The histopathologic spectrum of decorative tattoo complications

Tattooing for ornamental purposes is an ancient practice that remains popular in modern times. Tattoos are encountered by the dermatopathologist either as incidental findings on skin biopsies or because of complications specific to the tattoo. A range of neoplasms and inflammatory conditions are seen in association with tattoos, many of which may be attributed to hypersensitivity to tattoo inks. The composition of tattoo inks is highly variable, and inks can contain numerous potentially allergenic or carcinogenic compounds. Infections with bacterial, viral and fungal species can occur after tattooing, sometimes after substantial delay. Atypical mycobacterial infections in particular are increasingly reported; special stains for mycobacteria should be performed and cultures recommended particularly when dense, mixed or granulomatous infiltrates are present.     

Facial tuberculoid leprosy: case report

Leprosy is a chronic, infectious, systemic disease caused by Mycobacterium leprae and is classified as paucibacillary and multibacillary types. It is contagious and has an insidious onset. Clinical presentation is characterised by hypopigmented skin lesions with reduced sensation. Presence of acid-fast bacilli in tissue specimens is regarded as a gold standard for diagnosis. Treatment is based on multi-drug regimens. We report a case of borderline tuberculoid leprosy in a 31-year-old woman.     

Fluorescent quantitative PCR of Mycobacterium tuberculosis for differentiating intestinal tuberculosis from Crohn's disease

Intestinal tuberculosis (ITB) and Crohn''s disease (CD) are granulomatous disorders with similar clinical manifestations and pathological features that are often difficult to differentiate. This study evaluated the value of fluorescent quantitative polymerase chain reaction (FQ-PCR) for Mycobacterium tuberculosis (MTB) in fecal samples and biopsy specimens to differentiate ITB from CD. From June 2010 to March 2013, 86 consecutive patients (38 females and 48 males, median age 31.3 years) with provisional diagnoses of ITB and CD were recruited for the study. The patients'' clinical, endoscopic, and histological features were monitored until the final definite diagnoses were made. DNA was extracted from 250 mg fecal samples and biopsy tissues from each patient. The extracted DNA was amplified using FQ-PCR for the specific MTB sequence. A total of 29 ITB cases and 36 CD cases were included in the analysis. Perianal disease and longitudinal ulcers were significantly more common in the CD patients (P<0.05), whereas night sweats, ascites, and circumferential ulcers were significantly more common in the ITB patients (P<0.05). Fecal FQ-PCR for MTB was positive in 24 (82.8%) ITB patients and 3 (8.3%) CD patients. Tissue PCR was positive for MTB in 16 (55.2%) ITB patients and 2 (5.6%) CD patients. Compared with tissue FQ-PCR, fecal FQ-PCR was more sensitive (X²=5.16, P=0.02). We conclude that FQ-PCR for MTB on fecal and tissue samples is a valuable assay for differentiating ITB from CD, and fecal FQ-PCR has greater sensitivity for ITB than tissue FQ-PCR.     

高分辨率CT在非结核分枝杆菌肺病诊治中的价值

搜集广州市胸科医院2015年3月至2017年9月经菌种鉴定为非结核分枝杆菌(NTM)肺病、并行胸部高分辨率CT(HRCT)检查的99例患者,回顾性分析肺内病灶部位、数量、形态、大小、密度、边界、分布及淋巴结的CT征象;评估99例NTM肺病患者治疗前后HRCT的表现。HRCT检查表现为斑点片状渗出、实变93例,支气管扩张78例,空洞60例,纤维条索状影59例,结节状影58例,树芽征55例,磨玻璃样影46例,淋巴结肿大45例,胸膜增厚18例,胸腔积液9例,病变通常以多种形态混杂存在。91例NTM肺病患者经克拉霉素、利福平、乙胺丁醇及阿米卡星治疗6~18个月后CT复查,显示病灶均减少;8例患者经HRCT精准定位后行患侧肺叶切除术,术后肺部病灶明显减少,但均出现术侧胸膜肥厚粘连。胸部HRCT对NTM肺病的诊断及治疗具有重要的应用价值。     

Early Effects Triggered by Larrea divaricata Cav. on Murine Macrophages at Apoptotic Concentrations

Decoction and infusion of Larrea divaricata were tested at apoptotic concentrations (1 and 4 mg/ml) on peritoneal murine macrophages. Consistent changes were observed after incubation with 4 mg/ml decoction. Phagocytosis of zymosan, lysosomal enzyme activity, nitric oxide production, TNF-α release, and expression of CD14, TLR4, and CR3 increased significantly. Decoction at 1 and 4 mg/ml increased the binding of LPS-FITC. Apoptosis triggered by L. divaricata decoction is consequence of cell activation. The effects are independent of nordihydroguaiaretic acid. This “activation and death” could be the mechanism of L. divaricata to exert the antituberculosis effect known in folk medicine.     

