LASSO regression yielded a 13-protein model which, when added to a clinical model inclusive of NT-proBNP, improved the AUC from 0.82 to 0.92 (p = 1.5 × 10–4). Five proteins were associated with incident HF hospitalization, four with HFpEF hospitalization and eleven with mortality (p < 0.05). We identified and validated multiple circulating biomarkers associated with HFpEF as well as HF outcomes. These biomarkers added incremental discriminative capabilities beyond clinical factors and NT-proBNP.There is increasing evidence that HFpEF is a heterogeneous clinical entity and distinct molecular pathways may contribute to pathophysiology. Leveraging unbiased proteomics to identify novel biomarkers, this study seeks to understand the underlying molecular mechanisms of HFpEF. The discovery cohort consisted of HFpEF cases and non-HF controls from the CATHGEN study (N = 176); the validation cohort consisted of participants from the TECOS trial of patients with diabetes (N = 109). Proteins associated with HFpEF were included in a LASSO model to create a discriminative multi-protein model and assessed in the validation cohort. Survival models and meta-analysis were used to test the association of proteins with incident clinical outcomes, including HF hospitalization, mortality and HFpEF hospitalization in CATHGEN, TECOS and the Jackson Heart Study. In the derivation set, 190 proteins were associated with HFpEF in univariate analysis, of which 65 remained significant in the multivariate model. Twenty (30.8%) of these proteins validated in TECOS, including LCN2, U-PAR, IL-1ra, KIM1, CSTB and Gal-9 (OR 1.93–2.77,  Patients with HFpEF comprise about 50% of HF patients and HFpEF is responsible for approximately half of HF hospitalizations2. As the global burden of obesity and diabetes increases, the prevalence of HFpEF is expected to continue to rise disproportionately to that of heart failure with reduced ejection fraction (HFrEF)3.Heart failure with preserved ejection fraction (HFpEF) is a clinical syndrome of impaired diastolic function with symptoms of dyspnea, congestion and exercise intolerance with a preserved left ventricular ejection fraction 6. Importantly, the mechanisms underlying the development of diastolic dysfunction and progression of HFpEF also remain poorly understood.HFpEF continues to pose significant diagnostic and therapeutic challenges, in part because of marked heterogeneity in defining the clinical syndrome across studies, but also because non-cardiac causes of exertional dyspnea can be difficult to distinguish from HFpEF in compensated patients without invasive hemodynamic testing. Despite the common symptomatology and risk of adverse clinical outcomes in HFpEF and HFrEF, therapeutic strategies that improve morbidity and mortality in patients with HFrEF have not proven to be effective in HFpEF, suggesting that the molecular mechanisms of the two clinical entities may be distinct7. Previous studies have identified distinct profiles of inflammation and remodeling in HFpEF, but further investigation is needed to better understand the underlying pathophysiological mechanisms10.Only BNP and NT-proBNP are routinely used in clinical practice as HF protein biomarkers for presence and severity of HF, agnostic of HF phenotype, in both the acute and ambulatory settings. Despite the identification of promising candidates, including markers of fibrosis , myocardial injury (cardiac troponins), inflammation (GDF-15) and kidney injury (LCN2) for both prediction of and prognosis in HF, these biomarkers have not yet been translated into clinical practice and have limited support specifically for HFpEFLeveraging a high throughout proteomic profiling platform measuring 459 analytes, we investigated the association of protein biomarkers with HFpEF, validated these findings in an independent patient population and examined their prognostic utility in overall clinical events to identify novel molecular mechanisms that differentiate HFpEF from patients without HF.11. HFpEF cases were defined as having a history of HF, EF ≥ 45% and diastolic dysfunction class ≥ 1 on echocardiogram; controls were those with no reported history of HF, EF ≥ 45% and no diastolic dysfunction13. HF hospitalizations were defined using HF-associated International Classification of Diseases, Ninth Revision (ICD-9) codes for emergency room visits or hospitalizations > 30 days from study enrollment  and established cardiovascular disease to the dipeptidyl peptidase 4 inhibitor sitagliptin or placebo. From a previously defined nested major adverse cardiovascular event (MACE) case–control subset of TECOS placebo-arm participants, HFpEF cases (defined as history of HF at enrollment and echocardiographic assessment available with EF ≥ 55%) and non-HF controls  were identified; Twenty participants with HFrEF (EF < 40%) were also compared to non-HF controls; N = 440 participants had proteomic data available and were analyzed for incident events HF hospitalization and mortality.The validation cohort consisted of participants from the placebo arm of the Trial Evaluating Cardiovascular Outcomes with Sitagliptin (TECOS)15. For the present analyses, patients with prevalent HF were excluded from analyses of time to first HFpEF hospitalization. In JHS, mortality and HF adjudication have been described previously16.The Jackson Heart Study (JHS) is a prospective population-based cohort of Black adults residing in Jackson, MS, designed to investigate cardiovascular disease risk factors. Details of JHS have been published previouslyAll methods were carried out in accordance with relevant guidelines and regulations. All study participants in CATHGEN, TECOS and JHS gave written informed consent for participation in the parent study and for use of their stored biospecimens for future use. The institutional review board at Duke University and Beth Israel Deaconess Medical Center approved the studies. A comparison of the three study cohorts included is shown in Supplemental Table 17. Five Olink panels were used  which measured relative expression of a total of 459 unique proteins. CATHGEN and TECOS samples were run in the Shah Lab at the Duke Molecular Physiology Institute on the Olink 1200 platform. JHS samples were run in the Gerszten Lab at the Beth Israel Deaconess Medical Center on the Olink 1500 platform.Proteomic profiling was conducted in stored frozen plasma using the Olink platform, which combines an immunoassay with an oligonucleotide for greater specificity and multiplexing using the proximity extension assay p < 0.05)18. Significant proteins from the univariate model were then tested in step 2 using a multivariate adjusted model adjusted for age, race, sex, body mass index (BMI), systolic blood pressure (SBP), DM and creatinine . Sensitivity analyses included the addition of history of hyperlipidemia and enrollment serum hemoglobin to multivariate models.The overall analysis plan is shown in Fig. Results from step 2 informed three parallel analyses in step 3: (a) pathway analysis using Gene Set Enrichment Analysis (GSEA); (b) individual protein validation for association with HFpEF in the TECOS cohort; and (c) creation of a sparse protein model for HFpEF discrimination in CATHGEN using an iterative LASSO-based approach, and validation of this LASSO model by logistic regression modeling of individual LASSO model proteins in TECOS . Significant individual proteins were then tested in HFrEF compared to non-HF participants in TECOS to test specificity of protein associations for HFpEF.p < 0.05. To understand effects independent of prevalent HF, models in CATHGEN and TECOS were also tested stratified by prevalent HF. Likelihood ratio tests (LRT) were used to compare model fit of the sparse protein model to a single protein with the strongest association with clinical outcomes. All analyses were performed using R versions 4.0.3 and 4.1.2.Finally, in step 4, we evaluated the addition of the LASSO-derived protein model to clinical covariates from the multivariate model as well as NT-proBNP for prediction of HFpEF and association with time to clinical events in both CATHGEN and TECOS. Incremental improvement in prediction of HFpEF status was assessed using change in the AUC after addition of individual proteins identified in the LASSO model or HFpEF-associated proteins from the validation cohort. In exploratory analyses in CATHGEN, evaluation of the discriminatory performance of the LASSO model in sex-stratified analyses was also performed. Clinical events considered were any HF hospitalization (CATHGEN/TECOS), adjudicated HFpEF hospitalization (JHS), and all-cause mortality (CATHGEN/TECOS/JHS). Models for time to hospitalization were adjusted for the same clinical variables as previous models; Fine-Gray competing risk models were used in CATHGEN and TECOS, with all-cause mortality considered as a competing risk, while Cox proportional hazards models were used in JHS. Cox models were used to test for association with time to all-cause mortality and were adjusted for coronary artery disease, low density lipoprotein, history of hyperlipidemia and smoking history. The proportional hazards assumption was tested for all Cox models. Follow-up time in CATHGEN was truncated at 5 years to match the maximum follow-up time in TECOS. Results from survival models were combined using fixed effects inverse-variance weighted meta-analysis; proteins were considered significant at nominal 20, in the CATHGEN discovery cohort HFpEF cases were older , and had higher BMI , SBP (mean mmHg: 151 ± 30 vs. 138 ± 25), creatinine  and prevalence of DM (34% vs. 13%) compared with non-HF controls. In TECOS, similar patterns were observed  and TECOS (N = 109) study populations are shown in Table 76 and TEp < 0.05, Supplemental Table p < 0.05, Supplemental Table Of 459 unique proteins assayed in CATHGEN, 446 had < 25% of values below level of detection (LOD) and thus were analyzed as continuous variables; 12 had 25–75% of values below LOD and were thus analyzed as binary traits; and one protein had > 75% of values below LOD and was removed from analysis. In univariate models, 190 proteins were associated with HFpEF , U-PAR , IL-1Ra  and KIM1 . Of note, four of these 20 proteins are also included in the LASSO proteomic model: KIM1, Gal-9 , NEMO  and SUMF2 . In TECOS participants with HFrEF, five proteins  were significant .Of the 65 proteins significant in the CATHGEN discovery cohort multivariate analyses, 20 (30.8%) validated for association with HFpEF in TECOS multivariate analyses  AUC: 0.81 (0.067), mean (SD) accuracy: 0.72 (0.068)). This model included metabolic markers , markers of angiogenesis , and markers of inflammation . The AUC in TECOS of the 13-protein model was 0.80 (95% CI 0.70–0.90). This model yielded a leave-one-out cross-validation accuracy of 0.76 in CATHGEN and 0.70 in TECOS for discrimination of HFpEF.p = 1.5 × 10–4) and TECOS   in CATHGEN and 0.94 (p = 4.7 × 10–5) in TECOS. In exploratory sex-stratified analyses, the selected set of proteins performed equally well in males and females in CATHGEN  Fig. a. SimilaThe median follow-up time was 1825 days in CATHGEN and 1054 days in TECOS, with 32 incident HF hospitalizations and 45 deaths, and 23 incident HF hospitalization and 54 deaths, respectively. The baseline characteristics of JHS participants are shown in Supplemental Table p-value of 0.056, indicating a marginally improved fit after including the twelve additional proteins. For HF hospitalization in TECOS, the 13-protein model fit the data better than CLSTN2 (p = 0.046). HFpEF-specific hospitalization could only be evaluated in JHS, where four of the 29 HFpEF-associated proteins were significant . Eleven proteins were significantly associated with all-cause mortality . Mortality outcomes stratified by prevalent HF status are shown in Supplemental Table p < 0.05). A Kaplan–Meier survival curve for KIM1  in CATHGEN is shown in Fig. p = 0.016). For all-cause mortality in TECOS, the 13-protein model did not significantly improve the model fit compared to KIM1 (p = 0.52).Of the 29 HFpEF-associated proteins that were either selected by the LASSO model or