Analysis of xanthyletin and secondary metabolites from Pseudomonas stutzeri ST1302 and Klebsiella pneumoniae ST2501 against Pythium insidiosum.

BACKGROUND: Pythium insidiosum is a member of the oomycetes class of aquatic fungus-like microorganisms. It can infect humans and animals through skin wounds and the eyes, causing pythiosis, an infectious disease with high morbidity and mortality rates. Antifungal agents are ineffective as pythiosis treatments because ergosterol, the target site of most antifungal agents, is not found in the P. insidiosum cytoplasmic membrane. The best choice for treatment is surgical removal of the infected organ. While natural plant products or secretory substances from bacterial flora have exhibited in vitro anti-P. insidiosum activity, their mechanism of action remains unknown. Therefore, this study hypothesized that the mechanism of action could be related to changes in P. insidiosum biochemical composition (such as lipid, carbohydrate, protein or nucleic acid) following exposure to the inhibitory substances. The biochemical composition of P. insidiosum was investigated by Synchrotron radiation-based Fourier-transform infrared (FTIR) microspectroscopy. RESULTS: Fraction No.6 from the crude extract of P. stutzeri ST1302, fraction No.1 from the crude extract of K. pneumoniae ST2501 and xanthyletin were used as anti-P. insidiosum substances, with MFCs at 3.125, 1.57-1.91, 0.003 mg/ml, respectively. The synchrotron FTIR results show that the deconvoluted peak distributions in the amide I, amide II, and mixed regions were significantly different between the treatment and control groups. CONCLUSIONS: Xanthyletin and the secondary metabolites from P. stutzeri ST1302 and K. pneumoniae ST2501 exerted anti-P. insidiosum activity that clearly changed the proteins in P. insidiosum. Further study, including proteomics analysis and in vivo susceptibility testing, should be undertaken to develop a better understanding of the mechanism of anti-P. insidiosum activity.

Bactericidal efficacy of alcohol solution in community hospital and health centers.

OBJECTIVE: To evaluate bactericidal efficacy of alcohol solution during actual use and typical storage conditions in community hospital and health centers. MATERIAL AND METHOD: The alcohol samples were collected immediately after the first bottle-opening (day 0) and on day 3, 7, 14, 21 and 30 from 10 stations in hospital and community health centers in Pone-na-kaew district, Sakon Nakhon province, Thailand, during May-July 2011. Bactericidal efficacy of these samples against Staphylococcus epidermidis, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa and Klebsiella pneumoniae was evaluated. Ethanol concentration was quantified by a gas chromatography method. RESULTS: Bactericidal efficacy of the alcohol samples still remained on day 30 with ethanol concentration range of 60.91-65.99% v/v. CONCLUSION: This finding should be considered as a cost-benefit model for using alcohol solution in community hospital and health centers.

Evaluation for the clinical diagnosis of Pythium insidiosum using a single-tube nested PCR.

Pythiosis is a rare infectious disease caused by Pythium insidiosum, which typically occurs in tropical and subtropical regions. The high mortality rate may be in consequence of the lack of diagnosis. The objective of this study was to evaluate reliability of a new single-tube nested PCR for detection of P. insidiosum DNA. A total of 78 clinical isolates of various fungi and bacteria, 106 clinical specimens and 80 simulated positive blood samples were tested. The developed primer pairs CPL6-CPR8 and YTL1-YTR1 are located on 18S subunit of the rRNA gene of P. insidiosum. The specificity, negative and positive predictive values were 100, 100 and 87.5 %, respectively, as compared with direct microscopy and cultivation. The detection limit of the single-tube nested PCR was 21 zoospores corresponding to 2.7 pg of the DNA. The results demonstrate that the new single-tube nested PCR offers a highly sensitive, specific and rapid genetic method for detecting P. insidiosum.

Increased HIV-DNA load in CCR5-negative lymphocytes without viral phenotypic change.

We have previously described a selective increase in HIV-DNA content in CCR5-negative lymphocytes from late stage HIV-infected patients. Here, we show that this increase occurred even in the absence of viral phenotypic switching from CCR5- to CXCR4-tropic. This leads us to hypothesize that early and late CCR5-tropic viruses might be different in the ability to infect CCR5-low or -negative cells. We compared a set of early CCR5-tropic viruses with low viral DNA content in CCR5-negative cells to a set of late CCR5-tropic viruses with high viral DNA content in CCR5-negative cells. We could not find any significant differences between the two sets of viruses in the aspects of relative infectivity in CCR5-low cells and the level of inhibition by beta-chemokine. This suggested that there may be some changes in cellular phenotype or environment that allows an expansion of susceptible cell population in late stages HIV infection. Understanding these changes may provide a novel approach for HIV therapy.