16S Ribosomal DNA Research Articles

Diagnostic performance and comparison of ultrasensitive and conventional rapid diagnostic test, thick blood smear and quantitative PCR for detection of low-density Plasmodium falciparum infections during a controlled human malaria infection study in Equatorial Guinea

BackgroundProgress towards malaria elimination has stagnated, partly because infections persisting at low parasite densities comprise a large reservoir contributing to ongoing malaria transmission and are difficult to detect. This study compared the performance of an ultrasensitive rapid diagnostic test (uRDT) designed to detect low density infections to a conventional RDT (cRDT), expert microscopy using Giemsa-stained thick blood smears (TBS), and quantitative polymerase chain reaction (qPCR) during a controlled human malaria infection (CHMI) study conducted in malaria exposed adults (NCT03590340).MethodsBlood samples were collected from healthy Equatoguineans aged 18–35 years beginning on day 8 after CHMI with 3.2 × 103 cryopreserved, infectious Plasmodium falciparum sporozoites (PfSPZ Challenge, strain NF54) administered by direct venous inoculation. qPCR (18s ribosomal DNA), uRDT (Alere™ Malaria Ag P.f.), cRDT [Carestart Malaria Pf/PAN (PfHRP2/pLDH)], and TBS were performed daily until the volunteer became TBS positive and treatment was administered. qPCR was the reference for the presence of Plasmodium falciparum parasites.Results279 samples were collected from 24 participants; 123 were positive by qPCR. TBS detected 24/123 (19.5% sensitivity [95% CI 13.1–27.8%]), uRDT 21/123 (17.1% sensitivity [95% CI 11.1–25.1%]), cRDT 10/123 (8.1% sensitivity [95% CI 4.2–14.8%]); all were 100% specific and did not detect any positive samples not detected by qPCR. TBS and uRDT were more sensitive than cRDT (TBS vs. cRDT p = 0.015; uRDT vs. cRDT p = 0.053), detecting parasitaemias as low as 3.7 parasites/µL (p/µL) (TBS and uRDT) compared to 5.6 p/µL (cRDT) based on TBS density measurements. TBS, uRDT and cRDT did not detect any of the 70/123 samples positive by qPCR below 5.86 p/µL, the qPCR density corresponding to 3.7 p/µL by TBS. The median prepatent periods in days (ranges) were 14.5 (10–20), 18.0 (15–28), 18.0 (15–20) and 18.0 (16–24) for qPCR, TBS, uRDT and cRDT, respectively; qPCR detected parasitaemia significantly earlier (3.5 days) than the other tests.ConclusionsTBS and uRDT had similar sensitivities, both were more sensitive than cRDT, and neither matched qPCR for detecting low density parasitaemia. uRDT could be considered an alternative to TBS in selected applications, such as CHMI or field diagnosis, where qualitative, dichotomous results for malaria infection might be sufficient.

Biocontrol potential of plant growth-promoting rhizobacteria (PGPR) against Sclerotiorum rolfsii diseases on sugar beet (Beta vulgaris L.)

A total of 49 bacterial isolates were collected from the sugar beet rhizosphere in two regions of Morocco (Kenitra and Meknes). These bacteria were tested for their inhibitory activity against Sclerotium rolfsii Sacc., the causative pathogen of damping-off and root rot of sugar beet. Ten isolates showed a high ability to inhibit the mycelial growth of the pathogenic fungus in dual culture bioassays. The molecular identification based on 16S ribosomal DNA gene revealed that these isolates belong to the Bacillus genus. In addition, these bacterial isolates have also been characterized in terms of the production of certain antifungal products as well as for their capacities as plant growth-promoting traits. Under greenhouse conditions, sugar beet seeds treated with the each bacterium showed significant reduction in damping-off disease. Plants from treated seeds showed significant increases in both the length of shoots and roots, and the plant fresh and dry weight. Iinterestingly, these selected bacterial isolates displayed high antagonistic activity against S. rolfsii, and significantly reduced the severity and incidence of root rot disease. Therefore, these results highlighted that the soil harbors antagonistic bacteria offering several plant growth-promoting (PGP) traits, which can be exploited as a powerful growth promoter and/or biological control agent in sugar beet plants.

Pyropia yezoensis porphyran alleviates metabolic disorders via modulating gut microbiota in high-sucrose-fed Drosophila melanogaster.

