Adjusting intracellular metabolic pathways and adopting suitable live state such as biofilms, are crucial for bacteria to survive environmental changes. Although substantial progress has been made in understanding how the histone-like nucleoid-structuring (H-NS) protein modulates the expression of the genes involved in biofilm formation, the precise modification that the H-NS protein undergoes to alter its DNA binding activity is still largely uncharacterized. This study revealed that acetylation of H-NS at Lys19 inhibits biofilm development in Shewanella oneidensis MR-1 by downregulating the expression of glutamine synthetase, a critical enzyme in glutamine synthesis. We further found that nitrogen starvation, a likely condition in biofilm development, induces deacetylation of H-NS and the trimerization of nitrogen assimilation regulator GlnB. The acetylated H-NS strain exhibits significantly lower cellular glutamine concentration, emphasizing the requirement of H-NS deacetylation in Shewanella biofilm development. Moreover, we discovered in vivo that the activation of glutamine biosynthesis pathway and the concurrent suppression of the arginine synthesis pathway during both pellicle and attached biofilms development, further suggesting the importance of fine tune nitrogen assimilation by H-NS acetylation in Shewanella. In summary, posttranslational modification of H-NS endows Shewanella with the ability to respond to environmental needs by adjusting the intracellular metabolism pathways.

The filamentous 'Pf' bacteriophages of Pseudomonas aeruginosa play roles in biofilm formation and virulence, but mechanisms governing Pf prophage activation in biofilms are unclear. Here, we identify a prophage regulatory module, KKP (kinase-kinase-phosphatase), that controls virion production of co-resident Pf prophages and mediates host defense against diverse lytic phages. KKP consists of Ser/Thr kinases PfkA and PfkB, and phosphatase PfpC. The kinases have multiple host targets, one of which is MvaU, a host nucleoid-binding protein and known prophage-silencing factor. Characterization of KKP deletion and overexpression strains with transcriptional, protein-level and prophage-based approaches indicates that shifts in the balance between kinase and phosphatase activities regulate phage production by controlling MvaU phosphorylation. In addition, KKP acts as a tripartite toxin-antitoxin system that provides defense against some lytic phages. A conserved lytic phage replication protein inhibits the KKP phosphatase PfpC, stimulating toxic kinase activity and blocking lytic phage production. Thus, KKP represents a phosphorylation-based mechanism for prophage regulation and antiphage defense. The conservation of KKP gene clusters in >1000 diverse temperate prophages suggests that integrated control of temperate and lytic phage infection by KKP-like regulatory modules may play a widespread role in shaping host cell physiology.

Abstract The coral reef microbiome plays a vital role in the health and resilience of reefs. Previous studies have examined phage therapy for coral pathogens and for modifying the coral reef microbiome, but defence systems against coral‐associated bacteria have received limited attention. Phage defence systems play a crucial role in helping bacteria fight phage infections. In this study, we characterized a new defence system, Hma (HmaA‐HmaB‐HmaC), in the coral‐associated Halomonas meridiana derived from the scleractinian coral Galaxea fascicularis. The Swi2/Snf2 helicase HmaA with a C‐terminal nuclease domain exhibits antiviral activity against Escherichia phage T4. Mutation analysis revealed the nickase activity of the nuclease domain (belonging to PDD/EXK superfamily) of HmaA is essential in phage defence. Additionally, HmaA homologues are present in ~1000 bacterial and archaeal genomes. The high frequency of HmaA helicase in Halomonas strains indicates the widespread presence of these phage defence systems, while the insertion of defence genes in the hma region confirms the existence of a defence gene insertion hotspot. These findings offer insights into the diversity of phage defence systems in coral‐associated bacteria and these diverse defence systems can be further applied into designing probiotics with high‐phage resistance.

Filamentous bacteriophages play a critical role in biofilm formation and virulence in the opportunistic pathogen Pseudomonas aeruginosa. Here, studies of the filamentous Pf4 prophage life cycle within P. aeruginosa biofilms revealed that the prophage-encoded reverse transcriptase (RT) regulates phage genome dynamics. The RT and the non-coding RNA PhrD collaborate to edit the Pf4 phage genome to generate superinfective Pf4 variants capable of rapid propagation within biofilms by preserving genes essential for virion assembly and reconstituting a promoter for the phage excisionase gene. Mutant cells emerge in biofilms where intact Pf4 prophages are replaced by these reduced-genome phage variants, further enhancing virion production. The discovery of RT’s role in phage genome reduction expands understanding of RT functions and of the versatility of phage biology and its impact on microbial community dynamics within biofilms.

Although the gut microbiota has been well reported to contribute to the pest insecticidal activity of Bacillus thuringiensis (Bt) toxin, the underlying mechanism remains largely unknown. Here, bioassay, RNA-seq, and 16S rRNA sequencing were employed to explore the potential mechanism by which gut microbiota enhance the virulence of Bt maize to Mythimna separata. Two groups of neonate larvae were fed with the leaves of conventional maize (CK group) and insect-resistant transgenic maize (Bt group), respectively. The results indicated that body weight, body length, and survival rate of M. separata in the Bt group were significantly reduced. Additionally, the induced growth retardation further led to normal mating failure and fecundity deprivation of M. separata in Bt group. The 16S rRNA sequencing results indicated that exposure to insect-resistant transgenic maize significantly decreased the diversity of the bacterial community and increased the relative abundance of Enterobacter cloacae and Enterococcus mundtii by 153.02-folds and 2.08-folds, respectively. Reintroduction of these two bacteria significantly increased the sensitivity of M. separata to Bt toxin. Transcriptome results revealed that numerous genes related to Notch, Hippo, Wnt, amino acid synthesis, and fatty acid metabolism pathways were significantly down-regulated in the Bt group, whereas almost all Bt toxin receptor aminopeptidase N genes were up-regulated. This study revealed that exposure to insect-resistant transgenic maize reduced gut bacterial diversity and altered the gene expression of M. separata. Furthermore, E. cloacae and E. mundtii enhanced the susceptibility of M. separata to Bt toxin. Our findings on the interaction between Bt toxins, gut microbiota, and the host insect will contribute to the development of pest management strategies and the promotion of transgenic crops in China.

