The response of pigeon pea (Cajanus cajan) to the application of fertilisers

The effects of fertilisers on the performance of three cultivars of pigeon pea, Cita-1, Cita-2 and Cita-4, were measured in a pot experiment and a field trial from February to August 1982. The fertilisers were either applied as complete NPK (15-15-15) to the soil at a rate of 50 kg/ha in single or split doses within 4 wk of sowing, or as solutions containing 100 mg/litre each of N, P and K sprayed on to the plants 4 wk after sowing or at anthesis. The growth of pigeon pea was only enhanced significantly by fertiliser application in the field experiment but yields and their components were changed by fertilisers in both experiments. The magnitudes of the effects on growth and yield were similar with both methods of application.     

Cellular and molecular adaptations to injurious mechanical stress

Transient disruptions of plasma membrane integrity--'wounds'--are frequently suffered by cells of gut, skin, muscle and the aorta, organs that are normally subjected to mechanical stress in vivo.As a protection against such potentially fatal mechanically induced injuries, cells may employ specialized submembranous proteins that mechanically reinforce the plasma membrane and thus prevent wounding or, should wounding occur, they may assemble a cytoskeletal structure to aid wound healing. Membrane wounds may provide a route out of the cytoplasm for basic fibroblast growth factor, explaining how a growth factor that lacks a conventional signal peptide sequence can act extracellularly.     

Responses of Cajanus cajan and rhizospheric N-cycling communities to bioinoculants

Background and aims

Bioinoculants are commonly used for enhancing crop productivity but little information is available on their effect on key microbial communities such as those involved in the cycling of nitrogen, a major plant nutrient. Here we developed a formulation combining different bioinoculants (Bacillus megaterium, Pseudomonas fluorescens and Trichoderma harzianum) and examined their effects on both Cajanus cajan growth and N-cycling microorganisms.

Methods

Seven bioinoculant combinations were evaluated in pots under field conditions, and their effects on plant growth were measured using various biometric parameters. The abundances of the total bacterial and crenarchaeal communities along with those involved in N-cycling were monitored by qPCR at vegetative, pre-flowering, flowering and maturity stages of the crop.

Results

A significant increase in growth of C. cajan was observed when treated with mixture of three bioinoculants with dry biomass and grain yield increase by 330?% and 238?%, respectively. The combination of three bioinoculants also increased the abundance of nitrogen fixers and denitrifiers towards the flowering and maturity stages.

Conclusions

The consortium of three bioinoculants increased plant growth and grain yield of C. cajan. These bioinoculants also had a positive effect on the abundance of several N-cycling microbial communities stressing the importance of understanding non-target effects of bioinoculants together with their impact on plant growth.     

Regeneration of pigeonpea (Cajanus cajan) from cotyledonary node via multiple shoot formation

Summary Plant regeneration, which is the major limiting factor for transformation of Cajanus cajan, has been obtained via multiple shoot formation from the cotyledonary node region of seedlings germinated on MS medium containing 2 mgl^–1 6-benzylaminopurine. A mass of multiple shoot-initials formed at the axillary bud region of the cotyledonary node of the seedlings within two weeks. The cotyledonary node along with the mass of shoot-initials excised from the seedling, continued to form new shoot-initials on MS medium containing 6-benzylaminopurine (2 mgl^–1) and supplemented topically with indole-3-acetic acid. The formation of new shoot-initials was also observed from the cotyledonary nodal explant, after cutting off its surface layers to completely remove the pre-existing shoot-initials and culturing it on 6-benzylaminopurine (2 mgl^–1) containing medium. The shoots elongated rapidly on basal MS medium and rooted efficiently in MS medium supplemented with indole-3-butyric acid (0.5 mgl^–1). The procedure described is efficient, and highly reproducible and a common response was observed for all the six varieties tested.Abbreviations BAP 6-benzylaminopurine - IAA indole-3-acetic acid - IBA indole-3-butyric acid - NAA -naphthaleneacetic acid - MS medium, Murashige and Skoog (1962) medium - CN cotyledonary node     

Thidiazuron-induced shoot regeneration in pigeonpea (Cajanus cajan L.)

