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preface
Mycobacterium nontuberculous (NTM) refers to all mycobacteria except Mycobacterium tuberculosis and Mycobacterium leprosy, also known as environmental Mycobacterium[1
In recent years, with the change of the environment, the update of detection technology and the increased awareness of the disease among doctors, the incidence of NTM lung disease has been increasing, and it is rapidly becoming a serious public health problem, endangering human health
Case after
The patient, female, 56 years old, cough for half a year, aggravated for 3 weeks
The illness is still acceptable to eat, sleep is possible, the stool is as usual, and there is no obvious change in
On 15 June, BNP, PCT, erythrocyte sedimentation rate, myocardial markers, G test, GM test, and aspergillus antibodies were normal, and there were no significant abnormalities
Case studies
Clinical case studies
1.
NTM is widely found in natural environments such as water, soil, and dust, and can be infected by humans and certain animals [2
1.
Mycobacterium avium complex infection in the middle lobe of the right lung and the left tongue lobe is called Lady Wen syndrome
Tradition holds that excessive cough suppression by "noble women" leads to Lady Wen syndrome
1.
Lymphadenopathy is a common clinical symptom caused by proliferation of cells inside the lymph nodes or infiltration of tumor cells due to a variety of causes
Inspection case studies
How can laboratories challenge long-standing confused clinicians for NTM diagnosis?
1.
Various specimens, such as sputum, induced sputum, bronchial irrigation fluid, bronchoalveolar lavage fluid, as well as biopsy tissues such as lungs, lymph nodes, liver, kidneys, and spleen, as well as blood, bone marrow secretions, body fluids, and feces, can be tested
2.
As with Mycobacterium tuberculosis, smear microscopy with fluorescent staining is recommended
3.
Isolate culture
The positive rate of acid-resistance staining is very low and cannot be distinguished from tuberculosis, and culture is still one
of the most sensitive techniques for detecting NTM.
Both solid and liquid cultures can be used for the culture of NTMs, and a combination of the two is recommended to improve the positive rate
of culture.
Liquid cultures have a higher positive rate, especially for fast-growing Mycobacteria
.
The advantage of solid culture is that it can directly observe colony morphology and growth rate, which is easy to quantify bacteria and facilitates the detection of the presence of
mixed infections.
Obtaining a large number of colonies on solid medium often indicates clinical value, and if only sporadic colonies are present, it mostly means contamination or transient colonization
without clinical value.
The culture of some NTM strains requires special media, or a specific culture temperature, or a prolonged culture time, such as bloodthirsty Mycobacteria can grow on the medium containing iron ions, the subspecies of Mycobacterium ornithus subspecies needs to add mycobacterium mycobacterium to the medium, and Mycobacterium ulcerans needs to add egg yolks to the medium to be more successfully cultured
.
The optimal growth temperature for Mycobacterium ulcerans is 25 to 33 °C, mycobacterium sea is 28 to 30 °C, and the best growth temperature for Mycobacterium toad is 45 °C
.
For samples of joint fluid, skin, and bone tissue, it is recommended to conduct parallel cultures of 2 temperature gradients of 28°30°C and 35°37°C to improve the positivity rate
.
Slow-growing mycobacteria culture typically takes 2 to 3 weeks to form visible colonies on solid medium, while Mycobacterium ulcerans takes 8 to 12 weeks
.
The patient underwent sputum cultures during admission, all of which suggested normal throat flora growth, until BALF's NTM-PCR (+) and metagenomic sequencing suggested intracellular Mycobacterium avium, and we were able to catch the "real culprit"
.
Looking back at this case, the media currently used in our laboratory cannot cover all TTM growth, and there is a lack of culture and identification conditions for rare NTM cultures
.
This also suggests that we should improve the awareness of the culture of NTM, such as extending the culture time, adding special media, etc.
, expanding the scope of laboratory testing, and providing more basis
for clinical pathogen diagnosis.
