Emergence of Amoxicillin Resistance and Identification of Novel Mutations of the pbp1A Gene in Helicobacter pylori in Vietnam

Background Amoxicillin resistant Helicobacter pylori (H. pylori) strains seem to have increased over time in Vietnam. This threatens the effectiveness of H. pylori eradication therapies with this antibiotic. This study aimed to investigate the prevalence of primary resistance of H. pylori to amoxicillin and to assess its association with pbp1A point mutations in Vietnamese patients. Materials and Methods Naive patients who presented with dyspepsia undergoing upper gastrointestinal endoscopy were recruited. Rapid urease tests and PCR assays were used to diagnose H. pylori infection. Amoxicillin susceptibility was examined by E-tests. Molecular detection of the mutant pbp1A gene conferring amoxicillin resistance was carried out by real-time PCR followed by direct sequencing of the PCR products. Phylogenetic analyses were performed using the Tamura-Nei genetic distance model and the neighbour-joining tree building method. Results There were 308 patients (46.1% men and 53.9% women, p = 0.190) with H. pylori infection. The mean age of the patients was 40.5 ± 11.4 years, ranging from 18 to 74 years old. The E-test was used to determine the susceptibility to amoxicillin (minimum inhibitory concentration (MIC) ≤ 0.125 µg/ml) in 101 isolates, among which the rate of primarily resistant strains to amoxicillin was 25.7%. Then, 270 sequences of pbp1A gene fragments were analysed. There were 77 amino acid substitution positions investigated, spanning amino acids 310–596, with the proportion varying from 0.4–100%. Seven amino acid changes were significantly different between amoxicillin-sensitive (AmoxS) and amoxicillin-resistant (AmoxR) samples, including Phe366 to Leu (p < 0.001), Ser414 to Arg (p < 0.001), Glu/Asn464−465 (p = 0.009), Val469 to Met (p = 0.021), Phe473 to Val (p < 0.001), Asp479 to Glu (p = 0.044), and Ser/Ala/Gly595−596 (p = 0.001). Phylogenetic analyses suggested that other molecular mechanisms might contribute to amoxicillin resistance in H. pylori in addition to the alterations in PBP1A. Conclusions We reported the emergence of amoxicillin-resistant Helicobacter pylori strains in Vietnam and new mutations statistically associated with this antimicrobial resistance. Additional studies are necessary to identify the mechanisms contributing to this resistance in Vietnam.

Splitting Choice and Computational Complexity Analysis of Decision Trees

Some theories are explored in this research about decision trees which give theoretical support to the applications based on decision trees. The first is that there are many splitting criteria to choose in the tree growing process. The splitting bias that influences the criterion chosen due to missing values and variables with many possible values has been studied. Results show that the Gini index is superior to entropy information as it has less bias regarding influences. The second is that noise variables with more missing values have a better chance to be chosen while informative variables do not. The third is that when there are many noise variables involved in the tree building process, it influences the corresponding computational complexity. Results show that the computational complexity increase is linear to the number of noise variables. So methods that decompose more information from the original data but increase the variable dimension can also be considered in real applications.

Rapid Identification of Common Poisonous Plants in China Using DNA Barcodes

Toxic plants have been a major threat to public health in China. However, identification and tracing of poisoned species with traditional methods are unreliable due to the destruction of plant morphology by cooking and chewing. DNA barcoding is independent of environmental factors and morphological limitations, making it a powerful tool to accurately identify species. In our study, a total of 83 materials from 26 genera and 31 species of 13 families were collected and 13 plant materials were subjected to simulated gastric fluid digestion. Four markers (rbcL, trnH-psbA, matK, and ITS) were amplified and sequenced for all untreated and mock-digested samples. The effectiveness of DNA barcoding for the identification of toxic plants was assessed using Basic Local Alignment Search Tool (BLAST) method, PWG-Distance method, and Tree-Building (NJ) method. Except for the matK region, the amplification success rate of the remaining three regions was high, but the sequencing of trnH-psbA and ITS was less satisfactory. Meanwhile, matK was prone to be more difficult to amplify and sequence because of simulated gastric fluid. Among the three methods applied, BLAST method showed lower recognition rates, while PWG-Distance and Tree-Building methods showed little difference in recognition rates. Overall, ITS had the highest recognition rate among individual loci. Among the combined loci, rbcL + ITS had the highest species recognition rate. However, the ITS region may not be suitable for DNA analysis of gastric contents and the combination of loci does not significantly improve species resolution. In addition, identification of species to the genus level is sufficient to aid in the clinical management of most poisoning events. Considering primer versatility, DNA sequence quality, species identification ability, experimental cost and speed of analysis, we recommend rbcL as the best single marker for clinical identification and also suggest the BLAST method for analysis. Our current results suggest that DNA barcoding can rapidly identify and trace toxic species and has great potential for clinical applications. In addition, we suggest the creation of a proprietary database containing morphological, toxicological and molecular information to better apply DNA barcoding technology in clinical diagnostics.

