Tracking the progeny of bacterial persisters using a CRISPR-based genomic recorder.

The rise of antimicrobial failure is a global emergency, and causes beyond typical genetic resistance must be determined. One probable factor is the existence of subpopulations of transiently growth-arrested bacteria, persisters, that endure antibiotic treatment despite genetic susceptibility to the drug. The presence of persisters in infected hosts has been successfully established, notably through the development of fluorescent reporters. It is proposed that infection relapse is caused by persisters resuming growth after cessation of the antibiotic treatment, but to date, there is no direct evidence for this. This is because no tool or reporter currently exists to track the extent to which infection relapse is initiated by regrowth of persisters in the host. Indeed, once they have transitioned out of the persister state, the progeny of persisters are genetically and phenotypically identical to susceptible bacteria in the population, making it virtually impossible to ascertain the source of relapse. We designed pSCRATCH (plasmid for Selective CRISPR Array expansion To Check Heritage), a molecular tool that functions to record the state of antibiotic persistence in the genome of Salmonella persisters. We show that pSCRATCH successfully marks persisters by adding spacers in their CRISPR arrays and the genomic label is stable in persister progeny after exit from persistence. We further show that in a Salmonella infection model the system enables the discrimination of treatment failure originating from persistence versus resistance. Thus, pSCRATCH provides proof of principle for stable marking of persisters and a prototype for applications to more complex infection models and other pathogens.

Viral-bacterial co-infections screen in vitro reveals molecular processes affecting pathogen proliferation and host cell viability.

The broadening of accessible methodologies has enabled mechanistic insights into single-pathogen infections, yet the molecular mechanisms underlying co-infections remain largely elusive, despite their clinical frequency and relevance, generally exacerbating symptom severity and fatality. Here, we describe an unbiased in vitro screening of pairwise co-infections in a murine macrophage model, quantifying pathogen proliferation and host cell death in parallel over time. The screen revealed that the majority of interactions are antagonistic for both metrics, highlighting general patterns depending on the pathogen virulence strategy. We subsequently decipher two distinct molecular interaction points: Firstly, murine Adenovirus 3 modifies ASC-dependent inflammasome responses in murine macrophages, altering host cell death and cytokine production, thereby impacting secondary Salmonella infection. Secondly, murine Adenovirus 2 infection triggers upregulation of Mprip, a crucial mediator of phagocytosis, which in turn causes increased Yersinia uptake, specifically in virus pre-infected bone-marrow-derived macrophages. This work therefore encompasses both a first-of-its-kind systematic assessment of host-pathogen-pathogen interactions, and mechanistic insight into molecular mediators during co-infection.

[Rapid serotyping of Salmonella based on matrix assisted laser desorption ionization-time of flight mass spectrometry].

Objective: To establish a matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) assay for the identification of common Salmonella serotypes and provide etiology evidence for the early precise treatment of salmonellosis. Methods: A total of 500 strains were collected from different regions and sources and five predominant Salmonella serotypes (Salmonella Typhi, Salmonella Paratyphi A, Salmonella Typhimurium, Salmonella Enteritidis, and Salmonella Indiana) of each strain was identified by agglutination test and whole-genome sequencing. The protein complex of the strains was extracted by using optimized pretreatment method to establish the fingerprint database of peptides for each Salmonella serotype. The new serotyping assays were established by using different modules based on the mass spectra database. Additional 155 strains with specified serotypes and variant sources were used to test and evaluate the accuracy of the new typing assays. Results: Five MALDI-TOF MS databases were established, and two new serotyping assays were established via peptide fingerprint mapping/matching and machine learning of the neuronal convolutional network respectively based on the databases. The results showed that the fingerprint matching approach could quickly identify five common Salmonella serotypes in clinical practice compared with the machine learning method, the accuracy of fingerprint matching assay to identify five Salmonella serotypes reached 100.00% and the serotyping can be conducted within a short time (15-20 minutes) and had a good reproducibility, while the machine learning method could not completely identify these serotypes. Moreover the sensitivity and specificity of fingerprint matching assay were all 100.00% respectively, while they were only 82.23% and 95.81% for machine learning method. Conclusion: The established Salmonella serotyping assay based on MALDI-TOF MS in this study can easily, rapidly and accurately identify different serotypes of Salmonella.

