Infection of wild-type mice by SARS-CoV-2 B.1.351 variant indicates a possible novel cross-species transmission route

AbstractCOVID-19 is identified as a zoonotic disease caused by SARS-CoV-2, which also can cross-transmit to many animals but not mice. Genetic modifications of SARS-CoV-2 or mice enable the mice susceptible to viral infection. Although neither is the natural situation, they are currently utilized to establish mouse infection models. Here we report a direct contact transmission of SARS-CoV-2 variant B.1.351 in wild-type mice. The SARS-CoV-2 (B.1.351) replicated efficiently and induced significant pathological changes in lungs and tracheas, accompanied by elevated proinflammatory cytokines in the lungs and sera. Mechanistically, the receptor-binding domain (RBD) of SARS-CoV-2 (B.1.351) spike protein turned to a high binding affinity to mouse angiotensin-converting enzyme 2 (mACE2), allowing the mice highly susceptible to SARS-CoV-2 (B.1.351) infection. Our work suggests that SARS-CoV-2 (B.1.351) expands the host range and therefore increases its transmission route without adapted mutation. As the wild house mice live with human populations quite closely, this possible transmission route could be potentially risky. In addition, because SARS-CoV-2 (B.1.351) is one of the major epidemic strains and the mACE2 in laboratory-used mice is naturally expressed and regulated, the SARS-CoV-2 (B.1.351)/mice could be a much convenient animal model system to study COVID-19 pathogenesis and evaluate antiviral inhibitors and vaccines.

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Nanobody cocktails potently neutralize SARS-CoV-2 D614G N501Y variant and protect mice

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2101918118 ◽

2021 ◽

Vol 118 (19) ◽

pp. e2101918118

Author(s):

Phillip Pymm ◽

Amy Adair ◽

Li-Jin Chan ◽

James P. Cooney ◽

Francesca L. Mordant ◽

...

Keyword(s):

Neutralizing Antibodies ◽

Human Populations ◽

Wild Type ◽

Angiotensin Converting Enzyme 2 ◽

Viral Loads ◽

X Ray Crystallography ◽

Binding Domains ◽

Cryo Electron Microscopy ◽

Antigenic Sites ◽

Human Receptor

Neutralizing antibodies are important for immunity against SARS-CoV-2 and as therapeutics for the prevention and treatment of COVID-19. Here, we identified high-affinity nanobodies from alpacas immunized with coronavirus spike and receptor-binding domains (RBD) that disrupted RBD engagement with the human receptor angiotensin-converting enzyme 2 (ACE2) and potently neutralized SARS-CoV-2. Epitope mapping, X-ray crystallography, and cryo-electron microscopy revealed two distinct antigenic sites and showed two neutralizing nanobodies from different epitope classes bound simultaneously to the spike trimer. Nanobody-Fc fusions of the four most potent nanobodies blocked ACE2 engagement with RBD variants present in human populations and potently neutralized both wild-type SARS-CoV-2 and the N501Y D614G variant at concentrations as low as 0.1 nM. Prophylactic administration of either single nanobody-Fc or as mixtures reduced viral loads by up to 104-fold in mice infected with the N501Y D614G SARS-CoV-2 virus. These results suggest a role for nanobody-Fc fusions as prophylactic agents against SARS-CoV-2.

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Roasted coffee wastes as a substrate for Escherichia coli to grow and produce hydrogen

FEMS Microbiology Letters ◽

10.1093/femsle/fnaa088 ◽

2020 ◽

Vol 367 (11) ◽

Cited By ~ 1

Author(s):

Hripsime Petrosyan ◽

Liana Vanyan ◽

Satenik Mirzoyan ◽

Armen Trchounian ◽

Karen Trchounian

Keyword(s):

