A new species of semiterrestrial gecarcinucid freshwater crab, Terrathelphusa secula, is described from Danum Valley in Sabah, East Malaysia, eastern Borneo. In the form of its carapace, third maxilliped, and male second gonopod, it most closely resembles T. ovis Ng, 1997, and T. telur Ng, 1997, from eastern Sarawak and Brunei, respectively. It differs markedly from these and other congeners in its proportionately much wider carapace, and a male first gonopod that is strongly curved and sickle-shaped.
Corynebacterium pseudotuberculosis is the etiological agent of caseous lymphadenitis that affects sheep and goats. This study was designed to determine the presence of the causative organism in the female reproductive organs and associated lymph nodes in non-pregnant does experimentally inoculated through intradermal route in the chronic form.
Gastrointestinal helminthosis is a global problem in small ruminant production. Most parasites have developed resistance to commonly available anthelminthic compounds, and there is currently an increasing need for new compounds with more efficacies. This study evaluated the in vitro effects of ethanolic extract of Curcuma longa (EECL) as a biological nematicide against third stage Haemonchus larvae (L3) isolated from sheep.
The productivity of smallholder sheep and goat flocks is constrained by high morbidity and mortality of young stock due to helminthosis and coccidiosis. We hypothesized that gastrointestinal parasites are prevalent and may cause severe infections amongst small ruminants in Malaysia. A cross-sectional survey was conducted between March and December 2019 to investigate the prevalence, risk factors, and levels of infection with gastrointestinal strongyle and coccidia in selected smallholder goat flocks in Negeri Sembilan, Malaysia. A total of 257 blood and fecal samples and management data were collected from four farms in Negeri Sembilan. Gastrointestinal parasites were detected by routine sodium chloride floatation, and the McMaster technique was used to quantify the fecal eggs/oocysts per gram outputs (EPG/OPG). The severity of infection was classified as mild (50-799), moderate (800-1200), or severe (>1200). The packed cell volume (PCV) was determined by microhematocrit centrifugation and classified as anemic or non-anemic. Coprological examination revealed an overall prevalence of 78.6% (CI = 72.74-83.44) and 100% flock level prevalence of strongyle and coccidia infection among goats from Negeri Sembilan with a higher infection in flock A-Lenggeng (95.6%) than B-Senawang (87.3%), D-Mendom (80.6%), or C-Seremban (60.0%). The co-infections of strongyle + Eimeria (50.6; CI = 44.50 to 56.64) were more common than single infections of either strongyle (16.7%; CI = 12.66 to 21.78) or Eimeria (4.3%; CI = 2.41 to 7.50). Quantitative analysis has revealed different (p < 0.05) patterns of EPG/OPG in various categories of goats. In total, there were 49.8% mild, 8.6% moderate, and 13.6% severe infections of strongyle and 40.1% mild, 6.6% moderate, and 19.8% severe infections of coccidia among goats. The mean PCV of goats with severe strongyle infection (24.60 ± 0.85) was significantly (p < 0.05) lower than the moderate (26.90 ± 1.15), or mild (28.23 ± 0.50) infections and the uninfected (30.4 ± 0.71). There were increased odds of infection with strongyle and coccidia among female (OR = 3.2) and adult (OR = 11.0) goats from smallholder flocks in Negeri Sembilan. In conclusion, gastrointestinal strongyles and coccidia occur at high frequency among smallholder goats, and there is a higher risk of infection amongst the adult and female stock.
Although Brachiaria decumbens was not toxic when fed to cattle, the infusion of rumen liquor from B. decumbens intoxicated sheep into the rumen of cattle produced evidence suggesting hepatic and renal dysfunction. Several biochemical changes were observed including increases in serum aspartate amino transferase, serum creatinine and blood urea nitrogen and a marked reduction in the plasma bromosulphthalein clearance.
