红蓝光光强对多年生黑麦草生长产量与品质的影响
English
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受传统粮食观影响，牧草产业一直作为草食畜牧业的附属产业未受到重视[1]。随着草食畜牧业快速发展，优质牧草产品已不能满足需求因而大量进口[2]。当前，居民膳食消费水平与结构正在由温饱型向全面小康型转变，表现为由粮菜型向粮肉菜果多元型转变[3]，对草食动物肉奶制品刚性需求持续增加。国务院《关于促进畜牧业高质量发展的意见》指出，到2025年我国牛羊肉自给率保持在85%左右，奶源自给率保持在70%以上。为此，要健全饲草料供给体系，提高紧缺饲草自给率。设施牧草能通过环境控制技术，实现牧草周年和反季节生产，为解决牧草缺口问题提供支持[4]。其中，人工光植物工厂是牧草生产的重要设施，具有可周年生产，立体多层栽培等诸多优势[4-5]。Asseng等[6]研究表明，在1 hm2的10层垂直农场中小麦(Triticum aestivum)产量可达到世界平均产量(3.2 t·hm−2)的220～600倍。使用人工光植物工厂不仅能在有限的土地上生产出更多的产品，同时还能为特定区域稳定供应，可解决牧草种植区和养殖密集区远距离运输的问题[7-8]。
合理的光照是保证人工光植物工厂优质高产的重要环境因素。光照可调控植物的生长和发育[9]，包括光强、光质和光周期等属性[10]。确定适宜的光强可以促进光合作用，有利于生物量的积累[11]，节省电能投入。研究表明，光强通过调节植物中叶绿素、可溶性糖和可溶性蛋白等的含量影响植物的品质，当光强为100～800 μmol·(m2·s)−1时，生菜(Lactuca sativa)的叶绿素含量随着光强的升高而降低[12]。在薄荷(Mentha arvensis)叶片中可溶性糖含量随着光强的增加而升高[13]。番茄(Lycopersicon esculentum)幼苗在250、300和350 μmol·(m2·s)−1的光强下，虽然可溶性蛋白含量没有显著差异，但300 μmol·(m2·s)−1时的游离氨基酸含量较高[14]。此外，光强也会影响丙二醛(malondialdehyde, MDA)含量和抗氧化酶活性等植物生理特性[14-15]。当前，有关人工光植物工厂牧草栽培光强调控效应的研究较少，适宜的光强参数需要逐种类探明。为此，本研究以多年生黑麦草(Lolium perenne)为材料，在人工光植物工厂可控环境下，探究光强对黑麦草生长、产量和品质的影响，以期为黑麦草生产的光照环境调控提供科学依据。
1. 材料与方法
1.1 试验材料与方法
黑麦草(宽叶四倍体黑麦草)购于市场，使用 54 cm × 28 cm的32孔育苗穴盘播种，在中国农业科学院农业环境与可持续发展研究所试验用人工光植物工厂中进行栽培。该工厂内温度为(24 ± 2) ℃，相对湿度为35%～50%，CO2浓度为(500 ± 50) mL·m−3。选用规格为49 cm × 49 cm红蓝光组合面板灯给予光照，面板灯安装在穴盘上方40 cm处。参考陈艳琦和刘文科[16]的方法制备基质，将基质均匀装于穴盘中，每穴播种5粒大小均一、颗粒饱满的种子。
1.2 试验设计
LED红蓝光组合灯中红灯(600～700 nm)波峰为655 nm，蓝灯(400～500 nm)波峰为437 nm，红蓝比为4 ꞉ 1。试验共设150、250、350、450和550 μmol·(m2·s)−15个光强处理，分别标记为L1、L2、L3、L4和L5，光周期16 h，光照时段为08:00－24:00。于2021年11月16日播种，播种后避光培养，11月21日给予光照，光照培养期间向育苗托盘补充Hoagland营养液[17]。当黑麦草平均高度达到灯板高度时进行刈割，分别于12月14日、12月31日刈割，留茬高度(2.5 ± 0.5) cm。
1.3 指标测定及方法
刈割时随机选取4穴植株，统计每穴平均单株分蘖数。分别使用游标卡尺和直尺测量茎粗和株高，称量鲜重，后分装至信封，置于70 ℃烘干至恒重，称量干重，计算鲜干比。另取4穴植株用锡箔纸包裹，迅速液氮处理后保存–80 ℃冰箱中。
取新鲜叶片0.1 g剪碎，置于10 mL离心管中，浸入5 mL的95%乙醇溶液，避光室温浸泡48 h，于UV-1800紫外分光光度计测定吸光度，计算叶绿素含量[18]，公式如下：
$ 叶绿素\text{a}=\left (13.95\times {D}_{665}-6.88\times {D}_{649}\right)\times 0.05 \text{；} $
$ 叶绿素\text{b}=\left (24.96\times {D}_{649}-7.32\times {D}_{665}\right)\times 0.05 \text{；} $
