Antimicrobial Resistance of Streptococcus pneumoniae Clinical Serotypes between 2017 and 2022 in Crete, Greece.

BACKGROUND: Pneumococcal disease is still considered a global problem. With the introduction of pneumococcal conjugate vaccines (PCVs) serotype epidemiology changed, but antimicrobial resistance persists constituting a serious problem. The current study aimed to determine the serotype distribution and the antimicrobial susceptibility of recent Streptococcus pneumoniae isolates, following implementation of the 13-valent conjugate vaccine (PCV13). MATERIALS AND METHODS: From January 2017 to December 2022 we evaluated 116 nonduplicate S. pneumoniae isolates collected from adult patients (21 - 98 years) cared for in the University Hospital of Heraklion, Crete, Greece. Pneumococcal isolates were serotyped by the Quellung reaction, and antimicrobial susceptibility testing was performed using E-test. Multidrug resistance (MDR) was defined as non-susceptibility to at least one agent in ≥3 classes of antibiotics. RESULTS: Among the 116 isolates, 31% were recognized as invasive pneumococcal strains, while 69% were non-invasive. The isolates tested belonged to 25 different serotypes. The most prevalent serotypes were 11A (10.3%), and 35B (10.3%), followed by 3 (9.5%), 15A (7.8%), 25F (6.9%), 19A (5.3%), 35F (5.3%), and others (44.6%). The coverage rates of PCV13 and the pneumococcal polysaccharide vaccine (PPSV23) were 26.7% and 57.8%, respectively. PCV13 and PPSV23 serotypes decreased between 2017 - 2019 and 2020 - 2022, with a parallel increase in the non-vaccine types. Resistance rates to erythromycin, clindamycin, trimethoprim/sulfamethoxazole, penicillin, levofloxacin, and ceftriaxone, were 40.5%, 21.6%, 13.8%, 12.1%, 3.4%, and 0%, respectively. All isolates were susceptible to vancomycin, linezolid, and daptomycin. MDR was observed among 36 (31%) S. pneumoniae isolates. CONCLUSION: The increasing levels of resistance in S. pneumoniae in Crete, Greece, highlight the need for continuous surveillance of antimicrobial resistance and development of strategies for its reduction, including antimicrobial stewardship programs, increased pneumococcal vaccination, and development of next generation PCVs with a wider serotype coverage.

In Vitro Activities of Ceftobiprole, Dalbavancin, Tedizolid and Comparators against Clinical Isolates of Methicillin-Resistant Staphylococcus aureus Associated with Skin and Soft Tissue Infections.

Skin and soft tissue infections (SSTIs) are associated with significant morbidity and healthcare costs, especially when caused by methicillin-resistant Staphylococcus aureus (MRSA). Vancomycin is a preferred antimicrobial therapy for the management of complicated SSTIs (cSSTIs) caused by MRSA, with linezolid and daptomycin regarded as alternative therapeutic options. Due to the increased rates of antimicrobial resistance in MRSA, several new antibiotics with activity against MRSA have been recently introduced in clinical practice, including ceftobiprole, dalbavancin, and tedizolid. We evaluated the in vitro activities of the aforementioned antibiotics against 124 clinical isolates of MRSA obtained from consecutive patients with SSTIs during the study period (2020-2022). Minimum inhibitory concentrations (MICs) for vancomycin, daptomycin, ceftobiprole, dalbavancin, linezolid and tedizolid were evaluated by the MIC Test Strip using Liofilchem strips. We found that when compared to the in vitro activity of vancomycin (MIC90 = 2 µg/mL), dalbavancin possessed the lowest MIC90 (MIC90 = 0.094 µg/mL), followed by tedizolid (MIC90 = 0.38 µg/mL), linezolid, ceftobiprole, and daptomycin (MIC90 = 1 µg/mL). Dalbavancin demonstrated significantly lower MIC50 and MIC90 values compared to vancomycin (0.064 vs. 1 and 0.094 vs. 2, respectively). Tedizolid exhibited an almost threefold greater level of in vitro activity than linezolid, and also had superior in vitro activity compared to ceftobiprole, daptomycin and vancomycin. Multidrug-resistant (MDR) phenotypes were detected among 71.8% of the isolates. In conclusion, ceftobiprole, dalbavancin and tedizolid exhibited potent activity against MRSA and are promising antimicrobials in the management of SSTIs caused by MRSA.

