Climate change, human health, and resilience in the Holocene.

Climate change is an indisputable threat to human health, especially for societies already confronted with rising social inequality, political and economic uncertainty, and a cascade of concurrent environmental challenges. Archaeological data about past climate and environment provide an important source of evidence about the potential challenges humans face and the long-term outcomes of alternative short-term adaptive strategies. Evidence from well-dated archaeological human skeletons and mummified remains speaks directly to patterns of human health over time through changing circumstances. Here, we describe variation in human epidemiological patterns in the context of past rapid climate change (RCC) events and other periods of past environmental change. Case studies confirm that human communities responded to environmental changes in diverse ways depending on historical, sociocultural, and biological contingencies. Certain factors, such as social inequality and disproportionate access to resources in large, complex societies may influence the probability of major sociopolitical disruptions and reorganizations-commonly known as "collapse." This survey of Holocene human-environmental relations demonstrates how flexibility, variation, and maintenance of Indigenous knowledge can be mitigating factors in the face of environmental challenges. Although contemporary climate change is more rapid and of greater magnitude than the RCC events and other environmental changes we discuss here, these lessons from the past provide clarity about potential priorities for equitable, sustainable development and the constraints of modernity we must address.

3D printed chamber for live cell imaging on an upright epifluorescence microscope.

Live cell imaging is a standard technique in experimental biology that enables the observation of isolated cells and tissue slices in real time; and the testing of cellular responses to changes in buffer composition. However, most live cell imaging devices require the use of dedicated microscopes and/or specialized stage adaptors, and come at a reasonably high cost. We employed 3D printing technology to create a low-cost imaging chamber with side ports to exchange fluids, to be used on upright microscopes. The chamber increased the functionality of a standard upright epifluorescent microscope to allow dynamic, real-time calcium imaging of cultured hypothalamic astrocytes from mice, and to test the effects of ATP stimulation upon calcium signaling. It was also used on slices obtained from mouse brain using a brain matrix slicer. The advantages of this chamber include a very simple design that can be used with upright epifluorescence microscopes, does not require any special stage adaptor, and includes ports to permit fluid exchange during imaging. This chamber is ideal for educational settings with undergraduate laboratories that do not have access to dedicated inverted fluorescent microscopes for tissue culture experiments.

Rho-family G-proteins are required for the recovery of traumatized hair bundle mechanoreceptors in the sea anemone, Nematostella vectensis.

Immersing anemones in calcium-free seawater disorganizes hair bundle mechanoreceptors on tentacles of sea anemones while causing a loss of vibration sensitivity. Remarkably, anemone hair bundles recover after being returned to calcium-containing seawater. Reorganization of actin in stereocilia likely follows during the recovery of normal morphology of hair bundles after such immersion. Previous studies have reported that Rho G-proteins are located in the stereocilia of hair bundles in sea anemones where they participate in polymerizing actin in stereocilia upon activation of specific chemoreceptors. We here find that immersing anemones in calcium-free seawater significantly reduces the abundance of hair bundles. A partial recovery of abundance of hair bundles occurs within 3 h post-immersion, but a full recovery of abundance does not occur even 6 h after specimens are returned to calcium-containing seawater. Anemones recovering from immersion in calcium-free seawater feature hair bundles that are significantly wider at their tips than in controls. The hair bundles subsequently narrow at their tips, becoming comparable to those of untreated controls within 6 h. Stereocilia of hair bundles are significantly longer in experimental animals than in controls at 2 h of recovery before shortening to lengths comparable to untreated controls at 6 h. In the presence of Rho inhibitors, the recovery in abundance of hair bundles through 6 h is delayed or inhibited. Likewise, in the presence of Rho inhibitors, stereocilia fail to significantly elongate within 2 h of recovery. These data suggest that Rho G-proteins participate in the normal recovery of abundance and recovery of normal morphology of experimentally damaged hair bundle mechanoreceptors.

Evidence for two populations of hair bundles in the sea anemone, Nematostella vectensis.