Government-owned small-ruminant breeding farms in Malaysia provide the source of sheep and goats to smallholder farmers in the country. In the eastern Malaysian state of Sabah, high-level stock losses have been recorded on these farms for several years, frequently accompanied by clinical signs indicating pathogenic levels of infections with the nematode parasite Haemonchus contortus. This suggests that their dependence on chemotherapy to control parasite infections had failed. Accordingly, tests for anthelmintic efficacy using the faecal egg count reduction test (FECRT) on the range of drugs used to control nematode parasites were carried out on the five government small-ruminant breeding farms in Sabah. These tests showed a total failure of the benzimidazole, imidothiazole, macrocyclic lactone and salicylanilide groups of anthelmintics to control H. contortus infections of sheep and goats on all farms. Drastic changes in animal management need to be made in an attempt to deal with this situation, for which suggestions are made.
The objective of this study was to investigate association between GDF9 and BMP15 gene polymorphism and litter size in fat-tailed sheep, a total of 97 mature ewes from four breeds (Afshari=19; Baluchi=18; Makui=30 and Mehraban=30) were genotyped for the BMP15 HinfI and GDF9 HhaI polymorphisms by PCR-RFLP technique. The highest and lowest mutant allele frequencies were found in Makui (0.27) and Afshari (0.10) sheep for the BMP15 gene and in Afshari (0.24) and Mehraban (0.18) sheep for the GDF9 gene, respectively. Litter size was significantly influenced by genotype of the ewe for two genes (P < 0.01). Heterozygous genotypes for both loci showed higher litter size than homozygous genotypes (P < 0.01). None of the individuals carried homozygous genotype for both of the GDF9 and BMP15 variants in these breeds. The individuals carrying the mutant allele for one of the investigated candidate gene still showed fertile phenotype. Thus, existence of homozygosity at one of the BMP15 and GDF9 variant is not probably able to block normal hormonal pathway of reproduction in fat-tailed sheep.
Approximately 2,800 fresh dung samples from animals, mainly ruminant livestock, were screened for the presence of nematophagous fungi in Malaysia. Arthrobotrys spp. was noted on numerous occasions, but only one isolate of Duddingtonia flagrans was made. For the purposes of producing sufficient quantities of this fungus for feeding trials in sheep, various, commonly available, cheap plant materials were tested as possible growth substrates. This showed that cereal grains (wheat, millet and rice) were the best media for fungal growth. Pen feeding trials were carried out using sheep, both naturally and experimentally infected with nematode parasites (predominantely Haemonchus contortus), to test the efficiency of D. flagrans when administered either in a grain supplement, or incorporated into a feed block. These showed that the fungus survived gut passage in sheep and that dose rates of approximately 1 x 10(6) D. flagrans spores / animal / day, reduced the percentage of infective larvae developing in faecal cultures by more than 90%. These results indicate that using D. flagrans as a biological control agent of nematode parasites, is a promising alternative to nematode parasite control of small ruminants in Malaysia, where anthelmintic resistance is now a major problem.
Limited information is available on tropical ticks and tick-borne bacteria affecting the health of humans and animals in the Southeast Asia region. Francisella tularensis is a tick-borne bacterium which causes a potentially life-threatening disease known as tularemia. This study was conducted to determine the occurrence of Francisella spp. in questing ticks collected from Malaysian forest reserve areas. A total of 106 ticks (mainly Dermacentor and Haemaphysalis spp.) were examined for Francisella DNA using a Polymerase chain reaction (PCR) assay targeting the bacterial 16S rDNA. Francisella DNA was detected from 12 Dermacentor ticks. Sequence analysis of the amplified 16S rDNA sequences (1035 bp) show >99% identity with that of Francisella endosymbiont reported in a tick from Thailand. A dendrogram constructed based on the bacterial 16S rDNA shows that the Francisella spp. were distantly related to the pathogenic strains of F. tularensis. Three Francisella-positive ticks were identified as Dermacentor atrosignatus, based on sequence analysis of the tick mitochondrial 16S rRNA gene. Further screening of cattle and sheep ticks (Haemaphysalis bispinosa and Rhipicephalus microplus) and animal samples (cattle, sheep, and goats) did not yield any positive findings. Our findings provide the first molecular data on the occurrence of a Francisella strain with unknown pathogenicity in Dermacentor questing ticks in Malaysia.
