LowTempGAL: a highly responsive low temperature-inducible GAL system in Saccharomyces cerevisiae.

Temperature is an important control factor for biologics biomanufacturing in precision fermentation. Here, we explored a highly responsive low temperature-inducible genetic system (LowTempGAL) in the model yeast Saccharomyces cerevisiae. Two temperature biosensors, a heat-inducible degron and a heat-inducible protein aggregation domain, were used to regulate the GAL activator Gal4p, rendering the leaky LowTempGAL systems. Boolean-type induction was achieved by implementing a second-layer control through low-temperature-mediated repression on GAL repressor gene GAL80, but suffered delayed response to low-temperature triggers and a weak response at 30°C. Application potentials were validated for protein and small molecule production. Proteomics analysis suggested that residual Gal80p and Gal4p insufficiency caused suboptimal induction. 'Turbo' mechanisms were engineered through incorporating a basal Gal4p expression and a galactose-independent Gal80p-supressing Gal3p mutant (Gal3Cp). Varying Gal3Cp configurations, we deployed the LowTempGAL systems capable for a rapid stringent high-level induction upon the shift from a high temperature (37-33°C) to a low temperature (≤30°C). Overall, we present a synthetic biology procedure that leverages 'leaky' biosensors to deploy highly responsive Boolean-type genetic circuits. The key lies in optimisation of the intricate layout of the multi-factor system. The LowTempGAL systems may be applicable in non-conventional yeast platforms for precision biomanufacturing.

The Development of Cortical Responses to the Integration of Audiovisual Speech in Infancy.

In adults, the integration of audiovisual speech elicits specific higher (super-additive) or lower (sub-additive) cortical responses when compared to the responses to unisensory stimuli. Although there is evidence that the fronto-temporal network is active during perception of audiovisual speech in infancy, the development of fronto-temporal responses to audiovisual integration remains unknown. In the current study, 5-month-olds and 10-month-olds watched bimodal (audiovisual) and alternating unimodal (auditory + visual) syllables. In this context we use alternating unimodal to denote alternating auditory and visual syllables that are perceived as separate syllables by adults. Using fNIRS we measured responses over large cortical areas including the inferior frontal and superior temporal regions. We identified channels showing different responses to bimodal than alternating unimodal condition and used multivariate pattern analysis (MVPA) to decode patterns of cortical responses to bimodal (audiovisual) and alternating unimodal (auditory + visual) speech. Results showed that in both age groups integration elicits cortical responses consistent with both super- and sub-additive responses in the fronto-temporal cortex. The univariate analyses revealed that between 5 and 10 months spatial distribution of these responses becomes increasingly focal. MVPA correctly classified responses at 5 months, with key input from channels located in the inferior frontal and superior temporal channels of the right hemisphere. However, MVPA classification was not successful at 10 months, suggesting a potential cortical re-organisation of audiovisual speech perception at this age. These results show the complex and non-gradual development of the cortical responses to integration of congruent audiovisual speech in infancy.

Toll-Like Receptor 4 in Pain: Bridging Molecules-to-Cells-to-Systems.

Pain impacts the lives of billions of people around the world - both directly and indirectly. It is complex and transcends beyond an unpleasant sensory experience to encompass emotional experiences. To date, there are no successful treatments for sufferers of chronic pain. Although opioids do not provide any benefit to chronic pain sufferers, they are still prescribed, often resulting in more complications such as hyperalgesia and dependence. In order to develop effective and safe medications to manage, and perhaps even treat pain, it is important to evaluate novel contributors to pain pathologies. As such, in this chapter we review the role of Toll-like receptor 4, a receptor of the innate immune system, that continues to gain substantial attention in the field of pain research. Positioned in the nexus of the neuro and immune systems, TLR4 may provide one of the missing pieces in understanding the complexities of pain. Here we consider how TLR4 enables a mechanistical understanding of pain as a multidimensional biopsychosocial state from molecules to cells to systems and back again.

Cytoplasmic delivery of quantum dots via microelectrophoresis technique.

