Aquaculture ranks among the largest global food production industries, with Nile tilapia (Oreochromis niloticus) being one of the most widely farmed species. However, increasing consumer demand and higher stocking densities place considerable stress on aquaculture systems, often leading to a rise in fish diseases. Probiotics have emerged as valuable tools in this sector, promoting fish health by modulating physiological functions such as metabolism, digestion, immune responses, stress tolerance, and disease resistance. Here, the probiotic potential of Cetobacterium sp. nov C33, an anaerobic bacterium isolated from the intestine of Nile tilapia, was evaluated on short-term dietary supplementation in fingerlings in laboratory conditions. Using 16S rRNA amplicon sequencing, we assessed the impact of Cetobacterium sp. nov C33 on gut microbiota, while transcriptomic analysis of the head kidney provided insights into immune system modulation. Results indicate that dietary inclusion of this anaerobic bacterium significantly alters the gut microbiota structure in tilapia fingerlings and regulates genes associated with key metabolic pathways, including the immune system, underscoring its potential as a probiotic for enhancing tilapia health. Our results offer promising evidence of its potential as a probiotic to improve tilapia health.

This study uses high-throughput sequencing of the 16S rRNA gene and specific PCR to analyze the microbiome and identify secondary endosymbionts in sand flies from the Amazon region, important vectors of parasitic and viral diseases. Specimens of Psychodopygus, Trichophoromyia, Nyssomyia, Trichopygomyia and Brumptomyia were collected and analyzed. The results revealed that the richness, diversity, and composition of the microbiome are influenced by several factors, such as insect species specific composition, and insect sex. The core microbiome community was represented by 18 genera, with Novosphingobium, Cutibacterium, Methylobacterium and Staphylococcus being the most prevalent. The highest diversity at the genus level was observed in sand flies of epidemiological relevance as Psychodopygus and Nyssomyia, dominated by Novosphingobium (66.5%), Cutibacterium (29.4%) and Methylobacterium (20.4%), while in non-vectors such as Trichophoromyia, Delftia predominated (59.9%). Endosymbiont analysis showed a high prevalence of Cardinium (20%) and Wolbachia (33%), as well as the presence of Spiroplasma, Arsenophonus and Rickettsia. In addition, some bacterial genera related to the inhibition of parasite development, which have entomopathogenic activity and are involved in the degradation of insecticides were identified. Our results are relevant and contribute to the knowledge of the characterization of the microbiome and the endosymbionts in leishmaniasis vectors in the Amazon region and show promise for improving vector management, highlighting the importance of investigating their interaction with pathogens and their impact on vector biology.

IntroductionMosquitoes of the subfamily Culicinae transmit pathogens of major medical and veterinary importance, particularly in tropical regions where urbanization and ecological change promote arbovirus circulation. In Colombia, rural Culicinae species are diverse and harbor microbiomes that may influence vector competence, yet their bacterial communities remain poorly characterized.MethodsWe characterized the bacterial microbiota of multiple Culicinae species and morphotypes collected from two rural localities in Antioquia, Colombia, using an integrated metagenomic approach. Ribosomal 16S rRNA sequences were extracted from total RNA-seq datasets to infer bacterial community composition and assess α- and β-diversity. Diversity metrics (Chao1 and Shannon indices), Discriminant Analysis of Principal Components (DAPC), and Bray–Curtis ordination were used to evaluate community structure. In parallel, de novo assembled contigs were taxonomically annotated against the NCBI NR bacterial database to obtain complementary taxonomic and functional insights.ResultsCulex morphotypes exhibited the highest richness and evenness, whereas Aedes and Trichoprosopon showed lower diversity. Ordination and DAPC analyses revealed partial clustering by species and tribe. Both the 16S and assembly-based analyses showed complex bacterial assemblages dominated by Wolbachia (up to 60% of reads in several Aedes and Culex morphotypes), followed by environmental genera such as Pseudomonas and Acinetobacter (10–20%). Lower-abundance taxa of medical and veterinary importance—including Salmonella, Borrelia, and Clostridium (<5%)—were also detected. Bacterial community structure differed among mosquito species; Aedes albopictus was enriched in lactic acid bacteria, while Culex morphotypes exhibited broader environmental and endosymbiotic profiles.DiscussionThis study provides the first comprehensive metagenomic description of bacterial communities associated with rural Culicinae mosquitoes in Colombia. The predominance of symbionts such as Wolbachia and Spiroplasma, coupled with distinct bacterial signatures among host species, highlights the ecological complexity of these microbiomes and their potential relevance for microbiome-based strategies in sustainable arboviral disease management.

