Genetic improvement of legumes using somatic cell and molecular techniques

Summary The merits and limitations of somatic cell techniques involving Agrobacterium-mediated transformation, direct gene transfer and protoplast fusion, are discussed in relation to the genetic improvement of forage and grain legumes. Whilst progress with legumes is limited compared to that with plants of other families such as the Solanaceae, the fact that many legumes are readily amenable to tissue culture now permits somatic cell techniques to be targetted to these species. Future development of the subject will necessitate close collaboration between molecular biologists and plant breeders to enable novel plants generated by in vitro technologies to be incorporated into conventional breeding programmes.     

Estimation of gut contents of freshwater mussel, Lamellidens marginalis L.

This study examined the food organisms found in the gut of freshwater mussels, Lamellidens marginalis L. A total of 34 taxa of food organisms were recorded, out of which 30 taxa belonged to phytoplankton and four taxa to zooplankton. Both the groups comprised three families each: Cyanophyceae (blue‐green algae), Chlorophycea (green algae) and Bacillariophyceae (diatoms) constituted phytoplankton, whereas Euglenaceae, Chlamydodontidae and Brachionidae belonged to zooplankton. The total number of frequencies of phytoplankton (n=537) were almost nine times more than that of zooplankton (n=60). When blue‐green algae, green algae, diatoms and zooplankton (all the three families together) were tested for significant difference (P<0.05) following Duncan's multiple range test, the result showed only two groups. Blue‐green algae, green algae and zooplankton were not significantly different (P<0.05), forming group A, whereas diatoms were significantly different from others, forming group B. The present study showed that the maximum number of organisms that mussels feed upon belong to green algae, followed by diatoms, blue‐green algae and few taxa of zooplankton.     

Autochthonous halotolerant plant growth-promoting rhizobacteria promote bacoside A yield of Bacopa monnieri (L.) Nash and phytoextraction of salt-affected soil

Phytoremediation is a promising approach for reclamation of salt-affected soil. Phytoextraction is the most commonly used process, which exploits plants to absorb, immobilize, and accumulate salt in their shoots. In this study, halotolerant plant growth-promoting rhizobacteria (PGPR) were isolated from the rhizosphere of wild grasses growing naturally in salt-affected areas of Lucknow, Uttar Pradesh (India) and were tested for their efficacies of salt-tolerance and plant growth-promoting (PGP) abilities. Based on 16S rRNA sequences, the most efficient halotolerant isolates possessing PGP traits were identified as Pseudomonas plecoglossicida (KM233646), Acinetobacter calcoaceticus (KM233647), Bacillus flexus (KM233648), and Bacillus safensis (KM233652). Application of these isolates as bio-inoculants significantly (P < 0.05) increased the growth and bacoside A yield of a medicinal plant, Bacopa monnieri (L.) Nash, grown on natural salt-affected soil. The phytoremediation of salt-affected soil was evident by the substantial increase in shoot Na⁺:K⁺ ratio of bio-inoculant-treated plants. When compared to un-inoculated control plants, the soil physico-chemical properties of bio-inoculant-treated plants were improved. The shoot and root biomass (fresh and dry weights), soil enzymes, and soil nutrient parameters showed significant positive correlations with the shoot Na⁺:K⁺ ratio. Consequently, the halotolerant PGPR screened in this study could be useful for the reclamation of saline soils concomitant with improved plant growth and bacoside A yield.     

Effect of azadirachtin on haematological and biochemical parameters of Argulus-infested goldfish Carassius auratus (Linn. 1758)

