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Forensic Science International Nov 2017Molecular analyses in a post-mortem setting are becoming increasingly common, particularly in cases of sudden unexplained death, with the aim of identifying genetic... (Review)
Review
Molecular analyses in a post-mortem setting are becoming increasingly common, particularly in cases of sudden unexplained death, with the aim of identifying genetic mutations which may be responsible for causing death. In retrospective investigations, the access to suitable autopsy biological samples may be limited, and often formalin fixed paraffin embedded (FFPE) tissue is the only sample available. The preservation of tissue in formalin is known to damage DNA through crosslinking activity. This results in the extraction of severely fragmented DNA of variable yields, which subsequently reduces the ability to perform downstream molecular analyses. Numerous studies have investigated possible improvements to various aspects of the DNA extraction and amplification procedures from FFPE tissue and this review aims to collate these optimization steps in a cohesive manner. A systematic review was performed of three major databases, which identified 111 articles meeting the inclusion criteria. Five main areas for optimization and improvements were identified in the workflow: (1) tissue type, (2) fixation process, (3) post-fixation, (4) DNA extraction procedure and (5) amplification. It was found that some factors identified, for example tissue type and fixation process, could not be controlled by the researcher when conducting retrospective analyses. For this reason, optimization should be performed in other areas, within the financial means of the laboratories, and in accordance with the purposes of the investigation. Implementation of one or more of the optimization measures described here is anticipated to assist in the extraction of higher quality DNA. Despite the challenges posed by FFPE tissue, it remains a valuable source of DNA in retrospective molecular forensic investigations.
Topics: DNA; DNA Fingerprinting; Fixatives; Forensic Medicine; Formaldehyde; Humans; Paraffin Embedding; Polymerase Chain Reaction; Preservation, Biological
PubMed: 29078160
DOI: 10.1016/j.forsciint.2017.09.020 -
BMC Infectious Diseases May 2020The burden of drug resistant tuberculosis in Africa is largely driven by the emergence and spread of multidrug resistant (MDR) and extensively drug resistant (XDR)...
BACKGROUND
The burden of drug resistant tuberculosis in Africa is largely driven by the emergence and spread of multidrug resistant (MDR) and extensively drug resistant (XDR) Mycobacterium tuberculosis strains. MDR-TB is defined as resistance to isoniazid and rifampicin, while XDR-TB is defined as MDR-TB with added resistance to any of the second line injectable drugs and any fluoroquinolone. The highest burden of drug resistant TB is seen in countries further experiencing an HIV epidemic. The molecular mechanisms of drug resistance as well as the evolution of drug resistant TB strains have been widely studied using various genotyping tools. The study aimed to analyse the drug resistant lineages in circulation and transmission dynamics of these lineages in Africa by describing outbreaks, nosocomial transmission and migration. Viewed as a whole, this can give a better insight into the transmission dynamics of drug resistant TB in Africa.
METHODS
A systematic review was performed on peer reviewed original research extracted from PubMed reporting on the lineages associated with drug resistant TB from African countries, and their association with outbreaks, nosocomial transmission and migration. The search terms "Tuberculosis AND drug resistance AND Africa AND (spoligotyping OR molecular epidemiology OR IS6110 OR MIRU OR DNA fingerprinting OR RFLP OR VNTR OR WGS)" were used to identify relevant articles reporting the molecular epidemiology of drug resistant TB in Africa.
RESULTS
Diverse genotypes are associated with drug resistant TB in Africa, with variations in strain predominance within the continent. Lineage 4 predominates across Africa demonstrating the ability of "modern strains" to adapt and spread easily. Most studies under review reported primary drug resistance as the predominant type of transmission. Drug resistant TB strains are associated with community and nosocomial outbreaks involving MDR- and XDR-TB strains. The under-use of molecular epidemiological tools is of concern, resulting in gaps in knowledge of the transmission dynamics of drug resistant TB on the continent.
CONCLUSIONS
Genetic diversity of M. tuberculosis strains has been demonstrated across Africa implying that diverse genotypes are driving the epidemiology of drug resistant TB across the continent.
