Fauzia Yusuf Hafeez

Enhancing crop growth, nutrients availability, economics and beneficial rhizosphere microflora through organic and biofertilizers

Field experiment was conducted on fodder maize to explore the potential of integrated use of chemical, organic and biofertilizers for improving maize growth, beneficial microflora in the rhizosphere and the economic returns. The treatments were designed to make comparison of NPK fertilizer with different combinations of half dose of NP with organic and biofertilizers viz. biological potassium fertilizer (BPF), Biopower, effective microorganisms (EM) and green force compost (GFC). Data reflected maximum crop growth in terms of plant height, leaf area and fresh biomass with the treatment of full NPK; and it was followed by BPF+full NP. The highest uptake of NPK nutrients by crop was recorded as: N under half NP+Biopower; P in BPF+full NP; and K from full NPK. The rhizosphere microflora enumeration revealed that Biopower+EM applied along with half dose of GFC soil conditioner (SC) or NP fertilizer gave the highest count of N-fixing bacteria (Azotobacter, Azospirillum, Azoarcus andZoogloea). Regarding the P-solubilizing bacteria,Bacillus was having maximum population with Biopower+BPF+half NP, andPseudomonas under Biopower+EM+half NP treatment. It was concluded that integration of half dose of NP fertilizer with Biopower+BPF / EM can give similar crop yield as with full rate of NP fertilizer; and through reduced use of fertilizers the production cost is minimized and the net return maximized. However, the integration of half dose of NP fertilizer with biofertilizers and compost did not give maize fodder growth and yield comparable to that from full dose of NPK fertilizers.

Effect of salinity and inoculation on growth, nitrogen fixation and nutrient uptake ofVigna radiata (L.) Wilczek

This study reports the effect of salinity and inoculation on growth, ion uptake and nitrogen fixation byVigna radiata. A soil ECe level of 7.5 dS m−1 was quite detrimental causing about 60% decline in dry matter and grain yield of mungbean plants whereas a soil ECe level of 10.0 dS m−1 was almost toxic. In contrast most of the studied strains of Rhizobium were salt tolerant. Nevertheless, nodulation, nitrogen fixation and total nitrogen concentration in the plant was drastically affected at high salt concentration. A noticeable decline in acetylene reduction activity occurred when salinity level increased to 7.5 dS m−1.

Rhizobial inoculation improves seedling emergence, nutrient uptake and growth of cotton

Experiments were conducted to determine the growth promoting activities of various rhizobia in cotton (Gossypium hirsutum L.) under growth room conditions. Seeds of 4 cotton cultivars were inoculated with 4-indole-3-acetic acid producing selected (Brady) rhizobium strains and Azotobacter plant growth promoting rhizobacteria strains, included as a positive control. Growth responses to inoculation exhibited bacterial strain-cotton cultivar specificity and also included increase in rate of seedling emergence by 3–9%. Shoot dry weight, biomass and N uptake were increased by 48, 75 and 57%, respectively, due to inoculation with both the Rhizobium leguminosarum bv. trifolii E11 and Azotobacter sp. S8, whereas, strain E11 also increased root dry weight, root length and area by 248, 332 and 283%, respectively. K+ and Ca2+ uptake was also increased by 2–21% and 9–14%, respectively, due to rhizobial inoculation. The results also showed that (Brady) rhizobium strains promoted cotton growth through efficient nutrient uptake, which was mainly related to increased root growth due to the effect of IAA produced by these strains. However, growth promotion by Azotobacter sp. S8, in addition to 4-indole-3-acetic acid production, might also involve biological N2 fixation by this rhizobacterial strain at some stage during its growth.

Rhizobium, Bradyrhizobium and Agrobacterium strains isolated from cultivated legumes

