Menstrual knowledge and practices of Pakistani girls: A multicenter, cross-sectional study.

Information regarding the menstrual knowledge and practices of Pakistani girls is sparse. Therefore, we aimed to evaluate the knowledge and practices of Pakistani girls regarding menstruation. This cross-sectional study was conducted among four categories of university female students of medical, pharmacy, nursing, and arts & humanities during a period of 5 months (November 2016-March 2017). The data were acquired using a self-administered questionnaire from 3 medical institutes, 2 pharmacy institutes, 3 nursing institutes and 2 arts & humanities institutes at Lahore, Pakistan. The mean age of participants (N = 1777) was 20.38 ± 2.39 years. Overall 39.5%, 76.1% and 29.3% respondents' provided correct answers to the questions concerning menstrual cycle length, duration of menstrual bleeding and source of bleeding, respectively. Regarding the understanding of menstruation, around 27% understood it as monthly bleeding happening with every woman, whereas 18% stated that it was a sign of adulthood. However, a small proportion believed that it was removal of dirt from the body. Regarding the practices-related to menstruation, there were some malpractices regarding diet and dietary intake (avoiding certain foods, eating less, and less fluid intake). However, the hygienic practices were found to be satisfactory. In conclusion, Pakistani university girls have some misconceptions, misperceptions and malpractices related to menstruation. Our findings draw attention to conduct awareness programs in order to eradicate these misconceptions, myths and malpractices.

Introduction

Menstruation is a natural phenomenon that is defined as “periodic discharge of blood from the uterus through vagina, occurring more or less at regular monthly intervals throughout the active reproductive life of a female (Critchley, 1986)”. There are several types of menstrual disorders (amenorrhea, premenstrual syndrome, dysmenorrhea, oligomenorrhea, polymenorrhea, menorrhagia etc.) but the most prevalent among young females are dysmenorrhea and premenstrual syndrome (PMS) (Rafique and Al-Sheikh, 2018; Amu and Bamidele, 2014; Karout et al., 2012; Anandha Lakshmi et al., 2011; Houston et al., 2006; Singh et al., 2008). Aforementioned menstrual disorders had a negative impact on the quality of life as it resulted in class absenteeism, social restrictions and limited academic, sports and daily activities (Unsal et al., 2010). An increased risk of menstrual disorders has been associated with obesity, smoking, coffee consumption, menstrual bleeding >7 days and a positive family history of dysmenorrhea and PMS (Bae et al., 2018; Unsal et al., 2010). Psychological stress is also known to be independently associated with premenstrual symptoms and the experience of irregular menstrual cycles among college students (Rafique and Al-Sheikh, 2018; Yamamoto et al., 2009). An earlier study from Karachi, Pakistan, revealed that there were misconceptions and unhygienic practices of menstruation among school-going girls requiring action by healthcare professionals (Ali and Rizvi, 2010). However, to our knowledge, menstrual knowledge and practices of Pakistani university going girls have not been assessed. Therefore, we aimed to evaluate the menstrual knowledge and practices of Pakistani university girls.

Subjects and methods

Study population and settings

A cross-sectional study design was used and the study was conducted in the second largest metropolitan city of Pakistan (Lahore, the capital city of Punjab Province) among four groups of university-going girls (medical, pharmacy, nursing, and arts & humanities students) during a period of 5 months (November 2016–March 2017). The data of medical students were gathered from Fatimah Jinnah Medical College, Ameer-Ud-Din Medical College and Services Institute of Medical Sciences. The data of female pharmacy students were gathered from Punjab University College of Pharmacy, University of the Punjab and Faculty of Pharmacy, The University of Lahore. The data of nursing students were collected from Schools of Nursing at Lahore General Hospital, The University of Lahore and Mayo Hospital. The data of female arts and humanities students were gathered from University Oriental College, University of the Punjab and University of the Education, Lahore, Pakistan. Investigators approached the university students at the aforementioned settings during university timing and briefed them about the purpose of this study. Those who consented to participate were administered the study instrument and data were collected. If needed, assistance was provided by the investigators.

Sample size

The sample size for this study was computed by Sample Size Calculator (Raosoft®), available at http://www.raosoft.com/samplesize.html. The margin of error was kept at 5%, confidence level of 95% and response distribution was assumed 50%. A minimum sample of 378 female students was required in each group. To avoid biases, we took extra samples from each group, and the maximum required sample size was 1800. A convenient sampling method was employed and all the university students were approached directly by the researchers within the university timings (8 a.m–4 p.m). Researchers explained the objectives and nature of this study and subjects who consented were administered the study instruments.

