Antimicrobial Activity of Different Antimicrobial Peptides (AMPs) Against Clinical Methicillin-resistant Staphylococcus aureus (MRSA).

BACKGROUND: Antimicrobial research is being focused to look for more effective therapeutics against antibiotic-resistant infections caused by methicillin-resistant Staphylococcus aureus (MRSA). In this direction, antimicrobial peptides (AMP) appear as promising tool. OBJECTIVES: This study evaluated the antimicrobial activity of different AMPs (Citropin 1.1, Temporin A, Pexiganan, CA(1-7)M(2-9)NH2, Pal-KGK-NH2, Pal-KKKK-NH2, LL-37) against human MRSA clinical isolates. METHODS: The Minimum Inhibitory Concentration (MIC) was assessed for each AMP; then, the most active ones (Citropin 1.1, Temporin A, CA(1-7)M(2-9)NH2 and Pal-KGK-NH2) were tested against selected MRSA strains by time-kill studies. RESULTS: The lowest MIC value was observed for Pal-KGK-NH2 (1 µg/ml), followed by Temporin A (4- 16 µg/ml), CA(1-7)M(2-9)NH2 (8-16 µg/ml) and Citropin 1.1 (16-64 µg/ml), while higher MICs were evidenced for LL-37, Pexiganan and Pal-KKKK-NH2 (> 128 µg/ml). In time-kill experiments, Citropin 1.1 and CA(1-7)M(2-9)NH2 showed a relatively high percentage of growth inhibition (>30 %) for all the tested MRSA clinical isolates, with a dose-dependent activity resulting in the highest percentage of bacterial growth inhibition (89.39%) at 2MIC concentration. CONCLUSION: Overall, our data demonstrated the potential of some AMPs against MRSA isolates, such as Citropin 1.1 and CA(1-7)M(2-9)NH2, that represents a promising area of development for different clinical applications.

In vitro activity of the protegrin IB-367 alone and in combination compared with conventional antifungal agents against dermatophytes.

The occurrence of resistance or side effects in patients receiving antifungal agents leads to failure in the treatment of mycosis. The aim of this experimental study was to investigate the in vitro effects of IB-367 alone and in combination with three standard antifungal drugs, fluconazole (FLU), itraconazole (ITRA) and terbinafine (TERB), against 20 clinical isolates of dermatophytes belonging to three species. Minimum inhibitory concentrations (MICs), minimal fungicidal concentrations (MFCs), synergy test, time-kill curves, fungal biomass (FB) and hyphal damage using 2,3-bis-(2-methoxy-4-nitro-5-sulfenylamino carbonil)-2H-tetrazolium hydroxide assay (XTT) were performed to study the efficacy of IB-367. In this study, we observed that TERB and ITRA had MICs lower values for all the strains compared to IB-367 and FLU. Synergy was found in 35%, 30% and 25% of IB-367/FLU, IB-367/ITRA and IB-367/TERB interactions respectively. IB-367 exerted a fungicidal activity against Trichophyton mentagrophytes, T. rubrum and Microsporum canis at concentrations starting from 1x MIC. At a concentration of 5x MIC, IB-367 showed the highest rates of hyphae damage for M. canis 53% and T. mentagrophytes 50%; against the same isolates it caused a reduction of 1 log of the total viable count cell hyphae damage. We propose IB-367 as a promising candidate for the future design of antifungal drugs.

Intestinal parasitosis: data analysis 2006-2011 in a teaching hospital of Ancona, Italy.

Intestinal parasites are a serious problem in developing countries, but should not be underestimated in industrialised countries either. Between January 2006 and December 2011, stool specimens and the scotch tests of 5323 Italian and non Italian patients (adults and children) attending the laboratory of our Infectious Diseases Clinic in a teaching Hospital at Ancona were analyzed specifically for intestinal parasites. The present study shows that, over a six-year period, of a total of 5323 patients 305 harboured at least one species of parasite (5.7%). Among the pathogenic protozoa Giardia lamblia was the most common, the overall prevalence of giardiasis being 1.8 % (99/5323). Helminths were found in 0.9% of the patients (48/5323). In particular, Hymenolepis nana, Strongyloides stercoralis and Trichuris trichiura were most commonly recovered in non-Italian children, suggesting that certain intestinal parasites are restricted to endemic areas in the tropics. Eighteen of the 305 infected patients had more than one parasite in their stools. Our study demonstrates that intestinal parasites must be considered even in industrialised areas and stool examination should be supported by epidemiological data and clinical features.

