Phylogenetic evidence of a possible Trichuris globulosa species complex in Arabian camels from Kuwait.

During a 1-year study, Trichuris adults were obtained after necropsy of Arabian camels (Camelus dromedarius) from a slaughterhouse in Kuwait. Morphological and molecular identification was performed to confirm the identity of the Trichuris specimens obtained from C. dromedarius. Fifteen male Trichuris specimens were selected, and molecular identification was performed using mitochondrial cytochrome c oxidase subunit I, 12S ribosomal RNA, 16S ribosomal RNA genes and the nuclear internal transcribed spacer 2 (ITS2) region. Through phylogenetic analysis, 2 distinct groups were obtained using the mitochondrial genes, where group 1 showed a close relationship to Trichuris globulosa while group 2 showed a close relationship to Trichuris ovis, providing molecular evidence of a possible T. globulosa species complex. Additionally, the nuclear ITS2 region did not provide enough resolution to distinguish between the 2 groups of Trichuris specimens. Observation of morphological characters revealed variations in the shape of the male spicule sheath, where specimens present either a globular posteriorly truncated swelling or the absence of posteriorly truncated swelling. Moreover, the variations in male spicule sheath does not corroborate with the results of molecular data, suggesting the limited use of this character for identification of T. globulosa. In conclusion, molecular analysis suggests a possible species complex in T. globulosa, with the mitochondrial genetic markers successfully differentiating between the 2 groups. The limited use of the male spicule sheath as a diagnostic character for identification of T. globulosa is suggested.

Discovery of Encyclometra bungara (Digenea: Encyclometridae) in a new host (Enhydris enhydris) from Thailand and Cambodia through morphological and molecular identification.

The genus Encyclometra is one of the two genera in family Encyclometridae, known for parasitising the oesophagus, stomach and intestine of snakes. Among Encyclometra, the species present are: Encyclometra colubrimurorum, Encyclometra japonica, Encyclometra asymmetrica and Encyclometra bungara. Species discrimination within Encyclometra has predominantly relied on morphological differences, such as the length of the caeca and the position of the testes. Morphological overlaps exist among these species making species discrimination challenging. Additionally, the use of molecular information has been limited for Encyclometra. To determine the Encyclometra species infecting Enhydris enhydris from Thailand and Cambodia, morphological and molecular identification was conducted. Morphological characters and measurements were obtained from 30 Encyclometra adults, and they were compared with previous studies of other Encyclometra species. Novel sequences of E. bungara were generated using the nuclear 18S and 28S ribosomal RNA genes, and the mitochondrial cytochrome c oxidase subunit 1 gene. Our results revealed that the specimens could be morphologically identified as E. bungara, with support from molecular information obtained from the phylogenies of the 3 genetic markers employed. Molecular analysis showed that the Encyclometra specimens were distinct from E. colubrimurorum and E. japonica. Through morphological and molecular identification of the Encyclometra specimens found in E. enhydris from Thailand and Cambodia, we describe and provide a record of E. bungara in a new host and new locality. Additionally, novel molecular sequences were generated, revealing the phylogenetic position of E. bungara within the superfamily Gorgoderoidea.

Managing suspected myasthenia gravis and myositis induced by pembrolizumab in a Jehovah's Witness sarcoma patient.

