Differences of bacterioplankton communities between the source and upstream regions of the Yangtze River: microbial structure, co-occurrence pattern, and environmental influencing factors.

The source area of the Yangtze River is located in the hinterland of the Qinghai-Tibet Plateau, which is known as the "Earth's third pole." It is the water conservation area and the natural barrier of the ecosystem of the Yangtze River basin. It is also the most sensitive area of the natural ecosystem, and the ecological environment is very fragile. Microorganisms play key roles in the biogeochemical processes of water. In this paper, the bacterioplankton communities in the source and upstream regions of the Yangtze River were studied based on 16S rRNA high-throughput sequencing, and their environmental influencing factors were further analyzed. Results showed that the upstream region had higher richness and diversity than the source region. The predominant bacterial phyla in the source and upstream regions were Proteobacteria, Firmicutes, and Actinobacteriota. The bacterial phyla associated with municipal pollution and opportunistic pathogen, such as Firmicutes and Actinobacteriota, were more abundant in the upstream. By contrast, distinct planktonic bacterial genera associated with mining pollution, such as Acidiphilium and Acidithiobacillus, were more abundant in the source region. The co-occurrence network showed that the interaction of bacterioplankton community is more frequent in the upstream. The bacterioplankton community compositions, richness, and functional profiles were affected by the spatial heterogeneity. Moreover, variation partitioning analysis further confirmed that the amount of variation in the source region independently explained by variables of altitude was the largest, followed by water nutrient. This paper revealed the spatial distribution of planktonic bacterial communities in the source and upstream regions of the Yangtze River and its correlation with environmental factors, providing information support for ensuring the health and safety of aquatic ecosystems in the Yangtze River Basin.

Effect of chlorine disinfectant influx on biological sewage treatment process under the COVID-19 pandemic: Performance, mechanisms and implications.

Since the onset of the COVID-19 Pandemic, large amounts of chlorine-containing disinfectants have been used to interrupt the spread of SARS-CoV-2 and residual chlorine eventually entered the hospital or municipal sewage treatment facilities. However, little is known about the effect of chlorine influx on the biological sewage treatment process. Here we investigated the effect of chlorine on the microbiome and the mechanism of microbial chlorine resistance in the activated sludge of the aerobic treatment process, using metagenomic and metatranscriptomic sequencing. We found that chlorine could negatively impact the aerobic treatment performance regarding nitrogen/COD removal with a dose-dependent effect, and the dual effects of chlorine dose and interaction time differentiated the microbial community in activated sludge. The decline of nitrogen/COD removal was attributed to the compressed activity of functional microorganisms, such as the ammonia oxidation bacteria, under chlorinated conditions, and the damage cannot be recovered in a short term. In addition, some microorganisms could survive in chlorinated conditions by up-regulating the chlorine resistance genes (CRGs) expression (approximately 1.5 times) and stimulating new CRGs expression. In particular, species Acinetobacter johnsonii could resist high concentrations of chlorine through various mechanisms, especially the overexpression of efflux pump function encoded by qac genes play a key role. Based on these results, considering the persistence of the epidemic and extensive use of chlorine disinfectants, it cannot be ignored that large amounts of residual chlorine are entering the biological treatment facility, and strictly de-chlorination measures or microbial chlorine resistance regulations before entering should be implemented.

Interaction of silver nanoparticles with marine/lake snow in early formation stage.

Marine and lake snows play an important ecological role in aquatic systems, and recent researches have also revealed their interactions with various pollutants. In this paper, the interaction of silver nanoparticles (Ag-NPs), a typical nano-pollutant, with marine/lake snow in the early formation stage was investigated by roller table experiments. Results indicated Ag-NPs promoted the accumulation of larger marine snow flocs while inhibited the development of lake snow. The promotion effect of AgNPs might be attributed to their oxidative dissolution into low-toxic silver chloride complexes in seawater, and the subsequent incorporation into marine snow, which would enhance the rigidity and strength of larger flocs and favor the development of biomass. Conversely, Ag-NPs mainly existed in the form of colloidal nanoparticles in lake water and their strong antimicrobial effect suppressed the growths of biomass and lake snow. In addition, Ag-NPs could also affect the microbial community of marine/lake snow, including impact on microbial diversity, and elevation on abundances of extracellular polymeric substances (EPS) synthesis genes and silver resistance genes. This work has deepened our understanding of the fate and ecological effect of Ag-NPs via the interaction with marine/lake snow in aquatic environments.

