Effect of harvesting strategy of second-cut orchardgrass silage on feed intake, digestion, and milk production in dairy cows.

We investigated the effects of regrowth interval and first-cut timing on the dietary characteristics of second-cut orchardgrass silage and feed intake and milk production in dairy cows fed second-cut orchardgrass silage. The second-cut grasses were harvested 7w after the first-cut at the early stage (E7w) or at the heading stage (H7w), or harvested 6w after the first-cut at the early stage (E6w) from orchardgrass sward, and then ensiled. We evaluated the effect of regrowth interval by comparing E7w and E6w, and the effect of first-cut timing by comparing E7w and H7w. Six multiparous Holstein cows were used in a replicated 3 × 3 Latin square design, with three dietary treatments: diets containing E7w, E6w, or H7w silage at 30% dietary dry matter. We observed that feeding E6w silage instead of E7w silage increased fiber digestibility, dry matter intake, and milk production; however, the first-cut timing (E7w vs. H7w) did not affect nutrient content and digestibility, feed intake, or lactation performance. These results show that harvesting at short regrowth intervals for second-cut orchardgrass can be an effective strategy for improving feed utilization and milk yield; however, the first-cut timing for second-cut orchardgrass has little impact.

Insulin-like growth factor-1 induces IRE1-XBP1-dependent endoplasmic reticulum biogenesis in bovine mammary epithelial cells.

Insulin-like growth factor-1 (IGF-1) plays a key role in proliferation and galactopoiesis in mammary epithelial cells (MEC), but its definitive functions on endoplasmic reticulum (ER) during protein synthesis remain unknown. The present study aimed to elucidate the effects of IGF-1 on ER biogenesis in MEC in vitro and examined the expression of ER biogenesis-associated genes in the mammary gland during early lactation. We treated mammary alveolar cells-large T antigen cells (immortalized bovine MEC line established via stable transfection with simian virus-40 large T-antigen) with IGF-1 and examined ER biogenesis using the fluorescence intensity of an ER tracker and quantitative real-time PCR. We found IGF-1 significantly increased ER tracker staining and upregulated mRNA levels of ER biogenesis-related genes, such as CHKA (choline kinase α), PCYT1A (choline-phosphate cytidylyltransferase A), and SURF4 (surfeit locus protein 4). We focused on unfolded protein response to explore molecular mechanisms by which IGF-1 induces ER biogenesis. We found IGF-1 significantly increased mRNA levels of the XBP1 splicing form (XBP1s). Based on western blot analysis, IGF-1 induced the expression of (inositol-requiring kinase 1 α) protein, upstream of XBP1s, and phosphorylated-IRE1α. The inhibition of IRE1 endoribonuclease activity with 4-methylumbelliferone 8-carbaldehyde (4µ8C) significantly suppressed the increase in ER tracker fluorescence and ER biogenesis-related gene expression induced by IGF-1. Also, IGF-1-induced XBP1s and ER biogenesis-associated gene expression was inhibited by rapamycin, an inhibitor of mTORC1 (mammalian target of rapamycin complex 1), indicating that IRE1-XBP1 activation by IGF-1 is mediated by mTORC1. Moreover, to clarify the expression of XBP1s and ER biogenesis-associated genes expression under normal physiological conditions, mammary gland tissue from biopsies of dairy cows during late gestation and lactation were analyzed. In vivo data highlighted the significant increases in the mRNA levels of XBP1s and ER biogenesis-related genes in mammary gland tissue immediately after calving through 6 wk of lactation. The mRNA levels of IGF1R (IGF-1 receptor) in mammary glands increased during 6 wk of lactation. Therefore, the present study indicated for the first time that IGF-1 induces ER biogenesis by activating the IRE1-XBP1 axis under the regulation of mTORC1 in bovine MEC line.

Effects of feeding early-harvested orchardgrass-perennial ryegrass mixed silage instead of heading stage harvested timothy silage on digestion and milk production in dairy cows.

