Prevalence and Drug Resistance Pattern of Mycobacterium tuberculosis Isolated from Tuberculosis Patients in Basra, Iraq

Tuberculosis (TB) is considered to be a highly contagious disease, causing thousands of deaths globally (Sousa et al. 2020). It is caused mainly by Mycobacterium tuberculosis (MTB), responsible for 1.5 million deaths occurring each year in current times (WHO 2020). Since the twentieth century, after the invention of the BCG vaccine (Bacillus Calmette-Guérin, 1921) and the use of anti-TB agents (isoniazid (INH), rifampicin (RIF), ethambutol (EMB), pyrazinamide (PZA)), the occurrence of TB dropped rapidly (Godreuil et al. 2007). However, the rate of TB amplified in the 1980s due to the emergence of the strains resistant to antituberculosis drugs. The increase of drug-resistant tuberculosis (DR-MTB) has become a significant public health concern that represents a risk to world TB control programs (WHO 2018). About 25% of TB cases recorded globally demonstrate resistance to one of the first-line drugs used to treat this disease. Approximately

5% of first-line DR-MTB cases progress to multi-drug resistant tuberculosis, which can progress to extreme drug-resistant tuberculosis (XDR-TB) (Madrazo-Moya et al. 2019).

The main two types of drug resistance in M. tuberculosis are primary (transmitted) resistance and secondary (acquired) resistance. Primary resistance is defined as drug resistance in people who have never taken antituberculosis medicines or have only had them for less than a month. These individuals are thought to have an initial infection with drug-resistant bacteria (new cases). Resistance to anti-tuberculosis medicines that develops during treatment or in a person who has been on therapy for at least one month is referred to as an acquired resistance (retreated cases) (WHO 2003).

Resistance to anti-tuberculosis drugs is mainly associated with the infective TB control programs. Consequently, this has affected control efforts made by countries with a lack of accessibility to second-line anti-TB drugs. The reasons were insufficient therapy, low patient compliance, reduced drug supply, and unsuitable treatment routines (Urassa et al. 2008).

According to the data released in 2018 by World Health Organization, approximately 10 million people developed TB in 2017. Three and half percent of newly diagnosed and 18% of retreated TB cases were identified as multi-drug resistant (MDR) TB (WHO 2018).

Iraq is one of the countries in the World Health Organization Eastern Mediterranean Region (WHO-EMR), which has a population of approximately 38 million with a TB incidence rate of 42 per 100,000 (WHO 2018). It places Iraq in the top seven countries within the WHO-EMR for TB incidence, i.e., 3% of total TB cases (WHO 2009).

The present study aimed to investigate the prevalence of TB to understand the problem of DR-MTB and the issues related to its spread. It would help determine the high-risk population for drug-resistant TB, develop procedures to stop the spread, and provide effective management for future TB control.

Study population and data collection. During the study period (2016–2020), M. tuberculosis was detected in 2,542 patients, of which 2,296 TB patients were newly diagnosed, and 246 were the retreated MTB cases. The patients were classified into different categories and subjected to epidemiological analysis.

Table I clarifies the clinical and demographic characteristics of the study enrolled patients. Out of 2,296 newly diagnosed patients, 54 (2.4%) were drug-resistant TB, whereas among retreated cases, 50 (20.3%) patients represented drug-resistant TB. Among the newly diagnosed drug-resistant TB cases, there were more females (53.7%) than males (46.3%), more non-smokers (75.9%) than smokers (24.1%), and more pulmonary TB (98.1%) than extrapulmonary (1.9%). Additionally, most newly diagnosed drug-resistant TB – 52 cases (96.3%) were from three age groups: 15–34 years – 21 (38.9%), 35–54 years – 12 (22.2%), and more than 55 years – 19 patients (35.2%). The distribution of age, sex, smoking, etc., among treated drug-resistant TB cases was similar to the newly diagnosed cases (Table I).

