RESUMO
We present evidence for multiple independent origins of recombinant SARS-CoV-2 viruses sampled from late 2020 and early 2021 in the United Kingdom. Their genomes carry single-nucleotide polymorphisms and deletions that are characteristic of the B.1.1.7 variant of concern but lack the full complement of lineage-defining mutations. Instead, the remainder of their genomes share contiguous genetic variation with non-B.1.1.7 viruses circulating in the same geographic area at the same time as the recombinants. In four instances, there was evidence for onward transmission of a recombinant-origin virus, including one transmission cluster of 45 sequenced cases over the course of 2 months. The inferred genomic locations of recombination breakpoints suggest that every community-transmitted recombinant virus inherited its spike region from a B.1.1.7 parental virus, consistent with a transmission advantage for B.1.1.7's set of mutations.
Assuntos
COVID-19/epidemiologia , COVID-19/transmissão , Pandemias , Recombinação Genética , SARS-CoV-2/genética , Sequência de Bases/genética , COVID-19/virologia , Biologia Computacional/métodos , Frequência do Gene , Genoma Viral , Genótipo , Humanos , Mutação , Filogenia , Polimorfismo de Nucleotídeo Único , Reino Unido/epidemiologia , Sequenciamento Completo do Genoma/métodosRESUMO
BACKGROUND: Trombiculid mites are globally distributed, highly diverse arachnids that largely lack molecular resources such as whole mitogenomes for the elucidation of taxonomic relationships. Trombiculid larvae (chiggers) parasitise vertebrates and can transmit bacteria (Orientia spp.) responsible for scrub typhus, a zoonotic febrile illness. Orientia tsutsugamushi causes most cases of scrub typhus and is endemic to the Asia-Pacific Region, where it is transmitted by Leptotrombidium spp. chiggers. However, in Dubai, Candidatus Orientia chuto was isolated from a case of scrub typhus and is also known to circulate among rodents in Saudi Arabia and Kenya, although its vectors remain poorly defined. In addition to Orientia, chiggers are often infected with other potential pathogens or arthropod-specific endosymbionts, but their significance for trombiculid biology and public health is unclear. RESULTS: Ten chigger species were collected from rodents in southwestern Saudi Arabia. Chiggers were pooled according to species and screened for Orientia DNA by PCR. Two species (Microtrombicula muhaylensis and Pentidionis agamae) produced positive results for the htrA gene, although Ca. Orientia chuto DNA was confirmed by Sanger sequencing only in P. agamae. Metagenomic sequencing of three pools of P. agamae provided evidence for two other bacterial associates: a spirochaete and a Wolbachia symbiont. Phylogenetic analysis of 16S rRNA and multi-locus sequence typing genes placed the spirochaete in a clade of micromammal-associated Borrelia spp. that are widely-distributed globally with no known vector. For the Wolbachia symbiont, a genome assembly was obtained that allowed phylogenetic localisation in a novel, divergent clade. Cytochrome c oxidase I (COI) barcodes for Saudi Arabian chiggers enabled comparisons with global chigger diversity, revealing several cases of discordance with classical taxonomy. Complete mitogenome assemblies were obtained for the three P. agamae pools and almost 50 SNPs were identified, despite a common geographic origin. CONCLUSIONS: P. agamae was identified as a potential vector of Ca. Orientia chuto on the Arabian Peninsula. The detection of an unusual Borrelia sp. and a divergent Wolbachia symbiont in P. agamae indicated links with chigger microbiomes in other parts of the world, while COI barcoding and mitogenomic analyses greatly extended our understanding of inter- and intraspecific relationships in trombiculid mites.
Assuntos
Borrelia , Microbiota , Orientia tsutsugamushi , Tifo por Ácaros , Trombiculidae , Wolbachia , Animais , Borrelia/genética , DNA , Tipagem de Sequências Multilocus , Orientia , Orientia tsutsugamushi/genética , Filogenia , RNA Ribossômico 16S/genética , Roedores/genética , Arábia Saudita , Tifo por Ácaros/epidemiologia , Tifo por Ácaros/microbiologia , Trombiculidae/genética , Trombiculidae/microbiologia , Wolbachia/genéticaRESUMO
A 2022 canine gastroenteritis outbreak in the United Kingdom was associated with circulation of a new canine enteric coronavirus closely related to a 2020 variant with an additional spike gene recombination. The variants are unrelated to canine enteric coronavirus-like viruses associated with human disease but represent a model for coronavirus population adaptation.
