MPH1 is a thylakoid membrane protein involved in protecting photosystem II from photodamage in land plants

Photosystem II (PSII) is highly susceptible to photoinhibition caused by environmental stimuli such as high light; therefore plants have evolved multifaceted mechanisms to efficiently protect PSII from photodamage. We previously published data suggesting that Maintenance of PSII under High light 1 (MPH1, encoded by AT5G07020), a PSII-associated proline-rich protein found in land plants, participates in the maintenance of normal PSII activity under photoinhibitory stress. Here we provide additional evidence for the role of MPH1 in protecting PSII against photooxidative damage. Two Arabidopsis thaliana mutants lacking a functional MPH1 gene suffer from severe photoinhibition relative to the wild-type plants under high irradiance light. The mph1 mutants exhibit significantly decreased PSII quantum yield and electron transport rate after exposure to photoinhibitory light. The mutants also display drastically elevated photodamage to PSII reaction center proteins after high-light treatment. These data add further evidence that MPH1 is involved in PSII photoprotection in Arabidopsis. MPH1 homologs are found across phylogenetically diverse land plants but are not detected in algae or prokaryotes. Taken together, these results suggest that MPH1 protein began to play a role in protecting PSII against excess light following the transition from aquatic to terrestrial conditions.     

Methylphenidate exposure induces dopamine neuron loss and activation of microglia in the basal ganglia of mice

Background

Methylphenidate (MPH) is a psychostimulant that exerts its pharmacological effects via preferential blockade of the dopamine transporter (DAT) and the norepinephrine transporter (NET), resulting in increased monoamine levels in the synapse. Clinically, methylphenidate is prescribed for the symptomatic treatment of ADHD and narcolepsy; although lately, there has been an increased incidence of its use in individuals not meeting the criteria for these disorders. MPH has also been misused as a “cognitive enhancer” and as an alternative to other psychostimulants. Here, we investigate whether chronic or acute administration of MPH in mice at either 1 mg/kg or 10 mg/kg, affects cell number and gene expression in the basal ganglia.

Methodology/Principal Findings

Through the use of stereological counting methods, we observed a significant reduction (∼20%) in dopamine neuron numbers in the substantia nigra pars compacta (SNpc) following chronic administration of 10 mg/kg MPH. This dosage of MPH also induced a significant increase in the number of activated microglia in the SNpc. Additionally, exposure to either 1 mg/kg or 10 mg/kg MPH increased the sensitivity of SNpc dopaminergic neurons to the parkinsonian agent 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Unbiased gene screening employing Affymetrix GeneChip® HT MG-430 PM revealed changes in 115 and 54 genes in the substantia nigra (SN) of mice exposed to 1 mg/kg and 10 mg/kg MPH doses, respectively. Decreases in the mRNA levels of gdnf, dat1, vmat2, and th in the substantia nigra (SN) were observed with both acute and chronic dosing of 10 mg/kg MPH. We also found an increase in mRNA levels of the pro-inflammatory genes il-6 and tnf-α in the striatum, although these were seen only at an acute dose of 10 mg/kg and not following chronic dosing.

Conclusion

Collectively, our results suggest that chronic MPH usage in mice at doses spanning the therapeutic range in humans, especially at prolonged higher doses, has long-term neurodegenerative consequences.     

Improving the Secretion of a Methyl Parathion Hydrolase in Pichia pastoris by Modifying Its N-Terminal Sequence

Pichia pastoris is commonly used to express and secrete target proteins, although not all recombinant proteins can be successfully produced. In this study, we used methyl parathion hydrolase (MPH) from Ochrobactrum sp. M231 as a model to study the importance of the N-terminus of the protein for its secretion. While MPH can be efficiently expressed intracellularly in P. pastoris, it is not secreted into the extracellular environment. Three MPH mutants (N₆₆-MPH, D₁₀-MPH, and N₉-MPH) were constructed through modification of its N-terminus, and the secretion of each by P. pastoris was improved when compared to wild-type MPH. The level of secreted D₁₀-MPH was increased to 0.21 U/mL, while that of N₉-MPH was enhanced to 0.16 U/mL. Although N₆₆-MPH was not enzymatically active, it was secreted efficiently, and was identified by SDS-PAGE. These results demonstrate that the secretion of heterologous proteins in P. pastoris may be improved by modifying their N-terminal structures.     