When comparing new drug-susceptible TB cases to newly diagnosed drug-resistant TB cases, it might be observed that more patients were at the age of 15–34 years old (OR: 0.788, 95% CI: 0.420–1.479), and more numerous were females (OR: 1.324, 95% CI: 0.771–2.275). Also, newly diagnosed drug-resistant TB were more likely to be pulmonary TB (OR: 52.062, 95% CI: 7.187–377.129, p > 0.05), and more patients came from urban areas (OR: 1.939, 95% CI: 1.12–3.358, p < 0.05). Among retreated TB cases, secondary drug-resistant cases were more likely to be at the age of 15–34 years old (OR: 1.622, 95% CI: 0.758–3.470), presented pulmonary TB (OR: 18.634, 95% CI: 2.51–138.31, p > 0.05), and were from urban areas (OR: 2.661, 95% CI: 1.412–5.015, p > 0.05). There was no significant difference between males and females (OR = 1, p = 1) in this group of patients.

New TB cases were more likely to be diagnosed within patients at the age of 15–34 years (OR: 1.613, 95% CI: 1.166–2.233, p > 0.05), younger than 14 years (OR: 7.748, 95% CI: 3.109–19.308, p > 0.05), and aged 35–54 years old (OR: 0.851, 95% CI: 0.753–0.962). New TB was diagnosed less likely in patients aged more than 55 years (OR: 1.309, 95% CI: 0.945–1.813), compared with retreated TB cases (Table I).

The cavitary disease was associated with retreated cases more often than in patients with new TB cases (OR: 0.372, 95% CI: 0.256–0.541 p > 0.05). The cavity has a higher risk and more probability in patients of acquired resistance (OR: 0.391, 95% CI: 0.161–0.948, p > 0.05) (Table I).

In new TB cases, pulmonary drug-resistant TB occurred with a higher probability and risk than extra-pulmonary drug-resistant TB (OR: 52.062, 95% CI: 7.187–377.129, p > 0.05). Also, in retreated cases, resistance developed more often in pulmonary TB that in extra-pulmonary TB (OR: 18.634, 95% CI: 2.51–138.31, p > 0.05). Furthermore, patients with pulmonary TB showed a higher risk of being retreated cases than those with extra-pulmonary TB (OR: 0.307, 95% CI: 0.225–0.418, p > 0.05).

The new patients from urban areas had a higher risk and more probability of developing resistance (OR: 1.939, 95% CI: 1.12–3.358, p > 0.05) than in retreated cases (OR: 2.661, 95% CI: 1.412–5.015, p > 0.05) when compared to patients from rural regions. New patients living in urban areas were more likely to be diagnosed (OR: 1.393, 95% CI: 1.059–1.832, p > 0.05) than retreated cases when compared to patients from rural regions. A higher frequency of retreated cases (64.6%) than new cases (56.7%) occurred in rural regions (Table I).

Resistance patterns. As demonstrated in Table II, among 2,296 newly diagnosed and 246 retreated MTB clinical isolates, drug resistance is significantly higher among the isolates from retreated patients than those from newly diagnosed cases (20.3% vs. 2.4%, p > 0.05). The frequency and percentage of the resistance to first-line drugs, involving INH, RIF, EMB, SM among the newly diagnosed and retreaded patients were as follows: 23 (1%) vs. 42 (17.1%), 18 (0.78%) vs. 39 (15.8%), 13 (0.56%) vs. 21 (8.5%), and 30 (1.3%) vs. 24 (9.75%), respectively. MTB’s most common drug resistance was against streptomycin with a rate of 1.3% in new TB patients and isoniazid (17.1%) in retreated TB patients. When compared to retreated TB cases, new cases have lower rate of MDR-TB (0.34% vs. 14.2%, p < 0.05), PDR-TB (0.78% vs. 4.1%, p = 0.257), but the occurrence of MR-TB was virtually similar in this group (1.22% vs. 2%, p = 0.564). Out of 28 newly diagnosed MR-TB cases, the highest rate of MTB resistance was against SM (13, 0.56%), followed by RIF (7, 0.3%), INH (4, 0.17%), and EMB (4, 0.17%). The results showed the prevalence of four different types of newly diagnosed MDR-TB, for example, MDR1 (INH + RIF), MDR2 (INH + RIF + EMB), MDR3 (INH + RIF + EMB + SM), and MDR4 (INH + RIF + SM). Likewise, newly diagnosed PDR-TB mostly consist of two (0.08%) PDR1 (INH + EMB), seven (0.3%) PDR2 (INH + SM), three (0.13%) PDR4 (RIF + SM), two (0.08%) PDR5

