[Objective] Phosphate-solubilizing bacteria (PSB) can increase available phosphorus by promoting the transformation of different phosphorus forms in soil. However, the phosphate-solubilizing ability of PSB is influenced by soil texture. Therefore, we studied the changes of phosphorus fractions in red soil with different textures and treated with PSB, with the aims of improving the efficient utilization of soil phosphorus. [Methods] We collected the rhizosphere soil samples of four main Camellia oleifera varieties (‘Huashuo’ ‘Huaxin’ ‘Huajin’ and ‘Changlin No. 40’) in Liuyang City, Hunan Province. PSB strains were isolated, purified, screened, and identified by plate coating, transparent circle method, molybdenum-antimony anti-spectrophotometric method, and 16S rRNA gene sequencing, and a batch of highly efficient PSB strains were obtained. The phosphate-solubilizing abilities of the obtained PSB strains were investigated with different phosphorus sources (iron phosphate, aluminum phosphate, calcium phosphate, and calcium phytate). PSB were inoculated into the red soil samples added with 0, 20% and 40% perlite (0%PR, 20%PR and 40PR%) to clarify the changes in phosphorus fractions in the red soil samples with different textures. [Results] A total of 57 strains of PSB were isolated from the rhizosphere of C. oleifera, of which strains CL37, HS5, and CL36 exhibited stronger phosphate-solubilizing abilities. The three strains were identified by 16S rRNA gene sequencing and named Pantoea sp. CL37, Burkholderia sp. HS5, and Burkholderia sp. CL36, respectively. The three strains showed significant differences in their ability to solubilize different phosphorus sources, with the highest solubilizing ability for calcium phytate and calcium phosphate. Compared with CK, the inoculation of PSB increased the available phosphorus (AP) in soil by 8.90%-54.60% and 1.90%-56.00% in 20%PR and 40%PR, respectively. The inoculation with PSB increased Fe-P, Resin-P, NaHCO₃-Pi, and NaOH-Pi, which showed a tendency of first increasing and then decreasing along with the increase in the addition of perlite in red soil. Meanwhile, PSB decreased the content of HCl-Pi and Residual-P, and the decrease in HCl-Pi was more pronounced in 20%PR than in 0%PR and 40%PR. Of all treatments, the inoculation with HS5 led to the highest increases in Fe-P, Resin-P, NaHCO₃-Pi, and NaOH-Pi in the soil samples analyzed. Correlation analysis and random forest analysis suggested that AP was mainly affected by Resin-P, Al-P, NaOH-Pi, acid phosphatase, urease, pH, and NaOH-Po. [Conclusion] The phosphate-solubilizing pathways of PSB vary in red soil with different textures. Strain HS5 has a strong phosphorus-transforming ability and is more conducive to phosphorus transformation in the red soil with 20%PR.

As autotoxic substances secreted by plant roots, phenolic acids such as p-hydroxybenzoic acid (PHBA), are the main factors causing continuous cropping obstacles. [Objective] To study the effects of Microbacterium aurantiacum on the growth of tomato plants and the microbial community structure in rhizosphere soil under PHBA stress. [Methods] We irrigated tomato rhizosphere with the suspension of M. aurantiacum GX14001 and then measured the growth traits of tomato plants and the soil microbial community changes in the rhizosphere soil. [Results] Under the PHBA treatment, GX14001 significantly promoted the growth of tomato plants, increasing the leaf area, stem diameter, and plant height by 244.0%, 156.5%, and 128.0%, respectively. GX14001 increased the richness but did not cause changes in the diversity of bacteria in the rhizosphere soil. Meanwhile, it decreased the richness and diversity of fungi in the rhizosphere soil. At the phylum level, compared with the control group, the GX14001 group showed increased relative abundance of Actinobacteriota, Chloroflexi, and Proteobacteria, with Ascomycetes as the dominant fungal phylum. [Conclusion] M. aurantiacum GX14001 promotes the growth of tomato plants by changing the microbial community structure in the rhizosphere soil. It increases the relative abundance of beneficial microorganisms in the soil to create a favorable environment for tomato growth.

