Interspecific Differences in Carbon and Nitrogen Metabolism and Leaf Epiphytic Bacteria among Three Submerged Macrophytes in Response to Elevated Ammonia Nitrogen Concentrations.

Submerged macrophytes in eutrophic aquatic environments adapt to changes in ammonia nitrogen (NH4-N) levels by modifying their levels of free amino acids (FAAs) and soluble carbohydrates (SCs). As symbionts of submerged macrophytes, epiphytic bacteria have obvious host specificity. In the present study, the interspecific differences in the FAA and SC contents of Hydrilla verticillata (Linn. f.) Roylep, Vallisneria natans Hara and Chara braunii Gmelin and their leaf epiphytic bacterial communities were assessed in response to increased NH4-N concentrations. The results revealed that the response of the three submerged macrophytes to NH4-N stress involved the consumption of SCs and the production of FAAs. The NH4-N concentration had a greater impact on the variation in the FAA content, whereas the variation in the SC content was primarily influenced by the species. At the phylum level, the relative abundance of Nitrospirota on the leaves exhibited specific differences, with the order H. verticillata > V. natans > C. braunii. The dominant genera of epiphytic bacteria with denitrification effects on V. natans, H. verticillata and C. braunii leaves were Halomonas, Acinetobacter and Bacillus, respectively. When faced with NH4-N stress, the variation in epiphytic bacterial populations associated with ammonia oxidation and denitrification among submerged macrophytes could contribute to their divergent responses to heightened nitrogen levels.

Cultivable epiphytic bacteria of the Chlorophyta Ulva sp.: diversity, antibacterial, and biofilm-modulating activities.

AIMS: Macroalgae harbor a rich epiphytic microbiota that plays a crucial role in algal morphogenesis and defense mechanisms. This study aims to isolate epiphytic cultivable microbiota from Ulva sp. surfaces. Various culture media were employed to evaluate a wide range of cultivable microbiota. Our objective was to assess the antibacterial and biofilm-modulating activities of supernatants from isolated bacteria. METHODS AND RESULTS: Sixty-nine bacterial isolates from Ulva sp. were identified based on 16S rRNA gene sequencing. Their antibacterial activity and biofilm modulation potential were screened against three target marine bacteria: 45%, mostly affiliated with Gammaproteobacteria and mainly grown on diluted R2A medium (R2Ad), showed strong antibacterial activity, while 18% had a significant impact on biofilm modulation. Molecular network analysis was carried out on four bioactive bacterial supernatants, revealing new molecules potentially responsible for their activities. CONCLUSION: R2Ad offered the greatest diversity and proportion of active isolates. The molecular network approach holds promise for both identifying bacterial isolates based on their molecular production and characterizing antibacterial and biofilm-modulating activities.

A pan-genomic assessment: Delving into the genome of the marine epiphyte Bacillus altitudinis strain 19_A and other very close Bacillus strains from multiple environments.

Marine macroalgae are the habitat of epiphytic bacteria and provide several conditions for a beneficial biological interaction to thrive. Although Bacillus is one of the most abundant epiphytic genera, genomic information on marine macroalgae-associated Bacillus species remains scarce. In this study, we further investigated our previously published genome of the epiphytic strain Bacillus altitudinis 19_A to find features that could be translated to potential metabolites produced by this microorganism, as well as genes that play a role in its interaction with its macroalgal host. To achieve this goal, we performed a pan-genome analysis of Bacillus sp. and a codon bias assessment, including the genome of the strain Bacillus altitudinis 19_A and 29 complete genome sequences of closely related Bacillus strains isolated from soil, marine environments, plants, extreme environments, air, and food. This genomic analysis revealed that Bacillus altitudinis 19_A possessed unique genes encoding proteins involved in horizontal gene transfer, DNA repair, transcriptional regulation, and bacteriocin biosynthesis. In this comparative analysis, codon bias was not associated with the habitat of the strains studied. Some accessory genes were identified in the Bacillus altitudinis 19_A genome that could be related to its epiphytic lifestyle, as well as gene clusters for the biosynthesis of a sporulation-killing factor and a bacteriocin, showing their potential as a source of antimicrobial peptides. Our results provide a comprehensive view of the Bacillus altitudinis 19_A genome to understand its adaptation to the marine environment and its potential as a producer of bioactive compounds.