Isolation and Characterization of Salmonella Bacteriophages as Potential Agents for Phage Therapy of Antibiotic-Resistant Intestinal Infections.

Two bacteriophages specifically active against to pathogenic strains of the Salmonella genus were isolated. The morphology of phage colonies (size, transparency, and shape of the plaque edge, and halo) and the spectrum of their lytic activity and interaction with microbial cells (adsorption rate, duration of the latency, and reproductive efficiency) were examined. Using genome-wide sequencing, we determined the taxonomic position of bacteriophages and verified the absence of unwanted genes encoding toxins, adhesins, and invasins, as well as pathogenicity islands responsible for antibiotic resistance. In addition, phage stability under different physical conditions and their productivity were studied.

Low protein diet protects the liver from Salmonella Typhimurium-mediated injury by modulating the mTOR/autophagy axis in macrophages.

Western diets are the underlying cause of metabolic and liver diseases. Recent trend to limit the consumption of protein-rich animal products has become more prominent. This dietary change entails decreased protein consumption; however, it is still unknown how this affects innate immunity. Here, we studied the influence of a low protein diet (LPD) on the liver response to bacterial infection in mice. We found that LPD protects from Salmonella enterica serovar Typhimurium (S. Typhimurium)-induced liver damage. Bulk and single-cell RNA sequencing of murine liver cells showed reduced inflammation and upregulation of autophagy-related genes in myeloid cells in mice fed with LPD after S. Typhimurium infection. Mechanistically, we found reduced activation of the mammalian target of rapamycin (mTOR) pathway, whilst increased phagocytosis and activation of autophagy in LPD-programmed macrophages. We confirmed these observations in phagocytosis and mTOR activation in metabolically programmed human peripheral blood monocyte-derived macrophages. Together, our results support the causal role of dietary components on the fitness of the immune system.

Prevalence, antimicrobial resistance, and genomic characterization of Salmonella strains isolated in Hangzhou, China: a two-year study.

This study explored the molecular epidemiology and resistance mechanisms of 271 non-duplicate Salmonella enterica (S. enterica) strains, isolated mainly from adults (209/271) in a tertiary hospital in Hangzhou between 2020 and 2021. Through whole-genome sequencing and bioinformatics, the bacterial strains were classified into 46 serotypes and 54 sequence types (ST), with S. Enteritidis, S. 1,4,[5],12:i:-, and S. Typhimurium being the most prevalent serotypes and ST11, ST34, and ST19 the most common STs. The strains isolated from adults were primarily S. Enteritidis (59/209), while from children were mainly S. 1,4,[5],12:i:- (20/62). Worryingly, 12.55% strains were multi-drug resistant (MDR), with resistance rates to cefepime (FEP), ceftazidime (CAZ), ceftriaxone (CRO) and cefotaxime (CTX) of 7.38%, 9.23%, 15.87% and 16.24%, respectively, and resistance rates to levofloxacin (LEV) and ciprofloxacin (CIP) of 8.49% and 19.19%, respectively. It is worth noting that the resistance rates of CRO and CTX in children reached 30.65%. A total of 34 strains carried extended-spectrum ß-lactamase (ESBL) genes, dominated by blaCTX-M-65 (13/34) and blaCTX-M-55 (12/34); it is notable that one strain of S. Saintpaul carried both blaCTX-M-27 and blaCTX-M-55. The resistance mechanism to cephalosporins was mainly due to ESBL genes (20/43), and other genes included AmpC and ß-lactamase genes. The strains resistant to quinolones mainly carried qnrS1 (27/53), and others included qnrB6, aac(6')-Ib-cr, and mutations in gyrA and parC. One strain did not carry common quinolone resistance genes but had a parC (p.T57S) mutation to cause CIP resistance. This research provides vital insights into the molecular epidemiology and resistance mechanisms of clinical S. enterica, implicating possible infection control strategies.