用耐药基因检测方法指导老年肺结核病人治疗的临床研究

目的：了解老年肺结核病人的结核分支杆菌耐药情况，评价它们的临床应用价值。方法：采用聚合酶链反应－单链构象多态性分析（PCR－SSCP）检测结核分支杆菌rpoB,katG,rpsL,pncA,embB基因突变和药物敏感试验（比例法），了解117例老年结核病人结核分枝杆菌耐药情况。探讨和比较2种耐药性检测方法的检测效果。结果：1／2上的肺结核病人至少耐2种抗结核药物，对RFP、INH、SM、PZA和EB总耐药率为82．1％、71．7％、63．2％、51．2％和35．0％，耐多药率为67．5％，rpoB,katG,rpsL,PpncA和embB基因突变率分别为72．8％、64．9％、59．8％、41．0％和30．7％，多基因突变株达76．0％，结核杆菌耐药基因突变与耐药水平联系密切，多数结核分支杆菌耐药基因突变易发生在高耐药区，也有少数基因突变发生在低耐药区。根据药敏试验和耐药基因检测，耐多药结核病人6个月治愈率分别达到54．8％和62．8％，治疗效果满意，两种方法没有显著性差异（P＞0．05）。结论：耐药基因检测指寻治疗是一种新探索，PCR－SSCP方法敏感，特异，可以快速检测结核菌rpoB,katG,rpsL,pncA和embB耐药基因突变，可能会成为临床指导用药的好方法。     

天津地区临床分离株结核分枝杆菌分子流行病学研究

目的探讨天津地区结核分枝杆菌临床分离株分子流行病学特征。方法连续收集天津市海河医院2005年8月16日～11月25日就诊病人痰培养阳性的结核分枝杆菌100株，采用间隔区寡核苷酸分型（Spoligotyping）和数目可变串联重复序列（VNTR）两种方法进行基因分型，并运用软件对二者的结果进行分析。依据北京分化支的定义。运用多重和实时定量PCR方法将其区分为W菌/典型北京家族菌株和非典型北京菌株，x^2检验分析两种亚群与患者年龄和耐药性之间的联系。结果排除污染菌株，共对96株结核分枝杆菌临床分离株进行两种方法的基因分型，Spoligotyping结果为91．7％为北京基因型（含3株类北京基因型）结核分枝杆菌（88／96）。VNTR分型可将北京基因型分为60种基因型。在北京分化支结核分枝杆菌中，W菌/典型北京家族菌株占93．2％（82／88）。两种北京分化支亚群与患者年龄和耐药性没有显著性统计学差异（P〉0.05）。结论天津地区结核病患者临床分离的结核分枝杆菌中，北京基因型呈现较为明显的优势。VNTR的分辨率明显高于Spoligotyping。北京分化支的两种亚群在天津地区临床结核病患者中均存在流行，但以W菌/典型北京家族菌株为主。     

氯法齐明抗结核分枝杆菌的体内外活性研究

目的 评价氯法齐明(clofazimine,CLF)在体外及体内抗结核分枝杆菌的活性,为临床应用提供依据.方法 应用微孔板指示剂法,测定氯法齐明对结核分枝杆菌标准株H37Rv及临床分离耐多药结核分枝杆菌(30株)的MIC;建立小鼠静脉感染H37Rv的结核病模型(105 CFU/只),根据氯法齐明的不同剂量,分为3个治疗组:20 mg/kg,每周给药5次(CLF-1组);10 mg/kg,每周给约5次(CLF-2组);20 mg/kg,每周给药2次(CLF-3组);治疗30 d后进行肺、脾组织活菌计数,应用单冈素方差分析,比较氯法齐明在小鼠体内的抗结核分枝杆菌活性.结果 氯法齐明对结核分枝杆菌H37Rv的MIC为0.12～0.24 μg/ml;对30株耐多药结核分枝杆菌临床分离株的MIC为0.12～1.92 μg/ml.在小鼠结核病治疗模型中,3个治疗组的小鼠肺活菌计数分别较空白对照组降低2.92、1.78和1.39 lg CFU,均具有统计学意义(F=74.09,P＜0.01).结论 氯法齐明具有较好的体外及体内抗结核分枝杆菌活性,值得进一步研究.     