Comparative Analysis of the Symbiotic Microbiota in the Chinese Mitten Crab (Eriocheir sinensis): Microbial Structure, Co-Occurrence Patterns, and Predictive Functions.

Symbiotic microorganisms in the digestive and circulatory systems are found in various crustaceans, and their essential roles in crustacean health, nutrition, and disease have attracted considerable interest. Although the intestinal microbiota of the Chinese mitten crab (Eriocheir sinensis) has been extensively studied, information on the symbiotic microbiota at various sites of this aquatic economic species, particularly the hepatopancreas and hemolymph, is lacking. This study aimed to comprehensively characterize the hemolymph, hepatopancreas, and intestinal microbiota of Chinese mitten crab through the high-throughput sequencing of the 16S rRNA gene. Results showed no significant difference in microbial diversity between the hemolymph and hepatopancreas (Welch t-test; p > 0.05), but their microbial diversity was significantly higher than that in the intestine (p < 0.05). Distinct differences were found in the structure, composition, and predicted function of the symbiotic microbiota at these sites. At the phylum level, the hemolymph and hepatopancreas microbiota were dominated by Proteobacteria, Firmicutes, and Acidobacteriota, followed by Bacteroidota and Actinobacteriota, whereas the gut microbiota was mainly composed of Firmicutes, Proteobacteria, and Bacteroidota. At the genus level, Candidatus Hepatoplasma, Shewanella, and Aeromonas were dominant in the hepatopancreas; Candidatus Bacilloplasma, Roseimarinus, and Vibrio were dominant in the intestine; Enterobacter, norank_Vicinamibacterales, and Pseudomonas were relatively high-abundance genera in the hemolymph. The composition and abundance of symbiotic microbiota in the hemolymph and hepatopancreas were extremely similar (p > 0.05), and no significant difference in functional prediction was found (p > 0.05). Comparing the hemolymph in the intestine and hepatopancreas, the hemolymph had lower variation in bacterial composition among individuals, having a more uniform abundance of major bacterial taxa, a smaller coefficient of variation, and the highest proportion of shared genera. Network complexity varied greatly among the three sites. The hepatopancreas microbiota was the most complex, followed by the hemolymph microbiota, and the intestinal microbiota had the simplest network. This study revealed the taxonomic and functional characteristics of the hemolymph, hepatopancreas, and gut microbiota in Chinese mitten crab. The results expanded our understanding of the symbiotic microbiota in crustaceans, providing potential indicators for assessing the health status of Chinese mitten crab.

Significant alterations of intestinal symbiotic microbiota induced by intraperitoneal vaccination mediate changes in intestinal metabolism of NEW Genetically Improved Farmed Tilapia (NEW GIFT, Oreochromis niloticus).