Hematophagous mosquitoes are major vectors of diverse pathogens and serve as bioindicators in tropical ecosystems, yet their virome in non-urban Neotropical regions remains poorly characterized. We analyzed the virome of 147 mosquitoes from two natural ecosystems in Colombia using a hybrid viral identification approach, combining high-confidence and less stringent methods. Most high-confidence viral contigs remained unclassified or unknown, as expected for metagenomic surveys in novel ecosystems. However, members for the Magrovirales and Ortervirales, and other six orders were detected at lower abundance. Using a complementary, less stringent approach, we identified 168 viral species from 68 genera and 22 families across four mosquito tribes (Aedini, Culicini, Orthopodomyiini, Sabethini), with dominance of Metaviridae, Retroviridae, Iridoviridae, and Poxviridae, though many sequences could not be taxonomically assigned. Insect-specific viruses predominated, while no medically relevant arboviruses were detected. Both methods consistently identified Trichoplusia ni TED virus, Cladosporium fulvum T-1 virus, Lymphocystis disease viruses, and Oryctes rhinoceros nudivirus among the most abundant and frequently detected taxa across samples. Alpha diversity indices revealed the highest virome diversity in Sabethini, followed by Orthopodmyiini, and substantially lower richness and diversity in Aedini and Culicini. These results provide a baseline for virome characterization in sylvatic mosquitoes from Colombia and highlight the need for further research on the ecological roles of the mosquito virome in pathogen transmission and microbiome evolution.

Malaria disease affects millions of people annually, making the Amazon Basin a major hotspot in the Americas. While traditional control strategies rely on physical and chemical methods, the Anopheles microbiome offers a promising avenue for biological control, as certain bacteria can inhibit parasite development and alter vector immune and reproductive systems, disrupting the transmission cycle. For this reason, this study aimed to explore the bacterial communities in An. darlingi and An. triannulatus s.l., including breeding sites, immature stages, and adults from San Pedro de los Lagos (Leticia, Amazonas) through next-generation sequencing of the 16S rRNA gene. The results revealed a higher bacterial genus richness in the L1–L2 larvae of An. triannulatus s.l. Aeromonas and Enterobacter were prevalent in most samples, with abundances of 52.51% in L3–L4 larvae and 48.88% in pupae of An. triannulatus s.l., respectively. In breeding site water, Verrucomicrobiota bacteria were the most dominant (52.39%). We also identified Delftia (15.46%) in An. triannulatus s.l. pupae and Asaia (98.22%) in An. triannulatus, linked to Plasmodium inhibition, and Elizabethkingia, in low abundances, along with Klebsiella and Serratia, known for paratransgenesis potential. Considering the high bacterial diversity observed across the different mosquito life stages, identifying bacterial composition is the first step towards developing new strategies for malaria control. However, the specific roles of these bacteria in anophelines and the malaria transmission cycle remain to be elucidated.

Fecal microbiota filtrate transfer is discussed as a safe alternative to fecal microbiota transfer (FMT) to treat ulcerative colitis. We investigated modulation of viral and bacterial composition during fecal microbiota filtrate transfer followed by FMT in six patients with active ulcerative colitis (where clinical activity improved in three patients after filtrate transfer) and combined 16S ribosomal RNA gene amplicon sequencing with a virome analysis pipeline including fast viral particle enrichment and metagenome mapping to detect frequencies of 45,033 reference bacteriophage genomes. We showed that after antibiotic treatment and during filtrate transfer, the bacterial community typically adopted a stable composition distinct to that before antibiotic treatment, with no change toward a donor community. FMT in contrast typically changed the bacterial community to a community with similarity to donor(s). There were no indications of an establishment of predominant donor viruses during filtrate transfer but a remodeling of the virome. In contrast, the establishment of donor viruses during FMT correlated with the predicted hosts established during such transfer. Our approach warrants further investigation in a randomized trial to evaluate larger therapeutic interventions in a comparable and efficient manner.

