Prevalence and associated risk factors of ixodid tick species infesting cattle and sheep in Balochistan, Pakistan.

Tick infestation poses a serious threat to animal health, leading to significant losses in terms of vector-borne disease transmission, reduced live weight, lower quality hides, decreased milk production, and impaired reproduction in tropical and subtropical regions worldwide. This study aimed to determine the prevalence, seasonal variation, distribution pattern, and associated risk factors of Ixodid family tick species in the cattle and sheep population of three different districts in Balochistan, Pakistan. This study employed a convenient sampling method, collecting 4080 adult ticks from 816 cattle and sheep of various breeds, ages, and sexes. Specific morphological keys were used to identify the ticks up to the genus and species level. Among cattle, the highest prevalence was recorded for R. (B) annulatus (27.01%), followed by R. (B) microplus (24.02%), and H. anatolicum (20.54%). H. dromedarii (5.29%) was the least prevalent species observed in cattle. In the sheep population, H. anatolicum (30.34%) showed the highest prevalence, followed by H. marginatium (22.99%), and R. (B) annulatus (20.88%). H. dromedarii (6.96%) was the least prevalent species observed in sheep. The prevalence of R. (B) decoloratus, H. anatolicum and H. dromedarii was found to be significantly associated (P < 0.05) with the breed, age, and sex of both cattle and sheep. However, the presence of R. (B) annulatus, R. (B) microplus and H. marginatium tick species showed no significant association (P > 0.05) with these factors. In addition, the prevalence of ticks was higher in younger, female, and crossbred Friesian cattle compared to adults, males, and other breeds. Conversely, the prevalence of ticks was higher in adult, female and Hernai breed of sheep in the studied area. In conclusion, R. (B) annulatus and H. anatolicum are the dominant tick species infesting the cattle and sheep population in Balochistan. Consequently, this study provides valuable insights for developing practical and effective control measures against ticks and tick-borne diseases in the sheep and cattle population of Balochistan, Pakistan.

Newly optimized ELISA kit and LAT reveal significantly higher seroprevalence in sheep raised in agro-ecological zone as against range-ecological zone, with a significant association of meteorological parameters.

BACKGROUND: Toxoplasma gondii is a zoonotic and foodborne intracellular parasite capable of inducing congenital infections, stillbirths and abortions in humans and animals, globally. The consumption of undercooked or raw mutton is "one of the vital risks" for acquiring toxoplasmosis: an asymptomatic condition in healthy persons, while life-threatening in immunodeficient individuals like "HIV/AIDS" patients. OBJECTIVES: The current study has multiple objectives: to optimize a newly ELISA kit for Sheep, to find out the seroprevalence of ovine toxoplasmosis of two ecological zones of the Punjab, Pakistan through LAT and newly Optimized Sheep ELISA kit, to do the comparison of efficacies of various tests (LAT with newly Optimized ELISA kit and newly Optimized ELISA kit with commercial ELISA kit) and to determine the different meteorological parameters as the risk factors for T. gondii infection in sheep. METHODS: A cross-sectional study was conducted on 400 sheep sera, 200 were collected from sheep raised on open grazing system by local farmers in the adjoining areas of Civil Veterinary Dispensaries (CVDs) of range-ecological zone i.e. tehsil Kot Chutta (Dera Ghazi khan). Similarly, the remaining 200 were collected from agro-ecological zone i.e. tehsil Sharaqpur (Sheikhupura), to evaluate the comparative efficacy of LAT with optimized ELISA kit and newly optimized ELISA kit with commercial ELISA kit. FINDINGS: The newly ELISA kit optimized against a commercial ELISA kit was found to have 100% sensitivity, 97.6% specificity with 98% Positive Predictive Value, 100% Negative Predictive Value, Cut off value = 0.505, 28.28 LR+, 0.0104 LR-, and 2719.23 DOR. Seroprevalence of toxoplasmosis was detected significantly (P < 0.01; χ2) higher in Sharaqpur (44.5% by LAT; 35.5% by ELISA) as compared to that in Kot Chutta (39.5% by LAT; 31% by ELISA). The highest seroprevalence was seen in the sheep of the 1-2 years age group (P < 0.01; χ2), whereas the lowest in the oldest animals (≥ 4 years). Investigation of meteorological data of both the regions reveals that the zone with higher seroprevalence has relatively higher rainfall, higher humidity, lower environmental temperatures, and higher altitude as the critical factors, potentially behind the significant difference seen in seroprevalence level. The partial correlation of both tests (newly optimized ELISA kit and LAT) was 0.991 at maximum temperature in Sharaqpur while it was 0.981 in Kot Chutta. INTERPRETATION: A novel significant correlation was found between the meteorological parameters (relative humidity, minimum, maximum, and average temperatures) divided into yearly units of both the ecological zones, and year-wise seroprevalence (birth years of age-wise groups) of the corresponding regions. We hypothesize that such environmental conditions increase the risk of toxoplasmosis in grazing sheep, owing to a more favorable environment for coccidian oocyst survival. The ELISA kit optimized in this study will be helpful for the detection of seroprevalence of ovine toxoplasmosis in other ecological zones of Pakistan as well as of any other country in the world. More studies are recommended involving regions from other ecological zones of Pakistan to further explore the seroprevalence of ovine toxoplasmosis and to ratify the novel correlation of meteorological parameters with seroprevalence.

