Plant-derived carbon (PDC) released by roots has a strong effect on root-associated bacterial community, which is critical for plant fitness in natural environments. However, the freshly exuded PDC can be diluted by the ancient soil-derived carbon (SDC) at a short distance from root apices. Thus, the rhizosphere C pools are normally dominated by SDC rather than PDC. Yet, how PDC and SDC interact to regulate root-associated bacterial community is largely unknown. In this study, a grass species and a legume species were planted in two contrasting matrixes, quartz sand and soil, to assess the role of PDC and SDC in regulating root-associated bacterial community, and to explore whether SDC affects the influence of PDC on bacterial community in soil. Our results indicated that the legume plant showed significantly positive priming effect on soil organic matter decomposition but the grass plant did not. PDC significantly shaped bacterial community in sand culture as indicated by PCR-DGGE and high-throughput sequencing of bacterial 16S rRNA gene. Intriguingly, we found that dissimilarity of bacterial communities associated with two plant species and the percentage of specific OTUs in quartz sand were significantly higher than those in soil. Moreover, several biomarkers enriched by plants in quartz sand turned to be general taxa in soil, which indicated that SDC attenuated the regulation of bacterial community by PDC. Taken together, these results suggest that SDC interacted with PDC and the root-associated microbial community, thus acted as soil buffering component of biological process contributing to soil resilience. The importance of PDC in structuring rhizosphere bacterial community needs to be reconsidered in the context of wider contribution of other C pool, such as SDC.

The mortality rate of hemorrhagic African swine fever (ASF), which targets domestic pigs and wild boars is caused by African swine fever virus (ASFV), can reach 100%. Since the first confirmed ASF outbreak in China on 3 August 2018, 156 ASF outbreaks were detected in 32 provinces. About 1,170,000 pigs were culled in order to halt further spread. There is no effective treatment or vaccine for it and the present molecular diagnosis technologies have trade-offs in sensitivity, specificity, cost and speed, and none of them cater perfectly to ASF control. Thus, a technology that overcomes the need for laboratory facilities, is relatively low cost, and rapidly and sensitively detects ASFV would be highly valuable. Here, we describe an RAA-Cas12a-based system that combines recombinase aided amplification (RAA) and CRISPR/Cas12a for ASFV detection. The fluorescence intensity readout of this system detected ASFV p72 gene levels as low as 10 aM. For on-site ASFV detection, lateral-flow strip readout was introduced for the first time in the RAA-Cas12a based system (named CORDS, Cas12a-based On-site and Rapid Detection System). We used CORDS to detect target DNA highly specifically using the lateral-flow strip readout and the assay displayed no cross-reactivity to other 13 swine viruses including classical swine fever (CSF). CORDS could identify the ASFV DNA target at femtomolar sensitivity in an hour at 37°C, and only requires an incubator. For ease of use, the reagents of CORDS were lyophilized to three tubes and remained the same sensitivity when stored at 4°C for at least 7 days. Thus, CORDS provide a rapid, sensitive and easily operable method for ASFV on-site detection. Lyophilized CORDS can withstand long-term transportation and storage, and is ready for field-based applications.

Soil microbes are driver of nutrient cycling, with microbial function affected by community composition and soil chemical property. Legume and grass are ubiquitous in many ecosystems, however, their differential effects on microbial function are less understood. Here we constructed compartmented rhizobox planted with stylo (Stylosanthes guianensis, legume) or bahiagrass (Paspalum natatum, grass) to compare their influences on bacterial function and to investigate the determinant of bacterial function. Soils in root compartment and in near (0-5 mm from root compartment) or far (10-15 mm from root compartment) rhizosphere were sampled. Soil chemical properties, bacterial community composition and function were characterized. Results indicate that plant species and distance significantly affected bacterial function. The activities of beta-xylosidase, nitrate reductase and phosphomonoesterase were higher in stylo soil than in bahiagrass soil, while leucine-aminopeptidase activity and nosZ abundance were vice versa. Rhizosphere effect was obvious for the activities of beta-glucosidase, beta-xylosidase, chitinase, and the abundances of AOB-amoA, nirS, nosZ. Statistical analysis revealed that soil chemical property was significantly associated with bacterial function, with a higher coefficient than bacterial community composition. These data suggest that stylo and bahiagrass differentially affect bacterial function, which is affected more strongly by soil chemical property than by community composition.