An atypical, pigment-producing Metschnikowia strain from a leukaemia patient.

A yeast strain was isolated from the sputum sample of a leukaemia patient in the Spirito Santo Hospital of Pescara, Italy. The fungus produced a pigment that formed a reddish halo around colonies, and was identified and deposited as a Metschnikowia spp. (accession number IHEM 25107-GenBank accession number JQ921016) in the BCCM/IHEM collection of biomedical fungi and yeasts (Bruxelles, Belgium). Although the physiology of the strain was close to that of Metschnikowia sinensis, the D1/D2 sequence did not correspond to any previously described Metschnikowia species. Phylogeny of the genus Metschnikowia is complex and requires far more analysis. We present the first non-M. pulcherrima Metschnikowia spp. isolate recovered from a human, and emphasize the role of man as a transient carrier of environmental yeasts, the pathogenicity of which still needs to be defined.

In vitro susceptibility of dermatophytes to conventional and alternative antifungal agents.

The minimum inhibitory concentrations (MICs), the minimal fungicidal concentrations (MFCs), the fungal biomass (FB) and hyphal viability employing the dye 3-4,5 dimethyl- 2-thiazolyl- 2,5- diphenyl- 2H tetrazolium bromide (MTT) were used to compare the in vitro effects of fluconazole (FLU) with those of the N-terminal palmitoyl-lipidated peptide, Pal-Lys-Lys-NH(2) (PAL), and a tea tree oil component, gamma-Terpinene (TER), against several clinical isolates of Microsporum canis and Trichophyton rubrum. In general, FLU and PAL MICs were significantly lower than those observed with TER, while no differences in the three drugs were found in the MFCs. However, they were from two to 16-times higher than their respective MICs. FB of M. canis treated with either FLU or PAL, but not with TER, was significantly reduced over untreated controls. Only PAL and TER, in a medium-dependent fashion, but not FLU, reduced the FB of T. rubrum. Finally, PAL was found to be significantly more active than FLU at reducing the hyphal viability against both genera of dermatophytes. This study shows that PAL exerts an in vitro activity against dermatophytes at least similar to that observed with FLU and suggests that this compound might be a promising candidate in the treatment of infections due to dermatophytes.

In vitro activity of the synthetic lipopeptide PAL-Lys-Lys-NH(2) alone and in combination with antifungal agents against clinical isolates of Cryptococcus neoformans.

The in vitro activity of the lipopeptide PAL-Lys-Lys-NH(2) (PAL), alone or combined with either fluconazole (FLU) or amphotericin B (AMB), was investigated against 14 Cryptococcus neoformans isolates. PAL MICs ranged from 1.0 to 4.0 microg/ml. Fungicidal activity was observed. Synergy, defined as a fractional inhibitory concentration (FIC) index of < or =0.5, was observed in 21.4% of PAL/AMB interactions. Antagonism (FIC index>4) was never observed. The broad antifungal activity and the positive interactions with AMB suggest that PAL can represent a promising candidate in infections due to C. neoformans.

Effects of caspofungin against Candida guilliermondii and Candida parapsilosis.

The in vitro activity of caspofungin (CAS) was investigated against 28 yeast isolates belonging to Candida albicans (n = 5), Candida guilliermondii (n = 10), and Candida parapsilosis (n = 13). CAS MICs obtained by broth dilution and Etest methods clearly showed a rank order of susceptibility to the echinocandin compound with C. albicans > C. parapsilosis > C. guilliermondii. Similarly, time-kill assays performed on selected isolates showed that CAS was fungistatic against C. albicans and C. parapsilosis, while it did not exert any activity against C. guilliermondii. In a murine model of systemic candidiasis, CAS given at doses as low as 1 mg/kg of body weight/day was effective at reducing the kidney burden of mice infected with either C. albicans or C. guilliermondii isolates. Depending on the isolate tested, mice infected with C. parapsilosis responded to CAS given at 1 and/or 5 mg/kg/day. However, the overall CFU reduction for C. guilliermondii and C. parapsilosis was approximately 100-fold less than that for C. albicans. Our study shows that CAS was active in experimental systemic candidiasis due to C. guilliermondii and C. parapsilosis, but this activity required relatively high drug dosages.

