ddPCR increases detection of SARS-CoV-2 RNA in patients with low viral loads.

RT-qPCR detection of SARS-CoV-2 RNA still represents the method of reference to diagnose and monitor COVID-19. From the onset of the pandemic, however, doubts have been expressed concerning the sensitivity of this molecular diagnosis method. Droplet digital PCR (ddPCR) is a third-generation PCR technique that is particularly adapted to detecting low-abundance targets. We developed two-color ddPCR assays for the detection of four different regions of SARS-CoV-2 RNA, including non-structural (IP4-RdRP, helicase) and structural (E, N) protein-encoding sequences. We observed that N or E subgenomic RNAs are generally more abundant than IP4 and helicase RNA sequences in cells infected in vitro, suggesting that detection of the N gene, coding for the most abundant subgenomic RNA of SARS-CoV-2, increases the sensitivity of detection during the highly replicative phase of infection. We investigated 208 nasopharyngeal swabs sampled in March-April 2020 in different hospitals of Greater Paris. We found that 8.6% of informative samples (n = 16/185, P < 0.0001) initially scored as "non-positive" (undetermined or negative) by RT-qPCR were positive for SARS-CoV-2 RNA by ddPCR. Our work confirms that the use of ddPCR modestly, but significantly, increases the proportion of upper airway samples testing positive in the framework of first-line diagnosis of a French population.

[Acute pancreatitis complicated by Clostridium tertium septicaemia]. / Pancréatite aiguë compliquée d'une septicémie a Clostridium tertium.

INTRODUCTION: Clostridium tertium septicaemia, although usually described in neutropenic patients, can also occur in the absence of neutropenia, as in our case report. OBSERVATION: A 61 year-old woman presented with pain in the right hypochondrium and epigastrium, constipation and vomiting that led to the diagnosis of acute pancreatitis of biliary origin. During septic shock, analysis of the liquid of an intra-abdominal mass and the results of the blood cultures revealed a Gram positive bacillus identified as C. tertium. During laparotomy, necrotic-hemorrhagic acute pancreatitis lesions were found with subtotal colic necrosis. Analysis of the peritoneal fluid revealed C. tertium. The patient died on D 46 after several nosocomial pneumopathies without identification of C. tertium. DISCUSSION: Three factors appear related to the onset of C. tertium septicaemia: neutropenia, lesions of the intestinal mucosa and previous treatment with 3rd generation, broad spectrum, cephalosporins. Such septicaemias are rare in immunocompetent patients and do not appear to have been reported before in a context of acute pancreatitis.