Escherichia Coli ◽

H2 Production ◽

High Rate ◽

Main Role ◽

Wild Type ◽

Roasted Coffee ◽

Energy Needs ◽

Genetic Modifications ◽

Coffee Grounds ◽

And Control

ABSTRACT After brewing roasted coffee, spent coffee grounds (SCGs) are generated being one of the daily wastes emerging in dominant countries with high rate and big quantity. Escherichia coli BW25113 wild-type strain, mutants with defects in hydrogen (H2)-producing/oxidizing four hydrogenases (Hyd) (ΔhyaB ΔhybC, ΔhycE, ΔhyfG) and septuple mutant (ΔhyaB ΔhybC ΔhycA ΔfdoG ΔldhA ΔfrdC ΔaceE) were investigated by measuring change of external pH, bacterial growth and H2 production during the utilization of SCG hydrolysate. In wild type, H2 was produced with rate of 1.28 mL H2 (g sugar)−1 h−1 yielding 30.7 mL H2 (g sugar)−1 or 2.75 L (kg SCG)−1 during 24 h. In septuple mutant, H2 production yield was 72 mL H2 (g sugar)−1 with rate of 3 mL H2 (g sugar)−1 h−1. H2 generation was absent in hycE single mutant showing the main role of Hyd-3 in H2 production. During utilization of SCG wild type, specific growth rate was 0.72 ± 0.01 h−1 with biomass yield of 0.3 g L−1. Genetic modifications and control of external parameters during growth could lead to prolonged and enhanced microbiological H2 production by organic wastes, which will aid more efficiently global sustainable energy needs resulting in diversification of mobile and fixed energy sources.

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Identification of prolyl carboxypeptidase as an alternative enzyme for processing of renal angiotensin II using mass spectrometry

AJP Cell Physiology ◽

10.1152/ajpcell.00346.2012 ◽

2013 ◽

Vol 304 (10) ◽

pp. C945-C953 ◽

Cited By ~ 28

Author(s):

Nadja Grobe ◽

Nathan M. Weir ◽

Orly Leiva ◽

Frank S. Ong ◽

Kenneth E. Bernstein ◽

...

Keyword(s):

Ph Control ◽

Mass Spectrometric ◽

Acidic Ph ◽

Ang Ii ◽

Converting Enzyme ◽

Wild Type ◽

Angiotensin Converting Enzyme 2 ◽

Genetic Mouse Models

Angiotensin-converting enzyme 2 (ACE2) catalyzes conversion of ANG II to ANG-(1–7). The present study uses newly established proteomic approaches and genetic mouse models to examine the contribution of alternative renal peptidases to ACE2-independent formation of ANG-(1–7). In situ and in vitro mass spectrometric characterization showed that substrate concentration and pH control renal ANG II processing. At pH ≥6, ANG-(1–7) formation was significantly reduced in ACE2 knockout (KO) mice. However, at pH <6, formation of ANG-(1–7) in ACE2 KO mice was similar to that in wild-type (WT) mice, suggesting alternative peptidases for renal ANG II processing. Furthermore, the dual prolyl carboxypeptidase (PCP)-prolyl endopeptidase (PEP) inhibitor Z-prolyl-prolinal reduced ANG-(1–7) formation in ACE2 KO mice, while the ACE2 inhibitor MLN-4760 had no effect. Unlike the ACE2 KO mice, ANG-(1–7) formation from ANG II in PEP KO mice was not different from that in WT mice at any tested pH. However, at pH 5, this reaction was significantly reduced in kidneys and urine of PCP-depleted mice. In conclusion, results suggest that ACE2 metabolizes ANG II in the kidney at neutral and basic pH, while PCP catalyzes the same reaction at acidic pH. This is the first report demonstrating that renal ANG-(1–7) formation from ANG II is independent of ACE2. Elucidation of ACE2-independent ANG-(1–7) production pathways may have clinically important implications in patients with metabolic and renal disease.

