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RESEARCH HIGHLIGHTS: First confirmation of AOAV-16 in domestic and wild birds in China.AOAV-16 are low virulent viruses for chickens.Co-circulation/co-infection of AOAV-16 and H9N2 subtype AIV enhanced pathogenicity.Different intergenic sequences and recombination events exist within AOAV-16.
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Caprine arthritis encephalitis (CAE) is a worldwide economically important disease of small ruminants particularly goats. CAE has been considered to be an emerging/re-emerging disease of goats and a notifiable disease in the Philippines. In this study, a nested-PCR method to detect CAE virus (CAEv) infection was conducted between January 2021 to December 2022. A total of 1334 goat blood samples were collected from 24 goat farms throughout Luzon, the Philippines. The over-all prevalence rate was 31.41% (419/1334) in goats and 91.67% (22/24) of goat farms. These results showed high positivity rate of CAEv and the disease may be widespread in Luzon, the Philippines.
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Objective: The aerolysin (aerA) is a virulence indicator used to identify the pathogenicity of the Aeromonas strain. Targeting a pathogen's crucial virulence gene for detection is essential, as it determines the potential threat to the host. This study aimed to develop a gold nanoparticle (AuNP) probe for detecting the gene aerA in Aeromonas hydrophila among field samples. Materials and Methods: Kidney samples among both healthy and sick Nile tilapias in five provinces of Luzon Island were collected for bacterial analysis. Screening using specific primers targeting aerA was conducted in parallel with testing the AuNPs probe on the same sample set. The positive control provided by BFAR-NFLD, confirmed by polymerase chain reaction (PCR) assay, was used as a positive sample containing the target gene. Results: The AuNP probe demonstrated a computed accuracy of 81.32%, sensitivity of 100%, and specificity of 81.26%. Among the 257 reactions, 59 were false positives, while no false negative results were observed. The AuNP probe could detect aerA at levels as low as 30 ng/µl. The low prevalence of the target gene may be attributed to the use of general media instead of specific media like Rimler-Shotts agar. Conclusion: The established colorimetric detection method for A. hydrophila with the aerA gene offers a swift alternative to PCR, negating the requirement for advanced equipment like a thermal cycler.
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The highly virulent Newcastle disease virus (NDV) isolates typically result in severe systemic pathological changes and high mortality in Newcastle disease (ND) illness, whereas avirulent or low-virulence NDV strains can cause subclinical disease with no morbidity and even asymptomatic infections in birds. However, understanding the host's innate immune responses to infection with either a highly virulent strain or an avirulent strain, and how this response may contribute to severe pathological damages and even mortality upon infection with the highly virulent strain, remain limited. Therefore, the differences in epigenetic and pathogenesis mechanisms between the highly virulent and avirulent strains were explored, by transcriptional profiling of chicken embryonic visceral tissues (CEVT), infected with either the highly virulent NA-1 strain or the avirulent vaccine LaSota strain using RNA-seq. In our current paper, severe systemic pathological changes and high mortality were only observed in chicken embryos infected with the highly virulent NA-1 strains, although the propagation of viruses exhibited no differences between NA-1 and LaSota. Furthermore, virulent NA-1 infection caused intense innate immune responses and severe metabolic disorders in chicken EVT at 36 h post-infection (hpi), instead of 24 hpi, based on the bioinformatics analysis results for the differentially expressed genes (DEGs) between NA-1 and LaSota groups. Notably, an acute hyperinflammatory response, characterized by upregulated inflammatory cytokines, an uncontrolled host immune defense with dysregulated innate immune response-related signaling pathways, as well as severe metabolic disorders with the reorganization of host-cell metabolism were involved in the host defense response to the CEVT infected with the highly virulent NA-1 strain compared to the avirulent vaccine LaSota strain. Taken together, these results indicate that not only the host's uncontrolled immune response itself, but also the metabolic disorders with viruses hijacking host cell metabolism, may contribute to the pathogenesis of the highly virulent strain in ovo.