The oriental armyworm, Mythimna separata, is an extremely destructive polyphagous pest with a broad host range that seriously threatens the safety of agricultural production. Here, a high-quality chromosome-level genome was assembled using Illumina, PacBio HiFi long sequencing, and Hi-C scaffolding technologies. The genome size was 706.30 Mb with a contig N50 of 22.08 Mb, and 99.2% of the assembled sequences were anchored to 31 chromosomes. In addition, 20,375 protein-coding genes and 258.68 Mb transposable elements were identified. The chromosome-level genome assembly of M. separata provides a significant genetic resource for future studies of this insect and contributes to the development of management strategies.

蓝细菌(cyanobacteria)是一类能进行放氧光合作用的原核生物.蓝细菌生长速度较快,几乎存在于所有的陆地和水生环境中.毒素-抗毒素(toxin-antitoxin,TA)系统在原核生物中分布十分广泛,在细菌的生命活动中扮演了重要的角色,如维持水平基因转移元件的稳定性以及应对环境胁迫压力等.已有的基因组分析表明,蓝细菌基因组中含有大量潜在的毒素-抗毒素系统,但是目前对蓝细菌毒素-抗毒素系统的实验鉴定仍较少.本文分别以淡水和海水代表菌株集胞藻PCC6803(Synechocystis sp.PCC6803)和聚球藻WH7803(Synechococcus sp.WH7803)为研究对象,对二者基因组上的毒素-抗毒素系统进行预测,并选取预测结果中的7对潜在的毒素-抗毒素系统进行实验验证.结果表明,集胞藻PCC6803的两对毒素-抗毒素系统BAD01932-1933和BAA18559-New ORF7中的毒素具有明显的细胞毒性.本文的研究结果有助于后续对蓝细菌毒素-抗毒素系统及其生态和生物学功能的研究.

Regulatory systems that maintain prophage quiescence integrate phage and host gene expression with environmental conditions 1,2 . In the opportunistic bacterial pathogen Pseudomonas aeruginosa , Pf filamentous bacteriophages play critical roles in biofilm formation and virulence 3-5 , but mechanisms governing Pf prophage activation in biofilms are largely unknown. Here, we report a new type of prophage regulatory module in a widely-distributed P. aeruginosa lineage that not only controls virion production of co-resident Pf prophages, but also mediates defense against diverse lytic phages. By comparing two lineages of the prototype P. aeruginosa strain PAO1 that harbor different Pf prophages, we identified a prophage-encoded kinase-kinase-phosphatase (KKP) system that controls Pf production in biofilms. KKP components exhibit dynamic stoichiometry, where high kinase levels in planktonic conditions maintain phosphorylation of the host H-NS protein MvaU, repressing prophage activation. During biofilm formation, phosphatase expression is heightened, leading to MvaU dephosphorylation and alleviating repression of prophage gene expression. KKP clusters are present in hundreds of diverse temperate prophages and other mobile elements across Gram-negative bacteria. Characterization of KKP modules from different species revealed that, in addition to regulating Pf phage lysogeny, KKP functions as a tripartite toxin-antitoxin system that mediates host defense from predatory lytic phages. KKP represents a new phosphorylation-based mechanism for prophage regulation and for phage defense. The dual function of this module raises the question of whether other newly described phage defense systems 6-9 also regulate intrinsic prophage biology in diverse hosts.

Lateral gene transfer (LGT) plays a key role in shaping the genome evolution and environmental adaptation of bacteria. Xenogeneic silencing is crucial to ensure the safe acquisition of LGT genes into host pre-existing regulatory networks. We previously found that the host nucleoid structuring protein (H-NS) silences prophage CP4So at warm temperatures yet enables this prophage to excise at cold temperatures in Shewanella oneidensis. However, whether H-NS silences other genes and how bacteria modulate H-NS to regulate the expression of genes have not been fully elucidated. In this study, we discovered that the H-NS silences many LGT genes and the xenogeneic silencing of H-NS relies on a temperature-dependent phosphorylation at warm temperatures in S. oneidensis. Specifically, phosphorylation of H-NS at Ser42 is critical for silencing the cold-inducible genes including the excisionase of CP4So prophage, a cold shock protein, and a stress-related chemosensory system. By contrast, nonphosphorylated H-NS derepresses the promoter activity of these genes/operons to enable their expression at cold temperatures. Taken together, our results reveal that the posttranslational modification of H-NS can function as a regulatory switch to control LGT gene expression in host genomes to enable the host bacterium to react and thrive when environmental temperature changes.

We report the complete sequence of the mitochondrial genome of Penicillium marneffei, the first complete mitochondrial DNA sequence of a thermal dimorphic fungus. This 35 kb mitochondrial genome contains the genes encoding ATP synthase subunits 6, 8, and 9 (atp6, atp8, and atp9), cytochrome oxidase subunits I, II, and III (cox1, cox2, and cox3), apocytochrome b (cob), reduced nicotinamide adenine dinucleotide ubiquinone oxireductase subunits (nad1, nad2, nad3, nad4, nad4L, nad5, and nad6), ribosomal protein of the small ribosomal subunit (rps), 28 tRNAs, and small and large ribosomal RNAs. Analysis of gene contents, gene orders, and gene sequences revealed that the mitochondrial genome of P. marneffei is more closely related to those of molds than yeasts.