Post Inoculation Research Articles

Sunflower resistance against Sclerotinia sclerotiorum is potentiated by selenium through regulation of redox homeostasis and hormones signaling pathways.

White mold of sunflower caused by Sclerotinia sclerotiorum is a devastating disease that causes serious yield losses. Selenium (Se) helps plants resist stress. In this study, the resistance of sunflower to S. sclerotiorum was improved after foliar application of selenite. Selenite sprayed on leaves can be absorbed by sunflowers and transformed to selenomethionine. Consequently, sunflowers treated with Se exhibited a delay in lesion development with decrease by 54% compared to mock inoculation at 36-h post inoculation (hpi). In addition, treatment with Se compromised the adverse effects caused by S. sclerotiorum infection by balancing the regulation of genes involved in redox homeostasis. In particular, cat expression on leaves treated with Se increased to 2.5-fold to alleviate the downregulation caused by S. sclerotiorum infection at 12 hpi. Additionally, apx expression on leaves treated with Se decreased by 36% to alleviate the upregulation caused by S. sclerotiorum infection at 24 hpi, whereas expressions of gpx, pox, and nox on leaves treated with Se also successively decreased by approximately 40-60% to alleviate the upregulation caused by S. sclerotiorum infection at 24 and 36 hpi, respectively. The use of Se also enhanced the regulation of genes involved in hormones signaling pathways, in which expressions of AOC and PAL increased to 2.0- and 1.5-fold, respectively, to enhance the upregulation caused by S. sclerotiorum infection at 12 hpi, whereas expressions of AOC and PDF1.2 increased to 2.7- and 1.8-fold at 24 hpi, respectively. In addition, EIN2 expression on leaves treated with Se increased to 1.8-, 2.0-, and 1.5-fold to alleviate the downregulation caused by S. sclerotiorum infection. These results suggest that Se can improve sunflower defense responses against S. sclerotiorum infection aiming a sustainable white mold management.

Variation in Peperomia pellucida growth and secondary metabolism after rhizobacteria inoculation.

Peperomia pellucida L. Kunth is a herb well-known for its secondary metabolites (SM) with biological potential. In this study, the variations in the SM of P. pellucida during association with rhizobacteria were evaluated. Plants were inoculated with Enterobacter asburiae and Klebsiella variicola, which were identified by sequencing of the 16S rRNA gene. The data were evaluated at 7, 21, and 30-day post inoculation (dpi). Plant-bacteria symbiosis improved plant growth and weight. Total phenolic content and phenylalanine ammonia lyase enzyme activity had a significant increase mainly at 30 dpi. P. pellucida was mainly composed of phenylpropanoids (37.30–52.28%) and sesquiterpene hydrocarbons (39.28–49.42%). The phenylpropanoid derivative 2,4,5-trimethoxy-styrene (ArC2), the sesquiterpene hydrocarbon ishwarane, and the phenylpropanoid dillapiole were the major compounds. Principal component analysis (PCA) of the classes and compounds ≥ 2.0% indicated that plants colonized by E. asburiae had a reduction in the content of sesquiterpene hydrocarbons and an increase in phenylpropanoids and derivatives. Plants treated with this bacterium also had an increase in the content of 2,4,5-trimethoxystyrene at 30 dpi. Plants inoculated with K. variicola had significant increases only in the content of the classes monoterpene hydrocarbons and ‘other compounds’ (hydrocarbons, esters, ketones, etc.). These data suggest that the production of plant secondary metabolites can be modified depending on the type of rhizobacteria inoculated.

Supplementation of oligosaccharide-based polymer enhanced growth and disease resistance of weaned pigs by modulating intestinal integrity and systemic immunity

