Morphological, morphometrical and molecular identification of root-knot nematode (Meloidogyne javanica) infecting banana in Assiut governorate, Egypt

Plant-parasitic nematodes are extremely dangerous pests in a variety of economically important crops. Root-knot nematodes (RKN) Meloidogyne spp. are between the major important pests causing serious crops havoc worldwide because of their wide geographical distribution and variety of hosts. Therefore, both of identification that is true and trustworthy of these pests is required for evaluating various suitable management strategies. This study, aimed to characterize morphological, morphometric and molecularly isolate of Meloidogyne related to banana plants. Second-stage juveniles (body length, tail length, stylet length, hyaline terminus length, and DEGO) were used in morphometric and morphological studies and female in perineal patterns. The results revealed that the identified nematode species, Meloidogyne javanica, is the most common root knot nematode species in all three localities. To prove the identification, a polymerase chain reaction (PCR)-based experiment using a species-specific sequence described amplified regions (SCAR) primer series was used. The Fjav/Rjav primer effectively enhanced SCAR markers of 670 bp previously identified in M. javanica. This study confirms the use of an effective and reliable diagnosis of RKN using the three approaches

Pore-forming pyocin S5 utilizes the FptA ferripyochelin receptor to kill Pseudomonas aeruginosa.

Pyocins are toxic proteins produced by some strains of Pseudomonas aeruginosa that are lethal for related strains of the same species. Some soluble pyocins (S2, S3 and S4) were previously shown to use the pyoverdine siderophore receptors to enter the cell. The P. aeruginosa PAO1 pore-forming pyocin S5 encoding gene (PAO985) was cloned into the expression vector pET15b, and the affinity-purified protein product tested for its killing activity against different P. aeruginosa strains. The results, however, did not show any correlation with a specific ferripyoverdine receptor. To further identify the S5 receptor, transposon mutants were generated. Pooled mutants were exposed to pyocin S5 and the resistant colonies growing in the killing zone were selected. The majority of S5-resistant mutants had an insertion in the fptA gene encoding the receptor for the siderophore pyochelin. Complementation of an fptA transposon mutant with the P. aeruginosa fptA gene in trans restored the sensitivity to S5. In order to define the receptor-binding domain of pyocin S5, two hybrid pyocins were constructed containing different regions from pyocin S5 fused to the C-terminal translocation and DNase killing domains of pyocin S2. Only the protein containing amino acid residues 151 to 300 from S5 showed toxicity, indicating that the pyocin S5 receptor-binding domain is not at the N-terminus of the protein as in other S-type pyocins. Pyocin S5 was, however, unable to kill Burkholderia cenocepacia strains producing a ferripyochelin FptA receptor, nor was the B. cenocepacia fptA gene able to restore the sensitivity of the resistant fptA mutant P. aeruginosa strain

Genetic and morpho-physiological analyses of the tolerance and recovery mechanisms in seedling stage spring wheat under drought stress

Drought is one of the complex abiotic stresses that affect the growth and production of wheat in arid and semiarid countries. In this study, a set of 172 diverse spring wheat genotypes from 20 different countries were assessed under drought stress at the seedling stage. Besides seedling length, two types of traits were recorded, namely: tolerance traits (days to wilting, leaf wilting, and the sum of leaf wilting), and recovery traits (days to regrowth, regrowth biomass, and drought survival rate). In addition, tolerance index, recovery index, and drought tolerance index (DTI) were estimated to select the most drought tolerant genotypes. Moreover, leaf protein content (P), amino acid (AM), proline content (PRO), glucose (G), fructose (F), and total soluble carbohydrates (TSC) were measured under control and drought conditions to study the changes in each physiological trait due to drought stress. All genotypes showed a high significant genetic variation in all the physio-morphological traits scored under drought stress. High phenotypic and genotypic correlations were found among all seedling morphological traits. Among the studied indices, the drought tolerance index (DTI) had the highest phenotypic and genotypic correlations with all tolerance and recovery traits. The broad-sense heritability (H<sup>2</sup>) estimates were high for morphological traits (83.85–92.27), while the physiological traits ranged from 96.41 to 98.68 under the control conditions and from 97.13 to 99.99 under drought stress. The averages of the physiological traits (proteins, amino acids, proline, glucose, fructose, and total soluble carbohydrates) denoted under drought stress were higher than those recorded under well-watered conditions except for proteins. In this regard, amino acids, glucose, and total soluble carbohydrates had a significant correlation with all morphological traits. The selection for drought tolerance revealed 10 tolerant genotypes from different countries (8 genotypes from Egypt, one from Morocco, and one from the United States). These selected genotypes were screened for the presence of nine specific TaDREB1 alleles. Six primers were polymorphic among the selected genotypes. Genetic diversity among the selected genotypes was investigated using 21,450 SNP markers. The results of the study shed light on the different mechanisms for drought tolerance that wheat plants use to tolerate and survive under drought stress. The genetic analysis performed in this study suggested the most suitable genotypes for selective breeding at the seedling stage under water deficit

