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Climate change may increase the vulnerability of aquifers to contamination through extreme precipitation and extended drought periods. Therefore, the understanding of groundwater ecosystem dynamics is crucial, with bacterial assemblages playing a major role in biogeochemical cycles. The present research describes a geospatial study targeting the bacterial community structure of groundwaters from the largest karst aquifer in Portugal (the Maciço Calcário Estremenho), integrating hydrogeochemical and bacterial diversity data. A total of 22 samples were analyzed from a set of 11 geographically sparsely distributed groundwater sources in dry vs. wet seasons. The 16S rRNA gene barcoding data revealed bacterial community variability across samples in space and time. The phylum Proteobacteria was dominant across all samples (from 44 to 92% of total sequence reads), mainly represented by the classes Alphaproteobacteria (orders Sphingomonadales, BD7–3, Rhizobiales and Rhodospirillales), Betaproteobacteria (orders Burkholderiales, Rhodocyclales, Nitrosomonadales), Gammaproteobacteria (orders Pseudomonadales, Xanthomonadales, Alteromonadales, Legionellales) and Deltaproteobacteria (orders Myxococcales, Spirobacillales). Variation in the bacterial community was primarily attributed to parameters such as redox conditions (DO, ORP), Fe, Mn, SO4, PO4, Sr and Cl, but also some minor and trace elements (Al, V, Cr, Cu, Pb). Our results provide novel insights into bacterial diversity in relation to groundwater hydrogeochemistry. The strong dominance of OTUs related to bacterial taxa associated with nitrification/denitrification also highlights a potentially important role of these assemblages on nutrients (nitrogen sources) and groundwater quality dynamics at this karstic aquifer system. Moreover, the integration of bacterial assemblages information is emphasized as central for water quality monitoring programs. [ABSTRACT FROM AUTHOR]

Of the estimated 55 Hawaiian honeycreepers (subfamily Carduelinae) only 17 species remain, nine of which the International Union for Conservation of Nature considers endangered. Among the most pressing threats to honeycreeper survival is avian malaria, caused by the introduced blood parasite Plasmodium relictum, which is increasing in distribution in Hawaiʻi as a result of climate change. Preventing further honeycreeper decline will require innovative conservation strategies that confront malaria from multiple angles. Research on mammals has revealed strong connections between gut microbiome composition and malaria susceptibility, illuminating a potential novel approach to malaria control through the manipulation of gut microbiota. One honeycreeper species, Hawaiʻi ʻamakihi (Chlorodrepanis virens), persists in areas of high malaria prevalence, indicating they have acquired some level of immunity. To investigate if avian host‐specific microbes may be associated with malaria survival, we characterized cloacal microbiomes and malaria infection for 174 ʻamakihi and 172 malaria‐resistant warbling white‐eyes (Zosterops japonicus) from Hawaiʻi Island using 16S rRNA gene metabarcoding and quantitative polymerase chain reaction. Neither microbial alpha nor beta diversity covaried with infection, but 149 microbes showed positive associations with malaria survivors. Among these were Escherichia and Lactobacillus spp., which appear to mitigate malaria severity in mammalian hosts, revealing promising candidates for future probiotic research for augmenting malaria immunity in sensitive endangered species. [ABSTRACT FROM AUTHOR]

Microbial communities provide essential information about host ecology and could be helpful as a tool to improve species conservation efforts. However, microbes can also infect and compromise the host development process and viability. Caretta caretta is the most widespread marine turtle species in the Mediterranean basin and is the only species of sea turtle nesting along the Italian coasts. Little is known about the microbiota composition of the nest of sea turtles and its correlation with hatching failures. In this study, the microbial composition of two nests of C. caretta featuring different rates of hatching success from a nesting beach in Lampedusa (Italy) was analyzed and compared. The bacterial community was determined using culture-dependent methods and next-generation sequencing based on 16S rRNA gene metabarcoding analysis. Our results showed five dominant bacterial phyla (Proteobacteria, Bacteroidetes, Actinobacteria, Verrucomicrobia, and Firmicutes) and indicated different bacterial families (Pseudomonadaceae and Brucellaceae) as likely causes of hatching failures. Besides, our findings demonstrated the nests' active role in modulating the sand's bacterial communities. This study suggests microbiological analysis could be a valuable tool in monitoring nests to take preventive actions and reduce hatching failures. [ABSTRACT FROM AUTHOR]

