Bacterial Growth Research Articles

Diagnostic and prognostic value of time to positivity in blood cultures. An opinion paper

Time to positivity (TTP) refers to the duration required for a microbiological culture test to indicate a positive result, marking the onset of detectable bacterial or fungal growth in the sample. Numerous variables, including patient characteristics, infection source, former antimicrobial therapy, blood sample volume, and sample transportation time can influence the value of TTP. Several studies have been conducted on bloodstream infections, whereas studies on the clinical significance of yeast TTP are quite limited in the literature. Furthermore, many studies are retrospective and have a small sample size. In this opinion paper, we have formulated some questions and attempted to provide answers based on the available literature and our perspective. The objective of this opinion paper is to summarise current knowledge based on the literature, aiming to offer a critical perspective, particularly on aspects with weaker evidence, which could guide future studies in this area. We believe that TTP of blood cultures appears to exhibit considerable potential and may prove to be a valuable tool in clinical practice for estimating patient mortality risk and guiding antimicrobial therapy choices. Topics discussed include the diagnostic and prognostic role of TTP in Gram-positive and Gram-negative bacteremias and in candidemias, and the significance of differential time to positivity (DTTP). In summary, our opinion is that, based on the available literature, it is not possible to determine whether TTP provides prognostic information, particularly concerning candidemia. Therefore, clinical decisions cannot be systematically based on this parameter.

Insights into interspecies protein binding variability using clindamycin as an example

Abstract Background In the preclinical development of new drugs, animal models are often employed to predict their efficacy in humans, relying on translational pharmacokinetic/pharmacodynamic (PK/PD) studies. We performed in vitro experiments focusing on the comparison of plasma protein binding (PPB) and bacterial growth dynamics of clindamycin, a commonly used antimicrobial agent, across a range of drug concentrations and plasma environments. Methods Human, bovine and rat plasma were used for determining PPB of clindamycin at various antibiotic concentrations in buffer and media containing 20% to 70% plasma or pure plasma using ultrafiltration (UF) and equilibrium dialysis (ED). Also bacterial growth and time–kill assays were performed in Mueller–Hinton broth (MHB) containing various percentages of plasma. Results Protein binding of clindamycin correlated well between UF and ED. Notably, clindamycin exhibited substantially lower protein binding to rat plasma compared with human and bovine plasma. Staphylococcus aureus growth was significantly reduced in 70% human, bovine, and rat plasma after 4, 8 and 24 h compared with standard MHB. Time–kill data demonstrated that bacterial counts at both 20% and 70% plasmas were less when compared with MHB at drug concentrations lower than MIC after 4 and 8 h of incubation. For rat plasma, the difference was maintained over 24 h of incubation. Furthermore, a complete bacterial killing at 16 mg/L was observed after 24 h in 20% and 70% human and bovine plasma, but not for rat plasma. Conclusions Recognizing interspecies differences in PB might be essential for optimizing the translational relevance of preclinical studies.

Isolation of Bacillus Velezensis from Leepuram Sea Sediment and Its Bioactive Compound and Antimicrobial Activity

Research into marine bacteria in India for their potential active chemicals is essential for the creation of medications derived from marine sources. The objective of this research was to collect Bacillus velezensis from Leepuram Sea Sediment, test their antibacterial efficacy with the agar diffusion method, and find secondary metabolites by GC-MS spectroscopy.Traditional microbiological methods and phylogenetic analysis of 16S rRNA sequences were used to successfully identify Bacillus velezensis from sediment samples obtained in the Leepuram Sea, Kanyakumari. Using the agar well diffusion method, we tested the isolated metabolites for their antibacterial activity against Gram-positive and Gram-negative bacteria. The results showed that the extract had a strong antibacterial effect, inhibiting the growth of harmful bacteria with zones of inhibition as large as 19 mm for E. coli and Pseudomonas.Bacillus velezensis produces secondary metabolites with pharmacological potential, according to this study. These metabolites may have useful uses in biocontrol, agriculture, and medicine. The implications of these results for human health and the creation of new antimicrobial drugs are likely to grow substantially when more worldwide research is carried out.

