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1.
Biosensors (Basel) ; 11(5)2021 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-34062874

RESUMO

Molecular diagnostics has been the front runner in the world's response to the COVID-19 pandemic. Particularly, reverse transcriptase-polymerase chain reaction (RT-PCR) and the quantitative variant (qRT-PCR) have been the gold standard for COVID-19 diagnosis. However, faster antigen tests and other point-of-care (POC) devices have also played a significant role in containing the spread of SARS-CoV-2 by facilitating mass screening and delivering results in less time. Thus, despite the higher sensitivity and specificity of the RT-PCR assays, the impact of POC tests cannot be ignored. As a consequence, there has been an increased interest in the development of miniaturized, high-throughput, and automated PCR systems, many of which can be used at point-of-care. This review summarizes the recent advances in the development of miniaturized PCR systems with an emphasis on COVID-19 detection. The distinct features of digital PCR and electrochemical PCR are detailed along with the challenges. The potential of CRISPR/Cas technology for POC diagnostics is also highlighted. Commercial RT-PCR POC systems approved by various agencies for COVID-19 detection are discussed.


Assuntos
Teste de Ácido Nucleico para COVID-19/instrumentação , COVID-19/diagnóstico , Testes Imediatos , Reação em Cadeia da Polimerase/instrumentação , SARS-CoV-2/isolamento & purificação , Animais , Teste de Ácido Nucleico para COVID-19/métodos , Sistemas CRISPR-Cas , Desenho de Equipamento , Humanos , Reação em Cadeia da Polimerase/métodos , SARS-CoV-2/genética
2.
Georgian Med News ; (313): 146-152, 2021 Apr.
Artigo em Russo | MEDLINE | ID: mdl-34103447

RESUMO

The rapid development of molecular biology and genetic engineering contributes to the creation of plants with desired properties in a short time. One of the aspects of the study of genetically modified organisms (GMO) and genetically modified products (GMP) is the study of their impact on humans, animals and the environment. Subject of research - changes in the morph functional indicators of the reproductive system of mice. The relevance of the chosen topic is due to the importance of the reproductive system for the reproduction of a healthy generation, capable of developing normally and continuing its race. Purpose of the study - to identify the effect of GMOs on the reproductive system of mice. Cultivation of three groups of laboratory mice using transgenic feed (genetically modified soybean meal) and obtaining biological material for research. Determination of the presence of genetically modified sources (GMS), genetically modified organisms (GMO) in food and feed products using the polymerase chain reaction (PCR). Morphometric study of the obtained material and biometric data processing. It has been shown that the consumption of feed by animals prepared on the basis of genetically modified plants does not affect the reproductive functions of the parental generation; but at the same time there was an inhibition of the growth rate and the process of formation of the gonads of the descendants of the first and, especially, the second generation; the second generation of offspring, eating only soybean meal, had defective reproductive qualities and high mortality. Second-generation mice eating genetically modified soybean meal are at greater risk than second-generation mice eating traditional diets.


Assuntos
Alimentos Geneticamente Modificados , Animais , Dieta , Genitália , Camundongos , Plantas Geneticamente Modificadas/genética , Reação em Cadeia da Polimerase
3.
Viruses ; 13(6)2021 05 28.
Artigo em Inglês | MEDLINE | ID: mdl-34071726

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in Wuhan, China, in late 2019 and is the causative agent of the coronavirus disease 2019 (COVID-19) pandemic. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) represents the gold standard for diagnostic assays even if it cannot precisely quantify viral RNA copies. Thus, we decided to compare qRT-PCR with digital polymerase chain reaction (dPCR), which is able to give an accurate number of RNA copies that can be found in a specimen. However, the aforementioned methods are not capable to discriminate if the detected RNA is infectious or not. For this purpose, it is necessary to perform an endpoint titration on cell cultures, which is largely used in the research field and provides a tissue culture infecting dose per mL (TCID50/mL) value. Both research and diagnostics call for a model that allows the comparison between the results obtained employing different analytical methods. The aim of this study is to define a comparison among two qRT-PCR protocols (one with preliminary RNA extraction and purification and an extraction-free qRT-PCR), a dPCR and a titration on cell cultures. The resulting correlations yield a faithful estimation of the total number of RNA copies and of the infectious viral burden from a Ct value obtained with diagnostic routine tests. All these estimations take into consideration methodological errors linked to the qRT-PCR, dPCR and titration assays.


