Association between <Emphasis Type="Italic">ATM</Emphasis> gene polymorphisms,lung cancer susceptibility and radiation-induced pneumonitis: a meta-analysis

Background

Previous studies have suggested that DNA double-strand break (DSB) repair is an important protective pathway after damage. The ataxia telangiectasia mutated (ATM) gene plays an important role in the DNA DSB repair pathway. DNA damage is a major cytotoxic effect that can be caused by radiation, and the ability to repair DNA after damage varies among different tissues. Impaired DNA repair pathways are associated with high sensitivity to radiation exposure. Hence, ATM gene polymorphisms are thought to influence the risk of cancer and radiation-induced pneumonitis (RP) risk in cancer patients treated with radiotherapy. However, the results of previous studies are inconsistent. We therefore conducted this comprehensive meta-analysis.

Methods

A systematic literature search was performed in the PubMed, Embase, China National Knowledge Internet (CNKI) and Wanfang databases to identify studies that investigated the association between the ATM gene polymorphisms and both lung cancer and RP radiotherapy-treated lung cancer (the last search was conducted on Dec.10, 2015). The odds ratio (OR) and 95% confidence interval (CI) were used to investigate the strength of these relationships. Funnel plots and Begg’s and Egger’s tests were conducted to assess the publication bias. All analyses were performed in STATA 13.0 software.

Results

Ten eligible case-control studies (4731 cases and 5142 controls) on lung cancer susceptibility and four (192 cases and 772 controls) on RP risk were included. The results of the overall and subgroup analyses indicated that in the ATM gene, the rs189037 (?111G?>?A, ?4519G?>?A), rs664677 (44831C?>?T, 49238C?>?T) and rs664143 (131,717 T?>?G) polymorphisms were significantly associated with lung cancer susceptibility (OR?=?1.21, 95% CI?=?1.04–1.39, P?=?0.01; OR?=?1.26, 95% CI?=?1.06–1.49, P?=?0.01; OR?=?1.43, 95% CI?=?1.15–1.78, P?<?0.01). Additionally, the rs189037 variant was significantly associated with RP risk (OR?=?1.74, 95% CI?=?1.02–2.97, P?=?0.04). No publication bias was found in the funnel plots, Begg’s tests or Egger’s tests.

Conclusions

The results indicate that the ATM rs189037, rs664677 and rs664143 gene polymorphisms are risk factors for lung cancer, while the ATM rs189037 variant was significantly associated with RP risk.

     

Effects of chocolate containing <Emphasis Type="Italic">Leuconostoc mesenteroides</Emphasis> strain NTM048 on immune function: a randomized,double-blind,placebo-controlled trial

Background

Previous reports showed that oral administration of Leuconostoc mesenteroides strain NTM048 increases IgA levels and CD4+ T cell population in feces and mice, respectively, as revealed by flow cytometric analysis of splenocytes. This study aimed to evaluate the effect of chocolate supplemented with L. mesenteroides strain NTM048 (>?1.00?×?10⁹?CFU/day, NTM048) on the immune parameters of healthy subjects, using a randomized, placebo-controlled, double-blinded study design.

Methods

Participants (mean age: 46.3?years) ingested 28?g of test food daily, at a time of their own choice, for 4?weeks. The immunological parameters of all participants were evaluated two times (pre- and post- ingestion). At the end of the study, various immunological parameters of the participants were measured and scoring of immunological vigor (SIV) was performed using a comprehensive algorithm.

Results

Ingestion of NTM048-supplemented chocolate significantly improved SIV in the NTM048 group (18.6?±?1.6) compared to that in the placebo group (17.8?±?2.0) after 4?weeks (p?=?0.049). Several immunological parameters (CD8⁺T cells, CD8⁺CD28⁺ T cells, and memory T cells) were significantly elevated in the NTM048 group as compared to the placebo group (all p?<?0.05). In addition, T cell proliferation index at post-ingestion significantly increased compared with that at pre-ingestion in the NTM048 (p?=?0.017) and placebo groups (p?=?0.037), although no differences were observed between the two groups.

Conclusion

Our results suggest that ingestion of chocolate supplemented with NTM048 is effective against the age-related decline in T cell-related immune functions.

