Further stratification analyses indicated that pretherapeutic patients bearing vimentin (Vim)+ CTECs  at baseline revealed a significantly shortened mPFS compared with patients with Vim– CTECs. Post‐therapeutic patients harboring epithelial cell adhesion molecule (EpCAM)+ CTCs and CTECs , regardless of Vim expression or not, showed a significantly reduced mPFS. Post‐therapeutic patients possessing de novo EpCAM+/Vim+ (hybrid E/M‐type) CTECs displayed the shortest mPFS. Patients harboring either pre‐ or post‐therapeutic EpCAM–/Vim– null CTECs (N‐type) exhibited a better response to therapy compared to patients harboring EpCAM+ and/or Vim+ CTECs. The presented results support the notion that baseline Vim+ CTECs and post‐therapeutic EpCAM+ CTCs and CTECs are predictive biomarkers for longitudinal monitoring of response to anti‐angiogenesis combination regimens in NSCLC patients.Prognosticating the efficacy of anti‐angiogenic therapy through longitudinal monitoring and early detection of treatment resistance in cancer patients remain highly challenging. In this study, co‐detection and comprehensive phenotypic and karyotypic molecular characterization of aneuploid circulating tumor cells (CTCs) and circulating tumor endothelial cells (CTECs) were conducted on non‐small cell lung cancer (NSCLC) patients receiving bevacizumab plus chemotherapy. Prognostic values of the cell‐based significant univariate risk factors identified by Cox regression analyses were progressively investigated. Subjects showing an increase in total post‐therapeutic platelet endothelial cell adhesion molecule‐1 (CD31) − CTCs and CD31+ CTECs, a pair of mutually related and interacting cellular circulating tumor biomarkers, play a critical role in tumorigenesis, neovascularization, and cancer metastasis. Longitudinally comprehensive co‐detection of diverse subtypes of CTCs and CTECs during therapy by iFISH may effectively prognosticate and real‐time monitor therapeutic efficacy or emerging resistance in cancer patients subjected to anti‐angiogenic combination regimens.Aneuploid CD31 CTCcirculating tumor cellCTECcirculating tumor endothelial cellEMTepithelial‐to‐mesenchymal transitionEndoMTendothelial‐to‐mesenchymal transitioniFISHin situ hybridizationimmunostaining fluorescence metsmetastasesTEMtumor endothelial cell‐specific marker1Tumor neovascularization is primarily composed of endothelium‐dependent angiogenesis and vasculogenesis . NeoangiEndothelial cells in tumor vasculature are known as tumor ECs (TECs) , 5, showBoth epithelial‐to‐mesenchymal transition (EMT) and endothelial‐to‐mesenchymal transition (EndoMT)  are cent®), the only approved monoclonal antibody for anti‐angiogenic therapy in first‐line treatment of eligible advanced lung cancer patients . L. L+/Vim−otential . An EpCA3.2t1 = (t1 − t0)/t0 vs ∆t2 = (t2 − t0)/t0. Among 23 patients, two subjects were excluded for follow‐up analysis because follow‐up time was < 6 months  and descending  cohorts, respectively. Subjects who did not have t2 values available were classified upon t1 > t0 (ascending) or t1 < t0 (descending) for CTCs or CTECs. As graphically depicted in Fig. Quantitative variations in CTCs, CTECs, and their subtypes expressing EpCAM or vimentin in patients throughout therapy were analyzed. Changes in CTC and CTEC numbers were analyzed by comparison of ∆ths Fig. . There wLongitudinal variation of cell numbers and percentages of the specific subtype of CTC and CTEC expressing EpCAM or vimentin in both ascending and descending cohort are displayed in the heattable in Fig. + and – CTCs) in the ascending cohort were 6 , 3 , and 12 cells , respectively. Although median values of CTCs in this cohort did not show a statistically significant change (P = 0.210), they all had the same upward variation trend in the ascending cohort of patients. Among all CTCs in the ascending cohort, the number of EpCAM+ CTCs (red) considerably increased from 0 (t0‐1) to 18 cells (t2). Regarding the descending cohort in Fig. t0), 14 , and 3 cells . The difference in median values of total CTCs between t1 and t2 was statistically significant (**P = 0.008). EpCAM+ CTCs (red) in the descending cohort showed a similar variation pattern to that of overall CTCs, increasing from 2 (t0) to 13 cells (t1), then decreasing to 0 (t2). Vim+ CTCs in the same cohort (blue) decreased from 3 cells (t0‐1) to 1 cell (t2).Further analysis of the total numbers of CTCs, CTECs, and their subtypes is depicted in Fig. t0), 2 , and 6 cells , P = 0.642. In the same ascending cohort, an inverse variation pattern was found in EpCAM+ and Vim+ CTECs detected from t0 to t2 with Vim+ CTECs increasing at t1 and decreasing again at t2, whereas EpCAM+ CTECs slightly decreased at t1 and increased again at the same time point of t2. Regarding the descending cohort, the number of total CTECs had a median value of 6 , 9 , and 1 cell , respectively. Unlike CTCs (P = 0.210) and CTECs (P = 0.642) in the ascending cohort, differences in the median values of total CTECs between t0 and t2 (*P = 0.013), and between t1 and t2 (**P = 0.002) were statistically significant. Similar to the ascending cohort, the quantitative variation of Vim+ CTECs in the descending cohort displayed a similar upward (t1)‐downward (t2) pattern showing 12 (t0), 16 (t1), and 1 cell (t2), whereas the amount of EpCAM+ CTECs steadily decreased, revealing from 14 (t0), 12 (t1), then to 2 cells (t2). The quantitative variation of the post‐therapeutic EpCAM+ CTCs and CTECs was in line with the cell number change in total CTCs and CTECs in both ascending and descending cohorts of patients.As shown in Fig. + CTECs distinctly exhibited a unified upward (t1)‐downward (t2) biphasic response pattern in both ascending and descending cohorts, which was also reported on breast cancer patients receiving neoadjuvant chemotherapy [+ CTCs and CTECs in both ascending and descending cohorts of NSCLC patients treated with anti‐angiogenic combination regimen consistently matched the total CTCs’ and CTECs’ fluctuation from t0 to t2.The results of Fig. otherapy . However+ or Vim+ CTCs and CTECs shown in Fig. + CTCs detected at t2 in the ascending cohort, 14 of them (14/18 = 77.8%) were small cell‐sized SCTCs (CD31−/CD45−) and all 14 (100%) were diploid or near‐diploid (nonaneuploid) SCTCsdi [t1, 12 out of 13 (12/13 = 92.3%) EpCAM+ CTCs were large cell‐sized LCTCs and 11 of them (11/12 = 91.7%) were multiploid LCTCsmulti. Regarding CTECs in the ascending cohort, all five baseline (t0) EpCAM+ CTECs were large cells (100%), with four being multiploid LCTECsmulti (4/5 = 80%). Four out of five (4/5 = 80%) baseline Vim+ CTECs in the ascending cohort were large cells, with 3 of them (3/4 = 75%) being multiploid LCTECsmulti. In the same ascending cohort at t1, nine out of 12 Vim+ CTECs (9/12 = 75%) were sCTECs and seven of the nine (7/9 = 77.8%) were diploid/near‐diploid sCTECsdi, whereas all the particular EpCAM+ CTECs (t1) were LCTECsmulti (3/3 = 100%). Detection at t2 in this cohort showed that all seven EpCAM+ CTECs (100%) were LCTECs with six of them (6/7 = 85.7%) being LCTECsmulti. In the descending cohort, all 14 (100%) baseline EpCAM+ CTECs were large cells and 12 of them (12/14 = 85.7%) were LCTECsmulti. Among another 12 baseline Vim+ CTECs, eight of them (8/12 = 66.7%) were LCTECs and all eight (100%) were LCTECsmulti. At t1, all 12 EpCAM+ CTECs (100%) were LCTECsmulti. Fourteen out of 16 Vim+ CTECs at t1 (14/16 = 87.5%) were LCTECs with half of them (7/14 = 50%) being LCTECsmulti. Obtained results indicated that in contrast to EpCAM+ CTCs heterogeneous in both cell size  and chromosome ploidy (diploid and multiploid chr8), most EpCAM+ CTECs homogeneously exhibited as multiploid large cells (LCTECsmulti).As illustrated in Table CTCsdi , 41. In 3.3The above analysis illustrates how quantities of CTCs and CTECs as well as cell numbers of their specific subtypes varied in the ascending and descending cohorts. Accordingly, correlation of quantitative variation of CTCs, CTECs, and their various subtypes with patients’ prognosis was analyzed in this section and in i.e., ascending vs descending) in patients following therapy was the cell‐based significant univariable risk factor, indicating the risk of rapid disease progression (shortened PFS) and rapid patients’ death (reduced OS) for the ascending cohort was 7  and 11.6  times higher than that of descending cohort, respectively. Because ascending and descending cohorts classified by CTC or CTEC variation trend had identical subjects due to the concurrent upward or downward changes in patients  had a shorter median PFS (mPFS) of 3.0 months (95% CI: 0.4–5.6 months) compared to 9.4 months (95% CI: 7.8–11.1 months) in the descending cohort . Eleven out of 21 subjects were fatal cases in this study. The ascending cohort (N = 7) had a median OS (mOS) of 7.0 months (95% CI: 0.8–13.2 months) compared to 24.3 months (95% CI: 13.6–34.8 months) mOS in the descending cohort . mPFS and mOS for both CTC‐ and CTEC‐categorized cohorts were identical as explained above. Obtained results indicated that the ascending cohort of patients had a poorer response to bevacizumab in terms of both reduced mPFS and mOS.A Cox proportional hazards regression model analysis was performed to identify significant univariable risk factors indicated in red font in Fig. 3.4+ and Vim+ CTCs and CTECs in the same cohort of patients correlated with patients’ response to anti‐angiogenesis regimen.The above analysis illustrates how quantitative changes in post‐therapeutic total CTCs and CTECs were relevant to prognosis in both ascending and descending cohorts of patients. Further analysis was performed to investigate whether the existence of the particular subtypes of EpCAMt0), consisting of EpCAM−/Vim+ , EpCAM−/Vim− , and hybrid EpCAM+/Vim+  CTCs. Following combination therapy, a cohort of patients having the de novo subtype of EpCAM+/Vim− (E‐type) CTCs emerged . The majority of these patients were subjects containing EpCAM−/Vim− N‐type null CTCs at baseline, and a minority possessed M‐type CTCs prior to therapy (white arrows). Most patients who contained the emerged E‐type CTCs (red arrow) and the subject possessing the E/M‐type CTCs (black arrow) following therapy (t1‐2) were toward poor prognosis (mPFS < 9.4 months). As depicted in Fig. t0. A new subtype of EpCAM+/Vim+ CTECs  was detected in all three baseline cohorts following therapy . All patients who had the de novo identified subtype of E/M‐type CTECs (red arrow) and a