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Background: Most schwannomas are isolated tumours occurring in otherwise healthy people. However, bilateral vestibular schwannomas (BVS) or multiple non-vestibular schwannomas indicate an underlying genetic predisposition. This is most commonly NF2 -related schwannomatosis (SWN), but when BVS are absent, this can also indicate SMARCB1 -related or LZTR1 -related SWN., Methods: We assessed the variant detection rates for the three major SWN genes ( NF2 , LZTR1 and SMARCB1 ) in 154 people, from 150 families, who had at least one non-vestibular schwannoma, but who did not meet clinical criteria for NF2 -related SWN at the time of genetic testing., Results: We found that 17 (11%) people from 13 families had a germline SMARCB1 variant and 19 (12%) unrelated individuals had a germline LZTR1 variant. 19 people had an NF2 variant, but 18 of these were mosaic and 17 were only detected when 2 tumours were available for testing. The overall detection rate was 25% using blood alone, but increased to 36% when tumour analysis was included. Another 12 people had a germline variant of uncertain significance (VUS)., Conclusions: There were similar proportions of LZTR1 , SMARCB1 or mosaic NF2 . However, since an NF2 variant was detected in tumours from 103 people, it is likely that further cases of mosaicism would be detected if more people had additional tumours available for analysis. In addition, if further evidence becomes available to show that the VUSs are pathogenic, this would significantly increase the proportion of people with a genetic diagnosis. Our results indicate the importance of comprehensive genetic testing and improved variant classification., Competing Interests: Competing interests: GE has received consultancy fees from AstraZeneca, Springworks and Everything Genetic., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Antarctica harbours some of the most isolated and extreme environments on Earth, concealing a largely unexplored and unique component of the global animal virosphere. To understand the diversity and evolutionary histories of viruses in these polar species, we determined the viromes of gill metatranscriptomes from 11 Antarctic fish species with 248 samples collected from the Ross Sea region spanning the Perciformes, Gadiformes, and Scorpaeniformes orders. The continent's shift southward and cooling temperatures >20 million years ago led to a reduction in biodiversity and subsequent radiation of some marine fauna, such as the notothenioid fishes. Despite decreased host species richness in polar regions, we revealed a surprisingly complex virome diversity in Ross Sea fish, with the types and numbers of viruses per host species and individuals sampled comparable to that of fish in warmer marine environments with higher host community diversity. We also observed a higher number of closely related viruses likely representing instances of recent and historic host-switching events among the Perciformes (all notothenioids) than in the Gadiformes, suggesting that rapid speciation events within this order generated closely related host species with few genetic barriers to cross-species transmission. Additionally, we identified novel genomic variation in an arenavirus with a split nucleoprotein sequence containing a stable helical structure, indicating potential adaptation of viral proteins to extreme temperatures. These findings enhance our understanding of virus evolution and virus-host interactions in response to environmental shifts, especially in less diverse ecosystems that are more vulnerable to the impacts of anthropogenic and climate changes., Competing Interests: None declared., (© The Author(s) 2024. Published by Oxford University Press.)

Background: New Zealand is home to over 120 native endemic species of skinks and geckos that radiated over the last 20-40 million years, likely driven by the exploitation of diverse habitats formed during the Miocene. The recent radiation of animal hosts may facilitate cross-species virus transmission, likely reflecting their close genetic relationships and therefore relatively low barriers for viruses to emerge in new hosts. Conversely, as animal hosts adapt to new niches, even within specific geographic locations, so too could their viruses. Consequently, animals that have niche-specialised following radiations may be expected to harbour genetically distinct viruses. Through a metatranscriptomic analysis of eight of New Zealand's native skink and gecko species, as well as the only introduced lizard species, the rainbow skink (Lampropholis delicata), we aimed to reveal the diversity of viruses in these hosts and determine whether and how the radiation of skinks and geckos in New Zealand has impacted virus diversity and evolution., Results: We identified a total of 15 novel reptilian viruses spanning 11 different viral families, across seven of the nine species sampled. Notably, we detected no viral host-switching among the native animals analysed, even between those sampled from the same geographic location. This is compatible with the idea that host speciation has likely resulted in isolated, niche-constrained viral populations that have prevented cross-species transmission. Using a protein structural similarity-based approach, we further identified a highly divergent bunya-like virus that potentially formed a new family within the Bunyavirales., Conclusions: This study has broadened our understanding of reptilian viruses within New Zealand and illustrates how niche adaptation may limit viral-host interactions., (© 2024. The Author(s).)

