Mycotoxins and flours: Effect of type of crop, organic production, packaging type on the recovery of fungal genus and mycotoxins.

Heat-stable mycotoxins are widely detected in flour and produced by Aspergillus spp., Fusarium spp. and Penicillium spp. Forty different flours purchased in Italy are used to assess potential risk factors via a systematically screening of a number of variables: the type of flour, organic, whole and white wheat, types of packaging (paper, plastic and weight). Fungal recovery and co-occurrence of specific mycotoxins was also assessed. The results showed that flour originated from fruits had a significant higher recovery of fungi, while seed/pseudocereals had the highest mycotoxins detection. Flours originating from organic agriculture are more prone to higher fungal recovery and mycotoxins detection when compared with not-organic flours. Packaging is also important: packaging weighting less than 376 g supports significantly more fungal recovery and the plastic packages was observed to retain more fungi and mycotoxins detection when compared with paper. Recovery measured as Log (CFU/g) of fungal genera is not directly proportional to the amount of mycotoxins. Finally, linear regression and mixed logit regression models show that the mean level of aflatoxins B1 (ng/g on the logarithmic scale) reduces by 0.485 when moving from an organic to a non-organic flour, while a significant increase of 0.369 when moving from paper to a plastic packaging.

Antifungal activity of different essential oils against Malassezia pathogenic species.

ETHNOPHARMACOLOGICAL RELEVANCE: Plants produce essential oils in response to physiological stresses, pathogen attacks and ecological factors. Nowadays, they are recognized as defence compounds and attractors of pollinators. Essential oils have been traditionally used in the past years in various cultures for medicinal and health purposes. In recent times due to their well-documented antimicrobial activities, essential oils have consolidated their use in raw and processed food preservation, health and clinical uses. AIMS OF THE REVIEW: The potential activity of essential oils against the largely diffused Malassezia species on the human skin, which can cause common infections or exacerbate multiple skin disorders, such as P. versicolor, folliculitis, seborrheic dermatitis and dandruff, atopic dermatitis and psoriasis. MATERIALS AND METHODS: Information on essential oils activity against Malassezia species was obtained from published materials, including books and electronic databases, such as SCI finder, PubMed, Web of Science, ACS, Science Direct, Wiley, Springer, Taylor, J-STAGE and Google Scholar. Search was conducted covering the period from January 2013 to December 2018. RESULTS: In the in vitro studies diverse methods were used to test the essential oils activity, namely broth microdilution method, which resulted the most used one, followed by agar disk diffusion and vapour phase methods. Essential oils obtained by steam distillation were from different plant genera, Thymus, Artemisia, Malaleuca, Cinnamomun, Ocimum, Zataria, Rosmarinus, Origanum, Syzigium, Foenicolum, Thapsia, Tachyspermum, Myrtus. MIC values were expressed as µg/mL, µL/mL and by inhibition zone (mm) or µL/cm3. All the investigated essential oils were active at the tested conditions. In addition, two clinical studies used essential oils from Cymbopogon citratus and C. flexuosus formulated in shampoo, cream or lotion for the successful treatment of dandruff and P. versicolor. CONCLUSIONS: Results of these studies indicate worthy prospects for clinical application of essential oils and there is an urgent need to conduct further in vivo studies with large number of patients in order to verify the clinical potential of essential oils against Malassezia species.

Liquid and Vapor-Phase Activity of Artemisia annua Essential Oil against Pathogenic Malassezia spp.

