South African rabies viruses originating from dogs and jackals (canid viruses) are closely related and highlight cross-species transmission events between the two canine species. Rabies due to the canid lyssavirus variant is a significant public health matter in this country. The complete coding sequences of 23 canid lyssaviruses from South Africa are reported here.
ABSTRACT
South African rabies viruses originating from dogs and jackals (canid viruses) are closely related and highlight cross-species transmission events between the two canine species. Rabies due to the canid lyssavirus variant is a significant public health matter in this country. The complete coding sequences of 23 canid lyssaviruses from South Africa are reported here.
ANNOUNCEMENT
Rabies virus (RABV) is the prototype species of the Lyssavirus genus (family Rhabdoviridae, order Mononegavirales) (1) and causes encephalitis in all warm-blooded vertebrates. In South Africa, RABV infects both domestic and wild carnivore species (2). The domestic dog is the main vector species responsible for the transmission of rabies to humans (3), resulting in at least 59,000 human deaths annually, and the majority of these (≥95%) are recorded in Africa and Asia (2).
Specimens were collected by state veterinarians from animals in northern South Africa (Limpopo and Mpumalanga provinces) showing typical signs of central nervous system infection, preserved in glycerol-saline, and transported to the laboratory (see Table 1 for epidemiological information on the samples). The composite brain tissues were then subjected to a direct fluorescent antibody test for lyssavirus antigen (4). Total viral RNAs were extracted from the original lyssavirus-infected brain tissues and prepared for next-generation sequencing (NGS) using the HiSeq (Illumina, San Diego, CA, USA) platform. Briefly, the TRIzol-extracted viral RNA was depleted of host genomic DNA and rRNA using the on-column DNase treatment in the RNeasy Plus kit (Qiagen) and Terminator 5′-phosphate-dependent exonuclease (Epicentre Biotechnologies), as described previously (5–7). Double-stranded (ds) cDNA was synthesized from 50 ng RNA using a random cDNA synthesis system kit (Roche, Basel, Switzerland) according to the manufacturer’s instructions. The ds cDNA was purified using AMPure XP magnetic beads (Beckman Coulter, Brea, CA, USA), and 1 ng was used for the Nextera XT DNA sample preparation kit (Illumina). A sequencing library was prepared for each sample according to the manufacturer’s instructions and sequenced on an Illumina HiSeq with 2 × 125-bp paired-end reads following standard Illumina protocols, resulting in between 0.002 and 5.6 Gb data per isolate (Table 1). For each of the isolates, contigs were generated through both de novo assembling of all the reads and mapping of the same reads to existing rabies genomes (GenBank accession numbers KT336433 to KT336436) using default settings in CLC Genomics Workbench v.9 (Qiagen). A single consensus sequence was produced for each sample from an assembly of all the contigs previously generated. The newly obtained consensus sequence was used as a template to map all the original reads in order to estimate the average coverage and possible variants (see Table 1). The genetic organization of the full genomes of the South African RABVs was consistent with that of other previously characterized lyssaviruses (5–7). The sequence length and percent G+C content of each of the samples are listed in Table 1. Each sequence contains the complete coding regions of the nucleoprotein (N), glycoprotein (G), matrix protein (M), phosphoprotein (P), and the RNA-dependent polymerase (L) found in rabies viruses. Genetic analysis of these canid RABVs demonstrated that they are of the Africa 1-b lineage and have a high degree of sequence similarity (≥96.5%) (mean distance, 0.023; standard error [SE], 0.001), irrespective of their host species and locality of origin (1, 7–9).
TABLE 1.
