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Background: Disinfectants are important in the food industry to prevent the transmission of pathogens. Excessive use of disinfectants may increase the probability of bacteria experiencing long-term exposure and consequently resistance and cross-resistance to antibiotics. This study aims to investigate the cross-resistance of multidrug-resistant, drug-resistant, and drug-susceptible isolates of Salmonella enterica serovar Typhimurium (S. Typhimurium) with different sequence types (STs) to a group of antibiotics after exposure to different food-grade disinfectants. Methods: A panel of 27 S. Typhimurium strains with different antibiograms and STs were exposed to increasing concentrations of five food-grade disinfectants, including hydrogen peroxide (H2O2), benzalkonium chloride (BAC), chlorine dioxide (ClO2), sodium hypochlorite (NaClO), and ethanol. Recovered evolved strains were analyzed using genomic tools and phenotypic tests. Genetic mutations were screened using breseq pipeline and changes in resistance to antibiotics and to the same disinfectant were determined. The relative fitness of evolved strains was also determined. Results: Following exposure to disinfectants, 22 out of 135 evolved strains increased their resistance to antibiotics from a group of 14 clinically important antibiotics. The results also showed that 9 out of 135 evolved strains had decreased resistance to some antibiotics. Genetic mutations were found in evolved strains. A total of 77.78% of ST34, 58.33% of ST19, and 66.67% of the other STs strains exhibited changes in antibiotic resistance. BAC was the disinfectant that induced the highest number of strains to cross-resistance to antibiotics. Besides, H2O2 induced the highest number of strains with decreased resistance to antibiotics. Conclusions: These findings provide a basis for understanding the effect of disinfectants on the antibiotic resistance of S. Typhimurium. This work highlights the link between long-term exposure to disinfectants and the evolution of resistance to antibiotics and provides evidence to promote the regulated use of disinfectants. [ABSTRACT FROM AUTHOR]

Background: This study aimed to comprehensively evaluate the efficacy and toxicity of afatinib in patients with sarcopenia, an important prognostic factor for treatment efficacy and toxicity in patients with cancer. Methods: The clinical features of patients with advanced NSCLC treated with frontline afatinib between 2014 and 2018 at a medical center in Taiwan were retrospectively reviewed. Sarcopenia was evaluated based on the total cross‐sectional area of skeletal muscles assessed by computed tomography (CT) imaging at the L3 level. Baseline characteristics, response rates, survival rates, and adverse events (AEs) were compared between sarcopenic and nonsarcopenic patients. Results: A total of 176 patients evaluated for sarcopenia by CT and treated with afatinib were enrolled in the current study. Sarcopenia was significantly associated with good performance status, low body mass index (BMI), low body surface area (BSA), and low total mass area (TMA). Sarcopenia did not influence the response rate (69.2% vs. 72.0%, p = 0.299), progression‐free survival (median 15.9 vs. 14.9 months, p = 0.791), or overall survival (median 26.5 vs. 27.2 months, p = 0.441). However, BSA ≤ 1.7 and the 40 mg afatinib dose were associated with dose reduction. TMA was the only independent factor for afatinib discontinuation due to AEs. Conclusion: Sarcopenia was not associated with treatment efficacy or toxicity among patients with NSCLC harboring common mutations treated with afatinib, indicating sarcopenic patients should not be excluded from afatinib treatment. Other factors, such as BSA and TMA, were associated with dose reduction and afatinib discontinuation, respectively, which may require additional evaluations in future studies. [ABSTRACT FROM AUTHOR]

