Pathomorphological lesions in the ovaries of water buffaloes

Background: Buffalo breeding is common in many countries. Buffalo's milk is used in the production of mozzarella, yoghurt, ice cream, and various dairy desserts; meat is preferred in sausage production. The female buffaloes are bred to benefit from their milk and to obtain offspring. These animals, which are not suitable for feeding in barns, generally live in pastures, especially on wet land, and are very difficult to follow. Therefore, diseases occur randomly in slaughterhouses. Studies on genital system problems are very limited. Water buffaloes (Bubalus bubalis) are animals with economic value as many buffalo products are provided. In this respect, inspection and control of female genital disorders is fundamental to ensure good reproductive performance of female buffaloes. The aim of the study was to investigate pathomorphological lesions occurring in the ovaries of water buffaloes which were sent to slaughterhouses. Materials, Methods & Results: A total of 198 ovaries of water buffaloes were collected from various slaughterhouses located in Adapazari, Istanbul, Kocaeli, Samsun and Trabzon cities of Turkey. After macroscopic examination; tissue samples were fixed in 10 % buffered formalin, processed routinely and were stained with hematoxylin-eosin (HE). As a histochemical staining, Masson's trichrome staining was applied to characterize the lesions. Immunohistochemistry (IHC) was performed on 10 % neutral formalin fixed, paraffin-embedded, 4-6-µm-thick sections from ovaries using progesterone receptor (PR) with streptavidin-biotin complex peroxidase (StrepABC-P) method. Histopathologically, follicular cysts (n:147) and luteal cysts (n:22) were seen. While the cyst lumens were sometimes surrounded by granulosa and/or luteal cells, most of them were limited by the connective tissue capsule structure. This capsule structure was shown in blue by Masson's trichrome staining. Hemorrhage was observed in some cystic corpus luteum, diagnosed as corpus hemorrhagicum. In addition, cysts giving papillary extension into the lumen and inflammation of some ovaries were observed. Immunohistochemically, the staining with PR antibody in ovarian cysts showed no immunolabelling around the follicular cyst, while the nuclei of some of the luteal cells forming the luteal cyst had strongly nuclear positivity and slightly cytoplasmic positivity. In the biochemical examination of the fluids obtained from cystic ovaries (n: 37), the average of estradiol was 2.84 ng/mL (min: 0.01 ng/mL, max: 4.30 ng/mL) and progesterone average is 49.09 ng/mL (min: 1.88 ng/mL, max: 254.2 ng/mL). Discussion: Ovarian cysts in buffaloes seem to be among the serious fertility problems as in cattle. Although the exact cause of ovarian cyts has not been determined yet, it is known that the hypothalamic-pituitary-gonadal (HPG) axis is disrupted in cyst formation in general. In this study, the mean estradiol value was within the standard range but close to the lower limit; progesterone value was above the limit. The increase in progesterone level was compatible with the pathogenesis of cyst genesis. Beside this result, staining with PR was positive in the luteal cells that formed the luteal cyst immunohistochemically. In addition, although the hemorrhages observed in the corpus luteum are considered physiological, it should not be forgotten that they can be vital if they rupture. All these results show us the animals sent for slaughtering mostly have serious genital problems threatening their fertility.

Diagnosis of endometritis in the bitch / Diagnóstico da endometrite em cadelas

Endometritis refers to an inflammation of the uterine mucosa that does not extend beyond the stratum spongiosum. Recent studies indicate it is a common finding in 30-50% of bitches suffering from infertility/subfertility. This disorder is subclinical, and its diagnosis involves sampling the uterus, whether it is by performing a cytology from flushing the organ or histology on collected uterine biopsies. Its pathophysiology remains unclear in 2021. Some cases are associated with cystic endometrial hyperplasia, which leads to disruption in the uterine clearance mechanisms after breeding ; while other appear as a pure inflammatory process. While we don’t have all the answers yet, there is no doubt today that these disorders must be included in the differential diagnosis of infertility in the bitch. The work that has already been done on this topic already offers some idea on how to approach these cases in our veterinary clinics.

Alterações histopatológicas testiculares associadas a infecção experimental por vírus Zika em macacos-de-cheiro / Testicular histopathological changes associated with experimental infection by zika virus in squirrel monkeys

The Zika virus belongs to the family Flaviviridae, genus Flavivirus. It is an arbovirus whose main vectors are mosquitoes of the genus Aedes. Studies in rodents have shown that the persistence of the virus in the testicles causes damage to the reproductive tissue. This work aimed to study the effect of experimental infection by the Zika virus on fertility in non-human primates of the species Saimiri collinsi. Five pre-pubertal males (≤2 years old) of the species Saimiri collinsi were used. Three animals were infected (infected group) with the strain of Zika virus BE H815744. Two other uninfected males were used as a negative control (uninfected group). Twenty-one days after infection, infected and uninfected males were euthanized. After euthanasia, they were referred for necroscopic examination for macroscopic and microscopic evaluation. During the necropsy, the testicles were collected and fixed in 10% formaldehyde. After fixation, the tissues were processed routinely and embedded in paraffin. The slides were stained with hematoxylin and eosin for histopathological evaluation. Histopathological changes were observed in the testis of three of the five animals. Different degrees of inflammation were identified, in addition to degeneration and/or necrosis. The three animals presented a reduced number of sperm cells, with no sperm and severe necrosis. The results obtained conclude that the Zika virus can cause pathological changes in the reproductive system of males of the species Saimiri collinsi.

