Serotype diversity and antimicrobial susceptibility profiles of Actinobacillus pleuropneumoniae isolated in Italian pig farms from 2015 to 2022.

Actinobacillus pleuropneumoniae (APP) is a bacterium frequently associated with porcine pleuropneumonia. The acute form of the disease is highly contagious and often fatal, resulting in significant economic losses for pig farmers. Serotype diversity and antimicrobial resistance (AMR) of APP strains circulating in north Italian farms from 2015 to 2022 were evaluated retrospectively to investigate APP epidemiology in the area. A total of 572 strains isolated from outbreaks occurring in 337 different swine farms were analysed. The majority of isolates belonged to serotypes 9/11 (39.2%) and 2 (28.1%) and serotype diversity increased during the study period, up to nine different serotypes isolated in 2022. The most common resistances were against tetracycline (53% of isolates) and ampicillin (33%), followed by enrofloxacin, florfenicol and trimethoprim/sulfamethoxazole (23% each). Multidrug resistance (MDR) was common, with a third of isolates showing resistance to more than three antimicrobial classes. Resistance to the different classes and MDR varied significantly depending on the serotype. In particular, the widespread serotype 9/11 was strongly associated with florfenicol and enrofloxacin resistance and showed the highest proportion of MDR isolates. Serotype 5, although less common, showed instead a concerning proportion of trimethoprim/sulfamethoxazole resistance. Our results highlight how the typing of circulating serotypes and the analysis of their antimicrobial susceptibility profile are crucial to effectively manage APP infection and improve antimicrobial stewardship.

Photo-Controlled Reversible Uptake and Release of a Modified Sulfamethoxazole Antibiotic Drug from a Pillar[5]arene Cross-Linked Gelatin Hydrogel.

Pillararene cross-linked gelatin hydrogels were designed and synthesized to control the uptake and release of antibiotics using light. A suite of characterization techniques ranging from spectroscopy (FT-IR, 1H and 13C NMR, and MAS NMR), X-ray crystallographic analysis, scanning electron microscopy (SEM), and thermogravimetric analysis (TGA) was employed to investigate the physicochemical properties of hydrogels. The azobenzene-modified sulfamethoxazole (Azo-SMX) antibiotic was noncovalently incorporated into the hydrogel via supramolecular host-guest interactions to afford the A-hydrogel. While in its ground state, the Azo-SMX guest has a trans configuration structure and forms a thermodynamically stable inclusion complex with the pillar[5]arene motif in the hydrogel matrix. When the A-hydrogel was exposed to 365 nm UV light, Azo-SMX underwent a photoisomerization reaction. This changed the structure of Azo-SMX from trans to cis, and the material was released into the environment. The Azo-SMX released from the hydrogel was effective against both Gram-positive and Gram-negative bacteria. Importantly, the A-hydrogel exhibited a striking difference in antibacterial activity when applied to bacterial colonies in the presence and absence of UV light, highlighting the switchable antibacterial activity of A-hydrogel aided by light. In addition, all hydrogels containing pillar[5]arenes have demonstrated biocompatibility and effectiveness as scaffolds for biological and medical purposes.

The fate of sulfamethoxazole and trimethoprim in a micro-aerated anaerobic membrane bioreactor and the occurrence of antibiotic resistance in the permeate.

This study investigates the effects, conversions, and resistance induction, following the addition of 150 µg·L-1 of two antibiotics, sulfamethoxazole (SMX) and trimethoprim (TMP), in a laboratory-scale micro-aerated anaerobic membrane bioreactor (MA-AnMBR). TMP and SMX were removed at 97 and 86%, indicating that micro-aeration did not hamper their removal. These antibiotics only affected the pH and biogas composition of the process, with a significant change in pH from 7.8 to 7.5, and a decrease in biogas methane content from 84 to 78%. TMP was rapidly adsorbed onto the sludge and subsequently degraded during the long solids retention time of 27 days. SMX adsorption was minimal, but the applied hydraulic retention time of 2.6 days was sufficiently long to biodegrade SMX. The levels of three antibiotic-resistant genes (ARGs) (sul1, sul2, and dfrA1) and one mobile genetic element biomarker (intI1) were analyzed by qPCR. Additions of the antibiotics increased the relative abundances of all ARGs and intI1 in the MA-AnMBR sludge, with the sul2 gene folding 15 times after 310 days of operation. The MA-AnMBR was able to reduce the concentration of antibiotic-resistant bacteria (ARB) in the permeate by 3 log.