We evaluated the effects of replacement of heading stage harvested timothy silage with early-harvested orchardgrass-perennial ryegrass mixed (OP) silage while maintaining or reducing concentrate input on dry matter intake (DMI), milk production, nutrient digestibility, and N balance in dairy cows. Nine multiparous Holstein cows were used in a replicated 3 × 3 Latin square design with three dietary treatments: TYL, a diet containing timothy silage where forage-to-concentrate ratio (FC) was 50:50; OPL, a diet containing OP silage where FC ratio was 50:50; and OPH, a diet containing OP silage where FC ratio was 60:40. We observed that an equal replacement of timothy with OP silage increased DMI, milk yield, milk protein production, and nutrient digestibility but decreased milk fat content (TYL versus OPL). We observed that replacing timothy with OP silage while reducing concentrate input increased milk fat and protein yield, nutrient digestibility, and feed efficiency and reduced urinary N loss with no effect on DMI or milk fat content (TYL versus OPH). These results show that replacing timothy with OP silage can be a good approach to improve milk production, feed efficiency, and N utilization and reduce concentrate input. However, milk fat depression should be considered when an equal substitution is performed.

Effects of rice conservation methods on lactation, blood metabolites, and rumen fermentation in dairy cows.

We investigated the effect of rice grain conservation methods on feed intake, milk production, blood metabolites, and rumen fermentation in dairy cows. Raw rice grain was dried before crushing (DRY), ensiled after crushing (ENS-A), or ensiled before crushing (ENS-B). Twelve multiparous Holstein cows were used in a replicated 3 × 3 Latin square design with three dietary treatments comprising ad libitum access to one of three total mixed rations (TMRs; containing DRY, ENS-A, or ENS-B at 17% of dietary dry matter) plus a standard allowance of 2.0 kg/day of dairy concentrates. The dietary treatments did not affect the feed intake, milk yield, or milk composition. The selected blood constituents were not influenced by the rice conservation method. The ruminal lactic acid and total volatile fatty acid (VFA) concentrations and the VFA proportion in the cows were not influenced by the rice conservation method. These results demonstrate that the rice grain conservation method has little impact on lactation performance when cows are fed a TMR containing 17% treated rice grain (dry matter basis).

Effect of different methods for conserving rice grain on in situ ruminal degradation and in vivo nutrient digestion and rumen fermentation in steers.

We investigated the effects of different rice conservation techniques on in situ ruminal degradation and in vivo nutrient digestibility and rumen fermentation in steers. Raw rice grain was dried before crushing (DRY), ensiled after crushing (ENS-A), or ensiled before crushing (ENS-B). Six ruminally cannulated steers were used in a replicated 3 × 3 Latin square design with three dietary treatments: diets containing DRY, ENS-A, or ENS-B at 36% of the dietary dry matter. The in situ rapidly degradable fraction and effective ruminal degradability were higher for ensiled rice than for DRY, and higher for ENS-A than for ENS-B. The ruminal pH was lower and the lactic acid and total volatile acid concentrations were higher for the steers fed ensiled rice than those fed the DRY diet, but a treatment effect was not observed in the comparison between ENS-A and ENS-B. The whole-tract digestibility of crude protein and ether extract was improved when the rice grain was ensiled, but there were no differences in nutrient digestibility between ensiling methods. These results show that ensiling treatment can be a strategy to improve the nutrient value of rice grain, but the ensiling method has little impact on in vivo digestion.

Anthocyanin and proanthocyanidin contents, antioxidant activity, and in situ degradability of black and red rice grains.

OBJECTIVE: An experiment was conducted to assess the antioxidant contents and activities of colored rice grains and to evaluate their nutritive characteristics in terms of chemical composition and in situ ruminal degradation. METHODS: Ten cultivars of colored rice grains (Oryza sativa L.) collected from several areas of Japan were studied, and control rice without pigment, maize, barley, and wheat grains were used as control grains. Their chemical compositions, pigment, polyphenol contents, total antioxidant capacity (TAC), and degradation characteristics were determined. RESULTS: The starch contents of the colored rice grains were in the range of 73.5% to 79.6%, similar to that of the control rice grain. The black and red rice grains contained anthocyanin (maximum: 5,045.6 µg/g) and proanthocyanidin (maximum: 3,060.6 µg/g) at high concentrations as their principal pigments, respectively. There were significantly (p<0.05) positive relationships among the pigment contents, polyphenol content, and TAC values in the colored and control rice grains, indicating that the increase in pigment contents also contributed to the increased polyphenol content and TAC values in the colored rice grains. The dry matter and starch degradation characteristics, as represented by c (fractional degradation rate of slowly degradable fraction) and by the effective degradability, of the colored rice grains and the control rice grain were ranked as follows among commonly used grains: wheat>barley ≥rice>maize. The colored rice grains also included the most-digestible starch, since their potential degradable fraction and actual degradability at 48 h incubation were almost 100%. CONCLUSION: Colored rice grains have high potential to be used as antioxidant sources in addition to starch sources in ruminants.

