Facilitating taxonomy and phylogenetics: An informative and cost-effective protocol integrating long amplicon PCRs and third-generation sequencing.

Phylogenetic inference has become a standard technique in integrative taxonomy and systematics, as well as in biogeography and ecology. DNA barcodes are often used for phylogenetic inference, despite being strongly limited due to their low number of informative sites. Also, because current DNA barcodes are based on a fraction of a single, fast-evolving gene, they are highly unsuitable for resolving deeper phylogenetic relationships due to saturation. In recent years, methods that analyse hundreds and thousands of loci at once have improved the resolution of the Tree of Life, but these methods require resources, experience and molecular laboratories that most taxonomists do not have. This paper introduces a PCR-based protocol that produces long amplicons of both slow- and fast-evolving unlinked mitochondrial and nuclear gene regions, which can be sequenced by the affordable and portable ONT MinION platform with low infrastructure or funding requirements. As a proof of concept, we inferred a phylogeny of a sample of 63 spider species from 20 families using our proposed protocol. The results were overall consistent with the results from approaches based on hundreds and thousands of loci, while requiring just a fraction of the cost and labour of such approaches, making our protocol accessible to taxonomists worldwide.

Spontaneous Bleeding in Vestibular Schwannoma in Patients on Oral Anticoagulant Therapy: Report of Two Cases and Review of Literature.

OBJECTIVE: Anticoagulant therapy is a risk factor for repeated intratumoral hemorrhage and acute enlargement of a vestibular schwannoma (VS) with neurological deficits. Therefore, we describe two cases of patients on oral anticoagulant therapy with intratumoral hemorrhage in which anticoagulant therapy prior to surgical resection was discontinued. We also discuss other similar cases from the literature since this is a rare event. CASE REPORTS: We described the two cases of intratumoral hemorrhage in acoustic neurinoma and conducted a literature review of similar cases of patients with intratumoral hemorrhage in acoustic neurinoma who were also on oral anticoagulants. Both patients presented with CN-VII palsy prior to surgery; both also fully re-covered after surgery except for hearing loss on the tumor side. Our literature review found 50 cases of VS (reported as vestibular schwannomas in the literature) with intratumoral hemorrhage. From this total, 11 patients used oral anticoagulant therapy with reported poor outcomes and high mortality; 9 of these 11 cases were reported in the past 20 years. The incidence is expected to rise due to increased use of anticoagulant therapy due to onset of atrial fibrillation, atherosclerosis, and thromboembolism from longer human lifespan. CONCLUSION: Anticoagulant therapy represents a risk factor for intratumoral hemorrhage and acute enlargement of VS tumor mass with neurological deficits.

Mediterranean vineyards and olive groves in Croatia harbour some rare and endemic invertebrates.

The Mediterranean is characterised by high biodiversity and numerous endemic species. These species are not only present in natural habitats, but also inhabit areas under human influence, such as agricultural lands. In the biodiversity assessment of Mediterranean vineyards and olive orchards within Zadar County, in Croatia, we identified eight endemic species with Mediterranean distribution, six with a Balkan Peninsula distribution, four with Dinaric Alps distribution and three species rare and endangered in Europe. Alongside these species, we have recorded five new species for Croatian fauna, many of those identified by combining morphological characteristics and the DNA barcoding tool. Araneae and Coleoptera contributed the highest number of endemic species and groups with new record were the following: Coleoptera, Diptera and Araneae. Compared to other sites, an olive orchard with ecological pest management (EPM), surrounded by natural ecosystems, had the highest ratio of endemic and rare species. Our findings emphasise that agricultural lands in the Mediterranean can be habitats for endemic and rare species and that future biodiversity research of these habitats is highly important, to monitor potential biodiversity changes and motivate future species and ecosystem conservation.

Wolbachia infection dynamics in a natural population of the pear psyllid Cacopsylla pyri (Hemiptera: Psylloidea) across its seasonal generations.