Ethical considerations

The study protocol was reviewed and approved by the Human Research Ethics Committee, Punjab University College of Pharmacy, University of the Punjab, Lahore, Pakistan (HEC/PUCP/1935). Moreover, written requests, along with required documents, were submitted to the relevant authorities at all the other aforementioned institutes and verbal approvals were received. A written informed consent was obtained from every student prior to their recruitment in the study.

Outcome measures

Menstrual knowledge and practices were assessed by a 37 item self-administered questionnaire that was designed by previous researches (Ali and Rizvi, 2010; Houston et al., 2006). We measured weight with an electronic weighing scale (model MS-100; Certeza, USA) and height to the nearest millimeter with a portable measuring tape.

Statistical analysis

Frequency and percentages were calculated for categorical variables whereas mean ± standard deviation (SD) for continuous variables. P-values were obtained by ANOVA (continuous variables) and Chi-Square test (categorical variables) to determine the significance of the results and a p-value < 0.05 was considered statistically significant. The data were analyzed using SPSS version 22.0 (SPSS Inc., Chicago, IL) for Windows.

Results

A total 1800 questionnaires were distributed and 1777 fully completed questionnaires were received, with a response rate of 98.7% (nursing students 99.8%, pharmacy students 96.9%, medical students 99.6% and arts & humanities students 98.2%).

Demographic characteristics

Demographic characteristics of the respondents’ are given in Table 1. The mean age of the study cohort was 20.38 ± 2.39 years and there was predominance of Muslim undergraduates from urban areas. Majority of the participants had a normal body mass index (64.0%) followed by underweight (23.9%). Moreover, around 39% reported that they had a health professional in their family.

Table 1

Demographic data of the study population (N = 1777).

Variable N (%)	Overall	Medical students	Pharmacy students	Nursing students	Arts & humanities
Age	20.38 ± 2.39	19.08 ± 1.12	20.11 ± 1.99	20.30 ± 2.21	22.05 ± 2.87
Body mass index
Underweight	424 (23.9)	100 (22.3)	118 (27.1)	120 (26.7)	86 (19.5)
Normal	1137 (64.0)	301 (67)	264 (60.6)	289 (64.2)	283 (64)
Overweight	182 (10.2)	40 (8.9)	46 (10.6)	36 (8)	60 (13.6)
Obese	34 (1.9)	8 (1.8)	8 (1.8)	5 (1.1)	13 (2.9)
Year of education
1st	686 (38.6)	261 (58.1)	181 (43.8)	155 (36.8)	89 (84.0)
2nd	353 (19.9)	132 (29.4)	73 (17.7)	135 (32.1)	13 (12.3)
3rd	170 (9.6)	44 (9.8)	57 (13.8)	66 (15.7)	3 (2.8)
4th	117 (6.6)	10 (2.2)	41 (9.9)	65 (15.4)	1 (0.9)
5th	63 (3.5)	2 (0.4)	61 (14.8)	--	--
Stratum
Rural	245 (13.8)	66 (14.7)	28 (6.4)	70 (15.6)	81 (18.3)
Urban	1532 (86.2)	383 (85.3)	408 (93.6)	380 (84.4)	361 (81.7)
Hosteller
Yes	725 (40.8)	282 (62.8)	113 (25.9)	227 (50.4)	103 (23.3)
No	1052 (59.2)	167 (37.2)	323 (74.1)	223 (49.6)	339 (76.7)
Economic class
Lower	409 (23.0)	32 (7.1)	43 (9.9)	194 (43.1)	140 (31.7)
Middle	832 (46.8)	185 (41.2)	216 (49.5)	217 (48.2)	214 (48.4)
Upper	536 (30.2)	232 (51.7)	177 (40.6)	39 (8.7)	88 (19.9)
Father's Occupation
Self employed	630 (35.5)	133 (29.6)	166 (38.1)	173 (38.4)	158 (35.7)
Govt. servant	467 (26.3)	166 (37)	107 (24.5)	103 (22.9)	91 (20.6)
Private employee	348 (19.6)	75 (16.7)	98 (22.5)	83 (18.4)	92 (20.8)
Unemployed	51 (2.9)	9 (2.0)	6 (1.4)	15 (3.3)	21 (4.8)
Father not alive	130 (7.3)	18 (4.0)	22 (5)	46 (10.2)	44 (10.0)
Retired	151 (8.5)	48 (10.7)	37 (8.5)	30 (6.7)	36 (8.1)
Mother's Occupation
House wife	1449 (81.5)	324 (72.2)	358 (82.1)	383 (85.1)	384 (86.9)
Working woman	328 (18.5)	125 (27.8)	78 (17.9)	67 (14.9)	58 (13.1)
Health Care Professional In family
Yes	684 (38.5)	180 (40.1)	175 (40.1)	201 (44.7)	128 (29.0)
No	1093 (61.5)	269 (59.9)	261 (59.9)	249 (55.3)	314 (71.0)