Prebiotics, such as algal polysaccharides, can be used to manage metabolic diseases by modulating gut microbiota. However, the effect of Pyropia yezoensis porphyran (PYP), a red algal polysaccharide, on gut microbiota has not been reported. Thus, the objective of this study was to determine effects of PYP on metabolic disorders caused by high sucrose (HS) and underlying mechanisms involved in such effects. Biochemical analysis demonstrated that an HS diet increased triglyceride and circulating sugar contents (metabolic abnormalities) in Drosophila larvae. It also increased the relative abundance of harmful microbiota within the larvae as identified by 16S ribosomal DNA analysis. PYP supplementation at 25 and 50 g kg-1 equivalently reduced metabolic abnormalities in the HS group. Therefore, 25 g kg-1 PYP was selected to investigate its effects on the metabolic pathway and gut microbiota of larvae in the HS group. The activity of PYP in ameliorating metabolic abnormalities by reverse transcription quantitative real-time polymerase chain reaction analysis was consistent with the expression trend of key factors involved in metabolism regulation. PYP reduced the relative abundance of bacteria causing metabolic abnormalities, such as Escherichia-Shigella and Fusobacterium, but increased the relative abundance of beneficial bacteria such as Bacillus and Akkermansia. However, PYP had no effect on triglyceride and circulating sugar contents in HS-fed larvae treated with a mixture of antibiotics designed to remove gut microbiota. PYP exhibits anti-metabolic disorder activity by modulating gut microbiota, thereby supporting the development of PYP as a functional prebiotic derived from red algae food. Copyright © 2022 John Wiley & Sons, Ltd. © 2022 Society of Chemical Industry.

Identification of gut microbiome and transcriptome changes in ulcerative colitis and pouchitis

Background Pouchitis is the common postoperative complication of ulcerative colitis (UC) and is also considered as inflammatory bowel disease. The aim was to investigate the microbiological and transcriptional differences between the two illnesses. Methods Eighty-five participants were enrolled (37 UC, 15 healthy UC pouches, 15 pouchitis and 18 healthy volunteers) and stool samples were collected. Microbial populations were analyzed by pyrosequencing of 16S ribosomal DNA. Furthermore, transcriptome data of 119 UC and 28 pouch patients were obtained from two data sets for bioinformatics analysis. Results The results of gut microbiota community analysis showed that with aggravation of UC, intestinal microorganisms were characterized by a gradual decreased in diversity and numbers of butyrate-producing bacteria and Bacteroides. Besides, in addition to the decrease of probiotics, the proliferation of Escherichia–Shigella and Ruminococcus gnavus was observed in pouchitis which is related to multiple infection pathways. The function enrichment of differential expression genes and hub genes, as well as the immunological condition was shown to be distinct using transcriptome bioinformatics analysis between UC and pouchitis. A stronger immune response occurs in UC and may be associated with high expression of tumor necrosis factor and interleukin, while multiple hub genes such as CDK1 in pouchitis are associated with cell cycle regulation. Conclusions The characteristics of gut microbiota disturbance and transcriptome alteration in UC and pouchitis are different. Our findings suggested that pouchitis may have a unique pathogenesis which was separated from UC.

Berberine Improves Vascular Dysfunction by Inhibiting Trimethylamine-N-oxide via Regulating the Gut Microbiota in Angiotensin II-Induced Hypertensive Mice

Berberine (BBR) has been demonstrated to exert cardiovascular protective effects by regulating gut microbiota. However, few studies examine the effect of BBR on the gut microbiota in hypertension. This study aims to investigate the role of BBR in regulating microbial alterations and vascular function in hypertension. C57BL/6 J mice were infused with Ang II (0.8 mg/kg/day) via osmotic minipumps and treated with BBR (150 mg/kg/day) or choline (1%) for 4 weeks. Blood pressure was detected by tail-cuff measurement once a week. Abdominal aorta pulse wave velocity (PWV) and endothelium dependent vasodilatation were measured to evaluate vascular function. Vascular remodeling was assessed by histological staining of aortic tissue. The fecal microbiota was profiled using 16S ribosomal DNA (rDNA) sequencing. Plasma trimethylamine (TMA)/trimethylamine-N-oxide (TMAO) and hepatic FMO3 expression were measured. We found that BBR treatment significantly alleviated the elevated blood pressure, vascular dysfunction, and pathological remodeling in Ang II-induced hypertensive mice, while choline treatment aggravated hypertension-related vascular dysfunction. 16S rDNA gene sequencing results showed that BBR treatment altered gut microbiota composition (reduced the Firmicutes/Bacteroidetes (F/B) ratio and increased the abundances of Lactobacillus). Moreover, BBR inhibited FMO3 expression and plasma TMA/TMAO production in hypertensive mice. TMAO treatment increased the apoptosis and oxidative stress of human aortic endothelial cells (HAECs) and aggravated Ang II-induced HAECs dysfunction in vitro. These results indicate that the protective effect of BBR in hypertension might be attributed (at least partially) to the inhibition of TMAO production via regulating the gut microbiota.