[Characteristics of Epiphytic Bacterial Community on Submerged Macrophytes in Water Environment Supplemented with Reclaimed Water].

Biofilms attached to submerged macrophytes play an important role in improving the water quality of the water environment supplemented with reclaimed water. In order to explore the effects of reclaimed water quality and submerged macrophyte species on the characteristics of an epiphytic bacterial community, different types of submerged macrophytes were selected as research objects in this study. 16S rRNA high-throughput sequencing technology was used on the epiphytic bacteria and the surrounding environmental samples to analyze the bacterial community structure and functional genes. The results showed that approximately 20%-35% of the nitrogen and phosphorus nutrients were absorbed and utilized in the water environment supplemented with reclaimed water. However, the COD, turbidity, and chroma of the downstream water were significantly increased. The bacterial community of the biofilms attached to submerged macrophytes was significantly different from that in the surrounding environment (soil, sediment, and water body) and in the activated sludge that was treated by reclaimed water. In terms of bacterial community diversity, the richness and diversity were significantly lower than those of soil and sediment but higher than those of plankton bacteria in water. In terms of bacterial community composition, dominant genera and corresponding abundances were also different from those of other samples. The main dominant bacterial genera were Sphingomonas, Aeromonas, Pseudomonas, and Acinetobacter, accounting for 7%-40%, respectively. Both macrophyte species and the quality of reclaimed water (BOD5, TN, NH4+-N, and TP) could affect the bacterial community. However, the effect of water quality of the bacterial community was greater than that of macrophytes species. Additionally, the quality of reclaimed water also affected the abundance of functional genes in the bacterial community, and the relative abundance of nitrogen and phosphorus cycling functional genes was higher in areas with higher nitrogen and phosphorus concentrations.

Analysis of diversity and function of epiphytic bacterial communities associated with macrophytes using a metagenomic approach.

Epiphytic bacteria constitute a vital component of aquatic ecosystems, pivotal in regulating elemental cycling. Despite their significance, the diversity and functions of epiphytic bacterial communities adhering to various submerged macrophytes remain largely unexplored. In this study, we employed a metagenomic approach to investigate the diversity and function of epiphytic bacterial communities associated with six submerged macrophytes: Ceratophyllum demersum, Hydrilla verticillata, Myriophyllum verticillatum, Potamogeton lucens, Stuckenia pectinata, and Najas marina. The results revealed that the predominant epiphytic bacterial species for each plant type included Pseudomonas spp., Microbacterium spp., and Stenotrophomonas rhizophila. Multiple comparisons and linear discriminant analysis effect size indicated a significant divergence in the community composition of epiphytic bacteria among the six submerged macrophytes, with 0.3-1% of species uniquely identified. Epiphytic bacterial richness associated with S. pectinata significantly differed from that of both C. demersum and H. verticillata, although no significant differences were observed in diversity and evenness. Functionally, notable variations were observed in the relative abundances of genes associated with carbon, nitrogen, and phosphorus cycling within epiphytic bacterial communities on the submerged macrophyte hosts. Among these communities, H. verticillata exhibited enrichment in genes related to the 3-hydroxypropionate bicycle and nitrogen assimilation, translocation, and denitrification. Conversely, M. verticillatum showcased enrichment in genes linked to the reductive citric acid cycle (Arnon-Buchanan cycle), reductive pentose phosphate cycle (Calvin cycle), polyphosphate degradation, and organic nitrogen metabolism. In summary, our findings offer valuable insights into the diversity and function of epiphytic bacteria on submerged macrophyte leaves, shedding light on their roles in lake ecosystems.

Metagenomic insights into the dynamic degradation of brown algal polysaccharides by kelp-associated microbiota.

