Extraction kinetics of Brazilian green propolis and chemical characterization of its volatiles.

Propolis is a natural resinous product produced by Apis mellifera bees from the exudates of various plants. The color of propolis (green) is a consequence of its botanical origin, as bees collect young tissues and leaves of Baccaris dracunculifolia. This study evaluated the chemical composition and extraction kinetics of essential oils obtained from Brazilian green propolis by hydrodistillation. Hydrodistillation was performed for 360 min and analyzed at different times (30, 60, 120, 240, and 360 min), allowing the calculation of the accumulated content (% w/w) and the identification of the essential oil chemical profile. The GC/FID and GC/MS analysis led to the annotation of 60 compounds with estragole (13.30%), benzyl propanoate (14.59%), and (E)-nerolidol (13.57%) as the main compounds. The optimum conditions for extraction of phenylpropanoids (PP), hydrocarbons (HD), monoterpenes (MT), and oxygenated monoterpenes (OMT) are between 30 and 120 min. In comparison, sesquiterpenes (ST) and oxygenated sesquiterpenes (OST) are extracted more efficiently between 240 and 360 min. The optimal extraction speed determination is essential for industrial-scale processing to obtain components such as sesquiterpenes, which have a high economic value in the cosmetic/perfumery and pharmaceutical industries.

Perceptions about Aging and Ageism from 14 Cross-sectional Cohorts of Undergraduate Dental Students.

BACKGROUND: Although positive and negative views of aging and older adults exist, how undergraduate dental students imagine their lives to be as they grow older remains to be fully explored. This study aimed at determining the self-perceived views of being 65, 75, or 85 y of age, as expressed by undergraduate dental students at the University of British Columbia (UBC), Vancouver, Canada. METHODS: A 14-y cross-sectional study design was utilized involving third-year undergraduate dental students at UBC's Faculty of Dentistry. Brief individual essays (150 words) encompassing students' self-perceived views were gathered as part of a dental geriatric course from 2009 to 2022; however, essays were not mandatory. Saldaña's inductive coding and thematic analysis of textual data were used. Themes and categories of information were identified and matched with their excerpts while aiming for data saturation. RESULTS: Over the 14-y period, 657 students were enrolled in UBC's undergraduate dental geriatric course, and 561 essays were collected. Inductive coding and thematic analysis identified 5 main themes and 11 categories. While themes included "oral health, general health, and the mind" and "me, myself, and familial relationships," the categories focused on "(un)able bodies" and "general health." Positive views about the aging process were shared, while less optimistic ideas-and even ageism-were apparent when students saw themselves as not employable or living in isolation. Positive and negative views were not bound by the students' academic year but might have been influenced by the COVID-19 pandemic. CONCLUSION: Although the number of older adults already surpasses the number of children in many countries, ageism appears to have permeated through students' views of 3 older ages. More positive yet realistic views of growing older were also shared. Follow-up studies are needed to explore the impact of dental education on decreasing ageism. KNOWLEDGE TRANSFER STATEMENT: As the proportion of older adults in the global population steadily grows, it is important to educate heath care providers about normal and pathologic aging to avoid ageism-stereotypes, prejudices, and discrimination against older adults. This cross-sectional study involved 14 cohorts of undergraduate dental students exploring their self-perceived views of growing older. Although positive and negative views of aging were shared, dental education must focus on decreasing ageism.

Evaluation of sperm and hormonal assessments in Wagyu, Nellore, and Angus bulls.

Wagyu bulls are known to have a highly exacerbated libido, as shown by the intense sexual interest of young calves. Therefore we believe that Wagyu male animals have specialized Sertoli and Leydig cells that are directly involved with the sexual precocity in this breed as mature bulls have a small scrotal circumference. This study aimed to evaluate whether there were differences in the hormone and sperm characteristics of Wagyu bulls compared with the same characteristics of subspecies Bos indicus and Bos taurus sires. Frozen-thawed semen from Wagyu, Nellore, and Angus sires were analyzed for sperm kinetics (computer-assisted sperm analysis), plasma membrane integrity, chromatin integrity, acrosome status, mitochondrial activity, lipid peroxidation and hormone [luteinizing hormone (LH) and testosterone] serum concentration. The results showed that Wagyu had lower total motility and an increased number of sperm with no motility when compared with Nellore and Angus bulls. Wagyu breed did not differ from those breeds when considering plasma and acrosome membranes integrity, mitochondrial potential, chromatin resistance, sperm lipid peroxidation or hormone (LH and testosterone) concentrations. We concluded that Wagyu sires had lower total motility when compared with Nellore and Angus bulls. Wagyu breed did not differ from these breeds when considering plasma and acrosome membranes integrity, mitochondrial potential, chromatin resistance, sperm lipid peroxidation, or hormone (LH and testosterone) concentrations.

