Prevalence of Plasmodium spp. in Gulshan-e-Iqbal, Sindh, Pakistan Vol. 13 (2), December 2022 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 R A D S J . B i o l . R e s . A p p l . S c i . 163 Op e n Ac c e s s F u l l L e n g t h A r t i c l e Prevalence of Plasmodium spp. in Gulshan-e-Iqbal, Sindh, Pakistan Aly Khan1,*, Iqra Sheikh2, Nasira Khatoon2, Samina Waheed1, S. Shahid Shaukat3, Adnan Khan4 1CDRI, Pakistan Agricultural Research Council, University of Karachi, Karachi-75270, Pakistan 2Department of Zoology, University of Karachi, Karachi-75270, Pakistan 3Institute of Environmental Studies, University of Karachi, Karachi-75270, Pakistan 4Department of Microbiology, University of Karachi, Karachi-75270, Pakistan A B S T R A C T Background: Malaria is caused by malaria parasite of Genus Plasmodium. It is transmitted by the bite of an infected Anopheles mosquito. Four species of Plasmodium namely P. malariae, P. falciparum, P. vivax and P. ovale are responsible for causing malaria in humans. The fifth one, P. knowlesiare is responsible to cause zoonotic infection in humans. Severe malaria may lead to death in humans. Objectives: To study the prevalence of Plasmodium spp. and to evaluate the percentage of infection in males, females and children of Gulshan-e-Iqbal, Karachi, Sindh, Pakistan. Methodology: Samples were collected from different laboratories and hospitals of Gulshan-e-Iqbal, Karachi, Sindh, Pakistan. Chi- square test for frequency data was performed. Results: A total of 411 cases were found to be positive out of 2096 suspected cases. P. vivaxwas more prevalent than P. falciparum. Out of 411 patients, 296 cases were of P. vivax, 112 were of P. falciparum, 2 were of P. malariaewhile one was of P. ovale. The frequency of different species was found to be highly significant (p<0.001). In age class <1-15 yrs, 16-40 yrs and 41->80 yrs the frequency differed significantly as indicated by chi-square test (p<0.001). Our results show that infection of malaria is more frequent in males as compared to females. Conclusion: Present survey shall be beneficial for the control of Plasmodium infection. Screening of patients who have malaria is important in other areas of Karachi as well. Additional studies with large sample sizes in other localities are required to fully understand malaria pathology in detail. The presence of P. ovaleand P. malariaecould be due to travel of subjects to African countries or Srilanka. Keywords Malaria, Plasmodium spp., Gulshan-e-Iqbal, Karachi, Pakistan. *Address of Correspondence aly.khan@hotmail.com Article info. Received: November 05, 2021 Accepted: November 21, 2022 Cite this article: Khan A, Sheikh I, Khatoon N, Waheed S, Shaukat SS, Khan A. Prevalence of Plasmodium spp. in Gulshan-e-Iqbal, Sindh, Pakistan. 2022; 13(2):163-167. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium provided the original work is properly cited. I N T R O D U C T I O N Malaria parasite (Genus Plasmodium) is a single celled microorganism which in human population inflicts huge burden in terms of complications leading to sudden unexplained death1. Plasmodium has many species out of which four species are responsible for inflicting humans (causing malaria) namely Plasmodium falciparum, P. malariae, P. ovaleand P. vivaxwhile the fifth one P. knowlesicauses zoonotic infection in humans2. The death rate due to malaria ranges between 1.5 and 2.7 million deaths each year3. It is endemic in 109 countries and widespread in the tropics and sub-tropics4. In Africa, Papua, New Guinea and Haiti P. falciparum is the most O R I G I N A L A R T I C L E Prevalence of Plasmodium spp. in Gulshan-e-Iqbal, Sindh, Pakistan Vol. 13 (2), December 2022 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 R A D S J . B i o l . R e s . A p p l . S c i . 