$ 总叶绿素=\left (18.08\times {D}_{649}-6.63\times {D}_{665}\right)\times 0.05 。 $
式中：D649和D665分别为叶绿素提取液在649和665 nm处的吸光度值。
取−80 ℃保存样品研碎后，称取0.1 g样品，使用无水乙醇提取，参考 Çoruh等[19]的公式计算DPPH自由基清除率，公式如下：
$ \text{DPPH}自由基清除率 = \left[\left ({A}_{空白}-{A}_{样品}\right) \div {A}_{空白}\right]\times 100{\text{%}} 。 $
式中：A空白和A样品分别为空白和样品在517 nm处的吸光度值。
可溶性糖、可溶性蛋白、氨基酸、丙二醛(MDA)和半胱氨酸含量采用试剂盒测定，试剂盒购于北京索莱宝科技有限公司。
1.4 数据分析
使用SPSS 23.0软件进行单因素方差分析(One-way ANOVA)和LSD差异显著性检验(P < 0.05)，采用Pearson相关进行相关性分析；采用GraphPad Prism 8软件制图。
2. 结果与分析
2.1 LED红蓝光光强对黑麦草农艺性状和产量的影响
5种LED红蓝光光强下两次刈割黑麦草外观形态如图所示(图1)，不同处理下株高、茎粗、单株分蘖数、产量以及鲜干比如表1、表2所列。黑麦草第1次刈割5种光强处理下的株高和茎粗均无显著差异(P > 0.05)，第2次刈割时在高光强L5处理下的株高低于低光强L1处理，但差异不显著(P > 0.05)，L1和L5处理下茎粗分别为最低和最高，且两处理间存在显著差异(P < 0.05)，两次刈割中，第2次刈割时茎粗和光强呈极显著正相关关系(P < 0.01, r = 0.598)。第1次刈割L4和L5处理下单株分蘖数显著高于其他处理(P < 0.05)，而第2次刈割时除低光强L1处理显著低于其他处理(P < 0.05)外，其他处理间无显著差异(P > 0.05)，且两次刈割的单株分蘖数均与光强呈极显著正相关关系(P < 0.01, r = 0.890和r = 0.719)。产量方面，第1次刈割各处理间鲜草产量差异显著(P < 0.05)，第2次刈割L3、L4和L5处理差异不显著(P > 0.05)；第1次刈割L4和L5处理的干草产量显著高于其他处理(P < 0.05)，第2次刈割干草产量与鲜草产量表现一致。低光强L1处理两次刈割的鲜干比较高，且第1次刈割时显著高于其他处理(P < 0.05)。鲜草产量与光强极显著正相关(P < 0.01, r = 0.980和r = 0.713)，干草产量与光强也呈极显著正相关关系(P < 0.01, r = 0.947和r = 0.739)，而鲜干比随着光强的增加而降低，二者呈极显著负相关关系(P < 0.01, r = −0.661和r = −0.696)。
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图 1 5种光强下两次刈割黑麦草外观形态Figure 1. Morphology of ryegrass grown under five light intensities for twice cuttings表 1 5种光强下第1次刈割黑麦草农艺性状和产量的差异Table 1. Differences in the agronomic traits and yield of ryegrass under five light intensities at the first cutting处理
Treatment株高
Plant height/cm茎粗
Stem diameter/mm单株分蘖数
Tillers per plant穴鲜草产量
Fresh yield/(g·hole−1)穴干草产量
Hay yield/(g·hole−1)鲜干比
Fresh to dry ratioL1 44.68 ± 2.35a 2.61 ± 0.11a 4.05 ± 0.34c 11.31 ± 0.75e 0.99 ± 0.08d 11.43 ± 0.38a L2 46.03 ± 3.93a 2.64 ± 0.26a 5.95 ± 0.72b 17.54 ± 1.09d 1.72 ± 0.22c 10.24 ± 0.59b L3 45.66 ± 2.89a 2.79 ± 0.29a 6.55 ± 0.10b 22.41 ± 0.69c 2.53 ± 0.25b 8.90 ± 0.64c L4 47.71 ± 1.36a 2.71 ± 0.11a 7.60 ± 0.37a 28.14 ± 1.24b 3.09 ± 0.23a 9.14 ± 0.69c L5 46.04 ± 4.35a 2.80 ± 0.22a 7.70 ± 0.93a 29.93 ± 0.87a 3.18 ± 0.26a 9.46 ± 0.86bc 同列不同小写字母表示不同光强处理间差异显著(P < 0.05)；下同。