The fishing and aquaculture industry is vital for global food security, yet viral diseases can result in mass fish die-off events. Determining the viromes of traditionally understudied species, such as fish, enhances our understanding of the global virosphere and the factors that influence virome composition and disease emergence. Very little is known about the viruses present in New Zealand's native fish species, including the shortfin eel (Anguilla australis) and the longfin eel (Anguilla dieffenbachii), both of which are fished culturally by Māori (the indigenous population of New Zealand) and commercially. Through a total RNA metatranscriptomic analysis of longfin and shortfin eels across three different geographic locations in the South Island of New Zealand, we aimed to determine whether viruses had jumped between the two eel species and whether eel virome composition was impacted by life stage, species, and geographic location. We identified nine viral species spanning eight different families, thereby enhancing our understanding of eel virus diversity in New Zealand and the host range of these viral families. Viruses of the family Flaviviridae (genus Hepacivirus) were widespread and found in both longfin and shortfin eels, indicative of cross-species transmission or virus-host co-divergence. Notably, both host specificity and geographic location appeared to influence eel virome composition, highlighting the complex interaction between viruses, hosts, and their ecosystems. This study broadens our understanding of viromes in aquatic hosts and highlights the importance of gaining baseline knowledge of fish viral abundance and diversity, particularly in aquatic species that are facing population declines., (© 2024. The Author(s).)

The lesser short-tailed bat ( Mystacina tuberculata ) and the long-tailed bat ( Chalinolobus tuberculatus ) are Aotearoa New Zealand's only native extant terrestrial mammals and are believed to have migrated from Australia. Long-tailed bats arrived in New Zealand an estimated two million years ago and are closely related to other Australian bat species. Lesser short-tailed bats, in contrast, are the only extant species within the Mystacinidae and are estimated to have been living in isolation in New Zealand for the past 16-18 million years. Throughout this period of isolation, lesser short-tailed bats have become one of the most terrestrial bats in the world. Through a metatranscriptomic analysis of guano samples from eight locations across New Zealand, we aimed to characterise the viromes of New Zealand's bats and determine whether viruses have jumped between these species over the past two million years. High viral richness was observed among long-tailed bats with viruses spanning seven different viral families. In contrast, no bat-specific viruses were identified in lesser short-tailed bats. Both bat species harboured an abundance of likely dietary- and environment-associated viruses. We also identified alphacoronaviruses in long-tailed bat guano that had previously been identified in lesser short-tailed bats, suggesting that these viruses had jumped the species barrier after long-tailed bats migrated to New Zealand. Of note, an alphacoronavirus species discovered here possessed a complete genome of only 22,416 nucleotides with entire deletions or truncations of several non-structural proteins, thereby representing what may be the shortest genome within the Coronaviridae identified to date. Overall, this study has revealed a diverse range of novel viruses harboured by New Zealand's only native terrestrial mammals, in turn expanding our understanding of bat viral dynamics and evolution globally., Competing Interests: None declared., (© The Author(s) 2024. Published by Oxford University Press.)

