Project description:Indispensable for shortening treatment of drug-susceptible tuberculosis (TB), pyrazinamide (PZA, Z) is also essential in the treatment of multidrug-resistant (MDR)-TB. While resistance to PZA in MDR-TB is associated with poor treatment outcome, bacillary susceptibility to PZA along with the use of fluoroquinolone (FQ) and second-line injectable drugs (SLIDs) may predict improved treatment success in MDR-TB. Despite a high prevalence of PZA resistance among MDR-TB patients (10%-85%), PZA susceptibility testing is seldom performed because of technical challenges. To improve treatment of MDR-TB, we propose to: (i) classify MDR-TB into PZA-susceptible MDR-TB (Z(S)-MDR-TB) and PZA-resistant MDR-TB (Z(R)-MDR-TB); (ii) use molecular tests such as DNA sequencing (pncA, gyrA, rrs, etc.) to rapidly identify Z(S)-MDR-TB versus Z(R)-MDR-TB and susceptibility profile for FQ and SLID; (iii) refrain from using PZA in Z(R)-MDR-TB; and (iv) explore the feasibility of shortening the treatment duration of Z(S)-MDR-TB with a regimen comprising PZA plus at least two bactericidal agents especially new agents like TMC207 or PA-824 or delamanid which the bacilli are susceptible to, with one or two other agents. These measures may potentially shorten therapy, save costs, and reduce side effects of MDR-TB treatment.
Project description:Whole genome sequencing (WGS) is revolutionary for diagnostics of TB and its mutations associated with drug-resistances, but its uptake in low- and middle-income countries is hindered by concerns of implementation feasibility. Here, we provide a proof of concept for its successful implementation in such a setting. WGS was implemented in the Kyrgyz Republic. We estimated needs of up to 55 TB-WGS per week and chose the MiSeq platform (Illumina, USA) because of its capacity of up to 60 TB-WGS per week. The project's timeline was completed in 93-weeks. Costs of large equipment and accompanying costs were 222,065 USD and 8462 USD, respectively. The first 174 WGS costed 277 USD per sequence, but this was skewed by training inefficiencies. Based on real prices and presuming optimal utilization of WGS capacities, WGS costs could drop to 167 and 141 USD per WGS using MiSeq Reagent Kits v2 (500-cycles) and v3 (600-cycles), respectively. Five trainings were required to prepare the staff for autonomous WGS which cost 48,250 USD. External assessment confirmed excellent performance of WGS by the Kyrgyz laboratory in an interlaboratory comparison of 30 M. tuberculosis genomes showing complete agreeance of results.
Project description:We report the first use of whole viral genome sequencing to identify nosocomial transmission of varicella-zoster virus with fatal outcome. The index case patient, nursed in source isolation, developed disseminated zoster with rash present for 1 day before being transferred to the intensive care unit (ICU). Two patients who had received renal transplants while inpatients in an adjacent ward developed chickenpox and 1 died; neither patient had direct contact with the index patient.
Project description:Multidrug resistance in M. tb has become a huge global problem due to drug resistance. Hence, the treatment remains a challenge, even though short term chemotherapy is available. Therefore, it is of interest to identify novel drug targets in M.tb through gene expression profiling complimented by a subtractive proteome model. WhiB6 is a transcriptional regulator protein and a known drug resistant marker that is critical in the secretion dependent regulation of ESX-1, which is specialized for the deployment of host membrane-targeting proteins. The WhiB6 protein structure was modelled ab initio and was docked with a library of 173 phytochemicals with potential antituberculosis activity to the identified drug marker to find novel lead molecules. UDP-galactopyranose and GDP-L-galactose were identified to be potential lead molecules to inhibit the target WhiB6. The results were compared with the first line drugs for MDR-TB by docking with WhiB6. Data showed that Ethambutol showed better binding ability to WhiB6 but the afore mentioned top ranked phytochemicals were found to be better candidate molecules. The chosen candidate lead molecules should be further validated by suitable in vitro or in vivo investigation.
Project description:Multidrug-resistant Klebsiella pneumoniae (MDR Kp), in particular carbapenem-resistant Kp (CR-Kp), has become endemic in Italy, where alarming data have been reported on the spread of colistin-resistant CR-Kp (CRCR-Kp). During the period 2013-2014, 27 CRCR-Kp nosocomial strains were isolated within the Modena University Hospital Policlinico (MUHP) multidrug resistance surveillance program. We retrospectively investigated these isolates by whole-genome sequencing (WGS) analysis of the resistome, virulome, plasmid content, and core single nucleotide polymorphisms (cSNPs) in order to gain insights into their molecular epidemiology. The in silico WGS analysis of the resistome revealed the presence of genes, such as blaKPC, related to the phenotypically detected resistances to carbapenems. Concerning colistin resistance, the plasmidic genes mcr1-9 were not detected, while known and new genetic variations in mgrB, phoQ, and pmrB were found. The virulome profile revealed the presence of type-3 fimbriae, capsular polysaccharide, and iron acquisition system genes. The detected plasmid replicons were classified as IncFIB(pQil), IncFIB(K), ColRNAI, IncX3, and IncFII(K) types. The cSNPs genotyping was consistent with the multi locus sequence typing (MLST) and with the distribution of mutations related to colistin resistance genes. In a nosocomial drug resistance surveillance program, WGS proved to be a useful tool for elucidating the spread dynamics of CRCR-Kp nosocomial strains and could help to limit their diffusion.