Alterações histopatológicas testiculares associadas a infecção experimental por vírus Zika em macacos-de-cheiro / Testicular histopathological changes associated with experimental infection by zika virus in squirrel monkeys

The Zika virus belongs to the family Flaviviridae, genus Flavivirus. It is an arbovirus whose main vectors are mosquitoes of the genus Aedes. Studies in rodents have shown that the persistence of the virus in the testicles causes damage to the reproductive tissue. This work aimed to study the effect of experimental infection by the Zika virus on fertility in non-human primates of the species Saimiri collinsi. Five pre-pubertal males (≤2 years old) of the species Saimiri collinsi were used. Three animals were infected (infected group) with the strain of Zika virus BE H815744. Two other uninfected males were used as a negative control (uninfected group). Twenty-one days after infection, infected and uninfected males were euthanized. After euthanasia, they were referred for necroscopic examination for macroscopic and microscopic evaluation. During the necropsy, the testicles were collected and fixed in 10% formaldehyde. After fixation, the tissues were processed routinely and embedded in paraffin. The slides were stained with hematoxylin and eosin for histopathological evaluation. Histopathological changes were observed in the testis of three of the five animals. Different degrees of inflammation were identified, in addition to degeneration and/or necrosis. The three animals presented a reduced number of sperm cells, with no sperm and severe necrosis. The results obtained conclude that the Zika virus can cause pathological changes in the reproductive system of males of the species Saimiri collinsi.(AU)

Métodos de coleta de amostra para exame de citologia endometrial em éguas com endometrite / Sample collection methods for endometrial cytology examination in mares with endometritis

A endometrite é definida como uma inflamação do endométrio, podendo ser aguda ou crônica, infecciosa ou não infecciosa. Essa afecção é de grande importância econômica na medicina veterinária, podendo resultar em morte embrionária, aborto e até tornar a égua infértil, sendo uma das afecções que mais comprometem a sanidade uterina, colocando em risco os índices de prenhez obtidos pós-cobertura, pós-inseminação, e pós-inovulação de embriões. Existem diversos métodos para diagnóstico e o mais utilizado a campo é a citologia endometrial/uterina. Também existem algumas formas de coleta de amostra para análise laboratorial pela citologia, tais como: escova ginecológica, swab uterino guiado por mandril metálico ou plástico descartável e lavado uterino de baixo volume, seguido de sedimentação ou centrifugação podendo ser complementada por citocentrifugação. O diagnóstico pela citologia endometrial por meio da técnica da escova ginecológica é a mais utilizada entre os médicos veterinários, pois trata-se de um método prático para execução e rápido quanto à obtenção dos resultados sobre a presença ou não da endometrite, facilitando a conduta terapêutica quanto à utilização das éguas para reprodução ou descarte. Objetivou-se com esse estudo citaras formas de diagnóstico da endometrite em éguas, enfatizando a citologia endometrial e técnicas para obtenção de amostra, identificando a técnica mais indicada por diversos autores.(AU)

Endometritis is defined as an inflammation of the endometrium, which may be acute or chronic, infectious or noninfectious. This condition is of great economic importance in veterinary medicine, and may result in embryonic death, abortion and even render the mare infertile, being one of the conditions that most compromise uterine health, putting at risk the pregnancy rates obtained post-coverage, postinsemination, and post-embryo transfer. There are several methods for diagnosis and the most used in the field is endometrial/uterine cytology. There are also some forms of sample collection for laboratory analysis by cytology, such as: gynecological brush, uterine swab guided by metallic mandrel or disposable plastic and low volume uterine lavage, followed by sedimentation or centrifugation and may be complemented by cytocentrifugation. Diagnosis by endometrial cytology using the gynecological brush technique is the most used among Veterinary Practitioners, because it is a practical method to execute and fast to obtain the results in the presence of endometritis, facilitating the therapeutic conduct regarding the use of mares for breeding or discarding. The objective of this study was to cite the diagnostic methods of endometritis in mares, emphasizing endometrial cytology and its techniques for obtaining a sample, identifying the most indicated technique by several authors.(AU)

