Host range of Phytophthora spp. from berry crops in Huelva, Spain.

Crop declines have been observed in raspberry and blueberry farms in the southwest region of Spain, which is the most important berry-producing area in the country. This study aimed to identify and characterize the pathogens associated with these diseases using molecular and morphological methods. Additionally, pathogenicity tests were performed on different raspberry, blueberry, and strawberry cultivars to determine possible susceptible hosts in the area. An isolate of P. cactorum was obtained from a symptomatic strawberry plant, an isolate of P. cinnamomi was obtained from a symptomatic blueberry plant, and isolates identified as P. rosacearum, P. rubi and a previously unknow speciesrecently named as P. sp. balkanensis were recovered from symptomatic raspberry plants. Results from the pathogenicity tests reported, for the first time, P. rubi causing root rot and wilting complex (RRWC) in Spanish raspberry crops. Additionally, P. cinnamomi was found to affect highbush blueberry production in Spain. Thus, this study provides valuable insights into the identification and characterization of Phytophthora spp. associated with the decline of blueberry and raspberry crops in Huelva. It also provides essential recommendations regarding the potential risks associated with the use of other types of berries as rotational crops and emphasizes the necessity for effective management strategies to mitigate crop losses. This is particularly critical given the limited soil disinfection alternatives available in Spain.

Expression of IZUMO1 and JUNO in the gonads of domestic cats (Felis catus).

Because of the time-consuming nature of surgical neutering and the rapid rate of reproduction among domestic cats, it is crucial to investigate alternative, nonsurgical methods of contraception for this species. Sperm protein IZUMO1 and its oocyte receptor JUNO have been proposed as potential targets for nonsurgical contraceptives. This study aimed to demonstrate (1) the protein coding sequence of feline IZUMO1 and JUNO, (2) gene expression in specific organs by measuring mRNA levels in different visceral tissues, and (3) the expression of IZUMO1 and JUNO during sperm maturation and folliculogenesis, respectively. Amplification for sequencing of feline IZUMO1 and JUNO was performed using the RT-PCR method. Levels of gene expression in different tissues were evaluated using real-time PCR. In situ hybridization was performed to localize JUNO mRNA in ovarian tissues. The complete coding sequences of IZUMO1 and JUNO were obtained and analyzed. A comparison between protein orthologs demonstrated the conservation of IZUMO1 and JUNO in Felidae. The real-time PCR results from various visceral organs indicated that IZUMO1 was significantly higher in the testis than in other organs, whereas JUNO was significantly higher in the ovary than in other organs. Expression of IZUMO1 was found to be higher in the testes than in the caput, corpus, and cauda of epididymides. In situ hybridization revealed that JUNO mRNA was in the ooplasm and nucleus of the primordial, primary, secondary, and antral follicles. Importantly, this was the first study to demonstrate the IZUMO1 and JUNO genes in the testis and ovary of cats. The results are useful for future research related to these genes and for developing contraceptives against these targets.

First report of root and crown rot caused by Fusarium oxysporum Schltdl. on Prunus cerasifera L. in Spain.

