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TaqMan Multiplex Real-time PCR Assay for Crimean-Congo Hemorrhagic Fever Virus Diagnosis Using Armored RNA Technology TaqMan多重实时PCR技术在克里米亚-刚果出血热病毒诊断中的应用
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-04-29 DOI: 10.5812/jjm-134188
Saleh Jamehdor, N. Hosseinirouzbahani, Seyed javad Hoseinishokoh, K. Ghorban, A. Teimoori, M. Gholami, Parisa Agahi, Ghazale Azizi, M. Mohammadimehr
Background: Crimean-Congo hemorrhagic fever virus (CCHFV) is a highly lethal virus that causes hemorrhagic fever in humans and is endemic in many countries, including Iran. Therefore, fast, accurate, and reliable diagnosis is crucial for patient management and outbreak control. Objectives: This study aims to optimize a TaqMan multiplex real-time RT-PCR for the rapid and specific diagnosis of CCHFV. Methods: In this study, the L (NC_005301.3) and S (NC_005302.1) fragments were used as reference sequences for blast analysis. The L and S sequence segments of CCHFV with more than 90% identity from different areas were downloaded from the Genbank database. Primers and probes were designed based on the best-conserved regions of CCHFV L and S sequence segments. To construct the plasmid, a 1751 bp fragment from the MS2 phage that was previously amplified using cloning primers was inserted into the pET-32a plasmid. The S and L segments of the CCHFV, which were 110 bp and 135 bp, respectively, were inserted downstream of the MS2 phage sequence from HindIII to NotI. The Viral-like particles (VLPs) were produced in Escherichia coli, strain BL-21(DE3), in the presence of 1 mM Isopropyl β-D-1-thiogalactopyranoside (IPTG). The stability of VLP particles was confirmed in the presence of the ribonuclease enzyme. The fabrication of VLPs was approved by transmission electron microscopy (TEM) with negative staining (1% phosphotungstic acid). To validate the specificity of the primers and probes sequences, we compared them to the NCBI database and tested them experimentally using extracted DNA and RNA samples from healthy subjects and an infectious panel. Results: The VLPs showed complete resistance in the presence of the ribonuclease enzyme, and the TEM results confirmed that the VLPs were correctly produced. The TaqMan multiplex real-time RT-PCR confirmed that the primers and probes were designed correctly and were completely specific to the CCHFV. The limit of detection (LOD) of the multiplex assay for the L and S genes was one copy of the VLPs per µL. Conclusions: This TaqMan assay is reliable for amplifying CCHFV due to its design on conserved regions of the CCHFV sequences, which have minimal variability and high specificity.
背景:克里米亚-刚果出血热病毒(CCHFV)是一种导致人类出血热的高致死性病毒,在包括伊朗在内的许多国家流行。因此,快速、准确和可靠的诊断对患者管理和疫情控制至关重要。目的:本研究旨在优化TaqMan多重实时RT-PCR用于CCHFV的快速和特异性诊断。方法:在本研究中,使用L(NC_005301.3)和S(NC_00530 2.1)片段作为参考序列进行爆破分析。从Genbank数据库中下载了来自不同地区的具有90%以上同一性的CCHFV的L和S序列片段。引物和探针是基于CCHFV L和S序列片段的最佳保守区设计的。为了构建质粒,将先前使用克隆引物扩增的来自MS2噬菌体的1751bp片段插入pET-32a质粒中。CCHFV的S和L片段分别为110bp和135bp,插入从HindIII到NotI的MS2噬菌体序列的下游。在1mM异丙基β-D-1硫代吡喃半乳糖苷(IPTG)存在下,在大肠杆菌BL-21(DE3)菌株中产生病毒样颗粒(VLP)。VLP颗粒的稳定性在核糖核酸酶的存在下得到证实。VLP的制备通过透射电子显微镜(TEM)和阴性染色(1%磷钨酸)获得批准。为了验证引物和探针序列的特异性,我们将它们与NCBI数据库进行了比较,并使用从健康受试者和感染小组提取的DNA和RNA样本对它们进行了实验测试。结果:在核糖核酸酶存在下,VLP表现出完全的抗性,TEM结果证实VLP是正确产生的。TaqMan多重实时RT-PCR证实引物和探针设计正确,对CCHFV完全特异。L和S基因多重检测的检测限(LOD)为每µL一个VLP拷贝。结论:该TaqMan分析法是扩增CCHFV的可靠方法,因为它设计在CCHFV序列的保守区,具有最小的变异性和高特异性。
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引用次数: 0
Parainfluenza Virus Type 3 Co-infection with Other Respiratory Pathogens Among Hospitalized Children with Acute Respiratory Infections in Wuhan, China 武汉市急性呼吸道感染住院儿童中3型副流感病毒与其他呼吸道病原体的联合感染
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-04-26 DOI: 10.5812/jjm-135823
Dan Z. Lu, Ying Cheng, Hongbo Hu
Background: Bacterial and viral co-infections are increasingly recognized as the cause of Acute Respiratory Infection (ARI). The role of co-infection in ARI patients with Parainfluenza Virus type 3 (PIV3) infection is unclear. Objectives: This study aimed to determine the prevalence of PIV3 co-infections in hospitalized children and assess the co-infections' role in ARI patients with PIV3 infections. Methods: Between January 2018 and December 2021, children were confirmed to have a PIV3 infection via throat swabs or nasopharyngeal aspirates. Some digital clinical data were analyzed, including demographic, epidemiological, diagnostic, and laboratory data. Results: During the study period from 2018 to 2021, 2,539 patients were hospitalized with ARI caused by PIV3. Of them, 34.0% had co-infection with other pathogens, and 2.4% had co-infection with more than two pathogens. Mycoplasma pneumoniae was the most common co-infecting pathogen (71.3%), followed by other bacteria (13.3%) and viruses (8.2%). A significantly higher proportion of patients with M. pneumoniae co-infection was found in girls (χ2 = 19.233, P < 0.001). Co-infections with M. pneumoniae were observed principally in patients aged 1 – 2 years (χ2 = 202.130, P < 0.001). In contrast, viral (56.3%) and bacterial (66.1%) co-infections occurred mainly in children younger than one year. The diagnosis of PIV3 as a single infection included pneumonia (41.2%), bronchitis (39.9%), upper respiratory tract infections (15.0%), and laryngitis (3.9%), which were distinguished from those with bacterial co-infections (χ2 = 16.424, P = 0.001) and co-infections with more than two pathogens (χ2 = 11.687, P = 0.010). Co-infections of PIV3 with any pathogen were not associated with admissions to intensive care units or ventilator support. However, the mean hospitalization was significantly higher in M. pneumoniae co-infections (t = 2.367, P = 0.018), bacterial co-infections (t = 2.402, P = 0.016), and co-infections with more than two pathogens (t = 2.827, P = 0.006) than in single PIV3 infection. Conclusions: Parainfluenza virus type 3 frequently occurs with other pathogens. The epidemiological and clinical characteristics of co-infections with different pathogens differed. Mycoplasma pneumoniae co-infections, bacterial co-infections, and co-infections with more than two pathogens lengthened the hospitalization. Bacterial co-infections and co-infections with more than two pathogens increased the severity of ARI and worsened the symptoms.