Trial registration

UMIN Clinical Trials Registry UMIN000021989. Registered 19 April 2016, https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000025321

     

BrFLC5: a weak regulator of flowering time in <Emphasis Type="Italic">Brassica rapa</Emphasis>

Key message

A splicing site mutation in BrFLC5, a non-syntenic paralogue of FLOWERING LOCUS C, was demonstrated to be related to flowering time variation in Brassica rapa.

Abstract

Flowering time regulation in Brassica rapa is more complex than in Arabidopsis, as there are multiple paralogues of flowering time genes in B. rapa. Brassica rapa contains four FLOWERING LOCUS C (FLC) genes, three of which are syntenic orthologues of AtFLC, while BrFLC5 is not. BrFLC1, BrFLC2, and BrFLC3 have been reported to be involved in flowering time regulation. However, BrFLC5 has thus far been deemed a pseudogene. We detected two alternative splicing patterns of BrFLC5 resulting from a nucleotide mutation (G/A) at the first nucleotide of intron 3 (named as Pi3+1(G/A)). Genotyping of BrFLC5Pi3?+?1(G/A) for 301 B. rapa accessions showed that this single nucleotide polymorphism was significantly related to flowering time variation (p?<?0.001). In the collection, the frequency of the functional G allele (35.2%) was much lower than that of the nonfunctional A allele (59.1%); however, the frequency of the G allele was very high among the turnips (83.6%). An F2 population segregating at this locus was developed to analyze the genetic effect of BrFLC5. The result showed that the G allele individuals began to bolt two days later than the A allele individuals, indicating that BrFLC5 is a weak regulator of flowering time. BrFLC5 was expressed at the lowest level among the three analyzed BrFLCs. The late allele (G allele) was dominant to the early allele (A allele) at the BrFLC5 locus, which was in contrast to that of BrFLC1 and BrFLC2. This characteristic suggests that BrFLC5 would be more efficient for breeding premature bolting resistance in B. rapa.

     

Decreased formation of branched-chain short fatty acids in <Emphasis Type="Italic">Bacillus amyloliquefaciens</Emphasis> by metabolic engineering

Objectives

To reduce the unpleasant odor during 1-deoxynojirimycin (DNJ) production, the genes of leucine dehydrogenase (bcd) and phosphate butryltransferase (ptb) were deleted from Bacillus amyloliquefaciens HZ-12, and the concentrations of branched-chain short fatty acids (BCFAs) and DNJ were compared.

Results

By knockout of the ptb gene, 1.01 g BCFAs kg^?1 was produced from fermented soybean by HZ-12Δptb. This was a 56% decrease compared with that of HZ-12 (2.27 g BCFAs kg^?1). Moreover, no significant difference was found in the DNJ concentration (0.7 g kg^?1). After further deletion of the bcd gene from HZ-12Δptb, no BCFAs was detected in fermented soybeans with HZ-12ΔptbΔbcd, while the DNJ yield decreased by 26% compared with HZ-12.

Conclusions

HZ-12Δptb had decreased BCFAs formation but also maintained the stable DNJ yield, which contributed to producing DNJ-rich products with decreased unpleasant smell.

     

Association between <Emphasis Type="Italic">TNF,IL1B,IL6, IL10</Emphasis> and <Emphasis Type="Italic">IFNG</Emphasis> polymorphisms and recurrent miscarriage: a case control study

Background

Approximately half of recurrent miscarriages have unexplained etiology. Recent evidences suggest that cytokines are important determinants in pregnancy maintenance and as such, cytokine gene polymorphisms, which can affect cytokine production and/or functionality, could play a role in the disorder. Thus, we aimed to investigate the association of selected cytokine gene polymorphisms with risk of recurrent miscarriage among Chinese.

Methods

TNF -238G > A, TNF -308G > A, IL1B -511 T > C, IL1B 3954C > T, IL6 -174G > C, IL6 -634C > G, IL10 -1082A > G and IFNG 874A > T polymorphisms were genotyped on 775 women with idiopathic recurrent miscarriage and 805 healthy parous control women. Logistic regression analysis was performed to determine the odds ratios (ORs) of the association between the polymorphisms and recurrent miscarriage risk.