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Enfermedades Metabólicas , Virus no Clasificados , Animales , Embrión de Pollo , Pollos , Biología Computacional , Virus ADN , Inmunidad Innata , Virus de la Enfermedad de Newcastle/genéticaRESUMEN
OBJECTIVES: Determinants showing plasmid-mediated quinolone resistance, which usually leads to antimicrobial ineffectiveness, have become an emerging clinical problem. In our previous study in the Philippines, a high prevalence of qnr determinants was found in clinical samples and food-producing animals and their food products. However, no qnr-carrying plasmids have been investigated in animals or animal-derived foods. Hence, in the present, we aimed to characterise qnr-carrying plasmids in Escherichia coli isolated from the food supply chain. METHODS: Plasmids from 44 qnr-positive isolates were assigned to incompatibility groups by Polymerase chain reaction (PCR)-based replicon typing, and the presence of ß-lactamase-encoding genes were investigated by PCR. Localisation of qnr in plasmids was determined by S1-PFGE and Southern blot hybridisation. The transferability of qnr-carrying plasmids was examined by conjugation analysis. RESULTS: Overall, 77.3% (95% confidence interval [CI]: 62.2-88.5) of the isolates harbouring qnr determinants were positive for seven plasmid types, and 56.8% concurrently harboured blaTEM-1. Plasmid IncFrepB was prevalent (65.9% [95% CI: 50.1-79.5]) among qnr determinants. Localisation of qnr determinants in IncFrepB and transferability of plasmids was further confirmed. CONCLUSION: The current study proved that qnr in E. coli isolated from food-producing animals and their food products could spread via plasmid IncFrepB upon selective pressure with quinolones or other antimicrobials. Therefore, to curb the emergence and spread of qnr-harbouring bacteria in the Philippines, prudent use of antimicrobials in animal production and stricter hygiene and food handling are recommended.
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Infecciones por Escherichia coli , Quinolonas , Animales , Escherichia coli/genética , Infecciones por Escherichia coli/microbiología , Filipinas , Plásmidos/genética , Quinolonas/farmacología , beta-Lactamasas/genéticaRESUMEN
Fasciolosis is considered as one of the leading causes of morbidity and mortality among ruminants in the Philippines. Though anthelmintic drugs are widely used to treat and control the condition, it is still worthwhile to search for alternative treatments especially when resistance to commonly used anthelmintic drugs has been reported. In this study, the ethanolic leaf extract of fringed spiderflower (Cleome rutidosperma) was evaluated for its in vitro anthelmintic activity against Fasciola spp. Specifically, the study compared the different concentrations of ethanolic leaf extract and the commonly used anthelmintic drug (albendazole) on the gross motility and histology of Fasciola spp. The study consisted of five treatments: treatment 1, 2, and 3 which contain 10%, 20%, and 40% leaf extract, respectively, treatment 4 with 10% albendazole as the positive control, and treatment 5 with nutrient broth as the negative control. The motility of the Fasciola spp in all treatments was visually analyzed based on the established criteria. In addition Fasciola spp. in different treatments were subjected to tissue processing and histological examination. Results showed that increasing concentrations of leaf extract resulted in a decreasing time for Fasciola spp. to have a motility score of zero. Specifically, 10%, 20%, and 40% leaf extract resulted in a cumulative time period of 55.00 ± 5.00 min, 26.67 ± 2.89 min, and 15.00 ± 0.00 min, respectively, for the Fasciola spp. to have a motility score of zero. On the other hand, albendazole resulted in a 240.00 min cumulative time before it can cause a motility score of zero. Histologic examination showed that the different concentrations of leaf extract affected the tegument and parenchyma of the Fasciola spp.
