Isolates Of Lineage Research Articles

Mechanisms of high-level fosfomycin resistance in Staphylococcus aureus epidemic lineage ST5

ObjectivesFosfomycin resistance has become a clinical concern. In this study, we analysed the dynamic change of fosfomycin MIC in the epidemic Staphylococcus aureus lineages in a teaching hospital in Shanghai for 12 years and sought to elucidate the major underlying mechanisms.MethodsMLST was conducted for 4580 S. aureus isolates recovered from 2008 to 2019. Fosfomycin MIC was determined by the agar dilution method. The genome data of 230 S. aureus epidemic lineage isolates were acquired from a next-generation sequencing (NGS) platform. Gene deletion and corresponding complementation mutants were constructed to confirm the mechanism of fosfomycin resistance.ResultsThe predominant S. aureus lineages during the past 12 years were ST5 and ST239 (45.6%; 2090/4580). However, ST5 has been spreading clinically, while ST239 has gradually disappeared recently. Consistent with epidemic trends, fosfomycin-resistant ST5 increased from 19.5% to 67.3%. Most fosfomycin-resistant ST5 isolates (92.7%; 647/698) possessed high-level resistance (MIC > 1024 mg/L) with combined mutations mainly in glpT and uhpT. In contrast, fosfomycin-resistant ST239 isolates (76.8%; 149/194) mainly acquired low-level resistance (MIC = 64–128 mg/L) with mutation primarily in hptA. Deletion of a single resistant gene merely resulted in low-level fosfomycin resistance, while double-gene mutants ΔglpTΔuhpT, ΔglpTΔhptA and ΔglpTΔhptR acquired high-level fosfomycin resistance.ConclusionsThe high-level fosfomycin resistance of S. aureus epidemic lineage ST5 is mainly due to the accumulation of mutations in the resistant genes related to membrane transporter systems, and partly contributes to its persistent prevalence under clinical antibiotic pressure.

Evaluation of immunogenicity and cross-reactive responses of vaccines prepared from two chimeric serotype O foot-and-mouth disease viruses in pigs and cattle

Foot-and-mouth disease (FMD) remains a very serious barrier to agricultural development and the international trade of animals and animal products. Recently, serotype O has been the most prevalent FMDV serotype in China, and it has evolved into four different lineages: O/SEA/Mya-98, O/ME-SA/PanAsia, O/ME-SA/Ind-2001 and O/Cathay. PanAsia-2, belonging to the O/ME-SA topotype, is prevalent in neighbouring countries and poses the risk of cross-border spread in China. This study aimed to develop a promising vaccine candidate strain that can not only provide the best protection against all serotype O FMDVs circulating in China but also be used as an emergency vaccine for the prevention and control of transboundary incursion of PanAsia-2. Here, two chimeric FMDVs (rHN/TURVP1 and rHN/NXVP1) featuring substitution of VP1 genes of the O/TUR/5/2009 vaccine strain (PanAsia-2) and O/NXYCh/CHA/2018 epidemic strain (Mya98) were constructed and evaluated. The biological properties of the two chimeric FMDVs were similar to those of the wild-type (wt) virus despite slight differences in plaque sizes observed in BHK-21 cells. The structural protein-specific antibody titres induced by the rHN/TURVP1 and wt virus vaccines in pigs and cows were higher than those induced by the rHN/NXVP1 vaccine at 28–56 dpv. The vaccines prepared from the two chimeric viruses and wt virus all induced the production of protective cross-neutralizing antibodies against the viruses of the Mya-98, PanAsia and Ind-2001 lineages in pigs and cattle at 28 dpv; however, only the animals vaccinated with the rHN/TURVP1 vaccine produced a protective immune response to the field isolate of the Cathay lineage at 28 dpv, whereas the animals receiving the wt virus and the rHN/NXVP1 vaccines did not, although the wt virus and O/GXCX/CHA/2018 both belong to the Cathay topotype. This study will provide very useful information to help develop a potential vaccine candidate for the prevention and control of serotype O FMD in China.

Defining the Skeletal Myogenic Lineage in Human Pluripotent Stem Cell-Derived Teratomas.