majority of subjects having the E‐type CTECs following therapy (black arrow) exhibited an inferior prognosis. Most patients who had post‐therapeutic aneuploid N‐type null CTECs during therapy showed a better response to treatment .Longitudinal analysis of subcategorized cohorts of subjects hosting diverse subtypes of CTCs and CTECs at the designated time intervals during therapy is graphically shown in Sankey diagrams Fig. Aa‐b. The+ CTCs or CTECs independently of vimentin expression showed a poorer prognosis (mPFS < 9.4 months), whereas the majority of those who possessed EpCAM− CTECs following therapy, exhibited an improved survival.Sankey diagram graphical analysis indicated that patients who had post‐therapeutic EpCAMP = 0.652). Post‐therapeutic CTCs revealed subtypes comprising all possible four combinations of EpCAM and vimentin expression phenotypes. Seven out of eight patients (7/8 = 87.5%) harboring EpCAM+ CTCs following therapy, regardless of vimentin expression or not, exhibited a poorer prognosis. Nine of the remaining 13 patients (9/13 = 69.2%) possessing post‐therapeutic EpCAM− CTCs had no new mets at PD. Correlation of post‐therapeutic EpCAM+ CTCs with patients’ poorer prognosis was statistically significant (*P = 0.037).Further quantitative analysis was performed to examine correlations of different cohorts of patients bearing stratified subtypes of CTCs and CTECs with disease progression. As illustrated in Fig. − CTECs, 10 of them (10/15 = 66.7%) showed no new mets at PD. Correlation of positive detection of baseline Vim+ CTECs with poorer prognosis was statistically significant (*P = 0.023). Concerning 21 follow‐up patients, 10 of 11 subjects harboring post‐therapeutic EpCAM+ CTECs (9/11 = 81.8%) revealed a poorer prognosis regardless of vimentin expression or not. Among remaining 10 patients bearing EpCAM− CTECs, nine subjects (8/10 = 80%) did not have new mets at PD. Correlation between the existence of post‐therapeutic EpCAM+ CTECs and the poorer prognosis was statistically significant (*P = 0.017).Regarding baseline CTECs, all six patients (100%) possessing M‐type CTECs at baseline had a poorer prognosis. Among the remaining 15 patients who possessed baseline Vim+ CTCs and CTECs as well as Vim+ CTECs were relevant to patients’ poor response to treatment. The observation was further confirmed by Cox regression analysis for risk stratification. As revealed in Fig. P < 0.05, red), showing that subjects bearing baseline Vim− CTECs , post‐therapeutic Vim− CTECs , post‐therapeutic EpCAM− CTECs  or EpCAM− CTCs , had a lower risk (HR < 1) for a rapid disease progression compared to cohorts containing Vim+ or EpCAM+ cells. Accordingly, dichotomized Kaplan–Meier survival analysis was performed to further investigate how Vim+ or EpCAM+ CTCs and CTECs in all recruited NSCLC patients responded to bevacizumab combination therapy  showed a reduced mPFS of 5.0 months (95% CI: 0–10.4 months) compared to 10.1 months (95% CI: 1.6–30.6 months) in those without baseline Vim+ CTECs . Although Cox regression analysis revealed post‐therapeutic Vim+ CTECs as a significant univariable risk factor, Kaplan–Meier survival analysis did not show a significant difference on mPFS between post‐therapeutic Vim+ and Vim− CTECs cohorts (P = 0.132), possibly due to small sample size. Further K‐M survival analysis was performed to examine post‐therapeutic EpCAM+vs EpCAM− cohorts, regardless of vimentin expression or not. As depicted in Fig. + CTECs following therapy (N = 11) showed a reduced mPFS of 5.6 months (95% CI: 1.6–9.4 months), whereas those who had no EpCAM+ CTECs (N = 14) revealed a prolonged mPFS of 18.3 months (95% CI: 2.4–34.2 months) . Regarding post‐therapeutic CTCs, as demonstrated in Fig. + CTCs (N = 9) exhibited a mPFS of 5.6 months (95% CI: 3.8–7.4 months) compared to 10.1 months (95% CI: 3.2–17.0 months) in patients without post‐therapeutic EpCAM+ CTCs .The above analyses demonstrated that EpCAMapy Fig. . As depiP = 0.027  to the vasculature of malignant tumors remains a challenging topic . Aneuplorognosis , 45. In rognosis , 12, 46,rognosis . In thist2 correlated with a prolonged mPFS and mOS. The obtained results implied that both the total amount of aneuploid CTECs and CTCs detected after four‐to‐six treatment cycles (t2) may be able to function as an indicative biomarker to timely index patients’ response to combination regimen.In contrast to markedly reduced overall circulating ECs in chemotherapeutic breast cancer patients at PD , the prein situ phenotypic and karyotypic characterization performed by iFISH was applied to investigate whether and how the prognosticators vimentin [+ CTECs with multiploid chr8 exhibited a predictive value for patients’ poor prognosis, showing that all patients harboring this subtype of CTECs prior to treatment had new mets at PD  in the ascending cohort, as revealed in Table + CTCs and CTECs in post‐therapeutic lung cancer patients is of particular clinical utility, in terms of timely detecting emerging therapeutic resistance and appraising patients’ poor response to combination therapy. The conclusion was supported by Kaplan–Meier survival analysis, demonstrating that post‐therapeutic EpCAM+ CTECs and CTCs significantly correlated with patients’ reduced mPFS  models [+ and/or Vim+ CTCs and CTECs in cancer patients during therapy.The Sankey diagram analysis provides a convenient graphical approach for retrospectively tracing and analyzing how different cohorts of patients are relevant to each other and to correlate with the ultimate outcome throughout therapy. Sankey analysis Fig.  displaye) models  treated n = 47) had aneuploid EpCAM−/vimentin− null ECs detected in peripheral blood with an average of 2.8 cells compared with 100% in all carcinoma patients (n = 133) with an average of 8.8 cells, and none of EpCAM+ or Vim+ circulating CD31+ cells were detected in healthy donors [−/Vim− circulating ECs observed in healthy donors might also exist in cancer patients. Following the emergence of clinical utilities of EpCAM+ and/or vimentin+ aneuploid CTEC subtypes revealed in this study, several intriguing questions require further single cell‐based molecular unraveling, such as whether aneuploid CECs in healthy subjects are precancerous‐related cells and can be homeostatically depleted by immune scavengers of the host defense system [We previously reported that about 66% healthy subjects  targeting the EGFR‐T790M resistance mutation [The present study reported the clinical relevance of CTCs and CTECs, particularly EpCAMmutation , and themutation , as wellmutation , 61.5− CTCs and CD31+ CTECs may function as a pair of cellular circulating tumor biomarkers in predicting and prognosticating NSCLC patients subjected to anti‐angiogenic combination therapy. An upward trend in terms of an increase in total quantified post‐therapeutic CTC and CTEC numbers correlates with patients’ poor response to bevacizumab, showing a reduced mPFS and mOS. Existence of the specific subtype of vimentin+ mesenchymal CTECs at baseline, correlating with poor response to anti‐angiogenic therapy, might be an adverse biomarker in appropriately selecting eligible subjects. Furthermore, compared to conventional clinical imaging evaluation on tumor mass, positively detected post‐therapeutic EpCAM+ CTECs and CTCs, particularly de novo EpCAM+/Vim+ E/M‐type CTECs, may be indicative biomarkers in terms of timely indexing therapeutic efficacy and detecting emerging resistance to treatment. A majority of patients who displayed either pre‐ or post‐therapeutic aneuploid EpCAM−/Vim− N‐type null CTECs during therapy exhibited a better response to the combination regimen. In contrast to CD31− CTCs, which are highly heterogeneous in both cell sizes and degrees of aneuploidy, the main population of CD31+ CTECs are homogeneous, large multiploid cells .The present study provides proof of the concept showing that aneuploid CD31® is the registered trademarks of Cytelligen. Dr. Peter P. Lin is the president at Cytelligen. None of authors owns Cytelligen's stock shares. No additional COI to be disclosed.i•FISHTZ contributed conceptualization, validation, formal analysis, investigation, resources, funding acquisition, and writing—original draft; LZ contributed conceptualization, methodology, validation, formal analysis, investigation, data curation, funding acquisition, writing—original draft, visualization, and funding acquisition; YG contributed validation, formal analysis, investigation, resources, and project administration; YW contributed investigation and data curation; YL contributed investigation and data curation; HZ contributed conceptualization and investigation; QW contributed conceptualization and investigation; FH contributed conceptualization and investigation; JL contributed conceptualization and investigation; JT contributed formal analysis and software; DDW contributed methodology and validation; OG contributed writing—original draft, review, editing, and revising manuscript critically for important intellectual content; PPL contributed conceptualization, visualization, writing—original draft, review, and editing; BL contributed conceptualization, resources, and supervision. All the authors read and approved the final version of manuscript.https://publons.com/publon/10.1002/1878‐0261.13092.The peer review history for this article is available at Fig. S1. Risk stratification by Cox proportional hazards regression model analysis. Risk factors including non‐cell‐based gender, age, staging, and cell‐based quantitative variation trend of CTCs or CTECs (ascending vs descending cohort), positive detection of EpCAM+ CTCs or CTECs either prior to or post‐therapy, vimentin+ CTCs or CTECs either prior to or post‐therapy, are analyzed by the Cox regression analysis. *Results of Cox regression analysis performed on ascending vs descending cohorts classified by CTECs are identical to CTCs. Significant univariable risk factors (P < 0.05) are indicated in red font. HR: hazard ratio, HR > 1: higher risk, HR < 1: lower risk; n/a: not available, no death occurred.Click here for additional data file.Fig. S2. Multistrata Kaplan–Meier survival analyses. (a) Baseline CTECs: patients with EpCAM−/Vim+ M‐type CTECs have a mPFS of 5.0 months compared to 9.2 months and 16.1 months in the cohorts of EpCAM+/Vim− E‐type and EpCAM−/Vim− null cell, *P = 0.027 (log‐rank test). (b) Post‐therapeutic CTECs: patients having EpCAM+/Vim+ (hybrid E/M‐type) CTECs reveal the shortest mPFS of 3.0 months. The other three cohorts including EpCAM+/Vim− (E‐type), EpCAM−/Vim+ (M‐type) and EpCAM−/Vim− null CTECs show the mPFS of 6.5, 9.2 and 18.3 months, respectively, **P = 0.004 (log‐rank test). Cohorts possessing either pretherapeutic or post‐therapeutic EpCAM−/Vim− null CTECs have a better response to treatment, displaying a prolonged mPFS.Click here for additional data file.