Exophiala dermatitidis Central Line-Associated Bloodstream Infection in a Child with Ewing's Sarcoma: Case Report and Literature Review on Paediatric Infections.

Exophiala dermatitidis is a dematiaceous, ubiquitous, dimorphic fungus, which can cause a wide range of invasive diseases in both immunocompromised and immunocompetent hosts. Bloodstream infections due to E. dermatitidis are rarely encountered in clinical practice, especially in pediatric patients. We describe a case of central line-associated bloodstream infection due to E. dermatitidis in a 4.5-year-old boy with Ewing's sarcoma. The fungus was isolated from blood specimens taken from the Hickman line. The isolate was identified by its phenotypic characteristics, by MALDI-TOF and by using molecular methods. The infection was successfully treated with voriconazole and catheter removal. The literature was also reviewed on pediatric infections caused by E. dermatitidis, focusing on clinical manifestations and challenges associated with diagnosis and management.

Antifungal activity of amniotic fluid against different Candida species.

BACKGROUND: Although Candida is a commensal of the urogenital tract, intrauterine fungal infections are extremely uncommon in clinical practice. AIMS: In the present work we evaluated whether amniotic fluid (AF) possesses direct antifungal activity against clinical isolates of Candidaalbicans and other Candida species. METHODS: A total of 23 AF samples from pregnant women with gestational age of 38-41 weeks were obtained under aseptic conditions by the aspiration of the amniotic sac during cesarean section. Different Candida species were inoculated in amniotic fluid and Sabouraud broth, used as control, and were incubated at 37°C for 48h. Quantitative cultures of test samples and controls were performed at 0, 4, 8, 12, 24, and 48h. RESULTS: AF collected from 23 pregnant women had consistent and significant inhibitory activity against all Candida isolates tested. Nonetheless, a complete inhibition of growth by all 23 AF samples tested was observed only against Candida glabrata. CONCLUSIONS: It is likely that the antifungal activity of the AF against C. albicans, C. glabrata and Candida parapsilosis observed in vitro also exists in vivo, contributing to protect against intrauterine fungal infections.

Oral Bacteria and Intestinal Dysbiosis in Colorectal Cancer.

The human organism coexists with its microbiota in a symbiotic relationship. These polymicrobial communities are involved in many crucial functions, such as immunity, protection against pathogens, and metabolism of dietary compounds, thus maintaining homeostasis. The oral cavity and the colon, although distant anatomic regions, are both highly colonized by distinct microbiotas. However, studies indicate that oral bacteria are able to disseminate into the colon. This is mostly evident in conditions such as periodontitis, where specific bacteria, namely Fusobacterium nucrelatum and Porphyromonas gingivalis project a pathogenic profile. In the colon these bacteria can alter the composition of the residual microbiota, in the context of complex biofilms, resulting in intestinal dysbiosis. This orally-driven disruption promotes aberrant immune and inflammatory responses, eventually leading to colorectal cancer (CRC) tumorigenesis. Understanding the exact mechanisms of these interactions will yield future opportunities regarding prevention and treatment of CRC.

Epidemiology and antifungal susceptibility patterns of Candida isolates from Greek women with vulvovaginal candidiasis.

