Endosymbiont and gut bacterial communities of the brown-banded cockroach, Supella longipalpa.

The brown-banded cockroach (Supella longipalpa) is a widespread nuisance and public health pest. Like the German cockroach (Blattella germanica), this species is adapted to the indoor biome and completes the entirety of its life cycle in human-built structures. Recently, understanding the contributions of commensal and symbiotic microbes to the biology of cockroach pests, as well as the applications of targeting these microbes for pest control, have garnered significant scientific interest. However, relative to B. germanica, the biology of S. longipalpa, including its microbial associations, is understudied. Therefore, the goal of the present study was to quantitatively examine and characterize both the endosymbiont and gut bacterial communities of S. longipalpa for the first time. To do so, bacterial 16S rRNA gene amplicon sequencing was conducted on DNA extracts from whole adult females and males, early instar nymphs, and late instar nymphs. The results demonstrate that the gut microbiome is dominated by two genera of bacteria known to have beneficial probiotic effects in other organisms, namely Lactobacillus and Akkermansia. Furthermore, our data show a significant effect of nymphal development on diversity and variation in the gut microbiome. Lastly, we reveal significant negative correlations between the two intracellular endosymbionts, Blattabacterium and Wolbachia, as well as between Blattabacterium and the gut microbiome, suggesting that Blattabacterium endosymbionts could directly or indirectly influence the composition of other bacterial populations. These findings have implications for understanding the adaptation of S. longipalpa to the indoor biome, its divergence from other indoor cockroach pest species such as B. germanica, the development of novel control approaches that target the microbiome, and fundamental insect-microbe interactions more broadly.

Characterization of an outbreak caused by Elizabethkingia miricola using Fourier-transform infrared (FTIR) spectroscopy.

Fourier-transform infrared (FTIR) spectroscopy has the potential to be used for bacterial typing and outbreak characterization. We evaluated FTIR for the characterization of an outbreak caused by Elizabethkingia miricola. During the 2020-2021 period, 26 isolates (23 clinical and 3 environmental) were collected and analyzed by FTIR (IR Biotyper) and core-genome MLST (cgMLST), in addition to antimicrobial susceptibility testing. FTIR spectroscopy and cgMLST showed that 22 of the isolates were related to the outbreak, including the environmental samples, with only one discordance between both methods. Then, FTIR is useful for E. miricola typing and can be easily implemented in the laboratory.

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

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

Aedes albopictus infection by the native bacterium Elizabethkingia anophelis in Southern Mexico.

OBJECTIVE: To evaluate the presence of Elizabethkingia anophelis infection in Aedes albopictus wild populations of Southern Mexico. MATERIALS AND METHODS: Eight sites were selected to collect Aedes albopictus in the Soconusco region, Chiapas, females were analyzed to amplify the Gyrase B gene by PCR, the minimum infection rate of E. anopheliswas calculated and its species was determined by sequencing and phylogeny. RESULTS: The presence of E. anophelis was only observed in Huehuetán with a minimum infection rate of 37.8%. CONCLUSION: A local strain of E. anophelis was detected for the first time in Ae. albopictus from Chiapas and this bacterium could be considered a candidate for study as a probable control agent or as a vehicle for transgenesis.

Flavobacterium sedimenticola sp. nov., isolated from sediment.

