Characterization, histopathology and immunogenicity of the lumpy skin disease virus isolated during 2019-20 in Bangladesh.

Introduction: Lumpy skin disease (LSD) is a highly contagious vector-borne viral disease of cattle. LSD has emerged in Bangladesh in 2019, causing significant economic losses due to its high morbidity and mortality. This research was designed to isolate, identify, and assess the immunogenicity of LSD virus (LSDV) using nodular tissue samples obtained from affected cattle during the 2019-20 outbreak across nine districts of Bangladesh. Methods: To determine the presence of LSDV in nodular tissues, we initially used iiPCR and PCR, followed by histopathological examination. 151 were positive via iiPCR and PCR among the 180 collected samples. The PCR positive 151 samples were then inoculated into 10-day-old embryonated chicken eggs via the CAM route to isolate LSDV, confirmed through PCR. Subsequently, partial sequencing and phylogenetic analysis of the P32 gene were performed to determine the origin of the circulating LSDV strain. The immunogenicity of selected LSDV strains was assessed through an ELISA test. Results: The PCR results revealed a distinct positive band at 192 bp in both the nodular tissue samples and the LSDV isolated from chicken embryo inoculations. Microscopic analysis of the nodular lesions revealed thickening of the epidermis, ballooning degeneration of keratinocytes, and proliferation of follicular epithelia. Additionally, mononuclear infiltration was observed at the demarcation line between infected and healthy tissue, with necrosis of muscular tissues beneath the epidermis. The LSDV isolate from Bangladesh exhibited a close genetic relationship with LSDV strains isolated from neighboring and other regional countries including India, Myanmar, and Mongolia. This observation strongly suggests the possibility of a transboundary spread of the LSD outbreak in Bangladesh during 2019-2020. The results of the immunogenicity test showed that the serum antibody titer remained at a protective level for up to 18 months following secondary immunization with inactivated LSDV antigen. This finding suggests that the inactivated LSDV antigen could be a potential vaccine candidate to protect cattle in Bangladesh against LSDV. Conclusion: In conclusion, our research successfully isolated, identified, and characterized LSDV in cattle nodular tissues from the 2019-20 outbreak in Bangladesh. Furthermore, it provided insights into the probable origin of the circulating strain and investigated a potential vaccine candidate to protect cattle in the region from LSDV.

Complete genome sequence analysis of the multidrug resistant Aeromonas veronii isolated for the first time from stinging catfish (Shing fish) in Bangladesh.

Objective: Whole genome sequencing (WGS) of Aeromonas veronii Alim_AV_1000 isolated from ulcerative lesions of Shing fish (stringing catfish; Heteropneustes fossilis) was performed during the outbreak year 2021. Materials and Methods: Using next-generation sequencing (Illumina) technology, WGS was accomplished, resulting in the sequencing, assembly, and analysis of the entire genome of the A. veronii strain. Moreover, the genomic features, virulence factors, antimicrobial resistome, and phylogenetic analysis for the molecular evolution of this strain were also examined. Results: The genome size of the A. veronii Alim_AV_1000 strain was 4,494,515 bp, with an average G+C content of 58.87%. Annotation revealed the known transporters and genes linked to virulence, drug targets, and antimicrobial resistance. Conclusion: The findings of the phylogenetic analysis revealed that the strain of the present study has a close relationship with the China strain TH0426 and strain B56. This study provides novel information on A. veronii isolated from Shing fish in Bangladesh.

Epidemiological investigation of lumpy skin disease outbreaks in Bangladeshi cattle during 2019-2020.

Lumpy skin disease (LSD) is an acute infectious viral disease of cattle with a wide distribution that emerged in Bangladesh in 2019, causing huge economic losses. This study was undertaken to investigate the epidemiological features of LSD emergence in nine districts of Bangladesh between December 2019 and December 2020. A total of 8215 cattle from 603 herds were investigated and LSD was diagnosed based on the characteristic clinical findings. A standard questionnaire was administered to collect herd-level data including location, herd size, number of LSD-infected cattle, number died due to LSD, farm type, season, house type, vector presence, sanitation and fly repellent use. Similarly, data on clinical signs, sex, age, animal class and breed of the LSD-infected cattle were also recorded. The herd-level attack risk (%) and mortality risk (%) were calculated based on the number of infected and dead cattle, respectively, as a proportion of total cattle. The herd-level risk factors for LSD were identified using a multivariable Poisson regression model. The most common clinical signs were skin nodules (100%), fever (97.9%) and depression with anorexia and weight loss (97.9%). Crossbred (84.9%) and female (72.2%) cattle were mostly affected by LSD. The overall LSD attack risk, mortality risk and case fatality were 26.5%, 0.26% and 0.97%, respectively. The LSD attack risk was significantly higher in small herds (risk ratio: [RR] 1.39; 95% CI: 1.27; 1.53) than large herds. In addition, significantly higher LSD attack risk was observed in semi-intensive management systems (RR = 1.29; 95% CI: 1.01; 1.64) than intensive management systems. Moreover, it was also significantly higher in hut (RR = 1.81; 95% CI: 1.12; 2.92), temporary (RR = 1.62; 95% CI: 1.21; 2.17) and tin-shed houses (RR = 1.29; 95% CI: 1.11; 1.51) than in semi-building houses. To the best of our knowledge, this is the first detailed epidemiological study of LSD emergence in South Asia. Female crossbred cattle in small herds under semi-intensive management should be prioritized for LSD surveillance and vaccination to prevent further outbreaks and control the impact of the disease in Bangladesh.

