Bacillus thuringiensis isolates from soil and diseased insects in Egyptian cotton fields and their activity against lepidopterous insects

The distribution of Bacillus thuringiensis in Egyptian soils of cotton cultivations represented by 11 governorates was studied. This study aimed to isolate indigenous strains that may be potent against some lepidopterous insect pests. Out of 45 isolates, only 10 were effective and they showed high levels of toxicity against the target insects at 500 μg/ml (80% larval mortality or higher). Among these 10 isolates, two isolates were potent against Spodoptera littoralis, two isolates were potent against Helicoverpa (= Heliothis) armigera, seven isolates were potent against Pectinophora gossypiella, but none were potent against Agrotis ypsilon. LC₅₀ and LC₉₀ values and the potency of the B. thuringiensis isolates have been determined. Trials were conducted to isolate B. thuringiensis from diseased lepidopterous insects collected from the same locations, but all isolates showed low potential activity against the target insects. These findings showed promise for the possible use of some indigenous B. thuringiensis strains in the control of lepidopterous pests in Egypt.     

Characterisation of lepidopteran-active Bacillus thuringiensis isolates recovered from infected larvae

Abstract

As a part of an ongoing nationwide programme focused on finding novel strains of Bacillus thuringiensis (Bt) that are toxic to some of the major pests that impact economically important crops, we initiated a search for Bt isolates native to Syria. We succeeded in assembling a collection of 40 Bt isolates recovered from infected larvae of Galleria mellonella, Helicoverpa armigera and Ephestia kuehniella. Light microscopy showed that all isolates produce bipyramidal and cuboidal crystal proteins. The 50% lethal concentration of the spore-crystal mixture of the 40 isolates against E. kuehniella larvae varied from 3 to more than 200 µg g^?1. A comparison of the LC₅₀ values of the tested isolates with the reference strain Bt kurstaki HD-1 (20.55 µg g^?1), showed that some of these isolates have a similar or up to six times higher toxicity potential. PCR screening revealed that all obtained isolates contain cry1 and cry2 genes, whereas only four contain cry9. Moreover, the proteins of 130 and 65/70 Kda encoded by these genes were detected in the SDS-PAGE of the purified parasporal bodies. Flagellar serotyping classified 30 as serovar kurstaki, six isolates serovar aizawai, one isolate cross-reacted with more than one H3 antisera and three were not typeable. Assays of toxicity of the aizawai isolates against third instar of G. mellonella showed that four, which contain cry9, have almost similar toxicity to the commercial strain Bt aizawai B401. Therefore, these isolates could be adopted for future applications to control G. mellonella. Moreover, this study contributes to our knowledge of Bt diversity in Syria where to date very few collections have been described.     

Characterization of native <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> strains and selection of an isolate active against <Emphasis Type="Italic">Spodoptera frugiperda</Emphasis> and <Emphasis Type="Italic">Peridroma saucia</Emphasis>

Twelve Bacillus thuringiensis (Bt) strains, isolated from larvae and soil samples in Argentina, were molecularly and phenotypically characterized and their insecticidal activities against Spodoptera frugiperda and Peridroma saucia were determined. One isolate—Bt RT—produced more than 93% mortality on first instar larvae of both species, which was higher than that produced by the reference strain Bt 4D1. Bt RT carried a different cry gene profile than Bt 4D1. Scanning electron microscopy showed the presence of bipyramidal and cuboidal crystals. Phenotypic characterization revealed lytic enzymes that could contribute to Bt pathogenicity.     

Biocontrol efficacy of protoplast fusants between Bacillus thuringiensis and Bacillus subtilis against Spodoptera litura Fabr.