Thidiazuron either alone or in combination with IAA induced high frequency shoot regeneration from primary leaf segments of three pigeonpea cultivars. Transfer of the cultures to medium with reduced concentration of thidiazuron resulted in further development of the shoots. The regenerated shoots were subsequently transferred to medium supplemented with BA, IAA and gibberellic acid where 5-10% of the shoots elongated further. Rooting of shoots could be obtained on half strength MS medium supplemented with NAA. Histological studies confirmed the mode of regeneration as shoot organogenesis. This revised version was published online in June 2006 with corrections to the Cover Date.     

Oxidative stress and inflammation: liver responses and adaptations to acute and regular exercise

The liver is remarkably important during exercise outcomes due to its contribution to detoxification, synthesis, and release of biomolecules, and energy supply to the exercising muscles. Recently, liver has been also shown to play an important role in redox status and inflammatory modulation during exercise. However, while several studies have described the adaptations of skeletal muscles to acute and chronic exercise, hepatic changes are still scarcely investigated. Indeed, acute intense exercise challenges the liver with increased reactive oxygen species (ROS) and inflammation onset, whereas regular training induces hepatic antioxidant and anti-inflammatory improvements. Acute and regular exercise protocols in combination with antioxidant and anti-inflammatory supplementation have been also tested to verify hepatic adaptations to exercise. Although positive results have been reported in some acute models, several studies have shown an increased exercise-related stress upon liver. A similar trend has been observed during training: while synergistic effects of training and antioxidant/anti-inflammatory supplementations have been occasionally found, others reported a blunting of relevant adaptations to exercise, following the patterns described in skeletal muscles. This review discusses current data regarding liver responses and adaptation to acute and regular exercise protocols alone or combined with antioxidant and anti-inflammatory supplementation. The understanding of the mechanisms behind these modulations is of interest for both exercise-related health and performance outcomes.     

Haploid embryogenesis in anther cultures of pigeon-pea (Cajanus cajan)

Summary Pollen embryogenesis and callus showing a wide range of ploidy is induced in the in vitro cultured anthers of pigeon-pea. A suspension of pollen from such anthers incubated in drop cultures on agar medium develops further to form embryoids and colonies of callus.     

Biochemical changes in developing seeds of pigeonpea (Cajanus cajan)

Soluble sugars, starch, soluble nitrogen and protein nitrogen were studied in developing seeds of 3 cultivars of pigeonpea. When expressed on a per seed basis soluble sugars increased up to 35 days after flowering and then declined slightly. Rapid starch accumulation was observed between 14 and 28 days after flowering. The levels of soluble nitrogen and protein nitrogen underwent rapid changes during the same period. Amino-acid composition of seed protein was also studied at different stages of maturation. Sodium dodecyl sulphate polyacrylamide gel electrophoresis of salt-soluble proteins revealed that seed storage globulins are formed after 14 days of flowering and do not change much during later stages of maturation.     

Influence of cadmium stress and arbuscular mycorrhizal fungi on nodule senescence in Cajanus cajan (L.) Millsp

Cadmium (Cd) causes oxidative damage and affects nodulation and nitrogen fixation process of legumes. Arbuscular mycorrhizal (AM) fungi have been demonstrated to alleviate heavy metal stress of plants. The present study was conducted to assess role of AM in alleviating negative effects of Cd on nodule senescence in Cajanus cajan genotypes differing in their metal tolerance. Fifteen day-old plants were subjected to Cd treatments--25 mg and 50 mg Cd per kg dry soil and were grown with and without Glomus mosseae. Cd treatments led to a decline in mycorrhizal infection (MI), nodule number and dry weights which was accompanied by reductions in leghemoglobin content, nitrogenase activity, organic acid contents. Cd supply caused a marked decrease in nitrogen (N), phosphorus (P), and iron (Fe) contents. Conversely, Cd increased membrane permeability, thiobarbituric acid reactive substances (TBARS), hydrogen peroxide (H2O2), and Cd contents in nodules. AM inoculations were beneficial in reducing the above mentioned harmful effects of Cd and significantly improved nodule functioning. Activities of superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) increased markedly in nodules of mycorrhizal-stressed plants. The negative effects of Cd were genotype and concentration dependent.     