A novel yellow-pigmented bacterial strain, designated YZ-48T, was isolated from the sediment of the Yangtze River, PR China. Cells were Gram-stain-negative, non-motile, rod-shaped, strictly aerobic, catalase-positive and oxidase-positive. The strain grew optimally on R2A medium at 37 °C, pH 7.0 and with 1.0 % (w/v) NaCl. Strain YZ-48T showed the closest 16S rRNA gene sequence similarity to Flavobacterium solisilvae SE-s27T (96.4 %) and F. dankookense DSM 25687T (96.2 %). The phylogenetic trees based on 16S rRNA gene sequences showed that strain YZ-48T belonged to the genus Flavobacterium but formed a distinct phylogenetic lineage. The obtained average nucleotide identity and digital DNA-DNA hybridization values between YZ-48T and the two closest strains were 75.0 and 74.5 % and 19.6 and 19.0 %, respectively. The sole respiratory quinone was MK-6. The major polar lipids were phosphatidylethanolamine, two unidentified aminolipids and three unidentified polar lipids. The major cellular fatty acids were iso-C16 : 0, iso-C15 : 0, iso-C15 : 1 G, iso-C17 : 0 3-OH, iso-C15 : 0 3-OH and iso-C16 : 0 3-OH. The DNA G+C content was 40.2 mol%. Based on the phenotypic, chemotaxonomic, phylogenetic and genomic data, strain YZ-48T represents a novel species of the genus Flavobacterium, for which the name Flavobacterium sedimenticola sp. nov. is proposed, with strain YZ-48T (=KCTC 82329T=CCTC AB 2023061T=MCCC 1K08804T) as the type strain.

Robiginitalea aurantiaca sp. nov. and Algoriphagus sediminis sp. nov., isolated from coastal sediment.

Two novel rod-shaped, Gram-stain-negative, aerobic and non-motile bacterial strains, designated M39T and C2-7T, were isolated from the coastal sediment of Xiaoshi Island, Weihai, PR China. Growth of strain M39T occurred at 15-37 °C, at pH 6.0-9.0 and in the presence of 1.0-9.0 % (w/v) NaCl. Strain C2-7T grew at 15-40 °C, at pH 6.0-8.0 and in the presence of 0.5-8.0 % (w/v) NaCl. Phylogenetic analysis based 16S rRNA gene sequences revealed that strains M39T and C2-7T belong to the phylum Bacteroidota. Based on the results of 16S rRNA gene sequence analysis, the closest relative of strain M39T was Robiginitalea marina KCTC 92035T (95.4 %), and the closest relative of strain C2-7T was Algoriphagus namhaensis DPG-3T (97.0 %). The percentage of conserved protein and average nucleotide identity values between strain M39T and some species of the genus Robiginitalea were 66.9-77.6% and 69.3-71.0 %, respectively, while those between strain C2-7T and some species of the genus Algoriphagus were 68.0-70.1% and 56.1-72.6 %, respectively. The major cellular fatty acids (>10 %) of strain M39T consisted of iso-C15 : 1 F, iso-C15 : 0 and iso-C17 : 0 3-OH, while those of strain C2-7T were iso-C15 : 0 and C16 : 1 ω7c/C16 : 1 ω6c. MK-6 was the only respiratory quinone that was compatible with the genus of strain M39T. The predominant menaquinone of strain C2-7T was MK-7. The major polar lipids of strain M39T were phosphatidylethanolamine and glycolipids, and those of strain C2-7T were phosphatidylethanolamine, one unidentified aminolipid and four unidentified lipids. The DNA G+C contents of strains M39T and C2-7T were 46.9 and 40.8 mol%, respectively. Based upon the results presented in this study, strains M39T and C2-7T represent novel species of the genera Robiginitalea and Algoriphagus, respectively, for which the names Robiginitalea aurantiaca sp. nov. and Algoriphagus sediminis sp. nov. are proposed with the type strains M39T (=MCCC 1H00498T=KCTC 92014T) and C2-7T (=MCCC 1H00414T=KCTC 92027T).

Complete genome sequence of carotenoid-producing Aestuariibaculum lutulentum L182T isolated from the tidal sediment.

