title: Expression of Mitochondrial Stress Protein (Cpn60) in in vitro Cultured Neonatal Porcine Islet Cells

authors: Munif, Farina

subjects: Mitochondria

abstract: Xenotransplantation of neonatal porcine islets have been demonstrated to be a viable alternative to exogenous insulin therapy for diabetes mellitus. The use of liberase has gained much success in islet isolation but factors such as batch-to-batch variation and deterioration of a batch with storage time have hampered the quality and reproducibility of tissue dissociation. Islet culture aims to optimise islet survival and insulin release in response to glucose challenge. However, it is difficult to recover and preserve islets in vitro.

Mitochondria play a key role in the secretion of insulin from pancreatic   islet cells in response to glucose stimulation. Mitochondrial dysfunction results in the induction (at mRNA and protein levels) of a molecular stress protein/heat chock protein called Cpn60. Since mitochondrial impairment will have a significant effect on the ability of in vitro cultured islet cells to function properly (i.e. release insulin in response to glucose stimulation), the expression of Cpn60 was investigated as a function of exposing neonatal porcine islet cells to various growth conditions.

The best choice of media to culture neonatal porcine islet cells was found to be not heated activated serum which showed the least levels of Cpn60 expression at mRNA levels suggesting that the cells had low levels of mitochondrial stress. Neonatal porcine islet cells would be best digested in cells digested with new liberase (QC 1050) while in 2% not heat inactivated porcine serum (NPS) as this gave the lowest levels of Cpn60 expression suggesting low levels of mitochondrial stress.

Although expression of Cpn60 at mRNA levels seems to be modulated during the growth of the porcine islet cells in media supplemented with different serum, heat treatment of serum and liberase content, no firm conclusion can be made with regard to the effect of the different treatments on mitochondrial health status until the porcine Cpn60 protein can be unequivocally identified.

text: Expression of Mitochondrial Stress Protein (Cpn60) in in vitro Cultured Neonatal Porcine Islet Cells Xenotransplantation of neonatal porcine islets have been demonstrated to be a viable alternative to exogenous insulin therapy for diabetes mellitus. The use of liberase has gained much success in islet isolation but factors such as batch-to-batch variation and deterioration of a batch with storage time have hampered the quality and reproducibility of tissue dissociation. Islet culture aims to optimise islet survival and insulin release in response to glucose challenge. However, it is difficult to recover and preserve islets in vitro.

Mitochondria play a key role in the secretion of insulin from pancreatic   islet cells in response to glucose stimulation. Mitochondrial dysfunction results in the induction (at mRNA and protein levels) of a molecular stress protein/heat chock protein called Cpn60. Since mitochondrial impairment will have a significant effect on the ability of in vitro cultured islet cells to function properly (i.e. release insulin in response to glucose stimulation), the expression of Cpn60 was investigated as a function of exposing neonatal porcine islet cells to various growth conditions.

The best choice of media to culture neonatal porcine islet cells was found to be not heated activated serum which showed the least levels of Cpn60 expression at mRNA levels suggesting that the cells had low levels of mitochondrial stress. Neonatal porcine islet cells would be best digested in cells digested with new liberase (QC 1050) while in 2% not heat inactivated porcine serum (NPS) as this gave the lowest levels of Cpn60 expression suggesting low levels of mitochondrial stress.

Although expression of Cpn60 at mRNA levels seems to be modulated during the growth of the porcine islet cells in media supplemented with different serum, heat treatment of serum and liberase content, no firm conclusion can be made with regard to the effect of the different treatments on mitochondrial health status until the porcine Cpn60 protein can be unequivocally identified.