Endophytic prokaryotes, bacteria, and archaea, are important microorganisms that benefit host plants by promoting plant growth and reducing stress. The objective of this study was to evaluate temporal shifts in the root endophytic prokaryotic communities associated with carrots (Daucus carota subsp. sativus) and the effect of organic matter application on them. Carrots were grown in a planter under five fertilizer treatments (weed compost, bark compost, cattle manure, chemical fertilizer, and no-fertilizer control) and the compositions of rhizosphere and root endosphere prokaryotic communities were determined via amplicon sequencing analysis targeting the 16S rRNA gene at 60 and 108 days after sowing. The results showed that the rhizosphere prokaryotic community compositions were stable despite different sampling times and fertilizer treatments; however, a greater temporal shift and an effect of the type of organic matter applied were observed in the endosphere prokaryotic communities. The differences in treatments resulted in significant differences in the abundance and Faith pyrogenetic diversity of the endosphere prokaryotic community. Genera, such as Burkholderia, Sphingomonas, and Rhodanobacter, that exhibit plant-growth-promoting and biocontrol activities, were detected regardless of the treatments, suggesting that they may play an important ecological role as the core endophytes in carrot roots. [ABSTRACT FROM AUTHOR]

Climate change may increase the vulnerability of aquifers to contamination through extreme precipitation and extended drought periods. Therefore, the understanding of groundwater ecosystem dynamics is crucial, with bacterial assemblages playing a major role in biogeochemical cycles. The present research describes a geospatial study targeting the bacterial community structure of groundwaters from the largest karst aquifer in Portugal (the Maciço Calcário Estremenho), integrating hydrogeochemical and bacterial diversity data. A total of 22 samples were analyzed from a set of 11 geographically sparsely distributed groundwater sources in dry vs. wet seasons. The 16S rRNA gene barcoding data revealed bacterial community variability across samples in space and time. The phylum Proteobacteria was dominant across all samples (from 44 to 92% of total sequence reads), mainly represented by the classes Alphaproteobacteria (orders Sphingomonadales, BD7–3, Rhizobiales and Rhodospirillales), Betaproteobacteria (orders Burkholderiales, Rhodocyclales, Nitrosomonadales), Gammaproteobacteria (orders Pseudomonadales, Xanthomonadales, Alteromonadales, Legionellales) and Deltaproteobacteria (orders Myxococcales, Spirobacillales). Variation in the bacterial community was primarily attributed to parameters such as redox conditions (DO, ORP), Fe, Mn, SO4, PO4, Sr and Cl, but also some minor and trace elements (Al, V, Cr, Cu, Pb). Our results provide novel insights into bacterial diversity in relation to groundwater hydrogeochemistry. The strong dominance of OTUs related to bacterial taxa associated with nitrification/denitrification also highlights a potentially important role of these assemblages on nutrients (nitrogen sources) and groundwater quality dynamics at this karstic aquifer system. Moreover, the integration of bacterial assemblages information is emphasized as central for water quality monitoring programs.

Climate change is enhancing the frequency of cyanobacterial blooms not only during summer but also in spring and autumn, leading to increased ecological impacts. The bacterioplankton community composition (BCC), in particular, is deeply affected by these blooms, although at the same time BCC can also play important roles in blooms' dynamics. However, more information is still needed regarding BCC during species-specific cyanobacterial blooms. The goal of this study was to assess BCC succession in a hypereutrophic shallow lake (Vela Lake, Portugal) during a warm spring using a metagenomic approach to provide a glimpse of the changes these communities experience during the dominance of Aphanizomenon-like bloom-forming species. BCC shifts were studied using 16S rRNA gene metabarcoding and multivariate analyses. A total of 875 operational taxonomic units (OTUs) were retrieved from samples. In early spring, the dominant taxa belonged to Proteobacteria (mainly Alphaproteobacteria—Rickettsiales) and Bacteroidetes (Saprospirales, Flavobacteriales and Sphingobacteriales). However, at the end of May, a bloom co-dominated by cyanobacterial populations of Aphanizomenon gracile, Sphaerospermopsis aphanizomenoides and Synechococcus sp. developed and persisted until the end of spring. This led to a major BCC shift favouring the prevalence of Alphaproteobacteria (Rickettsiales and also Rhizobiales, Caulobacteriales and Rhodospirillales) and Bacteroidetes (Saprospirales, followed by Flavobacteriales and Sphingobacteriales). These results contribute to the knowledge of BCC dynamics during species-specific cyanobacterial blooms, showing that BCC is strongly affected (directly or indirectly) by Aphanizomenon-Sphaerospermopsis blooms. [ABSTRACT FROM AUTHOR]

