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Antonio Carlos Maneira Godinho Netto,1,2 Túlio Gonçalves dos Reis,1,2 Cássia Franco Matheus,1,2 Beatriz Julião Vieira Aarestrup,3,4 Fernando Monteiro Aarestrup1,2,4 1School of Medical and Health Sciences – SUPREMA, 2Maternity Hospital Terezinha de Jesus, 3Morphology Department, Federal University of Juiz de Fora, Institute of Biological Sciences, 4Laboratory of Immunopathology and Experimental Pathology, Federal University of Juiz de Fora, Reproductive Biology Center (CBR), Juiz de Fora, Brazil Objective: To assess the value of fraction of exhaled nitric oxide (FeNO) measurements in the diagnosis of asthma in elderly patients. Methods: The clinical symptoms of 202 elderly patients were assessed with the asthma module of the International Study of Asthma and Allergies in Childhood test, which had been modified for the elderly patients, and the diagnostic routine for chronic obstructive pulmonary disease (COPD), which was based on the Global initiative for chronic Obstructive Lung Disease criteria. Of the 202 patients assessed, 43 were subjected to pulmonary function evaluations (spirometry) and FeNO measurements. Results: Of the 202 elderly patients, 34 had asthma (23 definite and eleven probable), 20 met COPD criteria, 13 presented with an overlap of asthma and COPD, and 135 did not fit the criteria for obstructive pulmonary disease. Among the 43 elderly patients who were subjected to FeNO measurements, ten showed altered results (23.2%) and 33 had normal results (76.7%). The average value of FeNO in patients with definite and probable asthma undergoing this procedure was 29.2 parts per billion whereas that in nonasthmatic patients was 17.5 parts per billion (P=0.0002). Conclusion: We show a clear relationship between FeNO levels and asthma symptoms and previous asthma diagnoses in elderly patients. Keywords: asthma, chronic obstructive pulmonary disease, elderly patients, nitric oxide

Objectives: Identification of antimicrobial resistance genes is important for understanding the underlying mechanisms and the epidemiology of antimicrobial resistance. As the costs of whole-genome sequencing (WGS) continue to decline, it becomes increasingly available in routine diagnostic laboratories and is anticipated to substitute traditional methods for resistance gene identification. Thus, the current challenge is to extract the relevant information from the large amount of generated data.Methods: We developed a web-based method, ResFinder that uses BLAST for identification of acquired antimicrobial resistance genes in whole-genome data. As input, the method can use both pre-assembled, complete or partial genomes, and short sequence reads from four different sequencing platforms. The method was evaluated on 1862 GenBank files containing 1411 different resistance genes, as well as on 23 de-novo-sequenced isolates.Results: When testing the 1862 GenBank files, the method identified the resistance genes with an ID = 100% (100% identity) to the genes in ResFinder. Agreement between in silico predictions and phenotypic testing was found when the method was further tested on 23 isolates of five different bacterial species, with available phenotypes. Furthermore, ResFinder was evaluated on WGS chromosomes and plasmids of 30 isolates. Seven of these isolates were annotated to have antimicrobial resistance, and in all cases, annotations were compatible with the ResFinder results.Conclusions: A web server providing a convenient way of identifying acquired antimicrobial resistance genes in completely sequenced isolates was created. ResFinder can be accessed at www.genomicepidemiology.org. ResFinder will continuously be updated as new resistance genes are identified. [ABSTRACT FROM AUTHOR]

The recent expansion of mpox in Africa is characterized by a dramatic increase in zoonotic transmission (clade Ia) and the emergence of a new clade Ib that is transmitted from human to human by close contact. Clade Ia does not pose a threat in areas without zoonotic reservoirs. But clade Ib may spread widely, as did clade IIb which has spread globally since 2022 among men who have sex with men. It is not clear whether controlling clade Ib will be more difficult than clade IIb. The population at risk potentially counts 100 million but only a million vaccine doses are expected in the next year. Surveillance is needed with exhaustive case detection, polymerase chain reaction confirmation, clade determination, and about severe illness. Such data is needed to identify routes of transmission and core transmitters, such as sex workers. Health care workers are vaccinated to ensure their protection, but this will not curb mpox transmission. With the recent inequitable distribution of COVID-19 vaccines in mind, it is a global responsibility to ensure that low-income nations in the mpox epicenter have meaningful access to vaccines. Vaccination serves not only to reduce mortality in children but limit the risk of future mpox variants emerging that may spread in human populations globally., Competing Interests: Declarations of competing interest The authors have no competing interests to declare., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