Wolbachia is one of the most abundant intracellular symbionts of arthropods and has profound effects on host biology. Wolbachia transmission and host phenotypes often depend on its density within the host, which can be affected by multiple biotic and abiotic factors. However, very few studies measured Wolbachia density in natural host populations. Here, we describe Wolbachia in the pear psyllid Cacopsylla pyri from three populations in the Czech Republic. Using phylogenetic analyses based on wsp and multilocus sequence typing genes, we demonstrate that C. pyri harbours three new Wolbachia strains from supergroup B. A fourth Wolbachia strain from supergroup A was also detected in parasitised immatures of C. pyri, but likely came from a hymenopteran parasitoid. To obtain insights into natural Wolbachia infection dynamics, we quantified Wolbachia in psyllid individuals from the locality with the highest prevalence across an entire year, spanning several seasonal generations of the host. All tested females were infected and Wolbachia density remained stable across the entire period, suggesting a highly efficient vertical transmission and little influence from the environment and different host generations. In contrast, we observed a tendency towards reduced Wolbachia density in males which may suggest sex-related differences in Wolbachia-psyllid interactions.

Microbiome of pear psyllids: A tale about closely related species sharing their endosymbionts.

Psyllids are phloem-feeding insects that can transmit plant pathogens such as phytoplasmas, intracellular bacteria causing numerous plant diseases worldwide. Their microbiomes are essential for insect physiology and may also influence the capacity of vectors to transmit pathogens. Using 16S rRNA gene metabarcoding, we compared the microbiomes of three sympatric psyllid species associated with pear trees in Central Europe. All three species are able to transmit 'Candidatus Phytoplasma pyri', albeit with different efficiencies. Our results revealed potential relationships between insect biology and microbiome composition that varied during psyllid ontogeny and between generations in Cacopsylla pyri and C. pyricola, as well as between localities in C. pyri. In contrast, no variations related to psyllid life cycle and geography were detected in C. pyrisuga. In addition to the primary endosymbiont Carsonella ruddii, we detected another highly abundant endosymbiont (unclassified Enterobacteriaceae). C. pyri and C. pyricola shared the same taxon of Enterobacteriaceae which is related to endosymbionts harboured by other psyllid species from various families. In contrast, C. pyrisuga carried a different Enterobacteriaceae taxon related to the genus Sodalis. Our study provides new insights into host-symbiont interactions in psyllids and highlights the importance of host biology and geography in shaping microbiome structure.

Winter activity of Clubiona spiders and their potential for pest control.

Winter-active arthropod predators (like vegetation-dwelling spiders) significantly suppress pest populations during winter in pome fruit orchards in Central Europe. Clubiona spiders are very abundant in orchards and have been observed to be active during winter. Here, we performed laboratory experiments to assess the movement and predation activity of clubionids at low temperatures. In addition, we also assessed prey survival (psyllids and crickets). We revealed that Clubiona spiders actively moved even at a temperature below 0 °C. Pest prey (Cacopsylla sp.) was able to survive at low temperatures, but crickets died at 3 and -1 °C. Overall Clubiona activity was very low but present during the whole observation period of five days. The predation activity of Clubiona declined with lower temperatures for both cricket and pest (Cacopsylla sp.) prey. Nevertheless, 44% and 25% of Clubiona individuals captured and consumed psyllid and cricket prey, respectively, even at the lowest temperature of -1 °C. Our results show that Clubiona spiders are active predators at low temperatures and, therefore, should contribute to the suppression of overwintering pest populations.

FORAMEN OVALE AND FORAMEN ROTUNDUM: CHARACTERIZATION OF POSTNATAL DEVELOPMENT.

