rRNA Nucleotide Research Articles

Mechanistic Insights into Clinically Relevant Ribosome-Targeting Antibiotics

Antibiotics targeting the bacterial ribosome are essential to combating bacterial infections. These antibiotics bind to various sites on the ribosome, inhibiting different stages of protein synthesis. This review provides a comprehensive overview of the mechanisms of action of clinically relevant antibiotics that target the bacterial ribosome, including macrolides, lincosamides, oxazolidinones, aminoglycosides, tetracyclines, and chloramphenicol. The structural and functional details of antibiotic interactions with ribosomal RNA, including specific binding sites, interactions with rRNA nucleotides, and their effects on translation processes, are discussed. Focus is placed on the diversity of these mechanisms and their clinical implications in treating bacterial infections, particularly in the context of emerging resistance. Understanding these mechanisms is crucial for developing novel therapeutic agents capable of overcoming bacterial resistance.

Enhancing the biotransformation of progesterone to the anticancer compound testololactone by Penicillium chrysogenum Ras3009: kinetic modelling and efficiency maximization

BackgroundBiotransformation of steroid compounds into therapeutic products using microorganisms offers an eco-friendly and economically sustainable approach to the pharmaceutical industry rather than a chemical synthesis way. The biotransformation efficiency of progesterone into the anticancer compound testololactone using Penicillium chrysogenum Ras3009 has been investigated. Besides, maximization of testololactone formation was achieved by studying the kinetic modelling and impact of some fermentation conditions on the biotransformation process.ResultsThe fungal strain Ras3009 was selected among twelve fungal strains as the most runner for the transformation of 81.18% of progesterone into testololactone. Ras3009 was identified phenotypically and genotypically as Penicillium chrysogenum, its 18 S rRNA nucleotide sequence was deposited in the GenBank database by the accession number OR480104. Studying the impact of fermentation conditions on biotransformation efficiency indicated a positive correlation between substrate concentration and testololactone formation until reaching the maximum velocity vmax. Kinetic studies revealed that vmax was \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\:0.0482$$\\end{document} gL− 1hr− 1 with high accuracy, giving R2 of 0.977. The progesterone transformation efficiency generally increased with time, reaching a maximum of 100% at 42 h with testololactone yield (Ypt/s) 0.8700 mg/mg. Moreover, the study indicated that the enzymatic conversion by P. chrysogenum Ras3009 showed high affinity to the substrate, intracellularly expressed, and released during cell disruption, leading to higher efficiency when using whole microbial cell extract.ConclusionsFungi can be promising biocatalysts for steroid transformation into valuable chemicals and pharmaceutical compounds. The study revealed that the new fungal isolate P. chrysogenum Ras3009 possesses a great catalytic ability to convert progesterone into testololactone. Kinetic modelling analysis and optimization of the fermentation conditions lead to higher transformation efficiency and provide a better understanding of the transformation processes.Graphical abstract

Intraspecies mitogenomic variation in the Black-throated Tit (Aegithalos concinnus): conserved structure, concerted evolution of duplicate control regions and multiple distinct evolutionary lineages

The mitochondrial genome is a prominent research topic due to its indispensable role in organisms and its application in many research disciplines. However, few studies have investigated intraspecies mitogenomic variation. In this study, 69 mitogenomes of the Black-throated Tit (Aegithalos concinnus) were assembled and annotated from a large number of short reads generated using high-throughput sequencing technology. Comparative analyses revealed that mitogenomic characteristics such as length, gene and nucleotide composition, codon usage, and duplicated control regions were relatively conserved despite substantial intraspecies morphological changes. Yet, all the individuals from the subspecies A. c. iredalei had one more nucleotide in the 12S rRNA than the other studied subspecies. Phylogenetic analyses showed five distinct lineages based on the complete mitogenomes and the 13 combined protein-coding genes, whereas only four lineages were observed when using the duplicate control regions. Most interestingly, each lineage had both copies of the control regions of the comprising individuals, indicating that the paralogous control regions were more similar than the orthologous sequences from the distinct lineages. This suggested the control regions had undergone concerted evolution. The Black-throated Tit has complex evolutionary history and needs further investigating the taxonomic status of these lineages, as well as the underlying evolutionary processes. Our findings call for more research on intraspecies mitogenomic variation.

