Circular replication-associated protein encoding DNA viruses identified in the faecal matter of various animals in New Zealand.

In recent years, innovations in molecular techniques and sequencing technologies have resulted in a rapid expansion in the number of known viral sequences, in particular those with circular replication-associated protein (Rep)-encoding single-stranded (CRESS) DNA genomes. CRESS DNA viruses are present in the virome of many ecosystems and are known to infect a wide range of organisms. A large number of the recently identified CRESS DNA viruses cannot be classified into any known viral families, indicating that the current view of CRESS DNA viral sequence space is greatly underestimated. Animal faecal matter has proven to be a particularly useful source for sampling CRESS DNA viruses in an ecosystem, as it is cost-effective and non-invasive. In this study a viral metagenomic approach was used to explore the diversity of CRESS DNA viruses present in the faeces of domesticated and wild animals in New Zealand. Thirty-eight complete CRESS DNA viral genomes and two circular molecules (that may be defective molecules or single components of multicomponent genomes) were identified from forty-nine individual animal faecal samples. Based on shared genome organisations and sequence similarities, eighteen of the isolates were classified as gemycircularviruses and twelve isolates were classified as smacoviruses. The remaining eight isolates lack significant sequence similarity with any members of known CRESS DNA virus groups. This research adds significantly to our knowledge of CRESS DNA viral diversity in New Zealand, emphasising the prevalence of CRESS DNA viruses in nature, and reinforcing the suggestion that a large proportion of CRESS DNA viruses are yet to be identified.

Novel myco-like DNA viruses discovered in the faecal matter of various animals.

A wide variety of novel single-stranded DNA (ssDNA) viruses have been found in faecal matter of chimpanzees, cows, rodents, bats, badgers, foxes and pigs over the last few years. Using a combination of rolling circle amplification coupled with restriction enzyme digests based approach as well as a next generation sequencing informed approach, we have recovered fourteen full genomes of ssDNA viruses which exhibit genomic features described for members of the recently proposed gemycircularvirus group from a wide variety of mammal and bird faecal samples across New Zealand. The fourteen novel ssDNA viruses (2122-2290nt) encode two major proteins, a replication associated protein (Rep) and a capsid protein (Cp) which are bi-directionally transcribed. Interestingly, the Rep of these novel viruses are similar to gemycircularviruses detected in insects, cassava leaves, and badger faecal matter, the novel viruses share sequence similarities with the mycovirus sclerotinia sclerotiorum hypovirulence-associated DNA virus 1 (SsHADV-1) and Rep-like sequences found in fungal genomes. Pairwise sequence similarities between the 14 novel genomes with other related viral isolates (gemycircularviruses) indicated that they share greater than 55.8% genome-wide identity. Additionally, they share between 55% and 59% pairwise identity with putative novel ssDNA virus genomes recently isolated from sewage baminivirus, niminivirus and nephavirus. Based on the similarities to SsHADV-1 and Rep-like sequences found in fungal genomes, these novel gemycircularviruses may infect fungi.

Identification of a Novel Circular DNA Virus in New Zealand Fur Seal (Arctocephalus forsteri) Fecal Matter.

Fur seal feces-associated circular DNA virus (FSfaCV) is a novel virus isolated from the fecal matter of New Zealand fur seals. FSfaCV has two main open reading frames in its 2,925-nucleotide (nt) genome. The replication-associated protein (Rep) of FSfaCV has similarity to Rep-like sequences in the Giardia intestinalis genome.

Novel ssDNA viruses discovered in yellow-crowned parakeet (Cyanoramphus auriceps) nesting material.

During routine monitoring of yellow-crowned parakeets in the Poulter Valley of the South Island of New Zealand, a dead parakeet chick was discovered in a nest. Known parrot-infecting viruses, such as beak and feather disease virus (BFDV), avian polyomavirus (APV), and parrot hepatitis B virus (PHBV), were not detected in the nesting material. However, we recovered two novel single-stranded DNA viruses (ssDNA), CynNCXV (2308 nt) and CynNCKV (2087 nt), which have genome architectures similar to those of circoviruses, characterised by circular genomes with two large bidirectional open reading frames (ORFs). Both contain a stem-loop element with a conserved nonanucleotide motif, known to be required for rolling-circle replication. The full genomes had no BLASTn similarity to known ssDNA viruses. However, in both genomes the larger ORFs have BLAST similarity to known replication-associated proteins (Reps). CynNCKV has 30 % similarity to picobiliphyte nano-like virus (Picobiliphyte M5584-5) with 66-88 % coverage (e-value of 5×10(-33)), whereas CynNCXV has 33 % similarity to rodent stool-associated virus (RodSCV M-45) with 92-94 % coverage (e-value of 5 × 10(-31)). Found within these ORFs were the rolling-circle replication motifs I, II, III and the helicase motifs Walker A and Walker B. Maximum-likelihood phylogenetic analysis of the Reps reveals that these are two novel ssDNA viruses. At this point, we are unable to attribute the death of the parakeet to these two new novel ssDNA viruses.

Discovery of a novel circular single-stranded DNA virus from porcine faeces.

A large number of novel single-stranded DNA (ssDNA) viruses have been characterised from various environmental sources in the last 5 years. The bulk of these have been from faecal sources, and faecal sampling is an ideal non-invasive pathogen sampling method. We characterised a novel ssDNA from a porcine faecal sample from Cass Basin of the South Island of New Zealand. The novel viral genome has two large open reading frames (ORFs), which are bidirectionally transcribed and separated by intergenic regions. The largest ORF has some degree of similarity (<30 %) to the putative capsid protein of chimpanzee stool-associated circular ssDNA virus (ChiSCV) and pig stool-associated single-stranded DNA virus (PigSCV), whereas the second-largest ORF has high similarity to the putative replication-associated protein (Rep) of ChiSCV (~50 %) and bovine stool-associated circular DNA virus (BoSCV; ~30 %). Based on genome architecture, location of putative stem-loop like elements, and maximum-likelihood phylogenetic analysis of the gene encoding the Rep protein, the novel isolate belongs to the same family of ssDNA viruses as ChiSCV and BoSCV.