Cytochalasin D (CD) was employed to disrupt F-actin within stereocilia of anemone hair bundles. CD treatment decreases the abundance of hair bundles (by 85%) while significantly impairing predation. The remaining hair bundles are 'CD-resistant.' Surprisingly, the morphology and F-actin content of resistant hair bundles are comparable to those of untreated controls. However, the resistant hair bundles fail to respond normally to the N-acetylated sugar, NANA, by elongating. Instead, they remain at resting length. Immediately after CD treatment, when only CD-resistant hair bundles are present, nematocyst discharge is normal into targets touched to tentacles in the absence of vibrations (i.e., baseline) but fails to increase normally in the presence of nearby vibrations at 56Hz, a key frequency. After CD treatment, the abundance of hair bundles recovers to control levels within three hours. At 2h after CD treatment, when CD-resistant and CD-sensitive hair bundles are both present, but a full-recovery is not yet complete, somewhat enhanced discharge of nematocysts occurs into targets touched to tentacles in the presence of nearby vibrations at 56Hz (at least as compared to the response of CD-treated animals to contact with test probes in the absence of vibrations). Additionally, at 2h after CD-treatment, prey capture recovers to normal. Thus, two populations of hair bundles may be present on tentacles of sea anemones: those that are CD-resistant and those that are CD-sensitive. The functions of these hair bundles may be distinct.

Repair of traumatized mammalian hair cells via sea anemone repair proteins.

Mammalian hair cells possess only a limited ability to repair damage after trauma. In contrast, sea anemones show a marked capability to repair damaged hair bundles by means of secreted repair proteins (RPs). Previously, it was found that recovery of traumatized hair cells in blind cavefish was enhanced by anemone-derived RPs; therefore, the ability of anemone RPs to assist recovery of damaged hair cells in mammals was tested here. After a 1 h incubation in RP-enriched culture media, uptake of FM1-43 by experimentally traumatized murine cochlear hair cells was restored to levels comparable to those exhibited by healthy controls. In addition, RP-treated explants had significantly more normally structured hair bundles than time-matched traumatized control explants. Collectively, these results indicate that anemone-derived RPs assist in restoring normal function and structure of experimentally traumatized hair cells of the mouse cochlea.

Mining-caused changes to habitat structure affect amphibian and reptile population ecology more than metal pollution.

Emissions from smelting not only contaminate water and soil with metals, but also induce extensive forest dieback and changes in resource availability and microclimate. The relative effects of such co-occurring stressors are often unknown, but this information is imperative in developing targeted restoration strategies. We assessed the role and relative effects of structural alterations of terrestrial habitat and metal pollution caused by century-long smelting operations on amphibian and reptile communities by collecting environmental and time- and area-standardized multivariate abundance data along three spatially replicated impact gradients. Overall, species richness, diversity, and abundance declined progressively with increasing levels of metals (As, Cu, and Ni) and soil temperature (T(s)) and decreasing canopy cover, amount of coarse woody debris (CWD), and relative humidity (RH). The composite habitat variable (which included canopy cover, CWD, T(s), and RH) was more strongly associated with most response metrics than the composite metal variable (As, Cu, and Ni), and canopy cover alone explained 19-74% of the variance. Moreover, species that use terrestrial habitat for specific behaviors (e.g., hibernation, dispersal), especially forest-dependent species, were more severely affected than largely aquatic species. These results suggest that structural alterations of terrestrial habitat and concomitant changes in the resource availability and microclimate have stronger effects than metal pollution per se. Furthermore, much of the variation in response metrics was explained by the joint action of several environmental variables, implying synergistic effects (e.g., exacerbation of metal toxicity by elevated temperatures in sites with reduced canopy cover). We thus argue that the restoration of terrestrial habitat conditions is a key to successful recovery of herpetofauna communities in smelting-altered landscapes.

Diagnostic criteria and core outcome set development for necrotising otitis externa: the COSNOE Delphi consensus study.

