Dissecting the microbial community structure of internal organs during the early postmortem period in a murine corpse model.

BACKGROUND: Microorganisms distribute and proliferate both inside and outside the body, which are the main mediators of decomposition after death. However, limited information is available on the postmortem microbiota changes of extraintestinal body sites in the early decomposition stage of mammalian corpses. RESULTS: This study investigated microbial composition variations among different organs and the relationship between microbial communities and time since death over 1 day of decomposition in male C57BL/6 J mice by 16S rRNA sequencing. During 1 day of decomposition, Agrobacterium, Prevotella, Bacillus, and Turicibacter were regarded as time-relevant genera in internal organs at different timepoints. Pathways associated with lipid, amino acid, carbohydrate and terpenoid and polyketide metabolism were significantly enriched at 8 h than that at 0.5 or 4 h. The microbiome compositions and postmortem metabolic pathways differed by time since death, and more importantly, these alterations were organ specific. CONCLUSION: The dominant microbes differed by organ, while they tended toward similarity as decomposition progressed. The observed thanatomicrobiome variation by body site provides new knowledge into decomposition ecology and forensic microbiology. Additionally, the microbes detected at 0.5 h in internal organs may inform a new direction for organ transplantation.

Analysis of Postmortem Intestinal Microbiota Successional Patterns with Application in Postmortem Interval Estimation.

Microorganisms play a vital role in the decomposition of vertebrate remains in natural nutrient cycling, and the postmortem microbial succession patterns during decomposition remain unclear. The present study used hierarchical clustering based on Manhattan distances to analyze the similarities and differences among postmortem intestinal microbial succession patterns based on microbial 16S rDNA sequences in a mouse decomposition model. Based on the similarity, seven different classes of succession patterns were obtained. Generally, the normal intestinal flora in the cecum was gradually decreased with changes in the living conditions after death, while some facultative anaerobes and obligate anaerobes grew and multiplied upon oxygen consumption. Furthermore, a random forest regression model was developed to predict the postmortem interval based on the microbial succession trend dataset. The model demonstrated a mean absolute error of 20.01 h and a squared correlation coefficient of 0.95 during 15-day decomposition. Lactobacillus, Dubosiella, Enterococcus, and the Lachnospiraceae NK4A136 group were considered significant biomarkers for this model according to the ranked list. The present study explored microbial succession patterns in terms of relative abundances and variety, aiding in the prediction of postmortem intervals and offering some information on microbial behaviors in decomposition ecology.

Evaluating the effects of causes of death on postmortem interval estimation by ATR-FTIR spectroscopy.

Estimating postmortem interval (PMI) is one of the most challenging tasks in forensic practice due to the effects of many factors. Here, attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy combined with chemometrics was utilized to evaluate the effects of causes of death when estimating PMI and to establish a partial least square (PLS) regression model, which can precisely predict PMI under different causes of death. First, the sensitivities to causes of death (brainstem injury, mechanical asphyxia, and hemorrhage shock) of seven kinds of organs were evaluated based on their degrees of cohesion and separation. Then, the liver was selected as the most sensitive organ to establish a PMI estimation model to compare the predicted deviations from different causes of death. It turns out that the cause of death has no significant effect on estimating PMI. Next, a PLS regression model was built with kidney tissues, which have the lowest sensitivity, and this model showed a satisfactory predictive ability and wide applicability. This study demonstrates the feasibility of using ATR-FTIR spectroscopy in conjunction with chemometrics as a powerful alternative for detecting changes in biochemistry and estimating PMI. A new perspective was also provided for evaluating the effect of causes of death when predicting PMI.

Atrial dysplasia in the atria of humans without cardiovascular disease.