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The Proinflammatory Response Induced by Wild-Type Yersinia pseudotuberculosis Infection Inhibits Survival of yop Mutants in the Gastrointestinal Tract and Peyer's Patches

Infection and Immunity ◽

10.1128/iai.74.3.1516-1527.2006 ◽

2006 ◽

Vol 74 (3) ◽

pp. 1516-1527 ◽

Cited By ~ 25

Author(s):

Lauren K. Logsdon ◽

Joan Mecsas

Keyword(s):

Yersinia Pseudotuberculosis ◽

Intestinal Inflammation ◽

Peyer's Patches ◽

Host Responses ◽

Infection Model ◽

Peyer’S Patches ◽

Wild Type ◽

Single Strain ◽

Infection Models ◽

Ifn Γ

ABSTRACT Single-strain infections and coinfections are frequently used to assess roles of virulence factors in infected tissues. After oral inoculation of mice, Yersinia pseudotuberculosis yopE and yopH mutants colonize the intestines and Peyer's patches in single-strain infections but fail to persist in competition with wild-type Y. pseudotuberculosis, indicating that these two infection models provide different insights into the roles of Yops. To determine how wild-type Y. pseudotuberculosis hinders yop mutant survival, yop mutant colonization and host responses were investigated in several different infection models that isolated specific features of wild-type Y. pseudotuberculosis infection. Infection with wild-type Y. pseudotuberculosis caused significantly more inflammation than yop mutants. Results from coinfections of gamma interferon (IFN-γ)−/− mice revealed that IFN-γ-regulated defenses target these mutants, suggesting that YopE and YopH protect Y. pseudotuberculosis from these defenses in BALB/c mice. We developed an oral-intraperitoneal infection model to evaluate the effects of spleen and liver colonization by Y. pseudotuberculosis on yop mutants in the intestines. Spleen and liver infection increased inflammation and decreased yop mutant survival in the intestines, indicating that infection of these organs has consequences in intestinal tissues. Finally, competition infections with Y. pseudotuberculosis mutants with various abilities to induce inflammation demonstrated that survival of the yopE, but not the yopH, mutant was consistently decreased in inflamed tissues. In summary, infection with Y. pseudotuberculosis in intestinal and systemic sites induces intestinal inflammation, which decreases yop mutant survival. Thus, competition studies with wild-type yersiniae reveal critical roles of Yops in combating host responses to a normal virulent infection.

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S19W, T27W, and N330Y mutations in ACE2 enhance SARS-CoV-2 S-RBD binding toward both wild-type and antibody-resistant viruses and its molecular basis

Signal Transduction and Targeted Therapy ◽

10.1038/s41392-021-00756-4 ◽

2021 ◽

Vol 6 (1) ◽

Author(s):

Fei Ye ◽

Xi Lin ◽

Zimin Chen ◽

Fanli Yang ◽

Sheng Lin ◽

...

Keyword(s):

Molecular Basis ◽

Structural Data ◽

Van Der Waals Interactions ◽

Side Chains ◽

Receptor Binding Domain ◽

Converting Enzyme ◽

Wild Type ◽

Angiotensin Converting Enzyme 2 ◽

Domain Protein ◽

Entry Inhibition

AbstractSARS-CoV-2 recognizes, via its spike receptor-binding domain (S-RBD), human angiotensin-converting enzyme 2 (ACE2) to initiate infection. Ecto-domain protein of ACE2 can therefore function as a decoy. Here we show that mutations of S19W, T27W, and N330Y in ACE2 could individually enhance SARS-CoV-2 S-RBD binding. Y330 could be synergistically combined with either W19 or W27, whereas W19 and W27 are mutually unbeneficial. The structures of SARS-CoV-2 S-RBD bound to the ACE2 mutants reveal that the enhanced binding is mainly contributed by the van der Waals interactions mediated by the aromatic side-chains from W19, W27, and Y330. While Y330 and W19/W27 are distantly located and devoid of any steric interference, W19 and W27 are shown to orient their side-chains toward each other and to cause steric conflicts, explaining their incompatibility. Finally, using pseudotyped SARS-CoV-2 viruses, we demonstrate that these residue substitutions are associated with dramatically improved entry-inhibition efficacy toward both wild-type and antibody-resistant viruses. Taken together, our biochemical and structural data have delineated the basis for the elevated S-RBD binding associated with S19W, T27W, and N330Y mutations in ACE2, paving the way for potential application of these mutants in clinical treatment of COVID-19.

