Overview of the Haematological Effects of COVID-19 Infection.

From its early origins, COVID-19 has spread extensively and was declared a global pandemic by the World Health Organization in March of 2020. Although initially thought to be predominantly a respiratory infection, more recent evidence points to a multisystem systemic disease which is associated with numerous haematological and immunological disturbances in addition to its other effects. Here we review the current knowledge on the haematological effects of COVID-19.

Effects of varying doses of ß-nerve growth factor on the timing of ovulation, plasma progesterone concentration and corpus luteum size in female alpacas (Vicugna pacos).

Ovulation in camelids is induced by the seminal plasma protein ovulation-inducing factor (OIF), recently identified as ß-nerve growth factor (ß-NGF). The present study measured the total protein concentration in alpaca seminal plasma using a bicinchoninic acid (BCA) protein quantification assay and found it to be 22.2±2.0mgmL(-1). To measure the effects of varying doses of ß-NGF on the incidence and timing of ovulation, corpus luteum (CL) size and plasma progesterone concentration, 24 female alpacas were synchronised and treated with either: (1) 1mL 0.9% saline (n=5); (2) 4µg buserelin (n=5); (3) 1mg ß-NGF protein (n=5); (4) 0.1mg ß-NGF (n=5); or (5) 0.01mg ß-NGF (n=4). Females were examined by transrectal ultrasonography at 1-2-h intervals between 20 and 45h after treatment or until ovulation occurred, as well as on Day 8 to observe the size of the CL, at which time blood was collected to measure plasma progesterone concentrations. Ovulation was detected in 0/5, 5/5, 5/5, 3/5 and 0/4 female alpacas treated with saline, buserelin, 1, 0.1 and 0.01mg ß-NGF, respectively. Mean ovulation interval (P=0.76), CL diameter (P=0.96) and plasma progesterone concentration (P=0.96) did not differ between treatments. Mean ovulation interval overall was 26.2±1.0h. In conclusion, buserelin and 1mg ß-NGF are equally effective at inducing ovulation in female alpacas, but at doses ≤0.1mg, ß-NGF is not a reliable method for the induction of ovulation.

Rumen-protected methionine supplementation and fibre production in alpacas (Vicugna pacos).

Sulphur-containing amino acids are a crucial requirement for fibre production and may be supplemented in the diet of fibre-producing animals to stimulate fibre growth. The alpaca fibre industry is a developing industry in Australia with high variability in fibre production. To date, there is no evidence whether supplementing the diet of alpacas with sulphur amino acids improves fibre production. We hypothesised that supplementation with the rumen-protected sulphur amino acid, methionine would increase fibre growth in alpacas. Three groups of eight huacaya alpaca wethers were fed daily a maintenance diet supplemented with 0, 2 or 4 g of rumen-protected methionine for 7 weeks. Fibre samples were taken at the beginning and end of the study with a blood sample taken by jugular venipuncture prior to feeding on the first day of each week. Methionine supplementation had no effect on fibre diameter (p = 0.92), fibre length (p = 0.91) or fibre yield (p = 0.33). The change of season over the study affected plasma glucose (p < 0.001), plasma urea nitrogen (p < 0.001) and fibre diameter (p < 0.001). The indifference between groups may be due to the maintenance diet supplying sufficient levels of methionine, the lack of genetic potential of the experimental animals to respond to additional methionine or that the supplemental methionine was not protected in alpacas and deaminated for glucose production.

Successful vitrification of dromedary camel embryos using a novel embryo vitrification kit.