Toxicity Evaluation and Genetic Improvement of Bacillus thuringiensis Isolated from Different Regions in Assiut, Egypt against Mosquito Larvae

This study aims to evaluate the toxicity and genetic improvement of Bacillus thuringiensis isolates. Isolates were obtained from soil, insect and water samples from different regions of Assiut, Egypt for biological control of mosquito larvae. B. thuringiensis colonies were identified based on morphological and then by PCR which detect the Cry toxic genes in the isolates. Bioassays were performed to evaluate the toxicity of different strains of B. thuringiensis against mosquito larvae such as (Culex spp). In general, 36 B. thuringiensis isolates were obtained (31 from soil, 4 from insects, and 1 from water). And they were all toxic to mosquito larvae with different mortality percentages from 7 to 97% after 48 hours. Isolate Am2 recorded the highest mortality percentage 97% and Mn3 lowest mortality percentage 7%. PCR revealed that Am2 isolate which caused the highest mortality encodes three different types of Cry toxins, Cyt1AA, Cry1Ac and Cry2Aa. This isolate Am2 was examined by scanning electron microscopy to observe the shape of the Cry proteins. The results showed that the Am2 isolate contained of spherical and cuboidal toxic proteins. Then UV-mutagenesis was performed on the Am2 isolate to improve its toxicity. Out of 30 obtained UV-mutants, only one mutant showed improvement in the mortality of mosquito larvae since it caused a mortality rate of 100%. The results of the present study revealed the larvicidal efficacy of B. thuringiensis (Am2) isolate found in the soil of Assiut, could be used in biological control program of mosquito larvae

The t6A Modification Acts as a Positive Determinant for the Anticodon Nuclease Prrc, and is Distinctively Nonessential in Streptococcus mutans

Endoribonuclease toxins (ribotoxins) are produced by bacteria and fungi to respond to stress, eliminate non-self competitor species, or interdict virus infection. PrrC is a bacterial ribotoxin that targets and cleaves tRNALys UUU in the anticodon loop. In vitro studies suggested that the post-transcriptional modification threonylcarbamoyl adenosine (t6A) is required for PrrC activity but this prediction had never been validated in vivo. Here, by using t6A-deficient yeast derivatives, it is shown that t6A is a positive determinant for PrrC proteins from various bacterial species. Streptococcus mutans is one of the few bacteria where the t6A synthesis gene tsaE (brpB) is dispensable and its genome encodes a PrrC toxin. We had previously shown using an HPLC-based assay that the S. mutans tsaE mutant was devoid of t6A. However, we describe here a novel and a more sensitive hybridization-based t6A detection method (compared to HPLC) that showed t6A was still present in the S. mutans ΔtsaE, albeit at greatly reduced levels (93% reduced compared to WT). Moreover, mutants in two other S. mutans t6A synthesis genes (tsaB and tsaC) were shown to be totally devoid of the modification thus confirming its dispensability in this organism. Furthermore, analysis of t6A modification ratios and of t6A synthesis genes mRNAs levels in S. mutans suggest they may be regulated by growth phase

Exploring the mechanisms of endophytic bacteria for suppressing early blight disease in tomato (Solanum lycopersicum L.)

