Ecology of the respiratory tract microbiome

A thriving multi-kingdom microbial ecosystem inhabits the respiratory tract: the respiratory tract microbiome (RTM). In recent years, the contribution of the RTM to human health has become a crucial research aspect. However, research into the key ecological processes, such as robustness, resilience, and microbial interaction networks, has only recently started. This review leans on an ecological framework to interpret the human RTM and determine how the ecosystem functions and assembles. Specifically, the review illustrates the ecological RTM models and discusses microbiome establishment, community structure, diversity stability, and critical microbial interactions. Lastly, the review outlines the RTM responses to ecological disturbances, as well as the promising approaches for restoring ecological balance.     

呼吸道合胞病毒下呼吸道感染与患儿鼻咽部定植菌群的关系

研究鼻咽部定植菌群与呼吸道合胞病毒（RSV）下呼吸道感染患儿病情特点的关系,从微生态角度寻找预防或治疗RSV下呼吸道感染的理论依据。

回顾性分析我院2020年8月1日到2021年10月31日收治的RSV下呼吸道感染患儿,根据鼻咽部分泌物培养结果分为阴性组、阳性组,对比两组患儿临床特点及潮气肺功能检测结果。

1 090例RSV下呼吸道感染患儿中培养阳性者290例（26.58%）,中位数年龄为8个月（3.24个月,14个月）。各年龄层患儿鼻咽部菌群定植率及定植菌群差异有统计学意义（均P＞0.05）。各年龄层患儿鼻咽部定植菌均以肺炎链球菌为主。金黄色葡萄球菌、大肠埃希菌及卡他莫拉菌定植的患儿发热率较低。与喘息相关定植菌为肺炎链球菌、流感嗜血杆菌及金黄色葡萄球菌。与重症相关的定植菌为肺炎链球菌、流感嗜血杆菌、金黄色葡萄球菌、大肠埃希菌。与大叶性肺炎发生相关的只有肺炎链球菌。肺炎链球菌及流感嗜血杆菌定植的达峰容积比（Z = −4.12,P＜0.001;Z = −1.99,P = 0.046）与达峰时间比（Z = −4.24,P＜0.001;Z = −1.98,P = 0.047）均低于阴性组。

特定鼻咽部定植菌的存在会影响RSV下呼吸道感染患儿的临床表现,同时降低婴幼儿潮气肺功能检测结果。

     

大气细颗粒物影响小鼠上、下呼吸道菌群结构

探讨影响人类健康的大气细颗粒物干扰小鼠上呼吸道和下呼吸道菌群定植及可能机制。

建立大气细颗粒物暴露的小鼠模型。BALB/c小鼠随机分为PM2.5暴露组和对照组, 每组10只。收集上呼吸道和下呼吸道灌洗液进行菌群测序分析, 并进行肺组织HE染色, 观察肺组织形态改变情况。

大气细颗粒物暴露小鼠, 破坏肺组织气血屏障结构。16S rRNA测序证实了上、下呼吸道微生态的组成因大气细颗粒物暴露而发生改变。明确了与细颗粒物暴露组相比较, 对照组的噬几丁质菌定植的丰度较高。

大气细颗粒物影响小鼠上、下呼吸道的微生态结构。研究发现的差异菌群改变将会成为新的肺损伤治疗的潜在菌群。

     

Characterization of the viral microbiome in patients with severe lower respiratory tract infections, using metagenomic sequencing

The human respiratory tract is heavily exposed to microorganisms. Viral respiratory tract pathogens, like RSV, influenza and rhinoviruses cause major morbidity and mortality from respiratory tract disease. Furthermore, as viruses have limited means of transmission, viruses that cause pathogenicity in other tissues may be transmitted through the respiratory tract. It is therefore important to chart the human virome in this compartment. We have studied nasopharyngeal aspirate samples submitted to the Karolinska University Laboratory, Stockholm, Sweden from March 2004 to May 2005 for diagnosis of respiratory tract infections. We have used a metagenomic sequencing strategy to characterize viruses, as this provides the most unbiased view of the samples. Virus enrichment followed by 454 sequencing resulted in totally 703,790 reads and 110,931 of these were found to be of viral origin by using an automated classification pipeline. The snapshot of the respiratory tract virome of these 210 patients revealed 39 species and many more strains of viruses. Most of the viral sequences were classified into one of three major families; Paramyxoviridae, Picornaviridae or Orthomyxoviridae. The study also identified one novel type of Rhinovirus C, and identified a number of previously undescribed viral genetic fragments of unknown origin.     

