Histopathology of the Tongue in a Hamster Model of COVID-19.

Objective With altered sense of taste being a common symptom of coronavirus disease 2019 (COVID-19), our objective was to investigate the presence and distribution of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) within the tongue over the course of infection. Methods Golden Syrian hamsters were inoculated intranasally with SARS-CoV-2 and tongues were collected at 2, 3, 5, 8, 17, 21, 35, and 42 days post-infection (dpi) for analysis. In order to test for gross changes in the tongue, the papillae of the tongue were counted. Paraffin-embedded thin sections of the tongues were labeled for the presence of SARS-CoV-2 antigen. Results There was no difference in fungiform or filiform papillae density throughout the course of infection. SARS-CoV-2 antigen was observed in the circumvallate papillae taste buds (3-35 dpi) and autonomic ganglia (5-35 dpi), as well as in the serous and mucous salivary glands of the posterior tongue (2-42 dpi). Conclusion The presence and distribution of SARS-CoV-2 suggest that the virus could cause taste disturbance by infecting the circumvallate taste buds. This effect could be exacerbated by a diminished secretion of saliva caused by infection of the serous salivary glands and the autonomic ganglia which innervate them.

Large Angiomatous Nasal Polyp Presents With Epistaxis Imitating Juvenile Nasopharyngeal Angiofibroma.

An angiomatous nasal polyp is a rare subtype of sinonasal polyp that is commonly found in the middle meatus and characterized by the presence of blood vessels within polyp tissue. It is a benign lesion but is prone to misdiagnosis as a malignant tumor because it typically grows larger and is more vascular than other types of polyps. In this report, a 16-year-old male with no significant past medical history presents with a six-month history of epistaxis and progressive nasal obstruction. Examination of the oral cavity showed a centrally located soft palate mass. CT maxillofacial with contrast showed a hypervascular 3.4 x 4.7 x 6.1 cm mass in the nasal cavity extending through the nasal choanae and down to the level of the tongue. MRI showed a heterogenous polypoid mass originating from the left middle meatus vs. nasal cavity, with characteristics favoring an aggressive tumor. The patient was taken for interventional radiology (IR) embolization and nasal endoscopy. Biopsy showed the left nasal mass contained granulation tissue and the palatal mass consisted of necrotic tissue. He was taken for second-stage endoscopic sinus surgery with plans for extensive reconstruction if necessary. Extensive polyposis was found without gross evidence of an aggressive tumor. The anterior polyposis was debulked and the polyp was cut at its root to allow for removal of the nasopharyngeal/oropharyngeal portion through the mouth. He was able to be discharged on the same day and his postoperative recovery was uncomplicated. Angiomatous nasal polyps are uncommon, share features of aggressive tumors on imaging, and require angiography and biopsy to safely rule out malignancy. Endoscopic surgical resection typically results in good outcomes and low recurrence rates.

A Case of Radiologically Compatible Chronic Lymphocytic Inflammation With Pontine Perivascular Enhancement Responsive to Steroids (CLIPPERS) With Demyelinating Lesions.

Chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids (CLIPPERS) is a recently identified diagnosis that can cause a variety of severe symptoms, including ataxia, dysarthria, diplopia, paraparesis, and vertigo. These symptoms rarely present in isolation but often accompany one another in various combinations. Magnetic Resonance Imaging (MRI) of the brain is critical for making the diagnosis and typically reveals scattered enhancement within the pons and adjacent structures. The syndrome responds well to high-dose steroids, and maintenance therapy is required to prevent a recurrence. In this report, we present a case of a 62-year-old man who developed CLIPPERS syndrome. The patient presented with hemiparesis and dysarthria, which developed over four months and then acutely worsened within 24 hours. After diagnosing CLIPPERS, the patient was placed on high-dose steroids and experienced rapid clinical improvement, as well as improvement on repeat MRI. The patient's treatment was complicated by an incidental diagnosis of tuberculosis, which required simultaneous management with isoniazid.