BACKGROUND: After millions of years of coevolution, symbiotic microbiota has become an integral part of the host and plays an important role in host immunity, metabolism, and health. Vaccination, as an effective means of preventing infectious diseases, has been playing a vital role in the prevention and control of human and animal diseases for decades. However, so far, minimal is known about the effect of vaccination on fish symbiotic microbiota, especially mucosal microbiota, and its correlation with intestinal metabolism remains unclear. METHODS: Here we reported the effect of an inactivated bivalent Aeromonas hydrophila/Aeromonas veronii vaccine on the symbiotic microbiota and its correlation with the intestinal metabolism of farmed adult Nile tilapia (Oreochromis niloticus) by 16S rRNA gene high-throughput sequencing and gas chromatography-mass spectrometry metabolomics. RESULTS: Results showed that vaccination significantly changed the structure, composition, and predictive function of intestinal mucosal microbiota but did not significantly affect the symbiotic microbiota of other sites including gill mucosae, stomach contents, and stomach mucosae. Moreover, vaccination significantly reduced the relative abundance values of potential opportunistic pathogens such as Aeromonas, Escherichia-Shigella, and Acinetobacter in intestinal mucosae. Combined with the enhancement of immune function after vaccination, inactivated bivalent Aeromonas vaccination had a protective effect against the intestinal pathogen infection of tilapia. In addition, the metabolite differential analysis showed that vaccination significantly increased the concentrations of carbohydrate-related metabolites such as lactic acid, succinic acid, and gluconic acid but significantly decreased the concentrations of multiple lipid-related metabolites in tilapia intestines. Vaccination affected the intestinal metabolism of tilapia, which was further verified by the predictive function of intestinal microbiota. Furthermore, the correlation analyses showed that most of the intestinal differential microorganisms were significantly correlated with intestinal differential metabolites after vaccination, confirming that the effect of vaccination on intestinal metabolism was closely related to the intestinal microbiota. CONCLUSIONS: In conclusion, this paper revealed the microbial and metabolic responses induced by inactivated vaccination, suggesting that intestinal microbiota might mediate the effect of vaccination on the intestinal metabolism of tilapia. It expanded the novel understanding of vaccine protective mechanisms from microbial and metabolic perspectives, providing important implications for the potential influence of vaccination on human intestinal microbiota and metabolism. Video Abstract.

Attenuated Streptococcus agalactiae WC1535 ∆Sia perturbs the gut microbiota of Oreochromis niloticus, massively colonizes the intestine, and induces intestinal mucosal immunity after intraperitoneal inoculation.

We previously developed and assessed the effectiveness of the attenuated Streptococcus agalactiae (Group B Streptococcus, GBS) strain WC1535 ∆Sia (with neuA-D gene cluster deletion) vaccine in tilapia (Oreochromis niloticus). In this study, we characterized the bacterial communities of the tilapia intestines by 16S rRNA high-throughput sequencing and assessed the serum antibody response, expression of immune-related genes, and histological changes following formalin-killed GBS vaccine (FKV) and the live attenuated vaccine ∆Sia (LAV). Results showed that FKV and LAV induced robust systemic and intestinal mucosal immune responses in tilapia without causing obvious pathological changes in the hindgut, spleen, and head kidney but exerted different effects on intestinal bacterial communities. The richness or diversity of the intestinal bacterial community of FKV tilapia showed no significant changes compared with that of the control fish (p > 0.05) at either day 21 post-initial vaccination (21 dpiv) or day 35 (day 14 after the second immunization) (35 dpiv). The community composition of FKV tilapia and controls was significantly similar, although the relative abundance of some genera was significantly altered. Relative to control fish, the gut ecosystem of LAV tilapia was significantly disturbed with a substantial increase in community diversity at 21 dpiv (p < 0.05) and a significant decrease at 35 dpiv in fish with high serum antibody response (ΔSia35H) (p < 0.05). However, there was no significant difference between ΔSia35H and ΔSia35L (low serum antibody response) fish (p > 0.05). Moreover, the community composition of LAV tilapia at 21 dpiv or 35 dpiv was considerably different from that of the controls. Particularly, GBS ∆Sia was found to be abundant in the intestine at 21 and 35 dpiv. This result suggested that the parenteral administration of the LAV (∆Sia) may also have the effect of oral vaccination in addition to the immune effect of injection vaccination. In addition, a significant correlation was found between the expression of immune-related genes and certain bacterial species in the intestinal mucosal flora. Our findings will contribute to a better understanding of the effects of inactivated and attenuated vaccines on gut microbiota and their relationship with the immune response.

Construction of Streptococcus agalactiae sialic acid mutant and evaluation of its potential as a live attenuated vaccine in Nile tilapia (Oreochromis niloticus).

