Investigating the relationship between heat load and shade seeking behaviour in dairy buffaloes.

The study presented in this Research Communication aimed to investigate the relationship between physiological responses, body surface temperature and shade-seeking behaviour in Nili Ravi dairy buffaloes during summer months. We enrolled 60 buffaloes, and each animal was observed for three consecutive days starting before sunrise until they moved towards the shade structures. A repeated measures ANOVA was employed to assess the changes in physiological parameters and body surface temperature between the early morning and the occurrence of shade-seeking behaviour. The average temperature humidity index and heat load index during the behavioural monitoring period (0400 to 1200 h) were 81.3 ± 6.5 and 92.9 ± 17, respectively (mean ± sd). There was no significant difference in core body temperature between sunrise and the time of shade-seeking event. However, the buffaloes had a slightly higher respiration rate at the time of shade-seeking (19.2 vs. 22.4 breaths/min). In addition, body surface temperature, measured at the flank region, shoulder, base of the ear and forehead was significantly higher at the occurrence of shade-seeking behaviour compared to the early morning. On average, the buffaloes sought shade when the surface temperature was 2°C higher than the temperature recorded before sunrise. Overall, the current findings suggest that body surface temperature, rather than core body temperature was strongly associated with shade-seeking behaviour in dairy buffaloes. These findings could be useful in developing strategies to mitigate the effects of heat stress in dairy buffalo herds and thereby improve animal welfare.

Endogenous and viral microRNAs in nasal secretions of water buffaloes (Bubalus bubalis) after Bubaline alphaherpesvirus 1 (BuHV-1) challenge infection.

Bubaline alphaherpesvirus 1 (BuHV-1) is a pathogen of water buffaloes responsible for economic loss worldwide. MicroRNAs (miRNAs) regulate gene expression produced by alphaherpesviruses and hosts. This study aimed at (a) unravelling the ability of BuHV-1 to produce miRNAs, including hv1-miR-B6, hv1-miR-B8, hv1-miR-B9; (b) measuring the host immune-related miRNAs associated to herpesvirus infection, including miR-210-3p, miR-490-3p, miR-17-5p, miR-148a-3p, miR-338-3p, miR-370-3p, by RT-qPCR; (c) identifying candidate markers of infection by receiver-operating characteristic (ROC) curves; (d) exploiting the biological functions by pathway enrichment analyses. Five water buffaloes BuHV-1 and Bovine alphaherpesvirus 1 (BoHV-1) free were immunized against Infectious Bovine Rhinotracheitis (IBR). Five additional water buffaloes served as negative controls. All animals were challenged with a virulent wild-type (wt) BuHV-1 via the intranasal route 120 days after the first vaccination. Nasal swabs were obtained at days (d) 0, 2, 4, 7, 10, 15, 30, and 63 post-challenge (pc). The animals of both groups shed wt BuHV-1 up to d7 pc. Results demonstrated that (a) miRNAs produced by the host and BuHV-1 could be efficiently quantified in the nasal secretion up to d63 and d15 pc, respectively; b) the levels of host and BuHV-1 miRNAs are different between vaccinated and control buffaloes; c) miR-370-3p discriminated vaccinated and control animals; d) host immune-related miRNAs may modulate genes involved in the cell adhesion pathway of the neuronal and immune system. Overall, the present study provides evidence that miRNAs can be detected in nasal secretions of water buffaloes and that their expression is modulated by BuHV-1.

Efficacy of Duddingtonia flagrans (Bioverm®) on the biological control of buffalo gastrointestinal nematodes.

We evaluated the efficacy of Bioverm®, a commercial product containing Duddingtonia flagrans, on the control of buffalo (Bubalus bubalis) gastrointestinal nematodes. We randomly divided 12 buffaloes into two groups of six animals. In the treated group, each animal received a Bioverm®`s single dose of 1g (105 chlamydospores of D. flagrans) to 10 kg of live weight; in the control group, each animal received 1g of corn bran for each 10 kg of live weight as a placebo. Fecal samples were individually collected from 12, 24, 36, 48, 60 and 72 h after treatments. To examine 1) viability of chlamydospores passed through the gastrointestinal tract, 2 g of faeces and 1000 infective larvae (L3) were added to Petri dishes with 2% water-agar, and 2) to examine larval predation by D. flagrans during fecal cultures, 2000 L3 were added. In the Petri dishes, were observed significant reductions (p < 0.01) in the treated group after 48 (56.7%) and 60 h (91.5%). In the fecal cultures, significant reductions (p < 0.01) occurred in the treated group from 36 h (75%), with larval reduction up to 72 h. High larval predation rate occurred 60 h after Bioverm® administration. Bioverm® maintained viability and predation capacity after passage through the buffalo's gastrointestinal tract, showing efficacy on gastrointestinal nematodes.