Tuatara (Sphenodon punctatus) are one of the most phylogenetically isolated vertebrate species and provide a unique host system to study virus evolution. While the tuatara genome, sequenced in 2020, revealed many endogenous viral elements, we know little of the exogenous viruses that infect tuatara. We performed a metatranscriptomics study of tuatara cloaca samples from a wild population on Takapourewa (Stephens Island), Aotearoa New Zealand. From these data we identified 49 potentially novel viral species that spanned 19 RNA viral families and/or orders, the vast majority (48) of which were likely dietary-related. Notably, using a protein structure homology search, we identified a highly divergent novel virus within the Picornaviridae which may directly infect tuatara. Additionally, two endogenous tuatara adintoviruses were characterised that exhibited long-term viral-host co-divergence. Overall, our results indicate that the tuatara cloacal virome is highly diverse, likely due to a large number of dietary-related viruses., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Heat shock protein 70s (Hsp70s) are a highly conserved class of molecular chaperones that fold a large proportion of the proteome. Nematostella vectensis (Nv) is an estuarine sea anemone that has emerged as a model species to characterize molecular responses to physiological stressors due to its exposure to diverse, extreme abiotic conditions. Previous transcriptional data has shown dramatic differences among expression profiles of three NvHsp70 isoforms (NvHsp70A, B and D) under stress but it is unknown if, and to what extent, the client proteins for these chaperones differ. In order to determine client specificity, NvHsp70A, B and D were expressed in Saccharomyces cerevisiae budding yeast lacking native Hsp70 and interacting proteins for each Hsp70 were determined with mass spectrometry in yeast ambient and heat shock conditions. Our analyses showed <50% of identified interacting proteins were common to all three anemone Hsp70s and 3-18% were unique to an individual Hsp70. Mapping of temperature induced interactions suggest that under stress a proportion of clients are transferred from NvHsp70A and NvHsp70D to NvHsp70B. Together, these data suggest a diverse set of interacting proteins for Hsp70 isoforms that likely determines the precise functions for Hsp70s in organismal acclimation and potentially adaptation. BIOLOGICAL SIGNIFICANCE: Although the Hsp70 family of molecular chaperones has been studied for >50 years, it is still not fully understood why organisms encode and express many highly-similar Hsp70 isoforms. The prevailing theory is that these isoforms have identical function, but are expressed under unique cellular conditions that include heat shock to cope with increased number of unfolded/misfolded proteins. The sea anemone Nematostella vectensis encodes three Hsp70 isoforms A, B and D that when expressed in yeast demonstrate unique functionalities. This study provides the interactome of NvHsp70s A, B and D and demonstrates that Hsp70 isoforms, while highly similar in sequence, have unique co-chaperone and client interactors., (Copyright © 2019. Published by Elsevier B.V.)

Marine organisms experience abiotic stressors such as fluctuations in temperature, UV radiation, salinity, and oxygen concentration. Heat shock proteins (HSPs) assist in the response of cells to these stressors by refolding and maintaining the activity of damaged proteins. The well-conserved Hsp70 chaperone family is essential for cell viability as well as the response to stress. Organisms possess a variety of Hsp70 isoforms that differ slightly in amino acid sequence, yet very little is known about their functional relevance. In this study, we undertook analysis of three principal Hsp70 isoforms NvHsp70A, B, and D from the starlet sea anemone Nematostella vectensis. The functionality of Hsp70 isoforms in the starlet sea anemone was assessed through transcriptional analysis and by heterologous expression in budding yeast Saccharomyces cerevisiae. Interestingly, these isoforms were found to not only differ in expression under stress but also appear to have functional differences in their ability to mediate the cellular stress program. These results contribute to an understanding of Hsp70 isoform specificity, their shared and unique roles in response to acute and chronic environmental stress, and the potential basis of local adaptation in populations of N. vectensis.