Topics: Africa; Antitubercular Agents; Drug Resistance, Multiple, Bacterial; Epidemics; Extensively Drug-Resistant Tuberculosis; Genotype; High-Throughput Nucleotide Sequencing; Humans; Molecular Epidemiology; Mycobacterium tuberculosis; Polymorphism, Restriction Fragment Length
PubMed: 32404119
DOI: 10.1186/s12879-020-05031-5 -
Frontiers in Cellular and Infection... 2021Osseointegration is a well-established concept used in applications including the percutaneous Bone-Anchored Hearing System (BAHS) and auricular rehabilitation. To date,...
Multimodal Analysis of the Tissue Response to a Bone-Anchored Hearing Implant: Presentation of a Two-Year Case Report of a Patient With Recurrent Pain, Inflammation, and Infection, Including a Systematic Literature Review.
Osseointegration is a well-established concept used in applications including the percutaneous Bone-Anchored Hearing System (BAHS) and auricular rehabilitation. To date, few retrieved implants have been described. A systematic review including cases where percutaneous bone-anchored implants inserted in the temporal bone were retrieved and analyzed was performed. We also present the case of a patient who received a BAHS for mixed hearing loss. After the initial surgery, several episodes of soft tissue inflammation accompanied by pain were observed, leading to elective abutment removal 14 months post-surgery. Two years post-implantation, the implant was removed due to pain and subjected to a multiscale and multimodal analysis: microbial DNA using molecular fingerprinting, gene expression using quantitative real-time polymerase chain reaction (qPCR), X-ray microcomputed tomography (micro-CT), histology, histomorphometry, backscattered scanning electron microscopy (BSE-SEM), Raman spectroscopy, and fluorescence hybridization (FISH). Evidence of osseointegration was provided micro-CT, histology, BSE-SEM, and Raman spectroscopy. Polymicrobial colonization in the periabutment area and on the implant, including that with and , was determined using a molecular analysis a 16S-23S rDNA interspace [IS]-region-based profiling method (IS-Pro). The histology suggested bacterial colonization in the skin and in the peri-implant bone. FISH confirmed the localization of and coagulase-negative staphylococci in the skin. Ten articles (54 implants, 47 patients) met the inclusion criteria for the literature search. The analyzed samples were either BAHS (35 implants) or bone-anchored aural epitheses (19 implants) between 2 weeks and 8 years. The main reasons for elective removal were nonuse/changes in treatment, pain, or skin reactions. Most samples were evaluated using histology, demonstrating osseointegration, but with the absence of bone under the implants' proximal flange. Taken together, the literature and this case report show clear evidence of osseointegration, despite prominent complications. Nevertheless, despite implant osseointegration, chronic pain related to the BAHS may be associated with a chronic bacterial infection and raised inflammatory response in the absence of macroscopic signs of infection. It is suggested that a multimodal analysis of peri-implant health provides possibilities for device improvements and to guide diagnostic and therapeutic strategies to alleviate the impact of complications.
Topics: Bone-Anchored Prosthesis; Hearing; Hearing Aids; Humans; In Situ Hybridization, Fluorescence; Inflammation; Pain; Staphylococcus aureus; X-Ray Microtomography
PubMed: 33859952
DOI: 10.3389/fcimb.2021.640899 -
Nutrients Aug 2017The mechanisms behind the efficacy of exclusive enteral nutrition (EEN) in Crohn's disease (CD) remain poorly understood, despite the high rate of treatment response.... (Review)
Review
The mechanisms behind the efficacy of exclusive enteral nutrition (EEN) in Crohn's disease (CD) remain poorly understood, despite the high rate of treatment response. Evidence accumulated in the last 20 years suggests that a positive shift of the disrupted microbiota is one of the treatment effects. The purpose of this study was to critically review and summarize data reporting the microbiological effects of EEN in patients with CD. Fourteen studies were considered in the review, overall involving 216 CD patients on EEN. The studies were heterogeneous in methods of microbiota analysis and exclusion criteria. The most frequently reported effect of EEN was a reduction in microbiota diversity, reversible when patients returned to a normal diet. The effect of EEN on specific bacteria was very variable in the different studies, partially due to methodological limitations of the mentioned studies. The EEN seem to induce some metabolomic changes, which are different in long-term responder patients compared to patients that relapse earlier. Bacterial changes can be relevant to explaining the efficacy of EEN; however, microbiological data obtained from rigorously performed studies and derived from last generation techniques are largely inconsistent.
