Search results for: sequencing air-lift bioreactor

Authors: Vyacheslav Furtak, Merja Roivainen, Olga Mirochnichenko, Majid Laassri, Bella Bidzhieva, Tatiana Zagorodnyaya, Vladimir Chizhikov, Konstantin Chumakov

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Molecular epidemiology and environmental surveillance are parts of a rational strategy to control infectious diseases. They have been widely used in the worldwide campaign to eradicate poliomyelitis, which otherwise would be complicated by the inability to rapidly respond to outbreaks and determine sources of the infection. The conventional scheme involves isolation of viruses from patients and the environment, followed by their identification by nucleotide sequences analysis to determine phylogenetic relationships. This is a tedious and time-consuming process that yields definitive results when it may be too late to implement countermeasures. Because of the difficulty of high-throughput full-genome sequencing, most such studies are conducted by sequencing only capsid genes or their parts. Therefore the important information about the contribution of other parts of the genome and inter- and intra-species recombination to viral evolution is not captured. Here we propose a new approach based on the rapid concentration of sewage samples with tangential flow filtration followed by deep sequencing and reconstruction of nucleotide sequences of viruses present in the samples. The entire nucleic acids content of each sample is sequenced, thus preserving in digital format the complete spectrum of viruses. A set of rapid algorithms was developed to separate deep sequence reads into discrete populations corresponding to each virus and assemble them into full-length consensus contigs, as well as to generate a complete profile of sequence heterogeneities in each of them. This provides an effective approach to study molecular epidemiology and evolution of natural viral populations.

Keywords: poliovirus, eradication, environmental surveillance, laboratory diagnosis

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Authors: Yi-Lin Chen, Sheng-Jou Hung, Tsunglin Liu

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Adaptive immune system recognizes a wide range of antigens via expressing a large number of structurally distinct T cell and B cell receptor genes. The distinct receptor genes arise from complex rearrangements called V(D)J recombination, and constitute the immune repertoire. A common method of profiling immune repertoire is via amplifying recombined receptor genes using multiple primers and high-throughput sequencing. This multiplex-PCR approach is efficient; however, the resulting repertoire can be distorted because of primer bias. To eliminate primer bias, 5’ RACE is an alternative amplification approach. However, the application of RACE approach is limited by its low efficiency (i.e., the majority of data are non-regular receptor sequences, e.g., containing intronic segments) and lack of the convenient tool for analysis. We propose a computational tool that can correctly identify non-regular receptor sequences in RACE data via aligning receptor sequences against the whole gene instead of only the exon regions as done in all other tools. Using our tool, the remaining regular data allow for an accurate profiling of immune repertoire. In addition, a RACE approach is improved to yield a higher fraction of regular T-cell receptor sequences. Finally, we quantify the degree of primer bias of a multiplex-PCR approach via comparing it to the RACE approach. The results reveal significant differences in frequency of VJ combination by the two approaches. Together, we provide a new experimental and computation pipeline for an unbiased profiling of immune repertoire. As immune repertoire profiling has many applications, e.g., tracing bacterial and viral infection, detection of T cell lymphoma and minimal residual disease, monitoring cancer immunotherapy, etc., our work should benefit scientists who are interested in the applications.

Keywords: immune repertoire, T-cell receptor, 5' RACE, high-throughput sequencing, sequence alignment

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Authors: Deybith Venegas-Rojas, Jens Budde, Dominik Nörz, Manfred Jücker, Hoc Khiem Trieu

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Microfluidics is a promising approach for biomedicine cell culture experiments with microfluidic bioreactors (MBR), which can provide high precision in volume and time control over mass transport and microenvironments in small-scale studies. Nevertheless, shear stress and oxygen concentration are important factors that affect the microenvironment and then the cell culture. It is presented a novel MBR design in which differences in geometry, shear stress, and oxygen concentration were studied and optimized for cell culture. The aim is to mimic the in vivo condition with biocompatible materials and continuous perfusion of nutrients, a healthy shear stress, and oxygen concentration. The design consists of a capture system of PDMS micropillars which keep cells in place, so it is not necessary any hydrogel or complicated scaffolds for cells immobilization. Besides, the design allows continuous supply with nutrients or even any other chemical for cell experimentation. Finite element method simulations were used to study and optimize the effect of parameters such as flow rate, shear stress, oxygen concentration, micropillars shape, and dimensions. The micropillars device was fabricated with microsystem technology such as soft-lithography, deep reactive ion etching, self-assembled monolayer, replica molding, and oxygen plasma bonding. Eight different geometries were fabricated and tested, with different flow rates according to the simulations. During the experiments, it was observed the effect of micropillars size, shape, and configuration for stability and shear stress control when increasing flow rate. The device was tested with several successful HepG2 3D cell cultures. With this MBR, the aforementioned parameters can be controlled in order to keep a healthy microenvironment according to specific necessities of different cell types, with no need of hydrogels and can be used for a wide range of experiments with cells.

Keywords: cell culture, micro-bioreactor, microfluidics, micropillars, oxygen concentration, shear stress

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Authors: Vytautas Galvanauskas, Rimvydas Simutis, Donatas Levisauskas, Vykantas Grincas, Renaldas Urniezius

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The paper deals with model-based development and implementation of efficient control strategies for recombinant protein synthesis in fed-batch E.coli cultivation processes. Based on experimental data, a kinetic dynamic model for cultivation process was developed. This model was used to determine substrate feeding strategies during the cultivation. The proposed feeding strategy consists of two phases – biomass growth phase and recombinant protein production phase. In the first process phase, substrate-limited process is recommended when the specific growth rate of biomass is about 90-95% of its maximum value. This ensures reduction of glucose concentration in the medium, improves process repeatability, reduces the development of secondary metabolites and other unwanted by-products. The substrate limitation can be enhanced to satisfy restriction on maximum oxygen transfer rate in the bioreactor and to guarantee necessary dissolved carbon dioxide concentration in culture media. In the recombinant protein production phase, the level of substrate limitation and specific growth rate are selected within the range to enable optimal target protein synthesis rate. To account for complex process dynamics, to efficiently exploit the oxygen transfer capability of the bioreactor, and to maintain the required dissolved oxygen concentration, adaptive control algorithms for dissolved oxygen control have been proposed. The developed model-based control strategies are useful in scale-up of cultivation processes and accelerate implementation of innovative biotechnological processes for industrial applications.

