Structural Characterization and Ligand/Inhibitor Identification Provide Functional Insights into the Mycobacterium tuberculosis Cytochrome P450 CYP126A1

The Mycobacterium tuberculosis H37Rv genome encodes 20 cytochromes P450, including P450s crucial to infection and bacterial viability. Many M. tuberculosis P450s remain uncharacterized, suggesting that their further analysis may provide new insights into M. tuberculosis metabolic processes and new targets for drug discovery. CYP126A1 is representative of a P450 family widely distributed in mycobacteria and other bacteria. Here we explore the biochemical and structural properties of CYP126A1, including its interactions with new chemical ligands. A survey of azole antifungal drugs showed that CYP126A1 is inhibited strongly by azoles containing an imidazole ring but not by those tested containing a triazole ring. To further explore the molecular preferences of CYP126A1 and search for probes of enzyme function, we conducted a high throughput screen. Compounds containing three or more ring structures dominated the screening hits, including nitroaromatic compounds that induce substrate-like shifts in the heme spectrum of CYP126A1. Spectroelectrochemical measurements revealed a 155-mV increase in heme iron potential when bound to one of the newly identified nitroaromatic drugs. CYP126A1 dimers were observed in crystal structures of ligand-free CYP126A1 and for CYP126A1 bound to compounds discovered in the screen. However, ketoconazole binds in an orientation that disrupts the BC-loop regions at the P450 dimer interface and results in a CYP126A1 monomeric crystal form. Structural data also reveal that nitroaromatic ligands "moonlight" as substrates by displacing the CYP126A1 distal water but inhibit enzyme activity. The relatively polar active site of CYP126A1 distinguishes it from its most closely related sterol-binding P450s in M. tuberculosis, suggesting that further investigations will reveal its diverse substrate selectivity.This work was supported by Biotechnology and Biological Research Council (BBSRC) Grants BB/I019227/1 (to A. W. M.) and BB/I019669/1 (to C. A.) underpinning this research program and supporting the research of A. J. C. and K. J. M

Detection of four patients who were infected by Schistosoma haematobium in Vietnam

Nguyen Van De,1 Truong La,2 Pham Ngoc Minh,1 Pham Thi Bich Dao,1 Le Van Duyet3 1Department of Parasitology, Hanoi Medical University, Hanoi, Vietnam; 2Department of Microbiology and Parasitology, The Western Highlands Agriculture and Forestry Science Institute (WASI), Hanoi, Vietnam; 3Department of Molecular Diagnosis, National Hospital for Tropical Diseases, Hanoi, Vietnam Introduction: Schistosoma lives as a parasite in the portal vein causing intestinal lesions. It also lives in the liver, spleen, and the vein of the urinary bladder causing lesions in the urinary system. Angola is an endemic area of Schistosoma haematobium, which causes lesions in the urinary system, including the urinary bladder. In this study, we aimed to identify and classify the parasites that were collected from four patients from Angola, who currently live in Vietnam, by morphological and molecular methods. Patients and methods: The main clinical symptoms of the patients were collected, and Schistosoma eggs were taken from urine by a centrifugal method from the four patients in 2016. Identification of the species by morphological method was taken using a microscope. The DNA of the Schistosoma was also isolated and was identified by cytochrome C oxidase subunit 1 (Cox1) sequence. Results: The four Vietnamese patients infected with S. haematobium in Angola returned to Vietnam. All the patients felt strange and had cystalgia and hematuria (blood urine), and one of them was diagnosed with urinary bladder cancer, where surgery was necessary for that patient. Schistosoma eggs, which were collected from the urine of the four patients, were identified as S. haematobium by morphological and molecular methods. These patients were the first reports of Schistosoma in Vietnam. Conclusion: Four Vietnamese schistosomiasis patients returned from Angola: three were diagnosed with schistosomiasis and one was diagnosed with urinary bladder cancer. They had similar symptoms including a strange feeling, cystalgia, hematuria, and eosinophilia and were detected with Schistosoma eggs in urine. Keywords: hematuria, Schistosoma haematobium, Vietna

Human infection with an unknown species of Dracunculus in Vietnam

Guinea worm (GW) disease, caused by Dracunculus medinensis, is an almost eradicated waterborne zoonotic disease. The World Health Organization (WHO) currently lists GW as endemic in only five African countries. In July 2020, the Vietnamese public health surveillance system detected a hanging worm in a 23-year-old male patient, who did not report any travel to Africa or any country previously endemic for GW. The patient was hospitalized with symptoms of fatigue, anorexia, muscle aches, and abscesses, with worms hanging out of the skin in the lower limbs. The worms were retrieved from the lesions and microscopically examined in Vietnam, identifying structures compatible with Dracunculus spp. and L1-type larvae. A section of this parasite was sent to the Centers for Disease Control and Prevention (CDC) in Atlanta, United States, for confirmatory diagnosis of GW. The adult worm had cuticle structures compatible with Dracunculus parasites, although the length of L1 larvae was about 339 μm, substantially shorter than D. medinensis. DNA sequence analysis of the 18S small subunit rRNA gene confirmed that this parasite was not GW, and determined that the sample belonged to a Dracunculus sp. not previously reported in GenBank that clustered with the animal-infective Dracunculus insignis and Dracunculus lutrae, located in a different clade than D. medinensis. This study highlights the importance of effective public health surveillance systems and the collaborative work of local public health authorities from Vietnam with the WHO and CDC in efforts to achieve the eradication of GW