Skeletal muscle stem cells are essential to muscle homeostasis and regeneration after injury, and have emerged as a promising cell source for treating skeletal disorders. An attractive approach to obtain these cells utilizes differentiation of pluripotent stem cells (PSCs). We recently reported that teratomas derived from mouse PSCs are a rich source of skeletal muscle stem cells. Here, we showed that teratoma formation is also capable of producing skeletal myogenic progenitors from human PSCs. Using single-cell transcriptomics, we discovered several distinct skeletal myogenic subpopulations that represent progressive developmental stages of the skeletal myogenic lineage and recapitulate human embryonic skeletal myogenesis. We further discovered that ERBB3 and CD82 are effective surface markers for prospective isolation of the skeletal myogenic lineage in human PSC-derived teratomas. Therefore, teratoma formation provides an accessible model for obtaining human skeletal myogenic progenitors from PSCs.

SARS-CoV-2 Kappa Variant Shows Pathogenicity in a Syrian Hamster Model.

Objectives: The emergence of SARS-CoV-2 lineage B.1.617 variants in India has been associated with a surge in the number of daily infections. We investigated the pathogenic potential of Kappa (B.1.617.1) variant in Syrian golden hamsters. Methods: Two groups of Syrian golden hamsters (18 each) were inoculated intranasally with SARS-CoV-2 isolates, B.1 (D614G) and Kappa variant, respectively. The animals were monitored daily for the clinical signs and body weight. Throat swab, nasal wash, and organ samples (lungs, nasal turbinate, trachea) were collected and screened using SARS-CoV-2-specific RT-qPCR. Histopathologic evaluation of the lung samples was performed. Results: The hamsters infected with the Kappa variant demonstrated increased body weight loss compared to the B.1 lineage isolate. The highest viral RNA load was observed in the nasal turbinate and lung specimens of animals infected with both variants. A significantly higher sgRNA load was observed in the nasal swabs (7 DPI), trachea (3 DPI), and lungs (3 DPI) of hamsters infected with the Kappa variant. Neutralizing antibody response generated in the B.1 lineage-infected hamster sera were comparable against both B.1 and Kappa variant in contrast to Kappa variant-infected hamsters, which showed lower titers against B.1 lineage isolate. Gross and microscopic evaluation of the lung specimens showed severe lung lesions in hamsters infected with Kappa variant compared to B.1. Conclusions: The study demonstrates pathogenicity of Kappa variant in hamsters evident with reduced body weight, high viral RNA load in lungs, and pronounced lung lesions. Both Kappa variant- and B.1-infected hamsters produced neutralizing antibodies against both variants studied.

Evolutionary history and introduction of SARS-CoV-2 Alpha VOC/B.1.1.7 in Pakistan through international travelers.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants continue to emerge, and their identification is important for the public health response to coronavirus disease 2019 (COVID-19). Genomic sequencing provides robust information but may not always be accessible, and therefore, mutation-based polymerase chain reaction (PCR) approaches can be used for rapid identification of known variants. International travelers arriving in Karachi between December 2020 and February 2021 were tested for SARS-CoV-2 by PCR. A subset of positive samples was tested for S-gene target failure (SGTF) on TaqPathTM COVID-19 (Thermo Fisher Scientific) and for mutations using the GSD NovaType SARS-CoV-2 (Eurofins Technologies) assays. Sequencing was conducted on the MinION platform (Oxford Nanopore Technologies). Bayesian phylogeographic inference was performed integrating the patients’ travel history information. Of the thirty-five COVID-19 cases screened, thirteen had isolates with SGTF. The travelers transmitted infection to sixty-eight contact cases. The B.1.1.7 lineage was confirmed through sequencing and PCR. The phylogenetic analysis of sequence data available for six cases included four B.1.1.7 strains and one B.1.36 and B.1.1.212 lineage isolate. Phylogeographic modeling estimated at least three independent B.1.1.7 introductions into Karachi, Pakistan, originating from the UK. B.1.1.212 and B.1.36 were inferred to be introduced either from the UK or the travelers’ layover location. We report the introduction of SARS-CoV-2 B.1.1.7 and other lineages in Pakistan by international travelers arriving via different flight routes. This highlights SARS-CoV-2 transmission through travel, importance of testing, and quarantine post-travel to prevent transmission of new strains, as well as recording detailed patients’ metadata. Such results help inform policies on restricting travel from destinations where new highly transmissible variants have emerged.