VanHAX -mediated glycopeptide resistance has been consistently high in one of the three main sewer systems in Copenhagen, Lynetten, for +20 years. To explore this for other glycopeptide resistance genes, and whether the colonization has resulted in establishment of multiple bacterial taxa, we mapped 505 shotgun metagenomic data sets from the inlet of three sewage treatment plants to 831 different glycopeptide resistance genes. Only vanHAX and vanHBX genes were differentially abundant in Lynetten. Analyses of eight contigs suggested limited variations in the flanking regions. Proximity ligation metagenomic analysis of 12 samples from Lynetten identified 441 and 5 paired reads mapping to vanHAX and vanHBX , respectively. The other end of these reads was mapped to generated metagenomic-assembled genomes and NCBI using BLAST. vanHBX could only be linked to the phylum level (Bacillota). Plasmid analysis of vanHBX Hi-C contigs showed that these were mainly located on plasmids reported found in enterococci species. Most vanHAX -linked reads could only be linked to phylum and class level, but some reads were assigned to Enterococcus faecium (7 reads), Enterococcus faecalis (4 reads), Paenibacillus apiarius (2 reads), and Paenibacillus thiaminolyticus (27 reads). Ten of the 20 Hi-C contigs-containing vanHAX were annotated as plasmid, all reported found in Enterococcus species. This study shows that while Hi-C technology is valuable for linking antimicrobial resistance genes to bacterial taxa, it suffers from challenges in reliably mapping the linked read to a genomic region with sufficient taxonomic information. Our results also suggest that over the +20 years of colonizing a sewer system, vanHAX has not become widespread across multiple taxa, remaining primarily in E. faecalis and E. faecium , with the exception of Paenibacillus .IMPORTANCELong-term colonization of microbial communities with antimicrobial-resistant bacteria is expected to result in sharing of the resistance genes between several different bacterial taxa of the communities. We investigated microbiomes from a sewer, which have been colonized with glycopeptide-resistant bacteria harboring the mobile vanHAX gene cluster for a minimum of 20 years, using metagenomics sequencing and Hi-C. We found that despite the long-term presence in the sewer, the vanHAX genes have seemingly not disseminated widely.

Staphyloccous aureus belonging to sequence type 5477 have recently been identified as a predominant clone causing bovine mastitis in Rwanda and Tanzania. We compared nine S. aureus ST5477 to 17 isolates belonging to other sequence types by their biochemical profile and ability to acidify milk and grow in minimum media containing lactose. We found that ST5477 isolates all were positive in ONPG (o-nitrophenyl-β-D-galactopyranoside) test and negative for mannitol fermentation potentially challenging the correct identification of this sequence type as S. aureus. In addition, ST5477 isolates were all much faster in acidifying milk and grew faster in minimal media with lactose compared to other strains suggesting an increased lactose utilization and thereby adaptation to the bovine udder environment as a possible reason for the recent successful emergence. Comparison of the lac gene region of the genome of a recently sequenced ST5477 and that of S. aureus reference genome showed that both strains contained the known lacABCD genes involved in the lactose degradation, but that ST5477 had a 12 amino-acid deletion and two amino-acid differences in the lac gene transcription regulator, suggesting that increased transcription might play a role. In conclusion, these preliminary data suggests that improved lactose utilization and the ability to grow faster in milk may have been a key feature for the recent success of ST5477 as a bovine adapted clone., (Copyright © 2024. Published by Elsevier GmbH.)

Salmonella Typhi (S. Typhi) is an important pathogen causing typhoid fever worldwide. The emergence of antibiotic resistance, including that of bla TEM genes encoding to TEM [Formula: see text]-lactamases has been observed. This study aimed to investigate the dynamics of bla TEM genes in S. Typhi by analyzing the phylogeny and flanking region patterns and phylogenetic associating them with metadata (year, country) and genomic data (genotypes, antibiotic resistance genes (ARGs), plasmids). Genomic sequences of publicly available S. Typhi harboring bla TEM (n = 6079), spanning from 1983 to 2023, were downloaded and analyzed using CSIPhylogeny for phylogeny, Flankophile for identifying genetic contexts around bla TEM genes and GenoTyphi for determining genotypes, ARGs and plasmid replicons. We found that bla TEM -positive isolates occurred most commonly in specific location, especially in Asia and Africa and clustered among a limited number of genotypes. Flankophile identified 740 isolates (12.2%) with distinct flanking region patterns, which were categorized into 13 patterns. Notably, 7 patterns showed a predominantly phylogenetic association with genotypes. Additionally, these 7 patterns exhibited relation to the country, ARGs and plasmid replicons. Further examination of the flanking region patterns provided association with mobile genetic elements (MGEs). Taken together, this study suggests that bla TEM has been acquired by S. Typhi isolates a limited number of times and subsequently spread clonally with specific genotypes., (© 2024. The Author(s).)

We describe cases with monkeypox virus (MPXV) Clade Ib in Burundi from their first detection in July until 20 August 2024. Testing 442 people with vesicular lesions confirmed 170 cases (98 male; 72 female), 82 (48%) being < 15 years old. Differential diagnosis of the first 30 individuals testing MPXV negative revealed chickenpox in 20. Cases occurred in 26 of 49 Burundi health districts, but mostly in Bujumbura Nord (88/170; 67%). Case-derived MPXV genetic sequences from Burundi and South-Kivu (Democratic Republic of the Congo), clustered together in phylogenetic analysis.