validated in the TECOS cohort, five were significant in a meta-analysis of HF hospitalization in CATHGEN and TECOS Fig. : PSGL-1,Using a high-throughput platform for discovery proteomics, we have identified novel biomarkers related to fibrosis, angiogenesis, inflammation, kidney injury and fatty acid metabolism that are independently associated with HFpEF Fig. . Importa23. Other key inflammatory HFpEF-associated proteins were U-PAR, IL-1ra, KIM1 and Gal-9; markers of angiogenesis and remodeling  and metabolism  as predictors of clinical outcomes; and KIM1, a marker of kidney and tubular injury with roles in immune function, was the strongest predictor of all-cause mortality even after adjustment for renal function. These protein biomarkers have potential for guiding more precise diagnostic and prognostic cardiovascular care for patients and highlight potential novel therapeutic targets by identifying relevant pathways dysregulated in human HFpEF.Specifically, we found higher levels of LCN2, a siderophore-associated protein for iron trafficking with roles in inflammation and kidney injury, to have the strongest association with prevalent HFpEF24. Differences in levels of remodeling proteins have recently been suggested to aid along with clinical phenogroups in early HFpEF10. A recent publication from the PROMIS-HFpEF study measured 248 proteins and identified inflammatory proteins to mediate comorbidity burden and cardiac structure and function in patients with HFpEF25. Key overlapping proteins between the PROMISE-HFpEF study and the present work include U-PAR and NEMO. Additionally, the KaRen study found that out of 98 biomarkers tested, inflammatory biomarkers were strongly associated with HFpEF severity and outcomes26. Here we extend the support of heightened inflammation as a pathophysiologic mechanism in HFpEF via the identification of inflammatory protein biomarkers, such as U-PAR and Gal-9. There is strong pathophysiologic overlap and feedback mechanisms linking inflammation to extracellular matrix remodeling and cardiac fibrosis. For example, in the TNFα pathways, NEMO was associated with HFpEF and was included in the sparse protein model, which may reflect not only pro-inflammatory signaling, but also myocardial remodeling28. Though the inflammatory biomarkers ST2 and Gal-3 have shown associations with HF including with pro-inflammatory comorbidities and HFpEF severity and outcomes29, these biomarkers were not associated with HFpEF in our study. Notably, we find distinct protein associations in HFrEF samples in the present analyses with U-PAR as the only protein with shared significance across HFpEF and HFrEF analyses.The present data highlight the importance of biomarkers of fibrosis, angiogenesis and extracellular matrix remodeling in HFpEF. The mechanisms by which comorbidities may drive cardiac remodeling through endothelial dysfunction and inflammation have been proposed previously23. LCN2 is also important for iron trafficking, which has implications for innate immune processes. Growing evidence of the value of iron repletion in HF patients may further support its importance in HF-relevant pathophysiologic processes31. KIM1, a transmembrane glycoprotein and marker of renal tubular damage that also plays a role in innate immunity, has previously been associated with adverse HF outcomes. The upregulation of both LCN2 and KIM1 in our study adds to the growing body of evidence around the importance of inflammation, comorbid chronic kidney disease and impaired renal sodium handling in HFpEF pathophysiology32. Gal-9, an inflammatory protein produced by the extracellular matrix that signals through regulatory T cells, is elevated in DM and chronic kidney disease33. Recent studies identified Gal-9 to be associated with incident HF hospitalizations35. The present findings highlight the complex interplay of multiple comorbidities and bring focus to Gal-9 as a shared biomarker reflecting inflammatory and remodeling mechanisms across disease states that may be key to HFpEF pathophysiology.Several specific remodeling and inflammatory proteins from our study warrant further investigation including LCN2, a neutrophil inflammatory lipoprotein and marker of tubular injury which is associated with cardiac hypertrophy and diastolic dysfunction, and predicts cardiovascular events in patients with chronic kidney disease36. Through its role as a cadherin and adhesion molecule, CLSTN2 may contribute to extracellular matrix remodeling following pro-angiogenic stimuli and therefore warrants further evaluation in preclinical and clinical HFpEF research37. VEGFD, a secreted regulator of angiogenesis, was associated with HFpEF and incident HF in the present analyses. VEGFD stimulates cardiac fibrosis and has a growing body of support in HF and pulmonary hypertension38. Incorporation of extracellular matrix degradation pathway and matrix biomarker profiling into HF and HFpEF prognostication has been proposed previously and the results of the present study support these efforts39.Assessment of the prognostic value of these HFpEF-associated proteins for clinical outcomes revealed markers of angiogenesis to be significant. CLSTN2 was associated with incident HF hospitalization, HFpEF hospitalization and mortality across our study cohorts. CLSTN2 is predominantly expressed in neurons and is involved in synaptic transmission, but is also expressed in adipose and cardiac tissue40. In the present study, metabolic proteins including AOC3, IGFBP3 and SERPINA12 were selected in the sparse protein model for discriminative capability of predicting HFpEF and AOC3 and SERPINA12 were associated with risk of incident clinical outcomes. Adding support to the importance of dysregulated metabolism in HFpEF, AOC3, also known as vascular adhesion protein, is involved in leukocyte trafficking in response to inflammation and adipogenesis41. AOC3 has marked homology with copper-dependent semicarbazide-sensitive amine oxidases (SSAO), whose products cause endothelial damage and oxidative stress43. Circulating activity of AOC3/SSAO increase during biological stress in multiple pathologic states including HF, obesity, atherosclerosis, diabetes and inflammatory liver diseases, and are an independent marker of mortality in patients with chronic HF48. Here for the first time, we show elevated levels of AOC3 to be associated with prevalent HFpEF and to have discriminative capabilities in HFpEF prediction and risk of incident HFpEF hospitalization. Lower levels of IGFBP3 were associated with HFpEF in CATHGEN49, as well as incident HF hospitalization across CATHGEN and TECOS. In endothelial cells, IGFBP3 has antiproliferative and antiapoptotic effects via inhibition of VEGF50. The pleiotropic effects of IGFBP3 on cell growth and metabolism via both IGF-independent and IGF-mediated pathways warrant future investigation in HFpEF. The present work shows that higher levels of pro-angiogenic proteins including VEGF-D and CLSTN2 and lower levels of anti-angiogenic IGFBP3, are associated with HFpEF and incident outcomes. Taken together, these results suggest the opportunity for earlier therapeutic interventions targeting metabolic and angiogenic pathways to prevent HFpEF morbidity and mortality.Several proteins associated with dysregulated fatty acid metabolism were significant in our study. Comorbid conditions of obesity, metabolic syndrome and diabetes are well described in HFpEF. Metabolomic analyses from the CATHGEN cohort have previously identified elevated levels of long-chain acylcarnitines, markers of impaired mitochondrial fatty acid oxidation, in HFpEF and HFrEF compared to controls51; and 11 of the proteins in the 13-protein model have known cis protein quantitative trait loci (pQTL)51 supporting the binding specificity of the assay for its cognate protein. Future work should validate levels of the protein findings here across alternative technologies and consider serial measurements in prospective studies. Diastolic dysfunction was assessed clinically in CATHGEN using the 2009 American Society of Echocardiographic guidelines21 given the time period of the cohorts, and further diastolic dysfunction was not available in TECOS. The appropriate cutoff for EF to study HFpEF is not well established and the data in the present study does not allow for ascertainment of whether some patients may have recovered EF from previously < 40%. In TECOS, only a subset of patients had an EF assessment available limiting our sample size in the validation cohort and EF was only reported as a binned value with cutoffs including 40–55% or > 55%. As all participants in the TECOS validation cohort have DM this may limit the ability to validate proteins that may play a role in HFpEF independent of DM and the generalizability of these data to patients with HFpEF without DM. Given the broad discovery proteomics approach, but limited number of HF events, we applied stringent adjustment for multiple comparisons and only tested proteins that either validated in TECOS or were selected by the LASSO model to limit type-1 error. Given the large number of proteins tested in our models there is mild overfitting in the LASSO models but similar to the clinical model , but we note that the primary goal of this discovery study was to identify potential HFpEF protein biomarkers as opposed to fitting sparse discriminative models. In CATHGEN, HF hospitalization was only able to be ascertained based on ICD diagnosis codes linked to hospitalizations, whereas events were adjudicated in TECOS and JHS, including HFpEF-specific hospitalization in JHS. These different methods for determination of HF outcomes may bias the results given limitations in specificity of the ascertainment in CATHGEN and TECOS. Only two proteins were significant across all three cohorts for HFpEF in CATHGEN and TECOS and HFpEF hospitalization in JHS. This suggests some proteins which discriminate HFpEF may not be prognostic and may reflect sample size limitations and cohort heterogeneity. However, overall the results of this study suggest the diagnostic and prognostic utility of these biomarkers and point to potential dysregulated biologic pathways in HFpEF pathophysiology. Finally, NT-proBNP was not significant in our validation cohort, likely due to the small sample size, however the direction of effect was as expected.There are several strengths to our discovery proteomics approach to diagnostic and prognostic markers of HFpEF. We coupled high-throughput proteomics with a discovery and validation cohort and extended prognostic findings in a JHS, a third independent cohort; in fact, our study represents the largest number of proteins analyzed in HFpEF thus far. Further, we utilized traditional single-marker analyses adjusted for multiple comparisons, but also used pathway analyses given the large number of proteomic pathways represented. Limitations to the study include that Olink protein panels include prespecified proteins of interested compared to other proteomic platforms which allow for the simultaneous measurement of a larger number of proteins, and Olink is relative quantification. However, as evidence for robustness of our assays in orthogonal assessments, we note that five of the 13 proteins in the HFpEF proteomic-model are on the SomaScan platform and are highly correlated with Olink valuesIn conclusion, in a large discovery proteomics study of 459 proteins, we have identified circulating biomarkers of fibrosis, inflammation, kidney injury and fatty acid metabolism to be altered in HFpEF and to improve discriminative prediction when added to clinical variables and NT-proBNP. Our findings strengthen the importance of cardiac remodeling and systemic inflammation in HFpEF compared to patients without HF. We have also shown the prognostic value of these proteins for incident HFpEF hospitalization, HF hospitalization and all-cause mortality. These results suggest the importance of the relevant pathways to HFpEF pathophysiology that also contribute to adverse outcomes and may offer novel diagnostic tools and highlight future therapeutic targets.Supplementary Information 1.Supplementary Information 2.