Keywords: adaptive algorithms, model-based control, recombinant E. coli, scale-up of bioprocesses

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Authors: W. Baumgartner, M. Welti, N. Hild, S. C. Hess, W. J. Stark, G. Meier Bürgisser, P. Giovanoli, J. Buschmann

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Perfusion bioreactors are used to solve problems in tissue engineering in terms of sufficient nutrient and oxygen supply. Such problems especially occur in critical size grafts because vascularization is often too slow after implantation ending up in necrotic cores. Biominerizable and biocompatible nanocomposite materials are attractive and suitable scaffold materials for bone tissue engineering because they offer mineral components in organic carriers – mimicking natural bone tissue. In addition, human adipose derived stem cells (ASCs) can potentially be used to increase bone healing as they are capable of differentiating towards osteoblasts or endothelial cells among others. In the present study, electrospun nanocomposite disks of poly-lactic-co-glycolic acid and amorphous calcium phosphate nanoparticles (PLGA/a-CaP) were seeded with human ASCs and eight disks were stacked in a bioreactor running with normal culture medium (no differentiation supplements). Under continuous perfusion and uniaxial cyclic compression, load-displacement curves as a function of time were assessed. Stiffness and energy dissipation were recorded. Moreover, stem cell densities in the layers of the piled scaffold were determined as well as their morphologies and differentiation status (endothelial cell differentiation, chondrogenesis and osteogenesis). While the stiffness of the cell free constructs increased over time caused by the transformation of the a-CaP nanoparticles into flake-like apatite, ASC-seeded constructs showed a constant stiffness. Stem cell density gradients were histologically determined with a linear increase in the flow direction from the bottom to the top of the 3.5 mm high pile (r2 > 0.95). Cell morphology was influenced by the flow rate, with stem cells getting more roundish at higher flow rates. Less than 1 % osteogenesis was found upon osteopontin immunostaining at the end of the experiment (9 days), while no endothelial cell differentiation and no chondrogenesis was triggered under these conditions. All ASCs had mainly remained in their original pluripotent status within this time frame. In summary, we have fabricated a critical size bone graft based on a biominerizable bone-biomimetic nanocomposite with preserved stiffness when seeded with human ASCs. The special feature of this bone graft was that ASC densities inside the piled construct varied with a linear gradient, which is a good starting point for tissue engineering interfaces such as bone-cartilage where the bone tissue is cell rich while the cartilage exhibits low cell densities. As such, this tissue-engineered graft may act as a bone-cartilage interface after the corresponding differentiation of the ASCs.

Keywords: bioreactor, bone, cartilage, nanocomposite, stem cell gradient

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Authors: D. Hammad, D. P. Tonge

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Rheumatoid arthritis (RA) is a disabling and common autoimmune disease during which the body's immune system attacks healthy tissues. This results in complicated and long-lasting actions being carried out by the immune system, which typically only occurs when the immune system encounters a foreign object. In the case of RA, the disease affects millions of people and causes joint inflammation, ultimately leading to the destruction of cartilage and bone. Interestingly, the disease mechanism still remains unclear. It is likely that RA occurs as a result of a complex interplay of genetic and environmental factors including an imbalance in the microorganism population inside our body. The human microbiome or microbiota is an extensive community of microorganisms in and on the bodies of animals, which comprises bacteria, fungi, viruses, and protozoa. Recently, the development of molecular techniques to characterize entire bacterial communities has renewed interest in the involvement of the microbiome in the development and progression of RA. We believe that an imbalance in some of the specific bacterial species in the gut, mouth and other sites may lead to atopobiosis; the translocation of these organisms into the blood, and that this may lead to changes in immune system status. The aim of this study was, therefore, to characterize the microbiome of RA serum samples in comparison to healthy control subjects using 16S rRNA gene amplification and sequencing. Serum samples were obtained from healthy control volunteers and from patients with RA both prior to, and following treatment. The bacterial community present in each sample was identified utilizing V4 region 16S rRNA amplification and sequencing. Bacterial identification, to the lowest taxonomic rank, was performed using a range of bioinformatics tools. Significantly, the proportions of the Lachnospiraceae, Ruminococcaceae, and Halmonadaceae families were significantly increased in the serum of RA patients compared with healthy control serum. Furthermore, the abundance of Bacteroides and Lachnospiraceae nk4a136_group, Lachnospiraceae_UGC-001, RuminococcaceaeUCG-014, Rumnococcus-1, and Shewanella was also raised in the serum of RA patients relative to healthy control serum. These data support the notion of a blood microbiome and reveal RA-associated changes that may have significant implications for biomarker development and may present much-needed opportunities for novel therapeutic development.

Keywords: blood microbiome, gut and oral bacteria, Rheumatoid arthritis, 16S rRNA gene sequencing

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Authors: Guilherme Gorgulho, Carlos Roberto Camello Lima

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Due to the competitive market in which companies are currently engaged, the constant changes require companies to react quickly regarding the variability of demand and process. The changes are caused by customers, or by demand fluctuations or variations of products, or the need to serve customers within agreed delivery taking into account the continuous search for quality and competitive prices in products. These changes end up influencing directly or indirectly the activities of the Planning and Production Control (PPC), which does business in strategic, tactical and operational levels of production systems. One area of concern for organizations is in the short term (operational level), because this planning stage any error or divergence will cause waste and impact on the delivery of products on time to customers. Thus, this study aims to optimize the efficiency of production scheduling, using different sequencing strategies in an automotive company. Seeking to aim the proposed objective, we used the computer simulation in conjunction with lean manufacturing to build and validate the current model, and subsequently the creation of future scenarios.

Keywords: computational simulation, lean manufacturing, production scheduling, sequencing strategies

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Authors: D. S. V. Barbieri, R. R. Gomes, G. D. Santos, P. F. Herkert, M. Moreira, E. S. Trindade, V. A. Vicente

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The importance of yeast infections has increased in recent decades. The monitoring of Candida yeasts has been relevant in the study of groups and populations. This research evaluated 31 Candida spp. isolates from oral microbiota of 12 Brazilian schoolchildren coinfected with Streptococcus mutans. The isolates were evaluated for their ability to form biofilm in vitro and molecularly characterized based on the sequencing of intergenic spacer regions ITS1-5,8S-ITS2 and variable domains of the large subunit (D1/D2) regions of the rDNA, as well as ABC system genotyping. The sequencing confirmed 26 lineages of Candida albicans, three Candida tropicalis, one Candida guillhermondii and one Candida glabrata. Genetic variability and differences on in biofilm formation were observed among Candida yeasts lineages. At least one Candida strain from each caries activity child was C.albicans genotype A or Candida non-albicans. C. tropicalis was associated with highest cavities rates. These results indicate that the presence of C. albicans genotype A or multi-colonization by non albicans species seem to be associates to the potentialization of caries risk.