Assuntos
COVID-19/terapia , COVID-19/virologia , Reação em Cadeia da Polimerase/métodos , RNA Viral/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , SARS-CoV-2/isolamento & purificação , Carga Viral/métodos , Animais , Células Cultivadas , Chlorocebus aethiops , Tecnologia Digital/métodos , Humanos , SARS-CoV-2/genética , Células Vero , Cultura de Vírus
4.
J Pak Med Assoc ; 71(4): 1189-1192, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-34125769

RESUMO

OBJECTIVE: To study the different epidemiological and polymerase chain reaction-based identification of vibrio cholera. METHODS: The cross-sectional study was conducted at the Center for Advanced Studies in Vaccinology and Biotechnology, University of Balochistan, Quetta, Pakistan, from January 5 to December 6, 2019, and comprised faecal / rectal swab samples from patients with a history of untreated severe diarrhoea of <12-hour duration. The samples were collected from suspected cholera patients at different hospitals of the province. The isolates were examined and identified on the basis of colony characters on thiosulfate-citrate-bile salts-sucrose agar. Susppected colonies were subjected to gram staining, biochemical tests and polymerase chain reaction-based identification. Data was analysed using SPSS 19. RESULTS: Of the 444 samples, 33(7.43%) were positive for vibrio cholera and 411(92.56%) were negative. The incidence was higher in individuals aged 1-20 years 12(2.7%); males 18(4.05%); Balochs 18(4.05%); lower socioeconomic class 18(4.05%); and illiterates 26(5.85%). The incidence was more in summer 19(4.27%) and spring 8(1.80%) seasons. Polymerase chain reaction was highly effective diagnostic approach, with findings showing clear bands of 588bp of ompW gene. CONCLUSIONS: Surveillance for diarrhoeal disorders is necessary to control future outbreaks of cholera in the region.


Assuntos
Epidemias , Vibrio cholerae , Estudos Transversais , Diarreia/epidemiologia , Surtos de Doenças , Hospitais , Humanos , Masculino , Paquistão/epidemiologia , Reação em Cadeia da Polimerase , Vibrio cholerae/genética
5.
Biomed Environ Sci ; 34(5): 364-371, 2021 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-34059173

RESUMO

Objective: To study the polymorphism in P66 and its human B-cell epitopes of Borrelia burgdorferi strains in China. Methods: Polymerase chain reaction (PCR) and sequencing were used to obtain the P66 sequences of 59 Chinese B. burgdorferi. Then the sequences were analyzed by MEGA 5.10 software and compared with the human B-cell epitope sequences from the Immune Epitope Database (IEDB) based on the reference strain of each genotype. Results: Results showed that genetic and amino acid diversity presented in the 66 kD protein of all 59 Chinese strains, especially in Borrelia garinii ( B.g) and Borrelia afzelii ( B.a) strains. B.g strains were divided into three subclusters and two scattered strains JC1-7 and JC2-2 according to the amino acid sequences of P66. The P66 sequences of 15 Xinjiang strains represented by XI91-12 in the B.g subcluster 1, changed from CAA to TAA codon at 508aa position, resulting in early termination. Bases A and C were inserted at sequence position 1 523 bp of strains FP1, LB20, LB21, and SZ21 in the B.a genotype, which resulted to early termination at position 511 aa. G base was inserted at 438 bp of LIP94-11 strain, which led to early termination at position 172 aa. Conclusion: In P66 of 59 Chinese strains, polymorphisms were widely distributed. More importantly, the P66 amino acid sequences of B.g strains had a certain regional character. One of the characteristics of Xinjiang B.g isolates might be the variation at the 508aa location in 15 Xinjiang B.g strains, which may be related to the strains' pathogenicity in this area.