This study reports the molecular detection of Theileria spp. from six cattle farms, a sheep farm and a goat farm located at different states in Peninsular Malaysia. Animal blood samples were screened for the presence of Theileria DNA using a conventional polymerase chain reaction (PCR) assay. A total of 155 (69.2%) of 224 cattle investigated were PCR-positive for Theileria DNA. The occurrences of Theileria spp. ranged from 17.5% to 100.0% across six cattle farms. Theileria DNA was detected from 90.0% of 40 sheep but none of 40 goats examined in this study. Sequence analyses of amplified 18S rRNA partial fragments (335-338bp) confirmed the identification of Theileria buffeli, Theileria sergenti, and Theileria sinensis in representative samples of cattle and ticks. T. luwenshuni was identified in the infected sheep. The high occurrences of Theileria spp. in our farm animals highlight the needs for appropriate control and preventive measures for theileriosis.
Haemonchus species are major gastro-intestinal parasites affecting ruminants across the world. The present study aimed to assess the sympatric species distribution, genetic diversity, population structure and frequency of β-tubulin isotype 1 alleles associated with benzimidazole resistance. Internal transcribed spacer 2 (ITS2) sequences revealed three sympatric species of Haemonchus, H. contortus, H. placei and H. longistipes with 12 distinct genotypes circulating among ruminant hosts in Pakistan. High genetic variability was observed in Pakistani Haemonchus isolates at nicotine amide dehydrogenase subunit 4 (ND4) and cytochrome oxidase subunit 1 (COI) gene loci. Intra-population diversity parameters were higher in H. contortus isolates than H. placei. Phylogenetic analysis of ND4 and COI sequences did not reveal clustering of haplotypes originating from a particular host indicating high rate of gene flow among Haemonchus parasites infecting sheep, goat and cattle in Pakistan. ND4 and COI haplotypes from Pakistan were compared to sequences of Haemonchus isolates from 11 countries to elucidate the population structure. Multidimensional scaling (MDS) plot of pairwise FST derived from 531 ND4 haplotypes revealed clustering together of H. contortus from Pakistan, China, Malaysia and Italy while the isolates from Yemen and United States were found to be genetically distinct. With respect to H. placei, isolates from Pakistan were found to be genetically differentiated from isolates of other countries. The tests for selective neutrality revealed negative D statistics and did not reveal significant deviations in Pakistani Haemonchus populations while significant deviation (P < 0.05) was observed in Brazilian and Chinese H. contortus populations. Median Joining (MJ) network of ND4 haplotypes revealed Yemenese H. contortus being closer to H. placei cluster. β-tubulin isotype 1 genotyping revealed 7.86% frequency of Y allele associated with benzimidazole resistance at F200Y locus in Pakistani Haemonchus isolates.
Recent reports indicate N. caninum has a possible role in causing abortions in sheep in New Zealand. Knowledge about the mode of transmission of neosporosis in sheep in New Zealand is limited. This study aimed to determine the rate of vertical transmission that would occur in lambs born from experimentally inoculated ewes and to determine if previous inoculation would protect the lambs from N. caninum infection. A group of 50 ewes was divided into 2 groups with one group being inoculated with 5×10(6) N. caninum tachyzoites prior to pregnancy in Year 1. In Year 2, each of these groups was subdivided into 2 groups with one from each original group being inoculated with 1×10(7) N. caninum tachyzoites on Day 120 of gestation. Inoculation of N. caninum tachyzoites into ewes prior to mating resulted in no congenital transmission in lambs born in Year 1 but without further inoculation, 7 out of 11 lambs in Year 2 were positive for N. caninum infection. Ewes that were inoculated in both years resulted in all 12 lambs born in Year 2 being positive for N. caninum infection. This indicates that previous inoculation in Year 1 did not result in any vertical transmission in that year but did not provide any protection against vertical transmission in Year 2. These results suggest that vertical transmission occurs readily once the ewe is infected.