Artemisia annua essential oil has given us many encouraging results for its numerous antimicrobial properties. In this study, the essential oil, both in liquid and in vapor phases, was tested against various Malassezia species closely related to many skin disorders in humans and animals. Malassezia treatment and eradication are mainly based on old azole drugs, which are characterized by poor compliance, unpredictable clinical efficacy, emerging resistance, and several side effects. Monoterpenes (ca. 88%) represent the most abundant group of compounds in the essential oil, mainly the oxygenated derivatives (ca. 74%) with camphor (25.2%), 1,8-cineole (20%), and artemisia ketone (12.5%). In vapor phase, monoterpenes represent more than 98% of the constituents, α-pinene being the main constituent (22.8%), followed by 1,8-cineole (22.1%) and camphene (12.9%). Essential oil of A. annua, both in vapor phase and liquid, showed strong antimicrobial activity towards almost the tested twenty strains of Malassezia analyzed. The minimum fungicidal concentrations from most of the strains tested were from 0.78 µL/mL to 1.56 µL/mL, and only three strains of Malassezia sympodialis required a higher concentration of 3.125 µL/mL. Overall, the minimal inhibitor concentrations obtained by vapor diffusion assay were lower than those obtained by the liquid method. The average values of minimal inhibitor concentrations obtained by the two methods at 72 h are 1.3 - 8.0 times higher in liquid compared to those in the vapor phase.

Enzymatic characterization of clinical and environmental Cryptococcus neoformans strains isolated in Italy / Caracterización enzimática de aislamientos clínicos y ambientales de Cryptococcus neoformans de Italia

Background. Cryptococcus neoformans is an encapsulated yeast causing mainly opportunistic infections. The virulence factors involved in cryptococcosis pathogenesis include the presence and the size of the polysaccharide capsule, the production of melanin by phenoloxidase, the growth at 37°C and the enzyme secretion like proteinase, phospholipase and urease. Many other enzymes are secreted by C. neoformans but their role in the fungus virulence is not yet known. Aims. In order to investigate this topic, we compared the phospholipase production between strains from patients and from bird droppings, and we examined its relationship to phenoloxidase production. We further characterized the strains by determining the activity of 19 different extracellular enzymes. Methods. Two hundred and five Italian C. neoformans clinical isolates and 32 environmental isolates were tested. Phenoloxidase production was determined by the development of brown colonies on Staib's agar. Extracellular phospholipase activity was performed using the semiquantitative egg-yolk plate method. API ZYM commercial kit was used to observe the production and the activity of 19 different extracellular enzymes. Results. Statistical analysis of the results showed a significantly higher phospholipase activity in the clinical isolates than in the environmental isolates. No significant difference about the phenoloxidase production between both groups was found. Regarding the 19 extracellular enzymes tested using the API ZYM commercial kit, acid phosphatase showed the highest enzymatic activity in both groups. Concerning the enzyme α-glucosidase, the clinical isolates presented a significantly higher positivity percentage than the environmental isolates. A hundred percent positivity in the enzyme leucine arylamidase production was observed in both groups, but the clinical isolates metabolized a significantly greater amount of substrate. Conclusions. The higher phospholipase production in the clinical isolates group confirms the possible role of this enzyme in the cryptococcosis pathogenesis. The extracellular activities of the enzymes acid phosphatase, α-glucosidase and leucine arylamidase, tested by means of the API ZYM commercial kit, appear to be very interesting. Many studies indicate that these enzymes are involved in the virulence of bacteria and parasites; our results suggest their possible role as virulence factors in Cryptococcus infections too (AU)

