Whole genome sequencing and phylogenetic analysis of SARS-CoV-2 strains isolated during the COVID-19 pandemic in Nigeria.

Objectives: The emergence and spread of SARS-CoV-2 have stimulated ongoing research into the virus transmission dynamics, circulating variants, and potential mutations. This study was conducted to understand the genomic dynamics of the epidemic in Nigeria. Design: Whole genome sequencing was conducted on SARS-CoV-2 samples collected during the first and second outbreaks using the Oxford Nanopore MinION sequencing platform. Phylogenetic analysis was conducted, and genomes were grouped into different pangolin lineages. Results: The study revealed four circulating SARS-CoV-2 variants. The Alpha (B.1.1.7) variant was the most prevalent (32.7%), followed by Beta (B.1 B.1.1, L.3, and B.1.1.318) (30.8%), Eta (B.1.525) (28.9%), and Delta (B.1.617, AY.1, AY.109, and AY.36) (7.7%). Phylogenetic analysis revealed three clusters with four Nextstrain clades (20I, 20B, 21D, and 21J). The Alpha lineages (B.1.1.7) clustered with references from Italy. The Beta lineages (Clade 20B) (B.11, B.11318, and L3) and sub-lineage B.11 were distinct. Sub-lineage B.11318 is clustered with references from the USA, whereas sub-lineage L3 is clustered with references from Russia, the Philippines, Australia, and Japan. The 21D and 21J, belonging to two Pango lineages, Eta (B.1525) and Delta (B.1.617 and AY.109), showed high genetic similarity. Conclusion: The phylogenetic relatedness of the lineages suggests multiple virus introduction, which could be a source of more virulent, locally adapted variants.

High genetic and haplotype diversity in vaccine candidate Pfceltos but not Pfrh5 among malaria-infected children in Ibadan, Nigeria.

Malaria remains a global public health challenge. The disease has a great impact in sub-Saharan Africa among children under five years of age and pregnant women. Malaria control programs targeting the parasite and mosquitoes vectors with combinational therapy and insecticide-treated bednets are becoming obsolete due to the phenomenon of resistance, which is a challenge for reducing morbidity and mortality. Malaria vaccines would be effective alternative to the problem of parasite and insecticide resistance, but focal reports of polymorphisms in malaria candidate antigens have made it difficult to design an effective malaria vaccine. Therefore, studies geared towards elucidating the polymorphic pattern and how genes targeted for vaccine design evolve are imperative. We have carried out molecular and genetic analysis of two genes encoding vaccine candidates-the Plasmodium falciparum cell traversal ookinetes and sporozoites (Pfceltos) and P. falciparum reticulocyte binding protein 5 (Pfrh5) in parasite isolates from malaria-infected children in Ibadan, Nigeria to evaluate their genetic diversity, relatedness and pattern of molecular evolution. Pfceltos and Pfrh5 genes were amplified from P. falciparum positive samples. Amplified fragments were purified and sequenced using the chain termination method. Post-sequence edit of fragments and application of various population genetic analyses was done. We observed a higher number of segregating sites and haplotypes in the Pfceltos than in Pfrh5 gene, the former also presenting higher haplotype (0.942) and nucleotide diversity (θ = 0.01219 and π = 0.01148). In contrast, a lower haplotype (0.426) and nucleotide diversity (θ = 0.00125; π = 0.00095) was observed in the Pfrh5 gene. Neutrality tests do not show deviation from neutral expectations for Pfceltos, with the circulation of multiple low frequency haplotypes (Tajima's D = -0.21637; Fu and Li's D = -0.08164; Fu and Li's F = -0.14051). Strong linkage disequilibrium was observed between variable sites, in each of the genes studied. We postulate that the high diversity and circulation of multiple haplotypes has the potential of making a Pfceltos-subunit vaccine ineffective, while the low genetic diversity of Pfrh5 gene substantiates its evolutionary conservation and potential as a malaria vaccine candidate.

Severity of Schistosoma haematobium co-infection with malaria in school-children is potentially modulated by host CD14 gene variants.

