Electron beam irradiation, without any addition of cross-linking agents, was investigated at varying
doses of EB-Irradiation to develop an environmentally friendly hybridized kenaf (bast)/ pineapple leaf
fibre (PALF) bio-composites. Improvement in tensile property of the hybrid was achieved with the result
showing a direct proportionality relationship between tensile properties and increasing radiation dose.
Statistical analysis software (SAS) was employed to validate the result. HDPE has been shown to have
self-cross-linked, enabling interesting tensile properties with irradiation. Statistical analysis validated
the results obtained and also showed that adequate mixing of fibres and matrix had taken place at 95%
confidence level. Hybridization and subsequent irradiation increased the tensile strength and modulus
of HDPE up to 31 and 185%, respectively, at about 100kGy. Meanwhile, SEM was used to view the
interaction between the fibres and matrix.
We used 40 ± 5 nm gold nanoparticles (GNPs) as colorimetric sensor to visually detect swine-specific conserved sequence and nucleotide mismatch in PCR-amplified and non-amplified mitochondrial DNA mixtures to authenticate species. Colloidal GNPs changed color from pinkish-red to gray-purple in 2 mM PBS. Visually observed results were clearly reflected by the dramatic reduction of surface plasmon resonance peak at 530 nm and the appearance of new features in the 620-800 nm regions in their absorption spectra. The particles were stabilized against salt-induced aggregation upon the adsorption of single-stranded DNA. The PCR products, without any additional processing, were hybridized with a 17-base probe prior to exposure to GNPs. At a critical annealing temperature (55 °C) that differentiated matched and mismatched base pairing, the probe was hybridized to pig PCR product and dehybridized from the deer product. The dehybridized probe stuck to GNPs to prevent them from salt-induced aggregation and retained their characteristic red color. Hybridization of a 27-nucleotide probe to swine mitochondrial DNA identified them in pork-venison, pork-shad and venison-shad binary admixtures, eliminating the need of PCR amplification. Thus the assay was applied to authenticate species both in PCR-amplified and non-amplified heterogeneous biological samples. The results were determined visually and validated by absorption spectroscopy. The entire assay (hybridization plus visual detection) was performed in less than 10 min. The LOD (for genomic DNA) of the assay was 6 µg ml(-1) swine DNA in mixed meat samples. We believe the assay can be applied for species assignment in food analysis, mismatch detection in genetic screening and homology studies between closely related species.
In this study, we report for the first time the development of a dry-reagent-based nucleic acid-sensing platform by combining a thermostabilised linear-after-the-exponential (LATE)-PCR assay with a one-step, hybridisation-based nucleic acid lateral flow biosensor. The nucleic acid-sensing platform was designed to overcome the need for stringent temperature control during transportation or storage of reagents and reduces the dependency on skilled personnel by decreasing the overall assay complexity and hands-on time. The platform was developed using toxigenic Vibrio cholerae as the model organism due to the bacterium's propensity to cause epidemic and pandemic cholera. The biosensor generates result which can be visualised with the naked eyes and the limit of detection was found to be 1pg of pure genomic DNA and 10CFU/ml of toxigenic V. cholerae. The dry-reagent-based nucleic acid-sensing platform was challenged with 95 toxigenic V. cholerae, 7 non-toxigenic V. cholerae and 66 other bacterial strains in spiked stool sample and complete agreement was observed when the results were compared to that of monosialoganglioside (GM1)-ELISA. Heat-stability of the thermostabilised LATE-PCR reaction mixes at different storage temperatures (4-56°C) was investigated for up to 90days. The dry-reagent-based genosensing platform with ready-to-use assay components provides an alternative method for sequence-specific detection of nucleic acid without any cold chain restriction that is associated with conventional molecular amplification techniques.