Research on atrial histology of humans without cardiovascular disease is scarce. Therefore, our aim was to study human atrial histology in subjects without cardiovascular disease. Histology of the right atrium, left atrium or atrial septum was studied in eight patients (one newborn infant and seven adults) who died of a non-cardiac cause and who were not known to suffer from any cardiovascular pathology. Staining with hematoxylin phloxine saffron or Masson's trichrome was performed to have a better identification of fibrosis and H&E for better identification of lymphocytes. Atrial histology was compared with the histology of the left ventricle and was taken from a collection of standard glass slides. Common light microscopic examination and numeric image processing was performed in all samples. Left atrial histology showed a substantial amount of adipocytes and interstitial fibrosis, associated with replacement fibrosis in some of these cases including one case of lymphocytic infiltrates, similar to the histologic changes of the right ventricle (RV) known in patients with arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVD). Furthermore, we identified a perpendicular orientation of atrial myocardial fibres, which is also a feature of the thin RV free wall. A similar histologic substrate to the RV myocardium known in ARVD is found in the atria of humans without an overt cardiovascular pathology. This may explain the high prevalence of atrial fibrillation in the general population.

Right atrial pathology in arrhythmogenic right ventricular dysplasia.

BACKGROUND: Atrial fibrillation (AF) is the most common atrial arrhythmia in arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVD). Considering the histologic changes known in the right ventricular (RV) in ARVD, the aim of the present study was to examine right atrial (RA) pathology in patients with ARVD. METHODS: Histology of RA and RV was assessed from autopsy material in 3 patients with ARVD without persistent atrial arrhythmia. RA histology in 3 patients with permanent AF without ARVD and 5 patients without cardiovascular disease was also studied. Staining with hematoxylin phloxine saffron was performed for the ARVD patients to identify fibrosis, and hematoxylin-eosin for identification of lymphocytes. Masson's trichrome staining was performed for control groups taken from a collection of standard glass slides. RESULTS: In all 3 ARVD cases, RA anomalies were observed that revealed a reduction of cardiomyocytes, the presence of adipocytes, some of them inside the mediomural atrial layer and interstitial fibrosis. In 2 ARVD cases, interstitial fibrosis was also associated with a focus of replacement fibrosis, which was also observed in patients with permanent AF without ARVD. The histologic specimen of the RA and RV from the control group without cardiovascular disease did not display any evidence of fat or fibrosis with a preserved cardiomyocyte architecture. CONCLUSIONS: A similar histopathological substrate, as can be observed in the RV of patients with ARVD can also be seen in the RA of these patients. This may explain the high prevalence of atrial arrhythmias, particularly AF, in patients with ARVD.

Human and non-human bone identification using FTIR spectroscopy.

Human and non-human identification of unknown skeletal remains is of great importance in forensic and anthropologic contexts. However, the traditional morphological methods for bone species identification are subjective or time-consuming. Here, we utilized Fourier transform infrared (FTIR) spectroscopy and chemometric methods to determinate the spectral variances between human and non-human (i.e., pig, goat, and cow) bones. To simulate real forensic situations as much as possible, fresh, boiled, and decomposed bones were included in this study. Principal component analysis (PCA) results illustrated pig bones were more sensitive to the environmental and external factors than other species studied in this work. Thus, pig bone might not be a suitable proxy for human bone in the study of postmortem changes. More importantly, score plots of PCA results showed clear separation with a slight overlap between the human and non-human fresh bones, but it failed to distinguish the boiled and decomposed bones. Then, partial least squares discriminant analysis (PLS-DA) was employed, and both internal and external validations were conducted to assess its classification ability, which resulted in 99.72 and 99.53% accuracy, respectively. According to the loading plots of PCA and PLS-DA, the spectral diversity was mainly due to the inorganic portion (i.e., carbonates and phosphates), which can remain relatively stable under various conditions. As such, our results illustrate that FTIR spectroscopy could serve as a reliable tool to assist in bone species determination and also has great potential in real forensic cases with natural conditions.

Species identification of bloodstains by ATR-FTIR spectroscopy: the effects of bloodstain age and the deposition environment.