OBJECTIVE: Schistosomiasis remains a chronic disease of global importance, especially in many rural areas of the world where co-infection with Plasmodium falciparum is common. It is critical to decipher the role of single or co-infected disease scenarios on immune system regulation in such individuals and how such co-infections can either ameliorate or complicate immune response and the consequent disease outcome. First, 10 ml of urine samples, collected between 10:00 am and 2:00 pm, was filtered for diagnosis of schistosomiasis, while egg count, indicative of disease severity, was determined by microscopy. Furthermore, genomic DNA samples extracted from dried blood spots collected on filter paper from one hundred and forty-four Schistosoma haematobium-infected school-children was tested for P. falciparum parasite positivity by an allele-specific nested-PCR analysis of merozoite surface protein (msp)-1 and -2 genes and a real-time PCR assay. In addition, among P. falciparum parasite-positive individuals, we carried out a Taqman SNP genotyping assay to extrapolate the effect of host CD14 (-159 C/T; rs2569190) genetic variants on schistosome egg count. RESULTS: Of the 144 individuals recruited, P. falciparum parasite positivity with msp-1 gene were 34%, 43% and 55% for MAD20, RO33 and K1 alleles respectively. Of the co-infected individuals, CD14 genetic variants ranged from 18.8% vs 21.5%, 33.3% vs 44.4%, 9.7% vs 11.8% for single versus schistosome co-infection for the wild type (CC), heterozygous (CT) and mutant (TT) variants respectively. Though the mean egg count for co-infected individuals with CD14 wild type (33.7 eggs per 10 ml of urine) and heterozygote variants (37.5 eggs per 10 ml of urine) were lower than that of schistosome infection alone (52.48 and 48.08 eggs/10 ml of urine respectively), it lacked statistical significance (p-value 0.12 and 0.29), possibly reflecting the benefit of the CD14 activation in schistosome plus malaria co-infection and not schistosome infection alone. In addition, the lower mean egg count in co-infected individuals reveal the benefit of downstream Th1 immune response mitigated by CD14 innate activation that is absent in schistosome infection alone.

Genetic diversity of Plasmodium falciparum among asymptomatic pregnant women on intermittent preventive treatment with sulfadoxine-pyrimethamine in Nigeria.

This study investigated the genetic diversity of Plasmodium falciparum among asymptomatic pregnant women on intermittent preventive treatment with sulfadoxine-pyrimethamine (IPTp-Sp) in Osogbo, southwest Nigeria. Blood sample was obtained from consenting pregnant women attending antenatal clinics. Microscopy and Polymerase chain reaction (PCR) were employed to diagnose and analyse genetic diversity. Of the 301 samples, 53 (18%) and 83 (28%) were positive for P. falciparum by microscopy and PCR, respectively. Using the merozoite surface protein (msp)-1, msp-2, and glutamate-rich protein (glurp) genes of P. falciparum as polymorphic markers, the msp-1 gene showed nine alleles with R033 (66.7%) being predominant, followed by K1 (45.5%) and MAD20 (33.3%). The msp-2 gene had 16 alleles (eight each for FC27 and 3D7). The 3D7 alleles (82.1%) was significantly more than FC27 alleles (48.2%) (p = 0.0093). Nine alleles were detected with glurp gene, presenting with the highest monoclonal and the lowest polyclonal infection. The multiplicity of infection (MOI) of 1.5, 1.8, and 1.2 were obtained for msp-1, msp-2 and glurp genes. In light of the high P. falciparum genetic diversity among pregnant women on IPT-Sp in this study, additional strategies for preventing and controlling malaria in pregnancy might be required.

Molecular genotyping reveals mixed bovine and human trypanosomiasis in cattle from West Africa.