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Evolution of ACE2 and SARS-CoV-2 Interplay Across 247 Vertebrates

10.1101/2021.01.28.428568 ◽

2021 ◽

Author(s):

Tao Zhang ◽

Qunfu Wu ◽

Yicheng Ma ◽

Wenjing Liu ◽

Chengang Zhou ◽

...

Keyword(s):

Acid Sites ◽

Rapid Expansion ◽

Structural Basis ◽

S Protein ◽

Angiotensin Converting Enzyme 2 ◽

Evolutionary Pattern ◽

High Affinity ◽

Treatment And Prevention ◽

Infection Models ◽

Potential Benefits

AbstractSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cause the most serious pandemics of Coronavirus Disease 2019 (COVID-19), which threatens human health and public safety. SARS-CoV-2 spike (S) protein uses angiotensin-converting enzyme 2 (ACE2) as recognized receptor for its entry into host cell that contributes to the infection of SARS-CoV-2 to hosts. Using computational modeling approach, this study resolved the evolutionary pattern of bonding affinity of ACE2 in 247 jawed vertebrates to the spike (S) protein of SARS-CoV-2. First, high-or-low binding affinity phenotype divergence of ACE2 to the S protein of SARS-CoV-2 has appeared in two ancient species of jawed vertebrates, Scyliorhinus torazame (low-affinity, Chondrichthyes) and Latimeria chalumnae (high-affinity, Coelacanthimorpha). Second, multiple independent affinity divergence events recur in fishes, amphibians-reptiles, birds, and mammals. Third, high affinity phenotypes go up in mammals, possibly implying the rapid expansion of mammals might accelerate the evolution of coronaviruses. Fourth, we found natural mutations at eight amino acid sites of ACE2 can determine most of phenotype divergences of bonding affinity in 247 vertebrates and resolved their related structural basis. Moreover, we also identified high-affinity or low-affinity-associated concomitant mutation group.The group linked to extremely high affinity may provide novel potentials for the development of human recombinant soluble ACE2 (hrsACE2) in treating patients with COVID-19 or for constructing genetically modified SARS-CoV-2 infection models promoting vaccines studies. These findings would offer potential benefits for the treatment and prevention of SARS-CoV-2.

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Antibody binding and ACE2 binding inhibition is significantly reduced for the Omicron variant compared to all other variants of concern

10.1101/2021.12.30.21267519 ◽

2022 ◽

Author(s):

Daniel Junker ◽

Matthias Becker ◽

Teresa Wagner ◽

Philipp D Kaiser ◽

Sandra Maier ◽

...

Keyword(s):