In this study, we investigated the potential of attenuated total reflection (ATR) Fourier transform infrared (FTIR) spectroscopy combined with advanced chemometrics for species identification of bloodstains similar to evidence obtained from real crime scenes. Two partial least squares-discriminant analysis classification models (a human-mammal-domestic fowl trilateral model and a species-specific model) were established. The models demonstrated complete separation among the three classes (human, mammal, and domestic fowl) and distinguished six species (human, rat, rabbit, dog, chicken, and duck). Validation was subsequently conducted to evaluate the robustness of these two models, which resulted in 100 and 94.2% accuracy; even human bloodstains placed in an outdoor environment for up to 107 days were successfully identified. Additionally, all bloodstains were positively identified as blood using the squared Euclidean cosine method by comparing the spectra with those of non-blood substances that had a similar appearance or easily produced false positives. These results demonstrate that ATR-FTIR spectroscopy combined with chemometrics can be a powerful tool for species identification of bloodstains.

Estimation of the late postmortem interval using FTIR spectroscopy and chemometrics in human skeletal remains.

Due to a lack of reliable and accurate methods, determining the postmortem interval (PMI) of human skeletal remains is one of the most important and challenging tasks in forensic medicine. In this paper, we studied the changes to bone chemistry with increasing PMI in two different experimental conditions using Fourier transform infrared (FTIR) spectroscopy in conjunction with chemometrics methods Paired bone samples collected from 56 human corpses were buried (placed in soil) and unburied (exposed to the air) for intervals between 76 and 552 days. The results of principle component analysis (PCA) showed the chemical differences of these two cases had a significant influence on the rate of decomposition of the remains. Meanwhile, satisfactory predictions were performed by the genetic algorithm combined with partial least-squares (GA-PLS) with the root mean square errors of prediction (RMSEP) of 50.93days for buried bones and 71.03days for unburied bones. Moreover, the amide I region of proteins and the area around 1390cm-1, which is associated with fatty acids, were identified with regular changes by GA-PLS and played an important role in estimating PMI. This study illustrates the feasibility of utilizing FTIR spectroscopy and chemometrics as an attractive alternative for estimating PMI of human remains and the great potential of these techniques in real forensic cases with natural conditions.

Aortic Dissection and Sudden Unexpected Deaths: A Retrospective Study of 31 Forensic Autopsy Cases.

Acute aortic dissection (AAD) is the most common cause of sudden unexpected death related to aortic diseases. A retrospective study of 31 sudden unexpected deaths caused by AAD was conducted at Xi'an Jiaotong University Forensic Center from 2001 to 2012. We summarized the forensic characteristics of AAD and assessed the clinically diagnostic accuracy of AAD. The characteristics of sudden unexpected death due to AAD were male predominant (male: female=6.7:1), relatively young with the mean age of 44, and predominance of type A dissection (77.4%). Cardiac tamponade was the most frequent cause of sudden death (87.1%). Of the 31 cases, 26 (83.9%) patients were not recognized clinically and were misdiagnosed with acute myocardial infarction, coronary artery disease, cholecystitis, acute gastroenteritis, renal/urinary lithiasis, or acute pancreatitis. In summary, AAD can be difficult to recognize, diagnosis is therefore sometimes delayed or missed. The medicolegal death investigation can help physicians have a better understanding of AAD.

Genetic diversity and haplotype structure of 24 Y-chromosomal STR in Chinese Hui ethnic group and its genetic relationships with other populations.