吸烟对人体呼吸道厌氧菌的影响

目的探究吸烟对人体内的呼吸道微生态的影响。方法采用细菌鉴定及药敏分析系统鉴定健康吸烟者与非吸烟者,患下呼吸道感染吸烟与非吸烟者咽后壁分泌物的细菌密度、细菌数量及种类的改变。结果在健康吸烟者与患有下呼吸道感染患者的呼吸道中,厌氧菌明显升高,其中以韦荣菌和消化链球菌为主。结论吸烟可使人体的呼吸道微生态中细菌密度,细菌数量及种类的改变,易患呼吸系统疾病。     

吸烟对人体呼吸道菌群的影响

目的 探讨吸烟对人体呼吸道需氧菌群的影响.方法 采用细菌鉴定及药敏分析系统鉴定健康吸烟者与非吸烟者,患下呼吸道感染与非吸烟者咽后壁分泌物菌群的变化.结果 患下呼吸道感染者较健康者细菌检出率明显增高(P＜0.05),患下呼吸道感染吸烟者细菌检出率较非吸烟者细菌检出率明显增加(P＜0.05),健康吸烟者细菌检出率较非吸烟者明显增加(P＜0.05).结论 吸烟可导致人体口咽部细菌增高致病菌增加以及细菌量的变化,导致呼吸道微生态失调.     

Discriminant analysis of lower respiratory tract components associated with cigarette smoking, based on quantitative sputum cytology

Cigarette smoking is the major contributor to chronic bronchial irritation, and tobacco smoke is the most common bronchial carcinogen. A newly developed quantitative sputum cytology test was applied to three-day-pooled, spontaneously produced sputa collected from 349 cigarette smokers and 93 patients who had never smoked; the samples were processed by the Saccomanno technique. In addition to identifying malignant and precancerous cells in sputum, this test identifies eight morphologic parameters of bronchial irritation. Cross-sectional analysis of the cytologic data indicated that, within a 99% confidence interval, cigarette smokers had statistically significantly higher levels of all eight components, producing a unique profile. Discriminant analysis revealed that pigmentation of macrophages, alveolar macrophages and mucous spirals (in that order) were the leading components differentiating smokers from nonsmokers. This new quantitative sputum cytology test correlates with the model of lung carcinogenesis. The detection of proliferative cellular alterations, such as benign and early atypical metaplasia, represent the detection of chronic persistent irritation, which is considered to be a reversible change. The ability to monitor individual responses to toxic inhalants (before the development of irreversible disease) by utilizing a noninvasive test represents a unique opportunity to impact the historic course of lung carcinogenesis.     

COPD的下呼吸道微生态研究进展

新一代测序技术的发展,拓展了人类对肺微生物组的认识,下呼吸道微生物组与呼吸系统疾病的关系日益受到关注。下呼吸道微生物组包含平衡的共生菌群和致病菌群,在慢性阻塞性肺疾病(chronic obstructive pulmonary disease,COPD)中,其疾病的严重性、炎症程度或抗生素等治疗因素均会影响肺部微生物群稳态,下呼吸道微生物群落的改变又可能通过操纵炎症或免疫过程,对COPD的病程产生影响。据此,本文通过回顾近年来对COPD下呼吸道微生态的研究,总结了COPD患者的下呼吸道微生态特征及菌群定植情况,探讨下呼吸道微生态与COPD发病机制之间的相关性和治疗对策,希望为COPD的诊断和治疗提供新思路。     

Selective infection of lower respiratory tract by respiratory viruses in children with recurrent respiratory tract infections.