Four antibiotics and copper interactive effects on the growth and physiological characteristics of Hydrilla verticillata (L.f.) Royle.

Co-pollution of antibiotics and heavy metal copper (Cu) is common in freshwater environments because of their wide use as antimicrobial agents, especially in aquaculture. However, the toxic effects of coexisting antibiotics and heavy metals on aquatic plants remain unclear. This study investigated the effect of four antibiotics (i.e., enrofloxacin, ENR; tetracycline, TC; sulfamethoxazole, SMX; erythromycin, ERY), Cu, and their mixture on the growth and physiological responses of Hydrilla verticillata (L.f.) Royle. Results showed that the four antibiotics exhibited toxic effects on the growth and physiological indicators of H. verticillata, and root elongation was the most sensitive endpoint of the phytotoxicity test. The median effect concentration (EC50) of root elongation indicated that TC (EC50 = 10.05 mg/L) has the highest level of growth toxicity, and the toxicity of ENR to aquatic plants was close to TC (EC50 = 10.44 mg/L), followed by SMX (EC50 = 20.08 mg/L). However, there was no significant toxic effect of 20 mg/L ERY on the root elongation. Hydrophobicity may be a key factor affecting the phytotoxicity of antibiotics. Moreover, antagonistic toxic effects were observed under ENR + Cu, TC + Cu, SMX + Cu, and ERY + Cu co-exposures at all the experimental concentrations (0.01-20 mg/L). Due to the concentrations of antibiotics in natural waters usually with ng/L levels, our results suggested that environmental antibiotic concentrations probably pose low ecological risk to aquatic plants and indicated the H. verticillata could be used as phytoremediation candidate to remove antibiotic or antibiotic-Cu pollutions in general nature water.

Responses of microbial interactions and functional genes to sulfamethoxazole in anammox consortia.

Sulfamethoxazole (SMX) has been widely detected in various environments and its potential environmental risks have caused great concerns. However, the impact mechanism of SMX on microbial interactions among anammox consortia remain unknown. A long-term exposure experiments (140 d) was carried out to systematically examine the influence of SMX (0-1000 µg/L) on the anammox system, especially microbial network dynamics and variations of key metabolic genes. Results showed that anammox system could adapt to SMX below 500 µg/L and maintain a high nitrogen removal efficiency (NRE) of 85.35 ± 2.42%, while 1000 µg/L SMX significantly decreased the abundance of functional microbes and deteriorated denitrification performance with NRE dropped to 36.92 ± 15.01%. Co-occurrence network analysis indicated that 1000 µg/L SMX decreased the interactions between Proteobacteria and Chloroflexi and limited AnAOB from playing an important role as central nodes in the subnetwork of Planctomycetes. Metagenomics analysis found that genes associated with nitrogen removal (i.e., hdh, hzs, nirS, and hao) showed lower expression level after addition of SMX, while SMX-related ARGs (sul1 and sul2) increased by 1.22 and 2.68 times. This study provided us a relatively comprehensive perspective in response of microbial interactions and metabolic activity to various SMX concentrations.

Microbial community and gene dynamics response to high concentrations of gadolinium and sulfamethoxazole in biological nitrogen removal system.

The impact of high antibiotic and heavy metal pollution levels on biological nitrogen removal in wastewater treatment plants (WWTPs) remains poorly understood, posing a global concern regarding the issue spread of antibiotic resistance induced by these contaminants. Herein, we investigated the effects of gadolinium (Gd) and sulfamethoxazole (SMX), commonly found in medical wastewater, on biological nitrogen removal systems and microbial characteristics, and the fate of antibiotic resistance genes (ARGs), metal resistance genes (MRGs), and mobile genetic elements (MGEs). Our findings indicated that high SMX and Gd(III) concentrations adversely affected nitrification and denitrification, with Gd(III) exerting a strong inhibitory effect on microbial activity. Metagenomic analysis revealed that high SMX and Gd(III) concentrations could reduce microbial diversity, with Thauera and Pseudomonas emerging as dominant genera across all samples. While the relative abundance of most ARGs decreased under single Gd(III) stress, MRGs increased, and nitrification functional genes were inhibited. Conversely, combined SMX and Gd(III) pollution increased the relative abundance of intl1. Correlation analysis revealed that most genera could host ARGs and MRGs, indicating co-selection and competition between these resistance genes. However, most denitrifying functional genes exhibited a positive correlation with MRGs. Overall, our study provides novel insights into the impact of high concentrations of antibiotics and heavy metal pollution in WWTPs, and laying the groundwork for the spread and proliferation of resistance genes under combined SMX and Gd pollution.

