Fluorescence localization of epidermal cathepsins L and B in the Japanese eel

Histochemical localization of proteolytic activities in the dorsal epidermis of Japanese eel was demonstrated by fluorescent microscopy utilizing 4-methoxy-2-naphthylamide (4M$\beta$NA) derivatives as substrates and 5-nitrosalicylaldehyde as a trapping agent. Carbobenzoxy-L-phenylalanyl-L-arginyl-4M$\beta$NA (Cbz-Phe-Arg-4M$\beta$NA) and Cbz-Arg-Arg-4M$\beta$NA were used for direct detection of cathepsins L and B activities, respectively, in fresh frozen sections and unfixed cells of the eel epidermis. The fluorescing areas, where Cbz-Phe-Arg-4M$\beta$NA was hydrolyzed by cathepsin L, were shown in mucus secretory cells and club cells and broadly around skin surface. The fluorescing areas due to Cbz-Arg-Arg-4M$\beta$NA hydrolysis by cathepsin B were localized similarly in these tissues. The fluorescing intensity for both catheptic activities in mucus secretory cells was higher than that in club cells, where small fluorescing granules were distributed. These results indicate that eel cathepsins L and B are stored in epidermal secretory cells at different levels and probably serve as defense factors before or after secretion by these cells. Abbreviations: Cbz – carbobenzoxy; 4M$\beta$NA – 4-methoxy-2-naphthylamide; NSA – 5-nitrosalicylaldehyde.     

Fluorescence of Extracts from Argentine Hake (Merluccius hubbsi) Muscle and Its Correlation with Quality Indices During Chilled Storage

Argentine hake (Merluccius hubbsi) was stored chilled for 8 days. The evolution of fluorescence of the aqueous and organic extracts of muscle during storage at 4°C was studied and correlated with common damage indices; pH, total volatile basic nitrogen, free fatty acids, and the fluorescence intensity of the aqueous phase showed a good correlation with storage time. The excitation-emission matrices of the aqueous extracts were also obtained. A fluorescence landscape that could be attributed to tryptophan decreased with storage time, meanwhile a peak (around 340 and 400 nm for excitation and emission wavelengths, respectively) is developed during chilled storage.     

尼罗罗非鱼六个性别相关标记的FISH分析

从尼罗罗非鱼（Oreochromis niloticus）BAC基因文库5个克隆中提取和纯化含有6个性别连锁或相关标记（CLC5，GM204，GM271，GM354，UNH995和UNH104）的重组质粒DNA作为模板，以简并核苷酸为引物，通过PCR制备原位杂交探针。探针用荧光素进行标记，并与尼罗罗非鱼中期相染色体进行荧光原位杂交以确定这些标记在尼罗罗非鱼染色体上的位置和分布。结果显示，这些性别连锁或相关标记都位于尼罗罗非鱼第一对染色体长臂近末端，从分子细胞学角度验证了第一对染色体是尼罗罗非鱼的性染色体。另外由于这些标记的荧光信号在XY个体的2条性染色体上都有，一方面说明这些标记在罗非鱼上还不是性别特异的；另一方面也验证了尼罗罗非鱼的性染色体还处于分化的早期阶段。【中国水产科学，2006，13（4）：525—529】     

半滑舌鳎创伤弧菌分离鉴定及其荧光定量PCR方法的建立

为确定患病半滑舌鳎的病原,从其体内分离到3株革兰氏阴性菌,将其分别命名为ST-1、ST-3和ST-6,进行细菌理化特性、药敏试验、16S rDNA测序以及组织病理学观察等方面的研究,并建立了外膜蛋白(OMP)基因的SYBR Green I荧光定量PCR检测方法。结果显示,3株分离菌株的理化特性符合创伤弧菌特征,16S rDNA与创伤弧菌标准菌株ATCC29307序列相似性达99%以上,系统发育树分析显示,3株分离株与创伤弧菌自然聚为一支,最终鉴定为创伤弧菌;药敏试验显示,3株菌对头孢克肟、头孢哌酮、链霉素、亚胺培南和氟苯尼考等药物高度敏感。人工回归感染试验表明,分离株具有致病性,且半滑舌鳎、斑马鱼的临床症状与自然患病鱼症状相似;组织病理学观察显示,鳃丝上皮细胞增生,鳃小片肿胀黏连、颗粒细胞和黏液细胞增多,肝脏中央静脉淤血,肾脏出血、坏死,肠道黏膜固有层萎缩、黏液细胞增多等。进一步建立OMP基因的荧光定量PCR方法,结果显示,标准曲线的相关系数为0.995,检测下限为1.88×10~2 CFU/mL。以上结果可为半滑舌鳎弧菌病的致病机理、分子诊断和防治提供科学参考。     