The sphenoid bone development occurs in both prenatal and postnatal periods. Sphenoid bone openings are used as surgical landmarks and are of great importance for neurosurgeons in everyday practice. The aim of this study was to identify morphological characteristics, postnatal development and remodeling, as well as clinical aspect of the sphenoid bone openings and to investigate their relationship and difference in size. The macerated sphenoid bones analyzed in this study were scanned by micro-computed tomography. Areas and distance in-between foramen ovale and foramen rotundum were measured. In addition, different shapes of foramen ovale were described. The most common shape of foramen ovale on both sides was oval, followed by the round, almond and elongated shapes. Modest to strong positive correlations between all foramina and age for the whole sample and both subsamples were presented, except for the right foramen rotundum area in the male subsample, which did not show significant correlation with age. Our study revealed changes in postnatal development and anatomy of foramen ovale and foramen rotundum, primarily in the aspects of size and shape, and should contribute to reducing the risk of damage to neurovascular structures during surgical procedures.

METEOROLOGICAL VARIABLES ASSOCIATED WITH SUBARACHNOID HEMORRHAGE: A SINGLE CENTER STUDY.

Spontaneous subarachnoid hemorrhage (SAH) can occur unexpectedly and independently of the classic risk factors. Several different factors could affect intracranial aneurysm (IA) rupture, such as morphological and hemodynamic factors. The aim of this study was to establish the potential association of meteorological data such as temperature, atmospheric pressure, and humidity, and the onset of clinical symptoms preceding hospital admission of patients with acute SAH due to IA rupture. This retrospective study included 130 consecutive patients admitted for non-traumatic SAH with a determinable onset of SAH symptoms. The effects of meteorological parameters of atmospheric pressure, ambient temperature, and relative air humidity on the day of acute SAH onset and 24 hours prior to the onset of symptoms were recorded and analyzed in each patient. Spearman rank correlation analysis was used to assess the risks of incident SAH on the basis of daily meteorological data. Seasonal incidence of acute SAH showed the peak incidence in winter and a trough in summer, with monthly incidence peak in January and December. The circadian rhythm analysis showed the peak incidence of SAH in the forenoon, followed by the evening. Acute SAH incidence showed moderate positive association with daily atmospheric pressure (p<0.05), while no association was found with ambient temperature and relative air humidity. Our results suggested no significant association of changes in ambient temperature and relative humidity with the risk of SAH. Increases in atmospheric pressure were weakly associated with a higher SAH risk. Additional studies are needed to establish in detail both meteorological and morphological factors important to predict IA rupture and SAH.

SPONTANEOUS RESOLUTION OF A NONFUNCTIONING PITUITARY ADENOMA OVER ONE-MONTH PERIOD: A CASE REPORT.

Spontaneous resolution of nonfunctioning pituitary adenoma after hemorrhagic apoplexy is a rare clinical entity of unknown etiology and is defined as disappearance of a tumor without any specific treatment. Here we present a 54-year-old male patient who presented with acute onset of severe headache, vomiting, photophobia, and sonophobia. He was referred to brain computed tomography, which showed a 16x12x16 mm tumor mass located in the sellar region with signs of hemorrhage. Endocrinologic evaluation was consistent with under-function of pituitary gonadotropic cells. Magnetic resonance imaging (MRI) performed ten days later was consistent with hemorrhagic apoplexy of the pituitary adenoma. The patient's symptoms resolved after conservative treatment with dexamethasone, but he was scheduled for elective pituitary surgery. Preoperative MRI was performed one month after the first one and disclosed normal pituitary gland without any signs of adenoma. Our case is remarkable due to the fact that spontaneous remission of pituitary adenoma occurred within the first month, which is the shortest interval reported to date. Our case highlights the importance of conservative therapy as the first-line treatment for pituitary apoplexy in the absence of neurological impairment, since spontaneous remission may occur in a short time interval.

Assessment of the biocontrol potential of natural enemies against psyllid populations in a pear tree orchard during spring.

