The COVID-19 pandemic has demonstrated the need for massively-parallel, cost-effective tests monitoring viral spread. Here we present SARSeq, saliva analysis by RNA sequencing, a method to detect SARS-CoV-2 and other respiratory viruses on tens of thousands of samples in parallel. SARSeq relies on next generation sequencing of multiple amplicons generated in a multiplexed RT-PCR reaction. Two-dimensional, unique dual indexing, using four indices per sample, enables unambiguous and scalable assignment of reads to individual samples. We calibrate SARSeq on SARS-CoV-2 synthetic RNA, virions, and hundreds of human samples of various types. Robustness and sensitivity were virtually identical to quantitative RT-PCR. Double-blinded benchmarking to gold standard quantitative-RT-PCR performed by human diagnostics laboratories confirms this high sensitivity. SARSeq can be used to detect Influenza A and B viruses and human rhinovirus in parallel, and can be expanded for detection of other pathogens. Thus, SARSeq is ideally suited for differential diagnostic of infections during a pandemic.

Azoles such as posaconazole (Posa) are highly potent against Trypanosoma cruzi. However, when tested in chronic Chagas disease patients, a high rate of relapse after Posa treatment was observed. It appears that inhibition of T. cruzi cytochrome CYP51, the target of azoles, does not deliver sterile cure in monotherapy. Looking for suitable combination partners of azoles, we have selected a set of inhibitors of sterol and sphingolipid biosynthetic enzymes. A small-scale phenotypic screening was conducted in vitro against the proliferative forms of T. cruzi, extracellular epimastigotes and intracellular amastigotes. Against the intracellular, clinically relevant forms, four out of 15 tested compounds presented higher or equal activity as benznidazole (Bz), with EC50 values ≤2.2 μM. Ro48-8071, an inhibitor of lanosterol synthase (ERG7), and the steroidal alkaloid tomatidine (TH), an inhibitor of C-24 sterol methyltransferase (ERG6), exhibited the highest potency and selectivity indices (SI = 12 and 115, respectively). Both were directed to combinatory assays using fixed-ratio protocols with Posa, Bz, and fexinidazole. The combination of TH with Posa displayed a synergistic profile against amastigotes, with a mean ΣFICI value of 0.2. In vivo assays using an acute mouse model of T. cruzi infection demonstrated lack of antiparasitic activity of TH alone in doses ranging from 0.5 to 5 mg/kg. As observed in vitro, the best combo proportion in vivo was the ratio 3 TH:1 Posa. The combination of Posa at 1.25 mpk plus TH at 3.75 mpk displayed suppression of peak parasitemia of 80% and a survival rate of 60% in the acute infection model, as compared to 20% survival for Posa at 1.25 mpk alone and 40% for Posa at 10 mpk alone. These initial results indicate a potential for the combination of posaconazole with tomatidine against T. cruzi.

Non-steroidal anti-inflammatory drug (NSAID)-exacerbated respiratory disease (N-ERD) comprises the triad of chronic rhinosinusitis with nasal polyps, asthma, and intolerance to NSAIDs. Dupilumab treatment, targeting the IL-4 receptor alpha, significantly reduces polyp burden as well as asthma symptoms. Here we aimed to investigate the effect of dupilumab on aspirin intolerance, burden of disease, as well as on nasal cytokine profiles in patients suffering from N-ERD. In this open-label trial, adult patients with confirmed N-ERD were treated with dupilumab for six months. Clinical parameters (e.g., total polyp scores, quality of life questionnaires, smell test, spirometry), oral aspirin provocation testing, blood, nasal and urine sampling were monitored up to six months after starting dupilumab therapy at regular intervals. Of the thirty-one patients included in the study, thirty completed both aspirin provocation testing. After six months of treatment with dupilumab, 23.3% (n=7/30) of patients developed complete aspirin tolerance and an additional 33.3% of patients (n=10/30) tolerated higher doses. Polyp burden was significantly reduced (total polyp score: −2.68±1.84, p<0.001) and pulmonary symptoms (asthma control test: +2.34±3.67, p<0.001), as well as olfactory performance improved (UPSIT: +11.16±9.54, p<0.001) in all patients after therapy. Patients with increased aspirin tolerance showed a significant decrease in urine leukotriene E4 levels and their improvement in clinical parameters was associated with the reduction of eotaxin-1, CCL17, IL-5, IL-17A and IL-6. 57% of N-ERD patients tolerated higher doses of aspirin under dupilumab therapy.