Assuntos
Infecções por Coronavirus , Surtos de Doenças , Doenças do Cão , Gastroenterite , Filogenia , Animais , Cães , Surtos de Doenças/veterinária , Doenças do Cão/virologia , Doenças do Cão/epidemiologia , Reino Unido/epidemiologia , Gastroenterite/virologia , Gastroenterite/epidemiologia , Gastroenterite/veterinária , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/virologia , Coronavirus Canino/genética , Coronavirus Canino/classificação , Humanos , Glicoproteína da Espícula de Coronavírus/genéticaRESUMO
BACKGROUND: Maternal breastmilk is a source of pre- and pro-biotics that impact neonatal gut microbiota colonization. Because oral rotavirus vaccines (ORVs) are administered at a time when infants are often breastfed, breastmilk microbiota composition may have a direct or indirect influence on vaccine take and immunogenicity. METHODS: Using standardized methods across sites, we compared breastmilk microbiota composition in relation to geographic location and ORV response in cohorts prospectively followed from birth to 18 weeks of age in India (n = 307), Malawi (n = 119), and the United Kingdom ([UK] n = 60). RESULTS: Breastmilk microbiota diversity was higher in India and Malawi than the UK across 3 longitudinal samples spanning weeks of life 1 to 13. Dominant taxa such as Streptococcus and Staphylococcus were consistent across cohorts; however, significant geographic differences were observed in the prevalence and abundance of common and rare genera throughout follow up. No consistent associations were identified between breastmilk microbiota composition and ORV outcomes including seroconversion, vaccine shedding after dose 1, and postvaccination rotavirus-specific immunoglobulin A level. CONCLUSIONS: Our findings suggest that breastmilk microbiota composition may not be a key factor in shaping trends in ORV response within or between countries.
Assuntos
Microbiota , Infecções por Rotavirus , Vacinas contra Rotavirus , Rotavirus , Recém-Nascido , Feminino , Humanos , Lactente , Leite Humano , Infecções por Rotavirus/epidemiologia , Infecções por Rotavirus/prevenção & controle , Estudos Prospectivos , Anticorpos Antivirais , Imunoglobulina A , Vacinas AtenuadasRESUMO
Here, we report the first complete genomes of three cultivable treponeme species from bovine digital dermatitis (DD) skin lesions, two comparative human treponemes, considered indistinguishable from bovine DD species, and a bovine gastrointestinal (GI) treponeme isolate. Key genomic differences between bovine and human treponemes implicate microbial mechanisms that enhance knowledge of how DD, a severe disease of ruminants, has emerged into a prolific, worldwide disease. Bovine DD treponemes have additional oxidative stress genes compared to nearest human-isolated relatives, suggesting better oxidative stress tolerance, and potentially explaining how bovine strains can colonize skin surfaces. Comparison of both bovine DD and GI treponemes as well as bovine pathogenic and human non-pathogenic saprophyte Treponema phagedenis strains indicates genes encoding a five-enzyme biosynthetic pathway for production of 2,3-diacetamido-2,3-dideoxy-d-mannuronic acid, a rare di-N-acetylated mannuronic acid sugar, as important for pathogenesis. Bovine T. phagedenis strains further differed from human strains by having unique genetic clusters including components of a type IV secretion system and a phosphate utilisation system including phoU, a gene associated with osmotic stress survival. Proteomic analyses confirmed bovine derived T. phagedenis exhibits expression of PhoU but not the putative secretion system, whilst the novel mannuronic acid pathway was expressed in near entirety across the DD treponemes. Analysis of osmotic stress response in water identified a difference between bovine and human T. phagedenis with bovine strains exhibiting enhanced survival. This novel mechanism could enable a selective advantage, allowing environmental persistence and transmission of bovine T. phagedenis. Finally, we investigated putative outer membrane protein (OMP) ortholog families across the DD treponemes and identified several families as multi-specific adhesins capable of binding extra cellular matrix (ECM) components. One bovine pathogen specific adhesin ortholog family showed considerable serodiagnostic potential with the Treponema medium representative demonstrating considerable disease specificity (91.6%). This work has shed light on treponeme host adaptation and has identified candidate molecules for future diagnostics, vaccination and therapeutic intervention.
Assuntos
Treponema/genética , Infecções por Treponema/genética , Animais , Bovinos , DNA Bacteriano , Dermatite Digital/microbiologia , Humanos , FilogeniaRESUMO
The immunogenicity and effectiveness of oral rotavirus vaccines (ORVs) against severe rotavirus-associated gastroenteritis are impaired in low- and middle-income countries (LMICs) where the burden of disease is highest. Determining risk factors for impaired ORV response may help identify strategies to enhance vaccine effectiveness. In this study, we use metagenomic sequencing to provide a high-resolution taxonomic analysis of stool samples collected at 6 weeks of age (coinciding with the first ORV dose) during a prospective study of ORV immunogenicity in India and Malawi. We then analyse the functional capacity of the developing microbiome in these cohorts. Microbiome composition differed significantly between countries, although functional capacity was more similar than taxonomic composition. Our results confirm previously reported findings that the developing microbiome is more diverse in taxonomic composition in ORV non-seroconverters compared with seroconverters, and we additionally demonstrate a similar pattern in functional capacity. Although taxonomic or functional feature abundances are poor predictors of ORV response, we show that skews in the direction of associations within these microbiome data can be used to identify consistent markers of ORV response across LMIC infant cohorts. We also highlight the systemic under-representation of reference genes from LMICs that limit functional annotation in our study (7% and 13% annotation at pathway and enzyme commission level, respectively). Overall, higher microbiome diversity in early life may act as marker for impaired ORV response in India and Malawi, whilst a holistic perspective of functional capacity may be hidden in the "dark matter" of the microbiome.