A novel electrochemical DNA biosensor for ultrasensitive and selective quantitation of Escherichia coli DNA based on aminated hollow silica spheres (HSiSs) has been successfully developed. The HSiSs were synthesized with facile sonication and heating techniques. The HSiSs have an inner and an outer surface for DNA immobilization sites after they have been functionalized with 3-aminopropyltriethoxysilane. From field emission scanning electron microscopy images, the presence of pores was confirmed in the functionalized HSiSs. Furthermore, Brunauer-Emmett-Teller (BET) analysis indicated that the HSiSs have four times more surface area than silica spheres that have no pores. These aminated HSiSs were deposited onto a screen-printed carbon paste electrode containing a layer of gold nanoparticles (AuNPs) to form a AuNP/HSiS hybrid sensor membrane matrix. Aminated DNA probes were grafted onto the AuNP/HSiS-modified screen-printed electrode via imine covalent bonds with use of glutaraldehyde cross-linker. The DNA hybridization reaction was studied by differential pulse voltammetry using an anthraquinone redox intercalator as the electroactive DNA hybridization label. The DNA biosensor demonstrated a linear response over a wide target sequence concentration range of 1.0×10-12-1.0×10-2 μM, with a low detection limit of 8.17×10-14 μM (R2 = 0.99). The improved performance of the DNA biosensor appeared to be due to the hollow structure and rough surface morphology of the hollow silica particles, which greatly increased the total binding surface area for high DNA loading capacity. The HSiSs also facilitated molecule diffusion through the silica hollow structure, and substantially improved the overall DNA hybridization assay. Graphical abstract Step-by-step DNA biosensor fabrication based on aminated hollow silica spheres.
The taxonomic status of a Nocardiopsis strain, designated H13T, isolated from a high altitude Atacama Desert soil, was established by using a polyphasic approach. The strain was found to have chemotaxonomic, cultural and morphological characteristics consistent with its classification within the genus Nocardiopsis and formed a well-supported clade in the Nocardiopsis phylogenomic tree together with the type strains of Nocardiopsis alborubida, Nocardiopsis dassonvillei and Nocardiopsis synnematoformans. Strain H13T was distinguished from its closest relatives by low average nucleotide identity (93.2-94.9 %) and in silico DNA-DNA hybridization (52.5-62.4 %) values calculated from draft genome assemblies and by a range of phenotypic properties. On the basis of these results, it is proposed that the isolate be assigned to the genus Nocardiopsis as Nocardiopsis deserti sp. nov. with isolate H13T (=CGMCC 4.7585T=KCTC 49249T) as the type strain.
A taxonomic study was conducted to clarify the relationships of two bacterial populations belonging to the genus Weissella. A total of 39 strains originating mainly from Malaysian foods (22 strains) and clinical samples from humans (9 strains) and animals (6 strains) were analysed using a polyphasic taxonomic approach. The methods included classical phenotyping, whole-cell protein electrophoresis, 16S and 23S rDNA RFLP (ribotyping), determination of 16S rDNA sequence homologies and DNA-DNA reassociation levels. Based on the results, the strains were considered to represent two different species, Weissella confusa and a novel Weissella species, for which the name Weissella cibaria sp. nov. is proposed. Weisella confusa possessed the highest 16S rDNA sequence similarity to Weisella cibaria, but the DNA-DNA reassociation experiment showed hybridization levels below 49% between the strains studied. The numerical analyses of Weisella confusa and Weisella cibaria strains did not reveal any specific clustering with respect to the origin of the strains. Based on whole-cell protein electrophoresis, and ClaI and HindIII ribotyping patterns, food and clinical isolates were randomly located in the two species-specific clusters obtained.
Thirteen strains of dengue type 1 were isolated from the lymphocyte fractions of 69 acute phase blood samples collected at Thursday Island Hospital during 1981 and 1982. One further strain of type 1 was isolated from 7 blood samples despatched by air from Cairns Base Hospital during 1982. Four of these Australian isolates representing the beginning, middle, and end of the epidemic were examined by restriction enzyme mapping and were found to be identical for the nine restriction enzymes used. The maps differed from those derived from two Malaysian dengue type 1 strains isolated during the epidemic of 1981-82 in that country. This suggests reliance on serological typing to establish global circulation patterns of epidemic dengue is insufficient and that more specific methods such as genome mapping are useful.
Nanotechnology is playing a major role in the field of medical diagnosis, in particular with the biosensor and bioimaging. It improves the performance of the desired system dramatically by displaying higher selectivity and sensitivity. Carbon nanomaterial, gold nanostructure, magnetite nanoparticle, and silica substrate are the most popular nanomaterials greatly contributed to make the affordable and effective biosensor at low-cost. This research work is introducing a new sensing strategy with graphene oxide-constructed triangular electrodes to diagnose Alzheimer's disease (AD). MicroRNA-137 (miRNA-137) was found as a suitable biomarker for AD, and the sensing method was established here to detect miRNA-137 on the complementary sequence. To enhance the immobilization of capture miRNA-137, gold nanostar (GNS) was conjugated with capture miRNA and immobilized on the GO-modified surface through an amine linker. This immobilization process enhanced the hybridization of the target and reaches the detection limit at 10 fM with the sensitivity of 1 fM on the linear curve with a regression coefficient of 0.9038. Further control sequences of miRNA-21 and single and triple base mismatched miRNA-137 did not show a significant response in current changes, indicating the specific miRNA-137 detection for diagnosing AD.