Molecular detection and assessment of risk factors for Theileria lestoquardi in sheep from Balochistan, Pakistan.

This study aimed to determine the molecular prevalence and associated risk factors of theileriosis in sheep from Balochistan, Pakistan. For this purpose, a total of 408 blood samples were collected from tick-infested sheep in three different zones of Balochistan (i.e., Quetta, Zhob, and Loralai). All the collected samples were analyzed using conventional microscopy techniques, polymerase chain reaction (PCR), 18S small subunit rRNA gene sequencing, and phylogenetic analysis. The results of the microscopy and PCR confirmed the highest prevalence of Theileria species in district Zhob (14.22% and 15.68%) followed by district Loralai (11.52% and 13.97%) and district Quetta (10.29% and 12.00%), respectively. In addition, the prevalence of T. lestoquardi was higher in female sheep (84.12%), followed by adult sheep (74.71%) and the Hernai breed of sheep (28.23%) in the studied area. Similarly, the prevalence of theileriosis was higher in the summer season (40.59%), followed by the spring, autumn, and winter seasons. However, numerous risk factors such as age, sex, area, season, and breeds of the sheep were not significantly correlated (P > 0.05) with the presence of T. lestoquardi, except tick abundance and feeding pattern of animals (P < 0.05). Furthermore, sequencing and phylogenetic analyses of the isolated T. lestoquardi displayed 99% sequence similarity with isolates from Germany, Egypt, Iraq, India, Iran, and Pakistan. Altogether these results showed that T. lestoquardi is the main species causing ovine theileriosis in Balochistan. As a result, large-scale studies are required to design practical control approaches to reduce the risk of theileriosis infection in Balochistan, Pakistan.

First Detection of Theileria sinensis-like and Anaplasma capra in Ixodes kashmiricus: With Notes on cox1-Based Phylogenetic Position and New Locality Records.

Ixodes ticks transmit Theileria and Anaplasma species to a wide range of animals. The spreading of ticks and tick-borne pathogens has been attributed to transhumant herds, and research on these uninvestigated issues has been neglected in many countries, including Pakistan. Recently, we used internal transcribed spacer (ITS) and 16S ribosomal DNA partial sequences to genetically characterize Ixodes kashmiricus ticks and their associated Rickettsia spp. However, the data on its cox1 sequence and associated Theileria spp. and Anaplasma spp. are missing. This study aimed to genetically characterize I. kashmiricus based on the cox1 sequence and their associated Theileria spp. and Anaplasma spp. The I. kashmiricus ticks were collected from small ruminants: sheep (Ovis aries) and goats (Capra hircus) of transhumant herds in district Shangla, Dir Upper and Chitral, Khyber Pakhtunkhwa (KP), Pakistan. Out of 129 examined hosts, 94 (72.87%) (56 sheep and 38 goats) were infested by 352 ticks, including adult females (175; 49.7%) followed by nymphs (115; 32.7%) and males (62; 17.6%). For molecular analyses, 121 ticks were subjected to DNA isolation and PCR for the amplification of the cox1 sequence for I. kashmiricus, 18S rDNA for Theileria spp. and 16S rDNA sequences for Anaplasma spp. The obtained cox1 sequence showed 89.29%, 88.78%, and 88.71% identity with Ixodes scapularis, Ixodes gibbosus, and Ixodes apronophorus, respectively. Phylogenetically, the present cox1 sequence clustered with the Ixodes ricinus complex. Additionally, the 18S rDNA sequence showed 98.11% maximum identity with Theileria cf. sinensis and 97.99% identity with Theileria sinensis. Phylogenetically, Theileria spp. clustered with the T. cf. sinensis and T. sinensis. In the case of Anaplasma spp., the 16S rDNA sequence showed 100% identity with Anaplasma capra and phylogenetically clustered with the A. capra. PCR-based DNA detection targeting the amplification of groEL and flaB sequences of Coxiella spp. and Borrelia spp., respectively, was unsuccessful. This is the first phylogenetic report based on cox1 and new locality records of I. kashmiricus, and the associated T. sinensis-like and A. capra. Significant tick surveillance studies are needed in order to determine the epidemiology of Ixodes ticks and their associated pathogens.

