Sequence type 88 (ST88) methicillin-resistant Staphylococcus aureus (MRSA) has been recognized as an important pathogen that causes infections in humans, especially when it has strong biofilm production and multidrug resistance (MDR). However, knowledge of the determinants of resistance or virulence and genomic characteristics of ST88 MRSA from China is still limited. In this study, we employed the antimicrobial resistance (AMR), biofilm formation, and genomic characteristics of ST88 MRSA collected from various foods in China and estimated the worldwide divergence of ST88 MRSA with publicly available ST88 genomes. All ST88 isolates studied were identified as having resistance genes, while 50% (41/82) harbored MDR genes. All isolates carried core virulence genes related to immune modulation, adherence, secreted enzymes, and hemolysin. In addition, all 20 Chinese ST88 isolates showed biofilm production capacity, three strongly so. Bayesian phylogenetic analysis showed that Chinese ST88 clones formed an independent MRSA lineage, with two subclades associated with acquisition of type IVc staphylococcal cassette chromosome mec (SCCmec) elements. In contrast, all African ST88 strains were subtyped as SCCmecIVa, where the African clades were mixed with a few European and American isolates, suggesting potential transmission from Africa to these regions. In summary, our results revealed the evolution of ST88 MRSA in humans, animals, and foods in Africa and Asia. The food-associated ST88 genomes in this study will remedy the lack of food-associated ST88 isolates, and the study in general will extend the discussion of the potential exchanges of ST88 between humans and foods or food animals. IMPORTANCE ST88 MRSA has frequently been detected in humans, animals, and foods mainly in Africa and Asia. It can colonize and cause mild to severe infections in humans, especially children. Several studies from African countries have described its genotypic characteristics but, limited information is available on the evolution and characterization of ST88 MRSA in Asia, especially China. Meanwhile, the molecular history of its global spread remains largely unclear. In this study, we analyzed the genomic evolution of global ST88 MRSA strains in detail and identified key genetic changes associated with specific hosts or regions. Our results suggested geographical differentiation between ST88 MRSA's evolution in Africa and its evolution in Asia, with a more recent clonal evolution in China. The introduction of ST88 MRSA in China was aligned with the acquisition of SCCmecIVc elements, specific virulent prophages, and AMR genes.

As a severe emerging shrimp disease, TPD has heavily impacted the shrimp aquaculture industry and resulted in serious economic losses in China since spring 2020. This study aimed to identify the key virulent factors and related genes of the Vp TPD, for a better understanding of its pathogenicity of the novel highly lethal infectious pathogen, as well as its molecular epidemiological characteristics in China. The present study revealed that a novel protein, Vibrio high virulent protein-2 (MW >100 kDa), is responsible to the lethal virulence of V. parahaemolyticus to shrimp post-larvae. The results are essential for effectively diagnosing and monitoring novel pathogenic bacteria, like Vp TPD, in aquaculture shrimps and would be beneficial to the fisheries department in early warning of Vp TPD emergence and developing prevention strategies to reduce economic losses due to severe outbreaks of TPD. Elucidation of the key virulence genes and genomics of Vp TPD could also provide valuable information on the evolution and ecology of this emerging pathogen in aquaculture environments.