INTRODUCTION: Immune checkpoint inhibitors (ICIs) can cause a spectrum of adverse events known as immune-related adverse events (irAEs) that resemble autoimmune responses. Immune-mediated myasthenia gravis (MG) is a rare and serious neurologic adverse event that has been associated with ICIs requiring prompt treatment. In the Jehovah's Witness population, typical management of these adverse events may not be options, and alternative treatment choices would be needed. CASE REPORT: 73-year-old Jehovah's Witness patient with high-grade undifferentiated pleiomorphic sarcoma who developed immune-mediated MG approximately 4 weeks after initiation of pembrolizumab. On the day of admission, the patient presented with a three-day history of worsening ptosis, right greater than left. He was later found to be seronegative for MG. MANAGEMENT AND OUTCOME: The patient required therapy with pyridostigmine, steroids, and agreed to plasma exchange (PLEX) prior to discharge. He achieved near resolution of his neurologic symptoms, and pembrolizumab was discontinued. He later underwent radical resection of the left thigh soft tissue sarcoma and superficial inguinal lymph node dissection. He is now on active surveillance. DISCUSSION: While neurologic adverse events typically present 6 weeks after initiation of ICIs, MG has been reported occurring as early as 4 weeks after initiation. This rare and serious adverse event requires prompt treatment, and clinicians need to be aware of the alternative treatment options in this unique patient population. Early conversations regarding blood products and factions must be had to develop a treatment plan in accordance with the patient's personal decisions.

The potential use of mitochondrial ribosomal genes (12S and 16S) in DNA barcoding and phylogenetic analysis of trematodes.

BACKGROUND: Genetic markers like the nuclear ribosomal RNA (rRNA) genes, internal transcribed spacer regions, mitochondrial protein-coding genes, and genomes have been utilized for molecular identification of parasitic trematodes. However, challenges such as the design of broadly applicable primers for the vast number of species within Digenea and the genetic markers' ability to provide sufficient species-level resolution limited their utility. This study presented novel and broadly applicable primers using the mitochondrial 12S and 16S rRNA genes for Digenea and aimed to show their suitability as alternative genetic markers for molecular identification of orders Plagiorchiida, Echinostomida, and Strigeida. RESULTS: Our results revealed that the mitochondrial 12S and 16S rRNA genes are suitable for trematode molecular identification, with sufficient resolution to discriminate closely related species and achieve accurate species identification through phylogenetic placements. Moreover, the robustness of our newly designed primers to amplify medically important parasitic trematodes encompassing three orders was demonstrated through successful amplification. The convenience and applicability of the newly designed primers and adequate genetic variation of the mitochondrial rRNA genes can be useful as complementary markers for trematode molecular-based studies. CONCLUSIONS: We demonstrated that the mitochondrial rRNA genes could be alternative genetic markers robust for trematode molecular identification and potentially helpful for DNA barcoding where our primers can be widely applied across the major Digenea orders. Furthermore, the potential of the mitochondrial rRNA genes for molecular systematics can be explored, enhancing their appeal for trematode molecular-based studies. The novelty of utilizing the mitochondrial rRNA genes and the designed primers in this study can potentially open avenues for species identification, discovery, and systematics in the future.

Insights into the genetic diversity of Angiostrongylus spp. causing human angiostrongyliasis and implications for molecular identification and diagnosis.

Angiostrongylus cantonensis and Angiostrongylus costaricensis are known human pathogens responsible for eosinophilic angiostrongyliasis and abdominal angiostrongyliasis, respectively. Humans are accidental hosts, where infection occurs through the consumption of the infective larva stage 3 in intermediate or paratenic hosts. The proven method for abdominal angiostrongyliasis diagnosis is the histological examination through tissue biopsy, while the diagnosis of eosinophilic angiostrongyliasis is the detection of larva in the cerebrospinal fluid. As there is molecular evidence of cryptic species within A. cantonensis and A. costaricensis lineages, along with morphological similarities within both lineages, accurate species identification and disease diagnosis may be challenging. Moreover, species within the lineages share similar intermediate and definitive hosts and geographic distribution. For example, both A. cantonensis and Angiostrongylus malaysiensis (a closely related species in A. cantonensis lineage) overlap in their geographic distribution in Southeast Asia. Additionally, variations in the molecular makeup of A. costaricensis and A. cantonensis lineages may impact the pathogenicity, infectivity, and disease severity of angiostrongyliasis. Understanding of the genetic diversity of both lineages is a cornerstone for improved diagnosis and disease intervention, especially in a changing global environment. To shed light and provide insights into the genetic diversity of the Angiostrongylus lineages causing human angiostrongyliasis, we aim to present an up-to-date review of the studies conducted and genetic markers used for A. costaricensis and A. cantonensis lineages. The implications for accurate molecular identification and diagnosis of human angiostrongyliasis are also discussed.