Coccidioides spp. spherules and endospores. A suspected diagnosis of spinal coccidioidomycosis was made; fluconazole (10.9 mg/kg PO q12h) treatment was initiated, and decompressive neurosurgery was performed. The granuloma was removed en bloc and histopathology revealed marked, chronic-active, pyogranulomatous myelitis with intralesional Coccidioides spp. spherules with endosporulation. Serum anti-Coccidioides spp. antibody titer results revealed a negative IgM and a positive IgG (1:4). The cat was treated with fluconazole for 445 days and examined at various time points, with the last examination 2 years after initial presentation. The cat returned to full ambulation with only mild functional deficits of the right hind limb. In conclusion, this report documents the diagnosis, treatment, and long-term follow up of a cat with a compressive Coccidioides spp. spinal cord granuloma. This case highlights the importance of including coccidioidomycosis as a differential diagnosis for cats with peracute hindlimb paraplegia that have lived in or traveled to regions where Coccidioides spp. is endemic, and demonstrates the potential for a good long-term outcome with decompressive neurosurgery and antifungal therapy.A 9-year-old, neutered male, domestic shorthair cat from Arizona, was presented for evaluation of a 7-day history of hind limb paraparesis that progressed to paraplegia. There was no history of respiratory abnormalities. Neurologic examination supported localization of a T3-L3 myelopathy. Computed tomography (CT) revealed an expansile widening of the spinal canal dorsal to L4 associated with a strongly contrast-enhancing mass. Moreover, CT series of the thorax revealed a diffuse miliary pulmonary pattern, as well as tracheobronchial, sternal, and cranial mediastinal lymphadenomegaly. Transthoracic lung lobe and sternal lymph node fine needle aspiration revealed pyogranulomatous inflammation with  Coccidioides immitis and C. posadasii, colloquially known as “Valley Fever”, are dimorphic, soil-dwelling fungi that are commonly found in the southwestern United States  that are aerosolized from soil  was presented to the Companion Animal Clinic at Midwestern University College of Veterinary Medicine (MWU-CVM) for evaluation of a 7-day history of progressive hind limb paraparesis. The cat lived inside the owner's residence and did not venture outside. Seven days before presentation to the MWU-CVM, the cat was evaluated by the primary care veterinarian for a 12-h history of right hind limb paresis. Physical examination revealed decreased patellar and withdrawal (flexor) reflexes of the right hind limb. The remainder of the examination was deemed unremarkable. The cat was administered dexamethasone (0.4 mg/kg subcutaneously once) and discharged with prednisolone (1.6 mg/kg PO q24h) for the purpose of empirically treating a presumed spinal cord disorder with suspected underlying spinal cord edema. The primary care veterinarian evaluated the cat 6 days later, at which time, the left hind limb had become moderately paretic and the right was unchanged. A complete blood count (CBC) and serum chemistry revealed no clinically relevant abnormalities.Dirofilaria immitis antigen. Tests for Toxoplasma gondii and Coccidioides spp. IgM and IgG serum antibody titers were submitted . The cat was discharged with prednisolone (unchanged dosage) as the only medication.Physical examination findings upon presentation to the MWU-CVM (day 1), included heart rate of 210 beats/minute, respiratory rate of 30 breaths/minute, and a normal temperature 100.5°F (38.1°C). Cardiothoracic auscultation did not reveal a murmur, arrhythmia, or abnormal lung sounds. There was no history of respiratory abnormalities . Neurologic examination performed by a board-certified veterinary neurologist (JE) found ambulatory hind limb paraparesis and intact withdrawal reflexes. There was severely decreased voluntary ambulation on the right hind limb and moderate decrease in the left hind limb, with severe gluteal and hamstring muscle atrophy. Hind limb spinal reflexes were intact with increased patellar reflexes. The cutaneous trunci reflex was normal, and the cat vocalized on palpation of L1-L4. The remainder of the examination was unremarkable and neurolocalization supported a T3-L3 myelopathy. Primary differential diagnoses considered included spinal neoplasia , intervertebral disc disease, a vascular event, infectious causes , coccidioidomycosis, toxoplasmosis), or another inflammatory disorder. Three-view spinal and thoracic radiographs were procured and showed a diffuse miliary pattern  consisting of numerous small, ill-defined soft tissue nodules (~3 mm) within all lung lobes . There wThe cat was presented again on day 4 for thoracic and spinal computed tomography . On examination, voluntary movement of the hind limbs was now absent. The previous paraparesis had progressed to paraplegia with decreased, but present deep pain sensation to both hind limbs. The diffuse miliary pulmonary pattern was confirmed with the CT scan . There wCoccidioides spp. spherules and endospores (Coccidioides spp. granuloma was made. Magnetic resonance imaging (MRI) of the thoracolumbar spinal cord with emergent surgical decompression at a veterinary referral hospital with MRI capabilities was recommended, but the owner declined and elected to have an exploratory spinal cord decompressive procedure performed at the MWU-CVM. The cat was discharged with the following medications: fluconazole (10.9 mg/kg PO q12h) and prednisolone (unchanged dosage).Cytologic interpretation of samples obtained via fine needle aspiration of the lung and sternal lymph node revealed pyogranulomatous inflammation with dospores . CisternScheduling conflicts resulted in the cat being presented for surgery 3 days later (day 7). Immediate surgical decompression is generally recommended for paraplegic patients or those with acute rapid deterioration of myelopathic signs as delay in surgical intervention could result in a poor prognosis . The neuCoccidioides spp.). Occasionally, the spherules were degenerate with externally released endospores. At the periphery of sections, residual neuroparenchyma contained gemistocytic astrocytes, acute hemorrhage, degeneration, and malacia (Toxoplasma gondii (IgM and IgG negative) and Coccidioides spp.  also returned. The cat was discharged (2 days after surgery) on day 10, at which time, there had been considerable improvement. The cat was mobile using the left hind limb, which demonstrated moderate paresis. The right hind limb had good extensor tone, but still was without voluntary movement. The medications prescribed at discharge included fluconazole (unchanged dosage) and prednisolone .The day after surgery (day 8), the urinary catheter was removed. Minimal voluntary movement was noted in the right hind limb with good voluntary movement in the contralateral limb. Postural reactions were decreased in both hind limbs, but were improved compared to before surgery when they were absent. The following day (day 9), the cat was able to use the left hind limb to stand and ambulate, but there was still minimal movement in the right hind limb though, there was improvement in muscle tone. There was normal tactile placing and proprioception in the left hind limb with absent postural reactions in the right. Histopathology of the excised mass showed that normal spinal cord architecture was obscured by mixed inflammation dominated by epithelioid macrophages, few multinucleated giant cells, neutrophils, and fewer, often perivascular, lymphocytes and plasma cells. Admixed within the inflammatory cells were frequent hyperemic reactive vessels and numerous circular spherules that measured up to 33 μm in diameter with a thin 1 μm refractile capsule that surrounded combinations of central basophilic stippling, vacuolation, and packed with 2–3 μm diameter endospores  and thoracic radiographs were performed on day 35. The Coccidioides spp. IgM remained negative and there was a one-fold reduction in IgG titer (1:2). Thoracic radiographs revealed an improvement to the miliary pulmonary pattern with static sternal lymphadenomegaly   and the CBC, chemistry, and urinalysis were unremarkable. Thoracic radiographs revealed improvement of the miliary pattern with fewer pulmonary nodules and the remaining soft tissue nodules were less distinct. The sternal lymphadenomegaly was still identifiable and similar to the previous study , urinalysis, and thoracic radiographs were performed. The Coccidioides spp. titers were negative and the CBC and urinalysis were unremarkable. There was a mild increase in serum alanine transaminase activity ; all other measured chemistry analytes were unremarkable. Fluconazole was decreased (12 mg/kg PO q24h) and then discontinued approximately 4 months later (day 452). Thoracic radiographs at this time showed resolution of the pulmonary nodules (Coccidioides spp. antibody titers (Protatek Reference Laboratory) were similar . No medications were prescribed at discharge.The cat was presented for an additional recheck evaluation on day 310. The owner reported that since the last examination, the cat ambulated normally, was able to jump to the second tier of the “cat tree”, and run, but occasionally limped on the right hind limb. Neurologic examination revealed continued mild improvement with ambulation to both hind limbs with mild paresis of the right hind limb that occasionally “knuckled” while walking. Postural reactions on the right hind limb were slightly delayed but normal in the remaining three limbs . Serum c nodules . A reche nodules . CoccidiCoccidioides spp infection . On examination, the cat was ambulatory with a moderate shuffling gait associated with the right hind limb, with occasional knuckling. Postural reactions and spinal reflexes to the right hind limb were delayed to absent. Evaluation of the left hind limb was unremarkable (Coccidioides spp. antibody titers (Protatek Reference Laboratory) were repeated and were unchanged from previous results . Given there were no significant clinical changes, other than what might be expected over time, no specific treatment was recommended.The last in-hospital recheck examination took place on day 739 (2 years). The owner reported that although there was some decrease in function of the right hind limb, it did not hinder normal activity. There were no systemic signs of illness that could be associated with markable . The funCoccidioides spp. infections begin with inhalation of aerosolized arthroconidia that grow from mycelia in the soil  . Shubitz et al.  . Coccidiz et al.  revealedz et al. . The difrganisms , 21. CytCoccidioides spp. granulomas and neoplastic masses is important because it reinforces that coccidioidomycosis should remain a differential diagnosis in cats that have lived in or traveled to regions where Coccidioides spp. is endemic.The characteristics of the spinal granuloma on CT were similar to MRI findings in cases of solitary CNS granulomas, as reported in Bentley et al. , where dCoccidioides spp. granulomas in companion animals are limited in the literature (Coccidioides spp. spinal granuloma treated with surgery and fluconazole reported in Foureman et al. (Treatment information for spinal terature . In our terature . Similarn et al.  greatly Coccidioides spp. spinal cord granuloma. This case highlights the importance of including coccidioidomycosis as a differential diagnosis for cats with peracute hindlimb paraplegia that have lived in or traveled to regions where Coccidioides spp. is endemic. This report also serves as an indication that cats with compressive Coccidioides spp. spinal granulomas have potential for a good long-term prognosis with decompressive surgery and fluconazole therapy.In conclusion, this report documents the diagnosis, treatment, and long-term follow up of a cat with a compressive The original contributions presented in the study are included in the article/Ethical review and approval was not required for the animal study because this case study used historical medical records and did not require owner consent.HD writing and editing of the manuscript and review of final submission. JE management of case, collection of data, writing and editing of the manuscript, figure preparation, and review of final submission. JJ, KW, JS, and EH writing and editing of the manuscript, figure preparation, and review of final submission. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Background: The proportion of older adults is increasing worldwide and, with it, the physical inactivity common to this age group. Therefore, the promotion of active aging is a strategic factor in health policies for older people. The aim of this study was to identify the benefits and viability of the strength training program (STRENOLD) in health-related quality of life in adults over 60. Methods: A controlled experimental study was carried out with a sample of 181 people over 60 years old from different European countries belonging to the European project IN COMMON SPORTS. A pair work strength program was administered (STRENOLD) over a period of 24 months, consisting of two single sessions per week. Their health status was evaluated (EQ-5D-5L) before and after the interventions. Results: The adherence rate was over 89% and the tolerability rate over 100% in all participating countries. Significant improvements in the participants’ health were demonstrated in the areas of mobility, usual activities, pain/discomfort, and anxiety/depression. Conclusions: The regular practice of physical exercise, through the partnered STRENOLD strength program, has benefits on mobility, usual activities, pain/discomfort, and anxiety/depression, in short, health benefits for older adults. The proportion of older adults is increasing rapidly worldwide, and the proportion of people over 65 is projected to rise from 8% to 16% by 2050 . All thiPhysical inactivity among older adults increases progressively with age despite the multiple health benefits of regular and systematized physical activity . The proThese low levels of physical activity can compromise functional capacity during the aging process . TherefoBorba et al.  have staFor these reasons, the objective of the present study was to identify which benefits can be brought to the health-related quality of life in European adults over 60 years old by undertaking the strength training program (STRENOLD).A sample of 181 participants (79.4% women) with an average age of 69.24(6.94) was recruited for the European Erasmus+ “IN COMMON SPORTS” project, in which four European countries participated: Bulgaria, Hungary, Italy, and Spain. This project promoted physical activity in 60+ year old residents of Europe, in line with the ethical principles of the Declaration of Helsinki. The following data were gathered to evaluate feasibility: recruitment rate (number of participants recruited vs. number of participants who met the inclusion criteria); participation rate ; adherence (rate of patients with 80% participation or higher); dropout (number of participants who could not complete the program); safety and tolerability .The 5-level EQ-5D version (EQ-5D-5L) was introduced by the EuroQol Group in 2009. It is a simple, generic, and standardized instrument for assessing the state of patients’ health. . The EQ-The STRENOLD Program is a training program designed specifically for older people. The main objective is strength training combined indirectly with other conditional abilities . What is relevant about this program is that it is carried out in pairs, using very little equipment  and maximizing the use of the bodyweight of the individual and partner. It is also important to highlight the social element of the pair work.-Warm up (10 min), comprised of: cardio exercises (6 min) and joint mobility (4 min).-Main part (45 min), comprised of: coordination exercises and aerobic games (10 min), strength exercises based on bodyweight and resistance (in pairs—15 min), balance (5 min), and Pilates/yoga (15 min).-Cool down (5 min), comprised of: stretching (3 min) and relaxation (2 min) exercises.The STRENOLD Program is divided into three parts: The STRENOLD program was carried out over 24 months with two 60-min sessions per week by graduates in physical activity and sports sciences specialized in training older people. It is worth noting that, prior to the intervention, a meeting of specialists was held to agree on the program and the types of exercises to be implemented, as well as the different adaptations to be used according to the physical condition and functionality of the participants. The training in pairs was carried out after having considered their bodyweight and the functional level of each component.The representatives of the European project “IN COMMON SPORTS” contacted the participants in each member country to make a first assessment (baseline) of the individuals in March 2018. Within a week, all subjects were summoned simultaneously in all the participating countries . The assessments were made in sports centers  where the subjects carried out their training. After the first evaluation, the STRENOLD program was carried out over a period of 24 months with two 60-min sessions per week. At the end of the STRENOLD program, a final assessment was carried out simultaneously in all the participating countries, and the data were dumped into the common SPSS statistical system, where the different statistical analyses were performed. In the statistical analysis, the sample was divided into groups comprising the four countries.p > 0.05). The statistical analyses were carried out using the statistical package IBM-SPSS v25 for Mac. The significance was considered as p < 0.05.A descriptive analysis was carried out through measurements of central tendency (mean and standard deviation) and percentages. The analysis was carried out globally, and also stratified by country, with the aim of defining the basic characteristics of the sample . We compared the basic characteristics of the sample by country to identify significant differences between countries using the one-way analysis of variance (quantitative variable) and chi-square test . To analyze the feasibility of the STRENOLD program, the following parameters were calculated using percentages: recruitment rate, participation rate, adherence, dropout, safety, and tolerability. The state of the sample’s health was described by means of percentages and an average of the index and the visual analogue scale (VAS). In order to understand the effect of the STRENOLD program on the sample, an analysis was carried out using the paired Student’s t-test (quantitative variable) or chi-square test . This analysis was carried out globally and stratified by country across the sample. Effect size (ES) was calculated for each comparison using Cohen’s d to evaluate the size of mean differences. The Bonferroni correction for multiple comparisons was applied. The normality of the variables within the study was verified using the Kolmogorov–Smirnov test (p = 0.01). The participation rate was 90.07%, showing an adherence of 91.14%. The drop-out rate was 3.09%, with Bulgaria (6.33%) and Italy (4.16%) showing the highest rates in this field. The safety and tolerability rate was 100% since no adverse effects deriving from the program were reported. These data are shown in A total of 181 people over 60 years old from different European countries were selected to participate in the training sessions of the European IN COMMON SPORTS project. There was an overall recruitment rate of 64.25%, with Hungary having the highest rate at 94.44% (Hungary and Bulgaria recorded the largest effect sizes (ES = 0.39 and ES = 0.29) in this parameter, and these countries were also the only ones to show significant improvements in pain/discomfort (ES = 0.18 and ES = 0.39). In the mobility parameter, Hungary, Italy, and Spain showed significant improvements with Spain recording the largest effect size here (ES = 0.29). These three countries also demonstrated significant improvements for usual activities with similar effect sizes (ES = 0.17–0.19). In the self-care parameter, only Spain showed significant improvements. Thus, significant improvements were observed in four parameters for Spain and Hungary, in three for Italy, and in two for Bulgaria.In terms of the index score, only Hungary and Spain showed significant improvements, while in the EQ-VAS score Hungary, Spain, and Bulgaria reflected significant improvements.The population of older people is growing significantly and, with it, the number of