OBJECTIVE: Evidence for necrotising otitis externa (NOE) diagnosis and management is limited, and outcome reporting is heterogeneous. International best practice guidelines were used to develop consensus diagnostic criteria and a core outcome set (COS). METHODS: The study was pre-registered on the Core Outcome Measures in Effectiveness Trials (COMET) database. Systematic literature review identified candidate items. Patient-centred items were identified via a qualitative study. Items and their definitions were refined by multidisciplinary stakeholders in a two-round Delphi exercise and subsequent consensus meeting. RESULTS: The final COS incorporates 36 items within 12 themes: Signs and symptoms; Pain; Advanced Disease Indicators; Complications; Survival; Antibiotic regimes and side effects; Patient comorbidities; Non-antibiotic treatments; Patient compliance; Duration and cessation of treatment; Relapse and readmission; Multidisciplinary team management.Consensus diagnostic criteria include 12 items within 6 themes: Signs and symptoms (oedema, otorrhoea, granulation); Pain (otalgia, nocturnal otalgia); Investigations (microbiology [does not have to be positive], histology [malignancy excluded], positive CT and MRI); Persistent symptoms despite local and/or systemic treatment for at least two weeks; At least one risk factor for impaired immune response; Indicators of advanced disease (not obligatory but mut be reported when present at diagnosis). Stakeholders were unanimous that there is no role for secondary, graded, or optional diagnostic items. The consensus meeting identified themes for future research. CONCLUSION: The adoption of consensus-defined diagnostic criteria and COS facilitates standardised research reporting and robust data synthesis. Inclusion of patient and professional perspectives ensures best practice stakeholder engagement.

Rho participates in chemoreceptor-induced changes in morphology to hair bundle mechanoreceptors of the sea anemone, Nematostella vectensis.

Adjustable hair bundle mechanoreceptors located on anemone tentacles detect movements of nearby, swimming prey. The hair bundles are formed by numerous actin-based stereocilia that converge onto a single, central kinocilium. Interestingly, morphological and functional changes to the hair bundles are induced by activating chemoreceptors that bind prey-derived N-acetylated sugars and proline, respectively. Morphological changes to the hair bundles involve alterations to the actin cytoskeleton of stereocilia. A pharmacological activation of Rho induces hair bundles to elongate to lengths comparable to those normally induced by exposure to N-acetylneuraminic acid (NANA) and prevents shortening of hair bundles normally induced by proline. Rho inhibition prevents NANA-induced elongation, but does not prevent proline-induced shortening of hair bundles. Western blots feature a band similar in mass to that predicted for a Rho homolog in the genome of Nematostella. Immunocytochemistry localizes Rho in stereocilia of the hair bundle. Anemone hair bundles arise from multicellular complexes. Data from experiments using heptanol, a gap junction uncoupler, indicate that cell-cell communication is required in order for activated chemoreceptors to induce morphological changes to the hair bundles.

Evidence for involvement of TRPA1 in the detection of vibrations by hair bundle mechanoreceptors in sea anemones.

A homolog of TRPA1 was identified in the genome of the anemone, Nematostella vectensis (nv-TRPA1a), and predicted to possess six ankyrin repeat domains at the N-terminus and an ion channel domain near the C-terminus. Transmembrane segments of the ion channel domain are well conserved among several known TRPA1 polypeptides. Inhibitors of TRPA1 including ruthenium red decrease vibration-dependent discharge of nematocysts in N. vectensis and Haliplanella luciae. Activators of TRPA1 including URB-597 and polygodial increase nematocyst discharge in the absence of vibrations. Co-immunoprecipitation yields a band on SDS-PAGE gels at the predicted mass of the nv-TRPA1a polypeptide among other bands. Co-immunoprecipitation performed in the presence of antigenic peptide decreases the yield of this and several other polypeptides. In untreated controls, anti-nv-TRPA1a primarily labels the base of the hair bundle with some labeling also distributed along the length of stereocilia. Tissue immunolabeled in the presence of the antigenic peptide exhibits reduced labeling. Activating chemoreceptors for N-acetylated sugars induce immunolabel to distribute distally in stereocilia. In anemones, activating chemoreceptors for N-acetylated sugars induce hair bundles to elongate among several other structural and functional changes. Taken together, these results are consistent with the possibility that nv-TRPA1a participates in signal transduction of anemone hair bundles.

Self/Non-Self Recognition Affects Cnida Discharge and Tentacle Contraction in the Sea Anemone Haliplanella luciae.

