Ancestral haplotype reconstruction in endogamous populations using identity-by-descent

In this work we develop a novel algorithm for reconstructing the genomes of ancestral individuals, given genotype or sequence data from contemporary individuals and an extended pedigree of family relationships. A pedigree with complete genomes for every individual enables the study of allele frequency dynamics and haplotype diversity across generations, including deviations from neutrality such as transmission distortion. When studying heritable diseases, ancestral haplotypes can be used to augment genome-wide association studies and track disease inheritance patterns. The building blocks of our reconstruction algorithm are segments of Identity-By-Descent (IBD) shared between two or more genotyped individuals. The method alternates between identifying a source for each IBD segment and assembling IBD segments placed within each ancestral individual. Unlike previous approaches, our method is able to accommodate complex pedigree structures with hundreds of individuals genotyped at millions of SNPs.We apply our method to an Old Order Amish pedigree from Lancaster, Pennsylvania, whose founders came to North America from Europe during the early 18th century. The pedigree includes 1338 individuals from the past 12 generations, 394 with genotype data. The motivation for reconstruction is to understand the genetic basis of diseases segregating in the family through tracking haplotype transmission over time. Using our algorithm thread, we are able to reconstruct an average of 224 ancestral individuals per chromosome. For these ancestral individuals, on average we reconstruct 79% of their haplotypes. We also identify a region on chromosome 16 that is difficult to reconstruct—we find that this region harbors a short Amish-specific copy number variation and the gene HYDIN. thread was developed for endogamous populations, but can be applied to any extensive pedigree with the recent generations genotyped. We anticipate that this type of practical ancestral reconstruction will become more common and necessary to understand rare and complex heritable diseases in extended families.     

Evaluation of the MODS culture technique for the diagnosis of tuberculous meningitis

Background

Tuberculous meningitis (TBM) is a devastating condition. The rapid instigation of appropraite chemotherapy is vital to reduce morbidity and mortality. However rapid diagnosis remains elusive; smear microscopy has extremely low sensitivity on cerebrospinal fluid (CSF) in most laboratories and PCR requires expertise with advanced infrastructure and has sensitivity of only around 60% under optimal conditions. Neither technique allows for the microbiological isolation of M. tuberculosis and subsequent drug susceptibility testing. We evaluated the recently developed microscopic observation drug susceptibility (MODS) assay format for speed and accuracy in diagnosing TBM.

Methodology/Principal Findings

Two hundred and thirty consecutive CSF samples collected from 156 patients clinically suspected of TBM on presentation at a tertiary referal hospital in Vietnam were enrolled into the study over a five month period and tested by Ziehl-Neelsen (ZN) smear, MODS, Mycobacterial growth Indicator tube (MGIT) and Lowenstein-Jensen (LJ) culture. Sixty-one samples were from patients already on TB therapy for >1day and 19 samples were excluded due to untraceable patient records. One hundred and fifty samples from 137 newly presenting patients remained. Forty-two percent (n = 57/137) of patients were deemed to have TBM by clinical diagnostic and microbiological criteria (excluding MODS). Sensitivity by patient against clinical gold standard for ZN smear, MODS MGIT and LJ were 52.6%, 64.9%, 70.2% and 70.2%, respectively. Specificity of all microbiological techniques was 100%. Positive and negative predictive values for MODS were 100% and 78.7%, respectively for HIV infected patients and 100% and 82.1% for HIV negative patients. The median time to positive was 6 days (interquartile range 5–7), significantly faster than MGIT at 15.5 days (interquartile range 12–24), and LJ at 24 days (interquartile range 18–35 days) (P<0.01).

Conclusions

We have shown MODS to be a sensitive, rapid technique for the diagnosis of TBM with high sensitivity, ease of performance and low cost (0.53 USD/sample).     

New records of the genus Iporhogas Granger (Hymenoptera,Braconidae, Rogadinae) from Vietnam,with description of four new species

The genus Iporhogas Granger, 1949 (Braconidae: Rogadinae) is recorded for the first time for Vietnam. Four new species of the genus Iporhogas, viz. Iporhogas albilateralis sp. n., I. contrastus sp. n., I. simulatus sp. n. and I. tricoloratus sp. n., from Vietnam are described and illustrated, and additionally, one species, Iporhogas guangxiensis Chen & He, 1997, is newly recorded for Vietnam’s fauna of the family Braconidae. A key to the five Vietnamese species of the genus Iporhogas and a checklist with distributions of the ten species are provided.     