Rhipidocotyle husi n. sp. and three known species of Bucephalidae Poche, 1907 from the East Asian Region: morphological and molecular data.

AbstractMorphological data and the first molecular data are provided for four species of the trematode family Bucephalidae Poche, 1907 from marine and freshwater teleost fish species of East Asia. A new species, Rhipidocotyle husi n. sp., was isolated from Huso dauricus from the Amur River, Russia. Adult worms of this species were distinguished from their congeners Rhipidocotyle illense and Rhipidocotyle kovalai by morphological analysis. Three other known species were identified: Bucephalus skrjabini and Prosorhynchus cf. squamatus were detected in Siniperca chuatsi from the Amur River and in Myoxocephalus spp. from the Okhotsk Sea, Russia, respectively, while Prosorynchoides karvei was extracted from Strongylura strongylura from Halong Bay, Vietnam. The 28S ribosomal DNA (rDNA)-based phylogenetic analysis showed that the new species formed a shared polytomy clade with Rhipidocotyle fennica. Phylogenetic analysis of all available molecular data showed that four genera, namely Rhipidocotyle, Bucephalus, Prosorynchoides and Prosorhynchus, are para- or polyphyletic. Molecular-based phylogenetic analysis of morphologically validated bucephalid species indicated that three genera – Rhipidocotyle, Bucephalus and Prosorynchoides – were monophyletic. The genus Prosorhynchus maintained paraphyly, and P. cf. squamatus was more closely related to Dollfustrema spp. than to other Prosorhynchus spp. These findings do not exclude the possibility that representatives of Dollfustrema and P. cf. squamatus belong to the same genus.

Alkaline Tolerance and Biofilm Formation of Root Canal Isolates of Enterococcus faecalis: An In Vitro Study

IntroductionThe aim of this study was to compare the effects of glucose and glycerol (provided as principal fermentable supplements) on alkaline tolerance and biofilm-forming capabilities of root canal–derived strains of Enterococcus faecalis and those from other environments. MethodsThe planktonic growth kinetics and the biofilm-forming capabilities of E. faecalis isolates (identified by 16S ribosomal DNA sequencing) were compared when supplied with glucose and glycerol at pH levels of 8 and 11 in a microtiter plate. The metabolic activity of the biofilms plate that formed at a neutral pH level (supplied with either glucose or glycerol) was measured after subsequent adjustment to a pH level of 11. ResultsTen isolates (7 from root canals and 3 from other sources) were examined. The lag phase and the doubling time increased under elevated alkalinity irrespective of either the fermentable supplement (glucose or glycerol) or the origin of the isolate. Biofilm formation and metabolic activity varied among strains, but neither was related to the source of isolation. In general, biofilm formation was enhanced when grown in glucose compared with glycerol and at a pH of 8 compared with a pH of 11 (irrespective of the fermentable supplement). The provision of glycerol did not increase either the planktonic growth rate or biofilm development compared with glucose but significantly increased the metabolic activity of biofilms, especially at a pH of 11 compared with a pH of 8. ConclusionsIn the nutrient-deprived environment of a necrotic or root-filled root canal, glycerol may be an alternative energy source that can promote increased metabolic activity of E. faecalis under alkaline treatment conditions.

Molecular screening for Sarcocystidae in muscles of wild birds from Brazil suggests a plethora of intermediate hosts for Sarcocystis falcatula.