背景:细菌和病毒合并感染越来越被认为是急性呼吸道感染(ARI)的原因。ARI患者合并副流感病毒3型(PIV3)感染的合并感染的作用尚不清楚。目的:本研究旨在确定住院儿童PIV3合并感染的患病率,并评估合并感染在伴有PIV3感染的ARI患者中的作用。方法:2018年1月至2021年12月期间,通过咽拭子或鼻咽吸痰确认儿童感染PIV3。分析了一些数字临床数据,包括人口统计、流行病学、诊断和实验室数据。结果:2018年至2021年研究期间,2539例因PIV3引起的ARI住院。其中34.0%的患者合并感染其他病原菌,2.4%的患者合并感染2种以上病原菌。肺炎支原体是最常见的共感染病原体(71.3%),其次是其他细菌(13.3%)和病毒(8.2%)。女孩合并肺炎支原体感染的比例明显高于女孩(χ2 = 19.233, P < 0.001)。肺炎支原体合并感染主要发生在1 ~ 2岁的患者中(χ2 = 202.130, P < 0.001)。相比之下,病毒(56.3%)和细菌(66.1%)合并感染主要发生在一岁以下的儿童中。PIV3为单一感染的诊断包括肺炎(41.2%)、支气管炎(39.9%)、上呼吸道感染(15.0%)和喉炎(3.9%),与合并细菌感染(χ2 = 16.424, P = 0.001)和合并两种以上病原体感染(χ2 = 11.687, P = 0.010)有明显区别。PIV3与任何病原体的合并感染与入住重症监护病房或呼吸机支持无关。然而,肺炎支原体合并感染(t = 2.367, P = 0.018)、细菌合并感染(t = 2.402, P = 0.016)和两种以上病原体合并感染(t = 2.827, P = 0.006)的平均住院率显著高于单一PIV3感染。结论:3型副流感病毒常与其他病原体一起发生。不同病原菌合并感染的流行病学和临床特征存在差异。肺炎支原体合并感染、细菌合并感染和两种以上病原体合并感染延长了住院时间。细菌共感染和两种以上病原体的共感染增加了ARI的严重程度并使症状恶化。
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引用次数: 1
Serogroup and Pathogenicity Island Marker Distributions Among Uropathogenic Escherichia coli Isolates in Rasht, Iran 伊朗拉什特市尿路致病性大肠杆菌分离株的血清群和致病性岛标记分布
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-04-10 DOI: 10.5812/jjm-132754
Ali Moradpoor Shamami, M. Anvari, H. Pourmoshtagh, T. Shafighi, Hadi Seddigh Ebrahim-Saraie
Background: Urinary tract infections (UTIs) are among the most prevalent infections in hospitals and communities worldwide. Objectives: Due to the medical importance of UTIs caused by uropathogenic Escherichia coli (UPEC), this study aimed to investigate pathogenicity island (PAI) markers, O-antigen serogroups, and resistance to antibiotic agents associated with UPEC isolates obtained from hospitalized patients in Rasht city hospitals. Methods: A total of 110 urine samples were taken from patients with UTI referred to selected hospitals in Rasht, Iran. The double-disk synergy test (DDST) was used to detect the isolate’s ability to produce extended-spectrum β-lactamase (ESBL). Using particular primers, eight PAIs were detected (ie, PAI I536, PAI II536, PAI III536, PAI IV536, PAI ICFT073, PAI IICFT073, PAI IJ96, and PAI IIJ96). Results: According to the antibiotic susceptibility pattern, a high level of antibiotic resistance was observed against nalidixic acid (81.8%) and co-trimoxazole (78.2%), while the most effective agent was amikacin (85.5%). Double-disk synergy test revealed that the incidence of ESBL-positive strains was 62.7% (69/110). Of the 110 UPEC isolates, 106 (96.4%) carried at least one of the investigated PAI markers. UPEC isolates with PAI IV536 (81.8%) had the highest prevalence, and PAI J196 (6.4%) had the lowest PAI marker. The most predominant serogroup O was O25 (36.4%), followed by O16 (17.3%), while the O4 and O7 serogroups (0.9%) were the lowest serogroups among UPEC isolates. Conclusions: The characterization of our strain revealed the co-occurrence of PAI and serogroups, confirming the importance of antibiotic resistance among the distinct serogroups and PAI markers. Our results have potential application for epidemiological studies and designing UTI treatment strategies against UTIs caused by UPEC.