4.
Strain identification
There are too many types of NTM, and those who cultivate positive try to identify the bacteria, otherwise it is difficult to accurately treat
the next step.
The purpose of strain identification is to accurately diagnose NTM disease, including the determination of the clinical relevance of NTM, and because of the different sensitivities of different strains to drugs, strain identification is also of great value
to the formulation of treatment plans.
Clinical significance is greater
for multiple samples or samples from multiple sites isolated into the same strain, when a large number of NTMs are detected (e.
g.
, if the specimen is positive for smear or a large number of colonies are cultured), from sterile sites or from blood to NTMs.
However, not all isolated NTMs need to be identified by subsequent strains, such as when culturing and isolating mycobacteria with low bacteria volume and color production, it can almost be determined that it is not a pathogenic bacteria, and further identification of bacteria is not necessary
.
5.
Molecular diagnostic technology
Next generation sequencing (NGS): Is the highest resolution means of strain identification and can also be used to track transmission
in specific populations caused by NTM.
With the increasing popularity of NGS technology and the reduction of costs, it will play an increasingly important role
in the diagnosis of NTM disease.
mNGS technology is a new nucleic acid detection method for the detection of unknown and difficult pathogenic microorganisms, whether it is for emerging infectious diseases or clinically difficult infectious diseases
.
It has unmatched advantages over other testing techniques for its unbiasedness, rapid test results, and suitability for a variety of clinical specimens
.
But clinical infectious etiology diagnosis is challenging work
.
mNGS only detects nucleic acids (including DNA and RNA) in the sample, reflects the patient's true infection status, and needs to combine the test results with the clinical situation and check and screen
.
mNGS technology detects nucleic acids, and cannot simply equate nucleic acids with pathogenic microorganisms
.
However, no technology can solve all the problems, and mNGS technology has not gotten rid of the limitations of nucleic acid detection, and the interpretation of test results needs to be combined with clinical
.
The patient underwent metagenomic sequencing of BALF, which coincided with the results of conventional PCR techniques in the
laboratory.
Routine PCR results in the laboratory suggest NTM-PCR(+), but the classification
of the strains is not possible.
The mNGS technique quickly filled the gap in the laboratory's routine work, and the analysis results suggested an intracellular mycobacterial complex
.
Both technical methods prompted NTM, providing a solid laboratory basis for clinical diagnosis
.
Knowledge development
1.
Classification of non-tuberculous mycobacteria
According to the growth rate of NTM, the Bergy's manual of systematic bacteriology (Bergy's manual of systematic bacteriology) divides it into two categories: fast-growing and slow-growing, and this classification method
is currently used internationally.
The Runyon taxonomy divides it into 4 groups according to the growth temperature, growth rate, colony morphology and the relationship between pigment production and light response in the test tube, the first 3 groups are actually slow-growing Mycobacterium, while the 4th group is rapid-growing Mycobacterium, and the two classifications are currently combined
in China.
Group I: Photochromogens: On solid medium, colonies are pale yellow when they are not seen and turn yellow or orange
after illumination.
This group is dominated
by Mycobacterium Kansasis, Mycobacterium sea and Mycobacterium ape.
Group II: Scotochromogens: Colonies produce yellow or red
when there is no light.
This group is dominated
by Mycobacterium fistula, Mycobacter gordonii and Mycobacter surga.
Group III: non-photochromogens: Colonies do not produce pigmentation regardless of light or not, and may appear off-white or yellowish
.
This group includes Mycobacterium avium complex, Bloodthirsty Mycobacterium, Mycobacterium ulcerans, Mycobacterium toad, Mycobacterium Marmo, Mycobacterium earth and Mycobacterium gastricus
.
Group IV: Rapidly growing mycobacteria (RGM): there are colonies visible to the naked eye within 3 to 5 days, and most of them grow vigorously
within 1 week.
