Bridging the divide: addressing discrepancies between clinical guidelines, policy guidelines, and biomarker utilization.

OBJECTIVES: This paper aims to identify and address gaps in cancer treatment and diagnosis within European health services, focusing specifically on discrepancies between clinical guidelines and policy guidelines. It seeks to highlight how the underutilization of advanced diagnostic techniques recommended by medical societies contributes to missed opportunities for improving patient outcomes. METHODS: A comprehensive analysis was conducted across multiple European countries to assess the compliance and integration of clinical guidelines with the availability of advanced diagnostic technologies. Secondary data related to clinical and policy guidelines in cancer care were collected and analyzed. Key indicators of adoption and utilization of next-generation sequencing and liquid biopsy were examined to evaluate their impact on health service efficiency and patient care. RESULTS: The analysis revealed significant discrepancies between the recommendations of medical societies regarding advanced diagnostic techniques and their adoption in health policy decisions across Europe. Country-specific assessments indicated varying levels of alignment between clinical guidelines and the availability of advanced diagnostics. These findings underscored missed opportunities for optimizing patient care and health service efficiency through better alignment and integration of clinical guidelines with policy decisions. CONCLUSIONS: This study concludes that there is a critical need for health policy decision-makers to prioritize the adoption of clinical guidelines in resource allocation and health service organization. Greater attention to the recommendations of medical societies regarding advanced diagnostic techniques could significantly enhance diagnostic accuracy, treatment efficacy, and overall patient outcomes in cancer care. The paper advocates for policy reforms that acknowledge and leverage the potential benefits of advanced diagnostics in improving health service performance and patient-centered care across Europe.

Advances in Hemoglobinopathies and Thalassemia Evaluation.

Hemoglobin (Hb) disorders are among the most prevalent inherited diseases. Despite a limited number of involved genes, these conditions represent a broad clinical and prognostic spectrum. The menu of laboratory tests is extensive. From widely available modalities, for example, complete blood count to rather sophisticated molecular technologies, the investigation of Hb disorders recapitulates an increasing complexity of laboratory workup in other medical fields. This review highlights a current state of biochemical and molecular investigation of Hb disorders and offers a glimpse on technologies that are yet to be fully embraced in clinical practice.

Eight new variants of HLA-DQB1*03 identified by next-generation sequencing.

Genomic sequence of HLA-DQB1*03:01:01:60, -DQB1*03:01:01:61, -DQB1*03:01:01:62, -DQB1*03:01:01:63, -DQB1*03:02:01:23, -DQB1*03:02:01:24, -DQB1*03:02:01:25 and -DQB1*03:03:02:14 alleles in Spanish individuals.

Identification of the novel HLA-DPB1*14:01:15 allele in a Brazilian bone marrow donor.

The novel HLA-DPB1*14:01:15 allele differs from DPB1*14:01:01:01 by change of C > T in exon 3.

Exploring Atrial Fibrillation: Understanding the Complex Relation Between Lifestyle and Genetic Factors.

Cardiovascular diseases (CVDs) are the leading cause of death worldwide across diverse ethnic groups. Among these, atrial fibrillation (AF) stands as one of the most prevalent types of arrhythmias and the primary cause of stroke. Risk factors associated with AF include alcohol consumption, aging, high blood pressure, hypertension, inflammation, and genetic factors. A family history of CVD could indicate an increased risk. Consequently, genetic, and genomic testing should be performed to identify the molecular etiology of CVDs and assess at-risk patients. It is important to note that CVDs are the results of the complex interplay of genes and environmental factors, including ethnicity. In this case, the proband's clinic story includes a history of smoking abuse for 10 years (10 cigarettes per day), obesity, hypertension, and an associated familial history. These risk factors, along with genetic variants, could trigger the early onset of AF. In recent years, genetic and genomic studies have significantly advanced our understanding of CVD etiology, given that next-generation sequencing (NGS) allows for the identification of genetic variants that could contribute to these pathologies. Furthermore, NGS facilitates early diagnosis, personalized pharmacological approaches, and identification of novel biomarkers. Thus, NGS is a valuable tool in CVD management. However, such studies are limited in Ecuador, a low- and middle-income country. Several challenges contribute to this gap, encompassing economic, infrastructural, and educational obstacles. Notably, the cost of genetic and genomic studies may also pose a barrier, restricting access to a portion of the population. In this case report, we present a 56-year-old Ecuadorian woman, who has been diagnosed with AF; however, after performing NGS no disease-associated variants were found, despite having strong clinical signs and symptoms. In summary, this case report contributes valuable insights into the complex interplay between genetic and lifestyle factors in the development and management of AF. The case report aims to underscore the potential impact of genetic variants on disease risk, even when classified as variants of uncertain significance, and the importance of an integral approach to patient care that includes genetic screening, lifestyle interventions, and tailored pharmacological treatment.

