Genomic Insights into the Increased Occurrence of Campylobacteriosis Caused by Antimicrobial-Resistant Campylobacter coli.

Campylobacter is the leading bacterial cause of diarrheal illnesses worldwide. Campylobacter jejuni and C. coli are the most common species accounting for campylobacteriosis. Although the proportion of campylobacteriosis caused by C. coli is increasing rapidly in China, the underlying mechanisms of this emergence remain unclear. In this study, we analyzed the whole-genome sequences and associated environments of 1,195 C. coli isolates with human, poultry, or porcine origins from 1980 to 2021. C. coli isolates of human origin were closely related to those from poultry, suggesting that poultry was the main source of C. coli infection in humans. Analysis of antimicrobial resistance determinants indicated that the prevalence of multidrug-resistant C. coli has increased dramatically since the 2010s, coinciding with the shift in abundance from C. jejuni to C. coli in Chinese poultry. Compared with C. jejuni, drug-resistant C. coli strains were better adapted and showed increased proliferation in the poultry production environment, where multiple antimicrobial agents were frequently used. This study provides an empirical basis for the molecular mechanisms that have enabled C. coli to become the dominant Campylobacter species in poultry; we also emphasize the importance of poultry products as sources of campylobacteriosis caused by C. coli in human patients. IMPORTANCE The proportion of campylobacteriosis caused by C. coli is increasing rapidly in China. Coincidentally, the dominant species of Campylobacter occurring in poultry products has shifted from C. jejuni to C. coli. Here, we analyzed the whole-genome sequences of 1,195 C. coli isolates from different origins. The phylogenetic relationship among C. coli isolates suggests that poultry was the main source of C. coli infection in humans. Further analysis indicated that antimicrobial resistance in C. coli strains has increased dramatically since the 2010s, which could facilitate their adaptation in the poultry production environment, where multiple antimicrobial agents are frequently used. Thus, our findings suggest that the judicious use of antimicrobial agents could mitigate the emergence of multidrug-resistant C. coli strains and enhance clinical outcomes by restoring drug sensitivity in Campylobacter.

Mucoid Acinetobacter baumannii enhances anti-phagocytosis through reducing C3b deposition.

Background: Multidrug resistant (MDR) Acinetobacter baumannii causes serious infections in intensive care units and is hard to be eradicated by antibiotics. Many A. baumannii isolates are identified as the mucoid type recently, but the biological characteristics of mucoid A. baumannii and their interactions with host cells remains unclear. Methods: The mucoid phenotype, antimicrobial susceptibility, biofilm-forming ability, acid resistance ability, peroxide tolerance, and in vivo toxicity of clinical ICUs derived A. baumannii isolates were first investigated. Secondly, the phagocytic resistance and invasive capacity of A. baumannii isolates to macrophages (MH-S, RAW264.7) and epithelial cells (A549) were analyzed. Furthermore, the abundance of C3b (complement factor C3 degradation product) deposition on the surface of A. baumannii was investigated. Last, the relationship between C3b deposition and the abundance of capsule in A. baumannii isolates were analyzed. Results: These A. baumannii strains showed different mucoid phenotypes including hyper mucoid (HM), medium mucoid (MM), and low mucoid (LM). All tested strains were MDR with high tolerance to either acid or hydrogen peroxide exposure. Notably, these mucoid strains showed the increase of mortality in the Galleria mellonella infection models. Besides, the HM strain exhibited less biofilm abundance, higher molecular weight (MW) of capsule, and greater anti-phagocytic activity to macrophages than the LM strain. Together with the increased abundance of capsule, high expression of tuf gene (associated with the hydrolysis of C3b), the HM strain effectively inhibits C3b deposition on bacterial surface, resulting in the low-opsonization phenotype. Conclusion: Capsular characteristics facilitate the anti-phagocytic activity in hyper mucoid A. baumannii through the reduction of C3b deposition. Mucoid A. baumannii exhibits high phagocytosis resistance to both macrophages and epithelial cells.

Collateral sensitivity to pleuromutilins in vancomycin-resistant Enterococcus faecium.