BackgroundThere is a great demand for antibiotic alternatives to maintain animal health and productivity. The objective of this experiment was to determine the efficacy of dietary supplementation of a blood group A6 type 1 antigen oligosaccharides-based polymer (Coligo) on growth performance, diarrhea severity, intestinal health, and systemic immunity of weaned pigs experimentally infected with an enterotoxigenic Escherichia coli (ETEC), when compared with antibiotics.ResultsPigs in antibiotic carbadox or Coligo treatment groups had greater (P < 0.05) body weight on d 5 or d 11 post-inoculation (PI) than pigs in the control group, respectively. Supplementation of antibiotics or Coligo enhanced (P < 0.05) feed efficiency from d 0 to 5 PI and reduced (P < 0.05) frequency of diarrhea throughout the experiment, compared with pigs in the control group. Supplementation of antibiotics reduced (P < 0.05) fecal β-hemolytic coliforms on d 2, 5, and 8 PI. Pigs in antibiotics or Coligo groups had reduced (P < 0.05) neutrophil counts and serum haptoglobin concentration compared to pigs in the control group on d 2 and 5 PI. Pigs in Coligo had reduced (P < 0.05) total coliforms in mesenteric lymph nodes on d 5 and 11 PI, whereas pigs in antibiotics or Coligo groups had reduced (P < 0.05) total coliforms in spleen on d 11 PI compared with pigs in the control group. On d 5 PI, pigs in the Coligo group had greater (P < 0.05) gene expression of ZO1 in jejunal mucosa, but less (P < 0.05) mRNA expression of IL1B, IL6, and TNF in ileal mucosa, in comparison with pigs in the control group. Supplementation of antibiotics enhanced (P < 0.05) the gene expression of OCLN in jejunal mucosa but decreased (P < 0.05) IL1B and IL6 gene expression in ileal mucosa, compared with the control. On d 11 PI, supplementation of antibiotics or Coligo up-regulated (P < 0.05) gene expression of CLDN1 in jejunal mucosa, but Coligo reduced (P < 0.05) IL6 gene expression in ileal mucosa compared to pigs in the control group.ConclusionsSupplementation of Coligo improved growth performance, alleviated diarrhea severity, and enhanced gut health in weaned pigs infected with ETEC F18 in a manner similar to in-feed antibiotics.

The Potential of Molecular Indicators of Plant Virus Infection: Are Plants Able to Tell Us They Are Infected?

To our knowledge, there are no reports that demonstrate the use of host molecular markers for the purpose of detecting generic plant virus infection. Two approaches involving molecular indicators of virus infection in the model plant Arabidopsis thaliana were examined: the accumulation of small RNAs (sRNAs) using a microfluidics-based method (Bioanalyzer); and the transcript accumulation of virus-response related host plant genes, suppressor of gene silencing 3 (AtSGS3) and calcium-dependent protein kinase 3 (AtCPK3) by reverse transcriptase-quantitative PCR (RT-qPCR). The microfluidics approach using sRNA chips has previously demonstrated good linearity and good reproducibility, both within and between chips. Good limits of detection have been demonstrated from two-fold 10-point serial dilution regression to 0.1 ng of RNA. The ratio of small RNA (sRNA) to ribosomal RNA (rRNA), as a proportion of averaged mock-inoculation, correlated with known virus infection to a high degree of certainty. AtSGS3 transcript decreased between 14- and 28-days post inoculation (dpi) for all viruses investigated, while AtCPK3 transcript increased between 14 and 28 dpi for all viruses. A combination of these two molecular approaches may be useful for assessment of virus-infection of samples without the need for diagnosis of specific virus infection.

Oral and ocular transmission of severe fever with thrombocytopenia syndrome virus

BackgroundSevere fever with thrombocytopenia syndrome virus (SFTSV) is a tick-borne bunyavirus that could cause a severe hemorrhagic fever termed SFTS with a high fatality rate of up to 30%. Importantly, SFTSV is frequently transmitted from person-to-person and patients’ blood or excreta are considered as the risk factors for transmission of SFTSV. However, the mechanism of person-to-person transmission of SFTSV is still elusive. MethodsIn this study, wild-type (WT) C57BL/6 J mice and a lethal SFTSV mouse model IFNAR−/− A129 mice were utilized to evaluate whether SFTSV could be transmitted via oral or ocular routes. C57BL/6 J mice were inoculated with cell-cultured SFTSV via oral and ocular inoculation. IFNAR−/− A129 mice were inoculated with cell-cultured SFTSV or SFTSV infected mouse acute sera via oral and ocular inoculation. ResultsWe found that SFTSV antibody positive rates in C57BL/6 J mice were 70% (7/10) and 30% (3/10) in the oral inoculation group and ocular inoculation group, respectively on day 21 post SFTSV inoculation. The mortality rates of IFNAR−/− mice with oral and ocular inoculation of cell-cultured SFTSV were 100% and 83.33% (5/6), respectively on day 6 post inoculation. The mortality rates of IFNAR−/− mice with oral and ocular inoculation of SFTSV infected mouse acute serum were 100% and 66.67% (4/6), respectively on day 9 post inoculation. ConclusionsTogether, our results show that SFTSV can be transmitted effectively through oral and ocular membrane, suggesting exposure to SFTS positive excreta may be a high-risk factor of nosocomial transmission of SFTSV in hospitals and/or families. Family members and healthcare workers should be protected properly during taking care of SFTS patients to prevent SFTSV nosocomial infection.

