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The epidemiology and pathology of 

Paranannizziopsis australasiensis in New Zealand 

reptiles 

  

  

  

  

A thesis presented in partial fulfilment of the requirements 

 for the degree of  

Master of Veterinary Science 

 In 

 Wildlife Health  

at Massey University,  

Palmerston North New Zealand 

  

  

  

 

 

 

 Rebecca Kirsty Elizabeth Webster 

2018 

 





 
i 

Abstract 

 

Paranannizziopsis australasiensis, has recently been diagnosed in tuatara at two 

captive facilities in New Zealand. This newly emerging fungal pathogen, is a member 

of the onygenalean fungal group formally known as Chrysosporium anamorph of 

Nannizziopsis vriesii (CANV). Fungi of this genera are thought to be obligate primary 

pathogens in reptiles, and closely related species such as Ophidiomyces ophiodiicola, 

and Nannizziopsis guarroi have caused significant morbidity and mortalities in captive 

and wild reptile populations. The detection of this disease raised concerns for wild and 

captive population health and resulted in a temporary cessation of tuatara breed and 

release programmes from affected facilities. Similar lesions have been reported in 

tuatara at multiple other captive facilities in New Zealand, but lack of veterinary 

assessment and, until recently, inadequate diagnostic capabilities has led to an 

inability to confirm the presence or absence of P. australasiensis in these populations.  

 

This research aimed to investigate the epidemiology of P. australasiensis in New 

Zealand wild and captive endemic reptiles. Skin samples were collected from nine 

captive, six wild and two ecosanctuary populations of tuatara across New Zealand. 

Skin samples from in contact geckos and skinks were opportunistically collected to 

determine the possible cross species infection of P. australasiensis. Samples were 

tested for presence of P. australasiensis by fungal culture followed by PCR, and by 

loop-mediated isothermal amplification (LAMP). Soil samples were collected from 

burrows, basking areas and captive enclosures and analysed by LAMP to determine 

the presence of P. australasiensis within the environment.  

 

Paranannizziopsis australasiensis was found to be wide spread in New Zealand 

captive and wild reptile populations. In populations where the pathogen was detected 

prevalence varied between 6.7% and 44.4% for tuatara, 3.8% and 40% for geckos 

and 6.7% and 66.7% for skinks. A low virulence of disease associated with infection 

was seen in tuatara across New Zealand, with many LAMP positive tuatara being 

asymptomatic. Increased severity of disease was seen in two captive tuatara, where 



 
ii 

other concurrent disease was present. One fatality was reported. In other reptile hosts, 

no disease was identified, and it is suspected these species act as reservoirs for the 

transmission of this organism to tuatara.  Paranannizziopsis australasiensis was 

detected multiple times in soil samples and may survive as an environmental 

saprophyte. 

 

Paranannizziopsis australasiensis appears to have a close association with New 

Zealand reptiles. The prevalence, distribution and pathology of P. australasiensis 

observed in this study suggests that this organism is not a threat to tuatara or other 

endemic reptile populations in New Zealand. The findings of this study have enabled 

restrictions placed on tuatara translocations, based on P. australasiensis status, to be 

removed.  

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 



 
iii 

Acknowledgements 

 

This project has been a huge collaborative effort and would not have been possible 

without the time and generosity of many people. I am particularly grateful for the 

assistance given by both Hye Jeong Ha and the staff at the Animal Health Laboratory, 

and Richard Winkworth, and associated staff at the Institute of Fundamental Sciences.  

Thank you for both providing countless hours, expertise and enthusiasm for this 

project, and for tolerating a never-ending number of samples.  Special thanks also to 

Kate McInnes, for organising the collection of a significant amount of the data used in 

this study, for coordinating discussions on this disease and for assisting in gaining 

funding and support for this project. 