Angiotensin Converting Enzyme ◽

Antibody Binding ◽

Large Cohort ◽

Converting Enzyme ◽

Wild Type ◽

Immune Control ◽

Angiotensin Converting Enzyme 2 ◽

Igg Binding ◽

Binding Inhibition ◽

Rapid Emergence

The rapid emergence of the Omicron variant and its large number of mutations has led to its classification as a variant of concern (VOC) by the WHO. Initial studies on the neutralizing response towards this variant within convalescent and vaccinated individuals have identified substantial reductions. However many of these sample sets used in these studies were either small, uniform in nature, or were compared only to wild-type (WT) or, at most, a few other VOC. Here, we assessed IgG binding, (Angiotensin-Converting Enzyme 2) ACE2 binding inhibition, and antibody binding dynamics for the omicron variant compared to all other VOC and variants of interest (VOI), in a large cohort of infected, vaccinated, and infected and then vaccinated individuals. While omicron was capable of binding to ACE2 efficiently, antibodies elicited by infection or immunization showed reduced IgG binding and ACE2 binding inhibition compared to WT and all VOC. Among vaccinated samples, antibody binding responses towards omicron were only improved following administration of a third dose. Overall, our results identify that omicron can still bind ACE2 while pre-existing antibodies can bind omicron. The extent of the mutations appear to inhibit the development of a neutralizing response, and as a result, omicron remains capable of evading immune control.

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A porcine ex vivo lung perfusion model to investigate bacterial pathogenesis

10.1101/708503 ◽

2019 ◽

Author(s):

Amy Dumigan ◽

Marianne Fitzgerald ◽

Joana Sá Pessoa Graca Santos ◽

Umar Hamid ◽

Cecilia M. O’Kane ◽

...

Keyword(s):

Inflammatory Responses ◽

Respiratory Infections ◽

Ex Vivo ◽

Macrophage Polarization ◽

Lung Perfusion ◽

Respiratory Pathogens ◽

Wild Type ◽

Pathogenic Potential ◽

Ex Vivo Lung Perfusion ◽

Infection Models

ABSTRACTThe use of animal infection models is essential to understand microbial pathogenesis and to develop and test treatments. Insects, and 2D and 3D tissue models are increasingly being used as surrogate for mammalian models. However, there are concerns whether these models recapitulate the complexity of host-pathogen interactions. Here, we developed the ex vivo lung perfusion (EVLP) model of infection using porcine lungs to investigate Klebsiella pneumoniae-triggered pneumonia as model of respiratory infections. The porcine EVLP model recapitulates features of K. pneumoniae-induced pneumonia lung injury. This model is also useful to assess the pathogenic potential of K. pneumoniae as we observed that the attenuated Klebsiella capsule mutant strain caused less pathological tissue damage with a concomitant decrease in the bacterial burden compare to lungs infected with the wild type. The porcine EVLP model allows assessment of inflammatory responses following infection; similar to the mouse pneumonia model, we observed an increase of il-10 in the lungs infected with the wild type and an increase of ifn-γ in lungs infected with the capsule mutant. This model also allows monitoring phenotypes at the single-cell level. Wild-type K. pneumoniae skews macrophages towards an M2-like state. In vitro experiments probing pig bone marrow-derived macrophages uncovered the role of the M2 transcriptional factor STAT6, and that Klebsiella-induced il10 expression is controlled by p38 and ERK. Klebsiella-induced macrophage polarization is dependent on the capsule. Altogether, this study support the utility of the EVLP model using pig lungs as platform to investigate the infection biology of respiratory pathogens.IMPORTANCEThe implementation of infection models that approximate human disease is essential to understand infections and for testing new therapies before they enter into clinical stages. Rodents are used in most of pre-clinical studies, although the differences between mouse and man have fuelled the conclusion that murine studies are unreliable predictors of human outcomes. Here, we have developed a whole lung porcine model of infection using the established ex vivo lung perfusion (EVLP) system established to re-condition human lungs for transplant. As a proof-of-principle, we provide evidence demonstrating that infection of the porcine EVLP with the human pathogen K. pneumoniae recapitulates the known features of Klebsiella-triggered pneumonia. Moreover, our data revealed the porcine EVLP model is useful to reveal features of the virulence of K. pneumoniae including the manipulation of immune cells. Altogether, this study supports the utility of the EVLP model using pig lungs as surrogate host for assessing respiratory infections.

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Association of PAX3 and TMTC2 gene polymorphism with the face morphology change after excision of skin tumors

Vojnosanitetski pregled ◽

10.2298/vsp180613128m ◽

2020 ◽

Vol 77 (6) ◽

pp. 631-636

Author(s):

Sasa Milicevic ◽

Zvonko Magic ◽

Gordana Supic ◽

Aleksandar Jevtic ◽

Stevo Jovandic ◽

...

