Traits of transgenic Atropa belladonna doubly transformed with different Agrobacterium rhizogenes strains

Hairy root cultures of Atropa belladonna L. were established by infection either with Agrobacterium rhizogenes ATCC 15834 or MAFF 03-01724, and transgenic plants were obtained from both hairy root cultures. Doubly transformed roots were induced by re-infection of the leaf segments of transgenic Atropa belladonna plants (A. rhizogenes 15834) with MAFF 03-01724. Shoots and viviparous leaves were regenerated from the doubly transformed roots. The genetic transformation was determined by the opine assay (agropine, mannopine and/or mikimopine) and polymerase chain reaction. Physiological changes and tropane alkaloid biosynthesis in the hairy roots (singly and doubly transformed) were investigated. The alkaloid content in the doubly transformed root strain was intermediate as compared to the root strains which were singly transformed. On the other hand endogenous IAA levels in doubly transformed roots were significantly decreased compared to both singly transformed roots.Abbreviations BA benzyladenine - IAA indoleacetic acid - NAA naphthaleneacetic acid - PCR polymerase chain reaction - t-ZR trans-zeatin     

Increased daylength stimulates plasma growth hormone and gill Na+, K+-ATPase in Atlantic salmon (Salmo salar)

Atlantic salmon juveniles reared at constant temperature (9–10°C) were exposed to four photoperiod treatment and sampled every 2 weeks from January through May. Fish reared under normal photoperiod exhibited eight-and three fold increases in plasma growth hormone and gill Na⁺, K⁺-ATPase activity, respectively, between January and April. Fish exposed to abrupt increases in daylength (LD 15:9) in February or March responded with earlier increases in plasma growth hormone and gill Na⁺, K⁺-ATPase activity, and earlier decreases in condition factor relative to fish in the normal photoperiod group. Fish maintained under short daylength (LD 9:15) from January to May exhibited delayed and muted increases in plasma growth hormone and gill Na⁺, K⁺-ATPase activity. Plasma thyroxine exhibited a 2.5-fold increase from February to late March in the normal photoperiod group, was generally lower in the LD 9:15 group, but exhibited no obvious response to abrupt increases in daylength. There was an increase in plasma 3,5,3-triiodo-l-thyronine with time in all groups (43–80%) but no significant response to photoperiod. Plasma levels of somatostatin-25 were highest in the LD 9:15 group, but there was no detectable response to increased daylength in any of the photoperiod treatments. The results indicate that plasma growth hormone is responsive to increased daylength and may be causally related to subsequent increases in gill Na⁺, K⁺-ATPase.Abbreviations ANOVA two-way analysis of variance - BCA bicinchoninic acid - BSA Bovine serum albumin - EDTA ethylene diamine tetraacetic acid - ELISA enzyme-linked immunosorbent assay - EST eastern standard time - GH growth hormone - GLU Glucagen - IgG Immunoglobulin G - INS Insulin - LDN Simulated natural photoperiod - RIA radio immuno assay - RIA radio immuno assay - SEI Sucrose EDTA imidazole - SS-25 somatostatin-25 - SW sea water - T ₃ 3,5,3 triiodo-l-thyronine - T ₄ thyroxine     

Localization of the F420-reducing hydrogenase in Methanococcus voltae cells by immuno-gold technique

The F₄₂₀-reducing hydrogenase of Methanococcus voltae, which takes part in the terminal reduction step of methanogenesis, was localized in situ in ultrathin sections. This result was obtained by the immuno-gold technique using a high titer antiserum raised against the purified enzyme. Its specifity for the hydrogenase was shown by Western blot analysis. The hydrogenase of M. voltae was found to be membrane-associated.Abbreviations ELISA Enzyme linked immuno sorbent assay - F₄₂₀ 8-hydroxy-5-deazaflavin     

A bioluminescence assay to detect nitrification inhibitors released from plant roots: a case study with Brachiaria humidicola