Thirty preschool children presenting with recurrent respiratory infections and their unaffected siblings were observed prospectively for a year. The index children experienced more episodes of acute respiratory infection than their siblings. Respiratory viruses were the major cause of respiratory infections. The index children had lower respiratory tract disease, predominantly wheeze, during 34% of proved respiratory virus infections compared with 11% of such infections experienced by the control children (p less than 0.02). Atopic children had an increased tendency to wheeze that did not reach significance, but atopy was not associated with increased susceptibility to respiratory infections.     

The role of neutrophils in the upper and lower respiratory tract during influenza virus infection of mice

Background

Neutrophils have been shown to play a role in host defence against highly virulent and mouse-adapted strains of influenza virus, however it is not clear if an effective neutrophil response is an important factor moderating disease severity during infection with other virus strains. In this study, we have examined the role of neutrophils during infection of mice with influenza virus strain HKx31, a virus strain of the H3N2 subtype and of moderate virulence for mice, to determine the role of neutrophils in the early phase of infection and in clearance of influenza virus from the respiratory tract during the later phase of infection.

Methods

The anti-Gr-1 monoclonal antibody (mAb) RB6-8C5 was used to (i) identify neutrophils in the upper (nasal tissues) and lower (lung) respiratory tract of uninfected and influenza virus-infected mice, and (ii) deplete neutrophils prior to and during influenza virus infection of mice.

Results

Neutrophils were rapidly recruited to the upper and lower airways following influenza virus infection. We demonstrated that use of mAb RB6-8C5 to deplete C57BL/6 (B6) mice of neutrophils is complicated by the ability of this mAb to bind directly to virus-specific CD8⁺ T cells. Thus, we investigated the role of neutrophils in both the early and later phases of infection using CD8⁺ T cell-deficient B6.TAP^-/- mice. Infection of B6.TAP^-/- mice with a low dose of influenza virus did not induce clinical disease in control animals, however RB6-8C5 treatment led to profound weight loss, severe clinical disease and enhanced virus replication throughout the respiratory tract.

Conclusion

Neutrophils play a critical role in limiting influenza virus replication during the early and later phases of infection. Furthermore, a virus strain of moderate virulence can induce severe clinical disease in the absence of an effective neutrophil response.     

吸烟对健康人群唾液微生物组的影响

【背景】吸烟是重要的公共健康问题,可能通过改变唾液微生物组影响口腔健康甚至全身健康。【目的】探究吸烟对中国健康人群唾液微生物组的影响,为研究吸烟在疾病发展中的作用提供线索。【方法】收集167位健康个体的唾液样本,利用HiSeq高通量测序技术对样品中16S rRNA基因V3-V4区进行测序,分析比较吸烟者和非吸烟者的唾液微生物组的多样性、群落结构和物种丰度,并预测功能组成,探究吸烟对唾液微生物组的影响。【结果】吸烟对唾液微生物组的α多样性影响较小,但是在微生物组组成上,吸烟组中奈瑟菌属、艾肯菌属、二氧化碳嗜纤维属等细菌明显减少,而普雷沃氏菌属、韦荣氏球菌属、放线菌属、奇异菌属和巨球型菌属等细菌明显增加。在功能上,PICRUSt的预测结果显示吸烟组富集不依赖氧气的代谢功能,非吸烟组富集需要氧气参与的代谢功能。【结论】吸烟可能会通过改变唾液微生物组,造成微生物组功能途径的变化,进而引发与吸烟相关的疾病。     

Cyclooxygenase metabolites are compartmentalized in the human lower respiratory tract

Cyclooxygenase metabolites of arachidonic acid are thought to play an important role in the regulation of diverse physiological functions in the lung. Although the concentration of these metabolites required to have effects is several orders of magnitude greater than the concentration of these mediators in the blood, it has been postulated that local concentrations within tissues are much higher. In a direct test of this hypothesis, the concentrations of the cyclooxygenase products of arachidonic acid including prostaglandin (PG) E, PGF2 alpha, 6-keto-PGF1 alpha, and thromboxane B2, were measured in a specialized tissue compartment, the epithelial surface of the lower respiratory tract. The concentration of these mediators within this compartment was 50- to 80-fold greater than concurrent blood levels and are sufficient to likely have physiological effects. Thus the epithelial surface of the lower respiratory tract represents a specialized compartment with high local levels of cyclooxygenase products of arachidonic acid.     