Temporal evaluation of follicular dynamics and endocrine patterns of Holstein (Bos taurus), Gir (Bos indicus), and Murrah (Bubalus bubalis) heifers kept under the same nutritional, management and environmental conditions.

The objective of the study was to simultaneously compare ovarian follicular dynamics and endocrine parameters of taurine (Holstein; n = 14), zebuine (Gir; n = 5), and bubaline (Murrah; n = 15) heifers kept under the same environmental, nutritional and management conditions. Heifers were synchronized with two PGF treatments 14 days apart. Ovaries of cyclic heifers were scanned daily during two consecutive ovulations and blood samples were collected every 24 h from each animal. No significant difference was found for length of interovulatory interval, however, zebuine heifers presented a greater number of follicular waves, number of antral follicles on day of ovulation, and higher insulin concentration than the other two breeds. Taurine heifers had highest maximal diameter of first wave dominant and ovulatory follicles and CL volume. Taurine and bubaline heifer's dominant follicle of first wave had longer static and regression phases than zebuine heifers. Bubaline heifers presented overall lowest progesterone concentrations and CL volume, but higher IGF1 levels. No difference was observed between taurine and zebuine heifers regarding IGF1 concentration. Despite higher CL volume found in taurine heifers, no difference in mean progesterone concentration was observed between them and zebuine heifers. Insulin and IGF1 concentrations were greater in follicular phase than in luteal phase when breed was not evaluated. After evaluating the three breeds simultaneously, at the same nutritional and management status it is possible to conclude that each genetic group has a specific follicular development and endocrinology of the estrous cycle.

Nutritional Status and Adverse Outcomes in Older Depressed Inpatients: A Prospective Study.

OBJECTIVES: Significant weight loss and/or loss of appetite is a criterion of a depressive episode. While malnutrition is associated with many adverse health outcomes, the impact of malnutrition in late-life depression has hardly been examined. The present study aims to (1) evaluate the prevalence of malnutrition in depressed older inpatients, and (2) whether and which indices of malnutrition predict adverse health outcomes in late-life depression. DESIGN: A prospective study at 6 months follow-up. SETTING: A University-based psychiatric hospital. PARTICIPANTS: 105 older adults (psychiatric inpatients suffering from unipolar MDD). MEASUREMENTS: Participants were evaluated according the Mini Nutritional Assessment (MNA) and anthropometric measures to assess their nutritional status. Multiple regression analyses were used to evaluate the association between the MNA score as well as anthropometric measures with either falls or rehospitalization for any reason. RESULTS: Based on the MNA score, 78 (74.3%) patients were at risk of malnutrition and 13 (12.4%) actually presented malnutrition. Malnutrition was associated with a higher age, frailty, lower body mass index, and smaller calf circumference. During follow-up, 21 (20%) patients fell, 27 (25.7%) were rehospitalized, and 3 died (2.9%). The MNA score was associated with adverse health outcomes, but a low calf circumference predicted falling (OR 4.93 [95% CI: 1.42-17.2], p=.012) and a higher calf circumference rehospitalization (OR 1.17 [95% CI: 1.01-1.35], p=.032). CONCLUSION: Malnutrition is prevalent in older depressed inpatients. In contrast to subjective proxies for malnutrition, which are common in depression, only objective measures of malnutrition predict adverse health outcomes such as falls and rehospitalization.

A new method for measuring pen pressure in forensic handwriting analysis - a proof of concept study.

This work describes a new method for the analysis of handwritten documents through a system composed of a pre-selector optical analyser equipped with light sources of different wavelengths coupled with bandpass filters combined with an optical coherence tomography (OCT) instrument. The optical analyser identifies regions with different pen pressures on the paper using specific wavelengths from ultraviolet (UV) to infrared (IR) and bandpass filters. Then the selected regions are analysed with a coherence tomography analyser to measure the depth of grooves and capture three-dimensional images. With this methodology, it is possible to identify similarities, or differences, between the pieces of evidence under investigation, increasing the possibility of correct attribution concerning the authorship of the signature and we also showed that this feature is independent of the paper substrate. In this work, a new strategy will be presented to categorize and quantify pen pressure in order to aid a better response for a forensic examiner. Thereby, from the observed areas that display higher pressures (more significant grooves), it is possible to determine the authorship of the signature.