A new cluster of West Nile virus lineage 1 identified in Spain

<h3>Purpose</h3> West Nile Virus (WNV) is a zoonotic arbovirus worldwide spread. In Spain, its active circulation provokes outbreaks in birds, horses and lately in humans. In September 2017, an outbreak affecting raptors, in the province of Málaga (Andalusia), was investigated. One northern goshawk (<i>Accipiter gentilis</i>) died, while two peregrine falcons (<i>Falco peregrinus</i>) recovered. Here we report the isolation and characterization of a new WNV lineage 1 variant from this outbreak. <h3>Methods & Materials</h3> Heart, feathers, brain and kidney samples from the goshawk were collected and firstly analysed by real-time RT-PCR for WNV (Jiménez-Clavero <i>et al</i>., 2006, https://doi.org/doi:10.1177/104063870601800505). Virus isolation was performed in BSR and Vero cells. The whole genome of the isolate was sequenced by Sanger method. Phylogenetic analyses were performed including a wide range of representatives of: 1) all described WNV lineages, and 2) the different clusters of WNV lineage 1. <h3>Results</h3> All samples from the goshawk were positive for WNV lineage 1 by RT-PCR. Virus isolation was successful in heart and brain samples. A full genome sequence of 10,919 nt was obtained for the WNV isolated from the brain (named Spain/2017/NG-b). Phylogenetic analyses confirmed that Spain/2017/NG-b belonged to WNV clade 1a. Surprisingly, this isolate was placed neither in the cluster 2 of this clade, which comprises most of the recent European and Mediterranean strains (including the preceding Spanish ones), nor in other closest clusters. When analysing the polyprotein, amino acid substitutions that define WNV clusters 1-4 were not found in Spain/2017/NG-b isolate (May <i>et al.</i>, 2011, https://doi.org/10.1128/jvi.01963-10). Moreover, it presented 12 unique residues compared to cluster 2 viruses. <h3>Conclusion</h3> These results strongly support that Spain/2017/NG-b represents a new, independent introduction of WNV into the Western Mediterranean since it showed no close relationship to other strains from this region. Therefore, we propose to associate this variant to a new cluster 7 within WNV clade 1a. Finally, a WNV outbreak occurred in equids in December 2017, nearby the raptors´ outbreak above mentioned, suggests the ability of this new variant to infect and cause disease also in horses.

A high-resolution melting assay to differentiate a peste des petits ruminants virus vaccine strain from field isolates in Turkey.

Peste des petits ruminants (PPR) mostly affects small ruminants. Vaccination with attenuated vaccines derived from PPR virus (PPRV) provides successful protection against the disease. However, current molecular methods are unable to distinguish vaccine strains from field strains. In this study, we used an EvaGreen-based high-resolution melting (HRM) assay to differentiate a PPRV vaccine strain (Nigeria 75/1) from Turkish field isolates of lineage IV based on melting peaks and melting profiles.

A Phylogeographical Analysis of the Beetle Pest Species Callosobruchus chinensis (Linnaeus, 1758) in China.

Simple SummaryCallosobruchus chinensis, a stored product pest, is difficult to control. In the study, the goal was to explore the causes of the demographic history, dispersal path and genetic variations underlying the spatial and temporal distribution of C. chinensis in China. The phylogeography of C. chinensis was analyzed by distribution modelling (SDM) under six periods and the least-cost path (LCP) based on combined mitochondrial DNA. Our data showed that the geographical isolation of the genetic lineages and the distribution range of C. chinensis were restricted by climate in different times. The landscape structure had influence on the genetic differentiation of C. chinensis. Although the migration ability of C. chinensis is limited, the development of transportation and trade is helping the insect spread, along with the beans of its host.Callosobruchus chinensis (Coleoptera Bruchidae), is a pest of different varieties of legumes. In this paper, a phylogeographical analysis of C. chinensis was conducted to provide knowledge for the prevention and control of C. chinensis. A total of 224 concatenated mitochondrial sequences were obtained from 273 individuals. Suitable habitat shifts were predicted by the distribution modelling (SDM). Phylogeny, genetic structure and population demographic history were analyzed using multiple software. Finally, the least-cost path (LCP) method was used to identify possible dispersal corridors and genetic connectivity. The SDM results suggested that the distribution of C. chinensis experienced expansion and contraction with changing climate. Spatial distribution of mtDNA haplotypes showed there was partial continuity among different geographical populations of C. chinensis, except for the Hohhot (Inner Mongolia) population. Bayesian skyline plots showed that the population had a recent expansion during 0.0125 Ma and 0.025 Ma. The expansion and divergent events were traced back to Quaternary glaciations. The LCP method confirmed that there were no clear dispersal routes. Our findings indicated that climatic cycles of the Pleistocene glaciations, unsuitable climate and geographic isolation played important roles in the genetic differentiation of C. chinensis. Human activities weaken the genetic differentiation between populations. With the change in climate, the suitable areas of C. chinensis will disperse greatly in the future.