Evaluation of Ovsynch versus modified Ovsynch program on pregnancy rate in water buffaloes: a meta-analysis.

Ovsynch is a widely accepted estrus synchronization protocol for improving the reproductive performance of water buffaloes who manifest low reproductive efficiency. Recently, some modified protocols based on Ovsynch such as 2 injections of prostaglandin 14 days apart following the Ovsynch are also introduced to enhance the reproductive potential of this species. In the present study, a meta-analytical assessment was performed with the objective to evaluate the reproductive performance of water buffaloes synchronized with Ovsynch or modified Ovsynch programs. Meta-analysis of the fixed or random effects model was determined by the heterogeneity among the studies. Reproductive outcome of interest was pregnancy per artificial insemination (P/AI) measured on day 25 (25-100). A total of 32 articles including 4003 buffaloes using either Ovsynch or modified Ovsynch protocol were reviewed. In the random effects model for buffaloes, the overall proportion of P/AI was 42.55% [95% confidence interval (CI): 37.48-47.70; n = 3,089] and 46.44% (95% CI: 39.63-53.31; n = 914) on day 25 after AI for Ovsynch and modified Ovsynch, respectively. Results for P/AI were then categorized by ovarian activity, where P/AI was available for 3575 cyclic buffaloes and 320 non-cyclic buffaloes. For cyclic buffaloes, the overall proportion of P/AI was 47.54% (95% CI: 42.72-52.38; n = 2911) and 57.97% (95% CI: 54.12-61.77; n = 664) on day 25 after AI for Ovsynch and modified Ovsynch, respectively. In the fixed effects model for non-cyclic buffaloes, the overall proportion of P/AI was 19.68% (95% CI: 13.48-26.58; n = 167) and 33.01% (95% CI: 25.50-40.94; n = 153) on day 25 after AI for Ovsynch and modified Ovsynch, respectively. In conclusion, a benefit for P/AI is detected in buffaloes with the modified Ovsynch protocol. Besides, whichever estrus synchronization protocols (Ovsynch or modified Ovsynch), cyclic buffaloes have higher P/AI compared with non-cyclic buffaloes.

Karyotypic and mtDNA based characterization of Malaysian water buffalo.

BACKGROUND: In Malaysia, the domestic water buffaloes (Bubalus bubalis) are classified into the swamp and the murrah buffaloes. Identification of these buffaloes is usually made via their phenotypic appearances. This study characterizes the subspecies of water buffaloes using karyotype, molecular and phylogenetic analyses. Blood of 105 buffaloes, phenotypically identified as swamp, murrah and crossbred buffaloes were cultured, terminated and harvested using conventional karyotype protocol to determine the number of chromosomes. Then, the D-loop of mitochondrial DNA of 10 swamp, 6 crossbred and 4 murrah buffaloes which were identified earlier by karyotyping were used to construct a phylogenetic tree was constructed. RESULTS: Karyotypic analysis confirmed that all 93 animals phenotypically identified as swamp buffaloes with 48 chromosomes, all 7 as crossbreds with 49 chromosomes, and all 5 as murrah buffaloes with 50 chromosomes. The D-loop of mitochondrial DNA analysis showed that 10 haplotypes were observed with haplotype diversity of 0.8000 ± 0.089. Sequence characterization revealed 72 variables sites in which 67 were parsimony informative sites with sequence diversity of 0.01906. The swamp and murrah buffaloes clearly formed 2 different clades in the phylogenetic tree, indicating clear maternal divergence from each other. The crossbreds were grouped within the swamp buffalo clade, indicating the dominant maternal swamp buffalo gene in the crossbreds. CONCLUSION: Thus, the karyotyping could be used to differentiate the water buffaloes while genotypic analysis could be used to characterize the water buffaloes and their crossbreds.