Detection of RAS p.Q61R by Immunohistochemistry in Practice: A Clinicopathologic Study of 217 Thyroid Nodules with Molecular Correlates.

RAS p.Q61R is the most prevalent hot-spot mutation in RAS and RAS-like mutated thyroid nodules. A few studies evaluated RAS p.Q61R by immunohistochemistry (RASQ61R-IHC). We performed a retrospective study of an institutional cohort of 150 patients with 217 thyroid lesions tested for RASQ61R-IHC, including clinical, cytologic and molecular data. RASQ61R-IHC was performed on 217 nodules (18% positive, 80% negative, and 2% equivocal). RAS p.Q61R was identified in 76% (n = 42), followed by RAS p.Q61K (15%; n = 8), and RAS p.G13R (5%; n = 3). NRAS p.Q61R isoform was the most common (44%; n = 15), followed by NRAS p.Q61K (17%; n = 6), KRAS p.Q61R (12%; n = 4), HRAS p.Q61R (12%; n = 4), HRAS p.Q61K (6%; n = 2), HRAS p.G13R (6%; n = 2), and NRAS p.G13R (3%; n = 1). RASQ61R-IHC was positive in 47% of noninvasive follicular thyroid neoplasms with papillary-like nuclear features (NIFTP; 17/36), 22% of follicular thyroid carcinomas (FTC; 5/23), 10% of follicular thyroid adenomas (FTA; 4/40), and 8% of papillary thyroid carcinomas (PTC; 9/112). Of PTC studied (n = 112), invasive encapsulated follicular variant (IEFVPTC; n = 16) was the only subtype with positive RASQ61R-IHC (56%; 9/16). Overall, 31% of RAS-mutated nodules were carcinomas (17/54); and of the carcinomas, 94% (16/17) were low-risk per American Thyroid Associated (ATA) criteria, with only a single case (6%; 1/17) considered ATA high-risk. No RAS-mutated tumors recurred, and none showed local or distant metastasis (with a follow-up of 0-10 months). We found that most RAS-mutated tumors are low-grade neoplasms. RASQ61R-IHC is a quick, cost-effective, and reliable way to detect RAS p.Q61R in follicular-patterned thyroid neoplasia and, when malignant, guide surveillance.

Role of Genetic Testing in Kidney Stone Disease: A Narrative Review.

PURPOSE OF REVIEW: Kidney stone disease (KSD) is a common and potentially life-threatening condition, and half of patients experience a repeat kidney stone episode within 5-10 years. Despite the ~50% estimate heritability of KSD, international guidelines have not kept up with the pace of discovery of genetic causes of KSD. The European Association of Urology guidelines lists 7 genetic causes of KSD as 'high risk'. RECENT FINDINGS: There are currently 46 known monogenic (single gene) causes of kidney stone disease, with evidence of association in a further 23 genes. There is also evidence for polygenic risk of developing KSD. Evidence is lacking for recurrent disease, and only one genome wide association study has investigated this phenomenon, identifying two associated genes (SLC34A1 and TRPV5). However, in the absence of other evidence, patients with genetic predisposition to KSD should be treated as 'high risk'. Further studies are needed to characterize both monogenic and polygenic associations with recurrent disease, to allow for appropriate risk stratification. Durability of test result must be balanced against cost. This would enable retrospective analysis if no genetic cause was found initially. We recommend genetic testing using a gene panel for all children, adults < 25 years, and older patients who have factors associated with high risk disease within the context of a wider metabolic evaluation. Those with a genetic predisposition should be managed via a multi-disciplinary team approach including urologists, radiologists, nephrologists, clinical geneticists and chemical pathologists. This will enable appropriate follow-up, counselling and potentially prophylaxis.

