title: Bioactive Chemicals of Importance in Endophyte-Infected Grasses

authors: Babu, Jacob

subjects: indole-diterpenoids

abstract: Janthitrems are believed to be involved in the observed sporadic cases of AR37-
infected perennial ryegrass staggers. Investigations into the role of janthitrems in
perennial ryegrass staggers are difficult as isolation of the compounds from the
ryegrass is hindered by the inherent instability of these compounds. Therefore
attempts were made to isolate janthitrems from an alternative source, allowing
these janthitrem analogues to be used as surrogates for endophyte produced
janthitrems.
Analysis of a series of Penicillium janthinellum cultures revealed the presence of
janthitrems in a number of strains, including janthitrem B, janthitrem C and two
novel janthitrem compounds. Detailed one- and two-dimensional NMR and mass
spectral techniques identified the two novel compounds as 11,12-
epoxyjanthitrems B and C, which were subsequently given the trivial names
janthitrems A and D, respectively. Janthitrems B and C were isolated and
identified by NMR and revisions of some previously reported chemical shift
assignments were proposed. In addition to the janthitrems, penitrems were also
identified in two strains of P. janthinellum.
The isolated janthitrem B was utilised for the development of efficient extraction
procedures, and for the determination of ideal storage conditions for janthitrem
compounds. A method for the extraction and isolation of janthitrem B from a P.
janthinellum culture was developed and optimised to yield 6 mg of janthitrem B
from 900 mL of fungal culture in two days. Stability studies of janthitrem B
indicated the ideal storage condition which minimised degradation was dry at
−80 C where only 7% sample loss was observed over 300 days.
Bioactivity studies of janthitrems A and B found these compounds to be
tremorgenic to mice, with janthitrem A (an epoxyjanthitrem) inducing more
severe tremors than janthitrem B. Insect testing also showed that both janthitrems
A and B displayed anti-insect activity to porina larvae. Since the
epoxyjanthitrems, which are associated with AR37 endophyte-infected ryegrass,
were also shown to be tremorgenic and to display anti-insect activity, the insect
resistance and the sporadic cases of ryegrass staggers displayed by AR37 may be
related to the presence of epoxyjanthitrem compounds.
LC-UV-MS analysis of janthitrems A-D, penitrems A-F, lolitrem B, paspalinine,
paxilline and terpendole C found these indole-diterpenoids to be more sensitive
by analysis using an APCI source as opposed to an ESI source. APCI negative
ion LC-UV-MS required source induced dissociation in combination with
increased collision energy to suppress an acetate adduct peak, sourced from the
acetic acid buffer. Negative ion MS2 and MS3 data produced more informative
fragments compared to the conventional positive ion MS2 and MS3 data. The
availability of both positive and negative ion LC-UV-MS methodologies will
allow future endophyte products to be more thoroughly screened for different
classes of secondary metabolites.
Extracts of mouldy walnuts were analysed for the presence of tremorgenic
mycotoxins after a dog was found to exhibit symptoms characteristic of
tremorgenic mycotoxicosis. LC-UV-MS analysis of the mouldy walnuts
identified the tremorgenic mycotoxins penitrems A-F, thus confirming the
veterinarian's tentative diagnosis of canine tremorgenic mycotoxicosis   the first
reported case in New Zealand.