Controlling early blight of tomatoes using endophytic bacteria is an eco-friendly and sustainable approach to manage this common fungal disease caused by Alternaria solani, Alternaria alternata, and Curvularia lunata. Endophytic bacteria are microorganisms that live inside plant tissues without causing harm and can help protect the host plant from pathogens. In this work, twenty endophytic bacterial isolates from tomato healthy plants were tested against pathogenic fungal isolates that caused early blight disease in vitro. Out of the 20 tested isolates, three (B4, B7, and B17) were considered effective isolates against the growth of fungal pathogens. The three isolates were recognized as Enterobacter cloacae HS-6 (B4), Pseudomonas gessardii HS-5 (B 7), and Pseudomonas mediterranea HS-4 (B17) using 16s-rDNA sequencing. Different concentrations of bacterial cultural diltrates at 20, 40, and 60% were tested for their antagonistic effects on the development of pathogenic fungi in vitro. The lowest dry weights of pathogenic isolates in all bacterial culture filtrates were discovered at 60%. In all culture filtrates, phenolic compounds showed the largest peak area. Under greenhouse conditions, the least disease severity of tomato early blight was found for E. cloacae and its culture filtrate compared to other treatments. Real-time PCR was used to examine the expression pattern of the defense response gene β-1.3 glucanase gene in infected tomato plants with pathogenic fungi (control) as well as its relations with efficient biocontrol agent (E. cloacae). The expression of the gene increased substantially and significantly after three days from the inoculation-infected plants with C. lunata and E. cloacae while it reached the maximum after five days from the inoculation with A. alternata, A. solani and E. cloacae. Our study concluded that the endophytic bacterial isolate E. cloacae can be considered a promising biocontrol agent for preventing tomato early blight

Data_Sheet_1_Exploring the mechanisms of endophytic bacteria for suppressing early blight disease in tomato (Solanum lycopersicum L.).zip

Controlling early blight of tomatoes using endophytic bacteria is an eco-friendly and sustainable approach to manage this common fungal disease caused by Alternaria solani, Alternaria alternata, and Curvularia lunata. Endophytic bacteria are microorganisms that live inside plant tissues without causing harm and can help protect the host plant from pathogens. In this work, twenty endophytic bacterial isolates from tomato healthy plants were tested against pathogenic fungal isolates that caused early blight disease in vitro. Out of the 20 tested isolates, three (B4, B7, and B17) were considered effective isolates against the growth of fungal pathogens. The three isolates were recognized as Enterobacter cloacae HS-6 (B4), Pseudomonas gessardii HS-5 (B 7), and Pseudomonas mediterranea HS-4 (B17) using 16s-rDNA sequencing. Different concentrations of bacterial cultural diltrates at 20, 40, and 60% were tested for their antagonistic effects on the development of pathogenic fungi in vitro. The lowest dry weights of pathogenic isolates in all bacterial culture filtrates were discovered at 60%. In all culture filtrates, phenolic compounds showed the largest peak area. Under greenhouse conditions, the least disease severity of tomato early blight was found for E. cloacae and its culture filtrate compared to other treatments. Real-time PCR was used to examine the expression pattern of the defense response gene β-1.3 glucanase gene in infected tomato plants with pathogenic fungi (control) as well as its relations with efficient biocontrol agent (E. cloacae). The expression of the gene increased substantially and significantly after three days from the inoculation-infected plants with C. lunata and E. cloacae while it reached the maximum after five days from the inoculation with A. alternata, A. solani and E. cloacae. Our study concluded that the endophytic bacterial isolate E. cloacae can be considered a promising biocontrol agent for preventing tomato early blight.</p

The t⁶A modification acts as a positive determinant for the anticodon nuclease PrrC, and is distinctively nonessential in <i>Streptococcus mutans</i>

<p>Endoribonuclease toxins (ribotoxins) are produced by bacteria and fungi to respond to stress, eliminate non-self competitor species, or interdict virus infection. PrrC is a bacterial ribotoxin that targets and cleaves tRNA^Lys _UUU in the anticodon loop. <i>In vitro</i> studies suggested that the post-transcriptional modification threonylcarbamoyl adenosine (t⁶A) is required for PrrC activity but this prediction had never been validated <i>in vivo</i>. Here, by using t⁶A-deficient yeast derivatives, it is shown that t⁶A is a positive determinant for PrrC proteins from various bacterial species. <i>Streptococcus mutans</i> is one of the few bacteria where the t⁶A synthesis gene <i>tsaE</i> (<i>brpB</i>) is dispensable and its genome encodes a PrrC toxin. We had previously shown using an HPLC-based assay that the <i>S. mutans tsaE</i> mutant was devoid of t⁶A. However, we describe here a novel and a more sensitive hybridization-based t⁶A detection method (compared to HPLC) that showed t⁶A was still present in the <i>S. mutans</i> Δ<i>tsaE</i>, albeit at greatly reduced levels (93% reduced compared with WT). Moreover, mutants in 2 other <i>S. mutans</i> t⁶A synthesis genes (<i>tsaB</i> and <i>tsaC</i>) were shown to be totally devoid of the modification thus confirming its dispensability in this organism. Furthermore, analysis of t⁶A modification ratios and of t⁶A synthesis genes mRNA levels in <i>S. mutans</i> suggest they may be regulated by growth phase.</p