Bacillus thuringiensis (Bt) is a microbial pesticide widely used to control crop pests. Its strains have good biocontrol activity against crop insect pest, but lack some desirable characteristics that are found in Bacillus subtilis. An attempt has been made to combine those desirable characteristics; we used a highly effective biocontrol strain of B. thuringiensis in protoplast fusions with a strain of B. subtilis. The fusants were identified through cell culture and stained with crystal violet. The Bt and B. subtilis protoplasts were induced to fuse by PEG 6000. The fusants were produced almost 95% mortality in first instar larvae of Spodoptera litura. The lethal doses (The LC₅₀ and LC₉₀) for mortality of S. litura values were significantly in lower level in the fusant-treated larvae, when compared with Bt and B. subtilis individual treatment. The consumption and digestion of S. litura significantly decreased after treatment with fusant. Also the approximate digestibility of S. litura increased significantly.     

Characterization of a strain of Serratia sp. with ixodicide activity against the cattle tick Rhipicephalus microplus

Cattle ticks are considered the most important ectoparasite in the livestock industry. Rhipicephalus microplus causes economic losses both through direct feeding on livestock and through disease transmission. Reports of the failure of chemical ixodicides to control this tick have led to a search for control alternatives, such as bacteria with ixodicide activity. The objective of this work was to select a bacterial strain with ixodicide activity against R. microplus. In total, 83 bacterial strains were isolated from soil and dead R. microplus specimens, and all strains were evaluated against larvae in a screening test. Bacteria with ixodicide activity were evaluated in larvae and engorged adult female ticks. The larvae were challenged using the larval immersion test (LIT) with 20 µg/mL total protein. The median lethal concentration (LC₅₀) for larvae was obtained by using nine total protein concentrations. Engorged adult female ticks were challenged using the adult immersion test (AIT) with six protein concentrations. We evaluated adult mortality on day 10, oviposition rate on day 14 and hatching rate on day 40 after challenge. Only one bacterial strain (EC-35) showed ixodicide activity against larvae and adult R. microplus. The highest larval mortality, 52.3%, occurred with a total protein concentration of 40 μg/mL, and the LC₅₀ was 13.9 µg/mL of protein. In adults, a total protein concentration of 10 µg/mL had the highest mortality (55%), oviposition inhibition (50.9%) and reproductive potential inhibition (52.5%). However, there was no significant effect on hatching. The 16S rRNA gene sequence showed 99% identity of EC-35 with Serratia sp.

     

Pathogenicity of the entomogenous,hyphomycete fungus,Metarhizium anisopliae against the chrysomelid beetles Psylliodes chrysocephala and Phaedon cochleariae

Susceptibility of the mustard beetle (Phaedon cochleariae) and the cabbage stem flea beetle (Psylliodes chrysocephala) to six isolates of the entomogenous, hyphomycete fungus Metarhizium anisopliae, was investigated. A farther six isolates were assayed against P. cochleariae only. The isolates originated from hosts of various insect orders. Five of the six isolates tested against P. chrysocephala and P. cochleariae were infective for both species whereas one isolate, V107, was non‐pathogenic to both. The level of virulence of different M. anisopliae isolates for these chrysomelid beetles varied considerably. Isolates V90 and V93 were highly virulent to P. chrysocephala and P. cochleariae respectively but were significantly less virulent against the alternate host species. The LT₅₀ of isolate V90 for P. chrysocephala was 7 days at 4 x 10⁷ conidia/ml and its LC₅₀ value was 16 x 10⁵ conidia/ml. The LT₅₀ of V93 for P. cochleariae was approximately 8 days at 4 X 10⁸ conidia/ml and its LC₅₀ value was 3 x 10⁷ conidia/ml. Following inoculation, germinating conidia of all isolates produced appressoria on the cuticular surface of both hosts suggesting that specificity is determined at later stages of infection.     

Effects of seed mixture sowing with transgenic Bt rice and its parental line on the population dynamics of target stemborers and leafrollers,and non‐target planthoppers