BACKGROUND: Modern pest control management systems are based on the support of naturally occurring arthropod predators, as it has been shown that such predators offer an important ecosystem service. However, most naturally occurring arthropod predators are generalists (euryphagous). Their role in the biological control of specific pests has been recognized but remains poorly studied. Here, we focused on the naturally occurring arthropod predators of psyllids - the main insect pest of pear trees. We investigated the abundance of psyllids and all of their potential enemies in an abandoned pear orchard on a weekly basis from early spring to early summer. In addition, employing polymerase chain reaction diagnostics and specific primers, we investigated the predation rate on psyllids in all predators collected. RESULTS: We found four predatory groups: spiders were the most abundant (60%, N = 756), followed by coccinellid beetles, anthocorid bugs and cantharid beetles. Anthocorids and spiders had the highest predation rates among the predatory groups. Among spiders, >50% of foliage-dwelling spiders (belonging to the genera Philodromus and Clubiona; N = 206) were positive for psyllids and showed a numerical response to the abundance of psyllids. CONCLUSION: We conclude that foliage-dwelling spiders are, of the four groups, the most important natural enemies of psyllids on pear trees during spring in Central Europe, as they outnumber specialized Anthocoris bugs. © 2021 Society of Chemical Industry.

Surgical anatomy of microsurgical 3-level anterior cervical discectomy and fusion C4-C7.

Anterior cervical discectomy and fusion (ACDF) is one of the most common spinal procedures, frequently used for the treatment of cervical spine degenerative diseases. It was first described in 1958. Interestingly, to our knowledge, 3-level ACDF has not been previously published as a peer-reviewed video case with a detailed description of intraoperative microsurgical anatomy. In this video, we present the case of a 33-year-old male who presented with a combination of myelopathy (hyperreflexia and long tract signs in the upper and lower extremities) and bilateral radiculopathy of the upper extremities. He had been previously treated conservatively with physical therapy and pain management for 6 months without success. We performed 3-level microsurgical ACDF from C4 to C7. All 3 levels were decompressed, and bone allografts were placed to achieve intervertebral body fusion. A titanium plate was utilized from C4 to C7 for internal fixation. The patient was discharged home on the first postoperative day. His pain, numbness and tingling resolved, as well as his myelopathy. No perioperative complications were encountered. Herein we present the surgical anatomy of our operative technique including certain technical tips. Written consent was obtained directly from the patient.

Should we treat pain in the elderly palliative care cancer patients differently?

Opioids are considered the cornerstone of pain management in palliative care. Available data suggest that older patients use different analgesics and lower opioid doses compared to younger patients. However, it has not been elucidated yet whether such dosing is associated with worse pain levels or shorter survival in the palliative care setting. We evaluated the relationship among pain scores, quality of life, opioid dose, and survival in palliative care cancer patients in a hospice setting. A total of 137 palliative care cancer patients were analyzed prospectively. We divided patients into two groups using the age of 65 as a cut-off value. Younger patients exhibited significantly higher pain ratings (5.14 vs. 3.59, p=0.01), although older patients used almost 20 mg less oral morphine equivalent (OME) on arrival (p=0.36) and 55 mg OME/day less during the last week (p=0.03). There were no differences in survival between the two groups (17.36 vs. 17.58 days). The elderly patients also used nonsteroidal analgesics less often and paracetamol more often. Hence, using lower opioid doses in older palliative care cancer patients does not result in worse pain rating, and could be a plausible approach for pain management in this patient group.

THE CROATIAN HEALTH INSURANCE FUND DOES NOT RECOGNIZE DIFFERENCES IN THE COST OF DIFFERENT TREATMENTS FOR REVISION TOTAL HIP ARTHROPLASTY.