A bioluminescence assay using recombinant Nitrosomonas europaea was adopted to detect and quantify natural nitrification inhibitors in plant–soil systems. The recombinant strain of N. europaea produces a distinct two-peak luminescence due to the expression of luxAB genes, introduced from Vibrio harveyi, during nitrification. The bioluminescence produced in this assay is highly correlated with NO₂⁻ production (r ² = 0.94). Using the assay, we were able to detect significant amounts of a nitrification inhibitor produced by the roots of Brachiaria humidicola (Rendle) Schweick. We propose that the inhibitory activity produced/released from plants be termed ‘biological nitrification inhibition’ (BNI) to distinguish it from industrially produced inhibitors. The amount of BNI activity produced by roots was expressed in units defined in terms of the action of a standard inhibitor allylthiourea (AT). The inhibitory effect from 0.22 μM AT in an assay containing 18.9 mM of NH₄⁺ is defined as one AT unit of activity. A substantial amount of BNI activity was released from the roots of B. humidicola (15–25 AT unit g⁻¹ root dry wt day⁻¹). The BNI activity released was a function of the growth stage and N content of the plant. Shoot N levels were positively correlated with the release of BNI activity from roots (r ² = 0.76). The inhibitor/s released from B. humidicola roots suppressed soil nitrification. Additions of 20 units of BNI per gram of soil completely inhibited NO₃⁻ formation in a 55-day study and remained functionally stable in the soil for 50 days. Both the ammonia monooxygenase and the hydroxylaminooxidoreductase enzymatic pathways in Nitrosomonas were effectively blocked by the BNI activity released from B. humidicola roots. The proposed bioluminescence assay can be used to characterize and determine the BNI activity of plant roots, thus it could become a powerful tool in genetically exploiting the BNI trait in crops and pastures.     

Tissue culture of the desert plant Anastatica hiërochuntica

Summary Callus derived from the winter annual desert plant Anastatica hiërochuntica was grown on different media, Murashige and Skoog (1962) medium giving the best results. Large amounts of lignified xylem elements were formed resulting in an extremely hard tissue. The growth responses to different auxins, cytokinins and abscisic acid were investigated. When salts (high Na⁺, Ca²⁺ and Cl^--contents) as they can be found in aqueous extracts of desert soils from a natural A. hiëerochuntica habitat were added to Abou-Mandour (1977) or MS-media, growth of callus was inhibited drastically. In the presence of abscisic acid, however, original growth was completely restored. In salt free control media on the other hand, ABA proved to be inhibitory. Drought stress caused a decrease of both cytokinins and indoleacetic acid in the callus while ABA levels were increased, but by far not as distinct as in intact plants. Proline level was not affected by stress.Abbreviations ABA abscisic acid - AM Abou-Mandour-medium - BAP benzylaminopurine - 2,4-D 2,4-dichlorophenoxyacetic acid - DHZR dihydroxyzeatinriboside - DW dry weight - ELISA enzyme linked immuno sorbent assay - FW freshweight - IAA indole-3-acetic acid - NAA 1-naphthaleneacetic acid - IBA indole-3-butyric acid - IPA isopentenyladenosine - Kin kinetin - MS Murashige and Skoog-medium     

Cytokinins in the perennial herb Urtica dioica L. as influenced by its nitrogen status