The genus Sarcocystis and the species Toxoplasma gondii are the most prevalent sarcocystid organisms found in birds. Molecular phylogenies based on the first internal transcribed spacer of the ribosomal coding DNA (ITS1) have been widely used to identify them. Here, pectoral muscles from 400 wild birds from Brazil were screened by means of molecular methods using nested PCR, and Sanger sequencing yielded amplicons. A pan-sarcocystid ITS1-directed nested PCR revealed 28 birds infected by Sarcocystis falcatula (ten Piciformes, eight Psittaciformes, five Columbiformes, two Accipitriformes, one Anseriformes, one Passeriformes and one Strigiformes); one infected by Sarcocystis halieti (one Accipitriformes); nine infected by unknown or undescribed Sarcocystis (six Passeriformes, one Piciformes, one Cathartiformes and one Cuculiformes); and six harboring Toxoplasma gondii DNA (three Pelecaniformes, two Falconiformes and one Columbiformes). Samples harboring S. falcatula-related ITS1 sequences were further characterized by means of PCR and sequencing of genetic sequences of three surface antigen coding genes (SAGs). From this, 10 new allelic combinations of SAGs (SAG2, SAG3 and SAG4) were identified, in addition to 11 SAG allelic combinations already found in Brazil. Samples with S. falcatula-unrelated ITS1 sequences were further characterized by means of PCR and sequencing of cytochrome c oxidase subunit I coding sequences (CO1) and 18S ribosomal DNA gene (18S rDNA). This study was the first extensive survey of wild birds in Brazil for Sarcocystidae species. It provides the first molecular evidence of natural S. falcatula infection in 14 species, including in the order Piciformes, and shows the high genetic diversity of S. falcatula in intermediate hosts in South America. Evidence of occurrence of at least three non-described species of Sarcocystis was also presented in this study. This survey corroborated the ubiquity of T. gondii infection but revealed surprisingly low prevalence of this parasite (1.5%).

Sympatric occurrence of sibling Phytophthora species associated with foot rot disease of black pepper in India.

Foot rot disease caused by Phytophthora capsici is a serious threat to black pepper cultivation in India and globally. High diversity exists among the Phytophthora isolates of black pepper and hence detailed investigations of their morphology and phylogenetic taxonomy were carried out in the present study. In order to resolve the diversity, 182 isolates of Phytophthora, collected from different black pepper-growing tracts of South India during 1998-2013 and maintained in the National Repository of Phytophthora at ICAR-Indian Institute of Spices Research, Kozhikode, were subjected to morphological, molecular and phylogenetic characterization. Morphologically all the isolates were long pedicellate with umbellate/simple sympodial sporangiophores and papillate sporangia with l/b ranging from 1.63 to 2.55µm. Maximum temperature for the growth was ~ 34°C. Chlamydospores were observed in "tropicalis" group, whereas they were absent in "capsici" group. Initial molecular studies using internal transcribed spacer (ITS) marker gene showed two clear cut lineages-"capsici-like" and "tropicalis-like" groups among them. Representative isolates from each group were subjected to host differential test, multilocus sequence typing (MLST) and phylogeny studies. MLST analysis of seven nuclear genes (60S ribosomal protein L10, beta-tubulin, elongation factor 1 alpha, enolase, heat shock protein 90, 28S ribosomal DNA and TigA gene fusion protein) clearly delineated black pepper Phytophthora isolates into two distinct species-P. capsici and P. tropicalis. On comparing with type strains from ATCC, it was found that the type strains of P. capsici and P. tropicalis differed from black pepper isolates in their infectivity on black pepper. The high degree of genetic polymorphism observed in black pepper Phytophthora isolates is an indication of the selection pressure they are subjected to in the complex habitat which ultimately may lead to speciation. So based on the extensive analysis, it is unambiguously proved that the foot rot disease of black pepper in India is predominantly caused by two species of Phytophthora, viz. P. capsici and P. tropicalis. Presence of multiple species of Phytophthora in the black pepper agro-ecosystem warrants a revisit to the control strategy being adopted for managing this serious disease. The silent molecular evolution taking place in such an ecological niche needs to be critically studied for the sustainable management of foot rot disease.

Gut dysbiosis induced by antibiotics is improved by tangerine pith extract in mice

The gut microbiota can affect the intestinal barrier and other physiological functions. Tangerine pith contains various citrus pectins, which modulate the gut microbiota in a structure-dependent manner. We hypothesized that tangerine pith extract (TPE) has prebiotic effects on gut dysbiosis in antibiotic-treated mice. TPE was prepared via acid hydrolysis and isolated using an ultrafiltration method. Gut dysbiosis was induced through treatment with antibiotic cocktail for 7 days in mice. After spontaneous recovery or being supplemented with 0.5 mL TPE for 14 days, the gut microbiota composition and other parameters were examined. The 16s ribosomal DNA high-throughput sequencing and analysis confirmed that TPE positively improved the gut microbiota. The relative abundance of some probiotics such as [Ruminococcus]_torques_group, [Clostridium]_innocuum_group, and Enterorhabdus were significantly increased and harmful bacteria such as Alistipes were decreased in TPE-supplemented mice. Furthermore, the production of propionate and butyrate in the colonic contents were significantly enhanced in the TPE-treated mice. Serum D-lactic acid and diamine oxidase activity were increased in the antibiotic treatment group and reversed in the TPE-treated group. Moreover, protein expression of intestinal occludin was significantly increased by TPE. In conclusion, TPE is effective in restoring the gut microbiota in mice after antibiotic treatment and may act as a prebiotic in combatting gut dysbiosis.

