Streptococcus pneumoniae (S. pneumoniae) represents a major human bacterial pathogen leading to high morbidity and mortality in children and the elderly. Recent research emphasizes the role of extracellular vesicles (EVs) in bacterial pathogenicity. However, the contribution of S. pneumoniae EVs (pEVs) to host-microbe interactions has remained unclear. Here, we observed that S. pneumoniae infections in mice led to severe lung injuries and alveolar epithelial barrier (AEB) dysfunction. Infections of S. pneumoniae reduced the protein expression of tight junction protein OCLN (occludin) and activated macroautophagy/autophagy in lung tissues of mice and A549 cells. Mechanically, S. pneumoniae induced autophagosomal degradation of OCLN leading to AEB impairment in the A549 monolayer. S. pneumoniae released the pEVs that could be internalized by alveolar epithelial cells. Through proteomics, we profiled the cargo proteins inside pEVs and found that these pEVs contained many virulence factors, among which we identified a eukaryotic-like serine-threonine kinase protein StkP. The internalized StkP could induce the phosphorylation of BECN1 (beclin 1) at Ser93 and Ser96 sites, initiating autophagy and resulting in autophagy-dependent OCLN degradation and AEB dysfunction. Finally, the deletion of stkP in S. pneumoniae completely protected infected mice from death, significantly alleviated OCLN degradation in vivo, and largely abolished the AEB disruption caused by pEVs in vitro. Overall, our results suggested that pEVs played a crucial role in the spread of S. pneumoniae virulence factors. The cargo protein StkP in pEVs could communicate with host target proteins and even hijack the BECN1 autophagy initiation pathway, contributing to AEB disruption and bacterial pathogenicity.

It is common knowledge that prolonged and excessive use of antibiotics can lead to antimicrobial resistance. However, the characteristics and mechanism of resistant-bacteria induced by clinically recommended and prophylactic dose drugs remain largely unclear. This study aimed to observe the trends of drug resistance of the bacitracin-susceptible Staphylococcus aureus strain FS127 under exposure to bacitracin (BAC), which were induced in vitro and in chicken gut. Antimicrobial susceptibility testing was used to detect the susceptibility of S. aureus induced in vitro and in the chicken gut to gentamicin, chloramphenicol, tetracycline, doxycycline, penicillin and chloramphenicol. The research results showed that bacitracin could induce drug resistance in S. aureus both in vitro and in vivo. The bacitracin-resistance rate of S. aureus isolated from chicken gut was positively correlated with the dose and time of bacitracin administration. The findings revealed that bacitracin-resistant S. aureus induced in vivo had enhanced susceptibility to chloramphenicol but no such change in vitro. Meanwhile, RT-qPCR assay was used to detect the expression levels of vraD, braD, braR and bacA in typical strains with different bacitracin-resistance levels. It was found that BacA may play a key role in the bacitracin resistance of S. aureus. In conclusion, this work reveals the characteristics and mechanism of bacitracin-resistant S. aureus induced by bacitracin in vivo and in vitro respectively.

细菌可以通过多种方式产生对抗菌药物的耐药性,其中最为常见的方式之一为获得携带有耐药基因的质粒.耐药质粒可以在菌株之间转移,导致耐药性的广泛传播,因此消除耐药质粒是缓解细菌耐药扩散的关键.本综述介绍了基于物理、化学以及生物学的细菌耐药质粒消除方法的研究进展,并阐述了质粒消除的机理,为今后探寻安全有效地消除耐药质粒的研究方法提供参考.

