Metagenomic insight into the pathogenic-related characteristics and resistome profiles within microbiome residing on the Angkor sandstone monuments in Cambodia.

To reveal the characteristics of indigenous microbiome including the pathogenic-related ones on Angkor monuments in Cambodia and the distribution pattern of resistome at different locations, several sites, namely Angkor Wat, Bayon of Angkor Thom, and Prasat Preah Vihear with different exposure levels to tourists were selected to conduct the metagenomic analysis in this study. The general characteristics of the microbiome on these monuments were revealed, and the association between the environmental geo-ecological feature and the indigenous microbiome was delineated. The most common microbial groups included 6 phyla, namely Acidobacteria, Actinobacteria, Gemmatimonadetes, Nitrospirae, Proteobacteria and Verrucomicrobia on the monuments, but Firmicutes and Chlamydiae were the most dominant phyla found in bats droppings. The taxonomic family of Chitinophagaceae could serve as a signature microbial group for Preah Vihear, the less visited site. More importantly, the pathogenic-related characteristics of the microbiome residing on Angkor monuments were uncovered. A set of specific antibiotic resistance genes (ARGs) with cross-niches dispersal capacity (between the environmental microbiome and the microbiome within warm blood fauna) was identified to be high by the source tracking analysis based on ARGs profile varies in this study. Among the 10 ARG-types detected in this study, 6 of them are confined to resistance mechanism of antibiotic efflux-pump. The findings of this study provide new a new direction on public health management and implication globally at archaeological sites for tourism.

Microbiome characteristics and the key biochemical reactions identified on stone world cultural heritage under different climate conditions.

The surfaces of historical stone monuments are visibly covered with a layer of colonizing microorganisms and their degradation products. In this study, a metadata analysis was conducted using the microbial sequencing data available from NCBI database to determine the diversity, biodeterioration potential and functionality of the stone microbiome on important world cultural heritage sites under four different climatic conditions. The retrieved stone microbial community composition in these metagenomes shows a clear association between climate types of the historical monuments and the diversity and taxonomic composition of the stone microbiomes. Shannon diversity values showed that microbial communities on stone monuments exposed to dry climate were more diverse than those under humid ones. In particular, functions associated with photosynthesis and UV resistance were identified from geographical locations under different climate types. The distribution of key microbial determinants responsible for stone deterioration was linked to survival under extreme environmental conditions and biochemical capabilities and reactions. Among them, biochemical reactions of the microbial nitrogen and sulfur cycles were most predominant. These stone-dwelling microbiomes on historical stone monuments were highly diverse and self-sustaining driven by energy metabolism and biomass accumulation. And metabolic products of the internal geomicrobiological nitrogen cycling on these ancient monuments play a unique role in the biodeterioration of stone monuments. These results highlight the significance of identifying the essential microbial biochemical reactions to advance the understanding of stone biodeterioration for protection management.

Performance Enhancement of All-Inorganic Carbon-Based CsPbIBr2 Perovskite Solar Cells Using a Moth-Eye Anti-Reflector.

All-inorganic carbon-based CsPbIBr2 perovskite solar cells (PSCs) have attracted increasing interest due to the low cost and the balance between bandgap and stability. However, the relatively narrow light absorption range (300 to 600 nm) limited the further improvement of short-circuit current density (JSC) and power conversion efficiency (PCE) of PSCs. Considering the inevitable reflectance loss (~10%) at air/glass interface, we prepared the moth-eye anti-reflector by ultraviolet nanoimprint technology and achieved an average reflectance as low as 5.15%. By attaching the anti-reflector on the glass side of PSCs, the JSC was promoted by 9.4% from 10.89 mA/cm2 to 11.91 mA/cm2, which is the highest among PSCs with a structure of glass/FTO/c-TiO2/CsPbIBr2/Carbon, and the PCE was enhanced by 9.9% from 9.17% to 10.08%. The results demonstrated that the larger JSC induced by the optical reflectance modulation of moth-eye anti-reflector was responsible for the improved PCE. Simultaneously, this moth-eye anti-reflector can withstand a high temperature up to 200 °C, and perform efficiently at a wide range of incident angles from 40° to 90° and under various light intensities. This work is helpful to further improve the performance of CsPbIBr2 PSCs by optical modulation and boost the possible application of wide-range-wavelength anti-reflector in single and multi-junction solar cells.

