14.36%、5.61%和6.74%,表明建立的模型可以快速准确的预测大菱鲆0~10℃贮藏过程中的鲜度变化和剩余货架期。     

冷却链大黄鱼货架期预报系统的开发与评估

通过使用Visual Basic编程语言,完成了基于0~10℃腐败希瓦氏菌生长动力学模型建立的冷却链大黄鱼货架期预报系统的设计,验证了货架期预报系统的准确性。采用以自然污染鱼得到的实验数据建立特定腐败菌生长动力学模型的方法,提高了预测的准确性,腐败希瓦氏菌和剩余货架期预测值与实测值的可靠性评估表明,其相关系数R2分别为0.936和0.935,显示开发的冷却链大黄鱼货架期预报系统可以快速可靠地预测冰鲜大黄鱼的鲜度与剩余高品质期和货架期。系统开发过程中采用免安装处理,方便用户使用。     

大黄鱼冷藏过程中的鲜度变化

研究了大黄鱼(Pseudosciaena crocea)贮藏于0℃(冰藏)、3℃、7℃和10℃过程中,在感官、化学和微生物品质方面的变化特性,并对其货架期进行分析,探讨了菌落总数、嗜冷菌、假单胞菌、产H2S菌、TVBN和TMA与感官评价的一致程度.结果表明,大黄鱼是海水鱼中鲜度下降较慢的种类,在O℃、3℃、7℃、10℃的贮藏过程中,大黄鱼的高品质期分别为330 h、208 h、114 h和87 h,货架期分别为523 h、368 h、197 h和150 h.各温度高品质期终点和货架期终点时菌落总数(CFU)/g的对数平均值分别为6.10±0.30和6.67±0.35,产H2S菌数(CFU/g)的对数平均值分别为5.82±0.33和6.37±0.22,TVBN均值分别为(14.77±0.5)mg/(100 g)和(28.02±1.19)mg/(100 g);TMA均值分别为(2.55+0.62)mg/(100 g)和(9.84±1.28)mg/(100 g),各温度下高品质期终点和货架期终点时各指标均值均无显著差异(P>0.05),表明菌落总数、产H2S菌数和TVBN值作为大黄鱼低温贮藏的鲜度指标与感官鲜度评价有较好的一致性.     

鱼类腐败菌不同生长动力学温度模型的可靠性评价和比较

探讨和比较了Arrhenius方程和Belehradek方程两种腐败菌生长动力学温度模型预测鱼鲜度和货架期的可靠性。结果表明，两种温度模型都能很好地预测罗非鱼中假单胞菌的生长动态和预报货架期，通过假单胞菌生长动力学模型的预测值，与罗非鱼中假单胞菌生长的实测值比较，偏差度在0．95～1．01之间，准确度在1．02～1．06之间，两种温度模型相比，Belehradek温度模型的可靠性更好。罗非鱼在3和8℃贮藏中货架期实测值与利用Arrhenius方程和Belehradek方程建立的假单胞菌生长动力学温度模型求得的预测值的比较，Arrhenius温度模型预测值和实测值的相对误差分别为11．8％和8．00％，Belehradek温度模型预测值和实测值的相对误差分别为3．47％和一7．91％，两种温度模型相比，Belehradek温度模型的结果更为理想。     

鲤鱼冷藏中的鲜度变化与货架期

以感官、化学和微生物为指标,研究了养殖鲤鱼在0℃(冰藏)、5℃、10℃和15℃贮藏过程中的鲜度变化和货架期,并对菌落总数、嗜冷菌、假单胞菌、挥发性盐基氮与感官评价的一致性程度进行了探讨。结果表明,在0℃、5℃、10℃、15℃的贮藏过程中,鲤鱼的高品质期分别为426 h、189 h、95 h和58 h,货架期分别为713 h、382 h、146 h和87 h。各温度高品质期终点和货架期终点时菌落总数分别为6.63±0.45 lg cfu/g、7.18±0.29 lg cfu/g,假单胞菌数分别为5.69±0.75 lg cfu/g、6.50±0.33 lg cfu/g,挥发性盐基氮均值分别为9.90±0.52 mg/100 g、20.52±0.34 mg/100 g。各温度下高品质期终点和货架期终点时各指标均值均无显著差异(P0.05),表明菌落总数、假单胞菌和挥发性盐基氮作为鲤鱼低温贮藏的鲜度指标与感官鲜度评价有较好的一致性。     