The utility of next-generation sequencing in challenging liver FNA biopsies.

BACKGROUND: Fine-needle aspiration (FNA) biopsy is increasingly used for the diagnosis of hepatocellular masses. Because distinguishing well differentiated hepatocellular carcinoma (HCC) from other well differentiated hepatocellular lesions (e.g., large regenerative nodules or focal nodular hyperplasia) requires an assessment of architectural features, this may be challenging on FNA when intact tissue fragments are not sampled. Poorly differentiated HCC and intrahepatic cholangiocarcinoma (ICC) may exhibit overlapping pathologic features. Molecular testing can be helpful, because mutations in TERT promoter and CTNNB1 (ß-catenin) are characteristic of HCC, whereas mutations in BAP1, IDH1/IDH2, and PBRM1 may favor ICC. The goal of this study was to assess the role of next-generation sequencing (NGS) in further subclassifying indeterminate liver lesions sampled by FNA. METHODS: A retrospective review of liver cytology cases with NGS on cell block material was performed. Age, radiologic features, background hepatic disease and treatment, outcome, and NGS data were obtained from the electronic medical record. RESULTS: Twelve FNA biopsies that had cell blocks from clinically suspected primary hepatic masses were identified. The presence of a TERT promoter mutation supported a diagnosis of HCC for one well differentiated neoplasm. For three patients, the presence of mutations, such as IDH1, CDKN2A/CDKN2B, and BRAF, supported a diagnosis of ICC. Of the eight poorly differentiated carcinomas, NGS helped refine the diagnosis in six of eight cases, with one HCC, three ICCs, and two that had combined HCC-ICC, with two cases remaining unclassified. CONCLUSIONS: Molecular diagnostics can be helpful to distinguish HCC and ICC on FNA specimens, although a subset of primary hepatic tumors may remain unclassifiable.

Utility of pan-bacterial and pan-fungal PCR in endophthalmitis: case report and review of the literature.

BACKGROUND: Endophthalmitis is a clinical diagnosis but identification of the disease-causing agent or agents allows for a more tailored treatment. This is routinely done through intraocular fluid cultures and staining. However, culture-negative endophthalmitis is a relatively common occurrence, and a causative organism cannot be identified. Thus, further diagnostic testing, such as pan-bacterial and pan-fungal polymerase chain reactions (PCRs), may be required. BODY: There are now newer, other testing modalities, specifically pan-bacterial and pan-fungal PCRs, that may allow ophthalmologists to isolate a causative agent when quantitative PCRs and cultures remain negative. We present a case report in which pan-fungal PCR was the only test, amongst quantitative PCRs, cultures, and biopsies, that was able to identify a pathogen in endophthalmitis. Pan-PCR has unique advantages over quantitative PCR in that it does not have a propensity for false-positive results due to contamination. Conversely, pan-PCR has drawbacks, including its inability to detect viruses and parasites and its increased turnaround time and cost. Based on two large retrospective studies, pan-PCR was determined not to be recommended in routine cases of systemic infection as it does not typically add value to the diagnostic workup and does not change the treatment course in most cases. However, in cases like the one presented, pan-bacterial and pan-fungal PCRs may be considered if empiric treatment fails or if the infective organism cannot be isolated. If pan-PCR remains negative or endophthalmitis continues to persist, an even newer form of testing, next-generation sequencing, may aid in the diagnostic workup of culture-negative endophthalmitis. CONCLUSION: Pan-bacterial and pan-fungal PCR testing is a relatively new diagnostic tool with unique advantages and drawbacks compared to traditional culturing and PCR methods. Similar to the tests' use in non-ophthalmic systemic infections, pan-bacterial and pan-fungal PCRs are unlikely to become the initial diagnosis test and completely replace culture methods. However, they can provide useful diagnostic information if an infectious agent is unable to be identified with traditional methods or if empiric treatment of endophthalmitis continues to fail.

Next generation plant biostimulants & genome sequencing strategies for sustainable agriculture development.