Carbohydrate Metabolism in Drought-Stressed Leaves of Pigeonpea (Cajanus cajan)

Pigeonpea is a tropical grain-legume, which is highly dehydrationtolerant. The effect of drought stress on the carbohydrate metabolismin mature pigeonpea leaves was investigated by withholding waterfrom plants grown in very large pots (50 kg of soil). The moststriking feature of drought-stressed plants was the pronouncedaccumulation of D-pinitol (1D-3-methyl-chiro-inositol), whichincreased from 14 to 85 mg g^–1 dry weight during a 27d stress period. Concomitantly, the levels of starch, sucroseand the pinitol precursors myo-inositol and ononitol all decreasedrapidly to zero or near-zero in response to drought. The levelsof glucose and fructose increased moderately. Drought stressinduced a pronounced increase of the activities of enzymes hydrolysingsoluble starch (amylases) and sucrose (invertase and sucrosesynthase). The two anabolic enzymes sucrose phosphate synthase(sucrose synthetic pathway) and myo-inositol methyl transferase(pinitol synthetic pathway) also showed an increase of activityduring stress. These results indicate that pinitol accumulatedin pigeonpea leaves, because the carbon flux was diverted fromstarch and sucrose into polyols. Key words: Drought, polyols, pinitol, sucrose, starch, pigeonpea     

Hypoglycemic activity of Cajanus cajan (seeds) in mice.

The effect of roasted and unroasted seeds of C. cajan on serum glucose levels of normal and alloxan diabetic mice were studied. Single doses of unroasted seeds (60% and 80%) on administration to normal as well as alloxanized mice showed significant reduction in the serum glucose levels after 1-2 hr and a significant rise at 3 hr. In case of roasted seeds, on other hand there was a significant increase in serum glucose levels during 3 hr experimental period. It is therefore concluded that roasting of seeds at high temperature for an half hour period resulted in the total loss of hypoglycemic principle but not the hyperglycemic principle present in the seeds.     

Genetic patterns of domestication in pigeonpea (Cajanus cajan (L.) Millsp.) and wild Cajanus relatives

Pigeonpea (Cajanus cajan) is an annual or short-lived perennial food legume of acute regional importance, providing significant protein to the human diet in less developed regions of Asia and Africa. Due to its narrow genetic base, pigeonpea improvement is increasingly reliant on introgression of valuable traits from wild forms, a practice that would benefit from knowledge of its domestication history and relationships to wild species. Here we use 752 single nucleotide polymorphisms (SNPs) derived from 670 low copy orthologous genes to clarify the evolutionary history of pigeonpea (79 accessions) and its wild relatives (31 accessions). We identified three well-supported lineages that are geographically clustered and congruent with previous nuclear and plastid sequence-based phylogenies. Among all species analyzed Cajanus cajanifolius is the most probable progenitor of cultivated pigeonpea. Multiple lines of evidence suggest recent gene flow between cultivated and non-cultivated forms, as well as historical gene flow between diverged but sympatric species. Evidence supports that primary domestication occurred in India, with a second and more recent nested population bottleneck focused in tropical regions that is the likely consequence of pigeonpea breeding. We find abundant allelic variation and genetic diversity among the wild relatives, with the exception of wild species from Australia for which we report a third bottleneck unrelated to domestication within India. Domesticated C. cajan possess 75% less allelic diversity than the progenitor clade of wild Indian species, indicating a severe "domestication bottleneck" during pigeonpea domestication.     

Cardiovascular adaptations to mechanical overload

The cardiac changes resulting from mechanical overload of the left ventricle have been well documented and a variety of compensatory mechanisms described. These include a decrease in maximum velocity (V₀) of shortening in the absence of reduction in active tension (P₀), and a reversible decrease in myofibrillar adenosine triphosphatase activity resulting from isoenzymic shift from, predominantly, a form of myosin with high ATPase activity (V₁) to another with low (V₃). The thermodynamic advantage of the transition is the hypertrophied muscle possesses a more energy-efficient form of contraction. These reversible transitions resulted from altered gene expression of isoenzymic forms of myosin heavy chain. It must be borne in mind that the adaptational modifications just described appear to occur only in smaller animals such as the rat, that possesses several myosin isozymes. In large mammals it is mainly the V₃ form of myosin that is present, which does not change with altered contractile state. Responses of the large arteries to hypertension have been poorly studied. This is surprising when one recalls that degenerative disease of such vessels, that include the aorta, carotids and ileo-femoral arteries is almost an obligatory concomitant of hypertension. Such studies as have been carried out indicate that hyperplasia is specific for abdominal aortic stenosis while hypertrophy is found in aortic smooth muscle in rats with systemic hypertension. Mechanically, an increase in V₀ with no change in P₀ have been reported; an increase in myofibrillar ATPase activity was also reported. Though two myosin heavy chain isozymes have been found in aortic smooth muscle densitometry did not reveal any difference in distribution between tissues from control and hypertensive rats. The cause of the increased ATPase activity must be in increased phosphorylation of the muscles' 20,000 dalton light chain.     