N-acetyl-L-cysteine, and, to a lesser extent, L-cysteine attenuated the inhibitory effects of cucurbitacin B on the TNF-α-induced expression of ICAM-1, suggesting that an α,β-unsaturated carbonyl moiety is essential for anti-inflammatory activity. The present results revealed that cucurbitacin B down-regulated the expression of TNF-R1 at the initial step in the TNF-α-dependent NF-κB signaling pathway.Pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), induce the expression of intracellular adhesion molecule-1 (ICAM-1) by activating the nuclear factor κB (NF-κB) signaling pathway. In the present study, we found that cucurbitacin B decreased the expression of ICAM-1 in human lung adenocarcinoma A549 cells stimulated with TNF-α or interleukin-1α. We further investigated the mechanisms by which cucurbitacin B down-regulates TNF-α-induced ICAM-1 expression. Cucurbitacin B inhibited the nuclear translocation of the NF-κB subunit RelA and the phosphorylation of IκBα in A549 cells stimulated with TNF-α. Cucurbitacin B selectively down-regulated the expression of TNF receptor 1 (TNF-R1) without affecting three adaptor proteins . The TNF-α-converting enzyme inhibitor suppressed the down-regulation of TNF-R1 expression by cucurbitacin B. Glutathione,  Pro-inflammatory cytokines, such as those belonging to the tumor necrosis factor (TNF) family and interleukin-1 (IL-1) family, are mainly produced by macrophages ,3. NF-κBTNF-α binds to TNF receptor 1 (TNF-R1), which is widely expressed, and TNF receptor 2 (TNF-R2), the expression of which is more limited . TNF-R1 β-methyl-19-norlanosta-5-ene) and are present in Cucurbitaceae and other plant families  vector encoding an ICAM-1 promoter-driven firefly luciferase reporter and a pCR3 expression vector encoding a CMV promoter-driven escribed . RelativTM Protease Inhibitor Cocktail (Sigma-Aldrich) on ice for 15 min. After centrifugation , supernatants were collected as cytoplasmic fractions. Precipitates were further treated with sonication, and centrifuged  to remove insoluble materials. Supernatants were collected as nuclear fractions.A549 cells were treated with Triton X-100 lysis buffer , 2 mM DTT, and 2 mM sodium vanadate) supplemented with cOmpleteWestern blotting was performed as previously described . ProteinStatistical analyses were performed using a one-way ANOVA and Tukey’s test by KaleidaGraph software version 4.5.1 .The present results demonstrated that cucurbitacin B down-regulated the expression of TNF-R1 and inhibited the TNF-α-dependent NF-κB signaling pathway in A549 cells. TNF-α is produced by activated macrophages and other types of cells, while TNF-R1 is ubiquitously expressed in various cells. TNF-α-induced NF-κB-dependent gene expression plays an essential role in inflammatory and immune responses. The acute and chronic activation of the NF-κB signaling pathway has been implicated in the pathogenesis of inflammatory diseases. Natural products, such as cucurbitacins, that target the TNF-α-induced NF-κB pathway have potential as anti-inflammatory agents. Further studies are needed to elucidate the mechanisms of action of cucurbitacins on the NF-κB signaling pathway and develop more selective anti-inflammatory agents. While the present study focused on the cellular response of non-immune cells stimulated with inflammatory cytokines, macrophages are involved in detrimental inflammatory diseases by producing excess inflammatory cytokines, such as TNF-α. Future research is warranted to investigate the effects of cucurbitacin B on primary macrophages and a TNF-α-induced inflammation in vivo model.


Deutsche Zeitschrift für die gesamte gerichtliche Medizin spanned 38 volumes. The 1762 papers contained in those volumes reflect contemporary interests and include many papers from peripheral fields and non-medico-legal disciplines. Publications concerned with issues outside core legal medicine fields in particular allow two distinct tendencies in the development of German institutes of legal medicine to be discerned. Firstly, there is a focus on the psychological and psychiatric aspects of the discipline. Secondly, there is tendency towards a scientific-criminalistic outlook. The fatal consequences of the Nazi seizure of power in 1933 did not spare the sciences. For legal medicine, a discipline with close links to the state, it is unsurprising that fundamental changes to the political system had a significant impact on subject matter. Leaving aside articles notable principally for their ideological content, our analysis of the 38 volumes shows that the papers examined contain new insights into many subjects, some of which are still valid today.From its launch in 1922 to the end of the Second World War, the  Vierteljahrsschrift für gerichtliche und öffentliche Medicin , first published in early 1852. By the time of his death in 1864, a total of 25 volumes had been published. While a number of similar periodicals came and went  would also make possible the destruction of lebensunwertes Leben (“life unworthy of life”). I do not believe any objections to this can be raised from either a racial or medical point of view” (24/114). Other articles on Nazi criminal law deal with other far-reaching new legislation, such as the Gesetz zur Verhütung erbkranken Nachwuchses (Law for the Prevention of Offspring with Hereditary Diseases) or the Verordnung gegen Volksschädlinge (roughly translatable as Regulation against Parasites on the Body of the People).Within the category Articles on other areas of law are primarily concerned with family law and medical law. A review entitled “Legal Issues in Road Traffic Accidents” (28/30) deals with current legislation and case law. There are also isolated essays on the Code of Civil Procedure, the Prison Act and social legislation. Papers on civil law include an essay by Heinrich Többen on “Farmer Eligibility under the Reich Farm Inheritance Act” (29/317), a characteristic piece of Nazi legislation.expert witness work can be divided into two groups. There is a general group comprising 10 articles on methodological problems in appraisals and on fundamental questions such as causality or expert witness professional confidentiality. There is also a more specific group of 39 articles concerned with the significance of medical opinions in specific questions. Most of the published expert witness reports deal with an assessment of the causal relationship between physical injuries and preceding trauma. Thirteen of these relate to insurance medicine, and there are a further 4 case reports on simulation. Three of the case studies on simulated symptoms of illness include descriptions of experiences from the First World War, which in some cases attained renewed significance in the context of insurance assessments.Papers on post-mortem inspection and forensic autopsy are concerned with the performance of these two basic post-mortem diagnostic techniques. Seven articles deal with domestic and foreign regulations on autopsy and the purpose and objectives of court-ordered forensic autopsies. A further 7 articles concern specific autopsy techniques. Additional investigations performed after autopsies span a wide spectrum of techniques and are accordingly found in a number of volumes. Most papers are concerned with either toxicology or histological examination. The 14 publications in this area include both microscopy techniques and forensically significant microscopy findings from histological examination of organ material. There are also 3 papers on bacteriological and 3 on biochemical analysis, 2 on X-ray examinations and 3 on a range of other techniques. Twelve papers discuss expert evaluation of autopsy results. Problems of competing causes of death and the capacity to act of the victim following various traumas are considered in a number of papers. Eight papers deal with exhumation, with repeated emphasis on its diagnostic value. Finally, in 1922 and 1936, several authors called for the introduction of coroner’s post-mortem examinations.The majority of papers on early post-mortem changes. The phenomenon of supravital reactions includes studies on early post-mortem pupil changes , on tissue changes as a result of autolysis , and individual studies on the pattern (11/317) and colour (13/261) of livor mortis. There are 17 articles on problems relating to rigor mortis, forming the largest group of articles relating to thanatology. The main topic considered is the physiology of rigor mortis and the lactic acid theory (2/1). There are also earnest discussions of the special case of catalepsy in both a detailed literature review (2/647) and a collection of case studies (3/349). In addition, there are 3 reports on cases of cataleptic spasm . There are a total of 28 papers on late post-mortem changes. Dictated by practical concerns, the dominant theme is decomposition of the body, with 7 papers primarily concerned with differentiating between putrefactive gases and air embolism. A series of publications  on this diagnostic difficulty were published by Berlin forensic pathologist Felix Dyrenfurth, whose earlier findings were nonetheless overlooked in a later study (6/379). There is also a paper on gas analysis in putrefying lungs (16/459). Three papers explore the aetiology of coffin birth, which at the time had not been definitively explained. There are 9 papers on preservative post-mortem changes and 5 papers on animal bites on bodies. In the field of thanatochemistry, there are 6 papers on the production and detection of putrefaction products such as sulphhaemoglobin and biogenic amines.Twenty-seven papers, almost half of all papers on thanatology, are concerned with time of death estimation. A review summarising the current state of knowledge was penned by Hermann Merkel in 1930 (15/285). Still in the 1930s appeared the first papers on correlations between post-mortem rectal temperature and time of death .The utility of individual post-mortem changes and the diagnostic value of stomach contents and stomach mucosa are discussed as methodologies for vital reactions, by far the largest number of papers are on general reactions. In particular there are 19 papers on embolism, with the largest number of papers on air embolism, followed by fat embolism, and finally tissue embolism. There is one review devoted to the medico-legal significance of these types of embolism when presenting as a paradoxical embolism (23/338). The general vital reactions described include individual studies on haemorrhage, shock, aspiration and swallowing. There is relatively little on local vital reactions. In addition to 2 papers on thrombosis , there are a variety of post-mortem biochemical analyses intended to identify vital changes in post-mortem blood, muscle tissue, bronchi, nerve cells and cerebrospinal fluid.With respect to the phenomenon of blunt force trauma are as varied as the mechanisms by which they are produced. It is not uncommon that external findings are able to shed light on the event that caused them. This is true for the shape of epithelial abrasions (37/33), the morphology of the wound margin (22/299) and “ischaemic impact patterns” (29/408). Scalp wounds also have practical diagnostic significance, and specific features of these wounds are presented in 3 papers. If we look at the distribution of violence by body region, records of head injuries are by far the most common. Of the 11 papers which focus specifically on the shape of skull fractures, one is concerned with the sequence of skull fractures (8/430) and one with the fracture mechanism in babies (32/461). The 30 papers on the wide variety of brain injuries range from concussion and intracranial haemorrhages to delayed consequences of brain injuries. The mechanism of coup-contrecoup injuries is the most frequent subject, discussed in 7 papers, with some debate over Lenggenhager’s centrifugal theory as a potential