We experimentally isolated and characterized a CFEM protein with putative GPI-anchored site BcCFEM1 in Botrytis cinerea. BcCFEM1 contains a CFEM (common in several fungal extracellular membrane proteins) domain with the characteristic eight cysteine residues at N terminus, and a predicted GPI modification site at C terminus. BcCFEM1 was significantly up-regulated during early stage of infection on bean leaves and induced chlorosis in Nicotiana benthamiana leaves using Agrobacterium infiltration method. Targeted deletion of BcCFEM1 in B. cinerea affected virulence, conidial production and stress tolerance, but not growth rate, conidial germination, colony morphology, and sclerotial formation. However, over expression of BcCFEM1 did not make any observable phenotype change. Therefore, our data suggested that BcCFEM1 contributes to virulence, conidial production, and stress tolerance. These findings further enhance our understanding on the sophisticated pathogenicity of B. cinerea beyond necrotrophic stage, highlighting the importance of CFEM protein to B. cinerea and other broad-host-range necrotrophic pathogens.

Genitourinary syndrome of menopause (GSM) is a chronic and progressive condition with a series of vulvovaginal, sexual, and lower urinary tract discomforts, mainly due to hypoestrogenism. Menopausal hormone therapy (MHT) has generally been considered as the most effective treatment for GSM. In addition, vaginal microbiota is of particular significance to gynecological and reproductive illnesses and potentially has some intimate connections with GSM. Consequently, we sought to evaluate how MHT impacts the composition and structure of vaginal microbiota while alleviating GSM in Chinese menopausal women aged 45-65 years, which has not been investigated previously. 16S rRNA gene sequencing was performed to analyze microbial diversity and composition using vaginal swabs obtained from 100 menopausal women, classified as MHT women who have been taking tibolone regularly (n = 50) and non-treated women who never received any treatment (n = 50). Vaginal Health Index Score (VHIS) and GSM symptoms inquiry were also performed. We found that the vaginal microbial diversity decreased and that the abundance of Lactobacillus increased to be the dominant proportion significantly in the MHT group, in considerable contrast to vaginal microbiota of the non-treated group, which significantly comprised several anaerobic bacteria, namely, Gardnerella, Prevotella, Escherichia-Shigella, Streptococcus, Atopobium, Aerococcus, Anaerotruncus, and Anaerococcus. In this study, women without any MHT had significantly more severe GSM symptoms than those receiving tibolone, especially with regard to vulvovaginal dryness and burning, as well as decreased libido (P < 0.01). However, there was no significant difference in the severity of urological symptoms between the groups (P > 0.05). Furthermore, Lactobacillus was demonstrated to be associated with VHIS positively (r = 0.626, P < 0.001) and with GSM negatively (r = -0.347, P < 0.001). We also identified Chlamydia (r = 0.277, P < 0.01) and Streptococcus (r = 0.270, P < 0.01) as having a prominent association with more serious GSM symptoms. Our study provided an elucidation that MHT could notably alleviate GSM and conspicuously reshape the composition of the vaginal microbiota, which is of extreme importance to clinical practice for the management of GSM.

The benthic bacterial community in Antarctic continental shelf ecosystems are not well-documented. We collected 13 surface sediments from the Ross Sea, a biological hotspot in high-latitude maritime Antarctica undergoing rapid climate change and possible microflora shift, and aimed to study the diversity, structure and assembly mechanism of benthic bacterial community using both culture-dependent and -independent approaches. High-throughput sequencing of 16S rRNA gene amplicons revealed 370 OTUs distributed in 21 phyla and 284 genera. The bacterial community was dominated by Bacteroidetes, Gamma- and Alphaproteobacteria, and constituted by a compact, conserved and positively-correlated group of anaerobes and other competitive aerobic chemoheterotrophs. Null-model test based on βNTI and RCBray indicated that stochastic processes, including dispersal limitation and undominated fractions, were the main forces driving community assembly. On the other hand, environmental factors, mainly temperature, organic matter and chlorophyll, were significantly correlated with bacterial richness, diversity and community structure. Moreover, metabolic and physiological features of the prokaryotic taxa were mapped to evaluate the adaptive mechanisms and functional composition of the benthic bacterial community. Our study is helpful to understand the structural and functional aspects, as well as the ecological and biogeochemical role of the benthic bacterial community in the Ross Sea.

Bacteriolytic myxobacteria are versatile micropredators and are proposed as potential biocontrol agents against diverse bacterial and fungal pathogens. Isolation of new myxobacteria species and exploration of effective predatory products are necessary for successful biocontrol of pathogens. In this study, a myxobacterium strain CY-1 was isolated from a soil sample of a pig farm using the Escherichia coli baiting method. Based on the morphological observation, physiological test, 16S rRNA gene sequence, and genomic data, strain CY-1 was identified as a novel species of the myxobacterial genus Archangium, for which the name Archangium lipolyticum sp. nov. was proposed. Subsequent predation tests indicated that the strain efficiently lysed drug-resistant pathogens, with a higher predatory activity against E. coli 64 than Staphylococcus aureus GDMCC 1.771 (MRSA). The lysis of extracellular proteins against ester-bond-containing substrates (tributyrin, tween 80, egg-yolk, and autoclaved drug-resistant pathogens) inspired the mining of secreted predatory products with lipolytic activity. Furthermore, a lipase ArEstA was identified from the genome of CY-1, and the heterologously expressed and purified enzyme showed bacteriolytic activity against Gram-negative bacteria E. coli 64 but not against Gram-positive MRSA, possibly due to different accessibility of enzyme to lipid substrates in different preys. Our research not only provided a novel myxobacterium species and a candidate enzyme for the development of new biocontrol agents but also reported an experimental basis for further study on different mechanisms of secreted predatory products in myxobacterial killing and degrading of Gram-negative and Gram-positive preys.