(INH + EMB + SM), and four other isolates (0.17%) (SM + EMB). There were no PDR3 (RIF + EMB) isolates. The most common type of drug resistance in retreated cases is MDR (14.2%), with the highest resistance rate (8.1%) against INH and RIF, followed by PDR (4.1%) with the highest resistance rate (1.6%) for INH, EMB, and SM, and MR-TB (2%) with the highest resistance rate for RIF (1.2%) (Table II).

Overall and yearly new diagnosed drug resistance rate. Table III shows the total and annual newly diagnosed drug resistance rate of many TB subcategories from 2016 to 2020. Throughout the study period, the rate of newly diagnosed drug resistance among females, the 15–34-year-old TB subcategories increased by 365 and 274%. It varied from 1.75 to 8.13% and from 1.9 to 7.1%, respectively. The increased TB occurrence was observed in males and the other age groups as illustrated in Table III. Moreover, the rate of total DR-TB, MDR-TB, MR-TB, and RFP-resistant TB among new tuberculosis cases increased from 2.2 to 6.7%, 0.17 to 1.6%, 0.85 to 4%, and 0.17 to 4%, with a percentage change of 205, 841, 371, and 23%, respectively. On the other hand, PDR-TB and EMB-resistant TB rates dropped by 15.9 and 0.7%, decreasing from 1.19 to 1% and from 1 to 0.3%, respectively.

Total and annual secondary drug resistance rates. Table IV shows the overall and annual secondary drug resistance rate of TB subcategories from 2016 to 2020. Through the study period, the secondary drug resistance rate amongst females, males, and patients of all ages increased, as illustrated in Table IV. Additionally, the rate of total DR-TB, MDR-TB, PDR-TB, and MR-TB also increased. Moreover, the first-line drug resistance INH, RIF, EMB, and SM-resistant TB was also elevated during this period.

Our study enrolled 2542 cases of newly and retreated TB patients in Basra from 2016 to 2020 to decipher the epidemiology and prevalence of drug resistance in M. tuberculosis isolated from clinical samples in the Basra province.

The present results show that during the study period (five years), M. tuberculosis was detected in 2,542 samples at an average of 508.4 per year. WHO stated tuberculosis case detection for the last five years at a rate of about 43/100,000 population in Iraq (WHO 2019).

The total population in Basra is estimated to be 2,985,073 as of 2019 (Central Statistical Organization 2019), then the predictable tuberculosis cases would be about 1,284 patients per year. There were 508 identified patients in this study, as the average of the last five years, less than a half of the WHO predicted number. It could be attributed to an inadequate screening procedure or the overestimated case finding the target.

Relationship of TB with age. The study revealed that patients from the 15–34 years old group comprise the most affected group, and this observation is similar to most developing countries (Dogar et al. 2012). It was followed by the patients of 35–54 years old. These findings might be attributed to many young people in Iraq. At this age, individuals are more active and have a higher level of social activities, interactions with specific environmental factors, or lifestyle characteristics. These results agree with national and regional studies carried out by Al-Rubayai (2011), Mokdad et al. (2014), Khdban (2018), and Merza et al. (2020) but in contrast with Jappar and Low (2015), and Karadakhy et al. (2016) who reported that the most affected age groups are the older people.