Moths passing in the night: Phenological and genomic divergences within a forest pest complex.

Temporal separation of reproductive timing can contribute to species diversification both through allochronic speciation and later reinforcement of species boundaries. Such phenological differences are an enigmatic component of evolutionary divergence between two major forest defoliator species of the spruce budworm complex: Choristoneura fumiferana and C. occidentalis. While these species interbreed freely in laboratory settings, natural hybridization rates have not been reliably quantified due to their indistinguishable morphology. To assess whether temporal isolation is contributing to reproductive isolation, we collected adult individuals throughout their expected zone of sympatry in western Canada at 10‐day intervals over two successive years, assigning taxonomic identities using thousands of single nucleotide polymorphisms. We found unexpectedly broad sympatry between C. fumiferana and C. occidentalis biennis and substantial overlap of regional flight periods. However, flight period divergence was much more apparent on a location‐by‐location basis, highlighting the importance of considering spatial scale in these analyses. Phenological comparisons were further complicated by the biennial life cycle of C. o. biennis, the main subspecies of C. occidentalis in the region, and the occasional occurrence of the annually breeding subspecies C. o. occidentalis. Nonetheless, we demonstrate that biennialism is not a likely contributor to reproductive isolation within the species complex. Overall, interspecific F1 hybrids comprised 2.9% of sequenced individuals, confirming the genomic distinctiveness of C. fumiferana and C. occidentalis, while also showing incomplete reproductive isolation of lineages. Finally, we used F ST‐based outlier and genotype–environment association analyses to identify several genomic regions under putative divergent selection. These regions were disproportionately located on the Z linkage region of C. fumiferana, and contained genes, particularly antifreeze proteins, that are likely to be associated with overwintering success and diapause. In addition to temporal isolation, we conclude that other mechanisms, including ecologically mediated selection, are contributing to evolutionary divergence within the spruce budworm species complex.

Population genomics indicates micro‐refuges and riverine barriers for a southern South American grassland nightshade

AbstractAimPleistocene climate oscillations influenced the species distribution and genetic diversity in grasslands, and such climatic changes promoted expansion and contraction cycles, leading to fragmentation and isolation of evolutionary lineages in refuges. This study aimed to infer the evolutionary processes that most influenced the genetic diversity of a South American nightshade Petunia inhabiting subtropical grasslands under the Pleistocene influence.LocationPampa and Chaco.TaxonThe widely distributed herb Petunia axillaris subsp. parodii (Solanaceae).MethodsWe obtained high‐density genome coverage throughout the geographical distribution of Petunia axillaris subsp. parodii. We estimated the genetic diversity and structure to evaluate population differentiation and applied a model‐based demographic analysis to investigate the scenarios that could have influenced the species' evolutionary history. We evaluated the association between neutral and outlier polymorphisms with environmental variables to distinguish the influence of the geographical distance and environmental differences between populations.ResultsThere are three evolutionary lineages in P. axillaris subsp. parodii. The origin and differentiation of these lineages were related to the Pleistocene refuges and rivers acting as barriers to gene flow. We also identified 496 outlier loci related to adaptation to environmental conditions.Main conclusionThe Pleistocene climate changes drove lineage diversification isolated in micro‐refugia. Rivers and changes in their courses may have also acted on the population divergence, serving as barriers to gene flow or even as corridors for species range expansion. These findings contribute to the knowledge of the evolutionary processes that influenced the lineage diversification of South American subtropical grassland species.