Endophytic prokaryotes, bacteria, and archaea, are important microorganisms that benefit host plants by promoting plant growth and reducing stress. The objective of this study was to evaluate temporal shifts in the root endophytic prokaryotic communities associated with carrots (Daucus carota subsp. sativus) and the effect of organic matter application on them. Carrots were grown in a planter under five fertilizer treatments (weed compost, bark compost, cattle manure, chemical fertilizer, and no-fertilizer control) and the compositions of rhizosphere and root endosphere prokaryotic communities were determined via amplicon sequencing analysis targeting the 16S rRNA gene at 60 and 108 days after sowing. The results showed that the rhizosphere prokaryotic community compositions were stable despite different sampling times and fertilizer treatments; however, a greater temporal shift and an effect of the type of organic matter applied were observed in the endosphere prokaryotic communities. The differences in treatments resulted in significant differences in the abundance and Faith pyrogenetic diversity of the endosphere prokaryotic community. Genera, such as Burkholderia, Sphingomonas, and Rhodanobacter, that exhibit plant-growth-promoting and biocontrol activities, were detected regardless of the treatments, suggesting that they may play an important ecological role as the core endophytes in carrot roots.

The present study aimed to characterize the genetic structure of Artemisia eriantha Ten. and the diversity of the rhizosphere microbiota. Plant leaves and rhizosphere soils were sampled from three areas of Central Italy, namely Monte Corvo, Monte Portella (both from the Gran Sasso massif), and Monte Focalone (Majella massif). The plant samples were subjected to genetic structure analysis by amplified fragment length polymorphism (AFLP) markers. The microbiota from the rhizosphere soils was investigated by 16S rRNA gene metabarcoding. The within and among population variability was typical of outbreeding species. The AFLP polymorphisms revealed a marked closeness among plant populations collected in Monte Focalone and Monte Corvo, despite the geographical proximity of the latter with Monte Portella, a result confirmed by cluster, STRUCTURE, and discriminant analyses. 16S rRNA gene metabarcoding showed higher values of diversity for Monte Corvo (H, 5.7; Chao1, 445) and Monte Focalone (H′, 5.57; Chao1, 446) than Monte Portella (H′, 5.3; Chao1, 275). At the phylum level, the communities were mainly represented by Proteobacteria, Actinobacteria, and Acidobacteria (>10%). At the genus level, the Monte Focalone and Monte Corvo microbiotas were closer than Monte Portella, thus confirming the results from the plant communities. The findings provided evidence for the first time of an association between the Artemisia eriantha plant and microbiota communities. The relevance of the results in terms of biodiversity and the conservation strategies of plant and microbiota communities in the Central Apennines are discussed. [ABSTRACT FROM AUTHOR]

The SARS-CoV-2 presence and the bacterial community profile in air samples collected at the Intensive Care Unit (ICU) of the Operational Unit of Infectious Diseases of Santa Caterina Novella Hospital in Galatina (Lecce, Italy) have been evaluated in this study. Air samplings were performed in different rooms of the ICU ward with and without COVID-19 patients. No sample was found positive to SARS-CoV-2, according to Allplex 2019-nCoV Assay. The airborne bacterial community profiles determined by the 16S rRNA gene metabarcoding approach up to the species level were characterized by richness and biodiversity indices, Spearman correlation coefficients, and Principal Coordinate Analysis. Pathogenic and non-pathogenic bacterial species, also detected in outdoor air samples, were found in all collected indoor samples. Staphylococcus pettenkoferi, Corynebacterium tuberculostearicum, and others coagulase-negative staphylococci, detected at high relative abundances in all the patients' rooms, were the most abundant pathogenic species. The highest mean relative abundance of S. pettenkoferi and C. tuberculostearicum suggested that they were likely the main pathogens of COVID-19 patients at the ICU ward of this study. The identification of nosocomial pathogens representing potential patients' risks in ICU COVID-19 rooms and the still controversial airborne transmission of the SARS-CoV-2 are the main contributions of this study. [ABSTRACT FROM AUTHOR]