text: Bioactive Chemicals of Importance in Endophyte-Infected Grasses Janthitrems are believed to be involved in the observed sporadic cases of AR37-
infected perennial ryegrass staggers. Investigations into the role of janthitrems in
perennial ryegrass staggers are difficult as isolation of the compounds from the
ryegrass is hindered by the inherent instability of these compounds. Therefore
attempts were made to isolate janthitrems from an alternative source, allowing
these janthitrem analogues to be used as surrogates for endophyte produced
janthitrems.
Analysis of a series of Penicillium janthinellum cultures revealed the presence of
janthitrems in a number of strains, including janthitrem B, janthitrem C and two
novel janthitrem compounds. Detailed one- and two-dimensional NMR and mass
spectral techniques identified the two novel compounds as 11,12-
epoxyjanthitrems B and C, which were subsequently given the trivial names
janthitrems A and D, respectively. Janthitrems B and C were isolated and
identified by NMR and revisions of some previously reported chemical shift
assignments were proposed. In addition to the janthitrems, penitrems were also
identified in two strains of P. janthinellum.
The isolated janthitrem B was utilised for the development of efficient extraction
procedures, and for the determination of ideal storage conditions for janthitrem
compounds. A method for the extraction and isolation of janthitrem B from a P.
janthinellum culture was developed and optimised to yield 6 mg of janthitrem B
from 900 mL of fungal culture in two days. Stability studies of janthitrem B
indicated the ideal storage condition which minimised degradation was dry at
−80 C where only 7% sample loss was observed over 300 days.
Bioactivity studies of janthitrems A and B found these compounds to be
tremorgenic to mice, with janthitrem A (an epoxyjanthitrem) inducing more
severe tremors than janthitrem B. Insect testing also showed that both janthitrems
A and B displayed anti-insect activity to porina larvae. Since the
epoxyjanthitrems, which are associated with AR37 endophyte-infected ryegrass,
were also shown to be tremorgenic and to display anti-insect activity, the insect
resistance and the sporadic cases of ryegrass staggers displayed by AR37 may be
related to the presence of epoxyjanthitrem compounds.
LC-UV-MS analysis of janthitrems A-D, penitrems A-F, lolitrem B, paspalinine,
paxilline and terpendole C found these indole-diterpenoids to be more sensitive
by analysis using an APCI source as opposed to an ESI source. APCI negative
ion LC-UV-MS required source induced dissociation in combination with
increased collision energy to suppress an acetate adduct peak, sourced from the
acetic acid buffer. Negative ion MS2 and MS3 data produced more informative
fragments compared to the conventional positive ion MS2 and MS3 data. The
availability of both positive and negative ion LC-UV-MS methodologies will
allow future endophyte products to be more thoroughly screened for different
classes of secondary metabolites.
Extracts of mouldy walnuts were analysed for the presence of tremorgenic
mycotoxins after a dog was found to exhibit symptoms characteristic of
tremorgenic mycotoxicosis. LC-UV-MS analysis of the mouldy walnuts
identified the tremorgenic mycotoxins penitrems A-F, thus confirming the
veterinarian's tentative diagnosis of canine tremorgenic mycotoxicosis   the first
reported case in New Zealand.

year: 2009

title: Disbudding of dairy goat kids: Refining protocols to reduce or eliminate pain

authors: Hempstead, Melissa Nicole

subjects: Disbudding

abstract: The aim of this thesis was to investigate potential improvements for disbudding dairy goat kids to eliminate or reduce pain. Behavioural and physiological measures were used to evaluate: (1) pain associated with four disbudding methods, and (2) the effect of pain mitigation on cautery disbudded goat kids. Caustic paste and cryosurgical disbudding and clove oil injection initially showed promise as less painful alternatives to cautery disbudding. However, caustic paste and cryosurgical disbudding were found to be more painful, as cortisol concentrations and the frequencies of head-directed behaviours were higher than for cautery disbudded kids. Clove oil injection appeared to cause no more pain than cautery disbudding based on similar cortisol concentrations and head-directed behaviour, but caused less tissue damage. Alternatives to current practices must be efficacious in preventing horn or scur growth, therefore the effect of clove oil injection and two cautery disbudding techniques (horn bud removed vs. left intact) on horn growth were evaluated. Clove oil was ineffective at preventing horns or scurs compared with cautery disbudding. In addition, cautery disbudding by removing the horn bud was more successful at preventing growth than leaving the horn bud intact. As using a cautery iron to remove the horn bud appeared to be the most efficacious method evaluated in this thesis, the effect of isoflurane and meloxicam on pain associated with cautery disbudding was investigated. Goat kids disbudded under isoflurane, with or without meloxicam, had lower cortisol concentrations and displayed fewer head-directed behaviours than goat kids that were cautery disbudded without pain relief, suggesting that isoflurane can reduce pain associated with cautery disbudding. Overall, the results of this thesis suggest that cautery disbudding is an effective practice but should ideally be carried out using isoflurane to reduce pain; also, complete horn bud removal lowers the risk of horn or scur development. These results support the continued use of cautery disbudding by the dairy goat industry but suggest pain relief could significantly improve the welfare of dairy goats on farm.