Molecular Paleoparasitological Hybridization Approach as Effective Tool for Diagnosing Human Intestinal Parasites from Scarce Archaeological Remains

Paleoparasitology is the science that uses parasitological techniques for diagnosing parasitic diseases in the past. Advances in molecular biology brought new insights into this field allowing the study of archaeological material. However, due to technical limitations a proper diagnosis and confirmation of the presence of parasites is not always possible, especially in scarce and degraded archaeological remains. In this study, we developed a Molecular Paleoparasitological Hybridization (MPH) approach using ancient DNA (aDNA) hybridization to confirm and complement paleoparasitological diagnosis. Eight molecular targets from four helminth parasites were included: Ascaris sp., Trichuris trichiura, Enterobius vermicularis, and Strongyloides stercoralis. The MPH analysis using 18^th century human remains from Praça XV cemetery (CPXV), Rio de Janeiro, Brazil, revealed for the first time the presence E. vermicularis aDNA (50%) in archaeological sites of Brazil. Besides, the results confirmed T. trichiura and Ascaris sp. infections. The prevalence of infection by Ascaris sp. and E. vermicularis increased considerably when MPH was applied. However, a lower aDNA detection of T. trichiura (40%) was observed when compared to the diagnosis by paleoparasitological analysis (70%). Therefore, based on these data, we suggest a combination of Paleoparasitological and MPH approaches to verify the real panorama of intestinal parasite infection in human archeological samples.     

Development of a Physiologically Based Model to Describe the Pharmacokinetics of Methylphenidate in Juvenile and Adult Humans and Nonhuman Primates

The widespread usage of methylphenidate (MPH) in the pediatric population has received considerable attention due to its potential effect on child development. For the first time a physiologically based pharmacokinetic (PBPK) model has been developed in juvenile and adult humans and nonhuman primates to quantitatively evaluate species- and age-dependent enantiomer specific pharmacokinetics of MPH and its primary metabolite ritalinic acid. The PBPK model was first calibrated in adult humans using in vitro enzyme kinetic data of MPH enantiomers, together with plasma and urine pharmacokinetic data with MPH in adult humans. Metabolism of MPH in the small intestine was assumed to account for the low oral bioavailability of MPH. Due to lack of information, model development for children and juvenile and adult nonhuman primates primarily relied on intra- and interspecies extrapolation using allometric scaling. The juvenile monkeys appear to metabolize MPH more rapidly than adult monkeys and humans, both adults and children. Model prediction performance is comparable between juvenile monkeys and children, with average root mean squared error values of 4.1 and 2.1, providing scientific basis for interspecies extrapolation of toxicity findings. Model estimated human equivalent doses in children that achieve similar internal dose metrics to those associated with pubertal delays in juvenile monkeys were found to be close to the therapeutic doses of MPH used in pediatric patients. This computational analysis suggests that continued pharmacovigilance assessment is prudent for the safe use of MPH.     

Generation of haploid embryonic stem cells from Macaca fascicularis monkey parthenotes

Recent success in the derivation of haploid embryonic stem cells (haESCs) from mouse via parthenogenesis and androgenesis has enabled genetic screening in mammalian cells and generation of gene-modified animals. However, whether haESCs can be derived from primates remains unknown. Here, we report the derivation of haESCs from parthenogenetic blastocysts of Macaca fascicularis monkeys. These cells, termed as PG-haESCs, are pluripotent and can differentiate to cells of three embryonic germ layers in vitro or in vivo. Interestingly, the haploidy of one monkey PG-haESC line (MPH1) is more stable compared with that of the other one (MPH2), as shown by the existence of haploid cells for more than 140 days without fluorescence-activated cell sorting (FACS) enrichment of haploid cells. Importantly, transgenic monkey PG-haESC lines can be generated by lentivirus- and piggyBac transposon-mediated gene transfer. Moreover, genetic screening is feasible in monkey PG-haESCs. Our results demonstrate that PG-haESCs can be generated from monkeys, providing an ideal tool for genetic analyses in primates.     