In this pilot study, we investigated the bacterial changes introduced on the subgingival, tongue, and saliva microbiota during fixed orthodontic treatment, with or without daily administration of nitrate-containing beet juice for 2 weeks in 22 individuals with good general health. We followed clinical parameters in combination with microbiota changes before, after 2 weeks, and after 6 months of treatment with fixed orthodontic appliances. In accordance with variations in community composition at the sampling sites, effects to orthodontic treatment differed. Subgingival communities responded promptly to orthodontic treatment with no additional structural changes over time, whereas saliva and tongue communities were affected only after extended treatment. Periodontal pathogens such as Selenomonas sputigena were enriched in subgingival communities, whereas Streptococcus mutans was enriched in saliva. Specifically, Rothia mucilaginosa increased tremendously in relative abundance in both tongue and saliva communities. The effect of beet juice on microbial composition was significant in subgingival samples even though the differences were not mirrored in single differentially distributed genera or species. This indicates changes in the complete subgingival microbial net of interacting species. However, the prevention of Corynebacterium matruchotii enrichment by beet juice may be important for prevention of biofilm formation. Enrichment of Neisseria flavescens group bacteria and Abiotrophia and depletion of different Actinomyces and Stomatobaculum were observed on tongue communities. We conclude that subgingival microbiota are rapidly affected by fixed orthodontic appliances and can be positively influenced by regular administration of nitrate-containing juice. • The subgingival site, tongue, and saliva contain different microbiota • The microbiota react differently to orthodontic treatment and beet juice • Key genera and species affected by treatments were identified

Aedes aegypti is a key vector in the transmission of arboviral diseases in the Colombian Amazon. This study aimed to characterize microbiota composition using DNA extracted from water in artificial breeding sites, immature stages, and adults of Ae. aegypti in Leticia, Amazonas. Additionally, the physicochemical water variables were correlated with the bacterial communities present. Eight artificial breeding sites were identified, with bucket, plant pot, and tire being the most frequent. The breeding sites exhibited similar physicochemical profiles, with significant temperature and salinity differences (p-value < 0.03). The most representative bacterial genera included Ottowia (82%), Xanthobacter (70.59%), and Rhodocyclaceae (92.78%) in breeding site water; Aquabacterium (61.07%), Dechloromonas (82.85%), and Flectobacillus (58.94%) in immature stages; and Elizabethkingia (70.89%) and Cedecea (39.19%) in males and females of Ae. aegypti. Beta diversity analysis revealed distinct clustering between adults and the water and immature communities (p-value < 0.001). Multivariate analysis showed strong correlations among bacterial communities, breeding sites, and physicochemical variables such as tire and drum cover which exhibited high levels of total dissolved solids, conductivity, and salinity associated with Flectobacillus, Leifsonia, Novosphingobium, Ottowia, and Rhodobacter. Bacterial genera such as Mycobacterium, Escherichia, Salmonella, and Clostridium, present in artificial breeding sites, are associated with public health relevance. This study provides insights into bacterial community dynamics across Ae. aegypti’s life cycle and underscores the importance of water physicochemical and biological characteristics for developing new vector control strategies.

Temperature is one of the most important factors influencing mosquito development, vector competence, and microbiome composition, affecting the disease transmission cycle by either facilitating or inhibiting pathogen establishment. This study aimed to estimate temperature preference and its correlation with bacterial communities in populations of Ae. aegypti and Ae. albopictus from the urban area of Leticia, Amazonas. Temperature preference profiles were obtained by introducing 50 individuals of each species and sex in six replicates for one hour into a device with a temperature gradient of 19-36 °C. Subsequently, a metataxonomic analysis of bacterial communities was performed using DNA extracted from whole mosquito body groups associated with temperature ranges defined by their statistical significance. First, it was determined that females of both species preferred temperatures between 23 and 28 °C. Meanwhile, Ae. aegypti males showed a larger temperature preference range (21-28 °C) than Ae. albopictus males (25-28 °C). Regarding bacterial communities, an increase in diversity was observed in males of both species with rising temperatures, whereas the opposite occurred in females. The Amplicon Sequence Variants (ASVs) associated with Asaia (2.88%), Cedecea (45.80%), Elizabethkingia (77.78%), and Klebsiella (8.90%) were the most abundant genera in Ae. aegypti. In comparison, Ae. albopictus exhibited a high abundance of Elizabethkingia (36.34%), Pseudomonas (95.08%), and Wolbachia (12.73%). Interestingly, Elizabethkingia varied in abundance according to sex and temperature in both species. It was determined that the structure of Aedes bacterial communities in the Amazon was influenced by sex, species, and temperature, indicating high plasticity and ubiquity according to temperature profiles.