Simple Summary: "Infernaccio" gorges are one of the Earth's hidden habitats in Central Italy. Beyond the deep incisions and high slopes, these gorges are characterized by black deposits in gorge walls and covering rock surfaces. Several geological events have shaped these unique geological formations and their microbiota. This study investigated microbial contribution to black deposit formation and isolating Mn-oxide-solubilizing bacteria. Our results provided evidence of the putative role of Bacteria and Archaea in forming manganese oxide deposits. Findings also showed that these deposits are a source of valuable strains with manganese oxide bioleaching properties, essential for bioremediation and metal recovery. The present study explored the microbial diversity of black deposits found in the "Infernaccio" gorge. X-ray Powdered Diffraction (XRPD) was used to investigate the crystallinity of the samples and to identify the minerals. Scanning electron microscope and energy-dispersive X-ray spectroscopy (SEM-EDS) were used to detect the bacterial imprints, analyze microbe–mineral interactions, and highlight the chemical element distribution in the black deposits. 16S rRNA gene metabarcoding allowed the study of Archaea and Bacteria communities. Mn-oxide-solubilizing isolates were also obtained and characterized by culturable and molecular approaches. The multidisciplinary approach showed the occurrence of deposits composed of birnessite, diopside, halloysite, and leucite. Numerous bacterial imprints confirmed the role of microorganisms in forming these deposits. The Bacteria and Archaea communities associated with these deposits and runoff waters are dynamic and shaped by seasonal changes. The uncultured and unknown taxa are the most common and abundant. These amplicon sequence variants (ASVs) were mainly assigned to Proteobacteria and Bacteroidetes phyla. Six isolates showed interesting Mn solubilization abilities under microaerophilic conditions. Molecular characterization associated isolates to Brevibacterium, Bacillus, Neobacillus, and Rhodococcus genera. The findings enriched our knowledge of geomicrobiological aspects of one of the Earth's hidden habitats. The study also unveiled the potential of this environment as an isolation source of biotechnologically relevant bacteria. [ABSTRACT FROM AUTHOR]

This study aimed to estimate the green formation lampenflora of “Stiffe” caves in order to evaluate their suitability as an isolation source of cyanobacteria useful for the production of polyhydroxyalkanoates (PHAs). The cave system was chosen as the sampling site due to its touristic use and the presence of high-impact illuminations. The biofilms and the mats of the illuminated walls were sampled. Samples were investigated by 16S rRNA gene analysis and culturable cyanobacteria isolation. The isolated strains were then screened for the production of PHAs under typical culturing and nutritional starvation. Cultures were checked for PHA accumulation, poly-β-hydroxybutyrate (PHB) presence (infrared spectroscopy), and pigment production. The 16S rRNA gene metabarcoding. Highlighted a considerable extent of the pressure exerted by anthropogenic activities. However, the isolation yielded eleven cyanobacteria isolates with good PHA (mainly PHB)-producing abilities and interesting pigment production rates (chlorophyll a and carotenoids). Under normal conditions (BG110), the accumulation abilities ranged from 266 to 1,152 ng mg dry biomass–1. The optimization of bioprocesses through nutritional starvation resulted in a 2.5-fold increase. Fourier transform infrared (FTIR) studies established the occurrence of PHB within PHAs extracted by cyanobacteria isolates. The comparison of results with standard strains underlined good production rates. For C2 and C8 strains, PHA accumulation rates under starvation were higher than Azospirillum brasilense and similar to Synechocystis cf. salina 192. This study broadened the knowledge of the microbial communities of mats and biofilms on the lightened walls of the caves. These findings suggested that these structures, which are common in tourist caves, could be used to isolate valuable strains before remediation measures are adopted. [ABSTRACT FROM AUTHOR]

This study aimed to estimate the green formation lampenflora of “Stiffe” caves in order to evaluate their suitability as an isolation source of cyanobacteria useful for the production of polyhydroxyalkanoates (PHAs). The cave system was chosen as the sampling site due to its touristic use and the presence of high-impact illuminations. The biofilms and the mats of the illuminated walls were sampled. Samples were investigated by 16S rRNA gene analysis and culturable cyanobacteria isolation. The isolated strains were then screened for the production of PHAs under typical culturing and nutritional starvation. Cultures were checked for PHA accumulation, poly-β-hydroxybutyrate (PHB) presence (infrared spectroscopy), and pigment production. The 16S rRNA gene metabarcoding. Highlighted a considerable extent of the pressure exerted by anthropogenic activities. However, the isolation yielded eleven cyanobacteria isolates with good PHA (mainly PHB)-producing abilities and interesting pigment production rates (chlorophyll a and carotenoids). Under normal conditions (BG110), the accumulation abilities ranged from 266 to 1,152 ng mg dry biomass–1. The optimization of bioprocesses through nutritional starvation resulted in a 2.5-fold increase. Fourier transform infrared (FTIR) studies established the occurrence of PHB within PHAs extracted by cyanobacteria isolates. The comparison of results with standard strains underlined good production rates. For C2 and C8 strains, PHA accumulation rates under starvation were higher than Azospirillum brasilense and similar to Synechocystis cf. salina 192. This study broadened the knowledge of the microbial communities of mats and biofilms on the lightened walls of the caves. These findings suggested that these structures, which are common in tourist caves, could be used to isolate valuable strains before remediation measures are adopted.