In 2021, Spain was the largest producer of cherries in Europe with a production of 125810 tons (FAOSTAT, 2021). In May 2022, in several production fields in Huelva (Spain), wilting was noted in 4-year-old cherry trees cv. Crystal Champaign grafted on rootstock cv. Adara (Prunus cerasifera L.). Gumming and wilting affected approx. 1% of the production area, leading to the eventual collapse and death of most affected trees within 2-3 years. Discoloration of the vascular system of the crown and roots was also noted. Symptomatic crown and root pieces (0.5 cm) were subjected to surface sterilization: immersed in 1% NaClO for 2 min, rinsed in sterile distilled water, and air-dried in a laminar flow cabinet. Then, plant tissues were placed on potato dextrose agar (PDA) amended with streptomycin and incubated in a lab bench at room temperature for a week. Cottony and pink colonies were observed growing from the tissues. Two single strains (F175 and F176) were obtained from each tree by excising single spores (Gordon and Okamoto 1991). Isolates produced sparse aerial mycelia with white to pinkish-orange pigmentation on Spezieller Nährstoffarmer Agar (SNA). Both isolates produced microconidia in false heads on short monophialides. Microconidia were hyaline and measured in the range of 5.0-17.5 × 2.5-3.8 µm for both isolates (n = 50). Macroconidia were less abundant, falciform, and hyaline. Morphological characteristics were consistent with identification as Fusarium spp. (Leslie and Summerell 2006). A portion of the translation elongation factor-1 alpha (EF-1α) gene was sequenced using EF1/2 primers (O'Donnell et al. 1998) (GenBank Accession Nos. OR733348 and OR733349). Based on a comparison of 619 base pairs (bp), both isolates exhibited different sequences, with a 99.5% similarity (616/619 bp). A comparison with previously described isolates revealed a 100% match with published F. oxysporum sequences in the GenBank database (KT323846 and MZ404079, respectively). Isolates were used to conduct pathogenicity tests on 1-year-old plants cv. Adara growing in 512 cm3 pots. Using a scalpel, a 6-7 mm-length wound (2-3 mm deep) was made 5 cm above the soil line in all plants. For each isolate, 5 plants were inoculated by placing a 5 mm plug containing 10-day-old mycelia grown in AMAP medium (Borrero et al. 2009) at the incision point. Non-colonized AMAP plugs were used to inoculate 5 control plants. The inoculated sites were sealed with parafilm. Plants were randomly placed in a growth chamber with a temperature of 28/24ºC and a 12-hour photoperiod. A reddish-brown vascular stem discoloration was noticed in all the inoculated plants 73 days after inoculation. On average, the length of the necrotic area was 12.73 cm for F175, 20.12 cm for F176, and 4.59 cm for the control plants. Fusarium oxysporum was successfully re-isolated from all the inoculated plants. Recovered isolates were confirmed to be the same as the inoculated ones by sequencing the EF-1α gene. A one-way ANOVA indicates that plants cv. Adara were susceptible to both F. oxysporum isolates (P < 0.05). This is particularly noteworthy as cherries are predominantly planted on rootstocks and cv. Adara is a widely used rootstock in Spain. While F. oxysporum has been reported as the cause of root and crown rot in sweet cherry (P. avium L.) in British Columbia (Úrbez-Torres et al. 2016), this is the first report of F. oxysporum causing root and crown rot in cherry rootstocks (P. cerasifera L.) in Spain.

Adenylate kinase 9 is essential for sperm function and male fertility in mammals.

Despite passing routine laboratory tests for semen quality, bulls used in artificial insemination exhibit significant variation in fertility. Routine analysis of fertility data identified a dairy bull with extreme subfertility (10% pregnancy rate). To characterize the subfertility phenotype, a range of in vitro, in vivo, and molecular assays were carried out. Sperm from the subfertile bull exhibited reduced motility and severely reduced caffeine-induced hyperactivation compared to controls. Ability to penetrate the zona pellucida, cleavage rate, cleavage kinetics, and blastocyst yield after IVF or AI were significantly lower than in control bulls. Whole-genome sequencing from semen and RNA sequencing of testis tissue revealed a critical mutation in adenylate kinase 9 (AK9) that impaired splicing, leading to a premature termination codon and a severely truncated protein. Mice deficient in AK9 were generated to further investigate the function of the gene; knockout males were phenotypically indistinguishable from their wild-type littermates but produced immotile sperm that were incapable of normal fertilization. These sperm exhibited numerous abnormalities, including a low ATP concentration and reduced motility. RNA-seq analysis of their testis revealed differential gene expression of components of the axoneme and sperm flagellum as well as steroid metabolic processes. Sperm ultrastructural analysis showed a high percentage of sperm with abnormal flagella. Combined bovine and murine data indicate the essential metabolic role of AK9 in sperm motility and/or hyperactivation, which in turn affects sperm binding and penetration of the zona pellucida. Thus, AK9 has been found to be directly implicated in impaired male fertility in mammals.

Optimization of the Use of Industrial Wastes in Anaerobic Soil Disinfestation for the Control of Fusarium Wilt in Strawberry.