[Objective] Vibrio parahaemolyticus is a major foodborne pathogen that causes acute gastroenteritis in humans. Here, we aim to decipher the mechanisms by which the histidine kinase EnvZ regulates the swarming motility and biofilm formation of V. parahaemolyticus. [Methods] The plasmids pBAD24 and pMal carrying inducible promoters were used to construct the plasmids carrying target genes for overexpression. The recombinant plasmids were introduced into the wild-type strain (WT) and envZ-deleted strain (ΔenvZ) of V. parahaemolyticus. Swarming plates were prepared to measure swarming motility, while biofilm formation was detected by crystal violet staining. The RT-qPCR and bioluminescence reporter assays were employed to explore the mechanisms by which EnvZ regulated the expression of downstream genes. [Results] The swarming motility of ΔenvZ was significantly lower than that of WT, while the introduction of the pBAD24-envZ plasmid into ΔenvZ restored its swarming ability. The transcription levels of the lateral flagellar genes were positively correlated with the swarming motility. The promoter activities of P _scrABC -lux in ΔenvZ and ΔenvZΔompR were lower than those in WT and ΔompR. The swarming ability of ΔenvZ was significantly increased when the Scr system was overexpressed via the pMal-scrABC plasmid, while the overexpression of EnvZ in the strain without scrABC did not change the swarming motility. In addition, ΔenvZ exhibited a significant decrease in biofilm formation compared with WT. The pMal-envZ plasmid restored the biofilm formation of ΔenvZ to the level of WT, whereas the pMal-scrABC plasmid did not have this effect. The promoter activity of the extracellular polysaccharide operon (epsA-J) and the transcription levels of the extracellular polysaccharide genes in ΔenvZ were both significantly lower than those in WT. [Conclusion] The histidine kinase EnvZ enhances the swarming motility of V. parahaemolyticus by regulating the expression of the Scr system and promotes the biofilm formation by regulating the expression of extracellular polysaccharides.

[Objective] To investigate the inhibitory effect and mechanism of thymol on the biofilm formation of Proteusmirabilis (PM). [Methods] The minimal inhibitory concentration (MIC) and minimal biofilm inhibitory concentration (MBIC) of thymol were determined by the broth micro-dilution method. The time-kill curve of thymol was established by the colony counting method. Crystal violet staining was performed to determine the inhibitory effect of thymol on the biofilm formation of PM. The effects of thymol on the swarming and swimming of PM and the extracellular polymeric substances (extracellular polysaccharides, extracellular proteins, and eDNA) of the biofilm were determined. RT-qPCR was employed to investigate the effects of thymol on the expression levels of PM biofilm-related genes: rsbA (encoding histidine-containing phosphotransmitter of the bacterial two-component system), flhD (encoding the flagellar regulon), and mrpA (encoding mannose-resistant Proteus-like fimbriae). [Results] The MIC of thymol against PM was 0.156 mg/mL. Thymol significantly inhibited the biofilm formation of PM, with the MBIC of 0.313 mg/mL. Thymol inhibited the motility of PM, reduced the formation of extracellular polysaccharides, proteins, and eDNA in PM biofilm, and decreased the expression levels of rsbA, flhD, and mrpA. [Conclusion] Thymol inhibits the biofilm formation of PM by suppressing the motility and formation of extracellular polymers and down-regulating the expression of biofilm-related genes.

[Objective] Staphylococcusepidermidis is a coagulase-negative, Gram-positive coccobacillus that is widely found in the skin, breast milk, and blood. Bacteria in breast milk play a crucial role in the establishment of the gut microbiota in the intestinal tract and in the enhancement of immunity of infants. We then performed comparative genomic analyses to understand the genetic diversity and functional genes of breast milk-derived S. epidermidis. [Methods] We used the Illumina NovaSeq platform to sequence the genomes of 110 strains of S. epidermidis preliminarily isolated from healthy breast milk by our research team. We then performed comparative genomic analyses for the 110 strains and 263 skin, blood, and breast milk-derived S. epidermidis strains publicly available from the NCBI. [Results] The genome size of the 373 strains of S. epidermidis was (2.50±0.33) Mb, with the G+C content was (32.0±0.1)%, and the number of coding sequences (CDs) being 2 331±368. Differences existed in genome size and number of CDs among breast, blood, and skin isolates (P<0.05), with blood isolates having the largest genome size and the highest number of CDs. The phylogenetic tree showed that the S. epidermidis isolates of the same source had obvious aggregation, and the breast milk isolates and blood isolates were more closely related. There were differences in the number of virulence factors and drug resistance genes among the strains of three sources (P<0.05). The blood-derived strains exhibited the highest diversity of virulence and resistance genes, whereas the breast milk-derived strains displayed the lowest diversity of such genes. [Conclusion] S. epidermidis has undergone adaptive evolution to different habitats. Compared with skin- and blood-derived strains, breast milk-derived strains carry few genes related to biofilm synthesis, drug resistance, and virulence. This study gives new insights into the adaptive evolution of S. epidermidis and provides a theoretical basis for subsequent research on the genetic background of breast milk isolates.