Evolution of molecular pathways of Tregs mediated Suppression

Treg suppression of conventional T cells is a fundamental step in regulating the adaptive immune system function. It is known that Treg first appeared in vertebrates. However little is known about the evolutionary history of suppression pathways mediated by Tregs. We employed AI text mining system to highlight the suppression pathways currently known to be utilized by Tregs. Our system identified various pathways such as CTLA4, induction of apoptosis, calcium signaling, inhibition of NfkB and NFAT. After that we employed phylogenetic analysis including multiple sequence alignment, phylogenetic tree building, ancestral sequence reconstruction, neutrality tests and positive selection test to investigate the evolutionary history of Treg mediated pathways in more details. We found that CTLA4 first appeared in vertebrates possibly rising from an IGV containing protein in cartilaginous fish. Conversely, we found that Tregs repurposed ancient pathways such as Calcineurin and CAMP Response Element Modulator that both exists as far as amoeba. Interestingly we found that these two pathways were highly conserved between vertebrates and lower invertebrates indicating conservation of function. Taken together, our research indicate that Tregs evolved its regulatory systems that evolved in vertebrates as well as reused conserved ancient regulatory systems that are related to the innate immune system.

Molecular Decoding of Identity of Some Commercially Important Vandaceous Orchids (Orchidaceae) Based on the Sequences of the Internal Transcribed Spacer (ITS) Sequences and Their Phylogeny

Indiscriminate, rampant collection and natural habitat destruction of Vandaceous orchids for their ornamental and medicinal value have resulted in threatened category to many of the important species and struggling for their survival. Unconfirmed commercially important Vandaceous orchids still continues as these orchids are similar in shaped, size and difficult to identify and classify, especially when they are not in flowering stage. To solve this problem molecular barcode can be an useful tool for rapid identification even from vegetative tissues for commercial purpose. The present study was carried out to test the discriminatory power of IT1 and IT2 region in barcoding and to ascertain their phylogenetic relationship among these 31 species (67) from 15 genera of vandaceous orchids. The sequences were aligned using ClustalW and genetic distances were computed using MEGA 7.0. Data analyses suggest that Internal Transcribed Spacer (ITS) of nuclear ribosomal DNA is a reliable marker which can be use as an efficient barcode to identify (species resolution at 95.52% by genetic distance, 79.40% by BLAST analysis and 95.52% by phylogenetic tree building method) and analyze their phylogenetic relationship among Vandaceous orchids.

Large-scale Phylogenomic Analysis Provides New Insights Into the Phylogeny of the Order Gadiformes and Evolution of Freshwater Gadiform Species Burbot (Lota lota)

Our understanding of phylogenetic relationships among Gadiformes fish is obtained through the analysis of a small number of genes, but uncertainty remains around critical nodes. A series of phylogenetic controversial exists at the suborder, family, subfamily, and species levels. A total of 1105 orthologous exon sequences and translated amino acid sequences from 36 genomes and 12 transcriptomes covering 33 species were applied to investigate the phylogenetic relationships within Gadiformes and address these problems. Phylogenetic trees reconstructed with the amino acid data set using different tree-building methods (RAxML and MrBayes) showed consistent topology. The monophyly of Gadifromes was confirmed in our study. However, the three suborders Muraenolepidoidei, Macrouroidei, and Gadoidei were not well recovered by our phy-logenomic study, rejecting the validity of suborder Muraenolepidoidei. Four major lineages were revealed in this study. The family Bregmacerotidae forming clade I was the basal lineage of Gadiformes. The family Merluciidae formed clade II. Clade III contained families Melanonidae, Muraenolepididae, Macrouridae (with subfamilies Trachyrincinae, Macrourinae, and Bathygadinae), and Moridae. Clade IV contained at least three families of suborder Gadoidei, i.e., Gadidae, Phycidae, and Ranicipitidae. The subspecies of Lota lota from Amur River were confirmed, indicating that exon markers were a valid high-resolution method for delimiting subspecies or distinct lineages within species level. The PSMC analysis of different populations of L. lota suggests a continuous decline since 2 Myr.

