Evidence that Tuber Respiration is the Pacemaker of Physiological Aging in Seed Potatoes (Solanum tuberosum L.)

Storage temperatures greater than 4 °C (that is, heat-unit accumulation) increase respiration and accelerate physiological aging of seed tubers. The degree of apical dominance is a good indicator of physiological age (PAGE). As seed age advances, apical dominance decreases, resulting in more stems, greater tuber set, and shifts in tuber size distribution. Herein we provide evidence that tuber respiration rate may constitute the “pacemaker” of aging. Tubers exposed to a brief high-temperature age-priming treatment initially in storage, followed by holding at 4 °C for the remainder of a 190–200-day storage period, maintained a higher basal metabolic (respiration) rate throughout storage compared with tubers stored the entire season at 4 °C. Tubers thus “remembered” the age-priming treatment as reflected by their elevated respiration rate. Moreover, reducing the respiration rate of age-primed seed by subsequently storing it at 3.5 % O₂ (4 °C) until planting significantly attenuated the effects of the aging treatment on apical dominance, tuber set, and size distribution. The effect of the age-priming treatment on the magnitude of the respiratory response was the same whether given at the beginning or toward the end of storage. However, moving the age-priming treatment progressively later in the storage season effectively decreased its impact on plant growth and development. These results underscore the importance of time in the aging process. Exposure of seed to a high-temperature age-priming treatment at the beginning or end of storage elevated respiration (the pacemaker) to the same extent; however, the timing of these treatments resulted in vastly different physiological ages. The longer the respiration rate of tubers remains at an elevated level, the greater their PAGE at planting. Thus, an accurate but impractical measure of PAGE may be the respiratory output from vine kill to subsequent planting. Respiration appears to be the pacemaker of PAGE and production, and storage conditions that affect respiration may “set the clock speed” that will ultimately determine the PAGE at planting.     

Low Root Temperature Retardation of the Mineral Nitrogen Induced Decline in N2Fixation by a Northern Ecotype of White Clover

Nodulated white clover plants (Trifolium repensL.) of a Norwegian ecotype from Pasvik (70°N) were grown in flowing solution culture. Root temperature was 17°C until 51d after sowing, when it was lowered decrementally over 5d to 7°C in four of the eight plant culture units. After a further 24h, mineral N was supplied automatically at 20μMNH₄NO₃in three culture units at each root temperature (7 and 17°C) over 17d. The remaining two units provided control plants solely dependent on N₂fixation at 7 and 17°C.The supply of NH₄NO₃greatly reduced the nodule biomass per plant at 17°C over 17d compared with control plants, but had little effect at 7°C. The nodule decline at 17°C accompanied an acute and progressive decrease in specific rate of N₂fixation, from 9mmolN d^-1g^-1nodule d.wt on day 0 to zero by day 10. Whilst initial rates of N₂fixation were lower at 7°C, the mineral N-induced decrease in fixation rates was also less severe than at 17°C and specific fixation rates recovered after reaching a minimum on day 11. N₂fixation accounted for 36% of the total uptake of N by +min.N plants during the treatment period at 7°C as opposed to only 13% at 17°C. The total N₂fixed at 7°C was 86% of that fixed at 17°C, although the specific growth rate (d.wt) at 7°C was only 55% of that at 17°C. Addition of NH₄NO₃at 7°C had little effect on the gross amount of N₂fixed subsequently. In contrast, total N₂fixation by +min.N plants at 17°C was only 24% of that fixed by the corresponding controls. The possible mechanisms by which mineral N affects N₂fixation are discussed.     

Genotypic influence on in vitro induction, dormancy length, advancing age and agronomical performance of potato microtubers (Solanum tuberosum L.)

Microtubers of 13 cultivars, largely grown in Italy and other European countries, were induced. They were stored in the dark at 3°C for different periods (28, 56, 84 and 105 days), prior to being transferred to 20°C for between 4 and 17 weeks. Following removal to room temperature, sprouting was recorded and dormancy duration quantified. Dormancy decreased from 28.1 to 19.9, 11.1 and 7.8 days with reduced time of storage. Cvs Arsy, Nicola and Jaerla took consistently more time for dormancy release. The dormancy duration was linearly and inversely correlated with the length of storage. After sprouting, tubers were held at 20°C for various intervals and a range of physiological ages (0, 368, 720 and 1008 degree days) were accumulated. The field comparison of microtubers evidenced a plant growth response and tuber yield/plant affected by the cultivar and physiological age. In early cultivars (Jaerla), a better performance was shown by younger tubers; the opposite trend was noted in Alpha (a later cultivar) with an increase in stems/plant, tubers/plant and tuber yield/plant for tubers with greater physiological age. Like conventional seed tubers, microtubers showed differences in optimum physiological age associated with cultivar earliness. This study has provided some indications on how to enhance emergence and haulm development of plants from microtubers.     