year: 2006

title: Determining the risk of phosphite tolerance in Phytophthora species in New Zealand and the United States: a case study on the implications of long-term use of  phosphite to control Phytophthora cinnamomi in avocado (Persea americana)

authors: Hunter, Shannon Renee

subjects: Phytophthora

abstract: Phytophthora species cause many major plant disease epidemics worldwide, impacting horticulture, plant trade and production planting industries through the loss of plants and the costs arising from disease management. Phytophthora species have specialised structures which enhance their dispersal, survival and infection. These include motile swimming spores, caducous sporangia and resting spores. Nurseries and orchards use integrated management to control phytophthora diseases, including cultural practises, biological control, resistant host plants and chemical control. The use of fungicides is a very effective control method; however, Phytophthora species can develop resistance to fungicides after prolonged exposure. For example, phosphite tolerant isolates of Phytophthora cinnamomi have been found in avocado orchards in Australia and South Africa.
Phosphite is widely used in nurseries and orchards in New Zealand as a preventative and control treatment. The avocado industry largely relies on it for the control of avocado root rot caused by P. cinnamomi. Phosphite is most commonly applied to avocado trees using passive trunk injections but can also be applied as foliar sprays, soil drenches, high-pressure injections and capsule implants.
To assess phosphite tolerance of P. cinnamomi isolates from New Zealand avocado orchards, a high-throughput optical density (OD) assay was developed to measure mycelial growth inhibition in the presence of phosphite. The OD assay was used to screen 24 P. cinnamomi isolates from four orchards never treated with phosphite and 32 isolates from eight orchards treated with phosphite for 15 – 37 years. Four isolates had increased tolerance to phosphite and two were intermediately tolerant. These six isolates were from phosphite managed orchards and five were isolated from unhealthy trees.
To test if in vitro tolerance would be expressed in planta, three tolerant and three sensitive isolates were tested for their ability to colonise phosphite treated lupin (Lupinus angustifolius) roots. The tolerant isolates were more extensive colonisers of lupins treated with 5 and 10 g/L phosphite. In the presence of phosphite the tolerant isolates were able to produce more sporangia and release viable swimming zoospores more often.
Internationally important Phytophthora species from culture collections in New Zealand and the United States were assessed for sensitivity to phosphite and a subset of 32 isolates from the Berkeley collection was tested for mefenoxam sensitivity. The causal agent of kauri (Agathis australis) dieback, P. agathidicida, was found to be highly sensitive to phosphite while the possibly New Zealand native P. kernoviae, was found to be relatively tolerant. Intraspecific variation was found in lineages of P. ramorum, the causal agent of sudden oak death in the United States and the United Kingdom, NA2 was more tolerant than NA1. One P. megasperma isolate was more tolerant to mefenoxam compared to the other isolates tested. Of nine P. cinnamomi isolates from avocado orchards in southern California, one was relatively more tolerant to phosphite.
This study has shown phosphite tolerant isolates of P. cinnamomi have developed in phosphite managed avocado orchards in New Zealand and interspecific variation in sensitivity to fungicides is present across Phytophthora species in New Zealand and the United states.