164 common species while in parts of S. America, Middle East, North Africa and Indian subcontinent P. vivaxis endemic. In Pakistan it is a big public health problem as both P. vivaxand P. falciparum occur simultaneously. Poor sanitation, drug resistance, climate change and improper development activities are the main causes of spread of malaria5. In a previous study from Agha Khan University Hospital from 1997 to 2001 it was observed that percentage of P. vivaxwas 51.8 while that of P. falciparum was 46.5 with 1.5-2% mortality6. Almost ninety five million people here live in areas with high malaria incidence7. Cerebral malaria may cause sudden death, it is common when P. falciparum infects an individual. Sequestration of infected RBC within cerebral vessels is an important pathological reason of this disease. In places with endemic malaria patients with delirium and malaria must have at least three negative blood smears 8-12 days apart with microscopy in order to be considered fully recovered, although PCR test has shown to be more excessively affected than microscopy but does not provide any information of parasite load and is very expensive8,9. In the present study in Gulshan-e-Iqbal, Karachi 411 infected cases were studied for the type of Palsmodiuminfection besides infection rate in three different age groups <1-15, 16-40, 41->80, both male and female individuals was recorded. M A T E R I A L S A N D M E T H O D S A cross-sectional survey was carried out in Karachi to determine the prevalence of Plasmodium spp. targeting population in Gulshan-e-Iqbal, Karachi. A total of 411 cases were considered in this study out of 2096 suspected cases from Gulshan-e-Iqbal, Karachi, Sindh, Pakistan. Gulshan-e-Iqbal is in district east of Karachi, most of its population is working class having residential and commercial neighbourhood. The clinics and hospitals visited were namely DarulSehat hospital, LNH Laboratory Service, Dow University of Health Sciences, Sindlab Clinical Laboratory, Chughtai Laboratory, Ibn-e-Sena hospital, Dr. Essa Laboratory and Patel hospital over a period of 1 year from February 2019 to January 2020. The patients name, sex, age, details of clinical examination findings, antimalarial treatment if previously taken, history of blood transfusion were recorded in requisition form. After acquiring consents, 3 ml of blood specimen were collected from the antecubital vein from all the patients by using disposable syringe. Both thick and thin smears were prepared. The remaining blood was stored in a deep freezer at each laboratory (- 8±2ºF). The thick smear was dehaemoglobinized, slide stained with Leishmans stain. After fully drying the slides were observed under oil-immersion lens of microscope in Parasitology Section, Department of Zoology, University of Karachi. If at least one asexual form of parasite was detected in 100 microscopic field in thick film it was considered positive otherwise the report was termed negative. While the thin blood smears were thoroughly examined for malarial parasite. In positive cases the Plasmodium spp. was identified by a well trainedparasitologist. The study was approved by the Chairperson of Parasitology section, Department of Zoology, University of Karachi, Pakistan. Chi-square test for frequency data was performed10. R E S U L T S A total of 411 cases were found to be positive out of 2096 suspected cases. Four species of Plasmodium were identified. P. vivaxwas more prevalent than P. falciparum. Out of 411 patients, 296 cases were of P. vivax, 112 were of P. falciparum, 2 were of P. malariaewhile one was of P. ovale. Maximum number of cases 191 (46.47%) were observed in the age group 16-40 years, 123 (29.92%) in 1-15 years and 97 (23.61%) in the age group 41-80 years. The overall number of infection in the males was 246 out of which 175 (59.13%) were P. vivax, 69 (61.61%) were P. falciparum, one was P. malariaeand one was P. ovale. The overall number of infection in the females was 165 out of which 121 (40.87%) were P. vivax, 43 (38.39%) were P. falciparum and 1 was P. malariae. Table 1.Number of male and female infection of different species of Plasmodium in Gulshan-e-Iqbal, Karachi. Total cases Gender P. vivax P. falciparum P. malariae P. ovale Male Female Total Male Female Total Male Female Total Male Female Total Male Female 411 246 165 296 175 121 112 69 43 2 1 1 1 1 0 Chi-square=504.1, df=3, p<0.001 Prevalence of Plasmodium spp. in Gulshan-e-Iqbal, Sindh, Pakistan Vol. 13 (2), December 2022 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 R A D S J . B i o l . R e s . A p p l . S c i . 