Nitrogenase Activity in the Lichen Stereocaulon paschale: Recovery after Dry Storage

The effect of prolonged storage on nitrogenase activity in Stereocaulon paschale (L.) Fr. was studied. The thalli had a very low water content during the storage over silica gel at 4°C in the dark. For C₂H₂-reduction measurements the lichen samples were remoistened and held at 15°C in the light for 1–36 h before the addition of C₂H₂. With the longest pre-incubation time preceding the nitrogenase activity measurements there was no significant decrease in nitrogenase activity during 75 weeks of storage. The method of storage was simple and inexpensive and the nitrogenase activity characteristic of each lichen batch, due to seasonal variation, was well preserved during the storage.     

Storage of Wheat FoliagePrior to Enzyme-Linked Immunosorbent Assay for Detection of Wheat Soil-Borne Mosaic Virus

Plant samples, collected at various times during a growing season, are frequently stored prior to evaluating resistance to wheat soil-borne mosaic virus (WSBMV) by enzyme-linked immunosorbent assay (ELISA). Leaves of winter wheat cvs Sage and Vona, showing symptoms of WSBMV infection, were cut in half along the midrib. Each half was either: 1) refrigerated at 4 °C, 2) frozen at ?20 °C, 3) frozen at –70 °C, or 4) desiccated with CaCl₂. Relative virus antigen titres were evaluated for individual leaf halves by ELISA. ELISA absorbance means from desiccated leaf halves were consistently higher than absorbance means from corresponding leaf halves that had been frozen. This distinction suggests that virus antigen decreases during freezing but is retained during chemical desiccation. All 4 methods of storage were found to be suitable for short-term storage prior to qualitative evaluations by ELISA, but chemical desiccation was the superior method for long-term storage and for storage of foliar samples prior to quantitative evaluations by ELISA.     

QTL mapping of potato chip color and tuber traits within an autotetraploid family

Cultivated potato (Solanum tuberosum L.) is a highly heterozygous autotetraploid crop species, and this creates challenges for traditional line development and molecular breeding. Recent availability of a single-nucleotide polymorphism (SNP) array with 8303 features and software packages for linkage and association mapping in autotetraploid species present new opportunities for the identification of genomic regions that contribute to high-value traits in cultivated potato. A biparental tetraploid potato population was evaluated across three field seasons and storage trials in order to identify quantitative trait loci (QTL) for multiple tuber traits including fried chip color after 5.5–7.2 °C storage. Tetra-allelic dosage information was used to construct a genetic linkage map that covered 1041 cM and contained 2095 SNP markers with a median marker interval of 0.4 cM. A total of 41 QTL were identified for flower color, tuber yield, tuber number per plant, tuber weight, tuber size, and chip color after various storage regimes. Moderate effect QTL for chip color at 3 months were identified that co-localized with candidate genes vacuolar invertase (VInv), invertase inhibitor (INH2), and apoplastic invertase (Inv _ap -b). A separate QTL for chip color after 6 months of storage was identified in the short arm of chromosome 2, and this locus may contribute to variation in senescent sweetening resistance. QTL for tuber weight, length, and width co-localized with a known QTL for plant maturity on chromosome 5. Genome-wide association mapping using a polyploid model detected the tuber size QTL and identified a number of candidate SNPs, but was unable to detect markers significantly associated with chip color.     

In-season heat stress compromises postharvest quality and low-temperature sweetening resistance in potato (Solanum tuberosum L.)

Key message

High soil temperature during bulking and maturation of potatoes alters postharvest carbohydrate metabolism to attenuate genotypic resistance to cold-induced sweetening and accelerate loss of process quality.