The effect of nitrogen on the cytokinin relations of Urtica dioica, the stinging nettle, has been investigated. The plants were grown in quartz sand and nutrient solutions providing levels of nitrate ranging from 1 to 22 mM. Nitrogen supply did not affect biomass production within the range of 3–15 mM NO ₃ ^- . However, the shoot: root ratio of biomass was significantly higher at 15 mM (standard plants) than at 3 mM (low-nitrogen plants) nitrate supply. The cytokinin patterns of the roots, stems and adult, as well as meristematic leaves of plants grown at these two levels of nitrate supply, were determined by means of high-performance liquid chromatography (HPLC) and immunoassays. Enzyme-linked immunosorbent assays (ELISAs) for zeatin riboside, dihydrozeatin riboside, isopentenyladenosine, benzyladenosine and o-hydroxybenzyladenosine enabled the quantification of 17 cytokinins, 13 of which were found in the various tissues of Urtica. trans-Zeatin and its conjugates were the predominant cytokinins in all examined samples. While the free base trans-zeatin and its O-glucoside were the major cytokinins in adult leaves, trans-zeatin riboside was prominent in the other tissues of at least the standard plants. Glucosides of the trans-zeatin type cytokinins were present only in lower amounts. However, considerable amounts of a compound, tentatively identified as cis-zeatin riboside-O-glucoside, were found, particularly in roots and meristematic leaves. Comparatively high amounts of trans-zeatin nucleotide as well as isopentenyladenosine phosphate were also demonstrated in these tissues. Analysis of the root-pressure exudates similarly showed trans-zeatin riboside and, at a lower concentration, trans-zeatin to be the only substantial components. In the low-nitrogen plants, shortage of nitrogen was manifest only in the roots; the nitrogen contents of the shoots did not respond to the nitrogen supply. Likewise, the total content of cytokinins in the shoots of the low-nitrogen plants equaled that of the standard-plant shoots, while it was lower by about 25% in the roots of the low-nitrogen plants. In the latter, the amounts of cytokinins exuded via the root-pressure fluid were also approximately 25% lower. Since the levels of only the trans-zeatin cytokinins in the roots showed a linear correlation with the shoot-to-root ratios, these cytokinins may play an important role in biomass partitioning in Urtica dioica.Abbreviations DHZ dihydrozeatin - ELISA enzyme-linked immunosorbent assay - -G glucoside - HPLC high-performance liquid chromatography - 2iP isopentenyladenine - 2iPA isopentenyladenosine - -N nucleotide (ribotide) - -OG O-glucoside - -R riboside - S/R shoot-to-root (ratio) - Z zeatin This work was supported by the Deutsche Forschungsgemeinschaft within the scope of the SFB 137. The authors wish to thank Mrs. A. Fischbach for skilful technical assistence and Dr. Paul Ziegler (Lehrstuhl für Pflanzenphysiologie, University of Bayreuth, FRG) for linguistic suggestions.     

Enhancement of ferric-mugineic acid uptake by iron deficient barley roots in the presence of excess free mugineic acid in the medium

To investigate the mechanism of mugineic acid-Fe^III uptake by barley roots, plasma membrane fractions were isolated from Fe-deficient barley roots using an aqueous two-phase partition method. Utilizing the plasma membrane vesicles, we developed an assay system for studying mugineic acid-⁵⁵Fe^III binding to the plasma membrane. However, no efficient active transport of mugineic acid-⁵⁵Fe^III into the plasma membrane vesicle was detected, because of large amount of non-specific adsorption of ⁵⁵Fe^III onto the vesicle. And the adsorption could be decreased by adding excess amount of free mugineic acid to the assay system. From the results it is speculated that an excess of free mugineic acids is necessary in the medium for effective uptake of mugineic acid-Fe^III by Fe-deficient barley roots. Support for this speculation came from a multi-compartment transport box experiment with excised roots of Fe-deficient barley.Abbreviations CCCP carbonylcyanide-m-chlorophenylhydrazone - MA mugineic acid     

Culture of transformed horseradish roots as a source of fusicoccin-like ligands

Endogenous fusicoccin (FC) or related substances were sought in horseradish (Armoracia rusticana P.) roots. An actively growing root culture was derived from plants transformed with Agrobacterium rhizogenes. The presence of FC-like substances in ethanolic extracts from roots was established in a radioreceptor binding assay with plasmalemmal FC receptors and in radioimmune analysis with an antiserum specific for FC A. FC-like ligands were found in the tissue and medium of aseptically grown culture.Abbreviations FC fusicoccin - GC/MS gas chromatography/mass spectrometry - RIA radioimmunoassay - RRA radioreceptor analysis - BSA bovine serum albumin - Mes 4-morpholineethanesulfonic acid - HPLC high performance liquid chromatography     

Effect of Lysophosphatidylethanolamine and Brassinosteroids on Development of <Emphasis Type="Italic">Arabidopsis</Emphasis> Roots