In the present study, 24 Y-chromosomal short tandem repeat (Y-STR) loci were analyzed in 115 unrelated Hui male individuals from Haiyuan county or Tongxin county, Ningxia Hui Autonomous Region, China, to evaluate the forensic application of the 24 STR loci and to analyze interpopulation differentiations by making comparisons between the Hui group data and previously published data of other 13 populations. A total of 115 different haplotypes were observed on these 24 Y-STR loci. The gene diversities ranged from 0.4049 (DYS437) to 0.9729 (DYS385a, b). The overall haplotype diversity was 1 at AGCU 24 Y-STR loci level, while the values were reduced to 0.999237, 0.996949, and 0.996644 at the Y-filer 17 loci, 11 Y-STR loci of extended haplotype and 9 Y-STR loci of minimal haplotype levels, respectively; whereas, haplotype diversity for additional 7 loci (not included in Y-filer 17 loci) was 0.995271. The pairwise FST , multidimensional scaling plot and neighbor-joining tree indicated the Hui group had the closest genetic relationship with Sala in the paternal lineage in the present study. In summary, the results in our study indicated the 24 Y-STRs had a high level of polymorphism in Hui group and hence could be a powerful tool for forensic application and population genetic study.

Polymorphic distribution and forensic effectiveness study of eight miniSTR in Chinese Uyghur ethnic group.

We obtained the allelic frequencies and forensic efficiency data for eight mini short tandem repeat loci including Penta E, D12S391, D6S1043, D2S1338, D19S433, CSF1PO, Penta D and D19S253 loci from a sample of 128 unrelated Uyghur individuals from China. The amplification products of the eight STR loci are <240 bp in size. A total of 94 alleles were observed and the corresponding allelic frequencies ranged from 0.0039 to 0.3438 in the present study. Observed genotype distributions for each locus do not show deviations from Hardy-Weinberg equilibrium expectations. The combined power of discrimination, combined power of exclusion and combined matching probability of the eight STR loci equaled to 0.999999999963373, 0.9997770 and 3.6627 × 10(-11), respectively. Because of the small fragment length of PCR products and the high degree of polymorphisms, the eight STR loci are highly beneficial for the forensic analysis of degraded DNA samples which are commonly observed in forensic cases. The STR data of the Uyghur group were compared with the previously published population STR data of other groups from different ethnic or areas, and significant differences were observed among these groups at some loci.

Allelic polymorphic investigation of 21 autosomal short tandem repeat loci in a Chinese Bai ethnic group.

Population genetic data of 21 autosomal short tandem repeats (STRs) were obtained in a sample of 106 unrelated healthy individuals of Bai ethnic minority born in the Dali Bai Autonomous Prefecture in Yunnan Province. We observed 138 alleles with corresponding allelic frequencies ranging from 0.005 to 0.575. The genotypic frequency distributions at those STR loci were consistent with Hardy-Weinberg equilibrium (Bonferroni's correction was used for Hardy-Weinberg equilibrium tests). The combined probability of exclusion, power of discrimination, probability of matching value for all 21 STR loci were 0.9999975729, 0.999999999999999999872 and 1.28×10(-19), respectively. The population data in this study showed significant differences from the previously published population data of Tibetan and Salar groups in some loci.

The distribution of genetic diversity of KIR genes in the Chinese Mongolian population.

Killer cell immunoglobulin-like receptors (KIRs) are expressed on natural killer cells and as such regulate their response against infection and malignancy. KIR genes are variable in gene content and type, which results in different KIR haplotypes, and can be used to discriminate individuals and populations from different regions or ethnic groups. In the present study, we represent the first report on the KIR gene frequency and content diversities of 14 KIR genes (KIR2DL1, 2DL2, 2DL3, 2DL4, 2DL5, 2DS1, 2DS2, 2DS3, 2DS4, 2DS5, 3DL1, 3DL2, 3DL3, 3DS1) and 2 pseudogenes (KIR3DP1 and 2DP1) in the Chinese Mongolian population. The 16 detected KIR genes were all observed. All the individuals were typed positive for the four framework genes KIR3DL3, 3DL2, 2DL4 and the pseudogene KIR3DP1, as well as for the pseudogene KIR2DP1. The observed carrier gene frequencies (OF) of the other KIR genes ranged from 16% at the KIR2DL2 locus to 93% at the KIR3DL1 locus. Over all, 48 different gene profiles were found in the study population and the most commonly observed KIR gene profile with a frequency of 14% consisted of KIR2DL4, 3DL2, 3DL3, 2DP1, 3DP1, 2DL1, 2DL3 and 3DL1 which belongs to the AA genotype. Principal component analysis (PCA) and the dendrogram illustrated the genetic distances between our study population and previously published populations from other ethnic groups or regions. The results of the present study show that the KIR gene family is highly polymorphic and can be a valuable tool for enriching the Chinese ethnical gene information resources, for anthropological studies, as well as for KIR gene related disease research.