Anaerobic soil disinfestation (ASD) is proposed as an alternative to the use of chemical fumigants against Fusarium wilt in strawberry crops. Different residual wastes (rice bran, fishmeal, and residual strawberry extrudate) were assessed as amendments for ASD. Two different concentrations and two incubation durations were tested in growth chamber trials. The abundance of several microbial groups was noted before and after the treatments. Strawberry plants were grown in the treated soils to record Fusarium wilt disease severity. The population density of F. oxysporum increased after ASD in most amendments with rice bran and residual strawberry extrudate. Changes in Trichoderma spp., copiotrophic bacteria, and Streptomyces spp. populations were observed after anaerobiosis treatments and plant trials. A reduction in the disease severity was achieved in ASD-treated soils with 20 t/ha of rice bran at both 25 and 60 days of incubation, but not when using a 13.5 t/ha dose. Similarly, treatments using 19.3 t/ha of fishmeal for both incubation durations were able to reduce disease severity. In contrast, a severity reduction was only obtained in soils treated with 25.02 t/ha of the residual strawberry extrudate and incubated for 60 days in anaerobic conditions. Two of the three by-products tested were able to reduce Fusarium wilt symptoms in strawberry plants after an ASD-treatment period of only 25 days. Accordingly, the technique seems promising for strawberry growers in Huelva, Spain, and highly sustainable by giving value to residues produced in surrounding areas.

Colostrum Features of Active and Recovered COVID-19 Patients Revealed Using Next-Generation Proteomics Technique, SWATH-MS.

Colostrum performs nutritional, anti-inflammatory and anti-infective functions and promotes immune system formation and organ development. The new coronavirus, SARS-CoV-2, has generated concerns about viral transmission through human milk, with a lack of evidence about human milk's protective effects against the infection. This study aimed at analyzing presence of the virus and at identifying the protein expression profile of human colostrum in active and COVID-19-recovered patients. Colostrum samples were collected from women with COVID-19 (n = 3), women recently recovered from the infection (n = 4), and non-infected women (n = 5). The samples were analyzed by means of RT-qPCR to determine presence of the virus and using SWATH-MS for proteomic analysis. Proteomic results were then analyzed using bioinformatic methods. The viral tests were negative for SARS-CoV-2 in the colostrum from COVID-19 patients. The proteomic analysis identified 301 common proteins in all samples analyzed. Nineteen proteins were upregulated and 7 were downregulated in the COVID-19 group versus the control samples, whereas 18 were upregulated and 7 were downregulated when comparing the COVID-19 group to the recovered group. Eleven proteins were biomarkers of active COVID-19 infection. Ten were upregulated: ACTN1, CD36, FAM3B, GPRC5B, IGHA2, IGK, PLTP, RAC1, SDCBP and SERPINF1, and one was downregulated: PSAP. These proteins are mainly related to immunity, inflammatory response and protein transport. In conclusion, the results of this study suggest that colostrum is not a vehicle for mother-to-child SARS-CoV-2 transmission. Moreover, the colostrum's proteome of active and recuperated patients indicate that it could provide immune benefits to infants.

Human Sperm Head Vacuoles Are Related to Nuclear-Envelope Invaginations.

Nuclear vacuoles are specific structures present on the head of the human sperm of fertile and non-fertile men. Human sperm head vacuoles have been previously studied using motile sperm organelle morphology examination (MSOME) and their origin related to abnormal morphology, abnormal chromatin condensation and DNA fragmentation. However, other studies argued that human sperm vacuoles are physiological structures and consequently, to date, the nature and origin of the nuclear vacuoles remains to be elucidated. Here, we aim to define the incidence, position, morphology and molecular content of the human sperm vacuoles using transmission electron microscopy (TEM) and immunocytochemistry techniques. The results showed that ~50% of the analyzed human sperm cells (n = 1908; 17 normozoospermic human donors) contained vacuoles mainly located (80%) in the tip head region. A significant positive correlation was found between the sperm vacuole and nucleus areas. Furthermore, it was confirmed that nuclear vacuoles were invaginations of the nuclear envelope from the perinuclear theca and containing cytoskeletal proteins and cytoplasmic enzyme, discarding a nuclear or acrosomal origin. According to our findings, these human sperm head vacuoles are cellular structures originating from nuclear invaginations and contain perinuclear theca (PT) components, allowing us to define a new term of 'nuclear invaginations' rather than 'nuclear vacuoles'.

Molecular Chaperone HSPA2 Distribution During Hyaluronic Acid Selection in Human Sperm.