High-throughput sequencing reveals the spatial distribution variability of microbial community in coastal waters in Shenzhen.

Seashore habitats are located between terrestrial and marine ecosystems, which are a hotspot for anthropogenic impacts. Shenzhen is one of the most developed cities in south China, but the microbial functions of its coastal ecosystems remain poorly understood. The study applied 16S rRNA gene sequencing methods to identify the bacterial community from twenty sites of Shenzhen inshore waters. The microbial structure of the samples between eastern Shenzhen and western Shenzhen seashores is notably different, suggesting the spatial variability. Proteobacteria, Cyanobacteria, Actinobacteria, and Bacteroidetes were dominant phyla in the community, and the relative abundance of Bacteroidetes was significantly higher in eastern seashores. Specifically, samples from western Shenzhen contained much more Prochlorococcus, while Synechococcus was more abundant in eastern samples. Moreover, the metabolism of terpenoids and polyketides, and transport and catabolism were significantly more abundant in eastern samples, while antibiotic-resistant pathways were enriched in western samples. The results have important significance to understand bacterial ecosystem of coastal water and promote water quality management and protection activity in Shenzhen. This study can also help developing an optimal strategy for the green economy development and the policy planning of Guangdong-Hong Kong-Macao Greater Bay Area.

Eukaryotic communities in coastal water from Shenzhen in South China.

Eukaryotic microorganisms are ubiquitous in the marine environment, and have a wide variety of ecosystem functions. Shenzhen is one of the most developed cities in South China, but the eukaryotic communities in the water along its coastlines remain poorly understood. The study applied 18S rRNA gene ITS (internal transcribed spacer) sequencing to identify the eukaryotic community from twenty sites of Shenzhen coast water. The alpha-diversity of the samples between these sites were significantly different, and the seawater of eastern coast had higher alpha-diversity compared to that of the western coast. The abundance of Chlorophyta was notably higher in the seawater of western coast, but Picozoa was relatively depleted. Specifically, Cryptocaryon, Pseudovorticella, and Cyclotella were significantly higher in the water of western coast, while Guinardia, Minutocellus, and Amoebophrya were increased in eastern samples. The spatially variations of eukaryotic microorganism community in the seawater of Shenzhen coast were associated with the water quality. The results have important significance for the understanding of coastal eukaryotic community, their interaction network, and build a foundation for future management and protection of coastal water quality.

Diversity, abundance, and distribution of anammox bacteria in shipping channel sediment of Hong Kong by analysis of DNA and RNA.

Anammox bacteria have been detected in various ecosystems, but their occurrence and community composition along the shipping channels have not been reported. In this study, anammox bacteria were recovered by PCR-amplified biomarker hzsB gene from the genomic DNA of the sediment samples. Phylogenetic tree revealed that Candidatus Scalindua and Ca. Brocadia dominated the anammox community of the Hong Kong channels; Ca. Scalindua spp. was present abundantly at the sites farther from the shore, whereas Ca. Jettenia and Ca. Kuenenia were detected as the minor members in the estuarine sediments near the shipping terminals. The highest values of Shannon-Wiener index and Chao1 were identified in the sediments along the Urmston road (UR), suggesting the highest α-diversity and species richness of anammox bacteria. PCoA analysis indicated that anammox bacterial communities along UR and Tai Hong (TH) channel were site-specific because these samples were grouped and clearly separated from the other samples. The maximum diversity of anammox bacteria was detected in UR samples, ranging from 6.28 × 105 to 1.28 × 106 gene copies per gram of dry sediment. A similar pattern of their transcriptional activities was also observed among these channels. Pearson's moment correlation and redundancy analysis indicated that NH4+-N was a strong factor shaping the community structure, which showed significant positive correlation with the anammox bacterial abundance and anammox transcriptional activities (p < 0.01, r > 0.8). Also, NH4+-N, (NO3- + NO2-)-N, and NH4+/NOX were additional key environmental factors that influenced the anammox community diversity and distribution. This study yields a better understanding of the ecological distribution of anammox bacteria and the dominant genera in selective niche.

Impacts of anthropogenic disturbances on microbial community of coastal waters in Shenzhen, South China.