Assuntos
Infecções por Rotavirus , Vacinas contra Rotavirus , Rotavirus , Humanos , Lactente , Rotavirus/genética , Malaui , Estudos Prospectivos , Imunogenicidade da Vacina , Infecções por Rotavirus/prevenção & controle , Índia , Vacinas Atenuadas , Anticorpos AntiviraisRESUMO
Discovering the mutational events that fuel adaptation to environmental change remains an important challenge for evolutionary biology. The classroom example of a visible evolutionary response is industrial melanism in the peppered moth (Biston betularia): the replacement, during the Industrial Revolution, of the common pale typica form by a previously unknown black (carbonaria) form, driven by the interaction between bird predation and coal pollution. The carbonaria locus has been coarsely localized to a 200-kilobase region, but the specific identity and nature of the sequence difference controlling the carbonaria-typica polymorphism, and the gene it influences, are unknown. Here we show that the mutation event giving rise to industrial melanism in Britain was the insertion of a large, tandemly repeated, transposable element into the first intron of the gene cortex. Statistical inference based on the distribution of recombined carbonaria haplotypes indicates that this transposition event occurred around 1819, consistent with the historical record. We have begun to dissect the mode of action of the carbonaria transposable element by showing that it increases the abundance of a cortex transcript, the protein product of which plays an important role in cell-cycle regulation, during early wing disc development. Our findings fill a substantial knowledge gap in the iconic example of microevolutionary change, adding a further layer of insight into the mechanism of adaptation in response to natural selection. The discovery that the mutation itself is a transposable element will stimulate further debate about the importance of 'jumping genes' as a source of major phenotypic novelty.
Assuntos
Elementos de DNA Transponíveis/genética , Mariposas/genética , Mariposas/fisiologia , Mutação/genética , Pigmentação/genética , Asas de Animais/fisiologia , Adaptação Fisiológica/genética , Alelos , Animais , Evolução Biológica , Ciclo Celular/genética , Cor , Genes de Insetos/genética , Haplótipos/genética , Íntrons/genética , Masculino , Melanose/genética , Melanose/veterinária , Mariposas/citologia , Mutagênese Insercional/genética , Fenótipo , Pigmentação/fisiologia , Seleção Genética/genética , Reino Unido , Asas de Animais/crescimento & desenvolvimentoRESUMO
BACKGROUND: Metronidazole is the first-line treatment for bacterial vaginosis, but cure rates are suboptimal and recurrence rates high. OBJECTIVES: To evaluate the impact of a standard course of oral metronidazole treatment (500 mg twice per day for 7 days) on the vaginal microbiota of Rwandan bacterial vaginosis patients using microscopy and 16S rRNA gene sequencing, and to evaluate correlates of treatment failure. STUDY DESIGN: HIV-negative, nonpregnant women aged 18-45 years with bacterial vaginosis and/or Trichomonas vaginalis (N=68) were interviewed and sampled before and after metronidazole treatment. They were also screened, and treated if applicable, for other urogenital infections. The vaginal microbiota was assessed by Gram stain Nugent scoring, Illumina 16S rRNA HiSeq sequencing (relative abundances), and BactQuant 16S gene quantitative polymerase chain reaction (estimated concentrations). Only women with a pretreatment Nugent score of 7-10 and a valid posttreatment Nugent score (N=55) were included in metronidazole treatment failure analyses, with treatment failure defined as a posttreatment Nugent score of 4-10. RESULTS: The bacterial vaginosis cure rate by Nugent scoring was 54.5%. The mean total vaginal bacterial concentration declined from 6.59 to 5.85 log10/µL (P<.001), which was mostly due to a reduction in mean bacterial vaginosis-associated anaerobes concentration (all bacterial vaginosis-associated anaerobe taxa combined) from 6.23 to 4.55 log10/µL (P<.001). However, only 16.4% of women had a bacterial vaginosis anaerobes concentration reduction of more than 50%, and only 3 women had complete eradication. The mean concentration of lactobacilli (all species combined) increased from 4.98 to 5.56 log10/µL (P=.017), with L. iners being the most common species pre- and posttreatment. The mean concentration of pathobionts (defined as Proteobacteria, streptococci, staphylococci, enterococci, and a few others) did not change significantly: from 1.92 log10/µL pretreatment to 2.01 log10/µL posttreatment (P=.939). Pretreatment pathobionts concentration, and having a pretreatment vaginal microbiota type containing more than 50% Gardnerella vaginalis (compared with less than 50%), were associated with increased likelihood of treatment failure, but the latter did not reach statistical significance (P=.044 and P=.084, respectively). CONCLUSIONS: Metronidazole alone may not cure women with high G. vaginalis relative abundance, potentially due to biofilm presence, and women with high pathobionts concentration. These women may benefit from additional biofilm-disrupting and/or pathobiont-targeting treatments.
