A Combined Approach for the Characterization of Small Ruminant Lentivirus Strains Circulating in the Islands and Mainland of Greece.

Small ruminant lentiviruses are a group of viruses infecting goat and sheep worldwide. These viruses exhibit an extraordinary degree of genetic and antigenic variability that severely influence in vivo and in vitro features, as well as diagnostic test results. Small ruminant farming is the most important animal farming business in Greece, with a high impact on the Greek primary economy. Although SRLV infection and its impact on animal production are well established in the country, little is known about the circulating SRLV strains and their prevalence. The aim of this study was to characterize SRLVs circulating in Greece with a combined serological and molecular approach, using the bulk milk matrix collected from 60 farms in different municipalities. This study allowed us to estimate a seroprevalence of around 52% at the herd level. The B1, B2 and A3 subtypes and a novel A viral cluster were identified. Moreover, the amplicon sequencing method allowed us to identify more than one viral subtype in a sample. These results again confirm the high variability of these viruses and highlight the importance of the constant monitoring of viral evolution, in particular in antigens of diagnostic interest.

In vitro effects of different ozone preparations on microorganisms responsible for endometritis in the mare.

Infectious endometritis is considered one of the major causes of infertility and it can affect up to 60% of barren mares. It is characterized by the presence of one or more microorganisms in the reproductive tract and it is treated with the administration of antibiotics, ecbolic agents and uterine lavages. Ozone, thanks to its antimicrobial properties that are based on its high oxidative potential, could represent an effective alternative treatment for endometritis. The aim of this study was to test in vitro the bactericidal and fungicidal properties of different ozone formulations, either as gas (experiment 1) or dissolved in two liquid matrices (experiment 2), specifically distilled water or oil (Neozone 4000, Cosmoproject, Parma, Italy), onto 6 different species of microorganisms isolated from mares with clinical endometritis, namely Escherichia coli, Staphylococcus aureus, Streptococcus equi subsp. Zooepidemicus, Pseudomonas aeruginosa, Klebsiella pneumoniae and Candida albicans. In the first experiment, 3 clinical antibiotic-resistant strains per each species were exposed to different conditions: to O2O3 gas mixtures (15 and 40 µg/ml for 1, 3 and 5 min), to 100 % O2 or left untreated. The results showed a reduction of the microbial count of over 99,9% for every pathogen, time and concentration of O2O3 gas mixtures tested. Furthermore, gaseous ozone showed both a time-dependant effect (5 vs 3 vs 1 min of exposure) and a concentration-dependant effect (40 vs 15 µg/ml) at 1 and 3 min, while after 5 min no differences were observed. In the second experiment, minimum inhibitory concentration (MIC), and minimum bactericidal/fungicidal concentration (MBC, MFC) of ozonated distilled water and ozonated oil were evaluated. Ozonated oil showed a bactericidal/fungicidal activity against all the strains tested (MIC range 12.5-25 % v/v, MBC/MFC range 12.5-50 % v/v) while ozonated distilled water didn't show an observable antimicrobial effect, discouraging its use as an antimicrobial agent for the treatment of endometritis. The results of this in vitro study indicate that both gaseous ozone and ozonated oil exerted remarkable antimicrobial activities and are promising alternative treatments for infectious endometritis, even when caused by antibiotic-resistant bacteria, and encourage further experiments in an effort to scale down or even prevent the use of antibiotics in equine reproduction.

Serological and Molecular Characterization of Small Ruminant Lentiviruses in Morocco.