Explore the Anti-Acne Mechanism of Licorice Flavonoids Based on Metabonomics and Microbiome.

Acne vulgaris is one of the most common inflammatory dermatoses in dermatological practice and can affect any gender or ethnic group. Although in previous studies, we had found that licorice flavonoids (LCF) play an anti-acne role by inhibiting PI3K-Akt signaling pathways and mitochondrial activity, the mechanism of LCF regulating skin metabolism, serum metabolism and skin microbes is still unclear. Here, we performed a full spectrum analysis of metabolites in the skin and serum using UHPLC-Triple TOF-MS. The results showed that LCF could treat acne by regulating the metabolic balance of amino acids, lipids and fatty acids in serum and skin. Similarly, we performed Illumina Hiseq sequencing of DNA from the skin microbes using 16S ribosomal DNA identification techniques. The results showed that LCF could treat acne by regulating the skin microbes to interfere with acne and make the microecology close to the normal skin state of rats. In summary, this study confirmed the anti-acne mechanism of LCF, namely by regulating metabolic balance and microbial balance. Therefore, this discovery will provide theoretical guidance for the preparation development and clinical application of the drug.

Gastrointestinal Autonomic Neuropathy Exacerbates Gut Microbiota Dysbiosis in Adult Patients With Type 2 Diabetes Mellitus.

ObjectiveThe diabetic autonomic neuropathy is one of the most common complications in type 2 diabetes mellitus (T2DM), especially gastrointestinal autonomic neuropathy (GAN), which occurs in up to 75% of patients. The study aimed to investigate the gut microbiota composition, structure, and function in T2DM patients with GAN (T2DM_GAN) and set up a link between gut microbiota and clinical characteristics of patients.MethodsDNA was extracted from fecal samples of three groups using the kit method: healthy volunteers (n = 19), the patients with T2DM (n = 76), and T2DM_GAN (n = 27). Sequencing of 16S ribosomal DNA was performed using the MiSeq platform.ResultsAccording to the clinical data, higher age, lower triglyceride, and lower body mass index were the main features of patients with T2DM_GAN. The gut microbiota analysis showed that Bacteroidetes, Firmicutes, and Proteobacteria constituted the three dominant phyla in healthy individuals. In addition, the gut microbiota structure and function of T2DM_GAN patients were clearly different from that of T2DM patients. T2DM patients were characterized by Fusobacteria, Fusobacteriia, Fusobacteriales, Fusobacteriaceae, Fusobacterium, Lachnoclostridium, and Fusobacterium_mortiferum. Those gut microbiota may be involved in carotenoid and flavonoid biosyntheses. Relatively, the Gammaproteobacteria, Enterobacteriales, Enterobacteriaceae, Escherichia-Shigella, Megasphaera, Escherichia_coli, and Megasphaera_elsdenii were characteristic in the T2DM_GAN patients. Those may be involved in bacterial invasion of epithelial cells and pathogenic Escherichia coli infection.ConclusionsGAN exacerbated gut microbiota dysbiosis in adult patients with T2DM. The findings indicated that phyla Fusobacteria and class Gammaproteobacteria were closely related to the occurrence of T2DM. Especially the latter may promote T2DM_GAN.

Molecular Characterization and Phylogenetic Analysis of Spirometra Tapeworms from Snakes in Hunan Province.