Antecedentes. Cryptococcus neoformans es una levadura encapsulada que produce infecciones oportunistas. Los factores de virulencia involucrados en la patogénesis de la criptococosis incluyen la existencia y el tamaño de la cápsula polisacarídica, la producción de melanina por medio de la enzima fenoloxidasa, el crecimiento a 37°C y la secreción de ciertas enzimas como proteinasa, fosfolipasa y ureasa. Existen otras enzimas que son secretadas por C. neoformans, pero su papel en la virulencia de este hongo aún no es conocido. Objetivos. Se investigó la producción de fosfolipasa tanto en aislamientos de C. neoformans obtenidos de pacientes como de aislamientos recuperados de deposiciones de aves, y se comparó el grado de producción con el de la síntesis de la enzima fenoloxidasa. Además, distingue las cepas mediante la definición de la actividad de 19 enzimas extracelulares diferentes. Métodos. Se estudiaron 205 aislamientos clínicos de C. neoformans y 32 ambientales. La producción de fenoloxidasa se determinó por el crecimiento de colonias de color marrón en medio de Staib. Para determinar la actividad fosfolipasa extracelular se utilizó el método semicuantitativo en placa con yema de huevo. Con el método comercial API ZYM se determinó la producción de otras 19 enzimas extracelulares. Resultados. El análisis estadístico de los resultados mostró una producción de fosfolipasa significativamente mayor entre los aislamientos clínicos en comparación con los ambientales. No se encontraron diferencias significativas entre ambos grupos en la producción de fenoloxidasa. En lo referente a las 19 enzimas extracelulares valoradas mediante el sistema API ZYM, la fosfatasa ácida mostró la mayor actividad en ambos grupos. Respecto a la enzima α-glucosidasa, nuevamente los aislamientos clínicos presentaron una actividad significativamente mayor. Todos los aislamientos de ambos grupos presentaron actividad leucina-arilamidasa, si bien los aislamientos clínicos procesaron mayor cantidad de sustrato de manera significativa. Conclusiones. La mayor producción de enzima fosfolipasa entre los aislamientos clínicos evidencia que esta enzima puede estar implicada en la patogénesis de la criptococosis. También es interesante la actividad extracelular de las enzimas fosfatasa ácida, α-glucosidasa y leucina-arilamidasa, valorada por medio del sistema comercial API ZYM. Diversos estudios apuntan a que estas enzimas están implicadas en la virulencia de bacterias y parásitos; nuestros resultados muestran también su posible implicación como factores de virulencia en las infecciones por Cryptococcus (AU)

Enzymatic characterization of clinical and environmental Cryptococcus neoformans strains isolated in Italy.

BACKGROUND: Cryptococcus neoformans is an encapsulated yeast causing mainly opportunistic infections. The virulence factors involved in cryptococcosis pathogenesis include the presence and the size of the polysaccharide capsule, the production of melanin by phenoloxidase, the growth at 37°C and the enzyme secretion like proteinase, phospholipase and urease. Many other enzymes are secreted by C. neoformans but their role in the fungus virulence is not yet known. AIMS: In order to investigate this topic, we compared the phospholipase production between strains from patients and from bird droppings, and we examined its relationship to phenoloxidase production. We further characterized the strains by determining the activity of 19 different extracellular enzymes. METHODS: Two hundred and five Italian C. neoformans clinical isolates and 32 environmental isolates were tested. Phenoloxidase production was determined by the development of brown colonies on Staib's agar. Extracellular phospholipase activity was performed using the semiquantitative egg-yolk plate method. API ZYM commercial kit was used to observe the production and the activity of 19 different extracellular enzymes. RESULTS: Statistical analysis of the results showed a significantly higher phospholipase activity in the clinical isolates than in the environmental isolates. No significant difference about the phenoloxidase production between both groups was found. Regarding the 19 extracellular enzymes tested using the API ZYM commercial kit, acid phosphatase showed the highest enzymatic activity in both groups. Concerning the enzyme α-glucosidase, the clinical isolates presented a significantly higher positivity percentage than the environmental isolates. A hundred percent positivity in the enzyme leucine arylamidase production was observed in both groups, but the clinical isolates metabolized a significantly greater amount of substrate. CONCLUSIONS: The higher phospholipase production in the clinical isolates group confirms the possible role of this enzyme in the cryptococcosis pathogenesis. The extracellular activities of the enzymes acid phosphatase, α-glucosidase and leucine arylamidase, tested by means of the API ZYM commercial kit, appear to be very interesting. Many studies indicate that these enzymes are involved in the virulence of bacteria and parasites; our results suggest their possible role as virulence factors in Cryptococcus infections too.