营养盐浓度对棕鞭藻叶绿素荧光参数、细胞密度和总脂含量的影响

利用生物化学方法和叶绿素荧光动力学分析技术,研究了棕鞭藻在一次性培养过程中,不同营养盐浓度(f/4、f/2、f、2f、4f)对其叶绿素荧光参数、叶绿素含量、细胞密度、单位体积干质量、总脂含量和总脂收获量的影响。试验结果表明,营养盐浓度为2f时,该藻的最终细胞密度、叶绿素含量、单位体积干质量、总脂含量和总脂收获量均显著高于其他各处理组,由此可以看出,在本试验条件下,最适合棕鞭藻生长和油脂积累的营养盐浓度是2f。不同营养盐浓度对棕鞭藻的叶绿素荧光参数有显著影响,培养6～10d,f/4处理组的叶绿素荧光参数(光系统Ⅱ的最大光能转换效率;光系统Ⅱ的潜在活性;光系统Ⅱ的实际光能转化效率;相对电子传递效率和光化学淬灭)最低,其次为f/2处理组(P0.05)。培养7～10d,2f处理组的光系统Ⅱ的实际光能转化效率、相对电子传递效率和光化学淬灭显著高于其他处理组(P0.05)。培养结束时(第10d),各处理组细胞密度和总脂含量依次为:2f4fff/2f/4。相关性分析结果表明,营养盐浓度为f/4～2f时,棕鞭藻的细胞密度、叶绿素含量、单位体积干质量和总脂含量与营养盐浓度均呈显著的正相关;营养盐浓度为2f～4f时,上述各值与营养盐浓度则均呈显著的负相关。     

Effects of shrimp pond water on phytoplankton: importance of salinity and trophic status of the receiving environment

Aquaculture generates a large load of effluents rich in organic matter and nutrients that may be introduced into the environment. This study aimed to assess in a microcosm experiment, the effect of shrimp pond water mixed with Patos Lagoon estuary water on phytoplankton chlorophyll a and primary production, simulating two salinities. Chlorophyll a, dissolved inorganic nutrients and primary production were measured in two experiments. In Harvest I, salinity of shrimp pond and environment water was similar, and chlorophyll a showed different trends over time, according to the amount of nitrogen available. In Harvest II, with different salinities and high nutrient concentrations in environment water, chlorophyll a levels showed a similar increasing trend over time in all mixtures. Net primary production showed differences among treatments in the first sampling in Harvest I, but not in the second, whereas no differences were observed among treatments in Harvest II. We conclude that shrimp pond effluent can lead to short‐term variations in chlorophyll a and primary production levels, with similar salinities. Salinity differences result in lower chlorophyll a and primary production values than expected according to the nutrient input. Differences in salinity can be an important management strategy to choose the best harvest period.     

黄金梨和鸭梨叶片光合作用的光抑制及其恢复的比较研究

 以黄金梨和鸭梨的连体和离体叶片为研究材料, 在自然光强和模拟光强下测定光合作用和叶绿素荧光参数。结果表明: 黄金梨和鸭梨连体叶片经午间强光处理后, 净光合速率( P_n) 、表观量子效率(AQY) 和光系统Ⅱ ( PSⅡ) 的光化学效率(F_v /F_m ) 降低, 初始荧光( F_o ) 升高, 且其降低或升高的幅度随着光强的增强而加大, 说明强光胁迫使叶片发生了光抑制; 黄金梨和鸭梨离体叶片的光抑制程度均随着强光照射时间的延长而加重, 但是在光强减弱或在弱光下恢复2～4 h, 均可基本恢复到正常值, 说明仅强光引起的光抑制是可逆的; 对DTT处理的叶片进行强光处理, 其F_v/F_m比未吸入DTT的叶片低, F_o比对照高, 说明依靠叶黄素循环进行热耗散是梨树叶片防御强光破坏光合机构的重要途径之一; 黄金梨对强光胁迫的忍耐能力和恢复能力较鸭梨弱, 说明梨树光抑制程度品种间有差异。     