CRISPR-Cas systems are an immune defense mechanism that is widespread in archaea and bacteria against invasive phages or foreign genetic elements. In the last decade, CRISPR-Cas systems have been a leading gene-editing tool for agriculture (plant engineering), biotechnology, and human health (e.g., diagnosis and treatment of cancers and genetic diseases), benefitted from unprecedented discoveries of basic bacterial research. However, the functional complexity of CRISPR systems is far beyond the original scope of immune defense. CRISPR-Cas systems are implicated in influencing the expression of physiology and virulence genes and subsequently altering the formation of bacterial biofilm, drug resistance, invasive potency as well as bacterial own physiological characteristics. Moreover, increasing evidence supports that bacterial CRISPR-Cas systems might intriguingly influence mammalian immune responses through targeting endogenous genes, especially those relating to virulence; however, unfortunately, their underlying mechanisms are largely unclear. Nevertheless, the interaction between bacterial CRISPR-Cas systems and eukaryotic cells is complex with numerous mysteries that necessitate further investigation efforts. Here, we summarize the non-canonical functions of CRISPR-Cas that potentially impact bacterial physiology, pathogenicity, antimicrobial resistance, and thereby altering the courses of mammalian immune responses.

产气荚膜梭菌是一种食源性致病菌,可存在于外界环境和健康家禽等动物的肠道中,在家禽肠道中增殖后可引起坏死性肠炎等疾病,对全世界的肉鸡和火鸡产业造成严重损害.此外,产气荚膜梭菌还可引起人类食物中毒、伤口的气性坏疽等疾病,危害人类健康.养殖业通常采用在饲料中添加抗生素的方法预防鸡坏死性肠炎,但自2017年开始的"减抗禁抗"政策的实施必然对临床细菌耐药性产生很大的影响,因此有必要了解当前鸡源产气荚膜梭菌的耐药情况.本文结合目前鸡产气荚膜梭菌的耐药现状、耐药基因检测和预防措施展开讨论,为养殖业合理用药控制其耐药性的产生和传播提供参考.

Multidrug resistance (MDR) in Enterobacteriaceae including resistance to quinolones is rising worldwide. The plasmid-mediated quinolone resistance (PMQR) gene qnrS is prevalent in Enterobacteriaceae. However, the qnrS gene is rarely found in Enterobacter hormaechei (E. hormaechei). Here, we reported one multidrug resistant E. hormaechei strain M1 carrying the qnrS1 and blaTEM-1 genes. This study was to analyze the characteristics of MDR E. hormaechei strain M1. The E. hormaechei strain M1 was identified as Enterobacter cloacae complex by biochemical assay and 16S rRNA sequencing. The whole genome was sequenced by the Oxford Nanopore method. Taxonomy of the E. hormaechei was based on multilocus sequence typing (MLST). The qnrS with the other antibiotic resistance genes were coexisted on IncF plasmid (pM1). Besides, the virulence factors associated with pathogenicity were also located on pM1. The qnrS1 gene was located between insertion element IS2A (upstream) and transposition element ISKra4 (downstream). The comparison result of IncF plasmids revealed that they had a common plasmid backbone. Susceptibility experiment revealed that the E. hormaechei M1 showed extensive resistance to the clinical antimicrobials. The conjugation transfer was performed by filter membrane incubation method. The competition and plasmid stability assays suggested the host bacteria carrying qnrS had an energy burden. As far as we know, this is the first report that E. hormaechei carrying qnrS was isolated from chicken feed. The chicken feed and poultry products could serve as a vehicle for these MDR bacteria, which could transfer between animals and humans through the food chain. We need to pay close attention to the epidemiology of E. hormaechei and prevent their further dissemination. IMPORTANCE Enterobacter hormaechei is an opportunistic pathogen. It can cause infections in humans and animals. Plasmid-mediated quinolone resistance (PMQR) gene qnrS can be transferred intergenus, which is leading to increase the quinolone resistance levels in Enterobacteriaceae. Chicken feed could serve as a vehicle for the MDR E. hormaechei. Therefore, antibiotic-resistance genes (ARGs) might be transferred to the intestinal flora after entering the gastrointestinal tract with the feed. Furthermore, antibiotic-resistant bacteria (ARB) were also excreted into environment with feces, posing a huge threat to public health. This requires us to monitor the ARB and antibiotic-resistant plasmids in the feed. Here, we demonstrated the characteristics of one MDR E. hormaechei isolate from chicken feed. The plasmid carrying the qnrS gene is a conjugative plasmid with transferability. The presence of plasmid carrying antibiotic-resistance genes requires the maintenance of antibiotic pressure. In addition, the E. hormaechei M1 belonged to new sequence type (ST). These data show the MDR E. hormaechei M1 is a novel strain that requires our further research.