Insects have a mutually dependent symbiotic relationship with their gut microbiota, which plays an important role in the insect metabolism, immunity, development, and pesticide resistance. Gut microbiota is influenced by factors such as diet, sex, and rearing environment. Currently, little is known about the gut microbiota differences between males and females of the phytophagous insect Dolycoris baccarum. [Objective] To study the effects of sex on the composition and abundance of gut microbiota in D. baccarum, explore sex-related microorganisms, and provide a basis for utilizing different sex-associated strains in the biocontrol of D. baccarum. [Methods] The PacBio platform was used for third-generation 16S rRNA gene amplicon sequencing and the bioinformatics analysis was performed to reveal the diversity and composition of gut microbiota in male and female D. baccarum. The bacterial isolation and culture method was employed to obtain gut microbiota strains from D. baccarum, which were then identified by morphological analysis and 16S rRNA gene sequencing. [Results] The gut microbiota of D. baccarum was diverse, including a total of 165 genera belonging to 109 families, 60 orders, 29 classes of 14 phyla. The gut microbiota was analyzed at six taxonomic levels (phylum, class, order, family, genus, and species). Only the phylum Deinococcota and the class Deinococci were specific to males, while the phyla Chloroflexi, Desulfobacterota, and Cyanobacteria, the class Cyanobacteriia, the genus Lacticaseibacillus, and the species Glutamicibactercreatinolyticus, Acinetobacter, and Lacticaseibacillus paracasei were specific to females. The relative abundance of Serratia marcescens was significantly higher in males than in females. In the guts of females and males, the relative abundance of Gammaproteobacteria, Enterobacterales, Yersiniaceae, Serratia, and S. marcescens was 51.223% and 95.512%, 49.784% and 95.492%, 2.385% and 10.377%, 2.383% and 10.372%, and 2.310% and 10.375%, respectively. All 28 strains isolated in vitro and identified based on morphological characteristics and 16S rRNA sequences belonged to Serratia. [Conclusion] There are significant differences in the gut microbiota composition between male and female D. baccarum. The diversity of gut microbiota in male adults of D. baccarum is significantly higher than that in female adults. All strains isolated belong to Serratia. This study provides theoretical support for investigating the potential functions of gut microbiota in male and female D. baccarum as well as for the biocontrol of D. baccarum.

Aeromonas hydrophila is a pathogen that can infect both fish and mammals, including humans. [Objective] To construct the tolA-deleted strain of A. hydrophila ATCC 7966 and use this strain to explore the biological functions of tolA. [Methods] We constructed the tolA-deleted strain AhΔtolA by homologous recombination and characterized the physiological phenotype of AhΔtolA. Transcriptome sequencing was performed to compare the gene expression between the wild type (WT) and AhΔtolA. [Results] The cell morphology of AhΔtolA was changed. The deletion of tolA significantly enhanced the sensitivity to sodium deoxycholate and oxidative stress, while significantly reducing the biofilm formation and the expression levels of several virulence genes. The yield of outer membrane vesicles was significantly increased in AhΔtolA. Transcriptomic analysis data showed that a total of 300 differentially expressed genes (DEGs) were screened between WT and AhΔtolA, including171 genes with up-regulated expression and 129 genes with down-regulated expression. GO enrichment analysis showed that the DEGs were mainly enriched in the oxidation-reduction process, metabolic process, outer membrane, and oxidoreductase activity. KEGG pathway enrichment analysis showed that the DEGs were mainly enriched in the biosynthesis of secondary metabolites, microbial metabolism in diverse environments, biosynthesis of cofactors, and biosynthesis of amino acids. [Conclusion] This study gives an insight into the roles of tolA in A. hydrophila and provides information about the metabolic pathways involving tolA. These results provide a theoretical reference for the prevention and control of A. hydrophila.