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Antihelmínticos , Cleome , Fasciola , Fascioliasis , Animales , Antihelmínticos/farmacología , Antihelmínticos/uso terapéutico , Fascioliasis/tratamiento farmacológico , Extractos Vegetales/farmacologíaRESUMEN
Methicillin-resistant Staphylococcus aureus (MRSA) from dairy animals could pose a public health concern in the population. The study was designed to determine the prevalence of S. aureus and MRSA associated with mastitis among water buffaloes in the central part of Luzon island, the Philippines, and to investigate its associated factors. Three hundred and eighty-four water buffaloes were examined for mastitis using California mastitis test (CMT). Composite milk samples (n = 93) were collected from buffaloes showing positive reaction with CMT. S. aureus was identified from milk samples using biochemical tests. Cefoxitin disk diffusion assay and PCR detecting mecA gene were performed to identify MRSA isolates. Disk diffusion assay was used to investigate the antimicrobial resistance against 9 antibiotics. The prevalence of S. aureus was 41.94% (39/93). MRSA isolates resistant to cefoxitin were at 25.81% (24/93) but only 37.5% (9/24) harbored the mecA gene. All 24 MRSA isolates were resistant to penicillin while the majority were susceptible to clindamycin, trimethoprim-sulfamethoxazole, gentamycin, tetracycline, rifampicin, ciprofloxacin and chloramphenicol with intermediate susceptibility to erythromycin. Furthermore, 37.5% of the isolates were found resistant to two or more antibiotics. Animal-level factor associated with MRSA infection was the history of mastitis (OR = 3.18, CI = 1.03-9.79, p = 0.040). Herd-level factors associated with the detection of MRSA in milk included herd size (OR = 4.24, CI = 1.05-17.07, p = 0.042) and the presence of other animals (OR = 0.15, CI = 0.04-0.58, p = 0.006). High prevalence of intramammary infection with S. aureus and MRSA in dairy buffaloes was observed in the region. This finding raises the concern of preventing zoonotic spread of MRSA.
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OBJECTIVE: Mastitis is considered as an economically important disease of dairy buffaloes in Asia. This study examined the mastitis milk and nasal swab samples for the detection and genotyping of methicillin-resistant Staphylococcus aureus (MRSA) in water buffaloes. MATERIALS AND METHODS: Staphylococcus aureus was identified based on biochemical tests and Polymerase Chain Reaction (PCR) detection of nuc gene, whereas MRSA on mecA gene. The disc diffusion test was used to determine the antibiotic resistance and staphylococcal cassette chromosome mec (SCCmec), spa, and multilocus sequence typing for the genotyping of isolates. RESULTS: Staphylococcus aureus was detected on 39/93 milk (41.94%) and 27/384 nasal swab (7.03%) samples. However, only nine isolates (23.08%) harbored the mecA gene from milk samples and three isolates (11.11%) from the nasal carriage. All MRSA isolates exhibited resistance to cefoxitin and penicillin, whereas 50% were found resistant to clindamycin. All these isolates were found susceptible to sulfa-trimethoprim and chloramphenicol, whereas the majority of the isolates were susceptible to gentamicin, ciprofloxacin, tetracycline, and rifampicin. The SCCmec types of the MRSA isolates were type IVc (50.00%), type II (8.33%), type I (8.33%), and non-typeable (33.33%). The spa types and sequence type (ST) identified were t019 (ST30), t701 (ST1649), t311 (ST5), t657 (ST1148), t015 (ST508), t1939 (ST12), t800 (ST9), t091 (ST2454), t138 (ST5991), and t1642 (ST5992). CONCLUSION: Milk and nasal swab samples from dairy water buffaloes were found positive for MRSA. The MRSA isolates were still susceptible to most antibiotics tested. Moreover, the genotypes of some MRSA isolates were found similar to some human MRSA strains, suggesting a possible human to animal transmission.