The widespread planting of insect‐resistant crops has caused a dramatic shift in agricultural landscapes, thus raising concerns about the potential impacts on both target and non‐target pests. In this study, we examined the potential effects of intra‐specific seed mixture sowing with transgenic Bt rice (Bt) and its parental non‐transgenic line (Nt) (100% Bt rice [Bt₁₀₀], 5% Nt+95% Bt [Nt₀₅Bt₉₅], 10% Nt+90% Bt [Nt₁₀Bt₉₀], 20% Nt+80% Bt [Nt₂₀Bt₈₀], 40% Nt+60% Bt [Nt₄₀Bt₆₀] and 100% Nt rice [Nt₁₀₀]) on target and non‐target pests in a 2‐year field trial in southern China. The occurrence of target pests, Sesamia inferens, Chilo suppressalis and Cnaphalocrocis medinalis, decreased with the increased ratio of Bt rice, and the mixture ratios with more than 90% Bt rice (Bt₁₀₀ and Nt₀₅Bt₉₅) significantly increased the pest suppression efficiency, with the lowest occurrences of non‐target planthoppers, Nilaparvata lugens and Sogatella furcifera in Nt₁₀₀ and Nt₀₅Bt₉₅. Furthermore, there were no significant differences in 1000‐grain dry weight and grain dry weight per 100 plants between Bt₁₀₀ and Nt₀₅Bt₉₅. Seed mixture sowing of Bt rice with ≤10% (especially 5%) of its parent line was sufficient to overcome potential compliance issues that exist with the use of block or structured refuge to provide most effective control of both target and non‐target pests without compromising the grain yield. It is also expected that the strategy of seed mixture sowing with transgenic Bt rice and the non‐transgenic parental line would provide rice yield stability while decreasing the insecticide use frequency in rice production.     

Comparative bioassay of different isolates of Bacillus thuringiensis subsp. kurstaki on the third larval instars of diamondback moth,Plutella xylostella (L.) (Lep.: Plutellidae)

The diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae) is one of the most important pests of cruciferous plants throughout the world. In recent years, this insect has been a serious pest for cabbage fields in Tehran province. Resistance of P. xylostella to all main groups of insecticides has been recorded and it is ranked in the 20 most resistant pest species reported up to now. According to many researchers, to eliminate the problem of pest resistance to chemical pesticides, an integrated pest management programme should be used. In line with this, the uses of microbial control agents (MCAs) are discussed. The bacterium, Bacillus thuringiensis (Bt) is one of microbial control agents of pests. It is characterised by its ability to produce proteic crystalline inclusions during sporulation. Cry1 protein has insecticidal activity and is highly specific to certain insects and not toxic to unrelated insects, plants or vertebrates. In this work, the pathogenicity of some Bt isolates, including Dipel, 20, 29, 79 and 87, was tested against P. xylostella and the lethal concentrations (LC₅₀) of their crystal proteins to P. xylostella third larval instar was determined. The experiment was designed in factorial in randomised complete design with 5 treatments (different concentrations including 10⁴, 10⁵, 10⁶, 10⁷, 10⁸ CFU/ml and 5 replications and with 10 third larval instars. Spore–crystal complex was applied to the surface of natural diets (cabbage leaves) and the mortality of P. xylostella larvae was assessed 120?h after exposure of Bt toxin in each treatment. Results showed that percentage of survival was significantly higher for control treatment. Results also showed that after 5?days, LC₅₀ for isolates of Dipel, 20, 29, 79 and 87 were equal to 1?×?10⁶, 1?×?10⁵, 5?×?10⁵, 4?×?10⁵ and 1?×?10⁴ CFU/ml, respectively. LT₅₀ were equal to 93.71, 48.04, 71, 40.49 and 75.28?h. Of and most the percentage larval mortality relate to attendance 87 and also at least percentage mortality is related to the groom Dipel.     

Lethal,sublethal and transgenerational effects of the novel chiral neonicotinoid pesticide cycloxaprid on demographic and behavioral traits of Aphis gossypii (Hemiptera: Aphididae)