mechanism . There are also several papers on the consequences of blunt force to the trunk and extremities. The most common subject, with 12 papers, is blunt abdominal trauma and resulting injuries to the parenchymal organs and gastrointestinal tract. There are detailed discussions of the difficulties of reconstructing events in the context of a fall in the mountains (28/90) and a fall from height (30/334). Boxing-related deaths form a special category, with 6 case studies published. The causes of death were intracranial haemorrhage , one case of “death from shock”, in the sense of a sudden cardiac event , and suffocation due to acute laryngeal oedema (1/695). Finally, a further example of death from blunt force is the legendary case report “A new method of insurance fraud: the Tetzner case” (21/112) by Richard Kockel. Of publications on the consequences of sharp force, by far the largest number are concerned with stab wounds. Almost all of the 21 papers on the subject deal with knife wounds and the possibility of using the morphology of the wound to draw conclusions on the implement used or the events of the crime. Far less common are cuts. Five of 6 case reports are concerned with the question of whether a death has occurred as a result of murder or suicide in cases where the throat has been slit. A further small group of cases consists of 5 papers on wounds inflicted with slashing weapons, in which once again the focus is on the question of whether the injuries were inflicted by the victim or a third party. In addition to 2 collections of case studies , there are case reports on 3 suicides involving a blow from an axe . A special form of penetrating trauma is bite wounds. Two of 3 papers on this subject are concerned with the value of bite wounds for identification purposes . There are 4 case studies on criminal corpse dismemberment with cutting tools, 2 of which deal with medico-legal aspects of serial murders by Karl Großmann (3/147) and Karl Denke (8/703). Other reports on high-profile murders committed using penetrating force include the widely reported sexually motivated murder of high school graduate Helmut Daube (14/158) and serial murders by sadist Peter Kürten (17/247).Injuries from asphyxiation, including a number of unusual case studies and several experimental studies discussing morphological features diagnostic of asphyxiation. Gerhard Schrader penned a review (28/134) in which he attempts to produce a systematic overview of then (mid-1930s) known diagnostic criteria for violent asphyxiation. Six of the 7 reports on homicides through manual strangulation concern crimes where the perpetrator has attempted to disguise the crime. One further paper deals with the significance of the carotid sinus reflex (Hering reflex) in strangulation. Following a comparative analysis of human corpses and experimental animal corpses, the author expresses the opinion “that a powerful stranglehold around the neck can lead to unconsciousness as a result of the carotid sinus reflex, without leaving localised signs of strangulation on the victim” (20/361). There are only 5 articles on ligature strangulation, with 3 case reports highlighting the possibility of self-strangulation. A report on a homicide triggered an expert debate on the carotid sinus reflex during ligature strangulation . There are 25 case reports on hanging. The most significant from a pathophysiological perspective are 3 extensive studies  on the mechanism of death by hanging. Further insights into the dying process are provided by “Observations on Death in Executions by Hanging” (22/192) from 1917/18.There are 13 works on death in water are concerned with drowning. These include 8 papers on the pathophysiology of drowning and 3 on morphological findings. Erich Fritz describes a new sign of death by drowning in the form of tears in the mucous membrane of the stomach in people who have died by drowning (18/285), and Gyula Balázs’ collection of case studies represents the first report on “ischaemic impact patterns” after falling into water (21/515). Nine papers describe microscopic and biochemical techniques for diagnosing death by drowning. There are 11 papers concerned with post-mortem changes in bodies recovered from water and the phenomenon by which a body will float back up to the surface some time after death. There are individual treatises on the causes of atypical drowning and death when diving. An instructive overview of death by drowning (18/557) was produced by Gerhard Buhtz.The vast majority of papers on gunshot wounds necessitates close cooperation with forensic investigators, with the result that the section on ballistics includes a number of such publications from the forensic science field. From a medico-legal perspective, the focus is on the effects of and expert assessment of gunshots in humans. A key diagnostic objective is recognising and distinguishing the entry and exit wounds. There are 17 papers on this topic. With respect to determining the range of fire, there are 13 articles on signs indicative of gunshot from close range, including Anton Werkgartner’s first paper in the Deutsche Zeitschrift on muzzle imprint in contact wounds (11/154). There is just one paper (23/375) on determining the direction of fire from the body. In terms of body region, the vast majority, 18 papers, deal with specific features of gunshot wounds to the skull, including one case of a Krönlein shot (1/141). There are just 5 articles on gunshots passing through the chest, causing fatal injuries to the heart and lungs. There are 10 case reports on homicides involving firearms, including an interpretation of findings relating to the shots that killed Communist Party of Germany co-founder Karl Liebknecht (5/247). From 1940, there are 3 extensive reports on large-scale investigations into multiple gunshot wounds. Two legal medicine specialists working separately detail their findings in relation to the victims of Bloody Sunday in Bromberg (34/7) and “from the hostage processions in the Warthegau” (34/54). A forensic science approach is taken in reporting the results of an “Investigation into Polish Atrocities against Ethnic German” (34/90). Seven papers describe rarely used weapon systems  and the characteristic wounds they produce. Finally, 3 reports on explosion injuries can also be considered to belong to this group of topics. These include just a single study on bomb and shrapnel injuries from hostilities during the Second World War (35/173).Investigation of death from electricity, encompassing accidents, suicides and homicides. There are also articles on non-fatal electrical injuries and on animal experiments to investigate electricity-specific effects. There are 5 papers on the appearance and identification of electrical burn marks from the electrothermal effect on the skin. The state of knowledge at the time is summarised in a review by leading expert Stefan Jellinek “On Electropathological Semiotics and Causality” (12/104) and by Friedrich Pietrusky on morphological findings after exposure to technical electricity (29/135). There is also a collection of case studies (32/407) concerned specifically with the consequences of lightning strike.There are 16 papers on heat, with a focus on morphological findings in burns victims, is the subject of 5 publications. Hermann Merkel published a survey of diagnostic options for burnt corpses (18/232). Of the other publications, there are 3 papers on microscopy findings in human corpses  and one study using animal experiments (20/445). There is also one paper on delayed death from burns (35/75) and one on the pathogenesis of scalding (3/401). In addition, there is a case report on a self-scalding by a psychiatric patient in a boiling pan (36/49). A collection of case studies discusses 3 homicides by burning (21/120). The disposal of corpses by burning is the subject of a further 2 papers, and there is also an article entitled “Self-immolation of the Human Body” (18/437) which takes a historical criticism approach. Two papers deal with injuries caused by cold. One reports on the clinical progression of fatal hypothermia (18/270), while the other reports on results from animal studies on general hypothermia (30/199).Thermal damage caused by starvation are described in detail in the context of a death in connection with a fasting regime (6/520).Physical changes in the corpse after various types of external violence are not easily assigned to any of the above groups. These 9 papers are therefore recorded separately. In addition there are two collections of case studies on fatal sport-related accidents occurring in the course of a wide range of different sport activities. Table Reports on single or multiple deaths involving Forensic obstetrics as a speciality is concerned with forensic medical problems relating to pregnancy and childbirth and to the physical condition of neonates. The first group of 14 papers deals with health risks posed by mechanical contraceptive devices, pregnancy testing, complications of pregnancy and indications for terminating pregnancy. Eleven articles on complications of childbirth affecting mother or child are primarily focused on reliably distinguishing spontaneous injury from the results of criminal acts. Six papers on equivocal findings in neonates concern the same objective. Section 90 of the German Code of Criminal Procedure (StPO), which sets out the regulations on autopsies on neonates, raises several questions for the pathologist performing the autopsy. These are discussed in a total of 38 publications. One recurring theme is determining whether an infant was alive at birth, whereby in addition to a number of papers on the lung float test , a cord vessel test (36/21) is cited as a novel method for answering this question. The then Sect. 218 of the German Criminal Code (StGB), which dealt with termination of pregnancy, meant that from the 1920s to the 1940s abortion was one of the primary issues in forensic obstetrics. Consequently, the 30 publications on the methods and consequences of illegal procedures on pregnant women represent a further major topic for the volumes of the Deutsche Zeitschrift examined by this review. While doctors were motivated to oppose illegal abortion because of the health risks for the pregnant women, the Nazi era saw the focus of opposition shift to population policy objectives. Thus we have a 1940 publication entitled, “The Fight against Abortion as a Political Task” (32/226). There were 18 publications on infanticide as defined in Sect. 217 of the German Criminal Code (StGB). These illustrate the difficulties of forensic diagnostics, which consisted primarily of using morphological features to distinguish maternal attempts to facilitate delivery from actions performed with the intention of killing the child.medical malpractice are concerned with medical interventions in both surgical and non-surgical specialities. The most common topic is reports on adverse drug events, of which there are 10 cases. There are 4 reports on anaesthesia-related adverse events during surgery and an equal number alleging post-operative medical errors. There are also 3 case reports on radiation injuries, 2 on complications during diagnostic procedures and 2 on vaccine-related injuries. There is also a review tackling the subject of the “History and Concept of Malpractice” (20/161), primarily from a legal perspective.Publications on sudden unexpected natural death, there is a dramatic imbalance in the number of cases concerning the two age groups generally analysed. While there are 65 publications on adults, there are only 13 papers on sudden unexpected natural death in children. The most frequently cited groups of fatal diseases in adults are diseases of the central nervous system, on which there are 15 publications, and cardiovascular disease with 13 articles. There were far fewer