During the urbanization, human activities have brought great changes to marine biodiversity and microbial communities of coastal water. Shenzhen is a coastal city that has developed rapidly over the past four decades, but the microbial communities and metabolic potential in offshore water are still not well characterized. Here, 16S rRNA gene V4-V5 sequencing was conducted to determine the microbial components from coastal waters in twenty selected areas of Shenzhen. The results showed a significant difference on the microbial composition between the western and eastern waters. Samples from western coast had more abundant Burkholderiaceae, Sporichthyaceae, Aeromonadaceae, and Methylophilaceae compared to eastern coast, and at the genus level, Candidatus Aquiluna, Aeromonas, Arcobacter, Ottowia and Acidibacter were significantly higher in western waters. There was also a notable difference within the western sample group, suggesting the taxa-compositional heterogeneity. Moreover, analysis of environmental factors and water quality revealed that salinity, pH and dissolved oxygen were relatively decreased in western samples, while total nitrogen, total phosphorus, chemical oxygen demand, and harmful marine vibrio were significantly increased compared to eastern waters. The results suggest the coastal waters pollution is more serious in western Shenzhen than eastern Shenzhen and the microbial communities are altered, which can be associated with anthropogenic disturbances.

Role of the Phytochemical Compounds like Modulators in Gut Microbiota and Oxidative Stress.

BACKGROUND: Currently, daily consumption of green herb functional food or medicinal herbs has increased as adopted by many people worldwide as a way of life or even as an alternative to the use of synthetic medicines. Phytochemicals, which are a series of compounds of relatively complex structures and restricted distribution in plants, usually perform the defensive functions for plants against insects, bacteria, fungi or other pathogenic factors. A series of studies have found their effectiveness in the treatment or prevention of systemic diseases such as autoimmune diseases, cancer, neurodegenerative diseases, Crohn's disease and so on. OBJECTIVE: This review systematizes the literature on the mechanisms of the phytochemicals that react against unique free radicals and prevent the oxidative stress and also summarizes their role in gut microbiota inhibiting bacterial translocation and damage to the intestinal barrier and improving the intestinal membrane condition. CONCLUSION: The gut microbiota modulation and antioxidant activities of the phytochemicals shall be emphasized on the research of the active principles of the phytochemicals.

Exploring possible associations of the intestine bacterial microbiome with the pre-weaned weight gaining performance of piglets in intensive pig production.

The pre-weaned weight gain is an important performance trait of pigs in intensive pig production. The bacterial microbiome inside the host is vital to host health and growth performance. The purpose of this study was to explore the possible associations of the intestinal microbiome with the pre-weaned weight gain in intensive pig production. In this study, several anatomical sites (jejunum, ileum, cecum, and colon) were examined for bacterial microbiome structure using 16S rRNA V4-V5 region sequencing with Illumina Miseq. The results showed that the microbial richness (estimated by Chao1 index) in jejunum was positively correlated with the pre-weaned weight gain. This study also revealed that the Firmicutes and Bacteroidetes in colon were the weight gaining-related phyla; while the Selenomonas and Moraxella in ileum and the Lactobacillus in both cecum and colon were the weight gaining-related genera for the pre-weaned piglets in intensive pig prodution. Several intra-microbial interactions within commensal microbiome correlated with the pre-weaned weight gain were excavated, as well. Overall, this study provides an expanded view of the commensal bacterial community inside four anatomical intestinal sites of the commercial piglets and the associations of the intestinal microbiome with the pre-weaned weight gaining performance in intensive pig production.

A More Comprehensive Community of Ammonia-Oxidizing Archaea (AOA) Revealed by Genomic DNA and RNA Analyses of amoA Gene in Subtropical Acidic Forest Soils.