Menstruation knowledge

Menstrual knowledge of the study population is shown in Table 2. Self-reported menarche age was 12–14 years (69.2%) followed by 15–16 years (22.4%). In majority of the respondents, the feelings/reaction at first menarche was a mixture of anxiety, pain, feeling bad, shy or crying. Around 40% of the respondents’ provided the correct answers to the question concerning menstrual cycle length, with significantly higher number of correct responses by medical students as compared to others (p < 0.001). Likewise, a considerably higher frequency of medical students gave the right answers regarding the duration of menstrual bleeding as compared to pharmacy, nursing and arts & humanities students (p < 0.001). Regarding the understanding of menstruation in our sample, around 27% understood it as monthly bleeding that happens with every woman and it was a sign of adulthood (18%). However, a small proportion (4.6%) believed that it was the removal of dirt from the body and this response was frequent in females having arts and humanities education (11.5%) than other participant categories (medical 2.7%; pharmacy 1.6%; nursing students 2.4%). Majority of the respondents had incorrect responses about the source of bleeding (vagina, 62.9%; stomach, 0.7%; any part of abdomen, 4.7%; urinary tract, 2.4%). Menstrual knowledge prior menarche was predominantly acquired from the mothers followed by elder sisters and friends.

Table 2

Menstruation knowledge of the study population among different participant categories.