During fertilization, sperm hyaluronidase activity is essential for spermatozoa to successfully penetrate the hyaluronic acid-enriched extracellular matrix of the cumulus cells. Since molecular chaperones, as the heat shock protein A2, are typically involved in bringing hyaluronic acid receptors to the cell surface, here we evaluated the presence and spatial location of HSPA2 on human spermatozoa based on its hyaluronic acid binding capacity. This study included 16 normozoospermic sperm samples from volunteering donors. The location of HSPA2 was studied in cells before and after 1-h incubation under capacitating conditions, as well as in spermatozoa selected according to their ability of binding to hyaluronic acid. Our results showed no significant differences in HSPA2 immunofluorescent cells before and after 1 h of incubation in capacitating conditions. Nevertheless, after hyaluronic acid selection, the percentage of HSPA2-labelled cells increased significantly, indicating that the interaction with hyaluronic acid may induce the unmasking of HSPA2 epitopes. Furthermore, after swim-up and hyaluronic acid selection, spermatozoa presented a highly immunostained equatorial band with a homogeneous fluorescence throughout the acrosomal region. This distribution has been previously suggested to have important implications in male fertility. Noteworthy, a homogeneous fluorescence among the acrosomal region with a more intense labelling at the apical region was observed only in hyaluronic acid bound sperm cells, which may be associated with primary gamete recognition. Our findings suggest that the hyaluronic acid selection technique and HSPA2 biomarker should be considered candidates to complement the classic seminal analysis before recommending an appropriate assisted reproduction technique.

Soilborne pathogens affect strawberry fruit flavor and quality.

Fusarium oxysporum f. sp. fragariae and Macrophomina phaseolina are soilborne fungi leading impactful economical losses to strawberry growers worldwide. Symptoms caused by both pathogens are very similar and include vascular discoloration, wilting, stunting, and dieback of plants, but no fruit damage. An extraction of phenolic and volatile compounds was performed on strawberry fruits from three different cultivars while being grown in a plant growth medium infested by each pathogen. Inoculated plants showed higher content of certain phenolic compounds which have antifungal and antioxidant activity and may have a positive impact on strawberry shelf life. On the other hand, root and vascular infections caused by F. oxysporum and M. phaseolina were able to significantly alter strawberry aroma by reducing or increasing the content of specific volatile compounds which also have an important impact on fruit quality. The changes induced in the aroma profiles of the three strawberry cultivars do not only have organoleptic and economic implications for strawberry growers but play an important role in the plant defense system against pathogens. The results indicate a potential of this line of research to develop new tools for the detection and control of soil pathogens.

Water Quality Assessment Bias Associated with Long-Screened Wells Screened across Aquifers with High Nitrate and Arsenic Concentrations.

Semi-arid regions with little surface water commonly experience rapid water table decline rates. To hedge against the falling water table, production wells in central Mexico are commonly installed to depths of several hundred meters below the present water table and constructed as open boreholes or perforated casings across their entire length. Such wells represent highly conductive pathways leading to non-negligible flow across chemically distinct layers of an aquifer-a phenomenon known as ambient flow. The objectives of this study were to estimate the rate of ambient flow in seven production wells utilizing an end-member mixing model that is constrained by the observed transient chemical composition of produced water. The end-member chemical composition of the upper and lower layers of an urban aquifer that overlies geothermal heat is estimated to anticipate the future quality of this sole source of water for a rapidly growing urban area. The comprehensive water chemistry produced by seven continuously perforated municipal production wells, spanning three geologically unique zones across the city of San Miguel de Allende in Guanajuato State, was monitored during one day of pumping. The concentration of conservative constituents gradually converged on steady-state values. The model indicates that, relative to the lower aquifer, the upper aquifer generally has higher specific conductance (SC), chloride (Cl), nitrate (NO3), calcium (Ca), barium (Ba) and magnesium (Mg). The lower aquifer generally has a higher temperature, sodium (Na), boron (B), arsenic (As) and radon (Rn). Ambient flow ranged from 33.1 L/min to 225.7 L/min across the seven wells, but this rate for a given well varied depending on which tracer was used. This new 3D understanding of the chemical stratification of the aquifer suggests that as water tables continue to fall, concentrations of geothermally associated contaminants of concern will increase in the near future, potentially jeopardizing the safety of municipal drinking water.

Analysis of Minor Proteins Present in Breast Milk by Using WGA Lectin.

