Pathogen Streptococcus Research Articles

Purification, Characterization and Antimicrobial Properties of Hemolymph Lectin from the Larva of Red Palm Weevil, Rhynchophorus ferrugineus

Lectins are renowned hemagglutinins and multivalent proteins with a well-known quality for sugar-binding specificity that participate significantly in invertebrate defense functions. Studies on biological activity of lectin from coleopteran insect are very scarce. A lectin with specific affinity for N-acetyl neuraminic acid was purified from the hemolymph of the larva of the red palm weevil, Rhynchophorus ferrugineus by biospecific adsorption using formalinized rabbit erythrocytes and affinity chromatography using PSM-linked cyanogen bromide activated Sepharose 4B. The specific activity of the lectin purified by affinity chromatography was much higher than the lectin purified by biospecific adsorption. The binding specificity of the weevil lectin distinguishes it from other known insect lectins. Like the crude agglutinin, the lectin purified by affinity chromatography also showed the same pattern of specificity towards erythrocytes. However, 4-to-8-fold decrease in HA titer was observed when tested with the purified lectin. In the same way, reduction is also observed in the HAI titer of the purified lectin with most of the inhibitors except PSM where the HAI titer was identical both in the crude agglutinin and purified lectin. Sugars N-acetyl neuraminic acid, N-acetyl mannosamine and N-acetyl-D-galactosamine inhibited the HA titer of the purified lectin with greater efficacy than the crude agglutinin. The sialic acid specificity of the lectin was confirmed by 16-fold reduction in HA titer with asialo rabbit erythrocytes and 32-fold reduction in HAI titer with desialylated PSM. The purified lectin is homogenous on sodium dodecyl sulphate-polyacrylamide electrophorogram with a molecular weight of about 60 kDa. The lectin showed antimicrobial activity against pathogenic bacteria Streptococcus mutans, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeroginosa and fungi Candida albicans and Aspergillus niger.

Complete genome sequencing and comparative genomic analysis of three donkey Streptococcus equi subsp. equi isolates.

Streptococcus equi subspecies equi (S. equi) is the causative agent of strangles, which is one of the most common and highly contagious respiratory infectious illnesses in horses. Streptococcus equi (S. equi) is a horse-specific pathogen that originated from the closely related zoonotic pathogen Streptococcus equi subspecies zooepidemicus (S. zooepidemicus). Despite decades of research, the movement of genetic material across host-restricted diseases remains a mystery. Three S. equi donkey isolates (HTP133, HTP232, and HT1112) were recently isolated from a strangles epidemic on donkey farms in China's Xinjiang Province. In this study, we performed a comprehensive comparative analysis of these isolates using whole genome sequencing and compared them to the published genomic sequences of equine strain S. equi 4047 to uncover evidence of genetic events that shaped the evolution of these donkey S. equi isolates' genomes. Whole genome sequencing indicated that both strains were closely related, with comparable gene compositions and a high rate of shared core genomes (1788-2004). Our comparative genomic study indicated that the genome structure is substantially conserved across three donkey strains; however, there are several rearrangements and inversions when compared to the horse isolate S. equi 4047. The virulence factors conveyed by genomic islands and prophages, in particular, played a key role in shaping the pathogenic capacity and genetic diversity of these S. equi strains. Furthermore, we discovered that the HT133 isolate had a strong colonization ability and increased motility; the HT1112 isolates had a significantly higher ability for antimicrobial resistance and biofilm formation, and the HT232 isolate gained pathogenic specialization by acquiring a bacteriophage encoding hyaluronate lyase. In summary, our findings show that genetic exchange across S. equi strains influences the development of the donkey S. equi genome, offering important genetic insights for future epidemiological studies of S. equi infection.