Vapour and Liquid-Phase Artemisia annua Essential Oil Activities against Several Clinical Strains of Candida.

Candida spp. are often the cause of infection in immune-compromised individuals. They are characterized by a strong resistance to antimicrobial drugs and disinfectants. The activity of Artemisia annua essential oil against Candida spp. was determined by vapour contact and microdilution assay. The oil was characterized by the presence of oxygenated monoterpenes (more than 75 % of the constituents), mainly represented by the irregular monoterpene artemisia ketone (ca. 22 %), and the widespread monoterpenes 1,8 cineole (ca. 19 %) and camphor (ca. 17 %). Other representative constituents were artemisia alcohol (5.9 %), α-pinene (5.7 %), and pinocarvone (3.0 %). Thujone, a typical toxic constituent of the Artemisia species, was not detected. The results are reported as minimum inhibitory concentration, minimum fungicidal concentration, and diameter of inhibition zone obtained by the vapour diffusion assay. We tested 10 clinical Candida strains, coming from both clinical samples and international collections. The results show that the antifungal activity of A. annua is influenced by the type of method adopted. The inhibitory action of the essential oil was, in fact, higher in the vapour than in the liquid phase. Our results show an average minimum inhibitory concentration in the liquid phase of 11.88 µL/mL, while in the vapour phase, the growth of all Candida strains tested at a concentration of 2.13 µL/cm(3) was inhibited. A strain of Candida glabrata was found to be less susceptible to the liquid medium than the vapour assay (50 µL/mL vs. 0.64 µL/cm(3), respectively). Candida albicans and Candida dubliniensis were the most susceptible to the vapour test, while Candida parapsilosis was the most resistant.

Fusariosis diseminada refractaria debida a Fusarium verticillioides en un paciente con leucemia mieloide aguda que experimentó recidiva después de un alotrasplante de células progenitoras hematopoyéticas: informe del caso y revisión de los estudios publicados / Refractory disseminated fusariosis by Fusarium verticillioides in a patient with acute myeloid leukaemia relapsed after allogeneic hematopoietic stem cell transplantation: A case report and literature review

Fundamento. Fusarium es uno de los principales patógenos fúngicos que provoca infecciones invasoras en pacientes portadores de neoplasias malignas hematopoyéticas. Las especies del género Fusarium implicadas habitualmente en las infecciones del ser humano son Fusarium solani, Fusarium oxysporum y Fusarium verticillioides. No obstante, la identificación es una tarea lenta y que consume mucho tiempo. Fusarium spp. es resistente in vitro a numerosos fármacos antimicóticos y el tratamiento de la fusariosis no está bien definido. Objetivos. Destacar las dificultades en la identificación de Fusarium spp. por los métodos convencionales y la necesidad de disponer de nuevas técnicas moleculares rápidas para obtener un diagnóstico más precoz y un tratamiento apropiado. Métodos. En un paciente portador de una leucemia mieloide aguda con neutropenia refractaria, que experimentó recidiva tras alotrasplante de células progenitoras hematopoyéticas se documentó una infección diseminada por Fusarium debida a Fusarium verticillioides. Resultados. A pesar de recibir un tratamiento combinado con anfotericina B y voriconazol liposómicos y de la sensibilidad in vitro de los preparados administrados, el paciente falleció. Sólo después de su muerte se obtuvo la identificación morfológica y molecular de Fusarium verticillioides. Conclusiones. El caso descrito en el presente informe destaca el desenlace desfavorable de las micosis invasivas debidas a Fusarium en pacientes con aplasia de médula ósea. La identificación de los miembros del género Fusarium sigue limitándose a laboratorios seleccionados y debe introducirse en el diagnóstico micológico sistemático. En el huésped inmunocomprometido el diagnóstico de fusariosis se relaciona directamente con el estado del paciente. Se describen los métodos diagnósticos y las opciones terapéuticas actuales(AU)