Ribosomal a site binding pattern differs between arm methyltransferases from clinical pathogens and a natural producer of aminoglycosides.

The extensive use of aminoglycosides to treat bacterial infections has led to significant resistance, posing a global health threat. Recent clinical reports highlight high levels of aminoglycoside resistance due to Arm/Kam methyltransferases, which methylate specific nucleotides in 16S rRNA, preventing antibiotic binding to the ribosome. This study compared the ribosomal A site binding patterns of Arm methyltransferases from clinical pathogens (ArmA, RmtB, RmtC, and RmtD) with those of the Sgm methyltransferase from a natural aminoglycoside producer. We introduced single mutations near the G1405 nucleotide in helix 44 of 16S rRNA to assess their impact on the methylation ability of Arm methyltransferases in E. coli cells with homogeneous mutant ribosomes. We evaluated how these mutations affected bacterial viability in cells with mixed and homogeneous ribosome populations and determined the minimal inhibitory concentration of kanamycin to assess their impact on Arm enzyme activity. Notably, Sgm methyltransferase exhibited a distinct methylation pattern compared to Arm methyltransferases from clinical strains. Structural comparisons of Sgm, RmtB, and RmtC revealed different spatial orientations of key amino acids involved in ribosomal binding, highlighting evolutionary differences. This research enhances understanding of Arm methyltransferases and lays the groundwork for designing inhibitors to combat this potent form of antibiotic resistance.

Mechanistic Basis for the Translation Inhibition of Cutibacterium acnes by Clindamycin

Inflammation and the Gram-positive anaerobic bacterium Cutibacterium acnes, which is implicated in acne pathogenesis and pilosebaceous unit inflammation, are the main targets of antibiotic-based therapy against acne vulgaris (acne). The most widely used antibiotics in acne therapy are tetracyclines, macrolides, and lincosamides. Unfortunately, C. acnes bacteria over the past several decades have demonstrated increased resistance to these antibiotics, particularly to clindamycin. The precise knowledge of how antibiotics interact with their clinical target is needed to overcome this problem. Toward this goal, we determined the structure of clindamycin in complex with the ribosome of Cutibacterium acnes at 2.53 Å resolution using cryogenic electron microscopy. The galactose sugar moiety of clindamycin interacts with nucleotides of the 23S rRNA directly or through a conserved network of water-mediated interactions. Its propyl pyrrolidinyl group interacts with the 23S rRNA through van der Waals forces. Clindamycin binding to the Cutibacterium acnes ribosome interferes with both: proper orientation of the aminoacyl group of the A-site bound tRNA that is needed for peptide bond formation and with the extension of the nascent peptide. Our data are important for advancing understanding of antibiotic resistance and development of narrow- spectrum antibacterial drugs, which is an urgent need for contemporary antibiotic stewardship.

Study on the ability of activated sludge bacteria to form biofilms <i>in vitro</i>

The study aims to comparatively characterize in vitro biofilm formation in bacterial cultures isolated from activated sludge, as well as archival cultures capable of xenobiotics biodegradation: Alcaligenes faecalis 2, Acinetobacter guillouiae 11h, Rhodococcus erythropolis ILBIO, and Achromobacter pulmonis PNOS. An analysis of the 16S rRNA nucleotide sequence identified strains isolated from activated sludge: Paenibacillus odorifer, Bacillus subtilis, Micrococcus yunnanensis, and Bacillus proteolyticus. The formation of biofilms by microorganisms was studied on LB medium and synthetic culture medium (with sodium acetate as a carbon source). With cell growth on LB medium, an increase in biofilm biomass was observed in Paenibacillus odorifer, Bacillus subtilis, Alcaligenes faecalis 2, and Achromobacter pulmonis PNOS. The cultivation stage duration (72 and 144 h), as well as the additional dosing of substrates, had an effect on the biofilm formation process: by 144 h of cultivation, the biomass values amounted to 0.6–1.3 optical units. An average 63–77% increase in biofilm biomass was noted for Bacillus subtilis and Paenibacillus odorifer cells as compared to the 72-hour process. At the final stage of cultivation (144 h), the values of exopolysaccharides in the matrix amounted to over 0.02 optical units for Bacillus subtilis and Paenibacillus odorifer. The metabolic activity of activated sludge bacteria forming the biofilm reached 628–3609 Fl./OD540. Thus, activated sludge microorganisms forming the biofilm were shown to retain viability and metabolic activity during growth under in vitro conditions.