背景:尿路感染是世界各地医院和社区最常见的感染之一。目的:鉴于尿路致病性大肠杆菌(UPEC)引起的尿路感染在医学上的重要性,本研究旨在调查致病性岛(PAI)标志物、O-抗原血清群以及与Rasht市医院住院患者的UPEC分离株相关的抗生素耐药性。方法:从伊朗拉什特选定医院转诊的尿路感染患者身上共采集110份尿液样本。采用双纸片协同试验(DDST)检测分离株产超广谱β-内酰胺酶(ESBL)的能力。使用特定的引物,检测到8种PAI(即,PAI I536、PAI II536、PAI III536、PAI IV536、PAI ICFT073、PAI IICFT073,PAI IJ96和PAI IIJ96)。结果:从药敏模式来看,萘啶酸和复方三恶唑的耐药率较高(81.8%),阿米卡星的耐药率最高(85.5%),双纸片协同试验显示ESBL阳性菌株的发生率为62.7%(69/110)。在110个UPEC分离株中,106个(96.4%)携带至少一种所研究的PAI标记。具有PAI IV536的UPEC分离株(81.8%)的患病率最高,PAI J196(6.4%)的PAI标记物最低。最主要的O血清群是O25(36.4%),其次是O16(17.3%),而O4和O7血清群(0.9%)是UPEC分离株中最低的血清群。结论:我们菌株的特征揭示了PAI和血清群的共存,证实了不同血清群和PAI标记物之间抗生素耐药性的重要性。我们的研究结果有可能应用于流行病学研究和设计针对UPEC引起的尿路感染的UTI治疗策略。
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引用次数: 0
Genomic Insights Into Molecular Characteristics and Phylogenetic Linkage Between the Cases of Carbapenem-Resistant Klebsiella pneumoniae From a Non-tertiary Hospital in China: A Cohort Study 中国一家非三级医院耐碳青霉烯肺炎克雷伯菌病例的分子特征和系统发育联系的基因组学研究:一项队列研究
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-03-29 DOI: 10.5812/jjm-133210
C. Shen, Tao Lv, Ge Huang, Xiaoxiang Zhang, Lisi Zheng, Yunbo Chen
Background: Carbapenem-resistant Klebsiella pneumoniae (CRKP) strains have been listed as one of the major clinical concerns. Objectives: We investigated CPKP isolates from non-tertiary hospitals to find disseminated clones and analyze extensive phenotypic and genetic diversity in this study. Methods: In this cohort study, a total of 49 CRKP isolates from 3 hospitals in the same region were collected in 2021. The prevalence and antimicrobial susceptibility patterns were analyzed. Clinical data were retrieved from electronic medical record systems. The molecular types, antimicrobial resistance (AMR) profiles, plasmid replicons, and virulence factors were analyzed. The maximum-likelihood phylogenetic tree and transmission networks were constructed using single-nucleotide polymorphisms (SNPs). Results: The median age of patients (N = 49) was 66.0 years, and 85.7% were male. The most common CRKP infection was nosocomial pneumonia (75.5%), followed by bacteremia (10.2%). More than 53% of isolates were resistant to ceftazidime-avibactam (CAZ/AVI). Forty-five isolates were successfully sequenced; the predominant carbapenem-resistant gene was blaKPC-2 (93.3%). The 30-day mortality in our cohort was 24.5%. The most dominant sequence type (ST) was ST11 (60.0%), followed by ST15 (13.3%). Whole genome sequencing (WGS) analysis exhibited dissemination of ST11 strain clones, ST420, and ST15 clones, both within and outside the given hospital. Conclusions: In this surveillance study, several dissemination chains of CRKP were discovered in the hospital and the region, as ST11 was the main epidemic clone. Our findings suggest that effective infection control practices and antimicrobial stewardship are needed in non-tertiary hospitals in China.