The best environment for plant growth and development contains certain essential metabolites. A broad category of metabolites known as "plant biostimulants" (PBs) includes biomolecules such as proteins, carbohydrates, lipids, and other secondary metabolites related to groups of terpenes, specific nitrogen-containing compounds, and benzene ring-conjugated compounds. The formation of biomolecules depends on both biotic and abiotic factors, such as the release of PB by plants, animals, and microorganisms, or it can result from the control of temperature, humidity, and pressure in the atmosphere, in the case of humic substances (HSs). Understanding the genomic outputs of the concerned organism (may be plants or others than them) becomes crucial for identifying the underlying behaviors that lead to the synthesis of these complex compounds. For the purposes of achieving the objectives of sustainable agriculture, detailed research on PBs is essential because they aid in increasing yield and other growth patterns of agro-economic crops. The regulation of homeostasis in the plant-soil-microbe system for the survival of humans and other animals is mediated by the action of plant biostimulants, as considered essential for the growth of plants. The genomic size and gene operons for functional and regulation control have so far been revealed through technological implementations, but important gene annotations are still lacking, causing a delay in revealing the information. Next-generation sequencing techniques, such as nanopore, nanoball, and Illumina, are essential in troubleshooting the information gaps. These technical advancements have greatly expanded the candidate gene openings. The secondary metabolites being important precursors need to be studied in a much wider scale for accurate calculations of biochemical reactions, taking place inside and outside the synthesized living cell. The present review highlights the sequencing techniques to provide a foundation of opportunity generation for agricultural sustainability.

Cancer treatment comes to age: from one-size-fits-all to next-generation sequencing (NGS) technologies.

Cancer is one of the leading causes of death worldwide and one of the greatest challenges in extending life expectancy. The paradigm of one-size-fits-all medicine has already given way to the stratification of patients by disease subtypes, clinical characteristics, and biomarkers (stratified medicine). The introduction of next-generation sequencing (NGS) in clinical oncology has made it possible to tailor cancer patient therapy to their molecular profiles. NGS is expected to lead the transition to precision medicine (PM), where the right therapeutic approach is chosen for each patient based on their characteristics and mutations. Here, we highlight how the NGS technology facilitates cancer treatment. In this regard, first, precision medicine and NGS technology are reviewed, and then, the NGS revolution in precision medicine is described. In the sequel, the role of NGS in oncology and the existing limitations are discussed. The available databases and bioinformatics tools and online servers used in NGS data analysis are also reviewed. The review ends with concluding remarks.

Severe community-acquired pneumonia caused by Chlamydia abortus in China: a case report.

Background: Chlamydia abortus causes abortions in ruminants; it can also cause miscarriages and stillbirths in pregnant women. However, it rarely causes pneumonia in humans. Here, we report a case of severe community-acquired pneumonia caused by C. abortus. Case presentation: On admission to our hospital, a 74-year-old woman reported that she had had a fever, cough, phlegm in her throat, and shortness of breath for 10 days. In the local hospital, she was initially diagnosed with community-acquired pneumonia and treated with piperacillin-tazobactam for 4 days. However, her condition worsened, and she was therefore transferred to our hospital. On arrival at our emergency department, she was diagnosed with severe community-acquired pneumonia and treated with a high-flow nasal cannula and meropenem; she was then transferred to the Department of Respiratory Medicine. There, her condition continued to worsen despite continued treatment with the high-flow nasal cannula and omadacycline. After 24 h and emergency tracheal intubation, the patient was sent to the intensive care unit (ICU) for further treatment. The doctors in the ICU again adjusted the treatment, this time to meropenem along with mechanical ventilation; they also instituted methylprednisolone, ulinastatin, nadroparin calcium, and human immunoglobulin. In addition, bronchoalveolar lavage fluid was sent for metagenomic next-generation sequencing (mNGS). Subsequent mNGS suggested the presence of C. abortus, sequence number 5072; we therefore discontinued the meropenem and implemented a combination of doxycycline and moxifloxacin. After 8 days of treatment in the ICU, the patient's condition improved; she was then extubated and, 3 days later, transferred back to the respiratory medicine department. The respiratory physician continued to administer doxycycline and moxifloxacin for 4 days, after which the patient was discharged with medication. A month later, a repeat computed tomography (CT) scan of the chest suggested that the lesions in both lungs had been largely absorbed. Conclusion: C. abortus can occasionally cause pneumonia in humans and, rarely, severe, life-threatening pneumonia. mNGS is uniquely suited for the early detection of this unusual infection. The combination of doxycycline and quinolones has been shown to be effective in severe pneumonia caused by C. abortus.