Senecavirus A- and Non-Infected Cells at Early Stage of Infection: Comparative Metabolomic Profiles

Senecavirus A (SVA), classified into the genus Senecavirus in the family Picornaviridae, causes an infectious disease in pigs. This virus can efficiently replicate in some non-pig-derived cells, such as the BHK cell line and its derivative (BSR-T7/5 cell line). We had recovered a wild-type SVA from its cDNA clone previously, and then uncovered the proteomic profile of SVA-infected BSR-T7/5 cells at 12 h post inoculation (hpi). In order to explore the cellular metabolomics further, the SVA-inoculated BSR-T7/5 cell monolayer was collected at 12 hpi for assay via liquid chromatography-tandem mass spectrometry (LC-MS/MS). The resultant data set was comprehensively analyzed using bioinformatics tools. A total of 451 metabolites were identified using in-house and public databases. Out of these metabolites, sixty-one showed significantly differential values (p value < 0.05). The Kyoto Encyclopedia of Genes and Genomes (KEGG) database was used to analyze metabolic pathways of the significantly differential metabolites. There were eighty-one identified KEGG pathways, out of which twenty-seven showed their p values < 0.05. The pyrimidine metabolism revealed the minimum p value and the maximum number of significantly differential metabolites, implying the pyrimidine played a key role in cellular metabolism after SVA infection. SVA replication must rely on the cellular metabolism. The present study on metabolomics would shed light on impacts of SVA-induced multiple interactions among metabolites on cells or even on natural hosts.

Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold Value.

Coronavirus disease 2019 (COVID-19) is an emerging life-threatening pulmonary disease caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which originated in Wuhan, Hubei Province, China, in December 2019. COVID-19 develops after close contact via inhalation of respiratory droplets containing SARS-CoV-2 during talking, coughing, or sneezing by asymptomatic, presymptomatic, and symptomatic carriers. This virus evolved over time, and numerous genetic variants have been reported to have increased disease severity, mortality, and transmissibility. Variants have also developed resistance to antivirals and vaccination and can escape the immune response of humans. Reverse transcription polymerase chain reaction (RT–PCR) is the method of choice among diagnostic techniques, including nucleic acid amplification tests (NAATs), serological tests, and diagnostic imaging, such as computed tomography (CT). The limitation of RT–PCR is that it cannot distinguish fragmented RNA genomes from live transmissible viruses. Thus, SARS-CoV-2 isolation by using cell culture has been developed and makes important contributions in the field of diagnosis, development of antivirals, vaccines, and SARS-CoV-2 virology research. In this research, two SARS-CoV-2 strains were isolated from four RT–PCR-positive nasopharyngeal swabs using VERO E6 cell culture. One isolate was cultured successfully with a blind passage on day 3 post inoculation from a swab with a Ct > 35, while the cells did not develop cytopathic effects without a blind passage until day 14 post inoculation. Our results indicated that infectious SARS-CoV-2 virus particles existed, even with a Ct > 35. Cultivable viruses could provide additional consideration for releasing the patient from quarantine. The results of the whole genome sequencing and bioinformatic analysis suggested that these two isolates contain a spike 68-76del+spike 675-679del double-deletion variation. The double deletion was confirmed by amplification of the regions spanning the spike gene deletion using Sanger sequencing. Phylogenetic analysis revealed that this double-deletion variant was rare (one per million in public databases, including GenBank and GISAID). The impact of this double deletion in the spike gene on the SARS-CoV-2 virus itself as well as on cultured cells and/or humans remains to be further elucidated.

Regulatory Mechanisms of the Resistance to Common Bacterial Blight Revealed by Transcriptomic Analysis in Common Bean (Phaseolus vulgaris L.).

Common bean blight (CBB), primarily caused by Xanthomonas axonopodis pv. phaseoli (Xap), is one of the most destructive diseases of common bean (Phaseolus vulgaris L.). The tepary bean genotype PI 319443 displays high resistance to Xap, and the common bean genotypes HR45 and Bilu display high resistance and susceptibility to Xap, respectively. To identify candidate genes related to Xap resistance, transcriptomic analysis was performed to compare gene expression levels with Xap inoculation at 0, 24, and 48 h post inoculation (hpi) among the three genotypes. A total of 1,146,009,876 high-quality clean reads were obtained. Differentially expressed gene (DEG) analysis showed that 1,688 DEGs responded to pathogen infection in the three genotypes. Weighted gene coexpression network analysis (WGCNA) was also performed to identify three modules highly correlated with Xap resistance, in which 334 DEGs were likely involved in Xap resistance. By combining differential expression analysis and WGCNA, 139 DEGs were identified as core resistance-responsive genes, including 18 genes encoding resistance (R) proteins, 19 genes belonging to transcription factor families, 63 genes encoding proteins with oxidoreductase activity, and 33 plant hormone signal transduction-related genes, which play important roles in the resistance to pathogen infection. The expression patterns of 20 DEGs were determined by quantitative real-time PCR (qRT-PCR) and confirmed the reliability of the RNA-seq results.

