Comparison of two methods for determining in vitro intestinal absorption of nutrients using rats fed different diets

Absorption of sucrose, glucose, leucine and aspartate was studied using intestinal everted sac of rats fed on french bean diets namely PDR-14, HUR-137 and HUR-15 using casein as a control. Absorption of nutrients was monitored spectrophotometrically and by¹⁴C radio assay of metabolites using scientillation counting. The absorption pattern of amino acids was found to be similar but of glucose and sucrose differed. Glucose was found to be more absorbed than sucrose in spectrophotometer assay and the pattern reversed in radio assay. Absorption of sucrose and leucine were higher by rats fed on HUR-137 diet and similarly, more aspartate was absorbed when fed on HUR-15 diet as demonstrated by both the methods. Rats fed on HUR-137 diet exhibited higher glucose absorption as shown by spectrophotometric assay, but rats on HUR-15 diet by radio assay. Absorption of nutrients differed significantly between casein and french beans.     

Influence of plant growth regulators on some enzymes of nitrogen assimilation in mustard seedlings

Effect of plant growth regulators (PGR) viz. gibberellic acid (GA), kinetin (KN), and abscissic acid (ABA) were investigated on growth and activities of nitrate reductase (NR) and glutamine synthetase (GS) in mustard (Brassica juncea) seedlings. All the PGRs tested promoted in vivo NR activity in cotyledons, but the magnitude differed with different treatments. Cytosolic GS in root and hypocotyl was promoted by GA treatment and inhibited by ABA and KN treatments although the latter showed slight promotion initially in hypocotyl; the trend was not clear in cotyledons. Determination of K_m value of GS extracted from 96‐h‐old cotyledons recorded lower K_m value in GA treatment (2.5 mM), while it increased in ABA treatment (4.35 mM), There was little change in K_m value in KN (3.03 mM) treatment. The kinetics of GS enzyme in cotyledons of different treatments showed marked variation in V_max . Both GA and ABA treatments inhibited GS activity while no significant effect by KN treatment was observed. It is argued that GA treatment inhibits chloroplastic GS (an enzyme which has higher K_m value), while ABA‐induced inhibition may not be specific to cytosolic or chloroplastic isoforms. Kinetin treatment was ineffective in promoting or inhibiting GS activity in cotyledons. The above conclusion is further supported by chloroplastic pigment data where inhibition is recorded in all the PGRs tested.     

Effect of Plant Growth Regulators and Different Nitrogen Sources on Glutamate Dehydrogenase Activity

Radish (Raphanus sativus L.) seedlings, treated with various plant growth regulators (PGRs) [viz. kinetin (KiN), gibberellic acid (GA), and abscisic acid(ABA)] were exposed to different nitrogen (N) sources in light and dark condition, and aminative (NADH) and deaminative (NAD⁺) glutamate dehydrogenase (GDH) activities were measured in cotyledons. A differential effect of nitrogen sources, plant growth regulators, and light or dark condition was observed in all the treatments. The NAD‐GDH (deaminating) activity in radish seedlings was only about 10% of aminating activity (irrespective of the PGR treatment). Except with abscisic acid, in all other treatments, either NAD‐GDH or NADH‐GDH activities were more in dark than in light. The amination and deamination reactions also showed different ratios of activity under different N sources (KNO3, NH₄Cl and NH₄NO₃). These data suggest the presence of isoenzymes or conformers of GDH, specific for each tissue, whose activities vary depending on the physiological condition of the tissue. Different energy status of the seedlings during light or dark condition or with PGR treatments may affect the GDH activity differently.     

Improving androgenesis‐mediated doubled haploid production efficiency of FCV tobacco (Nicotiana tabacum L.) through in vitro colchicine application

Production of doubled haploid plants through androgenesis in flue‐cured Virginia (FCV) tobacco is a promising and convenient alternative to conventional selfing techniques for the generation of absolute homozygous lines. Here, we show a robust in vitro haploid and doubled haploid development protocol in FCV tobacco with major emphasis on improving the efficiency of chromosome doubling using in vitro colchicine treatment. We used five FCV tobacco hybrids for comparison of colchicine treatments. The anther culture response varied with developmental stages of the buds, and the highest response was observed in stage 2 buds. The effect of cold pretreatment was significant, and 4 days of pretreatment was optimum for gametic embryogenesis. Among the methods used for determining the ploidy status of plants, flow cytometry was found to be easy, fast and reliable for high‐throughput screening of haploids. Doubled haploids regeneration percentage varied from 6.77 to 11.95 in in vivo treatment, while the range of variation was 22.11% to 28.40% in in vitro colchicine treatment. We observed a pronounced increase in plant survival and the proportion of doubled haploid plants in in vitro treatment compared with the standard in vivo approach.     