Comparison of the fungicidal activities of caspofungin and amphotericin B against Candida glabrata.

We investigated the fungicidal activity of caspofungin (CAS) and amphotericin B (AMB) against 16 clinical isolates of Candida glabrata. The minimum fungicidal concentrations (MFCs) of CAS were similar to those of AMB, ranging from 2.0 to >8.0 microg/ml. Time-kill assays performed on selected isolates showed that AMB was fungicidal at concentrations four times the MIC while CAS was not. A neutropenic-mouse model of disseminated infection was utilized to determine the residual fungal kidney burden. While doses as low as 0.3 and 1 mg/kg of body weight/day of CAS and AMB, respectively, were effective at reducing the counts with respect to controls, organ sterilization was reached when both drugs were administered at 5 mg/kg/day. Our study reveals that, similar to AMB, CAS has the potential for a fungicidal effect in vivo against this difficult-to-treat fungal pathogen.

Caspofungin in combination with amphotericin B against Candida glabrata.

The effects of caspofungin combined with amphotericin B were investigated with Candida glabrata. Although in vitro experiments showed an indifferent interaction, the combination regimen was the only therapeutic approach yielding organ sterilization in a murine candidemia model.

Sequential therapy with caspofungin and fluconazole for Candida albicans infection.

A sequential therapy of caspofungin (CAS) and fluconazole (FLC) administration for treatment of Candida albicans infection was investigated. Treatment with CAS followed by FLC was as effective as CAS treatment given alone for the same duration. Our data suggest that switching from CAS to FLC is a potentially explorable therapeutic option for treatment of systemic candidiasis.

Antifungal susceptibility patterns of yeast isolates causing bloodstream infections.

A broth microdilution method following the NCCLS recommendations was used for testing fluconazole, itraconazole, posaconazole, 5-fluorocytosine and amphotericin B against 83 yeast isolates causing bloodstream infections in 59 patients hospitalized between January 1998 and June 2001 at the University Hospital of Bari, Italy. Isolates belonged to four species of Candida and three other yeast genera. Of the isolates, 97%, 95% and 100% were susceptible to fluconazole, itraconazole and posaconazole, respectively. Similarly, 97% and 100% of the isolates were susceptible to 5-fluorocytosine and amphotericin B, respectively.

In vitro and in vivo anticryptococcal activities of a new pyrazolo-isothiazole derivative.

We investigated the activity of a pyrazolo-isothiazole derivative (G8) against Cryptococcus neoformans. A first screening test showed that G8 at 10 mg/L inhibited the growth of 14 of 15 clinical isolates tested. Killing experiments showed that fungicidal activity was achieved after 8 h of treatment with G8 at concentrations > or =10 mg/L. In a murine model of systemic cryptococcosis, G8 was effective at prolonging survival compared with the controls. Our data indicate that this new derivative has a potential therapeutic role in infections caused by C. neoformans.

Point prevalence, microbiology and fluconazole susceptibility patterns of yeast isolates colonizing the oral cavities of HIV-infected patients in the era of highly active antiretroviral therapy.

We conducted a prospective study to address the prevalence and microbiological characteristics of yeast isolates colonizing the oral cavities of HIV-infected patients undergoing highly active antiretroviral therapy. Sixty-eight patients (67%) from a total of 102 were found to be colonized with yeasts. Sixty-five patients carried a single species (60 Candida albicans, three Candida glabrata and two Candida krusei) and three patients had mixed colonization of C. albicans and C. krusei. The status of yeast carrier was not associated with the number of CD4 cells or the viral load. Similarly, the type of antiretroviral regimen was not associated with the carriage of Candida spp. The only predictor of Candida colonization was a previous history of oropharyngeal candidiasis (P = 0.009). Although many patients in this series had already been treated with repeated courses of fluconazole therapy for previous episodes of oropharyngeal candidiasis, fluconazole susceptibility patterns showed that 93% of yeasts were susceptible to this triazole in vitro (MIC < or = 8.0 mg/L).