Diversity distributions of killer cell immunoglobulin-like receptor genes and their ligands in the Chinese Shaanxi Han population.

In the present study, 17 killer cell immunoglobulin-like receptors (KIR) genes and KIR ligands (human leukocyte antigen [HLA] -A and -B) were detected by using a polymerase chain reaction-sequence-specific primer (PCR-SSP) method in 104 unrelated healthy Han individuals living in Shaanxi province, China. The observed carrier frequencies of the 12 KIR genes ranged from 0.14 to 0.96. KIR2DL4, 3DL2, 3DL3, 2DP1 and 3DP1 were found to be present in every individual. A total of 51 different KIR gene profiles were identified, in which 11 gene profiles exclusively belonged to the study population. Neighbor-joining phylogenetic tree between the studing population and its neighboring ethnic groups was constructed using the observed carrier frequencies of 13 KIR loci. The phylogenetic tree shows that the Shaanxi Han population, Han populations in different regions, Yi, Japanese, and Koreans were in the same cluster. KIR/HLA relationships show that KIR3DS1(-)/3DL1(+)/Bw4(+) was the most common association in the population. In conclusion, the present study findings reveal the high polymorphism of KIRs in the Shaanxi Han population, demonstrate the KIR/HLA association in the study population, and enrich the KIR and HLA gene resources. The obtained KIR data will further the understanding of genetic relationships among populations in different geographic areas, and assist in answering questions regarding KIR/HLA relationships.

[Pathological characteristics of sudden death caused by coronary thrombosis].

OBJECTIVE: To explore medico-legal characteristics of sudden death caused by coronary heart disease combined with coronary thrombosis. METHODS: Ninety-six cases of sudden death caused by coronary heart disease were collected and divided into two groups: thrombus positive and thrombus negative groups. The time onset, induction and pathological features of coronary artery disease were analyzed. RESULTS: Two groups showed man-dominant population. There were no statistical significant differences in season, circadian rhythm and induction factor. The thrombus positive group (age < 40) showed a higher disease incidence. Heart weight and degree of coronary stenosis were lower in thrombus positive group. However, there was no statistical difference in the number of atherosclerotic coronary arteries (> or = 2), the length of coronary lesions and myocardial infarct. But thrombosis positive group showed lower tendency. CONCLUSION: Two groups are man-dominant population and similar induction factor, lesion position, mechanism of death. But thrombus positive group appeared more in a younger population and the degree of coronary stenosis is milder than thrombus negative group. Forensic pathologists should pay more attention to these characteristics in death investigation.

Genetic polymorphism analysis of 15 STR loci in Chinese Hui ethnic group residing in Qinghai province of China.

In the present study, we investigated the diversity distributions of allelic frequencies of 15 short tandem repeats (STRs) loci in a sample of Chinese Hui ethnic group in the Ningxia Hui Autonomous Region. The allelic frequencies of the 15 STR loci (D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, vWA, TPOX, D18S51, D5S818 and FGA) were obtained from 2975 unrelated healthy Hui individuals. The STR genotyping data of all the samples were generated by DNA extraction, multiple amplification, GeneScan and genotype analysis. The genetic distances among different populations were calculated by using Nei's method and a phylogenetic tree was constructed based on the allelic frequencies of the same 15 STR loci using the neighbor-joining method. A total of 185 alleles were observed in the Hui population, with the corresponding allelic frequencies ranging from 0.0002 to 0.5322. Chi-Square tests showed that all STR loci were in Hardy-Weinberg equilibrium. The forensic statistical parameters of all the loci showed high values. The population data in this study were compared with the previously published population data from other ethnics or areas. The Hui population showed significant differences from the Minnan Han, Uigur, Ewenki, Yi, Tibetan, Maonan and Malay ethnic minority groups in some loci, and from the South Morocco population and the Moroccan population in all the loci. Our results are valuable for human individual identification and paternity testing in the Chinese Hui population and are expected to enrich the genetic information resources of Chinese populations.