Different lowercase letters within the same column indicate significant differences between different light intensities at the 0.05 level. This is applicable for the following tables and figures as well.表 2 5种光强下第2次刈割黑麦草农艺性状和产量的差异Table 2. Differences in the agronomic traits and yield of ryegrass under five light intensities at the second cutting处理
Treatment株高
Plant height/cm茎粗
Stem diameter/mm单株分蘖数
Tillers per plant穴鲜草产量
Fresh yield/(g·hole−1)穴干草产量
Hay yield/(g·hole−1)鲜干比
Fresh to dry ratioL1 46.51 ± 4.94bc 2.29 ± 0.14b 5.25 ± 0.50b 16.20 ± 3.16c 1.39 ± 0.34c 11.82 ± 1.15a L2 51.12 ± 2.25a 2.42 ± 0.14ab 7.80 ± 1.01a 27.00 ± 6.58b 2.84 ± 0.98b 9.95 ± 1.94ab L3 50.47 ± 1.24ab 2.48 ± 0.19ab 9.30 ± 1.60a 34.76 ± 6.74ab 4.07 ± 1.28ab 8.83 ± 1.24b L4 47.66 ± 2.91abc 2.42 ± 0.14ab 9.40 ± 0.94a 37.67 ± 6.84a 4.43 ± 0.74a 8.54 ± 1.19b L5 43.31 ± 2.40c 2.65 ± 0.21a 9.10 ± 1.00a 33.85 ± 3.80ab 4.20 ± 0.87a 8.21 ± 0.97b 2.2 LED红蓝光光强对黑麦草叶绿素含量的影响
5种LED红蓝光光强下两次刈割黑麦草叶片的叶绿素含量如图2所示。黑麦草第1次刈割5种光强处理下总叶绿素含量和叶绿素a含量表现一致，L4和L5处理含量较低，且显著低于其他处理(P < 0.05)，其中L5叶绿素b含量显著低于L1、L2和L3处理(P < 0.05)。第2次刈割总叶绿素含量除L1处理外，其他处理间无显著差异(P > 0.05)，且叶绿素a含量各处理间也无显著差异(P > 0.05)，而L5的叶绿素b含量显著高于L1和L2处理(P < 0.05)，L3、L4和L5处理间差异不显著(P > 0.05)。两次刈割中，第1次刈割总叶绿素、叶绿素a和叶绿素b含量均与光强呈极显著负相关关系(P < 0.01, r = −0.597, r = −0.597和r = −0.585)，而第2次刈割仅叶绿素b含量与光强呈极显著正相关关系(P < 0.01, r = 0.584)。
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图 2 5种光强下黑麦草叶片叶绿素含量的差异Figure 2. Differences in the chlorophyll content of ryegrass leaves under five light intensities2.3 LED红蓝光光强对黑麦草可溶性糖、可溶性蛋白含量的影响
5种LED红蓝光光强下两次刈割黑麦草的可溶性糖和可溶性蛋白含量如图3所示。5种光强下可溶性糖含量介于7.21～13.29 mg·g−1，其中第1次刈割L4含量最高，为13.29 mg·g−1，两次刈割中L3、L4和L5处理的可溶性糖含量均无显著差异(P > 0.05)，第2次刈割L5含量最高，显著高于L1和L2处理(P < 0.05)，且两次刈割中可溶性糖含量与光强极显著正相关关系(P < 0.01, r = 0.662和r = 0.622)，均随着光强的增加而增加。两次刈割可溶性蛋白的含量差异较大，第1次刈割介于3.17～5.52 mg·g−1，第2次刈割介于4.88～11.86 mg·g−1，L2处理第1次刈割中显著高于L5处理(P < 0.05)，其他处理间差异不显著(P > 0.05)，L4和L5处理第2次刈割中显著低于L1、L2和L3处理(P < 0.05)，仅第1次刈割与光强具有相关性，呈极显著负相关关系(P < 0.01, r = −0.764)。
