In this study, culture-based methods and quantitative PCR (qPCR) assays were compared with each other for the measurement of Escherichia coli and Enterococcus spp. in water samples collected from rainwater tanks in Southeast Queensland, Australia. Among the 50 rainwater tank samples tested, 26 (52%) and 46 (92%) samples yielded E. coli numbers as measured by EPA Method 1603 and E. coli 23S rRNA gene qPCR assay, respectively. Similarly, 49 (98%) and 47 (94%) samples yielded Enterococcus spp. numbers as measured by EPA Method 1600 and Enterococcus spp. 23S rRNA gene qPCR assay, respectively. The mean E. coli (2.49 ± 0.85) log(10) and Enterococcus spp. (2.72 ± 0.32) log(10) numbers as measured by qPCR assays were significantly (P < 0001) different than E. coli (0.91 ± 0.80) log(10) and Enterococcus spp. (1.86 ± 0.60) log(10) numbers as measured by culture-based method. Weak but significant correlations were observed between both EPA Method 1603 and the E. coli qPCR assay (r = 0.47, P = 0.0009), and EPA Method 1600 and the Enterococcus spp. qPCR assay (r = 0.42, P = 0.002). Good qualitative agreement was found between the culture-based method and the Enterococcus spp. qPCR assay in terms of detecting fecal pollution in water samples from the studied rainwater tanks. More research studies, however, are needed to shed some light on the discrepancies associated with the culture-based methods and qPCR assays for measuring fecal indicator bacteria.

Aims: In this study, faecal indicator bacteria (FIB) namely Escherichia coli and Enterococcus spp. were seeded into slurries of possum faeces and placed on the roof and in the gutter of a roof-captured rainwater (RCR) system. The persistence of FIB in these circumstances was determined under ambient climatic conditions. FIB persistence was also determined under in situ conditions in tank water using diffusion chambers. Methods and Results: The numbers of surviving FIB at different time intervals were enumerated using culture-based methods. Both FIB were rapidly inactivated on the roof under sunlight conditions (T 90 = 2 h) compared with shade conditions (T 90 = 9-53 h). Significant differences were observed between sunlight and shade conditions on the roof for both T 90 values of E. coli (P < 0Á001) and Enterococcus spp. (P < 0Á001). E. coli showed biphasic inactivation patterns under both clean and unclean gutter conditions. Enterococcus spp., however, showed rapid inactivation (T 90 = 2 h for the clean gutter and T 90 = 6 h for the unclean gutter) compared with E. coli (T 90 = 22 h for the clean gutter and T 90 = 20 h for the unclean gutter). Significant differences were also observed between the T 90 values of E. coli and Enterococcus spp. for both clean (P < 0Á001) and unclean (P < 0Á001) gutters. Both E. coli and Enterococcus spp. showed nonlinear biphasic inactivation in tank water. Significant difference was observed between the T 90 value of E. coli inactivation compared with Enterococcus spp. (P < 0Á001) in the tank water. Conclusions: In this study, FIB were observed to survive longer (T 90 = 9-53 h) on the roof under shade conditions compared with sunlight conditions (T 90 = 2 h). If there is a rainfall event within two to three days after the deposition of faecal maters on the roof, it is highly likely that FIB would be transported to the tank water. When introduced into the tank, a relatively slow inactivation process may take place (T 90 = 38-72 h). Significance and Impact of the Study: The presence of FIB in water indicates faecal pollution and potential presence of enteric pathogens. Therefore, the information on the resilience of FIB, as obtained in this study, can be used for indirect assessment of health risks associated with using roof-captured rainwater for potable and nonpotable purposes.

Four 1-N-aminohydroxy-alkyl derivatives of kanamycin A were prepared and their in vitro activities against aminoglycoside-sensitive and aminoglycoside-resistant organisms were compared with amikacin.1-N-[(S)-4-Amino-2-hydroxybutyl] kanamycin A (Fig. 1, compound 2, code no. UK-18,892) was equipotent to amikacin in all these tests and in mouse protection studies.A, has been reported1) to be highly active against both kanamycin-sensitive and kanamycin-resistant bacteria.In contrast, several other 1-N-acyl derivatives of kanamycin A have been shown to possess low antibacterial activity.21 We undertook a program of 1-N-alkyl modifications of kanamycin A to investigate structure-activity relationships in this area, and this paper describes the synthesis and antibacterial properties of some of the compounds produced. Materials and MethodsKanamycin A-1-N-Alkylated Derivatives 1-N-(S)-w-Amino-2-hydroxyalkyl derivatives of kanamycin A were prepared by diborane reduction of the corresponding 1-N-acyl derivatives, using the trifluoroacetate salts to improve solubility. The 1-N-acyl derivative (150 mg) was converted to its trifluoroacetate salt by dissolving in anhydrous trifluoroacetic acid followed by evaporation under reduced pressure. The resulting salt was dissolved in dry tetrahydrofuran (5 ml) and treated portionwise with a 1 M solution of diborane in tetrahydrofuran (20 ml), under a nitrogen atmosphere. After 6 hours at 50°C the reaction mixture was cooled and excess diborane was destroyed by the addition of a small amount of water. After evaporation under reduced pressure, the residue was taken up in water (10 ml), adjusted to pH 12 with 0.1 N sodium hydroxide solution and then to pH 5 using 2 N hydrochloric acid. The resulting solution was chromatographed on a column of Amberlite CG-50 ion-exchange resin (50 ml), in the ammonium ion form, eluting with dilute ammonia solution (linear increase, 0.1 to 1 M). Fractions containing the product were combined and evaporated under reduced pressure to yield the product as an amorphous solid (Table 1). The requisite 1-N-acyl compounds were synthesized by the method of NAITO et al.2,The structures of the products prepared in the present study have been confirmed by 13C-n.m.r. and/or mass spectral data. Microanalyses of the products reported are in agreement with the expected formulae calculated as the carbonate salts.Micro-organisms The bacteria designated 'aminoglycoside-sensitive' were pathogenic strains well adapted to growth under laboratory conditions. The aminoglycoside-resistant organisms were bacteria with proven inactivating mechanisms and were obtained from Prof. J. DAVIES, University of Wisconsin.M.I.C. Determinations Minimum inhibitory concentrations (M.I.C.) of the compounds were determined by a standard dilution method in Diagnostic Sensitivity Test Agar (DSTA-Oxoid). Serial dilutions (in 2-fold steps) of the compounds were prepared in the agar plates over the required concentration range and, after the