一品红不同品种叶片叶绿素荧光特性比较

 利用叶绿素荧光技术测定了一品红4个品种‘金多利’、‘圣诞天使’、‘索诺拉飞雪’、‘旗帜’叶片光强依赖的叶绿素荧光特性。结果表明: ‘金多利’初始荧光( F_o ) 、最大荧光( F_m ) 和可变荧光( F_v ) 值极显著高于其它3个品种。在各个光强( PAR, 0～2 171μmol·m^{- 2} ·s^{- 1} ) 下, 表观电子传递速率( rETR) 、实际光化学效率(Yield) 和光化学淬灭( q_P ) 的变化趋势一致, 均为‘金多利’ > ‘圣诞天使’ > ‘索诺拉飞雪’ > ‘旗帜’, 在高PAR下这种差异更明显。拟合参数最大潜在相对电子传递速率( rETR_max ) 、半饱和光强( Ik) 和光抑制参数(β) 也同rETR等具有一致变化。说明‘金多利’有较高的PSⅡ活性, 耐强光; 而‘旗帜’则光化学效率较弱, 高光强下易受光抑制。     

茄子离体叶片光系统Ⅱ的冷激胁迫效应及热力学分析

 以耐冷性较强的短把黑圆茄和耐热性较强的黑帅圆茄为试材,经冷激胁迫后采用植物效率仪PEA,进行快速叶绿素荧光参数测定,并利用最大光化学效率Fv/F m和活性中心RC/CSo的标准状态变性自由能变△GD进行PSⅡ冷稳定性的热力学分析。随着冷激胁迫温度的降低和时间的延长,PSⅡ的Fv/Fm、实际光化学效率ΔF/Fm′、RC/CSo呈"S"型下降趋势;单位反应中心DIo/RC和非光化学猝灭qN呈"S"型上升趋势;综合分析反映出冷激胁迫下PSⅡ反应中心的可逆失活和依赖于叶黄素循环的热耗散的双重机制在保护PSⅡ防止光抑制中起到重要作用。另外,通过两品种间PSⅡ行为差异,引入了两组新的参数作为便于直观比较冷胁迫的评价指标。黑帅圆茄Fv/Fm、ΔF/Fm＇的半衰温度T1/2和t1/2值分别高于短把黑圆茄1.4 ℃和0.8 ℃。茄子叶片Fv/Fm和RC/CSo的△GD随着冷激胁迫温度的降低而减少呈瀑布型的下降趋势。黑帅圆茄的△GD值和变性中点温度Tm都低于短把黑圆茄。它们均反映出黑帅圆茄的耐冷性低于短把黑圆茄。     

Evaluating foliar nitrogen compounds as indicators of nitrogen status in Prunus persica trees

Mobile nitrogen (N) forms may be better N indicators of the N status of trees than total nitrogen (TN) due to their higher sensitivity to increasing N supply. A field experiment was carried out over a 3-year period to compare foliar concentrations of total N (TN), soluble N (SN), chlorophyll (Minolta SPAD readings), NH₄–N and NO₃–N as indicators of soil N availability in nectarine, Prunus persica L. Batsch, cv. ‘Fantasia’ (grafted on ‘Nemaguard’ peach, P. persica × P. davidiana) trees. Young trees were exposed to a range of fertilizer-N application rates. Based on correlation analysis, the best association between leaf N compounds with soil N supply and trunk diameter and/or fruit yield was obtained with TN and chlorophyll SPAD readings. Leaf concentrations of mobile N compounds (NH₄–N and NO₃–N) increased more than any other N compound under high N supply; however, their inconsistency among years and low leaf concentration difficult their use as N indicators. The optimum foliar TN for growth decreased with tree age, 4.4%, 3.6% and 3.3% in non-bearing 1-year-old trees, non-bearing 2-year-old trees and 3.3 fruit-bearing 3-year-old trees. The optimum SPAD readings were 40 in 2-year-old trees and 42 in 3-year-old trees. Stable N compounds (TN and chlorophyll SPAD) could be used to N diagnosis in the zone of N deficiency, and soluble N compounds (NH₄–N and NO₃–N) to diagnoses N excess.