Differential effects of methylphenidate and cocaine on GABA transmission in sensory thalamic nuclei

Methylphenidate (MPH) is widely used to treat children and adolescents diagnosed with attention deficit/hyperactivity disorder. Although MPH shares mechanistic similarities to cocaine, its effects on GABAergic transmission in sensory thalamic nuclei are unknown. Our objective was to compare cocaine and MPH effects on GABAergic projections between thalamic reticular and ventrobasal (VB) nuclei. Mice (P18‐30) were subjected to binge‐like cocaine and MPH acute and sub‐chronic administrations. Cocaine and MPH enhanced hyperlocomotion, although sub‐chronic cocaine‐mediated effects were stronger than MPH effects. Cocaine and MPH sub‐chronic administration altered paired‐pulse and spontaneous GABAergic input differently. The effects of cocaine on evoked paired‐pulse GABA‐mediated currents changed from depression to facilitation with the duration of the protocols used, while MPH induced a constant increase throughout the administration protocols. Thalamic reticular nucleus GAD67 and VB Ca_V3.1 protein levels were measured using western blot to better understand their link to increased GABA release. Both proteins were increased by sub‐chronic administration of cocaine. MPH showed effects on GABAergic transmission that seems less disruptive than cocaine. Unique effects of cocaine on postsynaptic VB calcium currents might explain deleterious cocaine effects on sensory thalamic nuclei. These results suggest that cocaine and MPH produced distinct presynaptic alterations on GABAergic transmission.     

Chiral analysis of methylphenidate and dextromoramide by capillary electrophoresis

Capillary electrophoretic methods have been developed to separate the enantiomers of methylphenidate (MPH) and dextromoramide. For MPH separation was achieved with heptakis (2,6-di-O-methyl)-β-cyclodextrin (DMCD) as chiral selector in a 100 mM phosphoric acid buffer adjusted to pH 3.0 with triethanolamine. Commercial samples of d,l-erytho-MPH HCl and d,l-threo-MPH HCl were analysed using the method. There was no evidence of the presence of d,l-threo-MPH HCl in d,l-erytho-MPH HCl and vice versa. The ratio of the enantiomers was determined for each diastereoisomer. Hydroxypropyl-β-cyclodextrin was the chiral selector of choice for the chiral separation of the enantiomers of moramide. The separation which gave a resolution of about 3.5 was achieved in 4 min using only a 6 cm of length of capillary. In a sample of dextro-R-moramide tartrate only a small quantity (4.9% w/w) of levo-S-moramide was detected with this method.     

Molecular Analysis of Maltotriose Active Transport and Fermentation by Saccharomyces cerevisiae Reveals a Determinant Role for the AGT1 Permease

Incomplete and/or sluggish maltotriose fermentation causes both quality and economic problems in the ale-brewing industry. Although it has been proposed previously that the sugar uptake must be responsible for these undesirable phenotypes, there have been conflicting reports on whether all the known α-glucoside transporters in Saccharomyces cerevisiae (MALx1, AGT1, and MPH2 and MPH3 transporters) allow efficient maltotriose utilization by yeast cells. We characterized the kinetics of yeast cell growth, sugar consumption, and ethanol production during maltose or maltotriose utilization by several S. cerevisiae yeast strains (both MAL constitutive and MAL inducible) and by their isogenic counterparts with specific deletions of the AGT1 gene. Our results clearly showed that yeast strains carrying functional permeases encoded by the MAL21, MAL31, and/or MAL41 gene in their plasma membranes were unable to utilize maltotriose. While both high- and low-affinity transport activities were responsible for maltose uptake from the medium, in the case of maltotriose, the only low-affinity (K_m, 36 ± 2 mM) transport activity was mediated by the AGT1 permease. In conclusion, the AGT1 transporter is required for efficient maltotriose fermentation by S. cerevisiae yeasts, highlighting the importance of this permease for breeding and/or selection programs aimed at improving sluggish maltotriose fermentations.     