The acquisition of resistance to one antibiotic sometimes leads to collateral sensitivity to a second antibiotic. Here, we show that vancomycin resistance in Enterococcus faecium is associated with a remarkable increase in susceptibility to pleuromutilin antibiotics (such as lefamulin), which target the bacterial ribosome. The trade-off between vancomycin and pleuromutilins is mediated by epistasis between the van gene cluster and msrC, encoding an ABC-F protein that protects bacterial ribosomes from antibiotic targeting. In mouse models of vancomycin-resistant E. faecium colonization and septicemia, pleuromutilin treatment reduces colonization and improves survival more effectively than standard therapy (linezolid). Our findings suggest that pleuromutilins may be useful for the treatment of vancomycin-resistant E. faecium infections.

Genomic Insight into the Antimicrobial Resistance of Streptococcus Suis - Six Countries, 2011-2019.

WHAT IS ALREADY KNOWN ON THIS TOPIC?: Streptococcus suis (S. suis) is a zoonotic pathogen causing disease in humans and animals, and the emergence of its increased resistance to antimicrobial agents has become a significant challenge in many countries. WHAT IS ADDED BY THIS REPORT?: Using whole genome sequencing data to accurately predict antimicrobial resistance determinants, it was found that the prevalence of antimicrobial resistance genes was higher in the pig isolates of S. suis than in the human isolates and that the prevalence of these genes varied with serotype. WHAT ARE THE IMPLICATIONS FOR PUBLIC HEALTH PRACTICE?: The data regarding S. suis antimicrobial resistance will help guide rational drug use in the clinic to better protect the health of humans and animals.

Plant Natural Flavonoids Against Multidrug Resistant Pathogens.

The increasing emergence and dissemination of multidrug resistant (MDR) bacterial pathogens accelerate the desires for new antibiotics. Natural products dominate the preferred chemical scaffolds for the discovery of antibacterial agents. Here, the potential of natural flavonoids from plants against MDR bacteria, is demonstrated. Structure-activity relationship analysis shows the prenylation modulates the activity of flavonoids and obtains two compounds, α-mangostin (AMG) and isobavachalcone (IBC). AMG and IBC not only display rapid bactericidal activity against Gram-positive bacteria, but also restore the susceptibility of colistin against Gram-negative pathogens. Mechanistic studies generally show such compounds bind to the phospholipids of bacterial membrane, and result in the dissipation of proton motive force and metabolic perturbations, through distinctive modes of action. The efficacy of AMG and IBC in four models associated with infection or contamination, is demonstrated. These results suggest that natural products of plants may be a promising and underappreciated reservoir to circumvent the existing antibiotic resistance.

A Marine Antibiotic Kills Multidrug-Resistant Bacteria without Detectable High-Level Resistance.

Antibiotic resistance nowadays is spreading much faster than the introduction of new antibiotics into clinical practice. There is an urgent need for potential compounds to combat multidrug-resistant (MDR) bacteria. Marine fungi provide a promising source for chemical diversity with antibiotic-like molecules. To identify structurally distinct compounds that effectively eradicate MDR pathogens and to control the development of antibiotic resistance, we have reinvestigated equisetin, a previously reported meroterpenoid isolated from a marine sponge-derived fungus. Equisetin exerted efficient antibacterial activities against either MRSA or VRE without detectable high-level resistance. Meanwhile, equisetin, as an antibiotic adjuvant, restores colistin susceptibility to colistin-resistant bacteria toward diverse Gram-negative pathogens. Intriguingly, the low-level equisetin-resistant Staphylococcus aureus displayed collateral sensitivity to multiple classes of existing antibiotics with decreased capacity to produce biofilm. Lastly, equisetin showed efficacy with MRSA in three infected animal models. This work suggests that equisetin derived from marine natural products is a promising lead to overcome antibiotic resistance, providing new insight in future antibiotic discovery and development.

Sodium dehydroacetate induces cardiovascular toxicity associated with Ca2+ imbalance in zebrafish.