Relationship of TB with sex. The present study illustrated that the TB rate was higher in males than in females, which may be related to the social behavior of males in our society, who have more social activities, travel, and more work outside the home than females. Our results are similar to those obtained by Al-Khafaji (2014) and Bedewi (2011) in Baghdad, Iraq. WHO also reported that men are more affected than women in Iraq (WHO 2017).

On the other hand, MTB isolates from females presented primary drug resistance with a higher frequency rate than males. There were no such differences in retreated cases. It could be due to cultural reasons when women provide care to in-home patients with DR-TB. It puts women at increased risk for exposure and subsequent DR-TB development. Our results agree with the studies in former Soviet Union countries (Faustini et al. 2006) and Republic of Georgia (Lomtadze et al. 2009), which showed that women are more likely to have MDR-TB than men in new cases of TB. Our findings, however, did not match with the results of the Iranian study, which presented females as more resistant to MDR-TB than males (Jimma et al. 2017).

Relationship of TB with pulmonary cavity. In the case of pulmonary TB, the cavity was more associated with retreated cases than new cases, which may cause the prolonged time to sputum culture conversion (Hernandez-Romieu et al. 2019). The presence of initial pulmonary cavitation can be considered a predictor for retreated TB because of containing a high bacillary load, so it may associate with the progress of drug resistance throughout the time of treatment (Kempker et al. 2012).

Relationship of TB with type of infection. Our study showed that total pulmonary TB (PTB) (54%) is higher than total extra-pulmonary TB (EPTB) (45.9%). The present results are similar to those obtained by Ahmed (2018) but different from other studies carried out in Baghdad by Durib and Blinova (2020), who reported that PTB occurred in 47.64% and EPTB in 52.36% cases, respectively. The low rate of EPTB could be because of the absence or very low rate of HIV infection since it has been reported that the increase in EPTB rate is more than 50% if associated with HIV (Golden and Vikram 2005).

The present results showed a high frequency of drug resistance among PTB agreed with the study in Papua New Guinea (Diefenbach‐Elstob et al. 2018).

Relationship of TB with urban vs. rural regions. The present study showed that TB in rural settings is higher than in urban areas. The reason may be that rural patients seek health care from traditional healers which causes the delay in TB diagnosis, resulting in continuous transmission of the disease. Our results were similar to those obtained in China, which showed that approximately 80% of TB patients live in rural areas (Liu et al. 2005), but different from that what was found in Baghdad by Durib and Blinova (2020), who reported that the frequency of TB in urban populations was much higher than in rural ones. However, the global problem of TB infection is estimated to be higher in urban than in rural areas (Kapata et al. 2016).

Furthermore, we found that the total MTB drug resistance in urban regions was higher than in rural regions, similar to what was found in India (Almeida et al. 2003). There may be several reasons for these findings, such as those patients in rural settings have fewer admissions to different doctors, and rural areas are less crowded, so there is less transmission of drug resistance TB. The present results revealed that drug resistance was more likely found in retreated cases than in new TB patients. It was almost similar to the results reported in Tanzania (20.6% in retreated vs. 8.3% new cases) (Chonde et al. 2010) and lower than what was found in Sudan (62.8% retreated vs. 30.7% in new TB cases) (Hajissa et al. 2021), India (58.4% retreated vs. 24.9% in new TB cases) (Lohiya et al. 2020), and China (retreated 43.9% vs. 22% in new TB cases) (Yang et al. 2014). Our results also showed that retreated TB patients were significantly more likely to have MDR-TB than patients with primary TB, which may be due to inadequate use of treatment, incomplete treatments, or due lung cavities. These results were almost similar to a study carried out in Ethiopia, which showed the frequency of MDR-TB was 2% in primary cases and 15% in retreated cases (Eshetie et al. 2017), and lower than those obtained by Lomtadze et al. (2009), who found 6.8% vs. 27.4% of MDR-TB in new cases and retreated cases, respectively.

