Mutational and expressional association of the PIK3CA gene with the risk of breast cancer in the Pakistani population.

PI3K pathway is a very important pathway that is reported to be involved in breast cancer. Mutation of PI3K and p110 alpha-catalytic subunit of phosphatidylinositol 3-kinase (PIK3CA) is of high predictive and prognostic values in breast cancer. The purpose of the current study was to screen the hotspot mutations of the PIK3CA gene i.e. rs2677760, rs3806685, rs121913273 & rs121913279 along with expressional analysis of PI3K and PIK3CA genes in breast cancer female patients. For mutational analysis, TaqMan assay & Sanger sequencing were performed while for expressional analysis real-time PCR was carried out. Mutant allele C of rs2677760 was observed to be high in postmenopausal patients. The frequency of mutant allele G of rs3806685 was significantly high in breast cancer patients. All diseased and control samples were of wild type for the hotspot rs121913273 and rs121913279 with allele G for rs121913273 and A for rs121913279. Expression of the PI3K was high in breast cancer tissue samples as compared to the adjacent controls. While the expression of the thePIK3CA gene was significantly high in premenopausal breast cancer patients. It was concluded that the mutant allele C of rs2677760 might have some sort of association with the menopausal status and it could be used as a diagnostic marker in post-menopausal women if studied further. Mutant allele G of rs3806685 was also found to be associated with breast cancer. While multiallelic rs121913273 and rs121913279 showed a different trend for the studied population. For expressional analysis, PI3K showed over-expression in the cases while PIK3CA gene expression was observed to be significantly associated with pre-menopausal status.

Comparative Analysis of Different ELISA Methods for the Serodiagnosis of Przhevalskiana silenus Infestation in Goats.

Przhevalskiana silenus (warble fly) grubs cause myiasis in goats, in mountainous and semi-mountainous areas and different regions in Pakistan, and cause substantial losses to livestock. The palpation method for detecting warble flies generally neglects the infestation intensity; therefore, the development of a reliable and efficient diagnostic technique is extremely necessary. This study compared three indirect enzyme-linked immunosorbent assay (ELISA) methods for detecting anti-P. silenus antibodies using the hypodermin C (HyC) purified from Hypoderma spp. Larvae collected in cattle (local isolate, Microbiology Laboratory, PMAS-Arid Agriculture University, Rawalpindi), the crude antigen from the first instar stage of P. silenus, and a commercial Bovine Hypodermosis Antibody ELISA kit (IDEXX Laboratory), for accurately estimating the seroprevalence of goat warble fly infestation (GWFI) in the Pothwar plateau, Punjab, Pakistan. The ELISA with the crude antigen of P. silenus proved very sensitive and specific, 91% and 93%, respectively. The optical density exhibited a monthly variation, and the antibody titer began increasing from June, continually increased from July to December, and gradually decreased thereafter until March. The study confirmed the endemic status of GWFI in the Pothwar region and identified that ELISA based on the crude antigen of P. silenus was a more sensitive and specific immunodiagnostic method for determining seroprevalence, and could be employed for initiating nationwide eradication campaigns.

A classification of genes involved in normal and delayed male puberty.

Puberty is a pivotal biological process that completes sexual maturation to achieve full reproductive capability. It is a major transformational period of life, whose timing is strongly affected by genetic makeup of the individual, along with various internal and external factors. Although the exact mechanism for initiation of the cascade of molecular events that culminate in puberty is not yet known, the process of pubertal onset involves interaction of numerous complex signaling pathways of hypothalamo-pituitary-testicular (HPT) axis. We developed a classification of the mechanisms involved in male puberty that allowed placing many genes into physiological context. These include (i) hypothalamic development during embryogenesis, (ii) synaptogenesis where gonadotropin releasing hormone (GnRH) neurons form neuronal connections with suprahypothalamic neurons, (iii) maintenance of neuron homeostasis, (iv) regulation of synthesis and secretion of GnRH, (v) appropriate receptors/proteins on neurons governing GnRH production and release, (vi) signaling molecules activated by the receptors, (vii) the synthesis and release of GnRH, (viii) the production and release of gonadotropins, (ix) testicular development, (x) synthesis and release of steroid hormones from testes, and (xi)the action of steroid hormones in downstream effector tissues. Defects in components of this system during embryonic development, childhood/adolescence, or adulthood may disrupt/nullify puberty, leading to long-term male infertility and/or hypogonadism. This review provides a list of 598 genes involved in the development of HPT axis and classified according to this schema. Furthermore, this review identifies a subset of 75 genes for which genetic mutations are reported to delay or disrupt male puberty.