Marine bacteria play important roles in the degradation and cycling of algal polysaccharides. However, the dynamics of epiphytic bacterial communities and their roles in algal polysaccharide degradation during kelp decay are still unclear. Here, we performed metagenomic analyses to investigate the identities and predicted metabolic abilities of epiphytic bacterial communities during the early and late decay stages of the kelp Saccharina japonica. During kelp decay, the dominant epiphytic bacterial communities shifted from Gammaproteobacteria to Verrucomicrobia and Bacteroidetes. In the early decay stage of S. japonica, epiphytic bacteria primarily targeted kelp-derived labile alginate for degradation, among which the gammaproteobacterial Vibrionaceae (particularly Vibrio) and Psychromonadaceae (particularly Psychromonas), abundant in alginate lyases belonging to the polysaccharide lyase (PL) families PL6, PL7, and PL17, were key alginate degraders. More complex fucoidan was preferred to be degraded in the late decay stage of S. japonica by epiphytic bacteria, predominantly from Verrucomicrobia (particularly Lentimonas), Pirellulaceae of Planctomycetes (particularly Rhodopirellula), Pontiellaceae of Kiritimatiellota, and Flavobacteriaceae of Bacteroidetes, which depended on using glycoside hydrolases (GHs) from the GH29, GH95, and GH141 families and sulfatases from the S1_15, S1_16, S1_17, and S1_25 families to depolymerize fucoidan. The pathways for algal polysaccharide degradation in dominant epiphytic bacterial groups were reconstructed based on analyses of metagenome-assembled genomes. This study sheds light on the roles of different epiphytic bacteria in the degradation of brown algal polysaccharides.IMPORTANCEKelps are important primary producers in coastal marine ecosystems. Polysaccharides, as major components of brown algal biomass, constitute a large fraction of organic carbon in the ocean. However, knowledge of the identities and pathways of epiphytic bacteria involved in the degradation process of brown algal polysaccharides during kelp decay is still elusive. Here, based on metagenomic analyses, the succession of epiphytic bacterial communities and their metabolic potential were investigated during the early and late decay stages of Saccharina japonica. Our study revealed a transition in algal polysaccharide-degrading bacteria during kelp decay, shifting from alginate-degrading Gammaproteobacteria to fucoidan-degrading Verrucomicrobia, Planctomycetes, Kiritimatiellota, and Bacteroidetes. A model for the dynamic degradation of algal cell wall polysaccharides, a complex organic carbon, by epiphytic microbiota during kelp decay was proposed. This study deepens our understanding of the role of epiphytic bacteria in marine algal carbon cycling as well as pathogen control in algal culture.

Growth promotion of Sargassum fusiforme by epiphytic microbes is dependent on the extent of interspecific interactions of the microbial community.

Profound growth differences such as seedling length and biomass are often observed during the cultivation of Sargassum fusiforme despite the absence of detectable variance in abiotic factors that could have affected this process. This highlights the importance of biotic factors such as epiphytic microbiota in controlling seedling growth. Yet, how, and to what extent microbial activities can affect host growth in the presence of seawater flow and continuous erosion remains debatable. Particularly, the contribution of microbial network interactions to the growth of macroalgae remains poorly understood. This study aimed to compare the physicochemical properties of S. fusiforme seedlings via 16S rRNA gene Illumina sequencing-based profiling of the epiphytic microbial communities of seedlings with different lengths. Significantly different epiphytic bacterial communities were observed among S. fusiforme seedlings of different lengths. The result showed that community from longer seedlings maintained higher bacterial diversity with the taxa Gammaproteobacteria, Burkholderiales, Alteromonadales, Vibrionaceae, Ralstonia, Colwelliaceae, and Thalassotalea being selectively enriched. More importantly, microbial interspecific interactions, which were predominantly positive, were enhanced consistently in communities of the longer seedlings, indicative of reinforced prevalent and mutually cooperative relationships among the microorganisms associated with S. fusiforme seedlings of greater length. Furthermore, longer seedlings also displayed up-regulation of microbial functional potentials involved in N fixation and mineralization, P mineralization and transportation, and ion transportation compared with shorter ones. Lastly, stochastic processes dominated the community assembly of the epiphytic microorganisms. These findings could provide new insights into the relationship between microbial communities and growth in S. fusiforme seedlings and enable us to predict the community diversity and assembly of macroalgae-associated microbial communities. This could have important implications for linking microbial community diversity and network interactions to their host productivity.

Effects of eutrophic water with ammonium chloride, urea, potassium dihydrogen phosphate and sodium-ß-glycerophosphate on Myriophyllum verticillatum and epiphytic bacteria.