The unfolded protein response is involved in both differentiation and apoptosis of bovine mammary epithelial cells.

The unfolded protein response (UPR) describes a process involved in the homeostasis of endoplasmic reticulum (ER) and the differentiation of secretory cells. At present, the roles of UPR in the mammary gland tissue of dairy cattle are unknown. In the current study, we investigated the expression of UPR-related genes in Holstein cows during the developmental and lactating stages of the mammary gland tissue. To investigate the roles of UPR during the differentiation of mammary epithelial cells (MEC), we used MAC-T cells, a line of MEC. We collected samples of mammary gland tissue in dairy cows by biopsy during the late gestation and lactation periods and examined the expression of UPR-related genes by quantitative real-time PCR. Expression levels of the spliced X-box binding protein 1 (XBP1) and activating transcription factor 4 (ATF4) were found to be significantly higher in the mammary gland tissue 10 d before delivery compared with 40 d before delivery. An investigation before and after differentiation in MAC-T cells showed that the expression of ATF4 increased after differentiation of MEC, whereas that of the spliced XBP1 did not significantly change. Western blot analysis revealed that the differentiation-inducing stimulus induced phosphorylation of eukaryotic initiation factor 2α (eIF2α) but reduced that of protein kinase RNA-like endoplasmic reticulum kinase (PERK). Additionally, in ATF4-knockdown bovine MEC, differentiation was significantly suppressed; ATF4 knockdown also significantly suppressed the expression of glucocorticoid and insulin receptors. These results revealed that ER stress-independent ATF4 is involved in the cell differentiation mechanism, either directly or indirectly, via the control of the expression of lactogenic hormone receptors in bovine MEC. Immediately after parturition, gene expression levels of the spliced XBP1, ATF4, and C/EBP homologous protein (CHOP) markedly increased in mammary gland tissue, with a strong negative correlation between expression of CHOP and initial milk yield; CHOP is an apoptosis-related protein induced by ER stress. The above findings indicate that UPR is intrinsically associated with apoptosis of MEC, thus affecting the differentiation of these cells, as well as milk yield.

Effect of ensiling process of total mixed ration on fermentation profile, nutrient loss and in situ ruminal degradation characteristics of diet.

This experiment aimed to determine the changes in chemical composition, fermentation profile, in situ disappearance characteristics, and nutrient losses of ensiled total mixed ration (TMR) containing steam-flaked corn or brown rice (BR) during storage. TMRs for dairy cows, containing either steam-flaked corn or BR at 31.9% with 15.2% rye silage, 40.5% alfalfa silage, 5.0% beet pulp and 7.0% soybean meal, were prepared (dry matter (DM) basis). Each TMR was placed in a plastic drum silo, stored at 23°C in an air-conditioned room and sampled 0, 7, 14, 30, 90 and 210 days after preparation. In both grain sources, the fermentation products increased, while DM and starch storage losses increased and starch content greatly decreased during storage. The rapidly degradable fraction and effective ruminal degradability of DM, crude protein and starch increased during storage. These changes of dietary characteristics were large during 30 days of storage, but small after 90 days of storage. Replacing corn with BR led to increased fermentation products, starch loss and effective ruminal degradability of the ensiled TMR. These results indicate that the ensiling process of TMR changes the dietary characteristics and replacing corn with BR in TMR had a large impact on these dietary characteristics.

Lactation and digestion in dairy cows fed ensiled total mixed ration containing steam-flaked or ground rice grain.

The objective of this experiment was to investigate the effect of feeding ensiled total mixed ration (TMR) containing steam-flaked (SF) or ground brown rice (Oryza satira L.; BR) on feed intake, lactation performance, digestion, ruminal fermentation and nitrogen (N) utilization in dairy cows. Eight multiparous Holstein cows were used in a crossover design with two dietary treatments: diets containing either SF or fine ground (FG) BR at 24% of dietary dry matter. Dietary treatment did not affect dry matter intake or milk yield and composition. The whole-tract digestibility of organic matter and fiber decreased, and the digestibility of starch increased with the replacement of SF with FG in ensiled TMR, but these differences were small between diets. Crude protein digestibility was not different between diets. The processing method of BR did not affect ruminal pH, total volatile acid concentration, or volatile fatty acid proportion in cows. The N intake, milk N secretion, fecal and urinary N excretion and N retention were not influenced by dietary treatment. These results show that feeding ensiled TMR containing FG instead of SF reduces fiber digestibility but has little impact on lactation and N utilization when diets contained 24% on a dry matter basis.