Vulvovaginal candidiasis (VVC) is a common infection of the genital tract affecting millions of women worldwide. Data on epidemiological trends of VVC in Greece are scarce. This study was undertaken to evaluate the prevalence of VVC among symptomatic women in Crete, Greece, identify the Candida species involved and determine their susceptibility to antifungals. Over a 6-year period (2012-2017), 10 256 symptomatic women with vaginitis were evaluated. Isolation of yeasts was performed on Sabouraud dextrose agar with chloramphenicol, and the isolates were identified using the API 20 C AUX and/or the Vitek 2 YST card. Susceptibility of the isolates to amphotericin, fluconazole, voriconazole and flucytosine was determined by the Vitek 2 automated system. The results were interpreted according to Clinical and Laboratory Standards criteria. Vaginal swab cultures of 1217 (11.9%) women yielded Candida species. Recurrent VVC was documented in 62 (5.1%) of them. Candida albicans was the most frequently isolated species (75.6%), followed by Candida glabrata (13.6%). Overall, resistance rates to amphotericin B, fluconazole, voriconazole and flucytosine were 0.2%, 6.6%, 1.4% and 2.1%, respectively. Fluconazole resistance of C. albicans significantly increased in the second period of the study (2015-2017) (P = 0.031). This study demonstrated that VVC is a common infection among women in our region, with C. albicans being the predominant species involved. Although resistance to antifungals was infrequent, resistance to fluconazole among C. albicans isolates was found to significantly increase with time. Continued surveillance of changes in species distribution and susceptibility to antifungals are necessary to guide treatment.

The Evolving Epidemiology of Serotype Distribution and Antimicrobial Resistance of Streptococcus pneumoniae Strains Isolated from Adults in Crete, Greece, 2009-2016.

BACKGROUND: Pneumococcal disease is a major cause of morbidity and mortality worldwide, especially in patients with comorbidities and advanced age. This study evaluated trends in epidemiology of adult pneumococcal disease in Crete, Greece, by identifying serotype distribution and antimicrobial resistance of consecutive Streptococcus pneumoniae strains isolated from adults during an 8-year time period (2009-2016) and the indirect effect of the infant pneumococcal higher-valent conjugate vaccines 10-valent pneumococcal conjugate vaccine (PCV10) and 13-valent pneumococcal conjugate vaccine (PCV13). MATERIALS AND METHODS: Antimicrobial susceptibility was performed by E-test and serotyping by Quellung reaction. Multidrug resistance (MDR) was defined as non-susceptibility to penicillin (PNSP) combined with resistance to ≥2 non-ß-lactam antimicrobials. RESULTS: A total of 135 S. pneumoniae strains were isolated from adults during the study period. Twenty-one serotypes were identified with 17F, 15A, 3, 19A, and 11A, being the most common. The coverage rates of PCV10, and PCV13 were 17.8% and 37.8%, respectively. PCV13 serotypes decreased significantly from 68.4% in 2009 to 8.3% in 2016 (P = 0.002). The most important emerging non-PCV13 serotypes were 17F, 15A, and 11A, with 15A being strongly associated with antimicrobial resistance and MDR. Among all study isolates, penicillin-resistant and MDR strains represented 7.4% and 14.1%, respectively. Predominant PNSP serotypes were 19A (21.7%), 11A (17.4%), and 15A (17.4%). Erythromycin, clindamycin, tetracycline, trimethoprim-sulfamethoxazole, and levofloxacin resistant rates were 30.4%, 15.6%, 16.3%, 16.3%, and 1.5%, respectively. CONCLUSION: Although pneumococcal disease continues to be a health burden in adults in Crete, our study reveals a herd protection effect of the infant pneumococcal higher-valent conjugate vaccination. Surveillance of changes in serotype distribution and antimicrobial resistance among pneumococcal isolates are necessary to guide optimal prevention and treatment strategies.

Study on the comparative activity of echinocandins on murine gut colonization by Candida albicans.

Colonization of the gastrointestinal (GI) tract by Candida species is a principal pathogenetic event for development of invasive candidiasis. Importantly, the effect of echinocandins, the preferred antifungal agents for treatment of invasive candidiasis, on GI tract colonization by Candida spp. is currently unknown. Herein, we used an established model of persistent murine GI tract colonization by Candida albicans to test the ability of different echinocandins to eradicate the yeast from murine gut. Adult male Crl:CD1 (ICR) BR mice were fed with chow containing C. albicans and subsequently treated with different echinocandins or normal saline via daily intraperitoneal injections for 10 days. Quantitative stool cultures were performed immediately before (week one), and weekly for three months after discontinuation of treatment. Notably, treatment with all three echinocandins used (caspofungin, anidulafungin, and micafungin) resulted in eradication of Candida albicans from the stools, as evidenced by the significant reduction of yeast cells from a mean of 4.2 log10 CFU/g of stool before treatment (week one of colonization) to undetectable (<2 log10 CFU/g of stool) levels (week 12, P < 0.0001). In contrast, there was no significant reduction of Candida yeast cells in the stools of control mice. Collectively, the ability of echinocandins to eradicate C. albicans from the stools could have important implications in prophylaxis of high-risk patients for development of invasive candidiasis originating from the GI tract.