The environmental effects of additives have attracted increasing attention. Sodium dehydroacetate (DHA-S), as an approved preservative, is widely added in processed foods, cosmetics and personal care products. However, DHA-S has been recently reported to induce hemorrhage and coagulation aberration in rats. Yet little is known about the ecotoxicological effect and underlying mechanisms of DHA-S. Here, we utilized the advantage of zebrafish model to evaluate such effects. DHA-S induced cerebral hemorrhage, mandibular dysplasia and pericardial edema in zebrafish after 24 h exposure (48-72 hpf) at 50 mg/L. We also observed the defective heart looping and apoptosis in DHA-S-treated zebrafish through o-dianisidine and acridine orange staining. Meanwhile, DHA-S induced the deficiency of Ca2+ and vitamin D3 in zebrafish. We further demonstrated that DHA-S stimulated Ca2+ influx resulting in Ca2+-dependent mitochondrial damage in cardiomyocytes. Additionally, DHA-S inhibited glucose uptake and repressed the biosynthesis of amino acids. Finally, we identified that sodium bicarbonate could rescue zebrafish from DHA-S induced cardiovascular toxicity. Altogether, our results suggest that DHA-S is a potential risk for cardiovascular system.

Genomic and Phenotypic Analysis of Persistent Carbapenem-Resistant Klebsiella pneumoniae Isolates from a 5-Year Hospitalized Patient.

The emerging epidemic of carbapenem-resistant Klebsiella pneumoniae (CRKP) is a global public health crisis. However, the phylogenetic affiliation and pathotypic status of CRKP strains in the host colonization period under consistent antibiotic treatments are not well characterized. In this study, a 5-year tracking study was performed, in which a patient admitted to an intensive care unit was recruited and then screened for the carriage of CRKP based on microbiological culture. Nine isolates from the sputum or stool samples were acquired and subjected to real-time whole-genome sequencing, antimicrobial susceptibility testing, Galleria mellonella larval infection, and epithelial cell invasion assay. All nine isolates showed phenotypic resistance to carbapenems, quinolones, and aminoglycosides. Altogether, blaKPC-2 and 10 other antibiotic resistance genes were identified and all nine CRKP isolates exhibited low virulence with more than 38 virulence factors. All but one variant belonged to ST11 with a novel sequence type, differing at the phoE locus. The isolates shared the same plasmid replicon type, prophages, and capsular serotype (K47) with few single-nucleotide polymorphism variations, consistent with epidemiological clones. Furthermore, these CRKP isolates displayed the ability of moderate invasion of lung epithelial cells. Meanwhile, a deficiency of chromosomal type IV secretion system-related gene cluster was detected after 2 years of carriage. Our findings demonstrated that low-virulence CRKP clones could colonize in the gut and respiratory tract under multiple antibiotic stresses, suggesting the strong colonization adaptability of CRKP to the host. Ethical approval was given by The Second Affiliated Hospital of Zhejiang University, School of Medicine, (2018-039).

Sodium dehydroacetate exposure decreases locomotor persistence and hypoxia tolerance in zebrafish.

Environmental exposure to sodium dehydroacetate (DHA-S) is inevitable as DHA-S is a high-volume preservative widely used in cosmetics, processed foods and personal care products. DHA-S is absorbed rapidly when administered orally or on the skin and generally considered to be safe and well tolerated. However, DHA-S has recently been reported to induce weight loss and allergic contact dermatitis, yet little is known about how DHA-S affect the related biological processes. Here, we characterize the biological effects of DHA-S on zebrafish model by directly waterborne exposure. Zebrafish is susceptible to DHA-S exposure at early developmental stage. DHA-S decreased the hatch rate and locomotor persistence of zebrafish, and eventually induced lethality during the continuous exposure at relatively low concentrations of commonly addition. Acute DHA-S exposure decreased respiration capacity in larval zebrafish, promoted the expression of HIF-1α (hypoxia-inducible factor-1α) and caused rapid adult zebrafish death in 30 h. We further demonstrated that DHA-S inhibited the activity of succinate dehydrogenase (SDH) inducing respiratory chain interruption, energy deficiency and organic acids accumulation. These results suggest that the approved DHA-S may pose serious environmental/ecological pressures on the aquatic animal's migration.

Sublethal Levels of Antibiotics Promote Bacterial Persistence in Epithelial Cells.