Breast milk is a complex and dynamic biological fluid and considered an essential source of nutrition in early life. In its composition, the proteins have a relevant biological activity and are related to the multiple benefits demonstrated when compared with artificial milks derived from cow's milk. Understanding human milk composition provides an important tool for health care providers toward the management of infant feeding and the establishment of breastfeeding. In this work, a new technique was developed to increase the knowledge of human milk, because many of the components remain unknown. To isolate minor proteins present in breast milk by using WGA lectin, breast milk was centrifuged to remove cells and separate the fat phase from the serum phase. The serum obtained was separated into two groups: control (n = 3; whole serum sample from mature milk) and WGA lectin (n = 3; sample processed with WGA lectin to isolate glycosylated proteins). The samples were analyzed by high-performance liquid chromatography coupled to mass spectrometry (HPLC/MS). A total of 84 different proteins were identified from all of the samples. In the WGA lectin group, 55 different proteins were isolated, 77% of which had biological functions related to the immune response. Of these proteins, there were eight WGA lectin group exclusives, and two had not previously been described in breast milk (polyubiquitin-B and POTE ankyrin domain family member F). Isolation by WGA lectin is a useful technique to detect minor proteins in breast milk and to identify proteins that could not be observed in whole serum.

Feasibility of near infrared spectroscopy for estimating suppressiveness of carnation (Dianthus cariophyllus L.) fusarium wilt in different plant growth media.

Fusarium wilt is one of the most widespread diseases in carnation crops in a large number of countries. Plant protection products commonly used to remedy the disease have been considered ineffective and environmentally unsafe for commercial use. As an alternative, the use of suppressive growth media has been proposed. The aim of this study was to evaluate the use of a rapid method such as near infrared spectroscopy (NIRS) to monitor and evaluate suppressive media potential. The NIR spectra were collected from 6 plant growth media used in a series of trials to evaluate suppressiveness to carnation Fusarium wilt namely grape marc compost, cork compost, olive oil husk + cotton gin trash composted and mixed with rice husk, spent mushroom composted and mixed with peat, coir fiber and light peat. The NIR calibration models showed promising results for estimating pH, ß-glucosidase activity, disease severity (AUDPC and RLSBX) in the growth media evaluated, with coefficients of determination of 0.99, 0.98, 0.98 and 0.90; SECV of 0.09, 11.63, 0.05 and 0.10; and RPD values of 13.86, 6.62, 7.19 and 3.24; respectively. NIR spectroscopy could become a useful non-destructive and fast analytical tool for the identification of Fusarium wilt suppressive composts, avoiding the use of reagents.

The Campo de Dalias GNSS Network Unveils the Interaction between Roll-Back and Indentation Tectonics in the Gibraltar Arc.

The Gibraltar Arc includes the Betic and Rif Cordilleras surrounding the Alboran Sea; it is formed at the northwest-southeast Eurasia-Nubia convergent plate boundary in the westernmost Mediterranean. Since 2006, the Campo de Dalias GNSS network has monitored active tectonic deformation of the most seismically active area on the north coast of the Alboran Sea. Our results show that the residual deformation rates with respect to Eurasia range from 1.7 to 3.0 mm/year; roughly homogenous west-southwestward displacements of the northern sites occur, while the southern sites evidence irregular displacements towards the west and northwest. This deformation pattern supports simultaneous east-northeast-west-southwest extension, accommodated by normal and oblique faults, and north-northwest-south-southeast shortening that develops east-northeast-west-southwest folds. Moreover, the GNSS results point to dextral creep of the main northwest-southeast Balanegra Fault. These GNNS results thus reveal, for the first time, present-day interaction of the roll-back tectonics of the Rif-Gibraltar-Betic slab in the western part of the Gibraltar Arc with the indentation tectonics affecting the eastern and southern areas, providing new insights for improving tectonic models of arcuate orogens.

Protein Identification of Spermatozoa and Seminal Plasma in Bottlenose Dolphin (Tursiops truncatus).