Cyclosporine A increases the intensity of Toxocara canis infection in swiss mice.

Toxocariasis is a zoonotic disease of worldwide distribution. The connection between parasitic diseases and conditions that depress the immune system, such as the use of immunosuppressive drugs, has been studied. The purpose of this study was to evaluate the effect of Cyclosporine A (CsA) on the intensity of infection, humoral response and gene transcription of interleukins IL-4, IL-10 and IL-12 in mice experimentally infected with Toxocara canis. To this end, mice were divided into two groups treated with CsA (G1: 10 mg/Kg and G2: 50 mg/kg), the G3 and G4 group received PBS. After the last administration of the drug or PBS (orally every 48 hours for 15 days), groups G1, G2 and G3 were inoculated with 1200 eggs of T. canis. Was collected blood samples on days zero, 15 and 30 days post-inoculation (PI), for ELISA test and the mice were euthanized 30 days PI. The organs and striated muscle tissue were collected for the recovery of larvae. The splenocytes were analyzed by RT-PCR. The intensity of infection in the mice treated with 50 mg/kg of CsA was 65.5% higher than in the control group (p=0.001). An analysis of the kinetics of anti-Toxocara antibody revealed that the groups treated with CsA showed significantly higher mean levels of antibodies on day 15 PI. The transcription of the three tested interleukins showed no statistical difference between G2 and G3 (control). It was concluded that the immunosuppression triggered by CsA (50 mg/Kg) favored the establishment of a larger number of T. canis larvae without, however, altering immunoglobulin production and IL-4, IL-10 and IL-12 transcription on day 30 PI.

Trial to control infectious bursal disease virus using iron oxide chitosan nanocomposite in broiler chicken

The present study attempted to control infectious bursal disease virus (IBDV) with iron oxide chitosan nanocomposite as an antiviral compound. The iron oxide chitosan nanocomposite was prepared using the co-precipitation method and characterized by high-resolution transmission electron microscopy (HR-TEM) and cytotoxicity assay on Vero cells. To evaluate the effectiveness of iron oxide chitosan nanocomposite against IBDV, eighty 21-day-old Baladi broiler chicks were divided into four groups (G1-G4). Chicks kept in G1 served as a negative control, while chicks kept in G2 (challenged-non-treated) were challenged orally with IBDV 103EID\50/bird). Chicks kept in G3 (challenged-treated) were challenged orally with 1103EID\50 IBDV/bird and received 1 mL of iron oxide chitosan nanocomposite (1 mg dissolved in 100 mL water) for three days post-inoculation (PI) while chicks kept in G4 (non-challenged-treated) received the same dose of iron oxide chitosan nanocomposite for three successive days. The effectiveness of iron oxide chitosan nanocomposite was assessed based on clinical signs, mortalities, postmortem lesions, and viral RNA load in the bursa of Fabricius samples at the 3rd, 6th, and 9th days PI by isolation onto specific pathogen-free embryonated chicken eggs (SPF-ECE) and histopathology. Results revealed that the average particle size of iron oxide chitosan nanocomposite size was found to be 35.1±5 nm. The selected particles exhibited zeta potentials of 16.8±10.9 mV, polydispersity index of 0.91, and iron concentration of 2,625 mg/L. Chicks kept in G2 showed severe clinical signs, including anorexia, ruffled feathers with droopy wings, prostration, and whitish mucoid diarrhea with a mortality rate of 30%. Other groups showed no specific mortalities. The IBDV bursal load was significantly lower (P&lt;0.05) in the challenged-treated chicks (0.5-1 log10) compared to the challenged-non-treated group (3-6.4 log10). Histopathological lesions of the bursa, spleen, thymus, and kidney in the challenged-treated group showed hyperactivity of the lymphoid population compared to necrosis and depletion of lymphoid elements in the challenged control group at the 6th days PI. Based on the current study, iron oxide chitosan nanocomposite showed a promising antiviral activity that could significantly reduce IBDV load in the bursa and decrease pathological changes in lymphoid organs.