Morphologic and phylogenetic investigations revealed size-divergent clades in chelae morphotypes of freshwater prawn Macrobrachium vollenhovenii Herklots (Decapoda: Palaemonidae) in a lake and river system of Southwest Nigeria

BackgroundThe freshwater prawn Macrobrachium vollenhovenii is one of the largest Macrobrachium species, a biological agent against human schistosomiasis, and a cheap protein source in riverine communities in West Africa. However, its aquaculture development for sustainable utilization is challenged by cryptic identity amidst the presence of morphotypes of unknown size and genetic relatedness. This study aimed to investigate the maximum sizes and evolutionary links of chelae morphotypes in M. vollenhovenii for precise identification and utilization in a 3 × 2 randomized block experimental design. Ninety biggest encountered samples of M. vollenhovenii chelae morphotypes—those possessing equal left and right side chelae, longer left chelae, and shorter left chelae—were obtained from fisherfolks’ catches at each of Asejire Lake and Ogun River during peak seasons (July–September) bimonthly field survey, representing EAAL, LLAL, SLAL—GAALs, and EAOR, LLOR, SLOR—GAORs. These were analyzed for differences (p < 0.05) in size-linked parameters—length (L (cm)), weight (W (g)), and condition factor (K). Specimens’ 16S rRNA nucleotide sequences were utilized to infer phylogenetic linkages, single-nucleotide polymorphism (SNP), and amino acid translations alongside NCBI references (NCBIrefseq).ResultsWeight (W) and condition factor (K), among GAALs, for SLAL and LLAL were similar; EAAL was significantly lowest; among GAORs, SLOR and LLOR were similar; and EAOR was significantly lowest. In GAALs, EAAL, LLAL, and SLAL had higher L, W, and K than counterpart GAORs. Sequences formed two polyphyletic groups: EAOR branch from EAAL, in which 100.0%EA rooted 75.0% NCBIrefseqs, forming a clade; and GAOR-SLOR and LLOR branch form GAOR-LLAL rooted SLAL, in which 100.0%LL and SL rooted KJ463387.1 (Badagry), forming another clade. SNP Locus 91 separated 100.0%GAOR from 100.0%GAAL and 100.0%NCBIrefseqs translating to valine; SNP Locus 171 separated 100.0%EA and its co-rooted NCBIrefseqs from 100.0%LL, SL, and their co-rooted NCBIrefseq, translating to glycine/glutamic acid change.ConclusionThe equal left and right side chelae and the unequal left and right side chelae specimens are, respectively, small- and robust-sized, irrespective of habitat. They are divergent size-linked clades having protein translate differences, delineable at 16S rRNA SNP Locus 171; their size variant habitat strains are delineable at SNP Locus 91. These SNP markers will be useful for precision identification and selection of the size variant chelae morph strains for sustainable utilization.

