Search

The mosquitoes of the Anopheles and Aedes genus are some of the most deadly insects to humans because of their effectiveness as vectors of malaria and a range of arboviruses, including yellow fever, dengue, chikungunya, West Nile and Zika. The use of insecticides from different chemical classes is a key component of the integrated strategy against An. gambiae and Ae. aegypti, but the problem of insecticide resistance means that new compounds with different modes of action are urgently needed to replace chemicals that fail to control resistant mosquito populations. We have previously shown that feeding inhibitors of peptidyl dipeptidase A to both An. gambiae and Ae. aegypti mosquito larvae lead to stunted growth and mortality. However, these compounds were designed to inhibit the mammalian form of the enzyme (angiotensin-converting enzyme, ACE) and hence can have lower potency and lack selectivity as inhibitors of the insect peptidase. Thus, for the development of inhibitors of practical value in killing mosquito larvae, it is important to design new compounds that are both potent and highly selective. Here, we report the first structures of AnoACE2 from An. gambiae in its native form and with a bound human ACE inhibitor fosinoprilat. A comparison of these structures with human ACE (sACE) and an insect ACE homologue from Drosophila melanogaster (AnCE) revealed that the AnoACE2 structure is more similar to AnCE. In addition, important elements that differ in these structures provide information that could potentially be utilised in the design of chemical leads for selective mosquitocide development., (© 2019 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

A newly modified, semiautomated instrument (Bacteriuria detection device (BDD] designed to detect the presence of bacteriuria in less than 3 min was compared to quantitative urine culture plating techniques. The instrument consists of a self-enclosed vacuum-filtration-staining system in which a 1-ml urine filtrate is stained on a filter. The resulting color determines the quantitation. Of the total of 525 clinical urine specimens tested, 66 (12.6%) were uninterpretable due to pigment deposition or inability to complete the filtering process (clogging of the filter). Of the remaining 459 specimens, 93 (20.3%) had a plate quantitation colony count of 10(5) colony-forming units (CFU)/ml or more. The BDD detected 94.2% of these positive specimens if only significant pathogens were included (85% if specimens that were probably contaminated were also included). For specimens containing significant pathogens at 10(4)-10(5) CFU/ml, the BDD detection rate was 41%. The device detected most (94.3%) gram-negative bacilli and enterococci at colony counts of 10(5) CFU/ml or more. In addition, the BDD accurately detected 95.6% of specimens with no growth or fewer than 10(4) CFU/ml. With several proposed modifications, these results suggest that this instrument is potentially useful as a urine screening device in a select population.

The major coat protein of the bacteriophage f1 is synthesized during infection of Escherichia coli and becomes tightly associated with the host membrane. This synthesis was studied in conjunction with the strain BB26-36, a mutant defective in phospholipid synthesis, to investigate basic questions concerning membrane protein and phospholipid synthesis. Coat protein synthesis is decreased in the absence of net phospholipid synthesis. The coat protein produced under these conditions is still found tightly associated with the membrane. Resumption of phospholipid synthesis leads to an increase in the synthesis and accumulation of the coat protein. Therefore, a correlation between coat protein and phospholipid synthesis seems to exist. However, the packaging of phage deoxyribonucleic acid into phage particles proceeds in the absence of phospholipid synthesis, and the number of phage particles produced appears to depend only on the amount of coat protein in the membrane.

Sets of three Isolator blood culture tubes were seeded with low numbers of 96 strains of 26 bacterial species (fresh and stock clinical isolates). One tube was processed immediately, and the other two were held at 22 and 34 degrees C for 15 h before processing. Organism recovery was 99, 99, and 98%, respectively. Organism numbers increased at both 22 degrees C (60% of strains) and 34 degrees C (79% of strains). Especially notable was that the increases were seen with most strains of Staphylococcus aureus, streptococcal species, Pseudomonas, and all of the Enterobacteriaceae tested. Seven strains, including Streptococcus pneumoniae and Haemophilus influenzae, although viable, were recovered with a decreased number of organisms at each temperature. Acridine orange staining detected organisms in 53% of those Isolator tubes being held and 71% of those demonstrating a numerical increase, after incubation at 34 degrees C. In addition, it was noted that after processing, 48% of the strains that had increased in number while being held at 34 degrees C resulted in visible growth on agar media in 6 h. The results suggest that up to a 15-h delay in processing Isolator tubes may be possible and that acridine orange staining for the rapid detection of positive cultures may be useful in such a circumstance.

[3H]Uracil-pulse-labeled RNA from Escherichia coli infected with f1 bacteriophage was fractionated on polyacrylamide gels containing urea. Eight phage-specific RNA species were present with approximate lengths ranging from 2100 to 400 nucleotides. The amount of the seven largest species was increased when the infected bacteria were incubated at 41 degrees C. When the RNA was isolated and used as message in an in vitro protein-synthesizing system, most of the RNA species appeared to direct the synthesis of the phage gene VIII protein. The six largest species also directed the synthesis of the phage gene V protein. Some of the labeled smaller RNA species increased in amount after addition to rifampicin, suggesting that they may have resulted from cleavage of larger RNA species. These particular smaller RNA species also were present in infected bacteria containing a mutant RNase III. The data are discussed in terms of the regulation of synthesis of the phage-specific proteins.

We have analyzed eight major phage-specific mRNA species which are synthesized following infection of Escherichia coli with bacteriophage fl. The approximate half-lives of these RNAs appear to be inversely proportional to their lengths. Three species have the properties of primary transcripts. They are labeled very rapidly with [3H]uracil, and they co-migrate by sodium laruyl sulfate-urea polyacrylamide gel electrophoresis with the three major RNAs transcribed from replicative form DNA in vitro using [gamma-32P]GTP as the labeled ribonucleoside triphosphate. At least three of the remaining RNAs synthesized in vivo arise by some type of processing reaction. The processed species contain, as determined by oligonucleotide analysis, the information for the gene VIII protein and the 3'-terminal oligonucleotide expected for a transcription termination event at the rho-independent site following gene VIII. The two primary transcripts which could be analyzed also have this same 3'-terminal oligonucleotide. These results suggest that the processed mRNAs are the 3'-terminal cleavage products of primary transcripts which had terminated at the rho-independent site.

An assessment was made of the necessity of performing routine screening for Staphylococcus aureus in stool specimens. A total of 527 stool specimens were evaluated. Because of the rare incidence of staphylococcal enterocolitis and the high ($2.50) screening cost, the results of this evaluation suggest that such a screening need not be performed routinely.

Allopurinol therapy in man interferes with pyrimidine biosynthesis de novo by inhibition of one or both of the two enzymes, orotate phosphoribosyltransferase (OPRT) and orotidylic decarboxylase (ODC), responsible for the conversion of orotic acid to uridine-5'-monophosphate. Inhibition of this pathway in vivo is followed in 1-3 wk by an increase in the activity of both of these enzymes in erythrocytes and of ODC in circulating leukocytes. This drug-mediated increase in enzyme activity in erythrocytes could not be attributed to enzyme stabilization or induction in vivo but appeared to be due to enzyme "activation." "Activation" of the OPRT enzyme was directly demonstrated in erythrocytes studied in vitro after incubation with oxipurinol, and to a lesser extent, with allopurinol. No evidence for "activation" of the ODC enzyme was demonstrated in vitro. This response to allopurinol therapy provides an excellent model for examining the mechanism of increased enzyme activity in response to drug administration.