The Gram-positive bacterial species Streptococcus suis is an important porcine and human pathogen that causes severe life-threatening diseases associated with high mortality rates. However, the mechanisms by which S. suis evades host innate immunity remain elusive, so identifying novel virulence factors involved in immune evasion is crucial to gain control over this threatening pathogen. Our previous work has shown that S. suis protein endopeptidase O (SsPepO) is a novel fibronectin-binding protein. Here, we identified that recombinant SsPepO binds human plasminogen in a dose-dependent manner. Moreover, the binding of SsPepO and plasminogen, upon the activation of urokinase-type plasminogen activator, generated plasmin, which could cleave complement C3b, thus playing an important role in complement control. Additionally, a SspepO-deficient mutant showed impaired adherence to plasminogen as well as impaired adherence to and invasion of rat brain microvascular endothelial cells compared with the wildtype strain. We further found that the SspepO-deficient mutant was efficiently killed by human serum and blood. We also confirmed that the SspepO-deficient mutant had a lower mortality rate than the wildtype strain in a mouse model. In conclusion, these results indicate that SsPepO is a novel plasminogen-binding protein that contributes to S. suis immune evasion.

Stigma maydis polysaccharide (SMPS) is a plant polysaccharide that participates in immune regulation and gastrointestinal motility. Autism spectrum disorder (ASD) refers to a group of neurodevelopmental disorders, and ASD patients often present intestinal microflora imbalance problems; however, there is no effective treatment method. This study explores the effect of SMPS intervention on the gut microbiota in autism model rats as well as the potential action pathways. Female Wistar rats were intraperitoneally injected with sodium valproic acid (VPA) or normal saline at embryonic day 12.5 to establish an autism model or normal control in their offspring. The offspring prenatally exposed to VPA were randomly assigned to the VPA and the SMPS groups. The SMPS group was administered SMPS from E0.5 to postnatal day (PND) 21. We performed 16S rRNA and transcriptomics analyses to reveal the gut microbiota (GM) and differentially expressed genes in the autism model rats in response to SMPS intervention. SMPS intervention significantly improved the diversity and structure of the GM in autism model rats compared with the VPA rats. Moreover, the relative abundance of Prevotellaceae and Lachnospiraceae_NK4A136_group was increased after SMPS intervention. Transcriptome sequencing showed that 496 differentially expressed genes (DEGs) were identified after SMPS administration compared with the VPA group. Meanwhile, gene ontology (GO) enrichment analysis of DEGs was showed that the SMPS group had significant 653 GO terms. SMPS intervention had a major influence on oxidative phosphorylation, retrograde endocannabinoid signaling, thermogenesis, ribosome, protein digestion and absorption, renin-angiotensin system, calcium signaling pathway, glycosphingolipid biosynthesis-ganglio series, and propanoate metabolism pathways. Overall, this study suggests that SMPS interventions in early life may have an impact on gut microbiota, and then affect the transcriptomics levels of the hippocampal tissue in the VPA-induced autism model rats. It provides scientific evidence for the role of the microbe-gut-brain axis in ASD research.

Plants are the essential factors shaping soil microbial community (SMC) structure. When most studies focus on the difference in the SMC structure associated different plant species, the variation in the SMC structure associated with phylogenetically close species is less investigated. Legume (Fabaceae) and grass (Poaceae) are functionally important plant groups; however, their influences on the SMC structure are seldom compared, and the variation in the SMC structure among legume or grass species is largely unknown. In this study, we grew three legume species vs. three grass species in mesocosms, and monitored the soil chemical property, quantified the abundance of bacteria and fungi. The SMC structure was also characterized using PCR-DGGE and Miseq sequencing. Results showed that legume and grass differentially affected soil pH, dissolved organic C, total N content, and available P content, and that legume enriched fungi more greatly than grass. Both DGGE profiling and Miseq-sequencing indicated that the bacterial diversity associated with legume was higher than that associated with grass. When legume increased the abundance of Verrucomicrobia, grass decreased it, and furthermore, linear discriminant analysis identified some group-specific microbial taxa as potential biomarkers of legume or grass. These data suggest that legume and grass differentially select for the SMC. More importantly, clustering analysis based on both DGGE profiling and Miseq-sequencing demonstrated that the variation in the SMC structure associated with three legume species was greater than that associated with three grass species.

