Transmission of feline immunodeficiency virus in domestic cats via artificial insemination.

The objective of this study was to determine whether semen from male domestic cats infected with feline immunodeficiency virus (FIV) can transmit virus to females. Twelve inseminations were performed by an intrauterine laparoscopic technique with fresh or cryopreserved electroejaculates from asymptomatic males chronically infected with the NCSU1 strain of FIV. Of six inseminations performed with fresh semen, three resulted in infection of queens, as indicated by seroconversion, expression of FIV gag provirus in peripheral blood leukocytes, and reduced peripheral CD4+/CD8+ T-lymphocyte ratios. None of the six inseminates with thawed cryopreserved semen resulted in infection. Two infected queens and one uninfected queen became pregnant. Virus was not evident in the seven offspring. We conclude that FIV can be transmitted horizontally by artificial insemination with fresh semen.     

The role of international transport of equine semen on disease transmission.

Despite the numerous benefits of having the capability to transport semen internationally, there are serious potential ramifications if that semen is contaminated with a communicable disease. BACTERIA: Many commensal bacteria colonize the exterior of the stallion penis and are not regarded as pathogenic. They may be cultured from an ejaculate. Alterations of the normal bacterial flora on the exterior genitalia may cause the growth of opportunistic bacteria such as Klebsiella pneumonia, Pseudomonas aeruginosa, Streptococcus zooepidemicus, which, if inseminated, may cause infertility in susceptible mares. Contagious equine metritis (CEM), a highly transmissible, true venereal disease of horses, is caused by the gram-negative coccobacillis, Taylorella equigenitalis. Even with the use of rigorous testing protocols, the current techniques used may not ensure accuracy of results. VIRUSES: Equine coital exanthema (equine herpes virus type 3; EHV-3) is a highly contagious virus that causes painful lesions on the stallion's penis and mare's vulva. Although it is primarily transmitted through coitus, infected fomites have also been implicated in its spread. Therefore, it is possible that the virus can potentially be transmitted to the ejaculate through penile contact with an artificial vagina or sleeve. Equine arteritis virus appears to be becoming more prevalent in recent years. The most common method of transmission is through respiratory disease, but the organism can also be shed in the semen of asymptomatic stallions. Equine infectious anemia virus has also been found to be present in the semen of an infected stallion, although no evidence exists at this time that there is venereal transmission of this disease. PROTOZOA: Dourine, caused by Trympanosoma equiperidum, is a venereal disease found only in Africa, South and Central America and the Middle East. Serological testing using complement fixation is recommended for diagnosis. Piroplasmosis, a disease caused by Babesia equi or by a less severe strain, Babesia caballi, has received a great deal of attention in recent years due to the increased transfer of horses between countries. It is considered to be enzootic in many areas of the southern US, and is found throughout the world. The protozoal agent is most often spread by ticks, but mechanical transmission has also been documented; therefore, there is concern for venereal transmission if blood from an infected horse contaminates the semen.     

Is caprine arthritis encephalitis virus (CAEV) transmitted vertically to early embryo development stages (morulae or blastocyst) via in vitro infected frozen semen?

The aim of this study was to determine, in vivo, whether in vitro infected cryopreserved caprine sperm is capable of transmitting caprine arthritis-encephalitis virus (CAEV) vertically to early embryo development stages via artificial insemination with in vitro infected semen. Sperm was collected from CAEV-free bucks by electroejaculation. Half of each ejaculate was inoculated with CAEV-pBSCA at a viral concentration of 10⁴ TCID₅₀/mL. The second half of each ejaculate was used as a negative control. The semen was then frozen. On Day 13 of superovulation treatment, 14 CAEV-free does were inseminated directly into the uterus under endoscopic control with thawed infected semen. Six CAEV-free does, used as a negative control, were inseminated intrauterine with thawed CAEV-free sperm, and eight CAEV-free does were mated with naturally infected bucks. Polymerase chain reaction (PCR) was used to detect CAEV proviral-DNA in the embryos at the D7 stage, in the embryo washing media, and in the uterine secretions of recipient does. At Day 7, all the harvested embryos were PCR-negative for CAEV proviral-DNA; however, CAEV proviral-DNA was detected in 8/14 uterine smears, and 9/14 flushing media taken from does inseminated with infected sperm, and in 1/8 uterine swabs taken from the does mated with infected bucks. The results of this study confirm that (i) artificial insemination with infected semen or mating with infected bucks may result in the transmission of CAEV to the does genital tack seven days after insemination, and (ii) irrespective of the medical status of the semen or the recipient doe, it is possible to obtain CAEV-free early embryos usable for embryo transfer.     

