The rockefeller university

Microbes use signaling factors for intraspecies and interspecies communications. While many intraspecies signaling factors have been found and characterized, discovery of factors for interspecies communication is lagging behind. To facilitate the discovery of such factors, we explored the potential of a mixed microbial culture (MMC) derived from wheatgrass, in which heterogeneity of this microbial community might elicit signaling factors for interspecies communication. The stability of Wheatgrass MMC in terms of community structure and metabolic output was first characterized by 16S ribosomal RNA amplicon sequencing and liquid chromatography/mass spectrometry (LC/MS), respectively. In addition, detailed MS analyses led to the identification of 12hydroxystearic acid (12-HSA) as one of the major metabolites produced by Wheatgrass MMC. Stereochemical analysis revealed that Wheatgrass MMC produces mostly the (R)-isomer, although a small amount of the (S)isomer was also observed. Furthermore, 12-HSA was found to modulate planktonic growth and biofilm formation of various marine bacterial strains. The current study suggests that naturally derived MMCs could serve as a simple and reproducible platform to discover potential signaling factors for interspecies communication. In addition, the study indicates that hydroxylated long-chain fatty acids, such as 12-HSA, may constitute a new class of interspecies signaling factors.

Turnover of cellular organelles, including endoplasmic reticulum (ER) and mitochondria, is orchestrated by an efficient cellular surveillance system. We have identified a mechanism for dual regulation of ER and mitochondria under stress. It is known that AMFR, an ER E3 ligase and ER-associated degradation (ERAD) regulator, degrades outer mitochondrial membrane (OMM) proteins, MFNs (mitofusins), via the proteasome and triggers mitophagy. We show that destabilized mitochondria are almost devoid of the OMM and generate "mitoplasts". This brings the inner mitochondrial membrane (IMM) in the proximity of the ER. When AMFR levels are high and the mitochondria are stressed, the reticulophagy regulatory protein RETREG1 participates in the formation of the mitophagophore by interacting with OPA1. Interestingly, OPA1 and other IMM proteins exhibit similar RETREG1-dependent autophagosomal degradation as AMFR, unlike most of the OMM proteins. The "mitoplasts" generated are degraded by reticulo-mito-phagy - simultaneously affecting dual organelle turnover.

There is an urgent need for effective therapeutic interventions against SARS-CoV-2, including new variants that continue to arise. Neutralizing monoclonal antibodies have shown promise in clinical studies. We investigated the therapeutic efficacy of a combination of two potent monoclonal antibodies, C135-LS and C144-LS that carry half-life extension mutations, in the rhesus macaque model of COVID-19. Twelve young adult macaques (three groups of four animals) were inoculated intranasally and intra-tracheally with a high dose of SARS-CoV-2 and 24 hours later, treated intravenously with a high (40 mg/kg) or low (12 mg/kg) dose of the C135-LS and C144-LS antibody combination, or a control monoclonal antibody. Animals were monitored for 7 days. Compared to the control animals, animals treated with either dose of the anti-SARS-CoV-2 antibodies showed similarly improved clinical scores, lower levels of virus replication in upper and lower respiratory tract, and significantly reduced interstitial pneumonia, as measured by comprehensive lung histology. In conclusion, this study provides proof-of-concept in support of further clinical development of these monoclonal antibodies against COVID-19 during early infection. Author summary Monoclonal antibodies that neutralize SARS-CoV-2 have shown promise in treating recently infected individuals who are at high risk of progression to severe COVID-19 disease. Although several monoclonal antibodies are currently being used in the clinic, there is an ongoing need to develop additional antibodies. The ideal monoclonal antibodies, or combinations, should be potent and durable, and maintain activity against emerging viral variants. In this study, we tested a combination of two potent monoclonal antibodies, C135-LS and C-144-LS, engineered to have long half-lives, in the macaque model of SARS-CoV-2 infection. Animals treated early after infection fared better than placebo-treated controls, manifesting fewer clinical signs, less virus replication in the respiratory tract, and reduced lung inflammation. These promising data support clinical testing of these monoclonal antibodies in humans and further development of similar antibody-based prophylactic and therapeutic strategies.