Recent studies that investigated the origins of SRLV strains offered new insights into their distribution among domestic ruminants. The aim of the study was to investigate SRLV circulation in Morocco. A total of 51 farms were selected in different geographical locations and tested by screening and genotyping ELISA. Whole blood was used for DNA extraction and nested gag PCR. The sample size allowed for an estimation of prevalence lower than 20% (CI 95%). Surprisingly, a large proportion of screening-positive samples were not correctly serotyped. Sanger and NGS amplicon sequencing approaches allowed us to obtain new sequences even from difficult-to-amplify samples. The serological data support the evidence of an intrinsic difficulty of SRLV to spread, likely due to management practices. The low rate of success by genotyping ELISA led us to suppose that divergent strains might have escaped from diagnostic tools, as partially confirmed by the evidence of an A subtype carrying a mismatch in serotyping epitope. The sequence analysis revealed the circulation of novel B and recombinant A/B subtypes. This study highlights the importance of monitoring viral sequences and their evolution to develop specific diagnostic tests, particularly in countries where control measures are in place.

Development of molecular and antigenic-based rapid tests for the identification of African swine fever virus in different tissues.

African swine fever (ASF) is a severe haemorrhagic infectious disease affecting suids, thus representing a great economic concern. Considering the importance of the early diagnosis, rapid point of care testing (POCT) for ASF is highly demanded. In this work, we developed two strategies for the rapid onsite diagnosis of ASF, based on Lateral Flow Immunoassay (LFIA) and Recombinase Polymerase Amplification (RPA) techniques. The LFIA was a sandwich-type immunoassay exploiting a monoclonal antibody directed towards the p30 protein of the virus (Mab). The Mab was anchored onto the LFIA membrane to capture the ASFV and was also labelled with gold nanoparticles for staining the antibody-p30 complex. However, the use of the same antibody for capturing and as detector ligand showed a significant competitive effect for antigen binding, so required an experimental design to minimize reciprocal interference and maximize the response. The RPA assay, employing primers to the capsid protein p72 gene and an exonuclease III probe, was performed at 39 °C. The limit of detection of the method was assessed using a plasmid encoding the target gene and resulted in 5 copy/µL. The new LFIA and RPA were applied for ASFV detection in the animal tissues usually analysed by conventional assays (i.e., real-time PCR), such as kidney, spleen, and lymph nodes. A simple and universal virus extraction protocol was applied for sample preparation, followed by DNA extraction and purification for the RPA. The LFIA only required the addition of 3% H2O2 to limit matrix interference and prevent false positive results. The two rapid methods (25 min and 15 min were needed to complete the analysis for RPA and LFIA, respectively) showed high diagnostic specificity (100%) and sensitivity (93% and 87% for LFIA and RPA, respectively) for samples with high viral load (Ct < 27). False negative results were observed for samples with low viral load (Ct > 28) and/or also containing specific antibodies to ASFV, which decreased antigen availability and were indicative of a chronic, poorly transmissible infection. The simple and rapid sample preparation and the diagnostic performance of the LFIA suggested its large practical applicability for POC diagnosis of ASF.

Resistance to Critical Important Antibacterials in Staphylococcus pseudintermedius Strains of Veterinary Origin.

Staphylococcal infections represent a challenge in companion animals and hospitalized patients. This study aimed to assess the resistance of Staphylococcus pseudintermedius isolates, against a broad panel of antibacterials, including exclusive to human medicine. A total of 40 S. pseudintermedius were collected from clinical specimens of dogs (n = 31) and cats (n = 5). All strains were tested for 20 antibacterials, namely 14 Critical Important and eight Highly Important Antibacterials (CIA and HIA, respectively), indicative for 18 antimicrobial classes. All strains were susceptible to seven antibiotics (daptomycin, fosfomycin, fusidic acid, linezolid, quinupristin-dalfopristin, teicoplanin/vancomycin, tigecycline). The highest resistance was against penicillin (97.5% Confidence Interval [CI]: 83.8-100.0), whereas the lowest against telavancin (2.5%, CI: 0.0-16.2). Resistance versus Highest Priority CIA was observed, namely against macrolides (70.0, CI: 52.1-84.3), quinolones (62.5, CI: 44.5-78.3), 5th generation cephalosporins (7.5, CI: 1.3-21.6), and glycopeptides (2.5%, CI: 0.0-14.2). Among High Priority CIA, strains were resistant only to aminoglycosides (65.0, CI: 47.0-80.4) and ansamycins (12.5, CI: 3.8-28.1). We observed the highest resistance against veterinary medicine antibacterials, but there was also resistance against antibacterials exclusive to human medicine, namely ceftaroline (7.5, CI: 1.0-23.8) and telavancin. S. pseudintermedius zoonotic potential and its rate of acquisition of new resistance should encourage surveillance on a broad spectrum of antibacterials.