Aestuariibaculum lutulentum L182T (= KCTC 92530T = MCCC 1K08065T) was isolated from the tidal sediment collected in Beihai, People's Republic of China. The genome was sequenced and consisted of a single chromosome with the size of 3,782,725 bp and DNA G + C content of 35.1%. Genomic annotations demonstrated that it encoded 12 rRNA genes, 56 tRNA genes and 3210 ORFs. The percentages of ORFs assigned to CAZy, COG, and KEGG databases were 5.5, 86.2 and 45.5%, respectively. Comparative genomic analysis indicated that the pan- and core-genomes of the genus Aestuariibaculum consisted of 4826 and 2257 orthologous genes, respectively. Carbohydrate-active enzyme annotations of the genus Aestuariibaculum genomes revealed that they shared three polysaccharide lyase (PL) families including PL1, PL22 and PL42. Meanwhile, one carotenoid biosynthetic gene cluster related to biosynthesizing flexixanthin was found in the genus Aestuariibaculum. Furthermore, the core-genome of the genus Aestuariibaculum showed that this genus played a role in cleaving pectate, degrading ulvan, and biosynthesizing carotenoids. This study is a complete genomic report of the genus Aestuariibaculum and broadens understandings of its ecological roles and biotechnological applications.

Description of Flavobacterium agricola sp. nov., an auxin producing bacterium isolated from paddy field.

An auxin-producing bacterial strain, CC-SYL302T, was isolated from paddy soil in Taiwan and identified using a polyphasic taxonomic approach. The cells were observed to be aerobic, non-motile, non-spore-forming rods, and tested positive for catalase and oxidase. Produced carotenoid but flexirubin-type pigments were absent. Optimal growth of strain CC-SYL302T was observed at 25 °C, pH 7.0, and with 2% (w/v) NaCl present. Based on analysis of 16S rRNA gene sequences, it was determined that strain CC-SYL302T belongs to the genus Flavobacterium of the Flavobacteriaceae family. The closest known relatives of this strain are F. tangerinum YIM 102701-2 T (with 93.3% similarity) and F. cucumis R2A45-3 T (with 93.1% similarity). Digital DNA-DNA hybridization (dDDH) values were calculated to assess the genetic distance between strain CC-SYL302T and its closest relatives, with mean values of 21.3% for F. tangerinum and 20.4% for F. cucumis. Strain CC-SYL302T exhibited the highest orthologous average nucleotide identity (OrthoANI) values with members of the Flavobacterium genus, ranging from 67.2 to 72.1% (n = 22). The dominating cellular fatty acids (> 5%) included iso-C14:0, iso-C15:0, iso-C16:0, iso-C15:0 3-OH, iso-C17:0 3-OH, C16:1 ω6c/C16:1 ω7c and C16:0 10-methyl/iso-C17:1 ω9c. The polar lipid profile consisted of phosphatidylethanolamine, an unidentified aminolipid, an unidentified aminophospholipid, and nine unidentified polar lipids. The genome (2.7 Mb) contained 33.6% GC content, and the major polyamines were putrescine and sym-homospermidine. Strain CC-SYL302T exhibits distinct phylogenetic, phenotypic, and chemotaxonomic characteristics, as well as unique results in comparative analysis of 16S rRNA gene sequence, OrthoANI, dDDH, and phylogenomic placement. Therefore, it is proposed that this strain represents a new species of the Flavobacterium genus, for which the name Flavobacterium agricola sp. nov. is proposed. The type strain is CC-SYL302T (= BCRC 81320 T = JCM 34764 T).

Complete genome sequence of pectin-degrading Flavobacteriaceae bacterium GSB9.

Pectic oligosaccharides, which are considered to be potential prebiotics, may be generated by pectin-degrading enzymes. Here, we report the complete genome sequence of the pectin-degrading marine bacterium, Flavobacteriaceae bacterium GSB9, which was isolated from seawater of South Korea. The complete genome sequence revealed that the chromosome was 3,630,376 bp in size, had a G + C content of 36.6 mol%, and was predicted to encode 3100 protein-coding sequences (CDSs), 40 tRNAs, and six 16S-23S-5S rRNAs. Genome sequence analysis revealed that this strain possesses multiple genes predicted to encode pectin-degrading enzymes. Our analysis may facilitate the future application of this strain against pectin in various industries.