This study aimed to determine the effect of enrofloxacin (ENR) on the transfer of the plasmid-mediated quinolone resistance (PMQR) gene qnrS from opportunistic pathogen Escherichia coli (E2) to Salmonella Enteritidis (SE211) and to analyze the resistance characteristics of SE211-qnrS isolates. The plasmid carrying qnrS gene of E2 was sequenced by Oxford Nanopore technology. The plasmid carrying qnrS gene belonged to incompatibility group IncY. In vitro, the transfer experiment of IncY plasmid was performed by the liquid medium conjugation method. The conjugation transfer frequency of the IncY plasmid was 0.008 ± 0.0006 in the absence of ENR, 0.012 ± 0.003 in 1/32 MICENR, 0.01 ± 0.008 in 1/8 MICENR, and 0.03 ± 0.015 (Mean±SD) in 1/2 MICENR, respectively. After inoculation of E. coli E2 and SE211, chickens were treated with different doses of ENR (3.03, 10, and 50 mg/kg b.w.) for 7 days consecutively. To screen the SE211-qnrS strains from intestinal tract of chickens, the resistance genes and susceptibility of isolates were identified. The amount of E. coli E2 and the copy number of qnrS gene in the chicken intestinal tract were determined by colony counting and qPCR, respectively. In vivo, more SE211-qnrS strains were isolated from the treated group compared with the untreated group. SE211-qnrS strains not only obtained IncY plasmid, but also showed similar resistance phenotype as E2. In conclusion, ENR treatment can promote the spread of a IncY-resistance plasmid carrying the qnrS fluoroquinolone-resistance gene in Escherichia coli and the development of drug-resistant bacteria.

利用调查和数据统计报道我国近年来的兽用抗生素使用、养殖业用药背景、抗菌药物耐药、动物源细菌耐药性监测、动物源细菌耐药监测数据库研究及国家制定的相关政策等现状,并针对我国动物源细菌的耐药和监测现状,提出了我国政府与各地区相关部门应科学评估分析动物源细菌耐药性监测数据及变化趋势;借鉴发达国家的经验和方法,完善我国动物源细菌耐药性监测网络和监测体系;加强教育和宣传等应对措施,为解决全球动物源细菌耐药性问题作出贡献.

Bacterial resistance is a complex scientific issue. To manage this issue, we need to deeply understand the influencing factors and mechanisms. Based on the background of livestock husbandry, this paper reviews the factors that affect the acquisition of bacterial resistance. Meanwhile, the resistance mechanism is also discussed. “Survival of the fittest” is the result of genetic plasticity of bacterial pathogens, which brings about specific response, such as producing adaptive mutation, gaining genetic material or changing gene expression. To a large extent, bacterial populations acquire resistance genes directly caused by the selective pressure of antibiotics. However, mobile resistance genes may be co-selected by other existing substances (such as heavy metals and biocides) without direct selection pressure from antibiotics. This is because the same mobile genetic elements as antibiotic resistance genes can be co-located by the resistance determinants of some of these compounds. Furthermore, environmental factors are a source of resistance gene acquisition. Here, we describe some of the key measures that should be taken to mitigate the risk of antibiotic resistance. We call on the relevant governments or organizations around the world to formulate and improve the monitoring policies of antibiotic resistance, strengthen the supervision, strengthen the international cooperation and exchange, and curb the emergence and spread of drug-resistant strains.