Neutralization of SARS-CoV-2 Variants by Serum from BNT162b2 Vaccine Recipients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has evolved rapidly, leading to viral lineages characterized by multiple mutations in the spike protein, which could potentially confer to the virus the ability to avoid the vaccine-induced immune response, making the vaccines less effective or ineffective. Here, we initially evaluated the neutralization capabilities in vitro by serum neutralization (SN) of six serum samples collected from recipients of the BNT162b2 vaccine against 11 SARS-CoV-2 isolates belonging to the major SARS-CoV-2 lineages that had been circulating in Italy. Then, we considered 30 additional serum samples by SN assay against the dominant B.1.617.2 (Delta) variant. A B.1 lineage isolate was used as a reference. In the first analysis, significant differences when compared with the reference strain (p > 0.05) were not evidenced; instead, when the panel of 30 sera was tested against the B.1.617.2 (Delta) variant, a significant (p = 0.0015) 2.38-fold reduction in neutralizing titres compared with the reference after the first vaccine dose was demonstrated. After the second vaccine dose, the reduction was not significant (p = 0.1835). This study highlights that the BNT162b2 vaccine stimulates a humoral response able to neutralize all tested SARS-CoV-2 variants, thus suggesting a prominent role in mitigating the impact of the SARS-CoV-2 pandemic in real-world conditions. Long-term follow-up is currently ongoing.

Heuristic and Hierarchical-Based Population Mining of Salmonella enterica Lineage I Pan-Genomes as a Platform to Enhance Food Safety

The recent incorporation of bacterial whole-genome sequencing (WGS) into Public Health laboratories has enhanced foodborne outbreak detection and source attribution. As a result, large volumes of publicly available datasets can be used to study the biology of foodborne pathogen populations at an unprecedented scale. To demonstrate the application of a heuristic and agnostic hierarchical population structure guided pan-genome enrichment analysis (PANGEA), we used populations of S. enterica lineage I to achieve two main objectives: (i) show how hierarchical population inquiry at different scales of resolution can enhance ecological and epidemiological inquiries; and (ii) identify population-specific inferable traits that could provide selective advantages in food production environments. Publicly available WGS data were obtained from NCBI database for three serovars of Salmonella enterica subsp. enterica lineage I (S. Typhimurium, S. Newport, and S. Infantis). Using the hierarchical genotypic classifications (Serovar, BAPS1, ST, cgMLST), datasets from each of the three serovars showed varying degrees of clonal structuring. When the accessory genome (PANGEA) was mapped onto these hierarchical structures, accessory loci could be linked with specific genotypes. A large heavy-metal resistance mobile element was found in the Monophasic ST34 lineage of S. Typhimurium, and laboratory testing showed that Monophasic isolates have on average a higher degree of copper resistance than the Biphasic ones. In S. Newport, an extra sugE gene copy was found among most isolates of the ST45 lineage, and laboratory testing of multiple isolates confirmed that isolates of S. Newport ST45 were on average less sensitive to the disinfectant cetylpyridimium chloride than non-ST45 isolates. Lastly, data-mining of the accessory genomic content of S. Infantis revealed two cryptic Ecotypes with distinct accessory genomic content and distinct ecological patterns. Poultry appears to be the major reservoir for Ecotype 1, and temporal analysis further suggested a recent ecological succession, with Ecotype 2 apparently being displaced by Ecotype 1. Altogether, the use of a heuristic hierarchical-based population structure analysis that includes bacterial pan-genomes (core and accessory genomes) can (1) improve genomic resolution for mapping populations and accessing epidemiological patterns; and (2) define lineage-specific informative loci that may be associated with survival in the food chain.

An epidemic Zika virus isolate suppresses antiviral immunity by disrupting antigen presentation pathways

Zika virus (ZIKV) has emerged as an important global health threat, with the recently acquired capacity to cause severe neurological symptoms and to persist within host tissues. We previously demonstrated that an early Asian lineage ZIKV isolate induces a highly activated CD8 T cell response specific for an immunodominant epitope in the ZIKV envelope protein in wild-type mice. Here we show that a contemporary ZIKV isolate from the Brazilian outbreak severely limits CD8 T cell immunity in mice and blocks generation of the immunodominant CD8 T cell response. This is associated with a more sustained infection that is cleared between 7- and 14-days post-infection. Mechanistically, we demonstrate that infection with the Brazilian ZIKV isolate reduces the cross-presentation capacity of dendritic cells and fails to fully activate the immunoproteasome. Thus, our study provides an isolate-specific mechanism of host immune evasion by one Brazilian ZIKV isolate, which differs from the early Asian lineage isolate and provides potential insight into viral persistence associated with recent ZIKV outbreaks.

