ePosters - S518

Routine use of targeted NGS panel in a Dutch cardiomyopathy cohort
 
Mohamed Z. Alimohamed, Lennart F. Johansson, Ludolf G. Boven, Krista van Dijk, Paul van der Zwaag, Richard J. Sinke, Rolf H. Sijmons, Birgit Sikkema-Raddatz, Jan D H Jongbloed
Departments of Genetics, University Medical Center Groningen, Groningen, The Netherlands
 
Background:
Next generation sequencing is increasingly used for clinical evaluation of patients with cardiomyopathies because it allows for simultaneous evaluation of multiple genes known to be associated with the disease. However, the diagnostic yield is variable in routine clinical practice. Furthermore, analysis of copy number variations is still not routinely performed.
 
Objectives:
(1) To determine the diagnostic yield of our custom targeted NGS gene panel used in routine clinical diagnostics in a Dutch cohort of over 2000 cardiomyopathy patients. (2) Examine the impact on the yield of analyzing this cohort for single or multiple exon duplications and deletions.
 
Methods:
Up to 61 genes known to be implicated in cardiomyopathies were selected for enrichment and analysis on DNA isolated from peripheral blood from patients as a routine procedure. Patients directed for genetic testing to our clinical genetics laboratory having a referral diagnosis for various types of cardiomyopathies were included in this study [N=2002]. A written informed consent was obtained for all patients referred to our clinical genetics laboratory. Classification of variants was based on guidelines for variant interpretation recommended by the American College of Medical Genetics and Genomics. Diagnostic yield was calculated using cases where a likely pathogenic or pathogenic variant was detected. Putative exonic deletions/duplications were analyzed using CoNVaDING and XHMM tools using settings previously described.
 
Results and Conclusion:
An overall diagnostic yield of 23% was achieved. CNVs were detected in 16 patients. Variants of unknown clinical significance were identified in 39% patients. These results illustrate the need to further reassess disease variant classification.
 
Key words:
Cardiomyopathy, NGS-panel, Diagnostic-yield

Continuous Circulation of Human derived Pandemic H1N1pdm09 Influenza Virus in Nigerian Pigs: Implication for Public Health and Pandemic Preparedness
 
Ijomanta, J.^1,+, Asala, O.¹, Bitrus, M.¹, Musa, A.², Shittu, I.¹, Joannis, T.¹, Nwosuh, C.¹ and ¹Meseko, C.*
¹Regional Centre for Animal Influenza, National Veterinary Research Institute, Vom Nigeria; ²Veterinary Department, Plateau State Ministry of Agriculture, Jos Nigeria
+presenting author
*Corresponding author: cameseko@yahoo.com
 
Influenza virus A belongs to the family Orthomyxoviridae and is divided into subtypes H1-18. The virus affects a wide range of avian and mammalian hosts, subtypes H1; H3; H5; H7 and H9 have been reported to cause fatal infections that can adversely affect socio-economics and public health. In April 2009, WHO declared influenza pandemic when H1N1pdm09 spread to 214 countries and an estimated 151,700 -575 400 deaths were recorded. The virus was subsequently transmitted to pigs and had remained enzootic being detected in many countries including Nigeria.
 
In this study, a total of 143 tracheal swabs were collected during a targeted active surveillance of apparently healthy pigs in Jos abattoir, Plateau state from December 2017 through February 2018. The samples were analyzed for Influenza A by conventional One-step RT-PCR protocol targeting the matrix.  To subtype the Influenza A viruses, multiplex RT-qPCR assays targeting the Hemagglutinin (H1pdm, H1avian, H1human and H3) and neuraminidase (N1, N2, N1pdm) were used. In all, 51 (35%) samples were positive for Influenza A while four of the ten selected samples for subtyping were identified as H1N1pdm09.
 
In a previous study carried out at the human-animal interface, circulation of H1N1pdm09 was reported in southwest Nigeria. Also, follow up studies revealed seroprevalence ranging from 20-60% in pigs. Here, we report a more recent and continuous circulation of the virus in pigs in North central Nigeria since it was first declared a pandemic in 2009.  The virus in human is now seasonal and can easily be managed with vaccination. Currently, there is no vaccination programme for swine influenza in Nigeria therefore maintaining H1N1pdm09 in pigs poses great public health risk. Possible mutations and reassortments may result in emergence of novel virus with pandemic potential.
 
