Detection of mutations in Mycobacterium tuberculosis genome determining resistance to fluoroquinolones by hybridization on biological microchips

We developed a method of identification of Mycobacterium tuberculosis with simultaneous evaluation of the sensitivity to fluoroquinolones on a biological microchip array. The method of multiplex two-staged PCR followed by hybridization of a biochip makes it possible to detect 8 mutant variants of gyrA gene occurring in fluoroquinolone-resistant strains (∼85% all resistant forms) within 1 day. Using this method we analyzed 107 cultures isolated from patients with tuberculosis and 78 sputum samples. Mutations in gyrA gene were detected in 48 (92%) resistant strains. Natural S95T polymorphism in gyrA gene was detected in all resistant and in 76% sensitive strains. The sensitivity and specificity of the proposed method calculated on the basis of the analysis of sputum samples (n=78) were 94 and 100%, respectively. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 145, No. 1, pp. 115–120, January, 2008     

Exome sequencing identifies mutations in KIF14 as a novel cause of an autosomal recessive lethal fetal ciliopathy phenotype

Gene discovery using massively parallel sequencing has focused on phenotypes diagnosed postnatally such as well‐characterized syndromes or intellectual disability, but is rarely reported for fetal disorders. We used family‐based whole‐exome sequencing in order to identify causal variants for a recurrent pattern of an undescribed lethal fetal congenital anomaly syndrome. The clinical signs included intrauterine growth restriction (IUGR), severe microcephaly, renal cystic dysplasia/agenesis and complex brain and genitourinary malformations. The phenotype was compatible with a ciliopathy, but not diagnostic of any known condition. We hypothesized biallelic disruption of a gene leading to a defect related to the primary cilium. We identified novel autosomal recessive truncating mutations in KIF14 that segregated with the phenotype. Mice with autosomal recessive mutations in the same gene have recently been shown to have a strikingly similar phenotype. Genotype–phenotype correlations indicate that the function of KIF14 in cell division and cytokinesis can be linked to a role in primary cilia, supported by previous cellular and model organism studies of proteins that interact with KIF14. We describe the first human phenotype, a novel lethal ciliary disorder, associated with biallelic inactivating mutations in KIF14. KIF14 may also be considered a candidate gene for allelic viable ciliary and/or microcephaly phenotypes.     

Synthesis and antitumor activity of fluorinated derivatives of [<Emphasis Type="Italic">i,j</Emphasis>]-annelated quinolones

Tri- and pentacyclic fluoroquinolones were synthesized by intramolecular cyclization of the corresponding 3-hydrazinopolyfluorobenzoylacrylates followed by substitution of fluorine atoms by amine residues. The antitumor activity of the resulting compounds was studied at the National Cancer Institute using cultures of 60 cell lines of nine groups, including leukemia, lung tumor, large intestine tumor, CNS tumor, melanoma, ovary tumor, renal tumor, prostate tumor, and breast tumor. Relationships between structure and antitumor activity were analyzed. In vivo experimental data from hollow fiber tests are presented for two derivatives.     

Molecular genetic methods for the detection of rifampicin-resistant Mycobacterium tuberculosis strains

RCR-heteroduplex (GDA) and chip methods were used to detect rifampricin-resistant (RR) and rifampicin-sensitive (RS) Mycobacterium tuberculosis (MTB) in the samples from patients (sputum) and in the clinical isolates of MTB from these patients (MB/BacT liquid medium and Lowenstein Jensen's (LJ) solid medium. The efficiency of detecting RR and RS of MTB (from the sputum) is 100 and 92.3% in the chip and GDA tests, respectively. Correlations between GDA (sputum) and drug test (LJ) were 91.7%, that of chip (sputum) and drug test LJ, 88.5%, chip (sputum) and chip clinical isolates (LJ), 100%. The efficacy of GDA and chip in the detection of RR of MTB strains is under discussion.