title: Development of a qPCR method to measure mitochondrial and genomic DNA damage with application to chemotherapy-induced DNA damage and cryopreserved cells

authors: Evans, Stephen Owen

subjects: LORD-Q

abstract: DNA damage quantitation assays such as the comet assay have focused on the measurement of total nuclear damage per cell. The adoption of PCR-based techniques to quantify DNA damage has enabled sequence- and organelle-specific assessment of DNA lesions. Here we report on an adaptation of a qPCR technique to assess DNA damage in nuclear and mitochondrial targets relative to control. Novel aspects of this assay include application of the assay to the Rotor-Gene platform with optimized DNA polymerase/fluorophore/primer set combination in a touchdown PCR protocol. Assay validation was performed using ultraviolet C radiation in A549 and THP1 cancer cell lines. A comparison was made to the comet assay applied to peripheral blood mononuclear cells, and an estimation of the effects of cryopreservation on ultraviolet C-induced DNA damage was carried out. Finally, dose responses for DNA damage were measured in peripheral blood mononuclear cells following exposure to the cytotoxic agents bleomycin and cisplatin. We show reproducible experimental outputs across the tested conditions and concordance with published findings with respect to mitochondrial and nuclear genotoxic susceptibilities. The application of this DNA damage assay to a wide range of clinical and laboratory-derived samples is both feasible and resource-efficient.

text: Development of a qPCR method to measure mitochondrial and genomic DNA damage with application to chemotherapy-induced DNA damage and cryopreserved cells DNA damage quantitation assays such as the comet assay have focused on the measurement of total nuclear damage per cell. The adoption of PCR-based techniques to quantify DNA damage has enabled sequence- and organelle-specific assessment of DNA lesions. Here we report on an adaptation of a qPCR technique to assess DNA damage in nuclear and mitochondrial targets relative to control. Novel aspects of this assay include application of the assay to the Rotor-Gene platform with optimized DNA polymerase/fluorophore/primer set combination in a touchdown PCR protocol. Assay validation was performed using ultraviolet C radiation in A549 and THP1 cancer cell lines. A comparison was made to the comet assay applied to peripheral blood mononuclear cells, and an estimation of the effects of cryopreservation on ultraviolet C-induced DNA damage was carried out. Finally, dose responses for DNA damage were measured in peripheral blood mononuclear cells following exposure to the cytotoxic agents bleomycin and cisplatin. We show reproducible experimental outputs across the tested conditions and concordance with published findings with respect to mitochondrial and nuclear genotoxic susceptibilities. The application of this DNA damage assay to a wide range of clinical and laboratory-derived samples is both feasible and resource-efficient.

year: 2016

title: Alum dosing effects on fish and aquatic invertebrates: Utuhina Stream 2024

authors: Ling, Nicholas

abstract: To reduce external phosphorus loading to Lake Rotorua, the Bay of Plenty Regional Council has been granted resource consent to dose the Utuhina Stream with alum (aluminium sulphate). Aluminium binds phosphorus, preventing its uptake by phytoplankton and thereby inhibiting growth. Aluminium forms monomeric species outside of circum-neutral pH (pH 68), and these species can disrupt osmoregulation and respiration of aquatic organisms. The alum dose rate to the Utuhina Stream is dependent on discharge, with a maximum application rate of 1 mg Al L-1. This report presents the results of an on-going assessment of the fish and aquatic macroinvertebrate communities of the Utuhina Stream for 2024.  Macroinvertebrates, fish and kōura (freshwater crayfish; Paranephrops planifrons) were sampled from one control (site 1) and two treatment (sites 2 & 3) reaches of the Utuhina Stream. Common bully (Gobiomorphus cotidianus) were the dominant species of the fish community, with juvenile trout and kōaro (Galaxias brevipinnis) also captured. Kōura were present at all sites but variable in abundance. There was a notable decline in the semiquantitative macroinvertebrate community (for soft-bottomed streams; MCI-sb) score at Sites 1 and 3 in 2024 compared to long-term site averages, with Site 1 declining into the ‘poor’ category. Interannual variation in MCI-sb scores is typical for these reaches of the Utuhina Stream and have been attributed to flood-related disturbances to stream bank morphology and in-stream vegetative cover (Ling 2021). Tissue aluminium concentrations were determined from common bully (flesh, gill, liver) and kōura (flesh, gill, hepatopancreas) by inductively coupled plasma mass spectrometry (ICP-MS). There was some evidence of aluminium bioaccumulation in the liver tissues of common bully at Site 2, but overall concentrations were similar to long-term averages and there were no notable differences for the same tissues between sites. Kōura tissue aluminium concentrations for 2024 were similar to long-term averages across all tissues and sites. A small number of individuals had elevated gill concentrations at Site 3 compared to long-term average concentrations, but this is unlikely to result in harmful physiological impacts. The data presented in this report further supports the conclusion by Tempero and Ling (2024) that alum dosing does not appear to have a notable effect on the fish and kōura community and that site and interannual differences appear to be due to hydrological and habitat variability.