Proteins play an important role in many reproductive functions such as sperm maturation, sperm transit in the female genital tract or sperm-oocyte interaction. However, in general, little information concerning reproductive features is available in the case of aquatic animals. The present study aims to characterize the proteome of both spermatozoa and seminal plasma of bottlenose dolphins (Tursiops truncatus) as a model organism for cetaceans. Ejaculate samples were obtained from two trained dolphins housed in an aquarium. Spermatozoa and seminal plasma were analyzed by means of proteomic analyses using an LC-MS/MS, and a list with the gene symbols corresponding to each protein was submitted to the DAVID database. Of the 419 proteins identified in spermatozoa and 303 in seminal plasma, 111 proteins were shared by both. Furthermore, 70 proteins were identified as involved in reproductive processes, 39 in spermatozoa, and 31 in seminal plasma. The five most abundant proteins were also identified in these samples: AKAP3, ODF2, TUBB, GSTM3, ROPN1 for spermatozoa and CST11, LTF, ALB, HSP90B1, PIGR for seminal plasma. In conclusion, this study provides the first characterization of the proteome in cetacean sperm and seminal plasma, opening the way to future research into new biomarkers, the analysis of conservation capacity or possible additional applications in the field of assisted reproductive technologies.

Evidence of haptoglobin in the porcine female genital tract during oestrous cycle and its effect on in vitro embryo production.

Recent evidence supports involvement of the acute phase protein haptoglobin in numerous events during mammalian reproduction. The present study represents an in-depth investigation of haptoglobin expression and secretion in the porcine oviduct and uterus, and assesses its effect on porcine in vitro embryo production. A systematic study was made of sows in different oestrous stages: late follicular, early luteal and late luteal stages. Relative haptoglobin mRNA abundance was quantified by RT-qPCR. In addition, expression of the protein was analysed by immunohistochemistry and the results were complemented by Western-blot and proteomic analyses of the oviductal and uterine fluids. In vitro porcine fertilization and embryo culture were carried out in the presence of haptoglobin. The results indicate that haptoglobin mRNA expression in the porcine oviduct and uterus is most abundant during the late luteal stage of the oestrous cycle. By means of Western blot and proteomic analyses haptoglobin presence was demonstrated in the oviduct epithelium and in the oviductal and uterine fluids in different stages of the oestrous cycle. The addition of haptoglobin during gamete co-incubation had no effect on sperm penetration, monospermy or efficiency rates; however, compared with the control group, blastocyst development was significantly improved when haptoglobin was present (haptoglobin: 64.50% vs. control: 37.83%; p < 0.05). In conclusion, the presence of haptoglobin in the oviduct and uterus of sows at different stages of the oestrous cycle suggests that it plays an important role in the reproduction process. The addition of haptoglobin during in vitro embryo production improved the blastocyst rates.

New Insights into the Mammalian Egg Zona Pellucida.

Mammalian oocytes are surrounded by an extracellular coat called the zona pellucida (ZP), which, from an evolutionary point of view, is the most ancient of the coats that envelope vertebrate oocytes and conceptuses. This matrix separates the oocyte from cumulus cells and is responsible for species-specific recognition between gametes, preventing polyspermy and protecting the preimplantation embryo. The ZP is a dynamic structure that shows different properties before and after fertilization. Until very recently, mammalian ZP was believed to be composed of only three glycoproteins, ZP1, ZP2 and ZP3, as first described in mouse. However, studies have revealed that this composition is not necessarily applicable to other mammals. Such differences can be explained by an analysis of the molecular evolution of the ZP gene family, during which ZP genes have suffered pseudogenization and duplication events that have resulted in differing models of ZP protein composition. The many discoveries made in recent years related to ZP composition and evolution suggest that a compilation would be useful. Moreover, this review analyses ZP biosynthesis, the role of each ZP protein in different mammalian species and how these proteins may interact among themselves and with other proteins present in the oviductal lumen.

Horizontal chromosome transfer and independent evolution drive diversification in Fusarium oxysporum f. sp. fragariae.

The genes required for host-specific pathogenicity in Fusarium oxysporum can be acquired through horizontal chromosome transfer (HCT). However, it is unknown if HCT commonly contributes to the diversification of pathotypes. Using comparative genomics and pathogenicity phenotyping, we explored the role of HCT in the evolution of F. oxysporum f. sp. fragariae, the cause of Fusarium wilt of strawberry, with isolates from four continents. We observed two distinct syndromes: one included chlorosis ('yellows-fragariae') and the other did not ('wilt-fragariae'). All yellows-fragariae isolates carried a predicted pathogenicity chromosome, 'chrY-frag ', that was horizontally transferred at least four times. chrY-frag was associated with virulence on specific cultivars and encoded predicted effectors that were highly upregulated during infection. chrY-frag was not present in wilt-fragariae; isolates causing this syndrome evolved pathogenicity independently. All origins of F. oxysporum f. sp. fragariae occurred outside of the host's native range. Our data support the conclusion that HCT is widespread in F. oxysporum, but pathogenicity can also evolve independently. The absence of chrY-frag in wilt-fragariae suggests that multiple, distinct pathogenicity chromosomes can confer the same host specificity. The wild progenitors of cultivated strawberry (Fragaria × ananassa) did not co-evolve with this pathogen, yet we discovered several sources of genetic resistance.