AIMS: This study aimed to develop a live attenuated vaccine as an effective approach to prevent streptococcosis in tilapia (Oreochromis niloticus). METHODS AND RESULTS: We eliminated the virulence factor, sialic acid (Sia) encoded by the neuA-D gene cluster from the Group B Streptococcus (Streptococcus agalactiae, GBS) strain WC1535, to construct Sia-deficient S. agalactiae (ΔSia) mutant by homologous recombination. Results showed that the ΔSia mutant had higher adherence to HEp-2 cells and lower resistance to RAW264.7 cell phagocytosis than the wild-type S. agalactiae. The virulence of the ΔSia mutant to tilapia dramatically decreased with no virulence recovery. The relative percent survivals (RPSs) were 50.00% and 54.50% at 30 days when challenged at the wild-type WC1535 doses of 1.0 × 107 and 5.0 × 107  CFU fish-1 , respectively, via intraperitoneal (IP) injection. The tilapia vaccinated via IP injection with the ΔSia mutant induced strong antibody agglutination titers. The expression of IL-1ß, TNF-α, MHC-Iα, and MHC-IIß could be enhanced in the intestine, spleen, and head kidney for tilapia administered with the ΔSia mutant. CONCLUSIONS: GBS Sia plays a critical role in adherence to HEp-2 cells and resistance to the immune clearance of RAW264.7 cells. Moreover, the ΔSia mutant is a safe, stable, and immunogenic live attenuated vaccine candidate to protect tilapia against GBS infection. SIGNIFICANCE AND IMPACT OF STUDY: The results offer more evidence of the importance of Sia in GBS and may be instructive in the control of tilapia streptococcosis.

Establishment of Epidemiological Resistance Cut-Off Values of Aquatic Aeromonas to Eight Antimicrobial Agents.

The abuse of antibiotics in aquaculture has led to the increasing rate of antibiotic resistance of aquatic bacteria including Aeromonas, which is an increasing threat to environmental and human health. To date, no epidemiological cut-off values (COWT) for Aeromonas spp. have been established by the Clinical and Laboratory Standards Institute nor the European Commission on Antimicrobial Susceptibility Testing. In this study, commercially prepared minimum inhibitory concentration (MIC) test 96-well plates (dry-form plates) were used to determine the MIC of eight antimicrobial agents against 556 Aeromonas strains. The obtained MIC distributions were simulated and analyzed by NRI and ECOFFinder to obtain tentative COWT values for Aeromonas spp. The COWT values of eight kinds of representative antimicrobial agents including trimethoprim-sulfamethoxazole, erythromycin, doxycycline, neomycin, colistin, florfenicol, enrofloxacin, and ceftazidime for Aeromonas spp. were established and were 0.25, 64/32, 4/2, 8, 4, 1, 0.062/0.125, and 0.5 µg/mL, respectively. Results showed that Aeromonas spp. had a very high proportion of non-wild-type strains to enrofloxacin, florfenicol, and doxycycline, which are the most widely used antimicrobials in aquaculture. The COWT values for Aeromonas spp. obtained in this study can contribute to the final establishment of COWT for Aeromonas spp. internationally.

Enhanced removal efficiency of sulfamethoxazole by acclimated microalgae: Tolerant mechanism, and transformation products and pathways.

This study utilized sulfamethoxazole (SMX) acclimatization to enhance the tolerance and biodegradation capacity of Chlorella vulgaris. Compared to wild C. vulgaris, the growth inhibition and oxidative damage induced by SMX evidently decreased in acclimated C. vulgaris, and meanwhile photosynthetic and antioxidant activities were significantly promoted. The physiological analyses with the aid of principal component analysis revealed the increase of catalase and glutathione reductase activities was the critical tolerant mechanism of acclimated C. vulgaris. As the consequence, the acclimated C. vulgaris exhibited enhanced efficiency and (pseudo-first-order) kinetic rate for removal of SMX. The distribution analysis of residual SMX demonstrated the biodegradation was the major removal mechanism of SMX by C. vulgaris, while bioadsorption and bioaccumulation made pimping contributions. During the degradation process of SMX, nine transformation products (TPs) were identified. Based on the identified TPs, a possible transformation pathway was proposed.