Background and Aim: Animal trypanosomiasis is a major contributor to agricultural and economic losses, especially in sub-Saharan Africa. We have shown that some animal species expressed genes that are significant players in immune response to bovine trypanosomosis, impeding signs and symptoms of the disease. We hypothesize that such animals are contributors to disease transmission dynamics and severe outcomes. Therefore, this study aims to ascertain trypanosome species diversity in cattle and their potential role as reservoirs for the transmission of human disease. Materials and Methods: We performed a molecular genotyping of trypanosome internal transcribed spacer 1 (ITS-1) and 18S ribosomal RNA genes on genomic DNA extracts from randomly sampled N'Dama cattle from slaughterhouses in Nigeria. We identified trypanosome species circulating among the animals through polymerase chain reaction and genomic sequencing. We performed multiple sequence alignments as well as conducted a phylogenetic relationship between identified species. Results: In all, 9 of 127 (7.1%) samples were positively amplified (band sizes ranging from 250 bp to 710 bp), including an isolate with two distinct bands (700 and 710 bp), indicating two trypanosome types. Sequence similarity and homology analysis identified four species, namely: Trypanosoma vivax, Trypanosoma congolense forest type, T. congolense savannah type, and Trypanosoma brucei. Interestingly, one of the bands, additionally verified by nucleotide sequencing, was identified as a human trypanosome (Trypanosoma brucei gambiense), confirming our hypothesis that cattle are potential reservoir hosts for human trypanosomes. Conclusion: Overall, we observed different trypanosome species in our study area, with animals on the same farm infected with multiple species, which could complicate treatment and disease control strategies. Finding human trypanosome species strengthens the argument that disease transmission dynamics are modulated by other vertebrates, further complicating control programs.

Molecular Evolution and Inheritance Pattern of Sox Gene Family among Bovidae.

Sox genes are an evolutionarily conserved family of transcription factors that play important roles in cellular differentiation and numerous complex developmental processes. In vertebrates, Sox proteins are required for cell fate decisions, morphogenesis, and the control of self-renewal in embryonic and adult stem cells. The Sox gene family has been well-studied in multiple species including humans but there has been scanty or no research into Bovidae. In this study, we conducted a detailed evolutionary analysis of this gene family in Bovidae, including their physicochemical properties, biological functions, and patterns of inheritance. We performed a genome-wide cataloguing procedure to explore the Sox gene family using multiple bioinformatics tools. Our analysis revealed a significant inheritance pattern including conserved motifs that are critical to the ability of Sox proteins to interact with the regulatory regions of target genes and orchestrate multiple developmental and physiological processes. Importantly, we report an important conserved motif, EFDQYL/ELDQYL, found in the SoxE and SoxF groups but not in other Sox groups. Further analysis revealed that this motif sequence accounts for the binding and transactivation potential of Sox proteins. The degree of protein-protein interaction showed significant interactions among Sox genes and related genes implicated in embryonic development and the regulation of cell differentiation. We conclude that the Sox gene family uniquely evolved in Bovidae, with a few exhibiting important motifs that drive several developmental and physiological processes.

Population genetic analysis of Plasmodium falciparum cell-traversal protein for ookinetes and sporozoite among malaria patients from southern Nigeria.

Plasmodium falciparum immune escape mechanisms affect antigens being prioritized for vaccine design. As a result of the multiple surface antigens the parasite exhibits at different life cycle stages, designing a vaccine that would efficiently boost the immune system in clearing infections has been challenging. The P. falciparum cell-traversal protein for ookinetes and sporozoite (Pfceltos) is instrumental for ookinete traversal of the mosquito midgut and sporozoites invasion of the human liver cells. Pfceltos elicits both humoral and cellular immune response but has been reported with multiple single nucleotide polymorphisms in global isolates. A cross-sectional survey, conducted in southern Nigeria, between January-March 2021 recruited 283 individuals. Of this, 166 demonstrated P. falciparum infections (86 from Cross River and 80 from Edo), 48 (55.8%) while only 36 (45%) were amplified for Pfceltos gene from both sites respectively. Fifty amplified samples were sequenced and analysed for their diversity, polymorphisms and population structure of the gene. The number of segregating sites in Edo State was higher (34) than that of Cross River State. Though nucleotide diversity was higher for Edo compared to Cross River State (θw = 0.02505; π = 0.03993 versus θw = 0.00930; π = 0.01033 respectively), the reverse was the case for haplotype diversity (0.757 versus 0.890 for Edo and Cross River respectively). Of the twelve haplotypes observed from both states, only two (KASLPVEK and NAFLSFEK) were shared, with haplotype prevalence higher in Edo (16% and 36%) than Cross River (8% and 4%). The Tajima's D test was positive for both states, with Fst value showing a strong genetic differentiation (Fst = 0.25599), indicating the occurrence of balancing selection favoring haplotype circulation at a low frequency. The shared haplotypes, low Hst and Fst values presents a challenge to predict the extent of gene flow. High LD values present a grim public health consequence should a Pfceltos-conjugated vaccine be considered for prophylaxis in Nigeria.