In recent years, there has been increased interest in the cost of treatment for revision interventions for hip and knee prostheses. In all publications so far, the authors note the high cost of treatment for revision interventions, especially if infection is present. The aim of this study was to compare the cost of treatment and health insurance reimbursements between revision total hip arthroplasty (THA) for infection and revision for aseptic indications (aseptic instability and periprosthetic fracture). Hospital data on 168 patients having undergone revision THA between 2010 and 2018 at the Department of Traumatology, Sestre milosrdnice University Hospital Centre from Zagreb were analyzed. Financial data were collected from the Hospital Information System. Financial analysis included total cost per patient, Croatian Health Insurance Fund reimbursements, cost of implants, and length of hospital stay. The difference between the mean total cost per patient and the mean Croatian Health Insurance Fund reimbursements was -262.83 € (-6.08%) for aseptic instability, -1694.94 € (-17.25%) for infection and -916.49 € (-17.33%) for periprosthetic fracture. The Croatian Health Insurance Fund does not recognize differences in the cost of revision THA for aseptic instability, infection and periprosthetic fracture. Health insurance reimbursement is inadequate for centers that offer revision hip surgery.

THE ASSOCIATION OF SUBCLINICAL INSULIN RESISTANCE WITH THYROID AUTOIMMUNITY IN EUTHYROID INDIVIDUALS.

Hashimoto thyroiditis is characterized by anti-thyroid peroxidase (anti-TPO) and anti-thyroglobulin (anti-Tg) antibodies that gradually lead to thyroid cell destruction. As hypothyroidism has been associated with insulin resistance (IR), we aimed to investigate whether IR is associated with thyroid antibody presence and whether the degree of IR correlates with their concentration in euthyroid individuals. A total of 164 non-diabetic, euthyroid individuals, average age 34 years, were included in the study, divided into two groups according to Hashimoto thyroiditis and underwent 5-hour oral glucose tolerance test. The degree of IR was evaluated by the Homeostatic Model Assessment of Insulin Resistance (HOMA-IR). The Hashimoto thyroiditis group had higher HOMA-IR (p=0.003) and lower glucose levels (p=0.04). HOMA-IR correlated positively with anti-TPO (p<0.001). Linear logistic regression revealed that anti-TPO concentration increased by 18.13 (p=0.001) with each HOMA-IR unit. IR might trigger thyroid antibody production and Hashimoto thyroiditis development, which needs to be evaluated in further larger scale follow up studies.

BURKITT LYMPHOMA IN GASTROINTESTINAL TRACT: A REPORT OF TWO CASES.

Burkitt lymphoma, a type of non-Hodgkin B-cell lymphoma, is the fastest growing human cancer, presenting pathologically with a 'starry sky' pattern. It is most often found in the abdomen and the jaw, however, localization in the abdomen other than the ileocecal area is very rare and described only in a handful of cases. Standard treatment consists of initial tumor cytoreduction followed by intense chemotherapy. Most of the relapses occur within one year of the diagnosis, while the 5-year survival is around 80%. We present two cases which are specific for unusual location of Burkitt lymphoma in the colon and stomach, in immunocompetent patients with negative Epstein-Barr virus tests. Also, one of the patients presented is one of the oldest ever reported with abdominal Burkitt lymphoma, while the other patient is an example of diagnostic difficulties in distinguishing Burkitt lymphoma from similar lymphomas. Due to the rapidly growing tumors and urgent need for cytoreductive surgery, it is crucial to consider the diagnosis of Burkitt lymphoma even in atypical localizations or absence of the common risk factors associated with Burkitt lymphoma.

TREATMENT OF HUMERAL SHAFT FRACTURES: ANTEGRADE INTERLOCKING INTRAMEDULLARY NAILING WITH ADDITIONAL INTERLOCKING NEUTRALIZATION SCREWS THROUGH FRACTURE SITE.

The aim of this study was to compare union time between two different nail designs for the treatment of humeral shaft fracture, i.e. antegrade interlocking intramedullary nail with and without additional interlocking neutralization screws. The retrospective study included 51 patients treated with antegrade humeral intramedullary nailing between January 2015 and December 2017. The inclusion criteria of the study were proximal and middle third humeral shaft fractures. Fifty-one patients met the inclusion criteria; 23 patients were treated with antegrade intramedullary nail with additional interlocking neutralization screws through fracture site (group A) and 28 patients were treated with antegrade intramedullary nail without additional interlocking neutralization screws (group B). Medical documentation and radiographic images taken preoperatively and postoperatively were reviewed. Radiological union was defined as cortical bridging of at least three of four cortices in two-plane radiographs, with disappearance of the fracture gap. There were no significant differences in union time between the groups (p>0.05). To our knowledge, this is the first report of antegrade interlocking humeral nailing with additional interlocking neutralization screws through fracture site. Hypothetical advantages of fracture gap reduction by additional interlocking neutralization screws to promote union were not confirmed by this first clinical trial.