Assuntos
Proteínas de Bactérias/genética , Borrelia burgdorferi/genética , Epitopos de Linfócito B/genética , Polimorfismo Genético , Porinas/genética , Borrelia burgdorferi/classificação , China , Análise por Conglomerados , Marcadores Genéticos , Genótipo , Humanos , Mutação , Reação em Cadeia da Polimerase
6.
Appl Microbiol Biotechnol ; 105(11): 4761-4773, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34059942

RESUMO

The highly complex raw milk matrix challenges the sample preparation for amplicon-sequencing due to low bacterial counts and high amounts of eukaryotic DNA originating from the cow. In this study, we optimized the extraction of bacterial DNA from raw milk for microbiome analysis and evaluated the impact of cycle numbers in the library-PCR. The selective lysis of eukaryotic cells by proteinase K and digestion of released DNA before bacterial lysis resulted in a high reduction of mostly eukaryotic DNA and increased the proportion of bacterial DNA. Comparative microbiome analysis showed that a combined enzymatic and mechanical lysis procedure using the DNeasy® PowerFood® Microbial Kit with a modified protocol was best suitable to achieve high DNA quantities after library-PCR and broad coverage of detected bacterial biodiversity. Increasing cycle numbers during library-PCR systematically altered results for species and beta-diversity with a tendency to overrepresentation or underrepresentation of particular taxa. To limit PCR bias, high cycle numbers should thus be avoided. An optimized DNA extraction yielding sufficient bacterial DNA and enabling higher PCR efficiency is fundamental for successful library preparation. We suggest that a protocol using ethylenediaminetetraacetic acid (EDTA) to resolve casein micelles, selective lysis of somatic cells, extraction of bacterial DNA with a combination of mechanical and enzymatic lysis, and restriction of PCR cycles for analysis of raw milk microbiomes is optimal even for samples with low bacterial numbers. KEY POINTS: • Sample preparation for high-throughput 16S rRNA gene sequencing of raw milk microbiota. • Reduction of eukaryotic DNA by enzymatic digestion. • Shift of detected microbiome caused by high cycle numbers in library-PCR.


Assuntos
Microbiota , Leite , Animais , Bovinos , DNA Bacteriano/genética , Feminino , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética
7.
Viruses ; 13(5)2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-34064763

RESUMO

Infection with SARS-CoV-2 leading to COVID-19 induces hyperinflammatory and hypercoagulable states, resulting in arterial and venous thromboembolic events. Deep vein thrombosis (DVT) has been well reported in COVID-19 patients. While most DVTs occur in a lower extremity, involvement of the upper extremity is uncommon. In this report, we describe the first reported patient with an upper extremity DVT recurrence secondary to COVID-19 infection.


Assuntos
COVID-19/complicações , COVID-19/diagnóstico , Trombose Venosa/complicações , Trombose Venosa/diagnóstico , Idoso de 80 Anos ou mais , Transtornos da Coagulação Sanguínea/complicações , Humanos , Masculino , Recidiva Local de Neoplasia , Reação em Cadeia da Polimerase , RNA Viral , Recidiva , Fatores de Risco , Extremidade Superior/irrigação sanguínea , Trombose Venosa/terapia
8.
Biosensors (Basel) ; 11(5)2021 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-34069758

RESUMO

A polydimethylsiloxane (PDMS)-based self-priming microfluidic chip with cushion chambers is presented in this study for robust and easy-operation digital polymerase chain reaction (dPCR). The chip has only one inlet and can partition samples autonomously through negative pressure, provided by a de-gassed PDMS layer with a multi-level vertical branching microchannel design. Meanwhile, cushion chambers make the chip capable of very robust use for sample partitioning. Finally, the proposed microfluidic chip showed excellent performance in the absolute quantification of a target gene by performing quantitative detection of a 10-fold serial dilution DNA template. Owing to its characteristics of easy operation, low cost, and high robustness, the proposed dPCR chip is expected to further promote the extensive application of digital PCR, especially in resource-limited settings.


Assuntos
Dispositivos Lab-On-A-Chip , Técnicas Analíticas Microfluídicas , Reação em Cadeia da Polimerase/métodos , Técnicas Biossensoriais , Dimetilpolisiloxanos , Desenho de Equipamento , Microfluídica , Análise de Sequência com Séries de Oligonucleotídeos
11.
Braz J Biol ; 82: e241110, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34133560

RESUMO

Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3α and Pvmsp-3ßgenes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3α and Pvmsp3ß genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp-3α genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3ßgenes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3α and Pvmsp-3ß genes of P. vivax isolates by using PCR/RFLP from District Mardan and showed a remarkable level of genetic diversity in the studied genes of circulating parasites in the study area. The results of this study will contribute in future studies about the genetic structure of parasite and vaccine development against the malaria.