Exogenously applied lysophosphatidylethanolamine (LPE) increased the growth of primary roots and the formation of lateral roots in Arabidopsis thaliana. In the presence of brassinolide, lateral root formation induced by LPE was enhanced, implying that both LPE and brassinosteroids (BR) interact positively in the development of Arabidopsis roots. Co-treatment with LPE and BRs increased the bending activity in the rice lamina inclination assay compared to that when BRs were applied alone, suggesting that LPE seems to exert its activity via BRs activity. RT-PCR revealed that LPE did not alter the expressions of genes involved in the biosynthesis of BRs but did activate the expressions of BR signaling genes in A. thaliana. In a BR-insensitive mutant, bri1, enhanced gravitropic response by LPE in wild-type A. thaliana was diminished. In conclusion, LPE is a positive regulator for the growth and development of Arabidopsis roots, and this process seems to be enhanced by BR signaling rather than by increase in endogenous levels of BRs in A. thaliana.     

Antifungal properties of selected plants from the Yucatan peninsula,Mexico

A total of 20 plants belonging to different genera (Acalypha, Ageratum, Ambrosia, Bidens, Blechum, Caesalpinia, Calea, Carlowrightia, Croton, Eugenia (2), Furcraea, Stenandrium, Tephrosia, Trichilia (2), Randia (3), and Vitex) were selected from native flora of the Yucatan peninsula. All plants selected were collected and separated in to leaves, stems and roots. These were then extracted with ethanol and their crude extracts (54) were evaluated against Alternaria tagetica, Colletotrichum gloeosporioides, Fusarium oxysporum and Rhizopus sp. using the filter paper disc diffusion assay. Results lead to the selection of 33 crude extracts active against at least one of the target strains, which were assessed to determine their ability to inhibit the mycelial growth of the pathogenic fungi in a second antifungal assay. The results of this assay indicated that extracts from the roots of Croton chichenensis were the most promising, with a wide activity spectrum against all pathogens tested in both assays with inhibition percentages of greater than 60%. Furthermore, extracts from leaves of Ambrosia hispida, Trichilia minutiflora, and roots of Acalypha gaumeri were able to cause growth inhibition against two or three pathogen strains (≥50%). Studies of these active extracts should be continued at different levels. In general, results revealed a good bioactive potential of the flora from the Yucatan peninsula to produce metabolites with potential applications as botanical pesticides in the near future.     

Use of ELISA with Antiexopolysaccharide Antibodies to Evaluate Wheat-Root Colonization by the Rhizobacterium Paenibacillus polymyxa

Enzyme-linked immunosorbent assay with rabbit polyclonal antibodies developed to isolated exopolysaccharide of Paenibacillus polymyxa 1465 was used to evaluate the colonization of wheat-seedling roots by this bacterium. The assay conditions were optimized for detection of the P. polymyxa exopolysaccharide determinants forming part of the samples used (homogenates of inoculated roots). The dynamics of the immunoenzymatic revealing of specific polysaccharidic antigenic determinants in the samples’ composition correlated with an increase in P. polymyxa numbers on the roots found by estimation of colony-forming units.     

Interactive effects of inorganic phosphate nutrition and carbon dioxide enrichment on assimilate partitioning in barley roots

The combined effects of inorganic phosphate (Pi) insufficiency and CO₂ enrichment on metabolite levels and carbon partitioning were studied using roots of 9-, 13- and 17-day-old barley seedlings (Hordeum vulgare L. cv. Brant). Plants were grown from seed in controlled environment chambers providing 36 ± 1 Pa (ambient) or 100 ± 2 Pa (elevated) CO₂ and either 1.0 mM (Pi sufficient) or 0.05 mM (Pi insufficient) Pi. When values were combined for both Pi treatments, plants grown under enhanced CO₂ showed increased root dry matter, adenylates (ATP + ADP), glutamine and non- structural carbohydrates other than starch. In contrast with shoots, enhanced CO₂ partially reversed the inhibition of root dry matter formation imposed by Pi insufficiency. The Pi-insufficient treatment also increased sucrose, glucose and fructose levels in barley roots. The Pi and CO₂ treatments were additive, so that the highest soluble carbohydrate levels were observed in roots of Pi-insufficient plants from the elevated CO₂ treatment. Pi limitation decreased dry matter formation, acid-extractable Pi, nitrate, hexose-phosphates, glutamate, glutamine and acid invertase activity of barley roots in plants grown in both ambient and elevated CO₂. Adenylate levels in roots were unaffected by the moderate Pi insufficiency described here. Thus, the reduced hexose-phosphate levels of Pi-insufficient roots were not likely to be the result of low adenylate concentrations. The above results suggest that the capacity of barley roots to utilize carbohydrates from the shoot is inadequate under both Pi-insufficient and CO₂-enriched treatments. In addition, the Pi and CO₂ treatments used here alter the nitrogen metabolism of barley roots. These findings further emphasize the importance of avoiding nutrient stress during CO₂ enrichment experiments.     