Sparganosis is a neglected zoonotic parasitic disease that poses huge threats to humans worldwide. Snakes play an important role in sparganosis transmission because they are the most common second intermediate hosts for Spirometra parasites. However, the population genetics of Spirometra isolates from snakes is currently not well studied in China. The present study was performed to explore the molecular characteristics and phylogenetic analysis of Spirometra tapeworms from different species of snakes in Hunan Province. This study obtained 49 Spirometra isolates from 15 geographical areas in Hunan Province, Central China. Subsequently, the 18S and 28S ribosomal DNA (rDNA) fragments were amplified from the isolated parasites, and their sequences were analyzed to assess their genetic diversity. Phylogenetic analyses were performed using the maximum likelihood algorithm. The results showed that sequence variations among these isolates were 0–2.3% and 0–0.1% for 18S and 28S rDNA, respectively. The phylogenetic analysis showed that all Spirometra isolates from Hunan Province were clustered into the same branch with Spirometra erinaceieuropaei isolated from other areas (China, Vietnam, Australia). Moreover, the phylogenetic trees revealed that Spirometra is closely related to Adenocephalus, Pyramicocephalus, Ligula, Dibothriocephalus, Schistocephalus, and Diphyllobothrium. The Spirometra isolates of different hosts/regions in Hunan Province are not host segregated or geographically isolated, and support for the taxonomic status of Spirometra tapeworms in China has been added. These results provide reference values for future accurate identification and taxonomic status of Spirometra tapeworms in China.

Lung microbiota features of stage III and IV non-small cell lung cancer patients without lung infection.

BackgroundThe microbial community affects the occurrence, development, metastasis and treatment response of cancers. But the detailed role and characteristics of lung microbiota (LM) in non-small cell lung cancer (NSCLC) are not fully known. For NSCLC associated microbiota analysis, it is valuable to combine multiple levels of detection, e.g., tumor, blood plasma, and bronchoalveolar fluid (BALF), but not single tissues.MethodsThis study collected above three sample types from NSCLC patients free from lung infection and aimed to describe their LM features using sequencing techniques. All patients diagnosed at the Department of Oncology in Shijiazhuang People’s Hospital with stage III or IV NSCLC from May 2019 to April 2020 were enrolled. All 37 pieces of tumor tissues and 6 blood samples were sent for pathogen targeted sequencing; for the BALF samples, 4 were used for pathogen targeted sequencing and 2 were sent for 16S ribosomal DNA (rDNA) sequencing.ResultsWe detected 49 pathogenic microorganisms (PMs) in the 37 tumor samples, 28 PMs in the 4 BALF samples, and 14 PMs in the 6 plasma samples. Overall, there were 5 common PMs in 3 types of samples. Between the tumor and BALF samples, there were another 11 common elements. In the 5 tumor-plasma pairs, the presence of a specific PM in blood was not necessarily consistent with that in the tumor. In the tumor-BALF pairs, the PM diversity was dramatically higher in the BALF than in the tumor. The PMs detected in the BALF could largely cover the PMs in the tumor. In the BALF 16S rDNA sequencing, there were 82 common operational taxonomic units (OTUs), and the microbiota in the BALF of advanced NSCLC patients exhibited some similarity.ConclusionsThis study showed the unique features of LM. The amount of intra-tumoral PMs was not necessarily consistent with that in the blood, but there was an obvious correlation between the intra-tumoral microbiota and that in the BALF. It is convenient and non-invasive to obtain BALF. Detection of LM classification and abundance in the BALF may help evaluate the severity of NSCLC.

Call me by my name: unravelling the taxonomy of the gulper shark genus Centrophorus in the Mediterranean Sea through an integrated taxonomic approach

Abstract The current shift of fishery efforts towards the deep sea is raising concern about the vulnerability of deep-water sharks, which are often poorly studied and characterized by problematic taxonomy. For instance, in the Mediterranean Sea the taxonomy of genus Centrophorus has not been clearly unravelled yet. Since proper identification of the species is fundamental for their correct assessment and management, this study aims at clarifying the taxonomy of this genus in the Mediterranean Basin through an integrated taxonomic approach. We analysed a total of 281 gulper sharks (Centrophorus spp.) collected from various Mediterranean, Atlantic and Indian Ocean waters. Molecular data obtained from cytochrome c oxidase subunit I (COI), 16S ribosomal RNA (16S), NADH dehydrogenase subunit 2 (ND2) and a portion of a nuclear 28S ribosomal DNA gene region (28S) have highlighted the presence of a unique mitochondrial clade in the Mediterranean Sea. The morphometric results confirmed these findings, supporting the presence of a unique and distinct morphological group comprising all Mediterranean individuals. The data strongly indicate the occurrence of a single Centrophorus species in the Mediterranean, ascribable to C. cf. uyato, and suggest the need for a revision of the systematics of the genus in the area.

