Tuberculosis (TB), mainly caused by
Bacille Calmette-Guérin (BCG), an attenuated derivative of
It is widely accepted that protective immunity of humans against TB is mainly attributable to cell-mediated immunity, especially T lymphocytes. In addition, humoral immunity plays small role in the protective immunity to TB [
Even though BCG is the most widely used vaccine in human history, its mechanisms remain poorly understood. Additionally, there have been many daughter strains of BCG with varied protective efficacy [
The 15 genome sequences assessed in this study belonged to 13 BCG strains (BCG-Moreau, BCG-Phipps, BCG-Sweden, BCG-China, BCG-Danish, BCG-Prague, BCG-Russia, BCG-Tice, BCG-Glaxo, BCG-Mexico, BCG-Pasteur, BCG-Tokyo 172, and BCG-Frappier),
An online search in the Immune Epitope Database (IEDB) was performed, and peptidic epitopes were selected as targets in this study.
The sequences of peptidic epitopes were compared to the H37Rv proteome, using BLAST to confirm which genes encoded the amino acid sequences of various B cell epitopes [
The gene sequences encoding B cell epitopes were compared individually with the whole genomes of BCG strains and
A total of 399 experimentally verified peptidic human B cell epitopes were obtained in the IEDB. Since 2 epitopes (epitope IDs 94762 and 94763) shared the same nucleic acid sequence with only a few modified amino acids, epitope 94763 was not included in this study. Finally, 398 B cell epitopes involved were analyzed.
All peptidic epitopes were included in 81 genes according to
According to an analysis by Cole et al. in 1998, all
Distribution of epitopes in various functional categories.
DNA fragments encoding B cell epitopes, based on H37Rv gene sequence, were extracted from the 13 BCG genomes to assess the changes of B cell epitopes. A total of 398 B cell epitopes were analyzed. No SNP was identified in 321 epitopes of all BCGs (classified as Group 1); the remaining 77 epitopes varied in DNA levels among BCGs to different degrees. Of the 77 nucleic acid-altered epitopes, 15 presented the same nucleotide change in all BCGs (classified as Group 2). A total of 26 epitopes were lost in all BCGs because of gene deletion (classified as Group 3). There were 13 epitopes lost in 8 BCG strains, including BCG-Phipps, BCG-Danish, BCG-Prague, BCG-Tice, BCG-Glaxo, BCG-Mexico, BCG-Pasteur, and BCG-Frappier (classified as Group 4). The remaining 23 epitopes showed different SNPs or nucleotide deletions in different BCGs (classified as Group 5) (Figure
The composition of 398 human B cell epitopes distributed in BCGs. Five groups of B-cell epitopes in the 13 BCG strains. Epitopes shown in red were present, those shown in sky blue were absent, and epitopes labeled yellow were showing same SNPs in BCGs exclusively.
Group 1 epitopes were present in all BCGs, with no SNP in these strains. In comparison with
Group 2 included 15 epitopes showing the same SNPs in BCGs compared with
A total of 26 epitopes were classified as Group 3, involving 6 genes according to
The epitopes in Group 4 were involved in 4 genes, including
Group 5 covered 23 epitopes with unique performances in different BCGs.
In summary, the 398 epitopes could be divided into 2 categories, conserved and changed epitopes, based on nucleotide changes. The conserved epitopes contained 321 (80.7%) epitopes of Group 1. The changed ones included 77 (19.3%) epitopes of Groups 2, 3, 4, and 5. The epitopes encoding genes for “cell wall and cell processes” presented high variability (Figure
Variability of B cell epitopes in different functional categories.
Here we compared B cell epitopes in 13 BCGs. All conserved and changed epitopes were analyzed according to BCG strain genomes. Finally, BCG-Tokyo 172 and BCG-China had the highest number of conserved epitopes (357 (89.7%)) (Figure
B cell epitope numbers in BCGs.
Tuberculosis, a horrible disease, had plagued humankind throughout the history. According to WHO data, 9.6 million new TB cases were estimated as well as 1.5 million TB related deaths in the world in 2014. BCG remains the only available and widely used vaccine against TB, although its protective effect against TB is not optimal, even with no protection for adults, because new TB vaccines are continually declared to be more effective in TB prevention. To date, at least 3 billion individuals have been immunized with BCG, and over 100 million doses of BCG are administered annually [
This is the first study analyzing the protective differences of BCGs based on B cell epitopes distribution. It has been acknowledged that BCG is not a single strain and has lots of descendants worldwide. After the original BCG was disseminated around the world, it likely to undergo various selective pressures. As a result, different daughter-BCGs were produced, with variable protective efficacy. The BCG ancestor was serially cultured for at least 230 passages from 1908 to 1921, losing some important epitopes (Group 3, coding with RD1, RD3, and RD7); meanwhile, some point-mutations occurred in the epitopes (Group 2). Therefore, all 13 BCGs contained epitopes of Groups 1 and 2, losing those of Group 3. Then, some BCGs successfully retained Group 4 epitopes, including BCG-Moreau, BCG-Sweden, BCG-China, BCG-Russia, and BCG-Tokyo 172. Of these strains, BCG-Moreau, BCG-Sweden, and BCG-Russia produced more changes in the next few years. In addition, other daughter-BCGs underwent a period of continuous change, losing Group 4 epitopes (coding with RD2), including BCG-Prague, BCG-Phipps, BCG-Glaxo, BCG-Frappier, BCG-Danish, BCG-Tice, BCG-Mexico, and BCG-Pasteur. Of these strains, 4 BCGs (BCG-Prague, BCG-Phipps, BCG-Glaxo, and BCG-Tice) produced more change in B cell epitopes (Group 5) when they were disseminated to various parts of the world. The currently available BCGs exhibited distinct process of attenuation after 1924 [
Although the reasons for variable protective efficacy of BCGs are not fully understood, a key factor might be genetic differences in BCGs [
B cell epitope is the region of a pathogen that interacts with the host immune system. As shown above, most epitopes identified from genes encoding antigens on the pathogen surface were recognized by the host. Generally, antigen epitopes tend to be hypervariable to evade immune detection by the host [
The BCG vaccine, an attenuated strain of
In this study, a total of 398 experimentally verified human B cell epitopes were analyzed with 13 BCG genomes. Of these, 321 were conserved in all BCGs; the remaining 77 B cell epitopes, encoding Regions of Differences or other important genes, have been altered during the ongoing dissemination. Our analysis, based on B cell epitopes distribution, also revealed that BCG-China especially BCG-Tokyo 172 could be the most effective BCG vaccines used in the world. Restoration of important antigens in BCG or enhancing effective B cell epitopes in vaccines could be a useful strategy for vaccine development.
The authors declare that they have no competing interests.