Concurrent inhibition of oncogenic and wild-type RAS-GTP for cancer therapy.

RAS oncogenes (collectively NRAS, HRAS and especially KRAS) are among the most frequently mutated genes in cancer, with common driver mutations occurring at codons 12, 13 and 611. Small molecule inhibitors of the KRAS(G12C) oncoprotein have demonstrated clinical efficacy in patients with multiple cancer types and have led to regulatory approvals for the treatment of non-small cell lung cancer2,3. Nevertheless, KRASG12C mutations account for only around 15% of KRAS-mutated cancers4,5, and there are no approved KRAS inhibitors for the majority of patients with tumours containing other common KRAS mutations. Here we describe RMC-7977, a reversible, tri-complex RAS inhibitor with broad-spectrum activity for the active state of both mutant and wild-type KRAS, NRAS and HRAS variants (a RAS(ON) multi-selective inhibitor). Preclinically, RMC-7977 demonstrated potent activity against RAS-addicted tumours carrying various RAS genotypes, particularly against cancer models with KRAS codon 12 mutations (KRASG12X). Treatment with RMC-7977 led to tumour regression and was well tolerated in diverse RAS-addicted preclinical cancer models. Additionally, RMC-7977 inhibited the growth of KRASG12C cancer models that are resistant to KRAS(G12C) inhibitors owing to restoration of RAS pathway signalling. Thus, RAS(ON) multi-selective inhibitors can target multiple oncogenic and wild-type RAS isoforms and have the potential to treat a wide range of RAS-addicted cancers with high unmet clinical need. A related RAS(ON) multi-selective inhibitor, RMC-6236, is currently under clinical evaluation in patients with KRAS-mutant solid tumours (ClinicalTrials.gov identifier: NCT05379985).

Tumour-selective activity of RAS-GTP inhibition in pancreatic cancer.

Broad-spectrum RAS inhibition has the potential to benefit roughly a quarter of human patients with cancer whose tumours are driven by RAS mutations1,2. RMC-7977 is a highly selective inhibitor of the active GTP-bound forms of KRAS, HRAS and NRAS, with affinity for both mutant and wild-type variants3. More than 90% of cases of human pancreatic ductal adenocarcinoma (PDAC) are driven by activating mutations in KRAS4. Here we assessed the therapeutic potential of RMC-7977 in a comprehensive range of PDAC models. We observed broad and pronounced anti-tumour activity across models following direct RAS inhibition at exposures that were well-tolerated in vivo. Pharmacological analyses revealed divergent responses to RMC-7977 in tumour versus normal tissues. Treated tumours exhibited waves of apoptosis along with sustained proliferative arrest, whereas normal tissues underwent only transient decreases in proliferation, with no evidence of apoptosis. In the autochthonous KPC mouse model, RMC-7977 treatment resulted in a profound extension of survival followed by on-treatment relapse. Analysis of relapsed tumours identified Myc copy number gain as a prevalent candidate resistance mechanism, which could be overcome by combinatorial TEAD inhibition in vitro. Together, these data establish a strong preclinical rationale for the use of broad-spectrum RAS-GTP inhibition in the setting of PDAC and identify a promising candidate combination therapeutic regimen to overcome monotherapy resistance.

Uropathogenic Escherichia coli Subverts Host Autophagic Defenses by Stalling Preautophagosomal Structures to Escape Lysosome Exocytosis.

Urinary tract infections are primarily caused by uropathogenic Escherichia coli (UPEC). UPEC infects bladder epithelial cells (BECs) via fusiform vesicles and escapes into the cytosol by disrupting fusiform vesicle membrane using outer membrane phospholipase PldA, and establishes biofilm-like intracellular bacterial communities (IBCs) for protection from host immune clearance. Cytosolic UPEC is captured by autophagy to form autophagosomes, then transported to lysosomes, triggering the spontaneous exocytosis of lysosomes. The mechanism by which UPEC evades autophagy to recognize and form IBCs remains unclear. Here, we demonstrate that by inhibiting autophagic flux, UPEC PldA reduces the lysosome exocytosis of BECs. By reducing intracellular phosphatidylinositol 3-phosphate levels, UPEC PldA increases the accumulation of NDP52 granules and decreases the targeting of NDP52 to autophagy, hence stalling preautophagosome structures. Thus, our results uncover a critical role for PldA to inhibit autophagic flux, favoring UPEC escapes from lysosome exocytosis, thereby contributing to acute urinary tract infection.