While lab-scale synthesis of trigonal-Zr2N2S, hexagonal-Zr2N2S and hexagonal-Zr2N2Se has been reported, meaningful data on the photophysical properties of IV-nitride chalcogenides in general are scarcely available. The first-principles calculations and genetic algorithm modeling in our work reveal the existence of remarkably stable, indirect gap trigonal-Zr2N2Se and trigonal-Hf2N2Se phases, which progress to direct gap, monoclinic materials in monolayer form. These structures display the desired optoelectronic properties, such as exceptionally high visible-UV absorption spectra (105-106 cm-1) and exciton binding energy below 0.02 eV. Strong hybridization between the Zr-d, N-p and Se-p orbitals is accounted for by the polysilicon comparable Vickers hardness (10.64-12.77 GPa), while retaining ductile nature.
A novel aerobic, non-motile, Gram-positive, rod-shaped actinobacterium, designated YIM 100951(T), was isolated from the faeces of civets (Viverra zibetha) living in the National Nature Protect Region in Selangor, Malaysia. Strain YIM 100951(T) shows high similarities with Microbacterium barkeri DSM 20145(T) (97.6 %), Microbacterium oryzae MB10(T) (97.3 %), Microbacterium lemovicicum ViU22(T) (97.1 %) and Microbacterium indicum BBH6(T) (97.0 %) based on their 16S rRNA genes. However, phylogenetic analysis showed that strain YIM 100951(T) formed a clade with Microbacterium halotolerans YIM 70130(T) (96.7 %), Microbacterium populi 10-107-8(T) (96.7 %) and Microbacterium sediminis YLB-01(T) (96.9 %). DNA-DNA hybridization was carried out between strains YIM 100951(T) and M. barkeri DSM 20145(T), the result showed a value of 23.2 ± 4.5 %. In addition, some of the physiological, biochemical and chemotaxonomic characteristics of strain YIM 100951(T) are different from the closely related strains. Thus, we suggest that strain YIM 100951(T) represents a novel species of the genus Microbacterium, for which the name Microbacterium gilvum sp. nov. is proposed. The type strain is YIM 100951(T) (=DSM 26235(T) = CCTCC AB 2012971(T)).
Deoxyribonucleic acid (DNA) of twenty chronic myeloid leukemia (CML) and thirty acute lymphoblastic leukemia (ALL) patients were analysed by Southern hybridization. The DNA was digested with BglII and hybridized with a 4.5-kilobase (kb) ph1/bcr-3 DNA probe. All the 20 CML patients showed gene rearrangement within a 5.8-kb segment (the major breakpoint cluster region, M-bcr) of the breakpoint cluster region (bcr) gene of chromosome 22, indicating the presence of the Philadelphia chromosome. M-bcr rearrangement at the bcr gene of chromosome twenty-two was not detected in all the thirty ALL patients (nine adults and twenty-one children) and two normal controls.
Two Gram-stain-negative, non-fermentative bacterial strains, designated 11-0202(T) and 11-0607, were isolated from soil in South Korea, and four others, LUH 13522, LUH 8638, LUH 10268 and LUH 10288, were isolated from a beet field in Germany, soil in the Netherlands, and sediment of integrated fish farms in Malaysia and Thailand, respectively. Based on 16S rRNA, rpoB and gyrB gene sequences, they are considered to represent a novel species of the genus Acinetobacter. Their 16S rRNA gene sequences showed greatest pairwise similarity to Acinetobacter beijerinckii NIPH 838(T) (97.9-98.4 %). They shared highest rpoB and gyrB gene sequence similarity with Acinetobacter johnsonii DSM 6963(T) and Acinetobacter bouvetii 4B02(T) (85.4-87.6 and 78.1-82.7 %, respectively). Strain 11-0202(T) displayed low DNA-DNA reassociation values (<40 %) with the most closely related species of the genus Acinetobacter. The six strains utilized azelate, 2,3-butanediol, ethanol and dl-lactate as sole carbon sources. Cellular fatty acid analyses showed similarities to profiles of related species of the genus Acinetobacter: summed feature 3 (C16 : 1ω7c, C16 : 1ω6c; 24.3-27.2 %), C18 : 1ω9c (19.9-22.1 %), C16 : 0 (15.2-22.0 %) and C12 : 0 (9.2-14.2 %). On the basis of the current findings, it is concluded that the six strains represent a novel species, for which the name Acinetobacter kookii sp. nov. is proposed. The type strain is 11-0202(T) ( = KCTC 32033(T) = JCM 18512(T)).