Embryos (n = 87) collected 8 days after mating and 7 days after ovulation were vitrified using a camel-specific vitrification kit. Vitrification solutions (VS) containing 20% foetal calf serum, with or without 2% bovine serum albumin (BSA) were used to cryopreserve embryos, in three steps VS1 (5 min), VS2 (5 min) and VS3 (30-35 s) at room temperature (RT) before being loaded into open pulled straws and immediately frozen in liquid nitrogen. Embryos were subsequently thawed in warming solutions (WS) in three steps: WS1 at 37 °C (1 min), WS2 at RT (5 min) then into holding media at RT (5-60 min) prior to transfer, in pairs, into recipient camels 6 days after ovulation. There were 42 of 43 embryos viable after vitrification in media without BSA and these were subsequently transferred into 21 recipient females which resulted in ten pregnancies 60 days after transfer (48% pregnancy rate). There were 38 of 44 viable embryos vitrified in media containing BSA that were transferred in pairs into 19 recipient females which resulted in five pregnancies 60 days after transfer (26% pregnancy rate; P > 0.05). Of the total 15 foetuses that developed to 60 days of gestation after vitrification, 11 resulted from embryos of 200-499 µm diameter and four from embryos of 500-700 µm diameter (P > 0.05). There were encouraging results with use of this novel vitrification kit for the commercial application of cryopreservation of camel embryos with a diameter of 300-550 µm.

Effect of diluent type, cryoprotectant concentration, storage method and freeze/thaw rates on the post-thaw quality and fertility of cryopreserved alpaca spermatozoa.

This study compared protocols for cryopreservation of ejaculated, papain-treated alpaca spermatozoa. This included different concentrations of egg yolk (EY; 5, 10 or 15%) and glycerol (2, 5 or 10%), diluent types (SHOTOR, lactose, skim milk or INRA-96™), freeze rates (2, 4 or 8 cm above liquid nitrogen; LN), thaw rates (37 °C for 1 min or 42 °C for 20 sec) and storage vessels (pellets, 0.25 mL straws or 0.5 mL straws). Spermatozoa were assessed pre-freeze and 0, 30, 60 and 90 min post-thaw. Forty-one hembras were inseminated with either fresh, papain-treated or frozen-thawed spermatozoa. Motility was affected by EY concentration (P < 0.001), diluent type (P < 0.001), freeze rate (P = 0.003) and storage vessel (P = 0.001). Viability was affected by EY concentration (P < 0.001), diluent type (P < 0.001), storage vessel (P = 0.002) and thaw rate (P = 0.03). For artificial insemination (AI), semen was diluted 1:3 in a lactose-based diluent, with 5% EY and glycerol. Freezing was in 0.5 mL straws, 2 cm above LN for 4 min then thawing at 37 °C for 1 min. Pregnancy rates of those ovulated (n = 26) were not different (1/5 fresh, 1/4 papain-treated, 0/17 frozen-thawed; P = 0.10). Pregnancy can be achieved after AI with papain-treated spermatozoa. Further work is needed to determine the optimal dose, timing and location for insemination.

The prevalence of HIV seropositivity and associated cytopenias in full blood counts processed at an academic laboratory in Soweto, South Africa.

BACKGROUND: The HIV epidemic in South Africa (SA) has had a substantial impact on laboratory services, at least partially owing to the well-described propensity to cytopenias in HIV-positive patients. OBJECTIVES: (i) To formally gauge the impact of HIV infection on the state sector haematology services in SA by determining the HIV seropositivity rate among full blood counts (FBCs) performed at a large academic state sector laboratory; and (ii) to document the prevalence of cytopenias among HIV-positive patients in this setting. METHODS: Randomly selected FBCs submitted to the National Health Laboratory Service laboratory at Chris Hani Baragwanath Academic Hospital, Johannesburg, were extracted from the laboratory information system (LIS) and retrospectively reviewed. HIV test results and other pertinent information in the LIS were documented, as was the presence of any cytopenias. RESULTS: HIV status was documented in 561 of 1 006 samples (55.8%), with 307 (54.7%) of these being HIV-positive. Of the HIV-positive patients, 63.2% had one or more cytopenia/s. Anaemia was present in 183/307 (59.6%) of the HIV-positive patients, and was severe (haemoglobin <8 g/dL) in 32/307 (10.4%). Multivariate linear regression analysis showed significant independent associations between the presence of anaemia and both immunological AIDS (iAIDS) (p<0.0001) and male sex (p<0.025), but not HIV viral load (VL) (p=0.33) or antiretroviral therapy (ART) exposure (p=0.70). Thrombocytopenia and neutropenia were present in 37/307 (12.1%) and 11/51 (21.6%) of the HIV-positive patients, respectively, with no statistically significant association between either of these cytopenias and iAIDS, exposure to ART or VL. CONCLUSIONS: The findings reflect the substantial impact of the HIV epidemic on state sector laboratory resources, particularly the haematology service.