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OBJECTIVE: Major histocompatibility complex (MHC) is a set of molecular proteins on the surface of antigen presenting cells encoded by a large gene family which are important parts of the immune system. This study was conducted to convey information on the genetic characteristics of the MHC II DRB3 gene in riverine and swamp buffaloes. MATERIALS AND METHODS: Characterization of MHC II DRB3 gene was carried out using polymerase chain reaction (PCR)-based assay. Thirty-milliliter milk samples were collected from 10 swamp-type and 10 riverine-type buffaloes. RNA from milk samples were extracted using Trizol and then followed by reverse transcription-PCR (RT-PCR). RESULTS: The phylogenetic analysis with 1,000 bootstrap replications clearly showed complex parsimony in MHC II DRB3 gene between 10 riverine- and 10 swamp-type but also confirmed that the samples are similar to Bubalus bubalis. Aligned sequences of the 20 water buffaloes were compared with three other ruminants (Bos taurus, Ovis aries, and Capra hircus) and non-ruminant (Sus scrofa) that serve as an outgroup. MHC sequences from GenBank show that there was an average of 705 identical pairs, with 22 transitional pairs and 30 transversional pairs with a ratio of 0.7. CONCLUSION: Based on the molecular data, the current study conforms to other works of literature that this gene is highly polymorphic which can be due to its function in the immune responsiveness and disease resistance. Further study on the immunological response of MHC II DRB3 to infection may elucidate its underlying function and role in the protection against specific disease of animals.
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OBJECTIVE: This study aimed to measure the anthelmintic effects of betel nut (Areca catechu) and neem (Azadirachta indica) leaf extracts against Fasciola spp. in vitro in comparison with the commercial dewormer, Albendazole, and the negative control, nutrient broth. The study determined the extract concentration that produced the highest efficacy based on the average recorded mean motility time, gross, and microscopic changes of the flukes treated with different concentrations of plant extracts. MATERIAL AND METHODS: The study consisted of eight treatments. Every treatment consisted of 10%, 20%, and 40% concentrations of both betel nut extract (BNE) and neem leaf extracts, positive control treatment (Albendazole-treated) and negative control treatment (25 ml nutrient broth). The motility of the flukes on all treatments was based on the established motility criteria scoring. The flukes subjected to all treatments were processed for histopathological analysis. RESULTS: The result of the study revealed that after exposure of Fasciola spp. under 10%, 20%, and 40% extract concentrations, betel nut showed higher efficacy having the recorded mean motility time of 0.22, 0.07 min, and no movement upon contact, respectively, than Albendazole which produced mean motility time of 0.38 min. Nevertheless, the flukes treated with 10%, 20%, and 40% neem leaf extracts obtained the average mean motility time of 220, 151, and 98 min, respectively. CONCLUSIONS: The results gathered showed that 40% BNE concentration showed the highest efficacy based on the recorded mean motility time. All treatments of betel nut extract evidently showed marked changes in the gross and microscopic morphology of the flukes. However, the neem extract was ineffective in all concentrations although changes were observed microscopically. Furthermore, the nutrient broth was proven to be effective as a culture medium since the flukes remained active until 8 h of exposure.
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FoxP3 is a forkhead family member that plays an important role in the development and function of a type of CD4 + T cell called T regulatory cells. Molecular characterization of FoxP3 gene in swamp- and riverine-type water buffaloes was conducted to determine its homology and compare it to the FoxP3 gene of other animal species (cattle, goat, sheep, horse, pig, cat, and dog), determine its unique characteristics in water buffaloes, and provide a reference for future studies to analyze its immunological function. FoxP3 nucleotide sequence of swamp- and riverine-type water buffaloes was 99% identical, whereas its protein translation revealed 97% homology. FoxP3 of swamp- and riverine-type water buffaloes were compared to FoxP3 of other animal species and revealed a high degree of homology which suggests that they may have the same biological properties. This study is the first report that describes the genetic characteristic of FoxP3 gene in water buffalo.