The frequency of MR-TB in the present study was higher in retreated cases than in new cases, which was consistent with another study in the north of Iraq (Merza et al. 2011), but in contrast with the situation in Sudan, where MR-TB was more prevalent in new cases than in retreated ones (Hajissa et al. 2021).

Globally, the occurrence of INH resistance in TB is poorly understood; however, in our study, the frequency of INH resistance was 17.1% in retreated cases and 1% in new cases. It is much less than the global prevalence of INH (27.2%) in retreated, and new cases (10.7%), and less than the Eastern Mediterranean retreated (23.5%) and new cases (10.7%), but almost similar to African retreated cases 13.5% (Dean et al. 2020). The wide variation in the prevalence of INH-resistant TB between different countries could be due to the absence of appropriate diagnostic tools for screening INH resistance, genetic diversity of MTB isolates, or the association of INH resistance with other diseases HIV. However, it requires further studies to be confirmed.

Annual disparity of primary and secondary TB. There was an increase in total drug resistance in Basra during the five years from 2016 to 2020. This phenomenon is considered a public world health problem due to the rapid spread of MTB resistance worldwide. The annual trend of primary drug resistance revealed that the rate of MR-TB and MDR-TB sharply increased while PDR-TB decreased by 15.96%. The significant increase in MDR may be due to increased testing in recent years and the efficient spread of MDR strains. It alarms us about the challenge of eliminating the anti-mycobacterial drug resistance.

Furthermore, secondary resistance to the three types of MR-TB, MDR-TB, and PDR-TB, was markedly increased in the last five years, especially MDR, which showed a high percentage change (1,251%). The increase in secondary DR-TB was higher than in primary DR-TB, similar to studies carried out in China by Song et al. (2019) and He et al. (2016). It indicates that patients with prior TB infections are more likely to develop resistance.

Regarding the number of primary DR-TB, this parameter increased through the five years (2016–2020) in all age groups. This phenomenon was significantly visible in people 15–34 years old, for whom the percentage change was 274%. The number of secondary DR-TB was also elevated in all age groups, with the most significant increase in patients in age 35–54 years old with a distinct percentage change (2,079%). It is, therefore, also essential to concentrate on patients who are categorized as young and middle-aged. A similar finding was recorded in China (Song et al. 2019).

The number of infected females increased over the years studied more than infected males in primary and secondary DR-TB. According to the first-line drug in primary DR-TB, there was a decrease in EMB over the years by 0.7%, and an increase in INH, RIF, and SM, and RIF had the highest percentage change (22.5%). In secondary DR-TB, resistance to the four drugs has increased over time, so we need more focus on the drug regimen. It was the opposite of what was found in China, as there was a decline in drug resistance to these four drugs (Lan et al. 2019) but similar to those reported by Shamaei et al. (2009), in Iran, who stated that the resistance to the first-line drugs significantly increased.

The present study had defined the time-based trend of DR-TB in Basra province in Iraq from 2016 to 2020. The results showed that the prevalence of TB in Basra from 2016 to 2020 was at an average of 508.4. The most affected age group was young people, a high frequency of TB infection was detected in males, and rural regions. The pulmonary cavity was more associated with retreated cases than new cases. The prevalence of drug resistance in retreated cases was higher than in new TB cases. The highest percentage of resistance was observed against isoniazid among first-line drugs. The temporal trend showed an increasing rate of drug resistance since 2016, especially in the number of MDR-TB patients, and with dominance in female patients. Concerning the primary resistance, the resistance against ethambutol is decreasing. The same trend follows the number of PDR-TB cases. These findings would enhance public responsiveness to TB prevention and control. It is fair to say that they can be deemed necessary in the aim of TB elimination and reduction in Iraq. However, the sample size and geographical sources of clinical samples may not be sufficient to generalize the conclusions to the whole country. More studies are required to investigate a larger sample size from different geographical locations to generalize the outcome information for the entire country.