背景:耐碳青霉烯类肺炎克雷伯菌(CRKP)菌株已被列为临床关注的主要菌株之一。目的:在本研究中,我们调查了来自非三级医院的CPKP分离株,以寻找传播性克隆并分析广泛的表型和遗传多样性。方法:在本队列研究中,2021年共收集了来自同一地区3家医院的49株CRKP分离株。分析了流行率和耐药性模式。临床数据是从电子病历系统中检索的。分析了分子类型、抗微生物耐药性(AMR)谱、质粒复制子和毒力因子。利用单核苷酸多态性(SNPs)构建了最大似然系统发育树和传播网络。结果:患者的中位年龄(N=49)为66.0岁,其中85.7%为男性。最常见的CRKP感染是医院内肺炎(75.5%),其次是菌血症(10.2%)。超过53%的分离株对头孢他啶-阿维巴坦(CAZ/AVI)具有耐药性。45个分离株成功测序;主要的碳青霉烯抗性基因是blaKPC-2(93.3%)。我们队列中的30天死亡率为24.5%。最主要的序列类型(ST)是ST11(60.0%),其次是ST15(13.3%)。全基因组测序(WGS)分析显示,ST11菌株克隆、ST420和ST15克隆在医院内外传播。结论:在本监测研究中,由于ST11是主要的流行克隆,在医院和该地区发现了CRKP的几个传播链。我们的研究结果表明,中国非三级医院需要有效的感染控制措施和抗菌药物管理。
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引用次数: 0
Immunologic Profile of Severe COVID-19 Patients in Alborz Province, Iran 伊朗阿尔布尔茨省新冠肺炎重症患者免疫概况
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-03-27 DOI: 10.5812/jjm-134264
Arman Shafiee, S. Rezaian, Mansur Aliyu, Ali Shayeghpour, Z. Mokhames, Hamed Mohammadi, Somayeh Yaslianifard, A. Soleimani, Fatemeh Soleimanifar, Taranom Tojari, M. Qorbani, Sayed-Hamidreza Mozhgani
Background: The coronavirus disease 2019 (COVID-19) pandemic has prompted researchers to look for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pathogenicity in depth. Immune system dysregulation was one of the major mechanisms in its pathogenesis. The evidence regarding the levels of interferons (IFNs) and pro- and anti-inflammatory cytokines in COVID-19 patients is not well-established. Objectives: Therefore, this study evaluated the expression level of type-I, II, III IFNs, along with interleukin-1 (IL-1), interleukin-6 (IL-6), interleukin-10 (IL-10), and FOXP3 genes in patients with severe COVID-19 to provide additional insights regarding the regulation of these cytokines during COVID-19 infection. Methods: Peripheral blood mononuclear cells were isolated from two groups, including severe COVID-19 patients and healthy controls. Ribonucleic acid was extracted to evaluate the expression level of IFN-a, IFN-b, IFN-g, IFN-la, IL-1, IL-6, IL-10, and FOXP3 genes using real-time polymerase chain reaction. The correlations between the expression levels of these genes were also assessed. Results: A total of 40 samples were divided into two groups, with each group consisting of 20 samples. When comparing the severe COVID-19 group to the controls, the expression levels of IFN-g, tumor necrosis factor-alpha (TNF-α), IL-6, and IL-10 genes were significantly higher in the severe COVID-19 group. The two groups had no significant differences in IFN-a, IFN-b, IFN-la, IL-1, and FOXP3 expression. The correlation analysis revealed a negative correlation between type I and type III IFNs (i.e., IFN-a and IFN-la) and pro-inflammatory cytokines (i.e., IL-1 and IL-10). Conclusions: This study suggests the possible upregulation of IFN-g, IL-6, IL-10, and TNF-α during SARS-CoV-2 pathogenicity. The preliminary findings of this study and those reported previously show that the levels of IFNs and pro- and anti-inflammatory cytokines are not uniformly expressed among all COVID-19 patients and might differ as the disease progresses to the severe stage.
背景:2019冠状病毒病(新冠肺炎)大流行促使研究人员深入寻找严重急性呼吸综合征冠状病毒2(SARS-CoV-2)的致病性。免疫系统失调是其发病机制之一。关于新冠肺炎患者中干扰素(IFN)、促炎和抗炎细胞因子水平的证据尚不明确。目的:因此,本研究评估了重症新冠肺炎患者中I型、II型、III型IFN以及白细胞介素-1(IL-1)、白细胞介素-6(IL-6)、白介素-10(IL-10)和FOXP3基因的表达水平,以提供有关这些细胞因子在新冠肺炎感染期间的调节的更多见解。方法:从新冠肺炎重症患者和健康对照组中分离外周血单核细胞。提取核糖核酸,使用实时聚合酶链反应评估IFN-a、IFN-b、IFN-g、IFN-la、IL-1、IL-6、IL-10和FOXP3基因的表达水平。还评估了这些基因的表达水平之间的相关性。结果:将40份样本分为两组,每组20份。当将重症新冠肺炎组与对照组进行比较时,重症新冠肺炎组IFN-g、肿瘤坏死因子α(TNF-α)、IL-6和IL-10基因的表达水平显著升高。两组在IFN-a、IFN-b、IFN-la、IL-1和FOXP3表达方面没有显著差异。相关性分析显示,I型和III型IFN(即IFN-a和IFN-la)与促炎细胞因子(即IL-1和IL-10)之间呈负相关。结论:本研究提示IFN-g、IL-6、IL-10和TNF-α在严重急性呼吸系统综合征冠状病毒2型致病过程中可能上调。这项研究的初步结果和之前报道的结果表明,在所有新冠肺炎患者中,IFN以及促炎和抗炎细胞因子的水平并不一致,并且可能随着疾病发展到严重阶段而有所不同。
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引用次数: 0
Evaluation of Anti-bacterial, Anti-adenoviral, and Apoptosis-inducing Activity of Bacillus clausii Supernatant 幽闭芽孢杆菌上清液的抗菌、抗腺病毒和诱导细胞凋亡活性评价
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-03-25 DOI: 10.5812/jjm-132952
Z. Fateminasab, M. Shayestehpour, M. Zolfaghari
Background: Bacillus clausii is being studied as a probiotic candidate. There is insufficient information on the antimicrobial and anticancer effects of B. clausii. Objectives: The present investigation was designed to evaluate the anti-bacterial, anti-adenoviral, and apoptosis-inducing activity of B. clausii cell-free supernatant (CFS). Methods: First, the supernatant of B. clausii was collected after culture for 24 h. Then, its anti-bacterial impact on several genera of bacteria was assessed through the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). Adenovirus 5 (Ad5) was exposed to the CFS under four conditions, including pre-treatment: first infecting cells with CFS and then with the virus; pre-incubation: incubation of the supernatant and virus for 1.5 hours and then adding to the cells; competition: infection of cells with the simultaneous mixture of the supernatant and virus, and post-treatment: first infecting cells with the virus and then with CFS. The median tissue culture infectious dose (TCID50) technique determined the virus titer. Real-time PCR was performed to assess the E1A expression. After exposure to the CFS, real-time PCR was utilized to measure the expression of MicroRNA-145, BCL-2, and BAX in HeLa cancer cells. Results: Bacillus clausii supernatant showed an inhibitory effect on MRSA, Enterococcus faecalis, Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, and Acinetobacter. baumannii. The Ad5 titers were reduced by about 4.61, 4, 3.9, and 3.1 Log10 TCID50/mL in pre-treatment, pre-incubation, competition, and post-treatment tests (CFS dilution: 1/4), respectively. Similar results of the viral titration were seen when experimental and control E1A expression levels were compared. Also, B. clausii supernatant during 48 h exposure to HeLa cells increased the transcript of the BAX, BCL-2, and miR-145 genes to 9.1, 2.3, and 55 folds, respectively, compared to the untreated condition. Conclusions: Bacillus clausii can be a potent antimicrobial and anticancer agent. Further research is required to learn about the spectrum of anti-bacterial, antiviral, and anti-cancerous activities of B. clausii.
