Journal Articles
Permanent URI for this collectionhttps://mro.massey.ac.nz/handle/10179/7915
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Item Correction: Development of a non-infectious control for viral hemorrhagic fever PCR assays(PLOS, 2025-07-09) Knox MA; Bromhead C; Hayman DTSThe Funding statement for this article is incorrect. The correct Funding statement is as follows: EBO-SURSY project funded by the European Union via the World Organisation for Animal Health (Grant 3000034275; OIE Laboratory (or Collaborating Centre) Twinning Project: Enhancing capacity for early detection of viral haemorrhagic fevers in Liberia through epidemiological and laboratory training).Item First Detection and Genetic Characterization of Felis catus Papillomavirus Type 11, the First Treisetapapillomavirus Type to Infect Domestic Cats(MDPI (Basel, Switzerland), 2025-05-14) Munday JS; French AF; Broughton L; Lin X; Bond SD; Kraberger S; Knox MA; De Martino LDomestic cats are currently recognized to be infected by 10 different Felis catus papillomavirus (FcaPV) types that are classified into three genera. Examination of a skin sample from a cat with presumptive allergic dermatitis revealed clusters of large amphophilic intracytoplasmic bodies within epidermal cells. A 312 bp section of DNA from a novel PV type was amplified from the sample, while the entire 7569 bp genome was amplified and sequenced from a skin swab. The novel PV, which was designated FcaPV11, was predicted to contain coding regions for five early proteins and two late ones. Phylogenetic analysis of the L1 gene sequence showed FcaPV11 clusters with members of the Treisetapapillomavirus genus and shares less than 64% similarity with any of the previously fully sequenced FcaPV types. FcaPV11 DNA was not detected in a series of neoplastic and non-neoplastic skin samples from an additional 30 cats. These results show, for the first time, that cats can be infected by members of the Treisetapapillomavirus genus and suggest PVs in this genus may have co-evolved with a common Carnivora ancestor. While FcaPV11 was considered unlikely to have caused skin lesions in this cat, the prominent PV-induced cell changes indicate the PV can influence cell regulation. This suggests FcaPV11 may have the potential to cause skin disease in cats.Item Metabarcoding captures genetic diversity and links cases in outbreaks of cryptosporidiosis in New Zealand.(Elsevier Ltd on behalf of The British Infection Association, 2025-01-30) Ogbuigwe P; Biggs PJ; Garcia-Ramirez JC; Knox MA; Pita A; Velathanthir N; French NP; Hayman DTSCryptosporidiosis is a disease caused by the parasite Cryptosporidium. Globally, it is a leading cause of diarrhoea and a notifiable disease in New Zealand. Molecular analyses of Cryptosporidium isolated from notified cases do not always provide support for epidemiological links between individuals. We hypothesised this could be due to undetected diversity and the use of consensus Sanger sequence analyses. Here, we analysed 105 Cryptosporidium samples from outbreaks and sporadic cases occurring between 2010 and 2018 in New Zealand using both Next-Generation Sequencing (NGS) and Sanger sequencing of the glycoprotein 60 (gp60) locus. NGS metabarcoding at the gp60 locus uncovered significant intra- and inter-sample genotypic diversity in outbreaks and identified subtypes shared by epidemiologically linked cases, along with rare subtypes, suggesting it may be a useful tool for epidemiological investigations.Item Development of a non-infectious control for viral hemorrhagic fever PCR assays(2023-05-23) Knox MA; Bromhead C; Hayman DTSItem Canis Familiaris Papillomavirus Type 26: A Novel Papillomavirus of Dogs and the First Canine Papillomavirus within the Omegapapillomavirus Genus.(MDPI (Basel, Switzerland), 2024-04-12) Munday JS; Bond SD; Piripi S; Soulsby SJ; Knox MA; Christensen NDomestic dogs are currently recognized as being infected by 25 different canine papillomavirus (CPV) types classified into three genera. A short sequence from a novel CPV type was amplified, along with CPV1, from a papilloma (wart) from the mouth of a dog. The entire 7499 bp genome was amplified, and CPV26 contained putative coding regions that were predicted to produce four early proteins and two late ones. The ORF L1 showed less than 62% similarity for all previously sequenced CPV types but over 69% similarity to multiple Omegapapillomavirus types from a variety of Caniform species including the giant panda, Weddel seal, and polar bear. Phylogenetic analysis confirmed CPV26 clusters within the Omegapapillomavirus genus. Specific primers were used to investigate the presence of CPV26 DNA within a series of 37 canine proliferative lesions. CPV26 DNA was amplified from one lesion, a cutaneous papilloma that also contained CPV6. This is the first time a PV type within the Omegapapillomavirus genus has been detected in a non-domestic species and this provides evidence that the omegapapillomaviruses infected a common ancestor of, and then co-evolved with, the Caniform species. Whether CPV26 causes disease is uncertain, but the absence of an E7 protein may suggest low pathogenicity.Item Quantifying Replication Slippage Error in Cryptosporidium Metabarcoding Studies.