Model-informed drug development for antimicrobials: translational pharmacokinetic-pharmacodynamic modelling of apramycin to facilitate prediction of efficacious dose in complicated urinary tract infections.

The use of mouse models of complicated urinary tract infection (cUTI) has usually been limited to a single timepoint assessment of bacterial burden. Based on longitudinal in vitro and in vivo data, we developed a pharmacokinetic-pharmacodynamic (PKPD) model to assess the efficacy of apramycin, a broad-spectrum aminoglycoside antibiotic, in mouse models of cUTI. Two Escherichia coli strains were studied (EN591 and ATCC 700336). Apramycin exposure-effect relationships were established with in vitro time-kill data at pH 6 and pH 7.4 and in mice with cUTI. Immunocompetent mice were treated with apramycin (1.5-30 mg/kg) starting 24 h post-infection. Kidney and bladder tissue were collected 6-96 h post-infection for cfu determination. A PKPD model integrating all data was developed and simulations were performed to predict bacterial burden in humans. Treatment with apramycin reduced the bacterial load in kidneys and bladder tissue up to 4.3-log compared with vehicle control. In vitro and in vivo tissue time-course efficacy data were integrated into the PKPD model, showing 76%-98% reduction of bacterial net growth and 3- to 145-fold increase in apramycin potency in vivo compared with in vitro. Simulations suggested that an 11 mg/kg daily dose would be sufficient to achieve bacterial stasis in kidneys and bladder in humans. PKPD modelling with in vitro and in vivo PK and PD data enabled simultaneous evaluation of the different components that influence drug effect, an approach that had not yet been evaluated for antibiotics in the cUTI model and that has potential to enhance model-informed drug development of antibiotics.

A Comprehensive Review on the Antibacterial, Antifungal, Antiviral, and Antiparasitic Potential of Silybin

Silybin, a flavonolignan extracted from the seeds of the plant species Silybum marianum (L.) Gaertn., has a variety of pharmacological activities, including antimicrobial activity against several microorganisms of clinical interest. This review analyzes the existing studies on silybin’s antimicrobial activity and possible mechanisms of action. Silybin has been shown to inhibit the growth of Gram-positive and Gram-negative bacteria, as well as some fungi, viruses, and protozoa. In general, possible mechanisms of antimicrobial action include the inhibition of efflux pumps, prevention of biofilm formation, reduction of the expression of virulence factors, induction of apoptosis-like effects, and plasma membrane damage, as well as the inhibition of nucleic acid and protein synthesis. Silybin has been shown to have synergistic effects when combined with conventional antibiotics against both drug-sensitive and drug-resistant microorganisms. However, the low bioavailability observed for this flavonolignan has been a challenge to its clinical use. In this context, nanotechnology has been used to increase silybin’s bioavailability while enhancing its antimicrobial activity. Furthermore, certain structural modifications have been able to enhance its antimicrobial activity in comparison to that of the natural molecule. Overall, this review provides insights into the scientific understanding of the mechanism of action of silybin and its desired properties for the effective treatment of infections.

Double-Layered Microneedle Patch Integrated with Multifunctional Nanoparticles and Live Bacteria for Long-Term Treatment of Atopic Dermatitis.