Fatal Theileria orientalis N2 genotype infection among Asian water buffaloes (Bubalus bubalis) in a commercial dairy farm in Kerala, India.

Fifteen dairy buffaloes of a farm in the state of Kerala, India developed fatal oriental theileriosis within 2 months of their procurement. Typical piroplasms of Theileria orientalis were observed in the erythrocytes of all affected animals by Giemsa-Leishman staining of blood smears. Case fatality rate was 87·5% (seven out of eight) in the clinically progressed cases. Therapeutic management with anti-theilerial drugs buparvaquone and oxytetracycline led to recovery of seven other animals in less advanced stages of the disease. The aim of this study was to determine the reasons for increased virulence of this pathogen, hitherto considered to be benign. Acute haemolytic anaemia was the predominant haematological finding in the affected animals. Lymphocytic infiltration and degeneration of vital organs leading to functional derangement was the cause of the high mortality. Identification of T. orientalis was confirmed by polymerase chain reaction (PCR). DNA sequencing of the PCR products revealed close identity with already reported sequences of T. orientalis/buffeli N2 genotype. The sequences were deposited in GenBank with accession number KM609973 and KM043772. Rhipicephalus ticks, previously not reported as vectors for oriental theileriosis, were identified as the potential vectors. This is the first report of fatal oriental theileriosis in Asian water buffaloes.

First Molecular Characterization of Cryptosporidium spp. Infecting Buffalo Calves in Brazil.

With the aim of determining the occurrence of Cryptosporidium spp., 222 fecal samples were collected from Murrah buffalo calves aged up to 6 mo. Fecal DNA was genotyped with a nested polymerase chain reaction targeting the 18S rRNA gene and sequencing of the amplified fragment. Nested 18S PCR was positive for 48.2% of the samples. Sequence analysis showed that the most frequent species in these animals was Cryptosporidium ryanae, which was present in buffalo calves as young as 5 d. The zoonotic species Cryptosporidium parvum was detected in one animal. An uncommon Cryptosporidium 18S genotype was found in buffaloes.

Bovine Alphaherpesvirus 1, Bovine Alphaherpesvirus 5 and Bubaline Alphaherpesvirus 1 in Palatine Tonsils from Water Buffaloes in Northern Brazil and Possible Links with the Origin of Bovine Alphaherpesvirus Type 5.

Herpesviruses are significant pathogens of ruminants. In water buffaloes (Bubalus bubalis), however, herpesviruses have not been thoroughly studied. Although bubaline alphaherpesvirus 1 (BuAHV1) and bovine alphaherpesvirus 1 (BoAHV1) have already been recovered from water buffaloes, to date, no reports on the occurrence of bovine alphaherpesvirus 5 (BoAHV5) in these animals have been published. Therefore, the aim of this study was to search for BuAHV1, BoAHV1, and BoAHV5 in palatine tonsils of apparently healthy water buffaloes from the Pará state, Northern Brazil. Tissue samples of tonsils (n = 293) were screened by a nested PCR (nPCR) targeting a region of UL44 (gC coding gene), followed by sequencing, to detect and differentiate between the viral types. Viral genome segments were detected in 18 out of 293 (6.1%) of the palatine tonsil samples. Two animals carried genomes of BoAHV1 only, eleven animals carried BoAHV5 genomes only, and four animals carried BuAHV1 only. Another animal had both BoAHV1 and BoAHV5 genomes in its tonsils. No infectious virus could be recovered from any of the samples. The BuAHV1 sequences identified here were more closely related to BuAHV1 genomes identified in India. Phylogenetic analyses suggested a closer relationship between the recovered BoAHV5 and BuAHV1 genomes. Therefore, evidence is provided here to confirm that not only BoAHV1 and BuAHV1, but also BoAHV5, can infect water buffaloes. This report highlights (i) the first detection of BoAHV5 in water buffaloes and (ii) the occurrence of coinfections with BoAHV1 and BoAHV5 in that species. Such findings and the similarity of BoAHV5 to Indian herpesvirus genomes suggest that the origin of type 5 may be linked to recombinations between bovine and bubaline herpesviruses within bubalines, since the scenario for generation of recombinants in buffaloes is potentially present.