With the rapid development of industry and agriculture, excessive nitrogen and phosphorus released into natural surface water have caused eutrophication. Applying submerged plants to manage eutrophic water has attracted widespread attention. However, there are limited studies on the effects of different nitrogen and phosphorus in the water environment on submerged plants and their epiphytic biofilm. Therefore, this paper investigated the effects of eutrophic water with ammonium chloride (IN), urea (ON), potassium dihydrogen phosphate (IP), and sodium-ß-glycerophosphate (OP) on Myriophyllum verticillatum and epiphytic biofilms. The results showed that Myriophyllum verticillatum exhibited a good purification effect on the eutrophic water with inorganic phosphorus, the removal rates of IP were 68.0%, and the plants grew best in this condition. The fresh weight of the IN group and ON group increased by 12.24% and 7.12%, and the shoot length of the IN group and the ON group increased by 17.71% and 8.33%; the fresh weight of the IP group and OP group increased by 19.19% and 10.83%, the shoot length of the IP group and the OP group increased by 21.09% and 18.23%. In addition, the enzyme activities of superoxide dismutase, catalase, nitrate reductase, and acid phosphatase in plant leaves were significantly changed in eutrophic water with different forms of nitrogen and phosphorus. Finally, the analysis of the epiphytic bacteria showed that different forms of nitrogen and phosphorus nutrients could significantly alter the abundance and structure of microorganisms and microbial metabolism also had significant changes. This study provides a new theoretical basis for evaluating the removal of different forms of nitrogen and phosphorus by Myriophyllum verticillatum, and it also provides new insights for the subsequent engineering of epiphytic microorganisms to improve the capability of submerged plants to treat eutrophic water.

Protective Effect of Select Bacterial Species Representative of Fresh Produce on Human Norovirus Surrogates Exposed to Disinfecting Pulsed Light.

Contamination of berries and leafy greens with human norovirus (HuNoV) is a major cause of outbreaks of epidemic gastroenteritis worldwide. Using murine norovirus type 1 (MNV-1) and Tulane virus, we studied the possible extension of HuNoV persistence by biofilm-producing epiphytic bacteria on fresh produce. Nine bacterial species frequently found on the surface of berries and leafy greens (Bacillus cereus, Enterobacter cloacae, Escherichia coli, Kocuria kristinae, Lactobacillus plantarum, Pantoea agglomerans, Pseudomonas fluorescens, Raoultella terrigena, and Xanthomonas campestris) were evaluated for the ability to form biofilms in the MBEC Assay Biofilm Inoculator and in 96-well microplates. The biofilm-forming bacteria were further tested for binding MNV-1 and Tulane virus and the ability to protect them against loss of capsid integrity upon exposure to disinfecting pulsed light at a fluence of 11.52 J/cm2. Based on viral reductions, MNV-1 did not benefit from attachment to biofilm whereas Tulane virus was significantly more resistant than the control when attached to biofilms of E. cloacae (P ≤ 0.01), E. coli (P ≤ 0.01), K. kristinae (P ≤ 0.01), P. agglomerans (P ≤ 0.05), or P. fluorescens (P ≤ 0.0001). Enzymatic dispersion of biofilm and microscopic observations suggest that the biofilm matrix composition may contribute to the virus resistance. Our results indicate that direct virus-biofilm interaction protects Tulane virus against disinfecting pulsed light, and that HuNoV on fresh produce therefore might resist such treatment more than suggested by laboratory tests so far. IMPORTANCE Recent studies have shown that bacteria may be involved in the attachment of HuNoV to the surface of fresh produce. Because these foods are difficult to disinfect by conventional methods without compromising product quality, nonthermal nonchemical disinfectants such as pulsed light are being investigated. We seek to understand how HuNoV interacts with epiphytic bacteria, particularly with biofilms formed by bacterial epiphytes, with cells and extracellular polymeric substances, and to determine if it thus escapes inactivation by pulsed light. The results of this study should advance understanding of the effects of epiphytic biofilms on the persistence of HuNoV particle integrity after pulsed light treatment and thus guide the design of novel pathogen control strategies in the food industry.

Antimicrobial and quorum sensing inhibitory activity of epiphytic bacteria isolated from the red alga Halymenia durvillei.