Genetic analysis of failed male puberty using whole exome sequencing.

OBJECTIVES: Although at least 598 genes are involved in the development of the hypothalamo-pituitary-testicular (HPT) axis, mutations in only 75 genes have so far been shown to cause delayed puberty. METHODS: Six male patients with failed puberty, manifested as absence of pubertal changes by 18 years of age, underwent whole exome sequencing of genomic DNA with subsequent bioinformatics analysis and confirmation of selected variants by Sanger sequencing. Genes having plausibly pathogenic non-synonymous variants were characterized as group A (previously reported to cause delayed puberty), group B (expressed in the HPT-axis but no mutations therein were reported to cause delayed puberty) or group C (not reported previously to be connected with HPT-axis). RESULTS: We identified variants in genes involved in GnRH neuron differentiation (2 in group A, 1 in group C), GnRH neuron migration (2 each in groups A and C), development of GnRH neural connections with supra-hypothalamic and hypothalamic neurons (2 each in groups A and C), neuron homeostasis (1 in group C), molecules regulating GnRH neuron activity (2 each in groups B and C), receptors/proteins expressed on GnRH neurons (1 in group B), signaling molecules (3 in group C), GnRH synthesis (1 in group B), gonadotropins production and release (1 each in groups A, B, and C) and action of the steroid hormone (1 in group A). CONCLUSIONS: Non-synonymous variants were identified in 16 genes of the HPT-axis, which comprised 4 in group A that contains genes previously reported to cause delayed puberty, 4 in group B that are expressed along HPT-axis but no mutations therein were reported previously to cause delayed puberty and 8 in group C that contains novel candidate genes, suggesting wider genetic causes of failed male puberty.

Prevalence and Distribution of Hard Ticks and Their Associated Risk Factors in Sheep and Goats from Four Agro-Climatic Zones of Khyber Pakhtunkhwa (KPK), Pakistan.

Hard ticks are blood-sucking ectoparasites infesting all types of domestic ruminants throughout the world and serve as vectors in the dissemination of a wide variety of pathogens. Sheep and goat farming is a vital economic source for resource-poor farming communities in the Khyber Pakhtunkhwa (KPK) province of Pakistan. Aim: The aim of present study is to study the epidemiological profile of ticks in various agro-climatic zones of KPK. Materials and Methods: A total of 1500 (882 sheep and 618 goats) of all age groups and sexes were examined for the presence of ticks belonging to six districts in four agro-climatic zones of KPK province, Pakistan. Non-probability sampling was undertaken based on animal hosts' selection, collection of hard ticks, and epidemiological parameters. Ticks collected from sheep and goats were identified phenotypically using standard keys. Results: The results revealed that Rhipicephalus microplus (36.2%) was the predominant tick species followed by Hyalomma anatolicum (25.2%), Hyalomma trancatum (18.1%), Hyalomma aegyptium (11.3%), Hyalomma asiaticum (6.9%), and Haemephysalis bispinosa (2.4%). Different epidemiological parameters showed that weather, humidity, and host age negatively affect (p < 0.05) tick load while temperatures (Minimum and Maximum), sunshine (hrs), and precipitation positively affected (p < 0.05) tick load. Host sex only showed a positive association with tick load (p > 0.05). The highest value of tick diversity index (H) 0.36748 was noted for Hyalomma anatolicum as this tick species was commonly distributed in three agro-climatic zones except in the Suleiman Piedmont zone, while Haemaphysalis bispinosa has the lowest diversity index (0.0334) as it was only found in the Central Valley Plains zone of KPK. The Shannon diversity index of tick diversity was highest at Barikot while the lowest index value was at Kabal (2.407). Similarly, a higher Simpson's diversity index was at Barikot (0.9554) and the lowest hard index was noted at Takht bai (2.874). The dominance index showed that the tick species were more dominant at Takht bai (0.1251), while the lowest dominance was at Barikot (0.04465). Conclusions: It has been concluded that tick diversity and distribution, abundance, reproduction, development, and survival depend on prevailing climatic conditions. The present study would not only help to identify the tick species but also facilitate in devising effective control measures to minimize the transmission pathogens in sheep and goats being reared in the various agro-climatic zones of KPK province, Pakistan.

