Search

Abstract Background Exploring the dynamics of gut microbiome colonisation during early-life stages is important for understanding the potential impact of microbes on host development and fitness. Evidence from model organisms suggests a crucial early-life phase when shifts in gut microbiota can lead to immune dysregulation and reduced host condition. However, our understanding of gut microbiota colonisation in long-lived vertebrates, especially during early development, remains limited. We therefore used a wild population of common buzzard nestlings (Buteo buteo) to investigate connections between the early-life gut microbiota colonisation, environmental and host factors. Results We targeted both bacterial and eukaryotic microbiota using the 16S and 28S rRNA genes. We sampled the individuals during early developmental stages in a longitudinal design. Our data revealed that age significantly affected microbial diversity and composition. Nest environment was a notable predictor of microbiota composition, with particularly eukaryotic communities differing between habitats occupied by the hosts. Nestling condition and infection with the blood parasite Leucocytozoon predicted microbial community composition. Conclusion Our findings emphasise the importance of studying microbiome dynamics to capture changes occurring during ontogeny. They highlight the role of microbial communities in reflecting host health and the importance of the nest environment for the developing nestling microbiome. Overall, this study contributes to understanding the complex interplay between microbial communities, host factors, and environmental variables, and sheds light on the ecological processes governing gut microbial colonisation during early-life stages.

Simple Summary: Precise analysis of T-cell receptor clonality is essential in lymphoma diagnostics, particularly for cutaneous T-cell lymphomas like Mycosis fungoides (MF) and Sézary Syndrome (SS). The current gold standard, based on next generation sequencing (NGS), is precise and accurate but time- and cost-intensive. This study established a workflow for the analysis of the T-cell receptor repertoire using third generation nanopore sequencing. Results are similarly accurate to NGS for cutaneous T-cell lymphoma, but less time-consuming and cost-intensive. Background: Analysis of T-cell receptor (TCR) clonality is a major diagnostic tool for lymphomas, particularly for cutaneous T-cell lymphomas (CTCL) like Mycosis fungoides and Sézary syndrome. However, a fast and cost-effective workflow is needed to enable widespread use of this method. Methods: We established a procedure for TCR rearrangement analysis via Oxford Nanopore Technology (ONT) sequencing. TCR receptor rearrangements (TCR-gamma and TCR-beta chains) were analyzed in samples from 45 patients with various diagnoses: Mycosis fungoides (37/45), Sézary Syndrome (2/45), folliculotropic CTCL (1/45), and non-CTCL diagnoses as polyclonal controls (5/45). Sample types included formalin-fixed paraffin-embedded (FFPE) samples (27/45), fresh frozen samples (9/45), and CD3-isolated cells (9/45). In addition, DNA of a Jurkat cell line was used as a monoclonal control. TCR amplicons were generated employing an optimized version of the protocol from the Euro Clonality consortium. Sequencing was conducted on the ONT GridION and Illumina MiSeq platforms, followed by similar bioinformatic analysis protocols. The tumor clone frequency (TCF), a crucial prognostic factor for CTCL patients, was used for method comparison. Results: The use of an optimized amplicon protocol and adapted bioinformatic tools demonstrated a strong correlation in TCF values between both sequencing methods across all sample types (range R: 0.992–0.996; range r2: 0.984–0.991). Conclusions: In summary, ONT sequencing was able to detect TCR clonality comparable to NGS, indicating its potential as a faster and more cost-effective option for routine diagnostic use. [ABSTRACT FROM AUTHOR]