More than one year after its inception, the coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains difficult to control despite the availability of several working vaccines. Progress in controlling the pandemic is slowed by the emergence of variants that appear to be more transmissible and more resistant to antibodies(1,2). Here we report on a cohort of 63 individuals who have recovered from COVID-19 assessed at 1.3, 6.2 and 12 months after SARS-CoV-2 infection, 41% of whom also received mRNA vaccines(3,4). In the absence of vaccination, antibody reactivity to the receptor binding domain (RBD) of SARS-CoV-2, neutralizing activity and the number of RBD-specific memory B cells remain relatively stable between 6 and 12 months after infection. Vaccination increases all components of the humoral response and, as expected, results in serum neutralizing activities against variants of concern similar to or greater than the neutralizing activity against the original Wuhan Hu-1 strain achieved by vaccination of naive individuals(2,5-8). The mechanism underlying these broad-based responses involves ongoing antibody somatic mutation, memory B cell clonal turnover and development of monoclonal antibodies that are exceptionally resistant to SARS-CoV-2 RBD mutations, including those found in the variants of concern(4,9). In addition, B cell clones expressing broad and potent antibodies are selectively retained in the repertoire over time and expand markedly after vaccination. The data suggest that immunity in convalescent individuals will be very long lasting and that convalescent individuals who receive available mRNA vaccines will produce antibodies and memory B cells that should be protective against circulating SARS-CoV-2 variants.

A mainstay of personal protective equipment during the coronavirus disease 2019 pandemic is the N95 filtering facepiece respirator. N95 respirators are commonly used to protect healthcare workers from respiratory pathogens, including the novel coronavirus severe acute respiratory syndrome coronavirus 2, and are increasingly employed by other frontline workers and the general public. Under routine circumstances, these masks are disposable, single-use items, but extended use and reuse practices have been broadly enacted to alleviate critical supply shortages during the coronavirus disease 2019 pandemic. Although extended-time single use presents a low risk of pathogen transfer, repeated donning and doffing of potentially contaminated masks presents increased risk of pathogen transfer. Therefore, efficient and safe decontamination methods for N95 masks are needed to reduce the risk of reuse and mitigate local supply shortages. Here, we review the available literature concerning use of germicidal ultraviolet-C (UV-C) light to decontaminate N95 masks. We propose a practical method for repeated point-of-use decontamination using commercially available UV-C cross-linker boxes from molecular biology laboratories to expose each side of the mask to 800-1200 mJ/cm(2) of UV-C. We measure the dose that penetrated to the interior of the respirators and model the potential germicidal action on coronaviruses. Our experimental results, in combination with modeled data, suggest that such a UV-C treatment cycle should induce a >3-log-order reduction in viral bioburden on the surface of the respirators and a 2-log-order reduction throughout the interior. We find that a dose 50-fold greater does not impair filtration or fit of 3M 8210 N95 masks, indicating that decontamination can be performed repeatedly. As such, UV-C germicidal irradiation is a practical strategy for small-scale point-of-use decontamination of N95s.

Multidrug-resistant Streptococcus pneumoniae emerge through the modification of core genome loci by interspecies homologous recombinations, and acquisition of gene cassettes. Both occurred in the otherwise contrasting histories of the antibiotic-resistant S. pneumoniae lineages PMEN3 and PMEN9. A single PMEN3 Glade spread globally, evading vaccine-induced immunity through frequent serotype switching, whereas locally circulating PMEN9 clades independently gained resistance. Both lineages repeatedly integrated Tn916-type and Tn1207.1-type elements, conferring tetracycline and macrolide resistance, respectively, through homologous recombination importing sequences originating in other species. A species-wide dataset found over 100 instances of such interspecific acquisitions of resistance cassettes and flanking homologous arms. Phylodynamic analysis of the most commonly sampled Tn1207.1-type insertion in PMEN9, originating from a commensal and disrupting a competence gene, suggested its expansion across Germany was driven by a high ratio of macrolide-to-beta-lactam consumption. Hence, selection from antibiotic consumption was sufficient for these atypically large recombinations to overcome species boundaries across the pneumococcal chromosome.

Congenital heart disease (CHD) is the most common congenital malformation and the leading cause of mortality therein. Genetic etiologies contribute to an estimated 90% of CHD cases, but so far, a molecular diagnosis remains unsolved in up to 55% of patients. Copy number variations and aneuploidy account for similar to 23% of cases overall, and high-throughput genomic technologies have revealed additional types of genetic variation in CHD. The first CHD risk genotypes identified through high-throughput sequencing were de novo mutations, many of which occur in chromatin modifying genes. Murine models of cardiogenesis further support the damaging nature of chromatin modifying CHD mutations. Transmitted mutations have also been identified through sequencing of population scale CHD cohorts, and many transmitted mutations are enriched in cilia genes and Notch or VEGF pathway genes. While we have come a long way in identifying the causes of CHD, more work is required to end the diagnostic odyssey for all CHD families. Complex genetic explanations of CHD are emerging but will require increasingly sophisticated analysis strategies applied to very large CHD cohorts before they can come to fruition in providing molecular diagnoses to genetically unsolved patients. In this review, we discuss the genetic architecture of CHD and biological pathways involved in its pathogenesis.