A multi-provincial outbreak of Salmonella Newport infections associated with red onions: A report of the largest Salmonella outbreak in Canada in over 20 years.

An investigation into an outbreak of Salmonella Newport infections in Canada was initiated in July 2020. Cases were identified across several provinces through whole-genome sequencing (WGS). Exposure data were gathered through case interviews. Traceback investigations were conducted using receipts, invoices, import documentation, and menus. A total of 515 cases were identified in seven provinces, related by 0-6 whole-genome multi-locus sequence typing (wgMLST) allele differences. The median age of cases was 40 (range 1-100), 54% were female, 19% were hospitalized, and three deaths were reported. Forty-eight location-specific case sub-clusters were identified in restaurants, grocery stores, and congregate living facilities. Of the 414 cases with exposure information available, 71% (295) had reported eating onions the week prior to becoming ill, and 80% of those cases who reported eating onions, reported red onion specifically. The traceback investigation identified red onions from Grower A in California, USA, as the likely source of the outbreak, and the first of many food recall warnings was issued on 30 July 2020. Salmonella was not detected in any tested food or environmental samples. This paper summarizes the collaborative efforts undertaken to investigate and control the largest Salmonella outbreak in Canada in over 20 years.

Everything but the kitchen sink: The use of multiple hypothesis generation methods to investigate an outbreak of Salmonella Enteritidis associated with frozen profiteroles and eclairs.

In December 2018, an outbreak of Salmonella Enteritidis infections was identified in Canada by whole-genome sequencing (WGS). An investigation was initiated to identify the source of the illnesses, which proved challenging and complex. Microbiological hypothesis generation methods included comparisons of Salmonella isolate sequence data to historical domestic outbreaks and international repositories. Epidemiological hypothesis generation methods included routine case interviews, open-ended centralized re-interviewing, thematic analysis of open-ended interview data, collection of purchase records, a grocery store site visit, analytic comparison to healthy control groups, and case-case analyses. Food safety hypothesis testing methods included food sample collection and analysis, and traceback investigations. Overall, 83 cases were identified across seven provinces, with onset dates from 6 November 2018 to 7 May 2019. Case ages ranged from 1 to 88 years; 60% (50/83) were female; 39% (22/56) were hospitalized; and three deaths were reported. Brand X profiteroles and eclairs imported from Thailand were identified as the source of the outbreak, and eggs from an unregistered facility were hypothesized as the likely cause of contamination. This study aims to describe the outbreak investigation and highlight the multiple hypothesis generation methods that were employed to identify the source.

[The application and reflection of emerging molecular detection technologies in Salmonella detection].

Salmonella is an important foodborne pathogen and one of the main causes of diarrhea. Every year, about 550 million people suffer from diarrhea due to Salmonella infection, of which about 230 000 die. It has become a major global public safety issue. The application fields of Salmonella detection involve food safety, water quality monitoring, animal husbandry, public health monitoring, and medical diagnosis. The detection requirements mainly come from three aspects: pathogen identification, serotype identification, drug resistance and virulence identification. In recent years, the detection technology for Salmonella has made rapid progress, especially the emergence and development of emerging molecular detection technologies, providing new perspectives for Salmonella detection in different scenarios. However, due to the diversity of Salmonella serotypes and the complexity of detection scenarios, existing detection technologies still have some pain points (such as long detection time, cumbersome operation steps, low scene adaptability, etc.). This article will elaborate on the application of several emerging molecular detection technologies with distinct characteristics, such as CRISPR Cas technology, digital PCR technology, sequencing technology, and microfluidic technology, in Salmonella detection. It aims to provide a reference for the development and improvement of Salmonella detection technology and the establishment of infection warning and control systems.

Natural medicine can substitute antibiotics in animal husbandry: protective effects and mechanisms of rosewood essential oil against Salmonella infection.