reports on other groups of fatal diseases. There are 3 reports on deceased children, each of which describes malformations or infections as the cause of death. Other publications on childhood consist of individual case reports on children with rare diagnoses and reports on experience gleaned from large numbers of autopsies.With respect to papers on disease and clinical features of various types of poisoning, on which there are 125 papers. In addition to 9 collections of case studies on various poisons and 3 cases of combined poisoning, there are 68 papers on inorganic poisons and 45 on organic poisons. Of the inorganic poisons, by far the most widely discussed are carbon monoxide, on which there are 13 papers, arsenic, on which there are 9, and thallium, on which there are 7. Of the organic poisons, there are notable clusters of 17 papers on plant toxins and 13 reports on pharmaceutical poisoning. With respect to illegal drugs, widespread today, there is just one article on the suicide by cocaine of a male addict (4/40) and one on fatal poisoning with heroin after being mistaken for cocaine by a cocaine-dependent individual (12/112).The largest group of publications in this subject area consists of descriptions of the pathological changes in tissue and organ anatomy plays a role in the diagnosis of poisoning, though it does have some natural limitations. Of 28 papers on this subject, primarily on organ toxicity, 13 are concerned with the central nervous system, which is particularly vulnerable to poisoning. There are also 2 articles on characteristic morphological changes in specific types of poisoning. These are Mees’ lines and coma blisters, both first described in the Deutsche Zeitschrift (34/307). The second, smaller group on pathological anatomical changes consists of 8 reports on histological changes in various organs caused by one or more poisons.The identification of toxicological analysis. In the period under review, 68 papers were published on this subject, of which 59 are concerned with the detection of a single poison or group of substances. As might be expected given the prevalence of carbon monoxide poisoning in that era, by far the most common, with 16 examples, are papers on various techniques for detecting CO or CO-Hb. Of groups of substances, alkaloid detection is described in 4 papers. In contrast to analysis of specific substances, there are 9 papers on the technical details of a variety of analytical methods, including spectrophotometry (1/411) and microscopy-based melting point determination (34/353).The gold standard for diagnosing poisoning is toxicology of alcohol. Of 33 publications on alcohol metabolism, 26 studies are concerned with factors which might affect the absorption, distribution and elimination of alcohol, such as age, trauma and chemicals. Just 5 publications focus on the correlation between blood alcohol concentration and the effects of alcohol. These include a review entitled “On the Terminology and Forensic Assessment of Alcoholic Intoxication” (14/296), which outlines the state of knowledge in 1930. The only paper on fatal alcohol poisoning (33/44) is primarily concerned with developmental peculiarities of alcohol clearance in children. There are 2 papers specifically on methanol poisoning. Thirty-one publications deal with methods for determining alcohol concentration in body fluids and tissues. In addition to two reviews , there are papers on individual methods, including the Nicloux method (18/638) and the Widmark method (19/513). Between these papers and the end of the war, there are 18 further studies on Widmark’s micromethod. All of these papers deal with subtle technical points for overcoming shortcomings in the method and eliminating sources of error.One area of toxicology which has evolved into its own extensive speciality is the identification methods. Six publications in the field of osteology discuss ways of differentiating between species, sex diagnosis and age estimation from skeletal remains. One of these papers, which takes a detailed look at the forensic medical significance of the humerus, includes basic data on several identification procedures (22/332). There are also 4 papers specifically on skull identification, 2 of which include detailed descriptions of techniques for the superprojection of an unknown skull onto a photograph of a missing person . Five papers describe odontological features which can be used for identification, and 2 linked articles discuss identifying features visible using radiological techniques. A case study on the particular utility of the frontal sinuses for identification (7/625) is the first paper on X-ray comparison in the German-language literature (10/81). There are 2 papers dealing with methods for specific investigatory queries and combinations of methods required for problem cases.There are a total of 21 publications on then common medico-legal blood groups to appear in the Deutsche Zeitschrift was published in 1924 and was entitled “On the Forensic Significance of Isoagglutination of Red Blood Cells in Humans” (3/42). The papers on test methods discuss the production, use and storage of test sera and other reactants. The first publication on the prevalence and distribution of A subgroups in the Deutsche Zeitschrift was by Karl Landsteiner, the discoverer of the main blood groups. Another pioneer of blood group research, Berlin serologist Fritz Schiff [Table z Schiff , publishtrace evidence analysis. The contemporary arsenal of methods necessitated a hierarchical sequence of examinations: test for the presence of blood − determine species specificity – determine blood group. Four articles describe peroxidase and crystal tests for blood detection. The specificity of these tests varies and depending on the test method should be considered a presumptive test only. The Uhlenhuth test has been used to determine species specificity since 1901, predominantly to distinguish whether trace material is human or animal [The practical use of identificatory blood groups is the subject of medico-legal r animal . The preindividuation of bloodstains. Landsteiner and Richter’s agglutination test represents the beginning of the use of inherited blood groups for forensic purposes [The discovery of human blood groups was soon followed by the development of the first methodical approach to the purposes . The smapurposes  (16/445)bodily fluid traces. Epithelial glycogen content can be used to identify vaginal secretions (4/1). There are 4 papers on sperm detection, including a new microscopy-based detection method using Baecchi’s stain (27/143). There is also a review describing a number of older tests for detecting dried saliva (11/211). At the time, it was possible to determine AB0 groups both from semen and from other bodily fluids (23/186).Detection methods based on characteristic constituents of bodily fluids were developed for various hair traces is concerned with using morphological features for species differentiation. Other topics include determining developmental stage and sex, and demonstrating damage to the hair.The majority of papers on traces of faeces discuss serological identification based on coliform bacteria and a presumptive assignment of blood group.The papers on family relationship testing relied on fertility testing, assessment of the period of gestation, a comparison of morphological characteristics and blood typing. There is one paper on fertility testing (26/64) and one on the legal and medical problems involved in assessing the period of gestation (14/11). The papers on hereditary biology deal with various physical characteristics for assessing similarity and with dactyloscopic criteria. By far the largest number of articles are on the results of blood group testing. As well as helping to clarify cases of disputed paternity, blood group analysis also served as evidence in a case of babies switched at birth (25/79). There is also a report by a veterinarian on “Blood Groups and Paternity Determination in Horses” (25/231).In the period under review, There are numerous areas of overlap between legal medicine and scientific-technical criminalistics. In the period under review, these went well beyond the kindred forensic biology role performed by the two disciplines. Table In his article “Institutes of Legal Medicine and Criminalistic-Technical Activities Performed within Them” (14/411), Martin Nippe explicitly advocates for legal medical institutions to take over criminalistic tasks. His argument that the courts and police lacked the necessary expertise seems entirely reasonable in light of advances in forensic technology in Germany at the time. The urgency of this need is demonstrated by a collection of case studies from Göttingen College of Legal Medicine (14/26). It features cases which indisputably fall within the purview of forensic technology.theory of forensics, on which only a single paper (34/404) was published. This paper, by distinguished Berlin criminalist Hans Schneickert, deals with the key insight that the prevalence of an identifying feature determines its value for identification purposes.One field in which there were particular deficits was forensic biology. In Germany, this sub-specialism was not an established academic discipline in the period under review, so that in the majority of cases the investigation of biological trace evidence was contracted to institutes of legal medicine. Accordingly, the theory behind many of the methods used in forensic genetics is equally applicable in the field of forensic biology. Twelve publications relate to the key criminalistic task of finding and securing bloodstains at a crime scene and on objects. Two studies on the morphology of bloodstains deal with fundamental insights into blood splatter patterns (16/272) and drip patterns (22/387). A further 2 papers are concerned with non-human material.The picture is very different for the science-focused sub-disciplines of forensic technology. Almost exclusively as a result of research performed at institutes of legal medicine, a considerable body of knowledge had already been amassed in the field of forensic chemistry, there are 8 papers on chemical analysis for the purpose of determining the cause of fire. In relation to 2 attempted fatal poisonings, there is a report on demonstrating the presence of poison in food. In addition, there are papers on the chemical/physical chemical analysis of glass dust, metal, wax, paint and paper (one paper on each).Of the publications on ballistics. Of these, 20 papers deal with different methods for determining firing range, making this a major thematic focus in this area. The second large group is made up of 11 publications on firearm identification from firearm trace evidence. There are just single publications on determining bullet trajectory in long-range shots and on evidence on the firing hand.Of the technical sub-specialisms, by far the largest number of papers in this area are dedicated to writing examination are to determine the authenticity or otherwise of a document and to identify the person responsible for a piece of handwriting or typescript. Nine papers are concerned with identifying an author based on individual handwriting features. Three papers look at criteria which enable handwriting and typescript to be falsified. One paper looks at typewriter identification and one at determining the age of a piece of handwriting or typescript. There is also a critical analysis which calls for proof of competence to be required before a court accepts the evidence of a writing expert (27/364).The objectives of forensic document examination are concerned with the examination of paper scraps, betting slips and paintings.The publications on dactyloscopy is the comparative examination of fingerprints to identify individuals. Five papers describe the conditions required and evaluation