Composition and Functional Characteristics and Influencing Factors of Bacterioplankton Community in the Huangshui River, China.

Bacterial community plays a key role in environmental and ecological processes of river ecosystems. Given the special climatic and geographical conditions, studying the compositional characteristics of microorganisms in highland rivers and the relationship between such microorganisms and water physicochemical factors is important for an in-depth understanding of microbial ecological mechanisms. In the present study, high-throughput sequencing was used to investigate and study the bacterioplankton community of the Huangshui River in the ecotone zone of the Qinghai Plateau and Loess Plateau. The results showed that the Huangshui River had significantly lower alpha diversity than the plain rivers. Despite the similarity in their environmental conditions, the main taxonomic compositions of the bacterial communities were distinct between the Huangshui River and polar regions (the Arctic and Antarctica). Proteobacteria accounted for the largest proportion (30.79-99.98%) of all the sequences, followed by Firmicutes (0-49.38%). Acidiphilium was the most numerous genera, which accounted for 0.03-86.16% of the assigned 16S reads, followed by Acidocella (0-95.9%), both belonging to Alphaproteobacteria. The diverse taxa of potential pathogens, such as Acinetobacter, Pseudomonas, and Aeromonas, were also identified. A principal coordinates analysis, coupled with a canonical correspondence analysis, showed spatial variations in the bacterial community composition. The water physical properties (e.g., Cr6+, total phosphorus, and CODMn); altitude; and land use (e.g., urban land cover and aquaculture) determined the distribution of the bacterioplankton composition. PICRUSt2 revealed that the overall functional profiles of the bacterial communities in different samples were similar, and our results suggested the potential health risks of water sources in this area. This work provided valuable insight into the composition of the plankton bacterial community and its relationship with the environmental factors in the Huangshui River in the ecotone zone of the Qinghai Plateau and Loess Plateau and a theoretical foundation for ecological health management.

Taxonomic and Functional Characteristics of the Gill and Gastrointestinal Microbiota and Its Correlation with Intestinal Metabolites in NEW GIFT Strain of Farmed Adult Nile Tilapia (Oreochromis niloticus).

The gill and gastrointestinal tract are primary entry routes for pathogens. The symbiotic microbiota are essential to the health, nutrition and disease of fish. Though the intestinal microbiota of Nile tilapia (Oreochromis niloticus) has been extensively studied, information on the mucosa-associated microbiota of this species, especially the gill and gastrointestinal mucosa-associated microbiota, is lacking. This study aimed to characterize the gill and gastrointestinal mucosa- and digesta-associated microbiota, as well as the intestinal metabolite profiles in the New Genetically Improved Farmed Tilapia (NEW GIFT) strain of farmed adult Nile tilapia by high-throughput sequencing and gas chromatography/mass spectrometry metabolomics. The diversity, structure, composition, and predicted function of gastrointestinal microbiota were significantly different across gastrointestinal regions and sample types (Welch t-test; p < 0.05). By comparing the mucosa- and digesta-associated microbiota, linear discriminant analysis (LDA) effect size (LEfSe) analysis revealed that Pelomonas, Ralstoniapickettii, Comamonadaceae, and Staphylococcus were significantly enriched in the mucosa-associated microbiota, whereas many bacterial taxa were significantly enriched in the digesta-associated microbiota, including Chitinophagaceae, Cetobacterium, CandidatusCompetibacter, Methyloparacoccus, and chloroplast (LDA score > 3.5). Furthermore, Undibacterium, Escherichia-Shigella, Paeniclostridium, and Cetobacterium were dominant in the intestinal contents and mucosae, whereas Sphingomonasaquatilis and Roseomonasgilardii were commonly found in the gill and stomach mucosae. The Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt2) analysis revealed that the predictive function of digesta-associated microbiota significantly differed from that of mucosa-associated microbiota (R = 0.8152, p = 0.0001). In addition, our results showed a significant interdependence between specific intestinal microbes and metabolites. Notably, the relative abundance values of several potentially beneficial microbes, including Undibacterium, Crenothrix, and Cetobacterium, were positively correlated with most intestinal metabolites, whereas the relative abundance values of some potential opportunistic pathogens, including Acinetobacter, Mycobacterium, Escherichia-Shigella, Paeniclostridium, Aeromonas, and Clostridiumsensustricto 1, were negatively correlated with most intestinal metabolites. This study revealed the characteristics of gill and gastrointestinal mucosa-associated and digesta-associated microbiota of farmed Nile tilapia and identified a close correlation between intestinal microbes and metabolites. The results serve as a basis for the effective application of targeted probiotics or prebiotics in the diet to regulate the nutrition and health of farmed tilapia.