Temporal Regulation of the Metabolome and Proteome in Photosynthetic and Photorespiratory Pathways Contributes to Maize Heterosis

Heterosis or hybrid vigor is widespread in plants and animals. Although the molecular basis for heterosis has been extensively studied, metabolic and proteomic contributions to heterosis remain elusive. Here we report an integrative analysis of time-series metabolome and proteome data in maize (Zea mays) hybrids and their inbred parents. Many maize metabolites and proteins are diurnally regulated, and many of these show nonadditive abundance in the hybrids, including key enzymes and metabolites involved in carbon assimilation. Compared with robust trait heterosis, metabolic heterosis is relatively mild. Interestingly, most amino acids display negative mid-parent heterosis (MPH), i.e., having lower values than the average of the parents, while sugars, alcohols, and nucleoside metabolites show positive MPH. From the network perspective, metabolites in the photosynthetic pathway show positive MPH, whereas metabolites in the photorespiratory pathway show negative MPH, which corresponds to nonadditive protein abundance and enzyme activities of key enzymes in the respective pathways in the hybrids. Moreover, diurnally expressed proteins that are upregulated in the hybrids are enriched in photosynthesis-related gene-ontology terms. Hybrids may more effectively remove toxic metabolites generated during photorespiration, and thus maintain higher photosynthetic efficiency. These metabolic and proteomic resources provide unique insight into heterosis and its utilization for high yielding maize and other crop plants.     

Stress-Induced In Vivo Recruitment of Human Cytotoxic Natural Killer Cells Favors Subsets with Distinct Receptor Profiles and Associates with Increased Epinephrine Levels

Background

Acute stress drives a ‘high-alert’ response in the immune system. Psychoactive drugs induce distinct stress hormone profiles, offering a sought-after opportunity to dissect the in vivo immunological effects of acute stress in humans.

Methods

3,4-methylenedioxymethamphetamine (MDMA), methylphenidate (MPH), or both, were administered to healthy volunteers in a randomized, double-blind, placebo-controlled crossover-study. Lymphocyte subset frequencies, natural killer (NK) cell immune-phenotypes, and changes in effector function were assessed, and linked to stress hormone levels and expression of CD62L, CX3CR1, CD18, and stress hormone receptors on NK cells.

Results

MDMA/MPH > MDMA > MPH robustly induced an epinephrine-dominant stress response. Immunologically, rapid redistribution of peripheral blood lymphocyte-subsets towards phenotypically mature NK cells occurred. NK cytotoxicity was unaltered, but they expressed slightly reduced levels of the activating receptor NKG2D. Preferential circulation of mature NK cells was associated with high epinephrine receptor expression among this subset, as well as expression of integrin ligands previously linked to epinephrine-induced endothelial detachment.

Conclusion

The acute epinephrine-induced stress response was characterized by rapid accumulation of mature and functional NK cells in the peripheral circulation. This is in line with studies using other acute stressors and supports the role of the acute stress response in rapidly mobilizing the innate immune system to counteract incoming threats.     

Methylphenidate hydrochloride increases energy expenditure in healthy adults

Objective: To examine the effects of methylphenidate hydrochloride (MPH) on resting energy expenditure (REE) and postprandial energy expenditure (PEE) and substrate partitioning. Methods and Procedures: Seven healthy men and seven healthy women participated in this double‐blind, randomized, placebo‐controlled, crossover study. MPH (0.5 mg/kg) or placebo was administered orally in the fasting state, 60 min before a REE measurement, and 90 min before a standardized breakfast of ～650 kcal. REE, PEE, and respiratory exchange ratio (RER) were obtained from indirect calorimetry. Body composition was measured using DEXA. Vital signs (blood pressure (BP) and heart rate (HR)) were assessed pre‐ and post‐administration of MPH or placebo in every session. Results: During the, MPH condition, REE increased over values observed during the placebo session (7%, P < 0.001). No changes in fasting RER were noted. Although PEE continually decreased with time as expected, MPH treatment resulted in significantly greater PEE values at 90 min (5%, P < 0.01). No significant effects of MPH were found for vital signs (HR, systolic, and diastolic BP). Discussion: MPH causes a significant increase in both REE and PEE without the significant changes in HR and BP that are commonly associated with psychostimulant use.     