背景:幽闭芽孢杆菌正在作为一种候选益生菌进行研究。关于B.clausii的抗菌和抗癌作用的信息不足。目的:本研究旨在评估幽闭双歧杆菌无细胞上清液(CFS)的抗菌、抗腺病毒和诱导细胞凋亡的活性。方法:首先收集克劳氏芽孢杆菌培养24小时后的上清液,然后通过最低抑菌浓度(MIC)和最低杀菌浓度(MBC)评估其对几个属细菌的抗菌作用。腺病毒5(Ad5)在四种条件下暴露于CFS,包括预处理:首先用CFS感染细胞,然后用病毒感染;预孵育:将上清液和病毒孵育1.5小时,然后加入细胞中;竞争:用上清液和病毒的同时混合物感染细胞,以及后处理:首先用病毒感染细胞,然后用CFS感染细胞。中位组织培养感染剂量(TCID50)技术测定病毒滴度。进行实时PCR以评估E1A的表达。暴露于CFS后,实时PCR用于测量HeLa癌症细胞中MicroRNA-145、BCL-2和BAX的表达。结果:幽闭芽孢杆菌上清液对MRSA、粪肠球菌、金黄色葡萄球菌、铜绿假单胞菌、大肠杆菌和不动杆菌均有抑制作用。鲍曼尼。在预处理、预孵育、竞争和处理后测试(CFS稀释度:1/4)中,Ad5滴度分别降低了约4.61、4、3.9和3.1 Log10 TCID50/mL。当比较实验和对照E1A表达水平时,可以看到类似的病毒滴定结果。此外,与未处理的条件相比,在暴露于HeLa细胞的48小时期间,B.clausii上清液将BAX、BCL-2和miR-145基因的转录物分别增加到9.1、2.3和55倍。结论:幽闭芽孢杆菌是一种有效的抗菌、抗癌药物。需要进一步的研究来了解B.clausii的抗菌、抗病毒和抗癌活性。
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引用次数: 0
Frequency of Clostridioides difficile Infection Among Hospitalized Patients in Kerman City, Iran 伊朗克尔曼市住院患者中艰难梭菌感染频率分析
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-03-14 DOI: 10.5812/jjm-132262
M. Shojaei, Farokh Rokhbakhsh-Zamin, Ebrahim Rezazadeh Zarandi, F. Sarafzadeh, Sayed Mohammad Reza Khoshroo
Background: Clostridioides difficile is one of the major causes of nosocomial infections, being responsible for 15 to 25% of antibiotic-associated diarrhea. It is important to determine the epidemiology and prevalence of this bacterium at hospitals and healthcare centers. Objectives: This study aims to investigate the prevalence of C. difficile infection (CDI) by identifying toxigenic isolates of C. difficile in different wards of the hospital. Methods: A total of 417 diarrheal stool samples were taken from hospitalized patients in different wards of three educational hospitals in Kerman City, Iran from 2018 to 2020. The samples were cultured on cycloserine-cefoxitin fructose agar and C. difficile suspected colonies were isolated. Identification of the cdd-3 gene for definitive diagnosis of C. difficile and identification of toxin genes in the positive isolates was performed using the PCR method. Results: A total of 68 isolates (16.3%) of C. difficile were isolated from the specimens. Besides, 8.6% (36/417) and 7.6% (32/417) of the isolates were toxigenic and nontoxigenic, respectively; thus, the prevalence of CDI was 8.6%. Most of the toxigenic isolates had the A+B+CDT- toxin phenotype. The highest prevalence of CDI was observed in males, ICU ward, and age group of 41 - 60. Conclusions: A total of 8.6% of hospitalized patients with diarrhea were infected with C. difficile. The prevalence of CDI in Kerman City is lower than that in Europe, East Asia, and other parts of Iran, but it is almost the same as that in the Middle East.