Aniba rosaeodora essential oil (RO) has been traditionally used in natural medicine as a substitute for antibiotics due to its notable antidepressant and antibacterial properties. Salmonella, a prevalent pathogen in foodborne illnesses, presents a major challenge to current antibiotic treatments. However, the antibacterial efficacy and mechanisms of action of RO against Salmonella spp. remain underexplored. This study aims to elucidate the chemical composition of RO, evaluate its antibacterial activity and mechanisms against Salmonella in vitro, and further delineate its anti-inflammatory mechanisms in vivo during Salmonella infection. Gas chromatography-mass spectrometry (GC-MS) was utilized to characterize the chemical constituents of RO. The antibacterial activity of RO was assessed using minimal inhibitory concentration (MIC) and time-kill assays. Various biochemical assays were employed to uncover the potential bactericidal mechanisms. Additionally, mouse and chick models of Salmonella infection were established to investigate the prophylactic effects of RO treatment. RO exhibited significant antibacterial activity against both Gram-positive and Gram-negative bacteria, with an MIC of 4 mg·mL-1 for Salmonella spp. RO treatment resulted in bacterial damage through the disruption of lipid and purine metabolism. Moreover, RO reduced injury and microbial colonization in infected mice and chicks. RO treatment also modulated the host inflammatory response by inhibiting proinflammatory pathways. In conclusion, our findings demonstrate that RO is effective against Salmonella infection, highlighting its potential as an alternative to antibiotics for antibacterial therapy.

Human tetraspanin CD81 facilitates invasion of Salmonella enterica into human epithelial cells.

Human CD81 and CD9 are members of the tetraspanin family of proteins characterized by a canonical structure of four transmembrane domains and two extracellular loop domains. Tetraspanins are known as molecular facilitators, which assemble and organize cell surface receptors and partner molecules forming clusters known as tetraspanin-enriched microdomains. They have been implicated to play various biological roles including an involvement in infections with microbial pathogens. Here, we demonstrate an important role of CD81 for the invasion of epithelial cells by Salmonella enterica. We show that the overexpression of CD81 in HepG2 cells enhances invasion of various typhoidal and non-typhoidal Salmonella serovars. Deletion of CD81 by CRISPR/Cas9 in intestinal epithelial cells (C2BBe1 and HT29-MTX-E12) reduces S. Typhimurium invasion. In addition, the effect of human CD81 is species-specific as only human but not rat CD81 facilitates Salmonella invasion. Finally, immunofluorescence microscopy and proximity ligation assay revealed that both human tetraspanins CD81 and CD9 are recruited to the entry site of S. Typhimurium during invasion but not during adhesion to the host cell surface. Overall, we demonstrate that the human tetraspanin CD81 facilitates Salmonella invasion into epithelial host cells.

Infection of the ovarian cyst: a rare presentation of extraintestinal salmonellosis.

A woman in her early 20's presented with fever and unintentional weight loss of 4 kg over a period of 1 month and abdominal pain for 10 days. Empirical antibiotic therapy administered prior to hospitalisation was not successful. Evaluation for fever was unrewarding except for an abnormal ultrasound which showed two cysts with the largest dimension of 9 cm in the right adnexal region. All blood cultures were sterile. She underwent laparoscopic cystectomy. Bacterial culture of cyst fluid grew Salmonella enterica subspecies enterica serotype Typhi which was found to be resistant to fluoroquinolones. The case emphasises the fact that localised infection of the ovarian cyst can occur in extraintestinal salmonellosis that can have a negative blood culture and can mimic ovarian malignancy.

Diarrhoeagenic Escherichia coli and Salmonella spp. Contamination of Food and Water Consumed by Children with Diarrhoea in Maputo, Mozambique.