Results

Among the eight polymorphisms studied, only the IL1B -511 T > C and IL6 -634C > G polymorphisms showed statistically significant associations with recurrent miscarriage risk. For the former, a significantly increased risk of recurrent miscarriage was observed for the mutant (CC) genotype (OR: 1.377; 95% CI: 1.039–1.824; P?=?0.026). However, for the IL6 -634C?>?G polymorphism, a decreased recurrent miscarriage risk was observed for the heterozygous (CG) genotype (OR: 0.614; 95% CI: 0.493–0.765; P < 0.001) and the mutant (GG) genotype (OR: 0.414; 95% CI: 0.251–0.684; P?=?0.001).

Conclusions

The IL1B -511 T > C polymorphism may serve as important risk factor for recurrent miscarriage while the IL6 -634C > G polymorphism may protect against the risk of recurrent miscarriage.

     

Factors associated with caregiving burden and mental health conditions in caregivers of patients with anorexia nervosa in Japan

Background

There are no studies about the caregiving burdens in families of patients with eating disorders in Japan, and only limited studies on the role of caregivers’ stress coping, social support, and mental health. This study examines caregiving burdens, mental health conditions, and associated factors in caregivers of anorexia nervosa (AN) patients in Japan.

Methods

Seventy-nine principal caregivers (70 mothers, 5 fathers, 3 spouses and 1 grandmother; mean age 56.0?±?8.0 years) for outpatients with AN (all female; mean age 26.6?±?7.9 years; BMI 14.6?±?3.2 kg/m²) were evaluated using self-report questionnaires in a cross-sectional study. The questionnaires included caregiving burden (J-ZBI_8), mental health conditions (GHQ28), stress coping styles (CISS), social support (SNQ), severity of the patient’s symptoms from the family’s perspective (ABOS), and family functioning (GF-FAD). Clinical information about the patients was also obtained.

Results

Mean caregiving burden assessed by J-ZBI_8 score was 12.4?±?7.0 (SD). The total GHQ score was 31.6?±?13.7 (Likert scoring) and 9.2?±?7.0 (GHQ scoring). Of the respondents, 48 (60.7 %) indicated a high risk for mental health problems that exceeded the cutoff point of the GHQ. Significantly higher caregiving burden and poor mental health conditions were shown in the group who had contact with patients?>?6 h a day compared to the group with daily patient contact?<?3 h (F (2, 76)?=?3.19, p?=?0.047 and F (2, 76)?=?9.39, p?<?0.001, respectively). Stepwise multiple regression analysis indicated that the factors that significantly predicted the caregiving burden were severity of the patient’s symptoms from the family’s perspective (β?=?0.47, p?<?0.001) and Emotion-Oriented Coping (β?=?0.38, p?=?0.002) (R²?=?0.401), while predictors of mental health conditions were Emotion-Oriented Coping (β?=?0.522, p?<?0.001), Affective Support (β?=??0.419, p?<?0.001), and contact time with patient (β?=?0.201, p?=?0.042) (R²?=?0.602).

Conclusion

Caregivers of AN patients experienced heavy burdens and manifested poor mental health conditions. The severity of the patient’s symptoms from the family’s perspective and the greater use of emotion-oriented coping were associated with higher burdens. Greater use of emotion-oriented coping, less affective support and longer contact with patients were related to worse mental health conditions. Interventions to promote caregivers’ adaptive coping styles may help reduce their caregiving burden and improve their mental health.

     

Enhancement of Skin Permeation and Skin Immunization of Ovalbumin Antigen <Emphasis Type="Italic">via</Emphasis> Microneedles