Detailed epitope mapping of SARS-CoV-2 nucleoprotein reveals specific immunoresponse in cats and dogs housed with COVID-19 patients.

Since the initial emergence in December 2019, the novel Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has been reported in over 200 countries, representing an unprecedented challenge related to disease control worldwide. In this context, cases of human to animal transmission have been reported, raising concern about the potential role of companion animals in the pandemic and stressing the need for reliable animal testing. In the study, a detailed epitope mapping of SARS-CoV-2 nucleoprotein, using both human and pet sera, allowed the identification of the most antigenic region in the C-terminus domain of the protein, which was used to develop an experimental double antigen-based ELISA. A panel of pre-pandemic sera and sera of animals immunized against (or naturally infected with) related coronaviruses was used to assess assay specificity at 99.5%. Positive sera belonging to animals housed with COVID-19 patients were confirmed with the experimental double-antigen ELISA using Plaque Reduction Neutralization test (PRNT) test as gold standard. The availability of a serological assay that targets a highly specific viral antigen represents a valuable tool for multispecies monitoring of Coronavirus Disease 2019 (COVID-19) infection in susceptible animals.

The Inhibition of Non-albicans Candida Species and Uncommon Yeast Pathogens by Selected Essential Oils and Their Major Compounds.

The epidemiology of yeast infections and resistance to available antifungal drugs are rapidly increasing, and non-albicans Candida species and rare yeast species are increasingly emerging as major opportunistic pathogens. In order to identify new strategies to counter the threat of antimicrobial resistant microorganisms, essential oils (EOs) have become an important potential in the treatment of fungal infections. EOs and their bioactive pure compounds have been found to exhibit a wide range of remarkable biological activities. We investigated the in vitro antifungal activity of nine commercial EOs such as Thymus vulgaris (thyme red), Origanum vulgare (oregano), Lavandula vera (lavender), Pinus sylvestris (pine), Foeniculum vulgare (fennel), Melissa officinalis (lemon balm), Salvia officinalis (sage), Eugenia caryophyllata (clove) and Pelargonium asperum (geranium), and some of their main components (α-pinene, carvacrol, citronellal, eugenol, γ-terpinene, linalool, linalylacetate, terpinen-4-ol, thymol) against non-albicans Candida strains and uncommon yeasts. The EOs were analyzed by GC-MS, and their antifungal properties were evaluated by minimum inhibitory concentration and minimum fungicidal concentration parameters, in accordance with CLSI guidelines, with some modifications for EOs. Pine exhibited strong antifungal activity against the selected non-albicans Candida isolates and uncommon yeasts. In addition, lemon balm EOs and α-pinene exhibited strong antifungal activity against the selected non-albicans Candida yeasts. Thymol inhibited the growth of all uncommon yeasts. These data showed a promising potential application of EOs as natural adjuvant for management of infections by emerging non-albicans Candida species and uncommon pathogenic yeasts.

Antifungal Activity of Melaleuca alternifolia Essential Oil (TTO) and Its Synergy with Itraconazole or Ketoconazole against Trichophyton rubrum.