Variable N (%)	Overall	Medical students	Pharmacy students	Nursing students	Arts & humanities	p-value
Menstrual cycle length (days)
5-10	788 (44.3)	155 (34.5)	234 (53.7)	216 (48)	183 (41.4)	<0.001
10-12	71 (4.0)	6 (1.3)	8 (1.8)	33 (7.3)	24 (5.4)
21-40	702 (39.5)	267 (59.5)	159 (36.5)	153 (34)	123 (27.8)
Don't know	216 (12.2)	21 (4.7)	35 (8)	48 (10.7)	112 (25.3)
Menstrual bleeding duration (days)
1-2	163 (9.2)	14 (3.1)	29 (6.7)	45 (10)	75 (17)	<0.001
2-8	1352 (76.1)	405 (90.2)	370 (84.9)	334 (74.2)	243 (55)
8-12	126 (7.1)	25 (5.6)	25 (5.7)	53 (11.8)	23 (5.2)
Don't know	136 (7.7)	5 (1.1)	12 (2.8)	18 (4)	101 (22.9)
Understanding of menstruation
1. Monthly bleeding	474 (26.7)	110 (24.5)	113 (25.9)	117 (26.0)	134 (30.3)	<0.001
2. Dirty bleeding	81 (4.6)	12 (2.7)	7 (1.6)	11 (2.4)	51 (11.5)
3. Good for marriage	68 (3.8)	13 (2.9)	14 (3.2)	17 (3.8)	24 (5.4)
4. Sign of adulthood	319 (18.0)	67 (14.9)	78 (17.9)	103 (22.9)	71 (16.1)
5. Happens with every female	380 (21.4)	100 (22.3)	99 (22.7)	84 (18.7)	97 (21.9)
6. 1 & 4	60 (3.4)	17 (3.8)	19 (4.4)	19 (4.2)	5 (1.1)
7. 1 & 5	124 (7.0)	46 (10.2)	31 (7.1)	31 (6.9)	16 (3.6)
8. 4 & 5	229 (12.9)	69 (15.4)	72 (16.5)	14 (10.7)	40 (9.0)
9. 1, 2, 3, 4 & 5	42 (2.4)	15 (3.3)	3 (0.7)	20 (4.4)	4 (0.9)
Source of bleeding
Uterus	521 (29.3)	186 (41.4)	105 (24.1)	91 (20.2)	139 (31.4)	<0.001
Vagina	1118 (62.9)	253 (56.3)	315 (72.2)	341 (75.8)	209 (47.3)
Stomach	12 (0.7)	1 (0.2)	2 (0.5)	2 (0.4)	7 (1.6)
Any part of abdomen	83 (4.7)	3 (0.7)	5 (1.1)	12 (2.7)	63 (14.3)
Urinary tract	43 (2.4)	6 (1.3)	9 (2.1)	4 (0.9)	24 (5.4)
Knowledge given about menstruation before menarche
Yes	1636 (92.1)	398 (88.6)	380 (87.2)	428 (95.2)	430 (97.3)	<0.001
No	141 (7.9)	51 (11.4)	56 (12.8)	22 (4.9)	12 (2.7)
Source of knowledge of menstruation before menarche
Mother	864 (48.6)	188 (41.9)	220 (50.5)	228 (50.7)	228 (51.6)
Elder sister	324 (18.2)	67 (14.9)	61 (14.0)	111 (24.7)	85 (19.2)
Aunt	81 (4.6)	10 (2.2)	11 (2.5)	30 (6.7)	30 (6.8)
Friend	313 (17.6)	106 (23.6)	79 (18.1)	51 (11.3)	77 (17.4)
None	141 (7.9)	51 (11.4)	56 (12.8)	22 (4.9)	12 (2.7)
Others	54 (3.0)	27 (6.0)	9 (2.1)	8 (1.8)	10 (2.3)
Information given about menstruation before menarche
1. Physical changes	426 (24.0)	69 (15.4)	102 (23.4)	132 (29.3)	123 (27.8)
2. Social & religious restrictions	226 (12.7)	75 (16.7)	53 (12.2)	44 (9.8)	54 (12.2)
3. Bathing practices	50 (2.8)	10 (2.2)	6 (1.4)	23 (5.1)	11 (2.5)
4. Use of material for absorption	151 (8.5)	46 (10.2)	37 (8.5)	39 (8.7)	29 (6.6)
5. Blood is dirty	80 (4.5)	12 (2.7)	15 (3.4)	10 (2.2)	43 (9.7)
6. Blood comes from vagina	266 (15.0)	90 (20.0)	78 (17.9)	56 (12.4)	42 (9.5)
7. All of the above	26 (1.5)	7 (1.6)	10 (2.3)	8 (1.8)	1 (0.2)
8. None	552 (31.1)	140 (31.2)	135 (31.0)	138 (30.8)	139 (31.4)

Menstrual practices

Menstrual practices of the study population are shown in Table 3. Regarding the dietary modification during menstruation, majority of the study participants reported to avoid eating “too cold foods” (50.3%) whereas around 12% stated that they avoid eating “too hot foods”. Almost 56% of the respondents reported of eating same amount of food during menstruation. Surprisingly, 34% and 38% of the respondents reported to consume less food and reduced fluid intake during menstruation, respectively. Almost 78% of the respondents used sanitary pads for absorbing the blood and among them the highest percentage was that of medical students. Among the participants that reported using absorbent items other than sanitary pads, majority discarded the absorbing materials (Table 3). The absorbent material/item that was not discarded was usually washed and dried in sunlight (54.1%). Moreover, the washed clothes were stored in the clean and covered place so that these could be used again. Around 39% of the respondents reported to avoid bathing during menstruation, with no significant difference among participant categories (p = 0.069).

Table 3

Menstrual practices of the study population.