associated health problems. Age-related illnesses are of great concern because they result in a marked deterioration in people’s quality of life. Therefore, the practice of physical activity becomes an ally of older adults, as a significant relationship between the amount of physical activity practiced and the functional level and health of the elderly population has been demonstrated .Strength training has a positive influence on the quality of life and functionality of the elderly and is the most beneficial form of physical exercise . In factThe results of this study show that the same strength training program applied in four countries  generated different results across the parameters of health status (EQ-5D-5L). Hungary and Bulgaria presented significant improvements in the pain/discomfort parameter (ES = 0.39 and ES = 0.18), while Spain presented significant improvements in self-care (ES = 0.15). Three countries  showed significant improvements in the usual activities and mobility parameters. These results reflect that, despite the application of the same strength program, there are differences depending on the place of residence. This may be due to the fact that quality of life is multifactorial and, therefore, affected by physical exercise and lifestyle, social limitations , physical limitations (comorbidities), accessibility issues , and motivations and beliefs .It is noteworthy that all the countries shared a significant improvement in the anxiety/depression parameter, and thus, it is evident that the practice of physical activity by older adults improves states of anxiety or depression . These sWhen considering the significant improvements in the parameters of mobility and usual activities, the economic, health, and educational factors—and the reasons why they carry out physical activity—should be taken into account. The countries of Spain, Italy, and Hungary have an annual gross domestic product (GDP) and a per capita GDP higher than Bulgaria, a factor which affects education, health, and ultimately the welfare of their citizens .In keeping with the findings of the study by Lubs et al. , it coulAdditionally, the reasons why this age group engages in physical activity may also be an important factor in these significant improvements . It has been shown that the most common reasons to do sport or physical activity are to improve health (54%) and to improve physical fitness (47%) .This suggests that the STRENOLD program is very important for this population group because of the multiple benefits associated with doing it.However, at present, there is an increase in physical inactivity among European older adults . This caThe results suggest that engaging in regular physical activity using the STRENOLD strength program  can, albeit in a small way, bring about health benefits inasmuch as the participants reported improved feelings of well-being and emotional state. The results suggest that older adults show improvements in mobility, in self-care, and in the execution of daily activities and report decreasing levels of pain/discomfort and anxiety/depression. However, these benefits may be conditioned by external factors because of the variability of results in the countries where the program was implemented.


Alkenylbenzenes are naturally occurring secondary plant metabolites, primarily present in different herbs and spices, such as basil or fennel seeds. Thus, alkenylbenzenes, such as safrole, methyleugenol, and estragole, can be found in different foods, whenever these herbs and spices (or extracts thereof) are used for food production. In particular, essential oils or other food products derived from the aforementioned herbs and spices, such as basil-containing pesto or plant food supplements, are often characterized by a high content of alkenylbenzenes. While safrole or methyleugenol are known to be genotoxic and carcinogenic, the toxicological relevance of other alkenylbenzenes  regarding human health remains widely unclear. In this review, we will briefly summarize and discuss the current knowledge and the uncertainties impeding a conclusive evaluation of adverse effects to human health possibly resulting from consumption of foods containing alkenylbenzenes, especially focusing on the genotoxic compounds, safrole, methyleugenol, and estragole. Alkenylbenzenes primarily occur as secondary plant metabolites in various herbs and spices  but are also present—albeit at lower levels—in agricultural crops, e.g., in tomatoes and apples ,2. AlkenThe toxicity of alkenylbenzenes—especially their genotoxic and carcinogenic potential—is a controversially debated issue. Results of various toxicological studies demonstrated that single alkenylbenzenes, such as safrole, methyleugenol, and estragole, cause—amongst other things—genotoxic and carcinogenic effects in animal studies ,7,8. HowBeside the toxicity of single compounds, it has to be kept in mind that different foods may contain more than one alkenylbenzene, such as basil, which contains methyleugenol, estragole, and other compounds . This isOccurrence data of alkenylbenzenes in different foods are necessary to assess human exposure. However, there are significant variations in occurrence levels, depending on, e.g., the analyzed samples , methodology , etc. Due to these differences, it is often not possible to assess occurrence data of different origins in a comparative manner, which complicates conducting a reliable exposure assessment.trans-anethole  and some strengths as well as weaknesses of current experimental and analytical strategies regarding the risks possibly resulting from exposure to alkenylbenzenes in general.Consumers need to know which of their food consumption habits might result in high intake levels of genotoxic alkenylbenzenes in order to become able to draw informed decisions, whether to choose or not to choose a certain alkenylbenzene-containing aromatized or natural food.In the following part, some examples are described to shed more light on the complexity of alkenylbenzene composition in different herbs and spices.Foeniculum vulgare Mill., Umbelliferae or Apiaceae) is cultivated in many countries all over the world. Essential oils can be obtained by steam distillation of the dried ripe fruits or other parts of the plant such as leaves, stems, or roots, as described by Trenkle -L-cysteine—the mercapturic acid formed from 1′-sulfoxy estragole—has been detected in the urine of human volunteers after drinking fennel tea containing approximately 2 mg estragole -L-cysteine—the mercapturic acid formed from 1′-sulfoxy estragole—in the urine of human volunteers after drinking fennel tea -L-cysteine was described as the main MA, which is formed from estragole and its structural congener trans-anethole [n = 12) resulted in the excretion of N-acetyl-S-[3′-(4-methoxyphenyl)allyl]-L-cysteine in the urine samples within 24 h. The interindividual variation of total N-acetyl-S-[3′-(4-methoxyphenyl)allyl]-L-cysteine excreted (93–1076 ng) reflected the complexity of estragole/trans-anethole metabolism involving different enzyme families, i.e., CYP, SULT and alcohol dehydrogenases (ADH). It hinders an accurate estimation of the external exposure for individuals from the N-acetyl-S-[3′-(4-methoxyphenyl)allyl]-L-cysteine determined in 24 h urine samples. In addition, the biomarker is not specific; one cannot distinguish between N-acetyl-S-[3′-(4-methoxyphenyl)allyl]-L-cysteine formed from estragole or trans-anethole. These considerations also hold true for another biomarker of exposure to these compounds, the hemoglobin adduct N-(isoestragole-3-yl)-valine  -L-cysteine (24 h urine) and the adduct IES-Val (hemoglobin) can only be used as biomarkers for the internal exposure to the ultimate carcinogen 1′-sulfoxyestragole or to 3′-sulfoxyisoestragole, the reactive sulfate ester metabolites of estragole and trans-anethole, respectively -L-cysteine—the mercapturic acid formed from 1′-sulfoxy estragole—was also found in the urine of human volunteers after drinking fennel tea [Results of rodent studies regarding safrole showed that genotoxic effects were mediated via its active metabolites, such as the proximate carcinogen 1′-hydroxysafrole or, rather, 1′-sulfoxysafrole ,98,102. nnel tea , and DNAnnel tea ,85. ThisDifferences between rodent and human metabolism influencing potential genotoxic effects of alkenylbenzenes were also indicated by others. Sulfoconjugation-mediating SULTs are known to play an important role in the generation of ultimate carcinogenic metabolites of different compounds, including alkenylbenzenes  or heat-induced food contaminants, such as furfuryl alcohol ,89,188. Therefore, it has not yet been fully clarified to what extent carcinogenicity data from animal studies regarding alkenylbenzenes is transferable to humans.In this review, we summarized several aspects regarding the occurrence, toxicokinetics, and toxicity of alkenylbenzenes.The currently available information summarized in this article clearly show that a number of different alkenylbenzenes, such as safrole, methyleugenol, and estragole, have genotoxic and carcinogenic properties. Although the toxicological relevance for these well-investigated derivatives is still under discussion when these substances are taken up in low amounts via herbs and spices, it seems very clear, from a toxicological point of view, that high intake levels—as may result from specific plant food supplements, for example—should be avoided.valid occurrence data reflecting the occurrence of all toxicologically relevant alkenylbenzenes in different food productscomprehensive consumption data for such alkenylbenzene-containing products, which should be collected via appropriate consumption surveysdetermination of toxicological properties of yet insufficiently investigated derivatives, such as elemicin and apiol, via adequate studies designed according to international guidelines and taking into account the alkenylbenzene-specific bioactivation However, there are still several uncertainties impeding a reliable evaluation of the health risks that may result from the intake of different alkenylbenzenes via food. These uncertainties are based on the following data gaps, which need to be closed by appropriate research:The aforementioned uncertainties and associated discussions underline that it is currently not possible to perform a conclusive evaluation of possible adverse effects to human health related to the consumption of alkenylbenzene-containing foods.