Genomic mutation patterns and prognostic value in de novo and secondary acute myeloid leukemia: A multicenter study from China.

Acute myeloid leukemia (AML) can manifest as de novo AML (dn-AML) or secondary AML (s-AML), with s-AML being associated with inferior survival and distinct genomic characteristics. The underlying reasons for this disparity remain to be elucidated. In this multicenter study, next-generation sequencing (NGS) was employed to investigate the mutational landscape of AML in 721 patients from June 2020 to May 2023.Genetic mutations were observed in 93.34% of the individuals, with complex variations (more than three gene mutations) present in 63.10% of them. TET2, ASXL1, DNMT3A, TP53 and SRSF2 mutations showed a higher prevalence among older individuals, whereas WT1 and KIT mutations were more commonly observed in younger patients. BCOR, BCORL1, ZRSR2, ASXL1 and SRSF2 exhibited higher mutation frequencies in males. Additionally, ASXL1, NRAS, PPMID, SRSF2, TP53 and U2AF1 mutations were more common in patients with s-AML, which PPM1D was more frequently associated with therapy-related AML (t-AML). Advanced age and hyperleukocytosis independently served as adverse prognostic factors for both types of AML; however, s-AML patients demonstrated a greater number of monogenic adverse prognostic factors compared to dn-AML cases (ASXL1, PPM1D, TP53 and U2AF1 in s-AML vs. FLT3, TP53 and U2AF1 in dn-AML). Age and sex-related gene mutations suggest epigenetic changes may be key in AML pathogenesis. The worse prognosis of s-AML compared to dn-AML could be due to the older age of s-AML patients and more poor-prognosis gene mutations. These findings could improve AML diagnosis and treatment by identifying potential therapeutic targets and risk stratification biomarkers.

Bacillus coagulans-associated severe necrotizing scleral infection.

Purpose: We describe a case of severe scleritis possibly caused by Bacillus coagulans. Observations: Conventional laboratory evaluation was inconclusive. The associated organism was identified with metagenomic RNA deep sequencing (MDS). The infection resolved with trimethoprim-sulfamethoxazole treatment. Conclusions: This case demonstrates the utility of unbiased, high-throughput sequencing for infectious scleritis.

Case Report: Taking action or standing by: managing a preterm neonate at the risk of neonatal varicella by metagenomic next-generation sequencing.

Neonatal varicella is indeed a rare condition, and most infants born to mothers with varicella have a good prognosis. However, in exceptional cases, neonatal varicella can be life-threatening, particularly for preterm infants. Therefore, it is vital to make an early diagnosis or predict the risk of neonatal varicella to ensure prompt treatment and improve prognosis. This report made an effort to early predict neonatal vericalla by using metagenomic next-generation sequencing (mNGS) in a preterm infant who was at risk for vericalla infection. A preterm infant born from a mother with varicella with symptom onset at 8 days before delivery, putting the infant at risk for varicella infection. Importantly, the patient develop pneumonia and pneumothorax, and neonatal vericella was suspected. Fortunately, the use of mNGS for testing the varicella gene in the serum promptly ruled out varicella zoster virus (VZV) infection in the patient, as indicated by a negative mNGS result. Subsequent follow-up, which included a 14-day stay in the hospital followed by an additional 7 days at home, confirmed this finding. Throughout this period, the patient did not exhibit any rash or other symptoms associated with varicella. Therefore, the novel approach of using mNGS allows neonatologists to predict and promptly address potential neonatal infections. This early detection is crucial, as delayed diagnosis or treatment could pose life-threatening risks, as exemplified by the case of neonatal varicella. In such cases, neonatologists can take proactive measures instead of standing by for at-risk neonates. Furthermore, given the severity of neonatal varicella as a life-threatening condition, the early exclusion of subsequent varicella infection by mNGS can offer reassurance to both family members and healthcare professionals.

A Novel Frameshift Mutation of HBB Causing Dominant ß-Thalassemia in a Chinese Individual.

