Heterologous expression of cry1Ia12 insecticidal gene in cotton encodes resistance against pink bollworm, Pectinophora gossypiella (Lepidoptera: Gelechiidae); an alternate insecticidal gene for insect pest management.

BACKGROUND: Cotton is continuously exposed to sucking and chewing insect pest pressure since emergence to harvesting. Pink bollworm (Pectinophora gossypiella) has become major chewing insect pest to reduce the cotton yield and results in bad lint quality even in transgenic crops. The efficiency of insecticidal genes has been compromised due to extensive utilization of transgenic crops. METHODS AND RESULTS: The present study was conducted to evaluate the efficacy of an alternate cry1Ia12 insecticidal gene against pink bollworm (PBW) in cotton. Agrobacterium tumefaciens strain LBA4404 harboring pCAMBIA2300 expression vector containing cry1Ia12 gene under the control of 35S CaMV was used to transform a local cotton cultivar GS-01. The various molecular analyses revealed the transgene integration and expression in primary transformants. Among five selected transgenic plants, tcL-08 showed maximum (16.06-fold) mRNA expression of cry1Ia12 gene whereas tcL-03 showed minimum (2.33-fold) expression. Feeding bioassays of 2nd and 3rd instar pink bollworm (PBW) larvae on immature cotton bolls, flowers and cotton squares revealed up to 33.33% mortality on tcL-08 while lowest mortality (13.33%) was observed in tcL-03 and tcL-15. Furthermore, the average weight and size of survived larvae fed on transgenic plants was significantly lesser than the average weight of larvae survived on non-transgenic plants. CONCLUSIONS: The present study suggests the cry1Ia12 gene as an alternate insecticidal gene for the resistance management of cotton bollworms, especially PBW.

Early detection of MDR Mycobacterium tuberculosis mutations in Pakistan.

The result of improper treatment has led to the rise of Multidrug-resistant (MDR) strains. This concern still exists in Pakistan. In order to save energy, time and resources an early detection of resistant cases is imperative. Thus, a treated group of 100 isolates and a control group of 56 untreated isolates were studied. PCR and gene sequencing showed mutations at codon 531 and 513 in the rpoB gene. 12% of cases showed a double mutation in the rpoB gene. katG gene showed mutations at codon 315 and 299. 28.6% of the control group cases were positive for MDR whereas 100% of the treated group were positive for MDR. This study explores the significantly increasing ratio of MDR-TB among Pakistani population. This study provides prevalent MDR mutations among Pakistanis and suggests developing such molecular assays that are time and cost effective. Importance: Pakistan is a developing country and has fourth highest incidence rate of MDR-TB. The treatment of MDR-TB is the use of second line drugs that has severe side effects as well as it requires long time span. One of the strategies to control the spread of MDR-TB is to decipher the aberrations at molecular level in order to formulate potent drugs that can treat the patients within short span of time. Determining the mutation profile of MDR in Pakistani populations will open new horizons for the improvement of drug treatment regimens to make it more effective or for the development of novel potent drugs and vaccines to better treat the drug-resistant TB. Moreover, this study will be help in disease control program.

Novel approaches to circumvent the devastating effects of pests on sugarcane.

Sugarcane (Saccharum officinarum L.) is a cash crop grown commercially for its higher amounts of sucrose, stored within the mature internodes of the stem. Numerous studies have been done for the resistance development against biotic and abiotic stresses to save the sucrose yields. Quality and yield of sugarcane production is always threatened by the damages of cane borers and weeds. In current study two problems were better addressed through the genetic modification of sugarcane for provision of resistance against insects and weedicide via the expression of two modified cane borer resistant CEMB-Cry1Ac (1.8 kb), CEMB-Cry2A (1.9 kb) and one glyphosate tolerant CEMB-GTGene (1.4 kb) genes, driven by maize Ubiquitin Promoter and nos terminator. Insect Bio-toxicity assays were carried out for the assessment of Cry proteins through mortality percent of shoot borer Chilo infuscatellus at 2nd instar larvae stage. During V0, V1 and V2 generations young leaves from the transgenic sugarcane plants were collected at plant age of 20, 40, 60, 80 days and fed to the Chilo infuscatellus larvae. Up to 100% mortality of Chilo infuscatellus from 80 days old transgenic plants of V2 generation indicated that these transgenic plants were highly resistant against shoot borer and the gene expression level is sufficient to provide complete resistance against target pests. Glyphosate spray assay was carried out for complete removal of weeds. In V1-generation, 70-76% transgenic sugarcane plants were found tolerant against glyphosate spray (3000 mL/ha) under field conditions. While in V2-generation, the replicates of five selected lines 4L/2, 5L/5, 6L/5, L8/4, and L9/6 were found 100% tolerant against 3000 mL/ha glyphosate spray. It is evident from current study that CEMB-GTGene, CEMB-Cry1Ac and CEMB-Cry2A genes expression in sugarcane variety CPF-246 showed an efficient resistance against cane borers (Chilo infuscatellus) and was also highly tolerant against glyphosate spray. The selected transgenic sugarcane lines showed sustainable resistance against cane borer and glyphosate spray can be further exploited at farmer's field level after fulfilling the biosafety requirements to boost the sugarcane production in the country.