(Oxford University Press, 2024-07-15) Knox MA; Biggs PJ; Garcia-R JC; Hayman DTSGenetic variation in Cryptosporidium, a common protozoan gut parasite in humans, is often based on marker genes containing trinucleotide repeats, which differentiate subtypes and track outbreaks. However, repeat regions have high replication slippage rates, making it difficult to discern biological diversity from error. Here, we synthesized Cryptosporidium DNA in clonal plasmid vectors, amplified them in different mock community ratios, and sequenced them using next-generation sequencing to determine the rate of replication slippage with dada2. Our results indicate that slippage rates increase with the length of the repeat region and can contribute to error rates of up to 20%.Item Historical translocations by Māori may explain the distribution and genetic structure of a threatened surf clam in Aotearoa (New Zealand)(Springer Nature Ltd, 2018-11-22) Ross PM; Knox MA; Smith S; Smith H; Williams J; Hogg IDThe population genetic structure of toheroa (Paphies ventricosa), an Aotearoa (New Zealand) endemic surf clam, was assessed to determine levels of inter-population connectivity and test hypotheses regarding life history, habitat distribution and connectivity in coastal vs. estuarine taxa. Ninety-eight toheroa from populations across the length of New Zealand were sequenced for the mitochondrial cytochrome c oxidase I gene with analyses suggesting a population genetic structure unique among New Zealand marine invertebrates. Toheroa genetic diversity was high in Te Ika-a Māui (the North Island of New Zealand) but completely lacking in the south of Te Waipounamu (the South Island), an indication of recent isolation. Changes in habitat availability, long distance dispersal events or translocation of toheroa to southern New Zealand by Māori could explain the observed geographic distribution of toheroa and their genetic diversity. Given that early-Māori and their ancestors, were adept at food cultivation and relocation, the toheroa translocation hypothesis is plausible and may explain the disjointed modern distribution of this species. Translocation would also explain the limited success in restoring what may in some cases be ecologically isolated populations located outside their natural distributions and preferred nichesItem Diagnosis of protozoa diarrhoea in Campylobacter patients increases markedly with molecular techniques.(Public Library of Science (PLoS), 2023-05-30) Hayman DTS; Garcia-Ramirez JC; Pita A; Velathanthiri N; Knox MA; Ogbuigwe P; Baker MG; Rostami K; Deroles-Main J; Gilpin BJ; Standley CCryptosporidium and Giardia are major causes of diarrhoea globally, and two of the most notified infectious diseases in New Zealand. Diagnosis requires laboratory confirmation carried out mostly via antigen or microscopy-based techniques. However, these methods are increasingly being superseded by molecular techniques. Here we investigate the level of protozoa detection by molecular methods in campylobacteriosis cases missed through antigen-based assays and investigate different molecular testing protocols. We report findings from two observational studies; the first among 111 people during a Campylobacter outbreak and the second during normal surveillance activities among 158 people presenting with diarrhoea and a positive Campylobacter test, but negative Cryptosporidium and Giardia antigen-based test results. The molecular methods used for comparison were in-house end-point PCR tests targeting the gp60 gene for Cryptosporidium and gdh gene for Giardia. DNA extraction was performed with and without bead-beating and comparisons with commercial real-time quantitative (qPCR) were made using clinical Cryptosporidium positive sample dilutions down to 10-5. The Cryptosporidium prevalence was 9% (95% CI: 3-15; 10/111) and Giardia prevalence 21% (95% CI: 12-29; 23/111) in the 111 Campylobacter outbreak patients. The Cryptosporidium prevalence was 40% (95% CI: 32-48; 62/158) and Giardia prevalence 1.3% (95% CI: 0.2-4.5; 2/158) in the 158 routine surveillance samples. Sequencing identified Cryptosporidium hominis, C. parvum, and Giardia intestinalis assemblages A and B. We found no statistical difference in positive test results between samples using end-point PCR with or without bead-beating prior to DNA extraction, or between the in-house end-point PCR and qPCR. The qPCR Ct value was 36 (95% CI: 35-37) for 1 oocyst, suggesting a high limit of detection. In conclusion in surveillance and outbreak situations we found diagnostic serology testing underdiagnoses Cryptosporidium and Giardia coinfections in Campylobacter patients, suggesting the impact of protozoa infections may be underestimated through underdiagnosis using antigen-based assays.Item A novel, stain-free, natural auto-fluorescent signal, Sig M, identified from cytometric and transcriptomic analysis of infectivity of Cryptosporidium hominis and Cryptosporidium parvum.