This group includes Mycobacterium abscess complex, occasional mycobacteria, Mycobacterium turtle, Mycobacterium Margaret, Mycobacterium exogenes, Mycobacterium diuresis and Mycobacterium cow
.
2.
Differential diagnosis of non-tuberculous mycobacterium lung disease and tuberculosis
Non-tuberculous mycobacterial disease is mainly diagnosed with tuberculosis, clinical manifestations and imaging tests are very similar, the clinical manifestations of patients include cough, sputum production, a small amount of cough, and patients with a wide range of lesions can have chest tightness and systemic symptoms, such as low-grade fever, night sweats, and wasting
.
Imaging features are also very similar, with patients presenting with multiple patchy shadows
in the lungs.
The imaging manifestations of non-Mycobacterium tuberculosis lung disease are different from those of tuberculosis, non-mycobacterium tuberculosis lung disease is often secondary to structural lung disease, patients are prone to bronchiectasis, congenital lung cysts, cavities are generally thin-walled cavities, while tuberculosis is dominated by thick-walled cavities
.
However, the two are mainly confirmed by the relevant tests related to the identification of mycobacterial species, and cannot be confirmed by imaging tests and clinical manifestations
.
NTM lung disease has respiratory symptoms and/or systemic symptoms, chest imaging examination found cavitation shadows, multifocal bronchiectasis and multiple small nodular lesions, etc.
, has ruled out other lung diseases, under the premise of ensuring that the specimen is free of exogenous contamination, one of the following conditions can be diagnosed as NTM lung disease: (1) 2 sputum specimens sent separately are positive for NTM culture and identified as the same pathogen, and/or NTM molecular biology tests are the same consistent bacteria; (2) Bronchial irrigation fluid or bronchoalveolar lavage fluid NTM culture and (or) molecular biology test once positive; (3) Characteristic changes in histopathology of mycobacteriosis (positive for granulomatous inflammation or acid-resistance staining) found on transbronchoscopic or other pulmonary biopsy, and positive for NTM culture and/or molecular biology; (4) Characteristic changes in histopathology of mycobacteriosis (positive granulomatous inflammation or antacid staining) were found by bronchoscopic or other pulmonary biopsy, and the NTM culture and/or molecular biology test in sputum specimens, bronchial irrigation fluid or bronchoalveolar lavage fluid once or more were positive
.
3.
Are non-tuberculous mycobacteria infectious?
In 2000, China formulated the "Guidelines for the Diagnosis and Treatment of NTM Diseases", and there have been no guidelines for nearly 20 years since then, until November 2020, the "Chinese Journal of Tuberculosis and Respiratory Disease" published the "Guidelines for the Diagnosis and Treatment of Non-Tuberculous Mycobacteriosis (2020 Edition)", which shows that the diagnosis and treatment of this disease is difficult, and the progress in these years is slow
.
A disease that can be transmitted is not necessarily an infectious disease
.
Infectious disease is a human-defined sociological concept, usually this disease can be directly contacted by infected individuals, the body fluids and excrement of infected people, objects contaminated by infected people, can be transmitted by air, water, food, contact, soil, vertical transmission (mother-to-child transmission), body fluid transmission, fecal mouth transmission, etc
.
Sputum can be cultured with multidrug-resistant Klebsiella pneumoniae, and hospitals should implement bedside contact isolation to avoid cross-infection
.
If Klebsiella pneumonia is not contagious, why should doctors wash their hands? However, many infectious diseases are very contagious, and healthy people with normal immunity will not be infected, let alone spread globally like the new crown virus, so such infectious diseases are generally not infectious diseases
.
In recent years, studies have shown that mycobacterium abscess can be transmitted from person to person, especially in patients with cystic pulmonary fibrosis, possibly through aerosols or contaminants
.
However, in many countries, NTM disease is still not included in infectious disease management and does not need to be reported
.