Multivariate Analyses for Determining the Association of Field Porcine Fertility With Sperm Motion Traits Analysed by Computer‐Assisted Semen Analysis and With Sperm Morphology

This study investigates the association of semen traits with boar fertility. The fertility outcome (farrowing rate – FR and total piglets born – TB) of 14 boars was obtained from a field trial conducted during 10 week of breeding period on a commercial farm using multiparous sows (n = 948) through single‐sire mating with 2 × 10⁹ motile sperm cells per artificial insemination (AI) dose. Sperm motion parameters, evaluated with computer‐assisted semen analysis system in raw and stored semen at 17°C for 240 h, in addition to morphological sperm defects, measured on the collection day, were included in the analysis to determine which semen traits were important to discriminate the fertility potential of ejaculates from these boars. The data underwent multivariate cluster, canonical and discriminant analyses. Four clusters of boars were formed based on fertility outcome. One boar, with the lowest FR and TB values (89.7% and 11.98), and two boars, with the highest FR and TB values (97.8% and 14.16), were placed in different clusters. The other boars were separated in two distinct clusters (four and seven boars), including boars with intermediate TB (12.64 and 13.22) but divergent values for FR (95.9% vs 91.8%). Semen traits with higher discriminatory power included total motility, progressive motility, amplitude of lateral head displacement and cytoplasmatic droplets. Through multivariate discriminant analysis, more than 80% of the 140 ejaculates were correctly classified into their own group, showing that this analysis may be an efficient statistical tool to improve the discrimination of potential fertility of boars. Nevertheless, the validation of the relationship between fertility and semen traits using this statistical approach needs to be performed on a larger number of farms and with a greater number of boars.     

Naturally occurring and experimental Stachys arvensis (staggerweed) toxicity in lambs in New Zealand

Stachys arvensis (staggerweed) is a common, widely distributed weed of cultivated and waste land with the potential to intoxicate sheep. Two naturally occurring outbreaks of suspected staggerweed toxicity in the lower North Island were investigated. Affected lambs had been recently moved onto staggerweed-contaminated Brassica spp. crops. In total, 150/1,200 (13%) lambs developed hindlimb paresis, a fine generalised muscular tremor, and hunched posture. When forced to move, many became recumbent. Most lambs recovered within 48 h of removal from staggerweed, although a few developed clinical signs again when transported 2–3 weeks later. Grossly, affected lambs had large amounts of staggerweed plant material and seeds within the rumen. Histopathology showed mild, multifocal degeneration of the white matter tracts of the central nervous system (CNS), most commonly in the ventral funiculi of the spinal cord, and acute, mild to moderate, multifocal degeneration of skeletal muscles. Creatine kinase (CK) activity in serum was mildly to markedly elevated in affected lambs. In a feeding trial, ten 10-month-old Romney lambs were randomly assigned to equal treatment and control groups. Treated lambs were drenched with a liquid extract of staggerweed once daily for 7 days. Three of five treated lambs developed mild exercise intolerance, and 1/5 displayed mild paresis of the hindlimbs, slightly crouched hindlimb stance, and shortened gait, on days 6 and 7. Histologically, 4/5 treated lambs had degeneration in white matter tracts of the CNS, indistinguishable from those seen in the lambs in the outbreak, and in 1/5 lambs there was scattered regeneration of skeletal muscle. CK activity in serum in treated lambs was not significantly higher than that in control lambs. None of the control lambs developed significant clinical signs, histological changes or increases in CK activity in serum. The clinical signs and lesions observed in both the outbreaks and feeding trial were similar to those previously described in studies in Australia, with the exception that myodegeneration was more prominent in the outbreaks in New Zealand. Further characterisation of the pathogenesis of staggerweed toxicity and its potential role as a food safety hazard will be facilitated through identification of the toxic principle(s).     

Isolation and characterization of gut bacteria of fruit fly, Bactrocera tau (Walker)

Gut bacteria of fruit fly, Bactrocera tau (Walker) (Diptera: Tephritidae), were isolated and the isolates attractive to B. tau adults were characterized using morphological, biochemical and 16S rRNA analyses to determine their taxonomic position. Based upon morphological, biochemical and 16S rRNA sequences (on the basis of closest match), five gut bacterial species of B. tau were characterized as Delftia acidovorans, Pseudomonas putida, Flavobacterium sp., Defluvibacter sp. and Ochrobactrum sp., of which four bacterial isolates, viz., Delftia acidovorans, Flavobacterium sp., Defluvibacter sp. and Ochrobactrum sp. are new records from guts of the fruit fly species.