Métodos de coleta de amostra para exame de citologia endometrial em éguas com endometrite / Sample collection methods for endometrial cytology examination in mares with endometritis

A endometrite é definida como uma inflamação do endométrio, podendo ser aguda ou crônica, infecciosa ou não infecciosa. Essa afecção é de grande importância econômica na medicina veterinária, podendo resultar em morte embrionária, aborto e até tornar a égua infértil, sendo uma das afecções que mais comprometem a sanidade uterina, colocando em risco os índices de prenhez obtidos pós-cobertura, pós-inseminação, e pós-inovulação de embriões. Existem diversos métodos para diagnóstico e o mais utilizado a campo é a citologia endometrial/uterina. Também existem algumas formas de coleta de amostra para análise laboratorial pela citologia, tais como: escova ginecológica, swab uterino guiado por mandril metálico ou plástico descartável e lavado uterino de baixo volume, seguido de sedimentação ou centrifugação podendo ser complementada por citocentrifugação. O diagnóstico pela citologia endometrial por meio da técnica da escova ginecológica é a mais utilizada entre os médicos veterinários, pois trata-se de um método prático para execução e rápido quanto à obtenção dos resultados sobre a presença ou não da endometrite, facilitando a conduta terapêutica quanto à utilização das éguas para reprodução ou descarte. Objetivou-se com esse estudo citaras formas de diagnóstico da endometrite em éguas, enfatizando a citologia endometrial e técnicas para obtenção de amostra, identificando a técnica mais indicada por diversos autores.

Endometritis is defined as an inflammation of the endometrium, which may be acute or chronic, infectious or noninfectious. This condition is of great economic importance in veterinary medicine, and may result in embryonic death, abortion and even render the mare infertile, being one of the conditions that most compromise uterine health, putting at risk the pregnancy rates obtained post-coverage, postinsemination, and post-embryo transfer. There are several methods for diagnosis and the most used in the field is endometrial/uterine cytology. There are also some forms of sample collection for laboratory analysis by cytology, such as: gynecological brush, uterine swab guided by metallic mandrel or disposable plastic and low volume uterine lavage, followed by sedimentation or centrifugation and may be complemented by cytocentrifugation. Diagnosis by endometrial cytology using the gynecological brush technique is the most used among Veterinary Practitioners, because it is a practical method to execute and fast to obtain the results in the presence of endometritis, facilitating the therapeutic conduct regarding the use of mares for breeding or discarding. The objective of this study was to cite the diagnostic methods of endometritis in mares, emphasizing endometrial cytology and its techniques for obtaining a sample, identifying the most indicated technique by several authors.

Inflammation: friend or foe of bovine reproduction?

Inflammation is not only the first line of defense of the organism but is also required in many reproductive processes such as ovulation, corpus luteum development, luteolysis, uterine clearance after insemination and post partum. Nevertheless, if excessive or persistent, inflammation can switch from a positive mechanism to a deleterious process, impairing oocyte quality and embryo development. Not only uterine but also non genital inflammatory sites can depreciate reproductive performances, with a carry over effect of 2 to 4 months. Since the metabolic challenges of the peripartum transition period make difficult for the cow to control inflammation, dairy cows are frequently in a pro-inflammatory stage, suggesting that inflammation, rather than infection, is a limiting factor of fertility in modern dairy cows. Within the first week after calving, cows have to mount an intense inflammatory response to the bacterial invasion of the uterine cavity with the challenge of being able to switch it off in no more than 5-6 weeks. The absence of neutrophils on endometrial smear is associated with the highest success rate at insemination. Since a fine tuning – rather than an absence - of inflammation is required along the reproductive cycle, anti-inflammatory drugs do not allow any improvement of pregnancy rate, except in the specific case of embryo transfer. Appropriate management of the transition period (especially nutritional) and in a long term perspective, genetic selection contribute to improve the aptitude of cows to controls the intensity of inflammatory process.

Inflammation: friend or foe of bovine reproduction?

Inflammation is not only the first line of defense of the organism but is also required in many reproductive processes such as ovulation, corpus luteum development, luteolysis, uterine clearance after insemination and post partum. Nevertheless, if excessive or persistent, inflammation can switch from a positive mechanism to a deleterious process, impairing oocyte quality and embryo development. Not only uterine but also non genital inflammatory sites can depreciate reproductive performances, with a carry over effect of 2 to 4 months. Since the metabolic challenges of the peripartum transition period make difficult for the cow to control inflammation, dairy cows are frequently in a pro-inflammatory stage, suggesting that inflammation, rather than infection, is a limiting factor of fertility in modern dairy cows. Within the first week after calving, cows have to mount an intense inflammatory response to the bacterial invasion of the uterine cavity with the challenge of being able to switch it off in no more than 5-6 weeks. The absence of neutrophils on endometrial smear is associated with the highest success rate at insemination. Since a fine tuning rather than an absence - of inflammation is required along the reproductive cycle, anti-inflammatory drugs do not allow any improvement of pregnancy rate, except in the specific case of embryo transfer. Appropriate management of the transition period (especially nutritional) and in a long term perspective, genetic selection contribute to improve the aptitude of cows to controls the intensity of inflammatory process.(AU)