Deciphering inhibitory mechanism of coronavirus replication through host miRNAs-RNA-dependent RNA polymerase interactome.

Despite what we know so far, Covid-19, caused by SARS-CoV-2 virus, remains a pandemic that still require urgent healthcare intervention. The frequent mutations of the SARS-CoV-2 virus has rendered disease control with vaccines and antiviral drugs quite challenging, with newer variants surfacing constantly. There is therefore the need for newer, effective and efficacious drugs against coronaviruses. Considering the central role of RNA dependent, RNA polymerase (RdRp) as an enzyme necessary for the virus life cycle and its conservation among coronaviruses, we investigated potential host miRNAs that can be employed as broad-range antiviral drugs averse to coronaviruses, with particular emphasis on BCoV, MERS-CoV, SARS-CoV and SARS-CoV-2. miRNAs are small molecules capable of binding mRNA and regulate expression at transcriptional or translational levels. Our hypothesis is that host miRNAs have the potential of blocking coronavirus replication through miRNA-RdRp mRNA interaction. To investigate this, we retrieved the open reading frame (ORF1ab) nucleotide sequences and used them to interrogate miRNA databases for miRNAs that can bind them. We employed various bioinformatics tools to predict and identify the most effective host miRNAs. In all, we found 27 miRNAs that target RdRp mRNA sequence of multiple coronaviruses, of which three - hsa-miR-1283, hsa-miR-579-3p, and hsa-miR-664b-3p target BCoV, SARS-CoV and SARS-CoV-2. Additionally, hsa-miR-374a-5p has three bovine miRNA homologs viz bta-miR-374a, bta-miR-374b, and bta-miR-374c. Inhibiting the expression of RdRp enzyme via non-coding RNA is novel and of great therapeutic importance in the control of coronavirus replication, and could serve as a broad-spectrum antiviral, with hsa-miR-1283, hsa-miR-579-3p, and hsa-miR-664b-3p as highly promising.

Genetic Diversity and Phylogenetic Relatedness of Plasmodium ovale curtisi and Plasmodium ovale wallikeri in sub-Saharan Africa.

P. ovale was until recently thought to be a single unique species. However, the deployment of more sensitive tools has led to increased diagnostic sensitivity, including new evidence supporting the presence of two sympatric species: P. ovale curtisi (Poc) and P. ovale wallikeri (Pow). The increased reports and evolution of P. ovale subspecies are concerning for sub-Saharan Africa where the greatest burden of malaria is borne. Employing published sequence data, we set out to decipher the genetic diversity and phylogenetic relatedness of P. ovale curtisi and P. ovale wallikeri using the tryptophan-rich protein and small subunit ribosomal RNA genes from Gabon, Senegal, Ethiopia and Kenya. Higher number of segregating sites were recorded in Poc isolates from Gabon than from Ethiopia, with a similar trend in the number of haplotypes. With regards to Pow, the number of segregating sites and haplotypes from Ethiopia were higher than from those in Gabon. Poc from Kenya, had higher segregating sites (20), and haplotypes (4) than isolates from Senegal (8 and 3 respectively), while nucleotide from Senegal were more diverse (θw = 0.02159; π = 0.02159) than those from Kenya (θw = 0.01452; π = 0.01583). Phylogenetic tree construction reveal two large clades with Poc from Gabon and Ethiopia, and distinct Gabonese and Ethiopian clades on opposite ends. A similar observation was recorded for the phylogeny of Poc isolates from Kenya and Senegal. With such results, there is a high potential that ovale malaria control measures deployed in one country may be effective in the other since parasite from both countries show some degree of relatedness. How this translates to malaria control efforts throughout the continent would be next step deserving more studies.