Background. Fusarium species are among the leading fungal pathogens to cause invasive mould infections in patients with hematopoietic malignancy. The Fusarium species most frequently involved in human infections are Fusarium solani, Fusarium oxysporum and Fusarium verticillioides. However, identification is a cumbersome and time-consuming task. Fusarium is resistant in vitro to many of the antifungal agents and the management of fusariosis is not well defined. Objectives. To emphasise the difficulty of identifying Fusarium spp. by conventional methods and the need of new rapid molecular tests to achieve earlier diagnosis and appropriate therapy. Methods. A disseminated Fusarium infection due to F. verticillioides was documented in a neutropenic refractory patient with acute myeloid leukaemia, relapsed after allogeneic hematopoietic stem cell transplantation. Results. The patient died despite liposomal amphotericin B and voriconazole combination and “in vitro” susceptibility of agents employed. Morphological and molecular identification of F. verticillioides was obtained only after the death of the patient. Conclusions. This case highlights the poor outcome of an invasive fungal disease caused by Fusarium in aplastic patients. Identification of members of Fusarium genus remains restricted to selected laboratories and should be introduced into routine mycological diagnostics. In immunocompromised patients, diagnosis of fusariosis is directly related to prompt diagnosis and to patient's status. Current diagnosis methods and therapeutic options are discussed(AU)

Refractory disseminated fusariosis by Fusarium verticillioides in a patient with acute myeloid leukaemia relapsed after allogeneic hematopoietic stem cell transplantation: a case report and literature review.

BACKGROUND: Fusarium species are among the leading fungal pathogens to cause invasive mould infections in patients with hematopoietic malignancy. The Fusarium species most frequently involved in human infections are Fusarium solani, Fusarium oxysporum and Fusarium verticillioides. However, identification is a cumbersome and time-consuming task. Fusarium is resistant in vitro to many of the antifungal agents and the management of fusariosis is not well defined. OBJECTIVES: To emphasise the difficulty of identifying Fusarium spp. by conventional methods and the need of new rapid molecular tests to achieve earlier diagnosis and appropriate therapy. METHODS: A disseminated Fusarium infection due to F. verticillioides was documented in a neutropenic refractory patient with acute myeloid leukaemia, relapsed after allogeneic hematopoietic stem cell transplantation. RESULTS: The patient died despite liposomal amphotericin B and voriconazole combination and "in vitro" susceptibility of agents employed. Morphological and molecular identification of F. verticillioides was obtained only after the death of the patient. CONCLUSIONS: This case highlights the poor outcome of an invasive fungal disease caused by Fusarium in aplastic patients. Identification of members of Fusarium genus remains restricted to selected laboratories and should be introduced into routine mycological diagnostics. In immunocompromised patients, diagnosis of fusariosis is directly related to prompt diagnosis and to patient's status. Current diagnosis methods and therapeutic options are discussed.

[Methicillin and vancomycin-resistant staphylococci and vancomycin-resistant enterococci in a hematology ward]. / Diffusione di stafilococchi meticillino e vancomicino-resistenti ed enterococchi vancomicino-resistenti in un reparto ospedaliero di ematologia.

Microbiological sampling of surfaces and air was performed in a hematology department, to monitor the presence of methicillin and vancomycin-resistant staphylococci and of vancomycin-resistant enterococci. The hematology department was divided in two areas, one of which with controlled access.Staphylococci were detected equally in the two areas while vancomycin-resistant enterococci were isolated only occasionally. The results show that the implementation of strict protocols for accessing hospital wards is not justified if effective cleaning and disinfection practices are not adopted.

Extracellular phospholipase activity of Malassezia strains isolated from individuals with and without dermatological disease.