Variable N (%)	Overall	MedicalStudents	PharmacyStudents	NursingStudents	Arts & humanities Students	P- value
Dietary modifications during menstruation
Food Avoided
Too hot	207 (11.6)	25 (5.6)	36 (8.3)	60 (13.3)	86 (19.5)	<0.001
Too cold	893 (50.3)	227 (50.6)	235 (53.9)	240 (53.3)	191 (43.2)
None	677 (38.1)	197 (43.9)	165 (37.8)	150 (33.3)	165 (37.3)
Eating Pattern
Eat less	604 (34.0)	114 (25.4)	148 (33.9)	161 (35.8)	181 (41)	<0.001
Eat more	176 (9.9)	34 (7.6)	35 (8)	50 (11.1)	57 (12.9)
Eat same amount	997 (56.1)	301 (67)	253 (58)	239 (53.1)	204 (46.2)
Decreased fluid intake
Yes	676 (38.0)	140 (31.2)	164 (37.6)	188 (41.8)	184 (41.6)	0.003
No	1101 (62.0)	309 (68.8)	272 (62.4)	262 (58.2)	258 (58.4)
Sanitary and hygienic practices
Material usedFor Absorption
1. Sanitary pads	1377 (77.5)	405 (90.2)	362 (83)	294 (65.3)	316 (71.5)	<0.001
2. Cotton wool	152 (8.6)	12 (2.7)	34 (7.8)	76 (16.9)	30 (6.8)
3. Pieces of new cloth	96 (5.4)	3 (0.7)	8 (1.8)	47 (10.4)	38 (8.6)
4. Pieces of old cloth	109 (6.1)	14 (3.1)	21 (4.8)	27 (6)	47 (10.6)
5. Woolen cloth	7 (0.4)	2 (0.4)	1 (0.2)	2 (0.4)	2 (0.5)
6. Trousers/Nothing	3 (0.2)	--	1 (0.2)	--	2 (0.5)
7. Others	19 (1.1)	4 (0.9)	5 (1.1)	4 (0.9)	6 (1.4)
8. 1 & 4	14 (0.8)	9 (2)	4 (0.9)	--	1 (0.2)
Handling of absorbent items
Discard it	1675 (94.3)	436 (97.1)	411 (94.3)	426 (94.7)	402 (91)	0.001
Do not discard	102 (5.7)	13 (2.9)	25 (5.7)	24 (5.3)	40 (9)
If do not discard
Wash and discard	28 (27.5)	6 (46.2)	10 (40)	7 (29.2)	5 (12.5)	0.032
Wash and dry	74 (72.5)	7 (53.8)	15 (60)	17 (70.8)	35 (87.5)
Drying of clothes
In sunlight	40 (54.1)	4 (57.1)	10 (66.7)	8 (47.1)	18 (51.4)	0.701
In hiding	34 (45.9)	3 (42.9)	5 (33.3)	9 (52.9)	17 (48.6)
Storing of washed clothes
Clean and covered place	53 (3.0)	5 (71.4)	10 (66.7)	12 (70.6)	25 (74.3)	0.298
Unclean and covered place	10 (0.6)	2 (28.6)	4 (26.7)	1 (5.9)	3 (8.6)
Clean and open space	11 (0.6)	--	1 (6.7)	4 (23.5)	6 (17.1)
Bathing during menstruation
Yes	1087 (61.2)	266 (59.2)	288 (66.1)	277 (61.6)	256 (57.9)	0.069
No	690 (38.8)	183 (40.8)	148 (33.9)	173 (38.4)	186 (42.1)
Pain management
1. Rest	844 (47.5)	197 (43.9)	223 (51.1)	208 (46.2)	216 (48.9)	<0.001
2. Oil massage	50 (2.8)	4 (0.9)	1 (0.2)	15 (3.3)	30 (6.8)
3. Turmeric milk	166 (9.3)	11 (2.4)	36 (8.3)	62 (13.8)	57 (12.9)
4. Hot water bottle	125 (7.0)	24 (5.3)	21 (4.8)	43 (9.6)	37 (8.4)
5. Medicine	332 (18.7)	119 (26.5)	93 (21.3)	59 (13.1)	61 (13.8)
6. Others	65 (3.7)	14 (3.1)	20 (4.6)	12 (2.7)	19 (4.3)
7. Nothing	136 (7.7)	54 (12)	29 (6.7)	40 (8.9)	13 (2.9)
8. 1 & 5	59 (3.3)	26 (5.8)	13 (3)	11 (2.4)	9 (2.0)

Impact of menstruation on absenteeism and daily activities

In the present study, approximately 58% of the participants reported no absenteeism from the institute due to menstruation, with significantly higher proportion (72.8%) that of medical students as compared to others. Regarding the daily/routine activities during menstruation, 31.7% reported no impact on routine activities, whereas 13.6, 34.5 and 20.3% reported seldom, sometimes and always missing routine activities, respectively.