Keywords: biofilm, Candida albicans, oral microbiota, caries

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Authors: Karol Rodriguez-Peña, Martha L. Macias-Rubalcava, Leticia Rocha-Zavaleta, Sergio Sanchez

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A bioassay-guided study for anti-cancer compounds from endophytes of the Mexican medicinal plant Amphipteryygium adstringens resulted in the isolation of a streptomycete capable of producing a group of compounds with high cytotoxic activity. Microorganisms from surface sterilized samples of various sections of the plant were isolated and all the actinomycetes found were evaluated for their potential to produce compounds with cytotoxic activity against cancer cell lines MCF7 (breast cancer) and HeLa (cervical cancer) as well as the non-tumoural cell line HaCaT (keratinocyte). The most active microorganism was picked for further evaluation. The identification of the microorganism was carried out by 16S rDNA gene sequencing, finding the closest proximity to Streptomyces scabrisporus, but with the additional characteristic that the strain isolated in this study was capable of producing colorful compounds never described for this species. Crude extracts of dichloromethane and ethyl acetate showed IC50 values of 0.29 and 0.96 μg/mL for MCF7, 0.51 and 1.98 μg/mL for HeLa and 0.96 and 2.7 μg/mL for HaCaT. Scaling the fermentation to 10 L in a bioreactor generated 1 g of total crude extract, which was fractionated by silica gel open column to yield 14 fractions. Nine of the fractions showed cytotoxic activity. Fraction 4 was chosen for subsequent purification because of its high activity against cancerous cell lines, lower activity against keratinocytes. HPLC-UV-MS/ESI was used for the evaluation of this fraction, finding at least 10 different compounds with high values of m/z (≈588). Purification of the compounds was carried out by preparative thin-layer chromatography. The prevalent compound was Steffimycin B, a molecule known for its antibiotic and cytotoxic activities and also for its low solubility in aqueous solutions. Along with steffimycin B, another five compounds belonging to the steffimycin family were isolated and at this moment their structures are being elucidated, some of which display better solubility in water: an attractive property for the pharmaceutical industry. As a conclusion to this study, the isolation of endophytes resulted in the discovery of a strain capable of producing compounds with high cytotoxic activity that need to be studied for their possible utilization.

Keywords: amphipterygium adstringens, cytotoxicity, streptomyces scabrisporus, steffimycin

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Authors: Aishik Deb, Rituparna Sinha

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We have developed an algorithm to detect the abnormal segment/"structural variation in the genome across a number of samples. We have worked on simulated as well as real data from the BAM Files and have designed a segmentation algorithm where abnormal segments are detected. This algorithm aims to improve the accuracy and performance of the existing CNV-CH algorithm. The next-generation sequencing (NGS) approach is very fast and can generate large sequences in a reasonable time. So the huge volume of sequence information gives rise to the need for Big Data and parallel approaches of segmentation. Therefore, we have designed a map-reduce approach for the existing CNV-CH algorithm where a large amount of sequence data can be segmented and structural variations in the human genome can be detected. We have compared the efficiency of the traditional and map-reduce algorithms with respect to precision, sensitivity, and F-Score. The advantages of using our algorithm are that it is fast and has better accuracy. This algorithm can be applied to detect structural variations within a genome, which in turn can be used to detect various genetic disorders such as cancer, etc. The defects may be caused by new mutations or changes to the DNA and generally result in abnormally high or low base coverage and quantification values.

Keywords: cancer detection, convex hull segmentation, map reduce, next generation sequencing

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Authors: Akram Abi, Clarissa Müller, Hans-Joachim Jördening

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Laminaribiose is a beta-1,3-glycoside which is used in the medical field for the treatment of dermatitis and also can be used as a building block for new pharmaceutics. The conventional process of laminaribiose production is the uneconomical process of hydrolysis of laminarin extracted from natural polysaccharides of plant origin. A more economical approach however is attainable by enzymatically synthesis of laminaribiose via a reverse phosphorylase reaction catalyzed by laminaribiose phosphorylase (LP) from Euglena gracilis. Different cultivation methods of Euglena gracilis and the effect on LP production have been investigated. Buffered/unbuffered heterotrophic and mixotrophic cultivations of Euglena gracilis has been carried out. Changes of biomass and LP production, glucose level and pH, cell count and shape has been monitored in the course of time. The results obtained from experiments each in three repetitions, show that in the heterotrophic cultivation of Euglena gracilis not only more biomass is produced compared to mixotrophic cultivation, but also higher specific protein concentration is achieved. Furthermore, the LP activity test showed that the protein extracted from heterotrophically cultured cells has a higher LP activity. It was also observed that the cells develop in a distinctive different shape between these two cultures and have different length to width ratios. Taking the heterotrophic culture as the more efficient cultivation method in LP production, another comparative experiment between buffered and unbuffered heterothrophic culture was carried out that showed the unbuffered culture has advantages over the other one in respect of both LP production and resulting activity. A hetrotrophic cultivation of Euglena gracilis in a 5L bioreactor with controlled operating conditions showed a distinctive improvement of all the aspects of culture compared to the shaking flask cultivations. Biomass production was improved from 5 to more than 8 g/l (dry weight) which resulted in a specific protein concentration of 45 g/l in the heterotrophic cultivation in the bioreactor. In further attempts to improve LP production, different purification methods were tested and each method was checks through an activity assay. A laminaribiose yield of 35% was achieved which was by far the highest amount amongst different methods tested.

Keywords: euglena gracilis, heterotrophic culture, laminaribiose production, mixotrophic culture

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Authors: Maryam Zafar, Sajida Rasheed, Imran Hashmi

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Water as an environmental reservoir is reported to act as a habitat and transmission route to microaerophilic bacteria such as Heliobacter pylori. It has been studied that in biofilms are the predominant dwellings for the bacteria to grow in water and protective reservoir for numerous pathogens by protecting them against harsh conditions, such as shear stress, low carbon concentration and less than optimal temperature. In this study, influence of these and many other parameters was studied on H. pylori in stagnant and moving water biofilms both in surface and underground aquatic reservoirs. H. pylori were recovered from pipe of different materials such as Polyvinyl Chloride, Polypropylene and Galvanized iron pipe cross sections from an urban water distribution network. Biofilm swabbed from inner cross section was examined by molecular biology methods coupled with gene sequencing and H. pylori 16S rRNA peptide nucleic acid probe showing positive results for H. pylori presence. Studies showed that pipe material affect growth of biofilm which in turn provide additional survival mechanism for pathogens like H. pylori causing public health concerns.