Fine structure and transcription dynamics of bread wheat ribosomal DNA loci deciphered by a multi‐omics approach

Three out of four RNA components of ribosomes are encoded by 45S ribosomal DNA (rDNA) loci, which are organized as long head-to-tail tandem arrays of nearly identical units, spanning several megabases of sequence. Due to this structure, the rDNA loci are the major sources of gaps in genome assemblies, and gene copy number, sequence composition, and expression status of particular arrays remain elusive, especially in complex genomes harboring multiple loci. Here we conducted a multi-omics study to decipher the 45S rDNA loci in hexaploid bread wheat. Coupling chromosomal genomics with optical mapping, we reconstructed individual rDNA arrays, enabling locus-specific analyses of transcription activity and methylation status from RNA- and bisulfite-sequencing data. We estimated a total of 6,650 rDNA units in the bread wheat genome, with approximately 2,321, 3,910, 253, and 50 gene copies located in short arms of chromosomes 1B, 6B, 5D, and 1A, respectively. Only 1B and 6B loci contributed substantially to rRNA transcription at a roughly 2:1 ratio. The ratio varied among five tissues analyzed (embryo, coleoptile, root tip, primary leaf, mature leaf), being the highest (2.64:1) in mature leaf and lowest (1.72:1) in coleoptile. Cytosine methylation was considerably higher in CHG context in the silenced 5D locus as compared with the active 1B and 6B loci. In conclusion, a fine genomic organization and tissue-specific expression of rDNA loci were deciphered, for the first time, in a complex polyploid species. The results are discussed in the context of wheat evolution and transcription regulation.

Natural Herbal Remedy Wumei Decoction Ameliorates Intestinal Mucosal Inflammation by Inhibiting Th1/Th17 Cell Differentiation and Maintaining Microbial Homeostasis.

Evidence has shown that the traditional Chinese herbal medicine Wumei decoction (WMD) has a protective effect on ulcerative colitis. Here, we studied the anti-inflammatory effects and potential mechanisms of WMD on chronic colitis in mice. A dextran sulfate sodium (DSS)-induced chronic colitis model and CD45RBhighCD4+ T cell transfer model were established in mice. Body weight, Disease Activity Index, and colon length were assessed, and histopathology was confirmed by hematoxylin and eosin staining. Colon tissue samples were collected to detect the frequencies of various immune cells, expression of cytokines, and tight junction-related proteins using flow cytometry, quantitative real-time polymerase chain reaction, and enzyme-linked immunosorbent assay, respectively. 16S ribosomal DNA sequencing was performed to distinguish differential microbiota of fecal samples. Severe chronic colitis was observed in mice after DSS exposure and in Rag1-/- mice reconstituted with CD45RBhighCD4+ T cells, as manifested by weight loss, hematochezia, and shortening and thickening of the colon, which were reversed by WMD treatment. WMD markedly suppressed intestinal mucosal CD4+ T cell differentiation and the secretion of proinflammatory cytokines (eg, tumor necrosis factor α, interleukin-1β, interferon γ, and IL-17A) by flow cytometry, quantitative real-time polymerase chain reaction, and enzyme-linked immunosorbent assay, respectively. Moreover, WMD promoted the expression of occludin, zonula occludens-1, and E-cadherin, thereby maintaining the epithelial barrier function. Additionally, 16S ribosomal DNA sequencing revealed that WMD regulated the dysbiosis of gut microbiota in CD45RBhighCD4+ T cell-reconstituted Rag1-/- mice, evidenced by an increase of Allobaculum and Bacteroides and a decrease of Ileibacterium. WMD ameliorates chronic colitis in mice induced by DSS or reconstituted with CD45RBhighCD4+ T cells through suppressing Th1/Th17 cell differentiation and the secretion of proinflammatory cytokines, maintaining epithelial barrier function, and improving the dysbiosis.

Supplementary studies on Myxobolus talievi Dogiel, 1957 (Cnidaria, Myxozoa) from the skeletal muscle of the cottoid fish Cyphocottus eurystomus in Lake Baikal: Morphological, histological and molecular characterisations.

Myxobolus talievi Dogiel, 1957 was originally described from the eyes, skeletal muscles and body cavity of endemic cottoid fish from Lake Baikal. In the present study, we supply new information on the myxospore morphology and histopathology of M. talievi; furthermore, we complete the original species description by Dogiel and Bogolepova (1957) with 18S ribosomal DNA (rDNA) sequence data. Histopathological analysis showed that the plasmodium was encapsulated by a thin layer of connective tissue and located in the intermuscular connective tissue among muscle cells. No inflammation was observed. Phylogenetic analysis revealed that M. talievi clustered with Myxobolus sp. 2 (NCBI Acc. No. U13830), an unidentified Myxobolus species from cottoid fish studied by Smothers et al. (1994), and located in the sister clade of Myxobolus spp. developing spores in the nerves of salmonids.