Green synthesis of TiO2 nanoparticles using Terminalia chebula and its potent antibiofilm activity against dental caries-associated bacterium Streptococcus mutans

The production of eco-friendly nanoparticles with biocompatible properties is gaining prominence in global medical research. In this study, we explore the antibiofilm potential of Titanium dioxide nanoparticles (TiO2NPs) synthesized from Terminalia chebula against the oral pathogen Streptococcus mutans. Characterization of TiO2NPs revealed a UV–Vis peak at 322 nm. FT-IR spectrum identified Ti–O–Ti vibrations, hydroxyl, and carboxylic groups. SEM analysis revealed tetragonal TiO2NPs, with EDX confirming titanium. XRD confirmed TiO2 anatase crystallographic planes at 25.8° with an average grain size of the particle of 56 nm. Biofilm inhibitory activity indicated significant inhibition of biofilm with BIC50 of 65.71 μg/ml. Microscopic visualization substantiated the efficiency of TiO2NPs in impeding biofilm formation. Increased TiO2NPs concentrations showed a marked reduction in exopolysaccharide and increased protein leakage from S. mutans after prolonged exposure. These findings suggest TiO2NPs as potential agents for treating dental caries, though further validation is needed for therapeutic use in oral care.

Immunobiotic Ligilactobacillus salivarius FFIG58 Confers Long-Term Protection against Streptococcus pneumoniae.

Previously, we isolated potentially probiotic Ligilactobacillus salivarius strains from the intestines of wakame-fed pigs. The strains were characterized based on their ability to modulate the innate immune responses triggered by the activation of Toll-like receptor (TLR)-3 or TLR4 signaling pathways in intestinal mucosa. In this work, we aimed to evaluate whether nasally administered L. salivarius strains are capable of modulating the innate immune response in the respiratory tract and conferring long-term protection against the respiratory pathogen Streptococcus pneumoniae. Infant mice (3-weeks-old) were nasally primed with L. salivarius strains and then stimulated with the TLR3 agonist poly(I:C). Five or thirty days after the last poly(I:C) administration mice were infected with pneumococci. Among the strains evaluated, L. salivarius FFIG58 had a remarkable ability to enhance the protection against the secondary pneumococcal infection by modulating the respiratory immune response. L. salivarius FFIG58 improved the ability of alveolar macrophages to produce interleukin (IL)-6, interferon (IFN)-γ, IFN-β, tumor necrosis factor (TNF)-α, IL-27, chemokine C-C motif ligand 2 (CCL2), chemokine C-X-C motif ligand 2 (CXCL2), and CXCL10 in response to pneumococcal challenge. Furthermore, results showed that the nasal priming of infant mice with the FFIG58 strain protected the animals against secondary infection until 30 days after stimulation with poly(I:C), raising the possibility of using nasally administered immunobiotics to stimulate trained immunity in the respiratory tract.

Bacterial second messenger cyclic di-AMP in streptococci.

Cyclic dimeric adenosine monophosphate (c-di-AMP) has been well studied in bacteria, including those of the genus Streptococcus, since the first recognition of this dinucleotide in 2008. Streptococci possess a sole diadenylate cyclase, CdaA, and distinct c-di-AMP phosphodiesterases. Interestingly, cdaA is required for viability of some streptococcal species but not all when streptococci are grown in standard laboratory media. Bacteria of this genus also have distinct c-di-AMP effector proteins, diverse c-di-AMP-signaling pathways, and subsequent biological outcomes. In streptococci, c-di-AMP may influence bacterial growth, morphology, biofilm formation, competence program, drug resistance, and bacterial pathogenesis. c-di-AMP secreted by streptococci has also been shown to interact with the mammalian host and induces immune responses including type I interferon production. In this review, we summarize the reported c-di-AMP networks in seven species of the genus Streptococcus, which cause diverse clinical manifestations, and propose future perspectives to investigate the signaling molecule in these streptococcal pathogens.