Since we published the "IV Brazilian Consensus on Rhinitis", in2017, several advances have been achieved and have enabled a further understanding of the different aspects of "Rhinitis". This new guideline, developed jointly by ASBAI, SBP and SBORL, represents a relevant milestone in the updated and integrated management of the different forms of the disease, and it aims to unify evidence-based approaches to improve the diagnosis and treatment of this common and often underestimated condition. The document covers a wide range of topics, including clear definitions of the different phenotypes and endotypes of rhinitis, risk factors, updated diagnostic criteria, and recommended methods for clinical and laboratory investigation. We stress the importance of detailed clinical history and objective assessment, as well as tools for control and assessing severity tools an accurate diagnostic approach to the disease. Regarding treatment, it emphasizes the treatment customization, considering the severity of symptoms, the presence of comorbidities and the impact on the patient's quality of life. We discuss different drug treatment, in addition to non-pharmacological measures, such as environmental control and specific immunotherapy; and the possible role of immunobiological agents. Furthermore, the consensus addresses issues related to patient education, prevention and management of special situations, such as rhinitis in children, in pregnant women and in the elderly. In short, the "V Brazilian Consensus on Rhinitis" represents a comprehensive and updated guide for healthcare professionals involved in the diagnosis and management of rhinitis, aiming to improve patients' quality of life through an integrated and evidence-based approach., (Copyright © 2024 Associação Brasileira de Otorrinolaringologia e Cirurgia Cérvico-Facial. Published by Elsevier España S.L.U. All rights reserved.)

Sewage metagenomics has risen to prominence in urban population surveillance of pathogens and antimicrobial resistance (AMR). Unknown species with similarity to known genomes cause database bias in reference-based metagenomics. To improve surveillance, we seek to recover sewage genomes and develop a quantification and correlation workflow for these genomes and AMR over time. We use longitudinal sewage sampling in seven treatment plants from five major European cities to explore the utility of catch-all sequencing of these population-level samples. Using metagenomic assembly methods, we recover 2332 metagenome-assembled genomes (MAGs) from prokaryotic species, 1334 of which were previously undescribed. These genomes account for ~69% of sequenced DNA and provide insight into sewage microbial dynamics. Rotterdam (Netherlands) and Copenhagen (Denmark) show strong seasonal microbial community shifts, while Bologna, Rome, (Italy) and Budapest (Hungary) have occasional blooms of Pseudomonas-dominated communities, accounting for up to ~95% of sample DNA. Seasonal shifts and blooms present challenges for effective sewage surveillance. We find that bacteria of known shared origin, like human gut microbiota, form communities, suggesting the potential for source-attributing novel species and their ARGs through network community analysis. This could significantly improve AMR tracking in urban environments., (© 2024. The Author(s).)

The use of deep learning models in computational biology has increased massively in recent years, and it is expected to continue with the current advances in the fields such as Natural Language Processing. These models, although able to draw complex relations between input and target, are also inclined to learn noisy deviations from the pool of data used during their development. In order to assess their performance on unseen data (their capacity to generalize ), it is common to split the available data randomly into development (train/validation) and test sets. This procedure, although standard, has been shown to produce dubious assessments of generalization due to the existing similarity between samples in the databases used. In this work, we present SpanSeq, a database partition method for machine learning that can scale to most biological sequences (genes, proteins and genomes) in order to avoid data leakage between sets. We also explore the effect of not restraining similarity between sets by reproducing the development of two state-of-the-art models on bioinformatics, not only confirming the consequences of randomly splitting databases on the model assessment, but expanding those repercussions to the model development. SpanSeq is available at https://github.com/genomicepidemiology/SpanSeq., (© The Author(s) 2024. Published by Oxford University Press on behalf of NAR Genomics and Bioinformatics.)

Rapid advancements in sequencing technologies have led to significant progress in microbial genomics, yet challenges persist in accurately identifying microbial strain diversity in metagenomic samples, especially when working with noisy long-read data from platforms like Oxford Nanopore Technologies (ONT). In this article, we introduce NanoMGT, a tool designed to enhance marker gene typing in low-complexity mono-species samples, leveraging the unique properties of long reads. NanoMGT excels in its ability to accurately identify mutations amidst high error rates, ensuring the reliable detection of multiple strain-specific marker genes. Our tool implements a novel scoring system that rewards mutations co-occurring across different reads and penalizes densely grouped, likely erroneous variants, thereby achieving a good balance between sensitivity and precision. A comparative evaluation of NanoMGT, using a simulated multi-strain sample of seven bacterial species, demonstrated superior performance relative to existing tools and the advantages of using a threshold-based filtering approach to calling minority variants in ONT's sequencing data. NanoMGT's potential as a post-binning tool in metagenomic pipelines is particularly notable, enabling researchers to more accurately determine specific alleles and understand strain diversity in microbial communities. Our findings have significant implications for clinical diagnostics, environmental microbiology, and the broader field of genomics. The findings offer a reliable and efficient approach to marker gene typing in complex metagenomic samples., (© The Author(s) 2024. Published by Oxford University Press.)