Molecular insights versus morphological traits: rethinking identification of the closely related Angiostrongylus cantonensis and Angiostrongylus malaysiensis.

BACKGROUND: The closely related Angiostrongylus cantonensis and Angiostrongylus malaysiensis have been reported to coexist in Thailand and share similar hosts and life cycles. Recently, in an angiostrongyliasis outbreak in Thailand, both A. cantonensis and A. malaysiensis were found in the cerebrospinal fluid of affected patients. Morphological similarities, overlapping distribution, shared hosts and habitats, and the close genetics of the two Angiostrongylus species can complicate accurate species identification. Addressing these challenges, this study aims to evaluate whether a correlation between the morphological and genetic identities of A. cantonensis and A. malaysiensis can improve species identification accuracy. METHODS: Angiostrongylus spp. specimens from five zoogeographical regions in Thailand were subjected to morphological and molecular identification using the mitochondrial cytochrome b gene and the nuclear internal transcribed spacer 2 region (ITS2). The morphological characters for males and females were then validated using the species identity obtained from the nuclear ITS2 region. RESULTS: The results revealed that morphological misidentifications between these two closely related species are common due to overlapping morphological characters. Although certain male traits such as body length and width aided species differentiation, female traits were found to be less reliable. Furthermore, hybrid forms (8.2%) were revealed through the ITS2 results, which can further complicate morphological identification. Mito-nuclear discordance was also present in 1.9% of the Angiostrongylus specimens from Thailand, suggesting a complex historical interbreeding between the species. CONCLUSIONS: Based on our findings, we suggest that nuclear ITS2 is a reliable marker for species identification of A. cantonensis and A. malaysiensis, especially in regions where both species coexist. Additionally, the scope and consequences of hybridization between the two closely related Angiostrongylus species should be further investigated in Thailand.

Detection of Gnathostoma spinigerum larva in the brain with complete follow-up after surgical treatment of human neurognathostomiasis.

Human gnathostomiasis is a food-borne zoonotic helminthic infection widely reported in Latin America, Asia, and Southeast Asia. Consuming raw, or under-cooked fresh-water fish is the leading cause of this helminthic infection, which is clinically characterized by signs of inflammation, itching sensation, or irritation with migratory swelling. Neurological symptoms resulting from neurognathostomiasis vary, and there is scant information due to the rareness of patient brain samples. This study aimed to demonstrate the first evidence of human neurognathostomiasis by the detection of Gnathostoma spinigerum larva in patient's brain during craniotomy, supported by histopathological, immunological and proteomic evidence. Clinical symptoms were obtained from medical history and physical examination with laboratory investigations, including magnetic resonance imaging (MRI), left temporal craniotomy, histopathology of brain tissue, and Western blot analysis, were performed to elucidate the causative pathogens for diagnosis. In addition, the host-parasite interaction of the parasite invading the patient's brain was characterized through proteomics. Histopathology revealed worms with the characteristic cuticular spines of G. spinigerum which were detected and identified. These histopathological findings were consistent with a positive Western blot showing a 24-kDa reactive-band for gnathostomiasis. Proteomic analysis revealed the presence of G. spinigerum serpin and serine protease in the patient's serum. Moreover, the leucine-rich alpha-2-glycoprotein was indicated as a systemic biomarker of early brain injury related to invasion by G. spinigerum. Therefore, our study provides the initial evidence of human neurognathostomiasis due to G. spinigerum larval invasion along with successful craniotomy and proven larval detection including complete follow-up, and the disease prognosis after surgical treatment.

From past to present: opportunities and trends in the molecular detection and diagnosis of Strongyloides stercoralis.