A search for the natural host of Nipah virus has led to the isolation of a previously unknown member of the family Paramyxoviridae. Tioman virus (TiV) was isolated from the urine of fruit bats (Pteropus hypomelanus) found on the island of the same name off the eastern coast of peninsular Malaysia. An electron microscopic study of TiV-infected cells revealed spherical and pleomorphic-enveloped viral particles (100--500 nm in size) with a single fringe of embedded peplomers. Virus morphogenesis occurred at the plasma membrane of infected cells and morphological features of negative-stained ribonucleoprotein complexes were compatible with that of viruses in the family Paramyxoviridae. Serological studies revealed no cross-reactivity with antibodies against a number of known Paramyxoviridae members except for the newly described Menangle virus (MenV), isolated in Australia in 1997. Failure of PCR amplification using MenV-specific primers suggested that this new virus is related to but different from MenV. For molecular characterization of the virus, a cDNA subtraction strategy was employed to isolate virus-specific cDNA from virus-infected cells. Complete gene sequences for the nucleocapsid protein (N) and phosphoprotein (P/V) have been determined and recombinant N and V proteins produced in baculovirus. The recombinant N and V proteins reacted with porcine anti-MenV sera in Western blot, confirming the serological cross-reactivity observed during initial virus characterization. The lack of a C protein-coding region in the P/V gene, the creation of P mRNA by insertion of 2-G residues, and the results of phylogenetic analyses all indicated that TiV is a novel member of the genus Rubulavirus.
In this study, bacterial strains Ha5T, Ta1, and Jb2 were isolated from different colonies of weaver ant Oecophylla smaragdina. They were identified as bacterial symbionts of the ant belonging to family Acetobacteraceae and were distinguished as different strains based on distinctive random-amplified polymorphic DNA (RAPD) fingerprints. Cells of these bacterial strains were Gram-negative, rod-shaped, aerobic, non-motile, catalase-positive and oxidase-negative. They were able to grow at 15-37°C (optimum, 28-30°C) and in the presence of 0-1.5% (w/v) NaCl (optimum 0%). Their predominant cellular fatty acids were C18:1ω7c, C16:0, C19:0ω8c cyclo, C14:0, and C16:0 2-OH. Strains Ha5T, Ta1, and Jb2 shared highest 16S rRNA gene sequence similarity (94.56-94.63%) with Neokomagataea tanensis NBRC106556T of family Acetobacteraceae. Both 16S rRNA gene sequence-based phylogenetic analysis and core gene-based phylogenomic analysis placed them in a distinct lineage in family Acetobacteraceae. These bacterial strains shared higher than species level thresholds in multiple overall genome-relatedness indices which indicated that they belonged to the same species. In addition, they did not belong to any of the current taxa of Acetobacteraceae as they had low pairwise average nucleotide identity (< 71%), in silico DNA-DNA hybridization (< 38%) and average amino acid identity (< 67%) values with all the type members of the family. Based on these results, bacterial strains Ha5T, Ta1, and Jb2 represent a novel species of a novel genus in family Acetobacteaceae, for which we propose the name Oecophyllibacter saccharovorans gen. nov. sp. nov., and strain Ha5T as the type strain.