Discussion

This is the first study to evaluate menstrual knowledge and practices of Pakistani university girls. Similar to the findings of earlier studies involving Pakistani and Indian school girls (Ali and Rizvi, 2010; Kural et al., 2015), predominance of our study population also reported a menarche age of 12–14 years. Around 92% of the participants reported that they were well informed of menstruation before menarche and the source of knowledge was predominantly mothers which were consistent with the previously reported studies from India (Dasgupta and Sarkar, 2008; Khanna et al., 2005). Around 60% of our study participants did not know the correct menstrual cycle length which was significantly higher (39%) than the findings of previous study conducted in adolescent females of the United States (Houston et al., 2006). However, higher proportion of incorrect response regarding menstrual cycle length might either be due to the predominance of teen girls belonging to 1st year of university education or study subjects confusing menstrual cycle days with menstrual bleeding duration as 44.3% selected the option of 5–10 days (Table 2). Although 76.1% of our participants gave correct response regarding the normal menstrual bleeding duration, still it was significantly lower than the findings of Houston et al. (2006) who reported that 95.7% of American girls knew the correct menstrual bleeding duration. This might be attributable to the fact that adolescents in many cultures, including Pakistan, are not provided comprehensive information on sexual and reproductive health, considering discussions on such issues a taboo. A wide majority of the respondents’ had misconception regarding the source of bleeding as almost 70% stated the source of bleeding was vagina instead of uterus. These findings indicate the need to improve the menstrual knowledge of Pakistani girls by providing comprehensive menstrual education in schools, colleges and universities, conducting awareness programs regarding menstrual abnormalities and/or adding these topics in the curriculum/syllabus of college/university education.

Contrary to the findings of earlier study (Ali and Rizvi, 2010), eating patterns were less affected by the menstruation which was due to the fact that our study cohort comprised of girls acquiring tertiary education. However, it was worrisome to see that 31% respondents’ reported of eating lesser than normal during menstruation and this may cause iron-deficiency anemia as adolescent girls, particularly in developing countries, are more at risk because of inadequate diet, loss of substantial amount of iron during menstruation, and early marriages (Chaturvedi et al., 2017; Panat et al., 2013; Dixit et al., 2011). The myths (beliefs that certain foods will make them sick in the present or have deleterious effect in future) that prevailed among our study participants were avoiding “too cold foods” or “too hot foods” and decreased fluid intake which was consistent with the study conducted in Pakistani (Ali and Rizvi, 2010) and Saudi Arabian school girls (Moawed, 2001). Regarding hygienic practices, it was encouraging to see the majority of the study participants managed menstruation hygienically by using sanitary pads which was better than the findings of previous studies (Dasgupta and Sarkar, 2008; Khanna et al., 2005). Moreover, the absorbent that was not discarded was washed and dried in the sunlight (hygienic practice) in contrast to previously reported study in India (Khanna et al., 2005). In contrast to the previous study from Pakistan (Ali and Rizvi, 2010), majority of our study population took bath during menstruation. These good hygienic practices can be attributed to the better educational and economic status of the participants. It was also encouraging to observe that majority of study participants reported no absenteeism and impact on routine activities which was contradictory to the findings of earlier studies reporting absenteeism as well as social and religious restrictions (Houston et al., 2006; Moawed, 2001; El-Shazly et al., 1990). Our study had some limitations. The study included only the institutions of one city of Pakistan and study sample was comprised of university-going girls. Therefore, our findings may not be representative of entire population of Pakistani girls. Moreover, we recruited study subjects using a convenience sampling method so we might have drawbacks such as non-generalizability and selection bias. Yet our study provided an insight of menstrual knowledge and practices in Pakistani female university students.

Conclusions

Pakistani university-going girls have somewhat satisfactory menstrual knowledge, and dietary and hygienic practices. There are some misconceptions and malpractices related to menstruation and our findings highlight the need to conduct awareness programs in order to eradicate these misconceptions and malpractices.

Declarations

Author contribution statement

H. Mansoor: Conceived and designed the experiments; Performed the experiments; Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data.

M. Salman: Conceived and designed the experiments; Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data; Wrote the paper.

N. Asif: Performed the experiments; Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data.

Z. Mustafa: Analyzed and interpreted the data; Contributed reagents, materials, analysis tools or data; Wrote the paper.

A. Nawaz: Contributed reagents, materials, analysis tools or data; Wrote the paper.

J. Mohsin, B. Arif, A. Sheikh, N. Hira and A. Masood: Performed the experiments; Contributed reagents, materials, analysis tools or data.

N. Shehzadi and K. Hussain: Conceived and designed the experiments; Contributed reagents, materials, analysis tools or data; Wrote the paper.

Funding statement

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

Competing interest statement

The authors declare no conflict of interest.

Additional information

No additional information is available for this paper.