Reclassification of Sabulilitoribacter multivorans and Sabulilitoribacter arenilitoris as Flaviramulus multivorans comb. nov. and Wocania arenilitoris comb. nov., respectively, based on genome sequence analysis.

Rapid advancements in DNA sequencing technologies are providing new approaches for bacterial taxonomy. The genus Sabulilitoribacter is a member of the family Flavobacteriaceae, which consists of more than 150 genera. In this study, genome sequence analysis was conducted to revisit the taxonomic status of Sabulilitoribacter arenilitoris and Sabulilitoribacter multivorans, the only two species of this genus. Genome sequence based phylogeny analysis showed that the genus Sabulilitoribacter was non-monophyletic: S. multivorans, the type species of genus Sabulilitoribacter, was clustered with the type species of the genus Flaviramulus, whereas S. arenilitoris formed a robust cluster with the only two species of the genus Wocania. The values of average amino acid identity, genome-wide average nucleotide identity, alignment fractions and some phenotypic features showed that S. multivorans was more closely related with the type species of the genus Flaviramulus than with S. arenilitoris, and S. arenilitoris was more closely related with the only two species of the genus Wocania than with S. multivorans. Based on these results, we consequently propose that S. multivorans and S. arenilitoris should be reclassified as Flaviramulus multivorans comb. nov. and Wocania arenilitoris comb. nov. respectively.

Deciphering Interactions Within a 4-Strain Riverine Bacterial Community.

The dynamics of a community of four planktonic bacterial strains isolated from river water was followed in R2 broth for 72 h in batch experiments. These strains were identified as Janthinobacterium sp., Brevundimonas sp., Flavobacterium sp. and Variovorax sp. 16S rRNA gene sequencing and flow cytometry analyses were combined to monitor the change in abundance of each individual strain in bi-cultures and quadri-culture. Two interaction networks were constructed that summarize the impact of the strains on each other's growth rate in exponential phase and carrying capacity in stationary phase. The networks agree on the absence of positive interactions but also show differences, implying that ecological interactions can be specific to particular growth phases. Janthinobacterium sp. was the fastest growing strain and dominated the co-cultures. However, its growth rate was negatively affected by the presence of other strains 10 to 100 times less abundant than Janthinobacterium sp. In general, we saw a positive correlation between growth rate and carrying capacity in this system. In addition, growth rate in monoculture was predictive of carrying capacity in co-culture. Taken together, our results highlight the necessity to take growth phases into account when measuring interactions within a microbial community. In addition, evidence that a minor strain can greatly influence the dynamics of a dominant one underlines the necessity to choose population models that do not assume a linear dependency of interaction strength to abundance of other species for accurate parameterization from such empirical data.

No Transcriptional Compensation for Extreme Gene Dosage Imbalance in Fragmented Bacterial Endosymbionts of Cicadas.

Bacteria that form long-term intracellular associations with host cells lose many genes, a process that often results in tiny, gene-dense, and stable genomes. Paradoxically, the some of the same evolutionary processes that drive genome reduction and simplification may also cause genome expansion and complexification. A bacterial endosymbiont of cicadas, Hodgkinia cicadicola, exemplifies this paradox. In many cicada species, a single Hodgkinia lineage with a tiny, gene-dense genome has split into several interdependent cell and genome lineages. Each new Hodgkinia lineage encodes a unique subset of the ancestral unsplit genome in a complementary way, such that the collective gene contents of all lineages match the total found in the ancestral single genome. This splitting creates genetically distinct Hodgkinia cells that must function together to carry out basic cellular processes. It also creates a gene dosage problem where some genes are encoded by only a small fraction of cells while others are much more abundant. Here, by sequencing DNA and RNA of Hodgkinia from different cicada species with different amounts of splitting-along with its structurally stable, unsplit partner endosymbiont Sulcia muelleri-we show that Hodgkinia does not transcriptionally compensate to rescue the wildly unbalanced gene and genome ratios that result from lineage splitting. We also find that Hodgkinia has a reduced capacity for basic transcriptional control independent of the splitting process. Our findings reveal another layer of degeneration further pushing the limits of canonical molecular and cell biology in Hodgkinia and may partially explain its propensity to go extinct through symbiont replacement.