The present study was conducted to isolate and characterize rhizobial strains from root nodules of cultivated legumes, i.e. chickpea, mungbean, pea and siratro. Preliminary characterization of these isolates was done on the basis of plant infectivity test, acetylene reduction assay, C-source utilization, phosphate solubilization, phytohormones and polysaccharide production. The plant infectivity test and acetylene reduction assay showed effective root nodule formation by all the isolates on their respective hosts, except for chickpea isolate Ca-18 that failed to infect its original host. All strains showed homology to a typical Rhizobium strain on the basis of growth pattern, C-source utilization and polysaccharide production. The strain Ca-18 was characterized by its phosphate solubilization and indole acetic acid (IAA) production. The genetic relationship of the six rhizobial strains was carried out by random amplified polymorphic DNA (RAPD) including a reference strain of Bradyrhizobium japonicum TAL-102. Analysis conducted with 60 primers discriminated between the strains of Rhizobium and Bradyrhizobium in two different clusters. One of the primers, OPB-5, yielded a unique RAPD pattern for the six strains and well discriminated the non-nodulating chickpea isolate Ca-18 from all the other nodulating rhizobial strains. Isolate Ca-18 showed the least homology of 15% and 18% with Rhizobium and Bradyrhizobium, respectively, and was probably not a (Brady)rhizobium strain. Partial 16S rRNA gene sequence analysis for MN-S, TAL-102 and Ca-18 strains showed 97% homology between MN-S and TAL-102 strains, supporting the view that they were strains of B. japonicum species. The non-infective isolate Ca-18 was 67% different from the other two strains and probably was an Agrobacterium strain.

Observations on the ultrastructure ofFrankia sp. in root nodules ofDatisca cannabina L.

SummaryThe fine structures of the microsymbiont inside the root nodules ofDatisca cannabina have been studied by light, by transmission- and by scanning-electron microscopy. The endophyte is prokaryotic and actinomycetal in nature. The hyphae are septate and branched, diameter 0.3–0.5 μm. The tips of hyphae are swollen to form electron-dense, clubshaped to filamentous vesicles, ranging in diameter: 0.4–1.4 μm. The endophyte penetrates through walls of the cortial cells. The infected zone is kidney shaped and confined to one side of the acentric stele. The orientation of infection is reversed from other actinorhizae exceptCoriaria. The hyphae are near the host cell wall and vesicles are directed towards the central vacuole. Vesicles are aseptate and no collapsing of the vesicle cell wall (void area) has been observed. Vesicle clusters structures are globular with an opening at one side of the cluster. The host cell is multinucleate or contains a lobed nucleus. Groups of mitochondria are located in between the hyphae, suggesting a strong association between the host and the endophyte for energy supply and amino acid production. The consequences of the inability to separate the mitochondria from the vesicle clusters in nodule homogenates in physiological studies have been discussed.Isolated vesicles clusters showed dehydrogenase activity, indicated by the presence of formazan crystals, after incubation with NADH and NBT. Strongest reducing activity was found within the vesicles. The possible role of filamentous vesicles in nitrogen fixation has been discussed.

Morphology, physiology and infectivity of two Frankia isolates An 1 and An 2 from root nodules of Alnus nitida

SummaryTwo different strains, An 1 and An 2, were obtained from root nodules ofAlnus nitida Endl., collected from one locality in the area of its natural habitat near Bahrin, District Swat, Pakistan. The light and electron microscopy of the isolates revealed the occurrence of septate and branched hyphae bearing sporangia and vesicles. The strains differed in their growth requirements, nitrogen-fixing ability and production of extracellular pigments, thus indicating the existence of more than oneFrankia strain in the same locality. In the absence of combined nitrogen in the medium strain An 1 formed vesicles and fixed N2 (up to 200 nmol C2H4. mg protein−1.h−1), while strain An 2 under the experimental conditions formed only few vesicles and fixed N2 at a very low rate (ca 10 nmol C2H4. mg protein−1 .h−1). The nitrogenase activity of strain An 1 was strongly affected by the O2 concentration.Frankia An 1 and An 2 were infective and effective onA. nitida andA. glutinosa but not onDatisca cannabina andElaeagnus umbellata. Both An 1 and An 2 strains were more infective and effective onA. glutinosa thanFrankia strains AvcIl and CpI1.

Ochrobactrum ciceri sp. nov., isolated from nodules of Cicer arietinum

A Gram-staining-negative, aerobic, rod-shaped, non-spore-forming bacterial strain, Ca-34(T), was isolated from nodules of chickpea (Cicer arietinum) in Pakistan and studied for its taxonomic affiliation. The almost full-length 16S rRNA gene sequence showed highest similarities to those of strains of the genus Ochrobactrum. Based on results of MALDI-TOF MS and 16S rRNA gene sequence similarity (98.6 %), strain Ca-34(T) and Ochrobactrum intermedium LMG 3301(T) are phylogenetic neighbours; the two strains shared DNA-DNA relatedness of 64 %. The fatty acid profile [predominantly C(18 : 1)omega7c (67.7 %) and C(19 : 0) cyclo omega8c (19.6 %)] also supported the genus affiliation. Metabolically, strain Ca-34(T) differed from other type strains of Ochrobactrum in many reactions and from all type strains in testing positive for gelatin hydrolysis and in testing negative for assimilation of alaninamide and l-threonine. Based on phenotypic and genotypic data, we conclude that strain Ca-34(T) represents a novel species, for which we propose the name Ochrobactrum ciceri sp. nov. (type strain Ca-34(T) =DSM 22292(T) =CCUG 57879(T)).