Aphis gossypii Glover (Hemiptera: Aphididae) is a key pest in cotton crops, notably owing to its increasing resistance to commonly used pesticides. Such resistance prompts for the development of integrated pest management (IPM) programs that include novel pesticides being effective against the aphid. In the present study, we assessed lethal and sublethal effects of cycloxaprid, a novel chiral neonicotinoid pesticide developed in China, on A. gossypii. The lethal concentration at 50% (LC₅₀) value of cycloxaprid on A. gossypii was estimated, using the dipping method, at 7.73 mg/L. The impact of a sublethal concentration (LC₁₀) and a lethal concentration (LC₄₀) of cycloxaprid on A. gossypii population growth and feeding behavior (using electrical penetration graph technique [EPG]), and its transgenerational effect were further assessed. Adult longevity and fecundity significantly decreased after exposure to LC₄₀ or LC₁₀ of cycloxaprid. Cycloxaprid with sublethal concentrations (especially LC₄₀) had negative effects on phloem ingestion by A. gossypii. Additionally, the offspring of the adults exposed to LC₄₀ of cycloxaprid had shorter nymphal development duration and adult longevity than the control, and those from LC₁₀ and LC₄₀ treatments had lower adult fecundity and net productive rate. We demonstrated that cycloxaprid is a pesticide showing both lethal and sublethal activities, and transgenerational effects on A. gossypii; it may be useful for implementation in IPM programs against this aphid pest.     

Comparison of Metarhizium isolates for biocontrol of Helicoverpa armigera (Lepidoptera: Noctuidae) in chickpea

Metarhizium isolates from soil (53) and insect hosts (10) were evaluated for extracellular production of cuticle degrading enzyme (CDE) activities such as chitinase, chitin deacetylase (CDA), chitosanase, protease and lipase. Regression analysis demonstrated the relation of CDE activities with Helicoverpa armigera mortality. On basis of this relation, ten isolates were selected for further evaluation. Subsequently, based on LT₅₀ of the 10 isolates towards H. armigera, five isolates were selected. Out of these five isolates, three were selected on the basis of higher conidia production (60–75 g/kg rice), faster sedimentation time (ST₅₀) (2.3–2.65 h in 0.1% (w/v) Tween 80) and lower LC₅₀ (1.4–5.7×10³ conidia/mL) against H. armigera. Finally, three Metarhizium isolates were selected for the molecular fingerprinting using ITS sequencing and RAPD patterning. All three isolates, M34412, M34311 and M81123, showed comparable RAPD patterns with a 935G primer. These were further evaluated for their field performance against H. armigera in a chickpea crop. The percent efficacies with the three Metarhizium isolates were from 65 to 72%, which was comparable to the chemical insecticide, endosulfan (74%).     

Resistant cabbage cultivars change the susceptibility of Plutella xylostella to Bacillus thuringiensis

1 Laboratory studies demonstrated that the susceptibility of larvae of the lepidopteran crucifer pest Plutella xylostella to the insect pathogen Bacillus thuringiensis (Bt) was influenced by the host plant. 2 Larvae reared on the resistant cabbage cultivars Minicole F₁ and Red Drumhead were significantly more susceptible to Bt (the LC₅₀ fell to one half) than larvae fed leaves of susceptible cultivars. 3 However, a third resistant cultivar, Aquarius F₁, had no synergistic effect on Bt‐related mortality. 4 Actual uptake of Bt was monitored in the bioassays, as a preliminary experiment showed that the plant resistance reduced consumption of Bt‐treated leaf discs. However, differences in feeding rate did not explain the observed differences in mortality.     

Isolation and identification of indigenous entomopathogenic nematode (EPN) isolate from Egyptian fauna

Soil samples from different Egyptian Governorates as Giza, Behera, Alexandria, Sohag, Qulupeia, El-Sadat City, Sharkia, Bani-Swif, North Sinai (El-Arish) and South Sinai (Ras-Seder), Tanta, Fayoum and Suez have been surveyed for isolation of new entomopathogenic nematode isolates. One isolate coded as IB was isolated from Sharkia (Belbies). Based on morphometric characters, the isolate was found to belong to Heterorhabditidae and identified as Heterorhabditis indica. The isolate individuals were reproduced by in vivo method using sixth larval instars of Galleria mellonella. The survival of native isolate H. indica was better at low temperature (15 °C) compared to room temperature (27 °C).The bioassay results showed reduction in stored IJs activity against G. mellonella compared with fresh IJs where the median lethal concentration LC₅₀ which recorded to be 46.87 for fresh IJs/ml then increased to 571.38 and to 1181.87 IJs/ml after storage for 2 and 4 weeks, respectively.     