Case summary
The patient in this case experienced cough symptoms for up to half a year before admission, aggravated for 3 weeks, cough before going to bed and in the morning, aggravated after activity, occasionally coughed up sputum, white foamy sputum, no fever, night sweats, no chest pain, hemoptysis, wheezing
.
Physical examination: clear speech, no cyanosis on the lips, stable breathing, thick breathing sounds in both lungs, crackling sounds can be heard in both lungs, dry and wet rales are not heard, and there is no pleural friction sound
.
Transferred to the respiratory department of our hospital for diagnosis and treatment, the disease can eat and sleep, the stool is as usual, and there is no obvious change in
weight.
Outer hospital lung CT shows interstitial pneumonia changes in both lungs, and a ground glass density foci of the upper lobe of the left lung, which is admitted to our hospital
for further diagnosis and treatment.
The illness is still acceptable to eat, sleep is possible, the stool is as usual, and there is no obvious change in
weight.
The results of lung CT suggested: 1.
Interstitial inflammation of both lungs; 2.
Nodular foci of the middle lobe of the right lung and the left lung of the tongue, and the preliminary diagnosis was interstitial pneumonia
.
Sputum cultures suggest normal throat flora growth
.
Pulmonary function: pulmonary ventilation function is normal; The diffusion function is slightly reduced
.
Cefminol anti-infective therapy is given, supplemented by cough suppressant, acid-suppressing and gastric treatment
.
Ultrasound of the right axillary shows abnormal right axillary lymph nodes, and no abnormally enlarged lymph nodes are seen in the left
axillary.
PETCT results suggested that there were multiple lymph nodes with increased FDG metabolism in the left neck, right armpit, right hilar, mediastinum and around the pancreas, and splenomegaly with increased FDG metabolism
.
Histopathological results of lymph nodes suggest granulomas visible, and no tumor cells
are seen.
Tracheoscopic related examination, microscopic visible bronchoppurulent discharge of the upper lobe of the right lung, patency of the lumen after suction, and alveolar lavage
in each segment of the upper lobe of the right lung.
Bronchoalveolar lavage fluid was sent to the laboratory for pathogenic related testing, and the next day the laboratory fed back TB-PCR (-), NTM-PCR (+), acid-antacid staining (+), and metagenomic sequencing mNGS results suggested: intracellular Mycobacterium ornithus complex
.
Final diagnosis: intracellular mycobacterium pulmonary disease, transferred to the city tuberculosis hospital for continued treatment
.
The patient is currently in good condition and follow-up treatment is proceeding
smoothly.
Mycobacterium avium is the most common species of NTM on all continents in the world, and is also the main strain of NTM lung disease, lymphadenopathy and disseminated NTM disease
.
Some anti-mycobacterial drugs have strong antibacterial activity against intracellular mycobacteria, such as macrolides, rifamycin, quinolones and aminoglycosides, among which macrolides have a definite efficacy [3].
It is well known that epidemiological research on NTM disease is difficult, and the exact data and data of different countries or regions are difficult to grasp, because the report of NTM disease is not mandatory in most countries, and it is difficult to distinguish between NTM infection and incidence, and the incidence and prevalence of NTM infection vary significantly in different studies
.
However, from the available data, the incidence and prevalence of NTM disease show an increasing trend in some countries and regions, even exceeding the incidence and prevalence of tuberculosis
.
The results of the study showed that NTM may be contained in soils (acid pine forests, coastal swamp soils), indoor swimming pools, hot tubs, coastal swamp drainage systems, indoor humidifiers and showers, and dust from rural, garden, and potted
soils.
For example, intracellular mycobacteria have a certain tolerance to disinfectants and heavy metals, making it possible to survive
in the tap water system.
The patient in this case is an elderly female patient with menopause and a preference for bathing in the public bath, which increases the chance of infection with NTM
.
Preventing the spread of NTM from the environment to people, detecting infections earlier, and using drugs precisely after infection are important challenges for clinicians
.