text: Disbudding of dairy goat kids: Refining protocols to reduce or eliminate pain The aim of this thesis was to investigate potential improvements for disbudding dairy goat kids to eliminate or reduce pain. Behavioural and physiological measures were used to evaluate: (1) pain associated with four disbudding methods, and (2) the effect of pain mitigation on cautery disbudded goat kids. Caustic paste and cryosurgical disbudding and clove oil injection initially showed promise as less painful alternatives to cautery disbudding. However, caustic paste and cryosurgical disbudding were found to be more painful, as cortisol concentrations and the frequencies of head-directed behaviours were higher than for cautery disbudded kids. Clove oil injection appeared to cause no more pain than cautery disbudding based on similar cortisol concentrations and head-directed behaviour, but caused less tissue damage. Alternatives to current practices must be efficacious in preventing horn or scur growth, therefore the effect of clove oil injection and two cautery disbudding techniques (horn bud removed vs. left intact) on horn growth were evaluated. Clove oil was ineffective at preventing horns or scurs compared with cautery disbudding. In addition, cautery disbudding by removing the horn bud was more successful at preventing growth than leaving the horn bud intact. As using a cautery iron to remove the horn bud appeared to be the most efficacious method evaluated in this thesis, the effect of isoflurane and meloxicam on pain associated with cautery disbudding was investigated. Goat kids disbudded under isoflurane, with or without meloxicam, had lower cortisol concentrations and displayed fewer head-directed behaviours than goat kids that were cautery disbudded without pain relief, suggesting that isoflurane can reduce pain associated with cautery disbudding. Overall, the results of this thesis suggest that cautery disbudding is an effective practice but should ideally be carried out using isoflurane to reduce pain; also, complete horn bud removal lowers the risk of horn or scur development. These results support the continued use of cautery disbudding by the dairy goat industry but suggest pain relief could significantly improve the welfare of dairy goats on farm.

year: 2018

title: Editorial

authors: Clarkin-Phillips, Jeanette

abstract: As New Zealand contemplates wholesale review of the formal schooling system and the leadership implications thereof, the winds of educational change begin to swirl. Not the persistent, yet gentle flutter of change that evolves the educational landscape in almost imperceptible ways, but tumultuous paradigmatic gales that potentially change it beyond all recognition. Heralded by politicians, economists and capitalists, there is little doubt that the 1989 neoliberal tempest wrought havoc on notions of community and profession. The whirlwind shift from social democracy to market competition, managerialism and performativity saw critical voices silenced and educators disparagingly positioned as self-interested providers. In the rush to legislate Tomorrow’s Schools, 18,000 submissions remained unanalysed and therein lies a crucial lesson: we ignore research at our peril. As educators, we have a moral obligation to generate, disseminate and critique understandings of leadership, policy and practice; to remain open to counter perspectives; and to contribute positively and hopefully to research that empowers and emancipates. Consistent with JELPP’s commitment to honouring diversity in leadership focus, setting, and authorship, articles in this year’s general edition offer fresh insight into topics of interconnected and perennial importance: native knowledge, instructional leadership, indigenous teachers, appraisal, professional values, and safe schools.

text: Editorial As New Zealand contemplates wholesale review of the formal schooling system and the leadership implications thereof, the winds of educational change begin to swirl. Not the persistent, yet gentle flutter of change that evolves the educational landscape in almost imperceptible ways, but tumultuous paradigmatic gales that potentially change it beyond all recognition. Heralded by politicians, economists and capitalists, there is little doubt that the 1989 neoliberal tempest wrought havoc on notions of community and profession. The whirlwind shift from social democracy to market competition, managerialism and performativity saw critical voices silenced and educators disparagingly positioned as self-interested providers. In the rush to legislate Tomorrow’s Schools, 18,000 submissions remained unanalysed and therein lies a crucial lesson: we ignore research at our peril. As educators, we have a moral obligation to generate, disseminate and critique understandings of leadership, policy and practice; to remain open to counter perspectives; and to contribute positively and hopefully to research that empowers and emancipates. Consistent with JELPP’s commitment to honouring diversity in leadership focus, setting, and authorship, articles in this year’s general edition offer fresh insight into topics of interconnected and perennial importance: native knowledge, instructional leadership, indigenous teachers, appraisal, professional values, and safe schools.

year: 2019