Cloning and characterization of a novel cry8Ab1 gene from Bacillus thuringiensis strain B-JJX with specific toxicity to scarabaeid (Coleoptera: Scarabaeidae) larvae

Isolation of Bacillus thuringiensis (Bt) strain or its cry gene encoding insecticidal crystal protein (ICP) with specific toxicity is of great importance to biological control of insect pests. In this study, by screening 66 strains of Bt isolated from soil samples collected in Shandong Province, China, a new cry8-type gene from Bt strain B-JJX was identified via PCR-RFLP method. This novel gene, cry8Ab1, was cloned from the Bt strain B-JJX and expressed in an acrystalliferous mutant strain HD-73^?. The open reading frame of the cry8Ab1 gene consists of 3543 bp with a G + C content of 37.99% and encodes a protein of 1180 amino acids with a putative MW of 133.3 kDa which was confirmed by SDS-PAGE analysis. The Cry8Ab1 protein was expressed and released as spherical parasporal crystals from Bt acrystalliferous mutant strain HD-73^? along with the presence of spores. In bioassays, this protein was toxic to 3-day-old larvae of the scarabaeid pests, Holotrichia oblita and H. parallela, with an LC₅₀ of 5.72 and 2.00 μg toxin g^?1 soil, respectively. The results are in accordance with the insecticidal activities of the original Bt strain B-JJX, which had an LC₅₀ of 1.72 and 0.96 μg toxin g^?1 soil against H. oblita and H. parallela, respectively.     

The Streptomyces flora of Jordan and its' potential as a source of antibiotics active against antibiotic-resistant Gram-negative bacteria

Detection and identification of members of the genus Streptomyces are of great value because they provide a rich source of antibiotics. Toward the goal of identifying additional novel antibiotics, a total of 292 different Streptomyces isolates were recovered from 54 soil samples collected from 28 different locations in Jordan. These were then characterized by conventional methods and assessed for their activity against two antibiotic-resistant Gram-negative isolates of Escherichia coli and Klebsiella pneumoniae. Results revealed that grey, white and yellow series isolates were the most abundant, with 15% of the Streptomyces isolates active against at least one of the test pathogens. Most of the active isolates exhibited activity against E. coli (96%), while less activity was exhibited against K. pneumoniae (18%). Overall screening revealed the characterization of six Streptomyces isolates (I₇, AC₃₂, G₁₇, Z₁₁, Bb₃₆ and AQ₁₆) which inhibited the growth of both pathogens. All were obtained from a region characterized by low-nutrient soils and harsh conditions. The unusual antibiotic profile of these isolates stressed their potential as a source of novel antibiotics.     

Bt抗虫棉秸秆还田对土壤养分特征的影响

【目的】研究转基因作物秸秆或残茬还田可能对土壤养分特性造成的影响。【方法】以不同抗虫水平Bt棉花和常规棉花(泗棉3号)为研究材料,分别在经过一、二个生长周期后将秸秆机械粉碎后原位还田,40 d后测定分析土壤中Bt蛋白含量及肥力相关的养分含量变化。【结果】Bt棉秸秆还田后,所有品种棉花土壤中Bt蛋白含量与还田前无显著增加,且转Bt基因棉与非转基因棉还田对土壤Bt蛋白含量的影响并无显著差异。同时,棉秸秆还田可显著提高土壤有机质、速效磷、碱解氮、速效钾、全氮、全磷和全钾含量,提升土壤pH值;增加幅度在不同抗虫水平Bt棉花间及与非转基因常规棉花品种间皆无显著性差异。【结论】秸秆还田对土壤肥力的提升与Bt棉的抗虫水平无关。“转Bt基因”不成为Bt棉秸秆还田提高土壤肥力的限制性因素,其秸秆还田不会对土壤肥力质量产生负面影响,可使土壤养分含量增加,有效提升土壤肥力。秸秆原位还田简单、无害又提升肥力,有条件作为转Bt基因植物秸秆无害化处理的理想方式。     