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图 3 5种光强下黑麦草可溶性糖和可溶性蛋白含量的差异Figure 3. Differences in the soluble sugar and soluble protein contents of ryegrass under five light intensities2.4 LED红蓝光光强对黑麦草氨基酸、半胱氨酸含量的影响
5种LED红蓝光光强下两次刈割黑麦草的氨基酸和半胱氨酸含量如图4所示。5种光强下氨基酸含量介于11.87～19.48 mg·g−1，第1次刈割的氨基酸含量不同处理间无显著差异(P > 0.05)，第2次刈割L1处理含量最低，显著低于L3、L4和L5处理(P < 0.05)，氨基酸含量仅第2次刈割与光强呈显著正相关关系(P < 0.05, r = 0.474)。半胱氨酸含量介于1.19～1.92 μmol·g−1，两次刈割中L1和L2处理含量较低，显著低于含量最高的L4处理(P < 0.05)，半胱氨酸含量与光强呈显著正相关关系(P < 0.05, r = 0.465和r = 0.498)。第2次刈割中L4处理下，氨基酸和半胱氨酸含量均为最高。
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图 4 5种光强下黑麦草氨基酸和半胱氨酸含量的差异Figure 4. Differences in the amino acid and cysteine contents of ryegrass under five light intensities2.5 LED红蓝光光强对黑麦草DPPH自由基清除率和MDA含量
5种LED红蓝光光强下两次刈割黑麦草的DPPH自由基清除率和MDA含量如图5所示。5种光强下黑麦草无水乙醇提取物的DPPH自由基清除率介于23.97%～33.34%，第1次刈割不同光强处理的DPPH自由基清除率无显著差异(P > 0.05)，第2次刈割L5处理的清除率最高，显著高于其他处理(P < 0.05)，仅第2次刈割下DPPH自由基清除率与光强呈显著正相关关系(P < 0.05, r = 0.451)。第1次刈割的MDA含量较高，介于26.03～32.55 nmol·g−1，除L1处理外，其他处理间无显著差异(P > 0.05)，MDA含量随光强的升高而增加，呈显著正相关关系(P < 0.05, r = 0.482)，而第2次刈割下MDA含量与光强无显著相关性，L3处理含量最低，其他处理间无显著差异(P > 0.05)。
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图 5 5种光强下黑麦草DPPH自由基清除率和MDA含量的差异Figure 5. Differences in the DPPH free radical scavenging rate and MDA content of ryegrass under five light intensities3. 讨论与结论
人工光植物工厂中，LED提供的光照是植物光合作用唯一能量来源，也是唯一的光信号来源。本研究中光强对黑麦草的生长和产量影响显著。作为一种禾本科植物，黑麦草叶片是通过叶片基部的分生组织进行生长，因此第2次刈割的茎粗小于第1次刈割。光照处理23 d时，株高达到45 cm，刈割后继续光照进行仅17 d株高就可再次达到45 cm，虽然缩短了时间，但相同光强处理中第2次刈割的分蘖数和产量升高，这与刘春英等[20]提出适宜的刈割能刺激黑麦草加速生长的研究结果一致。两次刈割下单株分蘖数、鲜草和干草产量均随着光强的增强而增大，当光强为450 μmol·(m2·s)−1时，进一步增加光强干草产量升高不显著。鲜干比是反映牧草干物质率和适口性的重要指标之一，田间黑麦草鲜干比介于3～6 [21]，由于采用营养液 + 基质培的方法，培养环境始终含有充足的水分，鲜干比较高于田间。本研究还发现鲜干比随着光强的增加而降低，即光强越强植株含水量降低，干物率上升，此外还需要结合纤维化程度，进一步探讨光强和干物质积累的关系。
叶绿素具有捕获光能等作用，是光合作用的重要部分，光强是影响其含量的重要环境因子之一[22-23]。有研究报道在100～600 μmol·(m2·s)−1光强下水培7 d黑麦草的叶绿素a含量，呈现出先升高再降低变化[22]，本研究中总叶绿素和叶绿素a含量也呈现该变化规律，L1和L5处理的含量较低。值得注意的是，两次刈割中叶绿素b含量与光强的相关性相反。有研究表明在寡照环境下提高叶绿素b含量有利于提高水稻(Oryza sativa)的生物产量[24]，推测由于第2次刈割时黑麦草的分蘖数增加，导致叶片间的光能竞争增强，提高叶绿素b的含量可能有利于提高叶片的光捕获能力。