背景:艰难梭菌是院内感染的主要原因之一,占抗生素相关性腹泻的15%至25%。确定这种细菌在医院和保健中心的流行病学和流行程度是很重要的。目的:本研究旨在通过鉴定难辨梭菌在医院不同病房的产毒分离株,了解难辨梭菌感染(CDI)的流行情况。方法:采集伊朗克尔曼市3所教育医院2018 - 2020年不同病房住院患者的腹泻粪便样本417份。在环丝氨酸-头孢西丁-果糖琼脂培养基上培养,分离出疑似难辨梭菌菌落。采用PCR方法对艰难梭菌的cdd-3基因进行鉴定,并对阳性分离株的毒素基因进行鉴定。结果:从标本中分离出难辨梭菌68株(16.3%)。产毒株占8.6%(36/417),非产毒株占7.6% (32/417);因此,CDI患病率为8.6%。大多数产毒株具有A+B+CDT-毒素表型。CDI患病率最高的是男性、ICU病房和41 - 60岁年龄组。结论:住院腹泻患者中难辨梭菌感染率为8.6%。克尔曼市的CDI患病率低于欧洲、东亚和伊朗其他地区,但与中东地区几乎相同。
{"title":"Frequency of Clostridioides difficile Infection Among Hospitalized Patients in Kerman City, Iran","authors":"M. Shojaei, Farokh Rokhbakhsh-Zamin, Ebrahim Rezazadeh Zarandi, F. Sarafzadeh, Sayed Mohammad Reza Khoshroo","doi":"10.5812/jjm-132262","DOIUrl":"https://doi.org/10.5812/jjm-132262","url":null,"abstract":"Background: Clostridioides difficile is one of the major causes of nosocomial infections, being responsible for 15 to 25% of antibiotic-associated diarrhea. It is important to determine the epidemiology and prevalence of this bacterium at hospitals and healthcare centers. Objectives: This study aims to investigate the prevalence of C. difficile infection (CDI) by identifying toxigenic isolates of C. difficile in different wards of the hospital. Methods: A total of 417 diarrheal stool samples were taken from hospitalized patients in different wards of three educational hospitals in Kerman City, Iran from 2018 to 2020. The samples were cultured on cycloserine-cefoxitin fructose agar and C. difficile suspected colonies were isolated. Identification of the cdd-3 gene for definitive diagnosis of C. difficile and identification of toxin genes in the positive isolates was performed using the PCR method. Results: A total of 68 isolates (16.3%) of C. difficile were isolated from the specimens. Besides, 8.6% (36/417) and 7.6% (32/417) of the isolates were toxigenic and nontoxigenic, respectively; thus, the prevalence of CDI was 8.6%. Most of the toxigenic isolates had the A+B+CDT- toxin phenotype. The highest prevalence of CDI was observed in males, ICU ward, and age group of 41 - 60. Conclusions: A total of 8.6% of hospitalized patients with diarrhea were infected with C. difficile. The prevalence of CDI in Kerman City is lower than that in Europe, East Asia, and other parts of Iran, but it is almost the same as that in the Middle East.","PeriodicalId":17803,"journal":{"name":"Jundishapur Journal of Microbiology","volume":" ","pages":""},"PeriodicalIF":0.6,"publicationDate":"2023-03-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"42307494","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Loop-mediated Isothermal Amplification Test for Rapid Identification of Clinical Nocardia Isolates 快速鉴定临床诺卡菌分离株的环介导等温扩增试验
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-03-13 DOI: 10.5812/jjm-132432
Mojtaba Taghizadeh Armaki, Mohammad Hossein Tayefeh-Arbab, Jalal Jafarzadeh, Sina Nasrollahian, Z. Baseri, H. Mehdinezhad, R. Rajabnia, Abazar Pournajaf
Background: Nocardia is a Gram-positive and partially acid-fast bacterium. The species are widely distributed in the environment and cause severe human infections. Nocardiosis is not easily identifiable due to the lack of pathognomonic clinical signs. Objectives: The present study was designed to develop and evaluate a simple and quick method based on a loop-mediated isothermal amplification (LAMP) assay for detecting Nocardia spp isolated from bronchoalveolar lavage (BAL) samples. Methods: In this cross-sectional study, 357 BAL samples were collected from two teaching hospitals. The polymerase chain reaction (PCR) was performed using a set of species-specific primers for the 16S rRNA gene. Kinyoun acid-fast staining and culture were done on the Sabouraud dextrose plate. The optimal LAMP reaction condition was set at 65°C for 45 min, with the recognition limit as 1 pg DNA/tube and 100 CFU/reaction. In addition to calcein and manganous ions, agarose gel electrophoresis was used to visualize the amplified LAMP products. Results: Out of 357 BAL samples, 0 (0.0%), 4 (1.1%), 9 (2.5%), and 10 (2.8%) Nocardia strains were identified by direct staining of partial acid-fast, streak culture plate, PCR, and LAMP methods, respectively. Conclusions: We developed a new LAMP technique for the recognition of Nocardia, which is fast, very precise, simple, and low-cost. According to our knowledge, this is the first report of the LAMP method to detect Nocardia in clinical samples.