Natural plant extracts mediated expression regulation of TGF-ß receptors and SMAD genes in human cancer cell lines.

BACKGROUND: Transforming growth factor beta (TGF-ß) superfamily has key role in cell proliferation which leads to tumor promoting activities at metastatic stage of cancer. Inhibition of transforming growth factor beta receptor (TGFßR) signaling pathway can provide better therapeutic strategy to control cancer. Natural products are best known for their safety, less toxic nature, antioxidant characteristics making them a promising candidate to inhibit TGFßR signaling pathway. METHODS AND RESULTS: Crude methanolic extracts (CMEs) of 16 selected plants were prepared by using maceration method and subjected to phytochemical assays for identification of major phytometabolites particularly cancer chemopreventive antioxidant constituents. Total flavonoid content of all plants CME was > 0.6 mg/ml exhibiting the Cichorium intybus contains comparatively highest amount of total flavonoid content (0.53 mg/ml). Scanvenging activity of all plants was determined having IC50 ranges between 2 and 88 (µg/ml) while Moringa oleifera revealed the maximum scavenging activity (IC50 2.03 µg/ml). Comparative cytotoxicity of plant extracts was evaluated in HUH and MCF-7 cell lines using 3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide (MTT) colorimetric assay. The nine active plant extracts i.e. Fagonia cretica, Argemone Mexicana, Rubus fruticosus, M. oleifera, Punica granatum, Cichorium intybus, Xanthium strumarium, Carissa opaca, Cyperus rotundus were identified based on their high antiproliferative activity > 50% against cancer cell lines and subjected to relative expression studies. Modulation of TGFß signaling molecules (i.e.TGFßR1, 2 & 3, SMAD3, SMAD5) and ubiquitous proteins i.e. SMURF1 and SMURF2 genetic expression by potent extracts was determined by RT-PCR using GAPDH (housekeeping gene) as gene of reference. CONCLUSIONS: This present study revealed that CME of Fagonia cretica and Argemone mexicana significantly inhibit TGF beta mediated signaling cascade by downregulating the gene expression fold change > 1 of TGFßR 1, 2 & 3 and receptor associated complex protein SMAD3 as compared to control.

The association between chronic stress, hair cortisol, and angiographically documented coronary atherosclerosis, a case-control study.

Cardiovascular diseases remain the principal contributor to global mortality and morbidity. Chronic stress has emerged as a strong prognostic factor for cardiovascular diseases. We aimed to measure hair cortisol concentration (HCC) in patients with angiographically confirmed coronary atherosclerosis and find the association between HCC and classic cardiovascular risk factors in a case-control study. The study included 500 angiographically confirmed coronary atherosclerosis patients and 500 age and sex-matched controls having no coronary stenosis. Hair cortisol concentration was assessed from a single sample of scalp hair by ELISA. Perceived stress scale 10 was used to evaluate the stress level of the participants. Spearman rank correlation was applied to find the association between hair cortisol level and classic cardiovascular risk factors. Multivariate regression was executed to assess the independent contribution of hair cortisol concentration as a coronary atherosclerosis risk factor. Median hair cortisol concentration was significantly high in patients with angiographically documented coronary atherosclerosis compared to controls (158(17.6-1331.3) pg/mg versus 73 (13.4-889) pg/mg. Hair cortisol concentration showed a significant positive correlation with BMI r = 525 and Hb1Ac r = 665 (both, p = ≤0.0001). In multivariate logistic regression analysis after adjusting for all the confounding variables, HCC, matched odds ratio (MOR), 10.73;95%CI (1.60,42.09) p-value = 0.001 remained the most significant predictor of coronary atherosclerosis.LAY SUMMARYHair cortisol concentrations (HCC) were significantly high in patients with angiographically confirmed coronary atherosclerosis compared to controls. HCC showed a significant positive correlation with diabetes and obesity and remained a major predictor of coronary atherosclerosis in the final analysis.

Calcium based siRNA coating: a novel approach for knockdown of HER2 gene in MCF-7 cells using gold nanoparticles.

