The risk of transmission of the novel coronavirus (SARS-CoV-2) with human heart valve transplantation: evaluation of cardio-vascular tissues from two consecutive heart donors with asymptomatic COVID-19.

We report on two living donors of explanted hearts while receiving heart transplantation that tested positive for SARS-CoV-2 on the day of donation, although clinically asymptomatic. They underwent heart transplantation for ischaemic and hypertrophic obstructive cardiomyopathy, respectively. After evaluation of donor hearts, we cryopreserved and stored two pulmonary valves for clinical application and one aortic valve for research. Light microscopy of myocardium, mitral valve and aortic and pulmonary arterial wall and RT-PCR SARS-CoV-2 test of myocardium, mitral and tricuspid valve and aortic wall for detection of SARS-CoV-2 were performed. Presence of ACE2 in tissues was assessed with immunostaining. Light microscopy revealed a mild eosinophilic myocarditis in the ischemic cardiomyopathy heart, whereas enlarged cardiomyocytes with irregular nucleus and some with cytoplasmic vacuoles in the hypertrophic obstructive cardiomyopathy heart. Aortic and pulmonary wall were histologically normal. Immunostaining revealed diffuse presence of ACE2 in the myocardium of the heart with eosinophilic myocarditis, but only discrete presence in the hypertrophic cardiomyopathy heart. The RT-PCR SARS-CoV-2 test showed no presence of the virus in tested tissues. Despite eosinophilic myocarditis in the ischemic cardiomyopathy heart, no viral traces were found in the myocardium and valve tissues. However, ACE2 was present diffusely in the ischemic cardiomyopathy heart. SARS-CoV-2 could not be detected in the cardiac tissues of these COVID-19 asymptomatic heart donors. In our opinion, clinical application of the valves from these donors presents negligible risk for coronavirus transmission. Nonetheless, considering the uncertainty regarding the risk of virus transmission with the human tissue transplantation, we would not release in any case the pulmonary valve recovered from the eosinophilic myocarditis heart. In contrast, we may consider the release of the pulmonary valve from the dilated cardiomyopathy heart only for a life-threatening situation when no other similar allograft were available

Nucleic acid sequence-based amplification assay for human papillomavirus mRNA detection and typing: evidence for DNA amplification

Human papillomavirus (HPV) E6/E7 mRNA has been proposed as a more specific marker for cervical dysplasia and cancer than HPV DNA. This study evaluated the RNA specificity of nucleic acid sequence-based amplification (NASBA)-based HPV detection using HPV DNA plasmids (HPV type 16 [HPV16], HPV18, HPV31, HPV33, and HPV45) and nucleic acid extracts of several cell lines, which were systematically subjected to enzymatic treatments with DNase and RNase. HPV plasmid dilutions (10(6) to 10(0) copies/μl) and nucleic acid extracts (total DNA, RNA-free DNA, total RNA, and DNA-free RNA) of unfixed and fixed (PreServCyt and SurePath) HaCaT, HeLa, and CaSki cells were tested with the NucliSENS EasyQ HPV test. The RNA-free DNA extracts of HeLa and CaSki cells could be amplified by HPV18 and -16 NASBA, respectively. Fixation of the cells did not influence NASBA. All HPV plasmids could be detected with NASBA. Based on the plasmid dilution series, a lower detection limit of 5 × 10(3) HPV DNA copies could be determined. Our study identified viral double-stranded DNA as a possible target for NASBA-based HPV detection. The differences in diagnostic accuracy between the NASBA-based tests and conventional HPV DNA detection assays seem to be attributable not to the more specific amplification of viral mRNA but to the limited type range and the lower analytical sensitivity for HPV DNA