Nanoparticles with specific properties and functions have been developed for various biomedical research applications, such as in vivo and in vitro sensors, imaging agents and delivery vehicles of therapeutics. The development of an effective delivery method of nanoparticles into the intracellular environment is challenging and success in this endeavor would be beneficial to many biological studies. Here, the well-established microelectrophoresis technique was applied for the first time to deliver nanoparticles into living cells. An optimal protocol was explored to prepare semiconductive quantum dots suspensions having high monodispersity with average hydrodynamic diameter of 13.2-35.0 nm. Micropipettes were fabricated to have inner tip diameters of approximately 200 nm that are larger than quantum dots for ejection but less than 500 nm to minimize damage to the cell membrane. We demonstrated the successful delivery of quantum dots via small electrical currents (-0.2 nA) through micropipettes into the cytoplasm of living human embryonic kidney cells (roughly 20-30 µm in length) using microelectrophoresis technique. This method is promising as a simple and general strategy for delivering a variety of nanoparticles into the cellular environment.

Beatboxers and Guitarists Engage Sensorimotor Regions Selectively When Listening to the Instruments They can Play.

Studies of classical musicians have demonstrated that expertise modulates neural responses during auditory perception. However, it remains unclear whether such expertise-dependent plasticity is modulated by the instrument that a musician plays. To examine whether the recruitment of sensorimotor regions during music perception is modulated by instrument-specific experience, we studied nonclassical musicians-beatboxers, who predominantly use their vocal apparatus to produce sound, and guitarists, who use their hands. We contrast fMRI activity in 20 beatboxers, 20 guitarists, and 20 nonmusicians as they listen to novel beatboxing and guitar pieces. All musicians show enhanced activity in sensorimotor regions (IFG, IPC, and SMA), but only when listening to the musical instrument they can play. Using independent component analysis, we find expertise-selective enhancement in sensorimotor networks, which are distinct from changes in attentional networks. These findings suggest that long-term sensorimotor experience facilitates access to the posterodorsal "how" pathway during auditory processing.

How Auditory Experience Differentially Influences the Function of Left and Right Superior Temporal Cortices.

To investigate how hearing status, sign language experience, and task demands influence functional responses in the human superior temporal cortices (STC) we collected fMRI data from deaf and hearing participants (male and female), who either acquired sign language early or late in life. Our stimuli in all tasks were pictures of objects. We varied the linguistic and visuospatial processing demands in three different tasks that involved decisions about (1) the sublexical (phonological) structure of the British Sign Language (BSL) signs for the objects, (2) the semantic category of the objects, and (3) the physical features of the objects.Neuroimaging data revealed that in participants who were deaf from birth, STC showed increased activation during visual processing tasks. Importantly, this differed across hemispheres. Right STC was consistently activated regardless of the task whereas left STC was sensitive to task demands. Significant activation was detected in the left STC only for the BSL phonological task. This task, we argue, placed greater demands on visuospatial processing than the other two tasks. In hearing signers, enhanced activation was absent in both left and right STC during all three tasks. Lateralization analyses demonstrated that the effect of deafness was more task-dependent in the left than the right STC whereas it was more task-independent in the right than the left STC. These findings indicate how the absence of auditory input from birth leads to dissociable and altered functions of left and right STC in deaf participants.SIGNIFICANCE STATEMENT Those born deaf can offer unique insights into neuroplasticity, in particular in regions of superior temporal cortex (STC) that primarily respond to auditory input in hearing people. Here we demonstrate that in those deaf from birth the left and the right STC have altered and dissociable functions. The right STC was activated regardless of demands on visual processing. In contrast, the left STC was sensitive to the demands of visuospatial processing. Furthermore, hearing signers, with the same sign language experience as the deaf participants, did not activate the STCs. Our data advance current understanding of neural plasticity by determining the differential effects that hearing status and task demands can have on left and right STC function.

Computer simulations show that Neanderthal facial morphology represents adaptation to cold and high energy demands, but not heavy biting.

Three adaptive hypotheses have been forwarded to explain the distinctive Neanderthal face: (i) an improved ability to accommodate high anterior bite forces, (ii) more effective conditioning of cold and/or dry air and, (iii) adaptation to facilitate greater ventilatory demands. We test these hypotheses using three-dimensional models of Neanderthals, modern humans, and a close outgroup (Homo heidelbergensis), applying finite-element analysis (FEA) and computational fluid dynamics (CFD). This is the most comprehensive application of either approach applied to date and the first to include both. FEA reveals few differences between H. heidelbergensis, modern humans, and Neanderthals in their capacities to sustain high anterior tooth loadings. CFD shows that the nasal cavities of Neanderthals and especially modern humans condition air more efficiently than does that of H. heidelbergensis, suggesting that both evolved to better withstand cold and/or dry climates than less derived Homo We further find that Neanderthals could move considerably more air through the nasal pathway than could H. heidelbergensis or modern humans, consistent with the propositions that, relative to our outgroup Homo, Neanderthal facial morphology evolved to reflect improved capacities to better condition cold, dry air, and, to move greater air volumes in response to higher energetic requirements.