The purpose of this study was to evaluate the use of different types of microneedles and doses of ovalbumin antigen for in vitro skin permeation and in vivo immunization. In vitro skin permeation experiments and confocal laser scanning microscopy revealed that hollow microneedles had a superior enhancing effect on skin permeation compared with a solid microneedle patch and untreated skin by efficiently delivering ovalbumin-fluorescein conjugate into the deep skin layers. The flux and cumulative amount of ovalbumin-fluorescein conjugate at 8 h after administering with various conditions could be ranked as follows: hollow MN; high dose?>?medium dose?>?low dose?>?MN patch; high dose?>?medium dose?>?low dose?>?untreated skin; high dose?>?medium dose?>?low dose?>?without ovalbumin-fluorescein conjugate. As the dose of ovalbumin-fluorescein conjugate was increased to 500 μg, the antigen accumulated in the skin to a greater extent, as evidenced by the increasing green fluorescence intensity. When the hollow microneedle was used for the delivery of ovalbumin into the skin of mice, it was capable of inducing a stronger immunoglobulin G immune response than conventional subcutaneous injection at the same antigen dose. Immunoglobulin G levels in the hollow MN group were 5.7, 11.6, and 13.3 times higher than those of the subcutaneous injection group for low, medium, and high doses, respectively. Furthermore, the mice immunized using the hollow microneedle showed no signs of skin infection or pinpoint bleeding. The results suggest that the hollow MN is an efficient device for delivering the optimal dose of antigen via the skin for successful immunization.     

Interactive effects of multiple vernalization (<Emphasis Type="Italic">Vrn-1</Emphasis>)- and photoperiod (<Emphasis Type="Italic">Ppd-1</Emphasis>)-related genes on the growth habit of bread wheat and their association with heading and flowering time

Background

The precise identification of Winterness/Springness (growth habit) for bread wheat, which is determined by genes involved in vernalization and photoperiod, will contribute to the effective utilization of bread wheat varieties. Here, 198 varieties from the Yellow and Huai wheat production region (YHW) in China were collected to identify their vernalization (Vrn-1) and photoperiod (Ppd-1) gene composition via a series of functional markers and their association with vernalization and photoperiod requirements at three locations during two years of experiments. The growth habits were measured during the spring sowing season.

Results

The results showed that the semi-winter varieties (grades1–4) were most prevalent in the population. The relative effects of single Vrn alleles on the growth period, such as heading date (HD) and/or flowering date (FD), were as follows: Vrn-B1b?>?Vrn-B1a?>?Vrn-D1b?>?Vrn-D1a?>?vrn-D1?=?vrn-B1. The interactive effects of Vrn-B1 and Vrn-D1 on HD and FD were identical to those of Vrn-B1b. Approximately 35.3% of the cultivars carried Ppd-B1a (photoperiod-insensitive) and exhibited the earliest HD and FD. The Ppd-D1a-insensitive allele (Hapl II) was carried by just 0.5% of the varieties; however, the other two sensitive alleles were present at a higher frequency, and their effects were slightly weaker than those of Ppd-B1a. In addition, strong interactive effects between Ppd-B1 and Ppd-D1 were detected. In terms of mean values among various genotypes, the effects followed the order of Vrn-1?>?Ppd-1.

Conclusions

According to the results of ANOVA and least significant range (LSR) tests, we can conclude that Vrn-1 rather than Ppd-1 played a major role in controlling vernalization and photoperiod responses in this region. This research will be helpful for precisely characterizing and evaluating the HD, FD and even growth habit of varieties in the YHW at molecular levels.

     

Mutation Screening of the <Emphasis Type="Italic">CHCHD10</Emphasis> Gene in Chinese Patients with Amyotrophic Lateral Sclerosis

Mutations in the coiled-coil-helix-coiled-coil-helix domain-containing protein 10 gene (CHCHD10), involved in mitochondrial function, have recently been reported as a causative gene of amyotrophic lateral sclerosis (ALS). The aim of this study was to obtain the mutation prevalence of CHCHD10 and the phenotypes with mutations in Chinese ALS patients. A cohort of 499 ALS patients including 487 sporadic ALS (SALS) and 12 familial ALS (FALS), from the Department of Neurology, West China Hospital of Sichuan University, were screened for mutations of all exons of the CHCHD10 gene by Sanger sequencing. Novel candidate mutations or variants were confirmed by polymerase chain reaction-restriction fragment length polymorphism in 466 healthy individuals. All patients identified with mutations of CHCHD10 gene were screened for mutations of the common ALS causative genes including C9orf72, SOD1, TARDBP, FUS, PFN1, and SQSTM1. Three heterozygous variants, including two missense mutations (c.275A?>?G (p.Y92C) and c.306G?>?C (p.Q102H)) and a synonymous change c.306G?>?A (p.Q102Q), were found in exon 3 of CHCHD10 in three alive SALS individuals (with the longest disease duration of 8.6 years), all of which were not detected in healthy controls. No mutation in CHCHD10 was identified in FALS patients. No mutation was found in the aforementioned common ALS causative genes in the patients who carried CHCHD10 mutations. The mutation frequency of CHCHD10 (0.4 %, 2/487) in a Chinese SALS population suggests CHCHD10 gene mutation appears to be an uncommon cause of ALS in Chinese populations. CHCHD10 mutations are associated with a slow progression and long disease duration.     

