Anopheles subpictus Grassi: Observations on survivorship and population size using mark-release-recapture and dissection methods

A mark-release-recapture experiment to estimate population survivorship and absolute size was performed with wild-caught An. subpictus adults at the village of Khano-Harni, Lahore District, Punjab Province, Pakistan during September 1978, the end of the monsoon rainy season, when temporal population abundance was maximized. Daily survival rate estimated from the recapture sequence of marked adults was low, males=0.192 and females=0.343. Survivorship for females estimated by several vertical age-grading procedures ranged from 0.347 to 0.628. Both stage- and age-specific life tables were calculated from vertical age-grading data determined by the dilatation method. Female and male population size was estimated byBailey 's modification of theLincoln Index and was found to average 4478.4 and 6106.8, respectively. The bionomics, survivorship and population size of An. subpictus in the Lahore are indicated that this species was probably not important in the transmission of human malaria.     

Adult anopheline ecology and malaria transmission in irrigated areas of South Punjab, Pakistan

Surface irrigation in the Punjab province of Pakistan has been carried out on a large scale since the development of the Indus Basin Irrigation System in the late 19th century. The objective of our study was to understand how the population dynamics of adult anopheline mosquitoes (Diptera: Culicidae) could be related to malaria transmission in rural areas with intensive irrigation and a history of malaria epidemics. In this paper we present our observations from three villages located along an irrigation canal in South Punjab. The study was carried out from 1 April 1999 to 31 March 2000. Mosquitoes were collected from bedrooms using the pyrethroid spraycatch method and from vegetation and animal sheds using backpack aspirators. Overall, Anopheles subpictus Grassi sensu lato predominated (55.6%), followed by An. stephensi Liston s.l. (41.4%), An. culicifacies Giles s.l. (2.0%), An. pulcherrimus Theobald (1.0%) and An. peditaeniatus Leicester (0.1%). Most mosquitoes (98.8%) were collected from indoor resting-sites whereas collections from potential resting-sites outdoors accounted for only 1.2% of total anopheline densities, confirming the endophilic behaviour of anophelines in Pakistan. Anopheles stephensi, An. culicifacies and An. subpictus populations peaked in August, September and October, respectively. High temperatures and low rainfall negatively affected seasonal abundance in our area. There were interesting differences in anopheline fauna between villages, with An. culicifacies occurring almost exclusively in the village at the head of the irrigation canal, where waterlogged and irrigated fields prevailed. Monthly house-to-house fever surveys showed that malaria transmission remained low with an overall slide positivity rate of 2.4% and all cases were due to infection with Plasmodium vivax. The most plausible explanation for low transmission in our study area seems to be the low density of Pakistan's primary malaria vector, An. culicifacies. The role of other species such as An. stephensi is not clear. Our observations indicate that, in South Punjab, irrigation-related sites support the breeding of anopheline mosquitoes, including the vectors of malaria. As our study was carried out during a year with exceptionally hot and dry climatic conditions, densities and longevity of mosquitoes would probably be higher in other years and could result in more significant malaria transmission than we observed. To assess the overall importance of irrigation-related sites in the epidemiology of malaria in the Punjab, more studies are needed to compare irrigated and non-irrigated areas.     

Breeding of Anopheles mosquitoes in irrigated areas of South Punjab, Pakistan

As part of investigations on potential linkages between irrigation and malaria transmission, all surface water bodies in and around three villages along an irrigation distributary in South Punjab, Pakistan, were surveyed for anopheline mosquito larvae (Diptera: Culicidae) from April 1999 to March 2000. Samples were characterized according to exposure to sunlight, substratum, presence of vegetation, fauna, inorganic matter and physical water condition (clear/turbid/foul). Also water temperature, dissolved oxygen (DO), electroconductivity (EC) and pH of sites were recorded. A total of 37982 Anopheles larvae of six morphological types were collected from 2992 samples taken from irrigation/agricultural and village/domestic aquatic habitats. Anopheles subpictus Grassi sensu lato was by far the most abundant (74.3%), followed by An. culicifacies Giles s.l. (4.1%), An. stephensi Liston s.l. (2.6%), An. pulcherrimus Theobald (1.8%), An. peditaeniatus Leicester (0.3%) and An. nigerrimus Giles (0.1%). The four most abundant species were significantly associated with waterlogged fields and communal village drinking-water tanks. Habitat characteristics most correlated with occurrence of anophelines were the physical water condition and the absence/presence of fauna, particularly predators. Occurrence and abundance of Anopheles immatures were not significantly correlated with water temperature, DO, EC or pH. Malaria vectors of the Anopheles culicifacies complex occurred at relatively low densities, mainly in irrigated and waterlogged fields. In South Punjab, where rainfall is very low, it should be possible to reduce anopheline breeding through water management, as larvae develop mainly in water bodies that are directly or indirectly related to the extensive canal-irrigation system.     