Recent reports indicate Neospora caninum has a possible role in causing abortions in sheep in New Zealand. Knowledge about the epidemiology of neosporosis in sheep is limited. This study aimed to adapt and validate a commercially available ELISA assay as an IgG avidity assay to discriminate between acute (primary and re-inoculated) and chronic N. caninum infections in sheep. In addition, it was used to compare the antibody avidity values between lambs from ewes inoculated with N. caninum either during the pregnancy or in the previous year. The avidity assay was undertaken by using 6M urea for the first wash after incubation with the primary antibody in the commercial ELISA (Chekit* Neospora antibody test kit, IDEXX Laboratories, Australia). Sequential serum samples were obtained from naïve ewes (n=16) experimentally inoculated with live N. caninum tachyzoites. All ewes were seropositive by two weeks post-inoculation and remained seropositive for 20 weeks post-inoculation. There was a linear relationship between time after inoculation and avidity values (p<0.05) over the first 24 weeks. In Week 4, all animals had avidity values <35% and by Week 8, 8/16 animals had avidity values of >35%. These results suggest that an avidity value of <35% indicates a recent primary infection while a value of >35% is indicative of a chronic infection. The assay was then validated using samples from other groups of experimentally inoculated sheep as well as samples from naturally infected ewes. When comparing sample to positive ratio (S/P) and avidity values from lambs born from recently inoculated ewes with those from ewes inoculated the previous year and re-inoculated in the current year, it was possible to differentiate the lambs at 2 weeks of age. Lambs from recently inoculated ewes had low S/P and avidity values at 2 weeks of age which increased by 12 weeks of age. In comparison, lambs from re-inoculated ewes had high S/P and avidity values at 2 weeks of age, due to maternal antibody influence but values were similar to those from lambs that were born from recently inoculated ewes at 12 weeks of age. Avidity values for four naturally infected ewes were all >60% indicating chronic infection. These results suggest that the assay is able to discriminate between recent and chronic infection in sheep as well as able to differentiate lambs with maternal immunity compared to their own de novo immunity. As such it can be utilized to understand the kinetics of N. caninum infection in sheep.
Recent reports from New Zealand indicate Neospora caninum has a possible role in causing abortions in sheep. Transmission of N. caninum via semen has been documented in cattle. This study aimed to investigate if horizontal transmission through semen was also possible in sheep. Initially, 6-month old crossbred ram lambs (n=32), seronegative to N. caninum, were divided into 4 equal groups. Group 1 remained uninoculated whilst the remainder were inoculated with N. caninum tachyzoites intravenously as follows: Group 2 - 50 tachyzoites; Group 3 - 10(3) tachyzoites; Group 4 - 10(7) tachyzoites. Semen samples were collected weekly for 8 weeks for the detection of N. caninum DNA and quantified using quantitative PCR (qPCR). Plasma collected 1 month post-inoculation was subjected to ELISA (IDEXX Chekit) and Western blot. At 2 weeks post-infection, three rams from Group 1 (uninoculated) and three rams from Group 4 (10(7)tachyzoites/ml) were mated with two groups of 16 ewes over two oestrus cycles. Ewe sera collected 1 and 2 months post-mating were tested for seroconversion by ELISA and Western blot. All experimentally infected rams seroconverted by 1 month with ELISA S/P% values ranging from 11% to 36.5% in Group 2, 12-39.5% in Group 3 and 40-81% in Group 4. However, none of the ewes mated with the experimentally infected rams seroconverted. For the Western blot, responses towards immunodominant antigens (IDAs) were observed in ram sera directed against proteins at 10, 17, 21, 25-29, 30, 31, 33 and 37 kDa. Rams in Group 2, 3 and 4 were noted to have at least 3 IDAs present. None of the ewes showed any of the 8 prominent IDAs except for the one at 21 kDa which was seen in 30 out of 32 ewes in both groups. N. caninum DNA was detected intermittently in the ram's semen up to 5 weeks post-inoculation with the concentrations ranging from that equivalent to 1-889 tachyzoites per ml of semen. Low concentrations of N. caninum DNA were also detected in the brain tissue of two rams (Groups 1 and 4). These results suggest that although N. caninum DNA can be found in the semen of experimentally infected rams, the transmission of N. caninum via natural mating is an unlikely event.