In recognition of the need to better prepare doctoral candidates with teaching and learning competencies, we devised an innovative internship program in the form of a structured apprenticeship and trialed it in public health higher education. The paid internship was comprised of: (i) Mentoring from an experienced educator, (ii) Structured program of education in pedagogy and curriculum design, and (iii) Opportunities for applied experience. Eleven interns completed the apprenticeship in its first 2 years. The mixed method evaluation assessed the impact of the internship on knowledge, skills, and confidence of interns throughout the internship, and included a cost-consequence analysis. Data collection included surveys and face-to-face interviews with interns and mentors. Changes in intern knowledge and skills were analyzed by intern self-ratings pre- and post-internship on 11 performance descriptors. All interns indicated improvement in at least one area of teaching. Interviews indicated general satisfaction, however raised incompatibilities between the unstructured nature of mentoring and intern expectations and preferences. The economic analysis calculated a cost-offset associated with intern-delivered teaching activities of $58,820 . The total cost of the program was calculated to be $70,561 . This Internship is associated with positive impacts for interns across a range of domains at a net total investment of $11,741. The primary objective of doctoral training programs is the development of researchers; developing the craft of teaching is secondary and something that is hoped will come in due time . While tThere are, however, some initiatives to train doctoral students to teach in academia such as the Preparing Future Faculty initiative (PPF) by the Council of Graduate Schools, USA . Under tThe Educated Citizen and Public Health Initiative, a collaboration of arts and sciences and public health organizations organized in 2006, aims to integrate public health perspectives within a comprehensive liberal education framework and to foster interdisciplinary and inter-professional collaboration  meaning Therefore, the current paper describes the evaluation of a model pedagogy-preparation program in public health education run at the University of Newcastle, NSW, Australia.The aims of this mixed method evaluation were toAssess self-reported changes in intern knowledge, skills and confidence,Calculate the net budget impact of the internship program from an institutional perspective.This evaluation used a mixed method, pre-post assessment of intern and mentor experiences, including a cost consequence analysis .The internship program commenced in 2018, launched as an opportunity for PhD candidates to differentiate themselves and potentially increase their employability. There were over 350 PhD Candidates enrolled in the School of Medicine and Public Health during the timeframe of the internship. Australian PhD programs focus solely on research training and do not include training in teaching and learning. The internship was advertised to all fulltime candidates and recruited via a competitive process.Mentors were selected via a call for expressions of interest and were matched with interns on the basis of teaching content. All mentors were experienced educators active in Scholarship of Teaching and Learning.The interns worked alongside their mentors developing teaching activities and assessment items. They gained real-world experience facilitating tutorials (face to face), facilitating online discussions, and marking assessments, all under the supervision and guidance of mentors. Mentors and interns debriefed following teaching sessions. Where appropriate, interns observed their mentor facilitating a face-to-face tutorial prior to facilitating their own tutorial. Online mentoring and supervision also occurred via the learning management system.There were 11 interns and 9 mentors involved in the program across 2018 and 2019, from which there were seven interns and six mentors who agreed to be involved and provided data for the evaluation. Ethics requirements for the study meant that individuals who declined to take part did not have to provide reasons as to why.The Internship program design was informed by a review of the literature. In the United Kingdom (UK) and North America the “Teaching Assistant” model is common. Across Australasia, the entry level teaching role is referred to as a tutor. Teaching assistants and tutors obtain practical experience rather than a structured program on how to teach. Korpan et al.  developeMentoring from an experienced educator who helped the interns analyze their existing teaching experience, compare this to relevant sample position descriptions, identify gaps, and develop a personalized plan;Facilitating access to structured courses on teaching and learning to address the gaps, for example, a short course on writing learning objectives;Practical experience designing assessments and marking rubrics, delivering lectures and tutorials, facilitating online activities and online discussions, and marking assessments.Based on the Korpan Model, the newly developed internship program was designed as a structured apprenticeship that comprised:Evaluation data were collected via surveys and interviews. Two complementary surveys were developed, one each for interns and mentors, informed by several existing tools , 18, andData for the cost analysis were sourced from the same surveys.The interview guide was framed by Korpan's teaching assistant competencies. The semi-structured interview format explored internship elements such as intern knowledge, skills and confidence, importance of different internship program components, benefits and challenges, expectations, personal goals and relationships with others. Some interview items were targeted toward mentors, and others toward the interns .Ethical approval was granted from the University of Newcastle Human Research Ethics Committee at the start of the study before participant recruitment. All participants (interns and mentors) provided written informed consent and where relevant, consented explicitly to the interviews being audio recorded. The invitation to participate in the evaluation was carried out by administrative staff independent to the academic staff involved in the internship program. Completed surveys were returned to administrative staff to anonymise. Participants consenting to interviews, responded directly to author MT who was invited to collect and analyse qualitative data, and who had not been involved in the internship program development or recruitment.Due to the small number of participants the analysis of survey data was limited to comparing pre- and post-internship rating scores for individual interns. The survey data were also used for the identification and measurement of resource use to inform the economic analysis. Costs of the program were identified as the cost of the scholarship top up payment  and the cost of participants' time to engage with the program. Participants' time was identified and measured via the survey data. This labor time was valued using the human capital approach .The economic benefit of the program was identified as mentors' time freed up as a result of the interns' teaching. The benefits of the program were measured as valuation of the teaching time undertaken by interns, using the opportunity cost method . This beA “coding reliability thematic analysis” was applied to the interview data. The primary steps in this approach were: data collected and coded qualitatively, then verified by members of the research team and illustrated by participant's statements , 22.Intern surveys reported here are based on responses from seven of the eleven interns (63% participation rate). We compared pre- and post-internship ratings for the five interns who provided sufficient data. Each responding intern rated themselves higher on at least two performance descriptors post-internship. Some interns assessed themselves as having an improvement of 1-point on more than half of the applicable descriptors  whereas other interns felt they had a more marked improvement of 2-points in fewer areas (2019/2 in two descriptors related to assessment and 2019/3 in two descriptors related to teaching identity/philosophy). This may reflect differences in tasks chosen by interns or assigned by mentors that either addressed a broad range of skills or a narrower focus on particular teaching activities. A full summary of pre- and post-change is in Only one intern gave both pre- and post- ratings for preparing and executing a rubric, indicating that many PhD students may not have experienced this skill. There were also fewer ratings for classroom-related descriptors  possibly due to fewer opportunities for face-to-face teaching . Almost none of the pre-internship descriptors had ratings that were <3, possibly indicating reluctance to use the lower end of the scale, and many were rated at the maximum 5. There were no negative pre-post changes.Four interns and three mentors were available and consented for interview, from the 2018 cohort (when interviews were conducted). Interviews with interns and mentors were guided by questions from didn't have any” specific goals in mind as to how to improve either when they began the program, nor clear expectations of what to expect overall. One intern said “I didn't really have any expectations to be honest… I went into it thinking whatever I get out of it is fantastic” and others “thought it would be teaching.” Interns who had previously gained teaching experience outside of Australia were hopeful for “a new way of teaching… different culture, different way of doing things” to broaden their repertoire as educators. There were improvements to confidence: interns who provided lectures were able to showcase their own expertise, and then spoke of receiving praise from their mentor, such as: “I have changed a lot over this year… I have learned a lot… I feel confident I can be a very good teacher.”The competency elements most discussed were “abilities” and “skills”, though interns admitted they “not to overwhelm” the interns, while providing a range of key teaching experiences. One mentor focused on involving an intern in “development of course content… and face-to-face teaching” also saying they urged their interns toward “recording audio feedback” for students even though it meant some discomfort “…but we do it anyway, because it's such a time saver and we want (ed) them to have experience in all the key areas of a course.” One early career mentor described gaining confidence from the internship themself, having never “had a person who I was giving tasks to, responsible for deciding what they do… checking in to make sure it was done….” As a result of their first mentoring experience, the same educator said they “might change the way we mentor… just modifying it a little bit to suit each individual intern experience… what they need to grow.”At the start of the program mentors were required to meet their intern and discuss the course(s) with which they would be assisting. Mentors tried “flexible, goal-oriented and open to constructive criticism and open to learning opportunities, to learning suggestions.” They expected their mentor to be someone who “understands the intern's needs… paying attention and being responsive.” Styles differed between mentors, as did their contact with interns, with one intern feeling that the early phases of their experience required more structure. A reflective journal had been suggested as “some sort of template to jot down things” as a record of teaching tasks, and also as “some measure of pre-and post” for the intern.For a successful internship experience, interns said they should be “…managing the time allocated to teaching, if some periods were busier than others,” and in general finding that teaching could be “a lot more time intensive than [they] had anticipated.” Providing