Three strains of Gram-staining-positive, coccus-shaped, lactic acid bacteria, designated as HibF3T, HibF2 and HibF5 were isolated from fresh flowers of hibiscus, and a fourth, DF1T, was isolated from fresh flowers of durian tree, in Penang, Malaysia. Taxonomic characterisation was performed by polyphasic analysis. Sequence similarities of the 16S rRNA gene and the housekeeping rpoA and pheS genes of these strains with their closely-related lactococcal and streptococcal relatives were 92-94, 78 and 81 %, respectively. The results of phylogenetic analysis indicated that strains DF1T, HibF2, HibF5 and HibF3T were clustered together but were clearly separated from species of the genera Streptococcus and Lactococcus, indicating that they represent members of a novel genus of the family Streptococcaceae. Calculation of average nucleotide identity (ANI) values between the genomes of DF1T and HibF3T yielded values of 92.50-92.93 %. ANI values below the cut-off value and distinctive chemotaxonomic characteristics supported the hypothesis that these strains represented two novel species. Major cellular fatty acids in DF1T, HibF2 and HibF5 were C18 : 1ω7c and C16 : 0, while C12 : 0 and C14 : 0 were also dominant, in addition to C18 : 1ω7c and C16 : 0, in HibF3T. A novel genus is proposed with the name Floricoccus gen. nov. which consists of two species, Floricoccus tropicus sp. nov as the type species, and Floricoccus penangensis sp. nov. The respective type strains are DF1T (=LMG 29833T=JCM 31733T) and HibF3T (=LMG 29831T=DSM 31735T).
There are four species of mud crabs within the genus Scylla, and most of them live sympatrically in the equatorial region. Apart from a report in Japan about the finding of a natural Scylla hybrid more than a decade ago after the division of genus Scylla into four species by Keenan, Davie & Mann (1998), no subsequent sighting was found. Thus, this study investigates the possible natural occurrence of potential hybridization among Scylla species in the wild. A total of 76,211 individuals from mud crab landing sites around the Malacca Straits, South China Sea and Sulu Sea were screened. In addition to the four-purebred species, four groups (SH 1, n = 2, 627; SH 2, n = 136; SH 3, n = 1; SH 4, n = 2) with intermediate characteristics were found, mostly at Sulu Sea. Discriminant Function Analysis revealed that all Scylla species, including SH 1 - 4, are distinguishable via their morphometric ratios. The most powerful discriminant ratios for each character and the top five discriminant ratios of males and females were suggested. The carapace width of SH 1 males and females were significantly smaller than pure species. Based on the discriminant ratios and the description of morphological characters, we hypothesize that the additional four groups of Scylla with intermediate characteristics could be presumed hybrids. Future work at the molecular level is urgently needed to validate this postulate.
One of the causative agents of lympahtic filariasis is the nematode parasite Brugia malayi that requires a competent mosquito vector for its development and transmission. Armigeres subalbatus mosquitoes rapidly destroy invading B. malayi microfilariae via a defense response known as melanotic encapsulation. We have constructed a genetic linkage map for this mosquito species using RFLP markers from Aedes aegypti. This heterologous approach was possible because of the conserved nature of the coding sequences used as markers and provided an experimental framework to evaluate the hypothesis that linkage and gene order are conserved between these mosquito species. Of the 56 Ae. aegypti markers tested, 77% hybridize to genomic DNA digests of Ar. subalbatus under stringent conditions, with 53% of these demonstrating strain-specific polymorphisms. Twenty-six Ae. aegypti markers have been mapped using an F2- segregating Ar. subalbatus population derived from a cross of strains originating in Japan and Malaysia. Linear order of these marker loci is highly conserved between the two species. Only 1 of these markers, LF92, was not linked in the manner predicted by the Ae. aegypti map. In addition, the autosomal sex-determination locus that occurs in linkage group 1 in Ae. aegypti resides in group 3 in Ar. subalbatus. The Ar. subalbatus map provides a basic genetic context that can be utilized in further genetic studies to clarify the genetic basis of parasite resistance in this mosquito and is a necessary precursor to the identification of genome regions that carry genes that determine the encapsulation phenotype. [The composite map and sequence database information for Ae. aegypti markers can be retrieved directly from the Ae. aegypti Genome Database through the World Wide Web: http://klab.agsci.colostate.edu.]