Assuntos
Anti-Infecciosos/uso terapêutico , Metronidazol/uso terapêutico , Vaginite por Trichomonas/tratamento farmacológico , Vaginose Bacteriana/tratamento farmacológico , Adulto , Bactérias Anaeróbias , Biofilmes , Enterococcus , Feminino , Gardnerella vaginalis , Humanos , Lactobacillus , Proteobactérias , RNA Ribossômico 16S/genética , Ruanda , Staphylococcus , Streptococcus , Falha de Tratamento , Resultado do Tratamento , Vaginite por Trichomonas/complicações , Vaginite por Trichomonas/microbiologia , Vaginose Bacteriana/complicações , Vaginose Bacteriana/microbiologia , Adulto JovemRESUMO
Equine grass sickness (EGS) is a frequently fatal disease of horses, responsible for the death of 1 to 2% of the U.K. horse population annually. The etiology of this disease is currently uncharacterized, although there is evidence it is associated with Clostridium botulinum neurotoxin in the gut. Prevention is currently not possible, and ileal biopsy diagnosis is invasive. The aim of this study was to characterize the fecal microbiota and biofluid metabolic profiles of EGS horses, to further understand the mechanisms underlying this disease, and to identify metabolic biomarkers to aid in diagnosis. Urine, plasma, and feces were collected from horses with EGS, matched controls, and hospital controls. Sequencing the16S rRNA gene of the fecal bacterial population of the study horses found a severe dysbiosis in EGS horses, with an increase in Bacteroidetes and a decrease in Firmicutes bacteria. Metabolic profiling by 1H nuclear magnetic resonance spectroscopy found EGS to be associated with the lower urinary excretion of hippurate and 4-cresyl sulfate and higher excretion of O-acetyl carnitine and trimethylamine-N-oxide. The predictive ability of the complete urinary metabolic signature and using the four discriminatory urinary metabolites to classify horses by disease status was assessed using a second (test) set of horses. The urinary metabolome and a combination of the four candidate biomarkers showed promise in aiding the identification of horses with EGS. Characterization of the metabolic shifts associated with EGS offers the potential of a noninvasive test to aid premortem diagnosis.
Assuntos
Acetilcarnitina/urina , Cresóis/urina , Disbiose/diagnóstico , Hipuratos/urina , Doenças dos Cavalos/diagnóstico , Metilaminas/urina , Ésteres do Ácido Sulfúrico/urina , Acetilcarnitina/sangue , Animais , Bacteroidetes/classificação , Bacteroidetes/isolamento & purificação , Biomarcadores/sangue , Biomarcadores/urina , Clostridium botulinum/metabolismo , Clostridium botulinum/patogenicidade , Cresóis/sangue , Disbiose/sangue , Disbiose/microbiologia , Disbiose/urina , Fezes/microbiologia , Firmicutes/classificação , Firmicutes/isolamento & purificação , Microbioma Gastrointestinal , Hipuratos/sangue , Doenças dos Cavalos/sangue , Doenças dos Cavalos/microbiologia , Doenças dos Cavalos/urina , Cavalos , Espectroscopia de Ressonância Magnética , Metilaminas/sangue , RNA Ribossômico 16S/genética , Ésteres do Ácido Sulfúrico/sangueRESUMO
Symbiotic associations of cellulolytic eukaryotic protists and diverse bacteria are common in the gut microbial communities of termites. Besides cellulose degradation by the gut protists, reductive acetogenesis from H2 plus CO2 and nitrogen fixation by gut bacteria play crucial roles in the host termites' nutrition by contributing to the energy demand of termites and supplying nitrogen poor in their diet, respectively. Fractionation of these activities and the identification of key genes from the gut community of the wood-feeding termite Hodotermopsis sjoestedti revealed that substantial activities in the gut--nearly 60% of reductive acetogenesis and almost exclusively for nitrogen fixation--were uniquely attributed to the endosymbiotic bacteria of the cellulolytic protist in the genus Eucomonympha. The rod-shaped endosymbionts were surprisingly identified as a spirochete species in the genus Treponema, which usually exhibits a characteristic spiral morphology. The endosymbionts likely use H2 produced by the protist for these dual functions. Although H2 is known to inhibit nitrogen fixation in some bacteria, it seemed to rather stimulate this important mutualistic process. In addition, the single-cell genome analyses revealed the endosymbiont's potentials of the utilization of sugars for its energy requirement, and of the biosynthesis of valuable nutrients such as amino acids from the fixed nitrogen. These metabolic interactions are suitable for the dual functions of the endosymbiont and reconcile its substantial contributions in the gut.