Molecular genetic analysis of West Nile virus variants circulating in European Russia between 2010 and 2019

Aim. A study of the prevalence of West Nile virus (WNV) genetic lineages and genovariants in the south of European Russia between 2010 and 2019.Materials and methods. The study was carried out on 311 WNV containing biological samples from patients, vectors and reservoirs of infection. WNV typing was carried out using reverse transcription and real-time polymerase chain reaction with designed three pairs of primers and three probes and by the sequencing of the 277 bp WNV genome region corresponding to the 5'-untranslated region and locus of the polyprotein gene encoding the capsid protein C. Sequencing results were analyzed using the Nucleotide BLAST software (NCBI).Results. As a result of typing, out of 311 WNV RNA isolates taken for the study, 15 (4.82%) were assigned to lineage 1 (from Astrakhan and Volgograd regions, Krasnodar and Stavropol Territories, Republic of Tatarstan), 285 (91.64%) to lineage 2 (from Astrakhan, Volgograd, Voronezh, Kursk, Lipetsk, Penza, Rostov and Saratov regions, Krasnodar and Stavropol Territories, Republics of Kalmykia and Crimea), and 11 (3.54%) to lineage 4 (from the Volgograd region, Republics of Kalmykia and Crimea). The predominance of viral lineage 2 was demonstrated. The identified isolates of the viral lineage 1 belonged to the «Astrakhan» variant, isolates of lineage 2 belonged to «Russian» and «European» variants. Previously uncommon WNV variants of lineages 1 and 4 were also found.Conclusion. Lineage 2 of WNV prevailed in the south of European Russia in the last decade. The «Russian» variant is most common and its area is expanding. The circulation of various WNV genetic lineages in Russia indicates the need for further study of their spread and improving diagnostic methods and test systems for identifying and differentiating pathogen strains.

Characterization of beta-lactam-resistant Escherichia coli from Australian fruit bats indicates anthropogenic origins.

Antimicrobial-resistant Escherichia coli , particularly those resistant to critically important antimicrobials, are increasingly reported in wildlife. The dissemination of antimicrobial-resistant bacteria to wildlife indicates the far-reaching impact of selective pressures imposed by humans on bacteria through misuse of antimicrobials. The grey-headed flying fox (GHFF; Pteropus poliocephalus), a fruit bat endemic to eastern Australia, commonly inhabits urban environments and encounters human microbial pollution. To determine if GHFF have acquired human-associated bacteria, faecal samples from wild GHFF (n=287) and captive GHFF undergoing rehabilitation following illness or injury (n=31) were cultured to detect beta-lactam-resistant E. coli . Antimicrobial susceptibility testing, PCR and whole genome sequencing were used to determine phenotypic and genotypic antimicrobial resistance profiles, strain type and virulence factor profiles. Overall, 3.8 % of GHFF carried amoxicillin-resistant E. coli (wild 3.5 % and captive 6.5 %), with 38.5 % of the 13 GHFF E. coli isolates exhibiting multidrug resistance. Carbapenem (bla NDM-5) and fluoroquinolone resistance were detected in one E. coli isolate, and two isolates were resistant to third-generation cephalosporins (bla CTX-M-27 and ampC). Resistance to tetracycline and trimethoprim plus sulfamethoxazole were detected in 69.2% and 30.8 % of isolates respectively. Class 1 integrons, a genetic determinant of resistance, were detected in 38.5 % of isolates. Nine of the GHFF isolates (69.2 %) harboured extraintestinal virulence factors. Phylogenetic analysis placed the 13 GHFF isolates in lineages associated with humans and/or domestic animals. Three isolates were human-associated extraintestinal pathogenic E. coli (ST10 O89:H9, ST73 and ST394) and seven isolates belonged to lineages associated with extraintestinal disease in both humans and domestic animals (ST88, ST117, ST131, ST155 complex, ST398 and ST1850). This study provides evidence of anthropogenic multidrug-resistant and pathogenic E. coli transmission to wildlife, further demonstrating the necessity for incorporating wildlife surveillance within the One Health approach to managing antimicrobial resistance.