Distinct processing of ambiguous speech in people with non-clinical auditory verbal hallucinations.

Auditory verbal hallucinations (hearing voices) are typically associated with psychosis, but a minority of the general population also experience them frequently and without distress. Such 'non-clinical' experiences offer a rare and unique opportunity to study hallucinations apart from confounding clinical factors, thus allowing for the identification of symptom-specific mechanisms. Recent theories propose that hallucinations result from an imbalance of prior expectation and sensory information, but whether such an imbalance also influences auditory-perceptual processes remains unknown. We examine for the first time the cortical processing of ambiguous speech in people without psychosis who regularly hear voices. Twelve non-clinical voice-hearers and 17 matched controls completed a functional magnetic resonance imaging scan while passively listening to degraded speech ('sine-wave' speech), that was either potentially intelligible or unintelligible. Voice-hearers reported recognizing the presence of speech in the stimuli before controls, and before being explicitly informed of its intelligibility. Across both groups, intelligible sine-wave speech engaged a typical left-lateralized speech processing network. Notably, however, voice-hearers showed stronger intelligibility responses than controls in the dorsal anterior cingulate cortex and in the superior frontal gyrus. This suggests an enhanced involvement of attention and sensorimotor processes, selectively when speech was potentially intelligible. Altogether, these behavioural and neural findings indicate that people with hallucinatory experiences show distinct responses to meaningful auditory stimuli. A greater weighting towards prior knowledge and expectation might cause non-veridical auditory sensations in these individuals, but it might also spontaneously facilitate perceptual processing where such knowledge is required. This has implications for the understanding of hallucinations in clinical and non-clinical populations, and is consistent with current 'predictive processing' theories of psychosis.

Richer histories for more relevant policies: 42 years of tree cover loss and gain in Southeast Sulawesi, Indonesia.

Understandings of contemporary forest cover loss are critical for policy but have come at the expense of long-term, multidirectional analyses of land cover change. This is a critical gap given (i) profound reconfigurations in land use and land control over the past several decades and (ii) evidence of widespread 'woodland resurgence' throughout the tropics. In this study, we argue that recent advancements within the field of land change science provide new opportunities to address this gap. In turn, we suggest that multidecadal and multidirectional analyses of land cover change can facilitate richer social analyses of land cover change and more relevant conservation policies and practice. Our argument is grounded in a case study from Southeast Sulawesi, Indonesia. Using a novel analytical platform, Google Earth Engine, and open access to high-quality Landsat data, we map land cover change in Southeast Sulawesi, Indonesia, from 1972 to 2014. We find that tree cover loss constitutes the single largest net change over the period 1972-2014 but that gross rates of tree cover gain were three times higher than gross loss rates from 1972 to 1995 and equivalent to loss rates from 1995 to 2014. We suggest the smallholder tree crop economy likely produced both forest loss and Imperata grassland restoration in this region. This case points to the need to expand rather than collapse the baselines used to study carbon and biodiversity change in tropical regions. It also demonstrates the possible utility of applying such methods to other regions.

Getting the Cocktail Party Started: Masking Effects in Speech Perception.

Spoken conversations typically take place in noisy environments, and different kinds of masking sounds place differing demands on cognitive resources. Previous studies, examining the modulation of neural activity associated with the properties of competing sounds, have shown that additional speech streams engage the superior temporal gyrus. However, the absence of a condition in which target speech was heard without additional masking made it difficult to identify brain networks specific to masking and to ascertain the extent to which competing speech was processed equivalently to target speech. In this study, we scanned young healthy adults with continuous fMRI, while they listened to stories masked by sounds that differed in their similarity to speech. We show that auditory attention and control networks are activated during attentive listening to masked speech in the absence of an overt behavioral task. We demonstrate that competing speech is processed predominantly in the left hemisphere within the same pathway as target speech but is not treated equivalently within that stream and that individuals who perform better in speech in noise tasks activate the left mid-posterior superior temporal gyrus more. Finally, we identify neural responses associated with the onset of sounds in the auditory environment; activity was found within right lateralized frontal regions consistent with a phasic alerting response. Taken together, these results provide a comprehensive account of the neural processes involved in listening in noise.