Occurrence of the Hyperparasite Ampelomyces quisqualis on Golovinomyces neosalviae (Erysiphaceae), Causal Agent of Powdery Mildew on Common Sage (Salvia officinalis)

Hamzeh, S., W. Naffaa and M.F. Azmeh. 2022. Occurrence of the Hyperparasite Ampelomyces quisqualis on Golovinomyces neosalviae (Erysiphaceae), Causal Agent of Powdery Mildew on Common Sage (Salvia officinalis). Arab Journal of Plant Protection, 40(2): 158-163. https://doi.org/10.22268/AJPP-040.2.158163 Ampelomyces quisqualis, the oldest mycoparasite of powdery mildew (PM), has been widely studied due to its potential in biocontrol. Many strains of this hyperparasite have been experimented worldwide and some of them were successfully applied for biocontrol, but others have been less efficient. No previous identification of Ampelomyces strains has been done in Syria, but some isolates were morphologically identified in the coastal region. There was no indication of Ampelomyces occurrence in any other location in Syria. During this three years survey (2019–2021), 73 plant samples were collected from five governorates, including coastal and southern regions in Syria. Ampelomyces pycnidia were detected in five samples from the coastal area and a new unpredictable finding of Ampelomyces sp. was found in southern Syria. This new occurrence of Ampelomyces isolate (S.ham82) is documented on a new mycohost; Golovinomyces neosalviae, the causal agent of PM on common sage (Salvia officinalis). Successful isolation of S.ham82 on PDA was conducted and parasitic activity was assessed by artificial inoculation using In vitro detached leaf assay. Morphological characteristics of this isolate were determined and compared with isolate Bah1 from the coastal region. Ampelomyces sp. (S.ham82) pycnidia size were 77.44 (±17.16) x 25.28 (±6.12) µm in natural host, 125.27 (±42.34) x 115.95 (±40.14) µm, 189.51 (±60.06) x 167.64 (±52.41) µm on PDA media pre and post artificial inoculation, respectively, and 88.24 (±20.05) x 27.98 (±5.68) µm on inoculated detached leaves. Conidia were also morphologically characterized and measured 8.11 (±0.87) x 3.88 (±0.51) µm in natural host, 8.86 (±1.65) x 3.18 (±0.80) µm in PDA pre and post artificial inoculation, respectively, and 7.82 (±0.69) x 3.61 (±0.37) µm in inoculated detached leaves. To our knowledge, this is the first report of the natural occurrence of Ampelomyces sp. in G. neosalviae on Saliva officinalis. Keywords: Ampelomyces sp., Common sage, Golovinomyces neosalviae, Syria.

Wheat Lysin-Motif-Containing Proteins Characterization and Gene Expression Patterns under Abiotic and Biotic Stress

Lysin motif (LysM)-containing proteins (LYPs) are important pattern recognition receptors in plants. However, the evolutionary history and characteristics of LYP genes remain largely unclear in wheat. In this study, 62 LYPs were identified at genome wide in wheat. Based on phylogenetic and domain analysis, wheat LYPs were classified into 6 subgroups (group LysMe, LysMn, LYP, LYK, LysMFbox). Syntenic analysis showed the evolution of LYP genes in wheat. RNA-seq data showed that 22 genes were not expressed at any tissue or stress stimulation period. Some LYP and LYK genes were tissue- or stage- specific. The majority of TaLYK5s, TaLYK6s, TaLYP2s and TaLysMns genes were induced under chitin, flg22 and fungal treatment. qRT-PCR analysis showed that 4 genes were upregulated during Puccinia triticina infection with a peak at 18 h post inoculation. Our findings suggested that wheat LYPs may have specific roles in response to fungal infection and provided insights into the function and characteristics of wheat LYP genes.

RNAi Mediated Silencing of ftf1 Gene Enhances Resistance in Banana to Fusarium Wilt

Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc) is a devastating disease of banana. Race 1 of Fusarium wilt is a serious threat to Musa spp. cv. Rasthali (AAB, Silk subgroup), which is a choice variety traditionally grown in most of the South East Asian countries. The investigation was an attempt to study whether RNAi-mediated expression of small interfering RNAs (siRNAs) targeted against ftf1 (fusarium transcription factor1) gene in transgenic banana could impart resistance against Foc race 1. Partial sequences comprising sense and antisense sequences of ftf1 gene were assembled as siRNA construct in a binary vector background and transformed into embryogenic cell suspensions of banana cv. Rasthali by Agrobacterium mediated genetic transformation. Presence of RNAi construct in transgenic banana plants were confirmed by polymerase chain reaction assay. Forty- six transformed lines derived from RNAi-FTF1 were found to be free of external symptoms of Foc after 3 months of challenge inoculation. Seventeen transgenic lines continued to resist the Foc race 1 even after six months of post inoculation. The present study confirmed that the host induced RNAi-mediated gene silencing of a fungal gene could confer resistance against Foc in banana.