Assuntos
Ácido Acético/metabolismo , Isópteros/fisiologia , Fixação de Nitrogênio , Spirochaetales/metabolismo , Simbiose , Animais , Dióxido de Carbono/metabolismo , Evolução Molecular , Genoma , Intestinos/microbiologia , Isópteros/microbiologia , Dados de Sequência Molecular , Nitrogênio/química , Filogenia , RNA Ribossômico 16S/metabolismoRESUMO
A mycoplasma isolated from the liver of a dead Humboldt penguin (Spheniscus humboldti) and designated strain 56A97T, was investigated to determine its taxonomic status. Complete 16S rRNA gene sequence analysis indicated that the organism was most closely related to Mycoplasma gallisepticum and Mycoplasma imitans(99.7 and 99.9â% similarity, respectively). The average DNA-DNA hybridization values between strain 56A97T and M. gallisepticum and M. imitans were 39.5 and 30â%, respectively and the Genome to Genome Distance Calculator gave results of 29.10 and 23.50â%, respectively. The 16S-23S rRNA intergenic spacer was 72-73â% similar to M. gallisepticum strains and 52.2â% to M. imitans. A partial sequence of rpoB was 91.1-92â% similar to M. gallisepticum strains and 84.7â% to M. imitans. Colonies possessed a typical fried-egg appearance and electron micrographs revealed the lack of a cell wall and a nearly spherical morphology, with an electron-dense tip-like structure on some flask-shaped cells. The isolate required sterol for growth, fermented glucose, adsorbed and haemolysed erythrocytes, but did not hydrolyse arginine or urea. The strain was compared serologically against 110 previously described Mycoplasma reference strains, showing that, except for M. gallisepticum, strain 56A97T is not related to any of the previously described species, although weak cross-reactions were evident. Genomic information, serological reactions and phenotypic properties demonstrate that this organism represents a novel species of the genus Mycoplasma, for which the name Mycoplasma tullyi sp. nov. is proposed; the type strain is 56A97T (ATCC BAA-1432T, DSM 21909T, NCTC 11747T).
Assuntos
Fígado/microbiologia , Mycoplasma/classificação , Filogenia , Spheniscidae/microbiologia , Animais , Técnicas de Tipagem Bacteriana , DNA Bacteriano/genética , Mycoplasma/genética , Mycoplasma/isolamento & purificação , Hibridização de Ácido Nucleico , RNA Ribossômico 16S/genética , Análise de Sequência de DNARESUMO
BACKGROUND: In the last 5 years, the rapid pace of innovations and improvements in sequencing technologies has completely changed the landscape of metagenomic and metagenetic experiments. Therefore, it is critical to benchmark the various methodologies for interrogating the composition of microbial communities, so that we can assess their strengths and limitations. The most common phylogenetic marker for microbial community diversity studies is the 16S ribosomal RNA gene and in the last 10 years the field has moved from sequencing a small number of amplicons and samples to more complex studies where thousands of samples and multiple different gene regions are interrogated. RESULTS: We assembled 2 synthetic communities with an even (EM) and uneven (UM) distribution of archaeal and bacterial strains and species, as metagenomic control material, to assess performance of different experimental strategies. The 2 synthetic communities were used in this study, to highlight the limitations and the advantages of the leading sequencing platforms: MiSeq (Illumina), The Pacific Biosciences RSII, 454 GS-FLX/+ (Roche), and IonTorrent (Life Technologies). We describe an extensive survey based on synthetic communities using 3 experimental designs (fusion primers, universal tailed tag, ligated adaptors) across the 9 hypervariable 16S rDNA regions. We demonstrate that library preparation methodology can affect data interpretation due to different error and chimera rates generated during the procedure. The observed community composition was always biased, to a degree that depended on the platform, sequenced region and primer choice. However, crucially, our analysis suggests that 16S rRNA sequencing is still quantitative, in that relative changes in abundance of taxa between samples can be recovered, despite these biases. CONCLUSION: We have assessed a range of experimental conditions across several next generation sequencing platforms using the most up-to-date configurations. We propose that the choice of sequencing platform and experimental design needs to be taken into consideration in the early stage of a project by running a small trial consisting of several hypervariable regions to quantify the discriminatory power of each region. We also suggest that the use of a synthetic community as a positive control would be beneficial to identify the potential biases and procedural drawbacks that may lead to data misinterpretation. The results of this study will serve as a guideline for making decisions on which experimental condition and sequencing platform to consider to achieve the best microbial profiling.