Transcriptomic and Physiological Analysis of the Effects of Exogenous Phloretin and Pterostilbene on Resistance Responses of Stylosanthes against Anthracnose.

Anthracnose caused by Colletotrichum gloeosporioides is a destructive disease of Stylosanthes (stylo). Combination treatment of phloretin and pterostilbene (PP) has been previously shown to effectively inhibit the conidial germination and mycelial growth of C. gloeosporioides in vitro. In this study, the effects of PP treatment on the growth of C. gloeosporioides in vivo and the biocontrol mechanisms were investigated. We found that exogenous PP treatment could limit the growth of C. gloeosporioides and alleviate the damage of anthracnose in stylo. Comparative transcriptome analysis revealed that 565 genes were up-regulated and 239 genes were down-regulated upon PP treatment during the infection by C. gloeosporioides. The differentially expressed genes were mainly related to oxidative stress and chloroplast organization. Further physiological analysis revealed that application of PP after C. gloeosporioides inoculation significantly reduced the accumulation of O2•- level and increased the accumulation of antioxidants (glutathione, ascorbic acid and flavonoids) as well as the enzyme activity of total antioxidant capacity, superoxide dismutase, catalase, glutathione reductase, peroxidase and ascorbate peroxidase. PP also reduced the decline of chlorophyll a + b and increased the content of carotenoid in response to C. gloeosporioides infection. These results suggest that PP treatment alleviates anthracnose by improving antioxidant capacity and reducing the damage of chloroplasts, providing insights into the biocontrol mechanisms of PP on the stylo against anthracnose.

Abdominal obesity and digestive system cancer: a systematic review and meta-analysis of prospective studies.

BACKGROUND: The diagnostic criteria for abdominal obesity are usually waist circumference or waist-to-hip ratio. The magnitude of the risks for cancers of the digestive system and abdominal obesity is unknown. To assess whether abdominal obesity increases the risk of digestive cancer, we conducted a systematic review and meta-analysis of prospective cohort studies in a database. METHODS: PubMed, Embase, and Web of Science databases were searched from their inception to December 2022. The 9-star Newcastle Ottawa Scale was used to assess  study quality. Pooled relative risks and 95% confidence intervals were calculated using fixed or random effect models respectively. The stability of the results was explored by one-by-one exclusion. Subgroup analysis was conducted to explore sources of heterogeneity. Publication bias was evaluated by Begg's and Egger's tests. RESULTS: A total of 43 cohort studies were included. There were 42 and 31 studies in the meta-analysis of waist circumference and waist-to-hip ratio on digestive system cancer, respectively. The results of the meta-analysis revealed that the greater waist circumference and waist-to-hip ratio were correlated with increased incidence of digestive system cancers: waist circumference: RR 1.48, 95% CI 1.38-1.59, p < 0.001; waist-to-hip ratio: RR 1.33, 95% CI 1.28-1.38, p = 0.001. Subgroup analysis by cancer type showed that higher WC and WHR would increase the prevalence of LC, PC, GC, EC, and CRC. The sensitivity analysis was conducted by a one-by-one elimination method, and the results of the meta-analysis remained stable. It is proved that the results were robust by the trim-and-fill method. CONCLUSIONS: There was evidence to suggest that abdominal obesity increased the incidence of digestive cancer, it is necessary to take appropriate measures to reduce abdominal obesity. Waist circumference and waist-to-hip ratio may be better predictors of digestive system cancers. However, the association between waist circumference and digestive system cancer was greater, so more attention should be paid to measuring abdominal obesity with waist circumference.

