Impact of COVID-19 Pandemic on Ambulatory and Operating Room Rhinology Practice in the US.

INTRODUCTION: The pandemic caused by the novel coronavirus virus has altered all facets of clinical practice in the United States. The goal of this study is to better understand the impact of COVID-19 on rhinologic ambulatory and operative practice. METHODS: A 27-item survey to assess these objectives was created and approved by the Division of Rhinology faculty at Rush University Medical Center in April 2020. The survey was then distributed to rhinologists in a web based format via www.surveymonkey.com from April 10 through April 23, 2020. RESULTS: A total of 277 U.S based rhinologists responded to the survey (23.04%). The most common practice types were single specialty private (44.9%) and academic (24.6%). 90.2% practice in a state under a shelter in place order. Comparing pre-COVID baseline to during-COVID, there was statistically significant reduction in the number of patients of seen daily in clinic (p < 0.001). The number of nasal endoscopies in the office and surgical procedures fell dramatically. Overall, 5 respondent rhinologists have been infected with COVID-19 and 27 have been furloughed. CONCLUSION: COVID-19 has drastically affected rhinologic practice. There is a dramatic reduction of in person care in the office setting and surgical management of sinonasal and skull base disease. Enhanced PPE is being used in only half of potentially aerosolizing procedures which represents an area of further education. Novel approaches such as use of virtual encounters and point of care testing should be considered as options to facilitate care.

AoHapB, AoHapC and AoHapE, subunits of the Aspergillus oryzae CCAAT-binding complex, are functionally interchangeable with the corresponding subunits in Aspergillus nidulans.

Two genes, AohapB and AohapE, encoding subunits of the Aspergillus oryzae CCAAT-binding complex were cloned and sequenced. The polypeptides encoded by AohapB and AohapE were expressed in Escherichia coli and used to reconstitute a DNA-binding complex with recombinant AoHapC. The DNA-binding activity was observed only in the presence of all three subunits, indicating that AoHapB, AoHapE and AoHapC are essential for CCAAT-binding. Furthermore, introduction of the AohapB, AohapC and AohapE genes into the A. nidulans hapB delta, hapC delta and hapE delta strains, respectively, revealed that the A. oryzae Hap subunits are functionally interchangeable with the corresponding subunits in A. nidulans.

An Aspergillus oryzae CCAAT-binding protein, AoCP, is involved in the high-level expression of the Taka-amylase A gene.

Aspergillus oryzae contains a nuclear protein designated AoCP, which binds specifically to a CCAAT sequence in the promoter region of the A. oryzae Taka-amylase A gene. A gene encoding a homologue of Aspergillus nidulans HAPC, a subunit of the A. nidulans CCAAT binding complex, was isolated from A. oryzae and designated AohapC. AoHAPC comprises 215 amino acids and shows 84% identity to A. nidulans HAPC. Transformation of the A. nidulans hapC deletion strain with the AohapC gene restored the CCAAT binding activity, resulting in both enhancement of taa gene expression and complementation for the poor growth phenotype of this strain. Furthermore, introduction of the AohapC gene also restored the expression of the A. nidulans eglA gene, encoding an endo-beta-1,4-glucanase, in the deletion strain. These results indicate functional interchangeability of the genes from two species.

AnCF, the CCAAT binding complex of Aspergillus nidulans, contains products of the hapB, hapC, and hapE genes and is required for activation by the pathway-specific regulatory gene amdR.

CCAAT binding factors (CBFs) positively regulating the expression of the amdS gene (encoding acetamidase) and two penicillin biosynthesis genes (ipnA and aatA) have been previously found in Aspergillus nidulans. The factors were called AnCF and PENR1, respectively. Deletion of the hapC gene, encoding a protein with significant similarity to Hap3p of Saccharomyces cerevisiae, eliminated both AnCF and PENR1 binding activities. We now report the isolation of the genes hapB and hapE, which encode proteins with central regions of high similarity to Hap2p and Hap5p of S. cerevisiae and to the CBF-B and CBF-C proteins of mammals. An additional fungus-specific domain present in HapE was revealed by comparisons with the homologs from S. cerevisiae, Neurospora crassa, and Schizosaccharomyces pombe. The HapB, HapC, and HapE proteins have been shown to be necessary and sufficient for the formation of a CCAAT binding complex in vitro. Strains with deletions of each of the hapB, hapC, and hapE genes have identical phenotypes of slow growth, poor conidiation, and reduced expression of amdS. Furthermore, induction of amdS by omega amino acids, which is mediated by the AmdR pathway-specific activator, is abolished in the hap deletion mutants, as is growth on gamma-aminobutyric acid as a sole nitrogen or carbon source. AmdR and AnCF bind to overlapping sites in the promoters of the amdS and gatA genes. It is known that AnCF can bind independently of AmdR. We suggest that AnCF binding is required for AmdR binding in vivo.

The Aspergillus nidulans CCAAT-binding factor AnCP/AnCF is a heteromeric protein analogous to the HAP complex of Saccharomyces cerevisiae.

The Aspergillus nidulans hapC gene was expressed as a fusion protein with MalE or glutathione-S-transferase (GST) in Escherichia coli, and used for the purification of HapC and the preparation of anti-HapC antiserum. The CCAAT-binding factor AnCP/AnCF contains a component with an approximate molecular mass of 32 kDa that cross-reacts with the antibody. The MalE-HapC fusion protein was able to replace authentic HapC in AnCP when incubated under appropriate conditions. Furthermore, reconstitution experiments with recombinant HapC, yHAP2 and yHAP5 polypeptides showed that all three polypeptides were required for the assembly of a complex capable of binding to CCAAT-containing taaG2 promoter DNA. The relationship between AnCP/AnCF and the Saccharomyces cerevisiae HAP complex is discussed.

The hapC gene of Aspergillus nidulans is involved in the expression of CCAAT-containing promoters.

The 5' regulatory region of the amdS gene of Aspergillus nidulans, which encodes an acetamidase required for growth on acetamide as a carbon and nitrogen source, contains a CCAAT sequence which is required for setting the basal level of amdS expression. Mobility shift studies have identified a factor in A. nidulans nuclear extracts which binds to this CCAAT sequence. In Saccharomyces cerevisiae the HAP3 gene encodes one component of a multisubunit complex that binds CCAAT sequences. A search of the EMBL and SwissProt databases has revealed an A. nidulans sequence with significant homology to the HAP3 gene adjacent to the previously cloned regulatory gene amdR. Sequencing of the remainder of this region has confirmed the presence of a gene, designated hapC, with extensive homology to HAP3. The predicted amino acid sequence of HapC shows extensive identity to HAP3 in the central conserved domain, but shows little conservation in the flanking sequences. A haploid carrying a hapC deletion has been created and is viable, but grows poorly on all media tested. This null mutant grows especially slowly on acetamide as a sole carbon and nitrogen source, indicating that hapC plays a role in amdS expression. In agreement with this notion, it has been shown that the hapC deletion results in reduced levels of expression of an amdS::lacZ reporter gene and this effect is particularly evident under conditions of carbon limitation. Nuclear extracts prepared from the hapC deletion mutant show no CCAAT binding activity to the amdS or gatA promoters, indicating that hapC may encode a component of the complex binding at this sequence.