Routinely detected indicators in plasma have a predictive effect on the identification of HIV-infected patients with non-tuberculous mycobacterial and tuberculous infections
- Ren-tian Cai†1, 2,
- Feng-xue Yu†3,
- Zhen Tao1,
- Xue-qin Qian4,
- Jun Chen2, 7Email author and
- Hong-zhou Lu1, 5, 6, 7Email author
© The Author(s). 2017
Received: 30 March 2017
Accepted: 8 August 2017
Published: 2 November 2017
It is difficult to quickly distinguish non-tuberculous mycobacterial (NTM) infection from tuberculosis (TB) infection in human immunodeficiency virus (HIV)-infected patients because of many similarities between these diseases. A simple and effective way to determine the differences using routine blood tests is necessary in developing countries.
A retrospective cohort study was conducted to recruit HIV-infected patients with either NTM infection or TB infection diagnosed for the first time according to mycobacterial culture and microscopic identification from May 2010 to March 2016. These data included the analysis of blood cells, liver function, renal function, C-reactive protein (CRP), and erythrocyte sedimentation rate (ESR), and were compared between the HIV/TB and HIV/NTM groups.
A total of 240 patients were enrolled. The number of HIV/TB and HIV/NTM patients was 113 and 127, respectively. There were no significant differences in the CD4 T-cell count, age, sex, percentage of patients initiating antiretroviral therapy (ART) before the explicit diagnosis of TB or NTM infection. NTM infection was more likely to be restricted in the pulmonary while TB infection also involves extra-pulmonary sites. Both the leukocyte count(5.60 × 109/L) and the proportion of neutrophils in the leukocyte count (76.70%) in the HIV/TB group were significantly higher than those in the HIV/NTM group (4.40 × 109/L [P = 0.0014] and 69.30% [P < 0.001]. The analysis of liver function markers indicated that the concentration of albumin but not ALT and AST was significantly lower in the HIV/TB group than in the HIV/NTM group (P < 0.001). The creatinine and urea levels were not significantly different between the two groups. The ESR (84.00 mm/h) and the concentration of CRP (59.60 mg/L) were significantly higher in the HIV/TB group than in the HIV/NTM group (52.00 mm/h and 19.60 mg/L, respectively) (P < 0.001). To distinguish TB infection from NTM infection, the best cut-off value was 69.5 mm/h for ESR, with a positive predictive value (PPV) of 0.740 and negative predictive value (NPV) of 0.721, and 48.8 mg/L for CRP, with a PPV of 0.676 and NPV of 0.697.
The dissemination character as well as stronger immune response characterized by higher inflammation markers (e.g. WBC, ESR, CRP) can help distinguish TB from NTM infection in HIV-infected patients who need empirical therapy or diagnostic therapy immediately in low-income areas.
Please see Additional flie 1 translations of the abstracts into the six official working languages of the United Nations.
Human immunodeficiency virus (HIV)-infected patients, particularly those with acquired immune deficiency syndrome (AIDS), are commonly infected with several pathogenic microorganisms. Mycobacteria, including non-tuberculous mycobacteria (NTM) and mycobacterium tuberculosis (TB), are an important group of pathogens that infect HIV/AIDS patients more often than immunocompetent individuals [1–4]. The regimens and course of TB/NTM infection treatment are different. Anti-TB drug regimens include isoniazid, rifampicin, pyrazinamide, and ethambutol, etc. Drug regimens for NTM infections include kanamycin and rifabutin, or azithromycin and rifabutin [5, 6]. Although rifabutin has anti-TB action, the treatment of patients infected with TB or NTM is not directly analogous [5–7]. Both HIV/TB and HIV/NTM-infected patients may die if they are not treated promptly and accurately.
However, the clinical manifestations of NTM/HIV and TB/HIV are similar, including fever, cough, and fatigue [8, 9]. Therefore, it is difficult to distinguish the type of mycobacterial infection by the patient’s clinical status. The golden evidence is mycobacterial culture and microscopic identification of the bacterial strains. However, it takes 3 to 4 weeks to obtain the results of a liquid culture medium for mycobacteria , and the positivity rate of mycobacterial culture is low (less than 50%) [10, 11]. The MPB64 antigen has been shown to be specific for the TB  and used clinically for identification of TB after mycobacterium culture [13, 14]. Nevertheless, this period is too long for patients who need anti-NTM or anti-TB therapies.