BACKGROUND: The Malassezia genus includes mainly lipophilic yeasts belonging to the cutaneous microbiota of man and other mammals. Some Malassezia species have been associated with various dermatological diseases. The factors permitting the transformation of yeasts of the Malassezia genus from a commensal organism to a pathogenic agent are still little known but the production of various enzymes such as lipase, phospholipase and lipoxygenase could contribute to the pathogenic activity of these yeasts. AIMS: Here we have determined and compared the extracellular phospholipase activity of sixty human isolates of Malassezia so as to relate this feature to the species of Malassezia and to the origin (from dermatological diseases or not) of the strains examined. METHODS: Phospholipase production was determined using the semi-quantitative egg-yolk plate method. RESULTS AND CONCLUSIONS: Malassezia obtusa, Malassezia slooffiae, Malassezia globosa, Malassezia restricta had difficulty developing in the chosen culture medium so that it was not possible to measure phospholipasic activity. Malassezia pachydermatis showed the highest phospholipase activity. Twenty-nine Malassezia sympodialis strains produced phospholipase; the isolates from patients with pityriasis versicolor had significantly higher phospholipasic activity than those isolated from healthy individuals. This observation suggests that the phospholipasic activity of Malassezia may play a role in the onset of skin lesions, especially in the case of pityriasis versicolor.

Invasive pulmonary aspergillosis in neutropenic patients and the influence of hospital renovation.

To evaluate the effects of airborne Aspergillus contamination during and after the renovation work of a Florentine haematology unit, we conducted (November 2003-January 2005) a strict programme of environmental fungal surveillance. Air samples were taken from patients' rooms, along the corridors inside the wards, along the corridor between wards and outside the building. The concentration of Aspergillus fumigatus was high along the corridor between the two haematology wards (2.98 CFU m(-3)), lower in the non-neutropenic patients' rooms and outside the hospital building (1.53 and 1.42 CFU m(-3), respectively), very low in the neutropenic patients' rooms (0.09 CFU m(-3)). During this period, three proven cases (A. fumigatus), two probable ones and two possible cases of invasive pulmonary aspergillosis were documented in 97 patients with acute leukaemia (7%). The three cases of proven aspergillosis coincided with the period of renovation work and with the period in which we have found the maximum concentration of A. fumigatus along the corridor. These data suggest a possible relationship between environmental fungal contamination and the incidence of invasive aspergillosis, and underline the importance of environmental surveillance.

Isolation of Trichosporon in a hematology ward.

During mycologic monitoring of the air in a hematology ward, we found massive air contamination caused by Trichosporon asahii, both in the room where neutropenic patients were staying and the corridor immediately outside the room. This fungal species had never been isolated in previous samplings. The urine culture taken from one of the patients in this room, whose urinary catheter had been removed immediately prior to air sampling, resulted positive for T. asahii. Both macroscopic and microscopic morphologic observation was insufficient for confirming the hypothesis of a close relationship between the strains isolated from the patient, from the air in the room and corridor. Therefore, we used genomic typing with random amplified polymorphic DNA (RAPD). The five primers used, (GTG)(5), (GACA)(4), M13, OPE01, RC08, produced different patterns of polymerase chain reaction (PCR) products; the genomic profiles obtained with the same primer, however, resulted perfectly superimposable for all the strains. This result led us to conclude that the massive air contamination caused by T. asahii can have effectively been determined by the removal of the urinary catheter from the patient who presented an asymptomatic infection caused by this microorganism.

Two years of a fungal aerobiocontamination survey in a Florentine haematology ward.