Limited Polymorphism in the Dihydrofolate Reductase (dhfr) and dihydropteroate synthase genes (dhps) of Plasmodium knowlesi isolate from Thailand.

BACKGROUND: The 2022 malaria WHO reported around 4000 P. knowlesi infections in the South-East Asia region. In the same period, 72 positive cases were reported by the Department of Disease Control in Thailand, suggesting a persistent infection. Little is known about dihydrofolate reductase (pkdhfr) and dihydropteroate synthase (pkdhps), putative antimalarial resistance markers for P. knowlesi. The relevant amplification and sequencing protocol are presently unavailable. In this study, we developed a protocol for amplifying and evaluating pkdhps mutations. The haplotype pattern of pkdhfr-pkdhps in Thai isolates was analyzed, and the effects of these pkdhps mutations were predicted by using a computer program. METHODS: Pkdhps were amplified and sequenced from 28 P. knowlesi samples collected in 2008 and 2020 from nine provinces across Thailand. Combining pkdhfr sequencing data from previous work with pkdhps data to analyze polymorphisms of pkdhfr and pkdhps haplotype. Protein modeling and molecular docking were constructed using two inhibitors, sulfadoxine and sulfamethoxazole, and further details were obtained through analyses of protein-ligand interactions by using the Genetic Optimisation for Ligand Docking program. A phylogenetic tree cluster analysis was reconstructed to compare the P. knowlesi Malaysia isolates. RESULTS: Five nonsynonymous mutations in the pkdhps were detected outside the equivalence of the binding pocket sites to sulfadoxine and sulfamethoxazole, which are at N391S, E421G, I425R, A449S, and N517S. Based on the modeling and molecular docking analyses, the N391S and N517S mutations located close to the enzyme-binding pocket demonstrated a different docking score and protein-ligand interaction in loop 2 of the enzyme. These findings indicated that it was less likely to induce drug resistance. Of the four haplotypes of pkdhfr-pkdhps, the most common one is the R34L pkdhfr mutation and the pkdhps quadruple mutation (GRSS) at E421G, I425R, A449S, and N517S, which were observed in P. knowlesi in southern Thailand (53.57%). Based on the results of neighbor-joining analysis for pkdhfr and pkdhps, the samples isolated from eastern Thailand displayed a close relationship with Cambodia isolates, while southern Thailand isolates showed a long branch separated from the Malaysian isolates. CONCLUSIONS: A new PCR protocol amplification and evaluation of dihydropteroate synthase mutations in Knowlesi (pkdhps) has been developed. The most prevalent pkdhfr-pkdhps haplotypes (53.57%) in southern Thailand are R34L pkdhfr mutation and pkdhps quadruple mutation. Further investigation requires additional phenotypic data from clinical isolates, transgenic lines expressing mutant alleles, or recombinant proteins.

Supplementation of exogenous phytohormones for enhancing the removal of sulfamethoxazole and the simultaneous accumulation of lipid by Chlorella vulgaris.

In this study, the phytohormone gibberellins (GAs) were used to enhance sulfamethoxazole (SMX) removal and lipid accumulation in the microalgae Chlorella vulgaris. At the concentration of 50 mg/L GAs, the SMX removal achieved by C. vulgaris was 91.8 % while the lipid productivity of microalga was at 11.05 mg/L d-1, which were much higher than that without GAs (3.5 % for SMX removal and 0.52 mg/L d-1 for lipid productivity). Supplementation of GAs enhanced the expression of antioxidase-related genes in C. vulgaris as a direct response towards the toxicity of SMX. In addition, GAs increased lipid production of C. vulgaris by up-regulating the expression of genes related to carbon cycle of microalgal cells. In summary, exogenous GAs promoted the stress tolerance and lipid accumulation of microalgae at the same time, which is conducive to improving the economic benefits of microalgae-based antibiotics removal as well as biofuel production potential.

Microplastic-contaminated antibiotics as an emerging threat to mammalian liver: enhanced oxidative and inflammatory damages.