 

My sincere gratitude is expressed to my supervisor Professor Brett Gartrell, without 

whom this project would not have been possible. Thank you for helping me find and 

build a project that I have loved. Thank you for your patience and encouragement 

throughout both my residency and this project and thank you especially for your 

perseverance in the analysis stage and for the many hours spent decoding my data.  

 

Thank you to all of the PaNTS team, particularly the Auckland zoo vets Sarah 

Alexander, An Pas and James Chatterton, for sharing your knowledge and providing 

discussion and advice on all things fungal. Thank you to Karina Argandona-Gonzalez 

and Wellington Zoo for the permission to include the clinical cases used in this project, 

thank you to Nicola Nelson, and Stephanie Price for teaching me the tricks of tuatara 

hunting, and to Stuart Hunter and Adrienne French for the histology photos and 

assistance with the interpretation of these. 

 

I would like to express my appreciation to all the staff from the many zoos and 

sanctuaries across New Zealand for your permission and assistance in the sampling 

of your reptiles. In particular; Chye-mei Haung and Chris Jerrum (Wellington Zoo), 

Matu Booth, Rhys Mills and Dave Banks (Nga Manu), Jess Flamy and Todd Jenkinson 

(Pukaha), Jo Ledington and Danielle Shanahan (Zealandia) Lindsay Hazley 



 
iv 

(Southland Museum), Alyssa Salton (Orana), Paul Kavsey (Kiwi Birdlife Park), Sue 

Keall (VUW), Mark Paterson, Teresa McCauley, and Kelly-Anne Banks (Rainbow 

Springs), Tamsin Ward-Smith (Cape Sanctuary), and, Rob Chappell for making the 

island work possible. It has been a privilege to meet you all and witness the 

conservation work taking place across New Zealand. 

 

Thank you to all my wonderful volunteers, Alisdair Eddie, Brittany Whitford, Amy 

Kachurowski, Sarah Michael (also thank you for the beautiful photography and for the 

coffee dates), Sarah Vermeer, Emily Kay, Christine Tan, Jaime Park, Maxine Jenkins 

and Kayla Aitken. And special thanks to the triangle team; Karissa Bernoth, Chye-Mei 

Huang, and Xue Qi Soon, for sharing many nights filled with coffee and chocolate, 

terrible singing, aching legs, and sometimes even tuatara. Thank you for sharing the 

trip of a life time with me I couldn’t have hoped for a better team. 

 

Thank you to Scotty the weird little welsh man who came to my rescue and fixed my 

computer when it crashed just two weeks before submission. And to PJ, my life 

assistant, thank you for your friendship and for helping me find a world outside of my 

tunnel of research.  

 

Thank you to my wonderful family. My mummy, Jacqui, for always believing in me and 

for helping out on all the rainy days. Thank you to my boys Lachlan and Jai for putting 

up with long periods with an absent, busy and distracted mum. I am so proud of you 

both. And to my Loz, thank you for just being there, and for still being here. These last 

four years have been massive, and I feel blessed to have been on this roller coaster 

with you by my side.   

 

And finally thank you to all the tuatara, for letting me poke you and for just being the 

perfect little beings that you are. 

 



 
v 

Funding and ethics approval  

 

Funding for this project was provided in part by the Institute of Veterinary, Animal and 

Biomedical Sciences (IVABS) postgraduate fund, and the New Zealand Department 

of Conservation. All work was carried out with permission from the New Zealand 

Department of Conservation authorisation number 48536 – FAU and approved by the 

Massey University Animal Ethics Committee (MUAEC 55/99). 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 



 
vi 

Table of contents  

 

ABSTRACT .......................................................................................................................... i 

ACKNOWLEDGEMENTS .................................................................................................. iii 

FUNDING AND ETHICS APPROVAL .............................................................................. v 

TABLE OF CONTENTS ..................................................................................................... vi 

LIST OF FIGURES ............................................................................................................. ix 

LIST OF TABLES ............................................................................................................... xi 

 

CHAPTER ONE:  Introduction, literature review and research aims ..................... 1 