LUMBAR SPINE SYNOVIAL CYST: A CASE SERIES REPORT AND REVIEW OF SURGICAL STRATEGIES.

Lumbar spine synovial cysts are benign growths adjoining the facet joints that may induce low back pain, lumbar radiculopathy and neurological deficit. However, they are not well defined concerning their origin, cause and pathology, as well as available treatment strategies. The scope of different surgical procedures includes image-guided epidural steroid injection, direct cyst puncture by percutaneous epidural needle, spinal canal decompression and cyst resection, and spinal bone fusion with/without instrumentation. Hereby, we report institutional experience and discuss surgical strategies of lumbar spine synovial cyst treatment. Presenting symptoms, imaging findings and outcomes were retrospectively analyzed in 15 patients with lumbar spine synovial cyst, operated on during a one-year period. The leading presenting symptom was lumbar radicular pain, while the most commonly involved vertebral level was L5-S1. In a great majority of patients, a single-level interlaminectomy and cyst resection were performed. Most patients recovered without postoperative neurological and functional deficit, as well as surgery-related complications. No poor outcome was noticed in our series. Concerning our results and literature review, the optimal management for patients with symptomatic lumbar synovial cyst has to be highly personalized, which is essential to achieve a favorable outcome. Nonetheless, the best treatment strategy has yet to be affirmed.

Microsurgical Decompression of Trigeminal Neuralgia Caused by Simultaneous Double Arterial (SCA and AICA) and Petrosal Vein Complex Compression.

Trigeminal neuralgia is a chronic pain disorder affecting the face. In approximately 80% of cases, it is most commonly caused, when the root entry zone (REZ) of the trigeminal nerve is compressed by the superior cerebellar artery (SCA). The etiology of the remaining 20% of cases is distributed among venous, arteriovenous malformations, posterior fossa tumors, multiple sclerosis plaque compressions, and other pathologies. Combinations of those compressive factors are very rare. 1 2 3 4 Herein, we present a video clip of microvascular decompression (MVD) in a 73-year-old female, who has failed conservative treatment with 6 medications over 10 years. She was affected by a unique triple compression of the right REZ by the SCA, anterior inferior cerebellar artery (AICA), and petrosal vein complex ( Fig. 1A ). Right-sided microsurgical decompression of the REZ of the trigeminal nerve through standard retrosigmoid craniotomy was performed by the senior author (K.I.A.). The SCA and AICA were separated from the nerve using Teflon pledgets. The petrosal vein complex was coagulated and divided, freeing up the right trigeminal nerve ( Fig. 1B ). The patient was discharged home on the third postoperative day with complete resolution of trigeminal neuralgia. The link to the Video can be found at: https://youtu.be/PYVvImGW0yE .

CURRENT PERSPECTIVES OF POTTER'S GLOBAL BIOETHICS AS A BRIDGE BETWEEN CLINICAL (PERSONALIZED) AND PUBLIC HEALTH ETHICS.

In the context of modern scientific and technological developments in biomedicine and health care, and the potential consequences of their application on humans and the environment, Potter's global bioethics concept resurfaces. By actualizing Potter's original thoughts on individual bioethical issues, the universality of two of his books, which today represent the backbone of the world bioethical literature, "Bioethics--Bridge to the Future" and "Global Bioethics: Building on the Leopold Legacy", is emphasized. Potter's global bioethics today can legitimately be viewed as a bridge between clinical personalized ethics on the one hand and ethics of public health on the other.