Poor management and disposal of plastic materials and the accumulation of microplatics in the environment and foods are an issue of increasing public concern. The current understanding of the implications of microplastics for human health has been limited to the bioeffect of individual exposure. In the bigger view of microplastic contamination, however, toxic compounds, including antibiotics, harbored on active microplastics can be collectively transported through food chains, raising questions about the effect of their combined exposure on human health. By employing a mouse model for human physiology, we discovered that a concurrent exposure to the major types of antibiotics and microplastics, namely sulfamethoxazole (SMZ) and polystyrene microplastics, respectively, would result in evident accumulation in detoxification organs; specifically, liver could amass 41.70 µg kg-1 of SMZ, while 3.83% of microplastics was accumulated in the kidney. Insights into the occurrence of liver histopathological changes (e.g., amyloidosis and necrocytosis) revealed that compared with the individual treatment of SMZ, treatment by microplastic-contaminated SMZ elicited increases in the levels of malonaldehyde and NF-κß by 174% and 104%, respectively; while the activities of antioxidases investigated were depressed by up to 22% upon co-exposure. It is suggested that SMZ enriched on active microplastic surfaces causes enhanced hepatic damage. Profiling of the gene expression clarified the correlation of the exacerbated oxidative and inflammatory damages in the liver with the overexpression of Nrf2 to dysregulate the Keap1-Nrf2 pathway. This study acts as a reminder about the complexity of contamination and raises awareness of health issues that microplastics could cause public health through liver diseases.

Anti-Balamuthia mandrillaris and anti-Naegleria fowleri effects of drugs conjugated with various nanostructures.

Balamuthia mandrillaris and Naegleria fowleri are protist pathogens that can cause fatal infections. Despite mortality rate of > 90%, there is no effective therapy. Treatment remains problematic involving repurposed drugs, e.g., azoles, amphotericin B and miltefosine but requires early diagnosis. In addition to drug discovery, modifying existing drugs using nanotechnology offers promise in the development of therapeutic interventions against these parasitic infections. Herein, various drugs conjugated with nanoparticles were developed and evaluated for their antiprotozoal activities. Characterizations of the drugs' formulations were accomplished utilizing Fourier-transform infrared spectroscopy, efficiency of drug entrapment, polydispersity index, zeta potential, size, and surface morphology. The nanoconjugates were tested against human cells to determine their toxicity in vitro. The majority of drug nanoconjugates exhibited amoebicidal effects against B. mandrillaris and N. fowleri. Amphotericin B-, Sulfamethoxazole-, Metronidazole-based nanoconjugates are of interest since they exhibited significant amoebicidal effects against both parasites (p < 0.05). Furthermore, Sulfamethoxazole and Naproxen significantly diminished host cell death caused by B. mandrillaris by up to 70% (p < 0.05), while Amphotericin B-, Sulfamethoxazole-, Metronidazole-based drug nanoconjugates showed the highest reduction in host cell death caused by N. fowleri by up to 80%. When tested alone, all of the drug nanoconjugates tested in this study showed limited toxic effects against human cells in vitro (less than 20%). Although these are promising findings, prospective work is warranted to comprehend the mechanistic details of nanoconjugates versus amoebae as well as their in vivo testing, to develop antimicrobials against the devastating infections caused by these parasites.

Sulfamethoxazole impact on pollutant removal and microbial community of aerobic granular sludge with filamentous bacteria.

In this study, sulfamethoxazole (SMX) was employed to investigate its impact on the process of aerobic granule sludge with filamentous bacteria (FAGS). FAGS has shown great tolerance ability. FAGS in a continuous flow reactor (CFR) could keep stable with 2 µg/L of SMX addition during long-term operation. The NH4+, chemical oxygen demand (COD), and SMX removal efficiencies kept higher than 80%, 85%, and 80%, respectively. Both adsorption and biodegradation play important roles in SMX removal for FAGS. The extracellular polymeric substances (EPS) might play important role in SMX removal and FAGS tolerance to SMX. The EPS content increased from 157.84 mg/g VSS to 328.22 mg/g VSS with SMX addition. SMX has slightly affected on microorganism community. A high abundance of Rhodobacter, Gemmobacter, and Sphaerotilus of FAGS may positively correlate to SMX. The SMX addition has led to the increase in the abundance of the four sulfonamide resistance genes in FAGS.

Antibacterial, antifungal, and anti-biofilm effects of sulfamethoxazole-complexes against pulmonary infection agents.