Mechanism of carbendazim in treating pebrine disease of Bombyx mori based on GC/MS-based metabonomics.

Pebrine disease is caused by microporidia (Nosema bombycis) and is destructive to sericulture production. A carbendazim-based drug FangWeiLing (FWL) has a significant control effect on the disease, which is a successful example of drug treatment of microsporidia. In this study, the therapeutic effect and critical action time of FWL were investigated by silkworm rearing biological test. Besides, the hemolymph samples from silkworms in the control group, model group, and FWL group were analyzed by metabonomics based on gas chromatography-mass spectrometry (GC/MS). The results showed that FWL had a significant therapeutic effect on pebrine disease, and the critical action time was 24 ~ 48h post inoculation. Forty-seven different metabolites related to pebrine disease were screened out, and correlated with starch and sucrose metabolism; aminoacyl-tRNA biosynthesis; arginine biosynthesis; glycine, serine, and threonine metabolism; and phenylalanine, tyrosine, and tryptophan biosynthesis. After pretreatment with FWL, the metabolites were all effectively regulated, indicating productive intervention. Principal component analysis (PCA) also showed that the overall metabolic profile of the FWL group tended toward the control group. Compared with the control group, 16 different metabolites were obtained from the hemolymph of B.mori in FWL group, mainly involving aminoacyl-tRNA biosynthesis and taurine and hypotaurine metabolism. It indicated that FWL had some effect on silkworm metabolism, which might be related to the decrease in cocoon quality. In conclusion, combined with the life cycle of N. bombycis, the mechanism of carbendazim in the treatment of pebrine disease can be fully revealed. Carbendazim can effectively reduce the destruction of amino acid metabolism and carbohydrate metabolism by N. Bombycis infection by inhibiting the proliferation of the meronts in silkworms, thus maintaining the normal physiological state of B. mori and achieve therapeutic effects. GC/MS-based metabonomics is a valuable and promising strategy to understand the disease mechanism and drug treatment of pebrine disease.

Exogenous hexanoic acid induced primary defense responses in tomato (Solanum lycopersicum L.) plants infected with Alternaria solani

The effect of exogenous hexanoic acid on Alternaria solani-infected tomato plants was investigated in the present study. Tomato seedlings treated with different doses of hexanoic acid were inoculated with A. solani and their antioxidative, biochemical and molecular responses were studied. Results indicated that hexanoic acid application increased hydrogen peroxide content by 49% and peroxidase activity by 408% in the infected plants, while ascorbate peroxidase and catalase activities and total polyphenol content were not significantly changed. It was revealed that the expression of NAM, ATAF1and 2, and CUC2 family multiple stress-responsive transcription factor (SlNAC1), Mitogen-activated protein kinase 3 (SlMAPK3), increased 13 and 4 folds, respectively, while the expression increase for pathogenesis related proteins type1) (SlPR1) was 57% in A. solani-infected plants. It was also revealed that SlPR1 expression in plants treated with 1 mM hexanoic acid reached its highest level 72 h post inoculation while highest superoxide dismutase (SlSOD) expression rate was recorded 24 h after inoculation. It seems that exogenous application of hexanoic acid increased reactive oxygen species (ROS) content in A. solani-infected plants and subsequently induced systemic acquired resistance (SAR) pathways which in turn enhanced expression of superoxide dismutase (SOD), NAC1 and MAPK3 genes. These findings indicated that hexanoic acid induced resistance in tomato plants against A. solani by enhanced SAR signaling network leading to enhanced defense responses. While Ja/Et pathway is believed to be the prevalent biochemical pathway involved in plant resistance to necrotrophic pathogens, our findings revealed that hexanoic acid-treated tomato plants activated defense responses though SAR which is mainly a salicylic acid-dependent signaling pathway.

The Fungus Metarhizium sp. BCC 4849 Is an Effective and Safe Mycoinsecticide for the Management of Spider Mites and Other Insect Pests.