Native bacterial bioagents for management of potato soft rot disease caused by Pectobacterium carotovorum subsp. carotovorum

BackgroundThe reduced rate of potato production may occur under different conditions such as field, transit, storage, and marketing. Potato cultivation is frequently affected by various pathogens, among which Pectobacterium carotovorum subsp. carotovorum is a notorious bacterial pathogen responsible for causing bacterial soft rot disease. This pathogen poses a significant threat to potato production worldwide, resulting in substantial economic losses and food security concerns. This study aimed to investigate the effectiveness of three bacterial bioagents, namely Brevundimonas bullata, Bacillus siamensis, and Bacillus velezensis, against P. carotovorum subsp. carotovorum, a notorious bacterium responsible for causing potato tuber soft rot disease.ResultsFifteen isolates were isolated from rhizosphere of potato plants. Out of 15 isolates, 3 isolates, No. 1, 11, and 12, showed highly antagonistic property to control the growth of P. carotovorum subsp. carotovorum in vitro. They were identified as B. bullata, B. velezensis, and B. siamensis, by using 16S rRNA nucleotide sequence analysis. Results from these experiments revealed that three bioagents exhibited notable inhibitory effects on the growth of the pathogenic bacterium. However, B. velezensis stood out as the most effective inhibitor among the tested bioagents, showcasing the highest level of bacteriostatic activity (2.6 cm). The findings from the in vitro experiments provided promising insights into the potential of these bacterial bioagents as natural antagonists against potato tuber soft rot disease. Building upon these promising results, the study recommended the utilization of all tested bioagents for controlling soft rot disease in potato tubers, especially during storage conditions where the risk of bacterial proliferation is heightened. Treatment with B. bullata, B. siamensis, and B. velezensis demonstrates varying degrees of efficacy in controlling the progression of soft rot disease. Notably, B. velezensis shows promising results with relatively low disease severity scores across all time points compared to the other treatments. B. velezensis demonstrates the lowest disease severity percentage at 7.00% compared to the other treatments.ConclusionsThe study underscores the significant potential of B. bullata, B. siamensis, and B. velezensis as effective biological control agents against potato tuber soft rot disease. By targeting the causative agent directly, these bioagents offer a promising avenue for mitigating the detrimental impact of soft rot disease on potato production, thereby contributing to improved crop yield and quality.

Loss of Conserved rRNA Modifications in the Peptidyl Transferase Center Leads to Diminished Protein Synthesis and Cell Growth in Budding Yeast.

Ribosomal RNAs (rRNAs) are extensively modified during the transcription and subsequent maturation. Three types of modifications, 2'-O-methylation of ribose moiety, pseudouridylation, and base modifications, are introduced either by a snoRNA-driven mechanism or by stand-alone enzymes. Modified nucleotides are clustered at the functionally important sites, including peptidyl transferase center (PTC). Therefore, it has been hypothesised that the modified nucleotides play an important role in ensuring the functionality of the ribosome. In this study, we demonstrate that seven 25S rRNA modifications, including four evolutionarily conserved modifications, in the proximity of PTC can be simultaneously depleted without loss of cell viability. Yeast mutants lacking three snoRNA genes (snR34, snR52, and snR65) and/or expressing enzymatically inactive variants of spb1(D52A/E679K) and nop2(C424A/C478A) were constructed. The results show that rRNA modifications in PTC contribute collectively to efficient translation in eukaryotic cells. The deficiency of seven modified nucleotides in 25S rRNA resulted in reduced cell growth, cold sensitivity, decreased translation levels, and hyperaccurate translation, as indicated by the reduced missense and nonsense suppression. The modification m5C2870 is crucial in the absence of the other six modified nucleotides. Thus, the pattern of rRNA-modified nucleotides around the PTC is essential for optimal ribosomal translational activity and translational fidelity.