The LysR-Type Transcription Regulator YhjC Promotes the Systemic Infection of Salmonella Typhimurium in Mice.

Salmonella Typhimurium is a Gram-negative intestinal pathogen that can infect humans and a variety of animals, causing gastroenteritis or serious systemic infection. Replication within host macrophages is essential for S. Typhimurium to cause systemic infection. By analyzing transcriptome data, the expression of yhjC gene, which encodes a putative regulator in S. Typhimurium, was found to be significantly up-regulated after the internalization of Salmonella by macrophages. Whether yhjC gene is involved in S. Typhimurium systemic infection and the related mechanisms were investigated in this study. The deletion of yhjC reduced the replication ability of S. Typhimurium in macrophages and decreased the colonization of S. Typhimurium in mouse systemic organs (liver and spleen), while increasing the survival rate of the infected mice, suggesting that YhjC protein promotes systemic infection by S. Typhimurium. Furthermore, by using transcriptome sequencing and RT-qPCR assay, the transcription of several virulence genes, including spvD, iroCDE and zraP, was found to be down-regulated after the deletion of yhjC. Electrophoretic mobility shift assay showed that YhjC protein can directly bind to the promoter region of spvD and zraP to promote their transcription. These findings suggest that YhjC contributes to the systemic virulence of S. Typhimurium via the regulation of multiple virulence genes and YhjC could represent a promising target to control S. Typhimurium infection.

miR-31-5p from placental and peripheral blood exosomes is a potential biomarker to diagnose preeclampsia.

BACKGROUND: Preeclampsia, a multisystem disorder of unknown etiology, is one of the leading causes of maternal and perinatal morbidity and mortality. Identifying sensitive, noninvasive markers can aid its prevention and improve prognosis. microRNAs (miRs), which function as negative regulators of gene expression, are closely related to preeclampsia occurrence and development. Herein we investigated the relationship between the DLK1-Dio3 imprinted miR cluster derived from placental and peripheral blood exosomes of pregnant women with preeclampsia and routine clinical diagnostic indicators, and also determined its potential as a noninvasive diagnostic marker. METHODS: Exosomes were extracted from the placenta and peripheral blood of pregnant women with preeclampsia. RESULTS: qPCR data indicated that the expression level of miRs, such as miR-134, miR-31-5p, miR-655, miR-412, miR-539, miR-409, and miR-496, in pregnant women with preeclampsia was significantly lower than that in healthy controls; miR-31-5p expression was the most different. Gene ontology analysis predicted that genes negatively regulated by miR-31-5p were mainly enriched in cellular entity, cellular process, and binding; moreover, Kyoto Encyclopedia of Genes and Genomes pathway analyses indicated that genes were involved in gonadotropin-releasing hormone receptor pathway and other signaling pathways. Correlation analysis revealed that miR-31-5p was significantly negatively correlated with clinical indicators of preeclampsia, such as systolic and diastolic pressure, lactate dehydrogenase, and proteinuria. CONCLUSION: We believe that exosome-derived miR-31-5p can serve as an effective and sensitive biomarker to determine the course of preeclampsia in pregnant women.

Genome-wide analysis of tandem duplicated genes and their contribution to stress resistance in pigeonpea (Cajanus cajan).

Pigeonpea is the main protein source for more than one billion people, and it shows a strong adaptation to biotic stress and abiotic stress. Gene duplication is a fundamental process in genome evolution. Although the draft sequence of the pigeonpea genome has been available since 2011, further analysis of tandem duplicated genes (TDGs) and their contribution to the evolution of pigeonpea has not been reported. In this study, we identify 3211 TDGs in the pigeonpea genome and KEGG enrichment analysis of these genes shows that the TDGs are significantly enriched in resistance-related pathways. In addition, we find that TDGs are more abundant in retrotransposon-related genes in pigeonpea than in the other species included in our study. These results indicate that stress resistance in pigeonpea may be ascribed to resistance-related pathways and retrotransposons originating from tandem duplications. Our study will provide an important basis for further research in pigeonpea breeding.