Biological control of red rot in sugarcane by native pyoluteorin-producing Pseudomonas putida strain NH-50 under field conditions and its potential modes of action

BACKGROUND Rhizobacteria have a good potential to suppress soilborne diseases, but their efficacy against sugarcane pests is rarely reported. Bacterial strains isolated from the rhizosphere of sugarcane were evaluated for their potential to suppress red rot disease on two susceptible varieties, Co-1148 and SPF-234, under field conditions. The strains were also characterised for the production of secondary metabolites associated with their antagonistic activity. RESULTS One out of four strains, the Pseudomonas putida strain NH-50 (EU627168), reduced disease severity by 44-60% in different field trials. This potent antagonistic strain produced pyoluteorin antibiotic, as confirmed by high-performance liquid chromatography (HPLC). The PltB gene involved in pyoluteorin synthesis was amplified from the P. putida strain NH-50 and sequenced. The extracellular metabolites and volatile and diffusible antibiotics secreted by the tested strains inhibited mycelial growth of Glomerella tucumensis (Speg.) Arx & E Mull in vitro by 7-55%. CONCLUSION The pyoluteorin-producing bacteria P. putida strain NH-50 significantly reduced disease severity on both sugarcane varieties, irrespective of fungal inoculation, i.e. either inoculated through stem or through soil. This strain also possesses other plant growth characteristics and can be used as a biopesticide for sugarcane crop.

Biological control of potato black scurf by rhizosphere associated bacteria

The present work was carried out to study the potential of plant rhizosphere associated bacteria for the biocontrol of potato black scurf disease caused by Rhizoctonia solani Khun AG-3. A total of twenty-eight bacteria isolated from diseased and healthy potato plants grown in the soil of Naran and Faisalabad, Pakistan were evaluated for their antagonistic potential. Nine bacterial strains were found to be antagonistic in vitro, reduced the fungal growth and caused the lysis of sclerotia of R. solani in dual culture assay as well as in extracellular metabolite efficacy test. The selected antagonistic strains were further tested for the production and efficacy of volatile and diffusible antibiotics, lytic enzymes and siderophores against R. solani. Selected antagonistic bacteria were also characterized for growth promoting attributes i.e., phosphate solubilization, nitrogen fixation and indole acetic acid production. Biocontrol efficacy and percent yield increase by these antagonists was estimated in greenhouse experiment. Statistical analysis showed that two Pseudomonas spp. StT2 and StS3 were the most effective with 65.1 and 73.9 percent biocontrol efficacy, as well as 87.3 and 98.3 percent yield increase, respectively. Potential antagonistic bacterial strain StS3 showed maximum homology to Pseudomonas sp. as determined by 16S rRNA gene sequencing. These results suggest that bacterial isolates StS3 and StT2 have excellent potential to be used as effective biocontrol agents promoting plant growth with reduced disease incidence.

Competition between effective and less effective strains of Bradyrhizobium spp. for nodulation on Vigna radiata

Abstract. Local and exotic cowpea bradyrhizobial strains were examined for their effectiveness and competitive ability on the cultivated Vigna radiata (mungbean) variety NM-51. The seven most efficient N2-fixing strains were then evaluated for their competitiveness against two less effective strains in a pair-wise inoculation experiment. Nodule occupancy was determined using fluorescent antibodies. The most efficient strain, Vr-2, could not compete with less effective strains for nodule occupancy, so the biomass and N yield were low. Bradyrhizobial strain M-55 had good competitive ability, as it occupied almost 50% of nodules in comparison with less effective strains and improved the biomass and N yield of mungbean significantly. Competitiveness/compatibility is one of the vital parameters for the selection of inocula strains, therefore, M-55, K-92 and Vr-6 are recommended as good inocula of commercial quality because of their competitiveness along with their highly efficient N2-fixation.