Keywords: biofilm, gene sequencing, heliobacter pylori, pipe materials

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Authors: Abu Salim Mustafa

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Brucellosis is a zoonotic disease endemic in Kuwait. Human brucellosis can be caused by several Brucella species with Brucella melitensis causing the most severe and Brucella abortus the least severe disease. Furthermore, relapses are common after successful chemotherapy of patients. The classical biochemical methods of culture and serology for identification of Brucellae provide information about the species and serotypes only. However, to differentiate between relapse and reinfection/epidemiological investigations, the identification of genotypes using molecular methods is essential. In this study, four molecular methods [16S rRNA gene sequencing, real-time PCR, enterobacterial repetitive intergenic consensus (ERIC)-PCR and multilocus variable-number tandem-repeat analysis (MLVA)-16] were evaluated for the identification and typing of 75 strains of Brucella isolated in Kuwait. The 16S rRNA gene sequencing suggested that all the strains were B. melitensis and real-time PCR confirmed their species identity as B. melitensis. The ERIC-PCR band profiles produced a dendrogram of 75 branches suggesting each strain to be of a unique type. The cluster classification, based on ~ 80% similarity, divided all the ERIC genotypes into two clusters, A and B. Cluster A consisted of 9 ERIC genotypes (A1-A9) corresponding to 9 individual strains. Cluster B comprised of 13 ERIC genotypes (B1-B13) with B5 forming the largest cluster of 51 strains. MLVA-16 identified all isolates as B. melitensis and divided them into 71 MLVA-types. The cluster analysis of MLVA-16-types suggested that most of the strains in Kuwait originated from the East Mediterranean Region, a few from the African group and one new genotype closely matched with the West Mediterranean region. In conclusion, this work demonstrates that B. melitensis, the most pathogenic species of Brucella, is prevalent in Kuwait. Furthermore, MLVA-16 is the best molecular method, which can identify the Brucella species and genotypes as well as determine their origin in the global context. Supported by Kuwait University Research Sector grants MI04/15 and SRUL02/13.

Keywords: Brucella, ERIC-PCR, MLVA-16, RT-PCR, 16S rRNA gene sequencing

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Authors: A. Delimitsou, C. Gouedard, E. Konstanta, A. Koletis, S. Patera, E. Manou, K. Spaho, S. Murray

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Background: There remains a lack of International Standard Control Reference materials for Next Generation Sequencing-based approaches or device calibration. We have designed and validated dsDNA biomimetic reference materials for targeted such approaches incorporating proprietary motifs (patent pending) for device/test calibration. They enable internal single-sample calibration, alleviating sample comparisons to pooled historical population-based data assembly or statistical modelling approaches. We have validated such an approach for BRCA Copy Number Variation analytics using iQRS™-CNVSUITE versus Mixed Ligation-dependent Probe Amplification. Methods: Standard BRCA Copy Number Variation analysis was compared between mixed ligation-dependent probe amplification and next generation sequencing using a cohort of 198 breast/ovarian cancer patients. Next generation sequencing based copy number variation analysis of samples spiked with iQRS™ dsDNA biomimetics were analysed using proprietary CNVSUITE software. Mixed ligation-dependent probe amplification analyses were performed on an ABI-3130 Sequencer and analysed with Coffalyser software. Results: Concordance of BRCA – copy number variation events for mixed ligation-dependent probe amplification and CNVSUITE indicated an overall sensitivity of 99.88% and specificity of 100% for iQRS™-CNVSUITE. The negative predictive value of iQRS-CNVSUITE™ for BRCA was 100%, allowing for accurate exclusion of any event. The positive predictive value was 99.88%, with no discrepancy between mixed ligation-dependent probe amplification and iQRS™-CNVSUITE. For device calibration purposes, precision was 100%, spiking of patient DNA demonstrated linearity to 1% (±2.5%) and range from 100 copies. Traditional training was supplemented by predefining the calibrator to sample cut-off (lock-down) for amplicon gain or loss based upon a relative ratio threshold, following training of iQRS™-CNVSUITE using spiked iQRS™ calibrator and control mocks. BRCA copy number variation analysis using iQRS™-CNVSUITE™ was successfully validated and ISO15189 accredited and now enters CE-IVD performance evaluation. Conclusions: The inclusion of a reference control competitor (iQRS™ dsDNA mimetic) to next generation sequencing-based sequencing offers a more robust sample-independent approach for the assessment of copy number variation events compared to mixed ligation-dependent probe amplification. The approach simplifies data analyses, improves independent sample data analyses, and allows for direct comparison to an internal reference control for sample-specific quantification. Our iQRS™ biomimetic reference materials allow for single sample copy number variation analytics and further decentralisation of diagnostics to single patient sample assessment.

Keywords: validation, diagnostics, oncology, copy number variation, reference material, calibration

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Authors: Niamh Higgins, Dawn Howard

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The Irish agri-food sector is of major importance to Ireland’s manufacturing sector and to the Irish economy through employment and the exporting of animal products worldwide. Infectious diseases and parasites have an impact on farm animal health causing profitability and productivity to be affected. For the sustainability of Irish dairy farming, there must be the highest standard of animal health. There can be a lack of information in accounting for > 1% of complete microbial diversity in an environment. There is the tendency of culture-based methods of microbial identification to overestimate the prevalence of species which grow easily on an agar surface. There is a need for new technologies to address these issues to assist with animal health. Metagenomic approaches provide information on both the whole genome and transcriptome present through DNA sequencing of total DNA from environmental samples producing high determination of functional and taxonomic information. Nanopore Next Generation Technologies have the ability to be powerful sequencing technologies. They provide high throughput, low material requirements and produce ultra-long reads, simplifying the experimental process. The aim of this study is to use a metagenomics approach to analyze dairy cattle faecal samples using the Oxford Nanopore MinION Next Generation Sequencer and to establish an in-house pipeline for metagenomic characterization of complex samples. Faecal samples will be obtained from Irish dairy farms, DNA extracted and the MinION will be used for sequencing, followed by bioinformatics analysis. Of particular interest, will be the parasite Buxtonella sulcata, which there has been little research on and which there is no research on its presence on Irish dairy farms. Preliminary results have shown the ability of the MinION to produce hundreds of reads in a relatively short time frame of eight hours. The faecal samples were obtained from 90 dairy cows on a Galway farm. The results from Oxford Nanopore ‘What’s in my pot’ (WIMP) using the Epi2me workflow, show that from a total of 926 classified reads, 87% were from the Kingdom Bacteria, 10% were from the Kingdom Eukaryota, 3% were from the Kingdom Archaea and < 1% were from the Kingdom Viruses. The most prevalent bacteria were those from the Genus Acholeplasma (71 reads), Bacteroides (35 reads), Clostridium (33 reads), Acinetobacter (20 reads). The most prevalent species present were those from the Genus Acholeplasma and included Acholeplasma laidlawii (39 reads) and Acholeplasma brassicae (26 reads). The preliminary results show the ability of the MinION for the identification of microorganisms to species level coming from a complex sample. With ongoing optimization of the pipe-line, the number of classified reads are likely to increase. Metagenomics has the potential in animal health for diagnostics of microorganisms present on farms. This would support wprevention rather than a cure approach as is outlined in the DAFMs National Farmed Animal Health Strategy 2017-2022.