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Búfalos/genética , Factores de Transcripción Forkhead/genética , Análisis de Secuencia de ADN , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Animales , Búfalos/clasificación , Filogenia , Alineación de Secuencia , Homología de SecuenciaRESUMEN
Salmonella infection can cause septicemia, acute or chronic enteritis and wasting in weaned pigs, but may occur in other age groups. The bactericidal/permeability-increasing protein (BPI) gene plays an important role in the natural defense of the host and is found to be associated with resistance/susceptibility to Salmonella infection and identified as a candidate gene for disease resistance breeding in pig. This study was conducted to screen the resistance and/or susceptibility of pigs to Salmonella infection, to determine the genotype and evaluate presence of resistant allele of the BPI gene in population of pigs, and to establish genetic data for pig breeders for the improvement of Philippine pig industry. In this study, 389 blood samples from different pig breeds were collected from pig breeder farms in the Philippines. Genomic DNA was extracted from these samples and genotyping was done by PCR-RFLP analysis using AvaII restriction enzyme. Out of 389 pigs, the genotypic frequency showed that 98.4, 1.3, and 0.3% pigs are resistant (GG), heterozygous type (AG), and susceptible (AA), respectively. The application of BPI gene as marker for disease resistance will provide information to the pig industry to implement strategies for the identification of Salmonella infection-resistant pigs.
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Péptidos Catiónicos Antimicrobianos/genética , Proteínas Sanguíneas/genética , Resistencia a la Enfermedad/genética , Salmonelosis Animal/inmunología , Salmonella/fisiología , Sepsis/veterinaria , Alelos , Animales , Antiinfecciosos , Cruzamiento , Marcadores Genéticos/genética , Genotipo , Reacción en Cadena de la Polimerasa/veterinaria , Polimorfismo de Longitud del Fragmento de Restricción , Salmonelosis Animal/microbiología , Sepsis/inmunología , Sepsis/microbiología , PorcinosRESUMEN
This study detected and characterized the TevAT1 gene of Trypanosoma evansi isolates from Philippine water buffaloes (Bubalus bubalis). A total of 68 blood samples from Philippine water buffaloes were subjected to DNA extraction and PCR assay was performed using RoTat 1.2 gene to detect T. evansi. Those samples positive for T. evansi subsequently underwent another PCR assay to detect the presence of TevAT1 gene. Trypanosoma evansi was detected in 26.47% (18/68) blood samples in which distributed throughout the main islands of the country (4 from Luzon, 2 from Visayas and 12 from Mindanao). However, only 10 of these samples were positive for TevAT1 gene. Sequence alignment of the TevAT1 gene from local isolates showed no single nucleotide polymorphisms when compared to other strains in various countries. Those T. evansi without the gene of interest could be possibly resistant to some trypanocidal drugs but this needs to be further investigated in-vitro or in-vivo.
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Búfalos , Resistencia a Medicamentos , Proteínas de Transporte de Nucleósidos , Trypanosoma , Tripanosomiasis , Animales , Búfalos/parasitología , Resistencia a Medicamentos/genética , Proteínas de Transporte de Nucleósidos/química , Proteínas de Transporte de Nucleósidos/genética , Filipinas , Polimorfismo de Nucleótido Simple , Trypanosoma/genética , Tripanosomiasis/parasitologíaRESUMEN
Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of Johne's disease and a possible cause of Crohn's disease in humans. A total of 70 blood and fecal samples were collected from water buffaloes in selected municipalities of Nueva Ecija for ELISA and qPCR assay. Results revealed presence of antibodies of MAP in 3 serum samples for ELISA. The qPCR assay was carried out using standard curve method targeting the MAP specific insertion element IS900. Results revealed that 10 of the samples were positive for MAP DNA in qPCR. ELISA was able to detect antibodies for MAP showing 2.48% infection rate among the 70 buffaloes tested using blood serum samples. On the other hand, qPCR was able to detect MAP using IS900 showed 14.28% infection rate among buffaloes tested using fecal samples. Nucleotide sequence of isolated MAP showed high homology (99-100%) among the reported MAP isolates in the GenBank.