Abstract

The effects of soil temperature during tuber development on physiological processes affecting retention of postharvest quality in low-temperature sweetening (LTS) resistant and susceptible potato cultivars were investigated. ‘Premier Russet’ (LTS resistant), AO02183-2 (LTS resistant) and ‘Ranger Russet’ (LTS susceptible) tubers were grown at 16 (ambient), 23 and 29 °C during bulking (111–164 DAP) and maturation (151–180 DAP). Bulking at 29 °C virtually eliminated yield despite vigorous vine growth. Tuber specific gravity decreased as soil temperature increased during bulking, but was not affected by temperature during maturation. Bulking at 23 °C and maturation at 29 °C induced higher reducing sugar levels in the proximal (basal) ends of tubers, resulting in non-uniform fry color at harvest, and abolished the LTS-resistant phenotype of ‘Premier Russet’ tubers. AO02183-2 tubers were more tolerant of heat for retention of LTS resistance. Higher bulking and maturation temperatures also accelerated LTS and loss of process quality of ‘Ranger Russet’ tubers, consistent with increased invertase and lower invertase inhibitor activities. During LTS, tuber respiration fell rapidly to a minimum as temperature decreased from 9 to 4 °C, followed by an increase to a maximum as tubers acclimated to 4 °C; respiration then declined over the remaining storage period. The magnitude of this cold-induced acclimation response correlated directly with the extent of buildup in sugars over the 24-day LTS period and thus reflected the effects of in-season heat stress on propensity of tubers to sweeten and lose process quality at 4 °C. While morphologically indistinguishable from control tubers, tubers grown at elevated temperature had different basal metabolic (respiration) rates at harvest and during cold acclimation, reduced dormancy during storage, greater increases in sucrose and reducing sugars and associated loss of process quality during LTS, and reduced ability to improve process quality through reconditioning. Breeding for retention of postharvest quality and LTS resistance should consider strategies for incorporating more robust tolerance to in-season heat stress.     

Changes in the activity of hydroxycinnamyl CoA:quinate hydroxycinnamyl transferase and in the levels of chlorogenic acid in potatoes and sweet potatoes stored at various temperatures

A large increase in the activity of hydroxycinnamyl CoA:quinate hydroxycinnamyl transferase (CQT) occurred in potatoes stored at 0 and 2° and such an increase was prevented by storage at either 5 or 10°. The increase was most rapid in potatoes stored at 0° where it reached a maximum after 28 days and then declined slowly during storage for up to 6 months. Accompanying these changes in CQT were transitory increases in p-coumarate CoA ligase and PAL which occured during the first few weeks of storage at 0° and during this period there was nearly a two fold increase in the chlorogenic acid content of the tissue. The increase in chlorogenic acid did not occur at 10° when the increases in PAL, ligase and CQT were also prevented. The increase in CQT was reversed when tubers stored at 0° for 14 days were returned to 10° and this warming up period prevented further increase in CQT on return to 0°. The increase in CQT at 0° was prevented if the air in the storageatmosphere was replaced by N₂, 1 % O₂ or 10–15% CO₂. Similar increases in CQT, ligase and chlorogenic acid occurred in sweet potatoes stored at 7.5° but were prevented by storage at 15°. The role of PAL, ligase and CQT in the control of chlorogenic acid accumulation in these commodities and the significance of changes in their activities in relation to physiological changes at low temperatures are discussed.     

Storage of samples infected with lettuce or cucumber mosaic viruses prior to testing by ELISA

Methods of processing and storing lettuce mosaic and cucumber mosaic virus infected tissues prior to ELISA have been examined. Virus antigen detection was highest in samples homogenised in phosphate inoculation buffer (1% K₂HPO₄, 0.1% Na₂SO₃) and stored at either — 20°C or — 196°C. Detection was poor in samples homogenised in phosphate buffered saline prior to storage and in samples of all treatments stored at 4°C. Freeze-dried leaf segments retained a high concentration of antigen, as did freeze-dried homogenates of samples prepared in inoculation buffer. Higher levels of antigen were detected in samples stored as whole leaves at 4°C for 24 h before processing, than in samples stored as leaf segments during this period.     