The control of microbial air contamination in hospital wards has assumed great importance particularly for those hospital infections where an airborne infection route is hypothesised, such as aspergillosis. Invasive aspergillosis represents one of the most serious complications in immunocompromised patients. For some authors there is a direct association between this pathology and the concentrations of Aspergillus conidia in the air; in addition, reports of aspergillosis concurring during building construction have been frequent. In this study, two haematology wards were monitored for about 2 years in order to make both a qualitative and quantitative evaluation of fungal burden in the air, also in relation to major construction and demolition work taking place in the same building. Air samples were taken from the hospital rooms of neutropenic patients, in the corridors of their ward and outside the building. Total fungal concentration resulted higher outside (mean 572 Colony Forming Units/m3 of air), lower in the corridors (147 CFU/m3) and even lower in the rooms (50 CFU/m3). In all the samples we found the development of at least one fungal colony. Cladosporium was the most frequently isolated genus (57%), in contrast to Aspergillus spp. (2%). The average concentration of Cladosporium spp. was 24 CFU/m3 in the rooms, 78 CFU/m3 in the corridors and 318 CFU/m3 outside. The average concentration of Aspergillus spp. was 1.2 CFU/m3 in the rooms, 3.5 CFU/m3 in the corridors, 5.6 CFU/m3 outside. Our observations show low concentrations of Aspergillus fumigatus and A. flavus in all the environments examined and particularly in the rooms (0.09 and 0.10 CFU/m3 respectively); this observation could explain the absence of cases of invasive aspergillosis during the period of air monitoring in the two haematology wards.

Low vitamin D status, high bone turnover, and bone fractures in centenarians.

The oldest olds, including centenarians, are increasing worldwide and, in the near future, will represent a consistent part of the population. We have studied bone status and metabolism in 104 subjects over 98 yr of age to evaluate possible interventions able to avoid fragility fractures and disability. Ninety females and 14 males not affected by any acute disease were considered. After a complete clinical assessment, blood was drawn for evaluating bone turnover markers, and performance tests together with skeletal ultrasonography (either at the phalanges or at the heel) were performed. We found that 38 subjects had sustained a total of 55 fractures throughout their lives, and 75% of these were fragility fractures. Twenty-eight fractures occurred at the proximal femur, with 14 after the age of 94 yr. Serum 25-hydroxyvitamin D was undetectable in 99 of 104 centenarians. PTH and serum C-terminal fragment of collagen type I were elevated in 64 and 90% of centenarians, respectively, with a trend toward hypocalcemia. Bone alkaline phosphatase levels were close to the upper limit. Serum IL-6 was elevated in 81% of centenarians and was positively correlated with PTH and negatively correlated with serum calcium. Serum creatinine was not correlated with PTH. Bone ultrasonography showed that most centenarians had low values, and ultrasonographic parameters were correlated with resorption markers. We conclude that the extreme decades of life are characterized by a pathophysiological sequence of events linking vitamin D deficiency, low serum calcium, and secondary hyperparathyroidism with an increase in bone resorption and severe osteopenia. These data offer a rationale for the possible prevention of elevated bone turnover, bone loss, and consequently the reduction of osteoporotic fractures and fracture-induced disability in the oldest olds through the supplementation with calcium and vitamin D.

The MALVA (MAntova LongeVA) study: an investigation on people 98 years of age and over in a province of Northern Italy.

The MALVA (MAntova LongeVA) study aims at investigating the socio-demographic, clinical and genetic characteristics of all subjects over 98 years of age residing in the province of Mantova (Northern Italy). In this paper we present the study protocol and the main demographic results. Absolute number, prevalence ratio and female/male ratio of the subjects aged 98+ and of the centenarians in the Mantova province (370,645 inhabitants at 31st December 1997) were checked at the baseline of the study (31st March 1998) as well as in the two years preceding and following the study. A total of 117 subjects aged 98+ (including 39 centenarians) were traced at 31st March 1998; the prevalence ratio was 31.6 per 100000 (12.1 for centenarians), and the female/male ratio was 6.3 (6.5 for centenarians). The distribution of the oldest old according to places of birth and residence was non-homogeneous across the provincial territory. Seventy-seven subjects (66% of the identified subjects) were enrolled in the study and administered a protocol including an interview about socio-economic conditions, lifetime habits and pathological and pharmacological case histories, as well as medical examination, performance-based tests and blood sample collection. Data on the socio-demographic characteristics and the health status of very old people in the province of Mantova are discussed and compared to findings from previous studies on Italian centenarians.