Monkeypox virus (MPXV) is an emerging zoonotic pathogen with complex epidemiology necessitating rapid diagnosis and distinguishing between clades and subclades. The emerging Clade Ib lacks the genomic region used in the Clade I-specific assay from the Centers for Disease Control and Prevention. We report an MPXV real-time PCR to specifically detect Clade Ib. The assay demonstrated proficient sensitivity and specificity in 92 samples and can be included along other TaqMan-based assays to detect MPXV and distinguish between clades and subclades.

We report the discovery of a persistent presence of Vibrio cholerae at very low abundance in the inlet of a single wastewater treatment plant in Copenhagen, Denmark at least since 2015. Remarkably, no environmental or locally transmitted clinical case of V. cholerae has been reported in Denmark for more than 100 years. We, however, have recovered a near-complete genome out of 115 metagenomic sewage samples taken over the past 8 years, despite the extremely low relative abundance of one V. cholerae read out of 500,000 sequenced reads. Due to the very low relative abundance, routine screening of the individual samples did not reveal V. cholerae. The recovered genome lacks the gene responsible for cholerae toxin production, but although this strain may not pose an immediate public health risk, our finding illustrates the importance, challenges, and effectiveness of wastewater-based pathogen surveillance., (© 2024. The Author(s).)

Members of Bacillota and Pseudomonadota phyla are frequently considered bacterial infectious agents in humans. As part of a large sequencing project of clinically relevant pathogens, we hybrid-assembled complete genomes of Cytobacillus , Domibacillus , Enterobacter , Neisseria , Pseudomonas, and Streptococcus species isolated from clinical specimens., Competing Interests: The authors declare no conflict of interest.

The rapid spread of antimicrobial resistance (AMR) is a threat to global health, and the nature of co-occurring antimicrobial resistance genes (ARGs) may cause collateral AMR effects once antimicrobial agents are used. Therefore, it is essential to identify which pairs of ARGs co-occur. Given the wealth of next-generation sequencing data available in public repositories, we have investigated the correlation between ARG abundances in a collection of 214,095 metagenomic data sets. Using more than 6.76∙10 8 read fragments aligned to acquired ARGs to infer pairwise correlation coefficients, we found that more ARGs correlated with each other in human and animal sampling origins than in soil and water environments. Furthermore, we argued that the correlations could serve as risk profiles of resistance co-occurring to critically important antimicrobials (CIAs). Using these profiles, we found evidence of several ARGs conferring resistance for CIAs being co-abundant, such as tetracycline ARGs correlating with most other forms of resistance. In conclusion, this study highlights the important ARG players indirectly involved in shaping the resistomes of various environments that can serve as monitoring targets in AMR surveillance programs., Importance: Understanding the collateral effects happening in a resistome can reveal previously unknown links between antimicrobial resistance genes (ARGs). Through the analysis of pairwise ARG abundances in 214K metagenomic samples, we observed that the co-abundance is highly dependent on the environmental context and argue that these correlations can be used to show the risk of co-selection occurring in different settings., Competing Interests: The authors declare no conflict of interest.

The use of allergen immunotherapy (AIT) in Brazil has specific regional conditions owing to the pattern of allergen sensitization, as well as to genetic, socioeconomic, and cultural characteristics. This review article aims to discuss the clinical practice of AIT by the subcutaneous or sublingual route in Brazil, addressing the possibilities of transition between these forms of administration. A systematic review using the PubMed and Cochrane databases was performed, and the websites of major allergy and immunology organizations were consulted. Knowledge of the mechanism of action of subcutaneous immunotherapy and sublingual immunotherapy, together with Brazilian real-life experience, allowed us to establish recommendations regarding switching routes of AIT administration in selected cases. Careful analysis of each clinical situation is necessary to perform the transition between subcutaneous and sublingual allergen immunotherapy., Competing Interests: Disclosure of potential conflict of interest: The authors declare that they have no relevant conflicts of interest., (© 2024 The Authors.)