Halymenia durvillei is a red alga that is commonly utilized in the Philippines as food and as a source of high-value natural products for industrial applications. However, there are no studies regarding the microbial community associated with H. durvillei and its potential applications. This study aimed to isolate and identify the epiphytic bacteria of H. durvillei and determine their antimicrobial and quorum sensing inhibitory (QSI) effects. The thalli of H. durvillei were collected at the shores of Santa Fe, Bantayan, Cebu, Philippines. Bacterial isolates were identified using 16S rRNA, and their ethyl acetate (EtOAc) extracts were subjected to antimicrobial susceptibility tests against representative species of yeast and Gram-negative and Gram-positive bacteria. Their QSI activity against Chromobacterium violaceum was also determined. Fourteen distinct bacterial colonies belonging to four genera, namely Alteromonas (3), Bacillus (5), Oceanobacillus (1) and Vibrio (5), were successfully isolated and identified. All 14 bacterial isolates exhibited antibacterial effects. EPB9, identified as Bacillus safensis , consistently showed the strongest inhibition against Escherichia coli , Staphylococcus aureus and Staphylococcus epidermidis , with minimum inhibitory concentrations (MICs) ranging from 0.0625 to 1.0 mg ml-1. In contrast, all 14 isolates showed weak antifungal effects. Both B. safensis (EPB9) and Bacillus australimaris (EPB15) exhibited QSI effects at 100 mg ml-1, showing opaque zones of 3.1±0.9 and 3.8±0.4 mm, respectively. This study is the first to isolate and identify the distinct microbial epiphytic bacterial community of H. durvillei and its potential as an abundant resource for new antibacterial and QSI bioactives.

Exploring Diversity and Polymer Degrading Potential of Epiphytic Bacteria Isolated from Marine Macroalgae.

The macroalgae surface allows specific bacterial communities to colonize, resulting in complex biological interactions. In recent years, several researchers have studied the diversity and function of the epiphytic bacteria associated with algal host, but largely these interactions remain underexplored. In the present study we analysed the cultivable diversity and polymer degradation potential of epiphytic bacteria associated with five different marine macroalgae (Sargassum, Ulva, Padina, Dictyota and Pterocladia sp.) sampled from the central west coast of India. Out of the total 360 strains isolated, purified and preserved, about 238 strains were identified through 16S rRNA gene sequence analysis and processed for polymer (cellulose, pectin, xylan and starch) degrading activities. Phylogeny placed the strains within the classes Actinobacteria, Bacilli, Alpha-proteobacteria, and Gamma-proteobacteria and clustered them into 45 genera, wherein Vibrio, Bacillus, Pseudoalteromonas, Alteromonas, Staphylococcus and Kocuria spp. were the most abundant with 20 strains identified as potentially novel taxa within the genera Bacillus, Cellulosimicrobium, Gordonia, Marinomonas, Vibrio, Luteimonas and Pseudoalteromonas. In terms of polymer hydrolysis potential, 61.3% had xylanase activity, while 59.7%, 58.8%, and 52.2% had amylase, cellulase, and pectinase activity, respectively. Overall, 75.6% of the strains degraded more than one polysaccharide, 24% degraded all polymers, while nine strains (3.8%) degraded raw sugarcane bagasse. This study showed great potential for seaweed-associated bacteria in the bio-remediation of agro-waste based raw materials, which can be employed in the form of green technology.

Comparison of epiphytic and intestinal bacterial communities in freshwater snails (Bellamya aeruginosa) living on submerged plants.

The combination of submerged plants and snails can combat eutrophication of freshwater systems by suppressing algal growth and assimilating nutrients. By consuming epiphytes, snails can benefit the growth of submerged plants. However, the efficiency of this phytoremediation strategy may depend on the microbes associated with the plants and snails. In this study, we compared the epiphytic bacterial communities on submerged plants (Vallisneria natans and Cabomba caroliniana) and intestinal bacterial communities of a snail, Bellamya aeruginosa, found on these plants using 16S rRNA gene sequencing. Epiphytic bacterial communities were similar between the two plant species and snails shared a high proportion of snail intestinal bacterial OTUs (75%) and genera (85%) with plants they grazed on. However, significant variations of Bray-Curtis distances differentiated epiphytic and intestinal bacterial communities. In addition, between the top 50 genera shared by intestinal and epiphytic bacterial communities, more Spearman correlations were detected within bacterial communities associated with snails than between communities associated with plants (190 vs. 143), and the correlations in epiphytic bacterial networks were more concentrated on certain genera, indicating they possessed distinct bacterial networks. This suggests the bacterial communities associated with snails do not depend strongly on the plant they graze on, which may be important for better understanding the role of snails in aquatic eco-restoration.