Effect of time at pasture and herbage intake on profile of volatile organic compounds of dairy cow milk.

Volatile organic compounds (VOCs) in milk were investigated as quantitative markers of herbage intake (HI) at pasture. Eight Holstein cows were fed indoors with concentrate and conserved forages (grass silage, corn silage and hay) (NG), then were divided into three treatments according to the duration of access to pasture: 4 h (G4), 8 h (G8), and 20 h (G20) per day. The HIs were 4.3, 8.6, and 13.0 kg dry matter/day for the G4, G8 and G20 treatments, respectively. Milk from cows was sampled and analyzed VOCs by the steam distillation-extraction method and gas chromatography-mass spectrometry (GC-MS). From the intensity of the GC peak area, the levels of 1-phytene (3,7,11,15-tetramethyl-1-hexadecene) and 2-phytene (3,7,11,15-tetramethyl-2-hexadecene) were lowest in NG treatment and markedly increased with grazing time at pasture. With simple regression analysis on the HI to each diterpenoid, a strong correlation was found between the intensity of 1-phytene in the milk and the HI (r = 0.807, P < 0.001). 1-phytene content in milk could be useful as a quantitative marker of the HI of grazing cows.

Milk production, nutrient digestibility and nitrogen balance in lactating cows fed total mixed ration silages containing steam-flaked brown rice as substitute for steam-flaked corn, and wet food by-products.

The objective of this study was to evaluate the effect of substituting brown rice grain for corn grain in total mixed ration (TMR) silage containing food by-products on the milk production, whole-tract nutrient digestibility and nitrogen balance in dairy cows. Six multiparous Holstein cows were used in a crossover design with two dietary treatments: a diet containing 30.9% steam-flaked corn (corn TMR) or 30.9% steam-flaked brown rice (rice TMR) with wet soybean curd residue and wet soy sauce cake. Dietary treatment did not affect the dry matter intake, milk yield and compositions in dairy cows. The dry matter and starch digestibility were higher, and the neutral detergent fiber digestibility was lower for rice TMR than for corn TMR. The urinary nitrogen (N) excretion as a proportion of the N intake was lower for rice TMR than for corn TMR with no dietary effect on N secretion in milk and fecal N excretion. These results indicated that the replacement of corn with brown rice in TMR silage relatively reduced urinary N loss without adverse effects on feed intake and milk production, when food by-products such as soybean curd residue were included in the TMR silage as dietary crude protein sources.

Effect of replacing corn with brown rice in a total, mixed, ration silage on milk production, ruminal fermentation and nitrogen balance in lactating dairy cows.

Nine multiparous Holstein cows were used in a replicated 3 × 3 Latin square design to determine the effects of substituting corn grain with brown rice (BR) grain in total mixed ration (TMR) silage on milk yield, ruminal fermentation and nitrogen (N) balance. The TMR silages were made from the ensiling of TMR containing (dry matter basis) 50.1% forage in rice silage and corn silage combination, and 49.9% concentrate. The grain portion of the diets contained 31.2% steam-flaked corn, 31.2% steam-flaked BR or an equal mixture of corn and BR. Dietary treatments did not affect dry matter intake, milk yield and milk fat, protein and lactose yields. The ruminal pH and total volatile fatty acid concentrations were not affected by dietary treatment. The urinary N excretion decreased linearly (P < 0.01) in response to increased levels of BR, with no dietary effect on N intake, N secretion in milk and fecal N excretion. Our results indicate that steam-flaked BR is a suitable replacement for steam-flaked corn in dairy cow diets, and that it can be included in rations to a level of at least 31.2% of dry matter without adverse effects on milk production, when cows were fed rice silage and corn silage-based diets.

Digestibility, ruminal fermentation, nitrogen balance and methane production in Holstein steers fed diets containing soy sauce cake at 10 or 20%.