A SNP in the 5′ flanking region of the myostatin-1b gene is associated with harvest traits in Atlantic salmon (Salmo salar)

Background

Myostatin (MSTN) belongs to the transforming growth factor-β superfamily and is a potent negative regulator of skeletal muscle development and growth in mammals. Most teleost fish possess two MSTN paralogues. However, as a consequence of a recent whole genome-duplication event, salmonids have four: MSTN-1 (?1a and -1b) and MSTN-2 (?2a and -2b). Evidence suggests that teleost MSTN plays a role in the regulation of muscle growth. In the current study, the MSTN-1b gene was re-sequenced and screened for SNP markers in a commercial population of Atlantic salmon. After genotyping 4,800 progeny for the discovered SNPs, we investigated their association with eight harvest traits - four body-weight traits, two ratios of weight traits, flesh colour and fat percentage - using a mixed model association analysis.

Results

Three novel SNPs were discovered in the MSTN-1b gene of Atlantic salmon. One of the SNPs, located within the 5′ flanking region (g.1086C?>?T), had a significant association with harvest traits (p?<?0.05), specifically for: Harvest Weight (kg), Gutted Weight (kg), Deheaded Weight (kg) and Fillet Weight (kg). The haplotype-based association analysis was consistent with this result because the two haplotypes that showed a significant association with body-weight traits, hap4 and hap5 (p?<?0.05 and p?<?0.01, respectively), differ by a single substitution at the g.1086C?>?T locus. The alleles at g.1086C?>?T act in an additive manner and explain a small percentage of the genetic variation of these phenotypes.

Conclusions

The association analysis revealed that g.1086C?>?T had a significant association with all body-weight traits under study. Although the SNP explains a small percentage of the variance, our results indicate that a variation in the 5′ flanking region of the myostatin gene is associated with the genetic regulation of growth in Atlantic salmon.

     

Polymorphisms of caprine <Emphasis Type="Italic">GnRHR</Emphasis> gene and their association with litter size in West African Dwarf goats

Gonadotropin-releasing hormone receptor (GnRHR) gene is considered a candidate gene for litter size due to its critical role in regulating the activities of hypothalamo-pituitary-gonadal axis which synthesizes and releases gonadotropins. This study was designed to identify mutations within the caprine GnRHR gene and investigate their association with litter size at various parities. Polymorphisms scanning and genotyping of GnRHR gene in West African Dwarf (WAD) goats (n?=?226) revealed three single nucleotide polymorphisms (SNPs), one mutation (g.-29T?>?G) was detected within 5′UTR region while two others (g.48G?>?A and g.209T?>?G) were identified in exon 1. Mutation at g.209T?>?G locus resulted in amino acid change from Methionine to Arginine at position 70 on the polypeptide residue. Based on heterozygosity and polymorphism information content (PIC), WAD goat population diversity at the SNP loci was moderate. Strong linkage disequilibrium (LD) (r²?>?0.98) existed among the detected mutations resulting in three observed haplotypes, two (T-G-T and G-A-G) had cumulative frequency of >?97%. The mutation within 5′UTR region of GnRHR gene (g.-29T?>?G) is novel, being reported in goats for the first time. Association analysis revealed a significant (p?<?0.05) association between allele G at g.-29T?>?G with higher mean litter size for homozygous (GG) mutant does compared with heterozygotes (GT) or homozygotes (TT), while the relationship between SNPs at the two loci detected in exon 1 and litter size was not significant.     