Atopic dermatitis (AD) is a complex and prevalent chronic inflammatory skin disease that impacts a significant portion of the global population. Conventional treatments often focus on a singular pathogenic factor or suffer from limited skin penetration, resulting in unsatisfactory outcomes. Here, a multifunctional double-layered microneedle (MN) patch is proposed for long-term and effective treatment of AD by integrating therapeutic nanoparticles (NPs) and live bacteria. In the design, the MN tips are loaded with Prussian blue NPs encapsulating cetirizine hydrochloride (CET@PB NPs), while the patch backing incorporates Bacillus subtilis (B. subtilis). Upon skin insertion, the MN patch efficiently delivers CET@PB NPs into the skin and deposits live B. subtilis on the skin surface after fast dissolution. The delivered NPs not only scavenge reactive oxygen species (ROS) and improve oxidative stress microenvironments in the AD lesions, but also provide sustained release of the antihistamine CET in the skin for alleviating AD symptoms. Furthermore, B. subtilis survives on the skin for over 9 days and effectively inhibits the growth of the harmful bacteria Staphylococcus aureus. These features highlight the superior efficacy of the MN patch in long-term treatment of AD, offering a promising alternative for the management of skin disorders in clinics.

Combining the Powerful Antioxidant and Antimicrobial Activities of Pomegranate Waste Extracts with Whey Protein Coating-Forming Ability for Food Preservation Strategies

Different solvents water, ethanol and ethanol/water (6:4 v/v), were compared in the extraction of pomegranate peels and seeds (PPS) in terms of recovery yields, antioxidant properties, and antimicrobial action against typical spoilage bacterial and fungal species. The best performing extract (ethanol/water (6:4 v/v) was shown to contain mostly ellagic acid and punicalagin as phenolic compounds (5% overall) and hydrolysable tannins (16% as ellagic acid equivalents) and was able to inhibit the growth of the acidophilic Alicyclobacillus acidoterrestris at a concentration as low as 1%. The preservation of the organoleptic profile of A. acidoterrestris-inoculated apple juice with extract at 1% over 20 days was also observed thanks to the complete inhibition of bacterial growth, while the extract at 0.1% warranted a significant (40%) inhibition of the enzymatic browning of apple smoothies over the first 30 min. When incorporated in whey proteins’ isolate (WPI) at 5% w/w, the hydroalcoholic extract conferred well appreciable antioxidant properties to the resulting coating-forming hydrogel, comparable to those expected for the pure extract considering the amount present. The WPI coatings loaded with the hydroalcoholic extract at 5% were able to delay the browning of cut fruit by ca. 33% against a 22% inhibition observed with the sole WPI. In addition, the functionalized coating showed an inhibition of lipid peroxidation of Gouda cheese 2-fold higher with respect to that observed with WPI alone. These results open good perspectives toward sustainable food preservation strategies, highlighting the potential of PPS extract for the implementation of WPI-based active packaging.

Impact of chemical oxygen demand to nitrogen ratio on ANAMMOX bacterial growth in an up-flow anaerobic sludge blanket reactor

Impact of chemical oxygen demand to nitrogen ratio on ANAMMOX bacterial growth in an up-flow anaerobic sludge blanket reactor

Intercellular mosaic methylation in fast-growing Mycobacterium tuberculosis clinical isolates

Abstract Orphan DNA adenine methyltransferases (MTases) of Mycobacterium tuberculosis (Mtb) exhibit diversity across clinical isolates, but the forces driving this variation are not entirely clear. Recently, we observed several isolates exhibiting anomalous hypomethylation by Type I MTase Mycobacterial Adenine Methyltransferase C (MamC) despite a wild-type mamC genotype (‘MamC-anomalous’ isolates). Investigating this hypomethylation through multiple analyses revealed three key findings. First, heterogeneity analysis revealed intercellular mosaic methylation (IMM) in MamC-anomalous isolates. While they often exhibit phase-variable heterogeneity, this is the first report of IMM by a prokaryotic Type I MTase. Second, MamC-anomalous isolates exhibited a large, stable difference in chromosome copy number along the replication axis (a proxy for bacterial growth rate), suggesting a distinct growth phase accompanied by MamC hypomethylation. Third, MamC methylation efficiency decreased progressively with distance from origin of replication on both strands, with a marked exaggeration in MamC-anomalous isolates. In contrast, other Mtb MTases (MamA and MamB) exhibited lower methylation levels away from the origin only on the lagging strand, and without exaggeration in MamC-anomalous isolates. We conclude that, among Mtb MTases, MamC is uniquely linked to growth dynamics.