Two feeding trials were performed to evaluate the effect of soy sauce cake (SSC) on digestibility, ruminal fermentation and nitrogen (N) balance as well as methane production in Holstein steers. Six animals received a diet without or with the SSC at 10% (Experiment 1) and 20% (Experiment 2) in a 2 × 2 crossover design with a balance trial and respiratory exchange measurement. The SSC feeding showed a strongly inhibitory effect on methane production when its proportion was elevated up to 20%. The digestibility of nutrients with the 10% SSC treatment was similar to that of the control, whereas that for the 20% SSC treatment was significantly decreased in comparison with the control. The 20% SSC treatment also inhibited ruminal fermentation. A lower N partition to urine and a higher N partition to retention were observed in the animals receiving the 20% SSC treatment. These results suggested that SSC feeding at 20% suppressed methane production and changed the N balance; however, the feeding level of 20% caused deterioration in some productive aspects, such as nutrient digestibility and ruminal fermentation. In addition, a feeding level of SSC at 10% of the diet should be considered a reasonable level in cattle.

Factors relating to doctors' desire to change hospitals in Japan.

PURPOSE: This study seeks to investigate doctors' desire to change the hospital where they work to sustain higher quality care. DESIGN/METHODOLOGY/APPROACH: Self-administered questionnaires were sent to doctors in Aichi Prefecture, Japan. Data were analyzed using univariate and logistic regression analysis and recursive partitioning. FINDINGS: Factors related to doctors' desire to change hospitals, according to logistic regression, were interaction between working hours and satisfaction with the hospital, evaluation, local government hospitals versus private ones, small vs large hospitals, ophthalmology versus internal medicine, desire to continue working as a hospital doctor and age. Additionally, working hours were also found to be related, based on recursive partitioning. RESEARCH LIMITATIONS/IMPLICATIONS: The response rate was low and sampling bias was observed--therefore results need careful interpretation. Also, because this was a cross-sectional study, causal relationships could not be identified. Desire to change hospitals, but not actual behavior, was measured. PRACTICAL IMPLICATIONS: Efforts to prevent doctors from changing hospitals should include considering job satisfaction and workload, doctor evaluation methods, support for career progression and organizational management. ORIGINALITY/VALUE: As the hospital doctor shortage in rural areas becomes more serious, exploring doctors' desire to leave their current hospital is meaningful for Japanese hospital managers and hospitals worldwide aiming to provide sustainable and higher quality care.

Pathogenicity of Ochroconis gallopava isolated from hot springs in Japan and a review of published reports.

Four strains of Ochroconis gallopava from 3 out of 15 Japanese hot springs were isolated. Colonies of the hot spring isolates were uniformly floccose and dark olive green on the surface and dark brown on their reverse side on potato dextrose agar (PDA) plates, however, they became felty, flat, and brownish-black, and produced a reddish-brown pigment after several times of subculture at room temperature. Shapes and sizes of conidia of the four strains were individual, while the D1/D2 domain of the large subunit ribosomal RNA gene sequences showed 99.7% identity in the GenBank database. The DNA pattern of the hot spring isolates amplified by species specific loop mediated isothermal amplification method were as the same pattern as that of a clinical isolate. The minimum inhibitory concentrations of antifungal agents to O. gallopava isolated from the hot springs were ranged from 0.5 to 1 microg/ml in amphotericin B, 1 to 16 microg/ml in flucytosine, 0.125 to 0.25 microg/ml in itraconazole, 1 to 4 microg/ml in miconazole, 16 to 64 microg/ml in flconazole and 0.03 to 0.5 microg/ml in micafungin. The isolates had fatal outcome in experimentally infected mice intravenously with severe invasiveness to brains and kidneys. These findings suggested that O. gallopava habitats in hot springs could be one of sources for infection.