Isolation and characterization of parvalbumins from skeletal muscles of a tropical amphibian,Leptodactylus insularis

Summary Parvalbumins were isolated from skeletal muscles of a tropical amphibian, Leptodactylus insularis, and three new isotypes were identified. The total concentration of parvalbumins in L. insularis was the same as the total amounts found in an amphibian from the temperate or variable zone (Rana temporaria). Muscles of the thigh and foreleg had the maximum parvalbumin concentration (0.35 mmol · kg wet weight^-1). Samples from pectoralis and rectus abdominis muscles had significantly less (0.29 mmol · kg^-1). Three previously unknown parvalbumin isotypes (IV, IIIa, and IIIb) were isolated from the tropical amphibian. They were different from the isotypes (IVa and IVb) predominant in R. temporaria skeletal muscle. Parvalbumins are thought to have a role in the short-term removal of myoplasmic Ca²⁺ during muscle relaxation. Hence, the unique isotypes in L. insularis may reflect optimal molecular adaptations retained during the animal's evolution in a constantly warm environment.Abbreviations DEAE diethylaminoethyl - ELISA enzyme linked immuno sorbent assay - SPDP N-succininydyl-3-(2-pyridyldithio) propionate - SR sarcoplasmic reticulum     

Effect of Wild-Type and Mutant Plant Growth-Promoting Rhizobacteria on the Rooting of Mung Bean Cuttings

Mung bean cuttings were dipped in solutions of wild type and mutant forms of the plant growth-promoting rhizobacterium Pseudomonas putida GR12-2 and then incubated for several days until roots formed. The bacteria P. putida GR12-2 and P. putida GR12-2/aux1 mutant do not produce detectable levels of the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, whereas P. putida GR12-2/acd36 is an ACC deaminase minus mutant. All bacteria produce the phytohormone indole-3-acetic acid (IAA), and P. putida GR12-2/aux1 overproduces it. Treatment of cuttings with the above-mentioned bacteria affected the rates of ethylene production in the cuttings in a way that can be explained by the combined effects of the activity of ACC deaminase localized in the bacteria and bacterial produced IAA. P. putida GR12-2 and P. putida GR12-2/acd36-treated cuttings had a significantly higher number of roots compared with cuttings rooted in water. In addition, the wild type influenced the development of longer roots. P. putida GR12-2/aux1 stimulated the highest rates of ethylene production but did not influence the number of roots. These results are consistent with the notion that ethylene is involved in the initiation and elongation of adventitious roots in mung bean cuttings. Received October 21, 1998; accepted January 3, 1999     

The effect of salinity on lipid composition and on activity of Ca2+- and Mg2+-stimulated ATPases in salt-sensitive and salt-tolerant Plantago species

Lipid composition of the roots and the shoot of the salt-sensitive Plantago media L., the salt-tolerant P. maritima L. and the less salt-tolerant P. coronopus L. was followed under saline conditions. In the roots of P. media the level of phospho-, galacto- and sulpholipids decreased strongly with increased NaCl concentration, indicating decreased control of permeability of the root cell membranes. In the roots of the two salt-tolerant species the level of most lipid classes was maintained or even raised up to 75 mM NaCl, and a decrease was noted only at higher NaCl concentrations. In P. maritima, a species from relatively nutrient-rich habitats, decreased lipid levels in the roots and shoot were observed with increasing salinity in combination with a low nutrient availability. The Ca²⁺- and Mg²⁺-stimulated ATPase activities of the microsomal fraction of the roots of P. maritima was decreased at a salinity level in excess of 150 mM, while in P. coronopus they were decreased at all NaCl levels tested. The obtained results are discussed as part of the adaptation of the species to salinity.     