Bacteria are vital components of lake systems, driving a variety of biogeochemical cycles and ecosystem services. Bacterial communities have been shown to have a skewed distribution with a few abundant species and a large number of rare species. The contribution of environmental processes or geographic distance in structuring these components is uncertain. The discrete nature of lakes provides an ideal test case to investigate microbial biogeographical patterns. In the present study, we used 16S rRNA gene metabarcoding to examine the distribution patterns on local and regional scales of abundant and rare planktonic bacteria across 167 New Zealand lakes covering broad environmental gradients. Only a few amplicon sequence variants (ASVs) were abundant with a higher proportion of rare ASVs. The proportion of locally abundant ASVs was negatively correlated with the percentage of high productivity grassland in the catchment and positively with altitude. Regionally rare ASVs had a restricted distribution and were only found in one or a few lakes. In general, regionally abundant ASVs had higher occupancy rates, although there were some with restricted occupancy. Environmental processes made a higher contribution to structuring the regionally abundant community, while geographic distances were more important for regionally rare ASVs. A better understanding of the processes structuring the abundance and distribution of bacterial communities within lakes will assist in understand microbial biogeography and in predicting how these communities might shift with environmental change. [ABSTRACT FROM AUTHOR]

Bacteria are vital components of lake systems, driving a variety of biogeochemical cycles and ecosystem services. Bacterial communities have been shown to have a skewed distribution with a few abundant species and a large number of rare species. The contribution of environmental processes or geographic distance in structuring these components is uncertain. The discrete nature of lakes provides an ideal test case to investigate microbial biogeographical patterns. In the present study, we used 16S rRNA gene metabarcoding to examine the distribution patterns on local and regional scales of abundant and rare planktonic bacteria across 167 New Zealand lakes covering broad environmental gradients. Only a few amplicon sequence variants (ASVs) were abundant with a higher proportion of rare ASVs. The proportion of locally abundant ASVs was negatively correlated with the percentage of high productivity grassland in the catchment and positively with altitude. Regionally rare ASVs had a restricted distribution and were only found in one or a few lakes. In general, regionally abundant ASVs had higher occupancy rates, although there were some with restricted occupancy. Environmental processes made a higher contribution to structuring the regionally abundant community, while geographic distances were more important for regionally rare ASVs. A better understanding of the processes structuring the abundance and distribution of bacterial communities within lakes will assist in understand microbial biogeography and in predicting how these communities might shift with environmental change.

The present study explored the microbial diversity of black deposits found in the “Infernaccio” gorge. X-ray Powdered Diffraction (XRPD) was used to investigate the crystallinity of the samples and to identify the minerals. Scanning electron microscope and energy-dispersive X-ray spectroscopy (SEM-EDS) were used to detect the bacterial imprints, analyze microbe–mineral interactions, and highlight the chemical element distribution in the black deposits. 16S rRNA gene metabarcoding allowed the study of Archaea and Bacteria communities. Mn-oxide-solubilizing isolates were also obtained and characterized by culturable and molecular approaches. The multidisciplinary approach showed the occurrence of deposits composed of birnessite, diopside, halloysite, and leucite. Numerous bacterial imprints confirmed the role of microorganisms in forming these deposits. The Bacteria and Archaea communities associated with these deposits and runoff waters are dynamic and shaped by seasonal changes. The uncultured and unknown taxa are the most common and abundant. These amplicon sequence variants (ASVs) were mainly assigned to Proteobacteria and Bacteroidetes phyla. Six isolates showed interesting Mn solubilization abilities under microaerophilic conditions. Molecular characterization associated isolates to Brevibacterium, Bacillus, Neobacillus, and Rhodococcus genera. The findings enriched our knowledge of geomicrobiological aspects of one of the Earth’s hidden habitats. The study also unveiled the potential of this environment as an isolation source of biotechnologically relevant bacteria.