Expression of Methyl Parathion Hydrolase in <Emphasis Type="Italic">Pichia pastoris</Emphasis>

In the Pichia pastoris expression system, increasing the copy number of the expression cassette often has the effect of increasing the amount of protein expressed. To improve the expression level of methyl parathion hydrolase (MPH), we constructed two integration vectors with four and eight direct repeats of the expression cassette using an in vitro multimerization approach. After two successive integrations, at least 12 copies of the MPH expression cassette were integrated into the P. pastoris chromosome. Under shake-flask conditions, over 55 mg active MPH/l was secreted into the medium by the multicopy clones. The extracellular enzyme activity was about 10-fold higher for the multicopy clones than for clones containing a single copy of the gene. Further investigations revealed that the multicopy MPH expression cassette could remain stably integrated and functional over five generations. Note that the expression vector pRF constructed in our study can be not only used to construct multiple copies of the expression cassette in vitro, but also integrated into the P. pastoris genome without introducing any antibiotic resistance gene, which is desirable for production of biotherapeutic proteins.     

D-methylphenidate and D,L-methylphenidate are not developmental toxicants in rats and rabbits

BACKGROUND: D ,L ‐threo‐Methylphenidate (D ,L ‐MPH) is marketed currently for attention deficit hyperactivity disorder in children. D ‐threo‐methylphenidate (dexmethylphenidate; D ‐MPH) is a refined formulation of D ,L ‐methylphenidate containing only the active enantiomer and was recently approved in the U.S. for the same condition. D ‐Methylphenidate has been shown to be efficacious in patients at half the dose of D ,L ‐MPH with a potentially improved therapeutic profile. The developmental toxicity of both compounds was determined and compared in rats and rabbits according to current International Conference on Harmonization (ICH) guidelines. METHODS: Groups of pregnant rats were orally dosed twice daily 6 hr apart from Days 7 to 17 of presumed gestation (DG 7–17) for total daily doses of 2, 6 and 20 mg/kg D ‐MPH and 40 mg/kg D ,L ‐MPH. Groups of presumed pregnant rabbits were similarly dosed from DG 6 to 18 for total daily doses of 4, 20 and 100 mg/kg D ‐MPH and 200 mg/kg D ,L ‐MPH. Control groups for both studies were given water vehicle. Comprehensive clinical and developmental measurements were made. Satellite groups of animals were included in the main rat and rabbit studies for toxicokinetic assessment. RESULTS: No drug‐related mortality was seen in the F0 rats and rabbits. The number of rats with repetitive pawing, dilated pupil and aggression was significantly greater for the 40 mg/kg D ,L ‐MPH compared to the 20 mg/kg D ‐MPH dosed rats. Maternal body weight and body weight gain were significantly reduced for both D ‐MPH and D ,L ‐MPH groups compared to control. Maternal reproductive and litter parameters were unaffected by both drugs. No gross external, soft tissue, or skeletal alterations related to both compounds were seen in the fetuses. In rabbits, head‐bobbing and hyperpnea were significantly greater for the 200 mg/kg D ,L ‐MPH compared to 100 mg/kg D ‐MPH. No other maternal or fetal effects related to both compounds were seen. Exposure to D ‐MPH (as assessed by AUC) showed no teratogenic effects at exposures of up to 5.6 and 1.7 times for the rat and rabbit respectively compared to children taking the maximum therapeutic dose of 20 mg/day (10 mg twice a day). No teratogenic effects were seen for D ,L ‐MPH in rat and rabbit at exposures of up to 3.7 to 11.7 times that of the maximum therapeutic pediatric dose of 60 mg/day. CONCLUSIONS: Rats and rabbits dosed with D ,L ‐MPH exhibited significantly greater incidence of maternal clinical observations at twice the dose of D ‐MPH. Both D ‐MPH and D ,L ‐MPH were not teratogenic in rats and rabbits at higher exposure levels compared to humans. © 2003 Wiley‐Liss, Inc.     