Keywords: animal health, buxtonella sulcata, infectious disease, irish dairy cattle, metagenomics, minION, next generation sequencing

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Authors: Ilze Dimanta, Zane Rutkovska, Vizma Nikolajeva, Janis Kleperis, Indrikis Muiznieks

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Majority of word’s steadily increasing energy consumption is provided by non-renewable fossil resources. Need to find an alternative energy resource is essential for further socio-economic development. Hydrogen is renewable, clean energy carrier with high energy density (142 MJ/kg, accordingly – oil has 42 MJ/kg). Biological hydrogen production is an alternative way to produce hydrogen from renewable resources, e.g. using organic waste material resource fermentation that facilitate recycling of sewage and are environmentally benign. Hydrogen gas is produced during the fermentation process of bacteria in anaerobic conditions. Bacteria are producing hydrogen in the liquid phase and when thermodynamic equilibrium is reached, hydrogen is diffusing from liquid to gaseous phase. Because of large quantities of available crude glycerol and the highly reduced nature of carbon in glycerol per se, microbial conversion of it seems to be economically and environmentally viable possibility. Such industrial organic waste product as crude glycerol is perspective for usage in feedstock for hydrogen producing bacteria. The process of biodiesel production results in 41% (w/w) of crude glycerol. The developed lab-scale test system (experimental bioreactor) with hydrogen micro-electrode (Unisense, Denmark) was used to determine hydrogen production yield and rate in the liquid phase. For hydrogen analysis in the gas phase the RGAPro-100 mass-spectrometer connected to the experimental test-system was used. Fermentative bacteria strains were tested for hydrogen gas production rates. The presence of hydrogen in gaseous phase was measured using mass spectrometer but registered concentrations were comparatively small. To decrease the hydrogen partial pressure in liquid phase reactor with a system for continuous bubbling with inert gas was developed. H2 production rate for the best producer in liquid phase reached 0,40 mmol H2/l, in gaseous phase - 1,32 mmol H2/l. Hydrogen production rate is time dependent – higher rate of hydrogen production is at the fermentation process beginning when concentration increases, but after three hours of fermentation, it decreases.

Keywords: bio-hydrogen, fermentation, experimental bioreactor, crude glycerol

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Authors: Maryam Vaezjalali, Koroush Rahimian, Maryam Asli, Tahmineh Kandelouei, Foad Davoodbeglou, Amir H. Kashi

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Objectives: The present study was conducted to assess the HBV molecular profiles including genotypes, subgenotypes, subtypes & mutations in hepatitis B genes. Materials/Patients and Methods: This study was conducted on 229 intravenous drug users who referred to three Drop- in-Centers and a hospital in Tehran. HBV DNA was extracted from HBsAg positive serum samples and amplified by Nested PCR. HBV genotype, subgenotypes, subtype and genes mutation were determined by direct sequencing. Phylogenetic tree was constructed using neighbor- joining (NJ) method. Statistical analyses were carried out by SPSS 20. Results: HBV DNA was found in 3 HBsAg positive cases. Phylogenetic tree of derived HBV DNAs showed the existence of genotype D (subgenotype D1, subtype ayw2). Also immune escape mutations were determined in S gene. Conclusion: There were a few variations and genotypes and subtypes among infected intravenous drug users. This study showed the predominance of genotype D among intravenous drug users. Our study concurs with other reports from Iran, that all showing currently only genotype D is the only detectable genotype in Iran.

Keywords: drug users, genotype, HBV, phylogenetic tree

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Authors: Tanushree Jaitly, Shailendra Gupta, Leila Taher, Gerold Schuler, Julio Vera

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Genomics-based personalized medicine is a promising approach to fight aggressive tumors based on patient's specific tumor mutation and expression profiles. A remarkable case is, dendritic cell-based immunotherapy, in which tumor epitopes targeting patient's specific mutations are used to design a vaccine that helps in stimulating cytotoxic T cell mediated anticancer immunity. Here we present a computational pipeline for epitope-based personalized cancer vaccines using patient-specific haplotype and cancer mutation profiles. In the workflow proposed, we analyze Whole Exome Sequencing and RNA Sequencing patient data to detect patient-specific mutations and their expression level. Epitopes including the tumor mutations are computationally predicted using patient's haplotype and filtered based on their expression level, binding affinity, and immunogenicity. We calculate binding energy for each filtered major histocompatibility complex (MHC)-peptide complex using docking studies, and use this feature to select good epitope candidates further.

Keywords: cancer immunotherapy, epitope prediction, NGS data, personalized medicine

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Authors: Azila Adnan, Giridhar R. Nair, Mark C. Lay, Janis E. Swan

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The effect of aeration on bacterial cellulose (BC) production by Gluconacetobacter xylinus DSM46604 was studied in 5-L bioreactor. Four aeration rates were applied (0.3, 0.6, 1.0 and 1.5 vvm) in the fermentation media at constant agitation rate, 150 rpm. One vvm enhanced BC concentration (4.4 g/L) and productivity (0.44 g/L/day) while greater agitation rate (1.5 vvm) decreased BC concentration (4.0 g/L) and productivity (0.40 g/L/day).