1.1 INTRODUCTION  ............................................................................................. 2 

1.2 EMERGING MYCOTIC DISEASES ............................................................... 3 

1.3 MYCOTIC DISEASE IN REPTILES  .............................................................. 4 

1.3.1 Mycotic Disease in New Zealand Reptiles  ................................... 4 

1.4 REPTILE PATHOGENS OF THE GENERA NANNIZZIOPSIS,  

     PARANANNIZZIOPSIS AND OPHIDIOMYCES  ................................. 5 

1.4.1 Classification .................................................................................... 5 

1.4.2 Diagnostics ....................................................................................... 6 

1.4.2.i Loop mediated isothermal amplification ............................ 8 

1.4.3 Transmission .................................................................................... 9 

1.4.4 Clinical Presentation ...................................................................... 10 

1.4.5 Predisposing Factors to Disease ................................................. 11 

1.4.6 Treatment ....................................................................................... 12 

1.4.7 Paranannizziopsis australasiensis ............................................... 14 

1.4.7.i Paranannizziopsis australasiensis in tuatara ............. 14 

1.5 GENERAL BIOLOGY AND CONSERVATION OF TUATARA .................. 15 

1.6 ENVIRONMENTAL CHARACTERISTICS OF WILD TUATARA  

HABITAT ......................................................................................................... 17 

1.7 CAPTIVE HUSBANDRY OF TUATARA ...................................................... 17 

1.7.1 Diet .................................................................................................. 19 

1.8 RESEARCH AIMS AND THESIS STRUCTURE  ........................................ 20 

LITERATURE CITED ................................................................................................ 21 

 



 
vii 

CHAPTER TWO: The prevalence of Paranannizziopsis australasiensis in New 

Zealand reptiles and environmental sources  .......................................................... 27 

2.1 INTRODUCTION ............................................................................................ 28 

2.2 MATERIALS AND METHODS ...................................................................... 30 

2.2.1 Location, capture and sample technique ....................................... 30 

2.2.2 Culture and PCR .............................................................................. 32 

2.2.3 Loop mediated isothermal amplification ........................................ 32 

2.2.4 Sex and age determination ............................................................. 33 

2.2.5 Soil samples ..................................................................................... 34 

2.2.6 Statistical Methods .......................................................................... 34 

2.3 RESULTS ........................................................................................................ 35 

2.3.1Tuatara ............................................................................................... 35 

2.3.1.i Culture and PCR ................................................................ 35 

2.3.1.ii LAMP .................................................................................. 35 

2.3.1.iii Dermatitis  ......................................................................... 36 

2.3.2 Geckos and Skinks .......................................................................... 37 

2.3.3 Association between Paranannizziopsis australasiensis presence 

and dermatitis ............................................................................................ 39 

2.3.3.i All reptiles combined .......................................................... 39 

2.3.3.ii Tuatara only ....................................................................... 40 

2.3.4 Analysis of risk factors for presence of Paranannizziopsis 

australasiensis ........................................................................................... 40 

   2.3.4.i All reptiles combined .......................................................... 40 

   2.3.4.ii Tuatara only ....................................................................... 44 

2.3.5 Analysis of risk factors for presence of dermatitis ........................ 45 

2.3.5.i All reptiles combined .......................................................... 45 

2.3.5.ii Tuatara only ....................................................................... 50 

2.3.6 Soil .................................................................................................... 53 

2.4 DISCUSSION .................................................................................................. 53 

LITERATURE CITED ................................................................................................. 57 

 

CHAPTER THREE: The pathology of Paranannizziopsis australasiensis in 

tuatara (Sphenodon punctatus)  ................................................................................. 60 