The composition and structure of fungal communities on healthy and diseased fruits of Cinnamomum burmannii (Nees and Nees) Blume were characterized, with evaluation of the antibacterial activity of secondary metabolites from culturable fungi following the first identification of secondary metabolites in the fungus Medicopsis romeroi (Esf-14; GenBank accession number OK242756). These results are significant for understanding the functional variation in bioactivity in fungal communities and developing a broader range of bioactive resources. High-throughput sequencing results indicated that the fungal community in diseased fruit differed from that in healthy fruit at the phylum, class, order, or genus level, with significant differences in the species and relative abundance of the dominant flora. A total of 49 (healthy fruit) and 122 (diseased fruit) artificially cultivable endophytic fungi were isolated, and 41 different strains (11 from healthy fruit and 30 from diseased fruit) were successfully identified by morphological and molecular biological analyses, which were classified into 8 groups and 23 genera by phylogenetic tree analysis, with Pleosporales, Glomerellales, and Hypocreales being the dominant groups at the order level and Colletotrichum being the dominant group at the genus level. The results of the antibacterial assay demonstrated that the secondary metabolites of all strains had different degrees of antibacterial activity, while the secondary metabolites of endophytic fungi from diseased fruit were generally stronger than those of fungi from healthy fruit, with the active secondary metabolites dominated by small and moderately polar compounds. Combined analysis of fungal communities, phylogenetic tree analysis, and bioactivity analysis of culturable strains revealed strong antibacterial activity of both upregulated and downregulated flora in diseased fruit. Five compounds, including two new (5,6-dimethoxy-[1',1:4,1″-terphenyl]-2-ol [compound 1] and 5-(methoxycarbonyl)-2-methylbenzo[d][1,3]dioxole-2-carboxylic acid [compound 2]) and three known compounds (3,7-dihydroxy-1,9-dimethyldibenzofuran [compound 3], methyl 3-hydroxybenzoate [compound 4], and uracil [compound 5]), were isolated and identified for the first time from the endophytic fungus Medicopsis romeroi. In general, the diversity of fungal communities on diseased fruit was lower than that on healthy fruits, while the antibacterial activity of artificially cultured endophytic fungi on diseased fruits was generally stronger than that on healthy fruits, suggesting excellent promise for the development of secondary metabolites from active strains on diseased fruit as antibacterial agents. IMPORTANCE Powdery fruit disease is a notorious disease of Cinnamomum burmannii that causes severe loss in fruit production. Studies on the function of endophytic fungal communities in healthy plant tissues are not new, while little is known about the functional changes of fungal communities in disease-causing plant tissues. Our results demonstrate that fungal communities in diseased fruits differ from those in healthy fruits at the level of phylum, class, order, or genus, with significant differences in the species and relative abundance of dominant groups. Endophytic fungi in diseased fruits appeared to produce secondary metabolites with stronger antibacterial properties, although the community diversity was not as varied as that in healthy fruits. In addition, secondary metabolites of the Medicopsis romeroi strain from diseased fruits were identified for the first time. These results have important implications for understanding the functional variation of bioactivity in fungal communities and for developing a broader resource of bioactivity.

Since single nucleotide polymorphisms (SNPs) have attracted attention, there have been many explorations and improvements in screening and detection methods for SNPs. Traditional methods are complex and time-consuming and rely on expensive instruments. Therefore, there is an urgent need for a low-cost, simple, and accurate method that is convenient for use in resource-poor areas. Thus, a platform based on allele-specific PCR (AS-PCR) and a gold nanoparticle-based lateral flow assay (LFA) was developed, optimized, and used to detect the SNPs of the drug resistance gene pfmdr1. Subsequently, the system was assessed on clinical isolates and compared with nested PCR followed by Sanger sequencing. The sensitivity and specificity of the AS-PCR-LFA platform were up to 99.43% and 100%, respectively, based on the clinical isolates. The limit of detection is approximately 150 fg/μL for plasmid DNA as the template and 50 parasites/μL for dried filter blood spots from clinical isolates. The established and optimized AS-PCR-LFA system is more adaptable and rapidly translated to SNP analysis of other drug resistance genes and genetic diseases. In addition, while actively responding to the point-of-care testing policy, it also contributes to the Global Malaria Eradication Program. IMPORTANCE Rapid detection of single nucleotide polymorphisms (SNPs) is essential for malaria treatment. Based on the techniques of allele-specific PCR (AS-PCR) and lateral flow assay (LFA), an accurate and powerful platform for SNP detection of pfmdr1 was developed and evaluated with plasmid and clinical isolates. It offers a useful tool to identify antimalarial drug resistance and can support the effort to eliminate malaria globally.

House dust mites (HDMs) are a major source of indoor allergens that cause airway allergic disease. Dermatophagoides farinae, a predominant species of HDMs in China, has demonstrated pathogenic role in allergic disorders. Exosomes derived from human bronchoalveolar lavage fluid have been strongly associated with allergic respiratory diseases progression. However, the pathogenic role of D. farinae-derived exosomes in allergic airway inflammation has remained unclear until now. Here, D. farinae was stirred overnight in phosphate-buffered saline, and the supernatant was used to extract exosomes by ultracentrifugation. Then, shotgun liquid chromatography-tandem mass spectrometry and small RNA sequencing were performed to identify proteins and microRNAs contained in D. farinae exosomes. Immunoblotting, Western blotting, and enzyme-linked immunosorbent assay demonstrated the specific immunoreactivity of D. farinae-specific serum IgE antibody against D. farinae exosomes, and D. farinae exosomes were found to induce allergic airway inflammation in a mouse model. In addition, D. farinae exosomes invaded 16-HBE bronchial epithelial cells and NR8383 alveolar macrophages to release the inflammation-related cytokines interleukin-33 (IL-33), thymic stromal lymphopoietin, tumor necrosis factor alpha, and IL-6, and comparative transcriptomic analysis of 16-HBE and NR8383 cells revealed that immune pathways and immune cytokines/chemokines were involved in the sensitization of D. farinae exosomes. Taken together, our data demonstrate that D. farinae exosomes are immunogenic and may induce allergic airway inflammation via bronchial epithelial cells and alveolar macrophages. IMPORTANCE Dermatophagoides farinae, a predominant species of house dust mites in China, has displayed pathogenic role in allergic disorders, and exosomes derived from human bronchoalveolar lavage fluid have been strongly associated with allergic respiratory diseases progression. However, the pathogenic role of D. farinae-derived exosomes in allergic airway inflammation has remained unclear until now. This study, for the first time, extracted exosomes from D. farinae, and sequenced their protein cargo and microRNAs using shotgun liquid chromatography-tandem mass spectrometry and small RNA sequencing. D. farinae-derived exosomes trigger allergen-specific immune responses and present satisfactory immunogenicity, as revealed by immunoblotting, Western blotting, and enzyme-linked immunosorbent assay and may induce allergic airway inflammation via bronchial epithelial cells and alveolar macrophages. Our data provide insights into the mechanisms of allergic airway inflammation caused with D. farinae-derived exosomes and the treatment of house dust mite-induced allergic airway inflammation.