The Partial Nucleotide Sequences of the Mitochondrial Genes, COI and 16s rRNA, of Fireflies in the Genera Pygoluciola, Trisinuata, and Medeopteryx (Coleoptera: Lampyridae)

Fireflies in the genera Pygoluciola, Medeopteryx, and Trisinuata distribute throughout lower northern Thailand. Five morphospecies of these fireflies were collected and nucleotide sequences were analyzed. We sequenced the partial of two mitochondrial genes; Cytochrome c oxidase subunit I (COI) and 16s rRNA, and the phylogenetic trees were constructed and compared with the firefly nucleotide sequences in the NCBI database. The COI gene was amplified using LCO1490 and HCO2198 primers while specific primers of LR-J-13020a and LR-N-13398a were used to analyze amplicons of 16s rRNA. The results showed the average GC-contents in the nucleotide of five firefly morphospecies were found to average 30.28% and 22.15% in (COI) and 16s rRNA, respectively. The variable site in COI nucleotide sequences was 399 of the total 520 nucleotides (76.73%) whereas 91 of the total 418 nucleotides (21.77%) were found in 16s rRNA nucleotide sequences. Construction of the phylogenetic trees of those five firefly morphospecies with reference accession samples indicated five clades when analyzed by COI nucleotide sequences and two clades were revealed in 16s rRNA gene. According to these results, COI nucleotides are one of the most effective methods for differentiating between and within firefly species. The addition of the 16s rRNA gene analysis showed an increase in the precision of firefly identification. Those five firefly morphospecies were separated using both the COI and 16s rRNA nucleotide sequences.

Sphingomonas flavescens sp. nov., isolated from soil.

An aerobic bacterium, designated as PT-12T, was isolated from soil collected from agriculture field, and its taxonomic position was validated through a comprehensive polyphasic methodology. The strain was identified as Gram-stain-negative, non-motile, rod-shaped, and catalase- and oxidase-positive. The yellow-colored colonies showed growth ability at temperature range of 18-37°C, NaCl content of 0-1.0% (w/v), and at a pH of 6.0-8.0. The 16S rRNA gene and phylogenetic analysis showed that strain PT-12T affiliated with the genus Sphingomonas in the family Sphingomonadaceae, and displayed the highest 16S rRNA nucleotide sequence similarity with Sphingomonas limnosediminicola 03SUJ6T (98.4%). The genome size of strain PT-12T was 2,656,862bp and the DNA G + C content estimated from genome was 63.5%. The highest values of average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) were observed between strain PT-12T and Sphingomonas segetis YJ09T, accounting to 76.2% and 20.2%, respectively. In addition, both ANI and dDDH values between strain PT-12T and other phylogenetically related neighbors ranged between 69.6% and 76.2% and 18.4% and 20.2%, respectively. Chemotaxonomic features exhibited Q-10 as the only ubiquinone; homospermidine as the major polyamine; summed feature 8 (C18:1ω7c and/or C18:1ω6c), C16:0, and 10-methyl C18:0 as the notable fatty acids; and phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, phosphatidylcholine, and sphingoglycolipid as dominating polar lipids. Overall, the comprehensive polyphasic data supported that strain PT-12T represents a novel bacterial species within the genus Sphingomonas. Accordingly, we propose the name Sphingomonas flavescens sp. nov. The type strain is PT-12T (= KCTC 92114T = NBRC 115717T).

Characterization of the Beauveria bassiana ga fungal strain isolated from the muscardine infected silkworms (Bombyx mori L.)

In this study, the focus was on isolating and characterizing the fungal strain Beauveria bassiana from muscardine-infected silkworms (Bombyx mori L.). The fungal strain isolated from the muscardine-infected silkworm, designated as GA, closely resembled Beauveria bassiana isolate SBI-Bb04 based on morphological analysis and comparison of the 18S rRNA nucleotide sequence with sequences in the NCBI database. Beauveria bassiana GA demonstrates the ability to grow on Sabouraud Dextrose Agar (SDA) medium. Optimal conditions for its growth were determined to be a pH range of 6-7 and a temperature range of 20-25°C. The composition of SDA, containing glucose and peptone sources, was found to be ideal for both growth and sporulation of this fungal strain. Furthermore, Beauveria bassiana GA exhibits the capacity to synthesize various extracellular enzymes, including chitinase, protease, cellulase, and amylase. These enzymatic activities contribute to its pathogenicity and suggest its potential as a biocontrol agent against insect pests.