Exogenous GA<Subscript>3</Subscript> overcomes bud deterioration in tulip (<Emphasis Type="Italic">Tulipa gesneriana</Emphasis> L.) bulbs during dry storage by promoting endogenous IAA activity in the internodes

Endogenous free IAA was examined with an immunohistochemical method for its involvement in the reduction of bud deterioration after GA₃ was injected into the bulbs. We found that tulip bulbs stored at 20°C constantly developed severe bud deterioration, whereas the symptoms of deterioration was lighter in the bulbs with GA₃ injection and not observed in the bulbs with 4°C treatment. 73% success in overcoming bud deterioration was achieved in 20°C with GA₃ treatment after 8 weeks of bulb storage, and the success rate was 7% after 12 weeks of storage. IAA was detected in the parenchyma cells in the internodes of the shoot after the bulbs were stored at 4°C or at 20°C with GA₃ injection for 4 weeks, but little was detected in the bulbs stored at 20°C constantly. Moreover, a weak IAA signal was present in between the cells of the internodes irrespective of bulb treatment. After planting, the bulbs that had been treated differently exhibited different flowering ability. The bulbs stored at 4°C for 4, 8 and 12 weeks attained high flowering percentage, which was lower in the 20°C with GA₃ treatment and lowest in the 20°C treatment. It may be concluded that GA₃ injection decreases bud deterioration of tulip bulbs during dry storage at 20°C by promoting the endogenous IAA in the internodes.     

Increased shelf life of a bioherbicide through combining modified atmosphere packaging and low temperatures

The effects of air temperatures (4, 14 and 24°C) and modified atmosphere packaging (MAP) (0% CO₂/100% N₂; 20% CO₂/80% N₂ or 40% CO₂/60% N₂) on vigour of a Sclerotinia minor barley formulation during 6 months storage were evaluated. The study was performed using a multilevel factorial experimental design and response surface methodology (RSM) and aimed to determine the optimum combination of the above factors that resulted in retention of S. minor vigour during storage. Temperature and storage duration are the main factors that affect S. minor vigour. CO₂ concentration had no effect on S. minor vigour during storage. However, oxygen displacement from storage containers by CO₂ and N₂ resulted in significant decrease of vigour reduction of S. minor as compared to ambient air control. An acceptable level of S. minor vigour reduction (ALVR) during storage was developed and determined to be ALVR=31.7±14.8% (mean±95% CI). Contour plot analysis indicated that the S. minor barley formulation at 0.4 water activity could be stored for 6, 12 or 26 weeks without exceeding the upper ALVR threshold (ca. 46%) at air temperatures not higher than 20, 17 or 11°C, respectively.     

Standardization of seed storage conditions in chilgoza pine (Pinus gerardiana Wall.): an endangered pine of Hind Kush Himalaya

The seeds of chilgoza pine (Pinus gerardiana) show moderate germination and not retain better germinability under normal ambient storage. In the present study, five storage containers [polythene bags (C₁), plastic jars (C₂), canvas bags (C₃), earthen pots (C₄) and tin boxes (C₅)] and four temperature regimes (19–22 °C) (room temperature, T₁), 0 ± 1 °C (T₂), ?4 ± 1 °C (T₃) and 4 ± 1 °C (T₄) were tested and the suitable seed storage conditions of chilgoza pine to retain viability were standardized. The storage devices preserve and retain viability trend of C₄ > C₃ > C₁ > C₂ > C₅ and T₂ > T₃ > T₄ > T₁ in the species throughout the storage period. However, the interaction treatment (C₄T₂) of earthen pots under 0 ± 1 °C temperature regime maintained significantly (P < 0.05) maximum germinability after 9 months (58.3 %) followed by C₃T₂ and C₄T₃ (47.5 %) as compared to other storage conditions. Notably, a sharp decline in germinability was recorded in seeds stored in tin boxes placed at room temperature. The implementation of these results for conservation management, especially nursery development and sustainable utilization of P. gerardiana in Himalayan region, has been suggested.     

Storability, post-storage conversion and genetic stability assessment of alginate-encapsulated shoot tips of monopodial orchid hybrid Aranda Wan Chark Kuan ‘Blue’ × Vanda coerulea Grifft. ex. Lindl.