Ammonia-oxidizing bacteria (AOB) and archaea (AOA) are the main nitrifiers which are well studied in natural environments, and AOA frequently outnumber AOB by orders especially in acidic conditions, making AOA the most promising ammonia oxidizers. The phylogeny of AOA revealed in related studies, however, often varied and hardly reach a consensus on functional phylotypes. The objective of this study was to compare ammonia-oxidizing communities by amoA gene and transcript based on both genomic DNA and RNA in extremely acidic forest soils (pH <4.5). Our results support the numerical and functional dominance of AOA over AOB in acidic soils as bacterial amoA gene and transcript were both under detection limits and archaeal amoA, in contrast, were abundant and responded to the fluctuations of environmental factors. Organic matter from tree residues was proposed as the main source of microbial available nitrogen, and the potential co-precipitation of dissolved organic matter (DOM) with soluble Al3+ species in acidic soil matrix may further restrict the amount of nitrogen sources required by AOB besides NH3/NH4+ equilibrium. Although AOA were better adapted to oligotrophic environments, they were susceptible to the toxicity of exchangeable Al3+. Phylotypes affiliated to Nitrososphaera, Nitrososphaera sister group, and Nitrosotalea were detected by amoA gene and transcript. Nitrosotalea devantaerra and Nitrososphaera sister group were the major AOA. Compared to the genomic DNA data, higher relative abundances of Nitrososphaera and Nitrososphaera sister group were recognized in amoA transcript inferred AOA communities, where Nitrosotalea relative abundance was found lower, implying the functional activities of Nitrososphaera sister group and Nitrososphaera were easily underestimated and Nitrosotalea did not attribute proportionally to nitrification in extremely acidic soils. Further comparison of the different AOA community compositions and relative abundance of each phylotypes revealed by amoA genes and transcripts make it possible to identify the functional AOA species and assess their ecological role in extremely acidic soils.

Analytical validation of a reverse transcriptase droplet digital PCR (RT-ddPCR) for quantitative detection of infectious hematopoietic necrosis virus.

Infectious hematopoietic necrosis virus (IHNV) is an important pathogen of salmonid fishes. A validated universal reverse transcriptase quantitative PCR (RT-qPCR) assay that can quantify levels of IHNV in fish tissues has been previously reported. In the present study, we adapted the published set of IHNV primers and probe for use in a reverse-transcriptase droplet digital PCR (RT-ddPCR) assay for quantification of the virus in fish tissue samples. The RT-ddPCR and RT-qPCR assays detected 13 phylogenetically diverse IHNV strains, but neither assay produced detectable amplification when RNA from other fish viruses was used. The RT-ddPCR assay had a limit of detection (LOD) equating to 2.2 plaque forming units (PFU)/µl while the LOD for the RT-qPCR was 0.2 PFU/µl. Good agreement (69.4-100%) between assays was observed when used to detect IHNV RNA in cell culture supernatant and tissues from IHNV infected rainbow trout (Oncorhynchus mykiss) and arctic char (Salvelinus alpinus). Estimates of RNA copy number produced by the two assays were significantly correlated but the RT-qPCR consistently produced higher estimates than the RT-ddPCR. The analytical properties of the N gene RT-ddPCR test indicated that this method may be useful to assess IHNV RNA copy number for research and diagnostic purposes. Future work is needed to establish the within and between laboratory diagnostic performance of the RT-ddPCR assay.

Identification of Infectious Salmon Anaemia Virus from Imported Iced Atlantic Salmons.

Infectious Salmon anaemia virus (ISAV) has become a threat to the salmon industry worldwide and has caused considerable economic loss. In the present study, 9 suspect cases of ISAV infection were identified from iced Atlantic salmons imported from Norway in 2014 through Shenzhen port (Shenzhen, China) using methods recommended by the World Organization for Animal Health. However, the results of virus isolation were negative., Based on the sequence analysis of ISAV segment 6, the 9 ISAV isolates belonged to the HPRO type, had high homology (98.3%~100.0%) and closest relationship with Norway strains. We identified the 9 positive HPRO ISAVs from 491 iced Atlantic salmons (1. 8%). Therefore, we should strengthen the quarantine of iced Atlantic salmons from Norway in case of HPRO ISAV into China.

Determination of the complete genome sequence of infectious hematopoietic necrosis virus (IHNV) Ch20101008 and viral molecular evolution in China.