Two Stacked Coins Mimicking an Esophageal Button Battery: A Case Presentation and Review of the Literature.

As button battery (BB) ingestion has become a popular topic with growing public awareness in recent years, pediatric otolaryngologists maintain a high index of suspicion for this diagnosis. Several recent reports have revealed the possibility for benign objects to masquerade as BBs, such as two coins stacked together or a coin with different metals in concentric rings. A 4-year-old female presented to the ED after unwitnessed ingestion of a foreign body. The child was reportedly seen playing with her sister's coin collection prior to the acute onset of drooling and dysphagia. She was vitally stable and without any shortness of breath, stridor, or wheezing. Plain film X-ray revealed a round, metallic object with a double density on the frontal view and beveled step-off on the lateral view at the level of the thoracic inlet. Due to high radiographic concern for BB ingestion, the patient was taken emergently to the operating room for a rigid esophagoscopy. A metallic object was seen at the thoracic inlet and removed with Magill forceps. The object was found to be two coins stuck together, with a smaller coin in the center of a larger coin mimicking the shape of a BB. The patient was discharged the next day without complication. This case highlights stacked coins as a radiologic masquerade for BBs as well as the emphasis on prompt esophagoscopy for both identification and removal. Radiographic densities alone cannot be relied upon to distinguish BBs from more innocuous objects, and esophagoscopy remains the mainstay of management for pediatric esophageal foreign bodies.

Mycobacterium abscessus Causes Highly Resistant Infection as a Breast Abscess.

Mycobacterium abscessus is an increasing threat to public health due to its multi-drug resistance and increasing prevalence. The pathogen most commonly causes chronic respiratory infections, but it may also invade locally through the skin and soft tissue damage caused by trauma, piercings, or tattoos. A 58-year-old African American female presented with a five-month history of recurrent abscesses in the right breast. She had previously been treated with doxycycline and ceftriaxone injections at an outside clinic with minimal improvement. Following incision and drainage, cultures and susceptibilities showed M. abscessus infection with a high level of drug resistance. Due to financial barriers, the patient received a suboptimal antibiotic regimen and required multiple surgical procedures, resulting in only temporary wound healing. At the time of this report, the patient is recovering from her fourth incision and drainage, with cultures and susceptibilities pending and discussions of a total mastectomy. M. abscessus is a highly resistant bacteria capable of causing skin and soft tissue infections of the breast. Such infections may occur without an inciting event and require extensive surgical and antimicrobial management.

Turbinate hypertrophy following COVID-19 infection masks choanal atresia in adult.

A 34-year-old female 4 weeks post COVID-19 infection was diagnosed with inferior turbinate hypertrophy and a deviated nasal septum. The patient underwent inferior turbinate bone resection and nasal septoplasty with minimal improvement in nasal obstruction. Upon reevaluation, unilateral choanal atresia was discovered. Subsequent repair of the choanal atresia resolved all complaints of nasal obstruction. This unique presentation of a missed diagnosis of choanal atresia contributes to the idea that a variety of conditions may precede and at times delay the discovery of choanal atresia in adults.

An 18-Year-Old Female Experiences Unilateral Vocal Cord Paralysis during Mild COVID-19 Infection.

The COVID-19 pandemic has shown with certainty that SARS-CoV-2 can cause a variety of clinical findings, with some of the most notable being lasting chemosensory changes. Severe infections with SARS-CoV-2 can also lead to a variety of complications. For example, vocal cord paralysis can be caused by trauma sustained during intubation, which is a necessary procedure for many severe cases. Rarely, SARS-CoV-2 related vocal cord paralysis has occurred outside the context of intubation. These cases contribute to an emerging assortment of evidence supporting the neuropathic capacity of SARS-CoV-2. This report documents a case of COVID-19 related vocal cord paralysis in an 18-year-old female. The patient had a significant history of muscle tension dysphonia, chronic laryngitis, and vocal cord nodules. The patient developed vocal cord paralysis concurrently with the onset of mild viral symptoms and was never intubated or hospitalized. Based on the onset of symptoms and other causes being excluded with CT, a diagnosis of COVID-19-related vocal cord paralysis was performed.