Comparison of the Peel-Associated Epiphytic Bacteria of Anthocyanin-Rich "Sun Black" and Wild-Type Tomatoes under Organic and Conventional Farming.

Tomatoes are among the most consumed vegetables worldwide and represent a source of health-beneficial substances. Our study represents the first investigating the peel-associated epiphytic bacteria of red and purple (anthocyanin-rich) tomatoes subjected to organic and conventional farming systems. Proteobacteria was the dominant phylum (relative abundances 79-91%) in all experimental conditions. Enterobacteriaceae represented a large fraction (39.3-47.5%) of the communities, with Buttiauxella and Atlantibacter as the most represented genera. The core microbiota was composed of 59 operational taxonomic units (OTUs), including the majority of the most abundant ones. The occurrence of the most abundant OTUs differed among the experimental conditions. OTU 1 (Buttiauxella), OTU 2 (Enterobacteriales), and OTU 6 (Bacillales) were higher in red and purple tomatoes grown under organic farming. OTU 5 (Acinetobacter) had the highest abundance in red tomatoes subjected to organic farming. OTU 3 (Atlantibacter) was among the major OTUs in red tomatoes under both farming conditions. OTU 7 (Clavibacter) and OTU 8 (Enterobacteriaceae) had abundances ≥1% only in red tomatoes grown under conventional farming. PCA and clustering analysis highlighted a high similarity between the bacterial communities of red and purple tomatoes grown under organic farming. Furthermore, the bacterial communities of purple tomatoes grown under organic farming showed the lowest diversity and evenness. This work paves the way to understand the role of nutritional superior tomato genotypes, combined with organic farming, to modulate the presence of beneficial/harmful bacteria and supply healthier foods within a sustainable agriculture.

Comparative genomic analysis of the genus Marinomonas and taxonomic study of Marinomonas algarum sp. nov., isolated from red algae Gelidium amansii.

Members of the genus Marinomonas are known for their environmental adaptation and metabolically versatility, with abundant proteins associated with antifreeze, osmotic pressure resistance, carbohydrase and multiple secondary metabolites. Comparative genomic analysis focusing on secondary metabolites and orthologue proteins was conducted with 30 reference genome sequences in the genus Marinomonas. In this study, a Gram-stain-negative, rod-shaped, non-flagellated and strictly aerobic bacterium, designated as strain E8T, was isolated from the red algae (Gelidium amansii) in the coastal of Weihai, China. Optimal growth of the strain E8T was observed at temperatures 25-30 °C, pH 6.5-8.0 and 1-3% (w/v) NaCl. The DNA G + C content was 42.8 mol%. The predominant isoprenoid quinone was Q-8 and the major fatty acids were C16:0, summed feature 3 and summed feature 8. The major polar lipids were phosphatidylglycerol (PG) and phosphatidylethanolamine (PE). Based on data obtained from this polyphasic taxonomic study, strain E8T should be considered as a novel species of the genus Marinomonas, for which the name Marinomonas algarum is proposed. The type strain is E8T (= KCTC 92201T = MCCC 1K07070T).

Changes in Composition and Diversity of Epiphytic Microorganisms on Field Pea Seeds, Partial Crop Peas, and Whole Crop Peas during Maturation and Ensiling with or without Lactic Acid Bacteria Inoculant.