Comparative evaluation of cell-mediated immune response in calves immunized with live-attenuated and killed Theileria annulata vaccines.

Tropical theileriosis is a tick-borne disease caused by the protozoan Theileria annulata and transmitted by numerous species of Ixodid ticks of the genus Hyalomma. The main clinical signs are fever, lymphadenopathy, and anemia responsible for heavy economic losses, including mortality, morbidity, vaccination failure, and treatment cost. Development of poor cell-mediated immunity (CMI) has been observed in the case of many bovine pathogens (bacteria, viruses, and parasites). Quantification of CMI is a prerequisite for evaluating vaccine efficacy against theileriosis caused by T. annulata. The current study evaluated the CMI in calves administered with two types of T. annulata vaccine (live attenuated and killed). We prepared a live attenuated T. annulata vaccine by attenuation in a rabbit model and also prepared killed vaccine from non-attenuated T. annulata. For the evaluation of immune response in experimental groups including control, 20 calves were divided into four different groups (A, B, C, and D). They were either inoculated subcutaneously with live rabbit-propagated-Theileria-infected RBCs (5 × 106) (group A) or with killed T. annulata vaccine (2 × 109 schizonts) with Freund's adjuvant (group B), along with an infected group (group C) and a healthy control group (group D). The protection of vaccinated calves was estimated with challenge infection. Our results showed that with a single shot of live-attenuated and killed vaccine with a booster dose elicited cell-mediated immune responses in immunized calves. We observed a significant elevation in CD4 + and CD8 + T cells in immunized calves. A significant difference in the CD8 + T cell response between the post-challenge stage of killed and live vaccine (p < 0.0001) was observed, whereas no other difference was found at both pre- and post-immunization stages. A similar finding was recorded for the CD4 + T cells at a post-challenge stage, where a significant difference was seen between killed and live vaccine (p < 0.0001). Another significant difference was observed between the CD8 + T cells and CD4 + T cells at the post-challenge stage in the live vaccine group, where there was a significantly higher induction of CD4 + T cell response (p < 0.0001).

Seroprevalence of toxoplasmosis in humans and animals in Pakistan: a systematic review and metadata analysis.

OBJECTIVE: To review the seroprevalence of toxoplasmosis in Pakistan. METHODS: The systematic review comprised search on Science Direct, Google Scholar, PubMed and Scopus databases for studies related to the seroprevalence of toxoplasmosis in Pakistan published between 2006 and 2020 which used serological diagnostic tests to detect Toxoplasma gondii. Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines were used throughout the review and statistical analysis was done using forest plot and random effect model. RESULTS: Of the 7093 human studies initially found, 20(0.28%) were reviewed. Of the 16,432 animal studies, 16(0.09%) were selected for detailed review. The pooled seroprevalence of toxoplasmosis in humans, calculated in this review was found as (76%) (95% confidence interval: 69-83%). Seroprevalence of human toxoplasmosis was higher in Khyber Pakhtunkhwa (31.7%) than Punjab (20.4%). Pooled seroprevalence in animals calculated in this review was found as (69%) (95% confidence interval: 64-74%). Seroprevalence in animals was higher in Khyber Pakhtunkhwa (44.7%) than Punjab (29.4%). CONCLUSIONS: The seroprevalence of toxoplasmosis in both humans and animals should be studied it other parts of Pakistan as well.

IEg67 kDa Bovine Hydatid Cyst Antigen: A Candidate for Developing Sero-Diagnostic Assays for Cystic Echinococcosis, a Disease of One Health Importance.