Other immunological indicators such as interferon gamma release assays (IGRAs) can rapidly diagnose TB infection. However, the sensitivity and specificity of IGRAs are compromised in HIV-infected patients [15, 16] and these methods cannot distinguish latent from active TB infection . Second, several NTM (e.g. in Mycobacterium kansasii, M. szulgai, and M. marinum) also express the ESAT-6 and CFP-10 protein, which is detected by IGRAs . Furthermore, these immunological methods are expensive and difficult to implement in low-income countries with a high burden of infectious diseases, including HIV, TB, and NTM.
Therefore, it is necessary to implement a simple and effective strategy to distinguish NTM from TB infection in HIV infected patients in low-income area who need empirical therapy or diagnostic therapy immediately. The purpose of this study was to help chronic HIV-infected patients suspected with TB/NTM infection distinguish TB from NTM by comparing the levels of routinely detected indicators in blood.
Detection of routine indicators
Complete blood count was carried out using automated blood cell analyser. The output results included leukocyte count, neutrophils, lymphocytes percentage and haemoglobin (Hb). The liver function tests including alanine aminotransferase (ALT) aspartate amino trasferase (AST) and renal function tests including urea, creatinine and serum albumin were measured by colorimetry using automated analyser. CRP was tested by turbidimetric inhibition immunoassay and ESR was detected by Westergren method.
Data normality was assessed using the Shapiro-Wilk test. Normally distributed data are shown as the mean ± standard deviation, whereas non-normally distributed data are shown as the median and interquartile range (IQR). The Levene’s test was used to evaluate the variance homogeneity of the data. Student’s t-test was conducted to assess differences between the two groups in cases in which the data showed normal distribution and homogeneity of variance. The Wilcoxon rank-sum test was used to evaluate data with non-normal distribution or heterogeneity of variance. The chi-square (χ2) test was applied to analyze the categorical variables. The sensitivity and specificity of indicators significant differences were obtained by receiver operating characteristic (ROC) curve. The results were considered significant when the P-values were equal to or smaller than 0.05. All statistical analyses were conducted using IBM SPSS software version 19.0 (IBM SPSS, Inc., Armonk, NY, USA) and GraphPad Prism 6.0 (GraphPad Software Inc., San Diego, CA, USA).
Characteristics of the study population
General characteristics of the study subjects
Onset at diagnosis
TB or NTM infection
Pulmonary and extrapulmonary
CD4 T cell count
Group and subgroup of patients with active tuberculosis
Ordinary pulmonary TB
Blood disseminated pulmonary TB
Lymph node TB
Skin tuberculosis and tuberculous choroiditis
Pulmonary and extra-pulmonary TB
Pulmonary TB and tuberculous peritonitis
Pulmonary TB and lymph node TB
Pulmonary and intestinal TB
Pulmonary and liver TB
Pulmonary TB combined with tuberculous pleurisy and lymph node TB
Pulmonary and brain TB
Pulmonary TB combined with tuberculous meningitis and intestinal TB
Pulmonary and bronchial gland TB
Group and subgroup of patients with NTM infection
Blood disseminated pulmonary NTM
NTM spinal meningitis
NTM peritonitis and intestinal NTM
Pulmonary and extra-pulmonary NTM
Pulmonary and intestinal NTM
Pulmonary and lymph node NTM
Pulmonary and skin NTM
Pulmonary and stomach NTM
Pulmonary and NTM meningitis
Pulmonary NTM and NTM peritonitis
Pulmonary and bone NTM
Accessible blood index
Routine blood test
ESR and CRP
ESR and CRP had a moderate effect on the identification of TB from NTM co-infection with HIV
Sensitivity and specificity of test indicators for distinguishing TB from NTM infections
Maximum Youden index
Best cut-off point
5.55 × 109/L
65.05% in leukocytes
17.75% in leukocytes
Our results indicated that the prevalence of NTM and TB infection is similar in advanced HIV infection. Similarly, other studies reported that more people were infected with NTM than before [19, 20]. This result suggests that HIV-infected individuals may be infected with Mycobacterium and that NTM infections are more likely to be diagnosed than TB infections. Methods of identification of NTM include mycobacterial culture [10, 11] and then are distinguished by specific antibodies against TB [12, 13], or other immunological methods [14–16]. However, these take a long time to achieve definite results or are expensive and difficult to implement in low-income countries. Our results revealed that it was possible to identify co-infection with HIV and either NTM or TB by markers that are routinely detected in blood.