Diverse communities of bacterial endophytes inhabit plant tissues, and these bacteria play important roles for plant growth and health. Cherry laurel (Prunus laurocerasus L.) is a broadleaf evergreen shrub that is widely grown in temperate zones for its ornamental and medicinal properties, however virtually nothing is known about its associated bacterial community. In this study, we analysed the matured one-year-old leaves of this plant using Illumina-based 16S rRNA gene metabarcoding to reveal the community structure of endophytic bacteria and understand its shifts during the seasonal transition from winter dormancy to a spring vegetative state. The overall community was composed of four dominant phyla (Proteobacteria, Actinobacteria, Firmicutes, Bacteroidetes). Corynebacterium, Acinetobacter, and Chryseobacterium genera were the most prevalent bacteria, comprising 13.3%, 6.9%, and 6.8% of the amplicon sequence variants (ASVs), respectively. The ASV richness and diversity increased significantly in May as compared to other sampling months (February, March, and April). We observed high variation in the overall community structure of endophytic bacteria among collection dates. The variation was only reflected by a few core community members, suggesting that the changes of the endophytic community during winter/spring seasonal transition are mostly associated with the less abundant community members. We identified biomarker taxa for late winter, mid spring, and late spring collection dates. This study is the first one to report on the diversity and composition of bacterial endophytes in the leaves of cherry laurel and its shifts across the dormancy-to-vegetative seasonal transition.

Aim: The high-throughput sequencing methods have revolutionized the study of the microbiota in different matrices including those of the grapevine production chain. DNA extraction is a crucial step in the sample processing. In this study, we compared different DNA purification methods and two primer sets for 16S rRNA gene metabarcoding to evaluate the best protocol to explore the wine microbiota by metabarcoding. Methods and results: We collected a wine from Barbera grapes after malolactic fermentation previously inoculated by Oenococcus oeni starter. The same sample was used to evaluate the best performing protocol to study the wine microbiota. DNA was purified using nine different methods and then amplified for the 16S rRNA gene with two primer sets (according to Illumina or Earth Microbiome Project protocols). The obtained amplicons were then sequenced in a single sequencing session on an Illumina MiSeq. We evaluated the best protocol considering DNA concentration and purity, alpha (Observed species) and beta diversity from metabarcoding analysis. The sequencing generated 36,031,756 reads in total. Although no statistically significant difference was observed between purification methods or primer sets, better results were obtained with phenol-chloroform DNA purification combined to Earth Microbiome Project primers. Metabarcoding was able to highlight the domination of the inoculum, O. oeni, representing the main species of the analyzed wine microbiota. Conclusion: Our data show that, for the tested wine, metabarcoding output is more influenced by the primer set than by the DNA purification method. Moreover, the metabarcoding detected that O. oeni represents the main species, evidencing the domination of the inoculum done with lyophilized commercial preparation of this species. Other lactic acid bacteria are present at a much lower abundance. Significance and impact of the study: This is the first report applying the 16S rRNA gene metabarcoding to study the microbiota of wine. For this reason, the evaluation of alternative methods for DNA processing is essential for future research using this innovative methodology.

Soil bacterial community structure has traditionally been measured using phospholipid fatty acid (PLFA) profiling. However, with the development of high-throughput sequencing technologies and metabarcoding techniques, more studies are now using 16S rRNA gene metabarcoding to measure bacterial community structure. Metabarcoding provides a much greater level of detail than PLFA profiling does, but it remains unclear whether or not the two techniques have a similar ability to answer many of the common questions asked by ecologists. We test the relative ability of the two techniques to quantify differences in bacterial community structure among five land uses (natural and planted forest, unimproved and improved grasslands, and vineyards), and to predict ecosystem functions. We also test whether PLFA- and metabarcoding-based metrics indicative of microbial growth strategies are correlated to each other. We show that both techniques showed broadly similar patterns of bacterial community composition change with land use and a remarkably similar ability to predict a wide range of ecosystem functions (carbon and nutrient cycling, and responses to drought). However, they were also complementary, as each showed different strengths in discriminating land uses and predicting ecosystem functions. PLFA metrics (i.e. the gram-positive:gram-negative ratio and fungal:bacterial ratio) were strongly correlated with the equivalent 16S rRNA gene metabarcoding metrics (i.e. the gram-positive:gram-negative and oligotrophic:copiotrophic ratios), although PLFA metrics were less well correlated with the Proteobacteria:Acidobacteria ratio. For many ecological questions the two techniques thus give broadly comparable results, providing confidence in the ability of both techniques to quantify meaningful changes in bacterial communities. [ABSTRACT FROM AUTHOR]