Isolation, Whole-Genome Sequencing, and Annotation of Three Unclassified Antibiotic-Producing Bacteria, Enterobacter sp. Strain RIT 637, Pseudomonas sp. Strain RIT 778, and Deinococcus sp. Strain RIT 780.

We report the isolation, whole-genome sequencing, and annotation of Enterobacter sp. strain RIT 637, Pseudomonas sp. strain RIT 778, and Deinococcus sp. strain RIT 780. Disk diffusion assays using spent medium demonstrated that all bacteria produced bactericidal compounds against Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, and Staphylococcus aureus ATCC 25923.

Regulatory network of miRNA, lncRNA, transcription factor and target immune response genes in bovine mastitis.

Pre- and post-transcriptional modifications of gene expression are emerging as foci of disease studies, with some studies revealing the importance of non-coding transcripts, like long non-coding RNAs (lncRNAs) and microRNAs (miRNAs). We hypothesize that transcription factors (TFs), lncRNAs and miRNAs modulate immune response in bovine mastitis and could potentially serve as disease biomarkers and/or drug targets. With computational analyses, we identified candidate genes potentially regulated by miRNAs and lncRNAs base pair complementation and thermodynamic stability of binding regions. Remarkably, we found six miRNAs, two being bta-miR-223 and bta-miR-24-3p, to bind to several targets. LncRNAs NONBTAT027932.1 and XR_003029725.1, were identified to target several genes. Functional and pathway analyses revealed lipopolysaccharide-mediated signaling pathway, regulation of chemokine (C-X-C motif) ligand 2 production and regulation of IL-23 production among others. The overarching interactome deserves further in vitro/in vivo explication for specific molecular regulatory mechanisms during bovine mastitis immune response and could lay the foundation for development of disease markers and therapeutic intervention.

Prevalence of urogenital and intestinal schistosomiasis among school children in South-west Nigeria.

BACKGROUND: The risk of co-infection with Schistosoma haematobium and S. mansoni and the potential harmful effect on morbidity and control is enhanced by the overlapping distribution of both species in sub-Saharan Africa. Despite the reported high endemicity of both species in Nigeria, studies on the spread and effect of their mixed infection are limited. Therefore, a cross-sectional survey was conducted among school children in two communities in South-west Nigeria to investigate the prevalence of mixed human schistosome infection, intensity, and possible ectopic egg elimination. METHODS: Urine and stool samples were collected from consenting school children in Ilie and Ore communities of Osun State, Nigeria. Schistosoma haematobium eggs were detected in urine using the urine filtration technique, while S. mansoni eggs were detected in stool using the Kato-Katz thick smear technique. RESULTS: The study enrolled 466 primary and secondary school children (211; 45.3% males vs. 255; 54.7% females; mean age 11.6 ± 3.16 years). The overall prevalence of schistosomiasis was 40% (185/466), with 19% (89/466) recording single S. haematobium infection while 9% (41/465) had a single S. mansoni infection. The geometric mean egg count for S. haematobium was 189.4 egg/10ml urine; 95% CI: range 115.9-262.9, while for S. mansoni, it was 115.7 epg; 95% CI: range 78.4-152.9. The prevalence of ectopic S mansoni (S. mansoni eggs in urine) was 4.7%, while no ectopic S. haematobium (S. haematobium eggs in stool) was recorded. Mixed infection of S. haematobium/S. mansoni had a prevalence of 9.5% (44/466). More females (54.5%) presented with S. haematobium/S. mansoni co-infection. For both parasites, males had higher infection intensity, with a significant difference observed with S. haematobium (p = 0.0004). Hematuria was significant in individuals with single S. haematobium infection (p = 0.002), mixed ectopic S. haematobium/S. mansoni (p = 0.009) and mixed S. haematobium/S. mansoni/ectopic S. mansoni (p = 0.0003). CONCLUSIONS: These findings suggest the probability of interspecific interactions between S. haematobium and S. mansoni. Scaling up of mass administration of praziquantel and control measures in the study areas is highly desirable.

Transcriptome Profiling Reveals CD73 and Age-Driven Changes in Neutrophil Responses against Streptococcus pneumoniae.