Isolation of Elizabethkingia spp. from Diagnostic Specimens from Dogs and Cats, United States, 2019-2021.

We retrospectively reviewed Elizabethkingia spp. culture and susceptibility results from 86 veterinary diagnostic laboratory results from US dogs and cats. We noted 26 E. menigoseptica, 1 E. miricola, and 59 unspeciated Elizabethkingia isolates from 9 US states (2-22 isolates per state). Elizabethkingia infections in animals might increase risks to humans.

Atypical flavobacteria recovered from diseased fish in the Western United States.

Flavobacterial diseases, caused by bacteria in the order Flavobacteriales, are responsible for devastating losses in farmed and wild fish populations worldwide. The genera Flavobacterium (Family Flavobacteriaceae) and Chryseobacterium (Weeksellaceae) encompass the most well-known agents of fish disease in the order, but the full extent of piscine-pathogenic species within these diverse groups is unresolved, and likely underappreciated. To identify emerging agents of flavobacterial disease in US aquaculture, 183 presumptive Flavobacterium and Chryseobacterium isolates were collected from clinically affected fish representing 19 host types, from across six western states. Isolates were characterized by 16S rRNA gene sequencing and phylogenetic analysis using the gyrB gene. Antimicrobial susceptibility profiles were compared between representatives from each major phylogenetic clade. Of the isolates, 52 were identified as Chryseobacterium species and 131 as Flavobacterium. The majority of Chryseobacterium isolates fell into six clades (A-F) consisting of ≥ 5 fish isolates with ≥ 70% bootstrap support, and Flavobacterium into nine (A-I). Phylogenetic clades showed distinct patterns in antimicrobial susceptibility. Two Chryseobacterium clades (F & G), and four Flavobacterium clades (B, G-I) had comparably high minimal inhibitory concentrations (MICs) for 11/18 antimicrobials tested. Multiple clades in both genera exhibited MICs surpassing the established F. psychrophilum breakpoints for oxytetracycline and florfenicol, indicating potential resistance to two of the three antimicrobials approved for use in finfish aquaculture. Further work to investigate the virulence and antigenic diversity of these genetic groups will improve our understanding of flavobacterial disease, with applications for treatment and vaccination strategies.

Insights into Regulating Mechanism of Mutagenesis Strains of Elizabethkingia meningoseptica sp. F2 by Omics Analysis.

Vitamin K2 plays an important role in electron transport, blood coagulation, and calcium homeostasis, and researchers have been trying to use microbes to produce it. Although our previous studies have shown that gradient radiation, breeding, and culture acclimation can improve vitamin K2 production in Elizabethkingia meningoseptica, the mechanism is still unclear. This study is the first which performs genome sequencing of E. meningoseptica sp. F2 as a basis for subsequent experiments and further comparative analyses with other strains. Comparative metabolic pathway analysis of E. meningoseptica sp. F2, E. coli, Bacillus subtilis, and other vitamin K2 product strains revealed that the mevalonate pathway of E. meningoseptica sp. F2 is different in bacteria at the system level. The expressions of menA, menD, menH, and menI in the menaquinone pathway and idi, hmgR, and ggpps in the mevalonate pathway were higher than those in the original strain. A total of 67 differentially expressed proteins involved in the oxidative phosphorylation metabolic pathway and citric acid cycle (TCA cycle) were identified. Our results reveal that combined gradient radiation breeding and culture acclimation can promote vitamin K2 accumulation probably by regulating the vitamin K2 pathway, oxidative phosphorylation metabolism pathway, and the citrate cycle (TCA cycle).