Is Dexamethasone superior to Ketorolac in reducing pain, swelling and trismus following mandibular third molar removal? A split mouth triple-blind randomized clinical trial.

BACKGROUND: The preemptive use of anti-inflammatory drugs, such as corticosteroids and NSAIDs, has the potential to reduce pain, swelling and trismus following oral surgery. The aim of this study was to compare the efficacy of dexamethasone and ketorolac tromethamine in reducing pain, swelling and trismus after mandibular third molar removal. MATERIAL AND METHODS: The researches implemented a triple-blind, randomized clinical trial. The study was conducted with ASA I individuals aging between 18 and 35 years, which were randomized and submitted to two interventions, one with 8mg dexamethasone and the other with 20mg ketorolac tromethamine given 1h before the procedure. The primary predictor variable was the use of dexamethasone or ketorolac. The primary outcome variable was the postoperative pain level, measured with a Visual Analogue Scale. The secondary outcome variables were the amount of rescue analgesic consumed, swelling and trismus. Repeated-measures ANOVA and t-test for paired samples were used to compare the means. Significance was set at p < 0.05. RESULTS: Fifty individuals were randomized and allocated to intervention, and the sample was composed of 40 subjects who completed the study (27 female and 13 male). Dexamethasone, when compared to ketorolac tromethamine, showed a significantly higher reduction in pain level at 8h, 16h, 24h, 32h, 40h and 72h, in swelling and trismus at 24h, 48h, 72h and 7 days and in total number of rescue analgesics taken up to 72h postoperative (p < 0.05). CONCLUSIONS: The clinical performance of dexamethasone in controlling pain, swelling and trismus after mandibular third molar removal was superior to ketorolac tromethamine's.

Bacterial versus non-bacterial infections: a methodology to support use-case-driven product development of diagnostics.

Acute febrile illness (AFI) is one of the most common reasons for seeking medical care in low-income and middle-income countries. Bacterial infections account for a relatively small proportion of AFIs; however, in the absence of a simple diagnostic test to guide clinical decisions, healthcare professionals often presume that a non-malarial febrile illness is bacterial in origin, potentially resulting in inappropriate antibiotic use. An accurate differential diagnostic tool for AFIs is thus essential, to both limit antibiotic use to bacterial infections and address the antimicrobial resistance crisis that is emerging globally, without resorting to multiple or complex pathogen-specific assays. The Biomarker for Fever-Diagnostic (BFF-Dx) study is one of the largest fever biomarker studies ever undertaken. We collected samples and classified disease aetiology in more than 1900 individuals, distributed among enrolment centres in three countries on two continents. Identical protocols were followed at each study site, and the same analyses were conducted in each setting, enabling like-with-like comparisons to be made among the large sample set generated. The BFF-Dx methodology can act as a model for other researchers, facilitating wider utility of the work in the future. The established sample collection is now accessible to researchers and companies and will facilitate the development of future fever-related diagnostic tests. Here, we outline the methodology used to determine the sample populations and to differentiate bacterial versus non-bacterial AFIs. Future publications will set out in more detail the study's demographics, the causes of fever identified and the performance of selected biomarkers.

Records of breeding in Wilson's Plover Charadrius wilsonia with new localities for Brazil / Registros da reprodução de Charadrius wilsonia, com novas ocorrências para o Brasil

Abstract Wilson's Plover, Charadrius wilsonia, is widely distributed in coastal areas of the Americas. This report presents the first record of breeding in this species on Coroa do Avião Island, on the coast of Pernambuco, and in the estuary of the Cardoso and Camurupim rivers, on the coast of Piauí, in northeastern Brazil, extending the known area of reproduction of this species in this region. One breeding pair was observed on October 29th 2014 on Coroa do Avião Island, and a second pair was recorded on April 6th 2016 in the Cardoso/Camurupim estuary on the Piauí coast. Both the male and the female contributed to the incubation of the eggs. The nest on Coroa do Avião island was camouflaged by the local vegetation, but despite this, the eggs were attacked by a predator. Possible predators observed on the island included Caracara plancus and domestic cats and dogs.