Isonitrile natural products, also known as isocyanides, demonstrate potent antimicrobial activities, yet our understanding of their molecular targets remains limited. Here, we focus on the so far neglected group of monoisonitriles to gain further insights into their antimicrobial mode of action (MoA). Screening a focused monoisonitrile library revealed a potent S. aureus growth inhibitor with a different MoA compared to previously described isonitrile antibiotics. Chemical proteomics via competitive cysteine reactivity profiling, uncovered covalent modifications of two essential metabolic enzymes involved in the fatty acid biosynthetic process (FabF) and the hexosamine pathway (GlmS) at their active site cysteines. In-depth studies with the recombinant enzymes demonstrated concentration-dependent labeling, covalent binding to the catalytic site and corresponding functional inhibition by the isocyanide. Thermal proteome profiling and full proteome studies of compound-treated S. aureus further highlighted the destabilization and dysregulation of proteins related to the targeted pathways. Cytotoxicity and the inhibition of cytochrome P450 enzymes require optimization of the hit molecule prior to therapeutic application. The here described novel, covalent isocyanide MoA highlights the versatility of the functional group, making it a useful tool and out-of-the-box starting point for the development of innovative antibiotics.

Characterization of the temperature effects on the abundance and richness of the microbiota of Lutzomyia longipalpis, insect vector of Leishmania infantum in America, is an aspect of pivotal importance to understand the interactions between temperature, bacteria, and Leishmania infection. We developed and used a customized device with a temperature gradient (21-34 °C) to assess the temperature preferences of wild females of Lu. longipalpis collected in a rural area (Ricaurte, Cundinamarca, Colombia). Each replicate consisted of 50 females exposed to the gradient for an hour. At the end of the exposure time, insects were collected and separated by the temperature ranges selected varying from 21 °C to 34 °C. They were organized in 17 pools from which total DNA extracts were obtained, and samples were subjected to 16S rRNA amplicon sequencing analyzes. The most abundant phyla across the different temperature ranges were Proteobacteria (17.22-90.73 %), Firmicutes (5.99-77.21 %) and Actinobacteria (1.56-59.85 %). Results also showed an abundance (30 % to 57.36 %) of Pseudomonas (mainly at temperatures of 21-29 °C and 34 °C) that decreased to 6.55 %-13.20 % at temperatures of 31-33 °C, while Bacillus increase its abundance to 67.24 % at 29-33 °C. Serratia also had a greater representation (49.79 %), specifically in sand flies recovered at 25-27 °C. No significant differences were found at α-diversity level when comparing richness using the Shannon-Wiener, Simpson, and Chao1 indices, while β-diversity differences were found using the Bray-Curtis index (F-value of 3.5073, p-value < 0.013, R-squared of 0,4889), especially in the groups of Lu. longipalpis associated at higher temperatures (29-33 °C). It was also possible to detect the presence of endosymbionts such as Spiroplasma and Arsenophonus in the range of 29-33 °C. Rickettsia was only detected in Lu. longipalpis sand flies recovered between 25-27 °C. It was possible to characterize Lu. longipalpis microbiota in response to intraspecific temperature preferences and observe changes in bacterial communities and endosymbionts at different ranges of said environmental variable, which may be important in its vector competence and environmental plasticity to adapt to new climate change scenarios.