SIgA structures bound to Streptococcus pyogenes M4 and human CD89 provide insights into host-pathogen interactions

Immunoglobulin (Ig) A functions as monomeric IgA in the serum and Secretory (S) IgA in mucosal secretions. Host IgA Fc receptors (FcαRs), including human FcαR1/CD89, mediate IgA effector functions; however, human pathogen Streptococcus pyogenes has evolved surface-protein virulence factors, including M4, that also engage the CD89-binding site on IgA. Despite human mucosa serving as a reservoir for pathogens, SIgA interactions with CD89 and M4 remain poorly understood. Here we report cryo-EM structures of M4-SIgA and CD89-SIgA complexes, which unexpectedly reveal different SIgA-binding stoichiometry for M4 and CD89. Structural data, supporting experiments, and modeling indicate that copies of SIgA bound to S. pyogenes M4 will adopt similar orientations on the bacterium surface and leave one host FcαR binding site open. Results suggest unappreciated functional consequences associated with SIgA binding to host and bacterial FcαRs relevant to understanding host-microbe co-evolution, IgA effector functions and improving the outcomes of group A Streptococcus infection.

Dietary Administration Effects of Exopolysaccharide Produced by Bacillus tequilensis PS21 Using Riceberry Broken Rice, and Soybean Meal on Growth Performance, Immunity, and Resistance to Streptococcus agalactiae of Nile tilapia (Oreochromis niloticus).

Overuse of antibiotics in aquaculture has generated bacterial resistance and altered the ecology. Aquacultural disease control requires an environmentally sustainable approach. Bacterial exopolysaccharides (EPSs) as bioimmunostimulants have not been extensively explored in aquaculture. This study investigated EPS produced from 5% w/v riceberry broken rice as a carbon source and 1% w/v soybean meal as a nitrogen source by Bacillus tequilensis PS21 from milk kefir grain for its immunomodulatory, antioxidant activities and resistance to pathogenic Streptococcus agalactiae in Nile tilapia (Oreochromis niloticus). The FTIR spectrum of EPS confirmed the characteristic bonds of polysaccharides, while the HPLC chromatogram of EPS displayed only the glucose monomer subunit, indicating its homopolysaccharide feature. This EPS (20 mg/mL) exhibited DPPH scavenging activity of 65.50 ± 0.31%, an FRAP value of 2.07 ± 0.04 mg FeSO4/g DW, and antimicrobial activity (14.17 ± 0.76 mm inhibition zone diameter) against S. agalactiae EW1 using the agar disc diffusion method. Five groups of Nile tilapia were fed diets (T1 (Control) = 0.0, T2 = 0.1, T3 = 0.2, T4 = 1.0, and T5 = 2.0 g EPS/kg diet) for 90 days. Results showed that EPS did not affect growth performances or body composition, but EPS (T4 + T5) significantly stimulated neutrophil levels and serum lysozyme activity. EPS (T5) significantly induced myeloperoxidase activity, catalase activity, and liver superoxide dismutase activity. EPS (T5) also significantly increased the survival of fish at 80.00 ± 5.77% at 14 days post-challenge with S. agalactiae EW1 compared to the control (T1) at 53.33 ± 10.00%. This study presents an efficient method for utilizing agro-industrial biowaste as a prospective source of value-added EPS via a microbial factory to produce a bio-circular green economy model that preserves a healthy environment while also promoting sustainable aquaculture.

Development of engineered endolysins with in vitro intracellular activity against streptococcal bovine mastitis-causing pathogens.

Bacteriophage-derived endolysins are a novel class of antimicrobials known to rapidly kill bacteria, including antibiotic-resistant strains. We here engineered endolysins against the bovine mastitis pathogens Streptococcus uberis, Streptococcus agalactiae and Streptococcus dysgalactiae, also targeting intracellular survival and biofilm formation. For this purpose, high-throughput DNA assembly was used to create a library with >80,000 theoretical endolysin variants for screening of their bacteriolytic activity against Gram-positive isolates from (sub)clinically affected cows. This lytic activity was evaluated by turbidity reduction and time-kill assays in phosphate-buffered saline and pasteurized whole cow's milk to allow a rank up of the most potent leading candidates. A top candidate was selected with a 4.0 log killing efficacy against S. uberis, also showing similar activity against S. agalactiae and S. dysgalactiae. This top candidate eradicated S. uberis biofilm and showed intracellular activity in two bovine mammary epithelial cell lines as was confirmed by confocal microscopy. A potentiating effect on cloxacillin, a beta-lactam penicillin used to intramammarily treat bovine Gram-positive mastitis, was observed for this top candidate endolysin in raw cow's milk from (sub)clinically infected udders. Our in vitro results indicate that engineered endolysins may have a future role as add-on in the treatment of bovine streptococcal mastitis.