Pregnancy in the domestic cat after vaginal or transcervical insemination with fresh and frozen semen

The objective was to compare pregnancy rates in domestic cats using fresh semen for intravaginal artificial insemination (IVI), either at the time of hCG treatment for induction of ovulation, or 28 h later, and to compare pregnancy rates following IVI or transcervical intrauterine insemination (IUI) of frozen-thawed semen. Eighteen queens were inseminated during 39 estrus cycles. Fresh semen with 13.5+/-5.4 x 10(6) sperm (range, 6.8-22 x 10(6)) collected by electroejaculation from four male cats was used in Experiment 1, and cryopreserved semen (20 x 10(6) sperm, with 70+/-5% post-thaw motility) from one male cat was used in Experiment 2. Serum concentrations of estradiol-17beta and progesterone were determined in most queens on the day of AI and again 30-40 days later. Treatment with 100 IU of hCG 3 days after the onset of estrus induced ovulation in 95% of treated queens. Pregnancy rates to IVI with fresh semen at the time of hCG administration versus 28 h later were not different (P=0.58); overall 33% (5/15) of the queens became pregnant. For frozen-thawed semen, AI was consistently done 28h after hCG administration; IUI and IVI resulted in pregnancy rates of 41.7% (5/12), whereas no queen (0/12) became pregnant by IVI (P=0.0083). In conclusion, an acceptable pregnancy rate was obtained with frozen-thawed semen in the domestic cat by non-surgical transcervical IUI; this method might also be useful in other small felids.     

Deformed wing virus in western honey bees (Apis mellifera) from Atlantic Canada and the first description of an overtly-infected emerging queen

Deformed wing virus (DWV) in western honey bees (Apis mellifera) often remains asymptomatic in workers and drones, and symptoms have never been described from queens. However, intense infections linked to parasitism by the mite Varroa destructor can cause worker wing deformity and death within 67 h of emergence. Ten workers (eight with deformed wings and two with normal wings) and three drones (two with deformed wings and one with normal wings) from two colonies infected with V. destructor from Nova Scotia, Canada, and two newly-emerged queens (one with deformed wings and one with normal wings) from two colonies infected with V. destructor from Prince Edward Island, Canada, were genetically analyzed for DWV. We detected DWV in all workers and drones, regardless of wing morphology, but only in the deformed-winged queen. This is the first report of DWV from Atlantic Canada and the first detection of a symptomatic queen with DWV from anywhere.     

Detection of viral sequences in semen of honeybees (Apis mellifera): evidence for vertical transmission of viruses through drones

Honeybees (Apis mellifera) can be attacked by many eukaryotic parasites, and bacterial as well as viral pathogens. Especially in combination with the ectoparasitic mite Varroa destructor, viral honeybee diseases are becoming a major problem in apiculture, causing economic losses worldwide. Several horizontal transmission routes are described for some honeybee viruses. Here, we report for the first time the detection of viral sequences in semen of honeybee drones suggesting mating as another horizontal and/or vertical route of virus transmission. Since artificial insemination and controlled mating is widely used in honeybee breeding, the impact of our findings for disease transmission is discussed.     