Over the past 20-30 years, Trichophyton rubrum represented the most widespread dermatophyte with a prevalence accounting for 70% of dermatophytosis. The treatment for cutaneous infections caused by Trichophyton spp. are imidazoles (ketoconazole (KTZ)) and triazoles (itraconazole (ITZ)). T. rubrum can develop resistance to azoles after prolonged exposure to subinhibitory concentrations resulting in therapeutic failures and chronic infections. These problems have stimulated the search for therapeutic alternatives, including essential oils, and their potential use in combination with conventional antifungals. The purpose of this study was to evaluate the antifungal activity of tea tree oil (TTO) (Melaleuca alternifolia essential oil) and the main components against T. rubrum and to assess whether TTO in association with KTZ/ITZ as reference drugs improves the antifungal activity of these drugs. We used a terpinen-4-ol chemotype (35.88%) TTO, and its antifungal properties were evaluated by minimum inhibitory and minimum fungicidal concentrations in accordance with the CLSI guidelines. The interaction between TTO and azoles was evaluated through the checkerboard and isobologram methods. The results demonstrated both the fungicide activity of TTO on T. rubrum and the synergism when it was used in combination with azoles. Therefore, this mixture may reduce the minimum effective dose of azole required and minimize the side effects of the therapy. Synergy activity offered a promise for combination topical treatment for superficial mycoses.

Enhanced Killing of Candida krusei by Polymorphonuclear Leucocytes in the Presence of Subinhibitory Concentrations of Melaleuca alternifolia and "Mentha of Pancalieri" Essential Oils.

The aim of this study was to evaluate the influence of tea tree oil (TTO) and "Mentha of Pancalieri" essential oil (MPP) on intracellular killing of Candida krusei, often resistant to conventional drugs, by polymorphonuclear leucocytes (PMNs). Intracellular killing was investigated by incubating yeasts and PMNs with essential oils (EOs) at 1/4 and 1/8 × MIC (Minimal Inhibitory Concentration), in comparison with anidulafungin, used as a reference drug. Killing values were expressed as Survival Index (SI) values. The cytotoxicity of EOs was evaluated by 3-[4,-5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. Both EOs were more efficaceous at 1/8 × MIC than 1/4 × MIC, with killing values higher than observed in EO-free systems and in presence of anidulafungin, indicating that the decreasing concentrations did not cause lower candidacidal activity. This better activity at 1/8 × MIC is probably due to the EOs' toxicity at 1/4 × MIC, suggesting that at higher concentrations EOs might interfere with PMNs functionality. TTO and MPP at 1/8 × MIC significantly increased intracellular killing by PMNs through their direct action on the yeasts (both EOs) or on phagocytic cells (MPP), suggesting a positive interaction between EOs and PMNs to eradicate intracellular C. krusei. These data showed a promising potential application of TTO and "Mentha of Pancalieri" EO as natural adjuvants in C. krusei infection management.

Evaluation of the Antifungal Activity of Mentha x piperita (Lamiaceae) of Pancalieri (Turin, Italy) Essential Oil and Its Synergistic Interaction with Azoles.

The promising antimicrobial activity of essential oils (EOs) has led researchers to use them in combination with antimicrobial drugs in order to reduce drug toxicity, side effects, and resistance to single agents. Mentha x piperita, known worldwide as "Mentha of Pancalieri", is produced locally at Pancalieri (Turin, Italy). The EO from this Mentha species is considered as one of the best mint EOs in the world. In our research, we assessed the antifungal activity of "Mentha of Pancalieri" EO, either alone or in combination with azole drugs (fluconazole, itraconazole, ketoconazole) against a wide panel of yeast and dermatophyte clinical isolates. The EO was analyzed by GC-MS, and its antifungal properties were evaluated by minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) parameters, in accordance with the CLSI guidelines, with some modifications. The interaction of EO with azoles was evaluated through the chequerboard and isobologram methods. The results suggest that this EO exerts a fungicidal activity against yeasts and a fungistatic activity against dermatophytes. Interaction studies with azoles indicated mainly synergistic profiles between itraconazole and EO vs. Candida spp., Cryptococcus neoformans, and Trichophyton mentagrophytes. Thus, the "Mentha of Pancalieri" EO may act as a potential antifungal agent and could serve as a natural adjuvant for fungal infection treatment.