Hierarchical Organization of Auditory and Motor Representations in Speech Perception: Evidence from Searchlight Similarity Analysis.

How humans extract the identity of speech sounds from highly variable acoustic signals remains unclear. Here, we use searchlight representational similarity analysis (RSA) to localize and characterize neural representations of syllables at different levels of the hierarchically organized temporo-frontal pathways for speech perception. We asked participants to listen to spoken syllables that differed considerably in their surface acoustic form by changing speaker and degrading surface acoustics using noise-vocoding and sine wave synthesis while we recorded neural responses with functional magnetic resonance imaging. We found evidence for a graded hierarchy of abstraction across the brain. At the peak of the hierarchy, neural representations in somatomotor cortex encoded syllable identity but not surface acoustic form, at the base of the hierarchy, primary auditory cortex showed the reverse. In contrast, bilateral temporal cortex exhibited an intermediate response, encoding both syllable identity and the surface acoustic form of speech. Regions of somatomotor cortex associated with encoding syllable identity in perception were also engaged when producing the same syllables in a separate session. These findings are consistent with a hierarchical account of how variable acoustic signals are transformed into abstract representations of the identity of speech sounds.

Feel the Noise: Relating Individual Differences in Auditory Imagery to the Structure and Function of Sensorimotor Systems.

Humans can generate mental auditory images of voices or songs, sometimes perceiving them almost as vividly as perceptual experiences. The functional networks supporting auditory imagery have been described, but less is known about the systems associated with interindividual differences in auditory imagery. Combining voxel-based morphometry and fMRI, we examined the structural basis of interindividual differences in how auditory images are subjectively perceived, and explored associations between auditory imagery, sensory-based processing, and visual imagery. Vividness of auditory imagery correlated with gray matter volume in the supplementary motor area (SMA), parietal cortex, medial superior frontal gyrus, and middle frontal gyrus. An analysis of functional responses to different types of human vocalizations revealed that the SMA and parietal sites that predict imagery are also modulated by sound type. Using representational similarity analysis, we found that higher representational specificity of heard sounds in SMA predicts vividness of imagery, indicating a mechanistic link between sensory- and imagery-based processing in sensorimotor cortex. Vividness of imagery in the visual domain also correlated with SMA structure, and with auditory imagery scores. Altogether, these findings provide evidence for a signature of imagery in brain structure, and highlight a common role of perceptual-motor interactions for processing heard and internally generated auditory information.

Distinct neural systems recruited when speech production is modulated by different masking sounds.

When talkers speak in masking sounds, their speech undergoes a variety of acoustic and phonetic changes. These changes are known collectively as the Lombard effect. Most behavioural research and neuroimaging research in this area has concentrated on the effect of energetic maskers such as white noise on Lombard speech. Previous fMRI studies have argued that neural responses to speaking in noise are driven by the quality of auditory feedback-that is, the audibility of the speaker's voice over the masker. However, we also frequently produce speech in the presence of informational maskers such as another talker. Here, speakers read sentences over a range of maskers varying in their informational and energetic content: speech, rotated speech, speech modulated noise, and white noise. Subjects also spoke in quiet and listened to the maskers without speaking. When subjects spoke in masking sounds, their vocal intensity increased in line with the energetic content of the masker. However, the opposite pattern was found neurally. In the superior temporal gyrus, activation was most strongly associated with increases in informational, rather than energetic, masking. This suggests that the neural activations associated with speaking in noise are more complex than a simple feedback response.

Greenhouse gas mitigation on marginal land: a quantitative review of the relative benefits of forest recovery versus biofuel production.

Decisions concerning future land-use/land cover change stand at the forefront of ongoing debates on how to best mitigate climate change. In this study, we compare the greenhouse gas (GHG) mitigation value over a 30-year time frame for a range of forest recovery and biofuel production scenarios on abandoned agricultural land. Carbon sequestration in recovering forests is estimated based on a statistical analysis of tropical and temperate studies on marginal land. GHGs offset by biofuel production are analyzed for five different production pathways. We find that forest recovery is superior to low-yielding biofuel production scenarios such as oil palm and corn. Biofuel production scenarios with high yields, such as sugarcane or high-yielding energy grasses, can be comparable or superior to natural forest succession and to reforestation in some cases. This result stands in contrast to previous research suggesting that restoring degraded ecosystems to their native state is generally superior to agricultural production in terms of GHG mitigation. Further work is needed on carbon stock changes in forests, soil carbon dynamics, and bioenergy crop production on degraded/abandoned agricultural land. This finding also emphasizes the need to consider the full range of social, economic, and ecological consequences of land-use policies.