Strongyloides stercoralis is a soil-transmitted helminth that is mainly found in the tropical and subtropical regions and affects approximately 600 million people globally. The medical importance of strongyloidiasis lies in its capacity to remain asymptomatic and chronically unnoticed until the host is immunocompromised. Additionally, in severe strongyloidiasis, hyperinfection syndrome and larva dissemination to various organs can occur. Parasitological techniques such as Baermann-Moraes and agar plate culture to detect larvae in stool samples are the current gold standard. However, the sensitivity might be inadequate, especially with reduced worm burden. Complementing parasitological techniques, immunological techniques including immunoblot and immunosorbent assays are employed, with higher sensitivity. However, cross-reactivity to other parasites may occur, hampering the assay's specificity. Recently, advances in molecular techniques such as polymerase chain reaction and next-generation sequencing technology have provided the opportunity to detect parasite DNA in stool, blood, and environmental samples. Molecular techniques, known for their high sensitivity and specificity, have the potential to circumvent some of the challenges associated with chronicity and intermittent larval output for increased detection. Here, as S. stercoralis was recently included by the World Health Organization as another soil-transmitted helminth targeted for control from 2021 to 2030, we aimed to present a review of the current molecular techniques for detecting and diagnosing S. stercoralis in a bid to consolidate the molecular studies that have been performed. Upcoming molecular trends, especially next-generation sequencing technologies, are also discussed to increase the awareness of its potential for diagnosis and detection. Improved and novel detection methods can aid in making accurate and informed choices, especially in this era where infectious and non-infectious diseases are increasingly commonplace.

Predicting if Lung Cancer Will Relapse-The Role of Neutrophil/Lymphocyte Ratio.

OBJECTIVE/BACKGROUND: Baseline neutrophil/lymphocyte ratio (NLR), a surrogate marker for systemic inflammation and immunosuppression, is a well-studied prognostic marker in non-small cell lung cancer (NSCLC). This study tests if interim NLR is prognostic in NSCLC patients in remission. METHODS: This single-center, retrospective cohort study analyzed 131 NSCLC patients treated from 2010 to 2015 who achieved complete remission. NLR was calculated at baseline and from the first available blood sample during remission. Kaplan-Meier estimates of overall survival (OS) and time to recurrence were compared using the log-rank test for trend. Multivariable analysis was conducted using the Cox proportional hazards model. RESULTS: Of the 131 cases, 63 had subsequently recurred at the last follow-up. The mean age was 64 ± 10 years. Patients with stage I (35%), II (24%), and III (41%) were included. Histology were adenocarcinoma (60%), squamous cell (33%), and unspecified (7%). The majority of patients were smokers. For the univariate analysis interim NLR was binned into tertiles, NLR < 2, 2-4.08, and > 4.08. Of those with an interim NLR > 4.08, prognosis and recurrence risk were higher. In the multivariable analysis, remission NLR was strongly prognostic for OS ( p < .001) as did patient's age ( p = .002), but not stage, race, sex, and baseline NLR. CONCLUSIONS: Our study found that interim NLR, obtained in remission, was strongly prognostic for OS and recurrence.

Updates on the Treatment of Tenosynovial Giant Cell Tumor.

Tenosynovial giant cell tumor (TGCT) is a rare inflammatory disorder affecting the joint synovium, bursae, and tendon sheaths that causes non-specific and often insidious joint discomfort. The application of systemic chemotherapy has been limited due to poor and unsustained disease responses. Surgery with or without adjuvant radiation is the primary treatment modality for TGCT. With its locally destructive nature and increased recurrence, multiple surgical interventions become necessary throughout the course of the disease, leading to disfigurement, decreased quality of life, and increased mortality. However, owing to recent evidence demonstrating the overexpression of colony-stimulating factor 1 (CSF-1) in TGCT, selective tyrosine kinase inhibitors targeting CSF-1 receptors are being developed. Pexidartinib is the first CSF-1 receptor inhibitor approved for the treatment of TGCT. Here, we discuss various available treatment strategies and ongoing investigations and trials targeting diffuse TGCT, which include nilotinib, lacnotuzumab, cabiralizumab, vimseltinib, and emactuzumab.