Resumo Charadrius wilsonia (Wilson's Plover) está amplamente distribuído pela costa das Américas. Este é o primeiro registro de reprodução no litoral de Pernambuco, Coroa do Avião, e no estuário dos rios Cardoso e Camurupim, litoral do Piauí, Brasil, ampliando a área de reprodução no Nordeste do Brasil. Foi encontrado um casal em período reprodutivo em 23 de outubro de 2014 na ilha Coroa do Avião, litoral de Pernambuco e outro registro em 06 de abril de 2016 nos estuários Cardoso e Camurupim, litoral do Piauí. Foi observado que o macho e a fêmea contribuem na incubação dos ovos. A vegetação na ilha contribui para a camuflagem do ninho, bem como na proteção dos ovos pelos predadores. Apesar da proteção ocorreu a predação do ninho na ilha Coroa do Avião. Alguns possíveis predadores foram registrados na ilha, como Caracara plancus que diariamente frequentam a área e animais domésticos como cães e gatos.

Records of breeding in Wilson's Plover Charadrius wilsonia with new localities for Brazil.

Wilson's Plover, Charadrius wilsonia, is widely distributed in coastal areas of the Americas. This report presents the first record of breeding in this species on Coroa do Avião Island, on the coast of Pernambuco, and in the estuary of the Cardoso and Camurupim rivers, on the coast of Piauí, in northeastern Brazil, extending the known area of reproduction of this species in this region. One breeding pair was observed on October 29th 2014 on Coroa do Avião Island, and a second pair was recorded on April 6th 2016 in the Cardoso/Camurupim estuary on the Piauí coast. Both the male and the female contributed to the incubation of the eggs. The nest on Coroa do Avião island was camouflaged by the local vegetation, but despite this, the eggs were attacked by a predator. Possible predators observed on the island included Caracara plancus and domestic cats and dogs.

The Relationship between Physical Frailty and Mild Cognitive Impairment in the Elderly: A Systematic Review.

INTRODUCTION: Physical frailty (PF) appears to be associated with low cognitive performance and mild cognitive impairment (MCI). This review evaluated and synthesized the evidence of studies investigating the association between PF and MCI, the prevalence of both conditions and the rate of conversion of healthy older adult to one of them during the follow-up. METHODS: A systematic review was performed according to the PRISMA recommendations in the Pubmed, SciELO and LILACS databases. Five studies were eligible according to inclusion and exclusion criteria. RESULTS: Regarding the study design, cross-sectional studies prevailed. Most studies showed a positive association between PF and MCI. Moreover, PF seems to predict a worse cognitive trajectory among participants with MCI and it is associated to a higher risk of developing MCI. CONCLUSION: Our findings suggest a significant association between PF and MCI. Further longitudinal studies are needed to better explore causality.

Ankle torque steadiness and gait speed after a single session of robot therapy in individuals with chronic hemiparesis: a pilot study.

Background: Anklebot therapy has proven to be effective in improving hemiparetic gait. However, neither ankle torque steadiness nor the relationship between changes in force control and functional tasks after therapy with Anklebot were described.Objective: To assess whether a single session of robotic therapy promotes short-term ankle adaptations that influence ankle torque steadiness and walking speed in individuals with chronic hemiparesis.Methods: A sample of participants who had residual hemiparesis deficits (hemiparesis group; n = 13) and age- and sex-matched healthy control participants (control group; n = 13). For sample characterization, balance, mobility, sensorimotor impairment, and daily living activities performance were measured.Results: Differences in functional tests were identified only when the control and hemiparesis groups (F = 29.1; p = .001) were compared during the 10-metre Walking Test. Regarding the pre- and post-robotic assistance session, no significant difference was observed for any comparison (p > .05), except for the steadiness test, as demonstrated by the standard deviation (F = 7.10; p = .01) and coefficient of variation (F = 6.20; p = .02). The hemiparesis group showed better torque steadiness during dorsiflexion post-robotic assistance therapy (p ≥ 0.02) when compared with pre-robotic assessment. Correlations were identified between steadiness and walking speed variables.Conclusion:  People with chronic hemiparesis presented short-term performance gains in torque steadiness, especially during dorsiflexion, after a single robotic therapy session. The robotic therapy did not influence the walking speed, although low to moderate correlations between torque steadiness variables and walking speed were observed.