Streptococcus pyogenes emm98.1 variants activate inflammatory caspases in human neutrophils

ABSTRACT The human pathogen Streptococcus pyogenes (Group A Streptococcus, GAS) is responsible for invasive disease characterized by inflammation and tissue destruction. Inflammatory symptoms of invasive disease may be attributed to the neutrophil response during the early stages of infection. Here, the human neutrophil response to GAS was characterized in vitro using emm98.1 type covS mutant GAS strain NS88.2 (isolated from invasive infection) and avirulent variant NS88.2rep. NS88.2 was shown to resist phagocytic killing and proliferate in the presence of human neutrophils, where neutrophil antimicrobial defence through the production of reactive oxygen species was reduced compared with NS88.2rep. In the presence of NS88.2, neutrophil death was delayed compared with NS88.2rep. Infection with either GAS strain induced expression of inflammatory caspases-1 and -4 in neutrophils, with increased detection of activated inflammatory caspases in response to NS88.2rep compared with NS88.2. NS88.2 infection caused differential expression of cell-surface CD66b, CD16, and CD31, when compared to NS88.2rep. We conclude that the neutrophil response to NS88.2 promotes inflammation and may be a contributing factor to the severity of invasive GAS infections. Abbreviations CovRS, control of virulence regulatory system; GAS, Group A Streptococcus; PMN, polymorphonuclear leukocyte.

IdeS, a secreted proteinase of Streptococcus pyogenes, is bound to a nuclease at the bacterial surface where it inactivates opsonizing IgG antibodies

The important bacterial pathogen Streptococcus pyogenes secretes IdeS (immunoglobulin G-degrading enzyme of S.pyogenes), a proteinase that cleaves human immunoglobulin G (IgG) antibodies in the hinge region resulting in Fc (fragment crystallizable) and F(ab')2 (fragment antigen-binding) fragments and protects the bacteria against phagocytic killing. Experiments with radiolabeled IdeS and flow cytometry demonstrated that IdeS binds to the surface of S.pyogenes, and the interaction was most prominent in conditions resembling those in the pharynx (acidic pH and low salt), the habitat for S.pyogenes. SpnA (S. pyogenes nuclease A) is a cell wall-anchored DNase. A dose-dependent interaction between purified SpnA and IdeS was demonstrated in slot binding and surface plasmon resonance spectroscopy experiments. Gel filtration showed that IdeS forms proteolytically active complexes with SpnA in solution, and super-resolution fluorescence microscopy revealed the presence of SpnA-IdeS complexes at the surface of S.pyogenes. Finally, specific IgG antibodies binding to S.pyogenes surface antigens were efficiently cleaved by surface-associated IdeS. IdeS is secreted by all S.pyogenes isolates and cleaves IgG antibodies with a unique degree of specificity and efficiency. These properties and the finding here that the proteinase is present and fully active at the bacterial surface in complex with SpnA implicate an important role for IdeS in S.pyogenes biology and pathogenesis.

Different Group A Streptococcus pili lead to varying proinflammatory cytokine responses and virulence.