Topics: Bacteria; Crohn Disease; Enteral Nutrition; Gastrointestinal Microbiome; Humans; Intestines; Ribotyping; Treatment Outcome
PubMed: 28777338
DOI: 10.3390/nu9080832 -
Acta Dermato-venereologica Jan 2019The immune mechanisms involved in atopic dermatitis (AD) are complex and little is known about the possible role of the gut microbiota in the aetiopathogenesis of AD. A...
The immune mechanisms involved in atopic dermatitis (AD) are complex and little is known about the possible role of the gut microbiota in the aetiopathogenesis of AD. A systematic review of the literature was performed according to PRISMA guidelines, and included 44 of 2,199 studies (26 observational and 18 interventional studies). Detection of gut microbiota was performed by either 16s rRNA PCR, or by culture. Observational studies were diverse regarding the age of study participants and the bacterial species investigated. Overall, the results were conflicting with regard to diversity of the gut microbiota, specific bacterial colonization, and subsequent risk of AD. Nearly half of the included interventional studies showed that an altered gut microbial colonization due to use of probiotics had a positive effect on the severity of AD. The remaining studies did not show an effect of probiotics on the severity of AD despite an alteration in the gut microbial composition. The role of the gut microbiome for the onset and severity of pre-existing AD remains controversial.
Topics: Bacteria; Dermatitis, Atopic; Gastrointestinal Microbiome; Gastrointestinal Tract; Host-Pathogen Interactions; Humans; Probiotics; Ribotyping; Risk Factors; Severity of Illness Index
PubMed: 30085318
DOI: 10.2340/00015555-3008 -
Journal of Plastic, Reconstructive &... Jan 2019We present a case of skin allograft survival in a patient who previously received a bone marrow transplant from the same HLA-matched donor. DNA fingerprinting of skin...
BACKGROUND
We present a case of skin allograft survival in a patient who previously received a bone marrow transplant from the same HLA-matched donor. DNA fingerprinting of skin biopsies showed mixed cellularity originating from the donor and recipient (68% and 32% donor DNA in the allograft skin and the native recipient's skin, respectively). Histologic sections demonstrated both grade 3/4 rejection and graft-versus-host-disease. We have conducted a systematic review in search for other cases of donor skin allograft survival after a bone marrow or hematopoietic stem cell transplantation.
METHODS
All reported cases in English, Spanish, French, and German were captured using the electronic databases. Bibliographies of relevant articles were manually searched.
RESULTS
Nineteen patients (12 females) who received skin allografts from their bone marrow or hematopoietic stem cell donors were identified. Average age was 27.2 years (range: 5 months to 64 years). Skin allografts were used to treat graft-versus-host-disease, Herlitz junctional epidermolysis bullosa, and to test tolerance before a kidney transplantation from the same donor. Eight cases were not receiving immunosuppressive therapy. Allografts survived in all patients. In three patients, skin punch biopsies were taken, and these biopsies demonstrated mixed donor and recipient cellularity. The pathology result is specified in two more cases, with no signs of rejection.
CONCLUSIONS
The same donor skin allografts may be a safe option to treat severe cutaneous conditions in recipients of a bone marrow/hematopoietic stem cell transplantation. However, future studies are needed to confirm these results.
Topics: Adolescent; Adult; Allografts; Bone Marrow Transplantation; Child; Child, Preschool; Escherichia coli Infections; Fasciitis, Necrotizing; Fatal Outcome; Female; Graft Survival; Graft vs Host Disease; Humans; Infant; Living Donors; Male; Middle Aged; Skin Transplantation; Transplant Donor Site; Transplantation, Homologous; Wound Healing; Young Adult
PubMed: 29983364
DOI: 10.1016/j.bjps.2018.05.018 -
Anaerobe Aug 2019Clostridioides difficile is considered one of the main etiological agents of bacterial diarrhea associated with the use of antibiotics. It is an important nosocomial...