We investigated the possibility of bacterial symbiosis in Globigerina bulloides, a palaeoceanographically important, planktonic foraminifer. This marine protist is commonly used in investigations of climatically sensitive subpolar and temperate water masses and wind driven upwelling regions of the world's oceans. G. bulloides is unusual because it lacks the protist algal symbionts that are often found in other spinose species and has an atypical geochemical shell signature. This is suggestive of a divergent ecology, making it a good candidate for investigating 5 the potential for bacterial symbiosis as a contributory factor in shell calcification. Such ecological information is essential to evaluate fully the potential response of G. bulloides to ocean acidification and climate change. To investigate the ecological interactions between G. bulloides and bacterial populations in the water column, 18S rRNA gene sequencing, fluorescence microscopy, 16S rRNA gene metabarcoding and transmission electron microscopy (TEM) were performed on individual specimens of G. bulloides (Type IId) collected from two locations in the California Current. Intracellular DNA extracted from five G. bulloides specimens was subjected to 16S rRNA gene metabarcoding and, remarkably, 37-87 % of all 16S rRNA gene sequences recovered were assigned to operational taxonomic units (OTUs) of the picocyanobacterium, Synechococcus. This finding was supported by TEM observations of intact Synechococcus cells in both the cytoplasm and vacuoles of G. bulloides. Concentrations were up to four orders of magnitude greater inside the foraminifera than those reported for the Californian Current water column and approximately 5 % of the intracellular Synechococcus cells observed were dividing. This suggests that Synechococcus is an endobiont of G. bulloides Type IId, which is the first report of a bacterial endobiont in the planktonic foraminifera. We consider the potential roles of Synechococcus and G. bulloides within the relationship and the need to determine how widespread the possible symbiotic association is within the widely distributed G. bulloides morphospecies. The possible influence of Synechococcus respiration on G. bulloides shell geochemistry is also discussed. [ABSTRACT FROM AUTHOR]

• Microscopy and metabarcoding reveal similar cyanobacteria communities in lakes. • Methods agree most at broader levels of taxonomy and within eutrophic sites. • Metabarcoding is far more dominated by picocyanobacteria. • Communities generated by each method are complementary as opposed to identical. • Microcystis genotypes structure along a nutrient gradient. Accurately identifying the species present in an ecosystem is vital to lake managers and successful bioassessment programs. This is particularly important when monitoring cyanobacteria, as numerous taxa produce toxins and can have major negative impacts on aquatic ecosystems. Increasingly, DNA-based techniques such as metabarcoding are being used for measuring aquatic biodiversity, as they could accelerate processing time, decrease costs and reduce some of the biases associated with traditional light microscopy. Despite the continuing use of traditional microscopy and the growing use of DNA metabarcoding to identify cyanobacteria assemblages, methodological comparisons between the two approaches have rarely been reported from a wide suite of lake types. Here, we compare planktonic cyanobacteria assemblages generated by inverted light microscopy and DNA metabarcoding from a 379-lake dataset spanning a longitudinal and trophic gradient. We found moderate levels of congruence between methods at the broadest taxonomic levels (i.e., Order, RV=0.40, p < 0.0001). This comparison revealed distinct cyanobacteria communities from lakes of different trophic states, with Microcystis, Aphanizomenon and Dolichospermum dominating with both methods in eutrophic and hypereutrophic sites. This finding supports the use of either method when monitoring eutrophication in lake surface waters. The biggest difference between the two methods was the detection of picocyanobacteria, which are typically underestimated by light microscopy. This reveals that the communities generated by each method currently are complementary as opposed to identical and promotes a combined-method strategy when monitoring a range of trophic systems. For example, microscopy can provide measures of cyanobacteria biomass, which are critical data in managing lakes. Going forward, we believe that molecular genetic methods will be increasingly adopted as reference databases are routinely updated with more representative sequences and will improve as cyanobacteria taxonomy is resolved with the increase in available genetic information. [ABSTRACT FROM AUTHOR]