Seroprevalence of Schistosoma japonicum Infection Among Dogs and Water Buffaloes Using Recombinant Antigen ELISA in New Corella, Davao del Norte, Philippines.

PURPOSE: Animal reservoirs are important targets for controlling and eliminating schistosomiasis. Prevalence studies showed that dogs (Canis familiaris) and water buffaloes (Bubalus bubalis) are important reservoirs of S. japonicum. Previous studies have validated the use of the recombinant proteins Sj1TR, Sj7TR, and SjTPx-1 in ELISA as diagnostics for dogs and water buffaloes from schistosomiasis-endemic areas. In this study, we aimed to determine the seroprevalence of S. japonicum among dogs and water buffaloes in New Corella, Davao del Norte, Philippines, using the recombinant proteins Sj1TR, Sj7TR, and SjTPx-1 in ELISA format. METHODS: Fecal and serum samples were collected from dogs (n = 63) and water buffaloes (n = 57). Schistosome-positive samples were detected using formalin-ether concentration technique (FECT), stool polymerase chain reaction, and enzyme-linked immunosorbent assay (ELISA) using soluble egg antigen (SEA), rSj1TR, rSj7TR, and rSjTPx-1. Positivity rates, sensitivity, specificity, predictive values, accuracy, and kappa values were calculated. RESULTS: Mean positivity rates for schistosome infection were high for both dogs (x = 15.40%) and water buffaloes (x = 6.32%). On dogs, the sensitivity and specificity of the tests are as follows: 66.67% and 100% for rSj7TR-ELISA, while 100% and 93.33% for rSjTPx-1-ELISA, respectively. rSjTPx-1-ELISA showed the highest agreement with stool PCR among all diagnostic tests, with an overall kappa value of 0.824. On water buffaloes, the sensitivity and specificity of both rSj1TR-ELISA and rSjTPx-1-ELISA are 100.0% and 98.15%. Both tests also had an overall kappa value of 0.84. CONCLUSION: To ensure elimination and prevention of schistosomiasis in humans, the use of validated serological diagnostics such as recombinant antigen ELISA is preferable for field detection in animals, especially in resource-limited areas.

The Characterization of Pastures by Grazing Cycle and Evaluation of Supplementation with Agro-Industry Co-Products on the Performance of Buffaloes in the Humid Tropics.

The objective was to characterize the pastures by grazing cycle, as well as to evaluate the performance of buffaloes in intensive rotational grazing in a silvopastoral system in the eastern Amazon supplemented with agro-industry co-products in order to characterize the grazing cycles, the composition of the fractions, and the carcass yield. Fifteen non-castrated, crossbred water buffaloes (Murrah × Mediterranean) were used. All animals used in the study were clinically healthy and weighed approximately 458 kg. The animals were grazed in a single group, and supplementation (1% of live weight-LW/day) was divided into three treatments: control (control-conventional ingredients); Cocos nucifera coconut cake (Cocos nucifera) (coconut cake-70%); and palm kernel cake (Guinean elaeis) (palm kernel cake-70% palm kernel cake). The chemical composition of the forage is different in each part of the plant, with higher protein values in the leaves (11.40%) and higher acid detergent fiber (ADF) values in the stems (50.03%). Among the ingredients of the supplement, corn has the highest percentage of indigestible protein (35.57%), most of the protein in palm kernel cake is B3 (49.11%), and in Coco, B2 (51.52%). Mombasa grass has a higher percentage of B3 and B2 proteins; the indigestible fraction is lower in the leaves (17.16%). The leaf/stem ratio also varied between grazing cycles, being better in the second cycle (2.11%) and with an overall average of 1.71. Supplement consumption varied between cycles and was higher in the control treatment, with an overall mean of 4.74. There was no difference in daily weight gain and carcass yield, with an average of 1 kg/day and 49.23%, respectively. Therefore, it can be concluded that including supplements based on by-products from the coconut and palm oil agro-industries promotes performance and carcass yields compatible with conventional supplements. Their use reduces production costs, optimizes the utilization of forage mass, enhances the sustainability of the production chain, and, therefore, is recommended.