Assuntos
Plasmodium vivax , Proteínas de Protozoários , Variação Genética , Genótipo , Humanos , Paquistão , Plasmodium vivax/genética , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Proteínas de Protozoários/genética
12.
Nat Commun ; 12(1): 3431, 2021 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-34103499

RESUMO

The current COVID-19 pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We demonstrate that despite the large size of the viral RNA genome (~30 kb), infectious full-length cDNA is readily assembled in vitro by a circular polymerase extension reaction (CPER) methodology without the need for technically demanding intermediate steps. Overlapping cDNA fragments are generated from viral RNA and assembled together with a linker fragment containing CMV promoter into a circular full-length viral cDNA in a single reaction. Transfection of the circular cDNA into mammalian cells results in the recovery of infectious SARS-CoV-2 virus that exhibits properties comparable to the parental virus in vitro and in vivo. CPER is also used to generate insect-specific Casuarina virus with ~20 kb genome and the human pathogens Ross River virus (Alphavirus) and Norovirus (Calicivirus), with the latter from a clinical sample. Additionally, reporter and mutant viruses are generated and employed to study virus replication and virus-receptor interactions.


Assuntos
Genética Reversa , SARS-CoV-2/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Chlorocebus aethiops , Culicidae/virologia , Furina/metabolismo , Genoma Viral , Células HEK293 , Humanos , Camundongos , Mutação/genética , Células NIH 3T3 , Reação em Cadeia da Polimerase , Células RAW 264.7 , Receptores Virais/metabolismo , Células Vero , Proteínas Virais/química , Replicação Viral
13.
BMC Infect Dis ; 21(1): 464, 2021 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-34020601

RESUMO

BACKGROUND: Leishmaniasis is one of the most neglected tropical diseases in the world and remains endemic in some underdeveloped regions, including western China. The phylogeny and classification of Chinese Leishmania has not been completely clarified to date, especially within the Leishmania (L.) donovani complex, although phylogenetic analyses based on a series of gene markers have been performed. More analytic methods and data are still needed. Random amplified polymorphic DNA (RAPD) technology can sensitively identify slight intraspecific differences, and it is a powerful tool to seek species-specific markers. This work attempted to identify Chinese Leishmania isolates from diverse geographic regions at the genomic level. Meanwhile, specific markers of the L. donovani complex were also developed by RAPD. METHODS: RAPD was applied to 14 Chinese Leishmania isolates from diverse geographic regions and 3 WHO reference strains. The polymorphic sites of amplification were transformed into a data matrix, based on which genetic similarity was calculated, and a UPGMA dendrogram was constructed to analyse the genetic diversity of these Leishmania isolates. Meanwhile, the specific amplification loci of the L. donovani complex were TA-cloned, sequenced and converted into sequence characterized amplified region (SCAR) markers, which were validated preliminarily in 17 available Leishmania strains in this study and analysed by bioinformatics. RESULTS: The cluster analyses showed that the three Leishmania sp. isolates SC10H2, SD and GL clustered together and apart from others, the strains of the L. donovani complex clearly divided into two clades, and the three isolates Cy, WenChuan and 801 formed a subclade. Three specific SCAR markers of the L. donovani complex, i.e., 1-AD17, 2-A816 and 3-O13, were successfully obtained and validated on 17 available Leishmania strains in this study. Through bioinformatic analyses, Marker 1-AD17 may have more specificity for PCR detection of VL, and Marker 3-O13 has the potential to encode a protein. CONCLUSIONS: The RAPD results verified that the undescribed Leishmania species causing visceral leishmaniasis (VL) in China was a unique clade distinguished from L. donovani and revealed that there was genetic differentiation among Chinese L. donovani. The identification of L. donovani-specific markers may help to provide a foundation for future research attempting to develop new specific diagnostic markers of VL and identify specific gene functions.