Cystic echinococcosis (hydatidosis) is a world-wide zoonotic disease of mainly humans, livestock and dogs, caused by Echinococcus granulosus. The disease can negatively impact food production and animal welfare and causes socio-economic hardship. Here, we aimed to identify the local bovine hydatid cyst fluid (BHCF) antigen for developing a sero-diagnostic assay to be used for the pre-slaughter screening of food animals. In total, 264 bovines approved for slaughter in Pakistan were subjected to serum collection and post-mortem screening for hydatid cysts. These cysts were assessed microscopically to assess fertility and viability, and by PCR for molecular confirmation of species. A BHCF antigen was identified from positive sera via SDS-PAGE, confirmed by Western blot, and quantified via a bicinchoninic acid (BCA) assay. The quantified crude BHCF antigen (iEg67 kDa) was then used in ELISA screening to test all sera collected from known positive and negative animals based on hydatid cyst presence/absence. Of the 264 bovines examined, 38 (14.4%) showed hydatid cysts during post-mortem examination. All of these individuals, plus an additional 14 (total: 52; 19.6%) tested positive based on less time-consuming ELISA examination. Based on ELISA, occurrence in females (18.8%) was significantly higher than in males (9.2%) and was higher in cattle (19.5%) compared to buffalo (9.5%). The infection rate increased with age in both host species: cumulatively, 3.6% in animals aged 2-3 years, 14.6% in 4-5-year-olds and 25.6% in 6-7-year-olds. The occurrence of cysts in cattle was significantly higher in the lungs (14.1%) compared to their livers (5.5%), whereas the opposite was true in buffalo (6.6% livers, 2.9% lungs). For both host species, most cysts in the lungs were fertile (65%), while the majority in the liver were sterile (71.4%). We conclude that the identified iEg67 kDa antigen is a strong candidate for the development of a sero-diagnostic screening assay for the pre-slaughter diagnosis of hydatidosis.

DOCUMENTATION OF TRYPANOSOMA EVANSI IN CAPTIVE TIGERS AND LIONS IN PUNJAB (2016-2018), PAKISTAN.

Trypanosoma evansi is an important hemoparasite of a variety of animal species worldwide. This parasite is a threat to the health of domestic animals as well as wild animals, particularly those managed in captivity. The current study investigated the presence of T. evansi in captive tigers (Panthera tigris tigris) and lions (Panthera leo) in Pakistan. In total, 24 blood samples from 11 tigers and 3 lions (n = 14) were collected during the course of roughly 3 yr (2016-2018). Eighteen samples were subjected to both microscopic and molecular evaluation for the presence of T. evansi; the remaining 6 samples were processed for PCR only. Of the 18 samples tested by both methods, 3 (16%) and 8 (44%) were positive by microscopy and PCR, respectively. This highlights the higher sensitivity of PCR over microscopy for detection of trypanosomes. Of the 24 total samples evaluated by PCR, 12 (50%) were positive. The three sequences obtained showed 99% identity with variant surface glycoprotein genes of the different isolates of T. evansi. The sensitivity, specificity, positive predictive value, and negative predictive value of microscopy in identifying T. evansi was 37.5, 100, 100, and 66.7%, respectively, considering PCR as the gold standard. We recommend rigorous monitoring of captive tigers and lions for hemoparasites, particularly in winter and early spring in areas with high infection rate of this parasite, preferably via PCR.

Molecular epidemiology and characterization of Babesia bovis in cattle of Khyber Pakhtunkhwa province, Pakistan.

Babesiosis is a tick-borne disease found globally but most prominent in tropical and subtropical regions. It is responsible for huge mortality and morbidity, especially in developing countries like Pakistan. The current study was designed to determine the molecular epidemiology and characterization of Babesia bovis (B. bovis) infection in cattle populations of districts Mardan, Kohat and Swat of Khyber Pakhtunkhwa (KP) province of Pakistan. A total of 434 tick-infested animals were sampled. Blood samples were collected, processed and then examined initially by microscopy for the presence of Babesia and were later confirmed through PCR by targeting cytochrome b gene, and the PCR products were sequenced. Phylogenetic analysis of sequenced isolates of the current study showed close sequence similarity with the reported strain of China. A non-significant association (p > 0.05) was observed between the prevalence of infections and risk factors. The overall prevalence of infection in all three districts was 10.11%. In district Swat (12.61%), the prevalence was recorded as the highest for B. bovis infection followed by district Mardan (10.60%) and district Kohat (06.90%). The Friesian breed of cattle, females and adult animals were highly susceptible to B. bovis infection. The prevalence of infection was recorded highest during the summer season and lowest during the winter season. This study concludes that B. bovis infection is prevalent in three studied districts of KP province and the sequenced isolates of the current study showed close sequence similarity with the reported strain of China.