Catecholamine- and acetylcholinesterase-containing nerves in human lower respiratory tract

Summary The innervation of human lower respiratory tract was studied with special emphasis on airways with sodium-potassium glyoxylic acid (SPG) and acetylcholinesterase (AChE) methods to demonstrate catecholamine-containing and acetylcholinesterase-containing nerve fibers. AChE-method revealed a rich network of cholinesterase positive nerves both inside the bronchial glands where they run around and between the acini, and the airway smooth muscle from secondary bronchi to terminal bronchioli. No AChE-positive fibers were found in connection with the blood vessels or within the epithelium of bronchi or bonchioli. The AChE-positive nerve fibers in bronchial smooth muscle greatly outnumbered those containing catecholamine. The SPG-method revealed the presence of adrenergic nerves from the level of secondary bronchi to that of terminal bronchioli. These nerve fibers were most abundant in bronchial glands, where their amount was equal and distribution similar to those of AChE-containing nerve fibers. Outside the glands adrenergic fibers were constantly seen in connection with the bronchial blood vessels in connective tissues surrounding bronchi. A few nerve fibers were also present in airway smooth muscle from the secondary bronchi to terminal bronchioli.     

Microbial flora variations in the respiratory tract of mice.

A stable microbial system in the respiratory tract acts as an important defense mechanism against pathogenic microorganisms. Perturbations in this system may allow pathogens to establish. In an ecological environment such as the respiratory tract, there are many diverse factors that play a role in the establishment of the indigenous flora. In the present work we studied the normal microbial flora of different areas of the respiratory tract of mice and their evolution from the time the mice were born. Our interest was to know which were the dominant groups of microorganisms in each area, which were the first capable of colonizing and which dominated over time to be used as probiotic microorganisms. Our results show that Gram negative facultatively anaerobic bacilli and strict anaerobic microorganisms were the last ones to appear in the bronchia, while aerobic and Gram positive cocci were present in all the areas of the respiratory tract. The number of facultative aerobes and strict anaerobes were similar in the nasal passage, pharynx instilled and trachea, but lower in bronchia. The dominant species were Streptococcus viridans and Staphylococcus saprophyticcus, followed by S. epidermidis, Lactobacilli and S. cohnii I which were present on every studied days but at different proportions. This paper is the first part of a research topic investigating the protective effect of the indigenous flora against pathogens using the mice as an experimental model.     

院内呼吸科下呼吸道感染病原学及耐药性结果分析

目的分析大连市第五人民医院2009年呼吸科送检1160例次痰标本细菌检查情况,为临床提供感染数据以及合理使用抗生素提供依据。方法按常规方法将临床送检的痰标本进行细菌培养、分离。使用BIO-FOSUN微生物鉴定药敏分析系统进行细菌鉴定及抗生素敏感试验。结果一年来共收痰标本1160例次,检出病原菌253例次,阳性率为21.8%。病原菌依次为肺炎克雷伯菌83株,铜绿假单胞菌57株,乙酸钙不动杆菌29株,阴沟肠杆菌14株,粘质沙雷菌13株,大肠埃希菌10株,,金黄色葡萄球菌5株等。大多数革兰阴性菌对亚胺培南较敏感,对头孢菌素类均有不同程度的耐药。革兰阳性球菌对万古霉素敏感,对其他抗生素有不同程度的耐药。结论该院呼吸科患者下呼吸道感染以革兰阴性杆菌为主,常见病原菌耐药性未呈现高水平、多重耐药情况。提示临床医生应与检验科密切配合,关注细菌感染情况,根据药敏试验结果合理选用抗生素。     

Facilitated expansion of pneumococcal colonization from the nose to the lower respiratory tract in mice preinfected with influenza virus.