Diverse communities of bacterial endophytes inhabit plant tissues, and these bacteria play important roles for plant growth and health. Cherry laurel (Prunus laurocerasus L.) is a broadleaf evergreen shrub that is widely grown in temperate zones for its ornamental and medicinal properties, however virtually nothing is known about its associated bacterial community. In this study, we analysed the matured one-year-old leaves of this plant using Illumina-based 16S rRNA gene metabarcoding to reveal the community structure of endophytic bacteria and understand its shifts during the seasonal transition from winter dormancy to a spring vegetative state. The overall community was composed of four dominant phyla (Proteobacteria, Actinobacteria, Firmicutes, Bacteroidetes). Corynebacterium, Acinetobacter, and Chryseobacterium genera were the most prevalent bacteria, comprising 13.3%, 6.9%, and 6.8% of the amplicon sequence variants (ASVs), respectively. The ASV richness and diversity increased significantly in May as compared to other sampling months (February, March, and April). We observed high variation in the overall community structure of endophytic bacteria among collection dates. The variation was only reflected by a few core community members, suggesting that the changes of the endophytic community during winter/spring seasonal transition are mostly associated with the less abundant community members. We identified biomarker taxa for late winter, mid spring, and late spring collection dates. This study is the first one to report on the diversity and composition of bacterial endophytes in the leaves of cherry laurel and its shifts across the dormancy-to-vegetative seasonal transition. [ABSTRACT FROM AUTHOR]

The geothermal system of the Salento peninsula (Italy) is characterized by the presence of many hydrogen sulfide-rich underground waters and thermal springs. We focused our attention on the submerged section of Zinzulùsa (Castro, Italy), a cave of both naturalistic and archaeological interest. In pioneer studies, some hypotheses about the origin of the sulfurous waters of this area were proposed. The most accredited one is that sulfate-enriched waters of marine origin infiltrate deep along bands with greater permeability, and warm-up going upwards, due to the geothermal gradient. During their route, marine waters interact with organic deposits and generate hydrogen sulfide as a result of sulfate reduction. To date, no studies have been conducted to elucidate the hydrogen sulfide origin in this site. The nature of reducing power and energy sources supporting microbial life in this submerged habitat is currently unknown. Here we present a multidisciplinary experimental approach aimed at defining geochemical features and microbiological diversity of the submerged habitat of Zinzulùsa cave. Our integrated data provide strong evidence that the sulfate content of the marine water and the activity of sulfate-reducing bacteria may account for the hydrogen sulfide content of the thermal springs. Anaerobic, sulfate-reducing, thermophilic Thermodesulfovibrio and hyperthermophilic Fervidobacterium genera show a high percentage contribution in 16S rRNA gene metabarcoding analyses, despite the mesophilic conditions of the sampling site. Besides, supported by PICRUSt functional analysis, we propose a chemotrophic model in which hydrocarbon deposits, entrapped in the stratifications of the seafloor, may be exploited by anaerobic oil-degrading bacteria as carbon and energy sources to sustain efficient hydrogen, sulfur, and nitrogen biogeochemical cycles. The Zinzulùsa hydrothermal site represents an ecosystem useful to obtain new insights into prokaryotic mutual interactions in oligotrophic and aphotic conditions, which constitute the largest environment of the biosphere. Unlabelled Image • Geochemical parameters of Zinzulùsa cave are variable over a long and short time. • The microbial community of Zinzulùsa cave shows an overall structural and functional stability over time. • Hydrogen sulfide production in submerged cave habitat is bacterially-mediated. • Hydrocarbon deposits mobilization supports efficient biogeochemical cycles in Zinzulùsa cave. [ABSTRACT FROM AUTHOR]

• Use of intermittent aeration enhanced azo dye mineralization. • Negative effects of aeration were minimized by increasing glucose concentration. • Aeration strategy and glucose concentration shifted microbial community. • A correlation between microbial diversity and decolorization was found. • Intermittently aerated assay presented reduced toxicity to D. magna. Most solutions for biological treatment of azo dyes are based on conventional anaerobic-aerobic processes, but transition to full scale demands technology simplification and cost reductions. We suggest a new approach, in which aeration is intermittently supplied for simultaneous removal of color and toxic metabolites in a single compartment. Effects of aeration strategy and glucose concentration on decolorization and organic matter removal were assessed using factorial design (32) and response surface analysis. Bioreactors were inoculated with microorganisms previously acclimated to Direct Black 22 (DB22), which was the azo compound used in this study. Assays performed with synthetic textile wastewater showed that long-term decolorization was not impaired at a moderate level of aeration (4 hourly-cycles per day). Aerated batches presented lower color removal velocities, but these negative impacts were offset by increasing initial glucose concentration. Higher degrees of mineralization of the azo compound and higher organic matter removals were achieved in intermittently aerated experiments, which led to lower toxicity to Daphnia magna. Biomolecular analysis revealed that the microbial community structure was strongly associated with operational efficiency parameters. These findings suggest intermittent aeration can be implemented to accomplish enhanced azo dye biodegradation. [ABSTRACT FROM AUTHOR]