3.1 INTRODUCTION ............................................................................................ 61 



 
viii 

3.2 MATERIALS AND METHODS ...................................................................... 63 

3.2.1 Skin samples and lesions ............................................................... 64 

3.2.2 Treatment and quarantine of positive cases ................................. 65 

3.2.3 Morphometrics ................................................................................. 65 

3.2.4 Haematology .................................................................................... 66 

3.2.5 Statistical Methods .......................................................................... 66 

3.3 RESULTS ........................................................................................................ 66 

3.3.1 Lesions ............................................................................................. 66 

3.3.1.i Case 1 ................................................................................. 67 

3.3.1.ii Case 2  ............................................................................... 68 

3.3.1.iii Case 3 ............................................................................... 69 

3.3.1.iv Other lesions ..................................................................... 72 

3.3.2 Effect of presence of Paranannizziopsis australasiensis on 

morphometrics and haematological parameters in tuatara ................... 72 

3.4 DISCUSSION .................................................................................................. 76 

LITERATURE CITED ................................................................................................. 79 

 

CHAPTER FOUR: GENERAL DISCUSSION ............................................................... 82 

4.1 OVERVIEW OF RESEARCH AIMS AND CONCLUSIONS ........................... 83 

4.1.1 Prevalence of Paranannizziopsis australasiensis in wild and 

captive tuatara ........................................................................................... 84 

4.1.2 Pathology of Paranannizziopsis australasiensis in tuatara ......... 85 

4.1.3 Paranannizziopsis australasiensis in endemic geckos and skinks

 ....................................................................................................................... 86 

4.1.4 Paranannizziopsis australasiensis in soil samples ..................... 86 

4.1.5 Paranannizziopsis australasiensis on Stanley and Cuvier Island

 ....................................................................................................................... 87 

4.2 DERMATITIS IN TUATARA .................................................................................. 88 

4.3 LIMITATIONS OF THE STUDY ........................................................................... 89 

4.4 CONSERVATION MANAGEMENT IMPLICATIONS ................................ 90 

4.5 AREAS OF FURTHER RESEARCH .......................................................... 91 

4.6 CONCLUSIONS ........................................................................................... 93 

     LITERATURE CITED .................................................................................................. 95 

 



 
ix 

List of figures   

 

FIGURE 2.1. Prevalence of Paranannizziopsis australasiensis for tuatara geckos and skinks at wild and

  ecosanctuary locations by LAMP test ..................................................................................... 39 

FIGURE 2.2. Frequency of P. australasiensis detection in all reptiles in the study by LAMP test in 

  different locations in New Zealand  ......................................................................................... 43 

FIGURE 2.3. Frequency of P. australasiensis detection in all reptiles in the study by reptile  

family  ....................................................................................................................................... 44  

FIGURE 2.4. Frequency of P. australasiensis detection in tuatara by LAMP test in different locations

  in New Zealand ....................................................................................................................... 45 

FIGURE 2.5. Frequency of the presence of dermatitis lesions in all reptiles examined by  

location ..................................................................................................................................... 47 

FIGURE 2.6. Frequency of presence of dermatitis lesions by management type (captive,  

  ecosanctuary or wild population) in all reptiles examined. ..................................................... 48 

FIGURE 2.7. Frequency of presence of dermatitis lesions by reptile family in all reptiles  

examined.  ................................................................................................................................ 49 

FIGURE 2.8. Frequency of presence of dermatitis lesions by age. The juveniles (*) have a significantly 

higher odds ratio for dermatitis presence than the adults ..................................................................... 51 

FIGURE 2.9. Frequency of the presence of dermatitis lesions in tuatara examined by  

location ..................................................................................................................................... 52 

FIGURE 2.10. Frequency of the presence of dermatitis lesions in tuatara examined by age  

cohort. ...................................................................................................................................... 52  

FIGURE 3.1. Map of New Zealand showing sampling sites ................................................................. 64 

FIGURE 3.2. Mild dermatitis lesions. a. A focal area of skin discolouration and flaking on the ventrum

  of a tuatara from North Brothers Island b. Multifocal discolouration of scales on the ventrum

  of a tuatara from Curvier Island.  ............................................................................................. 67 