criteria for fingerprint identification. There are 3 articles  on methods for visualising latent fingerprints. Two articles examine fingerprint identification in dead bodies in an advanced stage of decomposition .One of the main areas of application of shape traces deals with various impressions on a wide range of media. The publications in this field deal with tool, shoe and bite impressions and methods for preserving a trace.The field of photography, it is essential that images are informative and authentic, as they constitute evidence admissible in court. The publications in this field therefore deal with photographic and technical requirements for producing usable photographs.In the field of forensic odorology is a case report on the evidentiary value of olfactory traces of specific wood shavings (6/1).The sole work on investigative techniques deal with the application of UV light, spectrography and X-rays.The papers on age estimation in living subjects, one exploring diagnostic criteria in minors and one exploring criteria in adults. Leningrad author W. A. Nadeshdin produced a detailed discussion of features of the face and dentition useful for age estimation in adults (6/121). In a second work on age estimation in minors, he proposes growth, tooth eruption and signs of sexual maturity as a reliable combination of features for this purpose (8/273).There are two papers on drug addiction. The first, a case study (12/285) on multiple drug use with a preference for chloroform in a pharmacist, was published in 1928. The second report (35/17) from 1941 discusses the medical history of a patient who, following morphine addiction, had developed pethidine addiction.The papers analysed include just 2 early reports on child abuse (13/159) is the only publication on this subject.A detailed discussion of the legal and social significance of self-inflicted injury always aims to reveal the motivation. The 2 published cases are both cases of attempted insurance fraud, in one by severing the left thumb (23/352), in the other through many years of self-harm involving repeated injury to the left arm (38/244).The investigation of psychopathology. Of these, most are concerned with individual forensically significant psychopathological phenomena, discussed in 26 articles. There are 11 papers specifically on the forensic significance of psychiatric disorders. These include various psychoses, paralysis and dementia. Four case studies consider previous trauma in relation to the assessment of psychotic disorders. There are 3 papers on the principles and diagnostic value of graphology. There are a further 3 case reports on the psychopathology of prominent figures, including the sensational case of railroad killer Szilveszter Matuska (20/53). Of a paper by Roland Freisler, then Secretary of State in the Reich Ministry of Justice, entitled “The Psychology Underlying Polish Atrocities, Illustrated through the Development of the Polish National Character”, only the title was reproduced (34/7). The paper takes a social psychology approach, and the full text was published in a legal journal.The vast majority of publications on this topic are concerned with psychology of testimony. Topics include the impairment of memory capacity, in particular through hypnosis and suggestion, recognising false testimony and assessing testimony given by children and adolescents.A themed group of 13 papers covers various subjects in the field of the suicidology is, in the selected context of forensic psychiatry and psychology, reduced to problems in this area. The publications classified as belonging to this field are therefore primarily concerned with motives and risk factors for suicide. There are 14 articles on these topics, which, in addition to the sequelae of diseases and head injuries, discuss individual constitution as a causal factor. Epidemiological surveys on suicide in Russia, in Finland and in Berlin are the subject of 3 papers. In a paper entitled, “Contribution on the Psychology of Suicide” (12/346), the phenomenon of imitation is discussed in the context of a popular suicide location.The interdisciplinary field of paraphilias—sexual perversions in the nomenclature of the time. Topics covered by the 23 publications in this area range from sexually motivated self-mutilation, to rare potentially criminal forms such as fetishism, to clearly criminal acts such as incest and paedophilia. There is also one article calling, as part of a pending reform of criminal law, for the abolition of Sect. 175 of the German Criminal Code (StGB), which at the time made consensual sexual acts between men a criminal offence. The first appeals for sterilisation for deviant sexual behaviour were published as early as the 1920s. The earliest work on “Castration and Sterilisation as a Means of Treatment” (3/162) from 1924 is concerned with applying these measures both in the event of a “mental disorder triggered by functions of the sexual organs” and for the elimination of “a criminal activity”. A later article (14/432) from 1930 rejects the idea of surgical castration of sex offenders, but advocates radiation treatment of recidivist sex offenders. These two publications are followed by 9 further papers, all relating to the “Law against Dangerous Habitual Criminals and on Measures of Reform and Prevention” (Gesetz gegen gefährliche Gewohnheitsverbrecher und über Maßregeln der Sicherung und Besserung) of November 24, 1933 and in particular to the newly introduced measure “castration of dangerous sex offenders” in Sect. 42a (5) of the German Criminal Code (StGB). The last of these papers, entitled “Observations on Castrated Sex Offenders” (33/248) concludes with the assertion that, “the success of castration has far exceeded expectations.” Two papers discuss the physical consequences of castration, and a further 2 papers fertility in the context of the Law for the Prevention of Offspring with Hereditary Diseases (Gesetz zur Verhütung erbkranken Nachwuchses).Of the many and varied references to human sex life subsumed into the category sexual medicine, one extensive group is the hymen examination. These publications illustrate the extraordinary variability of the healthy hymen and to an even greater extent the pathologically altered hymen. Depending on shape and elasticity, where sexual violence is suspected, consideration must be given to the possibility of vaginal penetration without rupture of the hymen (14/265).There are 7 papers on autoerotic fatalities. Classification of a death as an autoerotic fatality is solely dependent on the circumstances at the site of discovery and is independent of the cause of death. Of 6 cases reported, 3 were deaths by hanging, and of the remainder the cause of death was only able to be determined in one case.Fatal sexual activity without a sexual partner is the common element uniting the phenomenon of sexual dysfunction and 2 publications on intersexuality.Other sexual medicine topics are treated in 3 papers on male road traffic accidents. These include 7 papers on the biomechanics of road traffic accidents. Injuries to the head and extremities are discussed with respect to their value for investigative purposes. Two papers discuss trace evidence after road traffic accidents and their use in accident reconstruction. A further 7 articles are concerned with the effects of alcohol on driving performance and the resulting accident risk. Two papers are concerned with other causes of accidents. Just one publication, titled accordingly, discusses the important question in rail traffic accidents of whether a victim was “Alive or Dead when Run over by a Train” (25/147).Almost all publications in this field are concerned with the causes of and consequences of injuries incurred in occupational problems relating to diseases with social sequelae. There are also 3 articles concerned with public health and a positively classic social medicine paper entitled “The Effect of Social Situation on the Health Status of Welfare Recipients” (20/535).There are just 11 publications relating to social medicine. Seven articles are primarily concerned with criminal biology, originally known as criminal anthropology, make it the best represented. Six papers in the then emerging research field of “blood groups and criminality” (9/426) form part of a series with 6 other papers on constitutional biology, most of which are concerned with identifying physical signs of a tendency to criminal activity. A prime example of forensic genealogical research is a case study entitled “A Criminal Family” (25/7). The increased focus on criminal biology, while pre-dating 1933, reaches its climax in the Nazi era. On this subject, Theodor Viernstein, head of the Bayerische kriminalbiologische Sammelstelle  wrote, “The Law for the Prevention of Offspring with Hereditary Diseases of July 14, 1933 and the Law on Measures against Dangerous Habitual Criminals of November 23, 1933 are, for our question, two decisive landmarks on this road to the renewal of the living race of our people. What until the seizure of power remained, in the minds of scientists, psychiatrists, hereditary biologists and racial hygienists, a mere hope for the future, was given, at the very moment that the people faced their greatest hardship, practical form and legislative expression” (26/3).Of the sub-specialisms historically forming part of the field of criminology, 19 publications on criminal psychology. In addition to a review on the concept and scope of the discipline from a 1940s perspective (36/119), there are 9 articles dealing with the psychology of specific offences, the most common of these being arson, to which 4 articles are dedicated. Finally, there are 2 papers discussing crimes committed under hypnosis.There are 12 publications in the field of Criminal phenomenology encompasses publications on the phenomenology of various groups of offences. The only group on which multiple papers are published is youth criminality. There are 5 papers on this subject, in one of which the supposedly modern term “hooliganism” is used in a wide-ranging sense (12/361). Heroin misuse, already widespread in the mid-1920s, is used to illustrate the consequences of drug addiction (8/81). Other topics include homicide, sexual offences and arson. The 3 papers in the field of victimology look at the physical and psychological sequelae experienced by female minors who have been the victim of a sexual offence. There are 2 publications on crime prevention, both discussing the basic idea that social welfare is an important component of crime prevention. In the criminal aetiology field, there is one article on the effects of war on criminality (1/697) and another on “Reading as a Stimulus to the Commission of Crime” (13/209).prisons and imprisonment, 8 concern self-harming by prisoners. The incidents described range from inducing symptoms of illness and swallowing foreign objects, to significant self-mutilation. A number of papers are also dedicated to the topic of illness and its treatment in prisoners. The 6 articles on this issue are primarily concerned with physical illness. Four papers tackle the subject of the role of medical personnel in prisons. Two further articles discuss the particular problems faced by adolescents in prison (4/121) and support for prison leavers in Germany (11/423).Of 20 publications on Erb- und Rassenpflege” .The period under review commences in 1922, shortly after the establishment of the Weimar Republic, and ends in the spring of 1944, in the final phase of the Second World War. The Nazi seizure of power in early 1933 represents a profound political and historical rupture. For legal medicine, a discipline with close links to the state, this social upheaval inevitably had an impact on issues and outlooks within the discipline. A comparison of the 19 volumes published from 1922 to 1932 with the 19 volumes