Neutrophils are required for host resistance against Streptococcus pneumoniae, but their function declines with age. We previously found that CD73, an enzyme required for antimicrobial activity, is downregulated in neutrophils (also known as polymorphonuclear leukocytes [PMNs]) from aged mice. This study explored transcriptional changes in neutrophils induced by S. pneumoniae to identify pathways controlled by CD73 and dysregulated with age. Pure bone marrow-derived neutrophils isolated from wild-type (WT) young and old and CD73 knockout (CD73KO) young mice were mock challenged or infected with S. pneumoniae ex vivo. RNA sequencing (RNA-Seq) was performed to identify differentially expressed genes (DEGs). We found that infection triggered distinct global transcriptional changes across hosts that were strongest in CD73KO neutrophils. Surprisingly, there were more downregulated than upregulated genes in all groups upon infection. Downregulated DEGs indicated a dampening of immune responses in old and CD73KO hosts. Further analysis revealed that CD73KO neutrophils expressed higher numbers of long noncoding RNAs (lncRNAs) than those in WT controls. Predicted network analysis indicated that CD73KO-specific lncRNAs control several signaling pathways. We found that genes in the c-Jun N-terminal kinase (JNK)-mitogen-activated protein kinase (MAPK) pathway were upregulated upon infection in CD73KO mice and in WT old mice, but not in WT young mice. This corresponded to functional differences, as phosphorylation of the downstream AP-1 transcription factor component c-Jun was significantly higher in neutrophils from infected CD73KO mice and old mice. Importantly, inhibition of JNK/AP-1 rescued the ability of these neutrophils to kill S. pneumoniae. Together, our findings revealed that the ability of neutrophils to modify their gene expression to better adapt to bacterial infection is in part regulated by CD73 and declines with age.

Integrative Network Analysis of Predicted miRNA-Targets Regulating Expression of Immune Response Genes in Bovine Coronavirus Infection.

Bovine coronavirus (BCoV) infection that causes disease outbreaks among farm animals, resulting in significant economic losses particularly in the cattle industry, has the potential to become zoonotic. miRNAs, which are short non-coding segments of RNA that inhibits the expression of their target genes, have been identified as potential biomarkers and drug targets, though this potential in BCoV remains largely unknown. We hypothesize that certain miRNAs could simultaneously target multiple genes, are significantly conserved across many species, thereby demonstrating the potential to serve as diagnostic or therapeutic tools for bovine coronavirus infection. To this end, we utilized different existing and publicly available computational tools to conduct system analysis predicting important miRNAs that could affect BCoV pathogenesis. Eleven genes including CEBPD, IRF1, TLR9, SRC, and RHOA, significantly indicated in immune-related pathways, were identified to be associated with BCoV, and implicated in other coronaviruses. Of the 70 miRNAs predicted to target the identified genes, four concomitant miRNAs (bta-miR-11975, bta-miR-11976, bta-miR-22-3p, and bta-miR-2325c) were found. Examining the gene interaction network suggests IL-6, IRF1, and TP53 as key drivers. Phylogenetic analysis revealed that miR-22 was completely conserved across all 14 species it was searched against, suggesting a shared and important functional role. Functional annotation and associated pathways of target genes, such as positive regulation of cytokine production, IL-6 signaling pathway, and regulation of leukocyte differentiation, indicate the miRNAs are major participants in multiple aspects of both innate and adaptive immune response. Examination of variants evinced a potentially deleterious SNP in bta-miR-22-3p and an advantageous SNP in bta-miR-2325c. Conclusively, this study provides new insight into miRNAs regulating genes responding to BCoV infection, with bta-miR-22-3p particularly indicated as a potential drug target or diagnostic marker for bovine coronavirus.

Dissecting Transcription Factor-Target Interaction in Bovine Coronavirus Infection.