We reported a rare ß-thalassemia patient, a 41-year-old Chinese male with small cell hypopigmentation anemia, jaundice and splenomegaly as the main clinical symptoms. By using Next-Generation Sequencing (NGS), we identified a novel de novo HBB mutation(c.358_365dup, p.Phe123Alafs*39) which resulted in an abnormally prolonged ß-globin chain comprising 159 amino acid residues. The secondary and three-dimensional structures of the ß-globin predicted that the novel prolonged ß-globin chain has a considerable risk of instability in the hemoglobin, and leads to clinical phenotype. This study contributes to the enrichment of the genetic pathogenic mutation database for thalassemia and underscores the significance of NGS in the screening of mutations for thalassemia families.

Whole-genome sequencing for antimicrobial surveillance: species-specific quality thresholds and data evaluation from the network of the European Union Reference Laboratory for Antimicrobial Resistance genomic proficiency tests of 2021 and 2022.

As antimicrobial resistance (AMR) surveillance shifts to genomics, ensuring the quality of whole-genome sequencing (WGS) data produced across laboratories is critical. Participation in genomic proficiency tests (GPTs) not only increases individual laboratories' WGS capacity but also provides a unique opportunity to improve species-specific thresholds for WGS quality control (QC) by repeated resequencing of distinct isolates. Here, we present the results of the EU Reference Laboratory for Antimicrobial Resistance (EURL-AR) network GPTs of 2021 and 2022, which included 25 EU national reference laboratories (NLRs). A total of 392 genomes from 12 AMR-bacteria were evaluated based on WGS QC metrics. Two percent (n = 9) of the data were excluded, due to contamination, and 11% (n = 41) of the remaining genomes were identified as outliers in at least one QC metric and excluded from computation of the adjusted QC thresholds (AQT). Two QC metric correlation groups were identified through linear regression. Eight percent (n = 28) of the submitted genomes, from 11 laboratories, failed one or more of the AQTs. However, only three laboratories (12%) were identified as underperformers, failing across AQTs for uncorrelated QC metrics in at least two genomes. Finally, new species-specific thresholds for "N50" and "number of contigs > 200 bp" are presented for guidance in routine laboratory QC. The continued participation of NRLs in GPTs will reveal WGS workflow flaws and improve AMR surveillance data. GPT data will continue to contribute to the development of reliable species-specific thresholds for routine WGS QC, standardizing sequencing data QC and ensure inter- and intranational laboratory comparability.IMPORTANCEIllumina next-generation sequencing is an integral part of antimicrobial resistance (AMR) surveillance and the most widely used whole-genome sequencing (WGS) platform. The high-throughput, relative low-cost, high discriminatory power, and rapid turnaround time of WGS compared to classical biochemical methods means the technology will likely remain a fundamental tool in AMR surveillance and public health. In this study, we present the current level of WGS capacity among national reference laboratories in the EU Reference Laboratory for AMR network, summarizing applied methodology and statistically evaluating the quality of the obtained sequence data. These findings provide the basis for setting new and revised thresholds for quality metrics used in routine WGS, which have previously been arbitrarily defined. In addition, underperforming participants are identified and encouraged to evaluate their workflows to produce reliable results.

Cost-efficient PCR based DNA barcoding of marine invertebrate specimens with NovaSeq amplicon sequencing.

BACKGROUND: The marine environment harbors high biodiversity; however, it is poorly understood. Nucleotide sequence data of all marine organisms should be accumulated before natural and/or anthropogenic environmental changes jeopardize the marine environment. In this study, we report a cost-effective and easy DNA barcoding method. This method can be readily adopted without using library preparation kits. It includes multiplex PCR of short targets, indexing PCR, and outsourcing to a sequencing service using the NovaSeq system. METHODS AND RESULTS: We targeted four mitochondrial genes [cytochrome c oxidase subunit I (COI), COIII, 16S rRNA (16S), and 12S rRNA (12S)] and three nuclear genes [18S rRNA (18S), 28S rRNA (28S), internal transcribed spacer 2 (ITS2)] in 95 marine invertebrate specimens, which were primarily annelids. The primers, including adapters and indices for NovaSeq sequencing, were newly designed. Two PCR runs were conducted. The 1st PCR amplified specific loci with universal primers and the 2nd added sequencing adapters and indices to the 1st PCR products. The gene sequences obtained from the FASTQ files were subjected to BLAST search and phylogenetic analyses. One run using 95 specimens yielded sequences averaging 2816 bp per specimen for a total length of six loci. Nuclear genes were more successfully assembled compared with mitochondrial genes. A weak but significantly negative correlation was observed between the average length of each locus and success rate of the assembly. Some of the sequences were almost identical to the sequences obtained from specimens collected far from Japan, indicating the presence of potentially invasive species identified for the first time. CONCLUSIONS: We obtained gene sequences efficiently using next-generation sequencing rather than Sanger sequencing. Although this method requires further optimization to increase the success rate for some loci, it is used as a first step to select specimens for further analyses by determining the specific loci of the targets.