New evidence for refinement of anesthetic choice in procedures preceding the forced swimming test and the elevated plus-maze.

Previous studies indicated that some general anesthetics induce long-term antidepressant and/or anxiolytic-like effects. This raises the concern about the use of anesthesia in surgeries that precede psychopharmacological tests, since it may be a potential bias on results depending on the experimental design used. Thus, we evaluated whether general anesthetics used in surgeries preceding psychopharmacological tests would affect rats behavior in tests predictive of antidepressant or anxiolytic-like effects. We tested if a single exposure to sub-anesthetic or anesthetic doses of tribromoethanol, chloral hydrate, thiopental or isoflurane would change rats behavior in the forced swimming test (FST) or in the elevated plus-maze (EPM) test, at 2 h or 7 days after their administration. We also evaluated whether prior anesthesia would interfere in the detection of the antidepressant-like effect of imipramine or the anxiolytic-like effect of diazepam. Previous anesthesia with the aforementioned anesthetics did not change rats behaviors in FST per se nor it changed the antidepressant-like effect induced by imipramine treatment. Rats previously anesthetized with tribromoethanol or chloral hydrate exhibited, respectively, anxiogenic-like and anxiolytic-like behaviors in the EPM. Prior anesthesia with thiopental or isoflurane did not produce any per se effect in rats behaviors in the EPM nor disturbed the anxiolytic-like effect of diazepam. Our results suggest that, in our experimental conditions, tribromoethanol and chloral hydrate are improper anesthetics for surgeries that precede behavioral analysis in the EPM. Isoflurane or thiopental may be suitable for anesthesia before evaluation in the EPM or in the FST.

GloPID-R report on Chikungunya, O'nyong-nyong and Mayaro virus, part I: Biological diagnostics.

The GloPID-R (Global Research Collaboration for Infectious Disease Preparedness) Chikungunya (CHIKV), O'nyong-nyong (ONNV) and Mayaro virus (MAYV) Working Group is investigating the natural history, epidemiology and medical management of infection by these viruses, to identify knowledge gaps and to propose recommendations for direct future investigations and rectification measures. Here, we present the first report dedicated to diagnostic aspects of CHIKV, ONNV and MAYV. Regarding diagnosis of the disease at the acute phase, molecular assays previously described for the three viruses require further evaluation, standardized protocols and the availability of international standards representing the genetic diversity of the viruses. Detection of specific IgM would benefit from further investigations to clarify the extent of cross-reactivity among the three viruses, the sensitivity of the assays, and the possible interfering role of cryoglobulinaemia. Implementation of reference panels and external quality assessments for both molecular and serological assays is necessary. Regarding sero-epidemiological studies, there is no reported high-throughput assay that can distinguish among these different viruses in areas of potential co-circulation. New specific tools and/or improved standardized protocols are needed to enable large-scale epidemiological studies of public health relevance to be performed. Considering the high risk of future CHIKV, MAYV and ONNV outbreaks, the Working Group recommends that a major investigation should be initiated to fill the existing diagnostic gaps.

A feasibility study to explore the governance processes required for linkage between dental epidemiological, and birth cohort, data in the U.K.

Birth cohort initiatives, such as 'Born in Bradford', provide a unique opportunity to study the influence of socio-economic and environmental factors acting in pregnancy, birth and infancy on the development of dental caries in later life. This paper describes a feasibility study which established the processes required, and outcomes of, successful linkage of oral health data collected by the 2013 three-year-old national dental epidemiology survey with the Born in Bradford birth cohort database. The necessary processes included achieving research permissions and ethical approval; creation of a data sharing agreement; ensuring data security and encrypted data transfer. With regard to the outcomes, a robust a priori statistical plan was developed. 152 three-year-old children were examined for the 2013 dental epidemiology survey in Bradford, and of those, 69 parents consented to data linkage believing that their child was part of the Born in Bradford cohort. However, only 36 of these 69 children were participating in the cohort. Of these, six children had obvious dentinal caries experience (dmft ⟩0). There was insufficient power with such small numbers, to examine the association between birthweight and dental caries at the age of three-years-old. Key learning points from this feasibility study have informed the design of a larger study to link the 2014/5 five-year-old dental epidemiology surveys with the Born in Bradford cohort. This paper reveals the important methodological considerations for future data linkages between routine health data and research data.

