Evaluation of Wheat Chromosome Translocation Lines for High Temperature Stress Tolerance at Grain Filling Stage

High temperature (HT, heat) stress is detrimental to wheat (Triticum aestivum L.) production. Wild relatives of bread wheat may offer sources of HT stress tolerance genes because they grow in stressed habitats. Wheat chromosome translocation lines, produced by introgressing small segments of chromosome from wild relatives to bread wheat, were evaluated for tolerance to HT stress during the grain filling stage. Sixteen translocation lines and four wheat cultivars were grown at optimum temperature (OT) of 22/14°C (day/night). Ten days after anthesis, half of the plants were exposed to HT stress of 34/26°C for 16 d, and other half remained at OT. Results showed that HT stress decreased grain yield by 43% compared with OT. Decrease in individual grain weight (by 44%) was the main reason for yield decline at HT. High temperature stress had adverse effects on leaf chlorophyll content and Fv/Fm; and a significant decrease in Fv/Fm was associated with a decline in individual grain weight. Based on the heat response (heat susceptibility indices, HSIs) of physiological and yield traits to each other and to yield HSI, TA5594, TA5617, and TA5088 were highly tolerant and TA5637 and TA5640 were highly susceptible to HT stress. Our results suggest that change in Fv/Fm is a highly useful trait in screening genotypes for HT stress tolerance. This study showed that there is genetic variability among wheat chromosome translocation lines for HT stress tolerance at the grain filling stage and we suggest further screening of a larger set of translocation lines.


Introduction
High temperature (HT, heat) stress is one of the most important environmental stresses that adversely affects growth, development and yield of many field crops including wheat [1][2][3][4]. High temperature stress is a principal wheat yield-decreasing factor in Central Asia, North Africa, Europe, Australia, and the United States [5][6][7]. A simulation study in Australia showed wheat yield could decrease up to 50% when growing season temperature became 2°C warmer than the average [8]. Wheat is usually planted in late autumn to early winter and harvested to biotic stress such as powdery mildew, wheat curl mite, and leaf and strip rusts [29][30][31][32], but knowledge on performance of chromosome translocation lines under HT stress is limited. Because the translocation lines have chromosome segments from alien species native to stressed habitats, we hypothesize that genetic variability will exist among chromosome translocation lines for tolerance to HT stress. The objectives of this study were to evaluate wheat chromosome translocation lines for tolerance to HT stress at the grain filling stage and identify physiological and yield traits associated with the tolerance.
Seeds were sown in 1.6-L plastic pots of dimensions 14 cm (height) × 50 cm (top perimeter) × 36 cm (bottom perimeter) filled with a potting mix (Metro Mix 360; Hummert International, Topeka, KS) and 8 g of Osmocote Plus (N:P 2 O 5 :K 2 O = 15:9:12; Scotts, Marysville, OH), a slowrelease fertilizer. Six pots of plants per genotype were randomly divided into three identical growth chambers (two pots per chamber) (Conviron, Winnipeg, MB, Canada) set at an optimum temperature (OT) of 22/14°C day/night,~85% humidity, 16 h photoperiod, and photosynthetically active radiation (PAR) of 650 μmol m -2 s -1 provided by cool white fluorescent lamps (Philips Lighting Co., Somerset, NJ). Thirty days after seeding, plants were thinned and staked, leaving four plants per pot. Granular Marathon 1% pesticide (a. i.: Imidacloprid, 1-((6-Chloro-3-pyridinyl) methyl)-N-nitro-2-imidazolidinimine) was applied to avoid sucking insect pest infestation, and plants were randomly relocated weekly within the growth chamber to avoid positional effect within the chamber. Throughout the growing season, growth chambers' air temperature and relative humidity were monitored every 20 min with HOBO U14-001 LCD Temperature/Relative Humidity (RH) Data Logger (Onset Computer Corporation, Bourne, MA). Thick paper caps were used on the top of the HOBO data loggers to shield against direct heat radiation from the lamps. The PAR was monitored once a month with a Field Scout Light Sensor (Spectrum Technologies, Inc., Plainfield, IL).
At anthesis, two plants per pot were randomly selected and main stems were tagged. Ten days after anthesis, the temperature treatment was applied by moving one pot of each genotype from the OT regime (22/14°C day/night) to one of the growth chambers' set at HT of 34/26°C day/night,~85% humidity, 16 h photoperiod, and PAR of 650 μmole m -2 s -1 . The dates of movement (transfer) of pots to the HT chamber were different because of different anthesis dates. Another pot of each genotype remained at OT regime throughout the experiment and served as a control. The growth chambers' air temperature and relative humidity at HT were also constantly monitored with HOBO loggers with thick paper cap throughout the experiment period. The stress period was 16 d, after which pots were moved back to their original growth chambers. To avoid water stress, plants in both temperature regimes were fully irrigated by keeping pots in a tray filled with~2 cm water until physiological maturity. We recognized, however, that leaving plants sitting in water for so long may cause water-logging/anoxia; which should be documented or avoided in the future studies. Once a week until physiological maturity, plants were fertigated with Miracle-Gro water-soluble fertilizer (N:P 2 O 5 :K 2 O = 24:8:16; Scotts Miracle-Gro Products, Inc., Marysville, OH) per manufacturer's instruction. The daytime and nighttime temperature of all growth chambers were held for 12 and 8 h, respectively, with a 2-h transition period between them. During transition period, the rate of change in chamber temperature was set at 1°C/15 minutes.