165 Table 2.Percentage of infection in both male and female of three different age groups. Number of positive cases Gender Age group Percentage % 123 ♂ ♀ <1-15 yrs 29.92 191 ♂ ♀ 16-40 yrs 46.47 97 ♂ ♀ 41->80 yrs 23.61 Chi-square=80.2, df=2, p<0.001 The frequency of type of malaria namely Plasmodium ovale, P. malariae, P. vivaxand P. falciparum was found to be highly significantly different as disclosed by chi- square test (chi-square=504.1, df=3, p<0.001) (Table 1). With respect to age classes <1-15 yrs, 16-40 yrs and 41- >80 years of patients (malaria cases) the frequency differed significantly as indicated by chi-square test (chi- square=80.2, df=2, p<0.001) (Table 2). D I S C U S S I O N Malaria is endemic in the province of Sindh, Pakistan. More than five lac cases are reported only from two provinces of Pakistan namely Balochistan and Sindh Province11. All four type of malaria namely Plasmodium ovale, P. vivax, P. falciparum and P. malariaewere found in Gulshan-e-Iqbal, Karachi, Sindh with respect to age classes of malaria patients the frequency differed significantly. Screening of patients in Karachi revealed the presence of P. vivaxto be two time higher than P. falciparum12 similar to the study conducted at Ayub Teaching hospital, Abbotabad13. One year data from Korangi creek area, Karachi was collected and found among 481 infected individuals 82.32 percent P. vivaxand 17.6 percent P. falciparum infection5. In a cross sectional study of Swat of Malakand division and district lower Dir (K.P) both thin and thick films were observed for Plasmodium infection. Overall positive percentage was 12%, P. vivax(99.07%) and P. falciparum (0.92%). No other Plasmodium species or mixed infection was recorded. Both chloroquine and Artemether were used for treatment14. It was recorded that the infection rate in children (5-15 years) P. vivaxwas more frequent (2.69%) than P. falciparum (0.35%) in rural areas of Bannu15 which was in agreement to the findings where was of the total of 11,353 malaria suspected samples studied by microscopy 1829/11353 (16.11 percent) samples were positive amongst which P. vivaxwas observed in 1825 subjects while P. falciparum was recorded only in 4 cases (0.2 percent)16. It was stated that in Bangladesh malaria exhibited highly seasonal and hypodermic transmission in geographic hotspots. Chittagong Hill Tracts remained malaria hotspot for a period of four years examined17. It was reported that at Rural Health Center, Sinawan, Muzzafargarh, district Punjab, amongst 10,023 suspected malaria cases 208 were confirmed as P. falciparum cases18. 135 cases of P. vivaxand 108 cases of P. falciparum amongst 241 cases of children visiting Tertiary care hospital, Karachi was recorded19. It is stated that not more than 20 percent population is availing any Government health facilities and many programmes lack facilities as have no microbiologist/parasitologist to detect occurrenece of Plasmodium is available11. Karachi being the biggest city of Pakistan with half the population of Sindh has very low annual parasite index (API)20. It was suggested that chloroquine drug has become inaffective for the treatment of P. falciparum in many countries including Aligarh, India due to development of resistance by the parasite which could be due to overexposure to improper therapeutic regimes, over the counter availability of drugs and pressure by improper prescribing practices by private doctors21. Forty seven positive patients were recorded for Plasmodium out of 210 suspected cases in blood smears. Out of 28, 59.57% were identified as P. vivaxand the rest 40.43% were P. falciparum in district Malakand, Khyber Pakhtunkhwa, Pakistan22. The recommended drug for both P. falciparum and P. vivaxis Artemisinin, other treatments for malaria include chloroquine, quinine, amodiaquine and doxycycline. C O N C L U S I O N Frequency of malaria causing species of Plasmodium was observed in total four species including P. vivax, P. falciparum, P. malariaeandP. ovalein Gulshan-e-Iqbal, Karachi, Sindh, Pakistan. The presence of P. ovaleand P. malariaecould be due to travel of subjects to African countries or Srilanka. The frequency of type of malaria Prevalence of Plasmodium spp. in Gulshan-e-Iqbal, Sindh, Pakistan Vol. 13 (2), December 2022 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 R A D S J . B i o l . R e s . A p p l . S c i . 