qnrS是质粒介导的喹诺酮类药物的耐药基因,通过保护DNA回旋酶活性从而降低宿主菌对喹诺酮类药物的敏感性.这种保护作用依赖于qnrS基因的表达水平,同时,qnrS的表达主要受喹诺酮类药物浓度的影响,且σ-D调控因子(regulator of sigma D,Rsd)也参与该基因的表达调控.qnrS基因在沙门氏菌中高度流行,并且是大肠埃希氏菌中质粒介导的喹诺酮类耐药基因的优势基因.qnrS可以在细菌间进行传播,或与其他耐药基因共存于同一质粒上发生共转移,从而导致多重耐药的发生.论文对qnrS基因的发现、表达及其调控机制、在沙门氏菌和大肠埃希氏菌中的流行及传播的研究进展进行综述,以期为控制qnrS基因介导的耐药性提供参考.

Background: Salmonella enterica is one of the common pathogens in both humans and animals that causes salmonellosis and threatens public health all over the world. Methods and Results: Here we determined the virulence phenotypes of nine Salmonella enterica subsp. enterica (S. enterica) isolates in vitro and in vivo, including pathogenicity to chicken, cell infection, biofilm formation and virulence gene expressions. S. Enteritidis 211 (SE211) was highly pathogenic with notable virulence features among the nine isolates. The combination of multiple virulence genes contributed to the conferring of the high virulence in SE211. Importantly, many mobile genetic elements (MGEs) were found in the genome sequence of SE211, including a virulence plasmid, genomic islands, and prophage regions. The MGEs and CRISPR-Cas system might function synergistically for gene transfer and immune defense. In addition, the neighbor joining tree and the minimum spanning tree were constructed in this study. Conclusions: This study provided both the virulence phenotypes and genomic features, which might contribute to the understanding of bacterial virulence mechanisms in Salmonella enterica subsp. enterica. The first completed genomic sequence for the high virulent S. Enteritidis isolate SE211 and the comparative genomics and phylogenetic analyses provided a preliminary understanding of S. enterica genetics and laid the foundation for further study.

The purpose of this study was to investigate the changes of resistance phenotype and plasmid-mediated quinolone resistance genes (PMQRs) in Escherichia coli (E. coli) during enrofloxacin (ENR) administration in different breeding cycles. In 2020, 983 strains of E. coli were isolated from different samples in different cycles at the broiler farm with the largest single batch of slaughter capacity in Hebei Province, China. All samples were from chicken, environmental, and human sources. The sensitivity of the isolates to various antibiotics was determined by broth microdilution method. The findings of this study include: (1) the total isolation rate of E. coli in the four cycles was 63.83% (983/1540); (2) the average resistance rate of E. coli from 1-day-old chickens to enrofloxacin was as high as 75% in each cycle, and with the use of enrofloxacin, the resistance rate of E. coli from chickens gradually increased to 100%; (3) 107 strains of E. coli randomly selected from different cycles and sources demonstrated the multi-drug resistance phenotypes. The highest resistance rate was doxycycline (100%), and the lowest was erythromycin (54.21%); (4) the detection rate of PMQRs of E. coli from chickens in different cycles were always higher than that from environmental and human. In particular, the PMQRs pollution rate of chicken seedlings in each cycle were generally higher than that of other sources; (5) We used SPSS software to analyze the Kendall rank correlation of the experimental data. The resistance of E. coli isolated from this farm to ciprofloxacin (CIP) may increase along with the increase of resistance to enrofloxacin (Kendall’s tau-b = 0.190, p = 0.021). All these data highlight the serious problem of bacterial resistance in this farm. Therefore, it is urgent to provide guidance for the prevention and control of colibacillosis and drug resistance in this farm.