Transcriptomics and metabolomics reveal the induction of flavonoid biosynthesis pathway in the interaction of Stylosanthes-Colletotrichum gloeosporioides.

Colletotrichum, a hemibiotrophic fungal pathogen with a broad host range, causes a yield-limiting disease called anthracnose. Stylo (Stylosanthes) is a dominant pasture legume in tropics and subtropics, and anthracnose is one of its most destructive disease. Resistance mechanisms against anthracnose in stylo are poorly understood, thus hindering the development of resistant varieties. We performed time-resolved leaf transcriptomics, metabolomics and in vitro inhibition assay to investigate the defense responses against Colletotrichum gloeosporioides in stylo. Transcriptomics demonstrated that flavonoid biosynthetic genes were significantly induced during the infection. Consistently, metabolomics also showed the increased accumulation of flavonoid compounds. In vitro assays showed that phloretin and naringenin inhibited the mycelial growth, and apigenin, daidzein, quercetin and kaempferol suppressed conidial germination of Colletotrichum strains. Together, our results suggest that stylo plants cope with C. gloeosporioides by up-regulation of genes and compounds in flavonoid biosynthesis pathway, providing potential targets for resistance breeding.

Collaborative Action of Microglia and Astrocytes Mediates Neutrophil Recruitment to the CNS to Defend against Escherichia coli K1 Infection.

Escherichia coli K1 is a leading cause of neonatal bacterial meningitis. Recruitment of neutrophils to the central nervous system (CNS) via local immune response plays a critical role in defense against E. coli K1 infection; however, the mechanism underlying this recruitment remains unclear. In this study, we report that microglia and astrocytes are activated in response to stimulation by E. coli K1 and/or E. coli K1-derived outer membrane vesicles (OMVs) and work collaboratively to drive neutrophil recruitment to the CNS. Microglial activation results in the release of the pro-inflammatory cytokine TNF-α, which activates astrocytes, resulting in the production of CXCL1, a chemokine critical for recruiting neutrophils. Mice lacking either microglia or TNF-α exhibit impaired production of CXCL1, impaired neutrophil recruitment, and an increased CNS bacterial burden. C-X-C chemokine receptor 2 (CXCR2)-expressing neutrophils primarily respond to CXCL1 released by astrocytes. This study provides further insights into how immune responses drive neutrophil recruitment to the brain to combat E. coli K1 infection. In addition, we show that direct recognition of E. coli K1 by microglia is prevented by the K1 capsule. This study also reveals that OMVs are sufficient to induce microglial activation.

Nitrate Utilization Promotes Systemic Infection of Salmonella Typhimurium in Mice.

Salmonella Typhimurium is an invasive enteric pathogen that causes gastroenteritis in humans and life-threatening systemic infections in mice. During infection of the intestine, S. Typhimurium can exploit nitrate as an electron acceptor to enhance its growth. However, the roles of nitrate on S. Typhimurium systemic infection are unknown. In this study, nitrate levels were found to be significantly increased in the liver and spleen of mice systemically infected by S. Typhimurium. Mutations in genes encoding nitrate transmembrane transporter (narK) or nitrate-producing flavohemoprotein (hmpA) decreased the replication of S. Typhimurium in macrophages and reduced systemic infection in vivo, suggesting that nitrate utilization promotes S. Typhimurium systemic virulence. Moreover, nitrate utilization contributes to the acidification of the S. Typhimurium cytoplasm, which can sustain the virulence of S. Typhimurium by increasing the transcription of virulence genes encoding on Salmonella pathogenicity island 2 (SPI-2). Furthermore, the growth advantage of S. Typhimurium conferred by nitrate utilization occurred only under low-oxygen conditions, and the nitrate utilization was activated by both the global regulator Fnr and the nitrate-sensing two-component system NarX-NarL. Collectively, this study revealed a novel mechanism adopted by Salmonella to interact with its host and increase its virulence.