Antibiotic resistance associated with pulmonary infection agents has become a public health problem, being considered one of the main priorities for immediate resolution. Thus, to increase the therapeutic options in the fight against resistant microorganisms, the synthesis of molecules from pre-existing drugs has shown to be a promising alternative. In this sense, the present work reports the synthesis, characterization, and biological evaluation (against fungal and bacterial agents that cause lung infections) of potential metallodrugs based on sulfamethoxazole complexed with AuI, AgI, HgII, CdII, NiII, and CuII. The minimal inhibitory concentration (MIC) value was used to evaluate the antifungal and antibacterial properties of the compounds. In addition, it was also evaluated the antibiofilm capacity in Pseudomonas aeruginosa, through the quantification of its biomass and visualization using atomic force microscopy. For each case, molecular docking calculations were carried out to suggest the possible biological target of the assayed inorganic complexes. Our results indicated that the novel inorganic complexes are better antibacterial and antifungal than the commercial antibiotic sulfamethoxazole, highlighting the AgI-complex, which was able to inhibit the growth of microorganisms that cause lung diseases with concentrations in the 2-8 µg mL-1 range, probably at targeting dihydropteroate synthetase - a key enzyme involved in the folate synthesis. Furthermore, sulfamethoxazole complexes were able to inhibit the formation of bacterial biofilms at significantly lower concentrations than free sulfamethoxazole, probably mainly targeting the active site of LysR-type transcriptional regulator (PqsR). Overall, the present study reports preliminary results that demonstrate the derivatization of sulfamethoxazole with transition metal cations to obtain potential metallodrugs with applications as antimicrobial and antifungal against pulmonary infections, being an alternative for drug-resistant strains.

Insight into sulfamethoxazole effects on aerobic denitrification by strain Pseudomonas aeruginosa PCN-2: From simultaneous degradation performance to transcriptome analysis.

It is a well-established fact that aerobic denitrifying strains are profoundly affected by antibiotics, but bacterium performing simultaneous aerobic denitrification and antibiotic degradation is hardly reported. Here, a typical aerobic denitrifying bacterium Pseudomonas aeruginosa PCN-2 was discovered to be capable of sulfamethoxazole (SMX) degradation. The results showed that nitrate removal efficiency was decreased from 100% to 88.12%, but the resistance of strain PCN-2 to SMX stress was enhanced with the increment of SMX concentration from 0 to 100 mg/L. Transcriptome analysis revealed that the down-regulation of energy metabolism pathways rather than the denitrifying functional genes was responsible for the suppressed nitrogen removal, while the up-regulation of antibiotic resistance pathways (e.g., biofilm formation, multi-drug efflux system, and quorum sensing) ensured the survival of bacterium and the carrying out of aerobic denitrification. Intriguingly, strain PCN-2 could degrade SMX during aerobic denitrification. Seven metabolites were identified by the UHPLC-MS, and three degradation pathways (which includes a new pathway that has never been reported) was proposed combined with the expressions of drug metabolic genes (e.g., cytP450, FMN, ALDH and NAT). This work provides a mechanistic understanding of the metabolic adaption of strain PCN-2 under SMX stress, which provided a broader idea for the treatment of SMX-containing wastewater.

Host-dependent resistance of Group A Streptococcus to sulfamethoxazole mediated by a horizontally-acquired reduced folate transporter.

Described antimicrobial resistance mechanisms enable bacteria to avoid the direct effects of antibiotics and can be monitored by in vitro susceptibility testing and genetic methods. Here we describe a mechanism of sulfamethoxazole resistance that requires a host metabolite for activity. Using a combination of in vitro evolution and metabolic rescue experiments, we identify an energy-coupling factor (ECF) transporter S component gene (thfT) that enables Group A Streptococcus to acquire extracellular reduced folate compounds. ThfT likely expands the substrate specificity of an endogenous ECF transporter to acquire reduced folate compounds directly from the host, thereby bypassing the inhibition of folate biosynthesis by sulfamethoxazole. As such, ThfT is a functional equivalent of eukaryotic folate uptake pathways that confers very high levels of resistance to sulfamethoxazole, yet remains undetectable when Group A Streptococcus is grown in the absence of reduced folates. Our study highlights the need to understand how antibiotic susceptibility of pathogens might function during infections to identify additional mechanisms of resistance and reduce ineffective antibiotic use and treatment failures, which in turn further contribute to the spread of antimicrobial resistance genes amongst bacterial pathogens.