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Búfalos/microbiología , Mycobacterium avium subsp. paratuberculosis/genética , Paratuberculosis/microbiología , Animales , Anticuerpos Antibacterianos/sangre , ADN Bacteriano/sangre , Ensayo de Inmunoadsorción Enzimática , Heces/microbiología , Paratuberculosis/sangre , Filipinas , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
The viral agent of the porcine epidemic diarrhea (PED) was investigated during the reported 2014-2015 outbreaks in commercial farms in Central Luzon, Philippines. The study covered detection of PED virus (PEDV) in fecal and intestinal samples through reverse transcription PCR and sequence analysis of the nucleocapsid (N) gene. Results showed that 10 out of 34 fecal and intestinal samples examined were positive for PEDV. The partial nucleotide sequence of the N gene of the field samples showed 98-99% homologous to PEDV sequences registered in the GenBank. It was also noted that N gene sequences between field samples were 98% homologous. Interestingly, the partial sequences of the N genes of the field samples were genetically similar to the PEDV isolates from USA, China, Mexico, Canada and Japan. The phylogenetic tree analysis revealed that the Philippine samples clustered in group 2-1 of the PEDV, wherein the isolates of this group were responsible for the outbreaks in Asia and the USA. Analysis of the partial nucleotide and amino acid sequences revealed polymorphisms, deletions and insertions in the N-gene of the PEDV. Amino acid sequence alignment also showed deletions and insertion in the PEDV detected in the Philippines.
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In the Philippines, bovine ephemeral fever (BEF) is currently undetected and considered as an exotic disease of both cattle and water buffaloes. The Philippines until now has no official data regarding the occurrence of BEF. There were no existing control programs or vaccine used for the prevention of the disease. However, there are claims of BEF existence in different water buffalo and cattle farms based on the clinical signs but never confirmed using laboratory test yet. Detection of BEF virus in cattle and water buffalo blood samples was conducted using reverse-transcription PCR targeting the glycoprotein (G) gene, a conserved region in the BEF virus genome. The samples were collected from 22 cattle and 50 water buffaloes with clinical signs suggesting of BEF infection. All water buffalo blood samples were negative while four cattle blood samples turned positive for BEF virus. The G gene partial sequence analysis from two BEF virus positive samples showed close relationship to Australian isolates.
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Sixty suspected protozoan oocysts were demonstrated from 260 fecal samples collected from water buffaloes aged one month to seven years old with clinical signs of diarrhea in four provinces in the Philippines after conventional methods of isolation, sporulation, morphological characteristics and Kinyoun Acid Fast Staining techniques. The recovered protozoan oocysts were subjected to molecular analysis. Amplification of DNA extracted from recovered Eimeria oocysts using universal primers for the ITS-1 region of 18S rRNA revealed PCR products with 348 bp size demonstrated by samples collected from Benguet, La Union and Nueva Ecija provinces in the Philippines while DNA extracted from oocysts of suspected Cryptosporidium spp. samples that applied primers for the SSU of 18S rRNA registered PCR products but no genes were amplified from diarrheic water buffaloes from these provinces. Alignment of the DNA sequences of the suspected Eimeria and Cryptosporidium species revealed sequences for three isolates of Buxtonella sulcata with product lengths that varied from 235 to 252 bp. This is an initial observation on the involvement of B. sulcata in diarrhea condition of water buffaloes in the Philippines. Phylogenetic analysis of the three local isolates of B. sulcata revealed no similarity with other protozoan constructed according to Neighbor-Joining method.