Use of Pinus sylvestris L. (Pinaceae), Origanum vulgare L. (Lamiaceae), and Thymus vulgaris L. (Lamiaceae) essential oils and their main components to enhance itraconazole activity against azole susceptible/not-susceptible Cryptococcus neoformans strains.

BACKGROUND: Cryptococcal infections, besides being a problem for immunocompromised patients, are occasionally being a problem for immunocompetent patients. In addition, the lower susceptibility of this yeast to azoles is a growing problem in health care. To date, there are very few molecules with any activity towards Cryptococcus neoformans, leading to heightened interest in finding new alternatives or adjuvants to conventional drugs for the treatment of mycosis caused by this yeast. Since the essential oils (EOs) are considered as a potential rich source of bioactive antimicrobial compounds, we evaluated the antifungal activity of Origanum vulgare (oregano), Pinus sylvestris (pine), and Thymus vulgaris (thyme red) EOs, and their components (α-pinene, carvacrol, thymol) compared with fluconazole, itraconazole, and voriconazole, against C.neoformans clinical strains. Then, we investigated the effect of EOs and components in combination with itraconazole. METHODS: EO composition was analysed by Gas chromatography-mass spectrometry (GC-MS). A broth microdilution method was used to evaluate the susceptibility of C.neoformans to azoles, EOs and components. Checkerboard tests, isobolograms and time-kill assays were carried out for combination studies. RESULTS: Six C.neoformans isolates were susceptible to azoles, while one C.neoformans exhibited a reduced susceptibility to all tested azole drugs. All EOs exerted a good inhibitory activity against all C.neoformans strains. Pine EO was the most effective. Among components, thymol exerted the most remarkable activity. By checkerboard testing and isobolographic analysis, combinations of itraconazole with oregano, pine, or thyme EOs, and carvacrol were found to be synergistic (FICI≤0.5) against azole susceptible C.neoformans. Regarding the azole not susceptible C.neoformans strain, the synergistic effect with itraconazole was observed with thyme EO (chemotype: thymol 26.52%; carvacrol 7.85%), and carvacrol. Time-kill assays confirmed the synergistic effects of itraconazole and oregano or thyme EO against azole susceptible C.neoformans. Binary mixtures of itraconazole/thyme EO or carvacrol yielded additive effects on the azole not susceptible C.neoformans. CONCLUSIONS: Our findings highlight the potential effectiveness of thyme, oregano EOs, and carvacrol as natural and cost-effective adjuvants when used in combination with itraconazole. Identification of EOs exerting these effects could be one of the feasible ways to overcome drug resistance, reducing drug concentration and side effects.

The Microbiological@mind project: a public engagement initiative of Turin University bringing microbiology and health education into primary schools.

Despite ongoing global efforts, antimicrobial resistance continues to threaten the treatment of an ever-increasing range of bacterial infections. There is substantial evidence that public education programs that foster microbial literacy amongst young school audiences may improve correct knowledge of specific health issues, such as prevention of microbial infections and responsible use of antibiotics. The aim of the Microbiological@mind project was to engage primary school students with the subject of microbiology, to promote both scientific interest and awareness towards correct behaviors that may ensure a safer lifestyle. Interactive workshops based on a full ''hands-on'' approach were carried out by an expert team from the University of Turin to over 1200 children aged 9-11 years at primary schools in Turin. A questionnaire (pre- and post-activity test) on the main topic (i.e. antibiotics) was used to assess project effectiveness. The workshops provided a useful means to strengthen the understanding of basic microbiology concepts amongst students. Students' baseline knowledge of antibiotics was quite low, as low percentages of correct answers on antibiotic action and use (5.0% and 12.1%, respectively) were found in the pre-activity tests. A significant increase (P <0.0001) in correct knowledge was observed in the post-activity tests, after implementation of the teaching activity. Our findings support the idea that microbial literacy in early childhood through hands-on educational programs is of great importance to foster children's interest in science learning, and to provide young people with information about general and specific health-related issues, such as prudent antibiotic use, for a more responsible citizenship.