动物疼痛和炎症可影响其健康状况、养殖场经济效益等,预防和治疗动物疼痛、炎症具有重要意义.非甾体抗炎药可以减少动物疼痛和炎症的发生.近几年对非甾体抗炎药在动物中的应用研究越来越多,同时非甾体抗炎药的一些新功能被发现.论文对兽医常用非甾体抗炎药物的应用研究进展做一综述,以期为非甾体抗炎药在动物中的合理使用以及进一步研究提供参考.

Salmonella is recognized as one of the most common microbial pathogens worldwide. The bacterium contains the clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated (Cas) systems, providing adaptive immunity against invading foreign nucleic acids. Previous studies suggested that certain bacteria employ the Cas proteins of CRISPR-Cas systems to target their own genes, which also alters the virulence during invasion of mammals. However, whether CRISPR-Cas systems in Salmonella have similar functions during bacterial invasion of host cells remains unknown. Here, we systematically analyzed the genes that are regulated by Cas3 in a type I-E CRISPR-Cas system and the virulence changes due to the deletion of cas3 in Salmonella enterica serovar Enteritidis. Compared to the cas3 gene wild-type (cas3 WT) Salmonella strain, cas3 deletion upregulated the lsrFGBE genes in lsr (luxS regulated) operon related to quorum sensing (QS) and downregulated biofilm-forming-related genes and Salmonella pathogenicity island 1 (SPI-1) genes related to the type three secretion system (T3SS). Consistently, the biofilm formation ability was downregulated in the cas3 deletion mutant (Δcas3). The bacterial invasive and intracellular capacity of Δcas3 to host cells was also reduced, thereby increasing the survival of infected host cells and live chickens. By the transcriptome-wide screen (RNA-Seq), we found that the cas3 gene impacts a series of genes related to QS, the flagellum, and SPI-1-T3SS system, thereby altering the virulence phenotypes. As QS SPI-1-T3SS and CRISPR-Cas systems are widely distributed in the bacteria kingdom, our findings extend our understanding of virulence regulation and pathogenicity in mammalian hosts for Salmonella and potentially other bacteria.

Cyadox, a new antibacterial agent as the quinoxaline-1, 4-dioxides, has a good antibacterial and growth-promoting effect, and has the advantages of lower toxicity, adequate safety and faster absorption. Seven differential expressed genes (DEGs) induced by cyadox were screened in swine liver tissues, including Insulin-like Growth Factor-1 (IGF-1), Epidermal Growth Factor (EGF), Poly ADP-ribose polymerase (PARP), the Defender Against Apoptotic Death 1 (DAD1), Complement Component 3 (C3), Transketolase (TK) and cyadox-related novel gene (CRNG). To elucidate the signal mechanism that cyadox altered these genes expression, the time-effect relationship and signaling pathways related to 7 DEGs induced by cyadox were determined in Porcine Kidney-15 (PK-15) cells by RT-qPCR and the application of various signal pathway inhibitors. The phosphorylation levels of signal factors in PK-15 cells were detected by Western blot. The analyses demonstrated that, the mRNA expressions of 7 DEGs were significantly enhanced by cyadox mainly through the phosphoinositide 3-kinase (PI3K) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-кB) signaling pathways in PK-15 cells. Furthermore, EGF might be the early response gene of cyadox to activate downstream signaling pathways and regulates the expression of other related genes or directly exerting biological effects. In brief, cyadox mainly regulates the expression of these 7 genes by PI3K and NF-кB signaling pathways to exert it's antibacterial and growth-promoting activity in PK-15 cells.