Assuntos
Variação Genética , Leishmania donovani/classificação , Leishmania donovani/genética , Leishmaniose Visceral/epidemiologia , Técnica de Amplificação ao Acaso de DNA Polimórfico/métodos , Animais , Sequência de Bases , China/epidemiologia , Análise por Conglomerados , DNA de Protozoário/genética , DNA de Protozoário/isolamento & purificação , Marcadores Genéticos , Humanos , Leishmania donovani/isolamento & purificação , Leishmaniose Visceral/parasitologia , Filogenia , Reação em Cadeia da Polimerase , Especificidade da Espécie
14.
J Med Microbiol ; 70(5)2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-34038341

RESUMO

Pleural infections cause major morbidity and mortality, particularly amongst paediatric and elderly populations. The aetiology is broad, but pleural culture fails to yield a causative pathogen in approximately 40 % of cases. Alternative pathogen identification methods are therefore required. The aim of the study was to investigate the yield from and impact on patient care when performing 16S rRNA PCR on culture-negative pleural fluid specimens and to determine whether any individual laboratory parameters were associated with a positive 16S rRNA PCR result. We conducted a study on 90 patients with suspected pleural infection, who had a culture-negative pleural fluid specimen, which underwent 16S rRNA PCR analysis between August 2017 and June 2019. This study was undertaken at a large NHS Trust in London, UK. Thirty-one per cent of culture-negative pleural fluid specimens tested by 16S rRNA PCR yielded a positive PCR result. Our data demonstrated that 16S rRNA PCR detected a significantly higher proportion of Streptococcus pneumoniae (P<0.0001) and fastidious, slow-growing and anaerobic pathogens (P=0.0025) compared with culture-based methods. Of the 25 16S rRNA PCR results that were positive for a causative pathogen, 76 % had a direct impact on clinical management. No single laboratory variable was found to be associated with a positive 16S rRNA PCR result. The findings from our real-world evaluation highlight the importance of 16S rRNA PCR in confirming pleural infection when the aetiology is unknown, and its direct, positive impact on clinical management.


Assuntos
Pleura/microbiologia , Doenças Pleurais , Infecções Pneumocócicas , Streptococcus pneumoniae/isolamento & purificação , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Doenças Pleurais/diagnóstico , Doenças Pleurais/microbiologia , Infecções Pneumocócicas/diagnóstico , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/isolamento & purificação , Estudos Retrospectivos , Reino Unido , Adulto Jovem
15.
J Med Microbiol ; 70(5)2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-34038342

RESUMO

Introduction. Invasive mucormycosis (IM) is a life-threatening infection caused by fungi belonging to the order Mucorales. Histopathology, culture and radiology are the mainstay of diagnosis but lack sensitivity, leading to a delay in timely diagnosis and intervention. Recently, PCR-based approaches have been shown to be a promising method in diagnosing IM.Hypothesis/Gap Statement. Molecular-based approaches may be a valuable adjunct to standard conventional methods for diagnosing IM, especially among culture negatives and patients on antifungal therapy.Aim. In the present study we aimed to evaluate the clinical utility of panfungal and Mucorales-specific PCR for diagnosing IM from various clinical specimens.Methodology. This was a prospective study in which 239 clinically suspected cases of IM attending our tertiary care hospital from August 2015 to March 2018 were enrolled. All the cases were defined as 'proven', 'probable' or 'possible' based on EORTC/MSGERC guidelines. In addition to conventional diagnostics (KOH-calcofluor stain and culture), panfungal and Mucorales-specific PCR assays were also performed. The amplified products were sequenced for species identification. In vitro antifungal susceptibility was performed on all the culture-positive isolates.Results. Among 239 clinically suspected cases of IM, only 140 cases were diagnosed by the demonstration of aseptate ribbon-like hyphae on direct microscopy. Culture was positive in 35.7 % (54/140) of direct microscopy-positive samples. Among the proven cases (n=11), the sensitivity for both Mucorales-specific nested PCR and panfungal PCR was 100 %, but specificity was 91.9 and 73.7% respectively. In probable cases (n=129), the sensitivity of both the PCRs was 98.5 % and specificity for panfungal PCR was 73.7 and 91.9 % for Mucorales-specific PCR.Conclusion. Pan fungal PCR in combination with Mucorales-specific PCR, followed by sequencing, may play a significant role in IM diagnosis especially among those negative for both direct microscopy and culture.