光强会影响植物营养物质的含量，如人工光植物工厂生产的水培生菜(Lactuca sativa)，当光强从100 μmol·(m2·s)−1升高到150 μmol·(m2·s)−1时，可溶性糖含量显著增加[25]。本研究中第2次刈割的可溶性糖含量随着光强的增加而升高。相同光强下，第2次刈割的可溶性糖含量降低，付志慧等[21]的研究也发现，第2次刈割的黑麦草水溶性碳水化合物含量降低，而中性洗涤纤维含量升高。弱光会推迟植物的生育进程[26]，在黑麦草的生育期中粗蛋白含量表现为开花期 < 孕穗期 < 拔节期[20]，本研究中第2次刈割L4、L5处理可溶性蛋白含量较其他处理降低了约60%，这主要是因为第2次刈割高光强下部分黑麦草已进入抽穗期而弱光下黑麦草仍处于拔节期。
黑麦草叶片氨基酸种类齐全、含量丰富，是畜禽的优质蛋白源[27]。本研究分析发现，第2次刈割时氨基酸含量在150～450 μmol·(m2·s)−1下随光强增强而升高。黄勤楼等[28]发现黑麦草氨基酸含量和分蘖数呈极显著相关，本研究第2次刈割的氨基酸含量也与分蘖数呈显著正相关(P < 0.05, r = 0.499)；此外还与总叶绿素含量呈显著正相关(P < 0.05, r = 0.540)，类似的结果在羊草(Leymus chinensis)中也有发现[29]。作为一种重要的含硫氨基酸，半胱氨酸在动物生产中具有促进生长、抗氧化和解毒等作用[30]，因此本研究关注了光强对黑麦草中半胱氨酸含量的影响，结果表明，增强光强可以提高半胱氨酸含量。
DPPH自由基清除率是评价物质体外抗氧化活性的重要途径[31]。本研究中，第2次刈割时高光强促进了黑麦草醇提物的DPPH自由基清除率能力，这一结果在垂盆草(Sedum sarmentosum)的研究中也有类似发现[32]。连续光照光强为100～300 μmol·(m2·s)−1下培养的生菜MDA含量，与光强呈正相关关系[15]，虽然第1次刈割时黑麦草MDA含量随光照增长而升高，但L2～L5处理间差异不显著，并没有发生高光强伤害情况。刈割胁迫给黑麦草造成逆境条件，激活了叶片抗氧化保护酶系统抑制细胞膜脂过氧化、维护氧自由基代谢平衡[33]，因此第2次刈割的MDA含量下降。两次刈割黑麦草的DPPH自由基清除率和MDA含量并没有表现出高光强胁迫，这与前人提出的黑麦草是弱光植物产生了分歧[22]，还应该结合抗氧化酶的活性变化分析光强对黑麦草抗氧化系统的影响。
光强对黑麦草生长产量和品质的影响显著，第2次刈割收获的黑麦草产量、可溶性蛋白、氨基酸和半胱氨酸含量提高，且MDA含量降低，营养品质较好。L4和L5处理下黑麦草生产性能最好，可溶性糖及半胱氨酸含量以及DPPH自由基清除能力较佳，450～550 μmol·(m2·s)−1是人工光植物工厂营养液基质培黑麦草生长的适宜光照强度，在生产实际中同时考虑能源投入，因此若以低能耗高产出为收获目标，应以450 μmol·(m2·s)−1光强为宜。田间栽培黑麦草223 d刈割3次时，不同品种鲜草产量为96 747～114 008 kg·hm−2 [20]，本研究中在45 d的培养期中，450 μmol·(m2·s)−1光强下两次刈割每穴总鲜草产量约为65 g，折算单位面积产量为137 566 kg·hm−2，由于不存在越冬期，极大缩短了生长周期，若周年运行，一年可栽培8批黑麦草，年单位面积产量为1 100 t·hm−2，以6～8层层架培养计算，年产量在6 603～8 804 t·hm−2。此外，还可以根据食草动物对鲜干比、适口性和蛋白含量的需求，选择合适的光强，并在进入孕穗期前刈割。
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图 1 5种光强下两次刈割黑麦草外观形态
Figure 1. Morphology of ryegrass grown under five light intensities for twice cuttings
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图 2 5种光强下黑麦草叶片叶绿素含量的差异
Figure 2. Differences in the chlorophyll content of ryegrass leaves under five light intensities
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图 3 5种光强下黑麦草可溶性糖和可溶性蛋白含量的差异
Figure 3. Differences in the soluble sugar and soluble protein contents of ryegrass under five light intensities
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图 4 5种光强下黑麦草氨基酸和半胱氨酸含量的差异
