A Futile Metabolic Cycle of Fatty Acyl-CoA Hydrolysis and Resynthesis in Corynebacterium glutamicum and Its Disruption Leading to Fatty Acid Production.

Fatty acyl-CoA thioesterase (Tes) and acyl-CoA synthetase (FadD) catalyze opposing reactions between acyl-CoAs and free fatty acids. Within the genome of Corynebacterium glutamicum, several candidate genes for each enzyme are present, although their functions remain unknown. Modified expressions of the candidate genes in the fatty acid producer WTΔfasR led to identification of one tes gene (tesA) and two fadD genes (fadD5 and fadD15), which functioned positively and negatively in fatty acid production, respectively. Genetic analysis showed that fadD5 and fadD15 are responsible for utilization of exogenous fatty acids and that tesA plays a role in supplying fatty acids for synthesis of the outer layer components mycolic acids. Enzyme assays and expression analysis revealed that tesA, fadD5, and fadD15 were co-expressed to create a cyclic route between acyl-CoAs and fatty acids. When fadD5 or fadD15 was disrupted in wild-type C. glutamicum, both disruptants excreted fatty acids during growth. Double disruptions of them resulted in a synergistic increase in production. Additional disruption of tesA revealed a canceling effect on production. These results indicate that the FadDs normally shunt the surplus of TesA-generated fatty acids back to acyl-CoAs for lipid biosynthesis and that interception of this shunt provokes cells to overproduce fatty acids. When this strategy was applied to a fatty acid high-producer, the resulting fadDs-disrupted and tesA-amplified strain exhibited a 72% yield increase relative to its parent and produced fatty acids, which consisted mainly of oleic acid, palmitic acid, and stearic acid, on the gram scale per liter from 1% glucose.IMPORTANCE The industrial amino acid producer Corynebacterium glutamicum has currently evolved into a potential workhorse for fatty acid production. In this organism, we obtained evidence showing the presence of a unique mechanism of lipid homeostasis, namely, a formation of a futile cycle of acyl-CoA hydrolysis and resynthesis mediated by acyl-CoA thioesterase (Tes) and acyl-CoA synthetase (FadD), respectively. The biological role of the coupling of Tes and FadD would be to supply free fatty acids for synthesis of the outer layer components mycolic acids and to recycle their surplusage to acyl-CoAs for membrane lipid synthesis. We further demonstrated that engineering of the cycle in a fatty acid high-producer led to dramatically improved production, which provides a useful engineering strategy for fatty acid production in this industrially important microorganism.

Cerebrospinal fluid analysis is associated with enhancement on MRI in bacterial and tuberculous meningitis: A retrospective observational study.

OBJECTIVE: Along with cerebrospinal fluid (CSF) analysis, enhancement on contrast-enhanced MRI is useful to diagnose meningitis. However, the conditions for its appearance have not been clarified. This study aimed to investigate the association between CSF parameters and enhancement on contrast-enhanced head or spinal MRI in patients with bacterial meningitis (BM) or tuberculous meningitis (TM). METHODS: A total of 12 patients with BM and 23 patients with TM who underwent both CSF analysis and contrast-enhanced MRI were included. The correlation between CSF analysis and MRI findings has been examined using receiver operating characteristic (ROC) analysis. RESULTS: Contrast enhancement was found in 7 and 10 patients with BM and TM, respectively. Both CSF analysis and MRI were useful to distinct between BM and TM, and the enhancement implied the severity of them. In patients with BM, higher CSF protein and lower CSF glucose were associated with enhancement on MRI, while not only CSF protein and glucose but also leukocyte and lymphocyte counts were associated with it in TM. CSF adenosine deaminase (ADA) did not show discriminant ability of the MRI findings. CONCLUSIONS: CSF analysis is associated with enhancement on contrast-enhanced MRI both in BM and TM. Our findings indicate the importance of CSF analysis in assessing the need to perform contrast-enhanced MRI, which may be useful in diagnosis, distinction, and estimation of prognosis in those patients.

An Autopsy Report of a Case with Cerebral Infarction Complicated by Coronavirus Disease 2019 Infection.

We herein report a case of cerebral infarct in a patient with coronavirus disease 2019 (COVID-19) infection who died of aspiration pneumonia. The postmortem examination of the brain revealed embolic infarct with negative findings on quantitative reverse transcription polymerase chain reaction (qRT-PCR) as well as immunohistochemistry to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The systemic examination only revealed low copy numbers of SARS-CoV-2 in the bronchus. This is the first and so far only autopsy case of COVID-19 infection with pathologic and virologic findings of the postmortem brain in Japan.