Argulosis hampers aquaculture production and alters the host physiology and growth. Azadirachtin is recognized as a potential antiparasitic agent against Argulus sp. The present study aimed to investigate the effect of different concentration of azadirachtin solution on haematological and serum biochemical parameters of Argulus-infested goldfish Carassius auratus. Ninety Argulus-infested goldfish were randomly divided into six equal groups. Fish of group 1–5 were treated with azadirachtin solution through bath of 1, 5, 10, 15 and 20 mg L^?1 as T1, T2, T3, T4 and T5, respectively, and group 6 was exposed to 2 % DMSO solution without azadirachtin and considered as negative control T0^?. Along with six treatment groups, a positive control T0⁺ of healthy goldfish free from Argulus infestation was also maintained. Parasitic mortality was evaluated after 3 days of consecutive bath treatment. After 7 days of post-treatment, the blood and serum were drawn from each of the treatment groups and haematological and serum biochemical parameters were evaluated. Total leucocyte count (TLC), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), blood glucose, total protein (TP), globulin, serum glutamate oxaloacetate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT) were significantly (p < 0.05) high in negative control group when compared with positive control group. It could be concluded that Argulus infestation altered marked haematological and serum biochemical parameters. However, in treated groups complete elimination of Argulus was found in T4 and T5 groups. Also significant (p < 0.05) reduction in haematological and serum biochemical parameters of all the treatment groups were recorded in comparison with negative control group. In addition, T4 and T5 groups showed significantly (p < 0.05) high superoxide dismutase (SOD), catalase, total erythrocyte count (TEC) and haemoglobin (Hb). However, higher mean corpuscular haemoglobin concentration (MCHC), blood glucose and lactate dehydrogenase (LDH) levels in T5 group revealed that higher concentration of azadirachtin have notable effects on activity of vital tissues function and physiology of the host. Argulus spp. from infested goldfish could be eliminated using bath treatment with solution of azadirachtin having concentration of 15 mg L^?1 and that also shifted haematological and serum biochemical parameters towards homeostasis.     

Interactions amongst Mycorrhiza,Azotobacter chroococcum and Root Characteristics of Wheat Varieties

A field experiment was conducted in a randomized block design with three replications over 2 years to evaluate the effect of wheat cultivar and dual inoculation of Azotobacter chroococcum (Azc) and arbuscular mycorrhiza fungi (AMF, Glomus fasciculatum) on root characters and AMF infection in three crosses of wheat. The experimental material comprised four wheat parents, WH‐147, WH‐157, WH‐542 and PBW‐175, and three F₁ crosses, WH‐147 ×WH‐157, WH‐147 × WH‐542 and WH‐147 × PBW‐175. Comparison of treatment averages, i.e. control (mineral nutrients 60 kg N + 30 kg P₂O₅ + 12.5 kg ZnSO₄ ha^?1, as in other two treatments), AMF and AMF + Azc, revealed that inoculation of Azc led to an increase in AMF infection in roots. Maximum root biomass was obtained in F₁ hybrids WH‐147 × WH‐157 in the AMF treatment and in WH‐147 × PBW‐175 receiving AMF + Azc. Total root length and AMF infection of roots was maximum in WH‐147 × PBW‐175 for all the treatments during both years. A positive association between AMF infection in roots and Azotobacter survival in the rhizosphere was apparent. Similarly, maximum A. chroococcum counts were observed 80 and 120 days after sowing in the AMF + Azc treatment in cross WH147 × PBW175.     

An Analysis of Seasonal Effects on Leaf Nitrate Reductase Activity and Nitrogen Accumulation in Maize (Zea mays L.)

The important enzyme in nitrogen (N) assimilation, nitrate reductase (NR), is an inducible enzyme influenced by many external (light, temperature, etc.) and internal (genotype) factors. The efficiency of the N assimilation system may vary with genotype and season. In the present study, the effects of season on NR activity in relation to N accumulation in maize plants were investigated. Six different cultivars of maize, namely Ganga-11, Deccan-103, Hi-starch (hybrids), Arun, Manjari and Vijay (composites), were sown during the monsoon (88-day crop duration) and in winter (150-day crop duration). In vivo NR activity in the last fully expanded leaf (LFEL), and the N contents of the whole plant and the LFEL were estimated at seven phenological growth stages. Three different states of N metabolism in maize, namely (i) low NR activity per unit leaf area per unit time coinciding with high accumulation of N, (ii) high NR activity coinciding with low N accumulation, and (iii) low NR activity coinciding with low N accumulation, were noted. These results clearly demonstrate that the relationships between N uptake and accumulation parameters change with the season and crop growth stage and are subject to a genotypic influence. Thus it is necessary to evaluate genotypes under similar environments to select a genotype with high N use efficiency. As these relationships are growth dependent, care must be taken to evaluate them at a particular phenological stage rather than on the basis of days after sowing.     