背景:诺卡氏菌是一种革兰氏阳性且部分耐酸的细菌。该物种广泛分布于环境中,会引起严重的人类感染。由于缺乏典型的临床症状,因此不易识别无心律不齐。目的:本研究旨在开发和评估一种基于环介导等温扩增(LAMP)检测从支气管肺泡灌洗液(BAL)样本中分离的诺卡氏菌spp的简单快速方法。方法:在这项横断面研究中,从两家教学医院收集了357份BAL样本。聚合酶链式反应(PCR)是使用一组16S rRNA基因的物种特异性引物进行的。在沙氏葡萄糖平板上进行Kinyoun抗酸染色和培养。LAMP的最佳反应条件设定为65°C,持续45分钟,识别极限为1 pg DNA/管和100 CFU/反应。除了钙黄绿素和锰离子外,还使用琼脂糖凝胶电泳来观察扩增的LAMP产物。结果:在357份BAL样本中,通过部分抗酸直接染色法、条纹培养板法、PCR法和LAMP法分别鉴定出0株(0.0%)、4株(1.1%)、9株(2.5%)和10株(2.8%)诺卡氏菌。结论:我们开发了一种新的LAMP技术来识别诺卡氏菌,该技术快速、精确、简单、低成本。据我们所知,这是LAMP方法首次在临床样本中检测诺卡氏菌。
{"title":"Loop-mediated Isothermal Amplification Test for Rapid Identification of Clinical Nocardia Isolates","authors":"Mojtaba Taghizadeh Armaki, Mohammad Hossein Tayefeh-Arbab, Jalal Jafarzadeh, Sina Nasrollahian, Z. Baseri, H. Mehdinezhad, R. Rajabnia, Abazar Pournajaf","doi":"10.5812/jjm-132432","DOIUrl":"https://doi.org/10.5812/jjm-132432","url":null,"abstract":"Background: Nocardia is a Gram-positive and partially acid-fast bacterium. The species are widely distributed in the environment and cause severe human infections. Nocardiosis is not easily identifiable due to the lack of pathognomonic clinical signs. Objectives: The present study was designed to develop and evaluate a simple and quick method based on a loop-mediated isothermal amplification (LAMP) assay for detecting Nocardia spp isolated from bronchoalveolar lavage (BAL) samples. Methods: In this cross-sectional study, 357 BAL samples were collected from two teaching hospitals. The polymerase chain reaction (PCR) was performed using a set of species-specific primers for the 16S rRNA gene. Kinyoun acid-fast staining and culture were done on the Sabouraud dextrose plate. The optimal LAMP reaction condition was set at 65°C for 45 min, with the recognition limit as 1 pg DNA/tube and 100 CFU/reaction. In addition to calcein and manganous ions, agarose gel electrophoresis was used to visualize the amplified LAMP products. Results: Out of 357 BAL samples, 0 (0.0%), 4 (1.1%), 9 (2.5%), and 10 (2.8%) Nocardia strains were identified by direct staining of partial acid-fast, streak culture plate, PCR, and LAMP methods, respectively. Conclusions: We developed a new LAMP technique for the recognition of Nocardia, which is fast, very precise, simple, and low-cost. According to our knowledge, this is the first report of the LAMP method to detect Nocardia in clinical samples.","PeriodicalId":17803,"journal":{"name":"Jundishapur Journal of Microbiology","volume":" ","pages":""},"PeriodicalIF":0.6,"publicationDate":"2023-03-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"43278355","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
The Circulation of Common Respiratory Viruses and Their Co-infection with Severe Acute Respiratory Syndrome Coronavirus 2 Before and After Coronavirus Disease of 2019 Vaccination 2019冠状病毒病疫苗接种前后常见呼吸道病毒的传播及其与严重急性呼吸综合征冠状病毒2型的共感染
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-02-19 DOI: 10.5812/jjm-133326
S. Ebrahimi, M. Kalantar, Barat Barati, Nayeb Fadaei Dehcheshmeh, Zahra Najafimemar, T. Navidifar, F. Seif
Background: Respiratory viruses play important roles in respiratory tract infections; they are the major cause of diseases such as the common cold, bronchiolitis, pneumonia, etc., in humans that circulate more often in the cold seasons. During the COVID-19 pandemic, many strict public health measures, such as hand hygiene, the use of face masks, social distancing, and quarantines, were implemented worldwide to control the pandemic. Besides controlling the COVID-19 pandemic, these introduced measures might change the spread of other common respiratory viruses. Moreover, with COVID-19 vaccination and reducing public health protocols, the circulation of other respiratory viruses probably increases in the community. Objectives: This study aims to explore changes in the circulation pattern of common respiratory viruses during the COVID-19 pandemic. Methods: In the present study, we evaluated the circulation of seven common respiratory viruses (influenza viruses A and B, rhinovirus, and seasonal human Coronaviruses (229E, NL63, OC43, and HKU1) and their co-infection with SARS-CoV-2 in suspected cases of COVID-19 in two time periods before and after COVID-19 vaccination. Clinical nasopharyngeal swabs of 400 suspected cases of COVID-19 were tested for SARS-CoV-2 and seven common respiratory viruses by reverse transcription real-time polymerase chain reaction. Results: Our results showed common respiratory viruses were detected only in 10% and 8% of SARS-CoV-2-positive samples before and after vaccination, respectively, in which there were not any significant differences between them (P-value = 0.14). Moreover, common viral respiratory infections were found only in 12% and 32% of SARS-CoV-2-negative specimens before and after vaccination, respectively, in which there was a significant difference between them (P-value = 0.041). Conclusions: Our data showed a low rate of co-infection of other respiratory viruses with SARS-CoV-2 at both durations, before and after COVID-19 vaccination. Moreover, the circulation of common respiratory viruses before the COVID-19 vaccination was lower, probably due to non-pharmaceutical interventions (NPI), while virus activity (especially influenza virus A) was significantly increased after COVID-19 vaccination with reducing strict public health measures.