Keywords: aeration, bacterial cellulose, fermentation, gluconacetobacter xylinus

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Authors: Dalia. G. Aseel, E. E. Hafez, S. M. Hammad

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Viral RNAs of both potato leaf roll virus (PLRV) and potato virus Y (PVY) were extracted from infected potato leaves collected from different Egyptian regions. Differential Display Polymerase Chain Reaction (DD-PCR) using (Endogluconase, β-1,3-glucanases, Chitinase, Peroxidase and Polyphenol oxidase) primers (forward strand) for was performed. The obtained data revealed different banding patterns depending on the viral type and the region of infection. Regarding PLRV, a 58 up regulated and 19 down regulated genes were detected, while, 31 up regulated and 14 down regulated genes were observed in case of PVY. Based on the nucleotide sequencing, variable phylogenetic relationships were reported for the three sequenced genes coding for: Induced stolen tip protein, Disease resistance RPP-like protein and non-specific lipid-transfer protein. In a complementary approach, using the quantitative Real-time PCR, the expressions of PRs genes understudy were estimated in the infected leaves by PLRV and PVY of three potato cultivars (Spunta, Diamont and Cara). The infection with both viruses inhibited the expressions of the five PRs genes. On the contrary, infected leaves by PLRV or PVY elevated the expression of some defense genes. This interaction also may be enhanced and/or inhibited the expression of some genes responsible for the plant defense mechanisms.

Keywords: PLRV, PVY, PR genes, DD-PCR, qRT-PCR, sequencing

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Authors: Young Ah Kim, Hyunsoo Kim, Eun-Jeong Yoon, Young Hee Seo, Kyungwon Lee

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Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli from food animals are considered as a reservoir for transmission of ESBL genes to human. The aim of this study is to assess the prevalence and molecular epidemiology of ESBL-producing E. coli colonization in pigs, farm workers, and farm environments to elucidate the transmission of multidrug-resistant clones from animal to human. Nineteen pig farms were enrolled across the country in Korea from August to December 2017. ESBL-producing E. coli isolates were detected in 190 pigs, 38 farm workers, and 112 sites of farm environments using ChromID ESBL (bioMerieux, Marcy l'Etoile, France), directly (stool or perirectal swab) or after enrichment (sewage). Antimicrobial susceptibility tests were done with disk diffusion methods and blaTEM, blaSHV, and blaCTX-M were detected with PCR and sequencing. The genomes of the four CTX-M-55-producing E. coli isolates from various sources in one farm were entirely sequenced to assess the relatedness of the strains. Whole genome sequencing (WGS) was performed with PacBio RS II system (Pacific Biosciences, Menlo Park, CA, USA). ESBL genotypes were 85 CTX-M-1 group (one CTX-M-3, 23 CTX-M-15, one CTX-M-28, 59 CTX-M-55, one CTX-M-69) and 60 CTX-M-9 group (41 CTX-M-14, one CTX-M-17, one CTX-M-27, 13 CTX-M-65, 4 CTX-M-102) in total 145 isolates. The rectal colonization rates were 53.2% (101/190) in pigs and 39.5% (15/38) in farm workers. In WGS, sequence types (STs) were determined as ST69 (E. coli PJFH115 isolate from a human carrier), ST457 (two E. coli isolates PJFE101 recovered from a fence and PJFA1104 from a pig) and ST5899 (E. coli PJFA173 isolate from the other pig). The four plasmids encoding CTX-M-55 (88,456 to 149, 674 base pair), whether it belonged to IncFIB or IncFIC-IncFIB type, shared IncF backbone furnishing the conjugal elements, suggesting of genes originated from same ancestor. In conclusion, the prevalence of ESBL-producing E. coli in swine farms was surprisingly high, and many of them shared common ESBL genotypes of clinical isolates such as CTX-M-14, 15, and 55 in Korea. It could spread by horizontal transfer between isolates from different reservoirs (human-animal-environment).

Keywords: Escherichia coli, extended-spectrum β-lactamase, prevalence, whole genome sequencing

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Authors: Roman Matejka, Vaclav Prochazka, Tibor Izak, Jana Stepanovska, Martina Travnickova, Alexander Kromka

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Cell-based impedance spectroscopy is suitable method for electrical monitoring of cell activity especially on substrates that cannot be easily inspected by optical microscope (without fluorescent markers) like decellularized tissues, nano-fibrous scaffold etc. Special sensor for this measurement was developed. This sensor consists of corning glass substrate with gold interdigitated electrodes covered with diamond layer. This diamond layer provides biocompatible non-conductive surface for cells. Also, a special PPFC flow cultivation chamber was developed. This chamber is able to fix sensor in place. The spring contacts are connecting sensor pads with external measuring device. Construction allows real-time live cell imaging. Combining with perfusion system allows medium circulation and generating shear stress stimulation. Experimental evaluation consist of several setups, including pure sensor without any coating and also collagen and fibrin coating was done. The Adipose derived stem cells (ASC) and Human umbilical vein endothelial cells (HUVEC) were seeded onto sensor in cultivation chamber. Then the chamber was installed into microscope system for live-cell imaging. The impedance measurement was utilized by vector impedance analyzer. The measured range was from 10 Hz to 40 kHz. These impedance measurements were correlated with live-cell microscopic imaging and immunofluorescent staining. Data analysis of measured signals showed response to cell adhesion of substrates, their proliferation and also change after shear stress stimulation which are important parameters during cultivation. Further experiments plan to use decellularized tissue as scaffold fixed on sensor. This kind of impedance sensor can provide feedback about cell culture conditions on opaque surfaces and scaffolds that can be used in tissue engineering in development artificial prostheses. This work was supported by the Ministry of Health, grants No. 15-29153A and 15-33018A.

Keywords: bio-impedance measuring, bioreactor, cell cultivation, diamond layer, gold interdigitated electrodes, tissue engineering

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Authors: Barsha Saha, Shouvik Chakravarty, Sukanta Ray, Kshaunish Das, Nidhan K. Biswas, Srikanta Goswami

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Pancreatic Ductal Adenocarcinoma is a well-recognised cause of cancer death with a five-year survival rate of about 9%, and its incidence in India has been found to be increased manifold in recent years. Due to delayed detection, this highly metastatic disease has a poor prognosis. Several molecular alterations happen during the progression of the disease from pre-cancerous conditions, and many such alterations could be investigated for their biomarker potential. MicroRNAs have been shown to be prognostic for PDAC patients in a variety of studies. We hereby used NGS technologies to evaluate the role of small RNA changes during pancreatic cancer development from chronic pancreatitis. Plasma samples were collected from pancreatic cancer patients (n=16), chronic pancreatitis patients (n=8), and also from normal individuals (n=16). Pancreatic tumour tissue (n=5) and adjacent normal tissue samples (n=5) were also collected. Sequencing of small RNAs was carried out after small RNAs were isolated from plasma samples and tissue samples. We find that certain microRNAs are highly deregulated in pancreatic cancer patients in comparison to normal samples. A combinatorial analysis of plasma and tissue microRNAs and subsequent exploration of their targets and altered molecular pathways could not only identify potential biomarkers for disease diagnosis but also help to understand the underlying mechanism.

