Mycoplasma pneumoniae is an important pathogen causing upper and lower respiratory tract infections in children and other age groups. Macrolides are the recommended treatments of choice for M. pneumoniae infections. However, macrolide resistance in M. pneumoniae is increasing worldwide, which complicates the treatment strategies. The mechanisms of macrolide resistance have been extensively studied focusing on the mutations in 23S rRNA and ribosomal proteins. Since the secondary treatment choice for pediatric patients is very limited, we decided to look for potential new treatment strategies in macrolide drugs and investigate possible new mechanisms of resistance. We performed an in vitro selection of mutants resistant to five macrolides (erythromycin, roxithromycin, azithromycin, josamycin, and midecamycin) by inducing the parent M. pneumoniae strain M129 with increasing concentrations of the drugs. The evolving cultures in every passage were tested for their antimicrobial susceptibilities to eight drugs and mutations known to be associated with macrolide resistance by PCR and sequencing. The final selected mutants were also analyzed by whole-genome sequencing. Results showed that roxithromycin is the drug that most easily induces resistance (at 0.25 mg/L, with two passages, 23 days), while with midecamycin it is most difficult (at 5.12 mg/L, with seven passages, 87 days). Point mutations C2617A/T, A2063G, or A2064C in domain V of 23S rRNA were detected in mutants resistant to the 14- and 15-membered macrolides, while A2067G/C was selected for the 16-membered macrolides. Single amino acid changes (G72R, G72V) in ribosomal protein L4 emerged during the induction by midecamycin. Genome sequencing identified sequence variations in dnaK, rpoC, glpK, MPN449, and in one of the hsdS (MPN365) genes in the mutants. Mutants induced by the 14- or 15-membered macrolides were resistant to all macrolides, while those induced by the 16-membered macrolides (midecamycin and josamycin) remained susceptible to the 14- and 15-membered macrolides. In summary, these data demonstrated that midecamycin is less potent in inducing resistance than other macrolides, and the induced resistance is restrained to the 16-membered macrolides, suggesting a potential benefit of using midecamycin as a first treatment choice if the strain is susceptible.

IntroductionBipolaris sorokiniana is the popular pathogenic fungi fungus which lead to common root rot and leaf spot on wheat. Generally, chemical fungicides are used to control diseases. However, the environmental pollution resulting from fungicides should not be ignored. It is important to study the mode of antagonistic action between biocontrol microbes and plant pathogens to design efficient biocontrol strategies.ResultsAn antagonistic bacterium DB2 was isolated and identified as Bacillus amyloliquefaciens. The inhibition rate of cell-free culture filtrate (CF, 20%, v/v) of DB2 against B. sorokiniana reached 92.67%. Light microscopy and scanning electron microscopy (SEM) showed that the CF significantly altered the mycelial morphology of B. sorokiniana and disrupted cellular integrity. Fluorescence microscopy showed that culture filtrate destroyed mycelial cell membrane integrity, decreased the mitochondrial transmembrane potential, induced reactive oxygen species (ROS) accumulation, and nuclear damage which caused cell death in B. sorokiniana. Moreover, the strain exhibited considerable production of protease and amylase, and showed a significant siderophore and indole-3-acetic acid (IAA) production. In the detached leaves and potted plants control assay, B. amyloliquefacien DB2 had remarkable inhibition activity against B. sorokiniana and the pot control efficacy was 75.22%. Furthermore, DB2 suspension had a significant promotion for wheat seedlings growth.ConclusionB. amyloliquefaciens DB2 can be taken as a potential biocontrol agent to inhibit B. sorokiniana on wheat and promote wheat growth.

The use of synthetic fungicides against postharvest Alternaria rot adversely affects human health and the environment. In this study, as a safe alternative to fungicides, Bacillus subtilis strain Y17B isolated from soil exhibited significant antifungal activity against Alternaria alternata. Y17B was identified as B. subtilis based on phenotypic identification and 16S rRNA sequence analysis. To reveal the antimicrobial activity of this strain, a PCR-based study detected the presence of antifungal lipopeptide (LP) biosynthetic genes from genomic DNA. UPLC Q TOF mass spectrometry analysis detected the LPs surfactin (m/z 994.64, 1022.68, and 1026.62), iturin (m/z 1043.56), and fengycin (m/z 1491.85) in the extracted LP crude of B. subtilis Y17B. In vitro antagonistic study demonstrated the efficiency of LPs in inhibiting A. alternata growth. Microscopy (SEM and TEM) studies showed the alteration of the morphology of A. alternata in the interaction with LPs. In vivo test results revealed the efficiency of LPs in reducing the growth of the A. alternata pathogen. The overall results highlight the biocontrol potential of LPs produced by B. subtilis Y17B as an effective biological control agent against A. alternata fruit rot of cherry.