Arylsulfatase A Remodeling during Human Sperm In Vitro Capacitation Using Field Emission Scanning Electron Microscopy (FE-SEM).

Capacitation drives sperm biophysical and biochemical changes for sperm-oocyte interactions. It is a well-known fact that the molecular complex arylsulfatase A (ARSA), hyaluronidase sperm adhesion molecule 1 (SPAM1), and heat shock protein 2 (HSPA2) plays a significant role in sperm-zona pellucida (ZP) binding. However, the time-dependent capacitation effects on the sperm surface ARSA presence and specific topographic distributions remain to be elucidated. Here, we quantified the ARSA density and specific membrane domain locations before (US) and after in vitro capacitation (one and four hours; CS1-CS4) in human sperm using high-resolution field emission scanning electron microscopy (FE-SEM) and immunogold labeling. Our results showed a significant and progressive capacitation-mediated increase of labeled spermatozoa from the US (37%) to CS4 (100%) physiological conditions. In addition, surface mapping revealed a close relationship between the ARSA residues and their acrosomal repositioning. Compared with the ARSA surface heterogeneous distribution found in US, the CS1-4 conditions exhibited clustering on the peri-acrosomal region, showing that time-dependent capacitation also induced a ARSA residue dramatic translocation on sperm surfaces. Our findings provide novel insights into the molecular remodeling events preceding sperm-oocyte interactions.

Human sperm chaperone HSPA2 distribution during in vitro capacitation.

Human fertilization success depends on the ability of the spermatozoa to undergo capacitation. Even though this process can be conducted in vitro, the optimal time for a sperm cell to complete capacitation in vitro is still under discussion due to the lack of proper capacitation biomarkers. Here, we evaluated the influence of in vitro capacitation time on HSPA2 distribution over human sperm head testing this chaperone as a potential capacitation biomarker. The chaperone was assessed in human spermatozoa from 16 normozoospermic donors using indirect immunofluorescence in uncapacitated, one and four-hour capacitated spermatozoa. The percentage of HSPA2 immunofluorescent cells before and after one hour of capacitation did not differ significantly. However, after four hours of capacitation, we observed a significantly higher percentage of HSPA2 labelled cells. In fluorescent cells analysed before capacitation, we could not identify a predominant distribution pattern. Meanwhile, after capacitation, most sperm showed a highly labelled equatorial band accompanied by a homogeneous fluorescence throughout the acrosomal region. Our findings suggest that HSPA2 needs more than one hour of in vitro capacitation for being correctly distributed in the anterior region of the sperm head. In conclusion, the present study provides solid evidences for the utility of HSPA2 as a biomarker of human sperm in vitro capacitation. Due to its importance during egg-sperm recognition, the use of HSPA2 as a biomarker before an artificial reproduction technique may be suggested, in addition to a longer capacitation time during sperm preparation.

A Comparative View on the Oviductal Environment during the Periconception Period.

The oviduct plays important roles in reproductive events: sperm reservoir formation, final gamete maturation, fertilization and early embryo development. It is well known that the oviductal environment affects gametes and embryos and, ultimately, the health of offspring, so that in vivo embryos are better in terms of morphology, cryotolerance, pregnancy rates or epigenetic profile than those obtained in vitro. The deciphering of embryo-maternal interaction in the oviduct may provide a better understanding of the embryo needs during the periconception period to improve reproductive efficiency. Here, we perform a comparative analysis among species of oviductal gene expression related to embryonic development during its journey through the oviduct, as described to date. Cross-talk communication between the oviduct environment and embryo will be studied by analyses of the secreted or exosomal proteins of the oviduct and the presence of receptors in the membrane of the embryo blastomeres. Finally, we review the data that are available to date on the expression and characterization of the most abundant protein in the oviduct, oviductin (OVGP1), highlighting its fundamental role in fertilization and embryonic development.