Background: Programmed death‐ligand 1 (PD‐L1) expression is associated with clinical outcomes of epidermal growth factor receptor (EGFR) mutant lung adenocarcinoma (ADC) treated with tyrosine kinase inhibitors (TKIs). However, whether PD‐L1 expression plays a role in anaplastic lymphoma kinase (ALK)‐positive lung ADC is unknown. We aimed to evaluate the impact of PD‐L1 in patients with ALK‐positive lung ADC receiving crizotinib. Materials and Methods: PD‐L1 expression was identified by immunohistochemistry (IHC). Reverse transcriptase‐polymerase chain reaction was used for ALK variant detection, and immunofluorescence‐based multiplex staining was applied for exploring immune cells in tumor microenvironments. Results: A total of 78 patients with ALK‐positive advanced ADC were enrolled in our study, of whom 52 received crizotinib. Compared with EGFR/ALK wild‐type tumors, PD‐L1 expression was lower in ALK‐positive ADC. ALK fusion variants were identified in 32 patients, and those with variant 3 and 5 (short variants) had higher PD‐L1 expression than those with other variants. The crizotinib objective response rate (ORR) and progression‐free survival (PFS) was better in tumors with negative PD‐L1 expression (ORR/PFS in PD‐L1 0% vs. 1%–49% vs. 50%–100%: 60.7%/11.8 months vs. 38.5%/6.5 months vs. 36.4%/4.0 months, p =.007/.022). The multivariate Cox proportional hazards model revealed that PD‐L1 0% (vs. ≥1%) was an independent factor for longer PFS (adjusted hazard ratio 0.322, 95% confidence interval 0.160–0.650, p =.002). Multiplex IHC in three cases showed a varied extent of immune cell infiltrations in tumors with different PD‐L1 expression. Conclusion: Positive PD‐L1 expression was associated with unfavorable clinical outcomes in patients with ALK‐positive lung ADC receiving crizotinib. Implications for Practice: Not all lung adenocarcinoma with sensitizing driver mutations experienced durable responses to small‐molecule tyrosine kinase inhibitors (TKIs). Similar to the negative impact of programmed death‐ligand 1 (PD‐L1) in epidermal growth factor receptor mutant tumors treated with TKIs, this study demonstrated that positive PD‐L1 expression was also associated with worse response rate and shorter progression‐free survival of anaplastic lymphoma kinase (ALK)‐positive adenocarcinoma treated with crizotinib. Among different ALK fusion partners, tumors with short variants (V3 and V5) had higher PD‐L1 compared with long variants (V1, V2, and V6). Testing PD‐L1 before initiating crizotinib for ALK‐positive lung cancer could be a simple method to provide important prognostic information. This article focuses on the effect of PD‐L1 on ALK‐positive lung adenocarcinoma and examines the association of pretreatment tumor PD‐L1 and clinical outcomes of patients receiving the first approved ALK inhibitor, crizotinib. [ABSTRACT FROM AUTHOR]

Genetic factors are an important cause of idiopathic sensorineural hearing impairment (SNHI). From the epidemiological perspective, mutations of three deafness genes: GJB2, SLC26A4, and MT-RNR1, are much more prevalent than those of other genes worldwide. However, mutation spectra of common deafness genes differ remarkably across different populations. Here, we performed comprehensive genetic examination and haplotype analyses in 188 unrelated Mongolian families with idiopathic SNHI, and compared their mutation spectra and haplotypes to those of other European and Asian cohorts. We confirmed genetic diagnoses in 18 (9.6%) of the 188 families, including 13 with bi-allelic GJB2 mutations, three with bi-allelic SLC26A4 mutations, and two with homoplasmic MT-RNR1 m.1555A>G mutation. Moreover, mono-allelic mutations were identified in 17 families (9.0%), including 14 with mono-allelic GJB2 mutations and three with mono-allelic SLC26A4 mutations. Interestingly, three GJB2 mutations prevalent in other populations, including c.35delG in Caucasians, c.235delC in East Asians, and c.-23+1G>A in Southwest and South Asians, were simultaneously detected in Mongolian patients. Haplotype analyses further confirmed founder effects for each of the three mutations, indicating that each mutation derived from its ancestral origin independently. By demonstrating the unique spectra of deafness-associated mutations, our findings may have important clinical and scientific implications for refining the molecular diagnostics of SNHI in Mongolian patients, and for elucidating the genetic relationships among Eurasian populations. [ABSTRACT FROM AUTHOR]