The human pathogen Streptococcus pyogenes, or Group A Streptococcus (GAS), is associated with a variety of diseases ranging from mild skin and soft tissue infections to invasive diseases and immune sequelae such as rheumatic heart disease. We have recently reported that one of the virulence factors of this pathogen, the pilus, has inflammatory properties and strongly stimulates the innate immune system. Here we used a range of nonpathogenic Lactococcus lactis gain-of-function mutants, each expressing one of the major pilus types of GAS, to compare the immune responses generated by various types of fully assembled pili. In vitro assays indicated variability in the inflammatory response induced by different pili, with the fibronectin-binding, collagen-binding, T antigen (FCT)-1-type pilus from GAS serotype M6/T6 inducing significantly stronger cytokine secretion than other pili. Furthermore, we established that the same trend of pili-mediated immune response could be modeled in Galleria mellonella larvae, which possess a similar innate immune system to vertebrates. Counterintuitively, across the panel of pili types examined in this study, we observed a negative correlation between the intensity of the immune response demonstrated in our experiments and the disease severity observed clinically in the GAS strains associated with each pilus type. This observation suggests that pili-mediated inflammation is more likely to promote bacterial clearance instead of causing disruptive damages that intensify pathogenesis. This also indicates that pili may not be the main contributor to the inflammatory symptoms seen in GAS diseases. Rather, the immune-potentiating properties of the pilus components could potentially be exploited as a vaccine adjuvant.

Transcriptome Analysis of Streptococcus mutans Quorum Sensing-Mediated Persisters Reveals an Enrichment in Genes Related to Stress Defense Mechanisms.

Persisters are a small fraction of growth-arrested phenotypic variants that can survive lethal concentrations of antibiotics but are able to resume growth once antibiotics are stopped. Their formation can be a stochastic process or one triggered by environmental cues. In the human pathogen Streptococcus mutans, the canonical peptide-based quorum-sensing system is an inducible DNA repair system that is pivotal for bacterial survival. Previous work has shown that the CSP-signaling peptide is a stress-signaling alarmone that promotes the formation of stress-induced persisters. In this study, we exposed S. mutans to the CSP pheromone to mimic DNA damage conditions and isolated the antibiotic persisters by treating the cultures with ofloxacin. A transcriptome analysis was then performed to evaluate the differential gene expression between the normal stationary-phase cells and the persisters. RNA sequencing revealed that triggered persistence was associated with the upregulation of genes related to several stress defense mechanisms, notably, multidrug efflux pumps, the arginine deaminase pathway, and the Opu/Opc system. In addition, we showed that inactivation of the VicK kinase of the YycFG essential two-component regulatory system abolished the formation of triggered persisters via the CSP pheromone. These data contribute to the understanding of the triggered persistence phenotype and may suggest new therapeutic strategies for treating persistent streptococcal infections.

The leaderless communication peptide (LCP) class of quorum-sensing peptides is broadly distributed among Firmicutes

The human pathogen Streptococcus pyogenes secretes a short peptide (leaderless communication peptide, LCP) that mediates intercellular communication and controls bacterial virulence through interaction with its receptor, RopB. Here, we show that LCP and RopB homologues are present in other Firmicutes. We experimentally validate that LCPs with distinct peptide communication codes act as bacterial intercellular signals and regulate gene expression in Streptococcus salivarius, Streptococcus porcinus, Enterococcus malodoratus and Limosilactobacillus reuteri. Our results indicate that LCPs are more widespread than previously thought, and their characterization may uncover new signaling mechanisms and roles in coordinating diverse bacterial traits.

Population structure and genetic diversity of Streptococcus suis isolates obtained from the United States.

Diseases caused by the zoonotic pathogen Streptococcus suis are an extensive economic problem as well as an animal welfare concern for the global swine industry. Previous studies have evaluated the genomic diversity and population structure of S. suis isolates, however, the majority of these studies utilized isolates obtained from countries other than the U.S. This study applied whole genome sequencing and cgMLST-based typing to evaluate the population structure and genetic relatedness among S. suis isolates obtained within the U.S. The established high-resolution phylogenomic framework revealed extensive genomic variation and diversity among the sampled S. suis isolates, with isolates from the U.S. and from countries outside the U.S. found interspersed in the phylogeny. S. suis isolates obtained within the U.S. did not cluster by state or geographic location, however, isolates with similar serotypes, both obtained from within and outside the U.S., generally clustered together. Average nucleotide identity (ANI) values determined for the S. suis genomes were extensively broad, approaching the recommended species demarcation value, and correlated with the phylogenetic group distribution of the cgMLST-based tree. Numerous antimicrobial resistance (AMR) elements were identified among both U.S. and non-U.S. isolates with ble, tetO, and ermB genes identified as the most prevalent. The epf, mrp, and sly genes, historically used as markers for virulence potential, were also observed in the genomes of isolates that grouped together forming a subclade of clonal complex 1 (CC1) isolates. Collectively, the data in this report provides critical information needed to address potential biosurveillance needs and insights into the genetic diversity and population structure of S. suis isolates obtained within the U.S.