Accurate monitoring of gastro-enteric and other diseases in large populations poses a challenge for public health management. Sewage represents a larger population, is freely obtainable and non-subject to ethical approval. Metagenomic sequencing offers simultaneous, multiple-target analysis. However, no study has demonstrated the sensitivity of metagenomics for detecting bacteria in sewage. In this study, we spot-released 1013 colony-forming units (CFU) of Staphyloccus hyicus (non-pathogenetic strain 842J-88). The strain was flushed down a toilet into the sewer in the catchment area of a public wastewater treatment plant (WWTP), serving a population of 36,000 people. Raw sewage was continuously sampled at the WWTP's inlet over 30- and 60-minute intervals for a total period of seven hours. The experiment was conducted twice with one week in-between release days and under comparable weather conditions. For the metagenomics analyses, the pure single isolate of S. hyicus was sequenced, assembled and added to a large database of bacterial reference sequences. All sewage samples were analyzed by shotgun metagenome sequencing and mapped against the reference database. S. hyicus was identified in duplicate samples at both of two release days and these sequence fragment counts served as a proxy to estimate the minimum number of sick people or sensitivity required in order to observe at least one sick person at 95% probability. We found the sensitivity to be in the range 41-140 and 16-36 sick people at release days 1 and 2, respectively. The WWTP normally serves 36,000 people giving a normalized sensitivity in the range of one in 257 to 2,250 persons., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Kohle et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Antimicrobial resistance (AMR) poses an escalating global public health threat. Canals are essential in Thailand, including the capital city, Bangkok, as agricultural and daily water sources. However, the characteristic and antimicrobial-resistance properties of the bacteria in the urban canals have never been elucidated. This study employed whole genome sequencing to characterize 30 genomes of a causal pathogenic bacteria, Salmonella enterica, isolated from Bangkok canal water between 2016 and 2020. The dominant serotype was Salmonella Agona. In total, 35 AMR genes and 30 chromosomal-mediated gene mutations were identified, in which 21 strains carried both acquired genes and mutations associated with fluoroquinolone resistance. Virulence factors associated with invasion, adhesion, and survival during infection were detected in all study strains. 75.9% of the study stains were multidrug-resistant and all the strains harbored the necessary virulence factors associated with salmonellosis. One strain carried 20 resistance genes, including mcr-3.1 , mutations in GyrA, ParC, and ParE, and typhoid toxin-associated genes. Fifteen plasmid replicon types were detected, with Col(pHAD28) being the most common type. Comparative analysis of nine S . Agona from Bangkok and 167 from public databases revealed that specific clonal lineages of S . Agona might have been circulating between canal water and food sources in Thailand and globally. These findings provide insight into potential pathogens in the aquatic ecosystem and support the inclusion of environmental samples into comprehensive AMR surveillance initiatives as part of a One Health approach. This approach aids in comprehending the rise and dissemination of AMR and devising sustainable intervention strategies.IMPORTANCEBangkok is the capital city of Thailand and home to a large canal network that serves the city in various ways. The presence of pathogenic and antimicrobial-resistant Salmonella is alarming and poses a significant public health risk. The present study is the first characterization of the genomic of Salmonella strains from Bangkok canal water. Twenty-two of 29 strains (75.9%) were multidrug-resistant Salmonella and all the strains carried essential virulence factors for pathogenesis. Various plasmid types were identified in these strains, potentially facilitating the horizontal transfer of AMR genes. Additional investigations indicated a potential circulation of S . Agona between canal water and food sources in Thailand. The current study underscores the role of environmental water in an urban city as a reservoir of pathogens and these data obtained can serve as a basis for public health risk assessment and help shape intervention strategies to combat AMR challenges in Thailand., Competing Interests: The authors declare no conflict of interest.

Metagenomic sequencing has proven to be a powerful tool in the monitoring of antimicrobial resistance (AMR). Here, we provide a comparative analysis of the resistome from pigs, poultry, veal calves, turkey, and rainbow trout, for a total of 538 herds across nine European countries. We calculated the effects of per-farm management practices and antimicrobial usage (AMU) on the resistome in pigs, broilers, and veal calves. We also provide an in-depth study of the associations between bacterial diversity, resistome diversity, and AMR abundances as well as co-occurrence analysis of bacterial taxa and antimicrobial resistance genes (ARGs) and the universality of the latter. The resistomes of veal calves and pigs clustered together, as did those of avian origin, while the rainbow trout resistome was different. Moreover, we identified clear core resistomes for each specific food-producing animal species. We identified positive associations between bacterial alpha diversity and both resistome alpha diversity and abundance. Network analyses revealed very few taxa-ARG associations in pigs but a large number for the avian species. Using updated reference databases and optimized bioinformatics, previously reported significant associations between AMU, biosecurity, and AMR in pig and poultry farms were validated. AMU is an important driver for AMR; however, our integrated analyses suggest that factors contributing to increased bacterial diversity might also be associated with higher AMR load. We also found that dispersal limitations of ARGs are shaping livestock resistomes, and future efforts to fight AMR should continue to emphasize biosecurity measures.IMPORTANCEUnderstanding the occurrence, diversity, and drivers for antimicrobial resistance (AMR) is important to focus future control efforts. So far, almost all attempts to limit AMR in livestock have addressed antimicrobial consumption. We here performed an integrated analysis of the resistomes of five important farmed animal populations across Europe finding that the resistome and AMR levels are also shaped by factors related to bacterial diversity, as well as dispersal limitations. Thus, future studies and interventions aimed at reducing AMR should not only address antimicrobial usage but also consider other epidemiological and ecological factors., Competing Interests: The authors declare no conflict of interest.