Certain species of sea anemone live in tightly packed communities, among clonemates and non-clonemates. Competition for space leads to intraspecific and interspecific aggressive interactions among anemones. The initial aggressive interactions appear to involve reciprocal discharge of cnidae triggered by contact with non-self feeding tentacles. We asked whether molecules contained in anemone-derived mucus constituted an important cue alone or in combination with cell surface molecules in stimulating aggressive or avoidance behaviors. In this study, we found that self and non-self stimuli differentially influenced two effector systems: cnida discharge and tentacle contraction. Interspecific mucus enhanced nematocyst discharge by 44% and spirocyst discharge by 90%, as compared to baseline discharge obtained in seawater alone. Conspecific stimuli accompanying touch inhibited specific tentacle contractions occurring on the far side of anemones relative to the site of contact. The greatest tentacle contractions occurred with exposure to interspecific mucus and tissue. Thus, several receptor systems are involved that integrate chemical and mechanical cues in order to initiate appropriate and graded effector responses during competition for space.

The involvement of arl-5b in the repair of hair cells in sea anemones.

The subcellular processes involved in repair of hair cells are not well understood. Sea anemones repair hair bundle mechanoreceptors on their tentacles after severe trauma caused by 1-h exposure to calcium-depleted seawater. Repair is dependent on the synthesis and secretion of large protein complexes named "repair proteins." A cDNA library on traumatized anemone tissue was probed using polyclonal antibodies raised to a specific chromatographic fraction of the repair protein mixture. An ADP-ribosylation factor-like protein, Arl-5b, was identified. The amino acid sequence of the Arl-5b protein in sea anemones is similar to that among several model vertebrates and humans. A polyclonal antibody raised to a peptide of the anemone Arl-5b labels some but not all hair bundles in healthy control animals. The abundance of labeled hair bundles significantly increases above healthy controls after trauma and continuing through the first hour of recovery. Dilute anti-Arl-5b blocks the spontaneous repair of hair bundle mechanoreceptors, suggesting that Arl-5b acts on the extracellular face of the plasma membrane. Immunoelectron microscopy indicates that Arl-5b is located along the length of stereocilia including sites in the vicinity of tip links. We propose that Arl-5b is involved in installing replacement linkages into damaged hair bundle mechanoreceptors.

Acquired paediatric subglottic cysts: a series from Manchester.

UNLABELLED: Subglottic cysts (SGC) have long been regarded as a rare cause of airway obstruction but through increased awareness an increase in the number of cases of SGC has been reported. OBJECTIVE: This paper describes the pathogenesis and management of SGC. DESIGN: Retrospective case series. Ethical approval not sought. SETTING: Royal Manchester Children's Hospital. PATIENTS: Two hundred and six new referrals for direct laryngotracheobronchoscopy (DLTB) were identified from records between September 2003 and September 2005. MAIN OUTCOMES MEASURED: Age at birth, sex, length of intubation, presenting symptoms, age at presentation, DLTB findings, interventional procedures, and follow-up DLTBs. RESULTS: Fourteen out of 206 (6.8%) infants were diagnosed as with subglottic cysts. This represented the fourth most common cause of upper airway pathology. Thirteen out of 14 (93%) infants were preterm (26.8 weeks S.D. 25.3-28.3 weeks). All infants had been intubated ranging from 1 to 180 days (median 42 days). The onset of symptoms ranged from 1 to 13 months (median 4.25 months). Initially, 8/14 (57.2%) infants had SGC cysts marsupialised with microforceps. A further six cysts (50%) were decapped between 2 and 4 months and one between 6 and 12 months. CONCLUSION: The number of cases of SGC has been increasing over the last three decades and represents the fourth most common causes of airway obstruction in our series. There is a delay in onset of symptoms and high rate of recurrence in the first 4 months. It is therefore prudent to reschedule further endoscopic evaluation between 2 and 4 months and after 6 months should the clinical need arise.

Diagnosis and management of a tympanic membrane hemangioma.

Vascular tumors arising in the temporal bone represent 0.7% of all lesions in this area. Hemangiomas confined to the tympanic membrane are exceptionally rare. We report a new case of tympanic membrane hemangioma that arose in 59-year-old man who presented with a 2-month history of constant right-sided pulsatile tinnitus and associated neck discomfort. The lesion and a cuff of healthy tympanic membrane were excised, and the resulting defect was repaired with a temporalis fascia graft. At 3 months, the neotympanum was well healed and the patient's symptoms had resolved. We also review the limited number of previously published cases of tympanic membrane hemangioma, and we discuss the presentation and management of these lesions. Some tympanic hemangiomas are asymptomatic and others present as hearing loss, tinnitus, otalgia, and/or otorrhea. Pure-tone audiometry and high-resolution computed tomography of the temporal bones are essential to determine the extent of the lesion. An excisional biopsy is recommended because it is curative and it provides histologic confirmation of the diagnosis. Excision requires elevation of a tympanomeatal flap, en bloc resection, and reconstruction of the tympanic membrane defect. Small lesions can be excised via a permeatal or endaural approach, while larger lesions require a postauricular approach.