Simple SummaryThe spider mite is a destructive pest of various crops during warm and dry conditions in tropical countries, including Thailand. The pest is difficult to control despite synthetic acaricide use. In the field, insect populations gradually develop resistance to synthetic pesticides over long-term use. The use of entomopathogenic fungi is more human- and environmentally friendly. We searched and identified a potential fungal isolate from a culture collection, focusing on the genus Metarhizium. Metarhizium sp. BCC 4849 not only controls the spider mite but also plays a significant role as a natural regulator of other insect pests. Here, we investigated its infection process on the mite, optimized a conidial formulation for extended shelf life, and conducted toxicological assays in animals to assess its biosafety in humans. The fungal genome has been sequenced. The genomic data indicated that oxidoreduction proteins; zinc-, heme-, and iron-binding proteins; and transmembrane transporters are abundant in the genome.Five isolates of Metarhizium sp. were evaluated for their pathogenicity against the spider mite (Tetranychus truncatus Ehara) (Acari: Tetranychidae) and Metarhizium sp. BCC 4849 resulted in the highest mortality (82%) on the 5th day post-inoculation (DPI). Subsequent insect bioassay data indicated similar high virulence against five other insects: African red mites (Eutetranychus africanus Tucker) (Acari: Tetranychidae), bean aphid (Aphis craccivora Koch) (Hemiptera: Aphididae), cassava mealybug (Phenacoccus manihoti Matile-Ferrero) (Hemiptera: Pseudococcidae), sweet potato weevil (Cylas formicarius Fabricius) (Coleoptera: Brentidae), and oriental fruit fly (Bactrocera dorsalis Hendel) (Diptera: Tephritidae), at mortalities of 92–99%, on 3rd–6th DPI, and in laboratory conditions. The pathogenicity assay against E. africanus in hemp plants under greenhouse conditions indicated 85–100% insect mortality on 10th DPI using the fungus alone or in combination with synthetic acaricide. Genome sequencing of Metarhizium sp. BCC 4849 revealed the high abundance of proteins associated with zinc-, heme-, and iron-binding; oxidation-reduction; and transmembrane transport, implicating its versatile mode of interaction with the environment and adaptation to various ion homeostasis. The light and scanning electron microscopy indicated that at 24 h post inoculation (PI), adhesion and appressorial formation occurred, notably near the setae. Most infected mites had stopped moving and started dying by 48–72 h PI. Elongated hyphal bodies and oval blastospores were detected in the legs. At 96–120 h PI or longer, dense mycelia and conidial mass had colonized the interior and exterior of dead mites, primarily at the bottom than the upper part. The shelf-life study also indicated that conidial formulation combined with an oxygen-moisture absorber markedly enhanced the viability and germination after storage at 35 °C for four months. The fungus was tested as safe for humans and animals, according to our toxicological assays.

Gene deletion and constitutive expression of the pectate lyase gene 1 (MoPL1) lead to diminished virulence of Magnaporthe oryzae.

Phytopathogenic fungi are known to secrete specific proteins which act as virulence factors and promote host colonization. Some of them are enzymes with plant cell wall degradation capability, like pectate lyases (Pls). In this work, we examined the involvement of Pls in the infection process of Magnaporthe oryzae, the causal agent of rice blast disease. From three Plgenes annotated in the M. oryzae genome, only transcripts of MoPL1 considerably accumulated during the infection process with a peak at 72 h post inoculation. Both, gene deletion and a constitutive expression of MoPL1 in M. oryzae led to a significant reduction in virulence. By contrast, mutants that constitutively expressed an enzymatic inactive version of MoPl1 did not differ in virulence compared to the wild type isolate. This indicates that the enzymatic activity of MoPl1 is responsible for diminished virulence, which is presumably due to degradation products recognized as danger associated molecular patterns (DAMPs), which strengthen the plant immune response. Microscopic analysis of infection sites pointed to an increased plant defense response. Additionally, MoPl1 tagged with mRFP, and not the enzymatic inactive version, focally accumulated in attacked plant cells beneath appressoria and at sites where fungal hyphae transverse from one to another cell. These findings shed new light on the role of pectate lyases during tissue colonization in the necrotrophic stage of M. oryzae's life cycle.

Effects of a blend of essential oils, medium-chain fatty acids, and a toxin-adsorbing mineral on diarrhea and gut microbiome of weanling pigs experimentally infected with a pathogenic Escherichia coli.