Prevalence of Strongyloides in Southeast Asia: a systematic review and meta-analysis with implications for public health and sustainable control strategies.

BACKGROUND: Strongyloidiasis, caused by the nematodes Strongyloides stercoralis and Strongyloides fuelleborni, is estimated to affect over 600 million individuals worldwide. The disease is endemic in Southeast Asia, where a warm-humid climate and socio-economic conditions maintain the parasite's life cycle and transmission. However, the current diagnostic methods may not be sufficiently sensitive, suggesting that the true prevalence of strongyloidiasis could be seriously underestimated in this. This study aims to determine the prevalence of strongyloidiasis in Southeast Asia through a systematic review and meta-analysis and to discuss the implications of the estimated prevalence on diagnostic approaches and control strategies. METHODS: Following PRISMA guidelines, we conducted a systematic literature search in PubMed and Google Scholar databases to identify studies reporting Strongyloides prevalence data in the 11 Southeast Asian countries up to December 2022. A random effects model was employed to estimate the pooled prevalence of S. stercoralis at both regional and country levels. RESULTS: Out of 3722 articles identified, 224 met our inclusion criteria. For S. stercoralis specifically, we found 187 articles, of which 52.4% were from Thailand. All Southeast Asian countries, except Brunei, had at least one study on Strongyloides prevalence. The estimated pooled prevalence of S. stercoralis regionally was 12.7% (95% CI 10.70-14.80%), ranging from 0.4 to 24.9% at the country level. Cambodia had the highest pooled prevalence (24.9%, 95% CI 15.65-35.38%), followed by Lao PDR (16.5%, 95% CI 9.50-24.95%). Moreover, we obtained a pooled prevalence of 10% (95% CI 7.06-13.52%) in a group comprising immigrants, workers, and veterans from Southeast Asian countries. S. stercoralis infects various host types, including nonhuman primates, domestic dogs and cats, rodents, and transport carriers such as cockroaches and vegetables. CONCLUSIONS: A high prevalence of strongyloidiasis in Southeast Asia was revealed, highlighting the importance of the region's ongoing research, surveillance, and control efforts. Factors contributing to the strongyloidiasis transmission include the role of animal hosts, the impact of global connectivity, and the significance of the co-endemicity of other Strongyloides species. Based on these findings, a multi-pronged One-Health approach is essential for sustainable intervention and control.

Safety of immune checkpoint inhibitors in patients with preexisting autoimmune disorders.

In recent years, immune checkpoint inhibitors (ICIs) have been approved for a growing number of cancer types. Approximately a quarter of cancer patients have a concomitant diagnosis of autoimmune disorders (AID). Activation of the immune system with ICIs poses a potential risk of AID worsening, thus, the majority of the ICI clinical trials excluded these patients from the study. There is a paucity of data regarding the benefits and risks of ICIs in cancer patients with AIDs. The primary objectives of this study were to determine the incidence of immunotherapy-related AID worsening and all immunotherapy-related adverse events (irAEs). Secondary outcomes were time to AID worsening and survival difference. All adult patients (age >=18 years) with solid malignancy who received ICIs between Jan 2016 and June 2019 were identified using the University of Louisville pharmacy database. Medical records were reviewed to include all the patients with preexisting AIDs. Descriptive statistics were used to determine the incidence of AID worsening and all irAEs. Baseline characteristics were compared between cancer patients with vs without AID worsening using Pearson chi2 and Student's t-test, where appropriate. Multivariate Cox regression analysis was used to compare survival between the 2 groups. A total of 40 patients with AIDs were identified during the study period. The cancer types were melanoma (57.5%), lung (15%), breast (5%), and others (22.5%). AIDs were rheumatological (52.5%), dermatologic (20%), gastroenterological (12.5%), neurologic (12.5%), and hematological (2.5%). The incidence of all irAEs was 60% (grade >=3 in 20%) and AID worsening was 40% (grade >=3 in 15%). The median time from ICI initiation to AID worsening was 94.5 (range 21-431) days. In multivariate Cox regression analysis, adjusted for demographics, cancer type, and stage, survival was similar for patients who had AID worsening vs those who did not (HR 0.30 (95%CI 0.06-1.40, P = 0.128). In our single-institution study, cancer patients with preexisting AID do have an increased risk of irAEs with high-grade toxicities in 20% compared to historically reported 5% in the general cancer population. About 15% of patients reported grade >=3 worsening of their AIDs. Although the risk of irAEs is numerically higher in patients with AIDs, it may be acceptable to the patients if the potential benefits of ICIs outweigh the risks.