Musicians and non-musicians are equally adept at perceiving masked speech.

There is much interest in the idea that musicians perform better than non-musicians in understanding speech in background noise. Research in this area has often used energetic maskers, which have their effects primarily at the auditory periphery. However, masking interference can also occur at more central auditory levels, known as informational masking. This experiment extends existing research by using multiple maskers that vary in their informational content and similarity to speech, in order to examine differences in perception of masked speech between trained musicians (n = 25) and non-musicians (n = 25). Although musicians outperformed non-musicians on a measure of frequency discrimination, they showed no advantage in perceiving masked speech. Further analysis revealed that non-verbal IQ, rather than musicianship, significantly predicted speech reception thresholds in noise. The results strongly suggest that the contribution of general cognitive abilities needs to be taken into account in any investigations of individual variability for perceiving speech in noise.

Evolutionary conservation of Nkx2.5 autoregulation in the second heart field.

The cardiac homeobox gene Nkx2.5 plays a key and dosage-sensitive role in the differentiation of outflow tract and right ventricle from progenitors of the second heart field (SHF) and Nkx2.5 mutation is strongly associated with human outflow tract congenital heart disease (OFT CHD). Therefore defining the regulatory mechanisms controlling Nkx2.5 expression in SHF populations serves an important function in understanding the etiology of complex CHD. Through a comparative analysis of regulatory elements controlling SHF expression of Nkx2.5 in the chicken and mouse, we have found evidence that Nkx2.5 autoregulation is important for maintaining Nkx2.5 expression during SHF differentiation in both species. However the mechanism of Nkx2.5 maintenance differs between placental mammals and non-mammalian vertebrates: in chick Nkx2.5 binds directly to a genomic enhancer element that is required to maintain Nkx2.5 expression in the SHF. In addition, it is likely that this is true in other non-mammalian vertebrates given that they possess a similar genomic organization. By contrast, in placental mammals, Nkx2.5 autoregulation in the SHF functions indirectly through Mef2c. These data underscore a tight relationship in mammals between Nkx2.5 and Mef2c in SHF transcriptional regulation, and highlight the potential for evolutionary cis-regulatory analysis to identify core, conserved components of the gene networks controlling heart development.

No clear evidence of a difference between individuals who self-report an absence of auditory imagery and typical imagers on auditory imagery tasks.

Aphantasia is characterised by the inability to create mental images in one's mind. Studies investigating impairments in imagery typically focus on the visual domain. However, it is possible to generate many different forms of imagery including imagined auditory, kinesthetic, tactile, motor, taste and other experiences. Recent studies show that individuals with aphantasia report a lack of imagery in modalities, other than vision, including audition. However, to date, no research has examined whether these reductions in self-reported auditory imagery are associated with decrements in tasks that require auditory imagery. Understanding the extent to which visual and auditory imagery deficits co-occur can help to better characterise the core deficits of aphantasia and provide an alternative perspective on theoretical debates on the extent to which imagery draws on modality-specific or modality-general processes. In the current study, individuals that self-identified as being aphantasic and matched control participants with typical imagery performed two tasks: a musical pitch-based imagery and voice-based categorisation task. The majority of participants with aphantasia self-reported significant deficits in both auditory and visual imagery. However, we did not find a concomitant decrease in performance on tasks which require auditory imagery, either in the full sample or only when considering those participants that reported significant deficits in both domains. These findings are discussed in relation to the mechanisms that might obscure observation of imagery deficits in auditory imagery tasks in people that report reduced auditory imagery.

Integration of Yeast Episomal/Integrative Plasmid Causes Genotypic and Phenotypic Diversity and Improved Sesquiterpene Production in Metabolically Engineered Saccharomyces cerevisiae.