冷藏养殖大黄鱼指数腐败货架期模型的构建与评价

通过感官、化学、微生物学分析对0~10℃冷藏养殖大黄鱼鲜度和货架期进行研究,构建和验证了指数腐败货架期模型。结果表明,0~10℃冷藏养殖大黄鱼货架期终点时菌落总数、假单胞菌数、嗜冷菌数和产H2S细菌数分别为6.64~7.60、6.24~6.96、6.16~6.90、6.14~6.62 lg cfu/g,挥发性盐基氮和三甲胺分别为27.15~30.12 mg/100g和8.44~10.83 mg/100g。0、5、8和10℃冷藏大黄鱼的货架期分别为17.8±2.5、9.3±1.1、7.0和5.4±1.3 d,并用于构建指数腐败货架期模型。用0、3、7℃冷藏养殖大黄鱼货架期验证指数腐败货架期模型,相对误差为?6.1%~4.6%,显示该模型可以快速有效预测0~10℃冷藏养殖大黄鱼的剩余货架期。     

罗非鱼冷藏过程细菌种群的变化

对养殖尼罗罗非鱼(Oreochromis niloticus)0～10℃贮藏过程中感官、化学、微生物品质和细菌种群消长研究表明,初始样品挥发性盐基氮为(8.08±0.29)mg/(100g),菌落总数(CFU/g)的对数值为4.79±0.60;细菌种群复杂,种类繁多,分离到570株菌,其中80.5%为革兰氏阴性菌,13.6%为革兰氏阳性菌,优势菌是假单胞菌(Pseudomonas spp.)(36.5%)、肠杆菌科细菌(Enterobacteriaceae)(14.2%)和气单胞菌(Aeromonas spp.)(15.3%),同时检出了一定比例的不动杆菌和其他细菌。冷藏过程中肠杆菌科细菌、气单胞菌等生长受到抑制,细菌菌相组成逐渐变得单一,适应低温环境下革兰氏阴性菌比例不断增加,0℃、5℃、10℃贮藏至货架期终点时,阴性菌的比例分别达到86.4%、88.9%、100.0%;假单胞菌增殖显著,比例分别达到80.7%、68.1%、60.0%,可确认其为0～10℃贮藏罗非鱼的优势腐败菌。假单胞菌的种类变化趋势显示,优势顺序分别为荧光假单胞菌(P.uorescens)、恶臭假单胞菌(P.putida)、铜绿假单胞菌(P.aeruginosa)、产碱假单胞菌(P.alcaligenes),0℃、5℃、10℃贮藏至货架期终点时,假单胞菌中荧光假单胞菌分别占52.1%、53.1%、59.3%,可确认其为最优势种群。     

养殖大黄鱼冷藏过程中细菌菌相的变化

采用感官、挥发性盐基氮(TVBN)、菌落总数对大黄鱼(Pseudosciaena crocea)在0℃、5℃冷藏过程中的品质变化特征进行分析,并对细菌菌相进行定性和定量研究。结果表明,冷藏初期、高品质期和货架期终点菌落总数N(CFU/g)的对数值(lgN)分别为5.40±0.17、6.98±0.17、7.38±0.09,TVBN分别为(7.00±1.82)mg.100-1.g-1、(13.00±1.42)mg.100-1.g-1、(29.92±1.75)mg.100-1.g-1。冷藏初期分离获得211株菌株,84.8%是革兰氏阴性菌,出现少量革兰氏阳性菌(6.2%),优势菌群是肠杆菌科细菌(6.6%)、气单胞菌属(14.2%)、不动杆菌属(13.3%)、摩氏杆菌属(11.8%),并出现了一定比例的假单胞菌属、嗜麦芽窄食单胞菌和其他细菌。冷藏过程中细菌菌相逐渐变得单一,腐败希瓦氏菌上升趋势明显。高品质期时,0℃冷藏大黄鱼优势菌群为腐败希瓦氏菌(45.8%)和缺陷短波胞单胞菌(13.6%);5℃冷藏大黄鱼优势菌群为腐败希瓦氏菌(37.9%)和假单胞菌属(15.6%)。货架期终点时,0℃、5℃冷藏大黄鱼优势菌为腐败希瓦氏菌,比例分别为75.5%和59.6%。     