Highly Effective Frontal Stunning Procedure Using a Pneumatic Penetrating Captive Bolt in Water Buffaloes.

An effective frontal stunning procedure in water buffaloes was assessed using a pneumatic penetrating captive bolt (PPCB) with high air pressure. The study contemplated two phases; first, 352 buffaloes and 168 post-mortem heads were evaluated to determine the most effective anatomical site for stunning. Then, the second phase (n = 182) was used to validate the stunning procedure at the discovered anatomical site in the first phase, which was located 8 cm dorsal above the middle of the forehead on an "X" formed between the eyes and the base of the contralateral horns, and 2 cm lateral, avoiding the midline, where the skull tended to narrow. A total of 95.1% of buffaloes received effective stunning at the first shot with evidence of the presence of collapse, absence of rhythmic breathing, and absence of ocular reflexes (corneal and palpebral). There were no differences in the stunning efficacy by sex, breed, or skull thickness. These findings demonstrated that stunning with a PPCB at pressures of 1379-1516.8 KPa (200-220 pounds per square inch (psi)) in the site reported here produces a highly effective stunning at the first shot in water buffaloes.

Seroprevalence and associated risk factors for Toxoplasma gondii in water buffaloes (Bubalus bubalis) in Egypt.

Toxoplasmosis is a parasitic disease caused by Toxoplasma gondii and affecting all warm-blooded animals. The available data about the epidemiological situation of T. gondii in water buffaloes in Egypt are scarce. Thus, a cross-sectional study was conducted to determine the seroprevalence of T. gondii in water buffaloes in three Egyptian governorates and to evaluate the associated risk factors for the infection. A total of 430 sera samples were examined using commercial Indirect ELISA Multi-species kit. The overall seroprevalence rate of T. gondii in examined water buffaloes was 7.4 %, and the highest rate (9.3 %) was found in Kafr ElSheikh governorate. Multivariate logistic regression analysis showed that adult buffalo (OR = 7.10; 95 % CI: 0.87-57.68; P = 0.067) and small herds (OR = 8.42; 95 % CI: 1.07-66.02; P = 0.043) were more likely than young buffalo and large herds to become infected with T. gondii. Moreover, the risk of buffaloes contracting T. gondii infection was higher in winter and especially among animals contacted with cats. It is necessary to identify risk factors in order to determine what mitigation, control, and prevention strategies to implement in order to reduce, control, and prevent T. gondii infection in domestic animals, which will in turn reduce human infection with the disease.

Seroprevalence of Orientia tsutsugamushi and Rickettsia typhi in water buffaloes (Bubalus bubalis) from Southern Thailand.

Background and Aim: Scrub typhus and murine typhus are globally distributed zoonoses caused by the intracellular Gram-negative bacteria Orientia tsutsugamushi and Rickettsia typhi, respectively. Numerous studies have been undertaken on rickettsial illnesses in humans and animals, including arthropod vectors, in Thailand. However, the reports on the seroprevalence of antibodies to O. tsutsugamushi and R. typhi in buffaloes is extremely rare. Thus, this study aimed to estimate the seroprevalence of both rickettsial infections in water buffaloes (Bubalus bubalis) in Phatthalung Province, southern Thailand. Materials and Methods: From February to March 2023, a total of 156 serum samples were collected from 156 water buffaloes on 29 farms in Phatthalung province. The sera were screened for antibodies against O. tsutsugamushi and R. typhi using an indirect immunofluorescence assay. Results: The seroprevalence of antibodies against O. tsutsugamushi and R. typhi in individual water buffaloes was 4.49% (95% confidence interval [CI]: 2.19%-8.97%) and 3.85% (95% CI: 1.77%-8.14%), respectively, whereas 31% (9/29) of the herds had buffaloes with antibodies. The number of buffaloes with scrub typhus infection and ectoparasite infestation was statistically significant (p < 0.05; odds ratio = 6.25 [95% CI: 1.19-33.33]). Intriguingly, the prevalence of scrub typhus antibodies in buffaloes that were not infested with ectoparasites was much higher than those that were. Conclusion: This is the first report of O. tsutsugamushi and R. typhi antibodies in water buffalo sera in Southern Thailand. Two serum samples showed a high antibody titer against O. tsutsugamushi. Seroprevalence mainly occurred in non-ectoparasite-infested buffaloes, especially for O. tsutsugamushi antibodies. At the herd level, one-third of the studied farms showed seroprevalence. Additional research on the occurrence of these pathogens in vectors and in other animal reservoirs is necessary.