Meta-genomic next-generation sequencing in the diagnosis of brucellosis: Five cases from a non-endemic area.

Metagenomic next-generation sequencing (mNGS) in diagnosis of human brucellosis is comparatively unexplored. This report details five human brucellosis cases diagnosed using mNGS based on Illumina sequencing platform, comprising three females and two males, four with epidemiological exposure. In cases 1 and 2, plasma mNGS results showed one positive and one negative for Brucella melitensis, and subsequent blood cultures were both positive. Cases 3, 4 and 5 involved spinal brucellosis, some with paravertebral abscesses. mNGS from infectious tissue samples successfully detected Brucella, with read counts ranging between 30 and 1314, yet cultures were negative in cases 4 and 5. Following antibiotic and surgical treatments, all patients showed clinical improvement. This report shows mNGS testing enhances the detection sensitivity of brucellosis diagnosis.

What is this summary about? Brucella is a type of bacteria that can infect humans and animals. It causes a disease called brucellosis. Symptoms of brucellosis include fever and fatigue, among others. Meta-genomic next-generation sequencing (mNGS) is a tool for sequencing the DNA of bacteria. In this report, we use mNGS to diagnose human brucellosis in five cases.What were the results? Brucella was found in the blood of two infected people, but mNGS found Brucella in only one. Of three people with Brucella infection of the spine, mNGS found Brucella in the infected tissue but Brucella was only cultured in one case. Following antibiotic and surgical treatments, all five patients showed improvement of their symptoms.What do the results of the study mean? mNGS is a relatively rapid and effective diagnostic method that can improve the detection of Brucella in brucellosis.

Integrated assessment of the chemical, microbiological and ecotoxicological effects of a bio-packaging end-of-life in compost.

The present study aimed to i) assess the disintegration of a novel bio-packaging during aerobic composting (2 and 6 % tested concentrations) and evaluate the resulting compost ii) analyse the ecotoxicity of bioplastics residues on earthworms; iii) study the microbial communities during composting and in 'earthworms' gut after their exposure to bioplastic residues; iv) correlate gut microbiota with ecotoxicity analyses; v) evaluate the chemico-physical characterisation of bio-packaging after composting and earthworms' exposure. Both tested concentrations showed disintegration of bio-packaging close to 90 % from the first sampling time, and compost chemical analyses identified its maturity and stability at the end of the process. Ecotoxicological assessments were then conducted on Eisenia fetida regarding fertility, growth, genotoxic damage, and impacts on the gut microbiome. The bioplastic residues did not influence the earthworms' fertility, but DNA damages were measured at the highest bioplastic dose tested. Furthermore bioplastic residues did not significantly affect the bacterial community during composting, but compost treated with 2 % bio-packaging exhibited greater variability in the fungal communities, including Mortierella, Mucor, and Alternaria genera, which can use bioplastics as a carbon source. Moreover, bioplastic residues influenced gut bacterial communities, with Paenibacillus, Bacillus, Rhizobium, Legionella, and Saccharimonadales genera being particularly abundant at 2 % bioplastic concentration. Higher concentrations affected microbial composition by favouring different genera such as Pseudomonas, Ureibacillus, and Streptococcus. For fungal communities, Pestalotiopsis sp. was found predominantly in earthworms exposed to 2 % bioplastic residues and is potentially linked to its role as a microplastics degrader. After composting, Attenuated Total Reflection analysis on bioplastic residues displayed evidence of ageing with the formation of hydroxyl groups and amidic groups after earthworm exposure.