Changes induced by pathogens and metabolic stress on endometrial function in cattle: possible impacts of increased inflammation on fertility / Mudanças induzidas por patógenos e estresse metabólico na função endometrial de bovinos: possíveis impactos do aumento da inflamação na fertilidade

Harmonized embryo maternal relationships, are necessary at the beginning of pregnancy to insure the development of the young embryo and of the placenta. Specific mechanisms modulate the immune balance towards immune-tolerance for the embryo to be accepted by the endometrium. By the same time this tissue is submitted to intense remodelling, under complex signalling, allowing implantation. Some pathogens and metabolic stress, especially excessive mobilisation of fat tissue, disturb this delicate balance. This review describes how these stressors can alter endometrial function through pro-inflammatory mechanisms and by inducing changes of specific signals possibly altering the establishment of contacts and functional interactions between the embryo and the endometrium around time of implantation.(AU)

Changes induced by pathogens and metabolic stress on endometrial function in cattle: possible impacts of increased inflammation on fertility / Mudanças induzidas por patógenos e estresse metabólico na função endometrial de bovinos: possíveis impactos do aumento da inflamação na fertilidade

Harmonized embryo maternal relationships, are necessary at the beginning of pregnancy to insure the development of the young embryo and of the placenta. Specific mechanisms modulate the immune balance towards immune-tolerance for the embryo to be accepted by the endometrium. By the same time this tissue is submitted to intense remodelling, under complex signalling, allowing implantation. Some pathogens and metabolic stress, especially excessive mobilisation of fat tissue, disturb this delicate balance. This review describes how these stressors can alter endometrial function through pro-inflammatory mechanisms and by inducing changes of specific signals possibly altering the establishment of contacts and functional interactions between the embryo and the endometrium around time of implantation.

Genetic control of reproduction in dairy cows under grazing conditions

Fertility performance is a key driver of the efficiency and profitability of seasonal-calving pasturebased systems of milk production. Since the 1990’s and early 2000’s, most countries have placed varying levels of emphasis on fertility and survivability traits, and phenotypic performance has started to improve. In recent years, the underlying physiological mechanisms responsible for good or poor phenotypic fertility have started to be unravelled. It is apparent that poor genetic merit for fertility traits is associated with multiple defects across a range of organs and tissues that are antagonistic to achieving satisfactory fertility performance. The principal defects include excessive mobilisation of body condition score (BCS), unfavourable metabolic status, delayed resumption of cyclicity, increased incidence of endometritis, dysfunctional estrous expression, and inadequate luteal phase progesterone concentrations. At a tissue level, coordinated changes in gene expression in different tissues have been observed to orchestrate more favourable BCS, uterine environment and corpus luteum function. Interestingly, cows with poor genetic merit for fertility traits have up-regulated inflammation and immune response pathways in multiple tissues. Sire genetic merit for daughter fertility traits is improving rapidly in the dairy breeds, especially in the predominant Holstein and Friesian breeds. With advances in animal breeding, especially genomic technologies to identify superior sires, genetic merit for fertility traits can be improved much more quickly than they initially declined.

Genetic control of reproduction in dairy cows under grazing conditions

Fertility performance is a key driver of the efficiency and profitability of seasonal-calving pasturebased systems of milk production. Since the 1990s and early 2000s, most countries have placed varying levels of emphasis on fertility and survivability traits, and phenotypic performance has started to improve. In recent years, the underlying physiological mechanisms responsible for good or poor phenotypic fertility have started to be unravelled. It is apparent that poor genetic merit for fertility traits is associated with multiple defects across a range of organs and tissues that are antagonistic to achieving satisfactory fertility performance. The principal defects include excessive mobilisation of body condition score (BCS), unfavourable metabolic status, delayed resumption of cyclicity, increased incidence of endometritis, dysfunctional estrous expression, and inadequate luteal phase progesterone concentrations. At a tissue level, coordinated changes in gene expression in different tissues have been observed to orchestrate more favourable BCS, uterine environment and corpus luteum function. Interestingly, cows with poor genetic merit for fertility traits have up-regulated inflammation and immune response pathways in multiple tissues. Sire genetic merit for daughter fertility traits is improving rapidly in the dairy breeds, especially in the predominant Holstein and Friesian breeds. With advances in animal breeding, especially genomic technologies to identify superior sires, genetic merit for fertility traits can be improved much more quickly than they initially declined.(AU)