Disease transmission in horses

Bacterial, viral and protozoal infections may cause severe reproductive losses. The present paper reviews the risk factors, clinical signs and preventive measures for the most important venereal or potential sexually transmitted diseases in horses. The stallion and use of semen for artificial insemination represent major risk factors for the transmission of bacterial contaminants of the penis, including Streptococcus equi subspecies zooepidemicus, Pseudomonas aeruginosa and Klebsiella pneumoniae, known to cause endometritis and infertility in the mare. The role of the stallion in disease transmission is also due to the non-clinical manifestation of diseases such as contagious equine metritis and equine viral arteritis. Dourine has been eradicated from many countries, but continues to be a problem in other areas of the globe. Strategies for the prevention of introduction and transmission of diseases in breeding operation are discussed.     

Viruses associated with ovarian degeneration in Apis mellifera L. queens

Queen fecundity is a critical issue for the health of honeybee (Apis mellifera L.) colonies, as she is the only reproductive female in the colony and responsible for the constant renewal of the worker bee population. Any factor affecting the queen's fecundity will stagnate colony development, increasing its susceptibility to opportunistic pathogens. We discovered a pathology affecting the ovaries, characterized by a yellow discoloration concentrated in the apex of the ovaries resulting from degenerative lesions in the follicles. In extreme cases, marked by intense discoloration, the majority of the ovarioles were affected and these cases were universally associated with egg-laying deficiencies in the queens. Microscopic examination of the degenerated follicles showed extensive paracrystal lattices of 30 nm icosahedral viral particles. A cDNA library from degenerated ovaries contained a high frequency of deformed wing virus (DWV) and Varroa destructor virus 1 (VDV-1) sequences, two common and closely related honeybee Iflaviruses. These could also be identified by in situ hybridization in various parts of the ovary. A large-scale survey for 10 distinct honeybee viruses showed that DWV and VDV-1 were by far the most prevalent honeybee viruses in queen populations, with distinctly higher prevalence in mated queens (100% and 67%, respectively for DWV and VDV-1) than in virgin queens (37% and 0%, respectively). Since very high viral titres could be recorded in the ovaries and abdomens of both functional and deficient queens, no significant correlation could be made between viral titre and ovarian degeneration or egg-laying deficiency among the wider population of queens. Although our data suggest that DWV and VDV-1 have a role in extreme cases of ovarian degeneration, infection of the ovaries by these viruses does not necessarily result in ovarian degeneration, even at high titres, and additional factors are likely to be involved in this pathology.     

Detection of deformed wing virus in the greenhouse for possible horizontal transmission of virus in honey bee colony

The honey bee Apis mellifera L. is a crucial insect in the agricultural industry and natural ecosystem by being a major pollinator. Nevertheless, honey bee population has been recently facing a decline. Among the several factors responsible for this decline, deformed wing virus (DWV) is considered a primary cause that negatively affects honey bee health. DWV is a cosmopolitan honey bee pathogen and causes morphological disadvantages in individual honey bees and colony collapse. Regarding the horizontal transmission of DWV, in addition to Varroa destructor, a well-known major vector of DWV, flowers have recently been implied as a transmission route. Therefore, in this study, we detected DWV from various substances, including flowers, honey bee feces, pupa, larva, nurse bee, surface of nurse bee, pollen collected by forager bee, and forager bee samples in four strawberry greenhouses, which could suggest the potential for the horizontal transmission of DWV in the semi-field condition. We also detected DWV in pollen collected by DWV-negative forager bees, implying that flowers can serve as a potential source of virus infection. These findings suggest that the surrounding environment such as shared floral sources affects the spread of DWV.     