背景:呼吸道病毒在呼吸道感染中起着重要作用;它们是常见感冒、细支气管炎、肺炎等疾病的主要病因,这些疾病在寒冷季节更常见。在新冠肺炎大流行期间,世界各地实施了许多严格的公共卫生措施,如手卫生、口罩的使用、保持社交距离和隔离,以控制疫情。除了控制新冠肺炎大流行,这些引入的措施可能会改变其他常见呼吸道病毒的传播。此外,随着新冠肺炎疫苗接种和公共卫生协议的减少,其他呼吸道病毒在社区中的传播可能会增加。目的:本研究旨在探讨新冠肺炎大流行期间常见呼吸道病毒循环模式的变化。方法:在本研究中,我们评估了7种常见呼吸道病毒(流感病毒A和B、鼻病毒和季节性人类冠状病毒(229E、NL63、OC43和HKU1)在新冠肺炎疫苗接种前后两个时间段内的循环情况及其在疑似病例中与SARS-CoV-2的共同感染。通过逆转录实时聚合酶链反应对400例新冠肺炎疑似病例的临床鼻咽拭子进行了SARS-CoV-2和7种常见呼吸道病毒检测。结果:我们的结果显示,在接种疫苗前后,常见呼吸道病毒分别仅在10%和8%的严重急性呼吸系统综合征冠状病毒2型阳性样本中检测到,两者之间没有任何显著差异(P值=0.14),结论:我们的数据显示,在新冠肺炎疫苗接种前后,其他呼吸道病毒与SARS-CoV-2的共同感染率较低。此外,在新冠肺炎疫苗接种前,常见呼吸道病毒的传播较低,这可能是由于非药物干预(NPI),而在新冠肺炎疫苗接种后,随着严格的公共卫生措施的减少,病毒活性(尤其是流感病毒A)显著增加。
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引用次数: 2
Analysis of Microsatellite Length Polymorphism for Clinical Isolates of Candida albicans from Animals 动物白色念珠菌临床分离株微卫星长度多态性分析
IF 0.6 4区 医学 Q4 MICROBIOLOGY Pub Date : 2023-02-15 DOI: 10.5812/jjm-132587
F. Katiraee, Neda Kiasat, Anahita Kasmaie, Alireza Salimi, H. Shokri
Background: Candida albicans has been shown as the most common species of Candida collected from different animals. Objectives: This study aimed to evaluate the genetic diversity and genetic relationships among C. albicans isolates collected from clinical specimens of animals suffering from candidiasis using microsatellite length polymorphism (MLP). Methods: We used MLP for a group of 60 C. albicans strains isolated from various animal species (dog: 16, cat: 10, horse: 10, cow: 14, chicken: 10), previously defined as animal clinical isolates. Three loci, including EF3, CDC3, and HIS3, were amplified, and the products ran onto an ABI XL 370 genetic analyzer, and fragment sizes were determined. Results: Of the 60 clinical strains illustrated, 49 different genotypes were identified with a discriminatory power index of 0.991. A total of 17 alleles and 26 different combinations were identified for EF3 locus, six alleles and 13 combinations for CDC3 locus, and 17 alleles and 27 combinations for HIS3 locus. The most common genotypes were GP9 (four strains) and GP1 and GP33 (three strains). Wright’s fixation index (FST) values were calculated to assess inter-group genetic diversity for all pairwise combinations of the five sub-populations of C. albicans isolated from the different animal hosts. The highest FST values related to C. albicans isolated from chicken to three sub-populations of cats (FST: 0.1397), cows (FST: 0.0639), and horses (FST: 0.0585). Conclusions: The results indicated a moderate genetic differentiation (0.05 < FST < 0.15) between C. albicans strains isolated from cats, cows, and horses as a mammal vs. chickens.
背景:白色念珠菌是从不同动物身上采集的最常见的念珠菌。目的:本研究旨在利用微卫星长度多态性(MLP)评价从念珠菌感染动物临床标本中分离的白色念珠菌的遗传多样性和遗传关系。方法:我们使用MLP对从不同动物物种(狗:16,猫:10,马:10,牛:14,鸡:10)分离的60株白色念珠菌菌株进行研究,这些菌株以前被定义为动物临床分离株。扩增三个基因座,包括EF3、CDC3和HIS3,并将产物运到ABI XL 370遗传分析仪上,测定片段大小。结果:在所示的60株临床菌株中,鉴定出49种不同的基因型,判别力指数为0.991。共鉴定出EF3基因座的17个等位基因和26个不同的组合,CDC3基因座的6个等位突变和13个组合,HIS3基因座的7个等位变异和27个组合。最常见的基因型是GP9(4株)和GP1和GP33(3株)。计算Wright固定指数(FST)值,以评估从不同动物宿主分离的白色念珠菌五个亚群的所有成对组合的组间遗传多样性。从鸡分离的白色念珠菌的最高FST值与猫(FST:0.1397)、牛(FST:0.0639)和马(FST:0.0585)三个亚群有关。
{"title":"Analysis of Microsatellite Length Polymorphism for Clinical Isolates of Candida albicans from Animals","authors":"F. Katiraee, Neda Kiasat, Anahita Kasmaie, Alireza Salimi, H. Shokri","doi":"10.5812/jjm-132587","DOIUrl":"https://doi.org/10.5812/jjm-132587","url":null,"abstract":"Background: Candida albicans has been shown as the most common species of Candida collected from different animals. Objectives: This study aimed to evaluate the genetic diversity and genetic relationships among C. albicans isolates collected from clinical specimens of animals suffering from candidiasis using microsatellite length polymorphism (MLP). Methods: We used MLP for a group of 60 C. albicans strains isolated from various animal species (dog: 16, cat: 10, horse: 10, cow: 14, chicken: 10), previously defined as animal clinical isolates. Three loci, including EF3, CDC3, and HIS3, were amplified, and the products ran onto an ABI XL 370 genetic analyzer, and fragment sizes were determined. Results: Of the 60 clinical strains illustrated, 49 different genotypes were identified with a discriminatory power index of 0.991. A total of 17 alleles and 26 different combinations were identified for EF3 locus, six alleles and 13 combinations for CDC3 locus, and 17 alleles and 27 combinations for HIS3 locus. The most common genotypes were GP9 (four strains) and GP1 and GP33 (three strains). Wright’s fixation index (FST) values were calculated to assess inter-group genetic diversity for all pairwise combinations of the five sub-populations of C. albicans isolated from the different animal hosts. The highest FST values related to C. albicans isolated from chicken to three sub-populations of cats (FST: 0.1397), cows (FST: 0.0639), and horses (FST: 0.0585). Conclusions: The results indicated a moderate genetic differentiation (0.05 < FST < 0.15) between C. albicans strains isolated from cats, cows, and horses as a mammal vs. chickens.","PeriodicalId":17803,"journal":{"name":"Jundishapur Journal of Microbiology","volume":" ","pages":""},"PeriodicalIF":0.6,"publicationDate":"2023-02-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"42787495","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
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Jundishapur Journal of Microbiology
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