Serological Investigation of Bovine Toxoplasmosis Using Commercial and Indigenous ELISA Kits While Validating Cattle Toxo IgG ELISA Kit.

Toxoplasma gondii is a protozoan parasite that causes toxoplasmosis in warm-blooded vertebrates, globally. The main aims of this study were to assess the seropositivity to toxoplasmosis of an exotic breed of cattle (n = 400) from different farms using the Latex Agglutination Test and validate Cattle Toxo IgG ELISA kit. Of a total of 400 cattle sera that were evaluated by LAT, 143 (35.75%) were found positive. Based on these data, 90 samples (n = 60 seronegative by LAT; n = 30 seropositive by LAT) were elected for screening through a commercially available ELISA kit. The same 90 samples were screened through a Cattle Toxo IgG ELISA kit for validation purposes. Of 90 samples, 40 were seropositive in the Cattle Toxo IgG ELISA kit (100% sensitivity), and 38 were seropositive in a commercially available ELISA kit. All 50 samples in the Cattle Toxo IgG ELISA kit (96.15% specificity) were also seronegative in the commercially available ELISA kit. Hence, the sensitivity and specificity of the Cattle Toxo IgG ELISA kit came out to be 100% and 96.15%, and in LAT, it was found as 26.31% and 61.53%, respectively. Therefore, the Cattle Toxo IgG ELISA kit is a highly reliable serodiagnostic tool to diagnose bovine toxoplasmosis.

Assessment of Cytotoxic, Genotoxic, and Oxidative Stress of Dibutyl Phthalate on Cultured Bovine Peripheral Lymphocytes.

Recently, there have been numerous reports showing that phthalates have negative human health impacts and may cause several diseases such as asthma, breast cancer, obesity, type II diabetes, and male infertility. Animals are also exposed to phthalates through the environment and can cause adverse health effects on them. Several studies have been found on the cytogenetic effects of dibutyl phthalate (DBP) on different organisms but no documented evidence has been found on the cytotoxic and genotoxic effects of dibutyl phthalate (DBP) on bovine cultured lymphocytes. MTT assay was performed on different series of DBP concentrations (10 µM, 20 µM, 30 µM, 50 µM, 70 µM, 100 µM). A concentration-dependent decrease in cell viability was observed by the DBP. The LD50, LD50/2, and 2∗LD50 were found to be 50 µM, 30 µM, and 80 µM on bovine lymphocytes, respectively. Then, these concentrations of DBP were utilized to perform comet, micronucleus assays, and oxidative stress. A concentration-dependent increase in DNA damage, oxidative stress, and micronuclei formation was observed in lymphocytes by the DBP as compared to the control group. Highest genotoxic effects were observed at a concentration of 2∗LD50. Similarly, total oxidative stress was found higher, and antioxidative stress was lower in concentration-dependent manner by the DBP. The current study revealed a significant cytotoxic, genotoxic, and oxidative stress of DBP on cultured bovine lymphocytes.

Theileria annulata: Its Propagation in Rabbits for the Attenuation of Piroplasms in Cross-Bred Calves.

Tropical theileriosis caused by the protozoan; Theileria annulata is a tick-borne disease (TBD) transmitted by ticks of genus Hyalomma; is clinically characterized by fever, anemia, and lymphadenopathy; and is responsible for heavy economic losses in terms of high morbidity and mortality rates with reduced production. Infected red blood cells of T. annulata were inoculated into rabbits intraperitoneally, and propagation of T. annulata has been investigated. The current study has shown an association between induced tropical theileriosis and variation of body temperature in rabbits. A significant rise in temperature (39.92 ± 0.33 °C) was seen on day 8 onwards, with the maximum temperature (40.27 ± 0.44 °C) on day 14 post-inoculation. In the current study, in vivo trials in susceptible cross-bred calves to investigate the attenuation and comparison with the infected group were also conducted. All the infected calves (n = 5) showed a significant rise in temperature (40.26 ± 0.05 °C) on day 10 onwards, with the maximum temperature (40.88 ± 0.05 °C) on day 16. The temperature of inoculated calves increased gradually post-inoculation, but the difference was not significant. A maximum parasitemia of 20% was observed in infected calves, but no piroplasm parasitemia was observed in inoculated calves. The prescapular lymph nodes of infected calves were enlarged, while the lymph nodes of inoculated calves remained normal throughout the trial. Analysis of clinical and parasitological responses of infected and inoculated calves showed a significant difference (p ≤ 0.05) in terms of temperature, parasitemia, and lymph node scoring between two groups. The current study was primarily aimed to attenuate T. annulata in rabbit and to check its virulence in susceptible calves. It is concluded that propagation of Theileria annulata in rabbits made it attenuated. Rabbit can be used as an in vivo model to weaken the virulence of T. annulata.