Palliative Referrals in Advanced Cancer Patients: Utilizing the Supportive and Palliative Care Indicators Tool and Rothman Index.

BACKGROUND: Timely identification of palliative care needs can reduce hospitalizations and improve quality of life. The Supportive & Palliative Care Indicators Tool (SPICT) identifies patients with advanced medical conditions who may need special care planning. The Rothman Index (RI) detects patients at high risk of acutely decompensating in the inpatient setting. SPICT and RI among cancer patients were utilized in this study to evaluate their potential roles in palliative care referrals. METHODS: Advanced cancer patients admitted to an institution in Baltimore, Maryland in 2019 were retrospectively reviewed. Patient demographics, length of hospital stay (LOS), palliative care referrals, RI scores, and SPICT scores were obtained. Patients were divided into SPICT positive or negative and RI > 60 or RI < 60.Unpaired t-tests and chi-square tests were utilized to determine the associations between SPICT and RI and early palliative care needs and mortality. RESULTS: 227 patients were included, with a mean age of 68 years, 63% Black, 59% female, with the majority having lung and GI malignancies. Sixty percent were SPICT +, 21% had RI < 60. SPICT + patients were more likely to have RI < 60 (p = 0.001). SPICT + and RI < 60 patients were more likely to have longer LOS, change in code status, more palliative/hospice referrals, and increased mortality (p <0.05). CONCLUSIONS: SPICT and RI are valuable tools in predicting mortality and palliative/hospice care referrals. These can also be utilized to initiate early palliative and goals of care discussions in patients with advanced cancer.

Sensitive and accurate DNA metabarcoding of parasitic helminth mock communities using the mitochondrial rRNA genes.

Next-generation sequencing technologies have accelerated the pace of helminth DNA metabarcoding research, enabling species detection in bulk community samples. However, finding suitable genetic markers with robust species-level resolution and primers targeting a broad species range among parasitic helminths are some of the challenges faced. This study aimed to demonstrate the potential use of the mitochondrial 12S and 16S rRNA genes for parasitic helminth (nematodes, trematodes, cestodes) DNA metabarcoding. To demonstrate the robustness of the 12S and 16S rRNA genes for DNA metabarcoding, we determined the proportion of species successfully recovered using mock helminth communities without environment matrix and mock helminth communities artificially spiked with environmental matrices. The environmental matrices are human fecal material, garden soil, tissue, and pond water. Our results revealed the robustness of the mitochondrial rRNA genes, through the high sensitivity of the 12S rRNA gene, and the effectiveness of the 12S and 16S primers targeting platyhelminths. With the mitochondrial rRNA genes, a broad range of parasitc helminths were successfully detected to the species level. The potential of the mitochondrial rRNA genes for helminth DNA metabarcoding was demonstrated, providing a valuable gateway for future helminth DNA metabarcoding applications like helminth detection and biodiversity studies.

Assessing the suitability of mitochondrial and nuclear DNA genetic markers for molecular systematics and species identification of helminths.