Rapid metastasis contributes significantly to the high mortality rates in lung adenocarcinoma. The present study reports a rare case of primary lung adenocarcinoma with concomitant gastric, duodenal, bone, and mediastinal lymph node metastases. A large gene panel based on the next-generation sequencing was used to detect gene mutations in different metastatic sites and blood. The results showed that the gene mutation spectrums among different metastatic foci were roughly similar. The mutation abundance was highest in mediastinal lymph nodes. Unique mutation sites were detected only in mediastinal lymph nodes, bone, and gastric metastatic foci. Despite partial heterogeneity, there are currently no applicable targeted drugs to adopt. In addition, tumor mutation burden (TMB) showed that blood, gastric, bone, and mediastinal lymph node metastases were all TMB-High, while the duodenal metastasis was TMB-Low. This study is the first to report a rare case of newly diagnosed lung adenocarcinoma with concomitant gastrointestinal metastases and to perform mutation analyses on all metastatic foci. Large sample sizes with similar cases are required to gain deeper insights. [ABSTRACT FROM AUTHOR]

Objectives: To investigate speech and language outcomes in children with cochlear implants (CIs) who had mutations in common deafness genes and to compare their performances with those without mutations. Study Design: Prospective study. Methods: Patients who received CIs before 18 years of age and had used CIs for more than 3 years were enrolled in this study. All patients underwent mutation screening of three common deafness genes: GJB2, SLC26A4 and the mitochondrial 12S rRNA gene. The outcomes with CIs were assessed at post-implant years 3 and 5 using the Categories of Auditory Performance (CAP) scale, Speech Intelligibility Rating (SIR) scale, speech perception tests and language skill tests. Results: Forty-eight patients were found to have confirmative mutations in GJB2 or SLC26A4, and 123 without detected mutations were ascertained for comparison. Among children who received CIs before 3.5 years of age, patients with GJB2 or SLC26A4 mutations showed significantly higher CAP/SIR scores than those without mutations at post-implant year 3 (p = 0.001 for CAP; p = 0.004 for SIR) and year 5 (p = 0.035 for CAP; p = 0.038 for SIR). By contrast, among children who received CIs after age 3.5, no significant differences were noted in post-implant outcomes between patients with and without mutations (all p > 0.05). Conclusion: GJB2 and SLC26A4 mutations are associated with good post-implant outcomes. However, their effects on CI outcomes may be modulated by the age at implantation: the association between mutations and CI outcomes is observed in young recipients who received CIs before age 3.5 years but not in older recipients. [ABSTRACT FROM AUTHOR]

Background: A novel link between oncogenic KRAS signalling and WT1 was recently identified. We sought to investigate the role of WT1 and KRAS in proliferation and apoptosis. Methods: KRAS mutations and WT1 (cMyc) expression were detected using Sanger sequencing and real-time PCR in 77 patients with non-small cell lung cancer (NSCLC). Overexpression and knockdown of WT1 were generated with plasmid and siRNA via transient transfection technology in H1299 and H1568 cells. MTT assay for detection of cell proliferation, and TUNEL assay and proteomic profiler assay for apoptosis evaluation were carried out. Dual luciferase reporter assay and ChIP-PCR were performed to validate the effect of WT1 on the cMyc promoter. Results: KRAS mutations showed a negative impact on overall survival (OS). High expressions of WT1 and cMyc were associated with poor OS in KRAS mutant subgroup. The potential mechanisms that WT1 promotes proliferation and impedes apoptosis through affecting multiple apoptosis-related regulators in KRAS mutant NSCLC cells were identified. WT1 could activate cMyc promoter directly in KRAS mutant cells. Conclusion: The results suggest that WT1 and c-MYC expression is important for survival in KRAS mutant tumors as opposed to KRAS wild-type tumors. For treatment of KRAS mutant NSCLC, targeting WT1 and cMyc may provide alternative therapeutic strategies. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Aims Me CP2 gene mutations are associated with Rett syndrome and X-linked mental retardation ( XLMR), diseases characterized by abnormal brain development and function. Recently, we created a novel Me CP2 A140V mutation mouse model that exhibited abnormalities of cell packing density and dendritic branching consistent with that seen in Rett syndrome patients as well as other Me CP2 mutant mouse models. Therefore, we hypothesized that some deficits of neuronal and synaptic functions might also be present in the A140V mutant model. Methods Here, we tested our hypothesis in hippocampal slices using electrophysiological recordings. Results We found that in young A140V mutant mice (3- to 4-week-old), hippocampal CA1 pyramidal neurons exhibited more positive resting membrane potential, increased action potential ( AP) firing frequency induced by injection of depolarizing current, wider AP duration, and smaller after hyperpolarization potential compared to neurons prepared from age-matched wild-type mice, suggesting a neuronal hyperexcitation. At the synaptic level, A140V mutant neurons exhibited a reduced frequency of spontaneous IPSCs (inhibitory postsynaptic potentials) and an enhanced probability of evoked glutamate release, both suggesting neuronal hyperexcitation. However, hippocampal CA1 long-term potentiation was not significantly different between A140V and WT mice. In adult mice (11- to 13-month-old), in addition to neuronal hyperexcitation, we also found significant deficits of both short-term and long-term potentiation of CA3- CA1 synapses in A140V mice compared to WT mice. Conclusions These results clearly illustrate the age-dependent abnormalities of neuronal and synaptic function in the Me CP2 A140V mutant mouse model, which provides new insights into the understanding of the pathogenesis of Rett syndrome. [ABSTRACT FROM AUTHOR]