Overexpression of methyl parathion hydrolase and its application in detoxification of organophosphates

The coding region of mpd gene corresponding to mature methyl parathion hydrolase (MPH) was heterologously overexpressed in Escherichia coli BL21 (DE3) by using pET expression system. The lactose-induced expression yield of MPH is increased 2-fold compared with IPTG as inducer. Furthermore, it was found that specific activity of MPH increased 48% by reducing the induction temperature to 22°C. The addition of 25 mM lactose at 22°C, the MPH activity of fermentation broth had a specific activity of 1.4 × 10⁴ U/mg protein. Plasmid was no significant decrease in the modified medium. The optimal pH and temperature of MPH were 8.0 and 30°C, respectively. Over a period of 5 months, the dried cells showed no significant decrease in the activity of the detoxifying enzymes. The crude enzymes in 50 mM citrate-phosphate buffer (pH 8.0) were able to degrade about 98% of the organophosphate pesticides sprayed on cabbage. The detoxification efficiency was superior to that of the treatments of water, detergent, and a commercially available enzyme product. Additionally, the products of pesticide hydrolysis generated by treatment with the enzyme extract were determined to be virtually nontoxic.     

Developmental toxicity assessment of d,l-methylphenidate and d-methylphenidate in rats and rabbits

BACKGROUND: Previous investigations reported no teratogenicity for methylphenidate (MPH). These studies investigated potential teratogenicity of d‐MPH and d,l‐MPH as commitments to the FDA. METHODS: Rabbits received 15, 50, 150 mg/kg/day (mkd) d‐MPH or 20, 60, 200, 300 mkd d,l‐MPH on gestation days 7–20. Rats received 2.5, 10, 40 mkd d‐MPH, or 7, 25, 75, 80 mkd d,l‐MPH on gestation days 6–17. RESULTS: d‐MPH—In rabbits, mortality occurred at 150 mkd. Dilated pupils, increased activity, biting/chewing, respiration, and salivation occurred at ≥15 mkd in rabbits and ≥10 mkd in rats. Decreased food consumption occurred at 40 mkd in rats. Decreased body weight parameters occurred at 150 mkd in rabbits and ≥10 mkd in rats. There were no fetal findings in rabbits. In rats, skeletal variations occurred at 40 mkd. d,l‐MPH—In rabbits, mortality occurred at ≥200 mkd. Dilated pupils, increased activity, biting/chewing, respiration, and salivation occurred at ≥20 mkd in rabbits and ≥25 mkd in rats. Decreased food consumption occurred at ≥200 mkd in rabbits and ≥25 mkd in rats. Decreased body weight parameters occurred at ≥200 mkd in rabbits and ≥25 mkd in rats. In rabbits, two fetuses (separate litters) had spina bifida and malrotated hindlimbs at 200 mkd. In rats, skeletal variations occurred at ≥75 mkd. CONCLUSIONS: There was no teratogenicity with d‐MPH. There was a low teratogenic risk with d,l‐MPH in only the rabbit. Higher C_max may explain differences in results from previous studies. Birth Defects Res (Part B) 83:489‐501, 2008. © 2008 Wiley‐Liss, Inc.     