An efficient short-term storage system of synthetic seeds, produced using in vitro shoot tips of the monopodial orchid hybrid Aranda Wan Chark Kuan ‘Blue’ × Vanda coerulea Grifft. ex. Lindl. (AV), was developed. In vitro shoot tips (3–4 mm) were successfully encapsulated, resulting in uniform spherical beads (capsules), using 3 % sodium alginate with 75 mM CaCl₂·2H₂O. Maximum (~100 %) conversion (into plantlets with shoot and root) of capsules (or synthetic seeds) was achieved on quarter-strength Murashige and Skoog regrowth medium, while full-strength MS medium was required for effective conversion of non-encapsulated shoot tips. The capsules showed distinct difference in their response to temperature during storage. The conversion efficiency declined upon storage duration at both 4 and 25 °C, with those stored at 25 °C being more tolerant to storage. Capsules stored at 4 °C had rapid deterioration and faced complete death within 160 days while those stored for 200 days at 25 °C showed relatively high conversion (71.6 %). An inter-simple sequence repeats fingerprinting approach, employed on indiscriminately chosen plantlets from converted capsules (following 4 and 25 °C of storage), ensured the post-storage genetic stability.     

Relationship Between Protein Denaturation and Water Holding Capacity of Pork During Postmortem Ageing

This study investigated water holding capacity (WHC), water distribution, and protein denaturation of pork loin chops (longissimus lumborum) packaged in polyethylene bags throughout display at 4?±?1 °C for up to 1, 3, 5, 7 and 9 days. The drip loss of pork eventually increased following a decrease during the first 5 days of storage. Nuclear magnetic resonance (NMR) analysis revealed an increase in population of immobilized water P₂₂ from day 1 to day 5, meanwhile a sharp decrease after 9 days was noticed. However, an opposite trend was observed for the population of free water P₂₃. Correlation analysis indicated that myofibrillar protein solubility was negatively correlated with drip loss (p?<?0.05), whereas sarcoplasmic protein measurement were not related to drip loss (p?>?0.05). Furthermore, the content of α-helices increased during the first 5 days of storage (p?<?0.05), which suggested increased WHC during the earlier period of postmortem storage. During the subsequent postmortem storage, the content of α-helices decreased significantly (p?<?0.05), while the β-sheets and β-turns increased. The maximum temperatures (T_max) of three endothermic peaks were found to be 53.6 °C, 65.2 °C, and 77.6 °C at 1 day postmortem. A significant decrease were observed for T_{max peakI},T_{max peakII},T_{max peakIII} at 9 d when compared to 1 d postmortem (p?<?0.05), suggesting loss of thermal stability and protein denaturation.     

Mitochondria-targeted plastoquinone derivative SkQ<Subscript>1</Subscript> decreases ischemia-reperfusion injury during liver hypothermic storage for transplantation

The ability of the mitochondria-targeted plastoquinone derivative 10-(6′-plastoquinonyl)decyl triphenylphosphonium (SkQ₁) to decrease ischemia-reperfusion injury in isolated liver during hypothermic storage (HS) was studied. Rat liver was stored for 24 h at 4°C without or in the presence of 1 μM SkQ₁ with following reperfusion for 60 min at 37°C. The presence in the storage medium of SkQ₁ significantly decreased spontaneous production of reactive oxygen species and intensity of lipid peroxidation in the liver during HS and reperfusion. The GSH level after HS in solution with SkQ₁ was reliably higher, but reperfusion leveled this effect. At all stages of experiment the presence of SkQ₁ did not prevent the decrease of antioxidant enzyme activities such as catalase, GSH peroxidase, GSH reductase, and glucose-6-phosphate dehydrogenase. The addition of SkQ₁ to the storage medium improved energetic function of the liver, as was revealed in increased respiratory control index of mitochondria and ATP level. SkQ₁ exhibited positive effect on the liver secretory function and morphology after HS as revealed in enhanced bile flow rate during reperfusion and partial recovery of organ architectonics and state of liver sinusoids and hepatocytes. The data point to promising application of mitochondria-targeted antioxidants for correction of the ischemia-reperfusion injury of isolated liver during long-term cold storage before transplantation.     

Produktion von Ochratoxin A und B durch<Emphasis Type="Italic">Penicillium verrucosum</Emphasis> auf Gerste in Abhängigkeit der Parameter Wasseraktivität,Wassergehalt und Temperatur

The ochratoxin A and B (OTA, OTB) production by a toxigenic isolate ofPenicillium verrucosum grown on brewing barley up to six weeks was studied at a storage temperature of 25 °C and different moisture and water activity conditions. Sorption isothermes for barley were prepared at temperatures of 10°C, 15°C and 25°C. OTA was produced after 2 weeks of storage at moisture contents of ≥19%, which is equivalent to water activities (a_w) of 0.83 (adsorptive) and 0.82 (desorptive) at 25 °C. Increased OTA concentrations (5.8-fold and 16.1-fold) were noticed when the moisture contents were adjusted to 20% (a_{w [ads]} 25 °C=0.86) and 21% (a_{w [ads]} [ 25 °C=0.88), respectively. An increase was also shown during storage of 4 and 6 weeks (1.2-fold and 2.4-fold, respectively). Production of OTB was shown to occur at moisture contents ≥18% (a_{w [ads]} 25 °C=0.81). The findings document that OTA and OTB are not produced byP. verrucosum grown on barley stored below 18% moisture content.     