Epidemiological information of the canine viruses sequenced in this study
| Virus no. | Lab ref no.a | Yr of submission | Species from which sample originated | Size of raw data (GB) | No. (%) of reads mapped to new consensus sequence | Sequence length (bp) | % G+C content | BioProject accession no. | GenBank accession no. |
|---|---|---|---|---|---|---|---|---|---|
| 1 | 125/15 | 2015 | Canis familiaris | 2.62 | 159,506 (0.76) | 11,923 | 45.4 | SRR12012256 | MT454631 |
| 2 | 155/15 | 2015 | Proteles cristatus | 2.30 | 7,365 (0.04) | 11,902 | 45.5 | SRR12012255 | MT454632 |
| 3 | 361/15 | 2015 | Proteles cristatus | 2.73 | 4,365 (0.02) | 11,908 | 45.6 | SRR12012244 | MT454633 |
| 4 | 471/15 | 2015 | Canis familiaris | 2.65 | 72,025 (0.34) | 11,923 | 45.4 | SRR12012240 | MT454634 |
| 5 | 682/15 | 2015 | Canis familiaris | 5.63 | 31,529 (0.07) | 11,923 | 45.3 | SRR12012239 | MT454635 |
| 6 | 516/16 | 2016 | Canis familiaris | 1.73 | 217,555 (1.57) | 11,923 | 45.3 | SRR12012238 | MT454636 |
| 7 | 583/16 | 2016 | Canis familiaris | 2.53 | 309,360 (1.53) | 11,923 | 45.4 | SRR12012237 | MT454637 |
| 8 | 631/16 | 2016 | Otocyon megalotis | 4.16 | 6,656 (0.02) | 11,923 | 45.4 | SRR12012236 | MT454638 |
| 9 | 635/16 | 2016 | Canis familiaris | 1.39 | 166,479 (1.5) | 11,923 | 45.3 | SRR12012235 | MT454639 |
| 10 | 676/16 | 2016 | Otocyon megalotis | 3.53 | 5,640 (0.02) | 11,923 | 45.4 | SRR12012234 | MT454640 |
| 11 | 690/16 | 2016 | Canis familiaris | 0.002 | 37.321 (0.22) | 11,923 | 45.3 | SRR12012254 | MT454641 |
| 12 | 725/16 | 2016 | Proteles cristatus | 2.90 | 6,971 (0.03) | 11,923 | 45.5 | SRR12012253 | MT454642 |
| 13 | 269/17 | 2017 | Canis familiaris | 4.40 | 218,019 (0.62) | 11,922 | 45.6 | SRR12012252 | MT454643 |
| 14 | 400/17 | 2017 | Canis familiaris | 1.94 | 37,211 (0.24) | 11,922 | 45.6 | SRR12012251 | MT454645 |
| 15 | 454/17 | 2017 | Canis mesomelas | 2.80 | 123,359 (0.55) | 11,922 | 45.6 | SRR12012250 | MT454646 |
| 16 | 460/17 | 2017 | Canis mesomelas | 2.95 | 21,259 (0.09) | 11,922 | 45.6 | SRR12012249 | MT454647 |
| 17 | 466/17 | 2017 | Canis mesomelas | 4.43 | 10,620 (0.03) | 11,923 | 45.3 | SRR12012248 | MT454648 |
| 18 | 474/17 | 2017 | Canis mesomelas | 1.58 | 26,616 (0.21) | 11,922 | 45.6 | SRR12012247 | MT454649 |
| 19 | 477/17 | 2017 | Otocyon megalotis | 2.14 | 92,643 (0.54) | 11,922 | 45.6 | SRR12012246 | MT454650 |
| 20 | 480/17 | 2017 | Canis mesomelas | 2.90 | 53,270 (0.23) | 11,922 | 45.6 | SRR12012245 | MT454651 |
| 21 | 483/17 | 2017 | Canis mesomelas | 2.40 | 7,664 (0.04) | 11,923 | 45.3 | SRR12012243 | MT454652 |
| 22 | 502/17 | 2017 | Canis mesomelas | 2.26 | 81,394 (0.45) | 11,923 | 45.3 | SRR12012242 | MT454653 |
| 23 | 503/17 | 2017 | Canis mesomelas | 2.21 | 70,808 (0.4) | 11,923 | 45.3 | SRR12012241 | MT454654 |
Sample submitted to Agricultural Research Council—Onderstepoort Veterinary Institute (ARC-OVI) for laboratory confirmation of RABV.
Data availability.
The complete coding sequences were submitted to GenBank and are available under accession numbers MT454631 through MT454654 (Table 1) and BioProject accession number PRJNA638742.
ACKNOWLEDGMENTS
This study was partly supported with funding from the National Research Foundation (NRF) of South Africa (NRF unique grant number 114679, ARC grant number P10000130) and the EU Framework Horizon 2020 Innovation Grant, European Virus Archive (EVAg, grant number 653316).
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Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
The complete coding sequences were submitted to GenBank and are available under accession numbers MT454631 through MT454654 (Table 1) and BioProject accession number PRJNA638742.