Liquid and vapour-phase antifungal activities of essential oils against Candida albicans and non-albicans Candida.

BACKGROUND: The management of Candida infections faces many problems, such as a limited number of antifungal drugs, toxicity, resistance of Candida to commonly antifungal drugs, relapse of Candida infections, and the high cost of antifungal drugs. Though azole antifungal agents and derivatives continue to dominate as drugs of choice against Candida infections, there are many available data referring to the anticandidal activity of essential oils. Since we have previous observed a good antimicrobial activity of some essential oils against filamentous fungi, the aim of this study was to extend the research to evaluate the activity of the same oils on Candida albicans, C.glabrata and C.tropicalis clinical strains, as well as the effects of related components. Essential oils selection was based both on ethnomedicinal use and on proved antibacterial and/or antifungal activity of some of these oils. Fluconazole and voriconazole were used as reference drugs. METHODS: The minimum inhibitory concentration (MIC) and the minimal fungicidal concentration (MFC) of essential oils (thyme red, fennel, clove, pine, sage, lemon balm, and lavender) and their major components were investigated by the broth microdilution method (BM) and the vapour contact assay (VC). RESULTS: Using BM, pine oil showed the best activity against all strains tested, though C.albicans was more susceptible than C.glabrata and C.tropicalis (MIC50-MIC90 = 0.06 %, v/v). On the contrary, sage oil displayed a weak activity (MIC50-MIC90 = 1 %, v/v). Thyme red oil (MIC50-MIC90 ≤ 0.0038 %, v/v for C.albicans and C.tropicalis, and 0.0078- < 0.015 %, v/v for C.glabrata), followed by lemon balm, lavender and sage were the most effective by VC. Carvacrol and thymol showed the highest activity, whereas linalyl acetate showed the lowest activity both by two methods. α-pinene displayed a better activity by BM than VC. CONCLUSION: Results show a good activity of essential oils, mainly thymus red and pine oils, and their components carvacrol, thymol and α-pinene against Candida spp., including fluconazole/voriconazole resistant strains. These data encourage adequately controlled and randomized clinical investigations. The use in vapour phase could have additional advantages without requiring direct contact, resulting in easy of environmental application such as in hospital, and/or in school.

Advances in Microbiology, Infectious Diseases and Public Health: Fungal Occurrence in the Hair and Skin of Symptomatic Pets in Turin, Italy.

Companion animals, often asymptomatic reservoir of fungi, can be important sources of infection in humans, due to the close contact with their owners. The present study was aimed to assess the occurrence of dermatophytes and other fungi isolated from pet dermatological lesions in Turin, Italy. Dermatological specimens were examined for fungal elements by direct microscopy and cultured to detect dermatophytes, other filamentous fungi and yeasts: 247 pets (118 cats, 111 dogs and 18 dwarf rabbits) were positive for fungal detection in culture. Microsporum canis was the most frequent dermatophyte in cats and dogs, whereas Trichophyton mentagrophytes was the most common in rabbits. Among the other fungi, for all examined pets, dematiaceous fungi were the most isolated, followed by Mucorales, penicilli, yeasts and yeast-like fungi, and aspergilli. No gender predisposition was detected for dermatophyte growth; on the contrary, for the other fungi male cats were more susceptible than female. The highest fungal occurrence was recorded in <1-year-old cats for dermatophytes, and in <5-year-old cats and dogs for the other fungi. Autumn was the period associated with a relevant incidence of fungal infection. Finally, fungi were more frequent in non pure-breed cats and in pure-breed dogs. These data underline the importance to timely inform pet owners about the potential health risk of infection caused not only by dermatophytes but also by non-dermatophyte fungi, routinely considered to be contaminants or harmless colonizers, since their role as source of zoonotic infections is not to be excluded.