Mutations in the SLC26A4 gene are a common cause of human hereditary hearing impairment worldwide. Previous studies have demonstrated that different SLC26A4 mutations have different pathogenetic mechanisms. By using a genotype-driven approach, we established a knock-in mouse model (i.e., Slc26a4tm2Dontuh/tm2Dontuh mice) homozygous for the common p.H723R mutation in the East Asian population. To verify the pathogenicity of the p.H723R allele in mice, we further generated mice with compound heterozygous mutations (i.e., Slc26a4tm1Dontuh/tm2Dontuh) by intercrossing Slc26a4+/tm2Dontuh mice with Slc26a4tm1Dontuh/tm1Dontuh mice, which segregated the c.919-2A>G mutation with an abolished Slc26a4 function. Mice were then subjected to audiologic assessments, a battery of vestibular evaluations, inner ear morphological studies, and noise exposure experiments. The results were unexpected; both Slc26a4tm2Dontuh/tm2Dontuh and Slc26a4tm1Dontuh/tm2Dontuh mice showed normal audiovestibular phenotypes and inner ear morphology, and they did not show significantly higher shifts in hearing thresholds after noise exposure than the wild-type mice. The results indicated not only the p.H723R allele was non-pathogenic in mice, but also a single p.H723R allele was sufficient to maintain normal inner ear physiology in heterozygous compound mice. There might be discrepancies in the pathogenicity of specific SLC26A4 mutations in humans and mice; therefore, precautions should be taken when extrapolating the results of animal studies to humans. [ABSTRACT FROM AUTHOR]

Germline mutations in genes that cause hereditary syndromes are highly predisposed to familial pancreatic cancer. However, genetic susceptibility to sporadic pancreatic cancer is largely uncovered. We conducted a two-stage association study on pancreatic cancer that included 981 cases and 1991 controls in the first stage followed by a second stage (2603 cases and 2877 controls). Using an approach based on candidate genes whose roles in pancreatic cancer have been well known, we identified two new susceptibility loci. rs11571836 located in the BRCA2 3′-untranslated region was significantly associated with lower expression of BRCA2 transcript and increased pancreatic cancer risk [odds ratio = 1.30, 95% confidence interval = 1.14–1.47, P = 7.64 × 10–5] in a recessive manner. rs12939944 located in the MAP2K4 intron was associated with decreased risk (odds ratio = 0.82, 95% confidence interval = 0.74–0.91, P = 0.0001) in a dominant manner. Our results demonstrate for the first time that common variants in BRCA2 and MAP2K4 are susceptibility to sporadic pancreatic cancer. [ABSTRACT FROM AUTHOR]