Hybridization, ecogeographical displacement and the emergence of new lineages – A genotyping‐by‐sequencing and ecological niche and species distribution modelling study of Sempervivum tectorum L. (Houseleek)

Ecogeographical displacement of homoploid hybrid lineages from their parents is well documented and considered an important mechanism to achieve reproductive isolation. In this study, we investigated the origin of the flowering plant species Sempervivum tectorum in the Massif Central (France) through homoploid hybridization between lineages of the species from the Rhine Gorge area (Germany) and the Pyrenees (France). We used genotyping-by-sequencing genetic data as evidence for the hybrid origin of the Massif Central lineage, and WorldClim climatic data and soil pH and soil temperature data collected by us for ecological niche and species distribution modelling. We could show that the Massif Central lineage shows hybrid admixture and that the niche of this lineage is significantly different from those of the parental lineages. In comparison with the parental niches, different variables of the niche of the hybrid lineage are intermediate, parental-combined or extreme. The different niche of the Massif Central populations thus can plausibly be interpreted as hybridization-derived. Our species distribution modelling for the Last Glacial Maximum and Mid-Holocene showed that the potential distribution of the hybrid lineage at the likely time of its origin in the Quaternary possibly was parapatric in relation to the largely sympatric distributions of the parental lineages. We hypothesize that reproductive isolation of the hybrid lineage from the parental lineages resulted from the segregation of distribution ranges by a differential response of the three lineages to a warming climate.

A Sequential Study on the Pathology of Peste Des Petits Ruminants and Tissue Distribution of the Virus Following Experimental Infection of Black Bengal Goats.

We studied the sequential pathology of peste des petits ruminants (PPR) in Black Bengal goats and analyzed virus distribution in tissues and virus shedding following experimental infection with a Bangladeshi isolate of lineage IV PPR virus (PPRV). The early clinical signs like fever, depression, and ocular and nasal discharges first appeared at 4–7 days post-infection (dpi). Three out of eight inoculated goats died at 13, 15, and 18 dpi, and the rest were killed at different time points from 5 to 18 dpi. Initially, the virus multiplied mostly in the lymphoid organs of the pharyngeal region and caused extensive lymphoid destruction and hemorrhages. This was followed by viremia, massive virus replication in the lungs, and pneumonia along with the appearance of the clinical signs. Subsequently, the virus spread to other organs causing necrotic and hemorrhagic lesions, as well as the virus localized in the upper respiratory, oral and intestinal mucosa resulting in catarrhal, erosive, and ulcerative lesions. On hematological and biochemical investigation progressive leukopenia and hypoproteinemia, a gradual increase of serum metabolites and enzymes associated with liver and kidney damage, and electrolyte imbalance were observed. Seroconversion started at 7 dpi and all the surviving animals had serum antibodies at 14 dpi. Virus shedding was observed in nasal and ocular secretions at 4 dpi and in feces and urine at 14 dpi, which gradually increased and continued till the end of the experiment (18 dpi) despite seroconversion. Therefore, the virus shedding of naturally infected seroconverted goats should be monitored for effective control strategies.

Emerging variants of concern in SARS-CoV-2 membrane protein: a highly conserved target with potential pathological and therapeutic implications

ABSTRACT Mutations in the SARS-CoV-2 Membrane (M) gene are relatively uncommon. The M gene encodes the most abundant viral structural protein, and is implicated in multiple viral functions, including initial attachment to the host cell via heparin sulphate proteoglycan, viral protein assembly in conjunction with the N and E genes, and enhanced glucose transport. We have identified a recent spike in the frequency of reported SARS-CoV-2 genomes carrying M gene mutations. This is associated with emergence of a new sub-B.1 clade, B.1.I82T, defined by the previously unreported M:I82T mutation within TM3, the third of three membrane spanning helices implicated in glucose transport. The frequency of this mutation increased in the USA from 0.014% in October 2020 to 1.62% in February 2021, a 116-fold change. While constituting 0.7% of the isolates overall, M:I82T sub-B.1 lineage accounted for 14.4% of B.1 lineage isolates in February 2021, similar to the rapid initial increase previously seen with the B.1.1.7 and B.1.429 lineages, which quickly became the dominant lineages in Europe and California over a period of several months. A similar increase in incidence was also noted in another related mutation, V70L, also within the TM2 transmembrane helix. These M mutations are associated with younger patient age (4.6 to 6.3 years). The rapid emergence of this B.1.I82T clade, recently named Pangolin B.1.575 lineage, suggests that this M gene mutation is more biologically fit, perhaps related to glucose uptake during viral replication, and should be included in ongoing genomic surveillance efforts and warrants further evaluation for potentially increased pathogenic and therapeutic implications.