Resposta imune humoral na paracoccidioidomicose experimental em camundongos ddY / Humoral immune response in experimental ddY mice paracoccidioidomycosis

A paracoccidioidomicose (PCM) é uma micose sistêmica, restrita à América Latina, com maior incidência noBrasil. O camundongo ddY tem sido empregado como modelo murino de PCM e, no entanto, não há informaçõesa respeito da resposta imune desse animal frente à infecção. O presente estudo tem como objetivo avaliar aresposta imune humoral específica para o principal antígeno, gp43, do fungo Paracoccidioides brasiliensis,em camundongos ddY infectados com a cepa virulenta Pb 18. Foram realizadas análises da antigenemia ehistopatológico em vários órgãos e em diferentes tempos pós-infecção. Os resultados obtidos demonstraramaumento nos níveis de IgG anti-gp43 nos dias 14, 17, 21, 24, 28 e 56 pós-infecção e aumento no nível de gp43solúvel aos 28 dias pós-infecção. As células fúngicas foram detectadas em todos os órgãos analisados(cérebro, coração, pulmão, fígado, baço e rim) e em todos os períodos. As lesões granulomatosas tornaramsepredominantes 14 dias pós-infecção. Os resultados evidenciaram que o camundongo ddY produz respostaimune humoral frente ao principal antígeno de P. brasiliensis, apresentando-se elevado até 56 dias pósinfecção.A redução do nível de gp43 solúvel na fase crônica, supostamente devido ao início do controle dainfecção, requer estudos complementares adicionais.

Paracoccidioidomycosis (PCM) is a systemic mycosis, restrict to Latin America, with higher incidence inBrazil. ddY mice have been used as experimental PCM model, although there is no data regarding immuneresponse. The aim of the present study was evaluated specific humoral response against the main specificantigen of the fungal Paracoccidioides brasiliensis, the gp43, in ddY mice infected with virulent Pb 18.Antigenemia analysis and histophatological exam in several organs were performed in different time post-infection The results showed increased levels of anti-gp43 IgG on days 14, 17, 21, 24, 28 and 56 post-infectionand increased levels of soluble gp-43 on day 28 post-infection. The fungal cells were detected in all organsanalyzed (brain, heart, lung, liver, spleen and kidney) in all investigated periods. The granulomatous lesionsbecame predominant 14 days after infection. The results evidence that ddY mice produce humoral immuneresponse to main P. brasiliensis antigen, with high levels until 56 days after infection. Further studies areneeded to show that reduction of soluble gp43 in chronic phase correlates with infection control.

Molecular epidemiology of canine histoplasmosis in Japan.

A recent case of canine histoplasmosis, the first confirmed case of disseminated infection accompanied by carcinoma in Japan, was diagnosed by clinical characteristics, histopathological examination, chest radiographs, ocular fundoscopy and molecular biological data. The clinical manifestations were not limited to cutaneous symptoms but were referable to disseminated infection, similar to human autochthonous cases. The partial sequences of the internal transcribed spacer (ITS1/2) regions of the ribosomal DNA genes of this and other Japanese canine histoplasmosis strains were 99-100% identical to the sequence AB211551 derived from a human isolate in Thailand, and showed a close relationship to the sequences derived from Japanese autochthonous systemic and cutaneous human cases. The phylogenetic analysis of 97 sequences of the ITS1/2 region disclosed six genotypes. The genotypes derived from Japanese autochthonous human and dog cases belonged to the cluster consisting of Histoplasma capsulatum var. capsulatum and H. capsulatum var. farciminosum sequences, indicating that these varieties might cause not only cutaneous but also systemic histoplasmosis, regardless of their host species. The current status of the 3 varieties of Histoplasma capsulatum according to the host species remains a subject of further investigation.

Determinação da freqüência de Candida spp na cavidade oral de pacientes graves internados no Hospital das Clínicas - Unicamp, através de testes fenotípicos / Detection and frequency of Candida spp in oral cavity from serious patients hospitalized in Teaching Hospital of Unicamp, by phenotypic tests