Despite the clinical utility of genetic diagnosis to address idiopathic sensorineural hearing impairment (SNHI), the current strategy for screening mutations via Sanger sequencing suffers from the limitation that only a limited number of DNA fragments associated with common deafness mutations can be genotyped. Consequently, a definitive genetic diagnosis cannot be achieved in many families with discernible family history. To investigate the diagnostic utility of massively parallel sequencing (MPS), we applied the MPS technique to 12 multiplex families with idiopathic SNHI in which common deafness mutations had previously been ruled out. NimbleGen sequence capture array was designed to target all protein coding sequences (CDSs) and 100 bp of the flanking sequence of 80 common deafness genes. We performed MPS on the Illumina HiSeq2000, and applied BWA, SAMtools, Picard, GATK, Variant Tools, ANNOVAR, and IGV for bioinformatics analyses. Initial data filtering with allele frequencies (<5% in the 1000 Genomes Project and 5400 NHLBI exomes) and PolyPhen2/SIFT scores (>0.95) prioritized 5 indels (insertions/deletions) and 36 missense variants in the 12 multiplex families. After further validation by Sanger sequencing, segregation pattern, and evolutionary conservation of amino acid residues, we identified 4 variants in 4 different genes, which might lead to SNHI in 4 families compatible with autosomal dominant inheritance. These included GJB2 p.R75Q, MYO7A p.T381M, KCNQ4 p.S680F, and MYH9 p.E1256K. Among them, KCNQ4 p.S680F and MYH9 p.E1256K were novel. In conclusion, MPS allows genetic diagnosis in multiplex families with idiopathic SNHI by detecting mutations in relatively uncommon deafness genes. [ABSTRACT FROM AUTHOR]

Adaptive mutations that have contributed to the emergence of influenza A pandemic (H1N1) 2009 virus, which can replicate and transmit among humans, remain unknown. We conducted a large-scale scanning of influenza protein sequences and identified amino acid-conserving positions that are specific to host species, called signatures. Of 47 signatures that separate avian viruses from human viruses by their nonglycoproteins, 8 were human-like in the pandemic (H1N1) 2009 virus. Close examination of their amino acid residues in the recent ancestral swine viruses of pandemic (H1N1) 2009 virus showed that 7 had already transitioned to human-like residues and only PA 356 retained an avian-like K; in pandemic (H1N1) 2009 virus, this residue changed into a human-like R. Signatures that separate swine viruses from human viruses were also present. Continuous monitoring of these signatures in nonhuman species will help with influenza surveillance and with evaluation of the likelihood of further adaptation to humans. [ABSTRACT FROM AUTHOR]

Febrile seizures (FS) are common in children, and the incidence is 2–5% before the age of 5 years. A four-generation Chinese family with autosomal dominant febrile seizure and epilepsy was studied by genome-wide linkage analysis. Significant linkage was identified with markers on chromosome 3q26.2–26.33 with a maximum pairwise LOD score of >3.00. Fine mapping defined the new genetic locus within a 10.7-Mb region between markers D3S3656 and D3S1232. A maximum multipoint LOD score of 5.27 was detected at marker D3S1565. A previously reported CLCN2 gene for epilepsy was excluded as the disease-causing gene in the family by mutational analysis of all exons and exon–intron boundaries of CLCN2 and by haplotype analysis. Mutation analysis of KCNMB2 and KCNMB3, which were two potassium-channel genes in this linkage region, did not reveal a disease causing mutation. Our results identified another novel locus on chromosome 3q26.2–26.33, and future studies of the candidate genes at the locus will identify a new gene for combined FS and idiopathic epilepsies. [ABSTRACT FROM AUTHOR]