In Mozambique, about 500,000 cases of diarrhoea were caused by foodborne pathogens in 2018. A review of the epidemiology of diarrhoea in children under five showed a high disease burden. This study aimed to identify Diarrhoeagenic Escherichia coli (DEC) and Salmonella spp. contamination of food and water in urban and rural areas of Maputo consumed by children under five with diarrhoea. One hundred and eighty-six children with diarrhoea were selected from Primeiro de Maio and Marracuene Health Care Centres from the Kamaxakeni and Marracuene districts, respectively. Food (n = 167) and water (n = 100) samples were collected in children's households for diarrhoeagenic bacterial identification. Interviews were conducted using a semi-structured questionnaire to collect data about demographics and foods consumed a week before the children's diarrhoea episodes. The prevalence of both DEC and Salmonella spp. was 9.8% in food and 5.4% in water samples. DEC was most prevalent in cereals (urban = 2.8%; rural = 2.4%) and water samples (urban = 1.4%; rural = 3.3%). Salmonella spp. was mainly detected in cereals (urban = 0.7%; rural = 0.8%). Diarrhoeagenic pathogens were associated with the type of food frequently consumed by children under five years with diarrhoea (infant formula, fruit puree, ready-to-eat meals, and bottled water), while the association with demographics was absent. We found that the infant foods consumed by children with diarrhoea are associated with DEC and Salmonella spp., and the prevalence of these contaminants is higher in the rural (8.9%) than in the urban area (6.3%), showing the need for caregiver education on food handling practices.

Heterogeneity of Salmonella enterica lipopolysaccharide counteracts macrophage and antimicrobial peptide defenses.

Salmonella enterica is comprised of over 2,500 serovars, in which non-typhoidal serovars (NTS), Enteritidis (SE), and Typhimurium (STM) are the most clinically associated with human infections. Although NTS have similar genetic elements to cause disease, phenotypic variation including differences in lipopolysaccharide (LPS) composition may control immune evasion. Here, we demonstrate that macrophage host defenses and LL-37 antimicrobial efficacy against SE and STM are substantially altered by LPS heterogeneity. We found that SE evades macrophage killing by inhibiting phagocytosis while STM survives better intracellularly post-phagocytosis. SE-infected macrophages failed to activate the inflammasomes and subsequently produced less interleukin-1ß (IL-1ß), IL-18, and interferon λ. Inactivation of LPS biosynthesis genes altered LPS composition, and the SE LPS-altered mutants could no longer inhibit phagocytosis, inflammasome activation, and type II interferon signaling. In addition, SE and STM showed differential susceptibility to the antimicrobials LL-37 and colistin, and alteration of LPS structure substantially increased susceptibility to these molecules. Collectively, our findings highlight that modification of LPS composition by Salmonella increases resistance to host defenses and antibiotics.

Eosinophils respond to, but are not essential for control of an acute Salmonella enterica serovar Typhimurium infection in mice.

Eosinophils are a highly abundant cell type in the gastrointestinal tract during homeostatic conditions, where they have recently been reported to take on an activated phenotype following colonization by the bacterial microbiota. To date, there have been few studies investigating whether eosinophils respond to infection with enteric bacterial pathogens and/or investigating the requirements for eosinophils for effective bacterial pathogen control. In this study, we investigated the response of eosinophils to an acute enteric infection of mice with the bacterial pathogen Salmonella enterica serovar Typhimurium. We also assessed whether eosinophil deficiency impacted Salmonella burdens in the intestinal tract or impacted the systemic dissemination of Salmonella following an oral infection of littermate wild-type BALB/cJ and eosinophil-deficient ΔdblGATA BALB/cJ mice. We found comparable Salmonella burdens in the intestinal tract of wild-type and eosinophil-deficient mice and no significant differences in the levels of Salmonella disseminating to systemic organs within 3 days of infection. Despite our evidence suggesting that eosinophils are not an essential cell type for controlling bacterial burdens in this acute infection setting, we found higher levels of eosinophils in gut-draining lymph nodes following infection, indicating that eosinophils do respond to Salmonella infection. Our data contribute to the growing evidence that eosinophils are responsive to bacterial stimuli, yet the influence of and requirements for eosinophils during bacterial infection appear to be highly context-dependent.

Salmonella Typhimurium infection inhibits macrophage IFNß signaling in a TLR4-dependent manner.