N-Heterocyclic carbine-silver (Ag-NHC) complexes possessing excellent stability, water solubility, and bactericidal activity are antimicrobial candidates with great potential. [Objective] To study the inhibitory activity and mechanism of a novel Ag-NHC complex 1,3-dibenzyl-4,5-diphenylimidazol-2-ylidene silver (I) acetate (SBC3), synthesized by Matthias Tacke’s team against Escherichia coli. [Methods] Visible spectrophotometry was employed to examine the antimicrobial activity of SBC3. Transmission microscopy was employed to observe the morphological changes of DHB4 cells post SBC3 treatment. Flow cytometry was performed to detect the effect of SBC3 on the intracellular reactive oxygen species (ROS) content. The 5,5′- dithiobis-(2-nitrobenzoic acid) (DTNB) assay was used to determine the intracellular thioredoxin (Trx) and thioredoxin reductase (TrxR) activities and the content of glutathione (GSH) post SBC3 treatment. Dithiothreitol (DTT), the ROS scavenger, was added to rescue DHB4 from ROS. SBC3-resistant strains (SRSs) were obtained by successive passaging in the laboratory. The obtained strains showed the minimal inhibitory concentrations (MICs) against SBC3 being 24, 32, and 56 μg/mL, respectively, which were 3, 4, and 7 folds of the MIC of WT. The three strains were named SRS3, SRS4, and SRS7 and then used to retest the above indicators. Western blotting was performed to determine the expression levels of Trx1 and S-glutathionylated proteins (S-PSSG) post SBC3 treatment. [Results] The MIC values of SBC3 against tested pathogens were 8.0–30.0 μg/mL. The DHB4 cells treated with SBC3 underwent swelling, which was accompanied by contents leakage. SBC3 significantly inhibited the Trx and TrxR activities, reduced the GSH content, and elevated the ROS level in DHB4. SBC3 treatment decreased the Trx and TrxR activities, reduced the GSH content, and down-regulated the expression of S-PSSG in SRS3, SRS4, and SRS7. However, all the above indicators were increased to different extent compared with those in DHB4. [Conclusion] SBC3 can target the thiol-dependent redox system (TDRS) of E. coli to exert antibacterial effects. This study provides a new idea for the design of SBC3 as a novel antimicrobial agent.

Salmonella Enteritidis is a major foodborne pathogen that can cause gastrointestinal infections in both humans and animals. As one of the key genes encoding the iron-sulfur cluster assembly, iscA plays a role in the transport of iron ions and energy metabolism. IscA is a conserved A-type iron-binding protein. [Objective] To study the role of iscA in the infection process of Salmonella by constructing an iscA-deleted mutant (ΔiscA) of Salmonella Enteritidis Z11. [Methods] The unmarked in-frame gene deletion method was employed to construct ΔiscA from the laboratory-preserved Salmonella Enteritidis Z11 strain. The wild type (WT) and ΔiscA were compared in terms of motility and biofilm formation. Additionally, the impact of IscA on the virulence of Salmonella Enteritidis was explored in both RAW264.7 cells and a mouse model. [Results] The deletion mutant ΔiscA was successfully constructed. No significant difference in the growth or biofilm formation was observed between ΔiscA and WT, indicating that the deletion of iscA did not affect the normal growth or biofilm formation of Salmonella Enteritidis. However, ΔiscA exhibited a significantly smaller zone of motility than WT at the time point of 6 h, suggesting that the loss of iscA reduced the motility of Salmonella Enteritidis Z11. In RAW264.7 cells, the adhesion and invasion of ΔiscA significantly decreased to 37% and 20%, respectively, of those of WT. Furthermore, the proliferation rate of ΔiscA in the cells was significantly lower than that of WT. Mouse infection experiments revealed that ΔiscA demonstrated reduced colonization in the jejunum and cecum compared with WT. [Conclusion] iscA is closely associated with the virulence of Salmonella Enteritidis. Its deletion affects the motility, adhesion, invasion, and proliferation, ultimately reducing the colonization in the host intestine and influencing the infection process of Salmonella Enteritidis.

[Objective] Waste printed circuit boards (PCBs) contain about 40% non-ferrous metals, which is more than 40 times that of natural ores, and thus they are known as a veritable “urban mine”. To achieve efficient and green recycling of PCBs, this study developed a leaching system that can efficiently leach non-ferrous metals from PCBs. [Methods] Acidithiobacillus ferrooxidans MA-Y1 was used to construct a bioleaching system for PCBs. Additionally, an electric current was introduced to strengthen the Cu and Ni leaching from PCBs. [Results] The optimal leaching parameters were addition of PCBs at 80.0 g/L, particle size of 4.0 cm, liquid velocity of 2.0 L/min, and electric current of 70.0 mA. Under these conditions, the leaching ratios of Cu and Ni from PCBs were 84.0% and 75.3%, respectively, which represented increases of 15.8% and 17.1% compared with the case without electric current. [Conclusion] An electric current improves the ability of A. ferrooxidans MA-Y1 to recover Cu and Ni from PCBs.