text: Determining the risk of phosphite tolerance in Phytophthora species in New Zealand and the United States: a case study on the implications of long-term use of  phosphite to control Phytophthora cinnamomi in avocado (Persea americana) Phytophthora species cause many major plant disease epidemics worldwide, impacting horticulture, plant trade and production planting industries through the loss of plants and the costs arising from disease management. Phytophthora species have specialised structures which enhance their dispersal, survival and infection. These include motile swimming spores, caducous sporangia and resting spores. Nurseries and orchards use integrated management to control phytophthora diseases, including cultural practises, biological control, resistant host plants and chemical control. The use of fungicides is a very effective control method; however, Phytophthora species can develop resistance to fungicides after prolonged exposure. For example, phosphite tolerant isolates of Phytophthora cinnamomi have been found in avocado orchards in Australia and South Africa.
Phosphite is widely used in nurseries and orchards in New Zealand as a preventative and control treatment. The avocado industry largely relies on it for the control of avocado root rot caused by P. cinnamomi. Phosphite is most commonly applied to avocado trees using passive trunk injections but can also be applied as foliar sprays, soil drenches, high-pressure injections and capsule implants.
To assess phosphite tolerance of P. cinnamomi isolates from New Zealand avocado orchards, a high-throughput optical density (OD) assay was developed to measure mycelial growth inhibition in the presence of phosphite. The OD assay was used to screen 24 P. cinnamomi isolates from four orchards never treated with phosphite and 32 isolates from eight orchards treated with phosphite for 15 – 37 years. Four isolates had increased tolerance to phosphite and two were intermediately tolerant. These six isolates were from phosphite managed orchards and five were isolated from unhealthy trees.
To test if in vitro tolerance would be expressed in planta, three tolerant and three sensitive isolates were tested for their ability to colonise phosphite treated lupin (Lupinus angustifolius) roots. The tolerant isolates were more extensive colonisers of lupins treated with 5 and 10 g/L phosphite. In the presence of phosphite the tolerant isolates were able to produce more sporangia and release viable swimming zoospores more often.
Internationally important Phytophthora species from culture collections in New Zealand and the United States were assessed for sensitivity to phosphite and a subset of 32 isolates from the Berkeley collection was tested for mefenoxam sensitivity. The causal agent of kauri (Agathis australis) dieback, P. agathidicida, was found to be highly sensitive to phosphite while the possibly New Zealand native P. kernoviae, was found to be relatively tolerant. Intraspecific variation was found in lineages of P. ramorum, the causal agent of sudden oak death in the United States and the United Kingdom, NA2 was more tolerant than NA1. One P. megasperma isolate was more tolerant to mefenoxam compared to the other isolates tested. Of nine P. cinnamomi isolates from avocado orchards in southern California, one was relatively more tolerant to phosphite.
This study has shown phosphite tolerant isolates of P. cinnamomi have developed in phosphite managed avocado orchards in New Zealand and interspecific variation in sensitivity to fungicides is present across Phytophthora species in New Zealand and the United states.

year: 2018

title: Mapping Māori knowledge from the past to inform marine management futures

authors: Paul-Burke, Kura

subjects: mātauranga Māori

abstract: This article provides an overview of research which used mātauranga moana (Māori marine knowledge systems) to inform and assist Western science field research methods and surveys. Place-based Māori marine knowledge identified the traditional distribution range and sizing of taonga (culturally important) species in traditional coastal areas which had been fished and managed by consecutive generations of Māori; kina, Evechinus chloroticus, sea urchin; koura, Jasus edwardsii, red rock lobster; kūtai, Perna canaliculus, green lipped mussel; and pāua, Haliotis iris, abalone. This knowledge was then mapped and used as the baseline for sub-tidal marine science field research surveys. Findings from the transdisciplinary marine research was used to develop management actions to assist Māori and Government entities for improving, enhancing and safeguarding marine taonga species into the future. This article critically discusses and demonstrates the relevance and complementarity of mātauranga Māori and Western science, and the importance of kaupapa Māori strategies for empowering Māori collaboration and voices in marine research co-development, implementation and communication.

text: Mapping Māori knowledge from the past to inform marine management futures This article provides an overview of research which used mātauranga moana (Māori marine knowledge systems) to inform and assist Western science field research methods and surveys. Place-based Māori marine knowledge identified the traditional distribution range and sizing of taonga (culturally important) species in traditional coastal areas which had been fished and managed by consecutive generations of Māori; kina, Evechinus chloroticus, sea urchin; koura, Jasus edwardsii, red rock lobster; kūtai, Perna canaliculus, green lipped mussel; and pāua, Haliotis iris, abalone. This knowledge was then mapped and used as the baseline for sub-tidal marine science field research surveys. Findings from the transdisciplinary marine research was used to develop management actions to assist Māori and Government entities for improving, enhancing and safeguarding marine taonga species into the future. This article critically discusses and demonstrates the relevance and complementarity of mātauranga Māori and Western science, and the importance of kaupapa Māori strategies for empowering Māori collaboration and voices in marine research co-development, implementation and communication.

year: 2020