Characterization and genomic analysis of bacteriophage VT223 infecting Salmonella enterica subsp enterica serovar Enteritidis

Bacteriophages are increasingly considered a promising alternative to antibiotics for treating bacterial infections. For the bacteriophage VT223, which was isolated from shrimp pond wastewater, a thorough analysis of its host range, morphology, and genome sequencing was performed. Bacteriophage VT223 exhibits high specificity towards Salmonella spp. strains, highlighting its potential as a targeted therapy for Salmonella spp. infections. Electron microscopy revealed that VT223 belongs to the Caudoviricetes, Jerseyvirus, with an icosahedral head and a non-contractile tail. This phage can infect three species within the Salmonella spp., with a short latent period of 30 minutes and a burst size of 446 PFU/infected cells. Genome sequencing revealed a 43,062 bp double-stranded DNA genome with a GC content of 49.6%. Stability tests showed that VT223 is stable over various temperatures and pH levels. Biofilm formation inhibition testing revealed that phage VT223 reduced biofilm by up to 57.7% after a four-hour treatment. In vitro studies demonstrated the lytic activity of VT223 against Salmonella enterica subsp enterica serovar Enteritidis ATCC 49223. These findings provide valuable insights into the biological properties of bacteriophage VT223 and its potential use as a biocontrol agent in livestock production and aquaculture to combat bacterial growth. Published on November 15, 2024

Low-cost, real-time detection of bacterial growth via diffraction-based sensing.

The emergence of antibacterial resistance impacts healthcare networks globally, with mortality rates and linked burdens of infection disproportionately affecting the developing world. Rapid alternatives to antibiotic susceptibility testing (AST) allow for swifter, more effective treatment, though they are limited in use in low-resource settings due to significant cost barriers. Herein we demonstrate a simple, cost-effective diffraction sensing-based approach for rapidly detecting bacterial growth (a precursor to AST). Diffraction gratings (1D, lined) directly comprised of our test bacteria (Escherichia coli DH5α) were produced using soft agar-based gel templates designed to direct bacterial attachment and produce a near-zero background signal. The diffraction spot intensities from the live bacterial gratings were monitored in growth and no growth (ampicillin) conditions at room temperature, using a simple fixed laser and photodetector setup. Growth-induced differences in signal were observed within 10-20 minutes, highlighting the sensitivity of this approach and its potential to be adapted as a rapid and accessible AST alternative.

Gallic acid functionalized silk fibroin/gelatin composite wound dressing for enhanced wound healing

As the incidence of chronic wounds increases, the requirements for wound dressings are rising. The specific aim of this study is to propose a novel gallic acid (GA) functionalized silk fibroin (SF) and gelatin (Gel) composite wound dressing in which GA is used as an antibacterial and wound healing substance. Via electrospinning, SF, Gel, and GA mixed solutions could be conveniently fabricated into a composite nanofiber mat (SF-Gel-GA), consisting of uniform fibers with an average diameter around 134.57 ± 84 nm. The internal mesh structure of SF-Gel-GA provides sufficient drug loading capacity, proper moisture permeability, and proper degradation rate. SF-Gel-GA presents excellent biocompatibility. NIH-3T3 fibroblast cells could adhere and spread stably on the SF-Gel-GA surface with slightly promoted proliferation. In the presence of SF-Gel-GA, the growth of both Gram-positive and Gram-negative bacteria, includingStaphylococcus aureusandPseudomonas aeruginosa, is significantly inhibited in both plate and suspension cultures. A cutaneous excisional mouse wound model proves the efficient ability of SF-Gel-GA to promote wound healing. Compared with pure SF dressing and commercial Tegaderm Hydrocolloid3Mdressing, the wound closure rate with SF-Gel-GA treatment is significantly improved. The histological assessments further demonstrate SF-Gel-GA could facilitate collagen deposition, neovascularization, and epithelialization at wound sites to promote wound healing. In conclusion, a novel SF-Gel-GA composite wound dressing with efficient wound healing activities have been developed for chronic wound treatment with broad healing potential.