text: Alum dosing effects on fish and aquatic invertebrates: Utuhina Stream 2024 To reduce external phosphorus loading to Lake Rotorua, the Bay of Plenty Regional Council has been granted resource consent to dose the Utuhina Stream with alum (aluminium sulphate). Aluminium binds phosphorus, preventing its uptake by phytoplankton and thereby inhibiting growth. Aluminium forms monomeric species outside of circum-neutral pH (pH 68), and these species can disrupt osmoregulation and respiration of aquatic organisms. The alum dose rate to the Utuhina Stream is dependent on discharge, with a maximum application rate of 1 mg Al L-1. This report presents the results of an on-going assessment of the fish and aquatic macroinvertebrate communities of the Utuhina Stream for 2024.  Macroinvertebrates, fish and kōura (freshwater crayfish; Paranephrops planifrons) were sampled from one control (site 1) and two treatment (sites 2 & 3) reaches of the Utuhina Stream. Common bully (Gobiomorphus cotidianus) were the dominant species of the fish community, with juvenile trout and kōaro (Galaxias brevipinnis) also captured. Kōura were present at all sites but variable in abundance. There was a notable decline in the semiquantitative macroinvertebrate community (for soft-bottomed streams; MCI-sb) score at Sites 1 and 3 in 2024 compared to long-term site averages, with Site 1 declining into the ‘poor’ category. Interannual variation in MCI-sb scores is typical for these reaches of the Utuhina Stream and have been attributed to flood-related disturbances to stream bank morphology and in-stream vegetative cover (Ling 2021). Tissue aluminium concentrations were determined from common bully (flesh, gill, liver) and kōura (flesh, gill, hepatopancreas) by inductively coupled plasma mass spectrometry (ICP-MS). There was some evidence of aluminium bioaccumulation in the liver tissues of common bully at Site 2, but overall concentrations were similar to long-term averages and there were no notable differences for the same tissues between sites. Kōura tissue aluminium concentrations for 2024 were similar to long-term averages across all tissues and sites. A small number of individuals had elevated gill concentrations at Site 3 compared to long-term average concentrations, but this is unlikely to result in harmful physiological impacts. The data presented in this report further supports the conclusion by Tempero and Ling (2024) that alum dosing does not appear to have a notable effect on the fish and kōura community and that site and interannual differences appear to be due to hydrological and habitat variability.

year: 2025

title: A community psychology-based evaluation of the you good? You Good! program delivered by Jade speaks up educational trust in Aotearoa. Empowering tamariki emotional literacy, supporting mental health and encouraging well-being.