Purification,characterization and application of a novel antimicrobial peptide from Andrias davidianus blood

The Andrias davidianus has been known as a traditional Chinese medicine for a long time. Its blood is considered as a waste or by‐product of the meat production industry. Although there are reports on isolation of the antimicrobial peptides from different resources, there are no reports of their isolation from A. davidianus blood. In this work, an antimicrobial peptide, andricin B, was isolated from the blood of A. davidianus by an innovative method in which the magnetic liposome adsorption was combined with reversed‐phase high‐performance liquid chromatography. The structure, antimicrobial activity and safety of andricin B were further investigated. Amino acid sequence was determined by N‐terminal sequencing and found to be Gly‐Leu‐Thr‐Arg‐Leu‐Phe‐Ser‐Val‐Ile‐Lys. Circular dichroism (CD) spectra and prediction of three‐dimensional structure by bioinformatics software suggested the presence of a well‐defined random coil conformation. Andricin B was found to be active against all bacteria tested in this study as well as some fungi. The minimum inhibitory concentrations (MICs) were in the range 8–64 μg ml^?1. Moreover, the haemolytic testing also suggested that andricin B could be considered safe at the MICs. Finally, andricin B was shown to inhibit the growth of Staphylococcus aureus in the cooked meat of A. davidianus. This study shows that andricin B is a promising novel antimicrobial peptide that may provide further insights towards the development of new drugs.

Significance and Impact of the Study

This is the pioneer study on screening and isolation of antimicrobial peptide from the blood of Andrias davidianus. Here, we have developed a novel method by combining magnetic liposomes adsorption with reversed‐phase high‐performance liquid chromatography to purify and screen the antimicrobial peptides. From this screen, we identified a novel antimicrobial peptide which we name as andricin B. Andricin B is unique as it checks the growth of both Gram‐positive and Gram‐negative bacteria as well as few fungal species.     

2′-Modified Guanosine Analogs for the Treatment of HCV

Novel 2′-modified guanosine nucleosides were synthesized from inexpensive starting materials in 7–10 steps via hydroazidation or hydrocyanation reactions of the corresponding 2′-olefin. The antiviral effectiveness of the guanosine nucleosides was evaluated by converting them to the corresponding 5′-O-triphosphates (compounds 38–44) and testing their biochemical inhibitory activity against the wild-type NS5B polymerase.     

DNA-binding domain of human c-Myc produced in Escherichia coli.

We have identified the domain of the human c-myc protein (c-Myc) produced in Escherichia coli that is responsible for the ability of the protein to bind sequence-nonspecific DNA. Using analysis of binding of DNA by proteins transferred to nitrocellulose, DNA-cellulose chromatography, and a nitrocellulose filter binding assay, we examined the binding properties of c-Myc peptides generated by cyanogen bromide cleavage, of mutant c-Myc, and of proteins that fuse portions of c-Myc to staphylococcal protein A. The results of these analyses indicated that c-Myc amino acids 265 to 318 were responsible for DNA binding and that other regions of the protein (including a highly conserved basic region and a region containing the leucine zipper motif) were not required. Some mutant c-Mycs that did not bind DNA maintained rat embryo cell-cotransforming activity, which indicated that the c-Myc property of in vitro DNA binding was not essential for this activity. These mutants, however, were unable to transform established rat fibroblasts (Rat-1a cells) that were susceptible to transformation by wild-type c-Myc, although this lack of activity may not have been due to their inability to bind DNA.     