Sigma factors are transcriptional regulators that are part of complex regulatory networks for major cellular processes, as well as for growth phase-dependent regulation and stress response. Actinoplanes sp. SE50/110 is the natural producer of acarbose, an α-glucosidase inhibitor that is used in diabetes type 2 treatment. Acarbose biosynthesis is dependent on growth, making sigma factor engineering a promising tool for metabolic engineering. ACSP50_0507 is a homolog of the developmental and osmotic-stress-regulating Streptomyces coelicolor σHSc. Therefore, the protein encoded by ACSP50_0507 was named σHAs. Here, an Actinoplanes sp. SE50/110 expression strain for the alternative sigma factor gene ACSP50_0507 (sigHAs) achieved a two-fold increased acarbose yield with acarbose production extending into the stationary growth phase. Transcriptome sequencing revealed upregulation of acarbose biosynthesis genes during growth and at the late stationary growth phase. Genes that are transcriptionally activated by σHAs frequently code for secreted or membrane-associated proteins. This is also mirrored by the severely affected cell morphology, with hyperbranching, deformed and compartmentalized hyphae. The dehydrated cell morphology and upregulation of further genes point to a putative involvement in osmotic stress response, similar to its S. coelicolor homolog. The DNA-binding motif of σHAs was determined based on transcriptome sequencing data and shows high motif similarity to that of its homolog. The motif was confirmed by in vitro binding of recombinantly expressed σHAs to the upstream sequence of a strongly upregulated gene. Autoregulation of σHAs was observed, and binding to its own gene promoter region was also confirmed. [ABSTRACT FROM AUTHOR]

The Great Pacific Garbage Patch, a significant collection of plastic introduced by human activities, provides an ideal environment to study bacterial lifestyles on plastic substrates. We proposed that bacteria colonizing the floating plastic debris would develop strategies to deal with the ultraviolet‐exposed substrate, such as the production of antioxidant pigments. We observed a variety of pigmentation in 67 strains that were directly cultivated from plastic pieces sampled from the Garbage Patch. The genomic analysis of four representative strains, each distinct in taxonomy, revealed multiple pathways for carotenoid production. These pathways include those that produce less common carotenoids and a cluster of photosynthetic genes. This cluster appears to originate from a potentially new species of the Rhodobacteraceae family. This represents the first report of an aerobic anoxygenic photoheterotrophic bacterium from plastic biofilms. Spectral analysis showed that the bacteria actively produce carotenoids, such as beta‐carotene and beta‐cryptoxanthin, and bacteriochlorophyll a. Furthermore, we discovered that the genetic ability to synthesize carotenoids is more common in plastic biofilms than in the surrounding water communities. Our findings suggest that plastic biofilms could be an overlooked source of bacteria‐produced carotenoids, including rare forms. It also suggests that photoreactive molecules might play a crucial role in bacterial biofilm communities in surface water. [ABSTRACT FROM AUTHOR]

Pseudomonas species have become promising cell factories for the production of natural products due to their inherent robustness. Although these bacteria have naturally evolved strategies to cope with different kinds of stress, many biotechnological applications benefit from engineering of optimised chassis strains with specially adapted tolerance traits. Here, we explored the formation of outer membrane vesicles (OMV) of Pseudomonas putida KT2440. We found OMV production to correlate with the recombinant production of a natural compound with versatile beneficial properties, the tripyrrole prodigiosin. Further, several P. putida genes were identified, whose up‐ or down‐regulated expression allowed controlling OMV formation. Finally, genetically triggering vesiculation in production strains of the different alkaloids prodigiosin, violacein, and phenazine‐1‐carboxylic acid, as well as the carotenoid zeaxanthin, resulted in up to three‐fold increased product yields. Consequently, our findings suggest that the construction of robust strains by genetic manipulation of OMV formation might be developed into a useful tool which may contribute to improving limited biotechnological applications. [ABSTRACT FROM AUTHOR]

Streptococcus pneumoniae ( S.p. ) is the most common causative agent of community-acquired pneumonia worldwide. A key pathogenic mechanism that exacerbates severity of disease is the disruption of the alveolar-capillary barrier. However, the specific virulence mechanisms responsible for this in the human lung are not yet fully understood.In this study, we infected living human lung tissue with S.p. and observed a significant degradation of the central junctional proteins occludin and VE-cadherin, indicating barrier disruption. Surprisingly, neither pneumolysin, bacterial hydrogen peroxide nor pro-inflammatory activation were sufficient to cause this junctional degradation. Instead, pneumococcal infection led to a significant decrease of pH (approximately 6), resulting in acidification of the alveolar microenvironment, which was linked to junctional degradation. Stabilising the pH at physiological levels during infection reversed this effect, even in a therapeutic-like approach.Further analysis of bacterial metabolites and RNA sequencing revealed sugar consumption and subsequent lactate production were the major factors contributing to bacterially induced alveolar acidification, which also hindered the release of critical immune factors.Our findings highlight bacterial metabolite-induced acidification as an independent virulence mechanism for barrier disruption and inflammatory dysregulation in pneumonia. Thus, our data suggest that strictly monitoring and buffering alveolar pH during infections caused by fermentative bacteria could serve as an adjunctive therapeutic strategy for sustaining barrier integrity and immune response., (Copyright ©The authors 2024.)

