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Andrimid (Adm) synthase, which belongs to the type-II system of enzymes, produces Adm in Pantoea agglomerans. The adm biosynthetic gene cluster lacks canonical acyltransferases (ATs) to load the malonyl group to acyl carrier proteins (ACPs), suggesting that a malonyl-CoA: ACP transacylase (MCAT) from the producer organism's fatty acid synthase (FAS) complex provides the essential acyltransferase (AT) activity in Adm biosynthesis. Here we report that the MCAT is essential for catalysis of the transacylation of malonate from malonyl-CoA to AdmA polyketide synthase (PKS) ACP in vitro. Furthermore, a catalytic self-malonylation of the AdmA PKS ACP was not observed in reactions without MCAT, while many type II PKS ACPs are capable of catalyzing the self-acylation process. This lack of self-malonylation was validated by amino acid sequence analysis of the AdmA PKS ACP and the type II PKS ACPs. These results show that the MCAT from the producer organism's FAS complex provides the missing AT activity in trans, suggesting a functional protein-protein interaction between the fatty acid and polyketide synthases in the Adm assembly line.

Rapid identification of causative agents from positive blood culture media is a prerequisite for the timely targeted treatment of patients with sepsis. The GENECUBE (TOYOBO Co., Ltd.) is a novel, fully-automated gene analyzer that can purify DNAs and amplify target DNAs. In this study, we evaluated the ability of two newly developed GENECUBE assays to directly detect the nuc and mecA genes in blood culture medium; nuc is specific to Staphylococcus aureus, and mecA indicates methicillin resistance. We examined 263 positive blood culture samples taken at three hospitals from patients suspected of having staphylococcal bacteremia. The results were then compared with those obtained using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, antimicrobial susceptibility testing (Microscan system or Dry-plate EIKEN), and sequencing analysis. The GENECUBE assays had sensitivity and specificity of 100% in detecting both S. aureus and methicillin resistance in positive blood culture. The turnaround time of the examination was evaluated for 36 positive blood culture samples. The time between the initiation of incubation and completion of the GENECUBE examination was 23 h (interquartile range: IQR 21–37 h); the time between reporting of Gram stain examination and completion of the GENECUBE examination was 52 min (IQR 48–62 min). These findings show that the GENECUBE assays significantly reduce the assay time with no loss of sensitivity or specificity.