Hydrogen protects against hyperoxia-induced apoptosis in type II alveolar epithelial cells via activation of PI3K/Akt/Foxo3a signaling pathway

Oxidative stress is regarded as a key regulator in the pathogenesis of prolonged hyperoxia-induced lung injury, which causes injury to alveolar epithelial cells and eventually leads to development of bronchopulmonary dysplasia (BPD). Many studies have shown that hydrogen has a protective effect in a variety of cells. However, the mechanisms by which hydrogen rescues cells from damage due to oxidative stress in BPD remains to be fully elucidated. This study sought to evaluate the effects of hydrogen on hyperoxia-induced lung injury and to investigate the underlying mechanism. Primary type II alveolar epithelial cells (AECIIs) were divided into four groups: control (21% oxygen), hyperoxia (95% oxygen), hyperoxia + hydrogen, and hyperoxia + hydrogen + LY294002 (a PI3K/Akt inhibitor). Proliferation and apoptosis of AECIIs were assessed using MTS assay and flow cytometry (FCM), respectively. Gene and protein expression were detected by quantitative polymerase chain reaction (q-PCR) and western blot analysis. Stimulation with hyperoxia decreased the expression of P-Akt, P- FoxO3a, cyclinD1 and Bcl-2. Hyperoxic conditions increased levels of Bim, Bax, and Foxo3a, which induced proliferation restriction and apoptosis of AECIIs. These effects of hyperoxia were reversed with hydrogen pretreatment. Furthermore, the protective effects of hydrogen were abrogated by PI3K/Akt inhibitor LY294002. The results indicate that hydrogen protects AECIIs from hyperoxia-induced apoptosis by inhibiting apoptosis factors and promoting the expression of anti-apoptosis factors. These effects were associated with activation of the PI3K/Akt/FoxO3a pathway.     

α5-nAChR modulates melanoma growth through the Notch1 signaling pathway

The roles of α5-nicotinic acetylcholine receptors (α5-nAChRs) in various types of solid cancer have been reported; however, its role in melanoma remains unknown. We knocked down α5-nAChR expression in melanoma cells to investigate the role of α5-nAChR in the proliferation, migration, and invasion of melanoma cells, and its effect on downstream signaling pathways. Using immunohistochemical analysis, we determined that α5-nAChR expression is significantly increased in human melanoma tissues and cell lines compared with normal human skin tissues. Knocking down α5-nAChR expression in melanoma cells in culture significantly inhibited the proliferation, migration, and invasiveness of melanoma cell lines. Specifically, knockdown of α5-nAChR inhibited PI3K-AKT and ERK1/2 signaling activity. Moreover, we confirmed that the Notch1 signaling pathway is the downstream target of α5-nAChR in melanoma. Our findings suggest that α5-nAChR plays a critical role in melanoma development and progression, and that targeting α5-nAChR may be a strategy for melanoma treatment.     

Minimum population size,genetic diversity,and social structure of the Asian elephant in Cat Tien National Park and its adjoining areas,Vietnam, based on molecular genetic analyses

Vietnam’s elephant population that has suffered severe declines during the past three decades is now believed to number 60–80 individuals in the wild. Cat Tien National Park is thought to be one of the key areas for the recovery of Vietnam’s elephants. We carried out a molecular genetic study of elephants in Cat Tien National Park and its adjoining areas with the objectives of estimating minimum population size, assessing genetic diversity, and obtaining insights into social organization. We obtained a minimum population size of 11 elephants based on a combination of unique nuclear microsatellite genotypes and mitochondrial haplotypes. While mitochondrial diversity based on a 600-base pair segment was high in this small sample of individuals, the six microsatellite loci examined showed low diversity and the signature of a recent population bottleneck. Along with nuclear genetic depauperation of Cat Tien’s elephants, we also report disruption of normal social organization, with different matrilines having coalesced into a single social group because of anthropogenic disturbance. The results emphasize the critical condition of this elephant population and the need for urgent conservation measures if this population is to be saved.     

Cutting edge: regulation of uterine NKT cells by a fetal class I molecule other than CD1

The peri-implantation uterus contains an expanded population of NK1.1(+) V alpha 14(+) TCR(int) (NKT) lymphocytes. Although these cells bear the above features in common with other NKT cells populations in thymus, bone marrow, liver, and spleen, they differ from these other populations in terms of an altered V beta repertoire and absence of a CD4(+) component. In this study, we demonstrate that the uterine population also differs from other NKT cell populations because they recognize a class I/class I-like molecule other than CD1, whereas most previously described V alpha 14(+) NKT cells are CD1-restricted. Moreover, the class I/class I-like molecule leading to the uterine NKT cell expansion may be supplied by the fetus. These data demonstrate a novel mechanism whereby the fetus is capable of modulating the maternal immune system.