Keyword(s):

Skin Tumor ◽

Human Populations ◽

Wild Type ◽

Tumor Excision ◽

Pax3 Gene ◽

Significant Difference ◽

The Face ◽

Suture Closure ◽

The Impact ◽

Face Morphology

Background/Aim. The group of genes, known as PAX (paired box), has a great role in organogenesis, as well as in maintaining the normal function of certain cells after the birth. In addition to these genes, the impact on the organogenesis, at the cellular level, has a transmembrane tetratricopeptid group of genes (TMTC). The term polymorphism in the human genome implies variations in the hereditary basis that occur in human populations, the presence of two or more different alleles of one genome in the population. The aim of the work was to determine whether there is an association of PAX3 and TMTC2 genes polymorphism with changes of the face morphology after skin tumor excision and direct suture closure. Methods. The study included 130 patients of both sexes, older than 50 years, with the medical indication for the elliptical surgical excision of the skin tumor. DNA was isolated from 5 mL of peripheral blood. Gene polymorphisms were analyzed with pre-designed single nucleotide polymorphisms (SNP) assays, by allelic discrimination method on REAL-TIME apparatus. The patients were subjected to a laser scanning preoperatively, and 7 and 90 days postoperatively, in order to obtain x, y and z coordinates of 5 cephalometric points on the face, which determined the shape of the medial cheek region. The shape of the medial cheek region, as well as the coordinates of 5 cepahlometric points, were compared among genotypes of both genes preoperatively, as well as 7 days and 90 days postoperatively. Results. A statistically significant difference in the shape of the medial cheek region between wildtype and mutant of PAX3 gene was found preoperatively, while the statistically significant difference in the shape of the medial cheek region was not found between wild-type and heterozygote, nor between wild-type and heterozygote and mutant of PAX3 gene, nor among genotypes of TMTC2 gene. Seven days and 90 days postoperatively, there were no statistically significant differences in the shape of the examined region among genotypes of both genes. Conclusion. Polymorphisms of PAX3 and TMTC2 genes are not associated with the change in the face morphology after the skin tumor excision and direct suture closure of the defect.

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CRUNC: a cryopreservation method for unencapsulated gemmae of Marchantia polymorpha

PeerJ ◽

10.7717/peerj.10174 ◽

2020 ◽

Vol 8 ◽

pp. e10174

Author(s):

Hitomi Takahashi ◽

Yutaka Kodama

Keyword(s):

Liquid Nitrogen ◽

Marchantia Polymorpha ◽

Wild Type ◽

High Recovery ◽

Simple Method ◽

High Recovery Rate ◽

Genetic Modifications ◽

Transgenic Lines ◽

Cryopreservation Method

Genetic modifications such as mutation and transformation are powerful tools to study the function of genes and proteins in the model liverwort Marchantia polymorpha, but maintaining the resulting germplasm requires a practical, reliable method. Cryopreservation methods allow researchers to maintain mutant and transgenic lines of M. polymorpha. To date, two methods have been developed for cryopreservation of M. polymorpha gemmae: in the first method, unencapsulated gemmae are stored in liquid nitrogen at −196 °C, and in the second method, encapsulated gemmae are stored in liquid nitrogen at −196 °C or a deep freezer at −80 °C. In the present study, we developed a simple method named CRUNC (cr yopreservation of un en c apsulated gemmae), which can be used to store unencapsulated, dried gemmae of wild-type and transgenic M. polymorpha lines in liquid nitrogen and in freezers at −80 °C and −20 °C. Using the CRUNC method, we observed a high recovery rate (as high as 100%) and successful long-term (5 months) storage of the gemmae. Therefore, the CRUNC method is practical for maintaining valuable M. polymorpha germplasm.

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