Data Collection
The date of anthesis was recorded when 50% of plants in a pot reached Feekes growth stage 10.5.1; and the days to anthesis was calculated by subtracting planting date from anthesis date.

Superoxide Dismutase and Superoxide Anions
Leaf samples (flag leaf) collected from the secondary tiller of a tagged plant at 10 DAST were immediately frozen in liquid nitrogen and stored at-80°C until further analysis. Superoxide dismutase activity was assayed by measuring its ability to inhibit the photochemical reduction of nitroblue tetrazolium (NBT) at 560 nm following the protocol of [39] with slight modification. Frozen tissues were ground in a cold mortar and pestle using liquid nitrogen and suspended in 1.5 ml of ice-cold extraction buffer solution containing 50 mM Na-PO 4 buffer at pH 7.8, 1 mM EDTA.Na 2 and 2% (w/v) polyvinypyrrolidone. The suspension was centrifuged at 20000 × g at 4°C for 20 min. The supernatant was used to measure the enzyme activity. A reaction mixture of 3 ml containing 50 mM Na-PO 4 buffer at pH 7.8, 0.66 mM of EDTA.Na 2 , 10 mM of L-methionine, 33 μM of nitroblue tetrazolium and 0.0033 mM of riboflavin, and 50 μL of extraction buffer was added to the 150 μl of supernatant, and the tubes were illuminated with luminescent lamps for 10 min. The extinction was then read against a blank at 560 nm using a U-2000 double-beam UV/Vis spectrophotometer (Hitachi, Tokyo, Japan). One enzyme unit of SOD was defined as the amount required to inhibit photochemical reduction of NBT by 50% and was expressed as enzyme unit mg protein -1 . The concentration of superoxide anion was estimated according to Chaitanya and Naithani [40]. About 0.2 g of leaf samples were suspended in ice-cold 0.2 M sodium phosphate buffer at pH 7.2 containing diethyl dithiocarbamate (10 -3 M). The suspension was immediately centrifuged at 3000 × g at 4°C for 1 min. In 200 μl of supernatant, 2 mL of nitroblue tetrazolium (2.5 × 10 -4 M) was added and the ODA was recorded against a blank at 540 nm every 30 s for 150 s using a Hitachi U-2000 double-beam UV/Vis spectrophotometer. The superoxide anion was expressed as change in optical density min -1 g -1 fresh weight.

Yield and Yield Components
Plants were harvested by cutting at ground level. Spikes were counted, detached, and dried to constant weight in a 38°C incubator. The remaining aboveground biomass was oven-dried at 65°C to a constant weight. The dried spikes were hand-threshed. The spikes from primary tillers of two tagged plants were used to determine the number of grains per spike and grain weight per spike. Individual grain weight was estimated by dividing grain weight per spike by number of grains per spike and expressed in mg. Yield per plant was the mean grain weight in g from all the spikes of tagged plants.

Heat Susceptibility Index (HSI)
The heat susceptibility index (HSI) for grain yield was calculated according to Fischer and Maurer [41]: where Y is the average grain yield per plant of an accession at HT of 34/26°C, Yp is the average grain yield per plant of the same accessions at OT of 22/14°C, and D is the stress intensity, equal to 1-X/Xp, in which X is the mean Y of all accessions and Xp is the mean Yp of all accessions. The genotypes were classified as highly tolerant (HSI 0.5), tolerant (0.5 < HSI 0.75), moderately tolerant (0.75 < HSI 1.0), and highly susceptible. Similarly, HSIs for physiological and yield traits were also calculated and used in path coefficients to find relationships among heat tolerance of various traits including yield. They were also used to rank the genotypes for tolerance to HT stress.