Molecular characteristics of Staphylococcus aureus strains isolated from subclinical mastitis of water buffaloes in Guangdong Province, China.

Intramammary infections (IMI) in animals reared for milk production can result in large economic losses and distress to the animals. Staphylococcus aureus is an important causative agent of IMI in dairy cows, but its prevalence in water buffaloes has not been determined. Therefore, the current study was conducted to investigate the prevalence of subclinical mastitis in water buffaloes and the antimicrobial susceptibility, virulence genes and biofilm formation abilities of Staphylococcus aureus isolates recovered from water buffaloes in Guangdong, China. Staphylococcus aureus strains were isolated from milk samples of water buffaloes with subclinical mastitis, and twofold microdilution, PCR and crystal violet staining methods were used to determine antimicrobial susceptibility, distributions of virulence and antimicrobial resistance genes and biofilm formation ability, respectively. Our results indicated that 29.44% of water buffaloes were diagnosed with subclinical mastitis, and the most prevalent pathogens were Escherichia coli (96.17%), coagulase-negative staphylococci (CoNS) (67.60%) and S. aureus (28.57%). Most S. aureus isolates showed resistance to bacitracin, doxycycline, penicillin, florfenicol, and tetracycline but were susceptible to ciprofloxacin, ceftizoxime, cefoquinoxime, and ofloxacin. Moreover, 63.72% of S. aureus isolates were positive for tetM, and the prevalence of msrB, blaZ, mecA, fexA, and tetK ranged from 21.24 to 6.19%. All S. aureus isolates harbored clfB and icaA genes, and the virulence genes hla (93.8%), hld (91.15%), clfA (90.27%), fnbA (86.73%), and hlb (83.19%), and tsst, icaD, sec, see, fnbB, and sea showed a varied prevalence ranging from 3.5 to 65.49%. All S. aureus isolates possessed the ability to form biofilms, and 30.09% of isolates showed strong biofilm formation abilities, while 19.47% of isolates were weak biofilm producers. Our results indicated that subclinical mastitis is prevalent in water buffaloes in Guangdong, China, and S. aureus is prevalent in samples from water buffaloes with subclinical mastitis. Most S. aureus isolates were susceptible to cephalosporins and fluoroquinolones; thus, ceftizoxime and cefoquinoxime can be used to treat subclinical mastitis in water buffaloes.

First report on the molecular detection of Entamoeba bovis from the endangered wild water buffalo (Bubalus arnee) in Nepal.

BACKGROUND: The Asiatic wild water buffalo (Bubalus arnee) is an endangered species that is conserved in the Koshi Tappu Wildlife Reserve (KTWR), Nepal, and was recently translocated to the Chitwan National Park (CNP). Gastrointestinal (GI) parasites are the cause of significant negative health and production impacts on animals worldwide. METHODS: A coprological survey of GI parasites of wild water buffalo was carried out in the CNP in 2020. Fresh dung samples (n = 25) were collected from wild water buffaloes and analysed using sedimentation and flotation techniques for morphological identification of parasite cysts, oocysts and eggs. RESULTS: Nine different GI parasites were recorded of which Entamoeba spp. (20 samples, 80%) were the most common. The presence of Entamoeba spp. was further validated using polymerase chain reaction (PCR) analysis and DNA sequencing. The PCR results were positive for all of the microscopically positive samples, and the species was identified as Entamoeba bovis. Three samples were sequenced and formed a cluster of E. bovis, which was separated from other Entamoeba spp. in phylogenetic analysis. CONCLUSION: This is the first report for molecular detection of E. bovis from wild water buffaloes in Nepal. Future work should focus on the prevalence of such infections in water buffaloes in forest environments.