Serological evidence of Neospora caninum in alpacas from eastern Australia.

BACKGROUND: The aim of this study was to investigate if there was any serological evidence of Neospora caninum in alpaca populations in south-eastern Australia. METHODS AND RESULTS: Serum samples from 100 alpacas were collected from four farms. All serum samples were screened for N. caninum antibodies using a commercially available competitive ELISA. Of the 100 alpacas sampled, 3 were suspect seropositive for N. caninum. CONCLUSION: There is natural N. caninum seroprevalence in alpacas in south-eastern Australia; however, it remains undetermined whether or not this infection is currently contributing to reproductive failure in alpacas in Australia.

Prevalence and causes of thrombocytopenia in an academic state-sector laboratory in Soweto, Johannesburg, South Africa.

BACKGROUND: Causes of thrombocytopenia range from laboratory errors to life-threatening pathological conditions. To establish the cause, appropriate laboratory investigation is required. OBJECTIVES: To determine the prevalence and causes of platelet counts <100 × 10(9)/L in state health facilities in Johannesburg, South Africa, as well as the quality of the subsequent laboratory work-up in this setting. METHODS: Full blood counts (FBCs) performed on 7 randomly selected days at the National Health Laboratory Service laboratory at Chris Hani Baragwanath Academic Hospital were retrospectively reviewed. Samples with platelet counts <100 × 109/L were identified, and pertinent information was extracted from the laboratory database. RESULTS: Of 4 456 FBCs included, 381 (8.6%) had a platelet count of <100 × 10(9)/L. Thrombocytopenia prevalence rates were high in haematology/oncology wards (34.4%), intensive care units (20.5%) and medical wards (18.7%) and among neonatal inpatients (16.5%), and were lowest in outpatient clinics (1-2%). A cause was apparent in ~60% of patients, the commonest causes being chemotherapy and sepsis (each comprising >20% of the recognised causes). Spurious thrombocytopenia, disseminated tuberculosis, aplastic anaemia, immune thrombocytopenia and malignant marrow infiltration each accounted for 5 - 10% of the causes, while malaria, thrombotic thrombocytopenic purpura, HIV effect and liver disease were each identified in <5% of cases. HIV status was documented in ~70% of the patients, of whom ~50% tested positive. The quality of the laboratory work-up showed differences between specialties within the hospital setting, and was poorest in the primary healthcare clinic sector. CONCLUSION: Thrombocytopenia is common in hospitalised patients in the Johannesburg academic state sector. Differences in the quality of the laboratory work-up emphasise the need for a standardised approach to thrombocytopenia investigation and increased awareness among clinicians.

Ovarian follicular wave characteristics in alpacas.

The objectives were to describe in detail ovarian follicular growth characteristics and to establish the interval between successive large follicles in unmated alpacas. The ovarian follicular status of 16 non-pregnant, non-lactating mature alpacas was recorded using ultrasound every second day for between 46 and 100 days. An inverse relationship was observed between the diameter of the largest follicle and the total number of follicles indicating that follicular growth in alpacas occurs in waves. There were 15/38 (39%) inter-wave intervals of 12 days and 12/38 (32%) intervals of 16 days. The maximum follicular diameter in each follicular wave was 8.8 +/- 0.3 mm (n=38). Inter-wave intervals of longer duration were associated with a larger maximum follicle diameter (P<0.001). However, the growth rate of dominant follicles was consistent over the first 10 days after emergence. They reached a diameter capable of ovulation by this time, regardless of subsequent inter-wave interval. The latter observation suggested that the optimal time of mating might be predicted in alpacas, provided that the emergence of ovarian follicular waves was controlled.

Anthelmintic efficacy of ivermectin in naturally parasitized cats.