published from 1933 to 1944 shows that, in addition to the expected effects on legislation and jurisprudence, there was also a transformation in the sociological and even in some cases medical topics within the discipline. Particularly striking is the link between legal medicine and the Nazi creed of “The start of the period under review saw legal medicine as a discipline attains a long-awaited goal. Following repeated calls from prominent experts in the field , in 1924 the discipline was finally included in the state medical examination, having been a compulsory subject for students of medicine since 1901 . On the Deutsche Zeitschrift discuss a number of pieces of legislation which emphatically highlight the consequences of Nazi ideology for the judiciary. While the vast majority of laws were rendered legally void by Control Council Law No. 1—Repealing of Nazi Laws of September 20, 1945, the provisions introduced in 1934 by the Law against Dangerous Habitual Criminals and on Measures of Reform and Prevention—with the exception of the provision on castration in Sect. 42a of the old version of the German Criminal Code—remained largely unchanged. The provisions on preventive detention were incorporated into the Criminal Code in 1953 (BGBl. I p. 735) and tightened in 1969 (BGBl. I p. 645). As a result, the Law on Habitual Criminals introduced a dual system of sanctions into German criminal law which still exists today.Articles in the th congress of the scientific association held in Erlangen in 1921, the main topic was the introduction of coroners autopsies. Three speeches were dedicated to the reasons why it was necessary to have this option available . This unanimous appeal by all three speakers was further reinforced by the speech which followed them, entitled “Experiences with Coroners Autopsies in Hamburg” (1/17). Despite the endeavours of the scientific association, no uniform legislation on this was forthcoming, with the result that in 1936 calls for the introduction of coroners autopsies were again the topic of the opening speech (28/1) at the 25th congress in Dresden. The text drafted by speaker Hermann Merkel entitled, “Reich Legislation to Introduce Coroners Autopsies” was unanimously adopted by congress participants. Despite this, corresponding legislation was still not on the statute books by the end of the war. It was not until 1949 that legislation was passed in East Germany to enable coroners autopsies to be performed in cases where the cause of death was unclear. Forty years of positive experiences [At the 11eriences  did not,Post-mortem rectal temperature measurement has proven to be highly valuable for estimating time of death. The first studies on the rate of cooling of dead bodies were published in 1937. Successive rectal temperature measurement produced very regular exponential decay curves. The temperature initially drops only modestly, followed by a period in which it drops more quickly, followed by a further period of slower decline . Though first described many years ago, this sigmoidal cooling curve still forms the basis of standard methods of temperature-based time of death estimation even today .Through a case report on the examination of an exhumed skull (8/430), Kurt Walcher drew expert attention to the rule on the sequence of skull fractures first published by Georg Puppe a few years earlier. Evaluation of fracture lines showed that the victim had suffered at least one hard blow before being killed by a gunshot to the head. As in the above case, it has subsequently proved possible to use this principle to determine the sequence of multiple skull fractures, including fractures arising through different means. Puppe’s rule is now regarded as essentially proven .Deutsche Zeitschrift. Franz was of the opinion that the theory must be wrong, as the speed of the process meant that it was not possible for the posited outflow of blood and cerebrospinal fluid to occur (31/61). In his reply (31/278), Lenggenhager explained his theory using his original publication. Though in his concluding remarks (31/280) Franz maintained his objection, the suction theory is today considered the most scientifically reasonable explanation for contrecoup injuries [A wide variety of views have evolved on the mechanism of coup-contrecoup brain injuries. Lenggenhager  theoriseinjuries , 18.By the early 1920s, there had already been a number of works on the mechanism of death by hanging. Nonetheless, it remained unclear “to what moment—whether it be the closure of the airways, the obstruction of the cervical vasculature or the effect on the vagus nerve—is to be attributed the main role in death by hanging” (1/686). Georg Strassmann studied the effect of strangulation on the airways and found that, for normal hangings, it was impossible to overcome the obstruction of the airways by even “the most strenuous inhalation or exhalation”. Where the noose was applied in an atypical position, it appeared that there was at the least “greater difficulty in obtaining air” (4/165). Walther Schwarzacher performed experiments to determine the tensile force required to obstruct the cervical vasculature (11/145). With a standard symmetrical noose position and a constant internal vascular pressure of 170 mmHg, a force of 3.5 kg is sufficient to obstruct the common carotid arteries, while a compression force of 16.6 kg is required to obstruct the vertebral arteries. The results of these two series of experiments remain part of the corpus of knowledge on the mechanism of death by hanging even today.Techniques for diagnosing death by drowning were enriched by the repeated observation by Erich Fritz of a specific morphological finding (18/285). He identified tears in the mucous membrane of the stomach in several victims of drowning in which there was no evidence of any violence to the body prior to drowning. With respect to the origin of this indication of drowning, after some initial uncertainty the opinion prevailed that the tears are caused by overstretching of the stomach primarily through swallowing the drowning medium . Gyula BDeutsche Zeitschrift on muzzle imprint in contact wounds (11/154). A muzzle imprint proved to be a reliable sign of a contact wound, and its characteristic shape can also provide information on the type of weapon and the position of the weapon on firing [As with the numerous forensic science publications on ballistics, forensic traumatology papers have similarly enriched our understanding of the effects of gunshots. This is particularly true for the first paper by Anton Werkgartner in the n firing .In the forensic obstetrics field, most publications are concerned with questions arising in the context of Sect. 90 of the German Code of Criminal Procedure (StPO). The main focus here is on determining whether an infant was alive at birth. In addition to the lung float test, for the abolition of which Albin Haberda appeals in a piece unequivocally entitled, “Out with the Lung Float Test!” (14/7), János Prievara presented the cord vessel test 36/21) after Jankovich [ after JaThere are various, generally non-specific signs on a body that may lead to a suspicion of poisoning. In some cases, specific pathological anatomical changes may act as an initial indication of potential poisoning. In a paper entitled “Post-mortem Findings Following Sedative Poisoning” (34/307), Franz Josef Holzer reported his systematic observations on the topic. He identified characteristic violet patches, often symmetrical, especially in particular areas on the legs. These lesions, today known as coma blisters, are observed particularly in barbiturate poisoning.Deutsche Zeitschrift carried a paper entitled “On a Technique for Quantitative Determination of Blood Alcohol Levels after Widmark’s Method” (19/513). Validated by numerous studies, Widmark’s micromethod established itself as the standard technique for determining blood alcohol.In 1932, almost contemporaneously with the German edition of Erik M. P. Widmark’s ground-breaking monograph , the DeuOver the period under review, advances in methodologies for the identification of unknown bodies occurred primarily in the fields of anatomy, anthropology and osteology. In addition to publishing an article on plastic reconstruction of the physiognomy (8/365), anatomist Friedrich Stadtmüller also subsequently developed a graphical method for verifying skull identification  based on Welcker’s method . Dionys Following the discovery of the main blood groups in 1901, the discipline of forensic serology developed into a more and more significant field . In the The scientific disciplines of forensic technology have the greatest overlap with legal medicine, both in theory and in practice. Consequently, legal medicine specialists have published numerous scientific articles and case reports on forensic biology and forensic chemistry. The publications evaluated here include significant excursions into the quite separate field of criminalistics, for example papers on traces of blood in snow (31/213) and the analysis of incendiary agents (36/245). Of the scientific sub-disciplines, the most prominent is ballistics, on which there are many papers which belong thematically to the field of criminalistics. This is particularly clearly illustrated by publications on firearm identification from trace evidence on munitions (18/350) and on cartridge cases (21/190). Similarly, numerous publications on writing examination can without exception be filed under forensic technology. Martin Nippe’s call for legal medicine institutions to take over criminalistic tasks (14/411) reflected a contemporary imperative aimed at providing greater legal certainty, as there were simply not enough experts in forensic technology. In the long term, however, legal medicine needed to be liberated from such a non-medico-legal role, and this occurred only gradually in the post-war period.Clinical legal medicine is significantly underrepresented compared to more recent times. There are only a handful of publications on the subject, and there are no papers on a number of significant health-related issues. Particularly notable by their absence are articles on the physical examination of victims and perpetrators of violent and sexual offences. Child abuse is another area which is notably underrepresented, with insufficient consideration of the complex fabric of causation and the many different patterns of harm. In an extensive analysis (13/159), Ernst Ziemke laments the fact that “reports from experienced legal medicine specialists and my own experience suggest that it is relatively rare for cases of child abuse to be subject to a medico-legal assessment. Where they are, this is almost always cases in which the outcome of the abuse has been the death of the child.”The publications analysed here deal with a wide range of forensic psychiatry and psychology topics. Legal reforms and the effects of changes to our understanding of medical psychology mean that the technical content has changed repeatedly since. Terms such as neurosis (1/325) and hysteria (2/523) are now considered outdated, and significant progress has been made in areas such as intelligence testing (8/580). The psychology of testimony, one of the cornerstones of interrogation theory, has evolved into an extensive sub-specialism within the forensic psychology field.In sexual medicine, there has been a quite contradictory development. While the sexology side was working towards liberalisation, controversial legislation on sexual offences remained in place. The quite reasonable call for the repeal of Sect. 175 of the German Criminal Code (StGB) (13/59) was not heeded until the end of the war. Even more draconian were the newly introduced “Measures of Reform and Prevention”. Under the 1933 Law on Habitual Criminals, a court could order, under Sect. 42a (5) of the Criminal Code, the “castration