BACKGROUND: Genetic markers are employed widely in molecular studies, and their utility depends on the degree of sequence variation, which dictates the type of application for which they are suited. Consequently, the suitability of a genetic marker for any specific application is complicated by its properties and usage across studies. To provide a yardstick for future users, in this study we assess the suitability of genetic markers for molecular systematics and species identification in helminths and provide an estimate of the cut-off genetic distances per taxonomic level. METHODS: We assessed four classes of genetic markers, namely nuclear ribosomal internal transcribed spacers, nuclear rRNA, mitochondrial rRNA and mitochondrial protein-coding genes, based on certain properties that are important for species identification and molecular systematics. For molecular identification, these properties are inter-species sequence variation; length of reference sequences; easy alignment of sequences; and easy to design universal primers. For molecular systematics, the properties are: average genetic distance from order/suborder to species level; the number of monophyletic clades at the order/suborder level; length of reference sequences; easy alignment of sequences; easy to design universal primers; and absence of nucleotide substitution saturation. Estimation of the cut-off genetic distances was performed using the 'K-means' clustering algorithm. RESULTS: The nuclear rRNA genes exhibited the lowest sequence variation, whereas the mitochondrial genes exhibited relatively higher variation across the three groups of helminths. Also, the nuclear and mitochondrial rRNA genes were the best possible genetic markers for helminth molecular systematics, whereas the mitochondrial protein-coding and rRNA genes were suitable for molecular identification. We also revealed that a general gauge of genetic distances might not be adequate, using evidence from the wide range of genetic distances among nematodes. CONCLUSION: This study assessed the suitability of DNA genetic markers for application in molecular systematics and molecular identification of helminths. We provide a novel way of analyzing genetic distances to generate suitable cut-off values for each taxonomic level using the 'K-means' clustering algorithm. The estimated cut-off genetic distance values, together with the summary of the utility and limitations of each class of genetic markers, are useful information that can benefit researchers conducting molecular studies on helminths.

Co-occurrence of Angiostrongylus malaysiensis and Angiostrongylus cantonensis DNA in cerebrospinal fluid: Evidence from human eosinophilic meningitis after ingestion of raw snail dish in Thailand.

Angiostrongylus cantonensis, the main causative agent of human neuroangiostrongyliasis, is a food-borne parasitic zoonosis, particularly in Southeast Asia and Mainland China. Angiostrongylus malaysiensis, a cryptic species, has not been unequivocally identified as a causative agent for human angiostrongyliasis. Here, we investigated a local incidence of human angiostrongyliasis in Kalasin Province, northeastern part of Thailand. Field and laboratory investigations, clinical symptoms, and treatment of the disease are also discussed. Five sera and three cerebrospinal fluid samples were taken from each patient who displayed clinical symptoms of mild or severe headache without neck stiffness after ingesting a local dish containing Pila virescens. With molecular evidence using PCR and DNA sequencing approaches, we confirmed the presence of A. malaysiensis and A. cantonensis DNA in the patient samples. In addition, P. virescens and Pomacea canaliculata collected in the vicinity were also examined for the existence of angistrongylid larvae. The rate of infection in the snail population was 33.3% (18 infection out of 54 examined), with A. cantonensis as the predominant species. Notably, two snails were found to be co-infected with both A. malaysiensis and A. cantonensis. This discovery comes after several years of suspicion that it could be a zoonotic pathogen. Therefore, our findings are important for public health and clinical diagnosis since clinicians are not aware of the zoonotic potential of A. malaysiensis in humans.

Newly developed SYBR Green-based quantitative real-time PCRs revealed coinfection evidence of Angiostrongylus cantonensis and A. malaysiensis in Achatina fulica existing in Bangkok Metropolitan, Thailand.