Surface functionalization of nanoparticles (NPs) for therapeutic siRNA delivery into cancer cells has gained interest. The present study was designed for surface functionalization of gold nanoparticles (AuNPs) for efficient siRNA delivery and knockdown in cancer cells. In order to achieve this objective, AuNPs were coated with HER2-siRNA in the presence of 11-mercaptoundecanoic acid (11-MUA), calcium chloride (CaCl2) and polyethyleneimine (PEI) in alternate charge bearing successive layers. MCF-7 cells were cultured and transfected with fabricated assembly of AuNPs. Cytotoxicity analysis revealed that the half inhibitory concentration (IC50) for the formulation was 45.35 nM  . Total RNA was isolated from transfected cells, reverse transcribed into complementary DNA (cDNA) and real-time polymerase chain reaction (RT-PCR) was performed. The RT-PCR based delta-delta Ct analysis in treated cells revealed a significant 18.94 times decrease (p<0.001) in the expression of HER2 gene standardized with ACTB housekeeping gene as compared to untreated cells, which makes this formulation a potent approach for siRNA delivery and  gene knockout.

Seroepidemiology of human fascioliasis and its relationship with anti-Fasciola IgG and liver enzymes as biomarkers of pathogenicity.

BACKGROUND: Fascioliasis has never been considered a public health concern in Pakistan, although the increasing numbers of human cases reported in south Asia need a re-consideration in the country. The current study aimed to find the seroprevalence of human fascioliasis, associated risk factors and its relationship with liver enzymes as biomarkers of pathogenicity. METHODS: The cross-sectional study was conducted in different districts of Punjab region from May 2014 to August 2016. A total of 546 respondents were screened by using enzyme-linked immunosorbent assay (ELISA) and serum biochemical tests. RESULTS: Higher seroprevalence was recorded in Muzaffargarh (6.2%) and Bhara kahu (5.9%), while low infection rate in Gujranwala (1.1%) and Islamabad (1.5%). The results of multiple logistic regression analysis showed rural inhabitants (OR=7.9, 95%CI: 2.5-24.8), females (OR=3.5, 95%CI: 1.7-7.1), family size 3-7 (OR=1.7, 95%CI: 1.0-2.9) and socioeconomic condition (OR=3.9, 95%CI: 1.5-10.4) were the significantly (p<0.005) associated risk factors with disease. The results of liver enzymes i.e. aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, gamma-glutamyl transferase and cholesterol levels were significantly (p=0.001) elevated and associated with fascioliasis pathogenicity. CONCLUSION: The higher prevalence recorded may explain with Fasciola IgG antibodies for both active and past infections and cross reactivity of the assay with other helminthes.

Seroprevalence and associated risk factors of Entamoeba histolytica infection among gastroenteritis patients visiting the public healthcare system, Pakistan.

OBJECTIVES: To investigate the seroprevalence and associated risk factors of entamoeba histolytica among patients with gastrointestinal complaints, and to measure the eventual changes in serum biochemical parameters to reflect its pathogenicity. METHODS: The cross-sectional study was conducted in different hospitals of Potohar region in Punjab province and in the Khyber Pakhtunkhwa province of Pakistan from September 2015 to February 2017, and comprised individuals of either gender belonging to diverse backgrounds, inhabiting different areas of the country. The patients were enrolled from among those who visited outpatient departments with complaints of vague abdominal pain, nausea, vomiting, indigestion and diarrhoea. Blood samples were screened by using enzyme-linked immunosorbent assay and serum biochemical tests. Data was analysed using SPSS 20. RESULTS: Of the 356 subjects, 238(66.9%) were females and 118 (33.1%) were males. The overall mean age was 33.4}11.05 years. Seroprevalence of entamoeba histolytica was 356(73%). The infection rate did not differ significantly (p>0.05) among cities, while the highest infection was recorded in Islamabad 91(25.5%). The participants in rural areas had 2.16-fold higher risk of infection compared to urban areas, while the lowest risk of infection among people aged 50years compared to those aged 40-49 years (p=0.04). The amoebiasis was significantly associated with eating unwashed raw vegetables and average toilet facilities. Among clinical complications, haemodynamic changes, jaundice, vomiting, haemoglobin level, loose motion, intolerance to oral feeding, and history of antibiotics were significant associated variables (p<0.05 each). Significant elevation in alkaline phosphatase, aspartate aminotransferase, total protein and globulin levels were positively associated with amoebiasis (p<0.01 each). CONCLUSIONS: In entamoeba histolytica -positive patients ,serum biochemical level was found elevated and the risk factors determined were eating unwashed vegetables, toilet facilities, age, locality, jaundice, vomiting, haemoglobin level, loose motion, intolerance to oral feeding, and history of antibiotics.

An investigation on the prevalence and efficiency of immunochromatographic testing in suspected malarial patients of Rawalpindi and Islamabad, Pakistan.