Double heterozygous hemoglobin Q India/hemoglobin D Punjab hemoglobinopathy: Report of two rare cases

Cation exchange high performance liquid chromatography (CE HPLC) provides an excellent tool for accurate and reliable diagnosis of various hemoglobin (Hb) disorders. HbQ India is a rare alpha chain variant that usually presents in the heterozygous state. Its presence in double heterozygous state with HbD Punjab is extremely rare. The double heterozygosity for α and β chain variants leads to formation of abnormal heterodimer hybrids, which can lead to diagnostic dilemmas. We report two rare cases of double heterozygous HbQ India/HbD Punjab where the hybrid Hb was seen to elute at retention time similar to HbC on CE HPLC. The first case had unconjugated hyperbilirubinemia at presentation; while, the second case was asymptomatic.     

Effects of Migration on Genetic Variability and Parent–Offspring Correlation Coefficients in Anthropometric and Physiometric Phenotypes Among Three Groups of Population in Punjab, India

In the present study, we examined the migration effects on genetic variabilities and heritabilities patterns between three groups of population like parental population in Punjab, migrant from Pakistan, and migrant from other states of India in Punjab using anthropometric and physiometric traits. A total of 500 adult individuals from 300 families were studied. Statistical comparisons were carried out through mean coefficients, Student’s t test, heritability, and regression analysis. The results suggest a significant migration effect on almost all traits. Correlation coefficient for first-degree relatives, the slope factors, and heritabilities for almost all variables have been found significant among the three groups of populations. However, the discrimination is more prominent among migrant from other states of India because of more genetic heterogeneity.     

Influence of weather on the moth catches of Helicoverpa armigera at various cotton-based agro-ecological sites

Pheromones are innate products, exploited for direct population manipulation in the trapping of precise pest density. They demonstrate feasibility and efficiency against Helicopverpa armigera at five different cotton-based agro-ecological sites: river site, vegetable farm, orchard, clean cultivation site, and forest. The data were recorded at 15–20 day intervals from 8 September 2004 to 1 December 2006. The Number of moths caught during the study period demonstrated that the vegetable farm showed the maximum population of moths per trap (0.54), followed by the orchard (0.29), forest (0.19), river (0.15), and clean cultivation site (0.12). During 2004, rainfall (in the orchard and clean cultivation site) and relative humidity (in the clean cultivation site) showed positive and significant correlation with the number of moths caught. During 2005, temperatures played a significant and positive role in population fluctuation of the pest almost at all the selected sites; the minimum temperature was recorded at the vegetable farm and the average temperature was found at the forest site. Rainfall also showed a significant and positive correlation with trapped moths in the vegetable farm, orchard, and clean cultivation site. During 2006, relative humidity showed a significant and negative correlation with the trapped population at all the sites except the river site. Rainfall showed a negative and significant correlation with the pest at the forest site. Relative humidity was the most important aspect at 8.93% causing variation in the trapped population followed by maximum temperature (5.7%), rainfall (1.63%), average temperature (0.74%), and minimum temperature (0.56%).     

Effect of weather factors on populations of Helicoverpa armigera moths at cotton-based agro-ecological sites

Pheromone trapping was used to monitor populations of the moth Helicoverpa armigera at five cotton‐based agro‐ecological sites – river, vegetable, orchard, forest and clean cultivation (areas under only cotton cultivation) – in the Bahawalpur district, Pakistan. Three locations at each site were chosen and three pheromone traps at each location were installed in cotton fields. Moth catches were recorded at 15–20 day intervals from 24 October 2004 to 19 December 2006. In 2004, the river sites showed the maximum trapped population of H. armigera (0.22/trap) followed by 0.165 per trap at the vegetable sites. Orchard, clean cultivation and forest sites had zero moth catches. In 2005, the river sites again showed the highest trapped population (0.57/trap), followed by clean cultivation (0.45/trap), vegetable (0.44/trap), orchard (0.40/trap) and forest (0.29/trap). The moths appeared during July to December and March to May. In 2006, sites showed non‐significant difference, with a population range of 0.47 to 0.97 moths per trap. On average, river sites peaked at 0.49 per trap, followed by vegetable (0.38), clean cultivation (0.47), orchard (0.35) and forest (0.25) sites. The peak was observed on 3 April 2006, and moths appeared during February to July and October to December. The minimum temperature in river, forest and clean cultivation sites; the maximum temperature in orchard sites; and the average temperature in river, orchard, forest and clean cultivation sites showed significant positive correlations with trapped moth populations. Relative humidity showed significant negative correlation with population at the orchard sites in 2005. All weather factors during 2004 and 2006 showed non‐significant correlations with the moth populations. No model was found to be best fit by multiple linear regression analysis; however, relative humidity, minimum temperature, maximum temperature, minimum temperature and maximum temperature contributed 8.40, 10.23, 2.43, 4.53 and 2.53% to the population fluctuation of the moth at river, vegetable, orchard, forest and clean cultivation sites, respectively.     