Immune response against toxoplasmosis-some recent updates RH: Toxoplasma gondii immune response.

AIMS: Cytokines, soluble mediators of immunity, are key factors of the innate and adaptive immune system. They are secreted from and interact with various types of immune cells to manipulate host body's immune cell physiology for a counter-attack on the foreign body. A study was designed to explore the mechanism of Toxoplasma gondii (T. gondii) resistance from host immune response. METHODS AND RESULTS: The published data on aspect of host (murine and human) immune response against T. gondii was taken from Google scholar and PubMed. Most relevant literature was included in this study. The basic mechanism of immune response starts from the interactions of antigens with host immune cells to trigger the production of cytokines (pro-inflammatory and anti-inflammatory) which then act by forming a cytokinome (network of cytokine). Their secretory equilibrium is essential for endowing resistance to the host against infectious diseases, particularly toxoplasmosis. A narrow balance lying between Th1, Th2, and Th17 cytokines (as demonstrated until now) is essential for the development of resistance against T. gondii as well as for the survival of host. Excessive production of pro-inflammatory cytokines leads to tissue damage resulting in the production of anti-inflammatory cytokines which enhances the proliferation of Toxoplasma. Stress and other infectious diseases (human immunodeficiency virus (HIV)) that weaken the host immunity particularly the cellular component, make the host susceptible to toxoplasmosis especially in pregnant women. CONCLUSION: The current review findings state that in vitro harvesting of IL12 from DCs, Np and MΦ upon exposure with T. gondii might be a source for therapeutic use in toxoplasmosis. Current review also suggests that therapeutic interventions leading to up-regulation/supplementation of SOCS-3, IL12, and IFNγ to the infected host could be a solution to sterile immunity against T. gondii infection. This would be of interest particularly in patients passing through immunosuppression owing to any reason like the ones receiving anti-cancer therapy, the ones undergoing immunosuppressive therapy for graft/transplantation, the ones suffering from immunodeficiency virus (HIV) or having AIDS. Another imortant suggestion is to launch the efforts for a vaccine based on GRA6Nt or other similar antigens of T. gondii as a probable tool to destroy tissue cysts.

Lumpy skin disease is expanding its geographic range: A challenge for Asian livestock management and food security.

In recent years, lumpy skin disease virus has extended its geographical range outside of endemic sub-Saharan countries to the Middle East and Asia indicating transboundary spread. Recently, lumpy skin disease (LSD) outbreaks have been reported in Asian countries such as Bangladesh, India, China, Nepal, Bhutan, Vietnam, Myanmar, Sri Lanka, Thailand, Malaysia, Laos and for the first time and represent a cause of serious concern for their livestock and dairy industries. This report summarizes information on the recent outbreaks of LSD in southern Asia and emphasizes the threat it poses to neighbouring countries. Various strategies and actions needed to control outbreaks of this emerging disease in Asia are also suggested.

Development of Human Toxo IgG ELISA Kit, and False-Positivity of Latex Agglutination Test for the Diagnosis of Toxoplasmosis.

Toxoplasma gondii is an intracellular zoonotic parasite that causes infection in a wide range of warm-blooded animals and humans. The main aim of this study was to assess the diagnostic value of the recombinant SAG1 antigen (rSAG1) for T. gondii-IgG screening through the Human Toxo IgG ELISA Kit (K). The rSAG1 was expressed in E. coli (DE3), and it was purified through metal-affinity chromatography. The rSAG1 was confirmed by immunoblotting, and it had a band on 35 kDa. Total of 400 human sera were tested by LAT and K. One hundred and twenty-two (30.5%) sera were found positive by LAT and eighty-nine (22.25%) sera were found positive by K. Out of 400 samples, 80 were selected to evaluate the performance of K through commercial Toxoplasma gondii IgG ELISA Kit (C). Out of 80 human sera, 55 (68.75%) were found positive, 25 (31.25%) were found negative by K and C, respectively. The cut-off value for K was 0.398 and it was calculated through the receiver operator characteristic curve. The ELISA plates were coated at optimized concentration of rSAG1 = 0.125 µg/mL, and the test was performed by diluting the sera at 1:50. The sensitivity and specificity of K were observed to be 98.5% and 100%, respectively. The six sera (K-L+) were found positive through LAT and these human sera were later evaluated by Western blot analysis. These sera did not produce a band equivalent to 35 kDa on WB analysis thus, LAT produced false-positive results.