A proprietary antimicrobial feed additive comprised of essential oils, medium-chain fatty acids, and a toxin-adsorbing mineral showed promising bacteriostatic and bactericidal effects in vitro. This study investigated the impacts of supplementing this blend on growth, gut microbiome, and enteric disease resilience in weaned pigs experimentally challenged with an enterotoxigenic Escherichia coli (ETEC). Thirty-six weanling pigs (6.88 ± 0.30kg body weight) blocked by weight and gender were assigned to one of three dietary treatments: control or dietary supplementation with 0.25% or 0.50% of the antimicrobial blend. This study lasted 28 d with 7 d before and 21 d after the first ETEC inoculation (day 0). All pigs were orally inoculated with 1010 CFU F18 + ETEC/3-mL dose for 3 consecutive days. Growth performance data and diarrhea scores were recorded throughout the experiment. Fecal samples collected on days -7, 0, 7, and 21 post first inoculation (PI), and ileal digesta and mucosal tissue collected on day 21 PI were further analyzed for gut microbiome using 16S rRNA sequencing. All data, except for frequency of diarrhea and gut microbiome, were analyzed by ANOVA using the PROC MIXED of SAS. The chi-square test was used for analyzing frequency of diarrhea. Gut microbiome data were analyzed using QIIME2 and visualized using the R program. Dietary supplementation of 0.25% or 0.5% of the antimicrobial blend increased (P < 0.05) feed efficiency on days 14 to 21 PI of ETEC and reduced (P < 0.05) frequency of diarrhea during the study. Compared with the control group, adding 0.5% dietary antimicrobial blend increased (P < 0.05) relative abundance of Firmicutes but reduced (P < 0.05) Bacteroidetes and Proteobacteria in feces on day 7 PI. Pigs that received the antimicrobial blend also had higher (P < 0.05) relative abundance of Lactobacillaceae, but lower (P < 0.05) relative abundance of Enterobacteriaceae in feces on day 7 PI than pigs in control. In conclusion, supplementation of this antimicrobial blend at 0.5% reduced incidence of severe diarrhea in weaned pigs challenged with F18 ETEC and enhanced feed efficiency of weaned pigs at the last week of the experiment. Supplementation of this antimicrobial blend also modified the microbiota diversity in feces and ileal mucosa of weaned pigs.

Ambisome shows potent anti-Candida auris in vitro and in vivo activity

Candida auris is an emerging multidrug resistant Candida species that has been reported in many parts of the world for causing severe illness in hospitalized patients especially bloodstream infections. The challenge with this type of yeast is its resistance to commonly used antifungal drugs, thus identifying antifungals that are effective against this species is critical. In this study, we determined the in vitro activity of Ambisome against 35 clinical isolates of C. auris using minimum inhibitory concentration (MIC) assay as well as the efficacy of Ambisome compared to Amphotericin B and fluconazole using a C. auris murine disseminated model. Antifungal activity of C. auris against Ambisome and comparators was assessed using amicrodilution method performed according to the Clinical and Laboratory Standard Institute (CLSI) M27-A4 methodology. Mice were immunocompromised and challenged with 3 x 107 C. auris blastopores in 0.1 ml of normal saline (via the tail vein). Treatment efficacy was assessed by determining reductions in mortality as well as decrease in tissue fungal burden (CFUs). Ambisome showed lower MIC50 and MIC90 values (1 and 2 μg/mL, respectively) than the comparators tested. Significant efficacy was observed in the Ambisome 7.5 mg/kg -treated group (100% and 90% survival by day 7- and 14-days post inoculation, respectively). Additionally, Ambisome and fluconazole treated groups showed significant reduction in CFUs in the kidneys (P- values of 0.028 and 0.022, respectively) compared to the untreated group. Our data shows that Ambisome shows significant antifungal activity against C. auris in vitro as well as in vivo.

Biosolid-Amended Soil Enhances Defense Responses in Tomato Based on Metagenomic Profile and Expression of Pathogenesis-Related Genes.

Biosolid application is an effective strategy, alternative to synthetic chemicals, for enhancing plant growth and performance and improving soil properties. In previous research, biosolid application has shown promising results with respect to tomato resistance against Fusarium oxysporum f. sp. radicis-lycopersici (Forl). Herein, we aimed at elucidating the effect of biosolid application on the plant–microbiome response mechanisms for tomato resistance against Forl at a molecular level. More specifically, plant–microbiome interactions in the presence of biosolid application and the biocontrol mechanism against Forl in tomato were investigated. We examined whether biosolids application in vitro could act as an inhibitor of growth and sporulation of Forl. The effect of biosolid application on the biocontrol of Forl was investigated based on the enhanced plant resistance, measured as expression of pathogen-response genes, and pathogen suppression in the context of soil microbiome diversity, abundance, and predicted functions. The expression of the pathogen-response genes was variably induced in tomato plants in different time points between 12 and 72 h post inoculation in the biosolid-enriched treatments, in the presence or absence of pathogens, indicating activation of defense responses in the plant. This further suggests that biosolid application resulted in a successful priming of tomato plants inducing resistance mechanisms against Forl. Our results have also demonstrated that biosolid application alters microbial diversity and the predicted soil functioning, along with the relative abundance of specific phyla and classes, as a proxy for disease suppression. Overall, the use of biosolid as a sustainable soil amendment had positive effects not only on plant health and protection, but also on growth of non-pathogenic antagonistic microorganisms against Forl in the tomato rhizosphere and thus, on plant–soil microbiome interactions, toward biocontrol of Forl.