Advances in Microbiology, Infectious Diseases and Public Health: Refractory Trichophyton rubrum Infections in Turin, Italy: A Problem Still Present.

Dermatophytosis caused by Trichophyton rubrum is the most common cutaneous fungal infection in industrialized countries and worldwide with high recurrence and lack of treatment response. In addition, patients with cutaneous and concurrent toenail lesions are often misdiagnosed and therefore treated with an inappropriate therapy. In this study, we evaluated five previously misdiagnosed cases of T.rubrum chronic dermatophytosis sustained by two variants at sites distant from the primary lesion. Our patients were successfully treated by systemic and topical therapy, and 1 year after the end of therapy follow-up did not show any recurrence of infection.Our data indicate that the localization of all lesions, the isolation and the identification of the causative fungus are essential to establish the diagnosis and the setting of a correct therapeutic treatment to avoid recurrences.

Key Roles of Human Polymorphonuclear Cells and Ciprofloxacin in Lactobacillus Species Infection Control.

Lactobacilli have the potential to act as reservoirs of antibiotic resistance genes similar to those found in human pathogens, with the risk of transferring these genes to many pathogenic bacteria. In this study, we investigated the role of human polymorphonuclear cells (PMNs) against Lactobacillus spp. both resistant and susceptible to ciprofloxacin (a fluoroquinolone) and the effect of ciprofloxacin on the interaction between PMNs and three Lactobacillus spp. with different patterns of susceptibility to this drug. Hence, the primary functions of PMNs, such as phagocytosis and bacterial intracellular killing, against lactobacilli were investigated. The rate of PMN phagocytosis was high for ciprofloxacin-sensitive and ciprofloxacin-resistant strains. The patterns of intracellular killing of ciprofloxacin-sensitive and ciprofloxacin-resistant strains by PMNs underline that PMNs alone were able to kill lactobacilli. The addition of ciprofloxacin to PMNs did not result in a significant increase in the bacterial uptake by phagocytes. On the contrary, ciprofloxacin had a marked effect on PMN intracellular killing, resulting in increased numbers of killed ciprofloxacin-sensitive bacteria in comparison with antibiotic-free controls. Our data show that by itself, the profile of antibiotic resistance does not constitute an intrinsic factor of greater or lesser pathogenicity toward the host. The ability of PMNs to kill a diverse array of bacterial pathogens is essential for human innate host defense, primarily in immunocompromised patients.

In vitro antifungal activity of fluconazole and voriconazole against non-Candida yeasts and yeast-like fungi clinical isolates.

The risk of opportunistic infections caused by non-Candida yeasts and yeast-like fungi is increasingly common, mainly in immunocompromised patients. Appropriate first-line therapy has not been defined and standardized, mainly due to the low number of cases reported. To improve empirical treatment guidelines, we describe the susceptibility profile to fluconazole and voriconazole of 176 non-Candida yeasts and yeast-like fungi collected from hospitals in Piedmont, North West Italy from January 2009 to December 2013. The results showed that most isolates are susceptible to voriconazole (94%), but less susceptible to fluconazole (78%), suggesting that voriconazole could be used as first-line therapy in infections caused by these fungi.

Bacteriological findings in radicular cyst and keratocystic odontogenic tumour fluids from asymptomatic patients.