Development of a leukocytopenic baby guinea pig model with azathioprine and cyclophosphamide

Immunosuppressive agents were given to baby guinea pigs to develop a leukocytopenic baby guinea pig model. Azathioprine in different doses, and cyclophosamide and methylprednisolone combinations did not result in predictable leukocytopenia without significant mortality. A combination of azathioprine 20mg and cyclophosphamide 20mg administered once daily for 5 days to 42 baby guinea pigs resulted in a mean white blood cell count of 2531/mm³ ±1198 (a reduction of 3931 ± 1413 from pretreatment values). The mean counts for polymorphonuclear leukocytes and lymphocytes on day 6 were 398/mm³ and 2035/mm³, respectively. Only 7% mortality was seen in these animals. This model can be utilized to study a variety of bacterial and viral infections in “immunocompromised hosts”.     

Neuromuscular pedicle graft for restoration of arytenoid abductor function in dogs with experimentally induced laryngeal hemiplegia

Left laryngeal hemiplegia was induced by resection of the left recurrent laryngeal nerve in 12 dogs. A neuromuscular pedicle graft formed from the first cervical nerve and sternothyroideus muscle was transplanted after 1 week to the denervated cricoarytenoideus dorsalis muscle in 8 dogs. The remaining 4 dogs served as controls. Left arytenoid abduction was blindly evaluated by laryngoscopy with video photography at time 0, at 1 week, and at 19 weeks in all dogs. At 19 weeks, biopsy specimens of the left cricoarytenoideus dorsalis muscle and the neuromuscular pedicle were taken from 4 of the treatment dogs, and biopsy specimens of the left cricoarytenoideus dorsalis muscle were taken from the 4 control dogs. All biopsy specimens were blindly evaluated by histologic and histochemical examination. At 36 to 44 weeks, the remaining 4 treatment dogs, from which biopsy specimens had not been taken, were reevaluated by use of laryngoscopy with video photography. Complications and difficulties encountered during surgery included hemorrhage in the area of the cricoarytenoideus dorsalis muscle, location of a branch of the first cervical nerve that was long enough to prevent tension at the graft site, orientation of the muscle pedicle in the cricoarytenoideus dorsalis muscle without the use of an operating microscope, and preservation of the terminal portion of the first cervical nerve while forming the neuromuscular pedicle. Results of the arytenoid movement evaluations revealed improvement in arytenoid abductor function in the treatment group, compared with that in the control group at 19 weeks. Arytenoid abduction in the treatment group at this time, however, was still significantly decreased (P less than 0.05), compared with presurgical movement evaluations.(ABSTRACT TRUNCATED AT 250 WORDS)     

Interaction of bovine neutrophils in Pasteurella haemolytica mediated damage to pulmonary endothelial cells

The purpose of these studies was to determine mechanisms of pulmonary tissue damage mediated by Pasteurella haemolytica and interaction with bovine neutrophils. Bovine pulmonary artery endothelial cell monolayers were treated with various combinations of P. haemolytica factors including bacterial culture supernatant (CS) and purified LPS, with and without bovine neutrophils. Damage to endothelial cells was monitored by 51Cr release, cell detachment rate, and morphological changes. At 5 h post-treatment (PT) bacterial factors produced very little toxic change in cells, however, by 22 h PT both crude leukotoxin and LPS caused high levels of cytotoxicity and detachment. Neutrophils did not augment toxicity mediated by LPS, but actually protected endothelial cells from low levels of LPS. When the LPS component of CS was neutralized with polymyxin B, leukotoxin mediated neutrophil killing resulted in extensive endothelial cell damage. These results suggest that LPS may directly injure endothelial cells and this toxic effect may be reduced by neutrophils. However, neutrophil killing by leukotoxin may also contribute to endothelial cell damage in the absence of LPS.