The composition of fresh and stored oyster mushrooms (Pleurotus ostreatus)

Soluble carbohydrate, protein, polysaccharide and cell wall composition were assayed in freshly harvested Pleurotus ostreatus sporophores and those stored for 4 days at 2° or 18°. Mannitol and trehalose were present at 1.8 and 6.5% dry wt respectively in fresh sporophores, and at reduced levels in those stored at 18°. In sporophores stored at 2°, trehalose levels increased by up to 122%. Soluble polysaccharide appeared to be composed of glycogen-like material, which was susceptible to post-harvest breakdown, and components containing mannose and other sugars. The total protein content was 42% dry wt; no protein degradation was seen in sporophores stored at 2°, but about 25% of the protein disappeared during storage at 18°. Cell wall polysaccharide was utilised during storage. Respiration rate was about 8–10 ml CO₂/g dry wt/hr at harvest and declined to about 5 ml/g dry wt/hr after 40 hr storage at 18°.     

Production of mouse fetuses using spermatozoa exposed temporarily to high temperature or continuously to room temperature after freeze-drying in Na+-free/K+-rich EGTA buffer

Present study aimed to determine to what extent freeze-dried spermatozoa were able to withstand high-temperature conditions: transient increase in storage temperature and long-term exposure to room temperature. Mouse spermatozoa were freeze-dried in EGTA/Tris-HCl buffered solution alkalinized using KOH (K-ETBS, pH 7.7), and then stored for up to 7 months at 4 °C or 25 °C. After 2 months’ storage, some of the 4°C-stored spermatozoa were exposed to 40 °C for 1 week or 1 month, then again stored at 4 °C for the remaining storage period. Following storage, rehydrated spermatozoa were injected into mouse oocytes. The resulting zygotes were assessed for chromosome damage, in vitro development up to the blastocyst stage, and post-implantation development to normal fetuses on day 18 of gestation. In storage at 4 °C, one-week exposure to 40 °C had no adverse effect on the chromosome integrity and developmental competence compared to non-exposure to 40 °C (continuous storage at 4 °C). In contrast, one-month exposure to 40 °C caused an increasing level of chromosome damage (36%, P < 0.05) and reduced frequencies of blastocysts (54%, P < 0.05) and normal fetuses (36%, P < 0.05) compared to the frequencies obtained by continuous storage at 4 °C (15%, 82% and 52%, respectively). Storage at 25 °C resulted in accumulation of chromosome damage (27%, P < 0.05), leading to decreased blastocyst formation (63%, P < 0.05). But, the frequency of normal fetus (44%) was not significantly different from that obtained by continuous storage at 4 °C. Consequently, mouse spermatozoa freeze-dried in K-ETBS withstood temporary exposure to 40 °C for 1 week. Chromosome damage accumulated in spermatozoa during storage at 25 °C.     

The pH-dependent recovery of smooth muscle from storage at -13 degrees C in unfrozen media.

The function of isolated strips of smooth muscle from the guinea pig were assessed by isometric contractile responses to histamine before and after storage at ?13 °C in high potassium, Pipes-buffered solutions containing 30% (w/v) Me₂SO and having different pH values. The structural integrity of the tissue during and after cooling was also assessed by electron microscopy.The pH-dependence of tissue recovery in these muscles was clearly demonstrated after about 17-hr storage at ?13 °C. Despite the wide difference in functional recoveries between groups of muscles cooled in either “high” (7.7) or “low pH” (6.4) media, however, there were no apparent ultrastructural differences between the two differently treated groups of muscles. Muscles from both pH groups, fixed after rewarming to 37 °C, showed the morphological characteristics typically seen in noncooled muscles. An examination of muscle cells which were fixed at ?13 °C (w/v) Me₂SO revealed that the contractile myofibrils of specimens in both experimental groups had undergone a noticeable reorganization and aggregation.