Effects of Environmental Pesticides on the Health of Rural Communities in the Malwa Region of Punjab,India: A Review

The Malwa region of Punjab, India, is facing an unprecedented crisis of environmental health linked to indiscriminate, excessive, and unsafe use of pesticides, fertilizers, and poor groundwater quality. The region has been described as India's “cancer capital” due to abnormally high number of cancer cases, which have increased 3-fold in the last 10 years. Studies of this region have also highlighted a sharp increase in many other pesticide-related diseases, such as mental retardation and reproductive disorders. The most affected individuals are the agricultural workers who are directly exposed to pesticides. The Malwa region of Punjab, India, is less than 15% of the total area of Punjab (only 0.5% of the total geographical area of India), but it consumes nearly 75% of the total pesticides used in Punjab. The high use of pesticides, along with environmental and social factors, is responsible for the high concentration of pesticide residues in the food chain of this region. Moreover, many banned and restricted pesticides are still in use in this region, warranting strict periodical health checkups and other interventions. The present review describes occupational, environmental, and social factors associated with pesticide use in the Malwa region of Punjab, India, and proposes some risk reduction interventions.     

Evaluation of DNA damage in agricultural workers exposed to pesticides using single cell gel electrophoresis (comet) assay

BACKGROUND:

Pesticides are used in agriculture to protect crops, but they pose a potential risk to farmers and environment. The aim of the present study is to investigate the relation between the occupational exposure to various pesticides and the presence of DNA damage.

MATERIALS AND METHODS:

Blood samples of 210 exposed workers (after a day of intense spraying) and 50 control subjects belonging to various districts of Punjab (India) were evaluated using Comet assay. Sixty workers who showed DNA damage were selected for follow up at 5-6 months after the first sampling during a low or null spraying period.

RESULTS:

Significant differences were found in DNA damage between freshly exposed workers and controls and freshly exposed and followed up cases. There was significant increase in the comet parameters viz. mean comet tail length and frequency of cells showing migration in exposed workers as compared to controls (72.22 ± 20.76 vs. 46.92 ± 8.17, P<0.001; 31.79 vs. 5.77, P<0.001). In the second samples, followed up cases showed significant decrease in frequency of damaged cells as compared to freshly exposed workers of first sampling (P<0.05). The confounding factors such as variable duration of pesticide exposure, age, smoking, drinking and dietary habits etc which were expected to modulate the damage, were instead found to have no significant effect on DNA fragmentation.

CONCLUSION:

The evidence of a genetic hazard related to exposure resulting from the intensive use of pesticides stresses the need for educational programs for agricultural workers to reduce the use of chemicals in agriculture.     

An independent occurrence of the chimeric P450 enzyme CYP337B3 of Helicoverpa armigera confers cypermethrin resistance in Pakistan

The increasing resistance level of insect pest species is a major concern to agriculture worldwide. The cotton bollworm, Helicoverpa armigera, is one of the most important pest species due to being highly polyphagous, geographically widespread, and resistant towards many chemical classes of insecticides. We previously described the mechanism of fenvalerate resistance in Australian populations conferred by the chimeric cytochrome P450 monooxygenase CYP337B3, which arose by unequal crossing-over between CYP337B1 and CYP337B2. Here, we show that this mechanism is also present in the cypermethrin-resistant FSD strain from Pakistan. The Pakistani and the Australian CYP337B3 alleles differ by 18 synonymous and three nonsynonymous SNPs and additionally in the length and sequence of the intron. Nevertheless, the activity of both CYP337B3 proteins is comparable. We demonstrate that CYP337B3 is capable of metabolizing cypermethrin (trans- and especially cis-isomers) to the main metabolite 4'-hydroxycypermethrin, which exhibits no intrinsic toxicity towards susceptible larvae. In a bioassay, CYP337B3 confers a 7-fold resistance towards cypermethrin in FSD larvae compared to susceptible larvae from the Australian TWB strain lacking CYP337B3. Linkage analysis shows that presence of CYP337B3 accounts for most of the cypermethrin resistance in the FSD strain; up-regulation of other P450s in FSD plays no detectable role in resistance. The presence or absence of CYP337B3 can be easily detected by a simple PCR screen, providing a powerful tool to rapidly distinguish resistant from susceptible individuals in the field and to determine the geographical distribution of this resistance gene. Our results suggest that CYP337B3 evolved twice independently by unequal crossing-over between CYP337B2 and two different CYP337B1 alleles.