Antibiotic therapy and host cells frequently fail to eliminate invasive bacterial pathogens due to the emergence of antibiotic resistance, resulting in the relapse and recurrence of infections. Bacteria evolve various strategies to persist and survive in epithelial cells, a front-line barrier of host tissues counteracting invasion; however, it remains unclear how bacteria hijack cellular responses to promote cytoplasmic survival under antibiotic therapy. Here, it is demonstrated that extracellular bacteria show invasive behavior and survive in epithelial cells in both in vivo and in vitro models, to increase antibiotic tolerance. In turn, sublethal levels of antibiotics increase bacterial invasion through promoting the production of bacterial virulence factors. Furthermore, antibiotic treatments interrupt lysosomal acidification in autophagy due to the internalized bacteria, using Bacillus cereus and ciprofloxacin as a model. In addition, it is found that sublethal levels of ciprofloxacin cause mitochondrial dysfunction and reactive oxygen species (ROS) accumulation to impair lysosomal vascular tape ATPase (V-ATPase) to further promote bacterial persistence. Collectively, these results highlight the potential of host cells mediated antibiotic tolerance, which markedly compromises antibiotic efficacy and worsens the outcomes of infection.

Capsular serotypes, antimicrobial susceptibility, and the presence of transferable oxazolidinone resistance genes in Streptococcus suis isolated from healthy pigs in China.

Streptococcus suis is a pig pathogen and a vector of zoonotic diseases that can cause severe systemic infection in humans. S. suis can colonize the nasal cavity, tonsils, and upper respiratory, genital, and digestive tracts in healthy pigs. Here, to determine prevalence, serotype distribution, and antimicrobial susceptibility of S. suis in healthy pigs, we collected 1813 nasal cavity samples from healthy pigs raised on 17 independent farms in six Chinese provinces between 2016 and 2018. We obtained 223 S. suis isolates (12.3 %) and the antimicrobial susceptibility to a panel of 11 antimicrobial agents was measured by microbroth dilution. Most S. suis isolates (98.7 %) were resistant to at least three classes of antimicrobial agents. The optrA gene conferring resistance to oxazolidinones and phenicols was identified in the chromosome of 27 isolates and on a ∼40-kb plasmid in one isolate; to the best of our knowledge, this was the first report of plasmid-borne optrA gene in S. suis. The genetic environment of optrA showed substantial diversity and could be divided into eleven different types. Interestingly, some fragments of the 89 K pathogenicity island (PAI) were observed together with optrA in 3 isolates, which warrants further attention. Capsular serotypes of S. suis isolates were determined by multiplex PCR. Serotype 29 was the most prevalent, followed by serotype 7 and serotype 2. The presence of highly virulent serotype 2 strains may pose a threat to public health.

Characterization of Bacillus cereus in Dairy Products in China.

Bacillus cereus is a common and ubiquitous foodborne pathogen with an increasing prevalence rate in dairy products in China. High and unmet demands for such products, particularly milk, raise the risk of B. cereus associated contamination. The presence of B. cereus and its virulence factors in dairy products may cause food poisoning and other illnesses. Thus, this review first summarizes the epidemiological characteristics and analytical assays of B. cereus from dairy products in China, providing insights into the implementation of intervention strategies. In addition, the recent achievements on the cytotoxicity and mechanisms of B. cereus are also presented to shed light on the therapeutic options for B. cereus associated infections.

Prevalence of pathogens harbouring mobile antimicrobial resistance genes and virulence factors in retail beef and mutton.

Food safety is always a global issue, due to the increased dissemination of antimicrobial resistance and food poisoning related to foodborne bacterial pathogens. The purpose of this study was to assess the risk of potential foodborne bacteria of beef and mutton in retail stores. A total of 134 samples were collected from 24 local markets in Beijing, including raw and cooked beef or mutton, as well as samples derived from the corresponding environment and human beings. We obtained 674 isolates, of which Klebsiella spp. and Staphylococcus spp. were the dominant bacterial species in the meat samples and the environmental samples, respectively. Additionally, environmental bacteria are common in samples from different sources. Based on the results of antimicrobial sensitivity testing, resistance to tetracycline (with a resistance rate of 47.40%), amoxicillin + clavulanate (47.13%) and erythromycin (28.03%) were the major resistant phenotypes. According to the whole genome analysis, the extended spectrum beta-lactamase genes harboured by two K. pneumoniae strains isolated from cooked and raw beef were located on mobile elements. The major toxin genes of Bacillus cereus and adhesion- or invasion-related virulence factors were also shared among isolates from different sources. These factors pose potential risks to public health and need attention.