a balanced teaching experience was essential, to avoid “marking exhaustion.” One intern confided that “you just wish the marking was over…that's just part and parcel of teaching.” Certification courses were also offered as part of the internship, which upskilled interns in key teaching proficiencies such as providing constructive feedback. They proved both popular and challenging, with one intern saying, “I had to rewire everything I had to learn, for teaching… so many things have changed over the years and learning those new things took a bit of time.”Some of the more challenging aspects of the program, from the intern's perspectives included “help develop that pipeline” in terms of job-ready PhD candidates who have experience and confidence in teaching and learning. Interns suggested a more structured “feedback procedure” including more regular meetings with mentors. But also commented that “every PhD student should have the opportunity to teach somehow” and “everyone would benefit from being an intern.”Going forward there was support for the internship program. The mentors saw value in the program to “Sensitivity analysis, conducted by varying the wage rate used to value the opportunity cost of the teaching time performed by interns, generated a range in the net cost of the program between $2,574 and $25,348 AUD (2019). Additional economic consequences arising from the program in the short term (and unable to be quantified) included the capacity building of both interns and mentors. Medium term consequences included expected earnings uplift for participating interns.This study is a first of its kind that analyzed multiple components of a teaching and learning internship program, including its cost-impact. Previous studies assessed the impact of an internship program for PhD students only from a career decision marking point of view  or in teWhile generally very positive, qualitative data pointed toward the importance of communication between interns and mentors. A more structured approach to feedback was a common discussion thread by interns, perhaps reflective of the intern's feelings of vulnerability given their new teaching and learning role . Each inBeing accepted into the internship program held status, with interns proudly including this on their education and academic webpages. The competitive application process for the internship resulted in high achievers being selected—this left them looking for reassurance and a system of measurement for their performance, whereas the Internship program took a less structured, personalized approach to interactions and “assessment.” The selection of academically successful interns may have also led to them being reluctant to rate themselves in the lower half of the rating scale, limiting the capacity to measure improvement.In addition to seeking more concrete expectations on which to assess their achievements throughout the internship, the interns also sought more opportunities to interact with each other. Increasing opportunities for formal and informal interactions and encouraging the development of a community of practice will be prioritized moving forward. We have also sought to align the internship activities more formally with Fellowship requirements with the Higher Education Academy.Inclusion of the cost impact analysis provides reassurance to higher education managers that internship programs such as this can be justified from a managerial perspective.There were limitations to this study. This evaluation was limited to responses from a small number of interns at one school within one academic institution. At this stage we have not conducted any follow-up assessment to evaluate if the interns maintained or improved the knowledge and skills acquired during the program and if they had the opportunity to apply these skills/knowledges in another setting. There is scope to assess employer views on whether participation in the internship increases perceived or actual employability. Our eligibility criteria required the interns to be enrolled as full time PhD candidates who were receiving a scholarship stipend for their living experience, this may have excluded some students from applying and limits generalisability.A strength of this internship program design is the evidence informed selection of components and the ability for components to be personally tailored to each interns needs. A strength of the mixed methods evaluation was the inclusion of structured reflection from interns, that strengthened our ability to assess the extent of improvement in any personal or professional skills after participating in the internship program.As only few such programs exist in Australia, this program and the results support the positive benefits for students on both a personal and professional level and at the institutional level and contribute to the body of research on graduate student teaching and learning internship programs in Australian higher education sector by identifying and emphasizing the value of a systematic internship component during their academic career.Our internship program was a three-way partnership between the interns, the educators and the School of Medicine and Public Health (University of Newcastle) designed to provide an experiential learning opportunity. While these internships provided PhD students with practical and hands-on teaching and learning experiences, introducing them to teaching collaborations within our school, to work with and strengthen ties with experienced educators, continued assessment of this program is imperative. Therefore, there is a need to develop and implement systematic ways to assess the viability, uptake and effectiveness of these initiatives, especially taking into account the valuable insight provided by the students' perceptions of the internship and the role that both the educator and the school played in facilitating their learning. Secondly, the results indicate that there is a need for increased structure. Therefore, our future project and research will focus more on creating effective, structured and engaged professional and graduate school programs and initiatives with a focus on longer term evaluation.In conclusion, the teaching and learning internship was associated with positive impacts for interns across a range of domains. Other schools of public health should consider offering a similar program to graduate students.meredith.tavener@newcastle.edu.au.The datasets presented in this article are not readily available because specific informed consent, and ethics committee approval, would need to be sought and confirmed. Requests to access the datasets should be directed to Meredith Tavener, The studies involving human participants were reviewed and approved by Human Research Ethics Committee, University of Newcastle. The participants provided their written informed consent to participate in this study.EJ and TM led the conception and design of the study. EJ oversaw the manuscript completion. MT was responsible for the qualitative data collection and analysis and interpretation. PR and SD carried out the economic analysis. NW was responsible for checking and analyzing the survey data. TB provided literature review input and wrote sections of the discussion. All authors contributed to the article and approved the submitted version.The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: The internship program was supported by the School of Medicine and Public Health, University of Newcastle, NSW, Australia. The evaluation activities were funded by a grant from the Faculty of Health and Medicine Research Committee.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Secondary schools are increasingly supporting adolescents’ mental health and well-being, yet many teachers report that they lack the skills and confidence to do so. Building Educators’ skills in Adolescent Mental Health (BEAM) is a web-based training program developed to improve secondary school teachers’ knowledge and confidence in caring for students’ mental health.This pilot study examined the preliminary effectiveness and acceptability of the BEAM program for improving mental health knowledge, attitudes, confidence, helping behaviors, and psychological distress among secondary school teachers.A single-arm pilot trial was conducted from July to December 2019 among secondary school teachers located in New South Wales, Australia, who were employed in leadership positions responsible for managing student well-being . Participants had access to the BEAM program for 6 weeks. Self-report surveys, delivered at baseline, postintervention (6-weeks post baseline) and 3-month follow-up (19 weeks post baseline) were used to measure changes in training outcomes. Acceptability was assessed by program use, barriers, satisfaction, and participants’ perceptions of program effectiveness.A total of 70 secondary school teachers took part . Significant improvements in confidence were reported at postintervention and 3-month follow-up. Significant improvements in helping behaviors were reported at 3-month follow-up only. There was also a significant reduction in psychological distress at postintervention. Participants agreed that the program content was easy to understand and relevant, but program completion was challenged by lack of time, competing priorities, and forgetfulness.Findings indicated that a web-based training program may be beneficial for improving secondary school teachers’ abilities to care for students’ mental health; however, program modifications are required to increase training completions.Australian New Zealand Clinical Trials Registry (ANZCTR) ACTRN12619000821190, Universal Trial Number U1111-1232-7680; https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=377529 Adolescence is a key period for the development of mental illness, with over half of all mental health problems first emerging before the age of 18 years ,3. PopulThree recent systematic reviews on youth mental health training for secondary school teachers have indicated a lack of evidence-based programs -20. AndeThe Black Dog Institute has developed a web-based training program that aims to improve secondary school teachers’ knowledge, confidence, and skills in recognizing and supporting students with mental health problems. Delivered over 6 weeks, the Building Educators’ skills in Adolescent Mental Health (BEAM) program combines self-directed content with in-person peer coaching activities and printable resources. The program consists of 5 topics that include educational information, quizzes, blog-style story-sharing, and case studies . Each toBEAM was developed in partnership with a lived experience advisory group of 12 teachers ,26. HeadThis study examined the preliminary effectiveness and acceptability of the BEAM program for improving mental health knowledge, attitudes, confidence, and helping behaviors among secondary school teachers in NSW, Australia. It was hypothesized that participating teachers would report improvements in knowledge, attitudes, confidence, and helping behaviors at postintervention. This study also examined teachers’ psychological distress to determine whether the training program was associated with positive improvements in the mental health of teachers themselves. Program acceptability was examined by measures of program use, barriers to use, and training satisfaction. Our results provide important information about the feasibility of using a training platform that combines web-based and offline learning to address shortages in teachers’ professional development in student mental health.An uncontrolled, single-arm pilot study was conducted with outcome measures assessed at baseline, postintervention (6 weeks after baseline), and 3-month follow-up (19 weeks after baseline). The study was approved by the University of New South Wales (UNSW) Human Research Ethics Committee (HC190047), the NSW State Education Research Advisory Process (SERAP 2019048), and