This study determined the complete genomic sequence of the infectious hematopoietic necrosis virus (IHNV) strain Ch20101008 isolated from farmed brook trout (Salvelinus fontinalis) that died from a disease caused by the virus in northeast China. The sequence was determined from 10 overlapping clones obtained through RT-PCR amplification. The whole genome length of Ch20101008 comprised 11,129 nucleotides (nt), and the overall organization was typical of that observed for all other IHNV strains. The phylogenetic analysis results of the 65 IHNV glycoprotein genes and 47 IHNV partial nucleoprotein genes presented five major genogroups (J, U, L, E and M). The J genogroup included the J Nagano and J Shizuoka subgroups. The IHNV Ch20101008 strain belonged to the J Nagano subgroup of the J genogroup and was significantly related to other Chinese IHNV strains. All Chinese IHNV isolates are monophyletic, with a recent common ancestor, except for the BjLL strain. The N, P, M, G, NV and L genes of Ch20101008 were compared with the available IHNV sequences in GenBank. The results indicated that 198 nt were substituted, 53 of which exhibited amino acid change in the Ch20101008 genome. An adenine nucleotide deletion was found at position 4959 of the 5' UTR of the L gene. In the G gene, specific nucleotides and amino acid variations of the Chinese IHNV strains were observed when compared with 23 isolates from other countries. Of the 15 nucleotide sites that changed, seven resulted in amino acid substitution. The data further demonstrated that the J genogroup IHNV was introduced to and evolved in salmon farm environments in China.

Selection and characterization of single-chain recombinant antibodies against infectious haematopoietic necrosis virus from mouse phage display library.

Six single-chain fragment variable (scFv) antibodies against infectious haematopoietic necrosis virus (IHNV) were selected from an antibody phage display library by phage display technology. The soluble scFv antibodies showed a molecular weight 32kDa by Western blot. Dot blot analysis revealed that the six scFv antibodies could recognize IHNV. For enzyme linked immunosorbent assay (ELISA), four scFv antibodies (P1A4, P1A12, P1D5 and P3E2) showed cross-reactivity with spring viraemia of carp virus (SVCV). However, none of the six scFv antibodies had cross-reaction with Pike fry rhabdovirus (PFRV), Soft-shelled turtle iridovirus (STIV), viral haemorrhagic septicemia virus (VHSV), or viral nervous necrosis virus (VNNV). Indirect immunofluorescence results showed that all of these scFv antibodies reacted positively with virus in the IHNV-infected cells. These scFv antibodies will be useful in diagnostic test development and pathogenesis studies for IHNV.

Development and evaluation of a loop-mediated isothermal amplification assay for diagnosis of Cyprinid herpesvirus 2.

Goldfish Haematopoietic Necrosis caused by Cyprinid herpesvirus 2 (CyHV-2) is a severe fish disease with high level of mortality. This is the first study on detection of this disease by loop-mediated isothermal amplification (LAMP). A set of six primers targeting terminase gene (accession no. EU349285.1) was determined after a serial of tests. Detection limit was 1.09 × 10(-4)µg/µL, which was superior to conventional PCR and real-time PCR. No cross reaction with 28 other viruses or bacteria commonly found in fish was observed. The application of commercial kit and instrument for the LAMP assay could reduce the risk of cross contamination, which is suitable for detection of infection under field conditions.

Selection and characterization of single-chain recombinant antibodies against spring viraemia of carp virus from mouse phage display library.

Antibody-displaying phage library was selected after three rounds of panning against spring viraemia of carp virus (SVCV) by phage display technology. Eight positive clones which could produce soluble single-chain fragment variable (scFv) antibody induced by isopropyl-beta-d-thiogalactopyranoside (IPTG) were obtained. Dot blot results showed that the eight scFv antibodies could recognize SVCV. The soluble scFv antibodies showed a molecular weight 29 kD by Western blot. All scFv antibodies could recognize SVCV proteins specifically without cross-reaction with other virus proteins by ELISA. Indirect immunofluorescence results showed that all of these scFv antibodies reacted positively with virus in the SVCV-infected cells. These scFv antibodies will be useful tools to establish immunological detection methods for SVCV.

Development of a novel optical biosensor for detection of organophosphorus pesticides based on methyl parathion hydrolase immobilized by metal-chelate affinity.