Urine metabolites are associated with glomerular lesions in type 2 diabetes

Introduction

Little is known about the association of urine metabolites with structural lesions in persons with diabetes.

Objectives

We examined the relationship between 12 urine metabolites and kidney structure in American Indians with type 2 diabetes.

Methods

Data were from a 6-year clinical trial that assessed renoprotective efficacy of losartan, and included a kidney biopsy at the end of the treatment period. Metabolites were measured in urine samples collected within a median of 6.5 months before the research biopsy. Associations of the creatinine-adjusted urine metabolites with kidney structural variables were examined by Pearson’s correlations and multivariable linear regression after adjustment for age, sex, diabetes duration, hemoglobin A_1c, mean arterial pressure, glomerular filtration rate (iothalamate), and losartan treatment.

Results

Participants (n?=?62, mean age 45?±?10 years) had mean?±?standard deviation glomerular filtration rate of 137?±?50 ml/min and median (interquartile range) urine albumin:creatinine ratio of 34 (14–85) mg/g near the time of the biopsy. Urine aconitic and glycolic acids correlated positively with glomerular filtration surface density (partial r?=?0.29, P?=?0.030 and r?=?0.50, P?<?0.001) and total filtration surface per glomerulus (partial r?=?0.32, P?=?0.019 and r?=?0.43, P?=?0.001). 2-ethyl 3-OH propionate correlated positively with the percentage of fenestrated endothelium (partial r?=?0.32, P?=?0.019). Citric acid correlated negatively with mesangial fractional volume (partial r=-0.36, P?=?0.007), and homovanillic acid correlated negatively with podocyte foot process width (partial r=-0.31, P?=?0.022).

Conclusions

Alterations of urine metabolites may associate with early glomerular lesions in diabetic kidney disease.

     

Micro-NMR elucidates altered metabolites in the Parkinson’s disease-related <Emphasis Type="Italic">catp-6</Emphasis> genotype of <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Introduction

A severe form of Parkinson’s disease (PD) is the Kufor-Rakeb syndrome. Here mutations in the ATP13A2 (PARK9) gene lead to an early juvenile-onset Parkinsonism often accompanied by dementia. ATP13A2 encodes a lysosomal P-type ATPase. Its ortholog in Caenorhabditis elegans is the catp-6 gene where phenotypes with mutations in the alleles ok3473 and tm3190 show high mortality and low reproduction.

Objectives

Since PD is difficult to study in humans we wanted to investigate the potential to use C. elegans as model for the Kufor-Rakeb syndrome. As it is difficult to obtain enough catp-6 mutant worms for standard NMR metabolic profiling, we explored focused ultrasonication extraction and miniaturized NMR as techniques to overcome this limitation.

Methods

One- and two-dimensional NMR experiments (¹ H, JRES, TOCSY) were performed with a commercial high-resolution magic angle spinning (HR-MAS) probe (25 µL sample volume). Significant features were identified through analysis of variance (ANOVA, p?<?0.05), volcano plots (p?<?0.05, fold change?>1.5), PCA, and PLS-DA.

Results

Assignment of statistically relevant peaks resulted in the identification of twenty altered metabolites. Previous studies on catp-6 mutants identified strong morphological and functional changes in their mitochondria. Our findings of altered TCA metabolites (fumarate, succinate), branched-chain amino acids (leucine, isoleucine and valine) and nucleotides (AMP, ATP and GTP), formate and hypoxanthine appear to support these findings. Highest fold changes (< ?5) in wildtype relative to both catp-6 strains were found for GTP. Formic acid is known to inhibit the mitochondrial respiratory chain complex IV and high hypoxanthine in catp-6 indicates an increased nucleotide salvage pathway. Alterations in most of the remaining metabolites may be the result of the recently discovered activation of AMPK (AMP-activated protein kinase) and inhibition of mTOR (mechanistic target of rapamycin) pathways together with a catabolic response to recover energy production.

Conclusions

If the effect of the catp-6 mutation in C. elegans at the level of metabolites is correlated to the metabolic dysfunction in the human PARK9 ortholog, then it may be possible to uncover the molecular mechanism behind Parkinsonism and the Kufor-Rakeb syndrome.