BACKGROUND/AIM: The prevalences of Plasmodium falciparum and Plasmodium vivax are increasing rapidly in Pakistan, but recent data on the epidemiology of malaria are not properly reported with scarce diagnostic methods for quick diagnosis. This study was designed to determine the current prevalence and distribution of Plasmodium species in the vicinity of Rawalpindi and Islamabad and report on the validity of the immunochromatographic test (ICT) in diagnosing malarial infections. MATERIALS AND METHODS: A total of 1500 blood samples obtained from a local hospital were screened during the course of this study via microscopic examination and ICT. RESULTS: It was seen that malaria was highly endemic in this region. Both P. vivax and P. falciparum were prevalent in all age groups with high seasonal variations, showing a summer peak for P. vivax and a winter peak for P. falciparum. In a comparative study of the diagnostic methods it was observed that the ICT is 95% sensitive and 100% specific for both P. falciparum and P. vivax, while microscopic study was 100% sensitive and 96.8% specific. CONCLUSION: Epidemiological study of the malarial parasites showed that majority of the patients were from Rawalpindi as compared to Islamabad and that P. vivax was the dominant cause of malarial infection.

Risk mapping of bovine hypodermosis using geographical information system (GIS) in cattle of subtropical region, Pakistan.

INTRODUCTION: Hypodermosis is an ectoparasitic disease of cattle caused by Hypoderma lineatum and Hypoderma bovis. It is an important health problem of cattle, leading to considerable economic losses. There are various factors that are involved in the spread of this disease such as herd size, location, temperature, humidity, and precipitation. METHODOLOGY: Blood samples from 112 herds were collected to determine the presence of Hypoderma spp. infestation. For these herds, size and location were determined; temperature, humidity, and precipitation data were obtained from meteorological stations; and topographic features were obtained from existing maps and through field work. A regression analysis was then used to generate a risk factor analysis profile for hypodermosis and geographic information system (GIS) was used to map the risks. RESULTS: The GIS map developed showed the degree of infestation in different geographical locations at district and village levels. Cluster analysis demonstrated that hypodermosis prevalence varied within zones and across zones. The regression analysis showed that the temperature in the months of January, February, March, August, and November, and the precipitation in September and October had significant results (p < 0.05) when all the risks factors were analyzed. CONCLUSIONS: It is concluded that different ecological factors have an important impact on the intensity and infestation rate of hypodermosis across the globe. The present study might be used to control and eradicate the hypodermosis across the globe.

Anthelmintic activity of Artemisia vestita Wall ex DC. and Artemisia maritima L. against Haemonchus contortus from sheep.

Current study was designed to evaluate in vivo and in vitro anthelmintic activity of Artemisia vestita Wall ex DC. and Artemisia maritima L. against Haemonchus contortus in comparison with ivermectin to investigate the effect of plant extracts on survival of infective L3 and adults under in vitro condition. Plant extracts were given to H. contortus infected sheep orally and it was infected with L3 stage of H. contortus at dose of 5000 larvae/sheep. Total of 25-30 larvae were incubated with plant extracts in PBS alone and ivermectin at different concentration used as positive control. It was recorded that there is a significant decrease in fecal egg count (FEC) after post-treatment period for both plants. The highest fecal egg count reduction for A. vestita was 87.2% at 100mg/kg while for A. maritima it was 84.5% on day 28 post-treatment. Investigated extracts indicated significant activity against larvae and adult worms.

Empirical evidence of cold stress induced cell mediated and humoral immune response in common myna (Sturnus tristis).

Common myna (Sturnus tristis) is a bird indigenous to the Indian subcontinent that has invaded many parts of the world. At the onset of our investigation, we hypothesized that the immunological profile of myna makes it resistant to harsh/new environmental conditions. In order to test this hypothesis, a number of 40 mynas were caught and divided into two groups, i.e., 7 and 25 °C for 14 days. To determine the effect of cold stress, cell mediated and humoral immune responses were assessed. The macrophage engulfment percentage was significantly (P < 0.05) higher at 25 °C rather than 7 °C either co-incubated with opsonized or unopsonized sheep red blood cells (SRBC). Macrophage engulfment/cell and nitric oxide production behaved in a similar manner. However, splenic cells plaque formation, heterophil to lymphocyte (H/L) ratio, and serum IgM or IgG production remained non-significant. There was a significant increase of IgG antibody production after a second immunization by SRBC. To the best of our knowledge, these findings have never been reported in the progression of this bird's invasion in frosty areas of the world. The results revealed a strengthened humoral immune response of myna and made this bird suitable for invasion in the areas of harsh conditions.