Photosynthetic organisms often interact with heterotrophic microbes. We here report the complete genome sequences of the bacterial strains Asticcacaulis sp. MM231 and Brevundimonas subvibrioides MM232. Both bacteria were co-isolated from a single green colony originating from an aquatic sample taken from a pond at the Bielefeld University campus., Competing Interests: The authors declare no conflict of interest.

We cultivated bacteria contained in a sandy soil sample, isolated DNA from a single bacterial colony, and assembled from genomic reads the full genome sequence of Chitinophaga and Microbacterium strains, termed MM2321 and MM2322. Besides the genome sequences, the phylogenetic classifications of both strains are reported., Competing Interests: The authors declare no conflict of interest.

Head and neck squamous cell carcinoma (HNSCC) is a highly malignant disease, and death rates have remained at approximately 50% for decades. New tumor-targeting strategies are desperately needed, and a previous report indicated the triggered differentiation of HPV-negative HNSCC cells to confer therapeutic benefits. Using patient-derived tumor cells, we created a similar HNSCC differentiation model of HPV+ tumor cells from two patients. We observed a loss of malignant characteristics in differentiating cell culture conditions, including irregularly enlarged cell morphology, cell cycle arrest with downregulation of Ki67, and reduced cell viability. RNA-Seq showed myocyte-like differentiation with upregulation of markers of myofibril assembly. Immunofluorescence staining of differentiated and undifferentiated primary HPV+ HNSCC cells confirmed an upregulation of these markers and the formation of parallel actin fibers reminiscent of myoblast-lineage cells. Moreover, immunofluorescence of HPV+ tumor tissue revealed areas of cells co-expressing the identified markers of myofibril assembly, HPV surrogate marker p16, and stress-associated basal keratinocyte marker KRT17, indicating that the observed myocyte-like in vitro differentiation occurs in human tissue. We are the first to report that carcinoma cells can undergo a triggered myocyte-like differentiation, and our study suggests that the targeted differentiation of HPV+ HNSCCs might be therapeutically valuable., (© 2024. The Author(s).)

Four rod-shaped, non-motile, non-spore-forming, facultative anaerobic, Gram-stain-positive lactic acid bacteria, designated as EB0058 T , SCR0080, LD0937 T and SCR0063 T , were isolated from different corn and grass silage samples. The isolated strains were characterized using a polyphasic approach and EB0058 T and SCR0080 were identified as Lacticaseibacillus zeae by 16S rRNA gene sequence analysis. Based on whole-genome sequence-based characterization, EB0058 T and SCR0080 were separated into a distinct clade from Lacticaseibacillus zeae DSM 20178 T , together with CECT9104 and UD2202, whose genomic sequences are available from NCBI GenBank. The average nucleotide identity (ANI) values within the new subgroup are 99.9 % and the digital DNA-DNA hybridization (dDDH) values are 99.3-99.9 %, respectively. In contrast, comparison of the new subgroup with publicly available genomic sequences of L. zeae strains, including the type strain DSM 20178 T , revealed dDDH values of 70.2-72.5 % and ANI values of 96.2-96.6 %. Based on their chemotaxonomic, phenotypic and phylogenetic characteristics, EB0058 T and SCR0080 represent a new subspecies of L. zeae . The name Lacticaseibacillus zeae subsp. silagei subsp. nov. is proposed with the type strain EB0058 T (=DSM 116376 T =NCIMB 15474 T ). According to the results of 16S rRNA gene sequencing, LD0937 T and SCR0063 T are members of the Lacticaseibacillus group. The dDDH value between the isolates LD0937 T and SCR0063 T was 67.6 %, which is below the species threshold of 70 %, clearly showing that these two isolates belong to different species. For both strains, whole genome-sequencing revealed that the closest relatives within the Lacticaseibacillus group were Lacticaseibacillus huelsenbergensis DSM 11542 5 (dDDH 66.5 and 65.9 %) and Lacticaseibacillus casei DSM 20011 T (dDDH 64.1 and 64.9 %). Based on the genomic, chemotaxonomic and morphological data obtained in this study, two novel species, Lacticaseibacillus parahuelsenbergensis sp. nov. and Lacticaseibacillus styriensis sp. nov. are proposed and the type strains are LD0937 T (=DSM 116105 T =NCIMB 15471 T ) and SCR0063 T (=DSM 116297 T =NCIMB 15473 T ), respectively.

