Peanut Rhizosphere Achromobacter xylosoxidans Inhibits Aspergillus flavus Development and Aflatoxin Synthesis by Inducing Apoptosis through Targeting the Cell Membrane.

Contamination of crop seeds and feed with Aspergillus flavus and its associated aflatoxins presents a significant threat to human and animal health due to their hepatotoxic and carcinogenic properties. To address this challenge, researchers have screened for potential biological control agents in peanut soil and pods. This study identified a promising candidate, a strain of the nonpigmented bacterium, Achromobacter xylosoxidans ZJS2-1, isolated from the peanut rhizosphere in Zhejiang Province, China, exhibiting notable antifungal and antiaflatoxin activities. Further investigations demonstrated that ZJS2-1 active substances (ZAS) effectively inhibited growth at a MIC of 60 µL/mL and nearly suppressed AFB1 production by 99%. Metabolomic analysis revealed that ZAS significantly affected metabolites involved in cell wall and membrane biosynthesis, leading to compromised cellular integrity and induced apoptosis in A. flavus through the release of cytochrome c. Notably, ZAS targeted SrbA, a key transcription factor involved in ergosterol biosynthesis and cell membrane integrity, highlighting its crucial role in ZJS2-1's biocontrol mechanism. Moreover, infection of crop seeds and plant wilt caused by A. flavus can be efficiently alleviated by ZAS. Additionally, ZJS2-1 and ZAS demonstrated significant inhibitory effects on various Aspergillus species, with inhibition rates ranging from 80 to 99%. These findings highlight the potential of ZJS2-1 as a biocontrol agent against Aspergillus species, offering a promising solution to enhance food safety and protect human health.

The Autophagy-Related Protein ATG8 Orchestrates Asexual Development and AFB1 Biosynthesis in Aspergillus flavus.

Autophagy, a conserved cellular recycling process, plays a crucial role in maintaining homeostasis under stress conditions. It also regulates the development and virulence of numerous filamentous fungi. In this study, we investigated the specific function of ATG8, a reliable autophagic marker, in the opportunistic pathogen Aspergillus flavus. To investigate the role of atg8 in A. flavus, the deletion and complemented mutants of atg8 were generated according to the homologous recombination principle. Deletion of atg8 showed a significant decrease in conidiation, spore germination, and sclerotia formation compared to the WT and atg8C strains. Additionally, aflatoxin production was found severely impaired in the ∆atg8 mutant. The stress assays demonstrated that ATG8 was important for A. flavus response to oxidative stress. The fluorescence microscopy showed increased levels of reactive oxygen species in the ∆atg8 mutant cells, and the transcriptional result also indicated that genes related to the antioxidant system were significantly reduced in the ∆atg8 mutant. We further found that ATG8 participated in regulating the pathogenicity of A. flavus on crop seeds. These results revealed the biological role of ATG8 in A. flavus, which might provide a potential target for the control of A. flavus and AFB1 biosynthesis.

Nitric oxide-mediated regulation of Aspergillus flavus asexual development by targeting TCA cycle and mitochondrial function.

Nitric oxide (NO) is a signaling molecule with diverse roles in various organisms. However, its role in the opportunistic pathogen Aspergillus flavus remains unclear. This study investigates the potential of NO, mediated by metabolites from A. oryzae (AO), as an antifungal strategy against A. flavus. We demonstrated that AO metabolites effectively suppressed A. flavus asexual development, a critical stage in its lifecycle. Transcriptomic analysis revealed that AO metabolites induced NO synthesis genes, leading to increased intracellular NO levels. Reducing intracellular NO content rescued A. flavus spores from germination inhibition caused by AO metabolites. Furthermore, exogenous NO treatment and dysfunction of flavohemoglobin Fhb1, a key NO detoxification enzyme, significantly impaired A. flavus asexual development. RNA-sequencing and metabolomic analyses revealed significant metabolic disruptions within tricarboxylic acid (TCA) cycle upon AO treatment. NO treatment significantly reduced mitochondrial membrane potential (Δψm) and ATP generation. Additionally, aberrant metabolic flux within the TCA cycle was observed upon NO treatment. Further analysis revealed that NO induced S-nitrosylation of five key TCA cycle enzymes. Genetic analysis demonstrated that the S-nitrosylated Aconitase Acon and one subunit of succinate dehydrogenase Sdh2 played crucial roles in A. flavus development by regulating ATP production. This study highlights the potential of NO as a novel antifungal strategy to control A. flavus by compromising its mitochondrial function and energy metabolism.

Dysfunction of FadA-cAMP signalling decreases Aspergillus flavus resistance to antimicrobial natural preservative Perillaldehyde and AFB1 biosynthesis.