Ivermectin was administered per os [( PO]; n = 15) or subcutaneously [( SC]; n = 3) to naturally parasitized cats at 10 (n = 6), 100 (n = 6), or 300 (n = 6) micrograms/kg of body weight. Nontreated control cats were given sterile isotonic saline solution PO (n = 5) or SC (n = 1). Qualitative fecal examinations were performed on each cat 1 day before treatment and 14 days after treatment. Cats were euthanatized 14 days after treatment, at which time parasites from the gastrointestinal tracts were recovered, identified, and enumerated. Lungs and urinary bladders were examined histologically or by digestion (lungs only) for Capillaria spp and/or Aelurostrongylus abstrusus. Ivermectin was effective in removing Ancylostoma spp at all doses, but removal of Toxocara cati required 300 micrograms of ivermectin/kg. Efficacies against A abstrusus, Capillaria spp, and Physaloptera spp could not be determined definitively. Ivermectin had no effect on Dipylidium caninum, Hydatigera taeniaeformis, Spirometra mansonoides, or Isospora spp. Adverse reactions were not observed in cats given ivermectin PO; however, 3 cats given ivermectin SC reacted as though they experienced pain at the injection site.

Efficacy of lufenuron against developmental stages of fleas (Ctenocephalides felis felis) in dogs housed in simulated home environments.

Twenty-four, adult, female Beagles were arranged by body weight from greatest to least and allocated to 2 groups of 12 dogs, using random numbers. Dogs were housed collectively in 2 adjacent metal buildings, each divided into 4 rooms measuring 2.1 x 3.7 m. Each room was paneled and carpeted and had an access door to the outside with a connecting run that measured 2.1 x 9.1 m. Each run had a surface consisting of 5 cm of pea gravel overlaying 5 cm of sand, and was partially covered by an awning that provided shade at its proximal end. For placement in room/run units, dogs in each of the treated and control groups were alloted to 4 subgroups of 3 dogs each. Each subgroup of dogs was placed in a separate room/run unit. Units containing treatment or control subgroups were alternated to avoid placing identically treated subgroups adjacent to each other. Dogs of subgroups A, C, E, and G were treated with lufenuron monthly at a minimal target dosage of 10 mg/kg of body weight; those of subgroups B, D, F, and H were treated with excipient tablets. Dogs were treated on study days 7, 37, 68, and 98. Each dog was infested with 100 newly emerged, unfed, insectary-reared, adult Ctenocephalides felis on each of study days 0 and 2. Thereafter, infestations on all dogs were dependent on continued development of fleas either in the indoor or outdoor environment. Numbers of fleas on each of the treated and control dogs were determined, using a nondestructive counting technique on days 6, 14, 21, 28, 35, 56, 70, 84, 98, 112, and 119.(ABSTRACT TRUNCATED AT 250 WORDS)

Efficacy dosage titration of lufenuron against developmental stages of fleas (Ctenocephalides felis felis) in cats.

Thirty-two mixed-breed male and female cats were blocked by sex, arranged by body weight from greatest to least, and allocated to 4 groups of 8 (4 male, 4 female) cats, using random numbers. Cats in each of 3 groups were treated orally with a 7% suspension formulation of lufenuron at dosage of 15, 30, or 45 mg/kg of body weight. Cats in the fourth group were treated orally with an excipient suspension without lufenuron. Cats were infested with newly emerged, unfed cat fleas (Ctenocephalides felis felis) on days -7 and -3 before treatment and at approximately weekly intervals after treatment. Flea eggs were collected from beneath each cat on selected days before and after treatment and placed in an artificial rearing medium. Flea eggs and medium were kept for 35 days in an insectary to determine effects of lufenuron or excipient suspension on emergence of adults of the F1 generation. Lufenuron was 100% effective in inhibiting development of C felis at all dosages for 11 days after treatment. Thereafter, efficacy exceeded 92% in all dosages groups. On day 32, when the study was terminated, efficacy for each of the dosage groups was: 15 mg/kg, 95.2%; 30 mg/kg, 98.2%; and 45 mg/kg, 99.6%. Adverse reactions or side effects were not observed in cats, regardless of treatment dosage.

