The remodeling of active sites to generate novel biocatalysts is an attractive and challenging task. We developed a stepwise loop insertion strategy (StLois), in which randomized residue pairs are inserted into active site loops. The phosphotriesterase-like lactonase from Geobacillus kaustophilus (GkaP-PLL) was used to investigate StLois's potential for changing enzyme function. By inserting six residues into active site loop 7, the best variant ML7-B6 demonstrated a 16-fold further increase in catalytic efficiency toward ethyl-paraoxon compared with its initial template, that is a 609-fold higher, >10 fold substrate specificity shift relative to that of wild-type lactonase. The remodeled variants displayed 760-fold greater organophosphate hydrolysis activity toward the organophosphates parathion, diazinon, and chlorpyrifos. Structure and docking computations support the source of notably inverted enzyme specificity. Considering the fundamental importance of active site loops, the strategy has potential for the rapid generation of novel enzyme functions by loop remodeling.

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Engineered bacteria with synthetic gene circuits are attractive tools to detect environmental contaminants. However, their applications in realistic settings are hindered by its relatively low sensitivity, long response time, and limited portability. Here, we present a synthetic bacterial consortium-based system for detecting organophosphorus pesticides (OPs). The system consists of two Escherichia coli strains with divided tasks, including one for hydrolyzing OPs to p-nitrophenol (PNP) and the other for converting the PNP signal into β-galactosidase production for colorimetric detection. Upon optimization, the system was able to detect ethyl-paraoxon at the concentration of 1 × 10 M within 3.5 h of induction at 28 °C, which is approximately 200-fold more sensitive than single-cell based whole-cell sensing. In addition, it was capable of detecting several OPs, commonly used in agriculture. Furthermore, the system showed promise for on-site detection through the demonstration of a paper-based setting and real apple and soil samples. This study provides a rapid, sensitive, and portable biosensing platform for contaminant detection and also demonstrates the utility of engineered microbial ecosystems for novel environmental applications.

Among hospitalized patients, the most common nosocomial infection is Urinary tract infection (UTI). The knowledge about the type of pathogens responsible for UTI and susceptibility and resistance pattern of the causative agents at a specific area may help the doctors to choose correct treatment regimen. This study was aimed to investigate the antibiotic susceptibility and resistance pattern of isolated urinary pathogens. This study was done at Anwer Khan Modern Medical College Hospital, Dhaka during January- June, 2011. Out of 498 clinical samples of urine collected, 245 (49.19%) showed significant bacterial growth. The most common pathogens isolated were Escherichia coli (142, 58.0%), Streptococcus feacalis (38, 15.5%), Pseudomonus (20, 8.2%), Klebsiella species (20, 8.2%) and Staphylococcus epidermidis (14, 5.7%). Members of the Enterobacteriaceae were 75%-100%sensitive to Amikacin and Nitrofurantoin while they were found variably sensitive to other commonly used antibiotics. Pseudomonas species were found 90% sensitive to Meropenem and 70% to Amikacin. Strep. feacalis were found 94.7% sensitive to Amoxicillin, 84.2% to Amoxiclave and 78.9% to Ciprofloxacin, 65.5% to Cephalexin, 50% to Ceftriaxone. The clinicians should use Meropenem and Amikacin selectively in cases of un-responsiveness to commonly used antibiotics. Anwer Khan Modern Medical College Journal Vol. 4, No. 2: July 2013, Pages 23-27 DOI: http://dx.doi.org/10.3329/akmmcj.v4i2.16938

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