Pathogenesis of uterine diseases in dairy cattle and implications for fertility

Uterine diseases in cattle occur at all stages of the reproduction cycle but the majority of cases is found in the postpartum period. The inflammation of the uterus is generally defined as metritis or endometritis, with several graduations, e.g. puerperal metritis, clinical metritis, clinical or subclinical endometritis. Whether uterine diseases have a negligible, moderate or detrimental effect on fertility is still under discussion and depends on definitions and classification. In the past, it was assumed that the pregnant uterus is free of pathogens, but recent studies found several species including pathogens in the uterus and endometrium of pregnant cows. After parturition, a broad diversity of bacteria with >200 different species has been found in the early postpartum period. Not all of these bacteria, however, are considered as pathogens. Furthermore, bacteriological findings provide only evidence for infection but not for inflammation. For some bacteria, particularly Escherichia coli and Trueperella pyogenes pathogenic mechanism resulting in metritis and endometritis have been elucidated in detail. The role of bacteria that can be regarded as opportunistic or potential pathogens, e.g. Bacillus pumilus, is still under investigation. The understanding of the uterine microbiota and its interactions is increasing with the use of modern high-resolution techniques such as Fouriertransform infrared spectroscopy. Endometrial cytology provides additional information about alterations in the endometrium. Knowledge of innate uterine defense mechanism in cattle has increased a lot in the recent past. It can be speculated that improving or modulating uterine defense mechanism will be part of future prevention and treatment approaches beyond the use of antimicrobials. In this context, cellular and molecular defense mechanisms have been in the focus of interest, e.g. the role of interleukins or mucins. This review gives a short overview on some aspects of recent research on uterine diseases in cattle.

Pathogenesis of uterine diseases in dairy cattle and implications for fertility

Uterine diseases in cattle occur at all stages of the reproduction cycle but the majority of cases is found in the postpartum period. The inflammation of the uterus is generally defined as metritis or endometritis, with several graduations, e.g. puerperal metritis, clinical metritis, clinical or subclinical endometritis. Whether uterine diseases have a negligible, moderate or detrimental effect on fertility is still under discussion and depends on definitions and classification. In the past, it was assumed that the pregnant uterus is free of pathogens, but recent studies found several species including pathogens in the uterus and endometrium of pregnant cows. After parturition, a broad diversity of bacteria with >200 different species has been found in the early postpartum period. Not all of these bacteria, however, are considered as pathogens. Furthermore, bacteriological findings provide only evidence for infection but not for inflammation. For some bacteria, particularly Escherichia coli and Trueperella pyogenes pathogenic mechanism resulting in metritis and endometritis have been elucidated in detail. The role of bacteria that can be regarded as opportunistic or potential pathogens, e.g. Bacillus pumilus, is still under investigation. The understanding of the uterine microbiota and its interactions is increasing with the use of modern high-resolution techniques such as Fouriertransform infrared spectroscopy. Endometrial cytology provides additional information about alterations in the endometrium. Knowledge of innate uterine defense mechanism in cattle has increased a lot in the recent past. It can be speculated that improving or modulating uterine defense mechanism will be part of future prevention and treatment approaches beyond the use of antimicrobials. In this context, cellular and molecular defense mechanisms have been in the focus of interest, e.g. the role of interleukins or mucins. This review gives a short overview on some aspects of recent research on uterine diseases in cattle.(AU)

Impact and mechanisms of inflammatory diseases on embryonic development and fertility in cattle

Inflammatory diseases are prevalent in cattle and impair fertility. Cows affected by inflammatory disease from parturition to the day before breeding have reduced fertilization of oocytes, reduced survival of zygotes to the morula stage, impaired development to early stages of conceptus elongation, reduced secretion of interferon during the period of pregnancy recognition, altered transcriptome of preimplantation conceptus cells, and increased pregnancy loss. Consequently, these cows have reduced pregnancy and calving per breeding. Reduced oocyte competence is alikely reason for the carryover effects of diseases on developmental biology, But impaired uterine environment is also involved. Effects on pregnancy survival are observed up to 5 months after the diagnosis and treatment of disease, and effects on developmental biology seem to be extended into postnatal life in pregnancies that survive until term. Although the biological mechanism mediating the effects of inflammatory diseases are still not completely understood, control of inflammation during the clinical presentation of diseases seems to alleviate the negative effects on reproductive biology. It is increasingly evident that animal health, not only at the time of breeding or pregnancy development but also in the period preceding breeding, is imperative for optimal reproduction in cattle and should always be considered in herd evaluations and managerial decisions.