Statistical Analyses
The experimental design was a split plot with three replications (three growth chambers under each temperature regime). The temperature treatment was randomly assigned to the growth chambers and considered as the main plot. The sub-plot comprised genotypes. SAS 9.2, PROC GLIMMIX (SAS Institute Inc., Cary, NC, 2003), was used to analyze the data with block, temperature, and genotypes as class variables; block and block × temperature as random effects; and all other variables as fixed effects [42]. The least square means were separated at p < 0.05 after Tukey-Kramer adjustment. The time series data on leaf chlorophyll, flag leaf temperature, and Fv/Fm were analyzed using the REPEATED statement and Type = CS, a covariance structure of compound-symmetry type. PROC REG in SAS was used to determine the relationship between two variables, and PROC CALIS in SAS was used to conduct path coefficient analysis. At the end, genotypes were ranked on the bases of heat susceptibility indices of physiological traits, yield, and yield components using the RANK function in Microsoft Excel (2010).

Results
The average day/night temperature recorded during the treatment period was 33.7±0.7/26.1±0.5°C in growth chambers set for the HT regime and 21.4±0.3/14.3±0.5°C in growth chambers set for the OT regime. As expected, there was no significant effect of temperature and interaction effect of temperature × genotype on days to anthesis; however, genotypes varied for this trait. The days to anthesis ranged from 88 in TA5599 to 114 in TA5088; and the mean, median, and mode were 98, 97, and 96 respectively. There were significant effects of temperature, genotype, and temperature × genotype interaction on physiological and yield traits, unless stated otherwise ( Table 2).
Leaf temperature and leaf chlorophyll as a function of DAST are shown in Fig. 2. The relationship between leaf temperature and DAST under HT stress was significant and positive, with a slope of 0.2°C when averaged across the genotypes ( Fig. 2A), and the relationship between leaf chlorophyll and DAST under HT stress was significant and negative, with a slope of-2.03 SPAD unit (Fig. 2B). At OT, the relationship was either not significant (leaf temperature: slope = 0.02°C, R 2 = 0.007, and p = 0.46) or very weak (leaf chlorophyll: slope = -0.3 SPAD unit, R 2 = 0.10, p = 0.003); therefore, OT data are not presented or discussed further. An analysis of genotypic behavior at HT stress showed that the change in leaf temperature over the time period was not significant in TA4350-119, TA5594, or TA3008; however, TA5617, TA4350-118, and TA5639 had the highest rate of increase in leaf temperature (slope = 0.32 to 0.34°C; Table 3). Similarly, under HT stress, the rate of decrease in leaf chlorophyll content throughout the experiment was < 1 SPAD unit in TA4350-119, TA5594, and TA5617; between 1 and 2 in TA5088, TA5608, TA5636, TA4305-118, and TA5595; and >3 in TA5638 and TA5639.
The difference between temperature regimes for maximum quantum yield of PSII (Fv/Fm, unitless) was significant at p < 0.001. High temperature stress at the grain filling stage decreased Fv/Fm by 38% (Fig. 1C) compared to OT when averaged across the genotypes and over the first 14 d of readings. Genotypes behaved differently according to the temperature treatment at p < 0.001 (Table 2). When averaged across the days, HT stress had no effect on TA4350-119, TA5594, TA5617, TA5088, and TA5599 for Fv/Fm compared with OT. However, Fv/Fm decreased by < 35% in TA5608, TA5595, TA5602, and TA4305-118 and by 60-64% in TA5640, TA5624-4, and TA5639. Fv/Fm as a function of DAST is shown in Fig. 2. A significant negative relationship was evident between Fv/Fm and DAST under HT stress when averaged across the genotypes, with a slope of-0.04 unit (Fig. 2C). At OT, the relationship of Fv/Fm with DAST was very weak (slope = -0.005 unit, R 2 = 0.08); therefore, data are not presented or discussed. Further analysis of genotypic behavior at HT stress showed that TA4350-119, TA5594, and TA5617 had the lowest rate of decrease in Fv/Fm (slope < -0.02 unit), and TA5638 and TA5601 had the highest rate of decrease (slope > -0.06 unit) during the time period (Table 3).   Relationships among leaf temperature, leaf chlorophyll, and Fv/Fm are presented in Fig. 3. At OT, there was no relationship between leaf chlorophyll and leaf temperature (R 2 = 0.12, p > 0.05) (Fig. 3A), and a very weak positive relationship between Fv/Fm and leaf temperature (R 2 = 0.19, p = 0.05) (Fig. 3B). The relationship between Fv/Fm and leaf chlorophyll content was non-significant (R 2 = 0.0002, p > 0.05) (Fig. 3C). At HT, there was a highly negative relationship between leaf chlorophyll and leaf temperature (R 2 = 0.58, p < 0.001) (Fig. 3D); and Fv/Fm and leaf temperature (R 2 = 0.52, p < 0.001) (Fig. 3E). The relationship between Fv/Fm and leaf chlorophyll at HT stress was strongly positive (R 2 = 0.67, p < 0.001) (Fig. 3F).