Levels of SARS-CoV-2 Lineage P.1 Neutralization by Antibodies Elicited after Natural Infection and Vaccination

Background: A new SARS-CoV-2 lineage, named P.1 (20J/501Y.V3), has recently been detected in Brazil. Mutations accrued by the P.1 lineage include amino acid changes in the receptor-binding domain of the spike protein that also are reported in variants of concern in the United Kingdom (B.1.1.7) and South Africa (B.1.325).Methods: We isolated two P.1-containing specimens from nasopharyngeal and bronchoalveolar lavage samples of patients of Manaus, Brazil. We measured neutralization of the P.1 virus after incubation with the plasma of 19 COVID-19 convalescent blood donors and recipients of the chemically-inactivated CoronaVac vaccine and compared these results to neutralization of a SARS-CoV-2 B-lineage previously circulating in Brazil.Findings: The immune plasma of COVID-19 convalescent blood donors had 6-fold less neutralizing capacity against the P.1 than against the B-lineage. Moreover, five months after booster immunization with CoronaVac, plasma from vaccinated individuals failed to efficiently neutralize P.1 lineage isolates.Interpretation: These data indicate that the P.1 lineage may escape from neutralizing antibodies generated in response to polyclonal stimulation against previously circulating variants of SARS-CoV-2.Funding: São Paulo Research Foundation, MCTI/FINEP, Medical Research Council, National Council for Scientific and Technological Development, National Institutes of Health.Conflict of Interest: M.S.D. is a consultant for Inbios, Vir Biotechnology, NGMBiopharmaceuticals, and Carnival Corporation, and on the Scientific Advisory Boards of Moderna and Immunome. The Diamond laboratory has received funding support in sponsored research agreements from Moderna, Vir Biotechnology, and Emergent BioSolutions.Ethical Approval: All procedures followed the ethical standards of the responsible committee on humanexperimentation and approved by the ethics committees from the University of Campinas, Brazil (Approval number CONEP 4.021.484 for plasma collection of blood donors, CAEE32078620.4.0000.5404 and 30227920.9.0000.5404 for the sampling of vaccinated and viral genome sequencing, respectively). All patient data were anonymized before study inclusion.Informed consent was obtained from all subjects for being included in the study.

Two pleosporalean root-colonizing fungi, Fuscosphaeria hungarica gen. et sp. nov. and Delitschia chaetomioides, from a semiarid grassland in Hungary

In this study, we investigated two unidentified lineages of root-colonizing fungi belonging to the order Pleosporales (Dothideomycetes), which were isolated from Festuca vaginata (Poaceae), a dominant grass species in the semiarid sandy grasslands of Hungary. For molecular phylogenetic studies, seven loci (internal transcribed spacer, partial large subunit and small subunit region of nrRNA, partial transcription elongation factor 1-α, RNA polymerase II largest subunit, RNA polymerase II second largest subunit, and ß-tubulin genes) were amplified and sequenced. Based on morphology and multilocus phylogenetic analyses, we found that one lineage belonged to Delitschia chaetomioides P. Karst. (Delitschiaceae), and the isolates of the other lineage represented a novel monotypic genus in the family Trematosphaeriaceae (suborder Massarineae). For this lineage, we proposed a new genus, Fuscosphaeria, represented by a single species, F. hungarica. In both lineages, only immature and degenerated sporocarps could be induced. These were sterile, black, globose, or depressed globose structures with numerous mycelioid appendages submerged in culture media or on the surface of autoclaved plant materials. Both species are first reported here as root-colonizing fungi.