Efficacy of bi-valent whole cell inactivated bacterial vaccine against Motile Aeromonas Septicemia (MAS) in cultured catfishes (Heteropneustes fossilis, Clarias batrachus and pangasius pangasius) in Bangladesh.

The Motile Aeromonas Septicemia (MAS) is an important disease of cultured catfishes (Heteropneustes fossilis, Clarias batrachus and Pangasius pangasius), caused by different species of Aeromonas bacteria which have been documented to be higher death rates (≤70%) in Bangladesh since 2016. Present study was conducted to develop bi-valent vaccine using A. hydrophila and A. veronii, and to validate their efficacy via intra-muscular (IM) and oral-routes of immunization in selected species of fishes. Brood fishes of the three species were immunized with three doses of inactivated vaccine (107 CFU /2.3 mg/ml). Hematological parameters of brood fishes and antibody levels (IgM) of broods, their larvae and eggs were determined by ELISA. Additionally, Relative Percent Survivability (RPS) and the IgM levels of the larvae after challenge with virulent A. hydrophila and A. veronii were also evaluated. Findings of this study showed that the lymphocytes, monocytes, granulocytes counts and antibody (IgM) titre of brood fishes, larvae and eggs from the vaccinated fishes were found significantly higher (p< 0.05) compared to the un-vaccinated control groups. Alternatively, antibody levels (IgM) in the larvae of vaccinated group of brood fishes fed with antigen coated feed was exhibited to be remarkably higher (p< 0.05) than the antigen non-fed group. The RPS of larvae of Shing (91.24 ± 2.00%), Magur (88.09 ± 2.88%) and Pangas (93.17 ± 1.52%) was found to be higher in the larvae at 20-day age of vaccinated group compared to non-vaccinated brood fishes group. Findings of this study indicated that the active immunization of brood fishes followed by oral immunization of their larvae feeding with antigen coated feed showed synergistic effect in protecting cultured Shing, Magur and Pangas fishes from frequent attack with Aeromonas spp at any age of their lifetime.

Isolation and molecular detection of Streptococcus agalactiae from popped eye disease of cultured Tilapia and Vietnamese koi fishes in Bangladesh.

OBJECTIVE: Present research aims to isolate, identify, and determine the virulence of the Streptococcus agalactiae (group B Streptococcus; GBS), isolated from popped eye disease affected Tilapia and Vietnamese Koi (V. Koi) fishes. MATERIALS AND METHODS: A total of 330 fish samples were collected, of which Tilapia (n = 180) and V. Koi (n = 150), were collected from 35 affected ponds of four selected districts of Bangladesh. Isolation of the bacterium was done using different culture media (Nutrient broth, Plate count agar, Tryptic Soy Agar, and Blood agar), and identification by using various biochemical tests (conventional and using API 20 Strep kit) and polymerase chain reaction (PCR) using primers against 16S rRNA gene of S. agalactiae. Antibiotic susceptibility of the bacteria was performed using seven different antibiotics disc (Tetracycline, Oxytetracycline, Chlortetracycline, Streptomycin, Ciprofloxacin, Gentamicin, and Neomycin). Virulence of the isolated S. agalactiae was determined by infecting healthy Tilapia and V. Koi fishes through experimental infection. RESULTS: Isolated bacteria were found Gram-positive paired and chained cocci, ß-hemolytic and non-motile. Findings of biochemical and serological tests indicate that the isolated bacterium belongs to Group B Streptococcus of Lancefield classification. PCR result also confirmed that the bacteria were S. agalactiae. The bacterial isolates possessed resistance property against all the seven antibiotics used in this study. The isolated GBS was found highly virulent and showed 80%-90% mortality for Tilapia and V. Koi fishes in experimental infection within 1-6 days of post-infection. CONCLUSION: From the findings of this study, it may be concluded that isolated GBS from the Tilapia and V. Koi fishes were highly virulent and possessed multidrug-resistance properties.