of dangerous sex offenders”. As long as certain statutory conditions were met, Sect. 42 k (1) of the Criminal Code extended the groups covered by this provision to include sex offenders with repeated convictions for exhibitionism. While there were many publications on the draconian measure of castration and its consequences, there is by contrast a notable absence of publications on sexual therapy for the afflicted.Increasing motorisation of road traffic saw the emergence of traffic medicine as a new sub-specialism within the legal medicine field. Publications on road traffic accidents are concerned with either the causes of accidents or the consequences of injuries. With the advent of motorised road traffic, it was not long before alcohol consumption by drivers (29/11) and pedestrians (32/312) was identified as a notable cause of accidents. There followed experimental studies aimed at obtaining concrete data on how alcohol affects driving performance, for example the effects of alcohol on eye and ear function (32/301). Results from these studies also represented an academic foundation on which to base road traffic legislation. With respect to the consequences of injuries, there was a need to clarify common patterns of injury and their origin . Insights gained into mechanisms of trauma offered an opportunity to use medico-legal findings to contribute to accident reconstruction.Given that for many years the name of the German scientific association included the words “social medicine”, the paucity of publications in this field is particularly surprising. The small number of works present are not even vaguely representative of the interdisciplinary field of social medicine.In keeping with the concerns of the period, most of the papers in the field of criminology are related to criminal biology. Studies of offenders inspired by constitutional biological investigations claimed to find features characteristic of born criminals and thereby to justify a robust approach to the fight against crime. In addition to studies on morphological features, efforts were also made to identify serological markers. A significant field of research at the time was the relationship between blood groups and crime. Results from such studies were, however, somewhat contradictory. In a study on more than 1000 convicts, Kurt Böhmer found “a general increase in blood group B. This increased further in recidivists and the irreformable” (9/426). Augustin Foerster’s accurate conclusion unambiguously disagreed. He was unable to confirm “that blood group B is more common in the prison inmates I examined. It was not possible to demonstrate any link between blood group and criminality” (11/487). A key area in criminal psychology is arson, the explanation for which in one case shifted from the classic pyromania to a “simmering subconscious sexuality” (33/52). To obtain usable insights into significant phenomenologies of criminality, criminal phenomenology requires up-to-date studies. Only if this condition is met can results from studies be used to formulate policy on crime or be applied to forensic science investigations. For this reason, publications on a range of phenomenological criminology topics are of historical interest only. Articles on female minors who have been the victim of a sexual offence represent the beginnings of victimology, which evolved into an academic discipline only after the Second World War.In conclusion, it is worth noting that, despite the many papers from a variety of fields closely allied with but distinct from legal medicine, most of the published papers are concerned with the traditional key fields of traumatology and toxicology. There are also large numbers of articles on various topics in the thanatology and forensic obstetrics fields. An emerging sub-specialism is the study of blood groups, which found increasing use in trace evidence analysis and in family relationship evaluation.


In this study, we aimed to position the Hungarian Merino among other Merino-derived sheep breeds, explore the characteristics of our sampled animals' genetic similarity network within the breed, and highlight single nucleotide polymorphisms (SNPs) associated with daily weight-gain.Hungarian Merino (n = 138) was genotyped on Ovine SNP50 Bead Chip  and positioned among 30 Merino and Merino-derived breeds (n = 555). Population characteristics were obtained via PLINK, SVS, Admixture, and Treemix software, within-breed network was analysed with python networkx 2.3 library. Daily weight gain of Hungarian Merino was standardised to 60 days and was collected from the database of the Association of Hungarian Sheep and Goat Breeders. For the identification of loci associated with daily weight gain, a multi-locus mixed-model was used.ST values are 0.035 and 0.036, respectively). Among Hungarian Merino, a highly centralised connectedness has been revealed by network analysis of pairwise values of identity-by-state, where the animal in the central node had a betweenness centrality value equal to 0.936. Probing of daily weight gain against the SNP data of Hungarian Merinos revealed five associated loci. Two of them, OAR8_17854216.1 and s42441.1 on chromosome 8 and 9  and one locus on chromosome 20, s28948.1 , were close to the markers reported in other breeds concerning daily weight gain, six-month weight, and post-weaning gain.Supporting the breed's written history, the closest breeds to Hungarian Merino were Estremadura and Rambouillet (pairwise FThe position of Hungarian Merino among other Merino breeds has been determined. We have described the similarity network of the individuals to be applied in breeding practices and highlighted several markers useful for elevating the daily weight gain of Hungarian Merino. Merino is a globally distributed fine-wool sheep breed representing about a quarter of the global sheep population . The appThe Industrial Revolution in Britain led to considerable improvements in weaving efficiency. The development of the wool textile industry resulted in an increasing global demand for wool. The selection process for wool quantity and quality resulted in many types of Merino sheep worldwide . The HunAt the end of the 19th century, in spite of the Western European meat-breeding trend, wool was still the main production objective in Hungary ,9. In thIn the 1980s, wool production was pushed into the background, and Hungarian Merino was considered a dual-purpose (meat and milk producing) breed . A drastIn the 2010s, due to considerable declines in breeding activity, Hungarian Merinos became classified as endangered. Subsequently, the Hungarian government began to provide subsidies for the conservation of genetic resources as part of the coordinated breeding program. In 2020, the Hungarian Merino population amounted to 8,167 ewes. At present, only 11 Hungarian genealogical lines are noted [We had three aims in this study: i) Since the genetic position of the Hungarian Merino is not known based on single nucleotide polymorphism (SNP) markers, our samples were compared to the database of Ciani et al  to deterApproval from the animal research ethics committee was not necessary as the ear tag application is part of the routine breeding procedure.30–80/d, where BM30–80 is the body mass of the lamb measured between 30 and 80 days of age, d is the actual number of days after birth on the day of the measurement.The sampling locations covered Hungary, which has a quite uniform, continental climate. One hundred thirty-eight Hungarian Merino ewes were included in this study. They were born in different years spanning 10 years. Their daily weight gain was standardised to 60 days as follows: 60 BMEar cartilage samples were collected by the TypiFix  tissue sampling system  during rQuality control of data included filtering for mapped and polymorphic markers  with a call rate higher than 0.95. Duplicated samples were not detected >0.95). Call rates of the genotypes were over 0.95 for all Hungarian Merino samples. The matched database, which included both the Sheep Consortium  and HungIn case of within-breed network and association analyses 46,906 SNPs were kept after data filtering.IS), genetic distance (FST), and principal component analysis (PCA) were performed by SNP & Variation Suite 8.1.1 program . The inbreeding coefficient is the (observed - expected number of homozygous markers)/(number of genotyped markers - expected number of homozygous markers). Expected number of homozygous markers ST is explained in Weir and Cockerham [Determination of observed and expected heterozygosity , subsequent calculation of inbreeding coefficient  from the value of cross-validation error in each Ks . Before Treemix 1.13  softwareNetwork analysis concerning the relation of the Hungarian Merinos was based on the identity-by-state value of pairs calculated by SVS program .Identity-by-state pairwise distance between any two individuals is .st is the total number of shortest paths from node s to node t and σst is the number of those paths that pass through the node v. The strength of the connections (or weight of the edges) is defined by the IBS value of the pairs. IBS values above the 85 percentile were visualised by thick black lines.Based on IBS values, the net properties of the Hungarian Merino samples, such as diameter and betweenness centrality, were calculated and visualised by Python 3.6 software using the libraries networkx 2.3 and matplotlib 3.1.1. Diameter is the largest distance observed between the pairs of nodes or the distance between the two furthest nodes . BetweenFor the identification of loci associated with daily weight gain, a multi-locus mixed-model was used . The modIS value (0.341), whereas, in Hungarian Merino, this value was 0.054. Pairwise values of FST calculation fell in the range of 0.004 to 0.313  above 0.73 have been maintained. In the analysed population, this value provided an interconnected network of all nodes . The number of edges was 341, the average degree was 5.014, net diameter was 4 . The fiv10P>10) on chromosome 2, 3, 8, 9, and 20 with false discovery rates lower than 4.1e–11.The values of daily weight gain ranged from 261.6 to 494.7 grams. In multi-locus mixed-model analysis, several loci were identified to be associated with the daily weight gain . The topIS value calculated from observed and expected heterozygosity of Hungarian Merinos is below 0.055. Ten out of the thirty-one studied breeds have small FIS values, indicating that there are not considerably more inbred animals than expected.FST value of the Spanish Cordoba and Hungarian Merino breed was the highest (FST = 0.184), ruling out the Cordoba breed as an origin breed of Hungarian Merino. The phylogenetic study placed Estremadura Merino the fourth closest to Hungarian Merino, while the Rambouillet breed remained the closest  was 19 ; howeverNetwork analysis of the individuals revealed  a spokedConcerning daily weight gain, we have found five highly correlated loci  using 13This study gives insight into the overall genetic state of the Hungarian Merino in contrast to other breeds. The results might be useful for other research conducted on a wider set of Merino breeds.The within-breed network analysis of the animals based on their genetic similarity and its visualisation could provide an intuitive tool to the breeders of any species to easily have a glimpse of the genetic structure of the population and the most important animals. Later on, when more within-breed structures could be analysed and compared, we could dissect different breeding practices among different breeders or countries, giving further insight into the influence of the breeding policy on the structure of a given breed.Some of the loci associated with a standardized daily weight gain were very close to the markers presented in other studies on 2.4 and 3.4 higher number of animals . Applica