Marine Bacteroidetes enzymatically digest xylans from terrestrial plants.

Marine Bacteroidetes that degrade polysaccharides contribute to carbon cycling in the ocean. Organic matter, including glycans from terrestrial plants, might enter the oceans through rivers. Whether marine bacteria degrade structurally related glycans from diverse sources including terrestrial plants and marine algae was previously unknown. We show that the marine bacterium Flavimarina sp. Hel_I_48 encodes two polysaccharide utilization loci (PULs) which degrade xylans from terrestrial plants and marine algae. Biochemical experiments revealed activity and specificity of the encoded xylanases and associated enzymes of these PULs. Proteomics indicated that these genomic regions respond to glucuronoxylans and arabinoxylans. Substrate specificities of key enzymes suggest dedicated metabolic pathways for xylan utilization. Some of the xylanases were active on different xylans with the conserved ß-1,4-linked xylose main chain. Enzyme activity was consistent with growth curves showing Flavimarina sp. Hel_I_48 uses structurally different xylans. The observed abundance of related xylan-degrading enzyme repertoires in genomes of other marine Bacteroidetes indicates similar activities are common in the ocean. The here presented data show that certain marine bacteria are genetically and biochemically variable enough to access parts of structurally diverse xylans from terrestrial plants as well as from marine algal sources.

Agarolytic Pathway in the Newly Isolated Aquimarina sp. Bacterial Strain ERC-38 and Characterization of a Putative ß-agarase.

Marine microbes, particularly Bacteroidetes, are a rich source of enzymes that can degrade diverse marine polysaccharides. Aquimarina sp. ERC-38, which belongs to the Bacteroidetes phylum, was isolated from seawater in South Korea. It showed agar-degrading activity and required an additional carbon source for growth on marine broth 2216. Here, the genome of the strain was sequenced to understand its agar degradation mechanism, and 3615 protein-coding sequences were predicted, which were assigned putative functions according to their annotated functional feature categories. In silico genome analysis revealed that the ERC-38 strain has several carrageenan-degrading enzymes but could not degrade carrageenan because it lacked genes encoding κ-carrageenanase and S1_19A type sulfatase. Moreover, the strain possesses multiple genes predicted to encode enzymes involved in agarose degradation, which are located in a polysaccharide utilization locus. Among the enzymes, Aq1840, which is closest to ZgAgaC within the glycoside hydrolase 16 family, was characterized using a recombinant enzyme expressed in Escherichia coli BL21 (DE3) cells. An enzyme assay revealed that recombinant Aq1840 mainly converts agarose to NA4. Moreover, recombinant Aq1840 could weakly hydrolyze A5 into A3 and NA2. These results showed that Aq1840 is involved in at least the initial agar degradation step prior to the metabolic pathway that uses agarose as a carbon source for growth of the strain. Thus, this enzyme can be applied to development and manufacturing industry for prebiotic and antioxidant food additive. Furthermore, our genome sequence analysis revealed that the strain is a potential resource for research on marine polysaccharide degradation mechanisms and carbon cycling.

Characterization of an endo-1,3-fucanase from marine bacterium Wenyingzhuangia aestuarii: The first member of a novel glycoside hydrolase family GH174.

Sulfated fucans are important marine polysaccharides with various biological and biomedical activities. Fucanases are favorable tools to establish the structure-activity relationships of sulfated fucans. Herein, gene fun174A was discovered from the genome of marine bacterium Wenyingzhuangia aestuarii OF219, and none of the pre-defined glycosidic hydrolase domains were predicted in the protein sequence of Fun174A. Recombinant Fun174A demonstrated a low optimal reaction pH at 5.5. It might degrade sulfated fucans in an endo-processive manner. Glycomics and NMR analyses proved that it specifically hydrolyzed α-1,3-l-fucoside bonds between 2-O-sulfated and non-sulfated fucose residues in the sulfated fucan from sea cucumber Isostichopus badionotus. D119, E120 and E218 were critical for the activity of Fun174A, as identified by site-directed mutagenesis. Three homologs of Fun174A were confirmed to exhibit endo-1,3-fucanase activities. The novelty on sequences of Fun174A and its homologs reveals a new glycoside hydrolase family, GH174.

