Incidence and Drug Resistance of Zoonotic Mycobacterium bovis Infection in Peshawar, Pakistan

Abstract

Prevalence of zoonotic Mycobacterium bovis (bTB) disease in human population is underreported from the North of Pakistan. Here, we report on the proportion of human bTB disease among the overall TB patients, drug resistance pattern of bTB isolates, and knowledge, attitude, and practices (KAP)-based analysis of bTB disease. For this purpose, sputum samples from a total of 300 clinically diagnosed TB patients and 100 randomly selected school children suspected of pulmonary TB were processed by culture as well as polymerase chain reaction (PCR) for isolation, identification, and confirmation of Mycobacterium tuberculosis (mTB) and bTB species. Isolates of bTB were processed for drug susceptibility tests. Data on KAP regarding TB were obtained on a pretested questionnaire. Sputum-based PCR results indicated that 288/300 (96%) were confirmed as mTB, while 12/300 (4%) were found as bTB diseases. Interestingly, none of the school child was declared positive for either mTB or bTB. Notably, 274/300 (91.3%) positively cultured samples were identified as mTB, 13/300 (4.3%) as bTB, while 5/300 (1.7%) as mixed containing both. Importantly, except one, all of the bTB isolates were found resistant to pyrazinamide. Surprisingly, most of the bTB isolates (~70%) were found resistant to a broad range of first- and second-line anti-TB drugs. SplitsTree and recombination analysis indicated no evidence of intergenic recombination. Finally, residence, occupation, presence of animals at home, and sleeping alongside animals were found significantly associated with occurrence of bTB disease. To the best of our knowledge, we report for the first time on the high (4%) burden of bTB disease in human TB patients in Peshawar, Pakistan.

Appendix A

1.1 Questionnaire used for Human Tuberculosis Patients

M. bovis

M. tuberculosis

Name		CNIC or cell#
Address				Urban	Rural
Age in years	<20	21–30	31–40	41–50	Above 50
Education	Illiterate	Middle	Secondary	Graduate
Socioeconomic status	V. high	High	Moderate	Low	V. low
Marital status	Single	Married	Divorced	Widower

1. Nationality:Pakistani Afghani Others (specify) __________________

2. Milk consumption habits:

Always use boiled milk
Always use raw milk
Routinely use boiled milk and occasionally raw milk

3. Occupation

(a) Livestock farmer (b) Abattoir worker/butcher/veterinarian (c) Others (specify)

__________________

4. Animals at home?

If yes: (a) Cattle/buffalo (b) Others (specify) ___________ (c) none

5. Do you sleep in animal shed? Yes No

6. Have you ever contracted tuberculosis before? Yes No

7. Is there any tuberculosis-infected person at your home? Yes No

8. Have you ever had close contact with anyone who was sick with tuberculosis? Yes No

9. Is TB a communicable disease? Yes 248 No Don’t know 16

10. Cause of TB? (a) Microorganisms (b) Hereditary (c) Don’t know

11. Health status of the patient

SNO	Signs and symptoms	Yes	No
1	Chronic cough lasts for more than 3 weeks
2	Pain in the chest
3	Coughing up blood or sputum
4	Weakness or fatigue
5	Weight loss
6	No appetite
7	Chills
8	Fever
9	Sweating at night

12. Tuberculosis is contracted by:

	Yes	No	Don’t know
Cough or sneeze
Handshake
Sexual relations
Sharing food
Living with patient Tb
Excess work

13. Tuberculosis can be transmitted from animals to human through:

	Yes	No	Don’t know
Consumption of unboiled milk
Consumption of raw meat
Through aerosol
Others (boiled milk, cooked meat, riding, contact with feces)

14. Is TB preventable? (a) Yes (b) No (c) Don’t Know

15. Tuberculosis can be prevented by:

	Yes	No	D/K
Good nutrition
Closing windows
Covering your mouth when sneezing or coughing
Not sharing food
Avoiding use of utensils used by patient TB

16. IS TB treatable? Yes No Don’t know

17. People affected by TB can be cured by:

	Yes	No	Don’t know
Adhering to treatment provided by physician
Herbal cures/consulting hakeem
Resting but not taking medications
Prayers/eating well/others

18. What feelings do you have when presented with a diagnosis of TB? (a) Fear(b) Surprise(c) Shame(d) Sadness/hopelessness(e) Others (hurt, do not fear, etc.)

19. With whom a diagnosis of TB would be shared (can choose multiple answer)? (a) Physician(b) Partner(c) Parent(d) Offspring(e) Friends or relation(f) No person

20. How serious a disease is TB?(a) Very serious(b) Somewhat serious(c) Not very serious(d) Don’t know

21. From what sources did you get information about tuberculosis? (can choose multiple answer) (a) Newspapers(b) Radio(c) TV(d) Internet(e) Pamphlets and posters(f) Health worker(g) Family member, friend, or neighbor(h) Others (specify)

22. Community attitudes toward patient tuberculosis (can choose multiple answer)(a) People keep distance from those with TB(b) People do not want those with TB playing with their children(c) People do not want to talk to others with TB(d) People try not to touch others with TB(e) People think that those with TB are unclean(f) Others (specify)

23. Do you know that TB treatment is free?(a) Yes (b) No