The present study was conducted under the hypothesis that, in field peas, type of plant material, stage of maturity, ensiling, silage additive, and aerobic stress affect the composition and diversity of epiphytic microbial communities. Epiphytic microbial composition and diversity of pea seeds, partial crop peas, and whole crop peas was analyzed at different stages of late maturity, before and after ensiling, and with or without the use of lactic acid bacteria (LAB) as inoculant. Suitable combinations among pea crop variants, maturity stages, and inoculant use for the production of stable silages with sufficient aerobic stability after opening and during feed-out were identified. Genomic DNA was extracted, and 16S and 18S rRNA gene amplicons were sequenced. To assess the quality of the various silages, nutrient concentration, pH value, concentration of lactic acid, short chain fatty acids, and alcohols, and aerobic stability were determined. Pea seeds were barely colonized by epiphytic microorganisms. In partial and whole crop peas, composition and α-diversity (Shannon index) of bacterial communities did not differ between crop variants but differed among maturity stages. Epiphytic eukaryotes were rarely found on partial and whole crop peas. Bacterial composition and α-diversity were affected by ensiling and subsequent aerobic storage. In partial and whole crop peas, plant maturation caused an increase of the relative abundance of naturally occurring LAB (Weissella, Pediococcus, and Lactobacillus spp.). As a possible result, natural LAB support stable ensiling conditions even without the use of inoculants beginning with a maturity of 78 on the BBCH scale. This corresponded with a dry matter (DM) concentration of 341 and 363 g/kg in partial and whole crop peas, respectively. Addition of LAB inoculants, however, reduced ammonia, acetic acid, and butanol concentrations, and supported aerobic stability. Earlier stages of plant maturity (BBCH 76 and 77, 300 g DM/kg or less) were more prone to microbial spoilage. Stable pea seed silages can be produced at a maturity between BBCH 78 (427 g DM/kg) and 79 (549 g DM/kg), but they undoubtedly require LAB inoculation or application of other ensiling agents. IMPORTANCE Field peas are important protein suppliers for human and animal nutrition. They can be grown in many areas of the world, which may reduce imports of protein plants and has beneficial economic and ecological effects. Ensiling is a method of preserving feed that can be implemented easily and cost-effectively at the farm. Peas harvested as seeds, partial crop, or whole crop at different maturities enable a wide range of applications. The study characterized epiphytic microbial communities on peas in terms of composition and diversity depending on the maturity of the plants and feed conservation by ensiling as they play an essential role for the production of silages. Even if this study did not consider year, site, or cultivar effects, the results would show which part of the plant is probably well suited for the production of stable and high-quality silages and at which stage of maturity.

Diversity of epiphytic bacterial communities on male and female Sargassum thunbergii.

The epiphytic bacteria are the most abundant microorganisms on marine macroalga. However, there are few studies on the distribution of these epiphytic bacteria on male and female Sargassum thunbergii. In this study, the composition and diversity of epiphytic bacterial communities on male and female S. thunbergii were investigated by using the traditional culture-based method and 16S rDNA high-throughput sequencing. The results showed that the dominant bacterial phyla and genera were the same on both male and female S. thunbergii. However, there were significant differences in the relative abundance of epiphytic bacteria at the genus level. Furthermore, male and female S. thunbergii had their own indicative species and specific bacteria. In addition, the predicted functions of the epiphytic bacteria mainly differed in transport and metabolism, environmental adaptation and spore development. This study enriches the baseline knowledge of epiphytic bacteria related to dioecious algae and paves the way for further studies of the relationships between epiphytic microbial communities and the sex of algae.

Bacterial Transformation and Processing of Diatom-Derived Organic Matter: A Case Study for Skeletonema dohrnii.

Bacterial transformation and processing of phytoplankton-derived organic matter are extremely important for the formation of ubiquitous organic matter (OM) in aquatic ecosystems. Heterotrophic bacteria convert OM into biomass and recycle inorganic components, contributing to the production of microbial food webs. While phytoplankton-derived organic matter is commonly studied, the transformation and processing of dissolved OM (DOM) and lysate OM (LOM) by culturable epiphytic bacteria remains poorly understood. In this study, cultivable epiphytic bacteria from the marine diatom, Skeletonema dohrnii, were isolated, purified, and identified. Three bacteria, Roseobacteria sp., Marinobacter sp., and Bacillus sp., were selected to study the transformation and processing of S. dohrnii-derived DOM and LOM using excitation-emission matrix (EEM) fluorescence methods, and bacterial abundance, dissolved organic carbon (DOC) concentration, and transparent exopolymer particle (TEP) content were measured. Meanwhile, the bacterial transformation of DOM and LOM was further evaluated by the fluorescence index, biological index, ß/α, and humification index. The primary fluorophores, peak A (humic-like), peak C (humic-like), peak M (humic-like), peak B (protein-like), and peak T (tryptophan-like), were present in the sample. The fluorescence of DOM and LOM was dominated by protein-like signal that became increasingly humic-like over time, suggesting that more complex molecules (e.g., recalcitrant OM) are being produced. The fluorescence of DOM and LOM was dominated by a protein-like signal that became increasingly humic-like over time, suggesting that epiphytic bacteria produced more complex molecules. Results showed that the bacteria utilized LOM more rapidly than DOM. While the three bacteria transformed OM to different degrees, all were able to facilitate microbial reprocessing of OM into refractory OM.