Impact and mechanisms of inflammatory diseases on embryonic development and fertility in cattle

Inflammatory diseases are prevalent in cattle and impair fertility. Cows affected by inflammatory disease from parturition to the day before breeding have reduced fertilization of oocytes, reduced survival of zygotes to the morula stage, impaired development to early stages of conceptus elongation, reduced secretion of interferon during the period of pregnancy recognition, altered transcriptome of preimplantation conceptus cells, and increased pregnancy loss. Consequently, these cows have reduced pregnancy and calving per breeding. Reduced oocyte competence is alikely reason for the carryover effects of diseases on developmental biology, But impaired uterine environment is also involved. Effects on pregnancy survival are observed up to 5 months after the diagnosis and treatment of disease, and effects on developmental biology seem to be extended into postnatal life in pregnancies that survive until term. Although the biological mechanism mediating the effects of inflammatory diseases are still not completely understood, control of inflammation during the clinical presentation of diseases seems to alleviate the negative effects on reproductive biology. It is increasingly evident that animal health, not only at the time of breeding or pregnancy development but also in the period preceding breeding, is imperative for optimal reproduction in cattle and should always be considered in herd evaluations and managerial decisions.(AU)

Postpartum uterine infection and endometritis in dairy cattle

After parturition, uterine involution, regeneration of the endometrium, return of ovarian cyclic activity, and the control of pathogenic bacteria in the uterus is required before cows are likely to conceive again. However, pathogenic bacteria often cause uterine disease in modern dairy cattle, leading to decreased productivity and reduced fertility. This review aims to provide an overview of postpartum uterine infection and disease in dairy cattle. Metritis and endometritis are the main postpartum clinical conditions; although, subclinical endometritis is an emerging issue. Postpartum uterine disease is associated with the isolation of Escherichia coli, Trueperella pyogenes, and anaerobic pathogenic bacteria. Sensing of bacteria or their pathogen-associated molecules, such as lipopolysaccharide, by the innate immune system generates inflammatory responses. Endometrial inflammation includes increased expression of complement, calgranulins, interleukins and acute phase proteins, as well as the chemotaxis of neutrophils and macrophages to the site of infection. Uterine disease is also characterised by tissue damage, including endometrial cytolysis caused by the cholesterol-dependent cytolysin, pyolysin. The responses to pathogens are energetically expensive, and depletion of the key cellular nutrients, glucose or glutamine, impairs inflammatory responses by endometrial tissues. For sustainable intensification of the dairy industry over the next 50 years, it is vital to understand why high-milk-yield cows are so susceptible to uterine pathology and develop new ways to prevent uterine disease.

Postpartum uterine infection and endometritis in dairy cattle

After parturition, uterine involution, regeneration of the endometrium, return of ovarian cyclic activity, and the control of pathogenic bacteria in the uterus is required before cows are likely to conceive again. However, pathogenic bacteria often cause uterine disease in modern dairy cattle, leading to decreased productivity and reduced fertility. This review aims to provide an overview of postpartum uterine infection and disease in dairy cattle. Metritis and endometritis are the main postpartum clinical conditions; although, subclinical endometritis is an emerging issue. Postpartum uterine disease is associated with the isolation of Escherichia coli, Trueperella pyogenes, and anaerobic pathogenic bacteria. Sensing of bacteria or their pathogen-associated molecules, such as lipopolysaccharide, by the innate immune system generates inflammatory responses. Endometrial inflammation includes increased expression of complement, calgranulins, interleukins and acute phase proteins, as well as the chemotaxis of neutrophils and macrophages to the site of infection. Uterine disease is also characterised by tissue damage, including endometrial cytolysis caused by the cholesterol-dependent cytolysin, pyolysin. The responses to pathogens are energetically expensive, and depletion of the key cellular nutrients, glucose or glutamine, impairs inflammatory responses by endometrial tissues. For sustainable intensification of the dairy industry over the next 50 years, it is vital to understand why high-milk-yield cows are so susceptible to uterine pathology and develop new ways to prevent uterine disease.(AU)