Angiostrongylus cantonensis is a well-known pathogen causing eosinophilic meningitis associated with angiostrongyliasis. Humans, as accidental hosts, are infected by consuming undercooked snails containing third-stage larvae. A. malaysiensis is closely related to A. cantonensis and has been described as a potential human pathogen. The two species distribution was recently reported to overlap in the same endemic area, particularly in the Indochina Peninsula. Similar morphological characteristics of the third-stage larva in the snail-intermediate host often lead to misidentification of the two species. Thus, we aimed to develop a sensitive and specific method to detect and discriminate Angiostrongylus third-stage larva by designing species-specific primers based on the mitochondrial cytochrome b gene. We developed the SYBR Green quantitative real-time PCR (qPCR) method for two species-specific detection assays, which could be conducted simultaneously. The method was subsequently employed to detect and identify third-stage larvae of Angiostrongylus isolated from infected Achatina fulica collected from six public parks in Bangkok Metropolitan, Thailand. The method was also a preliminary applied to detect parasite tissue debris in the patients' cerebrospinal fluid (CSF). SYBR Green qPCRs quantitatively detected approximately 10-4 ng of genomic DNA from one larva, facilitating species-specific detection. Based on the pools of third-stage larvae isolated individually from the tissue of each infected A. fulica collected from the public parks, the qPCR results revealed that A. malaysiensis was the predominant species infecting 5.26% of the collected snails. In comparison, coinfection between A. malaysiensis and A. cantonensis was 5.97%, and no single infection of A. cantonensis was detected in A. fulica. Our SYBR Green qPCR method is a useful and inexpensive technique for A. cantonensis and A. malaysiensis discrimination, and the method has sufficient sensitivity to detect isolated larvae from a snail-intermediate host. The ratio of A. cantonensis and A. malaysiensis larvae infecting the snails can also be estimated simultaneously. Our qPCRs can be employed in a molecular survey of A. cantonensis and A. malaysiensis within intermediate hosts and for clinical diagnosis of angiostrongyliasis with CSF specimens in future studies.

Outcome reporting in therapeutic mammaplasty: a systematic review.

BACKGROUND: Therapeutic mammaplasty (TM) is an oncological procedure which combines tumour resection with breast reduction and mastopexy techniques. Previous systematic reviews have demonstrated the oncological safety of TM but reporting of critically important outcomes, such as quality of life, aesthetic and functional outcomes, are limited, piecemeal or inconsistent. This systematic review aimed to identify all outcomes reported in clinical studies of TM to facilitate development of a core outcome set. METHODS: Medline, EMBASE, CINAHL and Web of Science were searched from inception to 5 August 2020. Included studies reported clinical outcomes following TM for adult women. Two authors screened articles independently for eligibility. Data were extracted regarding the outcome definition and classification type (for example, oncological, quality of life, etc.), time of outcome reporting and measurement tools. RESULTS: Of 5709 de-duplicated records, 148 were included in the narrative synthesis. The majority of studies (n = 102, 68.9 per cent) reported measures of survival and/or recurrence; approximately three-quarters (n = 75, 73.5 per cent) had less than 5 years follow-up. Aesthetic outcome was reported in half of studies (n = 75, 50.7 per cent) using mainly subjective, non-validated measurement tools. The time point at which aesthetic assessment was conducted was highly variable, and only defined in 48 (64.0 per cent) studies and none included a preoperative baseline for comparison. Few studies reported quality of life (n = 30, 20.3 per cent), functional outcomes (n = 5, 3.4 per cent) or resource use (n = 28, 18.9 per cent). CONCLUSION: Given the oncological equivalence of TM and mastectomy, treatment decisions are often driven by aesthetic and functional outcomes, which are infrequently and inconsistently reported with non-validated measurement tools.

A Case of COVID-19 Complicated by Venous Thromboembolism Despite Being on Prophylactic Anticoagulation.

The coronavirus disease 2019 (COVID-19) presents with variable clinical syndromes, from asymptomatic disease to acute respiratory failure. Complications such as acute respiratory distress syndrome (ARDS), secondary infection, acute cardiac injury, liver failure, renal failure, coagulable disorders and in many cases, death have been recognized. Venous thromboembolism (VTE) was found to be present in up to one-third of critically ill COVID-19 patients. Our case demonstrated a patient with COVID-19 who developed a massive pulmonary embolism leading to cardiac arrest, despite having an initial normal D-dimer level, normal chest imaging, and while maintained on appropriate VTE prophylaxis during hospitalization.