Superoxide Dismutase (SOD) and Superoxide Anion
There was a significant difference between temperature regimes for SOD activities and concentration of the superoxide anion, a ROS. High temperature stress decreased SOD activity by 8% and increased superoxide anion concentration by 44% (0.12 changes in OD min -1 g -1 FW) compared to OT when averaged across the genotypes. Genotypes behaved differentially according to the temperature treatment for SOD at p < 0.001 (Table 4). The effect of HT stress was not evident in 17 genotypes for SOD, but SOD activity decreased by 29% in TA5638 and TA5600 and by 52% in TA4305-118. Genotype and temperature × genotype interaction was not significant for the superoxide anion, so data for this trait were not presented in further detail (p > 0.05) ( Table 2).

Spike Number, Grain Number, Individual Grain Weight, and Grain Yield
Effects of temperature and interaction of temperature × genotype were not significant for spike number per plant and grain number per spike; however, a significant variation among genotypes was evident at p < 0.001 ( Table 2). The average spike number per plant ranged from 1.2 in TA5088 to 2.9 in TA5608; and grain number per spike ranged from below 33 in TA5600,  TA5601, TA5602, TA5624-4, TA4350-119, and TA4305-118 to more than 66 in TA5594,  TA5595, TA5638, and TA5088. There was a significant difference between temperature regimes for IGW and grain yield per plant, and genotypes behaved differently according to temperature treatment for these traits at p 0.004 (Table 2). High temperature stress decreased IGW and grain yield by~43% compared with OT when averaged across the genotypes. As a result of HT, IGW decreased by 30% in TA5594, TA5088, and TA5608 and by 71% in TA5640 (Fig. 4A). High temperature stress significantly decreased grain yield in TA5595, TA5615, TA5636, TA5637, TA5638, TA5640, TA3008, and TA4350-118, and the decrease ranged from about 41% in TA5595 to 77% in TA5640. The decrease in grain yield because of HT stress was not statistically significant in other genotypes (Fig. 4B).  Fig. 5. The genotypes TA5617 and TA4350-119 had 0.5 HSI for leaf chlorophyll; and TA5594, TA5599, TA5088, TA5608, TA4350-118 had HSI between 0.5 and 0.75. The genotypes TA5594, TA5617, TA5088, TA4350-119 had 0.5 HSI for Fv/Fm; and TA5599, TA4305-118 had HSI between 0.5 and 0.75. The genotypes TA5594, TA5088, TA5608 had 0.75 HSI for individual grain weight and all other genotypes had > 0.75 HSI. The genotypes TA5617 and TA5088 had < 0.5 HSI for grain yield, and TA5594, TA5639, TA5599, TA5608 had HSI between 0.5 and 0.75. The genotypes TTA5637, TA5638, TA5640, and TA3008 had > 1.0 HSI for all traits.