FATORES QUE INTERFEREM NA PREVALÊNCIA DE ENDOMETRITE CITOLÓGICA E PRODUÇÃO DE EMBRIÕES IN VIVO EM VACAS GIROLANDO

Existem vários entraves que podem prejudicar a lucratividade da produção leiteira, entre eles estão as doenças uterinas que acometem os animais durante o período pós parto. Elas podem ser ocasionadas pela entrada de microrganismos patogênicos, capazes de romper as barreiras físicas de defesa do útero e adentrar o tecido endometrial liberando toxinas e ativando os mecanismos da imunidade inata, que leva a uma resposta inflamatória. Entre essas doenças que estão relacionadas a presença de microrganismos estão a metrite que pode ocorrer nos primeiros 10 dias após o parto, o animal pode apresentar uma descarga vermelho acastanhada com odor muito fétido, geralmente pode desenvolver sintomas clínicos sistêmicos como febre, infecção generalizada e se não receber o tratamento adequando pode vir a óbito e a endometrite clínica que é uma inflamação do endométrio que pode ocorrer após 21 dias pós parto, caracterizada por descarga muco purulenta ou purulenta. Ambas as doenças são capazes de comprometer a fertilidade, e eficiência reprodutiva dos rebanhos. A endometrite citológica apesar de não possuir sinais clínicos e o animal apresentar descarga vaginal com muco limpo, está relacionada com o desequilíbrio da resposta imune e permanência de células polimorfonucleares no tecido endometrial após 21 dias pós parto. O limiar para considerar os animais positivos pode variar de 4 a 18%. Essas células ao exercerem sua função de defesa, modificam o PH uterino quando liberam espécies reativas de oxigênio que comprometem as taxas de fecundação, qualidade de oocitos e embriões, impactando diretamente nos índices de taxa de prenhez. O diagnóstico da metrite e endometrite clínica pode ser feito por meio da avaliação da descarga vaginal com o dispositivo metricheck®.

There are several obstacles that can harm the profitability of dairy production, among them are the uterine diseases that affect animals during the postpartum period. They can be caused by the entry of pathogenic microorganisms, capable of breaking the physical barriers of defense of the uterus and entering the endometrial tissue releasing toxins and activating the mechanisms of innate immunity, which leads to an inflammatory response. Among these diseases that are related to the presence of microorganisms are metritis that can occur in the first 10 days after birth, the animal may present a brownish red discharge with a very foul odor, generally it can develop systemic clinical symptoms such as fever, generalized infection and if not receiving adequate treatment may result in death and clinical endometritis, which is an inflammation of the endometrium that can occur after 21 days postpartum, characterized by purulent or purulent mucus discharge. Both diseases are capable of compromising fertility, and reproductive efficiency of herds. Cytological endometritis, despite having no clinical signs and the animal has vaginal discharge with clean mucus, is related to the imbalance of the immune response and the permanence of polymorphonuclear cells in the endometrial tissue after 21 days postpartum. The threshold for considering animals positive can vary from 4 to 18%. These cells, when exercising their defense function, modify the uterine PH when they release reactive oxygen species that compromise fertilization rates, quality of oocytes and embryos, directly impacting the pregnancy rate indexes. The diagnosis of metritis and clinical endometritis can be made by assessing vaginal discharge with the Metricheck® device. For the diagnosis of cytological endometritis, techniques with gynecological brush, low volume washing, or endometrial biopsy can be used

Modulação da resposta inflamatória no útero de éguas tratadas com plasma rico em plaquetas

A endometrite é a causa mais comum de subfertilidade em éguas. Todas as éguas apresentam uma inflamação endometrial fisiológica pós-cobertura; entretanto, 15-20% das éguas têm endometrite persistente pós-cobertura (EPPC). Recentemente, o plasma rico em plaquetas (PRP) vêm se tornando popular no manejo reprodutivo de éguas susceptíveis a EPPC. Atualmente, não existe uma padronização dos métodos de preparação do PRP para uso intrauterino em éguas. Desta forma, este estudo teve como objetivo comparar três métodos de preparação de PRP para infusão intrauterina em éguas e avaliar a resposta inflamatória pós-cobertura, cultura endometrial e recuperação embrionária em éguas suscetíveis a EPPC tratadas com PRP. No primeiro estudo, o PRP foi produzido por três métodos. Método 1, o sangue foi coletado em bolsa de transfusão, transferido para tubos Falcon e duplamente centrifugadas; Método 2, sangue foi coletado em tubos vacutainer e centrifugado uma única vez; Método 3 sangue foi coletado em uma seringa de 60 mL contendo anticoagulante, a qual foi mantida em posição vertical para sedimentar por 4h. Após o processamento, a contagem de células e a viabilidade plaquetária foram avaliadas. Em um subgrupo de éguas (n = 6), o PRP foi avaliado 6 e 24h pós-refrigeração a 5 °C. No estudo 2, éguas (n = 12) suscetíveis a EPPC tiveram três ciclos atribuídos aleatoriamente para receber infusões intrauterinas de LRS (controle), ou PRP autólogo ou plasma pobre em plaquetas (PPP) antes (48 e 24h) e após a inseminação artificial (IA) (6 e 24h). Tanto o PRP quanto o PPP foram obtidos pelo Método 1 no experimento 2. As éguas foram inseminadas com sêmen fresco de um garanhão com fertilidade conhecida. O acúmulo de líquido intrauterino (IUF) e as células polimorfonucleares endometriais (PMNs) foram avaliados a cada 24h até 96h pós-inseminação. Citocinas uterinas (Il, IL6, CXCL8 e IL10) foram avaliadas antes (0h), 6 e 24h após IA, e cultura endometrial três e nove dias após IA. A lavagem do embrião foi realizada 8 dias após a ovulação. Os dados foram analisados com modelo misto, teste post-hoc de Tukeys e regressão multivariada. No estudo 1, o Método 1 resultou no maior e o método 3 nas menores concentrações de plaquetas, e o último teve maior concentração de leucócitos que os outros (P<0,05). A viabilidade das plaquetas foi semelhante entre os tratamentos. O resfriamento por 24h não afetou a contagem de plaquetas. No entanto, a viabilidade plaquetária foi reduzida após o resfriamento do PRP produzido pelo método 3. Estudo 2, o tratamento com PRP reduziu os PMNs endometriais, IUF pós-reprodução e citocinas pró-inflamatórias quando comparados aos ciclos atribuídos pelo controle, mas não significativamente diferente do PPP. Os controles tiveram uma porcentagem significativamente maior de culturas bacterianas positivas (33%) em comparação com os ciclos atribuídos a PRP (0%), enquanto os ciclos tratados com PPP não foram significativamente diferentes dos outros grupos (25%). Os ciclos atribuídos ao PRP tiveram taxas de recuperação embrionária significativamente maiores (83%) do que o controle (33%), embora não significativamente diferente do PPP (60%). Em conclusão, o Método 1 resultou nas maiores concentrações de plaquetas, enquanto o método 3 resultou em maiores contagens de leucócitos. O resfriamento afetou a viabilidade das plaquetas no PRP obtido com o método 3. Resta determinar se os diferentes métodos e o resfriamento afetariam a eficácia clínica do PRP. A infusão de plasma reduziu a duração e intensidade da resposta inflamatória pós-IA e melhorou a recuperação embrionária em éguas suscetíveis a EPPC. As plaquetas parecem ter uma propriedade antimicrobiana e anti-inflamatória dependente da dose