Killer cell immunoglobulin-like receptor gene diversity in the Tibetan ethnic minority group of China.

The aim of this study was to analyze killer immunoglobulin-like receptor (KIR) gene polymorphisms in the Tibetan ethnic minority of China. To that purpose, we have studied KIR gene frequencies and genotype diversities of 16 KIR genes and three pseudogenes (2DL1, 2DL2, 2DL3, 2DL4, 2DL5A, 2DL5B, 2DS1, 2DS2, 2DS3, 2DS4*001/002, 2DS4*003-007, 2DS5, 3DL1, 3DL2, 3DL3, 3DS1, 2DP1, 3DP1*001/002/004, and 3DP1*003) in a population sample of 102 unrelated healthy individuals of the Tibetan population living in Lhasa city, Tibet Autonomous Region of China. Tibetans mainly live in "the roof of the world," the Qinghai-Tibet Plateau of China and surrounding areas stretching from central Asia in the North and West to Myanmar and mainland China in the East, and India, Nepal, and Bhutan to the south. KIR gene frequencies and statistical parameters of Tibetan ethnic minority were calculated. Fifteen KIR genes were observed in the 102 tested Tibetan individuals with different frequencies. The allelic frequencies of the 15 KIR genes ranged from 0.06 to 0.86. In addition, KIR 2DL1, 2DL4, 3DL2, and 3DL3 were found to be present in every individual. Variable gene content, together with allelic polymorphisms, can result in individualized human KIR genotypes and haplotypes, with the A haplotypes being predominantly observed. The results of tested linkage disequilibrium (LD) among KIR genes demonstrated that KIR genes present a wide range of linkage disequilibrium. Moreover, a comparison of the population data of our study with previously published population data of other ethnic groups or areas was performed. The differences of allelic frequency distribution in KIR2DL2, 2DL3, 2DL5, 3DL1, 2DS1, 2DS2, 2DS3, 3DS1, and 2DP1 were statistically significant among different populations using the statistical method of the standard χ(2) test. In conclusion, the results of the present study can be valuable for enriching the Chinese ethnical gene information resources of the KIR gene pool and for anthological studies, as well as for KIR-related disease research.

Population genetic analysis of 15 autosomal STR loci in the Russian population of northeastern Inner-Mongolia, China.

In the present study, we investigated the genetic polymorphisms of 15 autosomal STR loci in the Russian population of northeastern Inner-Mongolia, China as well as its genetic relationships with other populations. DNA typing for 15 autosomal STR loci was performed on 148 randomly selected healthy individuals from the Russian population living in Eerguna, northeastern Inner-Mongolia. Allelic frequencies of these loci were calculated by direct counting. The genotype data of this Russian population was moreover compared to other populations using neighbor-joining method, as such constructing a phylogenic tree. A total of 143 alleles were found in the Russian population with corresponding allele frequencies in the range from 0.0034 to 0.5372. Among all the 15 loci, D18S51 had the highest polymorphism (PIC = 0.8632), whereas TPOX had the lowest (PIC = 0.5179). In the phylogenic tree, this Russian population has a close relationship with the populations of South Siberia and northeastern Asia. This study may increase our understanding of the genetic background of the Russian population in Eerguna, China as such providing useful information for anthropological research, forensic sciences as well as disease-association studies.