Correction: Signal transduction pathway mediated by the novel regulator LoiA for low oxygen tension induced Salmonella Typhimurium invasion.

[This corrects the article DOI: 10.1371/journal.ppat.1006429.].

PagR mediates the precise regulation of Salmonella pathogenicity island 2 gene expression in response to magnesium and phosphate signals in Salmonella Typhimurium.

To establish systemic infections, Salmonella enterica serovar Typhimurium (S. Typhimurium) requires Salmonella pathogenicity island 2 (SPI-2) to survive and replicate within macrophages. High expression of many SPI-2 genes during the entire intracellular growth period within macrophages is essential, as it contributes to the formation of Salmonella-containing vacuole and bacterial replication. However, the regulatory mechanisms underlying the sustained induction of SPI-2 within macrophages are not fully understood. Here, we revealed a time-dependent regulation of SPI-2 expression mediated by a novel regulator PagR (STM2345) in response to the low Mg2+ and low phosphate (Pi ) signals, which ensured the high induction of SPI-2 during the entire intramacrophage growth period. Deletion of pagR results in reduced bacterial replication in macrophages and attenuation of systemic virulence in mice. The effects of pagR on virulence are dependent on upregulating the expression of slyA, a regulator of SPI-2. At the early (0-4 hr) and later (after 4 hr) stage post-infection of macrophages, pagR is induced by the low Pi via PhoB/R two-component systems and low Mg2+ via PhoP/Q systems, respectively. Collectively, our findings revealed that the PagR-mediated regulatory mechanism contributes to the precise and sustained activation of SPI-2 genes within macrophages, which is essential for S. Typhimurium systemic virulence.

Bacterial and Microfauna Mechanisms for Sludge Reduction in Carrier-Enhanced Anaerobic Side-Stream Reactors Revealed by Metagenomic Sequencing Analysis.

Packing carriers into the anaerobic side-stream reactor (ASSR) can enhance sludge reduction and save footprint by investigating ASSR-coupled membrane bioreactors (AP-MBRs) under different hydraulic residence times of the ASSR (HRTSR). Three AP-MBRs and an anoxic-aerobic MBR (AO-MBR) showed efficient chemical oxygen demand (>94.2%) and ammonium nitrogen removal (>99.3%). AP-MBRs have higher (p < 0.05) total nitrogen (61.4-67.7%) and total phosphorus (57.5-63.8%) removal than AO-MBRs (47.8 and 47.7%). AP-MBRs achieved sludge reduction efficiencies of 11.8, 31.8, and 36.2% at HRTSR values of 2.5, 5.0, and 6.7 h. Packing carriers greatly improved sludge reduction under low HRTSR and is promising for accelerating sludge reduction in compact space. Metagenomic sequencing analysis showed that genes responsible for metabolism were enriched in AO-MBRs, while genes related to cellular motility and cell signaling were more abundant in the AP-MBRs. A longevity-regulating pathway showed that long lifespan provided more opportunities for worms to prey bacteria. Microscopic examination revealed that some specific protozoa (Arcella, Clathrulina, Aspidisca, Litonotus, Chiloclonella, and Vorticella) and metazoa (Rotaria and Aeolosoma hemprichi) were enriched in ASSRs. Aeolosoma hemprichi was only detected in ASSRs, and unique Cylops appeared on carriers. These results contribute to growing understanding of micrometabolic mechanisms including functional genes and microfauna-driving sludge reduction.

Comparative metabolomics analysis reveals the metabolic regulation mechanism of yellow pigment overproduction by Monascus using ammonium chloride as a nitrogen source.