The variability in phenotypic outcomes among biological replicates in engineered microbial factories presents a captivating mystery. Establishing the association between phenotypic variability and genetic drivers is important to solve this intricate puzzle. We applied a previously developed auxin-inducible depletion of hexokinase 2 as a metabolic engineering strategy for improved nerolidol production in Saccharomyces cerevisiae, and biological replicates exhibit a dichotomy in nerolidol production of either 3.5 or 2.5 g L-1 nerolidol. Harnessing Oxford Nanopore's long-read genomic sequencing, we reveal a potential genetic cause─the chromosome integration of a 2µ sequence-based yeast episomal plasmid, encoding the expression cassettes for nerolidol synthetic enzymes. This finding was reinforced through chromosome integration revalidation, engineering nerolidol and valencene production strains, and generating a diverse pool of yeast clones, each uniquely fingerprinted by gene copy numbers, plasmid integrations, other genomic rearrangements, protein expression levels, growth rate, and target product productivities. Τhe best clone in two strains produced 3.5 g L-1 nerolidol and ∼0.96 g L-1 valencene. Comparable genotypic and phenotypic variations were also generated through the integration of a yeast integrative plasmid lacking 2µ sequences. Our work shows that multiple factors, including plasmid integration status, subchromosomal location, gene copy number, sesquiterpene synthase expression level, and genome rearrangement, together play a complicated determinant role on the productivities of sesquiterpene product. Integration of yeast episomal/integrative plasmids may be used as a versatile method for increasing the diversity and optimizing the efficiency of yeast cell factories, thereby uncovering metabolic control mechanisms.

Glyphosate-based herbicide has contrasting effects on prey capture by two co-occurring wolf spider species.

Anthropogenic substances have the potential to affect animal behavior either because they present a novel stimulus or because they interfere with natural chemical communication pathways. Such shifts can alter the dynamic between predators and potential prey, which might affect population success as well as the strength of food web linkages. We examined the foraging of two wolf spiders, Tigrosa helluo and Pardosa milvina (Araneae, Lycosidae), that are abundant in agroecosystems where they are routinely exposed to herbicides. We tested the hypothesis that the presence of a commercial formulation of a glyphosate-based herbicide would affect the prey capture behavior of these two wolf spiders. We tested the larger Tigrosa foraging on Pardosa or crickets (Acheta domesticus) and the smaller Pardosa foraging on crickets. Tigrosa subdued crickets more quickly and with fewer lunges than it took them to capture Pardosa. The presence of herbicide allowed Tigrosa to orient toward and capture both prey species more quickly but it did not affect the number of lunges required to subdue either prey. Herbicide did not affect the timing of prey capture for Pardosa but it did cause them to use more lunges in the process. Thus, herbicide had contrasting effects on foraging behavior of these two agrobiont predators, which means that it could shift the direction and strength of food web linkages in complex ways.

Development of a methodology for the standardisation and improvement of 'Smartphone' photography of patterned bruises and other cutaneous injuries.

Human bite-mark analyses can play a prominent role in forensic case investigations, including those involving sexual assault. High-quality photographs routinely secure a link between a bite-mark and an individual's dentition. Access to around the clock forensic photography, however, is often limited, resulting in delay and/or missed opportunities to record valuable evidence. The emergence of Smartphone high-quality photographic technology now provides a previously unimagined opportunity to gather timely forensic photographic evidence. Problems can arise, however, due to the relatively poor quality of the photographs, as a result of many of those taking photographs having received little or no forensic photography training. This study compares unassisted photography with assisted photography, by a specifically developed camera application (App), to provide a standardised method for taking forensic photographs. An App, written in Java, was hosted on the Google Android Operating System, on a Samsung Galaxy SII Smartphone. Twenty-four volunteers participated in a study to photograph a pseudo bite-mark using three methods, (1) unassisted (as a control), (2) assisted by an ABFO No.2 right-angled photographic reference scale and (3) assisted by the App. The App, method (3), was shown to consistently outperform methods (1) and (2), demonstrating greater standardisation and precision (p<0.001). Analysis of the data showed the extent to which acquiring an accurate photograph depends on the image being orthogonal to the camera. It appears likely that the relatively inaccurate photographs acquired by methods (1) and (2), were as a result of deviation from the plane, orthogonal to the bite-mark. Therefore, the App was successful in ensuring that the camera was both orthogonal and at an appropriate distance, relative to the bite-mark. Thus, the App enhanced the abilities of non-experts to acquire more accurate photographs and created the potential to significantly improve the quality of forensic photographs.