Interrelations between hydraulic and mechanical stress adaptations in woody plants

The fields of plant water relations and plant biomechanics have traditionally been studied separately even though often the same tissues are responsible for water transport and mechanical support. There is now increasing evidence that hydraulic and mechanical adaptations may influence one another. We studied the changes in the hydraulic and mechanical properties of the wood along lateral roots of two species of buttressed trees. In these roots, the mechanical contstraints quantified by strain measurements are known to decrease distally. Further, we investigated the effect of mechanical loading on the vessel anatomy in these and four other species of tropical trees. We found that as the strain decreased, the wood became progressively less stiff and strong but the conductivity increased exponentially. This was reflected in that adaptations towards re-enforcing mechanically loaded areas resulted in xylem with fewer and smaller vessels. In addition a controlled growth experiment on three tree species showed that drought adaptation may results in plants with stronger and stiffer tissue. Our results indicate that hydraulic and mechanical stress adaptations may be interrelated, and so support recent studied suggesting that physiological responses are complex balances rather than pure optimisations.Key words: conductivity, modulus of elasticity, strain, tree ecophysiology, tropical trees, wood anatomy, yield stressIt is well known that the woody tissue of plants is responsible for carrying out several functions simultaneously, of which the two most important may be water transport and mechanical support. In spite of this, the fields of plant water relations and plant biomechanics have traditionally been studied separately (however, see refs. ^1–4). An increasing number of studies now indicate that there may be interrelations between hydraulic and mechanical stress adaptations, both in the form of positive interactions and trade-offs. In the former case, adaptations with respect to one of the two stresses positively affect the plants ability to withstand the other. Drought adaptation, for instance, is associated with an increase in the density of the tissue, which may result in stiffer and stronger wood.^5,6 Further, increasing the transectional area of the sapwood increases the conductivity as well as the rigidity of the plant and may occur as an adaptation to either drought- or mechanical stress.^7,8 In such cases, because of the importance of hydraulic sufficiency as well as adequate mechanical support, biomass must be partitioned to allow for both functions simultaneously even if this results in a surplus allocation with respect to one. In the case of trade-offs, the hydraulic or mechanical adaptations are detrimental to the plants'' capabilities with respect to the other. Within the woody tissue, for instance, larger and more numerous vessels increase the conductivity but may weaken the wood.^1–3,9–11 Analogously, hydraulic optimisation dictates an increase in the transectional sapwood area up though the plant, so that the summed area of all branches at a given height would be greater than that of the trunk¹² but mechanical optimisation a decrease.¹³ In the case of physiological parameters within which there are trade-offs, achieving adequate design whilst minimising biomass allocation becomes complex,⁴ and a number of possible but less optimal solutions exist.In our study,¹⁴ we looked at how mechanical as well as hydraulic parameters changed along the lateral roots of two tropical tree species, Tachigali melinonii and Xylopia nitida, both of which produce buttress roots. Along the roots of these species there is a strong distal decrease in the magnitude of the locally supported mechanical loads,¹⁵ making them ideal model organisms for investigating mechanical adaptation of the tissue and the impact this has on hydraulic parameters. We measured the density, conductivity, strength, stiffness, sapwood area and second moment of area at various points distally along the roots as well as in the lower trunk, and compared the values to those for strain. In both species, the strength and stiffness of the tissue decreased distally along the roots as the strain dropped, and the conductivity concurrently increased exponentially (Fig. 1). This appeared related to changes in the density of the wood; in both species, the density increased towards the bole and was positively correlated with mechanical properties but negatively with conductivity. As in previous studies, the distal most roots had a higher conductivity and lower stiffness and strength than that of the trunk. The proximal roots, however, had a lower conductivity but a greater strength and density. This indicates that the general pattern that roots have a higher conductivity than the stem cannot be explained by the water potential gradient from the soil to the leaves as often assumed, but instead by the differing mechanical requirements on these structures.Open in a separate windowFigure 1The modulus of elasticity in bending, E, (solid lines) and the specific conductivity, K_s, (dashed lines) of the woody tissue of the lateral roots shown as a function of the distance from the bole. The trend-lines are shown for the buttressed tree species Tachigali melinonii (black) and Xylopia nitida (grey), and are based on data presented in Christensen-Dalsgaard, et al, 2007a.These hydraulic and mechanical adaptations were well in accordance with anatomical adaptations seen for these as well as four other species from the same area representing two different rooting morphologies, for which the strain patterns differ. We found that the vessels were significantly smaller and less numerous in the xylem tissue of highly mechanically loaded compared to less loaded sections of the tree¹⁵ (Fig. 2). Further, the generally observed pattern^16,17 that vessel size increases radially from the pit to the bark was not observed throughout the structure.