Haemonchus contortus, an obligatory haematophagus worm infection in small ruminants: Population genetics and genetic diversity.

Haemonchus contortus, a stomach worm, is prevalent in ruminants worldwide. They particularly hamper profitable small ruminant production. Here, we estimate the genetic variation of H. contortus collected from slaughtered goats and sheep from various geographic zones of Bangladesh using multiple genes. To perform this, adult parasites were isolated from the abomasum of slaughtered animals (sheep and goats). Among them, 79 male H. contortus were identified by microscopy. Following the extraction of DNA, ITS-2 and cox1 genes were amplified and subsequently considered for sequencing. After alignment and editing, sequences were analyzed to find out sequence variation, diversity pattern of genes, and population genetics of isolates. Among the sequence data, the analyses identified 19 genotypes of ITS-2 and 77 haplotypes of cox1 genes. The diversity of nucleotides was 0.0103 for ITS-2 and 0.029 for cox1 gene. The dendogram constructed by the genotype and haplotype sequences of H. contortus revealed that two populations were circulating in Bangladesh without any demarcation of host and geographic regions. Analysis of population genetics demonstrated a high flow of genes (89.2 %) within the population of the worm in Bangladesh. The Fst value showed very little amount of genetic difference among the worm populations of Bangladesh but marked genetic variation between different continents. The findings are expected to help explain the risks of anthelmintic resistance and the transmission pattern of the parasite, and also provide a control strategy against H. contortus.

Ex-vivo efficacy of commercially available anthelmintics against blood feeding stomach worm Haemonchus contortus of ruminants.

Haemonchus contortus is the most prevalent and pathogenic gastrointestinal nematodes (GINs) in ruminants causing extensive economic losses. It is essential to estimate the efficacy of common commercially available anthelmintics against Haemonchus contortus parasite. Here, we standardized an ex-vivo culture platform for H. contortus and evaluated the efficacy of commonly used anthelmintics namely, albendazole (ABZ), levamisole (LVM), ivermectin (IVM), closantel (CLS) and rafoxanide (RFX). Adult worms were collected from abomasa of slaughtered animals, cultured in MEM, DMEM, M199 or RPMI with or without 20% FBS for up to 72 h. Cultured worms were incubated with ABZ, LVM, IVM, RFX or CLS in DMEM supplemented with 20% FBS at different concentrations (0.5-50 µg/ml) in triplicates and examined at 0, 3, 6, 12, 24, 36 and 48 h post treatment. Of the culture conditions, DMEM supplemented with 20% FBS supported the survival of H. contortus for (P < 0.001) longer period of time which was used in the evaluation of anthelmintics. The efficacy of CLS and RFX were significantly (P < 0.001) higher than other drugs and 100% mortality was observed at 2 µg/ml of CLS and RFX within 12 h post treatment. However, ABZ, LVM, and IVM showed significant effect at the concentration of 50 µg/ml with 48, 36, and 24 h, respectively. Morphological changes included severe cuticle disruption around the buccal cavity, posterior region and vulva as well as loss of cuticle structure integrity coupled with expulsion and fragmentation of digestive components of parasites when treated with 50 µg/ml of ABZ, LVM, and IVM and 2 µg/ml of RFX and CLS. Collectively, DMEM supplemented with 20% FBS can be used as ex-vivo culture platform for maintenance of H. contortus, and RFX and CLS can be used as the promising drugs for the prevention, control and treatment of H. contortus infections.

Prevalence and risk factors of Toxocara vitulorum infection in buffalo calves in coastal, northeastern and northwestern regions of Bangladesh.

Buffalo (Bubalus bubalis) is popularly known as the black gold of South Asia, consisting of 97% of the world buffalo population. Among the parasitic infections, Toxocara vitulorum is one of the most common and harmful parasites of buffalo calves in Bangladesh. A cross-sectional study was conducted to explore the prevalence and associated risk factors of T. vitulorum infection of buffalo calves in four regions of Bangladesh. A total of 1751 fecal samples were collected and examined using flotation followed by the McMaster technique for counting the eggs per gram of feces (EPG) of T. vitulorum. The overall prevalence of T. vitulorum infection in buffalo calves was 22.9%. Significantly (p < 0.001) higher prevalence was found in the Barishal coastal area (35.7%) followed by Chattogram coastal area (29.2%), northeastern region (15.5%) and northwestern region (8.3%). Buffalo calves aged 1-3 months were heavily infected with T. vitulorum (51.7%) which was statistically different (p < 0.001) compared to those >3-6 months (27.6%) and > 6-12 months (6.5%). According to univariate analysis, coastal regions, rainy season, young age, gender, indigenous river type, buffalo calves with poor body condition and soft feces were found significantly associated with T. vitulorum infections. Coastal regions, rainy season and young age were identified as the risk factors of T. vitulorum infection in buffalo calves by final logistic regression model. This study reveals that T. vitulorum infection is endemic in Bangladesh and widely distributed in the coastal regions. Therefore, attention to buffalo calves regarding in-depth clinical effects and current therapeutic approaches against this nematode should be evaluated along with the economic impact of infection. Exploring the genetic diversity of T. vitulorum may help to reveal the host-parasite relationship in the future.