By investigating wet and dry age-related ripening of beef, Pseudomonas strains V3/3/4/13 T and V3/K/3/5 T were isolated. Strain V3/3/4/13 T exhibited more than 99 % 16S rRNA gene-based similarity to Pseudomonas fragi and other members of this group, while isolate V3/K/3/5 T was very close to Pseudomonas veronii and a number of relatives within the Pseudomonas fluorescens group. Additional comparisons of complete rpoB sequences and draft genomes allowed us to place isolate V3/3/4/13 T close to Pseudomonas deceptionensis DSM 26521 T . In the case of V3/K/3/5 T the closest relative was P. veronii DSM 11331 T . Average nucleotide identity (ANIb) and digital DNA-DNA hybridization (dDDH) values calculated from the draft genomes of V3/3/4/13 T and P. deceptionensis DSM 26521 T were 88.5 and 39.8 %, respectively. For V3/K/3/5 T and its closest relative P. veronii DSM 11331 T , the ANIb value was 95.1 % and the dDDH value was 60.7 %. The DNA G+C contents of V3/3/4/13 T and V3/K/3/5 T were 57.4 and 60.8 mol%, respectively. Predominant fatty acids were C 16 : 0 , C 18 : 1  ω7 c , C 17 : 0 cyclo and summed feature C 16 : 1 ω7 ct /C 15 : 0 iso 2OH. The main respiratory quinones were Q9, with minor proportions of Q8 and, in the case of V3/K/3/5 T , additional Q10. The main polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and, in the case of V3/K/3/5 T , additional phosphatidylcholine. Based on the combined data, isolates V3/3/4/13 T and V3/K/3/5 T should be considered as representatives of two novel Pseudomonas species. The type strain of the newly proposed Pseudomonas kulmbachensis sp. nov. is V3/3/4/13 T (=DSM 113654 T =LMG 32520 T ), a second strain belonging to the same species is FLM 004-28 (=DSM 113604=LMG 32521); the type strain for the newly proposed Pseudomonas paraveronii sp. nov. is V3/K/3/5 T (=DSM 113573 T =LMG 32518 T ) with a second isolate FLM 11 (=DSM 113572=LMG 32519).

Importance: Histone chaperones are proteins that are involved in nucleosome assembly and disassembly and can therefore influence all DNA-dependent processes including transcription, DNA replication, and repair. ASF1 is a histone chaperone that is conserved throughout eukaryotes. In contrast to most other multicellular organisms, a deletion mutant of asf1 in the fungus Sordaria macrospora is viable; however, the mutant is sterile. In this study, we could show that the histone-binding ability of ASF1 is required for fertility in S. macrospora , whereas the function of ASF1 in maintenance of genome stability does not require histone binding. We also showed that the histone modifications H3K27me3 and H3K56ac are misregulated in the Δasf1 mutant. Furthermore, we identified a large duplication on chromosome 2 of the mutant strain that is genetically linked to the Δasf1 allele present on chromosome 6, suggesting that viability of the mutant might depend on the presence of the duplicated region., Competing Interests: The authors declare no conflict of interest.