A strain of Streptococcus pneumoniae, when inoculated intranasally in 2 microl of suspension into BALB/c mice preinfected with influenza virus, colonized first in the nose, and several days thereafter also colonized significantly in the trachea and lungs with purulent inflammation. Pneumoccocal colonization was also observed in the noses of normal mice after the same bacterial inoculation, but not apparently in the lower respiratory tract. These results suggest that pneumococcal infection may develop from the upper to the lower respiratory tract as a possible sequence preferentially in influenza virus-infected subjects.     

下呼吸道感染流行菌及其耐药性

目的:研究本院下呼吸道感染流行菌及其耐药性,为合理使用抗生素提供依据.方法:对下呼吸道感染流行菌进行统计分析.结果:检出病原菌716株,革兰阴性杆菌为主要菌种,而G杆菌中铜绿假单胞菌占13.69%,肺炎克雷伯菌占8.8%,不动杆菌属占8.4%.除嗜麦芽寡养单胞菌外,G杆菌对亚胺培南、阿米卡星敏感性高.G 球菌以表皮葡萄球菌、溶血葡萄球菌为主要菌种,耐甲氧西林表皮葡萄球菌(MRSS)的检出率为85.3%,耐甲氧西林溶血葡萄球菌的检出率为91.6%.真菌分离呈增多趋势,占所有病原菌的23.28%.结论:下呼吸道感染流行菌药敏显示耐药水平较多,临床医生应根据药敏结果合理选择抗生素,以利于疾病的治疗.     

下呼吸道非发酵菌的分布及其耐药性研究

目的了解2000～2003年安微医科大学第一附属医院下呼吸道非发酵菌的分布及其耐药性,为临床合理选用抗生素提供依据。方法细菌鉴定、药敏试验和诱导β-内酰胺酶(IB)检测采用WalkAway-40全自动微生物分析仪,超广谱β-内酰胺酶(ESBb)检测采用纸片确证试验。结果2000～2003年下呼吸道非发酵菌的检出率为28．2％。其构成以铜绿假单胞菌为最多(56．6％),其次为洛菲不动杆菌(9．3％)、嗜麦芽窄食单胞菌(8．6％)和鲍曼或溶血不动杆菌(8．5％)。对复方新诺明,嗜麦芽窄食单胞菌的耐药率最低(14．3％)。对头孢他啶、亚胺培南、哌拉西林或他巴坦,铜绿假单胞菌的耐药性较低,分别为20．3％、20．8％和28．3％;洛菲不动杆菌耐药率比鲍曼或溶血不动杆菌普遍偏低。铜绿假单胞菌ESBLs和IB的检出率分别为15．7％和84．2％。结论下呼吸道非发酵菌的检出率较高,主要为铜绿假单胞菌。非发酵菌耐药现象严重,不同非发酵菌的耐药谱差异较大。铜绿假单胞菌IB发生率高。     

PM_(2.5)暴露对ICR小鼠上呼吸道菌群的影响

目的建立长期暴露高浓度PM_(2.5)的ICR小鼠动物模型,探索长期暴露于PM_(2.5)对呼吸道微生物组成的影响。方法以沈阳市黄河北大街为采样点,收集空气中细颗粒物,用浓度为200μg/mL的PM_(2.5)建立染尘染毒ICR小鼠模型,染尘周期为6个月,用无菌棉拭子进行咽拭取材后,使用靶向16S rRNA基因的V3-V4区域进行高通量测序来研究它们的微生物群。结果染尘组小鼠呼吸道菌群OTU总数大于对照组,物种分类分析后主要优势菌门为拟杆菌门、厚壁菌门和变形菌门。2组差异物种分析得出拟杆菌门和螺旋体菌门丰度染尘组大于对照组,变形菌门丰度染尘组小于对照组。此外α多样性分析Chao1指数和Ace指数差异具有统计学意义(均P0.05)。结论染尘组与对照组小鼠呼吸道菌群具有明显差异性,证实长期高浓度PM_(2.5)暴露导致呼吸道菌群失调。