Assuntos
Infecções Fúngicas Invasivas/diagnóstico , Mucorales/isolamento & purificação , Mucormicose/diagnóstico , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , DNA Fúngico/isolamento & purificação , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Estudos Prospectivos , Sensibilidade e Especificidade , Adulto Jovem
16.
Int J Food Microbiol ; 349: 109233, 2021 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-34022616

RESUMO

Escherichia coli O157:H7, a Shiga-producing E. coli is a major pathogenic E. coli strain which since the early 1980s has become a crucial food and water-borne pathogen. Several management strategies can be applied to control the spread of infection; however early diagnosis represents the optimum preventive strategy to minimize the infection. Therefore, it is crucial to detect this pathogen in a fast and efficient manner in order to reduce the morbidity and mortality. Currently used gold standard tests rely on culture and pre-enrichment of E. coli O157:H7 from the contaminated source; they are time consuming and laborious. Molecular methods such as polymerase chain reaction are sensitive; however, they require expensive instrumentation. Therefore, there is a requirement for Accurate, Sensitive, Specific, User friendly, Rapid, Equipment free and Deliverable (ASSURED) detection methods for use in the laboratory and in the field. Emerging technologies such as isothermal amplification methods, biosensors, surface enhanced Raman Spectroscopy, paper-based diagnostics and smartphone-based digital methods are recognized as new approaches in the field of E. coli O157:H7 diagnostics and are discussed in this review. Mobile PCR and CRISPR-Cas diagnostic platforms have been identified as new tools in E. coli O157:H7 POC diagnostics with the potential for implementation by industry. This review describes advances and progress in the field of E. coli O157:H7 diagnosis in the context of food and water industry. The focus is on emerging high throughput point-of-care (POC) E. coli O157:H7 diagnostics and the requirement for the transformation to service routine diagnostics in the food and water industry.


Assuntos
Técnicas Bacteriológicas , Escherichia coli O157/isolamento & purificação , Microbiologia de Alimentos , Sistemas Automatizados de Assistência Junto ao Leito , Microbiologia da Água , Técnicas Biossensoriais , Sistemas CRISPR-Cas , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/prevenção & controle , Humanos , Reação em Cadeia da Polimerase
17.
BMC Infect Dis ; 21(1): 493, 2021 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-34044785

RESUMO

INTRODUCTION: Cronobacter sakazakii is an opportunistic Gram-negative, rod-shaped bacterium which may be a causative agent of meningitis in premature infants and enterocolitis and bacteremia in neonates and adults. While there have been multiple cases of C. sakazakii infections, there have been no acute cholangitis cases reported in humans. CASE PRESENTATION: An 81-year-old male with a past medical history of basal cell carcinoma, alcoholic liver cirrhosis, transjugular intrahepatic portosystemic shunt procedure, complicated by staphylococcus bacteremia, pituitary tumor, glaucoma, and hypothyroidism presented to the emergency room with the complaint of diffuse and generalized 10/10 abdominal pain of 1 day's duration. There was a concern for pancreatitis, acute cholangitis, and possible cholecystitis, and the patient underwent a percutaneous cholecystostomy tube placement. Blood cultures from admission and biliary fluid cultures both grew C. sakazakii. The patient was treated with a carbapenem and clinically improved. CONCLUSIONS: The case study described a patient with multiple medical comorbidities that presented with C. sakazakii bacteremia and cholangitis. While this bacterium has been implicated in other infections, we believe this is the first time the bacteria is being documented to have caused acute cholangitis.


Assuntos
Bacteriemia/diagnóstico , Colangite/diagnóstico , Cronobacter sakazakii/isolamento & purificação , Infecções por Enterobacteriaceae/diagnóstico , Idoso de 80 Anos ou mais , Antibacterianos/uso terapêutico , Bacteriemia/complicações , Bacteriemia/terapia , Carbapenêmicos/uso terapêutico , Colangite/microbiologia , Colangite/terapia , Colecistostomia/métodos , Cronobacter sakazakii/patogenicidade , Drenagem/métodos , Infecções por Enterobacteriaceae/complicações , Infecções por Enterobacteriaceae/terapia , Humanos , Masculino , Infecções Oportunistas/diagnóstico , Infecções Oportunistas/microbiologia , Infecções Oportunistas/terapia , Reação em Cadeia da Polimerase/métodos , Tomografia Computadorizada por Raios X/métodos , Resultado do Tratamento
18.
BMC Infect Dis ; 21(1): 492, 2021 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-34044786