Monascus yellow pigments (MYPs), as food colorants, are of great interest to the food industry, because of their beneficial biological activities. In this study, a comparative metabolomics strategy revealed the metabolic regulatory mechanism of MYP overproduction, comparing ammonium chloride with peptone as nitrogen sources. Metabolomics-based multivariate regression modeling showed that metabolic biomarkers/modules, such as glucose, lactate, and the pentose phosphate (PP) pathway, were closely associated with the biosynthesis of MYPs. Exogenous addition of glucose increased production of MYPs, whereas lactate reduced it. Inhibition of the PP pathway with dehydroepiandrosterone decreased MYP production, while increasing the shunting production of orange and red pigments. All these treatments significantly changed the expression profiles of the pigment biosynthetic gene cluster and the mycelial morphology. Overall, this study demonstrates the feasibility of elucidating the mechanism of MYP biosynthesis by comprehensive metabolomics analysis, as well as discovering potential engineering targets of efficiency improvements to commercial MYP production. KEY POINTS: • Comparative metabolomics revealed the biomarkers/modules of MYP production. • A rational exogenously adding strategy was implemented to regulate MYP synthesis. • Expression profiles of gene cluster and mycelial morphology were characterized.

High-resolution mass spectrometry-based methodology for the identification of the metabolites of pterostilbene produced by rat, dog and human hepatocytes.

Pterostilbene, a natural bibenzjyl compound, has been demonstrated to have pleiotropic anticancer effects against a variety of cancer types. The aim of this study was carried out to disclose the metabolic profiles of pterostilbene using rat, dog and human hepatocytes. Metabolites were characterized by ultra-high-performance liquid chromatography/quadrupole Orbitrap mass spectrometry with electrospray ionization interface operating in positive ion mode. The structures of the metabolites were proposed by accurate MS, MS/MS spectra and based on their fragmentation patterns. A total of 12 metabolites, including six new ones, were detected and identified. M10 and M12 were unambiguously identified as pinostilbene and 3'-hydroxy-pterostilbene, respectively, by using reference standards. Our results revealed that pterostilbene was metabolized through the following pathways: (a) hydroxylation to form 3'-hydroxy-pterostilbene (M12), which further undergoes glucuronidation (M9), demethylation (M7) and GSH conjugation through the ortho-quinone intermediate; (b) demethylation to produce desmethyl-pterostilbene (M10), which is further subject to glucuronidation (M4); (c) direct conjugation with glucuronide (M11); and (d) direct sulfation (M8). Among the tested species, no significant species difference was observed. The current study provides valuable information on the metabolism of pterostilbene, which is helpful for us to understand the action of this compound.

Salmonella enterica Serovar Typhi Induces Host Metabolic Reprogramming to Increase Glucose Availability for Intracellular Replication.

Salmonella enterica serovar Typhi (S. Typhi) is a human-limited intracellular pathogen and the cause of typhoid fever, a severe systemic disease. Pathogen-host interaction at the metabolic level affects the pathogenicity of intracellular pathogens, but it remains unclear how S. Typhi infection influences host metabolism for its own benefit. Herein, using metabolomics and transcriptomics analyses, combined with in vitro and in vivo infection assays, we investigated metabolic responses in human macrophages during S. Typhi infection, and the impact of these responses on S. Typhi intracellular replication and systemic pathogenicity. We observed increased glucose content, higher rates of glucose uptake and glycolysis, and decreased oxidative phosphorylation in S. Typhi-infected human primary macrophages. Replication in human macrophages and the bacterial burden in systemic organs of humanized mice were reduced by either the inhibition of host glucose uptake or a mutation of the bacterial glucose uptake system, indicating that S. Typhi utilizes host-derived glucose to enhance intracellular replication and virulence. Thus, S. Typhi promotes its pathogenicity by inducing metabolic changes in host macrophages and utilizing the glucose that subsequently accumulates as a nutrient for intracellular replication. Our findings provide the first metabolic signature of S. Typhi-infected host cells and identifies a new strategy utilized by S. Typhi for intracellular replication.

SoxS is a positive regulator of key pathogenesis genes and promotes intracellular replication and virulence of Salmonella Typhimurium.