Salmonella infection is an important foodborne consumer health concern that can be mitigated during food processing. Bacteriophage therapy imparts many advantages over conventional chemical preservatives including pathogen specificity, natural derivation, potency, and providing a high degree of safety. The objective of this study aimed to isolate and characterize a phage that effectively control Salmonella food contamination. Out of 35 isolated phages, LPSE1 demonstrated a broad Salmonella host range, robust lytic ability, extensive pH tolerance, and prolonged thermal stability. The capacity for phage LPSE1 to control Salmonella Enteritidis-ATCC13076 in milk, sausage, and lettuce was established. Incubation of LPSE1 at 28°C in milk reduced recoverable Salmonella by approximately 1.44 log10 CFU/mL and 2.37 log10 CFU/mL at MOI of 1 and 100, respectively, as relative to the phage-excluded control. Upon administration of LPSE1 at an MOI of 1 in sausage, Salmonella count decreased 0.52 log10 at 28°C. At MOI of 100, the count decreased 0.49 log10 at 4°C. Incubation of LPSE1 on lettuce reduced recoverable Salmonella by 2.02 log10, 1.71 log10, and 1.45 log10 CFU/mL at an MOI of 1, 10, and 100, respectively, as relative to the negative control. Taken together, these findings establish LPSE1 as an effective weapon against human pathogenic Salmonella in various ready to eat foods.

Previous studies have identified oral administration of antibiotics and gut-impacting drugs as critical drivers for fecal antibiotic resistance (AR) and microbiome disruption in lab mice, but the practical implications of these findings have yet to be validated in hosts nurtured in conventional environment. Using ampicillin (Amp) as a way to extrapolate the general effect of antibiotics, this project examined the impact of drug administration routes on fecal microbiota and resistome using poultry raised in a teaching farm. AR genes were found to be abundant in the feces of young Leghorn chicks without previous antibiotic treatment. In chickens seeded with bla CMY-2 + Escherichia coli, 300 mg/kg body weight of Amp was orally administered for 5 days. This led to the fecal microbiota switching from Firmicutes occupied (95.60 ± 2.62%) and Lactobacillus rich, to being dominated by Proteobacteria (70.91 ± 28.93%), especially Escherichia/Shigella. However, when Amp was given via muscle injection, Firmicutes was mostly retained (i.e., from 83.6 ± 24.4% pre- to 90.4 ± 15.2% post-treatment). In control chickens without seeding with bla CMY-2 + E. coli, oral Amp also led to the increase of Proteobacteria, dominated by Klebsiella and Escherichia/Shigella, and a reduction of Firmicutes. Specifically within Firmicutes, Enterococcus, Clostridium, etc. were enriched but Lactobacillus was diminished. The fecal resistome including Ampr genes was more abundant in chickens receiving oral Amp than those treated with muscle injection, but the difference was primarily within 1 log. The data illustrated that both drug administration routes and pre-existing gut microbiota have profound impacts on gut microbiome disruption when antibiotic treatment is given. In hosts nurtured in a conventional environment, drug administration route has the most evident impact on gut microbiota rather than the size of the targeted bla CMY-2 + gene pool, likely due to the pre-existing bacteria that are (i) less susceptible to Amp, and/or (ii) with Ampr- or multidrug resistance-encoding genes other than bla CMY-2 +. These results demonstrated the critical interplay among drug administration routes, microbiota seeded through the gastrointestinal tract, AR, gut microbiota disruption, and the rise of common opportunistic pathogens in hosts. The potential implications in human and animal health are discussed.

Aeromonas hydrophila is an emerging foodborne pathogen causing human gastroenteritis. Aeromonas species isolated from food such as seafood presented multidrug-resistance (MDR), raising serious concerns regarding food safety and public health. The use of phages to infect bacteria is a defense against drug-resistant pathogens. In this study, phage ZPAH34 isolated from the lake sample exerted lytic activity against MDR A. hydrophila strain ZYAH75 and inhibited the biofilm on different food-contacting surfaces. ZPAH34 has a large dsDNA genome of 234 kb which belongs to a novel jumbo phage. However, its particle size is the smallest of known jumbo phages so far. Based on phylogenetic analysis, ZPAH34 was used to establish a new genus Chaoshanvirus. Biological characterization revealed that ZPAH34 exhibited wide environmental tolerance, and a high rapid adsorb and reproductive capacity. Food biocontrol experiments demonstrated that ZPAH34 reduces the viable count of A. hydrophila on fish fillets (2.31 log) and lettuce (3.28 log) with potential bactericidal effects. This study isolated and characterized jumbo phage ZPAH34 not only enriched the understanding of phage biological entity diversity and evolution because of its minimal virion size with large genome but also was the first usage of jumbo phage in food safety to eliminate A. hydrophila.