RESUMO

BACKGROUND: Group B streptococcus (GBS) is the leading cause of early-onset neonatal sepsis. However, GBS was infrequently reported in the developing world in contrast to western countries. This study assessed the prevalence of GBS colonization among pregnant women in Jiangsu, East China, and revealed the difference of GBS infection between culture and PCR. METHODS: A total of 16,184 pregnant women at 34 to 37 weeks' gestation aged 16-47 years were recruited from Nanjing Kingmed Center for Clinical Laboratory. Nine thousand twenty-two pregnant women received GBS screening by PCR detection only. Seven thousand one hundred sixty-two pregnant women received GBS screening by bacterial culture and GBS-positive samples were tested for antibiotic resistance. RESULTS: The overall GBS positive rate was 8.7% by PCR and 3.5% by culture. Colonization rate was highest in the "25-29 years" age group. The 249 GBS-positive samples which detected by culture were all sensitive to penicillin. The prevalence of resistance to erythromycin, clindamycin, and levofloxacin was 77.5, 68.3, and 52.2%, respectively. CONCLUSIONS: This study revealed the data on the prevalence of GBS colonization in pregnant women at 34 to 37 weeks' gestation in Jiangsu, East China. It compared the difference of the sensitivity to detect GBS between PCR and culture. PCR was expected to become a quick method in pregnancy women conventional detection of GBS infection.


Assuntos
Complicações Infecciosas na Gravidez/epidemiologia , Infecções Estreptocócicas/epidemiologia , Streptococcus agalactiae/isolamento & purificação , Adolescente , Adulto , Antibacterianos/uso terapêutico , China/epidemiologia , Clindamicina/uso terapêutico , Farmacorresistência Bacteriana , Eritromicina/uso terapêutico , Feminino , Humanos , Recém-Nascido , Levofloxacino/uso terapêutico , Pessoa de Meia-Idade , Sepse Neonatal/epidemiologia , Penicilinas/uso terapêutico , Reação em Cadeia da Polimerase , Gravidez , Complicações Infecciosas na Gravidez/diagnóstico , Complicações Infecciosas na Gravidez/tratamento farmacológico , Complicações Infecciosas na Gravidez/microbiologia , Gestantes , Prevalência , Estudos Retrospectivos , Infecções Estreptocócicas/diagnóstico , Infecções Estreptocócicas/tratamento farmacológico , Streptococcus agalactiae/genética , Vagina/microbiologia , Adulto Jovem
19.
Zoolog Sci ; 38(3): 213-222, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34057345

RESUMO

Many plant-sucking stinkbugs possess a specialized symbiotic organ with numerous crypts in a posterior region of the midgut. In stinkbugs of the superfamily Pentatomoidea, specific γ-proteobacteria are hosted in the crypt cavities, which are vertically transmitted through host generations and essential for normal growth and survival of the host insects. Here we report the discovery of an exceptional gut symbiotic association in the saw-toothed stinkbug, Megymenum gracilicorne (Hemiptera: Pentatomoidea: Dinidoridae), in which specific γ-proteobacterial symbionts are not transmitted vertically but acquired environmentally. Histological inspection identified a very thin and long midgut symbiotic organ with two rows of tiny crypts whose cavities harbor rod-shaped bacterial cells. Molecular phylogenetic analyses of bacterial 16S rRNA gene sequences from the symbiotic organs of field-collected insects revealed that (i) M. gracilicorne is stably associated with Pantoea-allied γ-proteobacteria within the midgut crypts, (ii) the symbiotic bacteria exhibit a considerable level of diversity across host individuals and populations, (iii) the major symbiotic bacteria represent an environmental bacterial lineage that was reported to be capable of symbiosis with the stinkbug Plautia stali, and (iv) the minor symbiotic bacteria also represent several bacterial lineages that were reported as cultivable symbionts of P. stali and other stinkbugs. The symbiotic bacteria were shown to be generally cultivable. Microbial inspection of ovipositing adult females and their eggs and nymphs uncovered the absence of stable vertical transmission of the symbiotic bacteria. Rearing experiments showed that symbiont-supplemented newborn nymphs exhibit improved survival, suggesting the beneficial nature of the symbiotic association.


Assuntos
Bactérias/isolamento & purificação , Hemípteros/microbiologia , Simbiose , Animais , Bactérias/classificação , Bactérias/genética , Clonagem Molecular , DNA Bacteriano/genética , Microbiologia Ambiental , Reação em Cadeia da Polimerase , RNA Bacteriano/genética , RNA Ribossômico 16S/genética
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