Carbon-mediated modulation pathways of phytotoxicity in chicken manure composting.

Compost phytotoxicity affects the safety of organic fertilizers returned to the field, thus hindering the breeding cycle, so it is essential to reduce the compost phytotoxicity. The phytotoxicity of compost was estimated utilizing the germination index (GI) and the aqueous substances (organics and ions) present in compost correlated closely with GI. This study assessed the effect of carbon additives from different plant sources (mushroom substrates (MS), cornstalks (CS) and garden substrates (GS)) on maturity parameters (temperature, pH, EC, C/N), content of aqueous carbon and nitrogen matters, salt ions, heavy metal ions, and microbiome of piles when composting with chicken manure and especially focused on their effect on GI. Results showed that all additives significantly improved GI (85.25%-106.28%). The primary factors influencing seed germination were Mg2+ and SO42- in CM compost, acetic acid and NH4+ in CM+MS compost, humic acid in CM+CS compost, and dissolved total nitrogen in CM+GS compost. During composting, the growth of heavy metal passivating bacteria (Bacillus) and organic matter degrading bacteria (Desemzia and Turicibacter) can be promoted by decreasing aqueous carbon and nitrogen substances (volatile fatty acids, NH4+, dissolved total nitrogen, amino acids) and increasing the content of humic acid, which improved the composting environment and provided favourable conditions for the germination of seeds, thereby increasing GI. Therefore, GS showed the best potential for accelerating degradation of organic matter and improving GI during composting with chicken manure.

Towards Bacterial Resistance via the Membrane Strategy: Enzymatic, Biophysical and Biomimetic Studies of the Lipid cis-trans Isomerase of Pseudomonas aeruginosa.

The lipid cis-trans isomerase (Cti) is a periplasmic heme-c enzyme found in several bacteria including Pseudomonas aeruginosa, a pathogen known for causing nosocomial infections. This metalloenzyme catalyzes the cis-trans isomerization of unsaturated fatty acids in order to rapidly modulate membrane fluidity in response to stresses that impede bacterial growth. As a consequence, breakthrough in the elucidation of the mechanism of this metalloenzyme might lead to new strategies to combat bacterial antibiotic resistance. We report the first comprehensive biochemical, electrochemical and spectroscopic characterization of a Cti enzyme. This has been possible by the successful purification of Cti from P. aeruginosa (Pa-Cti) in favorable yields with enzyme activity of 0.41 µmol/min/mg when tested with palmitoleic acid. Through a synergistic approach involving enzymology, site-directed mutagenesis, Raman spectroscopy, Mössbauer spectroscopy and electrochemistry, we identified the heme coordination and redox state, pinpointing Met163 as the sixth ligand of the FeII of heme-c in Pa-Cti. Significantly, the development of an innovative assay based on liposomes demonstrated for the first time that Cti catalyzes cis-trans isomerization directly using phospholipids as substrates without the need of protein partners, answering the important question about the substrate of Cti within the bacterial membrane.

Combining multiple stressors blocks bacterial migration and growth.