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Búfalos/parasitología , Infecciones por Cilióforos/veterinaria , Cilióforos/genética , Diarrea/veterinaria , Animales , Infecciones por Cilióforos/epidemiología , Infecciones por Cilióforos/parasitología , ADN Intergénico/genética , ADN Protozoario/genética , Diarrea/parasitología , Filipinas/epidemiología , Filogenia , Reacción en Cadena de la PolimerasaRESUMEN
Gold nanoparticle (AuNP) is considered to be the most stable metal nanoparticle having the ability to be functionalized with biomolecules. Recently, AuNP-based DNA detection methods captured the interest of researchers worldwide. Paratuberculosis or Johne's disease, a chronic gastroenteritis in ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP), was found to have negative effect in the livestock industry. In this study, AuNP-based probes were evaluated for the specific and sensitive detection of MAP DNA. AuNP-based probe was produced by functionalization of AuNPs with thiol-modified oligonucleotide and was confirmed by Fourier-Transform Infrared (FTIR) spectroscopy. UV-Vis spectroscopy and Scanning Electron Microscopy (SEM) were used to characterize AuNPs. DNA detection was done by hybridization of 10⯵L of DNA with 5⯵L of probe at 63⯰C for 10â¯min and addition of 3⯵L salt solution. The method was specific to MAP with detection limit of 103â¯ng. UV-Vis and SEM showed dispersion and aggregation of the AuNPs for the positive and negative results, respectively, with no observed particle growth. This study therefore reports an AuNP-based probes which can be used for the specific and sensitive detection of MAP DNA.
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Colorimetría/métodos , ADN/genética , ADN/aislamiento & purificación , Oro/química , Nanopartículas del Metal/química , Mycobacterium avium/genética , Mycobacterium avium/aislamiento & purificación , Técnicas Biosensibles/métodos , Hibridación in Situ/métodos , Técnicas de Sonda Molecular , Sondas Moleculares/química , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
OBJECTIVE: This study assessed the applicability of loop-mediated isothermal amplification (LAMP) for the detection of leptospirosis among domesticated animals and sewage rats. Specifically, it evaluated the ability of LAMP to amplify Leptospira spp. targeting the 16s rRNA gene in boiled urine samples. MATERIALS AND METHODS: A total of 140 samples from different domestic animals were tested for the presence of the antigen. A nested-polymerase chain reaction (nPCR) protocol was used to compare and determine the sensitivity of LAMP in detecting Leptospira spp. The LAMP was also evaluated by comparing its amplification result using agarose gel electrophoresis and color change using dye. RESULTS: Positivity rate of Leptospira spp. antigen was 29.0% (40/140) for LAMP and 9.3% (13/140) for nPCR. Also, LAMP results for gel electrophoresis and dye color change varied in some samples that may be due to the interpretation of the result in dye color change. CONCLUSION: Overall, LAMP is a rapid, sensitive, and cost-effective diagnostic method compared with nPCR. Also, LAMP has a potential application as pen-side screening, surveillance, and clinical diagnostic kits of infectious diseases without requiring advance equipment and skilled personnel.
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Phospholipase C zeta, a novel sperm-specific protein which is widely known to induce oocyte activation following fertilization, had already been characterized in various mammalian species, but not in water buffaloes thus far. The present study was conducted to initially characterize and compare the sequences of PLCZ1 gene of swamp and riverine buffaloes. Semen samples were collected; total RNA was extracted and reverse-transcribed. PLCZ1 cDNA was then amplified, and submitted for sequencing. Buffalo PLCZ1 gene yielded a sequence of 1905 base pair nucleotides translated into 634 bp amino acids. In general, the buffalo PLCZ1 gene was found to have high sequence identity with cattle and other domestic species. Similarly, significant residues and motifs in PLCZ1 gene sequence are found conserved in water buffaloes. However, there are variations in sequences identified between types of water buffaloes that may play a role in species-specific differences in terms of gene and protein expression, physiological mechanisms, and biological functions. The molecular information on buffalo PLCZ1 gene is highly valuable in subsequent works such as correlation studies on the identified gene variations with semen quality and fertility, and the development of biomarkers for bull fertility.