Proteomic identification of hair cell repair proteins in the model sea anemone Nematostella vectensis.

Sea anemones have an extraordinary capability to repair damaged hair bundles, even after severe trauma. A group of secreted proteins, named repair proteins (RPs), found in mucus covering sea anemones significantly assists the repair of damaged hair bundle mechanoreceptors both in the sea anemone Haliplanella luciae and the blind cavefish Astyanax hubbsi. The polypeptide constituents of RPs must be identified in order to gain insight into the molecular mechanisms by which repair of hair bundles is accomplished. In this study, several polypeptides of RPs were isolated from mucus using blue native PAGE and then sequenced using LC-MS/MS. Thirty-seven known polypeptides were identified, including Hsp70s, as well as many polypeptide subunits of the 20S proteasome. Other identified polypeptides included those involved in cellular stress responses, protein folding, and protein degradation. Specific inhibitors of Hsp70s and the 20S proteasome were employed in experiments to test their involvement in hair bundle repair. The results of those experiments suggested that repair requires biologically active Hsp70s and 20S proteasomes. A model is proposed that considers the function of extracellular Hsp70s and 20S proteasomes in the repair of damaged hair cells.

Outcomes in Stapedotomy Surgery: The Learning Curve Redefined.

OBJECTIVE: The aim of this study is to determine if the cumulative summation test for the learning curve (LC-CUSUM) and the cumulative summation graph (CUSUM) can be used to demonstrate landmark points of competence and maintenance of proficiency in stapes surgery over a continuous time period. STUDY DESIGN: Retrospective review from January 1999 until August 2014. SETTING: Tertiary referral hospital. PATIENTS: All adult patients with confirmed otosclerosis. INTERVENTION(S): Two-hundred and four primary and revision stapedotomy. MAIN OUTCOME MEASURE(S): Learning curves were constructed using the CUSUM and LC-CUSUM. Failure was defined as closure of the ABG >10 dB in less than 10% of patients to demonstrate the landmark point of competency and to highlight any fluctuations over a prolonged period. RESULTS: When the failure rate was defined as closure of the ABG >10 dB, it was not possible to create useful LC-CUSUM and CUSUM graphs, but by redefining the failure rate as > 15 dB, competency was reached at case 43 and maintained with natural fluctuations occurring between cases 137 and 149 and again at case 196. CONCLUSIONS: LC-CUSUM and CUSUM are a more robust analytical method of illustrating the learning curve and suggest that the traditional benchmark of closure of the ABG ≤10 dB in more than 90% of patients may need reconsideration. It can also be used as standardized audit tools when monitoring results and used to plan future training programs as they clearly define a point when novice trainees become competent.

Rapid recovery of sensory function in blind cave fish treated with anemone repair proteins.

Blind cave fish employ superficial neuromasts to detect currents [Baker, C.F. and J.C. Montgomery, J. Comp. Physiol. A 184 (1999) 519-527]. Briefly exposing fish to calcium-free water significantly reduces the ability of the fish to perform rheotaxis (i.e., to orient properly in currents). Spontaneous recovery to control levels of rheotaxis requires 9 days. However, if the fish are treated with fraction beta immediately after exposure to calcium-free water, recovery to control levels of rheotaxis occurs within 1.3 h, the first time point tested. Fraction beta is a chromatographic fraction of 'repair proteins' isolated from sea anemones. The benefits of fraction beta on restoring rheotaxis exhibit dose dependency with the minimum effective dose estimated at 1 ng/ml. Exogenously supplied ATP augments the efficacy of fraction beta. Such augmentation is abolished by PPADS, an inhibitor of purinoceptors. Immunocytochemistry confirms the presence of purinoceptors in superficial neuromasts. The present results suggest that 'repair proteins' obtained from anemones significantly augment intrinsic repair mechanisms in fish. Furthermore, the data obtained in the fish system strongly parallel our previously published findings on sea anemones, raising the possibility that mechanisms of hair bundle repair may be evolutionarily conserved.