[Historical Antibiotic Stress Changed the Effects of Sulfamethoxazole and Trimethoprim on Activated Sludge: ARGs and Potential Hosts].

The co-exposure of antibiotics has important effects on antibiotic resistance genes (ARGs) and microbial community aggregation in wastewater treatment plants (WWTPs). However, it is unclear whether differences in historical antibiotic exposure stress can determine responses of microbes and ARGs to combined antibiotics. By selecting a high concentration (30 mg·L-1) of sulfamethoxazole (SMX) and trimethoprim (TMP) as historical exposure stress conditions, the effects of SMX and TMP-combined pollution on ARGs, bacterial communities, and their interactions were explored in short-term experiments. Based on high-throughput quantitative PCR, a total of 13 ARGs were detected, and the absolute abundance was 2.21-5.42 copies·µL-1 (logarithm, DNA, the same below). Among them, sul2, ermB, mefA, and tetM-01 were the main subtypes in the samples, and the absolute abundance was between 2.95 and 5.40 copies·µL-1. The combined contamination of SMX and TMP could cause the enrichment of ARGs and mobile genetic elements (MGEs); however, their effects on each subtype were different, and the historical legacy effect of SMX was higher than that of TMP. Under the different exposure histories, the co-occurrence and co-exclusion patterns existed between ARGs. Moreover, MGEs (especially intI-1) were significantly correlated with sulfonamides (sul1 and sul2), tetracyclines[tet(32)], and macrolide-lincosamide-streptogramin (MLSB) resistance genes (ermB). Based on the full-scale classification of microorganisms, it was found that the microbial community structure of various groups responded differently to combined pollution, and the conditionally abundant taxa (CAT) were obviously enriched. Thauera, Pseudoxanthomonas, and Paracoccus were the dominant resistant bacterial genera. Furthermore, a total of 31 potential hosts of ARGs were identified with network analysis, which were dominated with conditionally rare taxa (CRT). Particularly, Candidatus_Alysiosphaera and Fusibacter were positively correlated with most of the ARGs, being the common protentional hosts. Importantly, some rare genera (RT, Variibacter, Aeromonas, Cloacibacterium, etc.) were potential hosts of transposon IS613, which played an important role in the proliferation and spread of ARGs. In conclusion, this study revealed the legacy effects of historical antibiotic stress on ARGs and their hosts, which could provide new ideas and theoretical basis for reducing ARGs pollution in WWTPs.

Genomic Characterization of Antimicrobial-Resistant Salmonella enterica in Duck, Chicken, and Pig Farms and Retail Markets in Eastern China.

Antimicrobial-resistant Salmonella enterica poses a significant public health concern worldwide. However, the dissemination of Salmonella enterica among food animals in eastern China has not been fully addressed. Here, we demonstrated the antimicrobial resistance (AMR) patterns and the whole-genome characterization of 105 S. enterica isolates from 1,480 fecal samples and anal swabs collected from 22 different farms (chickens, ducks, and pigs) and two live animal markets located in Zhejiang and Fujian Provinces in eastern China in 2019. The prevalence of isolates in duck farms (19.17%, 23/120) was statistically significantly higher (P < 0.001) than that in chicken farms (6.61%, 37/523) and pig farms (3.50%, 7/200). Among these isolates, 75.26% (79/105) were multidrug resistant, with the highest rates of resistance to tetracycline (76.20%) and ampicillin (67.62%) and the lowest resistance rate to meropenem (0.00%). The serotypes were consistent with sequence types and were closely related to the sampling animal species and sites. S. enterica serotype Kentucky (20.95%, 22/105) was the most frequent serotype and harbored more AMR patterns and genes than others. Furthermore, IncFII(S) and IncHI2 were the most prevalent replicons. A total of 44 acquired AMR genes were found. Among those genes, aac(6')-Iaa, blaTEM-1B, floR, dfrA14, fosA7, mph(A), qnrS1, sul1, tet(A), and ARR-3 were the dominant AMR genes mediating the AMR toward aminoglycosides, ß-lactams, phenicol, trimethoprim, fosfomycin, macrolide, quinolone, sulfonamides, tetracycline, and rifampin, respectively. The consistency of acquired AMR genes with AMR phenotypes for ampicillin, ceftiofur, ceftazidime, meropenem, sulfamethoxazole-trimethoprim, and tetracycline was >90%. Together, our study highlights the application of whole-genome sequencing to assess veterinary public health threats. IMPORTANCE Public health is a significant concern in China, and the foodborne pathogen Salmonella, which is spread via the animal-borne food chain, plays an important role in the overall disease burden in China annually. The development of advanced sequencing technologies has introduced a new way of understanding emerging pathogens. However, the routine surveillance application of this method in China remains in its infancy. Here, we applied a pool of all isolates from the prevalence data in Zhejiang and Fujian for whole-genome sequencing and combined these data with the cutting-edge bioinformatic analysis pipeline for one-step determination of the complete genetic makeup for all 105 genomes. The illustrated method could provide a cost-effective approach, without labor-intensive laboratory characterization, for predicting serotypes, genotypes, plasmid types, antimicrobial resistance genes, and virulence genes, and thus would provide essential knowledge for emerging pathogens. Our findings and perspectives are essential for delivering updated knowledge on foodborne pathogens in an understudied region in China.