Dose titration of an injectable formulation of lufenuron in cats experimentally infested with fleas.

OBJECTIVES: To identify the lowest single dose of lufenuron injected s.c. that results in a 90% disruption of the flea (Ctenocephalides felis) life cycle for 6 months in cats. ANIMALS: 40 domestic shorthair cats (20 males, 20 females) between 5 and 7 months old. PROCEDURE: Cats were randomly assigned to 1 of 5 eight-cat groups and experimentally infested with C. felis on days -8, -7, -6, and -4. On day 0, cats in the 4 treatment groups were treated with an injectable formulation of lufenuron at doses of 2.5, 5, 10, or 20 mg/kg of body weight, respectively. Control cats received the injectable formulation without lufenuron. Experimental infestations were repeated and flea eggs collected at various intervals for 196 days after treatment. Eggs were placed in media and incubated in an insectary for 28 days to determine effects of injectable lufenuron on egg and larval development. Number of adults that emerged from eggs were compared among groups. RESULTS: Lufenuron injected once at a dose of 10 or 20 mg/kg, but not at 2.5 or 5 mg/kg, resulted in a 90% decrease in number of adult fleas emerging from eggs for 196 days after treatment. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that control of flea egg and larval development for at least 6 months can be achieved in cats with a single s.c. injection of lufenuron (10 mg/kg). The injectable formulation may provide veterinarians and cat owners an alternative to the tablet formulation of lufenuron.

Efficacy of milbemycin oxime against naturally acquired or experimentally induced Ancylostoma spp and Trichuris vulpis infections in dogs.

The efficacy of milbemycin oxime was evaluated at dosages of 0.25, 0.50, and 0.75 mg/kg of body weight in dogs naturally infected with mature Ancylostoma spp, at a dosage of 0.50 mg/kg in dogs with experimentally induced immature and mature A caninum, and at dosages of 0.55 to 0.86 mg/kg in dogs naturally infected with mature Trichuris vulpis. Milbemycin oxime was 95 and 99% effective against mature Ancylostoma spp at dosages of 0.50 and 0.75 mg/kg, respectively, but only 49% effective at a dosage of 0.25 mg/kg. Efficacy was 49% against pulmonary L3-L4 stages of A caninum (36 hours after inoculation), greater than 80% against L4 (120 hours after inoculation) and early L5 stages (216 hours after inoculation), and greater than 90% against experimentally induced mature stages (360 hours after inoculation). Milbemycin oxime was also 97% effective in the removal of mature Tr vulpis from naturally infected dogs. Adverse reactions were not observed following treatment in any of the dogs.

Fasciola hepatica infection in farmed emus (Dromaius novaehollandiae).

OBJECTIVE: To describe two cases of infection with Fasciola hepatica in young farmed emus, subacute and chronic fasciolosis and a response to treatment of the flock with albendazole. PROCEDURE: Gross lesions were found at necropsy and hepatic lesions in microscopic examination. The parasite recovered from one emu was identified by its morphological characteristics and an egg count reduction test was carried out after treatment of the flock with albendazole. RESULTS: Hepatic lesions resembling subacute and chronic fasciolosis of ruminants were identified. An adult fluke was recovered from the liver of one of the birds and was identified as F hepatica. The eggs of the fluke were irregular in shape and size. No fluke eggs were identifiable in faeces of live emus 10 days after treatment of the flock with albendazole at a dose of 10 mg/kg. CONCLUSIONS: This is the first reported case of infection with F hepatica in farmed emus and the first report of the occurrence of Fasciola infection is the class Aves. The irregular shape and size of the eggs may be attributable to infection of an aberrant host. Treatment with albendazole eliminated eggs from the faeces of the flock.

Congenital caudal vertebral malformations in the alpaca (Lama pacos).