Bacillus thuringiensis Isolates from Great Nicobar Islands

Bacillus thuringiensis (Bt) strains were isolated from soil samples of Great Nicobar Islands, one of the “hottest biodiversity hotspots,” where no collection has been characterized previously. The 36 new Bt isolates were obtained from 153 samples analyzed by crystal protein production with light/phase-contrast microscopy, determination of cry gene profile by SDS-PAGE, evaluation of toxicity against Coleopteran, and Lepidopteran insect pests, finally cloning and sequencing. Majority of the isolates showed the presence of 66–35 kDa protein bands on SDS-PAGE while the rest showed >130, 130, 73, and 18 kDa bands. The variations in crystal morphology and mass of crystal protein(s) purified from the isolates of Bt revealed genetic and molecular diversity. Based on the toxicity test, 50 % of isolates were toxic to Ash weevils, 16 % isolates were toxic to cotton bollworm, 38 % isolates were toxic both to ash weevil as well as cotton bollworm, while 11 % of the isolates did not exhibit any toxicity. PCR analysis unveiled prepotency of cry1B- and cry8b-like genes in these isolates. This study appoints the first isolation and characterization of local B. thuringiensis isolates in Great Nicobar Islands. Some of these isolates display toxic potential and, therefore, could be adopted for future applications to control some agriculturally important insect pests in the area of integrated pest management for sustainable agriculture.     

Molecular and biological characterization of native <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> strains for controlling tomato leafminer (<Emphasis Type="Italic">Tuta absoluta</Emphasis> Meyrick) (Lepidoptera: Gelechiidae) in Colombia

Twenty-eight soil samples were obtained from open fields and greenhouses used for tomato cultivation in various regions of Colombia. For functional characterization, 99 Bacillus thuringiensis (Bt) strains were isolated and characterized by abundance and morphology of microscopic crystals, SDS–PAGE of protein extracts and M-PCR analyses of genes of the cry1 family, as well as for their insecticidal activity against Tuta absoluta second instar larvae. Native Bt strains had amorphous (5%), bi-pyramidal (27%), square (8%), spherical (38%) and triangular (22%) crystal forms. Based on the presence of 1–4 different crystal forms, 18 different profiles were established. The SDS–PAGE analyses of protein extracts established ten different strain groups based on their protein band weight and potential biological activity. The M-PCR technique identified 35 native Bt strains based on the presence of the 6 genes cry1Aa, cry1Ab, cry1Ac, cry1B, cry1C and cry1D, whose frequency of occurrence was 76, 26, 21, 35, 32 and 8.8%, respectively. Thirteen different PCR profiles were found in native Bt strains. Several gene combinations tended to co-occur with elevated frequency, such as the pairs cry1Ac/cry1C, cry1Ab/cry1Ac and cry1Ab/cry1B, for which Pearson correlation coefficients were 0.69, 0.52 and 0.54, respectively. Native strains ZBUJTL39 and ZCUJTL11 had up to three times higher biological activity against T. absoluta second instar larvae than the reference strain Bt var. kurstaki HD1, with an LD₅₀ of 2.4 μg/ml (P < 0.05) for native Bt strain ZCUJTL11. This study suggests a high biodiversity of native Bt strains from tomato growing regions in Colombia, which has important implications for designing biological control strategies for T. absoluta.     

Distribution and diversity of Dipteran-specific cry and cyt genes in native Bacillus thuringiensis strains obtained from different ecosystems of Iran