First Report of Pectobacterium aroidearum Causing Soft Rot on Zamioculcas zamiifolia

&lt;i&gt;Zamioculcas zamiifolia&lt;/i&gt; is a popular indoor ornamental plant in Korea. In August 2021, a severe outbreak of soft rot disease affected &lt;i&gt;Z. zamiifolia&lt;/i&gt; in Emseong, Chungcheongbuk-do, Korea. Infected plants displayed wilting, water-soaked lesions, stem collapse, and green-brown discoloration. The bacterial strain KNUB-05-21 was isolated from infected stems and identified as &lt;i&gt;Pectobacterium aroidearum&lt;/i&gt; using 16S rRNA nucleotide sequencing and multilocus sequence analysis based on partial sequences of &lt;i&gt;dnaX, leuS&lt;/i&gt;, and &lt;i&gt;recA&lt;/i&gt; genes. Confirmation of its affiliation with &lt;i&gt;P. aroidearum&lt;/i&gt; was also obtained through biochemical and morphological characterization. To confirm the pathogenicity of strain KNUB-05-21, its suspension was injected into &lt;i&gt;Z. zamiifolia&lt;/i&gt; stems. Within a week, soft rot developed on the stems, exhibiting symptoms similar to those observed in field-infected plants. The reisolated strain was identical to those of &lt;i&gt;P. aroidearum&lt;/i&gt;. Before this study, &lt;i&gt;P. aroidearum&lt;/i&gt; was not reported as a causative pathogen of &lt;i&gt;Z. zamiifolia&lt;/i&gt; soft rot in Korea.

The RNA helicase Dbp10 coordinates assembly factor association with PTC maturation during ribosome biogenesis.

During ribosome biogenesis a plethora of assembly factors and essential enzymes drive the unidirectional maturation of nascent pre-ribosomal subunits. The DEAD-box RNA helicase Dbp10 is suggested to restructure pre-ribosomal rRNA of the evolving peptidyl-transferase center (PTC) on nucleolar ribosomal 60S assembly intermediates. Here, we show that point mutations within conserved catalytic helicase-core motifs of Dbp10 yield a dominant-lethal growth phenotype. Such dbp10 mutants, which stably associate with pre-60S intermediates, impair pre-60S biogenesis at a nucleolar stage prior to the release of assembly factor Rrp14 and stable integration of late nucleolar factors such as Noc3. Furthermore, the binding of the GTPase Nug1 to particles isolated directly via mutant Dbp10 bait proteins is specifically inhibited. The N-terminal domain of Nug1 interacts with Dbp10 and the methyltransferase Spb1, whose pre-60S incorporation is also reduced in absence of functional Dbp10 resulting in decreased methylation of 25S rRNA nucleotide G2922. Our data suggest that Dbp10's helicase activity generates the necessary framework for assembly factor docking thereby permitting PTC rRNA methylation and the progression of pre-60S maturation.

Standard- and large-sized eggs of Trichuris trichiura in the feces of schoolchildren in the Yangon Region, Myanmar: Morphological and molecular analyses.

Standard- and large-sized eggs of Trichuris trichiura were found in the feces of schoolchildren in Yangon, Myanmar during epidemiological surveys and mass deworming with albendazole in 2017-2019. The standard-sized eggs were identified as those of T. trichiura, but it was necessary to exclude the possibility of the large-sized eggs belonging to Trichuris vulpis, a dog whipworm. We conducted morphological and molecular studies to determine the species of the 2 types of Trichuris eggs. Individual eggs of both sizes were isolated from Kato-Katz fecal smears (n=20) and mechanically destroyed using a 23G injection needle. Nuclear DNA was extracted, and the 18S rRNA region was sequenced in 15 standard-sized eggs and 15 large-sized eggs. The average size of standard-sized eggs (T. trichiura) was 55.2×26.1 μm (range: 51.7-57.6×21.3-28.0 μm; n=97), whereas the size of large-sized eggs was 69.3×32.0 μm (range: 65.1-76.4×30.1-34.5 μm; n=20), slightly smaller than the known size of T. vulpis. Regarding standard-sized eggs, the 18S rRNA nucleotide sequences exhibited 100% homology with T. trichiura deposited in GenBank and 88.6-90.5% homology with T. vulpis. Regarding large-sized eggs, the nucleotide sequences showed 99.8-100% homology with T. trichiura in GenBank and 89.6-90.7% homology with T. vulpis. Both standard- and large-sized eggs of Trichuris spp. found in Myanmar schoolchildren during 2017-2019 were morphologically and molecularly confirmed to belong to T. trichiura. The conversion of eggs from smaller to large sizes might be due to anthelmintic treatments with albendazole.