Geographic origin of Explanatum explanatum (Creplin, 1847) Fukui, 1929 detected from domestic water buffaloes in Sri Lanka.

The adult stage of Explanatum explanatum has economic importance in the production of ruminants, especially water buffaloes. This species has been widely reported in the Indian sub-continent. Recently, molecular analyses to reveal the dispersal route of this species were performed in Bangladesh, Nepal, and India. In the present study, we focused on E. explanatum distributed in Sri Lanka. A total of 52 flukes were collected from water buffaloes in Sri Lanka and identified as E. explanatum based on the internal transcribed spacer 2 (ITS2) region of nuclear ribosomal DNA. Analysis of the mitochondrial NADH dehydrogenase subunit 1 (nad1) gene from DNA samples detected 18 haplotypes, and five of them were identical to those from the Indian E. explanatum. The pairwise fixation index value indicated that the Sri Lankan population had a comparatively closer relationship with the Indian population than with the Bangladeshi or Nepalese populations. The Sri Lankan population showed significantly lower genetic variability than the Indian population, suggesting that the Indian population was the ancestor of the Sri Lankan population. The movement of host ruminants, including water buffaloes, was probably involved in the introduction of the fluke into Sri Lanka. The results of our study provide useful information for elucidating the geographic origin of E. explanatum distributed in the Indian subcontinent.

The first report of seroprevalence of Q fever in water buffaloes (Bubalus bubalis) in Phatthalung, Thailand.

BACKGROUND AND AIM: Q fever is a worldwide zoonosis caused by the intracellular bacterium, Coxiella burnetii. A few studies focused on the occurrence of Q fever infection in water buffaloes in Thailand have been conducted; however, little is known regarding the seroprevalence of C. burnetii antibodies in buffaloes. In the present study, we describe the prevalence of Q fever infection in water buffaloes (Bubalus bubalis) in Phatthalung, Thailand. MATERIALS AND METHODS: A total of 421 samples (156 blood, 156 sera, and 109 ectoparasites [lice]) were collected from 156 water buffaloes from 29 farms of the Phatthalung Province from January 22, 2021, to March 26, 2021. The blood and ectoparasite samples were screened for C. burnetii DNA using a polymerase chain reaction assay and the sera were tested for C. burnetii antibody using an indirect immunofluorescence assay. RESULTS: C. burnetii DNA was not detected in blood or ectoparasites; however, the seroprevalence of individual water buffaloes was 4.49% (95% CI: 2.19-8.99%), whereas that of the herd was 13.79%. There was a significant difference between abortion history and Q fever infection at 29 farms (p=0.005; OR=33.55 [95%CI: 156-722.38]). CONCLUSION: This is the first report describing the low seroprevalence of C. burnetii antibodies in water buffaloes in Phatthalung Province, Thailand. The occurrence of this pathogen in buffaloes with reproductive disorders and people working with buffaloes warrant further investigation. Animal health authorities should inform farmers to effectively prevent and control this zoonosis.

Lepra Bubalorum, a Potential Reservoir of Mycobacterium leprae.

In 1926, a mycobacterial skin disease was observed in water buffaloes by researchers in Indonesia. The disease was designated as skin tuberculosis, though it was hypothesized that it might be a form of leprosy or a leprosy-like disease. In a follow-up study (Ph.D. thesis Lobel, 1934, Utrecht University, Netherlands) a similar nodular skin disease was described in Indonesian water buffaloes and named "lepra bubalorum" or "nodular leprosy." Two decades later Kraneveld and Roza (1954) reported that, so far, the diagnosis lepra bubalorum had been made in 146 cases in Indonesia. After a final series of research reports by Indonesian veterinarians in 1961, no subsequent cases were published. Based on information from these reports, it can be concluded that, even though evidence of nerve involvement in buffaloes was not reported, similarities exist between lepra bubalorum and Hansen's disease (leprosy), i.e., nodular skin lesions with a chronic course and microscopically granulomatous reactions with AFB in globi in vacuoles. This raises the question as to whether these historical cases might indeed have been caused by Mycobacterium leprae, Mycobacterium lepromatosis or another representative of the M. leprae complex. The future use of state-of-the-art molecular techniques may answer this question and may also help to answer the question whether water buffaloes should be considered as a potential natural reservoir of the causative pathogen of Hansen's disease.