Treatment of relapsed/refractory multiple myeloma with thalidomide-based regimens: identification of prognostic factors.

We evaluated the predictive value of several parameters, including the International Staging System (ISS) for myeloma, in patients with advanced disease treated with thalidomide-based regimens (TBR). We analyzed 119 patients, from 3 phase II studies. Patients with pretreatment beta2 microglobulin<3.5 mg/l and albumin 3.5 g/dl were scored ISS stage 1, patients with beta2 microglobulin<3.5 mg/l and albumin<3.5 g/dl or beta2 microglobulin 3.5-5.5 mg/l regardless of albumin levels were scored ISS stage 2, patients with beta2 microglobulin>5.5 mg/l ISS stage 3. ISS stage was 1, 2 and 3 in 45, 32 and 23% of patients respectively. Seventy-four patients (62%) achieved at least partial response. Median progression-free and overall survival were 8 months and 19.5 months respectively. ISS stage, serum LDH and performance status were independent predictive factors for survival. Based on these 3 variables a scoring system was developed with survival times of 38.1, 28.8 and 5.8 months for scores 0, 1 and 2 respectively. The ISS staging system was highly predictive for overall survival of patients with advanced myeloma treated with TBR. With the addition of performance status and serum LDH, a simple scoring system was developed which may help select patients likely to benefit from TBR.

Hormonal therapy with letrozole for relapsed epithelial ovarian cancer. Long-term results of a phase II study.

OBJECTIVE: We conducted a phase II trial to evaluate the activity of oral letrozole in women with relapsed or recurrent epithelial ovarian cancer. METHODS: Twenty-seven patients were treated with letrozole at a dose of 2.5 mg once a day. Patients with measurable or evaluable disease (n = 21) and those with only increasing CA 125 serum levels (n = 6) were eligible. Paraffin-fixed histological sections from tumor specimens resected at the initial laparotomy were assessed for the presence of estrogen, and progesterone receptors. RESULTS: Among the 21 patients with measurable or evaluable disease who were evaluated for response by WHO criteria, we observed one complete and two partial responses for an objective response rate of 15%. Using criteria for CA 125 response we obtained a marker response in 4 of 27 patients (15%), and the marker remained stable in 5 additional patients (18%). Letrozole treatment was generally well tolerated. No correlation was observed between tumor marker response or stabilization and either estrogen or progesterone receptor expression. CONCLUSION: The results of our study suggest that the aromatase inhibitor letrozole is an agent with some activity and limited toxicity for relapsed ovarian cancer. As we could not find any association between response and hormonal receptor expression, the underlying mechanisms of letrozole action have to be elucidated.

Discordant response or progression in patients with myeloma treated with thalidomide-based regimens.

UNLABELLED: Thalidomide-based regimens (TBR) are now widely used for the treatment of refractory multiple myeloma and have shown significant activity in newly diagnosed patients. In some patients with secretory disease, we observed discrepancies between the reduction of the monoclonal protein levels and the plasma cell infiltration in the bone marrow and/or extramedullary sites of relapse after treatment with TBR. The purpose of this study was to assess the incidence and analysis of this phenomenon in all myeloma patients treated with TBR in our Institution. PATIENTS AND METHODS: We studied all patients who received TBRs and had a follow up time of at least 6 months. Partial response (PR) was defined as at least 50% reduction of serum myeloma protein and soft tissue plasmacytomas and/or > 90% reduction of Bence Jones protein excretion and minor response as a > 25% reduction of the serum myeloma protein or > 50% reduction of the Bence Jones myeloma protein. RESULTS: Between July 1999 and July 2002 we treated 94 patients with advanced myeloma and 9 patients with newly diagnosed disease with TBR. Sixty-seven patients (66%) achieved either partial or minor response. In 4 patients (3 with advanced and 1 with newly diagnosed myeloma) the bone marrow was heavily infiltrated by plasma cells, despite a decrease of the paraprotein levels ranging from 38% to 68%. This discordance between monoclonal protein levels and bone marrow plasmacytosis was noted in 6% of patients rated as responders and in 11% of responding patients who actually had a repeat bone marrow assessment. Furthermore 6 responding patients, after achieving a PR which lasted between 5 and 9 months, relapsed with bone marrow (all cases), and extramedullary (2 cases) plasmacytosis, without increase of serum and/or urine monoclonal protein. This hyposecretory conversion was noted in 12.5% of relapsing patients. CONCLUSION: Our data indicate that after treatment with TBR some patients with myeloma show discordant responses of the monoclonal protein levels and the bone marrow or extramedullary plasmacytosis. If our data are confirmed, they may have practical implications for assessment of response and follow up of patients treated with TBR.