Salmonella enterica serovar Typhimurium (S. Typhimurium) is an important intracellular pathogen, causing gastroenteritis or severe systemic infection in a variety of hosts. During infection, S. Typhimurium must survive and replicate in host macrophages, which produce abundant oxidative compounds. SoxRS regulon is a well-known regulator that is activated in response to oxidative stress and promotes bacterial tolerance to oxidants in E. coli. However, the global regulatory function of SoxS in S. Typhimurium remains poorly characterized. Here, we used an RNA sequencing-based approach to investigate the role of SoxS in the expression of S. Typhimurium virulence genes. Besides the downregulation of genes related to resistance to oxidative stress, we found that in a soxS deletion mutant the expression of Salmonella pathogenicity island (SPI)-2 genes, which are crucial for replication within macrophages, was significantly repressed. Moreover, immunofluorescence and mice infection experiments showed that soxS deletion inhibited replication in macrophages and decreased virulence upon intraperitoneal inoculation in mice, respectively. Collectively, our findings demonstrate that SoxS is a positive regulator of SPI-2 genes and, therefore, plays a crucial role in S. Typhimurium intracellular replication and virulence.

The putative transcriptional regulator STM14_3563 facilitates Salmonella Typhimurium pathogenicity by activating virulence-related genes.

Salmonella enterica serovar Typhimurium (S. Typhimurium) is an important gram-negative intracellular pathogen that infects humans and animals. More than 50 putative regulatory proteins have been identified in the S. Typhimurium genome, but few have been clearly defined. In this study, the physiological function and regulatory role of STM14_3563, which encodes a ParD family putative transcriptional regulator in S. Typhimurium, were investigated. Macrophage replication assays and mice experiments revealed that S. Typhimurium showed reduced growth in murine macrophages and attenuated virulence in mice owing to deletion of STM14_3563 gene. RNA sequencing (RNA-Seq) data showed that STM14_3563 exerts wide-ranging effects on gene expression in S. Typhimurium. STM14_3563 activates the expression of several genes encoded in Salmonella pathogenicity island (SPI)-6, SPI-12, and SPI-13, which are required for intracellular replication of S. Typhimurium. Additionally, the global transcriptional regulator Fis was found to directly activate STM14_3563 expression by binding to the STM14_3563 promoter. These results indicate that STM14_3563 is involved in the regulation of a variety of virulence-related genes in S. Typhimurium that contribute to its growth in macrophages and virulence in mice.

Myo-inositol utilization by Citrobacter koseri promotes brain infection.

Citrobacter species are opportunistic bacterial pathogens that are implicated in both nosocomial and community-acquired infections. Among the Citrobacter species, Citrobacter koseri is often isolated from clinical material, and it can cause meningitis and brain abscesses in neonates and immunocompromised individuals, thus posing a great threat to human health. However, the virulence determinants of C. koseri remain largely unknown. Myo-inositol is an abundant carbohydrate in the environment and in certain organs of the human body, especially the brain. The C. koseri genome harbors a cluster of genes, QCQ70420.1 to QCQ70429.1 (named the Ino-cluster in this study), which encode IolBCDE, MmsA, and an ATP-binding cassette transporter. The gene cluster may be involved in the utilization of myo-inositol. To investigate the functions of the Ino-cluster in C. koseri, we constructed a mutant strain by deleting the Ino-cluster and found that the mutant could not use myo-inositol as the sole carbon source, confirming that this cluster is responsible for myo-inositol utilization. Moreover, we investigated the function of the Ino-cluster and myo-inositol utilization in C. koseri pathogenicity. Deletion of the Ino-cluster significantly impaired C. koseri colonization of the brain of infected Sprague-Dawley (SD) rats and BALB/c mice, and this increased the survival rate of the infected animals, indicating that the Ino-cluster and the ability to use myo-inositol are essential for C. koseri pathogenicity. Taken together, our findings suggest that using the Ino-cluster products, C. koseri can exploit the abundant myo-inositol in the brain as a carbon source for growth, thus promoting colonization and virulence.