Assuntos
Genoma Bacteriano , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Metagenômica , RNA Ribossômico 16S/genética , Bactérias/genética , Benchmarking , Filogenia , RNA Ribossômico 16S/classificaçãoRESUMO
Disruption in stable establishment of commensal gut microbiota by early weaning is an important factor in susceptibility of young animals to enteric disorders. The artificial sweetener SUCRAM [consisting of neohesperidin dihydrochalcone (NHDC) and saccharin] included in piglets' feed reduces incidence of enteric disease. Pyrosequencing of pig caecal 16S rRNA gene amplicons identified 25 major families encompassing seven bacterial classes with Bacteroidia, Clostridia and Bacilli dominating the microbiota. There were significant shifts in microbial composition in pigs maintained on a diet containing SUCRAM, establishing SUCRAM as a major influence driving bacterial community dynamics. The most notable change was a significant increase of Lactobacillaceae population abundance, almost entirely due to a single phylotype, designated Lactobacillus 4228. The sweetener-induced increase in Lactobacillaceae was observed in two different breeds of pigs signifying a general effect. We isolated Lactobacillus 4228, sequenced its genome and found it to be related to Lactobacillus amylovorus. In vitro analyses of Lactobacillus 4228 growth characteristics showed that presence of NHDC significantly reduces the lag phase of growth and enhances expression of specific sugar transporters, independently of NHDC metabolism. This study suggests that sensing of NHDC by a bacterial plasma membrane receptor underlies sweetener-induced growth of a health promoting gut bacterium.
Assuntos
Bactérias/metabolismo , Microbioma Gastrointestinal , Intestinos/microbiologia , Lactobacillus/crescimento & desenvolvimento , Edulcorantes/metabolismo , Suínos/microbiologia , Ração Animal/análise , Animais , Bactérias/genética , Bactérias/isolamento & purificação , Ceco/metabolismo , Ceco/microbiologia , Feminino , Mucosa Intestinal/metabolismo , Lactobacillus/metabolismo , Masculino , Sacarina/metabolismo , Suínos/metabolismo , DesmameRESUMO
Babesia spp. are tick-borne, intraerythrocytic hemoparasites that use antigenic variation to resist host immunity, through sequential modification of the parasite-derived variant erythrocyte surface antigen (VESA) expressed on the infected red blood cell surface. We identified the genomic processes driving antigenic diversity in genes encoding VESA (ves1) through comparative analysis within and between three Babesia species, (B. bigemina, B. divergens and B. bovis). Ves1 structure diverges rapidly after speciation, notably through the evolution of shortened forms (ves2) from 5' ends of canonical ves1 genes. Phylogenetic analyses show that ves1 genes are transposed between loci routinely, whereas ves2 genes are not. Similarly, analysis of sequence mosaicism shows that recombination drives variation in ves1 sequences, but less so for ves2, indicating the adoption of different mechanisms for variation of the two families. Proteomic analysis of the B. bigemina PR isolate shows that two dominant VESA1 proteins are expressed in the population, whereas numerous VESA2 proteins are co-expressed, consistent with differential transcriptional regulation of each family. Hence, VESA2 proteins are abundant and previously unrecognized elements of Babesia biology, with evolutionary dynamics consistently different to those of VESA1, suggesting that their functions are distinct.
Assuntos
Variação Antigênica , Babesia/genética , Evolução Molecular , Genes de Protozoários , Interações Hospedeiro-Parasita/genética , Pontos de Quebra do Cromossomo , Genoma de Protozoário , Proteínas de Protozoários/genética , Recombinação GenéticaRESUMO
The α-proteobacterium Wolbachia is probably the most prevalent, vertically transmitted symbiont on Earth. In contrast with its wide distribution in arthropods, Wolbachia is restricted to one family of animal-parasitic nematodes, the Onchocercidae. This includes filarial pathogens such as Onchocerca volvulus, the cause of human onchocerciasis, or river blindness. The symbiosis between filariae and Wolbachia is obligate, although the basis of this dependency is not fully understood. Previous studies suggested that Wolbachia may provision metabolites (e.g., haem, riboflavin, and nucleotides) and/or contribute to immune defense. Importantly, Wolbachia is restricted to somatic tissues in adult male worms, whereas females also harbor bacteria in the germline. We sought to characterize the nature of the symbiosis between Wolbachia and O. ochengi, a bovine parasite representing the closest relative of O. volvulus. First, we sequenced the complete genome of Wolbachia strain wOo, which revealed an inability to synthesize riboflavin de novo. Using RNA-seq, we also generated endobacterial transcriptomes from male soma and female germline. In the soma, transcripts for membrane transport and respiration were up-regulated, while the gonad exhibited enrichment for DNA replication and translation. The most abundant Wolbachia proteins, as determined by geLC-MS, included ligands for mammalian Toll-like receptors. Enzymes involved in nucleotide synthesis were dominant among metabolism-related proteins, whereas the haem biosynthetic pathway was poorly represented. We conclude that Wolbachia may have a mitochondrion-like function in the soma, generating ATP for its host. Moreover, the abundance of immunogenic proteins in wOo suggests a role in diverting the immune system toward an ineffective antibacterial response.