INAUGURAL ARTICLE by a Recently Elected Academy Member:Crystal structure and stability of gyrase–fluoroquinolone cleaved complexes from Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) infects one-third of the world’s population and in 2013 accounted for 1.5 million deaths. Fluoroquinolone antibacterials, which target DNA gyrase, are critical agents used to halt the progression from multidrug-resistant tuberculosis to extensively resistant disease; however, fluoroquinolone resistance is emerging and new ways to bypass resistance are required. To better explain known differences in fluoroquinolone action, the crystal structures of the WT Mtb DNA gyrase cleavage core and a fluoroquinolone-sensitized mutant were determined in complex with DNA and five fluoroquinolones. The structures, ranging from 2.4- to 2.6-Å resolution, show that the intrinsically low susceptibility of Mtb to fluoroquinolones correlates with a reduction in contacts to the water shell of an associated magnesium ion, which bridges fluoroquinolone–gyrase interactions. Surprisingly, the structural data revealed few differences in fluoroquinolone–enzyme contacts from drugs that have very different activities against Mtb. By contrast, a stability assay using purified components showed a clear relationship between ternary complex reversibility and inhibitory activities reported with cultured cells. Collectively, our data indicate that the stability of fluoroquinolone/DNA interactions is a major determinant of fluoroquinolone activity and that moieties that have been appended to the C7 position of different quinolone scaffolds do not take advantage of specific contacts that might be made with the enzyme. These concepts point to new approaches for developing quinolone-class compounds that have increased potency against Mtb and the ability to overcome resistance.The causative agent of tuberculosis (TB), Mycobacterium tuberculosis (Mtb), is one of the most important pathogens of humans, second only to the HIV in the number of deaths caused annually (1). Mtb is estimated to latently infect one-third of the world’s population (2), thereby also creating a huge reservoir for future disease. Particularly problematic are cases of multidrug-resistant (MDR)-TB, which is defined as resistance to two primary anti-TB drugs, rifampicin and isoniazid. MDR-TB now represents 3.5% of new TB cases; of these, 9% are classified as extensively drug-resistant (XDR)-TB (1), which is defined as MDR-TB with additional resistance to any fluoroquinolone and one injectable second-line drug (3). TB isolates are also being recovered that are totally drug-resistant (3–5). Thus, controlling TB, in particular drug-resistant TB, is a major health problem.Fluoroquinolones are one of the most successful classes of drugs against bacterial pathogens, accounting for 24% of the $10 billion antibiotic market (6). Fluoroquinolones are also currently receiving considerable attention in the treatment of TB, with two new C8-methoxy derivatives, moxifloxacin and gatifloxacin, currently under evaluation as promising first-line therapeutics (7–10). These compounds have been used to restrict the development of XDR-TB from MDR-TB; however, emerging resistance threatens both first-line and second-line use (11). The widespread testing of fluoroquinolones against TB has revealed considerable variation in efficacy of different drug variants against Mtb. For example, ciprofloxacin is only marginally active, and its early use with Mtb was halted in favor of ofloxacin and levofloxacin (7). These two agents are now proving to be less effective than moxifloxacin and gatifloxacin (7, 10); however, the newest two compounds also exhibit some nonideality. For example, gatifloxacin can elicit side effects such as hypo/hyperglycemia (12), whereas moxifloxacin has potential cardiovascular risks (13). Although recent clinical trials that have included fluoroquinolones as part of an alternative drug regimen have faltered (14), there are prospects for other nonfluoroquinolone molecules to make an impact on the treatment of MDR- and XDR-TB (15, 16). Moreover, a promising recent trial that uses moxifloxacin as part of a three-drug regimen together with pretomanid and pyrazinamide reports superior bactericidal activity against TB and MDR-TB versus current regimens (17). These new data make it clear that new quinolone-class agents, which are also capable of circumventing known resistance mutations, might be useful therapeutic agents in the treatment of TB.Progress is presently being made toward developing new fluoroquinolone derivatives. For