The 3C protease (3C[sup pro] ) of enterovirus 71 (EV71) is a good molecular target for drug discovery. Notably, this protease was found to possess RNA-binding activity. The regions responsible for RNA binding were classified as ‘KFRDI’ (positions 82–86) and ‘VGK’ (positions 154–156) in 3C[sup pro] by mutagenesis study. Although the RNA-binding regions are structurally distinct from the catalytic site of EV71 3C[sup pro] , mutations in the RNA-binding regions influenced 3C[sup pro] proteolytic activity. In contrast, mutations at the catalytic site had almost no influence on RNA binding ability. We identified certain mutations within 3C[sup pro] which abrogated both the RNA-binding activity of the expressed, recombinant, protease and the ability to rescue virus from an infectious full-length clone of EV71 (pEV71). Interestingly, mutation at position 84 from Arg(R) to Lys(K) was found to retain good RNA binding and proteolytic activity for the recombinant 3C[sup pro] ; however, no virus could be rescued when pEV71 with the R84K mutation was introduced into the infectious copy. Together, these results may provide useful information for using 3C[sup pro] as the molecular target to develop anti-EV71 agents. Copyright © 2004 National Science Council, ROC and S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Aurora A kinase (AURKA) is an important regulator in mitotic progression and is overexpressed frequently in human cancers, including hepatocellular carcinoma (HCC). Many AURKA mutations were identified in cancer patients. Overexpressing wild-type Aurka developed a low incidence of hepatic tumors after long latency in mice. However, none of the AURKA mutant animal models have ever been described. The mechanism of mutant AURKA-mediated hepatocarcinogenesis is still unclear. A novel AURKA mutation with a.a.352 Valine to Isoleucine (V352I) was identified from clinical specimens. By using liver-specific transgenic fish overexpressing both the mutant and wild-type AURKA, the AURKA(V352I)-induced hepatocarcinogenesis was earlier and much more severe than wild-type AURKA. Although an increase of the expression of lipogenic enzyme and lipogenic factor was observed in both AURKA(V352I) and AURKA(WT) transgenic fish, AURKA(V352I) has a greater probability to promote fibrosis at 3 months compared to AURKA(WT). Furthermore, the expression levels of cell cycle/proliferation markers were higher in the AURKA(V352I) mutant than AURKA(WT) in transgenic fish, implying that the AURKA(V352I) mutant may accelerate HCC progression. Moreover, we found that the AURKA(V352I) mutant activates AKT signaling and increases nuclear β-catenin, but AURKA(WT) only activates membrane form β-catenin, which may account for the differences. In this study, we provide a new insight, that the AURKA(V352I) mutation contributes to early onset hepatocarcinogenesis, possibly through activation of different pathways than AURKA(WT). This transgenic fish may serve as a drug-screening platform for potential precision medicine therapeutics. [ABSTRACT FROM AUTHOR]

Different amino acid sequences of influenza virus proteins contribute to different viral phenotypes. However, the diversity of the sequences and its impact on noncoding regions or splice sites have not been intensively studied. This study focuses on the sequences at alternative 5′ splice sites on M1 mRNA. Six different mutations at the splice sites were introduced, and viral growth characteristics for those mutants generated by reverse genetics with 12 plasmids were examined, for which G12C (the G-to-C mutation at the first nucleotide of the intron for the mRNA3 5′ splice site), C51G (at the 3′ end of the exon of the M2 mRNA 5′ splice site), and G146C (for the first nucleotide of the intron for mRNA4) are lethal mutations. On the other hand, mutants with the mutation G11C (at the 3′ end of exon of the mRNA3 5′ splice site), G52C (for the first nucleotide of the intron for M2 mRNA), or G145A (at the 3′ end of the exon of mRNA4) were rescued, although they had significantly attenuated growth rates. Notably, these mutations did not change any amino acids in M1 or M2 proteins. The levels of precursor (M1 mRNA) and spliced products (M2 mRNA, mRNA3, and mRNA4) from the recombinant mutant virus-infected cells were further analyzed. The production levels of mRNA3 in cells infected with G11C, G52C, and G145A mutant viruses were reduced in comparison with that in wild-type recombinant virus-infected ones. More M2 mRNA was produced in G11C mutant virus-infected cells than in wild-type-virus-infected cells, and there was little M2 mRNA and none at all in G145A and G52C mutant virus-infected ones, respectively. Results obtained here suggest that introducing these mutations into the alternative 5′ splice sites disturbed M1 mRNA splicing, which may attenuate viral growth rates. [ABSTRACT FROM AUTHOR]

The postnatal neurodevelopmental disorder Rett syndrome, caused by mutations in MECP2, produces a diverse array of symptoms, including loss of language, motor, and social skills and the development of hand stereotypies, anxiety, tremor, ataxia, respiratory dysrhythmias, and seizures. Surprisingly, despite the diversity of these features, we have found that deleting Mecp2 only from GABAergic inhibitory neurons in mice replicates most of this phenotype. Here we show that genetically restoring Mecp2 expression only in GABAergic neurons of male Mecp2 null mice enhanced inhibitory signaling, extended lifespan, and rescued ataxia, apraxia, and social abnormalities but did not rescue tremor or anxiety. Female Mecp2+/- mice showed a less dramatic but still substantial rescue. These findings highlight the critical regulatory role of GABAergic neurons in certain behaviors and suggest that modulating the excitatory/inhibitory balance through GABAergic neurons could prove a viable therapeutic option in Rett syndrome. [ABSTRACT FROM AUTHOR]