Humoral and Cell-Mediated Immune Response Validation in Calves after a Live Attenuated Vaccine of Babesia bigemina.

The current vaccines to control bovine Babesia bigemina (B. bigemina) infection are not fully protective and vaccination failures incur heavy losses to the cattle industry around the world. Using modified micro-aerophilous stationary phase, we developed a culture-derived attenuated live vaccine against B. bigemina and tested a single subcutaneous inoculation of 2 × 108 infected erythrocytes in calves. The protection was measured after a lethal intravenous challenge with 5 × 108 virulent calf-derived B. bigemina. Our results demonstrated that a single shot of attenuated vaccine was capable of inducing robust humoral and cell-mediated immune responses in calves. We found a significant increase in the IgG antibody titers post-challenge and a strong proliferation of both CD4+ and CD8+ T cells contributing towards the protection. Our vaccine provided complete protection and parasitic clearance, which was followed for more than 100 days post-challenge. This immunity against babesiosis was directly linked to strong humoral responses; however, the parasitic clearance was attributed to significant T cells effector responses in vaccinated calves as compared to the infected control calves. We anticipate that these results will be helpful in the development of more efficient culture-derived vaccines against Babesia infections, thus reducing significant global economic losses to farmers and the cattle industry.

Expiry of a completely splenectomised calf in post-operative period due to mixed piroplasm infection: a case report.

Bovine babesiosis is an infectious protozoan disease and causes significant economic losses in terms of production loss and mortality. The genus Babesia belongs to the family Babesiidae order piroplasmida and is transmitted by ticks globally. The signs of disease are particularly prominent in old or immuno-compromised animals. The spleen plays a vital role in defence against hemoparasites like Babesia. A young cross-bred cow calf of about 3 months of age was splenectomised to propagate Babesia in vivo experimentally. Prior to splenectomy, the calf was examined through microscopy and PCR analysis and was found negative for any kind of piroplasms including Babesia. The calf was completely splenectomised, but the calf was naturally infected during its postoperative period. The calf expired after naturally acquiring Babesia bigemina and Theileria annulata during the 11 th day of postoperative period owing to increased parasitaemia, exhibiting typical mixed parasitic infection stigmata e.g. reddish urine, elevated temperature up to 41.38°C. This study concluded that complete splenectomy along with dexamethasone administration in the postop period caused exceptional increase in parasitaemia. This parasitaemia couldn't be countered by any symptomatic treatment because of the absence of spleen and greatly reduced immunity of the animal.

An investigation on some toxic effects of pyriproxyfen in adult male mice.

OBJECTIVES: Pyriproxyfen as an insect growth regulator is widely used globally for pest management. There are reports on adverse effects of insecticides such as organ toxicity, endocrine disruptions, and teratogenicity in animals and humans. We aimed to investigate reproductive toxicity of pyriproxyfen in adult male mice. MATERIALS AND METHODS: 48 male Swiss albino mice were divided into eight groups and received the different 1200, 600, 320, 200, 100, 40, 20, 0 mg/kg/day doses orally, and body weights were accessed for 28 consecutive days. In the end, mice were sacrificed, testes were dissected and weighed. Probable testicular tissue alterations were examined by histopathological studies. In addition, the diameter of seminiferous tubules and Leydig cells distribution were assessed in all experimental and control groups. RESULTS: Pyriproxyfen treatment caused significant (P<0.05) reduction in body and organ weights in mice. However, the shrinkage and displacement of seminiferous tubules, reduced lumen diameter, and vacuolization occurred in seminiferous tubules in higher doses exposed animals in comparison to controls. The relative testis weights, mean diameter of seminiferous tubules, and Leydig cells distribution remained unchanged at low doses. CONCLUSION: These findings reveal that pyriproxyfen caused reduction in body weight gain as well as damage to the testicular architecture in mice and thus may potentially interfere with spermatogenesis. Findings in an outbred strain of mice can be extrapolated fairly reliably to the human model. The chemical can thus be further exploited to study its effects on impairment of fertility and as an endocrine disruptor.