冰鲜大黄鱼加工过程中的细菌学分析及关键控制点

探讨了冰鲜大黄鱼加工过程中鱼体温度变化及细菌总数和主要腐败菌假单胞菌的变化,分析了加工中的关键控制点。活鱼、加工结束后中心温度超过8℃的鱼的细菌总数、假单胞菌数分别为5.5×104、4.5×104cfu/g和3.3×104、1.4×103cfu/g;加工结束后中心温度未超过8℃的鱼,细菌总数和假单胞菌数分别为9.6×103、3.8×103cfu/g;冰藏2d、5d后鱼的细菌总数、假单胞菌数分别为1.1×104、8.6×103cfu/g和2.4×103、5.6×103cfu/g。并从细菌学角度确定了原料鱼、冰、预冷却、运输、加工是冰鲜大黄鱼加工过程中的关键控制点。     

Rare serotype occurrence and PFGE genotypic diversity of Streptococcus agalactiae isolated from tilapia in China

Previously, we reported 10 PEGE types of 85 tilapia Streptococcus agalactiae(GBS), which shifted from Streptococcus iniae in China, by using PEGE method. Presently, larger and more representative tilapia GBS were isolated, for the ?rst time in China, to characterize their serotypes and genetic diversities more precisely than had done before. 168 GBS strains were distributed in ?ve provinces of China, in which Guangdong, Guangxi and Hainan were the major ones, holding36.9%(62/168), 37.5%(63/168) and 19.6%(33/168), respectively. Serotypes, Ia, Ib and III, were observed in these strains and the most predominant one was Ia(95.2%), which mainly distributed in Guangdong, Guangxi and Hainan. Ia initially occurred in 2009, it shoot up to 32.1% in 2010,but decreased to 16.1% in 2011 before went up to 45.2% in 2012. Ib sporadically occurred during2007–2011, III onlyoccurred in 2012. 14 different PFGE types, including 4 new types(N, O,P and Q), were observed, in which B, D, F and G were the predominant types, holding 83.9%(141/168) of the total GBS strains. Ia corresponded to 11 PFGE types(A–H, N–P), in which type D predominated(51%). Ib represented 3 genotypes(I, J and Q) and III harbored only 2genotypes(N and F). Type N and Fsynchronously presented in Ia and III. In summary, the genetic diversity of tilapia GBS varied by serotypes and changed with geographical locations and years.Although Iastillpredominated, new rareserotypeIII alreadyoccurred in China.     

Growth hormone (GH) and reproduction: a review

Interaction between growth and reproduction occurs in many vertebrates and is particularly obvious at certain stages of the life cycle in fish. Endocrine interactions between the gonadotropic axis and the somatotropic axis are described, the potential role of GH being emphasised. A comparative analysis of these phenomena in mammals, amphibians and fish, suggests a specific role of GH in the physiology of puberty, gametogenesis and fertility. It also shows the original contribution made by studies on the fish model in this field of investigations.     