De novo Assembly and Analysis of Amur Sturgeon (Acipenser schrenckii) Transcriptome in Response to Mycobacterium Marinum Infection to Identify Putative Genes Involved in Immunity.

Fish mycobacteriosis is a common bacterial disease in many species of freshwater and marine fish and has caused severe loss of fish production. Mycobacterium marinum has been the most prevalent pathogen observed in several outbreaks of mycobacteriosis of farmed sturgeons in China. However, the immune responses and pathology of sturgeons in mycobacterial infection are rarely studied. Therefore, we used the Illumina RNA-seq method to analyze the transcriptome profile of Acipenser schrenckii challenged with Mycobacterium marinum. To begin, 168,220 non-redundant contigs were acquired from the infection and control groups, and among these, 33,225 contigs have acquired annotations. A total of 4,043 differently expressed (DE) contigs between the two groups were identified, and among these, 2479 were upregulated and 1564 were down-regulated in the infected fish. A total of 1,340 DE contigs with acquired annotations in KEGG were enriched for 124 pathways including the TNF signaling pathway, and the Toll-like receptor signaling pathway. The roles of DE genes involved in significant pathways and other processes were discussed. The 2,209 DE contigs that have yet to acquire proper annotation may represent candidate genes associated with infection in sturgeons and are expected to serve as immunogenetic resources for further study. To our best knowledge, this is the first transcriptome study on sturgeons under bacterial infection.

High-throughput sequencing reveals the gut and lung prokaryotic community profiles of the Chinese giant salamander (Andrias davidianus).

Increasing attention has been attracted to host microbiota, due to their vital impact on host health. Little is known about the microbiota of the Chinese giant salamander (Andrias davidianus), in spite of the high economic and scientific value of this endangered species. This study was designed to characterise and compare the gut and lung prokaryotic communities of the Chinese giant salamander by high-throughput sequencing. Our study showed that the giant salamander had a lung prokaryotic community that clustered separately from its intestinal microbiota. Statistical analysis (LEfSe) revealed that the bacterial populations were dominated by Geobacter, Sulfurimonas, and Dechloromonas from Proteobacteria phylum, and Corynebacterium from Actinobacteria phylum in the lung, while Parabacteroides, Bacteroides, and PW3 from Bacteroidetes phylum, and Oscillospira from Firmicutes phylum were predominant in the intestine. A particularly innovative finding was the fairly high abundance of Archaea, especially methanogenic Euryarchaeota. The gut dominant Archaea were Methanocorpusculum and Thermoplasmata vadinCA11, while Methanosaeta and Methanoculleus were the main Archaea in the lung. PICRUSt analysis revealed differentiated functional profiles between the intestinal miacrobiota and the lung microbiota. Specially, some microbial metabolic functions were significantly more active in the intestinal microbiota, while the functional genes involved in infectious diseases were much richer in the lung microbiota. This study characterized the prokaryotic microbial community profiles in the gut and lung of the Chinese giant salamander, providing foundational support for future study seeking to understand microbiota of the giant salamander and the role of its microbiota on infectious diseases.