The clinical diagnosis and treatment of patients in this case can provide a reference for future clinicians' treatment to improve the level
of diagnosis and treatment of NTM disease.
Expert reviews
NTM lung disease is the most common type of NTM disease, and the incidence of disease in China has shown a significant increase in recent years
.
Early identification and correct diagnosis are essential
to reduce transmissibility, improve cure rates, and reduce drug resistance rates.
The clinical presentation in patients with NTM lung disease is multifaceted, and some patients have no obvious symptoms
at the time of onset.
Therefore, early identification and health management of NTM risk factors and high-risk groups is an important part
of early diagnosis of NTM disease.
At present, it is considered that the high risk factors for NTM disease include host factors, drug factors, and environmental factors
.
Host factors mainly refer to people with underlying lung disease and immunodeficiency, and studies have shown that rheumatoid arthritis, gastroesophageal reflux, malnutrition, and vitamin D deficiency are also risk factors
for NTM disease.
The drug factor refers mainly to patients
who use glucocorticoids and immunosuppressants.
Environmental factors include indoor aerosols, soil dust and so on
.
Positive results of pathogenic microbial culture and molecular biology strain identification of respiratory or lung tissue specimens are necessary conditions
for the diagnosis of NTM lung disease.
Combined solid culture and liquid culture helped to increase the positive rate
of NTM culture.
The treatment regimen varies from NTM subspecies, so NTM strain identification is a guarantee
of precision treatment.
NTM is widespread in the environment, but the pathogenicity of different types of NTMs is different
.
In general, THE ISOLATION OF CLINICAL SAMPLES OF MAC, MYCOBACTERIUM ABSCES, MYCOBACTERIS KANSAS, MYCOBACTER MALMOX, MYCOBACTERIUM TOAD, MYCOBACTERIUM SCAB, MYCOBACTERIUM SCAB, MYCOBACTERIUM TURTLE, MYCOBACTERIUM OCCASIONALLY, AND MYCOBACTERIUM SEA, ETC.
ARE ISOLATED IN CLINICAL SAMPLES, WHILE THE MYCOBACTERIUM GORDONUM, MYCOBACTERIUM MUCINE, MYCOBACTERIUM NON-COLORUM, AND MYCOBACTERIUM SOIL ARE GENERALLY NON-PATHOGENIC OR WEAKLY PATHOGENIC, AND THE ISOLATES MAY BE CONTAMINATED OR TRANSIBITATED [4
].
Strengthen health education, understand the harm and transmission of NTM disease, and develop good hygiene habits
.
Timely detection and cure of the source of infection, reduce contact with patients with NTM disease, and do a good job in protecting
human-to-human transmission.
Increases the body's resistance and reduces susceptibility
to NTM.
It is worth noting that it is crucial to prevent NTM infection in hospitals, and the key is to do a good job in disinfecting hospital water and medical equipment
.
The preparation of disinfectant must be carried out in strict accordance with the requirements and standardized operation
.
While doing a good job in prevention, we should also pay attention to strengthening the detection of NTM, carry out NTM strain identification and drug sensitivity tests, and improve the level
of diagnosis and treatment of NTM disease.
References:
1.
Expert consensus on the principle of Chinese translation of mycobacterial species[J].
Chinese Journal of Tuberculosis and Respiratory.
2018 (07)
2.
Guidelines for the diagnosis and treatment of non-tuberculous mycobacterial disease (2020 edition).
Chinese Journal of Tuberculosis and Respiratory Disease.
2020.
43(11):918-946.
3.
Daley CL,Iaccarino JM,Lange C,et al.
Treatment of nontuberculous mycobacterial pulmonary disease:an official ATS/ERS/ESCMID/IDSA clinical practice guideline.
Eur Respir J.
2020.
56(1)
4.
Re-negotiate the harm of non-mycobacterium tuberculosis[J].
SHA Wei,XIAO Heping.
Chinese Journal of Tuberculosis and Respiratory.
2018 (02)