Purification and partial characterization of GtHs (cfLH and cfFSH) from Indian walking catfish (Clarias batrachus) (L.) and development of a homologous ELISA for cfLH

Two gonadotropins (GtH; Qa and Qb) were purified by gel filtration and ion exchange chromatography from the pituitaries of Indian walking catfish (Clarias batrachus). The presence of GtH during purification was assessed by in vitro oocyte maturation and in vivo steroidogenic activity, and their identities were determined by elution profiles, molecular weight, biological activities and yield. The molecular weights of Qa and Qb were 37 and 42 kDa, respectively, and composed of distinct subunits (Qa: 20 and 14 kDa and Qb: 26 and 18 kDa). Polyclonal antibodies raised against Qa immunostained Qa, Qb and pituitary GtH cells. A competitive Qa‐ELISA was developed whose sensitivity was 6.25 ng mL^?1 (1.25 ng well^?1) with intra‐ (3.5%) and inter‐ (12.4%) assay coefficients of variation. Displacement curves parallel to the standard were obtained with plasma and pituitary extracts of catfish, Qb and carp GtHII. The assay was validated by measuring the plasma Qa levels after LHRH treatment and in relation to ovarian growth in the female catfish during different reproductive phases. Based on the results, Qa and Qb corresponded to fish LH and FSH respectively. The findings will increase the knowledge of the mechanisms controlling fish reproduction and identification of sensitive phases in fish in captivity for hormonal manipulation.     

Controlled infection of Poecilia reticulata Peters (guppy) with Tetrahymena by immersion and intraperitoneal injection

Tetrahymena is a protozoan parasite, which infects guppy, Poecilia reticulata Peters, and causes substantial economical losses in commercial farms worldwide. Studies of guppy infected by Tetrahymena require standardized infection protocols. The LD50 for Tetrahymena infection of guppies by intraperitoneal (IP) injection was calibrated, and the level obtained was 946 parasites per fish. Guppy infection with Tetrahymena by immersion, imitating the natural route of infection via the integument, was studied under normal or stress conditions. Exposure to cold and netting (CNI) and to cold only (CI) followed by immersion exposure to 10 000 Tetrahymena per mL resulted in 22.5% and 19.2% mortality, respectively, as compared to 14.2% and 10% in groups that were netted only (NI) or non‐stressed (I). Histopathology revealed that immersion infection resulted in a systemic infection. Lysozyme levels, measured 3 weeks after infection, were significantly higher in the CNI group (288 μg per mg protein) compared with CI‐, NI‐ and I‐treated groups (94.5, 64 and 62.3 μg mg^?1, respectively). There was no evident parasite immobilization activity in body homogenates, suggesting no development of acquired immunity. Re‐infection by IP injection revealed no increase in protection in any of the treatment groups, mortality range of 56.3–75%, higher than in the non‐exposed control (40.6% mortality).     

Evaluation of a Fast Method Based on the Presence of Two Restriction Sites in the Mitochondrial ND5 (mt ND5) Gene for the Identification of Scomber Species

The purpose of this work was to evaluate the suitability of a method based on the presence of two restriction sites (for Hae III and Hindf I) in the mitochondrial NADH dehydrogenase subunit 5 (mt ND5) gene to identify Scomber species. The evaluation was performed on 144 reference and market samples by sequencing of the entire 505-bp fragment of the mt ND5 gene and of a 464-bp fragment of the Kocher fragment of the cytochrome b gene (mt Cytb). Sequence analysis of any of the two fragments allows the identification of each of the four Scomber species, but S. japonicus and S. colias had the same restriction sites at the ND5 amplicon and would not have been differentiated by this analysis. Similarly, loss of the Hae III site in some S. scombrus individuals would have misidentified them as not being Scomber. All the market products were correctly labeled except one acquired in Spain labeled as originating in the Atlantic and containing S. japonicus.     