Pearson's Correlation Coefficients and Path Coefficient Analysis
At OT, there was no significant correlation between individual grain weight and grain yield per plant; and the maximum quantum yield of PSII (Fv/Fm) had no significant correlation with leaf chlorophyll and individual grain weight. A significant positive correlation was evident between Fv/Fm and grain yield per plant; and leaf chlorophyll had significant positive correlation with individual grain weight and grain yield per plant (Table 5). At HT, there was significant positive correlation among Fv/Fm, individual grain weight and grain yield per plant. Although leaf chlorophyll had strong positive correlation with Fv/Fm and individual grain weight, there was no correlation with grain yield per plant. A path coefficient analysis performed to identify correlation or direct and indirect effects of heat responses and relationships of heat susceptibility i.e. heat susceptibility indices (HSIs) of physiological and yield traits to each other and to HSI of yield are presented in Fig. 6. There was a strong positive correlation between SPAD HSI and Fv/Fm HSI . The SPAD HSI had neither direct nor indirect effect on IGW HSI and Yield HSI ; however, the Fv/Fm HSI had significant direct effect on IGW HSI and indirect effect on Yiled HSI . The IGW HSI had a strong direct effect on Yield HSI .

Ranking of Genotypes Based on HSIs of Physiological, Yield Traits, and Yield
The rankings of genotypes on the bases of their response to HT stress for traits i.e. Fv/Fm HSI , IGW HSI , and Yield HSI are presented in Table 6. There was evidence of direct and indirect effects of these traits on Yield HSI (Fig. 6). On the basis of final ranking estimated from a mean of all   Evaluation of Wheat Translocation Lines under High Temperature Stress rankings, TA5594, TA5617, and TA5088 were the top three genotypes tolerant to HT stress; and TA5640 and TA5637 were highly susceptible genotypes with rankings of 19 and 20.