Distribution of deformed wing virus within honey bee (Apis mellifera) brood cells infested with the ectoparasitic mite Varroa destructor

The distribution of deformed wing virus (DWV) in adult female Varroa destructor and in their progeny in relation to the pupal host bee was investigated to evaluate acquisition and transfer of DWV by the mites. The results clearly show that adult female mites regularly act as competent vectors of DWV, however, they do not acquire or transfer virus on all possible occasions. Mother mites may contain DWV while the pupal host remains free from overt infection and both mother mites and mite progeny may not acquire detectable amounts of DWV from an infected host bee. However, a majority of mites feeding on pupae that emerge with deformed wings will contain DWV. The data also demonstrates that both adult and immature mite progeny most likely acquire DWV from DWV-infected host bees and not from their mother mites. Possible explanations for the obtained results are discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Detection of multiple viruses in queens of the honey bee Apis mellifera L

Individual honey bee Apis mellifera L. queens were examined for the presence of six honey bee viruses including acute bee paralysis virus, chronic bee paralysis virus, black queen cell virus, deformed wing virus, Kashmir bee virus, and sacbrood virus. All viruses, except ABPV, were detected in the samples. Among queens examined for virus infections, 93% had multiple virus infections. The detection of viruses in queens raises the possibility of a vertical transmission pathway wherein infected queens can pass virus through their eggs to their offspring.     

Cryosurvival of Trichomonas vaginalis during cryopreservation of human semen

Despite a 90% cryosurvival of Trichomonas vaginalis in their growth medium trypticase yeast maltose (TYM) with DMSO, none of these parasites have previously been observed to survive during cryopreservation of infected human semen with glycerol (Andrologia 18, 323 (1986)). This could have been due to the failure of the culture method used to detect low numbers of survivors. The prospects of possible transmission of T. vaginalis by artificial insemination with cryobanked (-196 degrees C) semen prompted an investigation of the cryosurvival of this parasite in the presence of semen with the cryoprotectant glycerol, using a more sensitive culture method for viability evaluation. Semen and seminal fluid from the same 23 ejaculates, as well as culture medium, were inoculated with small clinical numbers of T. vaginalis and evaluated as to their survival before and after cryopreservation. Results indicated: (i) The highest cryosurvival of T. vaginalis (4.5%) was in cryobanked (glycerolated) semen, (ii) semen, as well as glycerol, shows cryoprotective action, and (iii) glycerol reduced survival of parasites in semen, seminal fluid, and TYM medium during exposure prior to freezing. Clinical information on infectivity of small numbers of T. vaginalis and the data presented here suggests that these organisms could be transmitted by artificial insemination with infected cryobanked human semen.     

Cuticular lipids correlate with age and insemination status in queen honeybees

Eusocial insects exhibit reproductive division of labour, in which one or a few queens perform almost all of the reproduction, while the workers are largely sterile and assist in rearing their siblings. Consequently, many of the colony’s tasks (e.g. nest construction and brood rearing) should be modulated depending on whether the queen is fertile. Here, we tested whether queens’ cuticular lipids could provide reliable signals of fertility in the honeybee Apis mellifera, as they do in other social Hymenoptera. Specifically, we tested whether cuticular lipids differ between virgin queens of different ages, and between queens exposed to different artificial insemination treatments being semen (sperm and seminal fluid), seminal fluid only or saline control. Using gas chromatography–mass spectrometry, we found 27 lipids: 21 different hydrocarbons, namely alkanes, alkenes and dienes, as well as six wax esters. The lipid profile changed dramatically in the first 10 days after eclosion, and there were differences in lipid composition between virgin and artificially inseminated queens. Insemination with semen, seminal fluid or saline did not result in distinct chemical profiles. Our findings indicate that the physical stimulus of insemination was responsible for the observed changes in the cuticular profile in honeybee queens. Our results demonstrate that cuticular lipid profiles encode information on queen age, fertility and mating status, which could in principle be utilised by workers and rival queens.     

Effects of various breeding strategies on diploid drone frequency and quantitative traits in a honey bee population