Seroprevalence and risk factors for toxoplasmosis in large ruminants in northern Punjab, Pakistan.

INTRODUCTION: Toxoplasmosis is a widespread zoonosis that causes significant economic losses due to abortions and other developmental disabilities in livestock animals. The objectives of the current study were to determine the prevalence and associated risk factors of Toxoplasma gondii infection in cattle and buffaloes in northern parts of Punjab, Pakistan, where no such work previously existed. METHODOLOGY: Serum samples obtained from 400 cattle and 422 buffaloes present on different private and government-owned farms were tested for anti-Toxoplasma IgG and anti-Toxoplasma IgM antibodies by using enzyme linked immunosorbent assay (ELISA). Additional data, including water sources, hygienic status at the farm, management practices, size of the herd, and presence of cats in the vicinity of the farm were obtained using a questionnaire in surveys and interviews. RESULTS: The overall prevalence of infection was 19.75% (79/400) in cattle and 15.16% (64/422) in buffaloes. IgG antibodies were found in 75 (18.75%) cattle and 58 (13.74%) buffaloes, while IgM antibodies were found in 9 (2.25%) cattle and 10 (2.37%) buffaloes. Seroprevalence was significantly higher in females and older animals of both species. Seroprevalence was found to be associated with poor hygienic conditions, extensive management practices, and presence of cats in the surrounding areas. No difference of seroprevalence was observed with respect to different breeds, location, water source, and herd size. CONCLUSION: The present study found that T. gondii is prevalent in large ruminants in northern Punjab, and may have important implications for the livestock industry and public health.

Impact of climate change and man-made irrigation systems on the transmission risk, long-term trend and seasonality of human and animal fascioliasis in Pakistan.

Large areas of the province of Punjab, Pakistan are endemic for fascioliasis, resulting in high economic losses due to livestock infection but also affecting humans directly. The prevalence in livestock varies pronouncedly in space and time (1-70%). Climatic factors influencing fascioliasis presence and potential spread were analysed based on data from five meteorological stations during 1990-2010. Variables such as wet days (Mt), water-budget-based system (Wb-bs) indices and the normalized difference vegetation index (NDVI), were obtained and correlated with geographical distribution, seasonality patterns and the two-decade evolution of fascioliasis in livestock throughout the province. The combined approach by these three indices proved to furnish a useful tool to analyse the complex epidemiology that includes (i) sheep-goats and cattlebuffaloes presenting different immunological responses to fasciolids; (ii) overlap of Fasciola hepatica and F. gigantica; (iii) co-existence of highlands and lowlands in the area studied; and (iv) disease transmission following bi-seasonality with one peak related to natural rainfall and another peak related to man-made irrigation. Results suggest a human infection situation of concern and illustrate how climate and anthropogenic environment modifications influence both geographical distribution and seasonality of fascioliasis risks. Increased fascioliasis risk throughout the Punjab plain and its decrease in the northern highlands of the province became evident during the study period. The high risk in the lowlands is worrying given that Punjab province largely consists of low-altitude, highly irrigated plains. The importance of livestock in this province makes it essential to prioritise adequate control measures. An annual treatment scheme to control the disease is recommended to be applied throughout the whole province.

Isolation of antibacterial compounds from Quercus dilatata L. through bioassay guided fractionation.

BACKGROUND: Four medicinal plants (Chrozophora hierosolymitana Spreng, Chrysanthemum leucanthemum L., Ephedra gerardiana Wall. ex Stapf, and Quercus dilatata L.) used by indigenous healers to treat various infectious diseases were selected for the present study. The major objective of the present study was isolation and characterization of antimicrobial components from the crude plant extracts using bioassay guided fractionation. METHODS: Seven methanolic extracts of the four plants were screened to identify any antimicrobial agents present in them. The active crude plant extract was fractionated first by solvent partitioning and then by HPLC. Characterization of the active fractions was done by using spectrophotometer. RESULTS: All the seven methanolic extracts showed low antifungal activity, however, when these extracts were tested for antibacterial activity, significant activity was exhibited by two extracts. The extract of aerial parts of Q. dilatata was most active and therefore, was selected for further analysis. Initially fractionation was done by solvent-solvent partitioning and out of six partitioned fractions, ethanol fraction was selected on the basis of results of antibacterial activity and phytochemical analysis. Further, fractionation was carried out by RP-HPLC and purified active subfractions were characterized by comparing their absorption spectra with that of the known natural products isolated from the plants of Quercus genus. DISCUSSION AND CONCLUSION: The results suggest that this is the first report of the isolated antibacterial compounds from this genus.