¹⁸ In the proximal parts of the buttress roots, which are highly mechanically loaded throughout growth and development, the vessels maintained the small size found at the pith throughout the transect. In the distal parts of the buttress roots, in which the mechanical loading increases during growth, the vessels decreased rather than increased in size from the pith to the bark.¹⁸ Since the adaptation of the xylem tissue towards re-enforcing highly strained areas appear associated with changes in the vessel anatomy reducing the conductivity, radial changes in vessel size may not only be a function of cambial ageing, but also influenced by changes in stresses during growth.Open in a separate windowFigure 2Tissue sections from the trunk and roots of the buttressed tree species Xylopia nitida, left, and the taproot anchored Oxandra asbeckii, right. All tissue sections are imaged at the same magnification; the width of each image is 4, 6 mm. In O. asbeckii, where the main rigid element resisting overturning is provided by the taproot and the lower bole, the trunk has smaller and fewer vessels than either of the root sections, as often described in litterature. In buttressed species such as X. nitida, on the other hand, where the area subjected to the greatest longitudinal strains and stresses is found in the proximal part of the buttress roots (marked in grey), this part of the root system is instead associated with fewer and smaller vessels than the trunk.In addition to hydraulic effects of mechanical adaptations, the converse may occur; hydraulic adaptations could affect the mechanical properties of the tissue. Drought adapted plants have a greater resistance to cavitation,^19,20 which appears related to an increased relative thickness of the conduit or the fibre cell walls,^5,21 and so an increased wood density.^5,22 However, since the mechanical properties are determined by the micro fibril angle (MFA) of the S2 cell wall layer as well as density,²³ the data from the few studies measuring the mechanical effects of hydraulic adaptation have not found consistent effects.^24,25 In these studies, complex natural systems were investigated making it difficult to distinguish between various climatic effects. We therefore grew seedlings from the three tree species Ochroma lagopus, Acacia karroo and Betula pendula under well-watered and droughted conditions, respectively, and measured the effect on the modulus of elasticity, the yield stress and the density (unpublished results). The stems of the droughted plants had a higher stiffness and strength than that of the well-watered plants. Only in O. lagopus, however, did the stems of the droughted seedlings have denser tissue than that of the well-watered seedlings; in B. pendula and A. karroo the mechanical differences were probably due to adaptations in the MFA instead. This was supported by that the density/elasticity ratio, which may be a good indicator of the MFA,^23,26 was significantly higher in the well-watered than in the droughted plants.Our work adds to the growing body of evidence indicating that there may be interrelations between hydraulic and mechanical stress adaptations.^{1–3,5,9–12,25} Modifications of the xylem towards mechanically reinforcing stressed or strained areas simultaneously impacted the conductive physiology of the trees studied, and drought adaptations resulted in stiffer and stronger tissue. This provides further evidence that in order to fully understand plant physiology and ecology it is necessary to consider the various functions of wood simultaneously and attempt to unravel causal relationships between e.g., the hydraulic and mechanical functioning of the tissue. Rather than being a matter of simple optimisations, adaptations towards one environmental stress will affect how plants adapt to the others. Because of the complexity of this balance, however, interrelations between parameters are not always found. For instance, mechanical strengthening due to hydraulic adaptations could reduce hydraulic effects of mechanical stresses; trees growing in climates with winter frost may be stronger due to larger amounts of sap- or heart wood and drought adaptation may result in a stiffer and stronger plant.^21,24,27 This could explain why the relatively clear trade-off between hydraulic and mechanical parameters found in this and other studies have not always been found in non-tropical species.^{21,24,27–29} Deepening our knowledge on how multi-factor adaptation balances are affected by climatic conditions and the signalling processes involved in these complex processes would aid our understanding of ecophysiological responses in plants.     