BackgroundIncreasing evidence supports that gut microbiota plays an important role in the development of cardiovascular diseases. The prevalence of sarcopenia is increasing in patients with heart failure. Muscle wasting is an independent predictor of death in heart failure patients.AimsIn this study, we aimed to explore the characteristics of gut microbiota and metabolites in heart failure patients with or without sarcopenia.MethodsFecal samples of 33 heart failure patients without sarcopenia, 29 heart failure patients with sarcopenia, and 15 controls were collected. The intestinal microbiota was analyzed using 16S rRNA sequencing and the metabolites were detected using the gas chromatography-mass spectrometry method.ResultsThere were significant differences in the overall microbial community structure and diversity between control and heart failure patients with or without sarcopenia. However, no clear clustering of samples was observed in heart failure with and without sarcopenia patients. Several bacterial, particularly Nocardiaceae, Pseudonocardiaceae, Alphaproteobacteria, and Slackia were significantly enriched in the heart failure patients without sarcopenia, while Synergistetes was more abundant in the heart failure patients with sarcopenia. Isobutyric acid, isovaleric acid, and valeric acid were lower in heart failure patients with sarcopenia than that without sarcopenia but lacked significance.ConclusionsThis study demonstrates that there are differences in the gut microbiota between control individuals and heart failure patients with or without sarcopenia. Modulating the gut microbiota may be a new target for the prevention and treatment of sarcopenia in heart failure patients.

BackgroundThe etiology of allergic rhinitis (AR) is complicated. Traditional therapy of AR still has challenges, such as low long-term treatment compliance, unsatisfactory therapeutic outcomes, and a high financial burden. It is urgent to investigate the pathophysiology of allergic rhinitis from different perspectives and explore brand-new possible preventative or treatment initiatives.ObjectiveThe aim is to apply a multi-group technique and correlation analysis to explore more about the pathogenesis of AR from the perspectives of gut microbiota, fecal metabolites, and serum metabolism.MethodsThirty BALB/c mice were randomly divided into the AR and Con(control) groups. A standardized Ovalbumin (OVA)-induced AR mouse model was established by intraperitoneal OVA injection followed by nasal excitation. We detected the serum IL-4, IL-5, and IgE by enzyme-linked immunosorbent assay (ELISA), evaluated the histological characteristics of the nasal tissues by the hematoxylin and eosin (H&E) staining, and observed the nasal symptoms (rubs and sneezes) to evaluate the reliability of the AR mouse model. The colonic NF-κB protein was detected by Western Blot, and the colonic histological characteristics were observed by the H&E staining to evaluate inflammation of colon tissue. We analyzed the V3 and V4 regions of the 16S ribosomal DNA (rDNA) gene from the feces (colon contents) through 16S rDNA sequencing technology. Untargeted metabolomics was used to examine fecal and serum samples to find differential metabolites. Finally, through comparison and correlation analysis of differential gut microbiota, fecal metabolites, and serum metabolites, we further explore the overall impact of AR on gut microbiota, fecal metabolites, and host serum metabolism and its correlation.ResultsIn the AR group, the IL-4, IL-5, IgE, eosinophil infiltration, and the times of rubs and sneezes were significantly higher than those in the Con group, indicating the successful establishment of the AR model. No differences in diversity were detected between the AR and Con groups. However, there were modifications in the microbiota’s structure. At the phylum level, the proportion of Firmicutes and Proteobacteria in the AR group increased significantly, while the proportion of Bacteroides decreased significantly, and the ratio of Firmicutes/Bacteroides was higher. The key differential genera, such as Ruminococcus, were increased significantly in the AR group, while the other key differential genera, such as Lactobacillus, Bacteroides, and Prevotella, were significantly decreased in the Con group. Untargeted metabolomics analysis identified 28 upregulated and 4 downregulated differential metabolites in feces and 11 upregulated and 16 downregulated differential metabolites in serum under AR conditions. Interestingly, one of the significant difference metabolites, α-Linoleic acid (ALA), decreased consistently in feces and serum of AR. KEGG functional enrichment analysis and correlation analysis showed a close relationship between differential serum metabolites and fecal metabolites, and changes in fecal and serum metabolic patterns are associated with altered gut microbiota in AR. The NF-κB protein and inflammatory infiltration of the colon increased considerably in the AR group.ConclusionOur study reveals that AR alters fecal and serum metabolomic signatures and gut microbiota characteristics, and there is a striking correlation between the three. The correlation analysis of the microbiome and metabolome provides a deeper understanding of AR’s pathogenesis, which may provide a theoretical basis for AR’s potential prevention and treatment strategies.