Association between prenatal antibiotics exposure and measures of fetal growth: A repeated-measure study.

The abuse of antibiotics in animal husbandry has brought many public health problems, among which the passive use of antibiotics caused by eating food containing residual antibiotics has attracted the most attention. However, few studies have examined the possible adverse effects of prenatal antibiotics exposure on fetal growth and development. In this study, we investigated the associations between prenatal antibiotics exposure and measures of fetal growth. A total of 429 mother-newborn pairs from a birth cohort were enrolled and spot urine samples (N = 1287) were collected during each trimester of pregnancy. Sixteen antibiotics from 7 categories, were selected for the determination of the targeted antibiotics in maternal urines by UHPLC-MS/MS. Fetal growth indicators including newborn birth weight, birth length and gestational age (GA), were obtained from medical record. Sixteen antibiotics were found in 92.3% of the urine samples with detection frequencies ranging from 0.3% to 41.3%. Among the 16 antibiotics detected, we found that the exposure level of ciprofloxacin in the first trimester of pregnancy was negatively correlated with GA (ß = -0.17 day, 95% CI, -0.32 to -0.02 day), which would increase the risk of preterm birth (OR=1.05, 95% CI, 1.00, 1.09). The exposure level of norfloxacin in the second trimester of pregnancy was negatively correlated with fetal birth weight (ß = -17.56 g, 95% CI, -31.13 to -3.99 g) and birth length (ß = -0.05 cm, 95% CI, -0.08 to -0.02 cm), and the exposure level of sulfamethoxazole in the third trimester of pregnancy was negatively correlated with fetal birth length (ß = -0.15 cm, 95% CI, -0.29 to -0.02 cm). Our findings suggest that prenatal exposure to norfloxacin and sulfamethoxazole may adversely affect fetal growth and development.

Analysis of virulence genes, drug resistance detection, and pathogenicity in Enterococcus from farm animals.

This study aimed to investigate the presence of eight virulence genes (ace, asa1, esp, efaA, gelE, cylA, agg, fsr) in Enterococcus from a variety of animals and to explore the drug resistance and pathogenicity. This could provide a theoretical basis for clinical treatment of Enterococcus infections. Anal swabs from pigs, chickens, cattle, and dogs in farms and pet hospitals were collected for Enterococcus isolation and identification. Eight virulence genes were detected (PCR method), and drug resistance was assessed (drug-sensitive paper method). The strains containing different virulence genes were then divided into EV1, EV2, and EV3 groups. The LD50 and pathogenicity was examined by intra-peritoneal injection to infect mice. Differences were found in the detection rates of virulence genes in Enterococcus from the different animals. The highest overall detection rate was for the esp gene (78.0%), and the lowest for the cylA gene (15.5%). Eight genes were detected most frequently in Enterococcus from dogs and least frequently from cattle. Among the Enterococcus strains from four variety of animals, drug resistance was highest against sulfamethoxazole (100%), cefotaxime (>97%), and cefotaxitin (>93%). Drug resistance was lowest against vancomycin (0%), levofloxacin (<12%) and ciprofloxacin (<13%). The LD50 for each of the three groups was EV1LD50=8.71×109CFU， EV2LD50=2.34×1010CFU，and EV3LD50=9.33×1010CFU. The Enterococcus12LD50 dose group caused significant clinical symptoms in mice, with pathological effects on the heart, liver, lungs, and kidneys, and particularly on the urinary system. The abundance of Enterococcus virulence genes, drug resistance, and pathogenicity vary among different animal origins, and the pathology caused by Enterococcus requires effective treatment protocols based on species and regional characteristics.