Pulsed cyclophosphamide, thalidomide and dexamethasone: an oral regimen for previously treated patients with multiple myeloma.

INTRODUCTION: Thalidomide is an oral agent with significant activity in one-third of patients with refractory myeloma. However, long-term continuous administration of thalidomide can be associated with significant side effects such as deep-vein thrombosis and peripheral neuropathy. Furthermore, it is not clear whether continuous administration of thalidomide is necessary for its antimyeloma effect. We performed a phase II study with a combination that was based on the intermittent administration of thalidomide. MATERIALS AND METHODS: A total of 53 patients with previously treated myeloma received cyclophosphamide 150 mg/m(2) p.o. every 12 h before meals on days 1-5, thalidomide 400 mg p.o. in the evening on days 1-5 and 14-18 and dexamethasone 20 mg/m(2) in the morning after breakfast on days 1-5 and 14-18 (CTD). The CTD combination was repeated every 28 days for three courses. Subsequently, responding patients were scheduled to receive maintenance treatment with monthly courses of CTD administered only for the first five days of each month. RESULTS: On an intention-to-treat basis, 32 patients (60%) achieved a partial response with a median time to response of 1.5 months. Among the 43 thalidomide-naïve patients, 67% responded. Toxicities were mild or moderate and the cumulative incidence of deep-vein thrombosis and peripheral neuropathy was 4 and 2%, respectively. The median time to progression for responding patients was 12 months and the median overall survival for all patients was 17.5 months. CONCLUSION: The oral, outpatient pulsed CTD regimen is associated with significant activity in patients with previously treated multiple myeloma. The incidence of deep-vein thrombosis and peripheral neuropathy appears to be lower than expected when thalidomide is being administered on a continuous basis.

Treatment of Waldenstrom's macroglobulinemia with the combination of fludarabine and cyclophosphamide.

Fludarabine is an active agent for the treatment of Waldenstrom's macroglobulinemia (WM) and its combination with cyclophosphamide has been effective in many patients with low-grade lymphoma and chronic lymphocytic leukemia. Based on these data, we administered the combination of fludarabine (25 mg/m2 i.v. day 1-3) and cyclophosphamide (250 mg/m2 i.v. day 1-3,) to 11 patients with WM. Most patients had features indicating poor prognosis including median age of 73 years (range 60-84 years), hemoglobin <100 g/l in 73%, B2-microglobulin >3 mg/l in 64%, symptomatic hyperviscosity in 55% of patients. Only 2 patients were previously untreated, 7 were primary refractory and 2 were relapsing on treatment. The fludarabine-cyclophosphamide combination (FC) was administered every 4 weeks for a total of four courses. Partial response, defined by at least 50% reduction of serum monoclonal protein and of tumor infiltrate at all involved sites was documented in 6 patients (55%) (The median time to response was 4 months). Responding patients demonstrated resolution of disease-related symptoms and correction of anemia. Median time to progression for all patients was 24 months. With a mean follow-up of 28 months, two of six responding patients have progressed so far. The probability of 2-year survival is 70%. This regimen was relatively well tolerated. Complications included neutropenia grade 3 or 4 in 3 patients and thrombocytopenia grade 3 or 4 in 2 patients. There were five infectious episodes including two episodes of neutropenic fever. We conclude that the FC combination appears to be active in patients with WM most of whom were resistant to treatment and had poor prognostic factors. The addition of rituximab to FC requires further investigation.