Listeria monocytogenes is an important human pathogen with a high mortality rate. Consumption of contaminated ready-to-eat food is the main mode of transmission to humans. Disinfectant-tolerant L. monocytogenes have emerged, which are believed to have increased persistence potential. Elucidating the mechanisms of L. monocytogenes disinfectant tolerance has been the focus of previous studies using pure cultures. A limitation of such approach is the difficulty to identify strains with reduced susceptibility due to inter-strain variation and the need to screen large numbers of strains and genes. In this study, we applied a novel metagenomic approach to detect genes associated with disinfectant tolerance in mixed L. monocytogenes planktonic communities. Two communities, consisting of 71 and 80 isolates each, were treated with the food industry disinfectants benzalkonium chloride (BC, 1.75 mg/L) or peracetic acid (PAA, 38 mg/L). The communities were subjected to metagenomic sequencing and differences in individual gene abundances between biocide-free control communities and biocide-treated communities were determined. A significant increase in the abundance of Listeria phage-associated genes was observed in both communities after treatment, suggesting that prophage carriage could lead to an increased disinfectant tolerance in mixed L. monocytogenes planktonic communities. In contrast, a significant decrease in the abundance of a high-copy emrC-harbouring plasmid pLmN12-0935 was observed in both communities after treatment. In PAA-treated community, a putative ABC transporter previously found to be necessary for L. monocytogenes resistance to antimicrobial agents and virulence, was among the genes with the highest weight for differentiating treated from control samples. The undertaken metagenomic approach in this study can be applied to identify genes associated with increased tolerance to other antimicrobials in mixed bacterial communities., Competing Interests: Declaration of competing interest None., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Purpose of Review: To review recent evidence on allergen immunotherapy (AIT) as a model of personalized medicine in the treatment of children and adolescents with respiratory allergies., Recent Findings: Meta-analysis and systematic review studies continue to point out that AIT is an effective treatment for children with respiratory allergies. Molecular allergy allows the understanding of patient sensitization profiles that frequently change the prescription of AIT. There is still a lack of evidence showing that this personalized prescription of AIT is associated with better clinical outcomes. The nasal allergen challenge has extended the indications of AIT for a new group of subjects with local allergic rhinitis. Patient selection of allergens involved in the increasingly personalized composition of extracts to be used in AIT increasingly characterizes it as personalized medicine., Summary: Despite the numerous studies carried out to identify the best biomarker to evaluate the response to AIT, there is still much disagreement, and clinical assessment (symptoms, quality of life, among others) continues to be the best way to evaluate the therapeutic success of AIT., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Background: Antimicrobial resistance has been identified as a major threat to global health. The pig food chain is considered an important source of antimicrobial resistance genes (ARGs). However, there is still a lack of knowledge on the dispersion of ARGs in pig production system, including the external environment., Results: In the present study, we longitudinally followed one swine farm located in Italy from the weaning phase to the slaughterhouse to comprehensively assess the diversity of ARGs, their diffusion, and the bacteria associated with them. We obtained shotgun metagenomic sequences from 294 samples, including pig feces, farm environment, soil around the farm, wastewater, and slaughterhouse environment. We identified a total of 530 species-level genome bins (SGBs), which allowed us to assess the dispersion of microorganisms and their associated ARGs in the farm system. We identified 309 SGBs being shared between the animals gut microbiome, the internal and external farm environments. Specifically, these SGBs were characterized by a diverse and complex resistome, with ARGs active against 18 different classes of antibiotic compounds, well matching antibiotic use in the pig food chain in Europe., Conclusions: Collectively, our results highlight the urgency to implement more effective countermeasures to limit the dispersion of ARGs in the pig food systems and the relevance of metagenomics-based approaches to monitor the spread of ARGs for the safety of the farm working environment and the surrounding ecosystems., (© 2024. The Author(s).)

Enterococcus faecium ( Efm ) is a leading cause of hospital-associated (HA) infections, often enriched in putative virulence markers (PVMs). Recently, the Efm clade B was assigned as Enterococcus lactis ( Elts ), which usually lack HA- Efm infection markers. Available databases for extracting PVM are incomplete and/or present an intermix of genes from Efm and Enterococcus faecalis , with distinct virulence profiles. In this study, we constructed a new database containing 27 PVMs [ acm, scm, sgrA, ecbA, fnm, sagA, hylEfm, ptsD, orf1481, fms15, fms21-fms20 (pili gene cluster 1, PGC-1), fms14-fms17-fms13 (PGC-2), empA-empB-empC (PGC-3), fms11-fms19-fms16 (PGC-4), ccpA, bepA, gls20-glsB1, and gls33-glsB ] from nine reference genomes (seven Efm + two Elts ). The database was validated against these reference genomes and further evaluated using a collection of well-characterized Efm ( n = 43) and Elts ( n = 7) control strains, by assessing PVM presence/absence and its variants together with a genomic phylogeny constructed as single-nucleotide polymorphisms. We found a high concordance between the phylogeny and in silico findings of the PVM, with Elts clustering separately and mostly carrying Elts- specific PVM gene variants. Based on our validation results, we recommend using the database with raw reads instead of assemblies to avoid missing gene variants. This newly constructed database of 27 PVMs will enable a more comprehensive characterization of Efm and Elts based on WGS data. The developed database exhibits scalability and boasts a range of applications in public health, including diagnostics, outbreak investigations, and epidemiological studies. It can be further used in risk assessment for distinguishing between safe and unsafe enterococci.IMPORTANCEThe newly constructed database, consisting of 27 putative virulence markers, is highly scalable and serves as a valuable resource for the comprehensive characterization of these closely related species using WGS data. It holds significant potential for various public health applications, including hospital outbreak investigations, surveillance, and risk assessment for probiotics and feed additives., Competing Interests: The authors declare no conflict of interest.