OBJECTIVE: In this study the potential presence of bacteria in radicular cyst (RC) and keratocystic odontogenic tumour(KCOT) fluids from clinically asymptomatic patients was investigated. MATERIALS AND METHODS: Cyst fluids were collected by needle aspiration from 16 patients with asymptomatic osteolytic lesions (10 RCs and 6 KCOTs) undergoing surgery. All samples were transferred into tubes containing pre-reduced transport medium, delivered to the microbiology laboratory and processed within 1h. The cysts, surgically enucleated, were sent for standard histopathological examination. Cyst fluid samples were cultured on selective and differential media in anaerobic (for about 2 weeks) and aerobic (for 24-48 h) conditions to detect viable microorganisms. After incubation, the colonies were counted, Gram-stained and identified by biochemical tests. RESULTS: Cultures were positive for the presence of bacteria in 15 (9 RCs, 6 KCOTs) out of 16 cases. RCs and KCOTs generally yielded low bacterial counts (10(2)-10(4) CFU/ml) and were predominantly colonized by obligate anaerobes (64%), whereas less commonly by facultative anaerobes (36%). No significant differences in the detection frequencies of obligate and facultative anaerobes were evidenced between RCs and KCOTs. Propionibacterium acnes was the most common obligate anaerobe recovered both in RC and KCOT fluids. Among facultative anaerobes, Gemella morbillorum was more frequently isolated in KCOTs, whereas Staphylococcus spp. in RCs. CONCLUSIONS: Bacteria may be present and persist within fluids of clinically asymptomatic jaw cystic lesions. The influence of bacteria and latent bacterial infection within cystic jaw lesions should be reconsidered in odontogenic cyst progression.

Caspofungin benefit on phagocytes from patients with renal dysfunction infected with multidrug-resistant Candida glabrata.

AIM: We evaluated the potential impact of caspofungin (CAS) on the functional activities of polymorphonuclear leukocytes (PMNs) from hemodialyzed patients (HDs) and renal transplant recipients (RTRs) against a multidrug-resistant clinical strain of Candida glabrata compared with those of PMNs from healthy subjects (HSs). MATERIALS & METHODS: Effects of CAS on PMN phagocytosis and intracellular killing towards multidrug-resistant C. glabrata were evaluated in 66 HDs, 54 RTRs and 30 HSs in the absence and presence of CAS at MIC and sub-MICs. RESULTS: When HD PMNs and RTR PMNs were exposed to both MICs and sub-MICs of CAS, their fungicidal activity against the multidrug-resistant C. glabrata strain was significantly higher than that of drug-free controls, with survival index values that overlapped with those achieved by HS PMNs. CONCLUSION: The obtained results underline the role of CAS in the restoration of the impaired PMN functions in HDs and RTRs. CAS might still constitute an effective therapeutic option for the treatment of invasive fungal infections caused by multidrug-resistant C. glabrata in patients with altered phagocyte-dependent innate immunity.

Positive interaction of thyme (red) essential oil with human polymorphonuclear granulocytes in eradicating intracellular Candida albicans.

The essential oils have started to be recognized for their potential antimicrobial role only in recent years. Clinical experience showed that the efficacy of antimicrobial agents depends not only on their direct effect on a given microorganism but also on the functional activity of the host immune system. Since data on the effects of essential oils on the innate immune system are scanty and fragmentary, the aim of this study was to evaluate the influence of thyme (red) essential oil (EO), at subinhibitory/inhibitory concentrations, on intracellular killing activity by human polymorphonuclear granulocytes (PMNs) against Candida albicans. In order to provide a frame of reference for the activity of this EO, its in vitro killing activity in the absence of PMNs was also evaluated.Results showed that EO at subminimal inhibitory (subMIC)/minimal inhibitory (MIC) concentrations significantly enhanced intracellular killing of C. albicans in comparison with EO-free controls and was comparable to the positive control (fluconazole). In in vitro killing assays without PMNs, we observed progressive growth of the yeast cells in the presence of EO subMIC/MIC concentrations. A positive antifungal interaction with phagocytes could explain why this EO, which appeared to be only fungistatic in time-kill assays, had efficacy in killing yeast cells once incubated with PMNs.