Rituximab in combination with CNOP chemotherapy in patients with previously untreated indolent non-Hodgkin's lymphoma.

Rituximab, a chimeric monoclonal antibody, produces response rates of up to 73% in patients with previously untreated indolent non-Hodgkin's lymphoma (NHL), and has high activity when combined with chemotherapy. The purpose of this phase II study was to determine the efficacy and safety of rituximab plus cyclophosphamide, mitoxantrone, vincristine and prednisone (CNOP) chemotherapy in patients with indolent NHL. In all, 42 patients (median age 67 years) with previously untreated follicular, marginal zone or small lymphocytic/lymphoplasmacytic NHL received six infusions of rituximab (375 mg/m(2)) in combination with six cycles of CNOP. The overall response rate was 90% comprising 30 complete (71%) and eight partial (19%) responses. Although patients with marginal zone lymphoma or International Prognostic Index (IPI) score 3 had lower complete response rates, no significant difference in overall response rate was observed between the histological groups (P=0.24) or between patients stratified according to IPI score (P>0.05). Median overall survival, time-to-progression and response duration had not been reached after a median 19.5-month follow-up. In all, 31 patients (74%) are currently free from progression and 38 (90%) remain alive. Treatment was well tolerated. One patient (2%) experienced grade 3/4 infusion-related toxicity; 13 (31%) grade 3/4 leukopenia and 18 (43%) grade 3/4 neutropenia. Infection was observed in nine patients: eight (19%) grade 1/2 and one (2.4%) grade 3. This study demonstrates that combining rituximab with CNOP achieves high remission rates without significant additional toxicity in patients with previously untreated indolent NHL. Further follow-up will determine response duration and survival.

Solitary bone plasmacytoma and extramedullary plasmacytoma.

Solitary bone and extramedullary plasmacytomas are rare plasma cell proliferative disorders. Their diagnosis is based on histologic confirmation of monoclonal plasma cell infiltration of a single disease site and on the exclusion of systemic myeloma. For both entities, the treatment of choice is localized radiotherapy. With modern radiotherapy and with a total dose of at least 4000 cGy, the risk for local recurrence is less than 5%. There is no role for systemic chemotherapy in the management of these disorders. Approximately 30% of patients with solitary bone plasmacytoma (SBP) remain disease-free for several years; some of these patients may be cured. Patients with the best prognosis are those in whom the monoclonal protein disappears by 1 year after radiotherapy. The prognosis of patients with solitary extramedullary plasmacytoma (SEP) appears to be better than for patients with SBP because approximately 70% of patients with SEP remain disease-free at 10 years. With more sensitive staging procedures, the diagnosis of SBP and SEP may become less common, but the number of patients with prolonged stability and cure may increase.

Extended rituximab therapy for previously untreated patients with Waldenström's macroglobulinemia.

Waldenström's macroglobulinemia is a low-grade lymphoplasmacytoid lymphoma characterized by CD20 expression on malignant cells. Several studies have indicated that the anti-CD20 monoclonal antibody rituximab has activity against this disease. Thus, we performed a prospective study in which 17 previously untreated patients with symptomatic macroglobulinemia were treated with rituximab 375 mg/m2 intravenously for 4 weeks. Three months after completion of rituximab, patients without evidence of progressive disease received repeat 4-week courses of this agent. Six patients (35%) achieved a partial response after extended treatment with rituximab. Median time to response was 3 months. The median time to progression (TTP) for all patients was 13 months. One of 6 responding patients has progressed at 10 months, while the other 5 patients remain progression free with a follow-up range of > 22-40 months. Eight patients (47%) were rated as stable disease, and their median TTP was 9 months. Treatment with rituximab was well tolerated and was not associated with myelosuppression; one third of the patients experienced infusion-related toxicity, usually fever and chills of mild degree. Our prospective trial of extended rituximab therapy for previously untreated patients with Waldenström's macroglobulinemia indicates that this agent is active, well tolerated, and might be associated with a long period without the need for further treatment. Studies that will combine rituximab with chemotherapy or with other monoclonal antibodies might be of interest.