Project description:Here, we report the draft genome sequence and annotation of a multidrug resistant Mycobacterium tuberculosis strain PR10 (MDR-TB PR10) isolated from a patient diagnosed with tuberculosis. The size of the draft genome MDR-TB PR10 is 4.34 Mbp with 65.6% of G + C content and consists of 4637 predicted genes. The determinants were categorized by RAST into 400 subsystems with 4286 coding sequences and 50 RNAs. The whole genome shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession number CP010968.
Project description:Hemangioblastomas (HBs) are uncommon tumors characterized by the presence of inactivating alterations in the von Hippel-Lindau (VHL) gene in inherited cases and by infrequent somatic mutation in sporadic entities. We performed whole exome sequencing on 11 HB patients to further elucidate the genetics of HBs. A total of 270 somatic variations in 219 genes, of which there were 86 mutations in 67 genes, were found in sporadic HBs, and 184 mutations were found in 154 genes in familial HBs. C: G>T: A and T: A>C: G mutations are relatively common in most HB patients. Genes harboring the most significant mutations include PCDH9, KLHL12, DCAF4L1, and VHL in sporadic HBs, and ZNF814, DLG2, RIMS1, PNN, and MUC7 in familial HBs. The frequency of CNV varied considerably within sporadic HBs but was relatively similar within familial HBs. Five genes, including OTOGL, PLCB4, SCEL, THSD4, and WWOX, have CNVs in the six patients with sporadic HBs, and three genes, including ABCA6, CWC27, and LAMA2, have CNVs in the five patients with familial HBs. We found new genetic mutations and CNVs that might be involved in HBs; these findings highlight the complexity of the tumorigenesis of HBs and pinpoint potential therapeutic targets for the treatment of HBs.
Project description:BackgroundEarly-onset Parkinson's disease (PD) is the most common inherited form of parkinsonism, with the PRKN gene being the most frequently identified mutated. Exon rearrangements, identified in about 43.2% of the reported PD patients and with higher frequency in specific ethnicities, are the most prevalent PRKN mutations reported to date in PD patients.MethodsIn this study, three consanguineous families with early-onset PD were subjected to whole-genome sequencing (WGS) analyses that were followed by Sanger sequencing and droplet digital PCR to validate and confirm the disease segregation of the identified genomic variations and to determine their parental origin.ResultsFive different PRKN structural variations (SVs) were identified. Because the genomic sequences surrounding the break points of the identified SVs might hold important information about their genesis, these were also characterized for the presence of homology and repeated sequences.ConclusionWe concluded that all identified PRKN SVs might originate through retrotransposition events.
Project description:Common variable immunodeficiency (CVID) belongs to the primary immunodeficiency disorders (PIDs), presenting a profound heterogeneity in phenotype and genotype, with monogenic or complex causes. Recurrent respiratory infections are the most common clinical manifestations. CVID patients can also develop various autoimmune and lymphoproliferative complications. Genetic testing such as whole exome sequencing (WES) can be utilized to investigate likely genetic defects, helping for better clinical management. We described the clinical phenotypes of three sporadic cases of CVID, who developed recurrent respiratory infections with different autoimmune and lymphoproliferative complications. WES was applied to screen disease-causing or disease-associated mutations. Two patients were identified to have monogenic disorders, with compound heterozygous mutations in LRBA for one patient and a frameshift insertion in NFKB1 for another. The third patient was identified to be a complex form of CVID. Two novel mutations were identified, respectively, in LRBA and NFKB1. A combination of clinical and genetic diagnosis can be more extensively utilized in the clinical practice due to the complexity and heterogeneity of CVID.
Project description:The Union in collaboration with national TB programme (NTP) started the community-based MDR-TB care (CBMDR-TBC) project in 33 townships of upper Myanmar to improve treatment initiation and treatment adherence. Patients with MDR-TB diagnosed/registered under NTP received support through the project staff, in addition to the routine domiciliary care provided by NTP staff. Each township had a project nurse exclusively for MDR-TB and 30 USD per month (max. for 4 months) were provided to the patient as a pre-treatment support.To assess whether CBMDR-TBC project's support improved treatment initiation.In this cohort study (involving record review) of all diagnosed MDR-TB between January 2015 and June 2016 in project townships, CBMDR-TBC status was categorized as "receiving support" if date of project initiation in patient's township was before the date of diagnosis and "not receiving support", if otherwise. Cox proportional hazards regression (censored on 31 Dec 2016) was done to identify predictors of treatment initiation.Of 456 patients, 57% initiated treatment: 64% and 56% among patients "receiving support (n = 208)" and "not receiving support (n = 228)" respectively (CBMDR-TBC status was not known in 20 (4%) patients due to missing diagnosis dates). Among those initiated on treatment (n = 261), median (IQR) time to initiate treatment was 38 (20, 76) days: 31 (18, 50) among patients "receiving support" and 50 (26,101) among patients "not receiving support". After adjusting other potential confounders (age, sex, region, HIV, past history of TB treatment), patients "receiving support" had 80% higher chance of initiating treatment [aHR (0.95 CI): 1.8 (1.3, 2.3)] when compared to patients "not receiving support". In addition, age 15-54 years, previous history of TB and being HIV negative were independent predictors of treatment initiation.Receiving support under CBMDR-TBC project improved treatment initiation: it not only improved the proportion initiated but also reduced time to treatment initiation. We also recommend improved tracking of all diagnosed patients as early as possible.