166 and age classes of patients differed as well, so it is suggested that screening of patients who have malaria is important in other areas of Karachi. Additional studies with large sample sizes in other localities are required to fully understand malaria pathology in detail. E T H I C A L A P P R O V A L Ethical approval for cross-sectional survey was obtained from the Chairperson of Parasitology Section, Department of Zoology, University of Karachi. C O N F L I C T S O F I N T E R S T The authors declare no conflict of interest. F U N D I N G S O U R C E None. A C K N O W L E D G E M E N T S None. L I S T O F A B B R E V I A T I O N S spp. Species ml mili liter % Percentage df Degrees of freedom p Probability value R E F E R E N C E S 1. Menezes RG, Kanchan T, Rai S, Rao PPJ, Naik R, Shetty BSK, et al. An autopsy case of sudden unexplained death caused by malaria. J Forensic Sci. 2010; 55(3):835-838. 2. Lee K-S, Divis PCS, Zakaria SK, Matusop A, Julin RA, Conway DJ, et al.Plasmodiumknowlesi: Reservoir hosts and tracking the emergence in humans and macaques. PLoSPathog. 2011; 7(4):e1002015. DOI: 10.1371/journal.ppat.1002015. 3. Soyemi S, Faduyile A, Lawal O, Benebo A, Obafunwa J, Mordi V. Sudden death due to cerebral malaria in a Nigerian adult: a rare post mortem finding. World J Pathol.2013;2:48-52. 4. World Health Organization (WHO). World malaria report 2008. 2008. p. 190. 5. Khatoon N, Malik R, Khan A, Shaukat SS, Saifi ZS, Khan A. Incidence of malaria in the population of Korangi creek area, Karachi, Pakistan. Pak J Pharm Sci.2018; 31(6):2575-2578. 6. Zubairi ABS, Nizami S, Raza A, Mehraj V, Rasheed AF, Ghanchi NK, et al. Severe Plasmodium vivaxmalaria in Pakistan. Emerg Infect Dis.2013; 19(11):1851-1854. 7. Suwonkerd W, Ritthison W, Ngo CT, Tainchum K, Bangs MJ, Chareonviriyaphap T. Vector biology and malaria transmission in Southeast Asia. In: Manguin S, editors. Anopheles mosquitoes - new insights into malaria vectors, Vol. 10. InTech. 2013. DOI: 10.5772/56347. 8. Idro R, Jenkins NE, Newton CRJC. Pathogenesis, clinical features, and neurological outcome of cerebral malaria. Lancet Neurol. 2005; 4(12):827-840. 9. Olumese PE, Gbadegesin RA, Adeyemo AA, Brown B, Walker A. Neurological features of cerebral malaria in Nigerian children. Ann Trop Paediatr. 1999; 19(4):321-325. 10. Zar JH. Biostatistical analysis. 6th ed. Prentice Hall, Englewood Cliffs, N.J. 2008. 11. Nizamani MA, Kalar NA, Khushk IA. Burden of malaria in Sindh, Pakistan: A two years surveillance report. J LiaquatUniv Med Health Sci.2006; 5(2):76- 83. 12. Mahmood KH. Malaria in Karachi and other areas in Sindh. Pak Armed Forces Med J.2005; 55(4):345- 348. 13. Idris M, Sarwar J, Fareed J. Pattern of malarial infection diagnosed at Ayub Teaching hospital, Abbottabad. J Ayub Med Coll Abbottabad. 2007; 19(2):35-36. 14. Zaman N, Haq FU, Khan Z, Ullah W, Ualiyeva D, Waheed Y, et al. Incidence of malarial infection and response to antimalarial drugs at Districts of Lower Dir and Swat of Khyber Pakhtankhwa, Pakistan. Dialogues in Health. 2022; 1:100035. http://dx.doi.org/10.1016/j.dialog.2022.100035. 15. Awan Z, Shah HTA, Shah AH, Khan MA, Suleman M. Malaria among the students of religious schools of Bannu district Khyber Pakhtunkhwa, Pakistan. Pakistan J Zool.2012; 44(4):959-962. 16. Khan MW, Khan MN, Khan RA. Epidemiology and parasitological survey of malarial parasites in Khyber Pakhtunkhwa, Pakistan. J Pak Med Assoc. 2018; 68(1):145-146. 17. Noé A, Zaman SI, Rahman M, Saha AK, Aktaruzzaman MM, Maude RJ. Mapping the stability of malaria hotspots in Bangladesh from 2013 to 2016. Malar J.2018; 17:259. DOI: 10.1186/s12936-018- 2405-3. Prevalence of Plasmodium spp. in Gulshan-e-Iqbal, Sindh, Pakistan Vol. 13 (2), December 2022 ISSN (Print): 2305 – 8722 ISSN (Online): 2521 – 8573 R A D S J . B i o l . R e s . A p p l . S c i . 167 18. Sahar S, Akhtar T, Bilal H, Rana MS. Prevalence of Plasmodium falciparum, malarial parasites in Muzaffargarh district, Punjab-Pakistan: A two year study. Pak J Sci.2012; 64(1):64-66. 19. Maheshwari N, Shaikh M, Chand R, Maheshwari H, Yasir M. Malarial hepatopathy in children visiting a Tertiary Healthcare hospital in Karachi. Cureus.2020; 12(1):e6696. DOI: 10.7759/cureus.6696. 20. Prybylski D, Khaliq A, Fox E, Sarwari AR, Strickland GT. Parasite density and malaria morbidity in Pakistani Punjab. Am J Trop Med Hyg.1999; 61(5):791-801. 21. Shujatullah F, Khan HM, Khatoon A, Khan PA, Ashfaq M. In vitro chloroquine resistance in Plasmodium falciparum isolates from Tertiary care hospital. Malar Res Treat. 2012; 538481. doi: 10.1155/2012/538481. 22. Imran M, Ahmad I, Kalim M. Identification of Plasmodium vivax and Plasmodium falciparum in the Northern areas (District Malakand) of Khyber Pakhtunkhwa, Pakistan. PSM Microbiol. 2017; 2(3):59-62.