One hundred and twenty-eight Bacillus thuringiensis isolates from fields of different ecological regions of Iran were collected to study the distribution and diversity of Dipteran-specific cry and cyt genes. The percentage of samples with Bt showed significant differences between different regions and also between different fields. The most Bt frequency was observed in the soil samples collected from Caspianic zone (7%) and soils of cotton (17%). Characterization of isolates was based on morphological characteristics of crystals, plasmid profiles and protein band patterns as well as PCR analysis using general and specific primers for 22 different cry and cyt genes encoding proteins active against mosquitoes. Thirty-eight different cry gene profiles were detected in this collection. Several of them were found to be different from all previously published profiles and none of the previous researches reported these numbers of profiles. Strains containing cry2-type genes were the most abundant and represent 57.1% of the isolates. Strains harboring cry24 and cry10 genes were also highly abundant (38.7 and 32.8%, respectively). cry11, cry4, cry17, cry19, cry21, cry29, cyt1, and cry9 genes were less abundant, found in 25.7, 14.3, 11.4, 1.4, 4.3, 1.4, and 10% of the strains, respectively. Among the cry2 gene containing isolates, 37.5% strains harbored cry2Aa, 55% cry2Ab, 2.5% cry2Ac, and 5% other or novel cry2-type genes. Among the cry4 gene containing isolates, 0% strains harbored cry4A, 60% cry4B, and 40% cry4C, cry4D or novel cry4 type genes. Finally, based on crystal morphology, protein patterns and PCR, 21 strains were selected as potentially high Dipteran-active for bioassays. Also our results showed that some of the isolates may harbor minimum a putative novel cry gene.     

In vitro evaluation of some bacterial isolates as biofertilizers and biocontrol agents against the second stage juveniles of <Emphasis Type="Italic">Meloidogyne incognita</Emphasis>

Representative soil samples were collected from soil apart and the rhizosphere of different plant varieties grown in five Egyptian Governorates. The presence of plant parasitic nematodes (PPN) and free living nematodes (FLN) in the collected samples were estimated. In addition, one hundred and 65 bacterial cultures which well known as biofertilizers (some are able to fix nitrogen and the others solubilize either phosphate or potassium) were isolated. The isolated bacteria were screened depending on their rate of growth. Thirty-five cultures of fast growing nitrogen fixing bacteria (NFB), phosphate solubilizing bacteria (PSB) and potassium solubilizing bacteria (KSB) and their cultural filtrates were tested in vitro as biocontrol agents against the second stage juvenile (J_2s) of the Meloidogyne incognita. In general, higher mortality percentages of nematodes were recorded by bacterial cultures than their comparative cultural filtrates. The highest mortality percentages were recorded by cultures of NFB7, PSB2 and KSB2 (100% at dilution 1/10), while it was 99.3, 99 and 97.8% at dilution 1/100, respectively. NFB7 exhibited a high nitrogen fixation rate (4.2 μmole N₂/mL/h), while PSB2 and KSB2 effectively solubilized phosphate and potassium comparing with the control treatments (1.94 fold of available phosphate and 2.0 fold of available potassium, respectively). NFB7, PSB2 and KSB2 isolates showed the highest protease, gelatinase and chitinase activities which were thought responsible for their nematicidal effect. The three bacterial isolates were identified as Paenibacillus polymyxa, Bacillus megaterium and Bacillus circulans, respectively.     

Plasmid incidence in bacteria from agricultural and industrial soils

Heavy metal contents of agricultural and industrial soils were determined by atomic absorption spectrophotometry. The analysis of the samples collected from two different locations revealed significantly high levels of Fe, Zn, Cu, Cr and Ni. Certain microbiological parameters (total aerobic heterotrophs, asymbiotic N₂-fixers, total Actinomycetes and fungi) were also monitored from these soils. A total of 70 bacterial isolates from agricultural and industrial soils were examined for plasmid DNA content and resistance to the antibiotics amoxycillin, cloxacillin, chloramphenicol, doxycycline methicillin, nalidixic acid, and tetracycline. Minimum inhibitory concentrations (MICs) of Cu, Cr, Pb, Cd, Hg, Zn, and Ni for each isolate were also determined. Resistance was most frequent to methicillin (48.5%), cloxacillin (45.7%), and nalidixic acid (40%) for all isolates of bacteria. The highest MICs observed were 100 g/ml for mercury, 800 g/ml for Ni and 1600 g/ml for other metals. The incidences of metal resistance and MICs of metals for bacteria from industrial soil were significantly different to those of agricultural soil. On a percentage basis, 91.4% of the total bacterial isolates from industrial soil were found to harbour plasmids whereas 40% of the isolates from agricultural soil contained plasmids.