Background: Trichothecenes are a large family of chemically related mycotoxins produced by various species of Fusarium, which is the most economically important fungal strains. Trichothecenes have most often been associated with acute and chronic toxicoses in human and livestock. Trichothecenes are linked with human diseases like alimentary toxic aleukia and Kashin-Beck disease, and also with Fusarium head blight in plants. At molecular level, the mitochondrial toxicities induced by trichothecenes have been investigated, yet no efforts are made to summarize these information. Scope and approach: The present review provides the most recent knowledge on the mitochondrion as a new molecular target of trichothecenes based on findings that trichothecenes can cross the mitochondrial membranes and be absorbed by mitochondrion. Here, we discriminate the ROS production site in trichothecene-treated cells, and discuss the mitochondrion-dependent apoptosis and various mitochondrial toxicities induced by trichothecenes. Some mitochondrion-associated miRNAs and potential treatment strategies against trichothecenes are included, and questions not interpreted clearly and future perspective are recommended. Key findings and conclusions: Recent investigations support that mitochondrion is new molecular target of trichothecenes and should be emphasized. Namely, mitochondrion is the main site of ROS generation in trichothecene-treated cells. Trichothecenes induce collapse of mitochondrial membrane potential, and apoptosis by mitochondrial pathway. Trichothecenes inhibit mitochondrial biogenesis and respiration, ATP generation, and mitochondrial transcription and translation. Trichothecenes inhibit activity of mitochondrial electron transport chain (ETC) and change expression of ETC subunits. Trichothecenes change the expression of numbers of miRNAs and specific mitochondria-associated miRNAs which may be involved in mitochondria-dependent toxicity.

欧盟考虑到抗生素耐药性的发展和抗生素耐药基因可能从动物到人微生物的转移,决定从2006年1月1日起禁止抗生素作为生长促进剂,甚至还建议禁止在动物饲料中全面使用所有的抗菌剂.收集和整理欧盟饲料药物添加剂的禁用清单和法规,介绍欧盟禁用饲料药物添加剂的历史及其对动物健康的影响,以期为中国饲料药物添加剂的使用和管理提供借鉴和参考.

Our previous study has showed that a novel gene is differentially expressed in the liver of cyadox-fed piglets, but its sequence and function are unknown. Here, rapid amplification of cDNA ends (RACE) and bioinformatics analysis showed that the novel gene is 953 bp without protein-coding ability and locates in chromosome 11. Hence, we identified the novel gene as long non-coding RNA (lncRNA) and named it cyadox-related novel gene (CRNG). Fluorescence in situ hybridization (FISH) showed that CRNG mainly distributes in cytoplasm. Moreover, microarray assay in combination with CRNG interference and overexpression showed that the differential genes such as ANPEP, KITLG, STAT5A, FOXP3, miR-451, IL-2, IL-10, IL-6, and TNF-α are mainly involved in viral and pathogens infection and the immune-inflammatory responses in PK-15 cells. This work reveals that CRNG might play a role in preventing the host from being infected by pathogens and viruses and exerting immune regulatory effects in the cytoplasm, which may be involved in prophylaxis of cyadox in piglets.

欧盟允许在饲料和饮水中添加抗菌药和抗球虫药治疗、预防和防治动物疾病.本文收集和整理了欧盟常见的准用饲料药物添加剂清单,说明其适应症和用法用量,并对其授权程序和使用等管理现状进行简单概述,为中国饲料药物添加剂的使用和管理提供借鉴和参考.

Autophagy and inflammasomes are shown to interact in various situations including infectious disease, cancer, diabetes and neurodegeneration. Since multiple layers of molecular regulators contribute to the interplay between autophagy and inflammasome activation, the detail of such interplay remains largely unknown. Non-coding RNAs (ncRNAs), which have been implicated in regulating an expanding list of cellular processes including immune defense against pathogens and inflammatory response in cancer and metabolic diseases, may join in the crosstalk between inflammasomes and autophagy in physiological or disease conditions. In this review, we summarize the latest research on the interlink among ncRNAs, inflammasomes and autophagy and discuss the emerging role of these three in multiple signaling transduction pathways involved in clinical conditions. By analyzing these intriguing interconnections, we hope to unveil the mechanism inter-regulating these multiple processes and ultimately discover potential drug targets for some refractory diseases.