Community structure and function of epiphytic bacteria attached to three submerged macrophytes.

In aquatic ecosystems, large amounts of epiphytic bacteria living on the leaf surfaces of submerged macrophytes play important roles in affecting plant growth and biogeochemical cycling. The restoration of different submerged macrophytes has been considered an effective measure to improve eutrophic lakes. However, the community ecology of epiphytic bacteria is far from well understood for different submerged macrophytes. In this study, we used quantitative PCR, 16S rRNA gene high-throughput sequencing and functional prediction analysis to explore the structure and function of epiphytic bacteria in an aquatic ecosystem recovered by three submerged macrophytes (Hydrilla verticillata, Vallisneria natans and Potamogeton maackianus) during two growth periods. The results showed that the community compositions and functions of epiphytic bacterial communities on the submerged macrophyte hosts were different from those of the planktonic bacterial communities in the surrounding water. The alpha diversity of the epiphytic bacterial community was significantly higher in October than in July, and the community compositions and functions differed significantly in July and October. Among the three submerged macrophytes, the structures and functions of the epiphytic bacterial community exhibited obvious differences, and some specific taxa were enriched on the biofilms of the three plants. The alpha diversity and the abundance of functions related to nitrogen and phosphorus transformation were higher in the epiphytic bacteria of P. maackianus. In summary, these results provide clues for understanding the distribution and formation mechanisms of epiphytic bacteria on submerged macrophyte leaves and their roles in freshwater ecosystems.

Differential responses of dominant and rare epiphytic bacteria from a submerged macrophyte to elevated CO2.

Epiphytic bacteria develop complex interactions with their host macrophytes and play an important role in the ecological processes in freshwater habitats. However, how dominant and rare taxa respond to elevated atmospheric CO2 remains unclear. A manipulated experiment was carried out to explore the effects of elevated CO2 on the diversity or functional characteristics of leaf epiphytic dominant and rare bacteria from a submerged macrophyte. Three levels (high, medium, normal) of dissolved inorganic carbon (DIC) were applied to the overlying water. The physicochemical properties of the overlying water were measured. Elevated atmospheric CO2 significantly decreased the pH and dissolved oxygen (DO) of overlying water. Proteobacteria, Cyanobacteria, Bacteroidetes, Planctomycetes, and Actinobacteria are the dominant phyla of leaf epiphytic bacteria from Myriophyllum spicatum, occupying over 90% of the accumulated relative abundances. The aquatic DIC level and further pH significantly drove the epiphytic community composition differences among the three DIC levels. For dominant epiphytic bacteria, the functional potential of nutrient processes and mutualistic relationships were strongly affected by a high DIC level, while responses of rare epiphytic bacteria were more related to trace element processes, pathogens, and defense strategies under a high DIC level. Our results showed the responses of epiphytic bacteria to elevated CO2 varied across dominant and rare taxa.

Context-Dependent Benefits of Aphids for Bacteria in the Phyllosphere.

AbstractInsect herbivores, such as aphids, are common on plants, yet how they interact with plant microbiomes remains largely unknown. For instance, for the widespread bacterial epiphyte and potential aphid pathogen Pseudomonas syringae, aphids could impact bacterial populations by serving as secondary hosts or by altering the epiphytic habitat through feeding and/or waste secretion. Here, we examined whether the pea aphid, Acyrthosiphon pisum, could influence epiphytic populations of P. syringae. First, we quantified epiphytic growth ability without aphids and virulence to aphids across 21 diverse P. syringae strains. For eight strains that varied in these traits we then assessed the influence of aphid presence on epiphytic bacterial growth. In some cases P. syringae benefited significantly from the presence of aphids, with up to 3.8 times more cell doublings. This benefit was not correlated with strain traits but rather with initial population densities; smaller bacterial populations received relatively more benefit from aphids, and larger populations received less benefit. Honeydew, the sugary waste product of aphids, in the absence of aphids was sufficient to increase P. syringae density on leaves. We conclude that aphid honeydew can sometimes increase P. syringae epiphytic growth but that the bacteria may not benefit from using aphids as hosts.