Motivation: Analyzing metagenomic data can be highly valuable for understanding the function and distribution of antimicrobial resistance genes (ARGs). However, there is a need for standardized and reproducible workflows to ensure the comparability of studies, as the current options involve various tools and reference databases, each designed with a specific purpose in mind., Results: In this work, we have created the workflow ARGprofiler to process large amounts of raw sequencing reads for studying the composition, distribution, and function of ARGs. ARGprofiler tackles the challenge of deciding which reference database to use by providing the PanRes database of 14 078 unique ARGs that combines several existing collections into one. Our pipeline is designed to not only produce abundance tables of genes and microbes but also to reconstruct the flanking regions of ARGs with ARGextender. ARGextender is a bioinformatic approach combining KMA and SPAdes to recruit reads for a targeted de novo assembly. While our aim is on ARGs, the pipeline also creates Mash sketches for fast searching and comparisons of sequencing runs., Availability and Implementation: The ARGprofiler pipeline is a Snakemake workflow that supports the reuse of metagenomic sequencing data and is easily installable and maintained at https://github.com/genomicepidemiology/ARGprofiler., (© The Author(s) 2024. Published by Oxford University Press.)

Since the beginning of 2023, the number of people with suspected monkeypox virus (MPXV) infection have sharply increased in the Democratic Republic of the Congo (DRC). We report near-to-complete MPXV genome sequences derived from six cases from the South Kivu province. Phylogenetic analyses reveal that the MPXV affecting the cases belongs to a novel Clade I sub-lineage. The outbreak strain genome lacks the target sequence of the probe and primers of a commonly used Clade I-specific real-time PCR.

Poultry farms are hotspots for the development and spread of antibiotic resistance genes (ARGs), due to high stocking densities and extensive use of antibiotics, posing a threat of spread and contagion to workers and the external environment. Here, we applied shotgun metagenome sequencing to characterize the gut microbiome and resistome of poultry, workers and their households - also including microbiomes from the internal and external farm environment - in three different farms in Italy during a complete rearing cycle. Our results highlighted a relevant overlap among the microbiomes of poultry, workers, and their families (gut and skin), with clinically relevant ARGs and associated mobile elements shared in both poultry and human samples. On a finer scale, the reconstruction of species-level genome bins (SGBs) allowed us to delineate the dynamics of microorganism and ARGs dispersion from farm systems. We found the associations with worker microbiomes representing the main route of ARGs dispersion from poultry to human populations. Collectively, our findings clearly demonstrate the urgent need to implement more effective procedures to counteract ARGs dispersion from poultry food systems and the relevance of metagenomics-based metacommunity approaches to monitor the ARGs dispersion process for the safety of the working environment on farms., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

This study presents the first complete genome of Staphylococcus aureus ST5477, one of the most common sequence types (ST) from bovine in eastern Africa. The genome consists of a 2,723,132-bp circular chromosome and a 3,044-bp plasmid. This strain was collected in 2017 from cow milk in Tanzania., Competing Interests: The authors declare no conflict of interest.

The COVID-19 pandemic has seen large-scale pathogen genomic sequencing efforts, becoming part of the toolbox for surveillance and epidemic research. This resulted in an unprecedented level of data sharing to open repositories, which has actively supported the identification of SARS-CoV-2 structure, molecular interactions, mutations and variants, and facilitated vaccine development and drug reuse studies and design. The European COVID-19 Data Platform was launched to support this data sharing, and has resulted in the deposition of several million SARS-CoV-2 raw reads. In this paper we describe (1) open data sharing, (2) tools for submission, analysis, visualisation and data claiming (e.g. ORCiD), (3) the systematic analysis of these datasets, at scale via the SARS-CoV-2 Data Hubs as well as (4) lessons learnt. This paper describes a component of the Platform, the SARS-CoV-2 Data Hubs, which enable the extension and set up of infrastructure that we intend to use more widely in the future for pathogen surveillance and pandemic preparedness.