Screening large molecule libraries against pathogenic bacteria is often challenged by a low hit rate due to limited uptake, underrepresentation of antibiotic structural motifs, and assays that do not resemble the infection conditions. To address these limitations, we present a screen of a focused library of alkyl guanidinium compounds, a structural motif associated with antibiotic activity and enhanced uptake, under host-mimicking infection conditions against a panel of disease-associated bacteria. Several hit molecules were identified with activities against Gram-positive and Gram-negative bacteria, highlighting the fidelity of the general concept. We selected one compound (L15) for in-depth mode of action studies that exhibited bactericidal activity against methicillin-resistant Staphylococcus aureus USA300 with a minimum inhibitory concentration of 1.5 mu M. Structure-activity relationship studies confirmed the necessity of the guanidinium motif for antibiotic activity. The mode of action was investigated using affinity-based protein profiling with an L15 probe and identified the signal peptidase IB (SpsB) as the most promising hit. Validation by activity assays, binding site identification, docking, and molecular dynamics simulations demonstrated SpsB activation by L15, a recently described mechanism leading to the dysregulation of protein secretion and cell death. Overall, this study highlights the need for unconventional screening strategies to identify novel antibiotics.

Primary sclerosing cholangitis (PSC) is an inflammatory disease of the biliary tract eventually leading to bile duct destruction, liver failure, cholangiocellular adenocarcinoma and/or death. No disease modifying treatments are available. Especially cytotoxicity of bile acids, are discussed as potential driver of disease progression. Cholangiocytes are protected by a bicarbonate umbrella formed by the glycocalyx, a dense layer of membrane bound polyglycans extending into the extracellular space. Bile of PSC patients harbors a unique microbiome. Here we identified a new factor in the pathogenesis of PSC. The bacterial degradation of sialic acid and galactose are associated with a poor event free survival of PSC patients and could identify bacterial liberation of sialic acid as crucial element in cholangiocyte damage using cell culture experiments, individualized organoid models and liver biopsies. With this study the view on bacteria-host interactions in bile duct associated diseases is widened. Functional patterns of the bacterial community are crucial for bile duct destruction in PSC patients. This opens a new field of diagnostic tools, disease modifying treatment options and identification of patients at risk. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement The study was funded by the Deutsche Forschungsgemeinschaft (DFG German Research Foundation) under Germany Excellence Strategy EXC 2155 RESIST Project ID 390874280 and by the German Centre for Infection Research (DZIF e.V.). This work was supported by a grant from the German Federal Ministry of Education and Research (reference number: 01EO1302) and by the Helmholtz Association Initiative on Aging and Metabolic Programming. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The study had been approved by the ethics committee of the Hannover Medical School (approval no. 220- 2007, approval no. 3241- 2016, approval no. 9660\_BO\_K\_2021 and approval no. 10183\_BO\_K\_2022). I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Raw data from 16S amplicon sequencing, metagenomic sequence data, bacterial genome data and data from mass spectrometry will be available upon publication after peer-review. All additional data in the present study are available upon reasonable request to the authors.

BACKGROUND:Still, little is known about microbial dysbiosis in oropharyngeal and laryngeal tissue as risk factor for development of local squamous cell carcinoma. The site-specific microbiota at these regions in healthy and cancer tissue and their modulation by environmental factors need to be defined. METHODS:The local microbiota of cancer tissue and healthy controls was profiled by 16S rRNA gene amplicon sequencing and statistical analysis using 111 oropharyngeal and 72 laryngeal intraoperative swabs. RESULTS:Oropharynx and larynx harbor distinct microbial communities. Clear effects of both smoking and cancer were seen in the oropharynx whereas effects in the larynx were minor. CONCLUSION:The distinct microbial communities at larynx and oropharynx partially explain why the effects of cancer and smoking were distinct at those sites. Thus, the use of microbiota supposed to mirror community changes in another target location should be avoided and more studies on the actual cancerous environment are necessary.