In nature, organisms experience combinations of stressors. However, laboratory studies use batch cultures, which simplify reality and focus on population-level responses to individual stressors.1,2,3,4,5 In recent years, bacterial stress responses have been examined with single-cell resolution using microfluidics.6,7,8,9,10,11,12 Here, we use a microfluidic approach to simultaneously provide a physical stressor (shear flow) and a chemical stressor (H2O2) to the human pathogen Pseudomonas aeruginosa. By treating cells with levels of flow and H2O2 that commonly co-occur in human host tissues,13,14,15,16,17,18 we discover that previous reports significantly overestimate the H2O2 levels required to block bacterial growth. Specifically, we establish that flow increases H2O2 effectiveness 50-fold, explaining why previous studies lacking flow required much higher concentrations. Using natural H2O2 levels, we identify the core H2O2 regulon, characterize OxyR-mediated dynamic regulation, and demonstrate that multiple H2O2 scavenging systems have redundant roles. By examining single-cell behavior, we serendipitously discover that the combined effects of H2O2 and flow block pilus-driven surface migration. Thus, our results counter previous studies and reveal that natural levels of H2O2 and flow synergize to restrict bacterial motility and survival. By studying two stressors at once, our research highlights the limitations of oversimplifying nature and demonstrates that physical and chemical stress can combine to yield unpredictable effects.

Bacillus xiamenensis Inhibits the Growth of Moraxella osloensis by Producing Indole-3-Carboxaldehyde.

Moraxella osloensis, a gram-negative rod-shaped bacterium found on human skin, produces 4-methyl-3-hexenoic acid, contributing to clothing and body malodor. M. osloensis is resistant to UV light, drying, and antimicrobials, making its eradication challenging. As the skin is low in nutrients, commensal bacteria compete for resources and use diverse strategies to inhibit their competitors. Therefore, skin-derived bacteria that exhibited growth-inhibitory activity against M. osloensis were searched. Screening skin-derived bacteria using a coculture halo assay revealed that Bacillus xiamenensis formed an inhibition zone with M. osloensis. Coculture plates were extracted with ethyl acetate and fractionated using a silica gel column and preparative thin-layer chromatography to isolate the active compound from the B. xiamenensis metabolites. Nuclear magnetic resonance spectroscopy identified the active compound as indole-3-carboxaldehyde, which has low toxicity in humans. At soluble concentrations, indole-3-carboxaldehyde does not inhibit the growth of other bacteria, such as Staphylococcus aureus, Escherichia coli, and Bacillus subtilis, suggesting M. osloensis is highly sensitive to indole-3-carboxaldehyde. These findings highlight B. xiamenensis as a promising candidate for the development of a skin probiotic to promote skin health and combat malodor-causing bacteria.

Effects of Inoculation of Thermotolerant Bacillus Strains on Lignocellulose Degradation

Thise study investigated the effect of three lignocellulolytic thermophilic Bacillus strains (F11, Q1, and FP4) on lignocellulose degradation, enzymatic activities, and microbial community structure in composting. The lignin degradation rate reached 36% in the presence of the inoculant, the hemicellulose degradation rate ranged from 43% (F11) to 51% (Q1), and cellulose degradation rates reached 57% in F11 and in FP4, which were significantly higher than the control (CK). The inoculation treatment could explain 28% of the lignin degradation for all three strains. The contribution of FP4 to hemicellulose and cellulose degradation was 30% and 20%, respectively. Compared to CK, lignin peroxidase activity in the water extract of the compost had increased by 66~145% for inoculation treatments, and manganese peroxidase and laccase activity increased by 114% and 78% for Q1. The inoculation stimulated the growth of indigenous bacteria with stronger lignocellulolytic enzyme-producing ability; such shifts in microbial communities were most likely responsible for the improved lignocellulose degradation.

Presence of lactic acid bacteria in hydrogen production by dark fermentation: competition or synergy.

Dark fermentation in mixed cultures has been extensively studied due to its great potential for sustainable hydrogen production from organic wastes. However, microbial composition, substrate competition, and inhibition by fermentation products can affect hydrogen yield and production rates. Lactic acid bacteria have been identified as the key organisms in this process. On one hand, lactic acid bacteria can efficiently compete for carbohydrate rich substrates, producing lactic acid and secreting bacteriocins that inhibit the growth of hydrogen-producing bacteria, thereby decreasing hydrogen production. On the other hand, due to their metabolic capacity and synergistic interactions with certain hydrogen-producing bacteria, they contribute positively in several ways, for example by providing lactic acid as a substrate for hydrogen generation. Analyzing different perspectives about the role of lactic acid bacteria in hydrogen production by dark fermentation, a literature review was done on this topic. This review article shows a comprehensive view to understand better the role of these bacteria and their influence on the process efficiency, either as competitors or as contributors to hydrogen production by dark fermentation.