Contamination with food-borne pathogens, such as Listeria monocytogenes, remains a big concern for food safety. Hence, rigorous and continuous microbial surveillance is a standard procedure. At this point, however, the food industry and authorities only focus on detection of Listeria monocytogenes without characterization of individual strains into groups of more or less concern. As whole genome sequencing (WGS) gains increasing interest in the industry, this methodology presents an opportunity to obtain finer resolution of microbial traits such as virulence. Within this study, we therefore aimed to explore the use of WGS in combination with Machine Learning (ML) to predict L. monocytogenes virulence potential on a sub-species level. The WGS datasets used in this study for ML model training consisted of i) national surveillance isolates (n = 169, covering 38 MLST types) and ii) publicly available isolates acquired through the GenomeTrakr network (n = 2880, spanning 80 MLST types). We used the clinical frequency, i.e., ratio of the number of clinical isolates to total amount of isolates, as estimate for virulence potential. The predictive performance of input features from three different genomic levels (i.e., virulence genes, pan-genome genes, and single nucleotide polymorphisms (SNPs)) and six machine learning algorithms (i.e., Support Vector Machine with a linear kernel, Support Vector Machine with a radial kernel, Random Forrest, Neural Networks, LogitBoost, and Majority Voting) were compared. Our machine learning models predicted sub-species virulence potential with nested cross-validation F1-scores up to 0.88 for the majority voting classifier trained on national surveillance data and using pan-genome genes as input features. The validation of the pre-trained ML models based on 101 previously in vivo studied isolates resulted in F1-scores up to 0.76. Furthermore, we found that the more rapid and less computationally intensive raw read alignment yields comparably accurate models as de novo assembly. The results of our study suggest that a majority voting classifier trained on pan-genome genes is the best and most robust choice for the prediction of clinical frequency. Our study contributes to more rapid and precise characterization of L. monocytogenes virulence and its variation on a sub-species level. We further demonstrated a possible application of WGS data in the context of microbial hazard characterization for food safety. In the future, predictive models may assist case-specific microbial risk management in the food industry. The python code, pre-trained models, and prediction pipeline are deposited at (https://github.com/agmei/LmonoVirulenceML)., Competing Interests: Declaration of competing interest None., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: West Nile virus (WNV) outbreaks in birds, humans, and livestock have occurred in multiple areas in Europe and have had a significant impact on animal and human health. The patterns of emergence and spread of WNV in Europe are very different from those in the US and understanding these are important for guiding preparedness activities., Methods: We mapped the evolution and spread history of WNV in Europe by incorporating viral genome sequences and epidemiological data into phylodynamic models. Spatially explicit phylogeographic models were developed to explore the possible contribution of different drivers to viral dispersal direction and velocity. A "skygrid-GLM" approach was used to identify how changes in environments would predict viral genetic diversity variations over time., Findings: Among the six lineages found in Europe, WNV-2a (a sub-lineage of WNV-2) has been predominant (accounting for 73% of all sequences obtained in Europe that have been shared in the public domain) and has spread to at least 14 countries. In the past two decades, WNV-2a has evolved into two major co-circulating clusters, both originating from Central Europe, but with distinct dynamic history and transmission patterns. WNV-2a spreads at a high dispersal velocity (88km/yr-215 km/yr) which is correlated to bird movements. Notably, amongst multiple drivers that could affect the spread of WNV, factors related to land use were found to strongly influence the spread of WNV. Specifically, the intensity of agricultural activities (defined by factors related to crops and livestock production, such as coverage of cropland, pasture, cultivated and managed vegetation, livestock density) were positively associated with both spread direction and velocity. In addition, WNV spread direction was associated with high coverage of wetlands and migratory bird flyways., Conclusion: Our results suggest that-in addition to ecological conditions favouring bird- and mosquito- presence-agricultural land use may be a significant driver of WNV emergence and spread. Our study also identified significant gaps in data and the need to strengthen virological surveillance in countries of Central Europe from where WNV outbreaks are likely seeded. Enhanced monitoring for early detection of further dispersal could be targeted to areas with high agricultural activities and habitats of migratory birds., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Lu et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Antimicrobial resistance (AMR) is a major global health concern, further complicated by its spread via the microbiome bacterial members. While mathematical models discuss AMR transmission through the symbiotic microbiome, experimental studies are scarce. Herein, we used a gregarious cockroach, Pycnoscelus surinamensis, as an in vivo animal model for AMR transmission investigations. We explored whether the effect of antimicrobial treatment is detectable with metagenomic sequencing, and whether AMR genes can be spread and established in unchallenged (not treated with antibiotics) individuals following contact with treated donors, and under various frequencies of interaction. Gut and soil substrate microbiomes were investigated by metagenomic sequencing for bacterial community composition and resistome profiling. We found that tetracycline treatment altered the treated gut microbiome by decreasing bacterial diversity and increasing the abundance of tetracycline resistance genes. Untreated cockroaches that interacted with treated donors also had elevated tetracycline resistance. The levels of resistance differed depending on the magnitude and frequency of donor transfer. Additionally, treated donors showed signs of microbiome recovery due to their interaction with the untreated ones. Similar patterns were also recorded in the soil substrate microbiomes. Our results shed light on how interacting microbiomes facilitate AMR gene transmission to previously unchallenged hosts, a dynamic influenced by the interaction frequencies, using an in vivo model to validate theoretical AMR transmission models.IMPORTANCEAntimicrobial resistance is a rising threat to human and animal health. The spread of resistance through the transmission of the symbiotic gut microbiome is of concern and has been explored in theoretical modeling studies. In this study, we employ gregarious insect populations to examine the emergence and transmission of antimicrobial resistance in vivo and validate modeling hypotheses. We find that antimicrobial treatment increases the levels of resistance in treated populations. Most importantly, we show that resistance increased in untreated populations after interacting with the treated ones. The level of resistance transmission was affected by the magnitude and frequency of population mixing. Our results highlight the importance of microbial transmission in the spread of antimicrobial resistance., Competing Interests: The authors declare no conflict of interest.