Imported ornamental fish are colonized with antibiotic‐resistant bacteria

There has been growing concern about the overuse of antibiotics in the ornamental fish industry and its possible effect on the increasing drug resistance in both commensal and pathogenic organisms in these fish. The aim of this study was to carry out an assessment of the diversity of bacteria, including pathogens, in ornamental fish species imported into North America and to assess their antibiotic resistance. Kidney samples were collected from 32 freshwater ornamental fish of various species, which arrived to an importing facility in Portland, Oregon from Colombia, Singapore and Florida. Sixty‐four unique bacterial colonies were isolated and identified by PCR using bacterial 16S primers and DNA sequencing. Multiple isolates were identified as bacteria with potential to cause disease in both fish and humans. The antibiotic resistance profile of each isolate was performed for nine different antibiotics. Among them, cefotaxime (16% resistance among isolates) was the antibiotic associated with more activity, while the least active was tetracycline (77% resistant). Knowing information about the diversity of bacteria in imported ornamental fish, as well as the resistance profiles for the bacteria will be useful in more effectively treating clinical infected fish, and also potential zoonoses in the future.     

Effect of iodophor disinfection of non‐hardened Salmo trutta eggs on their bacterial and fungus load

This study investigated the efficiency of iodophor disinfection (135 ppm active iodine for 15–30 min) of non‐hardened Salmo trutta eggs against different groups of bacteria and against fungus. Egg samples were taken from non‐disinfected and from disinfected eggs, microorganisms were cultured on specific nutrient media and their mass was measured by turbidimetric methods. Bacteria and fungus mass of non‐hardened eggs could be reduced but not eliminated by iodophor disinfection with 135 ppm active iodine for 15 min. The extent of reduction was 47–65% (Experiment 1). The efficiency of disinfection increased with disinfection time as the reduction in bacteria and fungus mass was 40–55% after 15 min and 58–74% after 30 min (Experiment 2). Disinfection efficiency of iodophor solution diluted in water (reduction 49–57%) and of iodophor solution diluted in sodium chloride solution iso‐osmolar to the oocytes (reduction 52–61%) was similar (Experiment 3). The reduction in bacteria and fungus mass was persistent as it was 39–72% lower in embryos deriving from disinfected eggs than in embryos deriving from non‐disinfected ones (Experiment 4). In conclusion, the tested disinfection method is inadequate to eliminate pathogens completely but it could positively influence immune defence of eggs and embryos.     

Nitrogenous waste excretion and accumulation of urea and ammonia inChalcalburnus tarichi (Cyprinidae), endemic to the extremely alkaline Lake Van (Eastern Turkey)

The endemic, anadromous cyprinidChalcalburnus tarichi is the only fish species known to occur in alkaline Lake Van (Eastern Anatolia, Turkey). EightC. tarichi were maintained individually in Lake Van water (17 – 19°C; pH 9.8; 153 mEq·I^–1 total alkalinity; 22 total salinity) and tank water samples analyzed for 24 h in 2 to 4 h intervals. At zero time, < 1µM ammonia was present and urea was undetectable in the tank water; at 24 h, total ammonia and urea made up 114±32 and 35±25µM, respectively. Over the experimental period, ammonia-N and urea-N excretion averaged 1041±494 and 607±169moles·kg^–1 fish·h^–1, respectively. The extent of urea excretion was highly variable between specimens. Uric acid excretion was not detectable.Urea was present at high concentrations in all tissues and plasma (25 – 35moles·g^–1·ml^–1) of freshly caughtC. tarichi; total ammonia content of the tissues was by a factor of 1.9 (liver) to 3.0 (brain) lower. High arginase activity (2.4±0.2 U·min^–1·g^–1) was detected in the liver ofC. tarichi but ornithine carbamoylphosphate transferase, a key enzyme of the ornithine-urea-cycle, was absent. Ureagenesis is likely through degradation of arginine and/or uricolysis. High glutamine synthetase activity (11±0.6 U·min^–1·g^–1) and low ammonia content in brain suggest that, like other teleosts,C. tarichi has an efficient ammonia detoxification in the brain, but in no other tissue.Nitrogenous waste excretion at alkaline pH is discussed. The ability ofC. tarichi to excrete high levels of ammonia at extremely alkaline pH is unique among teleosts studied so far. The mechanism of ammonia excretion under Lake Van conditions remains to be elucidated.