A TaqMan-based multiplex real-time PCR assay for the rapid detection of tigecycline resistance genes from bacteria, faeces and environmental samples.

BACKGROUND: Tigecycline is a last-resort antibiotic used to treat severe infections caused by extensively drug-resistant bacteria. Recently, novel tigecycline resistance genes tet(X3) and tet(X4) have been reported, which pose a great challenge to human health and food security. The current study aimed to establish a TaqMan-based real-time PCR assay for the rapid detection of the tigecycline-resistant genes tet(X3) and tet(X4). RESULTS: No false-positive result was found, and the results of the TaqMan-based real-time PCR assay showed 100% concordance with the results of the sequencing analyses. This proposed method can detect the two genes at the level of 1 × 102 copies/µL, and the whole process is completed within an hour, allowing rapid screening of tet(X3) and tet(X4) genes in cultured bacteria, faeces, and soil samples. CONCLUSION: Taken together, the TaqMan-based real-time PCR method established in this study is rapid, sensitive, specific, and is capable of detecting the two genes not only in bacteria, but also in environmental samples.

A broad-spectrum antibiotic adjuvant reverses multidrug-resistant Gram-negative pathogens.

The rapid emergence and dissemination of multidrug-resistant (MDR) bacterial pathogens pose a serious threat to global healthcare. One particular concern is the carbapenem-resistant Enterobacteriaceae (CRE), a group of Gram-negative bacteria that have evolved resistance to all or nearly all available antibiotics. Coupled with the fact of barren antibiotic development pipeline nowadays, a critical approach is to revitalize existing antibiotics using antibiotic adjuvants. We found a short linear antibacterial peptide (SLAP)-S25 carrying four non-natural amino acids of 2,4-diaminobutanoic acid (Dab), which solely showed weak antibacterial activity but boosted the efficacy of antibiotics covering all major classes, including cefepime, colistin, ofloxacin, rifampicin, tetracycline and vancomycin, against MDR Gram-negative pathogens. Mechanistic studies showed that SLAP-S25 triggers membrane damage by binding to both lipopolysaccharide (LPS) in the outer membrane and phosphatidylglycerol (PG) in bacterial cytoplasmic membrane, to potentiate antibiotic efficacy through collaborative strategies. Lastly, SLAP-S25 effectively enhanced the activity of colistin against MDR Escherichia coli-associated infections in three animal models. Our findings provide a potential therapeutic option using existing antibiotics in combination with broad-spectrum antibiotic adjuvants, to address the prevalent infections caused by MDR Gram-negative pathogens worldwide.

Toxins and mobile antimicrobial resistance genes in Bacillus probiotics constitute a potential risk for One Health.

Probiotic microbes conferring health benefits to the hosts have attracted great attention. However, the safety of probiotics is not guaranteed, although the increasing widespread use of probiotics with excellent overall safety records. Here, we performed a systematic evaluation of the safety of commercial Bacillus probiotics intended for usage in humans, animals, plants, aquaculture and environment in China. Nearly half of the 65 isolated Bacillus spp. strains from these commercial probiotic products were capable of producing hazardous toxins. Infections with the representative isolates could cause sepsis, intestinal inflammation and liver injury in different mouse models. Additionally, these isolates harbor multiple antimicrobial resistance genes coupled with mobile genetic elements. Collectively, the capability for producing various toxins and harboring mobile antimicrobial resistance genes in Bacillus probiotics indicates a potential risk for One Health.

Multifaceted toxin profile, an approach toward a better understanding of probiotic Bacillus cereus.