the Sydney, Parramatta, and Maitland-Newcastle Catholic Dioceses. This trial was registered with the Australian New Zealand Clinical Trials Registry (ACTRN12619000821190) and assigned a Universal Trial Number (U1111-1232-7680).The current study targeted secondary school teachers from NSW, who were employed in a teaching role with increased responsibility for the mental health of students aged between 12 and 18 years . These teachers were identified as requiring significant skills in student mental health as they acted as informal case managers for students by offering support when needed and liaising with others about school performance. Secondary school teachers were eligible to participate in the current study if they were (1) employed full-time for the duration of the study in a teaching role that had responsibility for student well-being, (2) able to obtain support and signed consent from their school principal, and (3) able to participate alongside at least 1 other equivalent colleague from their school to enable completion of the peer coaching.Recruitment was undertaken between June 17 and July 19, 2019, in NSW. The study was advertised in 9 School-Link newsletters . The study was also advertised on the Black Dog Institute’s website and Facebook page. Teachers were directed to the study website where they completed the eligibility screening and registered with their name, school, and email address, and downloaded the participant information statement and consent forms (PISCFs). Eligible teachers were advised to consult with their school principal and colleagues and email the completed PISCFs to the research team.Once eligibility was confirmed and consent forms were returned, participants were invited to complete the baseline survey. Participants who did not attempt or complete the baseline survey within 7 days were withdrawn from the study. Upon completion of baseline, participants were given access to the BEAM program for 6 weeks. The BEAM program was accessible on internet-enabled computers only. Modules were completed sequentially, and all activities were mandatory. The program provided automatic feedback on incorrect responses and this content was not reviewed by the researchers. Responses to the peer coaching activities were submitted to the research team through the program and researchers provided feedback via email within 3 days of submission. Participants were sent 4 reminders to use the program. At 6 weeks post baseline, participants were invited to complete the postintervention survey. At 3-month follow up , participants were invited to complete the follow-up survey, and those who completed it were reimbursed with an Aus $50 (US $36.26) e-gift voucher, irrespective of whether they completed the BEAM program.At baseline, participants reported their gender and age, teaching experience, duration in current role, and employment at current school . Participants also reported the location of their current school , the school funder (government or nongovernment), and gender type . Participants reported their current level of training in student mental health . Participants were asked to rate the importance of receiving mental health training and their confidence in web-based programs for satisfying their training needs. Items were answered using a 5-point Likert scale ranging from not at all (1) to extremely (5).A 6-item adapted subscale of the Mental Health Knowledge Schedule (MAKS)  scale waA modified version of the Depression Stigma Scale–Personal Stigma subscale  was usedAn adapted version of the Confidence in Helping subscale  was usedA 14-item adapted version of the Help Provided to Students questionnaire  was usedThe self-report 5-item Distress Questionnaire-5 (DQ5)  was usedAt postintervention, participants were asked to rate (using 4 separate items) the extent to which they believed the BEAM program increased their confidence, skills, and approach to students’ mental health needs, and whether the program met their training needs. Items were answered using a 5-point Likert scale ranging from not at all (1) to entirely (5).Program use was measured by the number of topics completed. As topics were sequential, participants could not progress through the program without completing the peer coaching. Therefore, topics were deemed completed when a participant submitted the corresponding peer coaching activity. Barriers to program use were assessed at postintervention. Participants were asked to report whether they had experienced any items from a list of 11 barriers to program use .Acceptability was measured at postintervention using a 13-item questionnaire that was adapted for the BEAM program ,36. PartP<.05 (2-tailed). We conducted t tests to compare mean differences in acceptability, satisfaction, and perceived effectiveness scores between program completers  and noncompleters.Program and study data were collected and stored securely using the Black Dog Institute Research Engine hosted on the University of New South Wales servers in Australia. Data was then downloaded into Microsoft Excel and exported into SPSS  for analysis. The final sample for analysis consisted of the participants who completed the baseline assessment. At postintervention and 3-month follow-up, participants’ responses were included in the analyses regardless of whether they completed all questionnaires. Basic descriptive statistics, including means and standard deviations were calculated and reported for participant and school characteristics, and acceptability items. To estimate the pre- and postintervention effects of the program on participants’ mental health knowledge, confidence, attitudes, helping behaviors, and psychological distress, mixed models repeated measures analysis of variance were conducted, with time as a main effect. Statistical significance was set at A total of 134 teachers expressed interest in the study. Of these, 81 (60.4%) teachers from 28 schools obtained consent from their principal and were invited to complete the baseline survey . Table 1At baseline, 40% (28/70) of the sample reported having had “nil” or “limited” prior training in student mental health and 60% (42/70) reported having had “moderate” or “extensive” prior training. All participants believed that mental health training was “moderately”  or “extremely”  important. The majority  felt “moderately” or “extremely confident” that a web-based program could meet their mental health training needs: 19% (13/70) were “neutral” and 9% (6/70) were “somewhat confident.”d=0.60) and 3-month follow-up (Cohen d=0.53). There was also a significant increase in the frequency of helping behaviors at 3-month follow-up and a significant reduction in participants’ psychological distress at postintervention (Cohen d=0.33). There were no significant changes in mental health knowledge or attitudes at postintervention or 3-month follow-up  and 3. SWhen asked about their perceptions of training effectiveness at postintervention, the mean scores indicated that on average, participants rated the training program as “somewhat”  effective for improving their confidence, skills, and approach to student mental health .A total of 94% (66/70) of the baseline sample accessed the first topic and 50% (35/70) completed it. There was a steady decrease in topic completions over time: 16 (23.9%) participants completed half the program  and 11 (16%) completed the entire program . Lack oft tests indicated no significant differences in the mean scores of acceptability, satisfaction, and perceived effectiveness between participants who completed the entire program (n=9) and those who did not (n=17) (P=.08-.98).Most participants rated the program content as easy to understand, relevant, practical, realistic, and helpful . ParticiThis study examined the preliminary effectiveness and acceptability of a web-based training program for improving mental health knowledge, attitudes, confidence, and helping behaviors among secondary school teachers in NSW. Our findings indicate that the BEAM program may be beneficial for improving teachers’ confidence in, and the frequency of, helping behaviors toward their adolescent students with mental health problems, when measured with self-report scales. The BEAM program did not appear to be associated with improvements in mental health knowledge or attitudes, although this may be owing to the high baseline levels of experience and training within this cohort. The BEAM program may also be associated with a reduction in teachers’ psychological distress, although this would need to be replicated within a randomized controlled trial. Given that past evaluations of teacher training programs for mental health have not observed improvements in helping behavior or teachers’ well-being ,19, thesProgram acceptability was high among the teachers who remained in the study at postintervention. These teachers viewed the training content as relevant, helpful, and engaging but were less favorable toward the peer coaching activities. Only half of all participants continued beyond the first topic, suggesting that the compulsory completion of the peer coaching activity may have halted progress for many participants. While there was no evidence to suggest that program completion was associated with greater acceptability and satisfaction, it is difficult to examine overall levels owing to the high study attrition rate. Future trials may benefit from removing the mandatory completion of the peer coaching activities to encourage more training completions. This would allow participants to complete the program at their own pace and lead to greater exposure to training content. While it is not yet clear which persuasive techniques are associated with better outcomes for web-based programs , engagemA key aim of this project was to create a mental health training program for secondary school teachers, which was highly accessible, easily disseminated, and scalable but also engaging and effective. The use of a web-based platform enabled these goals; however, this study shows that 1 in 4 teachers were not confident that this type of program could meet their training needs. Some teachers may have felt that the skills needed to care for students’ mental health could not be taught (or learnt) via a self-directed web-based program. Given that other web-based workplace mental health training programs have been shown to be effective , some teThe study was impacted by high attrition, for both survey completion and program use, thus limiting firm conclusions about program effectiveness and acceptability. It remains unclear whether participants’ attitudes towards the program or other external factors influenced dropout. A larger sample size, with a control group, would allow for these analyses. Of the 11 participants who completed the program, 9 (82%) also completed the postintervention. This may suggest that improvements in program completions may also improve study retention. Future studies would likely benefit from using incentives for all study assessments, thus increasing the study duration to give participants more time to access and complete the program, and modifying the program in accordance with the user feedback to sustain engagement . The higThis study indicates that the BEAM program may improve teachers’ confidence in caring for students’ mental health and increase the frequency of helping behaviors. The training program may also reduce teachers’ psychological distress. The program content was easy to understand and relevant, although barriers related to time, competing priorities, and forgetfulness inhibited program use. The results have indicated that some program modifications are necessary to increase completion. Taken together, the findings demonstrate the potential of a web-based training program for improving helping behavior and related outcomes among Australian secondary school teachers. A randomized controlled trial of the BEAM program will help determine genuine effects.