Assuntos
Regulação Bacteriana da Expressão Gênica , Genoma Bacteriano , Onchocerca volvulus/microbiologia , Simbiose/genética , Wolbachia/genética , Animais , Antibacterianos/metabolismo , Cromatografia Líquida , Replicação do DNA , DNA de Helmintos/genética , Feminino , Masculino , Proteômica/métodos , Riboflavina/metabolismo , Análise de Sequência de RNA , Espectrometria de Massas em Tandem , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo , Transcriptoma , Regulação para Cima , Wolbachia/imunologiaRESUMO
The commensal bacteria Lactobacillus are widely used as probiotic organisms conferring a heath benefit on the host. They have been implicated in promoting gut health via the stimulation of host immunity and anti-inflammatory responses, as well as protecting the intestinalmucosa against pathogen invasion. Lactobacilli grow by fermenting sugars and starches and produce lactic acid as their primary metabolic product. For efficient utilisation of varied carbohydrates, lactobacilli have evolved diverse sugar transport and metabolic systems, which are specifically induced by their own substrates. Many bacteria are also capable of sensing and responding to changes in their environment. These sensory responses are often independent of transport or metabolism and are mediated through membrane-spanning receptor proteins. We employed DNA-based pyrosequencing technology to investigate the changes in the intestinal microbiota of piglets weaned to a diet supplemented with either a natural sugar, lactose or an artificial sweetener (SUCRAM®, consisting of saccharin and neohesperidin dihydrochalcone (NHDC); Pancosma SA). The addition of either lactose or saccharin/NHDC to the piglets' feed dramatically increased the caecal population abundance of Lactobacillus, with concomitant increases in intraluminal lactic acid concentrations. This is the first report of the prebiotic-like effects of saccharin/NHDC, an artificial sweetener, being able to influence the commensal gut microbiota. The identification of the underlying mechanism(s) will assist in designing nutritional strategies for enhancing gut immunity and maintaining gut health.
Assuntos
Ceco/microbiologia , Chalconas/farmacologia , Hesperidina/análogos & derivados , Lactobacillus/crescimento & desenvolvimento , Lactose/farmacologia , Prebióticos , Sacarina/farmacologia , Edulcorantes/farmacologia , Animais , Ceco/metabolismo , Suplementos Nutricionais , Fermentação , Hesperidina/farmacologia , Ácido Láctico/metabolismo , Suínos , DesmameRESUMO
BACKGROUND: Insect cell lines play a vital role in many aspects of research on disease vectors and agricultural pests. The tsetse fly Glossina morsitans morsitans is an important vector of salivarian trypanosomes in sub-Saharan Africa and, as such, is a major constraint on human health and agricultural development in the region. METHODS: Here, we report establishment and partial characterisation of a cell line, GMA/LULS61, derived from tissues of adult female G. m. morsitans. GMA/LULS61 cells, grown at 28 °C in L-15 (Leibovitz) medium supplemented with foetal bovine serum and tryptose phosphate broth, have been taken through 23 passages to date and can be split 1:1 at 2-week intervals. Karyotyping at passage 17 revealed a predominantly haploid chromosome complement. Species origin and absence of contaminating bacteria were confirmed by PCR amplification and sequencing of fragments of the COI gene and pan-bacterial 16S rRNA gene respectively. However, PCR screening of RNA extracted from GMA/LULS61 cells confirmed presence of the recently described Glossina morsitans morsitans iflavirus and Glossina morsitans morsitans negevirus, but absence of Glossina pallipides salivary gland hypertrophy virus. GMA/LULS61 cells supported infection and growth of 6/7 different insect-derived strains of the intracellular bacterial symbiont Wolbachia. CONCLUSIONS: The GMA/LULS61 cell line has potential for application in a variety of studies investigating the biology of G. m. morsitans and its associated pathogenic and symbiotic microorganisms.
Assuntos
Moscas Tsé-Tsé , Moscas Tsé-Tsé/parasitologia , Animais , Linhagem Celular , Feminino , RNA Ribossômico 16S/genética , Cariotipagem , Insetos Vetores/virologiaRESUMO
OBJECTIVE: Microbial keratitis (MK) is a significant cause of blindness in sub-Saharan Africa. We investigated the feasibility of using a novel corneal impression membrane (CIM) for obtaining and processing samples by culture, PCR and whole-genome sequencing (WGS) in patients presenting with suspected MK in Malawi. METHODS AND ANALYSIS: Samples were collected from patients presenting with suspected MK using a 12 mm diameter polytetrafluoroethylene CIM disc. Samples were processed using culture and PCR for Acanthamoeba, herpes simplex virus type 1 (HSV-1) and the bacterial 16S rRNA gene. Minimum inhibitory concentrations of isolates to eight antimicrobials were measured using susceptibility strips. WGS was used to characterise Staphylococcus aureus isolates. RESULTS: 71 eyes of 71 patients were included. The overall CIM isolation rate was 81.7% (58 positive samples from 71 participants). 69 (81.2%) of isolates were Gram-positive cocci. Coagulase-negative Staphylococcus 31.8% and Streptococcus species 14.1% were the most isolated bacteria. Seven (9.9%) participants were positive for HSV-1. Fungi and Acanthamoeba were not detected. Moxifloxacin and chloramphenicol offered the best coverage for both Gram-positive and Gram-negative isolates when susceptibility was determined using known antimicrobial first quartile concentrations and European Committee on Antimicrobial Susceptibility Testing breakpoints, respectively. WGS identified known virulence genes associated with S. aureus keratitis. CONCLUSIONS: In a resource-poor setting, a CIM can be used to safely sample the cornea in patients presenting with suspected MK, enabling identification of causative microorganisms by culture and PCR. Although the microbiological spectrum found was limited to the dry season, these preliminary results could be used to guide empirical treatment.