     

Impact of <Emphasis Type="Italic">Mycobacterium tuberculosis</Emphasis> complex lineages as a determinant of disease phenotypes from an immigrant rich moderate tuberculosis burden country

Background

Growing evidences suggested that the Mycobacterium tuberculosis complex (MTBC) lineages can determine the clinical outcome of pulmonary and extra-pulmonary tuberculosis. However, limited data only available revealing such association of bacterial genotypes and clinical phenotypes from immigrant rich countries.

Methods

A multicenter study has been carried out on a collection of 2092 (1003 extrapulmonary and 1089 pulmonary) MTBC isolates. Genotyping of all the isolates were carried out by spoligotyping and 24 loci based MIRU-VNTR typing.

Results

Demographically domination of young Saudi nationals (61.4%) and men (61.2%) were found in this cohort. Lymph nodes (62.4%) and gastrointestinal sites (16.7%) were the most common anatomical sites of infection. The predominant lineages were Delhi/CAS (26.9%), EAI (14.2%) and Ghana (9.9%). Mycobacterium africanum type I and II were reported for the first time in the country among extrapulmonary cases. ‘Ancestral’ lineages M.bovis (OR-5.22; 95% CI-2.23-8.22, p-?<?0.001) and Delhi/CAS (OR-0.57; 95% CI-0.411-0.734, p-?<?0.001) were directly associated with lymph node tuberculosis and gastrointestinal tuberculosis (M. bovis-OR-0.33; 95% CI-0.085-0.567, p-0.001 and Delhi/CAS-OR-1.87; 95% CI-1.22-2.53, p-?<?0.001) respectively. Among the ‘Modern’ lineages, EAI showed significant association to central nervous system tuberculosis (OR-1.98; 95% CI-0.76-3.19, p-0.04) and Uganda-I to gastrointestinal tuberculosis (OR-2.41; 95% CI-0.77-4.06, p-0.02).

Conclusions

The findings substantially contribute to the emerging evidences that MTBC lineages influence disease phenotypes and epidemiological consequences.

     

Phenotypes and <Emphasis Type="Italic">PRRT2</Emphasis> mutations in Chinese families with benign familial infantile epilepsy and infantile convulsions with paroxysmal choreoathetosis

Background

Mutations in the PRRT2 gene have been identified as the major cause of benign familial infantile epilepsy (BFIE), paroxysmal kinesigenic dyskinesia (PKD) and infantile convulsions with paroxysmal choreoathetosis/dyskinesias (ICCA). Here, we analyzed the phenotypes and PRRT2 mutations in Chinese families with BFIE and ICCA.

Methods

Clinical data were collected from 22 families with BFIE and eight families with ICCA. PRRT2 mutations were screened using PCR and direct sequencing.

Results

Ninety-five family members were clinically affected in the 22 BFIE families. During follow-up, two probands had one seizure induced by diarrhea at the age of two years. Thirty-one family members were affected in the eight ICCA families, including 11 individuals with benign infantile epilepsy, nine with PKD, and 11 with benign infantile epilepsy followed by PKD. Two individuals in one ICCA family had PKD or ICCA co-existing with migraine. One affected member in another ICCA family had experienced a fever-induced seizure at 7 years old. PRRT2 mutations were detected in 13 of the 22 BFIE families. The mutation c.649_650insC (p.R217PfsX8) was found in nine families. The mutations c.649delC (p.R217EfsX12) and c.904_905insG (p.D302GfsX39) were identified in three families and one family, respectively. PRRT2 mutations were identified in all eight ICCA families, including c.649_650insC (p.R217PfsX8), c.649delC (p.R217EfsX12), c.514_517delTCTG (p.S172RfsX3) and c.1023A?>?T (X341C). c.1023A?>?T is a novel mutation predicted to elongate the C-terminus of the protein by 28 residues.

Conclusions

Our data demonstrated that PRRT2 is the major causative gene of BFIE and ICCA in Chinese families. Site c.649 is a mutation hotspot: c.649_650insC is the most common mutation, and c.649delC is the second most common mutation in Chinese families with BFIE and ICCA. As far as we know, c.1023A?>?T is the first reported mutation in exon 4 of PRRT2. c.649delC was previously reported in PKD, ICCA and hemiplegic migraine families, but we further detected it in BFIE-only families. c.904_905insG was reported in an ICCA family, but we identified it in a BFIE family. c.514_517delTCTG was previously reported in a PKD family, but we identified it in an ICCA family. Migraine and febrile seizures plus could co-exist in ICCA families.