Aspergillus flavus is an opportunistic fungal pathogen that colonizes agriculture crops with aflatoxin contamination. We found that Perillaldehyde (PAE) effectively inhibited A. flavus viability and aflatoxin production by inducing excess reactive oxygen species (ROS). Transcriptome analysis indicated that the Gα protein FadA was significantly induced by PAE. Functional characterization of FadA showed it is important for asexual development and aflatoxin biosynthesis by regulation of cAMP-PKA signalling. The ΔfadA mutant was more sensitive to PAE, while ΔpdeL and ΔpdeH mutants can tolerate excess PAE compared to wild-type A. flavus. Further RNA-sequence analysis showed that fadA was important for expression of genes involved in oxidation-reduction and cellular metabolism. The flow cytometry and fluorescence microscopy demonstrated that ΔfadA accumulated more concentration of ROS in cells, and the transcriptome data indicated that genes involved in ROS scavenging were downregulated in ΔfadA mutant. We further found that FadA participated in regulating response to extracellular environmental stresses by increasing phosphorylation levels of MAPK Kinase Slt2 and Hog1. Overall, our results indicated that FadA signalling engages in mycotoxin production and A. flavus resistance to antimicrobial PAE, which provide valuable information for controlling this fungus and AF biosynthesis in pre- and postharvest of agricultural crops.

Prevalence and Drug Resistance of Salmonella in Dogs and Cats in Xuzhou, China.

INTRODUCTION: Salmonellosis is a zoonotic disease, and Salmonella spp. can sometimes be found in dogs and cats, posing a risk to human health. In this study, the prevalence and antimicrobial susceptibility of faecal Salmonella were investigated in pet dogs and cats in Xuzhou, Jiangsu Province, China. MATERIAL AND METHODS: Faecal samples from 243 dogs and 113 cats, at seven pet clinics, were tested between March 2018 and May 2019. Each Salmonella isolate was characterised using serotyping and antimicrobial susceptibility tests. RESULTS: The prevalence of Salmonella was 9.47% in dogs and 1.77% in cats. Among the 25 isolates, eight serotypes of Salmonella enterica subsp. enterica were detected, S. Kentucky (n = 11), S. Indiana (n = 5), and S. Typhimurium (n = 4) predominating. S. Derby, S. Toucra, S. Sandiego, S. Newport, and S. Saintpaul all occurred singly. The 23 Salmonella strains found in dogs were from seven different serovars, while the two strains in cats were from two. The highest resistance rates were found for tetracycline (92%), azithromycin (88%), cefazolin (84%), nalidixic acid (80%), ampicillin (80%), ceftriaxone (80%), and streptomycin (76%). Resistance to three or more antimicrobial agents was detected in 24 (96%) isolates. Most of the S. Kentucky and S. Indiana isolates were multi-drug resistant to more than 11 agents. CONCLUSION: The carriage rate was far higher in dogs than in cats from Xuzhou. Some isolated strains were highly resistant to antimicrobials used to treat infections in humans and pets, which may raise the risk of humans being infected with multi-drug resistant Salmonella via close contact with pets.

Quantitative Analysis of Salivary Oral Bacteria Associated with Severe Early Childhood Caries and Construction of Caries Assessment Model.

To construct a saliva-based caries risk assessment model, saliva samples from 176 severe early childhood caries (S-ECC) children and 178 healthy (H) children were screened by real-time PCR-based quantification of the selected species, including Streptococcus mutans, Prevotella pallens, Prevotella denticola and Lactobacillus fermentum. Host factors including caries status, dmft indices, age, gender, and geographic origin were assessed in their influence on abundance of the targeted species, which revealed host caries status as the dominant factor, followed by dmft indices (both P < 0.01). Moreover, levels of S. mutans and P. denticola in the S-ECC group were significantly higher than those in the healthy group (P < 0.001 for S. mutans and P < 0.01 for P. denticola). Interestingly, the co-occurrence network of these targeted species in the S-ECC group differed from that from the healthy group. Finally, based on the combined change pattern of S. mutans and P. pallens, we constructed an S-ECC diagnosis model with an accuracy of 72%. This saliva-based caries diagnosis model is of potential value for circumstances where sampling dental plague is difficult.

[Salivary microbial communities associated with severe early childhood caries].

OBJECTIVE: To compare the salivary microbial profiles of healthy subjects and those with severe early childhood caries (S-ECC) by using high-throughput sequencing. METHODS: Salivary samples were obtained from children with S-ECC (group C, n=24) and healthy children (group H, n=24). Total metagenomic DNA was extracted, and DNA amplicons of the V1-V3 hypervariable region of the 16S rRNA gene were generated and subjected to 454 sequencing. The characteristics of oral microbial communities from the two groups were compared based on microbial diversity and taxonomy assignment. RESULTS: First, the microbial richness was significantly higher in group C than group H (P<0.05). Second, the microbial community structure was significantly different for the groups H and C (P<0.01). In addition, caries microbiota was significantly conserved in group C (P<0.001). High expression of suspected cariogenic microorganisms in group C (P<0.1) and health related microorganisms in group H (P<0.1) were identified. Finally, models of caries risk assessment were proposed to distinguish caries from healthy subjects with over 70% accuracy. CONCLUSIONS: Salivary microbiota and certain taxa, such as caries-associated taxa (Prevotella), may be useful to screen/assess the children's risk of developing caries.