Clostridioides difficile is considered one of the main etiological agents of bacterial diarrhea associated with the use of antibiotics. It is an important nosocomial pathogen and the main cause of morbidity and mortality. In recent years, infections associated with C. difficile have led to numerous investigations. It is well known that C. difficile associated diarrhea (CDAD) is favored by the suppression or imbalance of the intestinal microbiome during or after antibiotic therapy. Other risk factors are, for instance, advanced age, long periods of hospitalization, chemotherapy, and other gastrointestinal infections. In the 2000's, the number of CDAD cases largely increased due to the emergence of the epidemic clone named BI/NAP1 ribotype 027, responsible for causing several outbreaks in developed countries, such as Canada, the United States, and the United Kingdom. The presence of the epidemic clone has been reported in Asia, Latin America and Australia, however, infections associated with C. difficile (CDI) in these geographic regions are usually caused by other ribotypes. In Brazil, for instance, epidemiological data on the incidence of CDI are still limited, especially regarding the spread of C. difficile within hospital units, the spectrum of toxigenic genes and the antimicrobial resistance profile. Some studies have demonstrated the importance of notifying cases related to CDI and taking special care measures in order to minimize the spread of epidemic strains in Brazil. Finally, epidemiological analysis of the prevalent and/or exclusive ribotypes circulating in Brazil can contribute to understand and to correlate characteristics associated with the biology of this pathogen with other globally circulating ribotypes. This review aimed to summarize all published work related to the isolation of C. difficile from human patients in Brazil, being the main focus, the methodologies used for identification of prevalent ribotypes, the antimicrobial susceptibility profile, and the diseases associated with the acquisition of CDI.
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Anti-Bacterial Agents; Brazil; Child; Child, Preschool; Clostridioides difficile; Clostridium Infections; Diarrhea; Female; Hospitalization; Humans; Incidence; Infant; Male; Middle Aged; Ribotyping; Risk Factors; Young Adult
PubMed: 30851427
DOI: 10.1016/j.anaerobe.2019.03.002 -
Anaerobe Apr 2020Clostridioides difficile typing is invaluable for the investigation of both institution-specific outbreaks as well as national surveillance. While the epidemic ribotype...
Clostridioides difficile typing is invaluable for the investigation of both institution-specific outbreaks as well as national surveillance. While the epidemic ribotype 027 (RT027) has received a significant amount of resources and attention, ribotype 106 (RT106) has become more prevalent throughout the past decade. The purpose of this systematic review was to comprehensively summarize the genetic determinants, antimicrobial susceptibility, epidemiology, and clinical outcomes of infection caused by RT106. A total of 68 articles published between 1999 and 2019 were identified as relevant to this review. Although initially identified in the United Kingdom in 1999, RT106 is now found worldwide and became the most prevalent strain in the United States in 2016. Current data indicate that RT106 harbors the tcdA and tcdB genes, lacks binary toxin genes, and does not contain any deletions in the tcdC gene, which differentiates it from other epidemic strains, including ribotypes 027 and 078. Interestingly, RT106 produces more spores than other strains, including RT027. Overall, RT106 is highly resistant to erythromycin, clindamycin, fluoroquinolones, and third-generation cephalosporins. However, the MIC in most studies are one to two fold dilutions below the epidemiologic cut-off values of metronidazole and vancomycin, suggesting both are acceptable treatment options from an in vitro perspective. The few clinical outcomes studies available concluded that RT106 causes less severe disease than RT027, but patients were significantly more likely to experience multiple CDI relapses when infected with a RT106 strain. Specific areas warranting future study include potential survival advantages provided by genetic elements as well as a more robust investigation of clinical outcomes associated with RT106.
Topics: Anti-Bacterial Agents; Clostridioides difficile; Clostridium Infections; Genes, Bacterial; Humans; Microbial Sensitivity Tests; Public Health Surveillance; Ribotyping; Spores, Bacterial; Virulence
PubMed: 32007682
DOI: 10.1016/j.anaerobe.2019.102142