A critical review of protocols for conventional microwave oven use for denture disinfection.

INTRODUCTION: The lack of proper denture hygiene may cause denture stomatitis and be detrimental to older adults' health. The cleaning of complete dentures should be quick, efficient, and easy to perform, although it might not guarantee disinfection. The use of a microwave oven has been suggested for aiding in the disinfection of complete dentures, but lacks a gold standard protocol. OBJECTIVES: To critically review the literature on protocols for complete denture disinfection using conventional microwave ovens. METHODS: A comprehensive literature search through PubMed Central, Cochrane Database of Systematic Reviews, and Ovid MEDLINE (R) In-Process focused on publications in English dealing with microwave therapy for denture disinfection, and on the protocols used. RESULTS: A total of 266 articles with the full-text available were found; 31 were included in this manuscript after 236 were excluded. The protocols for microwave oven use for disinfection of complete dentures varied in terms of oven potency, time used for microwaving, and solution in which the complete dentures were immersed. CONCLUSIONS: There is still no standardized protocol for microwave oven use for denture disinfection. Although underutilized in residential care, daily denture hygiene seems to still be the optimal method for controlling fungal infections and denture stomatitis.

Epidemiology of acute febrile illness in Latin America.

BACKGROUND: The causes of acute febrile illness (AFI) in Latin America are diverse and their complexity increases as the proportion of fever due to malaria decreases, as malaria control measures and new pathogens emerge in the region. In this context, it is important to shed light on the gaps in the epidemiological characteristics and the geographic range for many AFI aetiologies. OBJECTIVES: To review studies on community-acquired fever aetiology other than malaria in Latin America, and to highlight knowledge gaps and challenges needing further investigation. SOURCES: PubMed from 2012 to April 2018. CONTENT: We found 17 eligible studies describing 13 539 patients. The median number of pathogens tested per individual was 3.5 (range 2-17). A causative pathogen could be determined for 6661 (49.2%) individuals. The most frequently reported pathogen during the study periods was dengue virus (DENV) (14 studies), followed by chikungunya virus (nine studies) and Zika virus (seven studies). Among the studies reporting concurrent infections, 296 individuals (2.2%) were found to have co-infections. In-hospital mortality was reported in eight (47%) studies, ranging between 0% and 18%. IMPLICATIONS: DENV fever is the febrile illness most frequently reported, reflecting its importance, while chikungunya and zika viruses present increasing trends since their emergence in the region. Studies with systematic and harmonized approaches for detection of multiple pathogens are needed and would probably reveal a higher burden of neglected pathogens such as Rickettsia spp. and arenaviruses. The lack of point-of-care tests and harmonized approach limits the care provided by health professionals and the efficacy of surveillance for AFI in the region.

Effect of bovine sperm chromatin integrity evaluated using three different methods on in vitro fertility.

In vitro fertility potential of individual bulls is still relatively uncharacterized. Classical sperm analysis does not include the evaluation of all sperm characteristics and thus, some cell compartments could be neglected. In humans, sperm DNA integrity has already proven to have major influence in embryo development and assisted reproduction techniques successfully. In bovine, some studies already correlated chromatin integrity with field fertility. However, none of those have attempted to relate DNA assessment approaches such as chromatin deficiency (CMA3), chromatin stability (SCSA; AO+) and DNA fragmentation (COMET assay) to predict in vitro bull fertility. To this purpose, we selected bulls with high and low in vitro fertility (n = 6/group), based on embryo development rate (blastocyst/cleavage rate). We then performed CMA3, SCSA test and COMET assay to verify if the difference of in vitro fertility may be related to DNA alterations evaluated by these assays. For the three tests performed, our results showed only differences in the percentage of cells with chromatin deficiency (CMA3+; high: 0.19 ± 0.03 vs low: 0.04 ± 0.04; p = 0.03). No difference for chromatin stability and any of COMET assay categories (grade I to grade IV) was observed between high and low in vitro fertility bulls. A positive correlation between AO + cells and grade IV cells was found. Despite the difference between groups in CMA3 analysis, our results suggest that protamine deficiency in bovine spermatozoa may not have a strong biological impact to explain the difference of in vitro fertility between the bulls used in this study.