Type I Interferons (IFNs) generally have a protective role during viral infections, but their function during bacterial infections is dependent on the bacterial species. Legionella pneumophila, Shigella sonnei and Mycobacterium tuberculosis can inhibit type I IFN signaling. Here we examined the role of type I IFN, specifically IFNß, in the context of Salmonella enterica serovar Typhimurium (STm) macrophage infections and the capacity of STm to inhibit type I IFN signaling. We demonstrate that IFNß has no effect on the intracellular growth of STm in infected bone marrow derived macrophages (BMDMs) derived from C57BL/6 mice. STm infection inhibits IFNß signaling but not IFNγ signaling in a murine macrophage cell line. We show that this inhibition is independent of the type III and type VI secretion systems expressed by STm and is also independent of bacterial phagocytosis. The inhibition is Toll-like receptor 4 (TLR4)-dependent as the TLR4 ligand, lipopolysaccharide (LPS), alone is sufficient to inhibit IFNß-mediated signaling. Cells downregulated their surface levels of IFNα/ß receptor 1 (IFNAR1) in response to LPS, which may be mediating our observed inhibition. Lastly, we examined this inhibition in the context of TLR4-deficient BMDMs as well as TLR4 RNA interference and we observed a loss of inhibition with LPS stimulation as well as STm infection. In summary, we show that macrophages exposed to STm have reduced IFNß signaling via crosstalk with TLR4 signaling, which may be mediated by reduced host cell surface IFNAR1, and that IFNß signaling does not affect cell-autonomous host defense against STm.

Endoprotein-activating DNAzyme assay for nucleic acid extraction- and amplification-free detection of viable pathogenic bacteria.

Pathogenic bacteria in food or environment, can pose threats to public health, highlighting the requirement of tools for rapid and accurate detection of viable pathogenic bacteria. Herein, we report a sequential endoprotein RNase H2-activating DNAzyme assay (termed epDNAzyme) that enables nucleic acid extraction- and amplification-free detection of viable Salmonella enterica (S. enterica). The direct detection allows for a rapid detection of viable S. enterica within 25 min. Besides, the assay, based on sequential reporting strategy, circumvents internal modifications in the DNAzyme's active domain and improve its catalytic activity. The multiple-turnover DNAzyme cutting and the enhanced catalytic activity of DNAzyme render the epDNAzyme assay to be highly sensitive, and enables the detection of 190 CFU/mL and 0.1% viable S. enterica. The assay has been utilized to detect S. enterica contamination in food and clinical samples, indicating its potential as a promising tool for monitoring pathogen-associated biosafety.

Prevalence and characteristics of ESBL-producing Salmonella in Weifang, China.

This study examined the prevalence and antibiotic resistance pattern of blaCTX-M extended-spectrum ß-lactamase positive Salmonella species isolated from a hospital in Weifang. Salmonella strains were isolated from hospitalized patients from January 2018 to April 2023. Whole-genome sequencing was performed by Illumina platform. CTX-M-producing Salmonella were identified by Comprehensive Antibiotic Research Database (CARD). Strain susceptibility to six antimicrobial agents was assessed by BD Phoenix™ M50 System. MLST analysis confirmed sequence types and additionally, serotypes were determined by SeqSero2. Genetic environments of blaCTX-M genes were analyzed by Isfinder and BLASTn. Single nucleotide polymorphisms were used to construct a phylogenetic tree to analyze homology. A total of 34 CTX-M-producing Salmonella were detected. The most prevalent serotype was Salmonella enterica subsp. enterica 1,4,[5],12:i:- (14/34, 41.18%), belonging to ST34, followed by Salmonella Enteritidis (10/34, 29.41%), belonging to ST11. The highest resistance rate was detected to ampicillin (97.06%), followed by ceftriaxone (94.12%) and ceftazidime (58.83%). In CTX-M-producing Salmonella five types of blaCTX-M genes were identified, the most prevalent was blaCTX-M-55 (47.06%, 16/34), followed by blaCTX-M-14, blaCTX-M-65, blaCTX-M-125, and blaCTX-M-27 at 26.47% (9/34), 11.77% (4/34), 8.82% (3/34), and 5.88% (2/34), respectively. Apart from blaCTX-M, 40 antibiotic resistance genes were also detected, conveying resistance to multiple drugs and the most frequent genes were namely, mcr-1.1, aph(6)-Id, aph(3″)-Ib, oqxAB, qnrB6, qnrS1. According to genetic environment analysis, the insertion sequence ISEcp1 was prevalent upstream of the blaCTX-M gene. Our study demonstrates that multiple resistance genes are carried by clinical isolates of Salmonella spp. however, the dominant ESBL genotype is CTX-M-55, that is associated with ISEcp1.