A systematic review on modelling approaches for economic losses studies caused by parasites and their associated diseases in cattle.

Parasites reside inside or outside their hosts and get host nutrition and blood. Here, we have emphasized economic losses in cattle caused by parasitic diseases due to ecto- and endo- parasites (flies, ticks, mites and helminths). We have outlined different methods/models including economic evaluation techniques and dynamic analysis as a major class, used for the calculation of economic losses caused by parasites in cattle. According to already conducted studies, a decrease in production is mentioned in quantity and percentage while financial losses are expressed in the form of account with respect to per head, herd or for the specific study area. The parasites cause the reduced production and financial losses due to control, treatment and mortality costs. We calculated the average decrease in milk production and organ condemnation as 1.16 L animal-1 day-1 and 12.95%, respectively, from overall cattle parasitic infections. Moreover, the average calculated financial and percentage losses were US$ 50.67 animal-1 year-1 and 17.94%, respectively. Economically important parasitic diseases mentioned here are caused by specific spp. of protozoans and helminths according to data collected from the literature. Protozoan diseases include tick-borne diseases, coccidiosis, neosporosis, trypanosomiasis and cryptosporidiosis. Losses due to tick-borne infections were encountered for decreased milk production, mortality, treatment and control. Losses from coccidiosis were due to decreased weight gain, treatment costs and mortality. While abortion losses were encountered in neosporosis. Trypanosomiasis caused losses due to a decrease in milk yield. Moreover, only diagnostic (conventional or molecular techniques) cost was taken into account for cryptosporidiosis. Economically important nematode parasites are Oesophagostomum spp., Cooperia spp., Trichostrongylus spp., Strongyloides spp., Ostertagia spp. and Haemonchus placei. Due to the zoonotic importance of echinococcosis, Echinococcus granulosus is the most economically important cestode parasite. Losses caused by echinococcosis were due to organ condemnation, carcass weight loss and decreases hide value, milk production and fecundity. While, fascioliasis is one of the most economically important trematodal disease, which causes cirrhosis of the liver due to parasite migration, and thus, the organ becomes inedible. So, it would be helpful for farmers and researchers to approach these methods/models for calculation of parasitic losses and should adopt suitable measures to avoid long-term economic losses.

Assessing the prevalence and economic significance of coccidiosis individually and in combination with concurrent infections in Pakistani commercial poultry farms.

Coccidiosis is one of the most economically important diseases of the poultry, around the globe. In order to assess seasonal and age-wise prevalence of coccidiosis individually and with concurrent infections, this study was conducted on commercial poultry farms in and around Multan division, province of Punjab, Pakistan. A total of 28,126 boilers, 4,052 layers and 7,699 golden bird samples, provided by regional farmers and consultants, were examined by microscopy for the diagnosis of coccidiosis. Based on postmortem lesions, several infections including coccidiosis, viral, and miscellaneous diseases were catalogued, whereas samples were cultured to identify concurrent bacterial diseases. Cumulative analysis of this large set of samples revealed a prevalence of 14.16, 11.01, and 19.57% in broiler, layer and golden birds, respectively. Ascaridia galli (A. galli) showed a higher prevalence in layer (2.47%) compared to golden (0.21%) birds (P < 0.01). Amongst all concurrent infections investigated, bacterial infections were identified in highest percentage of samples (59.24%; P < 0.05). The age-wise and season-wise prevalence of coccidiosis and A. galli was found to be significant (P < 0.05). During the study period, an estimated of 10.69 (coccidiosis) and 0.29 (A. galli) million poultry birds were treated or provided prophylaxis and supportive therapy at Pakistani commercial poultry farms. Depending upon the prevailing market conditions, the total economic losses (treatment, prophylaxis, and supportive therapy) from coccidiosis and A. galli were estimated to be US $45,405.00 and 2,638.50, respectively, while production (weight and eggs) losses for broiler (US$ 2,750,779.00), layer and golden, young (US$ 13,974.98 and 50,228.76) and adult (US$ 104.74 and 203.77) were estimated. Unit loss of coccidiosis with concurrent diseases and intestinal worm was estimated to be US$ 0.005 and 0.01, respectively. These results highlight the potential impact of coccidiosis individually and with concurrent infections on the poultry productivity and will inform farmers, policy makers, and other governmental and non-governmental stakeholders on the use of control and management measures in containing these infections.