Mucosal Vaccines against Bacterial and Viral Pathogens

The mucosal membranes of the human body play a crucial role in the development, maintenance, and regulation of barrier functions and immune homeostasis, representing an integral component of the overall immune system. Mucosal vaccines elicit immune processes in the lymphoid tissue associated with the mucosal membranes. A critical objective of mucosal immunization is the identification of an antigen delivery vector capable of ensuring optimal vaccine efficacy. The authors of this article have conducted extensive research on the probiotic properties of enterococci over an extended period. They employ a safe and beneficial probiotic strain, Enterococcus faecium L3, as a delivery vector for vaccine antigens. Initially, the gene encoding the pathogenicity factor Bac, derived from group B streptococci (Streptococcus agalactiae), was successfully integrated into the genome of the probiotic strain E. faecium L3. Intravaginal, oral, and intranasal mucosal immunization methods utilizing the L3-Bac+ probiotic, which expresses antigenic determinants of pathogenic streptococci, were found to confer protection against bacterial infection in laboratory animals. Subsequently, recombinant technologies were refined, leading to the development of a universal method for incorporating a region of interest from the gene into the structure of the major pili protein gene of E. faecium L3. Using this technology, candidate vaccines against various infections, including Streptococcus pneumoniae, influenza A virus, and SARS-CoV-2 following the onset of the Covid-19 pandemic, have been obtained and tested. In this study, alongside the presentation of our own data, the challenges associated with utilizing recombinant probiotic bacteria as vectors for vaccine antigen delivery are discussed.

Probiotic Lactobacillus plantarum Ln4 Showing Antimicrobial and Antibiofilm Effect against Streptococcus mutans KCTC 5124 Causing Dental Caries.

Dental caries has known as an infectious disease that is considered a serious global public health problem. Recently, report indicate that probiotics play a vital role in maintaining oral health. Therefore, this study aimed to evaluate the prevention effects of Lactobacillus plantarum Ln4 against dental infection by the pathogenic bacterium Streptococcus mutans KCTC 5124 through biofilm formation inhibition. To evaluate such prevention effects against S. mutans KCTC 5124, antimicrobial activity, auto-aggregation, co-aggregation, cell surface hydrophobicity, total exopolysaccharide (EPS) production rate, and biofilm formation were analyzed. Results showed that L. plantarum Ln4 showed higher antimicrobial activity than L. rhamnosus GG (LGG). In the group treated with L. plantarum Ln4, the co-aggregation (58.85%), cell surface hydrophobicity (16.75%), and EPS production rate (73.29%) values were lower than those of LGG and the negative control. Additionally, crystal violet staining and confocal laser scanning microscopy (CLSM) revealed that L. plantarum Ln4 effectively inhibited biofilm formation in S. mutans KCTC 5124. Therefore, L. plantarum Ln4 could be used in the industry as a probiotics to prevent and improve oral health.

Whole Genomic Characterization of Streptococcus iniae Isolates from Barramundi (Lates calcarifer) and Preliminary Evidence of Cross-Protective Immunization.

Lates calcarifer, also known as Barramundi or Asian seabass, is a highly productive and fast-growing species that is well suited to large-scale aquaculture due to its attractive harvestable yields (premium fish). This fish has been envisioned as having the potential to be the "Salmon of Tropics". Cultivating Lates calcarifer in aquaculture poses challenges, as the dense populations that make such aquaculture commercially viable facilitate the rapid spread of infectious diseases, which in turn significantly impact yield. Hence, the immunization of juveniles is necessary, and the development of new immunization agents enhances the efficiency of aquaculture and improves food security. In our study, we characterize seven novel strains of the bacterial pathogen Streptococcus iniae that were collected from commercial fish farms in Singapore and Australia. We find that the capsular operon in our strains is highly conserved and identify a number of major surface antigens previously described in Streptococcus. A genome analysis indicates that the present strains are closely related but form distinct strains within the S. iniae species. We then proceed to demonstrate that inoculation with the inactivated strain P3SAB cross-protects Lates calcarifer against S. iniae infections in vivo from a variety of strains found in both Singapore and Australia.