Background:Spodoptera frugiperda (or fall armyworm, FAW) is a polyphagous pest native to Western Hemisphere and recently discovered in the Eastern Hemisphere. In Colombia, S. frugiperda is recognized as a pest of economic importance in corn. The species has genetically differentiated into two host populations named "corn" and "rice" strains. In 2012, a study made in central Colombia demonstrated that the corn strain is less susceptible to Bacillus thuringiensis (Bt) endotoxins (Cry1Ac and Cry 1Ab) than the rice strain. In this country, Bt transgenic corn has been extensively produced over the last 15 years. Since gut microbiota plays a role in the physiology and immunity of insects, and has been implicated in promoting the insecticidal activity of Bt, in this study an analysis of the interaction between Bt endotoxins and FAW gut microbiota was made. Also, the detection of endosymbionts was performed here, as they might have important implications in the biological control of a pest.Methods:The composition and diversity of microbiomes associated with larval specimens of S. frugiperda(corn strain) was investigated in a bioassay based on six treatments in the presence/absence of Bt toxins and antibiotics (Ab) through bacterial isolate analyses and by high throughput sequencing of the bacterial 16S rRNA gene. Additionally, species specific primers were used, to detect endosymbionts from gonads in S. frugiperda corn strain.Results:Firmicutes, Proteobacteria and Bacteroidota were the most dominant bacterial phyla found in S. frugiperda corn strain. No significant differences in bacteria species diversity and richness among the six treatments were found. Two species of Enterococcus spp., E. mundtii and E. casseliflavus were detected in treatments with Bt and antibiotics, suggesting that they are less susceptible to both of them. Additionally, the endosymbiont Arsenophonus was also identified on treatments in presence of Bt and antibiotics. The results obtained here are important since little knowledge exists about the gut microbiota on this pest and its interaction with Bt endotoxins. Previous studies made in Lepidoptera suggest that alteration of gut microbiota can be used to improve the management of pest populations, demonstrating the relevance of the results obtained in this work.

The probiotic potential of a designed bacterial consortia isolated from a competitive exclusion culture originally obtained from the intestinal contents of tilapia juveniles were evaluated on Nile tilapia alevins. The growth performance, intestinal histology, microbiota effects, resistance to Streptococcus agalactiae challenge, and immune response were assessed. In addition, the following treatments were included in a commercial feed: A12+M4+M10 (Lactococcus lactis A12, Priestia megaterium M4, and Priestia sp. M10), M4+M10 (P. megaterium M4, and Priestia sp. M10) and the single bacteria as controls; A12 (L. lactis A12), M4 (P. megaterium M4), M10 (Priestia sp. M10), also a commercial feed without any probiotic addition was included as a control. The results showed that all probiotic treatments improved the growth performance, intestinal histology, and resistance during experimental infection with S. agalactiae in comparison to the control fish. Also, the administration of probiotics resulted in the modulation of genes associated with the innate and adaptive immune systems that were non-dependent on microbial colonization. Surprisingly, L. lactis A12 alone induced benefits in fish compared to the microbial consortia, showing the highest increase in growth rate, survival during experimental infection with S. agalactiae, increased intestinal fold length, and the number of differentially expressed genes. Lastly, we conclude that a competitive exclusion culture is a reliable source of probiotics, and monostrain L. lactis A12 has comparable or even greater probiotic potential than the bacterial consortia.

Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with a dismal outcome. To improve understanding of sequential microbiome changes during PDAC development we analyzed mouse models of pancreatic carcinogenesis (KC mice recapitulating pre-invasive PanIN formation, as well as KPC mice recapitulating invasive PDAC) during early tumor development and subsequent tumor progression. Diversity and community composition were analyzed depending on genotype, age, and gender. Both mouse models demonstrated concordant abundance changes of several genera influenced by one or more of the investigated factors. Abundance was significantly impacted by gender, highlighting the need to further elucidate the impact of gender differences. The findings underline the importance of the microbiome in PDAC development and indicate that microbiological screening of patients at risk and targeting the microbiome in PDAC development may be feasible in future.

Microbiome components and bacterial isolates related to healthy and epitheliocystis states in aquaculture cycles of cobia fish were studied. We detected well-defined 16S rRNA amplicon gene sequence variants showing differential abundance in healthy or diseased cycles. Isolation trials were performed, and experimental tests were used to determine probiotic potential of the bacterial strains obtained from water, tissues or live food used in this aquaculture model. The taxonomic affiliation of these strains was cross-compared against microbiome components, finding that some of them had close or identical affiliation to the abundant types found in healthy cycles. Strains belonging to the groups already identified as predominant by culture-independent means were screened as potential probiotics based on desirable activities such as antagonism and antibiosis against marine pathogenic bacteria, quorum quenching, bile acid resistance, antibiotic sensitivity and enzymatic activities for improved nutrient digestion. We have also found that in the tracking of microbiome composition across different developmental stages of cobia, healthy cycles exhibited a consistent high relative abundance of a Mesobacillus sp., while in the diseased cycle the emergence of a Vibrio sp. was observed. Our study suggests that epithelocystis in cobia is associated with a displacement of a symbiotic microbiome community linked to the increase frequency of Vibrio species.