Year-Round Presence of Microcystins and Toxin-Producing Microcystis in the Water Column and Ice Cover of a Eutrophic Lake Located in the Continuous Permafrost Zone (Yakutia, Russia).

This study aimed to test the hypothesis of the year-round presence of toxigenic Microcystis and cyanotoxins in the water and ice of the shallow eutrophic Lake Ytyk-Kyuyol located in the continuous permafrost zone. Three independent approaches-mass-spectrometry, molecular methods and light microscopy-were applied in the study. The cyanobacterial biomass ranged from 1.0 × 10-4 to 4.8 mg L-1. Microcystis flos-aquae and M. aeruginosa were the dominant morphospecies in plankton throughout the observation. In environmental DNA, the presence of M. aeruginosa was supported and mcy gene regions responsible for microcystin biosynthesis were detected through a BLAST (Basic Local Alignment Search Tool) search and phylogenetic estimation based on newly obtained 16S rRNA, 16S-23S ITS rRNA, mcyA and mcyE nucleotide sequences. The intracellular microcystin concentration ranged from <0.1 to 803 ng L-1, and the microcystin quota in the Microcystis biomass was extremely low. For the first time, it was shown that Microcystis cells containing mcy genes and microcystins presented permanently in the water column, both during the ice-free period and under ice, as well as inside thick ice covers within 7 months of severe winter. We hypothesized that minor pelagic and ice populations of Microcystis could participate in increasing cell density in the spring. However, further studies are needed to confirm the viability of the overwintering Microcystis colonies in the water and inside the ice of Lake Ytyk-Kyuyol.

Genetic diversity and phylogenetic analysis of mayfly Caenis (Insecta: Ephemeroptera) using Cytochrome C Oxidase I (COI) and 12s rRNA genes from Thailand

Abstract. Prakrongrak N, Boonsoong B, Monthatong M. 2023. Genetic diversity and phylogenetic analysis of mayfly Caenis (Insecta: Ephemeroptera) using Cytochrome C Oxidase I (COI) and 12s rRNA genes from Thailand. Biodiversitas 24: 1989-1997. Mayflies in genus Caenisis one of the top fifth species richnessbelonged to Family Caenidae. The present study aimed to use the partial mitochondrial Cytochrome COxidase subunit I (COI) and 12s rRNA nucleotide sequences for molecular species identification, diversity and phylogenetic relationships of mayflies in genus Caenis Stephens, 1835 species from Thailand. From a total of 37 specimens, thirteen nymphs were morphologically identified into five species: Caenis nasuta Malzacher, C. Picea Kimmins, C. ulmeriana Malzacher, C. cornigera Kang and Yang and C. longiforcipata Malzacher. The other 24 samples were Caenis sp.1 to sp.5. Partial COI sequences of all samples were compared with BOLD and GenBank for species identification. However, the result showed only genus-level identification as Caenis with greater than 80% similarity. For species delimitation, interspecific genetic distance among these species ranged from 14.4 to 26.6% (COI) and from 7.36 to 22.4% (12s rRNA). Intraspecific divergence levels were 0.0% to 2.35%. ABGD analysis of both genes divided data into 10 groups corresponding to 10 morphospecies of Caenis. The phylogenetic relationships of Caenis, COI and 12s rRNA genes also classified each species to well supported clusters. In addition, we report C. cornigera from Thailand for the first time. The COI and 12s rRNA phylogenetic trees indicate a close relationship between C. cornigera and C. longiforcipata, which is supported by their similar morphology.