OBJECTIVE: To assess radiologically the caudal vertebrae of a male alpaca with an abnormal tail, and nine of his offspring. PROCEDURE: Right lateral and ventrodorsal radiographs were taken of the caudal vertebrae. RESULTS: The caudal vertebral abnormalities in these alpacas included vertebral subluxation, hemivertebrae, wedge hemivertebrae and block vertebrae. CONCLUSION: These malformations in offspring of one sire suggest a hereditary basis for congenital caudal vertebral malformations in the alpaca.

Effects of mating behaviour and the ovarian follicular state of female alpacas on conception.

OBJECTIVE: To determine relationships between mating behaviour, ovarian follicular state and successful conception in receptive female alpacas. PROCEDURE: Seventy pen matings were observed at a commercial alpaca stud in south-western Victoria. The behaviours observed included time taken to assume sternal recumbency, mating duration, and evidence of nonreceptive behaviour such as spitting, kicking and vocalisation. Ovarian follicular state was determined by ultrasonography, which was complemented by measuring plasma concentrations of oestradiol and progesterone. Pregnancies were confirmed by transabdominal ultrasonography between days 45 and 80 after mating. RESULTS: There were no significant differences between receptive females that conceived and those that failed to conceive in the time taken to adopt the copulation position of sternal recumbency, mating duration, or maximum follicle diameter. There was no significant relationship between time taken to assume sternal recumbency (log10) and maximum follicle diameter or plasma oestradiol (log10). However, there was a significant quadratic relationship between plasma oestradiol concentration (log10) and follicle diameter, and the probability of pregnancy increased as the plasma concentration of oestradiol (log10) at the time of mating increased. Females were sexually receptive most of the time in the absence of a corpus luteum, and regardless of size of the largest follicle or plasma concentration of oestradiol. Breed (Huacaya vs Suri), site of the dominant follicle (left or right ovary), lactation state, number of matings by the male (1 or 2), or interval between parturition and mating, did not affect pregnancy outcome. Follicles with a diameter less than 7 mm were able to ovulate in response to mating. This was smaller than previously reported. Thirty-four pregnancies (49% pregnancy rate) resulted in 30 (88%) births with a gestation length of 343 days (SEM +/- 2, range 316-367 days). There were 4 (12%) abortions between days 45 and 80 of gestation and full term. CONCLUSION: It was not possible to correlate mating behaviour and ovarian state with conception. To optimise pregnancy rates in receptive alpacas, matings need to occur in the presence of an oestrogenic follicle that is capable of ovulation in response to mating. A simple method of detecting alpacas with follicles in this state is not currently available and treatments that control ovarian follicular growth should therefore be investigated.

Relative kinetic frictional forces between sintered stainless steel brackets and orthodontic wires.

The level of kinetic frictional forces generated during in vitro translation at the bracket-wire interface were measured for two sintered stainless steel brackets as a function of two slot sizes, four wire alloys, and five to eight wire sizes. The two types of sintered stainless steel brackets were tested in both 0.018-inch and 0.022-inch slots. Wires of four different alloy types, stainless steel (SS), cobalt chromium (Co-Cr), nickel-titanium (Ni-Ti), and beta-titanium (beta-Ti), were tested. There were five wire sizes for the 0.018-inch slot and eight wire sizes for the 0.022-inch slot. The wires were ligated into the brackets with elastomeric ligatures. Bracket movement along the wire was implemented by means of a mechanical testing instrument, and time dependent frictional forces were measured by a load cell and plotted on an X-Y recorder. For most wire sizes, lower frictional forces were generated with the SS of Co-Cr wires than with the beta-Ti or Ni-Ti wires. Increase in wire size generally resulted in increased bracket-wire friction. There were no significant differences between manufacturer for the sintered stainless steel brackets. The levels of frictional force in 0.018-inch brackets ranged from a low of 46 gm with 0.016-inch Co-Cr wire to a high of 157 gm with 0.016 x 0.025-inch beta-Ti wire. In comparing the data from a previous study by Kapila et al. 1990 performed at OUHSC with the same apparatus, the friction of sintered stainless steel brackets was approximately 40% to 45% less than the friction of the conventional stainless steel brackets.