authors: Webbon, Abigail

abstract: This evaluation was completed for Jade Speaks Up (JSU) Educational Trust to provide feedback from school teachers and whānau who were involved in the You Good? You Good! (YG?YG!) program, delivered by JSU Educational Trust to schools across Aotearoa. The evaluation goal was to collate the narratives of participating school staff, whānau, and JSU Educational Trust trainers, contrasting experiences and perspectives to determine program effectiveness.
This program is for schools, management, and teachers who work with tamariki, to help teachers and students understand anxiety and develop skills and strategies to have self-agency. JSU Educational Trust developed YG?YG! in 2021, and in 2022 the YG?YG! pilot program was implemented.
This evaluation had three research objectives: 1) Identify and collate the experiences of school staff and other key stakeholders when implementing the YG?YG!, 2) Identify the strengths and barriers to inclusion for tamariki in schools where the program was delivered, and 3) Draw on the stakeholder narratives to assess the program's ability to achieve its goal: enable tamariki to be more agentic in responding to early signs of anxiety in themselves and others.
Information for this evaluation was collected from four sources: 1) Focus groups with JSU trainers, 2) Interviews with school staff, 3) Online surveys with school staff, 4) Archival supervision, and 5) Literature review. Data collected was interpreted through the use of thematic analysis, informed by Community Psychology principles, and the “Six C” framework, guided by He Awa Whiria.
Thematic analysis was completed by coding themes and categories from data sources. The findings from this empirical research engagement informed the analysis and discussion on program effectiveness. 
The findings and analysis found that forming a program design and approach informed with respect for the community’s needs and world-views can enable recognition, awareness and inclusion. A way for the program to begin to do so is to collaborate with communities.
Including researchers from a local community can enable empowerment, build social power, and create responses that promote the needs of the involved communities. The program can champion the field and those they work with by being accountable and enabling transparency.
The program can take an inclusive approach to sustaining its relationships, content relevance, and development for tamariki emotional well-being. The program can work on cultivating and empowering participants to be active partners, enabling the program to be shaped with equity in mind.
Key to program efficacy is acknowledging change and encompassing adaptation. The way to do this is to consider local health issues driven by who experience them, elevating the voice of the community.
Change and need cannot be determined without considering the voice of involved communities. By taking participants feedback, the program can create change for tamariki mental health, whilst acknowledging system and structural barriers, striving for community competence.
Importance lies within the work the program does to enable and support those affected by external factors. The program can acknowledge these challenges, and provide hope that community competence can be achieved.
To conclude, recommendations are presented for JSU Educational Trust to consider.

text: A community psychology-based evaluation of the you good? You Good! program delivered by Jade speaks up educational trust in Aotearoa. Empowering tamariki emotional literacy, supporting mental health and encouraging well-being. This evaluation was completed for Jade Speaks Up (JSU) Educational Trust to provide feedback from school teachers and whānau who were involved in the You Good? You Good! (YG?YG!) program, delivered by JSU Educational Trust to schools across Aotearoa. The evaluation goal was to collate the narratives of participating school staff, whānau, and JSU Educational Trust trainers, contrasting experiences and perspectives to determine program effectiveness.
This program is for schools, management, and teachers who work with tamariki, to help teachers and students understand anxiety and develop skills and strategies to have self-agency. JSU Educational Trust developed YG?YG! in 2021, and in 2022 the YG?YG! pilot program was implemented.
This evaluation had three research objectives: 1) Identify and collate the experiences of school staff and other key stakeholders when implementing the YG?YG!, 2) Identify the strengths and barriers to inclusion for tamariki in schools where the program was delivered, and 3) Draw on the stakeholder narratives to assess the program's ability to achieve its goal: enable tamariki to be more agentic in responding to early signs of anxiety in themselves and others.
Information for this evaluation was collected from four sources: 1) Focus groups with JSU trainers, 2) Interviews with school staff, 3) Online surveys with school staff, 4) Archival supervision, and 5) Literature review. Data collected was interpreted through the use of thematic analysis, informed by Community Psychology principles, and the “Six C” framework, guided by He Awa Whiria.
Thematic analysis was completed by coding themes and categories from data sources. The findings from this empirical research engagement informed the analysis and discussion on program effectiveness. 
The findings and analysis found that forming a program design and approach informed with respect for the community’s needs and world-views can enable recognition, awareness and inclusion. A way for the program to begin to do so is to collaborate with communities.
Including researchers from a local community can enable empowerment, build social power, and create responses that promote the needs of the involved communities. The program can champion the field and those they work with by being accountable and enabling transparency.
The program can take an inclusive approach to sustaining its relationships, content relevance, and development for tamariki emotional well-being. The program can work on cultivating and empowering participants to be active partners, enabling the program to be shaped with equity in mind.
Key to program efficacy is acknowledging change and encompassing adaptation. The way to do this is to consider local health issues driven by who experience them, elevating the voice of the community.
Change and need cannot be determined without considering the voice of involved communities. By taking participants feedback, the program can create change for tamariki mental health, whilst acknowledging system and structural barriers, striving for community competence.
Importance lies within the work the program does to enable and support those affected by external factors. The program can acknowledge these challenges, and provide hope that community competence can be achieved.
To conclude, recommendations are presented for JSU Educational Trust to consider.

year: 2024-11-14