We found NTM infection was more probably to be located in the pulmonary but TB infection was more distributed in other tissues and organs including peritoneum, lymph nodes, intestinal tract, liver, brain and bronchial gland. Our study confirmed that the most common clinical manifestation of NTM disease is lung disease, while lymphatic and skin/soft tissue involvement as well as disseminated disease are also reported [21, 22]. However, NTM infections of soft tissue, lymph node or bone are less prevalent . In contrast, tuberculosis easily disseminate to other organizations. One of the reasons is that tuberculosis granuloma play an important role in expansion and dissemination of tuberculosis infection .
There was no significant difference in the CD4 T cell count between the two study groups, and this result might suggest that the immunity of HIV/NTM or HIV/TB infection was similar. Although the guidelines from the Center for Disease Control and Prevention (CDC) emphasize that HIV-infected patients with a CD4 T-cell count lower than 50 cells/μl are more susceptible to infection with NTM but not with TB . We found that even in cases of which the CD4 cell count was higher than 100 cells/μl, the patients could still be infected with NTM. A similar result was found in previous studies [4, 8, 25, 26].
In line with the dissemination character of TB infection in HIV-infected patients, we found these patients also have higher inflammation markers. Both the leukocyte count and neutrophils percentage in the HIV/TB group were significantly higher than those in the HIV/NTM group. Several previous studies found signals from dead or dying granuloma macrophages infected by tuberculous recruit neutrophils and stimulate neutrophil proliferation, which then phagocytose infected macrophages [27, 28]. This may partially explain the phenomenon found in our study.
For the first time, this study showed that the ESR and the concentration of CRP were significantly higher in HIV/TB patients than in the HIV/NTM group. The ESR and CRP had a moderate effect on the identification of co-infection with HIV and either TB or NTM. The probable cause of this difference was that TB infection might induce a stronger inflammatory response than NTM infection. The CRP concentration in HIV(−) patients infected with TB was also reported to be higher than in non-HIV patients infected with NTM .
These results, combined with data on leukocyte count and neutrophils, allow us to speculate that HIV patients infected with TB might present a stronger inflammatory response than HIV patients infected with NTM. This result may be because of the characteristics of the two types of microorganisms. TB is not an opportunistic pathogen. The patients have to undergo anti-TB chemotherapy after the diagnosis of active TB infection. However, NTM are common opportunistic pathogens and their natural environment is larger than that of TB. Furthermore, NTM can survive in the soil, water, milk, food products, aerosols, and wild and domestic animals [30–32]. Individuals are easily exposed to NTM but do not present symptoms and do not need treatment, indicating that NTM infections are harmless to the human body in most situations [3, 23]. Therefore, the infectivity and pathogenicity of NTM are lower than those of TB [23, 33]. However, NTM is the name of a variety of non-tuberculous mycobacteria, and some of which may be higher pathogenic .
Our results indicated that the concentration of albumin in HIV/TB patients was lower than that in the HIV/NTM group. This may be a result of increased albumin consumption in HIV/TB group as they have stronger inflammation as compared with HIV/NTM group.
One of the limitations of our study was that the NTM was not classified into subtypes as they may have different characters. In addition, the retrospective nature of the current study also limited the degree to which these findings can apply to current practice.
The dissemination character as well as stronger immune response characterized by higher inflammation markers (e.g. WBC, ESR, CRP) can help distinguish TB from NTM infection in HIV-infected patients who need empirical therapy or diagnostic therapy immediately.
We are grateful to the staff from the Department of Infectious Diseases at the Shanghai Public Health Clinical Center.
This study was supported by grants from the fourth round public health 3-year action plan key disciplines construction project of infectious diseases and health microbiology (Grant No. 15GWZK0103).
Availability of data and materials
All data presented in this study are available in the Department of Infectious Diseases, Shanghai Public Health Clinical Center, China.
HZL and CJ conceived and designed the study. ZT and XQQ collected and analyzed the data. RTC and FXY prepared the first version of the manuscript. All authors read and approved the final draft of the manuscript.
Ethics approval and consent to participate
The study protocols were approved by the Research Ethics Committee of the Shanghai Public Health Clinical Center. This committee waived the need for written informed consent from the participants because the study was retrospective, anonymous, and only used currently existing data.
Consent for publication
All authors provided consent for publication.
The authors declare that there are no competing interests associated with this study.
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