CCL28 modulates neutrophil responses during infection with mucosal pathogens.

The chemokine CCL28 is highly expressed in mucosal tissues, but its role during infection is not well understood. Here, we show that CCL28 promotes neutrophil accumulation in the gut of mice infected with Salmonella and in the lung of mice infected with Acinetobacter. Neutrophils isolated from the infected mucosa expressed the CCL28 receptors CCR3 and, to a lesser extent, CCR10, on their surface. The functional consequences of CCL28 deficiency varied between the two infections: Ccl28-/- mice were highly susceptible to Salmonella gut infection but highly resistant to otherwise lethal Acinetobacter lung infection. In vitro, unstimulated neutrophils harbored pre-formed intracellular CCR3 that was rapidly mobilized to the cell surface following phagocytosis or inflammatory stimuli. Moreover, CCL28 stimulation enhanced neutrophil antimicrobial activity, production of reactive oxygen species, and formation of extracellular traps, all processes largely dependent on CCR3. Consistent with the different outcomes in the two infection models, neutrophil stimulation with CCL28 boosted the killing of Salmonella but not Acinetobacter. CCL28 thus plays a critical role in the immune response to mucosal pathogens by increasing neutrophil accumulation and activation, which can enhance pathogen clearance but also exacerbate disease depending on the mucosal site and the infectious agent.

Plasmid-mediated azithromycin resistance in non-typhoidal Salmonella recovered from human infections.

OBJECTIVES: Mechanisms of non-typhoidal Salmonella (NTS) resistance to azithromycin have rarely been reported. Here we investigate the epidemiology and genetic features of 10 azithromycin-resistant NTS isolates. METHODS: A total of 457 NTS isolates were collected from a tertiary hospital in Guangzhou. We performed antimicrobial susceptibility tests, conjugation experiments, efflux pump expression tests, whole-genome sequencing and bioinformatics analysis to conduct the study. RESULTS: The results showed that 10 NTS isolates (2.8%) were resistant to azithromycin with minimum inhibitory concentration values ranging from 128 to 512 mg/L and exhibited multidrug resistance. The phylogenetic tree revealed that 5 S. London isolates (AR1-AR5) recognized at different times and departments were closely related [3-74 single-nucleotide polymorphisms (SNPs)] and 2 S. Typhimurium isolates (AR7 and AR8) were clones (<3 SNPs) at 3-month intervals. The azithromycin resistance was conferred by mph(A) gene found on different plasmids, including IncFIB, IncHI2, InFII, IncC and IncI plasmids. Among them, IncFIB, InFII and IncHI2 plasmids carried different IS26-class 1 integron (intI1) arrangement patterns that mediated multidrug resistance transmission. Conjugative IncC plasmid encoded resistance to ciprofloxacin, ceftriaxone and azithromycin. Furthermore, phylogenetic analysis demonstrated that mph(A)-positive plasmids closely related to 10 plasmids in this study were mainly discovered from NTS, Escherichia coli, Klebsiella pneumonia and Enterobacter hormaechei. The genetic environment of mph(A) in 10 NTS isolates was IS26-mph(A)-mrx(A)-mphR(A)-IS6100/IS26 that co-arranged with intI1 harbour multidrug-resistant (MDR) gene cassettes on diverse plasmids. CONCLUSIONS: These findings highlighted that the dissemination of these plasmids carrying mph(A) and various intI1 MDR gene cassettes would seriously restrict the availability of essential antimicrobial agents for treating NTS infections.