Keywords: small RNA sequencing, pancreatic cancer, biomarkers, tissue sample

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Authors: J. Ritonja

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The biochemical technology has been developing extremely fast since the middle of the last century. The main reason for such development represents a requirement for large production of high-quality biologically manufactured products such as pharmaceuticals, foods, and beverages. The impact of the biochemical industry on the world economy is enormous. The great importance of this industry also results in intensive development in scientific disciplines relevant to the development of biochemical technology. In addition to developments in the fields of biology and chemistry, which enable to understand complex biochemical processes, development in the field of control theory and applications is also very important. In the paper, the control for the biochemical reactor for the milk fermentation was studied. During the fermentation process, the biophysical quantities must be precisely controlled to obtain the high-quality product. To control these quantities, the bioreactor’s stirring drive and/or heating system can be used. Available commercial biochemical reactors are equipped with open loop or conventional linear closed loop control system. Due to the outstanding parameters variations and the partial nonlinearity of the biochemical process, the results obtained with these control systems are not satisfactory. To improve the fermentation process, the self-tuning adaptive control system was proposed. The use of the self-tuning adaptive control is suggested because the parameters’ variations of the studied biochemical process are very slow in most cases. To determine the linearized mathematical model of the fermentation process, the recursive least square identification method was used. Based on the obtained mathematical model the linear quadratic regulator was tuned. The parameters’ identification and the controller’s synthesis are executed on-line and adapt the controller’s parameters to the fermentation process’ dynamics during the operation. The use of the proposed combination represents the original solution for the control of the milk fermentation process. The purpose of the paper is to contribute to the progress of the control systems for the biochemical reactors. The proposed adaptive control system was tested thoroughly. From the obtained results it is obvious that the proposed adaptive control system assures much better following of the reference signal as a conventional linear control system with fixed control parameters.

Keywords: adaptive control, biochemical reactor, linear quadratic regulator, recursive least square identification

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Authors: Yantao Li, Jun Zheng

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Food poison caused by consumption of contaminated food, especially seafood, is one of most serious public health threats worldwide. Vibrio parahaemolyticus is emerging bacterial pathogen and the leading cause of human gastroenteritis associated with food poison, especially in the southern coastal region of China. To successfully cause disease in host, bacterial pathogens need to overcome the host-derived stresses encountered during infection. One of the toxic chemical species elaborated by the host is nitric oxide (NO). NO is generated by acidified nitrite in the stomach and by enzymes of the inducible NO synthase (iNOS) in the host cell, and is toxic to bacteria. Bacterial pathogens have evolved some mechanisms to battle with this toxic stress. Such mechanisms include genes to sense NO produced from immune system and activate others to detoxify NO toxicity, and genes to repair the damage caused by toxic reactive nitrogen species (RNS) generated during NO toxic stress. However, little is known about the NO resistance in V. parahaemolyticus. In this study, a transposon coupled with next generation sequencing (Tn-seq) technology will be utilized to identify genes for NO resistance in V. parahaemolyticus. Our strategy will include construction the saturating transposon insertion library, transposon library challenging with NO, next generation sequencing (NGS), bioinformatics analysis and verification of the identified genes in vitro and in vivo.

Keywords: vibrio parahaemolyticus, nitric oxide, tn-seq, virulence

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Authors: Sijie Chen, Xiaofang Chen, Juan Wang, Yingying Zhang, Yu Hong, Wanyu Zhuang, Xinxin Huang, Ping Ou, Longsheng Huang

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Purpose: Autistic symptom improvement can be observed in children treated with acupuncture, but the mechanism is still being explored. In the present study, we used scalp acupuncture to treat autism rat model, and then their improvement in the abnormal behaviors and specific mechanisms behind were revealed by detecting animal behaviors, analyzing the RNA sequencing of the prefrontal cortex(PFC), and observing the ultrastructure of PFC neurons under the transmission electron microscope. Methods: On gestational day 12.5, Wistar rats were given valproic acid (VPA) by intraperitoneal injection, and their offspring were considered to be reliable rat models of autism. They were randomized to VPA or VPA-acupuncture group (n=8). Offspring of Wistar pregnant rats that were simultaneously injected with saline were randomly selected as the wild-type group (WT). VPA_acupuncture group rats received acupuncture intervention at 23 days of age for 4 weeks, and the other two groups followed without intervention. After the intervention, all experimental rats underwent behavioral tests. Immediately afterward, they were euthanized by cervical dislocation, and their prefrontal cortex was isolated for RNA sequencing and transmission electron microscopy. Results: The main results are as follows: 1. Animal behavioural tests: VPA group rats showed more anxiety-like behaviour and repetitive, stereotyped behaviour than WT group rats. While VPA group rats showed less spatial exploration ability, activity level, social interaction, and social novelty preference than WT group rats. It was gratifying to observe that acupuncture indeed improved these abnormal behaviors of autism rat model. 2. RNA-sequencing: The three groups of rats differed in the expression and enrichment pathways of multiple genes related to synaptic function, neural signal transduction, and circadian rhythm regulation. Our experiments indicated that acupuncture can alleviate the major symptoms of ASD by improving these neurological abnormalities. 3. Under the transmission electron microscopy, several lysosomes and mitochondrial structural abnormalities were observed in the prefrontal neurons of VPA group rats, which were manifested as atrophy of the mitochondrial membran, blurring or disappearance of the mitochondrial cristae, and even vacuolization. Moreover, the number of synapses and synaptic vesicles was relatively small. Conversely, the mitochondrial structure of rats in the WT group and VPA_acupuncture was normal, and the number of synapses and synaptic vesicles was relatively large. Conclusion: Acupuncture effectively improved the abnormal behaviors of autism rat model and the ultrastructure of the PFC neurons, which might worked by improving their abnormal synaptic function, synaptic plasticity and promoting neuronal signal transduction.