The mitogenome of Phytophthora agathidicida: Evidence for a not so recent arrival of the “kauri killing” Phytophthora in New Zealand

Phytophthora agathidicida is associated with a root rot that threatens the long-term survival of the iconic New Zealand kauri. Although it is widely assumed that this pathogen arrived in New Zealand post-1945, this hypothesis has yet to be formally tested. Here we describe evolutionary analyses aimed at evaluating this and two alternative hypotheses. As a basis for our analyses, we assembled complete mitochondrial genome sequences from 16 accessions representing the geographic range of P. agathidicida as well as those of five other members of Phytophthora clade 5. All 21 mitogenome sequences were very similar, differing little in size with all sharing the same gene content and arrangement. We first examined the temporal origins of genetic diversity using a pair of calibration schemes. Both resulted in similar age estimates; specifically, a mean age of 303.0–304.4 years and 95% HPDs of 206.9–414.6 years for the most recent common ancestor of the included isolates. We then used phylogenetic tree building and network analyses to investigate the geographic distribution of the genetic diversity. Four geographically distinct genetic groups were recognised within P. agathidicida. Taken together the inferred age and geographic distribution of the sampled mitogenome diversity suggests that this pathogen diversified following arrival in New Zealand several hundred to several thousand years ago. This conclusion is consistent with the emergence of kauri dieback disease being a consequence of recent changes in the relationship between the pathogen, host, and environment rather than a post-1945 introduction of the causal pathogen into New Zealand.

Uncovering feature interdependencies in high-noise environments with stepwise lookahead decision forests

Conventionally, random forests are built from “greedy” decision trees which each consider only one split at a time during their construction. The sub-optimality of greedy implementation has been well-known, yet mainstream adoption of more sophisticated tree building algorithms has been lacking. We examine under what circumstances an implementation of less greedy decision trees actually yields outperformance. To this end, a “stepwise lookahead” variation of the random forest algorithm is presented for its ability to better uncover binary feature interdependencies. In contrast to the greedy approach, the decision trees included in this random forest algorithm, each simultaneously consider three split nodes in tiers of depth two. It is demonstrated on synthetic data and financial price time series that the lookahead version significantly outperforms the greedy one when (a) certain non-linear relationships between feature-pairs are present and (b) if the signal-to-noise ratio is particularly low. A long-short trading strategy for copper futures is then backtested by training both greedy and stepwise lookahead random forests to predict the signs of daily price returns. The resulting superior performance of the lookahead algorithm is at least partially explained by the presence of “XOR-like” relationships between long-term and short-term technical indicators. More generally, across all examined datasets, when no such relationships between features are present, performance across random forests is similar. Given its enhanced ability to understand the feature-interdependencies present in complex systems, this lookahead variation is a useful extension to the toolkit of data scientists, in particular for financial machine learning, where conditions (a) and (b) are typically met.

A comparison of approaches to scaffolding multiple regions along the 16S rRNA gene for improved resolution

MotivationFull length, high resolution 16s rRNA marker gene sequencing has been challenging historically. Short amplicons provide high accuracy reads with widely available equipment, at the cost of taxonomic resolution. One recent proposal has been to reconstruct multiple amplicons along the full-length marker gene, however no barcode-free computationally tractable approach for this is available. To address this gap, we present Sidle (SMURF Implementation Done to acceLerate Efficiency), an implementation of the Short MUltiple Reads Framework algorithm with a novel tree building approach to reconstruct rRNA genes from individually amplified regions.ResultsUsing simulated and real data, we compared Sidle to two other approaches of leveraging multiple gene region data. We found that Sidle had the least bias in non-phylogenetic alpha diversity, feature-based measures of beta diversity, and the reconstruction of individual clades. With a curated database, Sidle also provided the most precise species-level resolution.Availability and ImplementationSidle is available under a BSD 3 license from https://github.com/jwdebelius/q2-sidle