Background: A 58-year-old man, previously diagnosed with Bartter's syndrome, presented with a short history of vomiting, diarrhea and weakness. He had severe hyperkalemia (serum potassium levels >10 mmol/l), which was successfully managed. Post hoc investigation suggested that the patient had Gitelman's rather than Bartter's syndrome., Investigations: Physical examination, urine and blood analyses, chest radiography, electrocardiogram, renal ultrasound, and genetic analysis focusing on the SLC12A3 gene, which encodes the thiazide-sensitive Na/Cl cotransporter., Diagnosis: Gitelman's syndrome and hyperkalemia secondary to acute renal failure plus exogenous potassium supplementation., Management: Intravenous calcium gluconate, insulin and dextrose administration. Temporary continuous venovenous hemodiafiltration. Genetic confirmation of the underlying molecular defect. Long-term treatment for Gitelman's syndrome with oral potassium and magnesium supplements and epithelial sodium channel-blocking drugs. Review of patient education regarding renal salt-wasting syndromes.

The effect of concurrent affective and alcohol use disorders on suicidal ideation and behavior was investigated. The Diagnostic Interview Schedule Version III-R (DIS) was administered to 307 adult veteran men ranging in age from 23 to 78. Participants were classified into one of four groups based on their final DIS diagnosis-lifetime unipolar depression and lifetime bipolar I disorder with or without a lifetime alcohol use disorder. Logistic regression analyses indicated that veterans with a major affective disorder were at greater risk for suicidality than veterans without an affective disorder. However, veterans with unipolar depression were at no greater risk for suicidality than those with bipolar I disorder. Unipolar and bipolar I disorders with a concurrent alcohol use disorder were always associated with an increased risk for suicidality.

1. Studies of the regulation of neurosecretory cell gene expression suffer from the lack of suitable cell lines. Two approaches have been used to overcome this deficit: transfection of neuropeptide genes into heterologous cell lines and generation of transgenic animals. 2. Studies with heterologous cell lines have revealed the potential involvement of nuclear hormone receptors, POU proteins, and fos/jun/ATF family members in the regulation of the vasopressin and oxytocin genes. Although limited in their scope, these studies have contributed greatly to the dissection of basic properties of elements in the vasopressin and oxytocin gene promoters. 3. Transgenic mice, and more recently rats, have been used to elucidate genomic regions governing cell specificity and physiological regulation of neurosecretory gene expression. The genes encoding the neuropeptides vasopressin and oxytocin have been used in many transgenic studies, due to the well-defined expression patterns and physiology of the endogenous neuropeptides. Cell-specific and physiologically regulated expression of these transgenes has been achieved, demonstrating the action of putative repressor elements and regulation of the expression of one gene by sequences present in the other gene. 4. Appropriate expression and translation of transgenes have resulted in the production of several useful systems. Expression of oncogene sequences in gonadotropin-releasing hormone neurons has allowed the development of cell lines from the resulting tumors, overproduction of corticotropin-releasing factor has produced animal models of anxiety and obesity, and directed ectopic expression of growth hormone has generated a potentially useful rat model of dwarfism. These and other animal models of human disease will provide important avenues for the development of therapeutic strategies.

Major improvements to the HPLC separation of fluorescent derivatives of excitatory sulphur-containing amino acids have been made. Quisqualate was used as the internal standard since no endogenous derivatives coeluted with it. The artefactual generation of sulphinic and sulphonic amino acids from the oxidation of cysteine (56 microM) and homocysteine (1.2 microM) has been investigated using deionised water, an acidic phosphate/methanol mixture, perchloric acid and trichloroacetic acid (TCA) as extraction media. Of the four extraction media examined, TCA in combination with ether extraction was shown to be the most potent oxidative treatment and resulted in 23% oxidation of original cysteine or homocysteine to sulphinic and sulphonic acids. This oxidation was partially resistant to the presence of physiological concentrations of glutathione (1.5 mM) such that in the case of cysteine, 6% oxidation was observed. A 10% (v/v) mixture of methanol in 75 mM phosphate solution (pH 4.6) was found to be the most artefact-free extraction method and in spinal cord tissue processed with this medium, cysteine sulphinic acid was the only excitatory sulphur-containing amino acid consistently detectable (0.24 +/- 0.01 pmol/mg wet weight, n = 6).