     

Novel mutation G324C in <Emphasis Type="Italic">WNT1</Emphasis> mapped in a large Pakistani family with severe recessively inherited <Emphasis Type="Italic">Osteogenesis Imperfecta</Emphasis>

Introduction

Osteogenesis imperfecta (OI) is a clinically and genetically heterogeneous disease with skeletal fragility and variable extra-skeletal manifestations. To date several point mutations in 18 different genes causing different types of OI have been identified. Mutations in WNT1 compromise activity of the osteoblasts leading to disturbed bone mass accrual, fragility fractures and progressive skeletal abnormalities. The present study was conducted to determine the underlying genetic cause of an autosomal recessive skeletal dysplasia in a large consanguineous family from Chinute, Pakistan.

Materials and methods

Blood was collected from 24 individuals of affected family along with clinical data. Homozygosity mapping was performed to confirm consanguinity. SNPs were identified, followed by whole exome and Sanger sequencing. In silico characterization of WNT1 mutation was performed using multiple platforms.

Results

Nine affected family members exhibited severe bone deformities, recurrent fractures, short stature and low bone mineral density. SNP array data revealed homozygous segments >?1 Mb in length accounting for 2.1–12.7% of the genome in affected individuals and their siblings and a single 6,344,821 bp homozygous region in all affected individuals on chromosome 12q12-q13. This region includes two potential OI candidate genes WNT1 and VDR. We did whole-exome sequencing for both genes in two patients and identified a novel damaging missense mutation in exon 4 of WNT1: c.1168G?>?T (NM_005430) resulting in p.G324C. Sanger sequencing confirmed segregation of mutation with the disease in family.

Conclusion

We report a novel mutation responsible for OI and our investigation expands the spectrum of disease-causing WNT1 mutations and the resulting OI phenotypes.

     

Changes in Immunity,Expression of some Immune-Related Genes of Shabot Fish, <Emphasis Type="Italic">Tor grypus</Emphasis>, Following Experimental Infection with <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>: Effects of Autochthonous Probiotics

In this study, the effects of orally administrated two native probiotics (Lactobacillus plantarum and Lactobacillus delbrueckii ssp. bulguricus), isolated from the intestine of Shabot fish, Tor grypus, on some immune response parameters and immune-related genes expression against Aeromonas hydrophila in T. grypus were evaluated. Four hundred and eighty juveniles weighing 45?±?10 g were randomly divided into four groups (with three replications) and fed with the experimental diet containing 5?×?10⁷ cfu g^?1 of L. plantarum (G1), Lactobacillus bulgaricus (G2), Lactobacillus casei (G3), and a control diet (without probiotics) for 60 continuous days. At the end of the dietary treatments, fish were challenged with a lethal concentration of A. hydrophila (5?×?10⁸ CFU ml^?1) via intra peritoneal (i.p) injection. Blood and head kidney samples were taken from six fish in each treatment before challenging and 6, 12, 24, and 48 h and also 7 days after injection. The results showed that lysozyme, complement, bactericidal, and NBT activity of probiotic-treated groups were significantly elevated (P?<?0.05). The IL-8, IL-1β, and TNF-α gene expressions were significantly higher in all probiotic-treated groups (P?<?0.05). Meanwhile, a high direct correlation was observed between serum immune parameters and expression of immune-related genes (P?<?0.0001); furthermore, the highest correlation (R ²?=?0.634, P?<?0.0001) was recorded between IL-1β expression and NBT activity. It can be concluded that not only two native probiotics strains stimulate serum immune responses parameters and immune-related gene expression in T. grypus, but also a high correlation was seen among these indices. The study suggests that gastrointestinal colonization is preferred for host specificity as the strain previously derived from shabot fish displayed better colonization than the non-indigenous bacteria strain such as L. casei. Therefore, these native probiotics bacteria can be accounted as suitable candidates to immune stimulation in fish.