Nanomolar competitive inhibitors of Mycobacterium tuberculosis and Streptomyces coelicolor type II dehydroquinase.

Isomeric nitrophenyl and heterocyclic analogues of the known inhibitor (1S,3R,4R)-1,3,4-trihydroxy-5-cyclohexene-1-carboxylic acid have been synthesized and tested as inhibitors of M. tuberculosis and S. coelicolor type II dehydroquinase, the third enzyme of the shikimic acid pathway. The target compounds were synthesized by a combination of Suzuki and Sonogashira cross-coupling and copper(I)-catalyzed 2,3-dipolar cycloaddition reactions from a common vinyl triflate intermediate. These studies showed that a para-nitrophenyl derivative is almost 20-fold more potent as a competitive inhibitor against the S. coelicolor enzyme than that of M. tuberculosis. The opposite results were obtained with the meta isomer. Five of the bicyclic analogues reported herein proved to be potent competitive inhibitors of S. coelicolor dehydroquinase, with inhibition constants in the low nanomolar range (4-30 nM). These derivatives are also competitive inhibitors of the M. tuberculosis enzyme, but with lower affinities. The most potent inhibitor against the S. coelicolor enzyme, a 6-benzothiophenyl derivative, has a K(i) value of 4 nM-over 2000-fold more potent than the best previously known inhibitor, (1R,4R,5R)-1,5-dihydroxy-4-(2-nitrophenyl)cyclohex-2-en-1-carboxylic acid (8 microM), making it the most potent known inhibitor against any dehydroquinase. The binding modes of the analogues in the active site of the S. coelicolor enzyme (GOLD 3.0.1), suggest a key pi-stacking interaction between the aromatic rings and Tyr 28, a residue that has been identified as essential for enzyme activity.

Crystal structures of Helicobacter pylori type II dehydroquinase inhibitor complexes: new directions for inhibitor design.

The crystal structures of the type II dehydroquinase (DHQase) from Helicobacter pylori in complex with three competitive inhibitors have been determined. The inhibitors are the substrate analogue 2,3-anhydroquinate (FA1), citrate, and an oxoxanthene sulfonamide derivative (AH9095). Despite the very different chemical nature of the inhibitors, in each case the primary point of interaction with the enzyme is via the residues that bind the C1 functionalities of the substrate, 3-dehydroquinate, i.e., N76, H102, I103, and H104. The DHQase/AH9095 complex crystal structure shows that sulfonamides can form a scaffold for nonsubstrate-like inhibitors and identifies a large conserved hydrophobic patch at the entrance to the active site as a locus that can be exploited in the development of new ligands.

Rational design of new bifunctional inhibitors of type II dehydroquinase.

Selective inhibitors of type II dehydroquinase were rationally designed to explore a second binding-pocket in the active-site. The molecular modelling, synthesis, inhibition studies and crystal structure determination are described.

Identification of 4-amino-4-deoxychorismate synthase as the molecular target for the antimicrobial action of (6s)-6-fluoroshikimate.

(6S)-6-Fluoroshikimate has antimicrobial activity. The molecular basis of this effect had not been identified, but there was speculation that (6S)-6-fluoroshikimate is first converted in vivo into 2-fluorochorismate, which then could inhibit 4-amino-4-deoxychorismate synthase (ADCS). 2-Fluorochorismate was prepared from E-fluorophosphoenolpyruvate and erythose-4-phosphate by the sequential reactions of DAHP synthase, dehydroquinate synthase, dehydroquinase, shikimate dehydrogenase, EPSP synthase, and chorismate synthase. Inhibition studies on ADCS showed that it was inhibited rapidly and irreversibly by 2-fluorochorismate. Electrospray mass spectrometry of the inactivated enzyme showed an additional mass of 198 +/- 10 Da. A novel peptide of 1087.6 Da was identified in the HPLC trace for the tryptic digest of 2-fluorochorismate-inactivated ADCS. Sequencing of this peptide by MS/MS showed that the peptide corresponded to residues 272-279 with a modification of 206.1 Da on Lys-274. This observation is particularly exciting in the context of a recent proposal for the catalytic mechanism of ADCS.