Keywords: surveillance; Influenza A virus; Nigeria; Pandemics; Pigs
 

 

Mitogenomic Analysis of Western Baggara Cattle Breed-Nyalawi Population Revealed High Level of Haplotype Diversity and Population Expansion
 
Sumaya Kambal¹, Bashir Salim^2
1Faculty of Animal Production Science and Technology, Sudan University of Science and Technology; ²Department of Parasitology, Faculty of Veterinary Medicine, University of Khartoum
  
Western Baggara cattle breed is naturally inhabiting the western regions of the Sudan, primarily raised by nomadic pastoralists in Darfur and Kurdofan states and ranked the best beef producing cattle breed in the country. This breed had structured phenotypically into three populations, Nyalawi, Rezaigi and Messairi. Decades to date, western Baggara cattle breed-Nyalawi population (WBCB-NP) is the most reputable beef cattle among the other types by its large size that contribute significantly to local meat consumption as well as to the export revenue. Knowledge-based breeding programs and conservation strategies are necessarily needed to aid in sustainable production and preservation of such population.
This study was undertaken to investigate the maternal genetic diversity and the demography dynamic by addressing whether this population had undergone demographic expansion, decline or equilibrium.
 
Genomic DNA was extracted from blood of 30 animals selected WBCB-NP and subjected to PCR targeting the entire mitochondrial DNA D-loop region and subsequently to DNA sequencing. This resulted in 43 polymorphic sites that defined 28 haplotypes of which 4 signatures of breed specific transition were detected when compared to bovine reference sequence. Genetic diversity indices revealed high level of haplotype diversity (1.000 ± 0.0091) and low level of nucleotide diversity (0.005 ± 0.0028) between these haplotypes that agreed with the Median Joining and Minimum Spanning networks' pattern, and the Neighbor Joining phylogenetic tree. The revealed unimodal pattern of the mismatch distribution among the haplotypes was in a good accordance with both the negative values of Tajima's D (-2.259; P= 0.000) and Fu's Fs (-25.671; P= 0.000) results that are significantly deviated from the neutral model, suggesting population demographic expansion event.
 
It is concluded that WBCB-NP is highly divers and had experienced a demographic growth. This study has raised important queries about the nature and the factors of this demographic expansion that can be compared with other populations of cattle for historical dynamics tracing purpose. Therefore, this research lays the groundwork for future research enhancing our understanding of Baggara breed as well as the other local cattle breeds. Further Bayesian analysis is required to trace the time series and nature of this population expansion.

 

Genetic Diversity of Plasmodium falciparum Parasites in Pregnant and Non-pregnant Women and Potential Resistance to Antimalarial Drugs in Kenya
 
Brenda Makena^1,2, Martha Nginya^1,2, Marcel Nyambute¹, Charles Okello¹, Edwin Mwakio¹, Gladys Chemwor¹, Raphael Okoth¹, Redemptah Yeda¹, Agnes Cheruiyot¹, Benjamin Opot¹, Dennis Juma¹, Victor Mobegi², Hosea Akala¹, Ben Andagalu^1
1United States Army Medical Research Directorate-Kenya (USAMRD-K), Kenya Medical Research Institute (KEMRI)
²Centre for Biotechnology and Bioinformatics (CEBIB). University of Nairobi, Nairobi, Kenya.
 
During pregnancy, malaria causes maternal anaemia, placental accumulation of parasites, low birth weight, infant mortality as well as maternal mortality. Pregnant women are presumed non-immune to malaria whereas the non-pregnant are thought to be semi-immune. Non-pregnant women therefore have faster parasite clearance compared to pregnant women. The non-immune environment in pregnant women may select for parasite populations that are associated with resistance to artemisinin. This study will determine genetic variations in Plasmodium falciparum parasites in pregnant versus non-pregnant women due to differences in their immunity. Samples were collected at hours 0, 8, 24 and at days 7 and 28 from 75 participants in an efficacy study. Rapid diagnostic test and microscopy will be done followed by 18s rRNA PCR for speciation analysis. Sanger sequencing and MassARRAY SNP genotyping will be done to determine mutations in parasites without fast clearance as well as microsatellite genotyping to determine genetic variations in the parasites between the two populations. Genotyping for both MSP1 and MSP2 will then follow to determine recrudescence and reinfection. Twelve microsatellite markers will be used to calculate allele frequencies, heterozygosity (H_e) values, molecular variance, haplotypes, principal coordinates and fixation indices using GenAIEx and Arlequin softwares. For MassARRAY SNP genotyping, genotype calls will be made using SpectroTyper 4.0 software (Agena). For the K13 and Sanger sequencing SNPs will be analyzed by doing multiple sequence alignment using CLC Genomics Workbench software. This study will contribute to knowledge on guiding the World Health Organization implementation of intermittent preventive treatment in pregnancy with sulfadoxine-pyrimethamine if found to influence parasites response to artemisinins.
Funding: Armed Forces Health Surveillance Branch (AFHSB) and Developing Excellence in Leadership and Genetics Training for Malaria Elimination in Sub-Africa (DELGEME)