Spontaneous bacterial peritonitis due to streptococcus pneumoniae: a case report

Despite not being a normal gastrointestinal flora, Streptococcus pneumoniae can be the causative agent of Spontaneous Bacterial Peritonitis (SBP) in clinical setting. Patient present with fever, progressive abdominal pain, swelling of limbs, abdomen or both. It is mainly diagnosed by ascitic fluid tapping and its cell count along with culture and sensitivity. This condition has very good response with antibiotics as per culture reports. Here, we present a common case of SBP caused by an uncommon pathogen Streptococcus pneumoniae.

Gut microbiota differences induced by Toxoplasma gondii seropositivity in stray cats in South Korea.

T. gondii is a highly prevalent parasite worldwide, with cats serving as its final host. However, few studies have investigated the impact of T. gondii infection on cat gut microbiota. Therefore, this study examined the influence of T. gondii infection on the gut microbiota of stray cats and identified potential pathogens in their feces. This study examined T. gondii infection through blood of stray cats and the influence of microbiota in their feces using 16S rRNA gene amplicon sequencing. The results revealed significant differences in gut microbiota composition and diversity between the T. gondii seropositive and seronegative groups. Seropositive samples displayed a lower number of operational taxonomic units and reduced Shannon index than the seronegative samples. The seropositive and seronegative groups exhibited enrichment of taxa, including Escherichia and Enterobacteriaceae and Collinsella, Bifidobacterium, and Roseburia, respectively. Furthermore, potential pathogen species, including Campylobacter, Escherichia, and Streptococcus, were identified in the fecal samples. These findings suggest that T. gondii infection significantly impacts gut microbiota composition and diversity in stray cats. Additionally, an increased potential pathogen load, represented by Escherichia spp., was observed. These results underscore the importance of monitoring the prevalence of zoonotic pathogens in stray cats, as they can serve as reservoirs for zoonotic diseases.

Lobosteroids A-F: Six New Highly Oxidized Steroids from the Chinese Soft Coral Lobophytum sp.

To explore the steroidal constituents of the soft coral Lobophytum sp. at the coast of Xuwen County, Guangdong Province, China, a chemical investigation of the above-mentioned soft coral was carried out. After repeated column chromatography over silica gel, Sephadex LH-20, and reversed-phase HPLC, six new steroids, namely lobosteroids A-F (1-6), along with four known compounds 7-10, were obtained. Their structures were determined by extensive spectroscopic analysis and comparison with the spectral data reported in the literature. Among them, the absolute configuration of 1 was determined by X-ray diffraction analysis using Cu Kα radiation. These steroids were characterized by either the presence of an α,β-α',β'-unsaturated carbonyl, or an α,β-unsaturated carbonyl moiety in ring A, or the existence of a 5α,8α-epidioxy system in ring B, as well as diverse oxidation of side chains. The antibacterial bioassays showed that all isolated steroids exhibited significant inhibitory activities against the fish pathogenic bacteria Streptococcus parauberis FP KSP28, Phoyobacterium damselae FP2244, and Streptococcus parauberis SPOF3K, with IC90 values ranging from 0.1 to 11.0 µM. Meanwhile, compounds 2 and 6-10 displayed potent inhibitory effects against the vancomycin-resistant Enterococcus faecium bacterium G7 with IC90 values ranging from 4.4 to 18.3 µM. Therefore, ten highly oxidized steroids with strong antibacterial activities were isolated from the Chinese soft coral Lobophytum sp., which could be developed as new chemotypes of antibacterial drug leads.