Beta (β)- thalassaemia is a public health problem in Malaysia. The carrier rate is estimated to be 4.5% by micro-mapping studies particularly among Malays who comprise 53.5% of the population in Malaysia. The common diagnostic method in Malaysia for mutation detection is by amplification refractory mutation system (ARMS). It allows single mutation detection in each reaction but is labour intensive and time consuming when many mutations need to be identified. The purpose of this study was to develop a diagnostic tool for effective mutation detection of beta thalassaemia in Malay patients and compare its efficacy with ARMS-PCR, the current method in use. Methods: Reverse dot blot hybridization (RDBH) technique was incorporated in the development of two strip assays [RDBH-Strip M(6) and RDBH-Strip C(6)] to identify common beta thalassaemia mutations in the Malays. The panels of selected mutations were based on the mutation frequencies in Malaysia reported in previous studies. RDBH-Strip M(6) was applied as step 1 and RDBH-Strip C(6) was applied as step 2 for unidentified mutations. The strips were validated with the gold standard method, ARMS- PCR. Results: One hundred and thirty seven Malay patients with 274 alleles were studied. In Step 1 mutation detection, 238 alleles (86.9%) were identified in 119 of patients by RDBH-Strip M(6). Step 2 resulted in a further detection of 20 alleles in another 10 patients by RDBH-Strip C(6). The combination of both strips resulted in the identification of 258 alleles in 129 (94.6%) of 137 Malay patients. The strip assays were 100% sensitive and specific when compared with ARMS-PCR method. Conclusion: Two strip assays utilising the RDBH technique were developed to identify common β-thalassaemia mutations in Malays. The RDBH Strip M(6) identified 86.9% of the mutations and the RDBH-Strip C (6) detected further 7.3% alleles. This two step strategy was found to be rapid and cost effective for the direct diagnosis of β-thalassaemia mutations in the Malays. The remaining unidentified mutations would require DNA sequencing. It can serve as a specific molecular diagnostic tool for effective diagnosis of
β-thalassaemia mutations in this ethnic group.
A Gram-negative, filamentous aerobic bacterium designated as strain Mgbs1T was isolated on 12 April 2017 from the subsurface soil and leaf litter substrate at the base of a Koompassia malaccensis tree in a tropical peat swamp forest in the northern regions of the state of Selangor, Malaysia (3° 39' 04.7' N 101° 17' 43.7'' E). Phylogenetic analyses based on the full 16S rRNA sequence revealed that strain Mgbs1T belongs to the genus Chitinophaga with the greatest sequence similarity to Chitinophaga terrae KP01T (97.65 %), Chitinophaga jiangningensis DSM27406T (97.58 %), and Chitinophaga dinghuensis DHOC24T (97.17 %). The major fatty acids of strain Mgbs1T (>10 %) are iso-C15 : 0, C16 : 1 ω5c and iso-C17 : 0 3-OH while the predominant respiratory quinone is menaquinone-7. Strain Mgbs1T has a complete genome size of 8.03 Mb, with a G+C content of 48.5 mol%. The DNA-DNA hybridization (DDH) score between strain Mgbs1T and C. jiangningensis DSM27406T was 15.9 %, while in silico DDH values of strain Mgbs1T against C. dinghuensis DHOC24T and C. terrae KP01T were 20.0 and 19.10% respectively. Concurrently, Average Nucleotide Identity (ANI) scores between strain Mgbs1T against all three reference strains are 73.2 %. Based on the phenotypic, chemotaxonomic, and phylogenetic consensus, strain Mgbs1T represents a novel species of the genus Chitinophaga, for which the name Chitinophaga extrema sp. nov. is proposed (=DSM 108835T=JCM 33276T).
The evolutionary history of the wolf-like canids of the genus Canis has been heavily debated, especially regarding the number of distinct species and their relationships at the population and species level [1-6]. We assembled a dataset of 48 resequenced genomes spanning all members of the genus Canis except the black-backed and side-striped jackals, encompassing the global diversity of seven extant canid lineages. This includes eight new genomes, including the first resequenced Ethiopian wolf (Canis simensis), one dhole (Cuon alpinus), two East African hunting dogs (Lycaon pictus), two Eurasian golden jackals (Canis aureus), and two Middle Eastern gray wolves (Canis lupus). The relationships between the Ethiopian wolf, African golden wolf, and golden jackal were resolved. We highlight the role of interspecific hybridization in the evolution of this charismatic group. Specifically, we find gene flow between the ancestors of the dhole and African hunting dog and admixture between the gray wolf, coyote (Canis latrans), golden jackal, and African golden wolf. Additionally, we report gene flow from gray and Ethiopian wolves to the African golden wolf, suggesting that the African golden wolf originated through hybridization between these species. Finally, we hypothesize that coyotes and gray wolves carry genetic material derived from a "ghost" basal canid lineage.