When selecting in a finite population of honeybees there is a conflict between gain in a quantitative trait and increasing homozygosity, and therefore the frequency of inviable diploid drones. The consequences when using different mating, import, and selection strategies on diploid drone frequency and genetic gain, was explored with Monte Carlo computer simulations.Within a closed population breeding structure, mass selection gave the highest genetic gain in the quantitative trait, but also the largest increase in percentage diploid drones and queens with unacceptably-low brood viability. Mass selection combined with truncation selection against queens having more than 15% diploid drones gave a comparable genetic gain and was the best strategy of the ones studied to avoid diploid drones. Within-family selection (one replacement per sib group) gave the least genetic gain, and a frequency of diploid drones comparable to random (no) selection. It was intermediate between mass selection and mass selection combined with viability selection concerning the frequency of diploid drones.Insemination with pooled and homogenized semen originating from all breeder queens (30), as compared to natural mating with 12 randomly-selected drones, had little effect on the genetic gain and on the overall frequency of diploid drones (10 to 15% by generation 20).The effect of opening the closed breeding population for the import of external queens every generation, by exchanging breeder queens of lowest performance with a corresponding number of new queens (5, 10and 15 out of 30), was also investigated. Under mass selection (natural mating as well as artificial insemination) the frequency of diploid drones and the proportion of queens discarded were reduced because of low brood viability. However, artificial insemination was superior to natural mating considering the latter criterion. If the imported queens were at the same genetic level for the quantitative trait under selection as the whole breeding population at that generation, or 10% better, the genetic gain was respectively slightly reduced and approximately maintained. If the imported queens were of inferior quality (equal to the initial population) the import of queens slowed genetic progress considerably.     

Bovine viral diarrhea virus (BVDV): epidemiologic concerns relative to semen and embryos

Artificial insemination and embryo transfer are used commonly in cattle production and exchange of germplasm between populations of cattle. If properly monitored, assisted reproductive techniques can be used to prevent the spread of infectious agents. However, these techniques potentially represent unnatural routes for transmission of diseases. Bovine viral diarrhea virus (BVDV) is broadly distributed among the world's populations of cattle. Fluids, gametes and somatic cells from infected animals are likely contaminated with the virus. Thus, use of semen or embryos from infected animals could result in spread of BVDV. This paper provides an overview of the risks of transmitting this virus by AI or production and transfer of embryos and summarizes the precautions needed to prevent such transmissions of disease from occurring.     

Queen reproductive state modulates pheromone production and queen-worker interactions in honeybees

The mandibular glands of queen honeybees produce a pheromone that modulates many aspects of worker honeybee physiology and behavior and is critical for colony social organization. The exact chemical blend produced by the queen differs between virgin and mated, laying queens. Here, we investigate the role of mating and reproductive state on queen pheromone production and worker responses. Virgin queens, naturally mated queens, and queens instrumentally inseminated with either semen or saline were collected 2 days after mating or insemination. Naturally mated queens had the most activated ovaries and the most distinct chemical profile in their mandibular glands. Instrumentally inseminated queens were intermediate between virgins and naturally mated queens for both ovary activation and chemical profiles. There were no significant differences between semen- and saline-inseminated queens. Workers were preferentially attracted to the mandibular gland extracts from queens with significantly more activated ovaries. These studies suggest that the queen pheromone blend is modulated by the reproductive status of the queens, and workers can detect these subtle differences and are more responsive to queens with higher reproductive potential. Furthermore, it appears as if insemination substance does not strongly affect physiological characteristics of honeybee queens 2 days after insemination, suggesting that the insemination process or volume is responsible for stimulating these early postmating changes in honeybee queens.     

Chill sensitivity of honey bee, Apis mellifera, embryos

Improved methods for preservation of honey bee, Apis mellifera L., germplasm would be very welcome to beekeeping industry queen breeders. The introduction of two parasites and the emergence of an antibiotic resistant disease have increased demands for resistant stock. Techniques for artificial insemination of queens are available, and semen has been cryopreserved with limited success. However, cryopreservation of embryos for rearing queens would mesh well with current practices and also provide drones (haploid males). Eggs at five ages between twenty-four hours and sixty-two hours were exposed to 0, -6.6, and/or -15 degrees C for various times, and successful hatch measured. Honey bee embryos show chill sensitivity as do other insect embryos, and the rate of chill injury increases dramatically with decrease in holding temperature. The 48 h embryos in both groups showed the greatest tolerance to chilling, although 44 h embryos were only slightly less so.