Os métodos fenotípicos tradicionais e os testes comerciais utilizados na identificação das espécies do gênero Candida são úteis na averiguação da colonização oral, especialmente se utilizados em conjunto. O presente estudo foi realizado no Hospital de Clínicas da Unicamp com o objetivo de averiguar a diversidade das espécies de Candida na cavidade oral de 225 pacientes gravemente enfermos, internados nas unidades de Moléstias Infecciosas, Cirurgia do Trauma, Emergência Clínica e Transplante de Medula Óssea. A coleta do material ocorreu quinzenalmente no período compreendido entre novembro de 2000 e dezembro de 2002. Foram utilizados cinco testes: temperatura de crescimento, assimilação da xilose, CHROMagar Candida®, Candida Check® e ID 32®. Dentre os 349 “swabs” orais colhidos, obtiveram-se 141/349 (40,4%) culturas positivas de 126/225 (56,0%) pacientes. Os resultados obtidos com o teste de CHROMagar Candida® deixaram dúvidas quanto à diferenciação entre as amostras de C. albicans e C. dubliniensis, mas os demais testes confirmaram a ausência dessa segunda espécie. A espécie predominante detectada no presente estudo foi C. albicans, com 91 (64,5%), seguida de C. tropicalis com 17 (12,0%), C. glabrata com 9 (6,4%), C. parapsilosis com 7 (5,0%), C. krusei com 6 (4,2%), C. lusitaniae 5 (3,5%) e C. humicola, C. norvegiensis e C. kefyr com 2 (1,4%) cada. Os dados obtidos indicam que o conjunto de testes utilizados foi útil e suficientemente discriminatório na determinação das espécies colonizadoras da cavidade oral dos pacientes em estudo.

HIV-1 anti-retroviral drug effect on the C. albicans hyphal growth rate by a Bio-Cell Tracer system

Declining incidence of oropharyngeal candidosis and opportunistic infections over recent years can be attributed to the use of highly active anti-retroviral therapy (HAART). Infection with C. albicans generally involves adherence and colonization of superficial tissues. During this process, budding yeasts are able to transform to hyphae and penetrate into the deep tissue. Using the biocell tracer system, C. albicans hyphal growth was dynamically observed at the cellular level. Ritonavir was effective in the inhibition of hyphal growth with growth rate of 0.8 mum/min. This study showed the in vitro effect of HIV anti-retroviral drug on the growth rate of the C. albicans hyphae.

O declínio na incidência de candidose orofaríngea e infecções oportunistas associadas a infecção pelo HIV tem sido atribuído a introdução da terapia antiretroviral combinada (HAART). Infecção por C. albicans envolve aderência e colonização da mucosa superficial. Durante este processo leveduras são capazes de transformar-se na forma de hifas e penetrar nos tecidos mais profundos. Usando o sistema "Bio-Cell Tracer", o crescimento de hifas de C. albicans foi observado dinamicamente a nível celular. Ritonavir, inibidor de protease do HIV, foi efetivo na inibição do crescimento de hifas com media de 0.8 mim/min.O presente estudo demonstrou o efeito in vitro de um agente anti-retroviral HIV sobre o crescimento de hifas de C. albicans.

Candida dubliniensis in a Brazilian family with an HIV 1- infected child: identification, antifungal susceptibility, drug accumulation and sterol composition

This study investigated the prevalence of C. dubliniensis in a Brazilian family with an HIV - infected child. A total of 42 oral isolates were obtained from eight family members. The identification of C. dubliniensis was performed by polymerase chain reactions (PCR) using primers against a specific sequence of the C. dubliniensis cytochrome b gene. Only the HIV-infected child and his grandmother were colonized by C. dubliniensis. In this study C. dubliniensis isolated from the HIV-infected child exhibited high susceptibility for azoles tested with MICs of 0.125 and 0.5 æg/mL for voriconazole and fluconazole, respectively. Accumulation of [³H] fluconazole in C. dubliniensis isolated from the HIV-infected child was slightly reduced in comparison to the reference susceptible strain. C. dubliniensis isolates had significantly lower ergosterol levels in comparison to C. albicans reference strains.

O presente estudo investigou a prevalência de C. dubliniensis em uma família brasileira com uma criança infectada pelo vírus HIV. Um total de 42 isolados orais foram obtidos de 8 membros da família. A identificação de C. dubliniensis foi realizada por polymerase chain reactions (PCR) usando primers contra a sequência específica para o gene C. dubliniensis cytochrome b. Apenas a criança infectada pelo vírus HIV e a avó estavam colonizados por C. dubliniensis. Neste estudo C. dubliniensis isolado da criança infectada pelo vírus HIV exibiu alta susceptibilidade para azoles com concentração mínima inibitória de 0.125 and 0.5 æg/mL para voriconazole and fluconazole respectivamente. Acúmulo de [³H] fluconazol intra-celular foi ligeiramente reduzido em C. dubliniensis isolado da criança infectada pelo vírus HIV em comparação com a cepa referência sensível ao fluconazole. Isolados de C. dubliniensis neste estudo apresentaram níveis significantemente reduzidos de ergosterol da membrane celular em comparação com C. albicans.