Mutation of conserved residues in the laminarinase Lam1092 increases the antioxidant activity of the laminarin product hydrolysates.

Laminarinases from the glycoside hydrolase 16 (GH16) family are hydrolases that break ß-1,3-glycosidic bonds in laminarin, which is the major storage polysaccharide present in brown algae or microalgae. We explored a laminarinase from the marine Flavobacteriaceae species Tamlana sp. PT2-4 at the structural and functional levels. Based on a homology model of Lam1092-substrate interactions, the large active groove crossing Lam1092 was deemed a reasonable pathway for the bent substrates for hydrolysis. Eight residues (Gly361, Asn364, Arg400, His466, Asp449, Glu452, Ser477 and Thr538) were selected for mutagenesis based on the interactions of Lam1092 in complex with Lam4/Lam6. Ultimately, we generated eight mutants of Lam1092, and the antioxidant activities of the hydrolysates of two mutants (G361A and H466A) showed significant improvement. These results show that the antioxidant activity of laminarin can be improved by laminarinase mutation, which will be beneficial for developing efficient approaches to engineer the substrate specificity of laminarinases and improve the application of bioactive laminarioligosaccharides.

Paracrocinitomix mangrovi gen. nov., sp. nov., isolated from a mangrove sediment: proposal of two new families, Phaeocystidibacteraceae fam. nov. and Owenweeksiaceae fam. nov., and emended description of the family Schleiferiaceae.

A Gram-stain-negative and short rod-shaped bacterial strain designated GM2-3-6-6T was obtained from a mangrove sediment. Cells were light yellow-pigmented, catalase-positive and oxidase-positive. Carotenoid pigment was produced. Phylogeny of the 16S rRNA gene showed that strain GM2-3-6-6T was affiliated to the family Crocinitomicaceae, sharing maximum sequence similarities with Crocinitomix algicola 0182T, C. catalasitica IFO 15977T, and Putridiphycobacter roseus SM1701T of 93.8%, 93.6%, and 92.5%, respectively. The average nucleotide identity values, digital DNA-DNA hybridization estimates and average amino acid identity values between strain GM2-3-6-6T and the three close relatives were 68.6-68.8%, 18.5-19.2%, and 59.0-62.3%, respectively. The complete circular genome of strain GM2-3-6-6T was 4,365,762 bp in length with a DNA G + C content of 35.0%. The respiratory quinone was MK-7. The major polar lipids consisted of phosphatidylethanolamine, two unidentified phospholipids, one unidentified aminoglycolipid, one unidentified aminolipid and four other unidentified lipids. The major fatty acids were iso-C15:0, iso-C15:1 G, summed feature 3 (C16:1ω7c and/or C16:1ω6c), and iso-C17:0 3-OH. Based on genomic, phenotypic, and chemotaxonomic characterizations, strain GM2-3-6-6T represents a novel species of a novel genus, for which the name Paracrocinitomix mangrovi gen. nov., sp. nov. is proposed. The type strain is GM2-3-6-6T (= MCCC 1K04831T = KCTC 82931T). Additionally, phylogenomic analysis of the type strains of the family Schleiferiaceae and family Cryomorphaceae related members including uncultivated bacteria, was performed using the Genome Taxonomic Database toolkit (GTDB-Tk). Based on 16S rRNA gene phylogeny and genomic features, two novel families, Phaeocystidibacteraceae fam. nov. and Owenweeksiaceae fam. nov. are proposed. An emended description of the family Schleiferiaceae is also proposed.