Coronaviruses are RNA viruses that cause significant disease within many species, including cattle. Bovine coronavirus (BCoV) infects cattle and wild ruminants, both as a respiratory and enteric pathogen, and possesses a significant economic threat to the cattle industry. Transcription factors are proteins that activate or inhibit transcription through DNA binding and have become new targets for disease therapies. This study utilized in silico tools to identify potential transcription factors that can serve as biomarkers for regulation of BCoV pathogenesis in cattle, both for testing and treatment. A total of 11 genes were identified as significantly expressed during BCoV infection through literature searches and functional analyses. Eleven transcription factors were predicted to target those genes (AREB6, YY1, LMO2, C-Rel, NKX2-5, E47, RORAlpha1, HLF, E4BP4, ARNT, CREB). Function, network, and phylogenetic analyses established the significance of many transcription factors within the immune response. This study establishes new information on the transcription factors and genes related to host-pathogen interactome in BCoV infection, particularly transcription factors YY1, AREB6, LMO2, and NKX2, which appear to have strong potential as diagnostic markers, and YY1 as a potential target for drug therapies.

Differential Evolution of α-Glucan Water Dikinase (GWD) in Plants.

The alpha-glucan water dikinase (GWD) enzyme catalyzes starch phosphorylation, an integral step in transitory starch degradation. The high phosphate content in stored starch has great industrial value, due to its physio-chemical properties making it more versatile, although the phosphate content of stored starch varies depending on the botanical source. In this study, we used various computational approaches to gain insights into the evolution of the GWD protein in 48 plant species with possible roles in enzyme function and alteration of phosphate content in their stored starch. Our analyses identified deleterious mutations, particularly in the highly conserved 5 aromatic amino acid residues in the dual tandem carbohydrate binding modules (CBM-45) of GWD protein in C. zofingiensis, G. hirsutum, A. protothecoides, P. miliaceum, and C. reinhardtii. These findings will inform experimental designs for simultaneous repression of genes coding for GWD and the predicted interacting proteins to elucidate the role this enzyme plays in starch degradation. Our results reveal significant diversity in the evolution of GWD enzyme across plant species, which may be evolutionarily advantageous according to the varying needs for phosphorylated stored starch between plants and environments.

Analysis of sulphadoxine-pyrimethamine resistance-associated mutations in Plasmodium falciparum isolates obtained from asymptomatic pregnant women in Ogun State, Southwest Nigeria.

Intermittent preventive treatment in pregnancy with sulphadoxine-pyrimethamine (IPTp-SP) is one of the main strategies for protecting pregnant women, fetus, and their new-born against adverse effects of P. falciparum infection. The development of the drug resistance linked to mutations in P. falciparum dihydrofolate reductase gene (pfdhfr) and P. falciparum dihydropteroate synthase gene (pfdhps), is currently threatening the IPTp-SP approach. This study determined the prevalence of pfdhfr and pfdhps mutations in isolates obtained from pregnant women with asymptomatic P. falciparum infection in Nigerian. Additionally, P. falciparum genetic diversity and multiplicity of infection (MOI) was assessed by genotyping the P. falciparum merozoite surface Protein 1 and 2 (pfmsp-1 and pfmsp-2) genes. The pfdhfr and pfdhps were genotyped by direct sequencing, and the pfmsp-1 and pfmsp-2 fragment analysis by polymerase chain reaction was used to determine P. falciparum genetic diversity. Of the 406 pregnant women recruited, 123 had P. falciparum infection by PCR, and of these, 52 were successfully genotyped for pfdhfr and 42 for pfdhps genes. The pfdhfr triple-mutant parasites (N51I, C59R, and S108N) or the IRN haplotype were predominant (98%), whereas pfdhfr mutations C50R and I164L did not occur. For pfdhps gene, the prevalence of A437G, A581G, A436A, and A613S mutations were 98, 71, 55, and 36%, respectively. Nineteen (44%) isolates with quintuple mutations (CIRNI- SGKGA) had the highest combined pfdhfr-pfdhps haplotype. Isolates with sextuple mutants; CIRNI- AGKAS and CIRNI- AGKGA had a prevalence of 29 and 14%, respectively. High genetic diversity (7 pfmsp-1 alleles and 10 pfmsp-2 alleles) and monoclonal infection rate (76%) was observed. This study demonstrated a continuous high prevalence of pfdhfr mutation and an increase in pfdhps mutations associated with SP-resistance in southwest Nigeria. Continuous surveillance of IPTp-SP effectiveness and consideration of alternative IPTp strategies is recommended.