Molecular Survey and Spatial Distribution of Rickettsia spp. in Ticks Infesting Free-Ranging Wild Animals in Pakistan (2017-2021).

Rickettsia spp. associated with ticks infesting wild animals have been mostly neglected in several countries, including Pakistan. To address this knowledge gap, ticks were collected during 2017 to 2021 from wild animals including cats (Felis chaus), Indian hedgehogs (Paraechinus micropus), and wild boars (Sus scrofa). The collected ticks were morpho-molecularly identified and screened for the detection of Rickettsia spp. Morphologically identified ticks were categorized into four species of the genus Rhipicephalus: Rhipicephalus haemaphysaloides, Rh. turanicus, Rh. sanguineus sensu lato (s.l), and Rh. microplus. Among 53 wild animals examined, 31 were infested by 531 ticks, an overall prevalence of 58.4%. Adult female ticks were predominant (242 out of 513 ticks collected, corresponding to 46%) in comparison with males (172, 32%), nymphs (80, 15%) and larvae (37, 7%). The most prevalent tick species was Rh. turanicus (266, 50%), followed by Rh. microplus (123, 23%), Rh. sanguineus (106, 20%), and Rh. haemaphysaloides (36, 7%). Among the screened wild animals, wild boars were the most highly infested, with 268 ticks being collected from these animals (50.4%), followed by cats (145, 27.3%) and hedgehogs (118, 22.3%). Tick species Rh. haemaphysaloides, Rh. turanicus, and Rh. sanguineus were found on wild boars, Rh. haemaphysaloides, and Rh. microplus on cats, and Rh. turanicus on hedgehogs. In a phylogenetic analysis, mitochondrial cytochrome C oxidase 1 (cox1) sequences obtained from a subsample (120) of the collected ticks clustered with sequences from Bangladesh, China, India, Iran, Myanmar, and Pakistan, while 16S ribosomal DNA (16S rDNA) sequences clustered with sequences reported from Afghanistan, Egypt, India, Pakistan, Romania, Serbia, and Taiwan. Among Rickettsia infected ticks (10/120, 8.3%), Rh. turanicus (7/10, 70%), and Rh. haemaphysaloides (3/10, 30%) were found infesting wild boars in the districts Mardan and Charsadda. The obtained rickettsial gltA gene sequences showed 99% and ompA gene sequences showed 100% identity with Rickettsia massiliae, and the phylogenetic tree shows ompA clustered with the same species reported from France, Greece, Spain, and USA. This study emphasizes the need for effective surveillance and control programs in the region to prevent health risks due to tick-borne pathogens, and that healthy infested wild animals may play a role in the spread of these parasites.

Prevalence of Antibiotic Resistant Bacteria in Ready to eat Food Sold by Street Vendors in Bangladesh

Bangladesh has a vibrant street food culture, with easily available ready-to-eat (RTE) foods. In this study, a total of 221 isolates were obtained from 141 Bangladeshi RTE food samples, such as fried, non-fried, and cooked foods; egg-, milk-, cereal-, and cream-based foods; pickles/achar; fruit; and RTE leaves, through culture on trypticase soy (for aerobic plate count), mannitol salt (MS for Staphylococcaceae), deoxycholate-hydrogen sulfide-lactose (DHL, for Enterobacteriaceae), and NaCl glycine Kim Goepfert agar (NGKG, for Bacillus cereus-like bacteria) agar plates. The aerobic plate counts ranged from undetectable to 8.5 log CFU/g. After enrichment with peptone water, contaminated bacteria detected on DHL, MS, and NGKG agar plates, from 77, 110, and 77 samples, respectively. Twenty out of 111 isolates on DHL agar, and 17 of 32 isolates on NGKG agar clearly showed resistance against three or more drugs. Through 16S ribosomal DNA sequencing analysis, six selected isolates from DHL agar were identified as Pseudomonas nitroreducens, Citrobacter braakii, Klebsiella pneumoniae subsp. pneumoniae, and Serratia marcescens. One selected isolate from NGKG agar was identified as Bacillus cereus-like bacteria. The results suggest that additional safety measures and regulations are necessary to ensure the quality and safety of the RTE foods in Bangladesh.