A comprehensive and single-use foot-and-mouth disease sero-surveillance prototype employing rationally designed multiple viral antigens.

Foot-and-mouth disease (FMD) is a great havoc in agri-business-based countries like Bangladesh, for which existing detection system limits the identification and differentiation of serotypes. In this study, an engineered platform was introduced incorporating serotype-specific FMDV VP1 (structural), serotype-independent VP2 (structural) and 3AB (non-structural) proteins for holistic detection. VP1 sequences were engineered combining sequences of BAN/TA/Dh-301/2016 (serotype O), BAN/CH/Sa-304/2016 (serotype A) and BAN/DH/Sa-318/2016 (serotype Asia1). Consensus 3AB sequence was constructed from the selected prevalent viral genomes. Both VP1 and 3AB along with designed VP2 sequences were optimized for codon usage bias, stable mRNA, secondary and tertiary protein structure. Proteins were synthesized in pET-21a ( +) plasmid vector followed by transformation of Escherichia coli BL21(DE3) and IPTG-induced- expression. The western blot analysis of engineered proteins showed that purified VP1 prominently bound to anti-VP1 antibodies in vaccinated sera, whereas 3AB and VP2 bound anti-3AB and anti-VP2 antibodies, respectively from infected cattle sera, all previously collected during epidemiological investigation. Furthermore, dot blot hybridization confirmed efficient antibody capture ability of the membrane-immobilized proteins. This holistic diagnostic platform justifies a comprehensive prototype diagnostic kit that would be cost-effective and efficient for serotype specific and non-specific FMDV sero-surveillance.

Emergence of a novel sublineage, MYMBD21 under SA-2018 lineage of Foot-and-Mouth Disease Virus serotype O in Bangladesh.

Foot-and-Mouth Disease (FMD) hinders the growth of the livestock industry in endemic countries like Bangladesh. The management and prevention of FMD are severely impacted by the high mutation rate and subsequent frequent generation of newer genotypes of the causative agent, Foot-and-Mouth Disease Virus (FMDV). The current study was conducted in nine districts of Bangladesh during 2019-21 to characterize the circulating FMDV strains based on the VP1 sequence analysis, the major antigenic recognition site providing serotype specificity and high variability of FMDV. This study detected the first emergence of the SA-2018 lineage in Bangladesh along with the predominance of Ind-2001e (or Ind-2001BD1) sublineage of ME-SA topotype under serotype O during 2019-21. The mutational spectrum, evolutionary divergence analysis and multidimensional plotting confirmed the isolates collected from Mymensingh districts, designated as MYMBD21 as a novel sublineage under the SA-2018 lineage. Analysis of the amino acid sequence revealed several changes in the G-H loop, B-C loop and C-terminal region of VP1, revealing a 12-13% divergence from the existing vaccine strains and a 95% VP1 protein homology, with most of the mutations potentially considerable as vaccine escape mutations, evidenced by three-dimensional structural analysis. This is the first report on the emergence of the SA-2018 lineage of ME-SA topotype of FMDV serotype O in Bangladesh, as well as a possible mutational trend towards the emergence of a distinct sublineage under SA-2018 lineage, which calls for in-depth genome-wide analysis and monitoring of the FMD situation in the country to implement a strategic vaccination and effective FMD control program.

A Global Survey of Hypervirulent Aeromonas hydrophila (vAh) Identified vAh Strains in the Lower Mekong River Basin and Diverse Opportunistic Pathogens from Farmed Fish and Other Environmental Sources.

Hypervirulent Aeromonas hydrophila (vAh) has emerged as the etiologic agent of epidemic outbreaks of motile Aeromonas septicemia (MAS) in high-density aquaculture of farmed carp in China and catfish in the United States, which has caused millions of tons of lost fish. We conducted a global survey to better understand the evolution, geographical distribution, and phylogeny of vAh. Aeromonas isolates were isolated from fish that showed clinical symptoms of MAS, and pure cultures were screened for the ability to utilize myo-inositol as the sole carbon source. A total of 113 myo-inositol-utilizing bacterial strains were included in this study, including additional strains obtained from previously published culture collections. Based on a gyrB phylogeny, this collection included 66 A. hydrophila isolates, 48 of which were vAh. This collection also included five new vAh isolates from diseased Pangas catfish (Pangasius pangasius) and striped catfish (Pangasianodon hypophthalmus) obtained in Cambodia and Vietnam, respectively. Genome sequences were generated from representative vAh and non-vAh isolates to evaluate the potential for lateral genetic transfer of the myo-inositol catabolism pathway. Phylogenetic analyses of each of the nine genes required for myo-inositol utilization revealed the close affiliation of vAh strains regardless of geographic origin and suggested lateral genetic transfer of this catabolic pathway from an Enterobacter species. Prediction of virulence factors was conducted to determine differences between vAh and non-vAh strains in terms of virulence and secretion systems. Core genome phylogenetic analyses on vAh isolates and Aeromonas spp. disease isolates (55 in total) were conducted to evaluate the evolutionary relationships among vAh and other Aeromonas sp. isolates, which supported the clonal nature of vAh isolates. IMPORTANCE This global survey of vAh brought together scientists that study fish disease to evaluate the evolution, geographical distribution, phylogeny, and hosts of vAh and other Aeromonas sp. isolates. In addition to vAh isolates from China and the United States, four new vAh isolates were isolated from the lower Mekong River basin in Cambodia and Vietnam, indicating the significant threat of vAh to modern aquaculture and the need for improved biosecurity to prevent vAh spread.