Strains of the Bacillus cereus group have been widely used as probiotics for human beings, food animals, plants, and environmental remediation. Paradoxically, B. cereus is responsible for both gastrointestinal and nongastrointestinal syndromes and represents an important opportunistic food-borne pathogen. Toxicity assessment is a fundamental issue to evaluate safety of probiotics. Here, we summarize the state of our current knowledge about the toxins of B. cereus sensu lato to be considered for safety assessment of probiotic candidates. Surfactin-like emetic toxin (cereulide) and various enterotoxins including nonhemolytic enterotoxin, hemolysin BL, and cytotoxin K are responsible for food poisoning outbreaks characterized by emesis and diarrhea. In addition, other factors, such as hemolysin II, Certhrax, immune inhibitor A1, and sphingomyelinase, contribute to toxicity and overall virulence of B. cereus.

Ca2+ protect zebrafish embryos from water acidification.

Ionizable strategies are routinely used to enhance the solubility and dissolution rates of various pharmaceuticals. These chemicals may directly affect aquatic environment once discharged from factories, hospitals or livestock farms. Here, we assessed the potential side effect of tetracyclines (TCs) on the development of zebrafish embryos. Tetracycline hydrochloride decreased water pH from 6.4 to 4.4 at 30 mg/L. Acidified water exceeded the tolerance of zebrafish embryos in pure water during the early ten hours post fertilization (hpf). Interestingly, we found that Ca2+ in the embryo medium could increase the tolerance of embryos to acidified water. Furthermore, we found that the protection of Ca2+ was not due to the formation of TCs-Ca2+ complexes under acidic condition, based on spectral analysis. Meanwhile we showed that exogenous addition of Ca2+ could inhibit the accumulation of Ca2+ from the cytoplasm to the surface of embryos. These results may shed light on the strategies for protecting aquatic animals from acidic environments.

Macrolides induce severe cardiotoxicity and developmental toxicity in zebrafish embryos.

Macrolide antibiotics (MALs) are widely used for both human and animal health. Most MALs and their metabolites transfer into aquatic organisms and environment resulting in violent consequences. Previous studies show that MALs cause cardiotoxicity in humans and mammals. However, the potential risk of these chemicals in aquatic organisms remains unclear. Here, we used zebrafish embryos as a model to evaluate the toxicity of MALs. Zebrafish embryos were exposed to four typical MALs including azithromycin (AZM), clarithromycin (CLR), tilmicosin (TMS) and tylosin (TYL) to study their cardiotoxicity. The heart rate of zebrafish embryos showed similar biphasic distribution in the presence of four MALs at 2 days post-fertilization (dpf). The heart rate increased significantly at low levels of MALs while decreased obviously at high levels. Subsequently, TMS was chose to study its acute toxicity and developmental toxicity, which caused pericardial edema and spinal curvature in zebrafish embryos at 4 dpf. Furthermore, we found that TMS triggered oxidative stress, with decreased SOD activities and increased MDA contents. Lastly, apoptosis was observed in zebrafish embryos under TMS treatment, with up-regulation of apoptosis associated genes such as p53, bcl 2, bax, caspase 3 and caspase 9, confirmed by increased protein expression based on Western blot analysis. Taken together, these data indicate that MALs can cause serious toxicity in the development of zebrafish. Great caution should be taken due to the huge consumption of MALs for food animal production and treatments with TMS for infections in aquaculture.

Nonribosomal antibacterial peptides that target multidrug-resistant bacteria.

Covering: 2000 to 2018, particularly from 2010 to early 2018 The increase in the incidence of antibiotic resistant infections is threatening to overwhelm healthcare practices worldwide. Most antibiotics in clinical use are becoming ineffective, so therefore it is imperative to develop new antibiotics and novel therapeutic strategies. Traditionally, the chemical and mechanistic diversity of nonribosomal antibacterial peptides (NRAPs) as lead compounds have meant that their structures are ideal for antibiotic discovery. Here, we summarize the state of our current knowledge about the mechanisms of antibiotic resistance, which can be used to guide the development of new antibiotics. Furthermore, we provide an overview of NRAPs for treating multi-drug resistant bacteria, including innovative approaches for screening NRAPs from new sources and the underlying mechanisms of antibacterial activity. Finally, we discuss the design of NRAP scaffolds for precise medicine and combinatorial NRAP therapies with existing antibiotics to overcome resistance, which will help to control infections in the post-antibiotic era.