example, a methoxy group at fluoroquinolone position C8 (Fig. 1A) increases activity against mycobacteria, particularly resistant mutants (18–20). Indeed, moxifloxacin, along with a more active C8-methyl derivative, retains high inhibitory activity against purified Mtb gyrase even when the enzyme contains commonly acquired fluoroquinolone-resistance substitutions [as described in the accompanying paper by Aldred et al. (21)]. In another example, quinazolinediones (diones) have been shown to have an ability to bypass existing resistance within mycobacteria and other bacterial species (20, 22–24). Thus, there still exist opportunities to design more effective quinolone-class molecules for treatment of TB.Open in a separate windowFig. 1.DNA cleavage by Mtb gyrase induced by fluoroquinolones. (A) Fluoroquinolones tested in this study. The constant quinolone core of each drug is highlighted in orange, numbered as shown around C8-Me-moxifloxacin. (B) DNA cleavage assays with full-length Mtb gyrase, using WT (upper gels) and a GyrA A90S (lower gels) sensitizing mutant. Each fluoroquinolone is titrated against a constant amount of protein (125 nM) and supercoiled (SC) plasmid DNA substrate (12.5 nM). The “no protein” control shows the supercoiled substrate DNA, along with a nicked (N) and linear (L) control lanes. Each gel is representative of triplicate data. (C) Graphical analysis of data in B. The relative amount of DNA cleavage is plotted as an increase in linear product (obtained by densitometry) compared with the zero drug control as a function of drug concentration. (Left) WT. (Right) A90S. Data points and error bars represent the mean and SD of triplicate data, respectively.DNA gyrase is a heterotetrameric (GyrA₂GyrB₂) enzyme that transiently catalyzes dsDNA breaks as it negatively supercoils DNA. Fluoroquinolones prevent the resealing of the ds breaks that normally follows DNA strand passage (25), generating persistent, covalent enzyme–DNA adducts called cleaved complexes. Cleaved-complex formation, which is reversible, blocks bacterial growth; at elevated fluoroquinolone concentrations, release of DNA breaks from the complexes leads to chromosome fragmentation and cell death (26). Although numerous crystallographic studies have defined the primary binding site of fluoroquinolones against gyrase (and against topoisomerase IV, a gyrase paralog) (27–30), there has been debate as to whether fluoroquinolones use a magnesium ion to bridge their interaction with the enzyme (28, 29). Resolving this question is important, because the intrinsic resistance of certain species of gyrase to quinolones (e.g., Mtb) has been proposed to result from natural sequence variation that leads to the loss of a magnesium-ion “bridge” formed between the drug and enzyme (31–33). At the same time, there currently exists no clear chemical explanation as to why certain fluoroquinolone derivatives act with different efficacies against Mtb gyrase.To better understand such questions, we carried out a structural and biochemical analysis of Mtb gyrase in the presence of a panel of four different, clinically used fluoroquinolones (ciprofloxacin, levofloxacin, gatifloxacin, and moxifloxacin) and one new fluoroquinolone derivative (C8-Me-moxifloxacin). X-ray crystallography revealed that Mtb gyrase indeed makes an intrinsically low number of interactions with a magnesium ion that accompanies fluoroquinolone binding, and that the introduction of a drug-sensitizing mutation (GyrA A90S) restores interactions seen in nonresistant gyrase homologs. In vitro, we find that C8-Me-moxifloxacin and moxifloxacin are most effective at promoting cleaved complex formation and inhibiting DNA supercoiling by Mtb gyrase, followed by gatifloxacin, and then ciprofloxacin and levofloxacin; however, crystal structures of the DNA-binding-and-cleavage core of Mtb gyrase with both DNA and drug surprisingly failed to reveal any substantial differences in the contacts formed between the protein and different drugs. Further biochemical investigations using an assay that monitors the stability of preformed cleaved complexes largely corroborated the rank order of inhibition seen in DNA cleavage experiments and moreover resulted in drug efficacy trends that closely accord with clinical effectiveness. Collectively, our data explain why Mtb gyrase is naturally quinolone-resistant and show that the relative activities of existing panels of anti-TB quinolone therapeutics are heavily influenced by base-stacking interactions and have yet to take advantage of direct gyrase contacts to maximize therapeutic potential.