Somatic embryogenesis in cell suspension cultures of pigeonpea (Cajanus cajan)

Suspension cultures of calli derived from seedling leaf explants of Cajanus cajan L. var. Vamban-1 produced somatic embryos. The highest embryogenic frequency was induced on semisolid MS (Murashige and Skoog, 1962) medium supplemented with 6.78 μM 2,4-dichlorophenoxyacetic acid (2,4-D). The maximum frequency of somatic embryogenesis was observed when this callus was transferred to MS liquid medium supplemented with 4.52 μM 2,4-D. Further studies on ontogeny of somatic embryos showed that the cells destined to become somatic embryos divided into spherical proembryos. Subsequent divisions in the proembryo led to globular, heart and torpedo-shaped somatic embryos. The germination of somatic embryos occurred on auxin-free MS basal medium. Effects of various auxins, cytokinins and carbohydrates on induction and frequency of somatic embryogenesis were studied. A medium supplemented with 4.52 μM of 2,4-D and 87.64 mM sucrose was effective in inducing a higher frequency of somatic embryos, whereas cytokinin had no effect and led to recallusing of embryos. About 5–6% of embryos converted into plants. This revised version was published online in June 2006 with corrections to the Cover Date.     

ACC deaminase positive Enterobacter-mediated mitigation of salinity stress,and plant growth promotion of Cajanus cajan: a lab to field study

Salinity is a major abiotic stress that negatively impacts plant health and soil microbiota. ACC (1-aminocyclopropane carboxylic acid) deaminase producing microorganisms act as natural stress busters that protect plants from different kinds of stresses. The study focused on the isolation of potent, indigenous, multi-trait ACC deaminase producers. The shortlisted ACC deaminase producers were checked for their ability to promote growth of Cajanus cajan, and mitigate stress under laboratory conditions followed by validation of their potency in naturally saline field conditions. Physiological stress markers were assessed to evaluate the impact of salinity in plants treated with ACC deaminase producer, compared to controls. Further, the contribution of ACC deaminase in stress mitigation was demonstrated by using a chemical inhibitor for ethylene biosynthesis. This study presents a polyphasic approach, transitioning from the rhizospheric soil to the laboratory to validation in the field, and puts forth a promising eco-friendly alternative for sustainable agriculture.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12298-021-01031-0.     

Sex allocation and mating systems in pigeonpea,Cajanus cajan (Fabaceae)

Sex allocation theory predicts that: (1) resources allocated to androecium should decrease with an increase in selfing, (2) a decrease in androecium biomass should be accompanied by an increase in the biomass of pistils, and (3) a decrease in androecium biomass should be coupled with a decrease in flower size, specifically corolla biomass. Another predicted change in reproductive traits associated with variation in selfing concerns seed to ovule ratios, but does not directly stem from sex allocation theory. It has been postulated that seed to ovule ratios should be positively correlated with the amount of selfing. These predictions were tested for six accessions of pigeonpea,Cajanus cajan L., that differed in selfing rates. The results were remarkably in accordance with the predictions. We conclude that sex allocation theory provides a powerful tool to understand the evolution of many reproductive traits in plants.     

Intracellular Distribution of Heat Shock Proteins in Pigeon Pea (Cajanus cajan)

The proteins synthesized In response to higher temperature In pigeon pea (Cajanus cajan) plants have been studied with respect to their Intracellular localization using root tissue. The heat shock proteins (hsps) of 18, 20, 22 and 24 kD were found to be associated with mitochondrial and membrane fractions, while the 60, 70 and 81 kD hsps were found In the soluble fraction. No evidence for the presence of hsps among the proteins synthesized in organello by isolated mitochondria could be obtained. Low molecular weight hsps (18, 20, 22 and 24 kD) were found associated with mitochondria Isolated from the heat shocked tissue suggesting that these hsps may have been transported post-translationally into mitochondria.