We have developed a novel optical biosensor device using recombinant methyl parathion hydrolase (MPH) enzyme immobilized on agarose by metal-chelate affinity to detect organophosphorus (OP) compounds with a nitrophenyl group. The biosensor principle is based on the optical measurement of the product of OP catalysis by MPH (p-nitrophenol). Briefly, MPH containing six sequential histidines (6 × His tag) at its N-terminal was bound to nitrilotriacetic acid (NTA) agarose with Ni ions, resulting in the flexible immobilization of the bio-reaction platform. The optical biosensing system consisted of two light-emitting diodes (LEDs) and one photodiode. The LED that emitted light at the wavelength of the maximum absorption for p-nitrophenol served as the signal light, while the other LED that showed no absorbance served as the reference light. The optical sensing system detected absorbance that was linearly correlated to methyl parathion (MP) concentration and the detection limit was estimated to be 4 µM. Sensor hysteresis was investigated and the results showed that at lower concentration range of MP the difference got from the opposite process curves was very small. With its easy immobilization of enzymes and simple design in structure, the system has the potential for development into a practical portable detector for field applications.

Microbial community analysis of fresh and old microbial biofilms on Bayon temple sandstone of Angkor Thom, Cambodia.

The temples of Angkor monuments including Angkor Thom and Bayon in Cambodia and surrounding countries were exclusively constructed using sandstone. They are severely threatened by biodeterioration caused by active growth of different microorganisms on the sandstone surfaces, but knowledge on the microbial community and composition of the biofilms on the sandstone is not available from this region. This study investigated the microbial community diversity by examining the fresh and old biofilms of the biodeteriorated bas-relief wall surfaces of the Bayon Temple by analysis of 16S and 18S rRNA gene sequences. The results showed that the retrieved sequences were clustered in 11 bacterial, 11 eukaryotic and two archaeal divisions with disparate communities (Acidobacteria, Actinobacteria, Bacteroidetes, Cyanobacteria, Proteobacteria; Alveolata, Fungi, Metazoa, Viridiplantae; Crenarchaeote, and Euyarchaeota). A comparison of the microbial communities between the fresh and old biofilms revealed that the bacterial community of old biofilm was very similar to the newly formed fresh biofilm in terms of bacterial composition, but the eukaryotic communities were distinctly different between these two. This information has important implications for understanding the formation process and development of the microbial diversity on the sandstone surfaces, and furthermore to the relationship between the extent of biodeterioration and succession of microbial communities on sandstone in tropic region.

Removal of methyl parathion from artificial off-gas using a bioreactor containing a constructed microbial consortium.

Methyl parathion (MP), a highly toxic organophosphorus pesticide, was widely used for agriculture crop protection. During the production of MP and the process of MP-containing wastewater treatment, MP can release into the atmosphere and will do great harm to adjacent communities. A consortium comprised of an engineered microorganism and a natural p-nitrophenol (PNP) degrader was assembled for complete mineralization of MP. We genetically engineered Escherichia coli BL21 (DE3) enabling the overexpression of methyl parathion hydrolase (MPH). In addition, we isolated Ochrobactrum sp. strain LL-1 that utilized PNP, a product of MP hydrolysis, as the sole carbon, nitrogen, and energy source. The coculture effectively hydrolyzed 0.2 mM MP and prevented the accumulation of PNP in suspended culture. A laboratory-scale bioreactor containing the dual-species consortium was developed for the treatment of artificial off-gas containing MP. The bioreactor maintained over 98% of average MP removal efficiency over a 75 day period, and PNP produced from hydrolysis of MP was degraded completely, indicating that complete mineralization of MP was achieved. The strategy of linking degrading consortium to a bioreactor may provide an alternative to physicochemical abatement technologies for the treatment of waste-gas streams containing MP as well as other PNP-substituted organophosphates.

Expression and characterization of carboxylesterase E4 gene from peach-potato aphid (Myzus persicae) for degradation of carbaryl and malathion.

An abridged carboxylesterase E4 (CbE E4) gene was cloned from the peach-potato aphid, Myzus persicae, by reverse transcription-PCR and subcloned into the expression vector pET28b. The abridged CbE E4 gene was successfully expressed in E. coli BL21 (DE3). The recombinant CbE E4 hydrolyzed beta-naphthyl acetate and Carbaryl by 64% within 2.5 h, Malathion by 80% within 1.25 h. However, the hydrolysis of other pesticides (Dichlorovos, Parathion, Pirimicarb and Deltamethrin) was not detected.