(Frontiers Media S.A., 2023-05-22) Ogbuigwe P; Roberts JM; Knox MA; Heiser A; Pita A; Haack NA; Garcia-Ramirez JC; Velathanthiri N; Biggs PJ; French NP; Hayman DTS; Xu RCryptosporidiosis is a worldwide diarrheal disease caused by the protozoan Cryptosporidium. The primary symptom is diarrhea, but patients may exhibit different symptoms based on the species of the Cryptosporidium parasite they are infected with. Furthermore, some genotypes within species are more transmissible and apparently virulent than others. The mechanisms underpinning these differences are not understood, and an effective in vitro system for Cryptosporidium culture would help advance our understanding of these differences. Using COLO-680N cells, we employed flow cytometry and microscopy along with the C. parvum-specific antibody Sporo-Glo™ to characterize infected cells 48 h following an infection with C. parvum or C. hominis. The Cryptosporidium parvum-infected cells showed higher levels of signal using Sporo-Glo™ than C. hominis-infected cells, which was likely because Sporo-Glo™ was generated against C. parvum. We found a subset of cells from infected cultures that expressed a novel, dose-dependent auto-fluorescent signal that was detectable across a range of wavelengths. The population of cells that expressed this signal increased proportionately to the multiplicity of infection. The spectral cytometry results confirmed that the signature of this subset of host cells closely matched that of oocysts present in the infectious ecosystem, pointing to a parasitic origin. Present in both C. parvum and C. hominis cultures, we named this Sig M, and due to its distinct profile in cells from both infections, it could be a better marker for assessing Cryptosporidium infection in COLO-680N cells than Sporo-Glo™. We also noted Sig M's impact on Sporo-Glo™ detection as Sporo-Glo™ uses fluoroscein-isothiocynate, which is detected where Sig M also fluoresces. Lastly, we used NanoString nCounter® analysis to investigate the transcriptomic landscape for the two Cryptosporidium species, assessing the gene expression of 144 host and parasite genes. Despite the host gene expression being at high levels, the levels of putative intracellular Cryptosporidium gene expression were low, with no significant difference from controls, which could be, in part, explained by the abundance of uninfected cells present as determined by both Sporo-Glo™ and Sig M analyses. This study shows for the first time that a natural auto-fluorescent signal, Sig M, linked to Cryptosporidium infection can be detected in infected host cells without any fluorescent labeling strategies and that the COLO-680N cell line and spectral cytometry could be useful tools to advance the understanding of Cryptosporidium infectivity.Item Abundant dsRNA picobirnaviruses show little geographic or host association in terrestrial systems.(Elsevier, 2023-08) Knox MA; Wierenga J; Biggs PJ; Gedye K; Almeida V; Hall R; Kalema-Zikusoka G; Rubanga S; Ngabirano A; Valdivia-Granda W; Hayman DTSPicobirnaviruses are double-stranded RNA viruses known from a wide range of host species and locations but with unknown pathogenicity and host relationships. Here, we examined the diversity of picobirnaviruses from cattle and gorillas within and around Bwindi Impenetrable Forest National Park (BIFNP), Uganda, where wild and domesticated animals and humans live in relatively close contact. We use metagenomic sequencing with bioinformatic analyses to examine genetic diversity. We compared our findings to global Picobirnavirus diversity using clustering-based analyses. Picobirnavirus diversity at Bwindi was high, with 14 near-complete RdRp and 15 capsid protein sequences, and 497 new partial viral sequences recovered from 44 gorilla samples and 664 from 16 cattle samples. Sequences were distributed throughout a phylogenetic tree of globally derived picobirnaviruses. The relationship with Picobirnavirus diversity and host taxonomy follows a similar pattern to the global dataset, generally lacking pattern with either host or geography.