Figure 4. Differences in the amino acid and cysteine contents of ryegrass under five light intensities
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图 5 5种光强下黑麦草DPPH自由基清除率和MDA含量的差异
Figure 5. Differences in the DPPH free radical scavenging rate and MDA content of ryegrass under five light intensities
表 1 5种光强下第1次刈割黑麦草农艺性状和产量的差异
Table 1 Differences in the agronomic traits and yield of ryegrass under five light intensities at the first cutting
处理
Treatment株高
Plant height/cm茎粗
Stem diameter/mm单株分蘖数
Tillers per plant穴鲜草产量
Fresh yield/(g·hole−1)穴干草产量
Hay yield/(g·hole−1)鲜干比
Fresh to dry ratioL1 44.68 ± 2.35a 2.61 ± 0.11a 4.05 ± 0.34c 11.31 ± 0.75e 0.99 ± 0.08d 11.43 ± 0.38a L2 46.03 ± 3.93a 2.64 ± 0.26a 5.95 ± 0.72b 17.54 ± 1.09d 1.72 ± 0.22c 10.24 ± 0.59b L3 45.66 ± 2.89a 2.79 ± 0.29a 6.55 ± 0.10b 22.41 ± 0.69c 2.53 ± 0.25b 8.90 ± 0.64c L4 47.71 ± 1.36a 2.71 ± 0.11a 7.60 ± 0.37a 28.14 ± 1.24b 3.09 ± 0.23a 9.14 ± 0.69c L5 46.04 ± 4.35a 2.80 ± 0.22a 7.70 ± 0.93a 29.93 ± 0.87a 3.18 ± 0.26a 9.46 ± 0.86bc 同列不同小写字母表示不同光强处理间差异显著(P < 0.05)；下同。
Different lowercase letters within the same column indicate significant differences between different light intensities at the 0.05 level. This is applicable for the following tables and figures as well.
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表 2 5种光强下第2次刈割黑麦草农艺性状和产量的差异
Table 2 Differences in the agronomic traits and yield of ryegrass under five light intensities at the second cutting
处理
Treatment株高
Plant height/cm茎粗
Stem diameter/mm单株分蘖数
Tillers per plant穴鲜草产量
Fresh yield/(g·hole−1)穴干草产量
Hay yield/(g·hole−1)鲜干比
Fresh to dry ratioL1 46.51 ± 4.94bc 2.29 ± 0.14b 5.25 ± 0.50b 16.20 ± 3.16c 1.39 ± 0.34c 11.82 ± 1.15a L2 51.12 ± 2.25a 2.42 ± 0.14ab 7.80 ± 1.01a 27.00 ± 6.58b 2.84 ± 0.98b 9.95 ± 1.94ab L3 50.47 ± 1.24ab 2.48 ± 0.19ab 9.30 ± 1.60a 34.76 ± 6.74ab 4.07 ± 1.28ab 8.83 ± 1.24b L4 47.66 ± 2.91abc 2.42 ± 0.14ab 9.40 ± 0.94a 37.67 ± 6.84a 4.43 ± 0.74a 8.54 ± 1.19b L5 43.31 ± 2.40c 2.65 ± 0.21a 9.10 ± 1.00a 33.85 ± 3.80ab 4.20 ± 0.87a 8.21 ± 0.97b
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