Extradiol dioxygenation is a key reaction in the microbial aerobic degradation of mono- and polycyclic aromatic hydrocarbon catecholic derivatives. It has been reported that many bacterial enzymes exhibiting such converging functions act on a wide range of catecholic substrates. The present study reports a new subfamily of extradiol dioxygenases (EXDOs) with broad substrate specificity, the HrbC EXDOs. The new clade belongs to the XII cluster within family 2 of the vicinal oxygen chelate superfamily (EXDO-VC2), which is typically characterized by a preference for bicyclic substrates. Coding hrbC orthologs were isolated by activity-based screening of fosmid metagenomic libraries from large DNA fragments derived from heavily PAH-contaminated soils. They occurred as solitary genes within conserved sequences encoding enzymes for amino acid metabolism and were stably maintained in the chromosomes of the Betaproteobacteria lineages harboring them. Analysis of contaminated aquifers revealed coexpression of hrbC as a polycistronic mRNA component. The predicted open reading frames were verified by cloning and heterologous expression, confirming the expected molecular mass and meta-cleavage activity of the recombinant enzymes. Evolutionary analysis of the HrbC protein sequences grouped them into a discrete cluster of 1,2-dihydroxynaphthalene dioxygenases represented by a cultured PAH degrader, Rugosibacter aromaticivorans strain Ca6. The ecological importance and relevance of the new EXDO genes were confirmed by PCR-based mapping in different biogeographical localities contaminated with a variety of mono- and polycyclic aromatic compounds. The cosmopolitan distribution of hrbC in PAH-contaminated aquifers supports our hypothesis about its auxiliary role in the degradation of toxic catecholic intermediates, contributing to the composite EXDO catabolic capacity of the world's microbiomes.

Microbiota of insect vectors is involved in processes of immune response, fitness,resistance to insecticides and environmental changes particularly linked to climatechange signals such as temperature shifts, influencing in turn the transmission rate ofhuman pathogens, as is the case of phlebotomine sand flies. Characterization of thetemperature effects in the abundance and richness of the microbiota of Lutzomyia longipalpis, insect vector of Leishmania infantum in America, is an aspect of pivotalimportance in order to understand how such changes can affect the potentialtransmission of Leishmania. We develop and used a customized device with atemperature gradient (21-34°C) to maintain two groups of wild females of Lu.longipalpis collected in a rural area (Ricaurte, Cundinamarca). Each replica consistedof 50 females who were exposed to such gradient for an hour. At the end of theexposure time, insects were collected and separated by temperature ranges 21-23 °C,23-25 °C, 25-27 °C, 27-29 °C, 29-31 °C, 31-33°C, and 34 °C. They were organized in17 pools, total DNA extracts were obtained and samples were subjected to 16Samplicon sequencing analyses. The most abundant phyla were Proteobacteria(90.26%), Firmicutes (73.52%) and Actinobacteriota (8.09%). Results also show anabundance of 57.36% in Pseudomonas at 25-27 °C that decreases to 6.55% atwarmer temperatures 29-31°C and 13.20% at 31-33°C (p-value < 0.055), while at thesame ranges Bacillus presents an abundance of 1.21%, 61.54% and 37.64%respectively (p-value < 0.096) with a Pearson Correlation Coefficient of -0.84 (p-value< 0.00007, and FDR of 0.015). No significant differences were found when comparingrichness using the Shannon-Weiner, Simpson and Chao1 indices, while -diversitydifferences were found using the Bray-Curtis index (p-value < 0.013) especially atwarm temperatures (29-33 °C). It was also possible to detect the presence ofendosymbionts such as Spiroplasma and Asaia, but with low prevalence, whileinfection of Arsenophonus was also present with positive interactions with Bacillus(PCC 0.77, p-value of 0.006, and FDR of 0.015). It was possible to perform thecharacterization of Lu. longipalpis microbiota in relation to the temperature preferencesof the insect vector and observe changes in bacterial communities at different rangesof said environmental variable