Assuntos
Infecções Oculares Bacterianas , Humanos , Projetos Piloto , Malaui/epidemiologia , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Infecções Oculares Bacterianas/microbiologia , Infecções Oculares Bacterianas/epidemiologia , Infecções Oculares Bacterianas/tratamento farmacológico , Adulto Jovem , Bactérias/isolamento & purificação , Bactérias/efeitos dos fármacos , Bactérias/genética , Testes de Sensibilidade Microbiana , Córnea/microbiologia , Ceratite/microbiologia , Ceratite/tratamento farmacológico , Ceratite/epidemiologia , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Idoso , Reação em Cadeia da Polimerase , Adolescente , Acanthamoeba/isolamento & purificação , Acanthamoeba/genética , Acanthamoeba/efeitos dos fármacos , RNA Ribossômico 16S/genéticaRESUMO
BACKGROUND: SARS-CoV-2 remains rapidly evolving, and many biologically important genomic substitutions/indels have characterised novel SARS-CoV-2 lineages, which have emerged during successive global waves of the pandemic. Worldwide genomic sequencing has been able to monitor these waves, track transmission clusters, and examine viral evolution in real time to help inform healthcare policy. One school of thought is that an apparent greater than average divergence in an emerging lineage from contemporary variants may require persistent infection, for example in an immunocompromised host. Due to the nature of the COVID-19 pandemic and sampling, there were few studies that examined the evolutionary trajectory of SARS-CoV-2 in healthy individuals. METHODS: We investigated viral evolutionary trends and participant symptomatology within a cluster of 16 SARS-CoV-2 infected, immunocompetent individuals with no co-morbidities in a closed transmission chain. Longitudinal nasopharyngeal swab sampling allowed characterisation of SARS-CoV-2 intra-host variation over time at both the dominant and minor genomic variant levels through Nimagen-Illumina sequencing. RESULTS: A change in viral lineage assignment was observed in individual infections; however, there was only one indel and no evidence of recombination over the period of an acute infection. Minor and dominant genomic modifications varied between participants, with some minor genomic modifications increasing in abundance to become the dominant viral sequence during infection. CONCLUSIONS: Data from this cohort of SARS-CoV-2-infected participants demonstrated that long-term persistent infection in an immunocompromised host was not necessarily a prerequisite for generating a greater than average frequency of amino acid substitutions. Amino acid substitutions at both the dominant and minor genomic sequence level were observed in immunocompetent individuals during infection showing that viral lineage changes can occur generating viral diversity.
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COVID-19 , Genoma Viral , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , COVID-19/transmissão , COVID-19/virologia , COVID-19/genética , Masculino , Adulto , Feminino , Pessoa de Meia-Idade , Variação Genética , Imunocompetência , Evolução Molecular , Filogenia , IdosoRESUMO
OBJECTIVES: To understand severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission risks, perceived risks and the feasibility of risk mitigations from experimental mass cultural events before coronavirus disease 2019 (COVID-19) restrictions were lifted. DESIGN: Prospective, population-wide observational study. SETTING: Four events (two nightclubs, an outdoor music festival and a business conference) open to Liverpool City Region UK residents, requiring a negative lateral flow test (LFT) within the 36 h before the event, but not requiring social distancing or face-coverings. PARTICIPANTS: A total of 12,256 individuals attending one or more events between 28 April and 2 May 2021. MAIN OUTCOME MEASURES: SARS-CoV-2 infections detected using audience self-swabbed (5-7 days post-event) polymerase chain reaction (PCR) tests, with viral genomic analysis of cases, plus linked National Health Service COVID-19 testing data. Audience experiences were gathered via questionnaires, focus groups and social media. Indoor CO2 concentrations were monitored. RESULTS: A total of 12 PCR-positive cases (likely 4 index, 8 primary or secondary), 10 from the nightclubs. Two further cases had positive LFTs but no PCR. A total of 11,896 (97.1%) participants with scanned tickets were matched to a negative pre-event LFT: 4972 (40.6%) returned a PCR within a week. CO2 concentrations showed areas for improving ventilation at the nightclubs. Population infection rates were low, yet with a concurrent outbreak of >50 linked cases around a local swimming pool without equivalent risk mitigations. Audience anxiety was low and enjoyment high. CONCLUSIONS: We observed minor SARS-CoV-2 transmission and low perceived risks around events when prevalence was low and risk mitigations prominent. Partnership between audiences, event organisers and public health services, supported by information systems with real-time linked data, can improve health security for mass cultural events.