Antimicrobial susceptibility and minimum inhibitory concentration distribution of common clinically relevant non-tuberculous mycobacterial isolates from the respiratory tract.

Objective: To determine the minimum inhibitory concentration (MIC) distribution of antibacterial drugs and the susceptibility of non-tuberculous mycobacterial (NTM) isolates to provide a reference basis for the clinical selection of an effective starting regimen.Methods: The common clinical isolates of NTM in the respiratory tract, which met the standards of the American Thoracic Society for NTM lung disease, were collected. The MICs of 81 isolates were determined using the microbroth dilution method (Thermo Fisher Scientific, USA), as recommended by the Clinical and Laboratory Standards Institute, USA.Results: Included were 43 Mycobacterium avium complex (MAC) strains, 24 M. abscessus complex (MAB) strains, and 14 M. kansasii strains. The sensitivity rates of MAC to clarithromycin and amikacin were 81.4% and 79.1%, respectively, while the sensitivity rates to linezolid and moxifloxacin were only 20.9% and 9.3%; the MIC of rifabutin was the lowest (MIC50% was just 2 µg/mL). After incubation for 3-5 days, the sensitivity rate of MAB to clarithromycin was 87.5%; this decreased to 50% after 14 days' incubation. Most of them were susceptible to amikacin (91.6%), and most were resistant to moxifloxacin (95.8%), ciprofloxacin (95.8%), imipenem (95.8%), amoxicillin/clavulanate (95.8%), tobramycin (79.1%), doxycycline (95.8%) and trimethoprim/sulfamethoxazole (95.8%). intermediate (83.3%) and resistant (16.7%) to cefoxitin. The susceptibility to linezolid was only 33.3%. The sensitivity and resistance breakpoints of tigecycline were set to ≤0.5 and ≥8 µg/mL, respectively, and the sensitivity and resistance rates were 50% and 0%, respectively. M. kansasii was susceptible to clarithromycin, amikacin, linezolid, moxifloxacin, rifampicin and rifabutin (100%).Discussion: In Wenzhou, clarithromycin, amikacin and rifabutin have good antibacterial activity against MAC, while linezolid and moxifloxacin have high resistance. Amikacin and tigecycline have strong antibacterial activity against MAB, while most other antibacterial drugs are resistant to varying degrees. Most antibacterial drugs are susceptible to M. kansasii and have good antibacterial activity.Conclusion: The identification of NTM species and the detection of their MICs have certain guiding values for the treatment of NTM lung disease.Key MessageThe three most common respiratory non-tuberculous mycobacterial (NTM) isolates with clinical significance in the Wenzhou area were tested for drug susceptibility. The broth microdilution method was used to determine the minimum inhibitory concentration distribution of antibacterial drugs and the susceptibility of NTM isolates to provide a reference basis for the clinical selection of an effective starting regimen.

Dissecting the roles of conductive materials in attenuating antibiotic resistance genes: Evolution of physiological features and bacterial community.

Supplying conductive materials (CMs) into anaerobic bioreactors is considered as a promising technology for antibiotic wastewater treatment. However, whether and how CMs influence antibiotic resistance genes (ARGs) spread remains poorly known. Here, we investigated the effects of three CMs, i.e., magnetite, activated carbon (AC), and zero valent iron (ZVI), on ARGs dissemination during treating sulfamethoxazole wastewater, by dissecting the shifts of physiological features and microbial community. With the addition of magnetite, AC, and ZVI, the SMX removal was improved from 49.05 to 71.56-92.27 %, while the absolute abundance of ARGs reducing by 26.48 %, 61.95 %, 48.45 %, respectively. The reduced mobile genetic elements and antibiotic resistant bacteria suggested the inhibition of horizontal and vertical transfer of ARGs. The physiological features, including oxidative stress response, quorum sensing, and secretion system may regulate horizontal transfer of ARGs. The addition of all CMs relieved oxidative stress compared with no CMs, but ZVI may cause additional free radicals that needs to be concerned. Further, ZVI and AC also interfered with cell communication and secretion system. This research deepens the insights about the underlying mechanisms of CMs in regulating ARGs, and is expected to propose practical ways for mitigating ARGs proliferation.