(1R,4S,5R)-3-Fluoro-1,4,5-trihydroxy-2-cyclohexene-1-carboxylic acid: the fluoro analogue of the enolate intermediate in the reaction catalyzed by type II dehydroquinases.

The fluoro analogue of the enolate intermediate in the reaction catalyzed by type II dehydroquinases has been prepared from naturally occurring (-)-quinic acid over seven steps and has been shown to be the most potent inhibitor reported to date of the type II enzyme from Mycobacterium tuberculosis.

The structure of Escherichia coli ATP-phosphoribosyltransferase: identification of substrate binding sites and mode of AMP inhibition.

ATP-phosphoribosyltransferase (ATP-PRT), the first enzyme of the histidine pathway, is a complex allosterically regulated enzyme, which controls the flow of intermediates through this biosynthetic pathway. The crystal structures of Escherichia coli ATP-PRT have been solved in complex with the inhibitor AMP at 2.7A and with product PR-ATP at 2.9A (the ribosyl-triphosphate could not be resolved). On the basis of binding of AMP and PR-ATP and comparison with type I PRTs, the PRPP and parts of the ATP-binding site are identified. These structures clearly identify the AMP as binding in the 5-phosphoribosyl-alpha-1-pyrophosphate (PRPP)-binding site, with the adenosine ring occupying the ATP-binding site. Comparison with the recently solved Mycobacterium tuberculosis ATP-PRT structures indicates that histidine is solely responsible for the large conformational changes observed between the hexameric forms of the enzyme. The role of oligomerisation in inhibition and the structural basis for the synergistic inhibition by histidine and AMP are discussed.

Parallel solid-phase synthesis and evaluation of inhibitors of Streptomyces coelicolor type II dehydroquinase.

A series of 1-substituted and 4-substituted benzyl analogues of the known inhibitor (1S,3R,4R)-1,3,4-trihydroxy-5-cyclohexene-1-carboxylic acid has been synthesized and tested as inhibitors of Streptomyces coelicolor type II dehydroquinase. The solid-phase syntheses of 18 new analogues are reported. The most potent inhibitor, 2-nitrobenzyloxy analogue 5i, has K(i) of 8 microM, more than 30 times lower than the K(M) of the substrate and approximately 4 times more potent than the original inhibitor. The binding modes of the synthesized analogues in the active site were studied by molecular docking with GOLD 2.0.

Enzymic synthesis of 3-[3- 13C]dehydroquinic acid.

The title compound has been prepared enzymically over four steps from commercially available D-[5- 13C]fructose.

Design, synthesis and evaluation of bifunctional inhibitors of type II dehydroquinase.

Inhibitors of type II dehydroquinase were designed to straddle the two distinct binding sites identified for the inhibitor (1S,3R,4R)-1,3,4-trihydroxy-5-cyclohexene-1-carboxylic acid and a glycerol molecule in a crystallographic study of the Streptomyces coelicolor enzyme. A number of compounds were designed to incorporate characteristics of both ligands. These analogues were synthesized from quinic acid, and were assayed against type I (Salmonella typhi) and type II (S. coelicolor) dehydroquinases. None of the analogues showed inhibition for type I dehydroquinase. Six of the analogues were shown to have inhibition constants in the micromolar to low millimolar range against the S. coelicolor type II dehydroquinase, while two showed no inhibition. The binding modes of the analogues in the active site of the S. coelicolor enzyme were studied by molecular docking with GOLD1.2. These studies suggest a binding mode where the ring is in a similar position to (1S,3R,4R)-1,3,4-trihydroxy-5-cyclohexene-1-carboxylic acid in the crystal structure and the side-chain occupies part of the glycerol binding-pocket.