ITS1-PCR based identification of chicken Eimeria species in poultry litter from Mymensingh district, Bangladesh.

OBJECTIVES: The purpose of this study was to determine the species composition of Eimeria circulating in Mymensingh district, Bangladesh, using Internal Transcribed Spacer 1 (ITS1) sequences in polymerase chain reaction (PCR) assay. MATERIALS AND METHODS: Coccidian oocysts were isolated and sporulated in a solution containing 2% potassium dichromate from litter slurry collected from 13 commercially active broiler farms in the research region. Genomic DNA was isolated from sporulated oocysts and used to amplify the Eimeria species-specific ITS1 gene by PCR amplification. Electrophoresis of 1.5% agarose gel was used to visualize the amplified PCR products. RESULTS: In the study samples from Mymensingh district, Bangladesh, the presence of Eimeria brunetti, Eimeria acervulina, Eimeria necatrix, Eimeria mitis, and Eimeria tenella was identified. CONCLUSIONS: The findings of this study may shed light on the zonal approach to chicken coccidiosis control. Additionally, it suggests that ITS1-based PCR might be used in the field to accurately identify Eimeria species.

Effects of single and multiple species inocula of arbuscular mycorrhizal fungi on the salinity tolerance of a Bangladeshi rice (Oryza sativa L.) cultivar.

Soil salinization due to sea level rise and groundwater irrigation has become an important agronomic problem in many parts of the world. Symbiosis between crop species and arbuscular mycorrhizal fungi (AMF) may alleviate salt stress-induced detrimental effects on crop growth and yield, for example, through helping the host plant to selectively absorb potassium while avoiding uptake of excessive sodium. Here, we performed a greenhouse experiment to evaluate growth, grain yield, and salt tolerance of a Bangladeshi rice cultivar under three levels of salt stress (0, 75, and 120 mM) after inoculation with three different AMF species from three different genera (Funnelliformis mosseae (BEG12), Acaulospora laevis (BEG13), and Gigaspora margarita (BEG34)), singly and in combination. We found that under salt stress, AMF inoculation enhanced total chlorophyll concentration, shoot K+/Na+ ratio, and lowered shoot Na+/root Na+ ratio, accompanied by increased root biomass, spikelet fertility, and grain yield compared with the non-inoculated control plants. Specifically, we found that the combination of BEG13 and BEG34 increased rice yield by 125 and 143% as compared with the non-inoculated controls, at the 75 and 120mM salt levels, respectively. In general, the low AMF diversity treatments (one species or a combination of two AMF species) were found to be the most effective in mediating salt stress tolerance for the majority of the measured crop performance variables. Overall, our results indicate that specific AMF species can promote the salt tolerance and productivity of rice, likely by increasing photosynthetic efficiency and restricting Na+ uptake and transport from root to shoot in AMF-inoculated plants.

Variation in arbuscular mycorrhizal fungal communities associated with lowland rice (Oryza sativa) along a gradient of soil salinity and arsenic contamination in Bangladesh.

Rice is an essential food crop that nourishes >50% of the world population. In many regions of Bangladesh rice production is constrained by high soil salinity and heavy metal contamination due to irrigation practices. Plants may naturally overcome such stress through mutualistic interactions with arbuscular mycorrhizal fungi (AMF). Yet, little is known regarding the diversity and composition of AMF communities in rice fields with high saline and arsenic concentration. Here we used high throughput Illumina sequencing to characterize AMF communities in rice roots from 45 Bangladeshi rice fields, along a large geographical gradient of soil salinity and arsenic contamination. We obtained 77 operational taxonomic units (OTUs, based on a sequence similarity threshold of 97%) from eight AMF families, and showed that high soil salinity and arsenic concentration are significantly associated with low AMF diversity in rice roots. Soil salinity and arsenic concentration also explained a large part of the variation in AMF community composition, but also soil pH, moisture, organic matter content and plant available soil phosphorus played an important role. Overall, our study showed that even at very high salinity and arsenic levels, some AMF OTUs are present in rice roots. Their potential role in mediating a reduction of rice stress and arsenic uptake remains to be investigated.