The response to auxin of rapeseed (Brassica napus L.) roots displaying reduced gravitropism due to transformation by Agrobacterium rhizogenes

It has recently been documented that, compared to untransformed controls, the roots of oilseed rape (Brassica napus L. CV CrGC5) seedlings transformed by Agrobacterium rhizogenes A4 show a reduced gravitropic reaction (Legué et al. 1994, Physiol Plant 91: 559–566). After stimulation at 90°C or 135°, the transformed root tips curve, but never reach a vertical orientation. In the present study, we investigated the causes of reduced gravitropic bending observed in stimulated transformed root tips. First, we localized the gravitropic curvature in normal and in transformed roots after 1.5 h of stimulation. The cells involved in root curvature (target cells) corresponded at the cellular level to the apical part of the zone of increasing cell length. In transformed roots grown in the vertical position, these cells showed a reduction in cell length compared to controls. Because auxin is considered to be the gravitropic mediator, the response of normal and transformed roots to exogenous auxin was studied. Indole-3-acetic acid (IAA) was applied along the first 3 mm using resin beads loaded with the hormone. In comparison to normal roots, transformed roots showed reduced bending toward the bead at all points of bead application. Moreover, the cells which responded to IAA corresponded to the target cells involved in the gravitropic reaction. The level of endogenous IAA was lower in transformed roots. Thus, it was concluded that the modified behavior of transformed roots during gravitropic stimulation could be due to differences either in IAA levels or in reactivity of the target cells to the message from the cap.Abbreviations DEZ distal elongation zone - ELISA enzymelinked immunosorbent assay - T-DNA DNA transferred from Agrobacterium rhizogenes to the plant genome This work was supported by the Centre National d'Etudes Spatiales.     

A comparison of the effects of auxin on cluster root initiation and development in Grevillea robusta Cunn. ex R. Br. (Proteaceae) and in the genus Lupinus (Leguminosae)

The effect of NAA (naphthaleneacetic acid) on the development of cluster roots in members of the Proteaceae and Leguminosae was investigated. The exogenous addition of NAA led to initiation of cluster roots in phosphate conditions normally inhibitory for their development, but initiation took place within the limits of the cluster pattern under –P conditions. There was no change in spacing within the cluster root nor between cluster roots in Grevillea robusta Cunn. ex R. Br. or in rootlet length or cluster root length. In Lupinus albus L., change in rootlet length and cluster root length was noted at 10^-10 and 10¹² M NAA. In L. albus, the length of time that roots were exposed to NAA does not appear to be important, with similar levels of cluster root initiation after 48 h and 7 days. Cluster root production in G. robusta differed from that in L. albus in terms of the concentration of NAA needed to induce initiation, and in the effects of extremely low levels of NAA on rootlet numbers and lengths. L. arboreus L. does not produce cluster roots under –P conditions. Furthermore, neither L. arboreus L., L. angustifolius L., L. luteus L. nor L. mutabilis L. were induced to produce cluster roots under –P conditions, nor under +P conditions in the presence of exogenous NAA. Thus, exogenous NAA only leads to the induction of cluster roots, at levels of P normally inhibitive of their development, in species of Lupinus that produce them under –P conditions. Auxin-induced cluster roots develop within the same constraints as those developing under –P conditions. NAA does not induce cluster roots in species of Lupinus that do not produce them under –P conditions.     

Evaluation of bacteria isolated from rice rhizosphere for biological control of charcoal rot of sorghum caused by <Emphasis Type="Italic">Macrophomina phaseolina</Emphasis> (Tassi) Goid.