Endometritis is the most common cause of subfertility in mares. All mares display a physiological post-breeding endometrial inflammation; however, 15-20% of mares have persistent breeding-induced endometritis (PBIE). Recently, platelet-rich plasma (PRP) is becoming popular in mare practice to mitigate post-breeding induced endometritis and improve fertility. Currently, there is no standardization of methods to prepare PRP for intrauterine use in mares. This study aimed to compare three methods to prepare PRP for intrauterine infusion in mares and to assess the post-breeding inflammatory response, endometrial culture and embryo recovery in mares susceptible to PBIE treated with PRP. In the first study, PRP was produce by three methods. Method 1, blood was collected in blood transfusion bag, transferred to Falcon tubes and double centrifuged; Method 2, blood was collected in vacutainer tubes and centrifuged once; Method 3, blood was collected in a syringe containing anticoagulant, which was kept in an upright-position to sediment for 4h. After processing, cell counts and platelet viability were assessed. In a subset of mares (n=6), PRP was evaluated at 6 and 24h post-cooling at 5°C. In study 2, mares (n=12) susceptible to PBIE had three cycles randomly assigned in a crossover design to receive intrauterine infusions of LRS (control), or autologous PRP or platelet-poor plasma (PPP) pre- (48 and 24h) and post-breeding (6 and 24h). Both PRP and PPP were obtained by Method 1. Mares were bred with fresh semen from one stallion. Intrauterine fluid accumulation (IUF) and endometrial polymorphonuclear cells (PMNs) were assessed every 24h up to 96h post-breeding. Uterine cytokines (Il, IL6, CXCL8 and IL10) were evaluated before (0h), 6 and 24h post-breeding, and endometrial culture three and nine-days after breed. Embryo flushing was performed 8d post-ovulation. Data were analyzed with mixed model, Tukeys post-hoc test, and multivariate regression. In study 1, Method 1 resulted in the greatest and method 3 in the fewest platelet concentrations, and the latter had greater WBC than the others (P<0.05). Platelet viability was similar across treatments. Cooling for 24h did not affect platelet counts. However, platelet viability was reduced after cooling PRP produced by method 3. Study 2, PRP treatment reduced endometrial PMNs, post-breeding IUF and pro-inflammatory cytokines when compared to control-assigned cycles, but not significantly different than PPP. Controls had a significantly higher percentage of positive bacterial cultures (33%) in comparison to PRP-assigned cycles (0%), whereas cycles treated with PPP were not significantly different from the other groups (25%). The PRP-assigned cycles had significantly higher embryo recovery rates (83%) than the control (33%), though not significantly different than PPP (60%). In conclusion, Method 1 resulted in the greatest platelet concentrations, while method 3 resulted had greater WBC counts. Cooling affected platelet viability in PRP obtained with method 3. It remains to be determined whether the different methods and cooling would affect PRP's clinical efficacy. Plasma infusion reduced the duration and intensity of the post-breeding inflammatory response and improved embryo recovery in mares susceptible to PBIE. Platelets incrementally downregulate PBIE and appear to have a dose-dependent antimicrobial property.