Structural basis for reduced ribosomal A-site fidelity in response to P-site codon–anticodon mismatches

Rapid and accurate translation is essential in all organisms to produce properly folded and functional proteins. mRNA codons that define the protein-coding sequences are decoded by tRNAs on the ribosome in the aminoacyl (A) binding site. The mRNA codon and the tRNA anticodon interaction is extensively monitored by the ribosome to ensure accuracy in tRNA selection. While other polymerases that synthesize DNA and RNA can correct for misincorporations, the ribosome is unable to correct mistakes. Instead, when a misincorporation occurs, the mismatched tRNA-mRNA pair moves to the peptidyl (P) site and, from this location, causes a reduction in the fidelity at the A site, triggering post-peptidyl transfer quality control. This reduced fidelity allows for additional incorrect tRNAs to be accepted and for release factor 2 (RF2) to recognize sense codons, leading to hydrolysis of the aberrant peptide. Here, we present crystal structures of the ribosome containing a tRNALys in the P site with a U•U mismatch with the mRNA codon. We find that when the mismatch occurs in the second position of the P-site codon-anticodon interaction, the first nucleotide of the A-site codon flips from the mRNA path to engage highly conserved 16S rRNA nucleotide A1493 in the decoding center. We propose that this mRNA nucleotide mispositioning leads to reduced fidelity at the A site. Further, this state may provide an opportunity for RF2 to initiate premature termination before erroneous nascent chains disrupt the cellular proteome.

Chordicoccus furentiruminis, gen. nov., sp. nov., a novel succinic acid producing bacterium isolated from a steer on a high grain diet.

This study presents MP1D12T (=NRRL B-67553T=NCTC 14480T), an isolate from the ruminal content of an Angus steer fed a high grain diet. Phenotypic and genotypic traits of the isolate were explored. MP1D12T was found to be a strictly anaerobic, catalase-negative, oxidase-negative, coccoid bacterium that frequently grows in chains. Analysis of metabolic products as a result of carbohydrate fermentation showed succinic acid as the major organic acid produced with lactic acid and acetic acid as minor products. Phylogenetic analysis of MP1D12T based on 16S rRNA nucleotide sequence and amino acid sequences from the whole genome presents a divergent lineage from other members in the family Lachnospiraceae. 16S rRNA sequence comparison, whole genome average nucleotide identity digital DNA-DNA hybridization and average amino acid identity results suggest that MP1D12T represents a novel species in a novel genus within the family Lachnospiraceae. We propose the creation of the genus Chordicoccus in which MP1D12T represents the type strain for the novel species Chordicoccus furentiruminis.

Isolation and Identification of New Soil Strains with Phosphate-Solubilizing and Exopolysaccharide-Producing Abilities in the Yellow River Wetland Nature Reserve of Luoyang City, China

The establishment of the Yellow River wetland nature reserves improves the local soil structure and fertility through the long-term succession of microorganisms. However, little is known about which indigenous microbial resources can accelerate the process of soil improvement and ecology restoration. To fill this gap, exopolysaccharides-producing bacteria and phosphate-solubilizing bacteria were isolated from soil samples of the wetland nature reserve with higher soil organic matter, available phosphorus, and available nitrogen content. 16S rRNA nucleotide sequence homology analysis and physiological-biochemical assay showed that the strain PD12 with the highest phosphate solubilization activity and higher EPS production was identified as Klebsiella variicola, and other high yield EPS-producing strains (EPS12, EPS15, EPS18, and EPS19) were identified as Pseudomonas migulae, Pseudomonas frederiksbergensis, Aeromonas media, and Pseudomonas vancouverensis, respectively. These results provided new potential microbial resources for the research and development of biofertilizers and added new insights into accelerating the restoration of physical, chemical, and biological properties of soil in the Yellow River basin.