The large stomach worm, Haemonchus contortus, commonly known as "the barber's pole worm", is a blood-sucking nematode found in the abomasa of sheep and goats. This work is the first documentation on the ND4 sequences of H. contortus from sheep and goats in Malaysia and Yemen and the results provide a preliminary insight on the genetic differences of H. contortus found in the two countries. In general, this study showed a high degree of diversity and low population structure of this species within the same country in comparison with higher genetic structuring at a wider geographical scale. The results also showed that the majority of genetic variance was within H. contortus populations. The Malaysian sheep and goat populations investigated appeared to share the same isolate of H. contortus while different isolates may be found in Yemen which must be taken into account in the design of an effective control strategy. Analysis of the internal transcribed spacer-2 (ITS-2) confirmed that all samples investigated in this study belonged to H. contortus. However presence of other Haemonchus species parasitizing these two hosts can only be confirmed by further detailed studies.
To determine if toltrazuril was effective in eliminating Neospora caninum infection from congenitally infected lambs. Twenty-eight ewes were allocated to 3 groups where animals in Groups A and B were inoculated with 1 × 10(7)N. caninum tachyzoites on Day 120 of gestation and Group C was maintained as a negative control group. Lambs born from ewes in Group A were treated with toltrazuril (20mg/kg) on Days 0, 7, 14 and 21 after birth. Lambs in Groups B and C were untreated. All lambs in Groups A and B were seropositive at 12 weeks of age. At 12 weeks of age, no differences between lambs in Group A and Group B were observed in serological results (ELISA and western blot), presence of N. caninum-related brain histopathological lesions or the number of organisms detected by qPCR. Group C remained negative for serology, detection of N. caninum DNA as well as histopathology throughout the study. Results indicate that N. caninum congenitally-infected lambs had a continuing infection with N. caninum despite being treated with toltrazuril.
The faecal egg count reduction test (FECRT) was conducted on 39 sheep farms and 9 goat farms located in Peninsular Malaysia. The anthelmintic groups used in these tests were the benzimidazoles, levamisole, the benzimidazole/levamisole combination, macrocyclic lactones and closantel. Results indicated that the prevalence of resistance to the benzimidazole group was high, with approximately 50% of the sheep farms and 75% of the goat farms having resistant nematode parasite populations present. Resistance to levamisole, closantel and ivermectin was also detected. Differentiation of the infective larvae derived from faecal cultures indicated that by far the most predominant parasite species was Haemonchus contortus.
Faecal egg counts were used to study patterns of trichostrongyle infections in sheep and goats according to season, age, pregnancy and lactation on traditional farms in west Malaysia. Haemonchus contortus and Trichostrongylus spp. were the most important strongyles in sheep and in goats, H. contortus, Trichostrongylus spp. and Oesophagostomum spp. were most prevalent. The faecal egg counts of sheep and goats were apparently not influenced by the small seasonal climatic variations. Strongyle infections were acquired at an earlier age in sheep than in goats. Mean faecal egg counts decreased from the age of 8 months onwards in sheep while in goats this occurred from 12-18 months onwards. A periparturient rise in strongyle egg counts was observed in both animal species. Haemonchus contortus was mainly responsible for this rise in faecal egg counts. The results are discussed with reference to control of gastrointestinal strongyle infections in sheep and goats.
The anthelmintic efficacy of benzimidazoles, levamisole, closantel, ivermectin and moxidectin was evaluated on an institutional farm in Malaysia using faecal egg count reduction tests, controlled slaughter trials and an in vitro egg hatch assay. The results of this study indicated simultaneous resistance of Haemonchus contortus against benzimidazoles and ivermectin and of Trichostrongylus colubriformis against benzimidazoles and levaminsole on the same farm. Moxidectin was effective against the ivermectin resistant H. contortus.
Albendazole, oxfendazole, fenbendazole, levamisole, closantel, ivermectin and febantel were administered to sheep on four farms and their efficacy assessed by faecal egg count reduction test. High level of resistance of Haemonchus contortus was found to benzimidazoles (albendazole, oxfendazole, fenbendazole) on all farms and to febantel on the one farm where it was tested. No resistance to closantel and levamisole was observed. Resistance to ivermectin was absent on the three farms examined under this study, but has been reported on the fourth farm in earlier work. It is concluded that anthelmintic resistance to benzimidazoles and the probenzimidazole, febantel, is a serious and widespread problem in H. contortus in sheep in Malaysia.