FIGURE 3.3.  Case 1.  Ulceration of the mid ventral dermis of an adult male tuatara associated with a

  positive LAMP test result for Paranannizziopsis australasiensis.  .......................................... 68 

FIGURE 3.4. Case 2. Multifocal granulomatous lesions on A. the dorsal and right lateral aspects of the

  tail base (arrows), and B. between digits 2 and 3 of the right front foot (arrow) of a juvenile



 
x 

  male tuatara with Paranannizziopsis australasiensis confirmed by lamp test, culture and DNA

  sequencing .............................................................................................................................. 69 

FIGURE 3.5. Case 3. A. Multifocal granulomatous lesions on the ventral chest, abdomen and tail (arrow

  heads) and B. transverse section through the palmar carpal aspect of the right front foot 

  showing multiple granulomas (arrow heads) adjacent to the carpal bones of a juvenile male 

 tuatara with  Paranannizziopsis australasiensis confirmed by LAMP test, culture and DNA

  sequencing .............................................................................................................................. 70 

FIGURE 3.6. A. Case 1: Hyperkeratosis (arrow) of the lesion edges, with gram negative and occasional

 gram-positive coccobacillus embedded in this material (arrow head). B. Case 2: Necrotic 

 dermis and epidermis infiltrated with moderate numbers of septate parallel walled hyphae with

 right angled branching (arrow heads) C – F. Case 3.  C. Granulomatous dermatitis and myositis

 with heterophilic infiltration (arrow head) and pyknotic debris (bold arrow). The dermis is intact

 with areas of hyperkeratosis (arrow). D. A granuloma containing fungal hyphae with occasional 

 branching (arrow heads) PAS stain. E and F: Right front foot, Case 3.  Granulomas extend 

 deep into the dermis and muscle, with large amounts of necrotic debris surrounding bones and 

 joints (bold arrows), with early cortical inflammation (arrow head). 

  ................................................................................................................................................. 71 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 



 
xi 

List of tables 

 

TABLE 2.1. Number of reptiles, organised by family, tested for P. australasiensis by LAMP and culture

 at each location ........................................................................................................................ 31 

TABLE 2.2. LAMP results for Paranannizziopsis australasiensis in tuatara by location within New 

 Zealand .................................................................................................................................... 36 

TABLE 2.3. LAMP results for Paranannizziopsis australasiensis in New Zealand geckos and skinks by 

 species and location ................................................................................................................. 38 

TABLE 2.4. Contingency table for presence or absence of dermatitis (lesions) and the result of the 

 LAMP analysis for Paranannizziopsis australasiensis in all reptiles studied ........................... 40 

TABLE 2.5. Contingency table for presence or absence of dermatitis (lesions) and the result of the 

 LAMP analysis for Paranannizziopsis australasiensis in tuatara only ..................................... 40 

TABLE 2.6. Results of a reduced model binomial logistic regression for risk factors associated with a 

 positive P. australasiensis LAMP test result in all reptiles in this study ................................... 42 

TABLE 2.7. Results of a reduced model binomial logistic regression for risk factors associated with the 

 presence of dermatitis lesions in all reptiles examined ............................................................ 46 

TABLE 2.8. Results of a reduced model binomial logistic regression for risk factors associated with the 

 presence of dermatitis lesions in tuatara ................................................................................. 49 

TABLE 3.1. Lesion presence in tuatara tested for Paranannizziopsis australasiensis (PA) on LAMP. 

 There was no significant association in the frequency of dermatitis lesions with a positive result 

 on the PA LAMP test (Chi-square 0.016, df = 1, p = 0.901) .................................................... 67 

TABLE 3.2. Descriptive statistics for morphological and haematological variables between tuatara 

 testing positive and negative for P. australasiensis by LAMP test........................................... 73 

TABLE 3.3 AND 3.4. Analysis of differences between morphological and haematological parameters

  in tuatara testing positive and negative for P. australasiensis by LAMP test ......................... 74