Biological monitoring of blood naphthalene levels as a marker of occupational exposure to PAHs among auto-mechanics and spray painters in Rawalpindi.

BACKGROUND: Routine exposure to chemical contaminants in workplace is a cause for concern over potential health risks to workers. In Pakistan, reports on occupational exposure and related health risks are almost non-existent, which reflects the scarce availability of survey data and criteria for determining whether an unsafe exposure has occurred. The current study was designed to evaluate blood naphthalene (NAPH) levels as an indicator of exposure to polycyclic aromatic hydrocarbons (PAHs) among automobile workshop mechanics (MCs) and car-spray painters (PNs). We further determined the relationship between blood NAPH levels and personal behavioural, job related parameters and various environmental factors that may further be associated with elevated risks of occupational exposures to PAHs. METHODS: Sixty blood samples (n = 20 for each group i.e. MC, PN and control group) were collected to compare their blood NAPH levels among exposed (MCs and PNs) and un-exposed (control) groups. Samples were analyzed using high pressure liquid chromatography (HPLC). Data regarding demographic aspects of the subjects and their socioeconomic features were collected using a questionnaire. Subjects were also asked to report environmental hygiene conditions of their occupational environment. RESULTS: We identified automobile work areas as potential sites for PAHs exposure, which was reflected by higher blood NAPH levels among MCs. Blood NAPH levels ranged from 53.7 to 1980.6 µgL(-1) and 54.1 to 892.9 µgL(-1) among MCs and PNs respectively. Comparison within each group showed that smoking enhanced exposure risks several fold and both active and passive smoking were among personal parameters that were significantly correlated with log-transformed blood NAPH levels. For exposed groups, work hours and work experience were job related parameters that showed strong associations with the increase in blood NAPH levels. Poor workplace hygiene and ventilation were recognized as most significant predictors related to differences among workplaces that may enhance the extent of exposure to chemical contaminants. CONCLUSIONS: It appeared that chemical exposure at the workplace may be influenced by multiple environmental factors, but poor workplace hygiene and duration of exposure (long work hours) were the most important factors. Smoking and negligence of workers regarding self protection were among some of the important personal behaviours than can be addressed with better training. There is also a need to improve workplaces hygiene and to rationalize work hours to minimize health risks. Since smoking was an important confounding factor that supplemented most of the actual occupational exposure, a study based on non-smoker subjects is needed to separate out the effects of smoking and other confounding factors that may obscure measurements of actual extent of occupational exposure.

Di- and triorganotin(IV) esters of 3,4-methylenedioxyphenylpropenoic acid: synthesis, spectroscopic characterization and biological screening for antimicrobial, cytotoxic and antitumor activities.

Nine biologically significant organotin(IV) esters of 3,4-Methylenedioxyphenylpropenoic acid (L) were synthesized with the general formulae [R2SnL2], where R includes Me(1), Et(3), But(4), Oct(5), Ph(8), and [R3SnL], in which R is Me(2), Cy(6), Ph(7), and But(9). The acid and its compounds were characterized by basic analytical techniques comprising elemental analysis, FTIR and mass spectrometry in solid state and by Multinuclear (1H, 13C and 119Sn) NMR in solution form, which provides some important information about the different coordination behaviors of metal in both solid and solution. Methylenedioxy moiety in these compounds enhances the biological activity of these compounds. These compounds were screened for a range of biological activities. Antibacterial activities were determined against six pathogenic bacterial strains, three gram-positive and three gram-negative, the activities were measured in terms of inhibition zones (mm). Results demonstrate that diorganotin derivatives are more active than triorganotin derivatives and ligand acid. Antifungal activity was determined against six pathogenic fungal strains, cytotoxicity by the brine shrimp lethality assay, and antitumor activity by crown gall tumor inhibition (potato disc) assay. Results for antifungal activity, cytotoxicity, and antitumor activity of these compounds demonstrate that triorganotin derivatives are more active than diorganotin derivatives and ligand. Finally, the results were compared with similar reports in the literature.