Autosomal recessive (AR) STAT1 deficiency is a severe inborn error of immunity disrupting cellular responses to type I, II, and III IFNs, and IL-27, and conferring a predisposition to both viral and mycobacterial infections. We report the genetic, immunological, and clinical features of an international cohort of 32 patients from 20 kindreds: 24 patients with complete deficiency, and 8 patients with partial deficiency. Twenty-four patients suffered from mycobacterial disease (bacillus Calmette-Guerin = 13, environmental mycobacteria = 10, or both in 1 patient). Fifty-four severe viral episodes occurred in sixteen patients, mainly caused by Herpesviridae viruses. Attenuated live measles, mumps, and rubella and/or varicella zoster virus vaccines triggered severe reactions in the five patients with complete deficiency who were vaccinated. Seven patients developed features of hemophagocytic syndrome. Twenty-one patients died, and death was almost twice as likely in patients with complete STAT1 deficiency than in those with partial STAT1 deficiency. All but one of the eight survivors with AR complete deficiency underwent hematopoietic stem cell transplantation. Overall survival after hematopoietic stem cell transplantation was 64%. A diagnosis of AR STAT1 deficiency should be considered in children with mycobacterial and/or viral infectious diseases. It is important to distinguish between complete and partial forms of AR STAT1 deficiency, as their clinical outcome and management differ significantly.

Using self-organizing human models of gastrulation, we previously showed that (1) BMP4 initiates the cascade of events leading to gastrulation, (2) BMP4 signal reception is restricted to the basolateral domain, and (3) in a human-specific manner, BMP4 directly induces the expression of NOGGIN. Here, we report the surprising discovery that in human epiblasts, NOGGIN and BMP4 were secreted into opposite extracellular spaces. Interestingly, apically presented NOGGIN could inhibit basally delivered BMP4. Apically imposed microfluidic flow demonstrated that NOGGIN traveled in the apical extracellular space. Our co-localization analysis detailed the endocytotic route that trafficked NOGGIN from the apical space to the basolateral intercellular space where BMP4 receptors were located. This apical-basal transcytosis was indispensable for NOGGIN inhibition. Taken together, the segregation of activator/inhibitor into distinct extracellular spaces challenges classical views of morphogen movement. We propose that the transport of morphogen inhibitors regulates the spatial availability of morphogens during embryogenesis.

Staphylococcus saprophyticus is a common pathogen of the urinary tract, a heavy metal-rich environment, but information regarding its heavy metal resistance is unknown. We investigated 422 S. saprophyticus isolates from human infection and colonization/contamination, animals, and environmental sources for resistance to copper, zinc, arsenic, and cadmium using the agar dilution method. To identify the genes associated with metal resistance and assess possible links to pathogenicity, we accessed the whole-genome sequence of all isolates and used in silico and pangenome-wide association approaches. The MIC values for copper and zinc were uniformly high (1,600mg/liter). Genes encoding copper efflux pumps (copA, copB, copZ, mco, and csoR) and zinc transporters (zinT, czrAB, znuBC, and zur) were abundant in the population (20 to 100%). Arsenic and cadmium showed various susceptibility levels. Genes encoding the ars operon (arsRDABC), an ABC transporter and a two-component permease, were linked to resistance to arsenic (MICs >= 1,600mg/liter; 14% [58/422]; P < 0.05). At least three cad genes (cadA or cadC and cadD-cadX or czrC) and genes encoding multidrug efflux pumps and hyperosmoregulation in acidified conditions were associated with resistance to cadmium (MICs >= 200mg/liter; 20% [85/422]; P < 0.05). These resistance genes were frequently carried by mobile genetic elements. Resistance to arsenic and cadmium were linked to human infection and a clonal lineage originating in animals (P < 0.05). Altogether, S. saprophyticus was highly resistant to heavy metals and accumulated multiple metal resistance determinants. The highest arsenic and cadmium resistance levels were associated with infection, suggesting resistance to these metals is relevant for S. saprophyticus pathogenicity.