A Defined Antigen Skin Test for Diagnosis of Bovine Tuberculosis in Domestic Water Buffaloes (Bubalus bubalis).

Bovine tuberculosis (bTB) remains endemic in domestic water buffaloes (Bubalus bubalis) in India and elsewhere, with limited options for control other than testing and slaughter. The prescribed tuberculin skin tests with purified protein derivative (PPD) for diagnosis of bTB preclude the use of Bacille Calmette-Guérin (BCG)-based vaccination because of the antigenic cross-reactivity of vaccine strains with Mycobacterium bovis and related pathogenic members of the M. tuberculosis complex (MTBC). For the diagnosis of bTB in domestic water buffaloes, we here assessed a recently described defined-antigen skin test (DST) that comprises overlapping peptides representing the ESAT-6, CFP-10 and Rv3615c antigens, present in disease-causing members of the MTBC but missing in BCG strains. The performance characteristics of three doses (5, 10 or 20 µg/peptide) of the DST were assessed in natural tuberculin skin test reactor (n = 11) and non-reactor (n = 35) water buffaloes at an organized dairy farm in Hisar, India, and results were compared with the single intradermal skin test (SIT) using standard bovine tuberculin (PPD-B). The results showed a dose-dependent response of DST in natural reactor water buffaloes, although the SIT induced a significantly greater (P < 0.001) skin test response than the highest dose of DST used. However, using a cut-off of 2 mm or greater, the 5, 10, and 20 µg DST cocktail correctly classified eight, 10 and all 11 of the SIT-positive reactors, respectively, suggesting that the 20 µg DST cocktail has a diagnostic sensitivity (Se) of 1.0 (95% CI: 0.72-1.0) identical to that of the SIT. Importantly, none of the tested DST doses induced any measurable skin induration responses in the 35 SIT-negative animals, suggesting a specificity point estimate of 1.0 (95% CI: 0.9-1.0), also identical to that of the SIT and compares favorably with that of the comparative cervical test (Se = 0.85; 95% CI: 0.55-0.98). Overall, the results suggest that similar to tuberculin, the DST enables sensitive and specific diagnosis of bTB in water buffaloes. Future field trials to explore the utility of DST as a defined antigen replacement for tuberculin in routine surveillance programs and to enable BCG vaccination of water buffaloes are warranted.

The first report of the seroprevalence of antibodies against Bartonella spp. in water buffaloes (Bubalus bubalis) from South Thailand.

BACKGROUND AND AIM: Bartonellosis is an emerging worldwide zoonosis caused by bacteria belonging to the genus Bartonella. Several studies have been conducted on the prevalence of Bartonella infections from animals and humans, including reports from wild and domestic ruminants. However, there has been only one report of Bartonella infection in water buffaloes from the northeastern part of Thailand. Moreover, the seroprevalence of Bartonella spp. in water buffaloes still remains unknown. This study was conducted to explore the prevalence of Bartonella spp. among water buffaloes from South Thailand using molecular and serological techniques. MATERIALS AND METHODS: A total of 312 samples (156 blood and 156 sera) of 156 water buffaloes from 29 farms in Phatthalung Province, South Thailand, were collected from January to March 2021. All samples were screened for Bartonella spp. using polymerase chain reaction and indirect immunofluorescence assay. RESULTS: The seroprevalence of antibodies against three Bartonella spp. was 16.03% (25/156, 95% confidence interval: 10.65-22.74%), and among 25 water buffaloes with seroprevalence, 56%, 20%, and 24% were positive for antibodies against Bartonella henselae, Bartonella vinsonii subspp. berkhoffii, and Bartonella tamiae, respectively. No significant difference was detected among seroprevalence, gender, age, and ectoparasite infestation. CONCLUSION: This is the first report of the seroprevalence of antibodies against B. henselae, B. vinsonii subspp. berkhoffii, and B. tamiae in water buffaloes from South Thailand. Further studies are required on the epidemiology of Bartonella infection among water buffaloes, related personnel, and ectoparasites.