A total of 360 bacteria, isolated from the rhizospheres of a system of rice intensification (SRI) fields, were characterized for the production of siderophore, fluorescence, indole acetic acid (IAA), hydrocyanic acid (HCN) and solubilization of phosphorus. Of them, seven most promising isolates (SRI-156, -158, -178, -211, -229, -305 and -360) were screened for their antagonistic potential against Macrophomina phaseolina (causes charcoal rot in sorghum) by dual culture assay, blotter paper assay and in greenhouse. All the seven isolates inhibited M. phaseolina in dual culture assay, whereas six isolates solubilized phosphorous (except SRI-360), all seven produced siderophore, four produced fluorescence (except SRI-178, -229 and -305), six produced IAA (except SRI-305) and five produced HCN (except SRI-158 and -305). In the blotter paper assay, no charcoal rot infection was observed in SRI-156-treated sorghum roots, indicating complete inhibition of the pathogen, while the roots treated with the other isolates showed 49–76% lesser charcoal rot infection compared to the control. In the antifungal activity test (in green house on sorghum), all the isolates increased shoot dry mass by 15–23% and root dry mass by 15–20% (except SRI-158 and -360), over the control. In order to confirm the plant growth-promoting (PGP) traits of the isolates, the green house experiment was repeated but, in the absence of M. phaseolina. The results further confirmed the PGP traits of the isolates as evidenced by increases in shoot and root dry mass, 22–100% and 5–20%, respectively, over the control. The sequences of 16S rDNA gene of the isolates SRI-156, -158, -178, -211, -229, -305 and -360 were matched with Pseudomonas plecoglossicida, Brevibacterium antiquum, Bacillus altitudinis, Enterobacter ludwigii, E. ludwigii, Acinetobacter tandoii and P. monteilii, respectively in BLAST analysis. This study indicates that the selected bacterial isolates have the potential for PGP and control of charcoal rot disease in sorghum.     

Immunodiagnosis of bancroftian filariasis—Problems and progress

The immunodiagnosis of bancroftian filariasis is a major challenge to the immunoparasitologist. Significant progress is yet to be made in developing convenient laboratory animal model and inin vitro cultivation of filarial parasites making it very difficult to obtain required amount of parasite material for research. Parasitological examination techniques are not useful in low microfilaraemia, occult or chronic.filarial infections. A precise and accurate immunodiagnostic technique is very much needed for successful filaria control programmes. Such a test will also avoid the need for laborious night blood examination in bancroftian filariasis. Due to comparatively easy availability, a good amount of work has been done to explore immunodiagnostic potential of heterologous filarial antigens isolated fromLitomosoide carinii, Dirofilaria immitis, Brugia malayi, Setaria digitata, Setaria cervi and number of other filarial species. However, there has been limited or no significant success due to number of false negative and false positive reactions. Extensive study has also been made with antigens isolated fromWuchereria bancrofti microfilariae. Soluble antigens of microfilariae have been used in different immunological techniques such as skin test, counter immuno electrophoresis, indirect haemagglutination test, indirect fluorescent antibody test and enzyme linked immunosorbent assay. Fractionation of Wuchereria bancrofti microfilarial soluble antigens yielded mfS3e antigen fraction which was found to be highly reactive in microfilaraemia by enzyme linked immunosorbent assay, but the yield of the purified antigen was not sufficient enough to make it a practical proposition for large scale isolation of antigen. Wuchereria bancrofti microfilarial excretory-secretory antigens were found to be specific and highly sensitive requiring as little as 0.35 ng antigen protein per well in penicillinase enzyme linked immunosorbent assay for detection of filarial antibody. One ml of culture fluid was found to be sufficient for 400,000 tests. Field evaluation of this test showed that it can replace laborious night blood examination. Assay systems have been developed for detection of filarial antigen in serum, urine, hydrocele fluid and immune complexes using immunoglobulins from chronic filarial sera and antisera to excretory filarial antigens. Further purification of excretory-secretory antigens by affinity chromatography and production of monoclonal antibodies should hopefully give suitable reagents for use in sensitive assays such as enzyme immuno assay and immuno radiometric assay, providing an ideal assay system for detection of active filarial infection in the not too distant future.