RNA-seq profiling of skin in temperate and tropical cattle.

Skin is a major thermoregulatory organ in the body controlling homeothermy, a critical function for climate adaptation. We compared genes expressed between tropical- and temperate-adapted cattle to better understand genes involved in climate adaptation and hence thermoregulation. We profiled the skin of representative tropical and temperate cattle using RNA-seq. A total of 214,754,759 reads were generated and assembled into 72,993,478 reads and were mapped to unique regions in the bovine genome. Gene coverage of unique regions of the reference genome showed that of 24,616 genes, only 13,130 genes (53.34%) displayed more than one count per million reads for at least two libraries and were considered suitable for downstream analyses. Our results revealed that of 255 genes expressed differentially, 98 genes were upregulated in tropically-adapted White Fulani (WF; Bos indicus) and 157 genes were down regulated in WF compared to Angus, AG (Bos taurus). Fifteen pathways were identified from the differential gene sets through gene ontology and pathway analyses. These include the significantly enriched melanin metabolic process, proteinaceous extracellular matrix, inflammatory response, defense response, calcium ion binding and response to wounding. Quantitative PCR was used to validate six representative genes which are associated with skin thermoregulation and epithelia dysfunction (mean correlation 0.92; p < 0.001). Our results contribute to identifying genes and understanding molecular mechanisms of skin thermoregulation that may influence strategic genomic selection in cattle to withstand climate adaptation, microbial invasion and mechanical damage.

Whole-Genome Sequencing of Pantoea sp. Strain RIT388, a Potential Oral Opportunistic Pathogen Isolated from a Chewing Stick (Distemonanthus benthamianus).

In this study, we report the isolation, identification, characterization, and whole-genome sequence of the endophyte Pantoea sp. strain RIT388, isolated from Distemonanthus benthamianus, a plant known for its antifungal and antibacterial properties that is commonly used for chewing sticks.

SNP Diversity in CD14 Gene Promoter Suggests Adaptation Footprints in Trypanosome Tolerant N'Dama (Bos taurus) but not in Susceptible White Fulani (Bos indicus) Cattle.

Immune response to infections has been shown to be mediated by genetic diversity in pattern recognition receptors, leading to disease tolerance or susceptibility. We elucidated naturally occurring variations within the bovine CD14 gene promoter in trypanosome-tolerant (N'Dama) and susceptible (White Fulani) cattle, with genomic and computational approaches. Blood samples were collected from White Fulani and N'Dama cattle, genomic DNA extracted and the entire promoter region of the CD14 gene amplified by PCR. We sequenced this region and performed in silico computation to identify SNP variants, transcription factor binding sites, as well as micro RNAs in the region. CD14 promoter sequences were compared with the reference bovine genome from the Ensembl database to identify various SNPs. Furthermore, we validated three selected N'Dama specific SNPs using custom Taqman SNP genotyping assay for genetic diversity. In all, we identified a total of 54 and 41 SNPs at the CD14 promoter for N'Dama and White Fulani respectively, including 13 unique SNPs present in N'Dama only. The significantly higher SNP density at the CD14 gene promoter region in N'Dama may be responsible for disease tolerance, possibly an evolutionary adaptation. Our genotype analysis of the three loci selected for validation show that mutant alleles (A/A, C/C, and A/A) were adaptation profiles within disease tolerant N'Dama. A similar observation was made for our haplotype analysis revealing that haplotypes H1 (ACA) and H2 (ACG) were significant combinations within the population. The SNP effect prediction revealed 101 and 89 new transcription factor binding sites in N'Dama and White Fulani, respectively. We conclude that disease tolerant N'Dama possessing higher SNP density at the CD14 gene promoter and the preponderance of mutant alleles potentially confirms the significance of this promoter in immune response, which is lacking in susceptible White Fulani. We, therefore, recommend further in vitro and in vivo study of this observation in infected animals, as the next step for understanding genetic diversity relating to varying disease phenotypes in both breeds.