Single-nucleotide polymorphisms and genotyping related to genetic detection are several of the focuses of contemporary biotechnology development. Traditional methods are complex, take a long time, and rely on expensive instruments. Therefore, there is an urgent need for a rapid, simple, and accurate method convenient for use in resource-poor areas. Thus, a platform based on allele-specific PCR (AS-PCR) combined with a lateral flow assay (LFA) was developed, optimized, and used to detect the genotype of the Plasmodium falciparum chloroquine transporter gene (pfcrt). Subsequently, the system was assessed by clinical isolates and compared with Sanger sequencing. The sensitivity and specificity of the AS-PCR-LFA platform were 95.83% (115/120) and 100% (120/120), respectively, based on the clinical isolates. The detection limit of plasmid DNA was approximately 3.38 × 105 copies/μL. In addition, 100 parasites/μL were used for the dried filter blood spots from clinical isolates. The established rapid genotyping technique is not limited to antimalarial drug resistance genes but can also be applied to genetic diseases and other infectious diseases. Thus, it has realized the leap and transformation from scientific research theory to practical application and actively responds to the point-of-care testing policy. IMPORTANCE Accurate recognition of the mutation and genotype of genes are essential for the treatment of infectious diseases and genetic diseases. Based on the techniques of allele-specific PCR (AS-PCR) and a lateral flow assay (LFA), a rapid and useful platform for mutation detection was developed and assessed with clinical samples. It offers a powerful tool to identify antimalarial drug resistance and can support malaria control and elimination globally.

Cronobacter is a foodborne pathogen associated with severe infections in restricted populations and particularly with high mortality in neonates and infants. The prevalence and antimicrobial resistance (AMR) phenotype of Cronobacter cultured from powdered infant formula and supplementary food were studied. The virulence factors, AMR genes, and genomic environments of the multidrug-resistant isolates were further studied. A total of 1,055 Cronobacter isolates were recovered from 12,105 samples of powdered infant formula and supplementary food collected from 29 provinces between 2018 and 2019 in China. Among these, 1,048 isolates were from infant supplementary food and 7 were from powdered infant formula. Regarding antimicrobial resistance susceptibility, 11 (1.0%) isolates were resistant and two showed resistance to four antimicrobials (ampicillin [AMP], tetracycline [TET], sulfamethoxazole-trimethoprim [SXT], and chloramphenicol [CHL]), defined as MDR. These two MDR isolates were subsequently identified as Cronobacter sakazakii sequence type 4 (ST4) (C. sakazakii Crono-589) and ST40 (C. sakazakii Crono-684). Both MDR isolates contain 11 types of virulence genes and 7 AMR genes on their genomes. Meanwhile, the IncFIB plasmids of both MDR C. sakazakii isolates also harbored 2 types of virulence genes. Results of the genomic comparative analysis indicated that food-associated C. sakazakii could acquire antimicrobial resistance determinants through horizontal gene transfer (HGT). IMPORTANCE As a foodborne pathogen, Cronobacter can cause serious infections in restricted populations and lead to death or chronic sequelae. Although a number of investigations showed that Cronobacter isolates are susceptible to most antimicrobial agents, MDR Cronobacter isolates, isolated mainly from clinical cases but occasionally from foods, have been reported in recent years. In this study, we successfully identified two MDR Cronobacter sakazakii isolates from infant foods based on nationwide surveillance and genome sequencing in China. Genomic analysis revealed that these two MDR C. sakazakii strains acquired resistance genes from other species via different evolution and transmission routes. It is important to monitor MDR C. sakazakii isolates in infant foods, and appropriate control measures should be taken to reduce the contamination with and transmission of this MDR bacterium.