Discussion
We evaluated 16 wheat chromosome translocation lines that had a small chromosome segment from different wild wheat species introgressed into spring wheat cultivars for HT stress tolerance at the grain filling stage. The effect of HT stress was highly significant for all physiological and yield traits except grain number per spike, and we demonstrated genotypic variability in chromosome translocation lines for physiological traits, yield components, and grain yield. This information can be used in breeding programs to develop stress-tolerant cultivars. Canopy temperature shows the efficiency with which evaporative cooling is occurring from the plant surface, and it has been used to estimate heat stress in field crops under fully or limited irrigated conditions [16,[43][44][45]. Ayeneh et al. [46] showed a strong positive correlation between canopy temperature and flag leaf temperature (r = 0.91) in 13 wheat genotypes grown under HT stress. In this study, the HT stress at the grain filling stage increased flag leaf temperature by 6°C as compared to OT (Fig. 1A); and there was genotypic variation in flag leaf temperature. Genotypic variation in flag leaf temperature under HT stress was evident in wild wheat relatives [1]; and the variability in flag leaf temperature at HT stress might be attributed to differential decrease in vascular capacity and impairment of cooling mechanisms [47,48]. High temperature stress increased superoxide anions by 44% and decreased SOD activity by 8% (Table 4). Reactive oxygen species like superoxide anions are a product of regular subcellular processes and are strictly regulated by the antioxidant enzyme SOD to avoid their detrimental effects on cellular structures. High temperature stress increases the formation of ROS in chloroplast owing to the over excitation of chlorophyll molecules and decreases in antioxidant enzyme activity [13,17]. High temperature stress decreased leaf chlorophyll and maximum quantum yield of PSII (Fig. 1B, and 1C). Genotypic variation for these attributes was evident. High temperature stress is well known to decrease leaf chlorophyll [1,10,13], which could be the result of thylakoid membrane damage [11,12], lipid peroxidation of chloroplast membranes [13], and inhibition of chlorophyll biosynthesis [14]. The study showed that the longer the duration of HT stress, the higher the leaf temperature but the lower the leaf chlorophyll and the maximum quantum yield of PSII (Fv/Fm) (Fig. 2). It has been reported that light is one of the primary stress parameters for PSII performance, and significant differences in Fv/Fm can be found even among upper and lower leaves of a wheat plant due to shading [49]. In this study, as the fluorescence data was collected from the upper most flag leaf only; and the light level in both HT and OT chambers were about 650 μmol m -2 s -1 provided with the same make and model of white fluorescent lamps, we assumed that the influence of light source on Fv/Fm was negligible; and difference in Fv/Fm among genotypes was due to their differential tolerance to high temperature. Genotypic variation for the rate of increase or decrease of respective physiological traits at HT stress was evident (Table 3). In addition, under OT, there were no evidence of a relationship between leaf temperature and leaf chlorophyll or Fv/Fm, and leaf chlorophyll content and Fv/ Fm (Fig. 3), whereas under HT stress, the relationships were either highly negative (between leaf temperature and leaf chlorophyll or Fv/Fm) or highly positive (between leaf chlorophyll and Fv/ Fm). In this study, under HT stress, for a unit increase in leaf temperature, there was an evidence of decrease in leaf chlorophyll by 5.82 SPAD unit, and Fv/Fm by 0.105 unit; and for a decrease in one SPAD unit there was an evidence of decrease in Fv/Fm by 0.016 unit (Fig. 3).
In this study, HT stress had no effect on grain number per spike ( Table 4). The absence of a treatment effect on grain number in this study contrasted with earlier findings from a controlled environment experiment, where winter wheat cultivar 'Karl 92' had a 12.5% reduction in grain number when temperature increased from 25/20°C to 35/20°C from 10 d after anthesis [18], and from a field experiment, where Zhong-hu and Rajaram [50] observed about a 13% decrease in grain number between late-and normal-planted wheat. Hays et al. [51] reported that HT stress at 10 d after anthesis induced ethylene production, resulting in kernel abortion in heat-susceptible winter wheat 'Karl 92' but not in heat-tolerant spring wheat cultivar 'Halberd'. This result showed that genotypes used in the current study were tolerant to HT stress for kernel abortion; however, we recommend further study of these genotypes to elucidate the mechanism of embryo tolerance to HT stress. High temperature stress decreased individual grain weight (Fig. 4A) by 43%, which is about 7% lower than the decline observed by Yang et al. [20] in 30 synthetic hexaploid wheat varieties and 13% higher than the decline observed by Gibson and Paulsen [18] in 'Karl 92' subjected to HT stress from 10 d after anthesis. The decrease in IGW under HT stress might be due to an increase in leaf senescence and decrease in grain-filling duration [8,10,20,21], and genotypic variation for this trait has been observed by other authors [15,[18][19][20]. High temperature stress decreased grain yield per plant by 44%. Yang et al. [20] and Gibson and Paulsen [18] observed about 54 and 78% decreases, respectively, in grain yield when HT stress was applied to wheat 10 d after anthesis. Grain number per spike and IGW are two important factors of grain yield per plant. The absence of treatment effect on grain number might be the reason for the smaller decrease in grain yield in this study compared with earlier studies. As in earlier work, genotypes differed significantly in their response to HT stress for grain yield.
A correlation and path diagram analyses to investigate correlations between heat responses of physiological and yield traits (HSIs) to each other and to grain yield HSI showed that there was a strong correlation between SPAD HSI and Fv/Fm HSI (Fig. 6); however, SPAd HSI had no significant effect on IGW HSI and Yield HSI , whereas Fv/Fm HSI had a strong correlation with IGW HSI and Yield HSI . There was evidence that the nature of the effects of Fv/Fm HSI on IGW HSI and of IGW HSI on Yield HSI were direct but the effect of Fv/Fm HSI on Yield HSI was indirect. These results provide evidence that there might be strong associations (cause-effect relationships) among HSIs of Fv/Fm, IGW, and grain yield; this implies that the physiological trait such as Fv/Fm is useful in screening genotypes for HT stress tolerance. However, we are of view that gas exchange traits would have added further valuable information on the behavior of genotypes at HT; and therefore we recommend the collection of gas exchange data in future studies. On the bases of above result, we used Fv/Fm HSI , IGW HSI, and Yield HSI in ranking the genotypes for HT stress tolerance instead of using HSIs of Yield and/or IGW only. The ranking showed that TA5594, TA5617, and TA5088 were highly tolerant (ranked 1to 3), and TA5637 and TA5640 were highly susceptible (ranked 19 and 20) to HT stress compared with other lines and the check cultivars.

Conclusions
This study revealed genetic variability among wheat chromosome translocation lines for HT stress tolerance at the grain filling stage. It illustrated that a decrease in individual grain weight was the main reason for yield decline and quantified that an increase in HT stress (high leaf temperature) adversely affected leaf chlorophyll and Fv/Fm; and a decrease in Fv/Fm might be one of the physiological reasons for subsequent decline in individual grain weight. In this study, we identified TA5594, TA5617, and TA5088, having small segment of chromosome from Haynaldia villosa or Aegilops speltoides, as highly tolerant to HT stress at grain filling stage. These tolerant genotypes may be used to improve HT stress tolerance of wheat cultivars. We recommend further screening of a larger population of translocation lines. Because other translocation lines might have chromosome segments from different wild relatives or different genomes, there are good possibilities of discovering more HT stress tolerant genotypes.