The hydrodynamic sizes of antibody/3H-dsDNA immune complexes are correlated with their ability to fix complement in a red blood cell-complement linked binding assay. The results with PM2 dsDNA and a sonicated preparation (with a molecular weight of approximately 5.10(5) suggest that packing considerations are also important in the complement binding reaction. The relevance of these findings with respect to the role of antibody/dsDNA immune complexes in the pathogenesis of glomerulonephritis in systemic lupus erythematosus is discussed.

The Farr assay and sucrose gradient ultracentrifugation were used to study the importance of kinetic factors on the size of antibody/DNA immune complexes prepared from SLE sera and dsDNA. We have found that significantly different numbers and sizes of antibody/DNA immune complexes can be formed by varying the time course of addition of a given amount of DNA to an SLE serum. For example, the introduction of additional DNA to a preincubated antibody-DNA system does not lead to reequilibration of that system with respect to the amount of DNA bound or the size of the complexes formed even after 1 hr at 37 degrees C. The potential implications of these observations with respect to the pathogenesis of SLE is discussed.

We describe a prototype quantitative automated assay for fibrin and fibrinogen degradation products, a particle-enhanced turbidimetric inhibition immunoassay (PETINIA) in the Du Pont aca discrete clinical analyzer. This assay involves a latex particle reagent with covalently bound fibrinogen and a polyclonal antiserum raised in rabbits against human fibrinogen. A special secondary sample-collection tube quantitatively removes fibrinogen from citrated plasma and inhibits further fibrinolysis, independent of heparin concentration. The assay range is 0-100 mg/L, in fibrinogen equivalents. The CV for the assay is less than 10% when performed with the aca. Nonclottable fibrin and fibrinogen fragments are measured by the assay, the greatest sensitivity being directed at the E domain of the fibrinogen molecule. We illustrate with case studies the potential of this assay for providing clinical information not obtainable with currently available qualitative and semi-quantitative assays.

Artificial cryoprecipitates have been prepared from SLE sera by incubation of anti-double stranded DNA with 3H-PM2 DNA. IgM rheumatoid factor, and complement to study further the relationships between natural cryoprecipitation and the DNA/anti-DNA system in this disorder. Addition of rheumatoid factor and/or complement to a mixture of SLE serum and 3H-DNA results in increased precipitation of the 3H-DNA following incubation at 4 degrees C. Precipitation formed with added complement or complement plus rheumatoid factor could be rapidly resolubilized or reprecipitated by warming to 37 degrees C or recooling to 4 degrees C, i.e., exhibited the behavior of natural cryoprecipitates. The resolubilized cryoprecipitates contained excess antibody, because they bound significant quantities of additional 3H-DNA in the Farr assay. Of special interest was the observation that DNA binding activity frequently could be recovered from solubilized cryoprecipitates even though 3H-DNA was not added during the original cryoprecipitate preparation. With certain cryoprecipitations prepared in this way, the DNA binding activity was further increased by deoxyribonuclease treatment, raising the possibility that endogenous DNA may be constituent of such cryoprecipitates. The implications of these findings with respect to the nature of antibody-DNA immune complexes in SLE are discussed.

A polyethylene glycol (PEG) radioimmunoprecipitation assay for human IgG is described that is sufficiently sensitive to detect 0.5 ng of IgG. This model antibody-antigen system was also used to study the stoichiometries of PEG-precipitated complexes. Our results suggest that the presence of PEG may affect the stoichiometry of the complexes which precipitate from solution.

The influence of rheumatoid factor (RF) on the complement mediated binding of antibody/double-stranded DNA immune complexes to red blood cells has been investigated. Our results indicate that RF enhances this binding reaction, apparently by fixing complement via its own Fc region. These findings suggest that under certain circumstances, RF may play an exacerbating role in the antibody/DNA induced glomerulonephritis of systemic lupus erythematosus.