Successful laparoscopic lavage and drainage for intestinal perforation in a patient with COVID-19: A case report.

INTRODUCTION: Some patients with coronavirus disease 2019 (COVID-19) have acute abdomen and need surgery. However, surgery in the acute phase of COVID-19 is associated with worse postoperative outcomes and an increased risk of mortality. We report a case of a patient with COVID-19 who developed intestinal perforation that was treated acutely with antibiotics and delayed surgical intervention. PRESENTATION OF CASE: A 79-year-old man with COVID-19 was treated with remdesivir and dexamethasone, and his respiratory symptoms and hypoxia improved. However, abdominal symptoms developed, and intestinal perforation occurred. As the nasopharyngeal swab PCR test was positive for SARS-CoV-2, conservative treatment with tazobactam/piperacillin was started to avoid surgery in the acute phase of COVID-19. An intraperitoneal abscess was confirmed on follow-up computed tomography. Emergent laparoscopic lavage and drainage, and transverse colon stoma construction were performed with medical staff using full personal protective equipment. Bacterial culture from the ascites detected Escherichia coli and Bacteroides. The SARS-CoV-2 PCR test of the ascites sample was negative. No infection was observed in the medical staff. DISCUSSION: COVID-19 has been associated with a higher perioperative risk and postoperative mortality. There has also been a report of ascitic fluid testing positive for SARS-CoV-2 on PCR, suggesting the possibility of intraoperative aerosolization. Avoiding surgical treatment in the acute phase of COVID-19 may reduce deaths from perioperative complications. CONCLUSION: Our case suggests that in acute COVID-19 lung infection, careful observation and delayed surgical treatment could prevent worsening of the COVID-19 and reduce the risk of infection to the medical staff.

Directed evolution of thermotolerant malic enzyme for improved malate production.

The directed evolution of the thermotolerant NADP(H)-dependent malic enzyme from Thermococcus kodakarensis was conducted to alter the cofactor preference of the enzyme from NADP(H) to NAD(H). The construction and screening of two generations of mutant libraries led to the isolation of a triple mutant that exhibited 6-fold higher kcat/Km with NAD(+) than the wild type. We serendipitously found that, in addition to the change in the cofactor preference, the reaction specificity of the mutant enzyme was altered. The reductive carboxylation of pyruvate to malate catalyzed by the wild type enzyme is accompanied by HCO(3)(-)-independent reduction of pyruvate and gives lactate as a byproduct. The reaction specificity of the triple mutant was significantly shifted to malate production and the mutant gave a less amount of the byproduct than the wild type. When the triple mutant enzyme was used as a catalyst for pyruvate carboxylation with NADH, the enzyme gave 1.2 times higher concentration of malate than the wild type with NADPH. Single-point mutation analysis revealed that the substitution of Arg221 with Gly is responsible for the shift in reaction specificity. This finding may shed light on the catalytic mechanisms of malic enzymes and other related CO2- and/or HCO(3)(-)-fixing enzymes.

Direct conversion of glucose to malate by synthetic metabolic engineering.

Synthetic metabolic engineering enables us to construct an in vitro artificial synthetic pathways specialized for chemical manufacturing through the simple heat-treatment of the recombinant mesophiles having thermophilic enzymes, followed by rational combination of those biocatalytic modules. In this work, we constructed a synthetic pathway capable of direct conversion of glucose to malate. The reversible carboxylation of pyruvate catalyzed by a malic enzyme derived from Thermococcus kodakarensis (TkME) (ΔG°'=+7.3kJmol(-1)) was coupled with a thermodynamically favorable non-ATP-forming Embden-Meyerhof pathway to balance the consumption and regeneration of redox cofactors and to shift the overall equilibrium toward malate production (glucose+2HCO3(-)+2H→2 malate+2H2O; ΔG°'=-121.4kJmol(-1)). TkME exhibited both pyruvate carboxylation (malate-forming) and pyruvate reduction (lactate-forming) activities. By increasing HCO3(-) concentration, the reaction specificity could be redirected to malate production. As a result, the direct conversion of glucose to malate was achieved with a molar yield of 60%.

The mechanism of fibril formation of a non-inhibitory serpin ovalbumin revealed by the identification of amyloidogenic core regions.

Ovalbumin (OVA), a non-inhibitory member of the serpin superfamily, forms fibrillar aggregates upon heat-induced denaturation. Recent studies suggested that OVA fibrils are generated by a mechanism similar to that of amyloid fibril formation, which is distinct from polymerization mechanisms proposed for other serpins. In this study, we provide new insights into the mechanism of OVA fibril formation through identification of amyloidogenic core regions using synthetic peptide fragments, site-directed mutagenesis, and limited proteolysis. OVA possesses a single disulfide bond between Cys(73) and Cys(120) in the N-terminal helical region of the protein. Heat treatment of disulfide-reduced OVA resulted in the formation of long straight fibrils that are distinct from the semiflexible fibrils formed from OVA with an intact disulfide. Computer predictions suggest that helix B (hB) of the N-terminal region, strand 3A, and strands 4-5B are highly ß-aggregation-prone regions. These predictions were confirmed by the fact that synthetic peptides corresponding to these regions formed amyloid fibrils. Site-directed mutagenesis of OVA indicated that V41A substitution in hB interfered with the formation of fibrils. Co-incubation of a soluble peptide fragment of hB with the disulfide-intact full-length OVA consistently promoted formation of long straight fibrils. In addition, the N-terminal helical region of the heat-induced fibril of OVA was protected from limited proteolysis. These results indicate that the heat-induced fibril formation of OVA occurs by a mechanism involving transformation of the N-terminal helical region of the protein to ß-strands, thereby forming sequential intermolecular linkages.

Changes in activities of enzymes in erythrocytes from ddY mice supplemented with dietary selenium.

Plasma and erythrocyte enzyme activities were measured in ddY mice supplemented with dietary selenium (Se) from baker's yeast (Saccharomyces serevisiae) or selenious acid at 0.3 ppm Se content. Glutathione peroxidase (GSHpx) activities increased significantly in erythrocytes from mice supplemented with dietary Se. It was concluded that addition of dietary Se as food additive is very effective for activation of GSHpx in mice.