Effects of salts on the function and conformational stability of shikimate kinase.

The unfolding of shikimate kinase (SK) from Erwinia chrysanthemi by urea and its subsequent refolding on dilution of the denaturing agent has been studied in detail [Eur. J. Biochem. 269 (2002) 2124]. Comparison of the effects of urea on the enzyme with those of guanidinium chloride (GdmCl) and NaCl indicated that chloride ions significantly weakened the binding of shikimate. This finding prompted a more detailed examination of the effects of salts on the structure, function and stability of the enzyme; the effects of NaCl and Na(2)SO(4) were investigated in detail. These salts have very small effects on the secondary structure as judged by far UV CD circular dichroism (CD), although the near UV CD spectra suggest that some limited changes in the environment of aromatic amino acids may occur. Both salts inhibit SK activity and analysis of the kinetic and substrate binding parameters point to a complex mechanism for the inhibition. Inclusion of salts leads to a marked stabilisation against unfolding of the enzyme by urea. When the enzyme is unfolded by incubation in 4 M urea, addition of NaCl or Na(2)SO(4) leads to a relatively slow refolding of the enzyme as judged by the regain of native-like CD and fluorescence. In addition, the refolded enzyme can bind shikimate, though more weakly than the native enzyme. However, the refolded enzyme does not appear to be capable of binding nucleotides, nor does it possess detectable catalytic activity. The refolding process brought about by addition of salt in the presence of 4 M urea is not associated with any change in the fluorescence of the probe 8-anilino-1-naphthalenesulfonic acid (ANS), indicating that an intermediate formed by hydrophobic collapse is unlikely to be significantly populated. The results point to both specific and general effects of salts on SK. These are discussed in the light of the structural information available on the enzyme.

Structures of shikimate dehydrogenase AroE and its Paralog YdiB. A common structural framework for different activities.

Shikimate dehydrogenase catalyzes the fourth step of the shikimate pathway, the essential route for the biosynthesis of aromatic compounds in plants and microorganisms. Absent in metazoans, this pathway is an attractive target for nontoxic herbicides and drugs. Escherichia coli expresses two shikimate dehydrogenase paralogs, the NADP-specific AroE and a putative enzyme YdiB. Here we characterize YdiB as a dual specificity quinate/shikimate dehydrogenase that utilizes either NAD or NADP as a cofactor. Structures of AroE and YdiB with bound cofactors were determined at 1.5 and 2.5 A resolution, respectively. Both enzymes display a similar architecture with two alpha/beta domains separated by a wide cleft. Comparison of their dinucleotide-binding domains reveals the molecular basis for cofactor specificity. Independent molecules display conformational flexibility suggesting that a switch between open and closed conformations occurs upon substrate binding. Sequence analysis and structural comparison led us to propose the catalytic machinery and a model for 3-dehydroshikimate recognition. Furthermore, we discuss the evolutionary and metabolic implications of the presence of two shikimate dehydrogenases in E. coli and other organisms.

Specificity of substrate recognition by type II dehydroquinases as revealed by binding of polyanions.

The interactions between the polyanionic ligands phosphate and sulphate and the type II dehydroquinases from Streptomyces coelicolor and Mycobacterium tuberculosis have been characterised using a combination of structural and kinetic methods. From both approaches, it is clear that interactions are more complex in the case of the latter enzyme. The data provide new insights into the differences between the two enzymes in terms of substrate recognition and catalytic efficiency and may also explain the relative potencies of rationally designed inhibitors. An improved route to the synthesis of the substrate 3-dehydroquinic acid (dehydroquinate) is described.