Keywords: autism spectrum disorder, acupuncture, animal behavior, RNA sequencing, transmission electron microscope

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Authors: Didem Güven, Oytun Hanhan, Elif Ceren Aksoy, Emine Ubay Çokgör

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This paper presents a biological treatability study ofpaintshopwastewaterof a bus factory by an anoxic/aerobic sequencing batch reactor.A lab scale 14L SBR system was implementedto investigate carbon and nitrogen removal performance frompaint shop waste streams combined with domestic and process wastewater of a bus production factory in Istanbul (Turkey).The wastewater collected from decanters of the paint boots and pre-treatmentplant was usedforthefeeding of SBR. The reactor was operated with a total hydraulic retention time of 24 hrs, and a total sludge age of 18.7 days. Initially the efficiency and stability of the reactor were studied when fed with main wastewater stream to simulate the current wastewater treatment plant. Removal efficiency of 57% nitrogen and 90% COD were obtained. Once the paint shop wastewater was introduced to mainstream feeding with a ratio of 1:5, nitrification completely, carbon removal were partially inhibited. SBR system was successful to handle even at very high COD concentrations of paint shop wastewater after feeding of 2 months, with an average effluent COD of 100 mg/L. For the determination of kinetic parameters, respirometric analysis was also conducted with/without paint shop wastewater addition. Model simulation indicated lower maximum specific growth and hydrolysis rates when paint shop wastewater was mixed with the mainstream wastewater of the factory.

Keywords: biological treatability, nitrogen removal, paint shop wastewater, sequencing batch reactor

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Authors: Petros Gkotsis, Anastasios Zouboulis, Manasis Mitrakas, Dimitrios Zamboulis, E. Peleka

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The most serious drawback in wastewater treatment using membrane bioreactors (MBRs) is membrane fouling which gradually leads to membrane permeability decrease and efficiency deterioration. This work is part of a research project that aims to develop an integrated methodology for membrane fouling control, using specific chemicals which will enhance the coagulation and flocculation of compounds responsible for fouling, hence reducing biofilm formation on the membrane surface and limiting the fouling rate acting as a pre-treatment step. For this purpose, a pilot-scale plant with fully automatic operation achieved by means of programmable logic controller (PLC) has been constructed and tested. The experimental set-up consists of four units: wastewater feed unit, bioreactor, membrane (side-stream) filtration unit and permeate collection unit. Synthetic wastewater was fed as the substrate for the activated sludge. The dissolved oxygen (DO) concentration of the aerobic tank was maintained in the range of 2-3 mg/L during the entire operation by using an aerator below the membrane module. The membranes were operated at a flux of 18 LMH while membrane relaxation steps of 1 min were performed every 10 min. Both commercial and composite coagulants are added in different concentrations in the pilot-scale plant and their effect on the overall performance of the ΜΒR system is presented. Membrane fouling was assessed in terms of TMP, membrane permeability, sludge filterability tests, total resistance and the unified modified fouling index (UMFI). Preliminary tests showed that particular attention should be paid to the addition of the coagulant solution, indicating that pipe flocculation effectively increases hydraulic retention time and leads to voluminous sludge flocs. The most serious drawback in wastewater treatment using MBRs is membrane fouling, which gradually leads to membrane permeability decrease and efficiency deterioration. This results in increased treatment cost, due to high energy consumption and the need for frequent membrane cleaning and replacement. Due to the widespread application of MBR technology over the past few years, it becomes clear that the development of a methodology to mitigate membrane fouling is of paramount importance. The present work aims to develop an integrated technique for membrane fouling control in MBR systems and, thus, contribute to sustainable wastewater treatment.

Keywords: coagulation, membrane bioreactor, membrane fouling, pilot plant

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Authors: Rozhgar A. Khailany, Mehri Igci, Emine Bayraktar, Sakip Erturhan, Metin Karakok, Ahmet Arslan

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Kidney cancer is the most lethal urological cancer accounting for 3% of adult malignancies. VHL, a tumor-suppressor gene, is best known to be associated with renal cell carcinoma (RCC). The VHL functions as negative regulator of hypoxia inducible factors. Recent sequencing efforts have identified several novel frequent mutations of histone modifying and chromatin remodeling genes in ccRCC (clear cell RCC) including PBRM1 and SETD2. The PBRM1 gene encodes the BAF180 protein, which involved in transcriptional activation and repression of selected genes. SETD2 encodes a histone methyltransferase, which may play a role in suppressing tumor development. In this study, RNAs of 30 paired tumor and normal samples that were grouped according to the types of kidney cancer and clinical characteristics of patients, including gender and average age were examined by RT-PCR, SSCP and sequencing techniques. VHL, PBRM1 and SETD2 expressions were relatively down-regulated. However, statistically no significance was found (Wilcoxon signed rank test, p > 0.05). Interestingly, no mutation was observed on the contrary of previous studies. Understanding the molecular mechanisms involved in the pathogenesis of RCC has aided the development of molecular-targeted drugs for kidney cancer. Further analysis is required to identify the responsible genes rather than VHL, PBRM1 and SETD2 in kidney cancer.

Keywords: kidney cancer, molecular biomarker, expression analysis, mutation screening

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Authors: M. Suwaibah, S. W. Tan, I. Aiini, K. Yusoff, A. R. Omar

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Respiratory diseases are the most important infectious diseases affecting poultry worldwide. One of the avian respiratory virus of global importance causing significant economic losses is Infectious Bronchitis Virus (IBV). The virus causes a wide spectrum disease known as Infectious Bronchitis (IB), affecting not only the respiratory system but also the kidney and the reproductive system, depending on its strain. IB and Newcastle disease are two of the most prevalent diseases affecting poultry in Malaysia. However, a study on the molecular characterization of Malaysian IBV is lacking. In this study, an IBV strain IBS130 which was isolated in 2015 was fully sequenced using next-gene sequencing approach. Sequence analysis of IBS130 based on the complete genome, polyprotein 1ab and S1 genes were compared with other IBV sequences available in Genbank, National Center for Biotechnology Information (NCBI). IBV strain IBS130 is characterised as QX-like strain based on whole genome and S1 gene sequence analysis. Comparisons of the virus with other IBV strains showed that the nucleotide identity ranged from 67% to 99.2%, depending on the region analysed. The similarity in whole genome nucleotide ranging from 84.9% to 90.7% with the least similar was from Singapore strains (84.9%) and highly similar with China QX-like strains. Meanwhile, the similarity in polyprotein 1ab ranging from 85.3% to 89.9% with the least similar to Singapore strains (85.3%) and highly similar with Mass strains from USA.

Keywords: infectious bronchitis virus, phylogenetic analysis, chicken, Malaysia

Procedia PDF Downloads 147