Enhancement of anaerobic digestion by adding elemental sulfur.

In this study, a new approach to enhance methane (CH4) production from organic substrates in anaerobic digestion (AD) has been discovered. That is, the addition of elemental sulfur (S0) particles into the AD system promotes the synergistic growth of elemental sulfur disproportionation bacteria, acidogenic bacteria and methanogenic archaea, thus facilitating hydrolysis, acidogenesis and methanogenesis. The efficacy of this AD enhancement pathway was confirmed in AD experiments with glucose as a model organic substrate. The results demonstrated that CH4 production in the AD system increased considerably with S0 dosages ranging from 20 mg/L to 300 mg/L. Two gas production peaks appeared at dosages of 20 mg/L and 180 mg/L, where the total CH4 production increased by 2.1 times and 2.5 times, respectively compared with the control group. However, inhibitory effect was observed for S0 dosages above 300 mg/L. The chemical states of S, the microbial community and the abundance of key functional enzymes in the AD system were analyzed. The results showed that S0 addition increased the relative abundance of Dethiobacteraceae, Caldatribacterium, Anaerolineaceae, Methanobacterium and Methanosaeta and considerably increased the abundance of key functional enzymes, such as dehydrogenase, D-glucosidic glucosidase, pyruvate synthase and acetyl-CoA deacetylase. The enrichment of these microorganisms and functional enzymes was strongly positively correlated with the production of volatile fatty acids and CH4, demonstrating that S0 addition effectively enhances methanogenesis during AD.

Geology of Mudstones Atop the Sangonghe Formation, Shuixigou Group, Turpan‐Hami Basin: New Insights and Their Implications for Petroleum Geology

ABSTRACTAs the exploration of deep‐seated oil and gas resources in the Turpan‐Hami Basin intensifies, there is an urgent need to thoroughly depict the geological characteristics of newly uncovered, underexplored strata in sub‐sag centers. This study undertakes the inaugural systematic geochemical analysis of mudstones atop the Sangonghe Formation, including palynological identification, maceral identification, biomarker analysis, total organic carbon (TOC) analysis, and pyrolysis. The findings reveal that the mudstone sequence, as the target layer, was deposited in a warm and moist paleoclimate and a weakly reducing to weakly oxidising saline water environment, fostering the growth of prolific algae and bacteria, thus ensuring substantial foundational materials for source rock formation. The organic matter in the mudstone sequence displays pronounced laminar accumulation. Despite the overall modest abundance, the organic matter features notable hydrocarbon‐generating potential per unit of organic carbon and favourable types. Humic‐sapropelic kerogens (type II1) are found in the mudstones, with organic matter generally reaching a mature to highly mature stage. These characteristics establish the mudstones as effective source rocks, furthermore, the hydrocarbon expulsion in the Xishanyao Formation precedes that of the Sangonghe Formation and that both formations constitute a sequential process in terms of hydrocarbon generation and expulsion timing and hydrocarbon contribution. Reanalyzing this mudstone sequence not only revises prior geological understanding of it as direct cap rocks, but also facilitates the reclassification of deep‐seated strata into three distinct petroleum systems. Centered around this source rock layer, dual modes, namely the “lower‐source rock and upper‐reservoir” and the “ lower‐reservoir and upper‐source rock” modes can be formed. These new insights will offer profound implications for hydrocarbon resource evaluation and future hydrocarbon exploration endeavours in the Shuixigou Group within the Taibei sag.