Plant height as a simple predictor of the root to shoot ratio: Evidence from alpine grasslands on the Tibetan Plateau

Question: Since increases in altitude and grazing intensity generally result in decreases in height growth of alpine grasslands, plant height may integrate effects of environmental stress and grazing disturbance and provide better assessments of the variation in root: shoot (R: S) biomass ratio than other variables. However, it is unclear if there is a general relationship between plant height and R:S ratio across grassland ecosystems. Such knowledge would be helpful for root biomass estimation in grasslands. Location: An altitudinal transect in the Gonghe Basin (2880–4040 m a.s.l.), northeast Tibetan plateau. Methods: We measured standing biomass both above‐ground and below‐ground, maximum plant height (MPH) and soil variables across 43 plots. Results: Climatic variables explained the variations in MPH and R: S ratio of undegraded grasslands better than soil variables (46–50% vs < 19%), while those of degraded grasslands generally showed insignificant correlations with climatic and soil variables. There was a general relationship between R: S ratio and MPH (negative, R²= 0.76, P< 0.001) across degraded and undegraded grasslands. The relationship was used to predict R: S ratio in 13 additional plots in steppe grasslands of Inner Mongolia, and good agreement of expected and observed values has been found (R²= 0.87, P < 0.001). Conclusions: MPH, that is relatively easy to measure, can be used to predict R:S ratio at plot to regional scales. It is promising to develop a new method for large‐scale estimation of root biomass in grasslands using MPH and shoot biomass avoiding tedious procedures of physical measuring of above and below‐ground biomass.     

Utilization of mustard waste isolates for improved production of astaxanthin by Xanthophyllomyces dendrorhous

Astaxanthin production in the wild strain Xanthophyllomyces dendrorhous TISTR 5730 was investigated using different mustard waste media, including mustard waste residue extract (MRE), mustard waste residue hydrolysate (MRH), mustard waste precipitated extract (MPE), and mustard waste precipitated hydrolysate (MPH). The growth of X. dendrorhous and the production of astaxanthin were dependent on the type and initial concentrations of mustard waste media. The MPH medium was the best substrate resulting in yields of biomass and astaxanthin of 19.6 g/L and 25.8 mg/L, respectively, under optimal conditions. MPH medium improved astaxanthin production 11-fold compared to the commonly used commercial yeast malt medium, and 1.3–2.1-fold compared to other mustard waste media.     

Population-based diallel analyses among nine historically recognized alfalfa germplasms

Identification of heterotic groups and patterns among breeding populations provides fundamental information to help plant breeders more knowledgeably manipulate heterosis. A diallel analysis was conducted among nine alfalfa (Medicago sativa L.) germplasms, commonly referred to as African, Chilean, Flemish, Indian, Ladak, M. falcata, M. varia, Peruvian, and Turkistan, which represent a significant proportion of the genetic diversity present in US cultivars. Heterotic responses were determined by evaluating forage yield of the germplasms and their 36 half-diallel hybrids in seeded plots that were harvested five times in each of 2 years. Commercially acceptable yields were obtained from some hybrids of unimproved parents, where at least one parent was adapted to the study environment. Variation among crosses was attributed primarily to general combining ability (GCA) effects; however, specific combining ability effects were also significant. GCA estimates for African, Chilean and Peruvian were positive, while those for Ladak, M. falcata, and M. varia were negative. Estimates for variety heterosis effects were positive for Peruvian and M. falcata and negative for Indian and M. varia. Significant mid-parent heterosis [(MPH) range of –21% to 55%] and high-parent heterosis [(HPH) range of –33% to 23%] was detected. M. falcata hybrids exhibited the highest MPH values. However, this likely reflects the poor yield of M. falcata per se in the study environment and consequently, low MPH values. Peruvian hybrids demonstrated the highest cross mean performance, significant positive MPH in all crosses, and positive HPH in five out of eight crosses. The results indicate that Peruvian should be recognized as a heterotic group. Alfalfa breeders may wish to explore opportunities for heterotic yield gains that are likely to exist in hybrids between the Peruvian germplasm and elite breeding populations, in particular, those adapted to the southwestern United States. MPH results suggest that alfalfa breeders may have capitalized on the heterotic response between Flemish and M. varia during past development of alfalfa synthetics adapted to the central and northern latitudes of the United States.